TWI885236B - Lipid compounds and lipid nanoparticle compositions - Google Patents
Lipid compounds and lipid nanoparticle compositionsDownload PDFInfo
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- TWI885236B TWI885236BTW111101514ATW111101514ATWI885236BTW I885236 BTWI885236 BTW I885236BTW 111101514 ATW111101514 ATW 111101514ATW 111101514 ATW111101514 ATW 111101514ATW I885236 BTWI885236 BTW I885236B
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- C07C219/06—Compounds containing amino and esterified hydroxy groups bound to the same carbon skeleton having esterified hydroxy groups and amino groups bound to acyclic carbon atoms of the same carbon skeleton the carbon skeleton being acyclic and saturated having the hydroxy groups esterified by carboxylic acids having the esterifying carboxyl groups bound to hydrogen atoms or to acyclic carbon atoms of an acyclic saturated carbon skeleton
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- C07C229/10—Compounds containing amino and carboxyl groups bound to the same carbon skeleton having amino and carboxyl groups bound to acyclic carbon atoms of the same carbon skeleton the carbon skeleton being acyclic and saturated having only one amino and one carboxyl group bound to the carbon skeleton the nitrogen atom of the amino group being further bound to acyclic carbon atoms or to carbon atoms of rings other than six-membered aromatic rings
- C07C229/14—Compounds containing amino and carboxyl groups bound to the same carbon skeleton having amino and carboxyl groups bound to acyclic carbon atoms of the same carbon skeleton the carbon skeleton being acyclic and saturated having only one amino and one carboxyl group bound to the carbon skeleton the nitrogen atom of the amino group being further bound to acyclic carbon atoms or to carbon atoms of rings other than six-membered aromatic rings to carbon atoms of carbon skeletons containing rings
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- C07C233/35—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by amino groups with the substituted hydrocarbon radical bound to the nitrogen atom of the carboxamide group by an acyclic carbon atom
- C07C233/36—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by amino groups with the substituted hydrocarbon radical bound to the nitrogen atom of the carboxamide group by an acyclic carbon atom having the carbon atom of the carboxamide group bound to a hydrogen atom or to a carbon atom of an acyclic saturated carbon skeleton
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- C07C271/20—Esters of carbamic acids having oxygen atoms of carbamate groups bound to acyclic carbon atoms with the nitrogen atoms of the carbamate groups bound to hydrogen atoms or to acyclic carbon atoms to carbon atoms of hydrocarbon radicals substituted by nitrogen atoms not being part of nitro or nitroso groups
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Abstract
Description
Translated fromChinese本揭示案總體上係關於脂質化合物，其可與其他脂質組分，諸如中性脂質、膽固醇及聚合物結合之脂質組合使用，以形成脂質奈米顆粒用於在活體外及活體內遞送治療劑(例如核酸分子，包括核酸模擬物，諸如鎖核酸(LNA)、肽核酸(PNA)及嗎啉核酸(morpholino))以達成治療或預防目的，包括疫苗接種。The present disclosure generally relates to lipid compounds that can be used in combination with other lipid components, such as neutral lipids, cholesterol, and polymer-bound lipids, to form lipid nanoparticles for in vitro and in vivo delivery of therapeutic agents (e.g., nucleic acid molecules, including nucleic acid mimetics, such as locked nucleic acids (LNA), peptide nucleic acids (PNA), and morpholinos) for therapeutic or preventive purposes, including vaccination.
本說明書係與序列表之電腦可讀形式(CRF)複本一起提呈，該序列表之全部內容併入本文作為參照。This specification is submitted together with a copy of the computer readable form (CRF) of the sequence listing, the entire contents of which are incorporated herein by reference.
治療性核酸具有徹底改變疫苗接種、基因療法、蛋白質替代療法及其他遺傳性疾病治療方法之潛力。自2000年代開始對治療性核酸進行首次臨床研究以來，核酸分子及其遞送方法之設計已取得重大進展。然而，核酸治療劑仍面臨若干挑戰，包括低細胞滲透性及對包括RNA在內之某些核酸分子降解之高敏感性。因此，仍需要開發新的核酸分子，以及促進核酸分子之活體外或活體內遞送以達成治療及/或預防目的之相關方法及組合物。脂質化合物可與其他脂質組分，諸如中性脂質、膽固醇及聚合物結合之脂質組合使用，以形成用於遞送治療劑之脂質奈米顆粒。需要開發新的脂質化合物(例如陽離子脂質化合物)，其提供治療劑之有效遞送、治療劑之足夠活性(例如遞送後mRNA之表現)、最佳藥代動力學及/或其他合適之生理、生物及/或治療特性。Therapeutic nucleic acids have the potential to revolutionize vaccination, gene therapy, protein replacement therapy, and other treatments for genetic diseases. Since the first clinical studies of therapeutic nucleic acids began in the 2000s, there have been significant advances in the design of nucleic acid molecules and methods for their delivery. However, nucleic acid therapeutics still face several challenges, including low cell permeability and high sensitivity to degradation of certain nucleic acid molecules, including RNA. Therefore, there is still a need to develop new nucleic acid molecules, as well as related methods and compositions that facilitate the delivery of nucleic acid molecules in vitro or in vivo for therapeutic and/or preventive purposes. Lipid compounds can be used in combination with other lipid components, such as neutral lipids, cholesterol, and polymer-bound lipids, to form lipid nanoparticles for delivery of therapeutic agents. There is a need to develop new lipid compounds (e.g., cationic lipid compounds) that provide effective delivery of therapeutic agents, adequate activity of therapeutic agents (e.g., expression of mRNA after delivery), optimal pharmacokinetics, and/or other suitable physiological, biological, and/or therapeutic properties.
在一個實施例中，本文提供脂質化合物，包括其醫藥學上可接受之鹽、前藥或立體異構物，其可單獨使用或與其他脂質組分，諸如中性脂質、帶電脂質、類固醇(包括例如所有固醇)及/或其類似物及/或聚合物結合之脂質及/或聚合物組合使用，以形成用於遞送治療劑(例如核酸分子，包括核酸模擬物，諸如鎖核酸(LNA)、肽核酸(PNA)及嗎啉核酸)之脂質奈米顆粒。在一些情況下，使用該等脂質奈米顆粒遞送核酸，諸如反義及/或信使RNA。亦提供使用此等脂質奈米顆粒治療各種疾病或疾患，諸如由感染物及/或蛋白質不足引起之疾病或疾患之方法。In one embodiment, provided herein are lipid compounds, including pharmaceutically acceptable salts, prodrugs or stereoisomers thereof, which can be used alone or in combination with other lipid components, such as neutral lipids, charged lipids, steroids (including, for example, all sterols) and/or analogs thereof and/or polymer-bound lipids and/or polymers, to form lipid nanoparticles for delivering therapeutic agents (e.g., nucleic acid molecules, including nucleic acid mimetics, such as locked nucleic acids (LNA), peptide nucleic acids (PNA) and morpholino nucleic acids). In some cases, the lipid nanoparticles are used to deliver nucleic acids, such as antisense and/or messenger RNA. Methods of using the lipid nanoparticles to treat various diseases or disorders, such as diseases or disorders caused by infectious agents and/or protein deficiencies are also provided.
在一個實施例中，本文提供一種式(I)之化合物：
在一個實施例中，本文提供一種奈米顆粒組合物，其包含本文所提供之化合物以及治療劑或預防劑。在一個實施例中，該治療劑或預防劑包含至少一種編碼抗原或其片段或抗原決定基之mRNA。In one embodiment, a nanoparticle composition is provided herein, which comprises a compound provided herein and a therapeutic agent or a preventive agent. In one embodiment, the therapeutic agent or preventive agent comprises at least one mRNA encoding an antigen or a fragment or antigenic determinant thereof.
熟習此項技術者在考慮以下對特定實施例之詳細闡述之後將對本揭示案之額外特徵顯而易知。Additional features of the present disclosure will become apparent to those skilled in the art after considering the following detailed description of specific embodiments.
本說明書與序列表之電腦可讀格式(CRF)拷貝一起提出申請。該CRF之標題為14639-019-146_SeqListing_ST25.txt，創建於2021年12月20日，大小為627位元組，且其全部內容以引用之方式併入本文中。This specification is filed with a copy of the sequence listing in computer readable format (CRF). The CRF is titled 14639-019-146_SeqListing_ST25.txt, created on December 20, 2021, is 627 bytes in size, and its entire contents are incorporated herein by reference.
5.1一般技術5.1General Technology
本文所闡述或引用之技術及程式包括熟習此項技術者使用習用方法一般很好理解及/或通常採用之技術及程式，諸如例如Sambrook等人，Molecular Cloning：A Laboratory Manual(第3版，2001)；Current Protocols in Molecular Biology(Ausubel等人編，2003)中所闡述之廣泛使用之方法。The techniques and procedures described or referenced herein include those that are generally well understood and/or commonly employed by those skilled in the art using customary methods, such as the widely used methods described in, for example, Sambrook et al.,Molecular Cloning: A Laboratory Manual (3rd ed., 2001);Current Protocols in Molecular Biology (Ausubel et al., eds., 2003).
5.2術語5.2Terminology
除非另有闡述，否則本文中使用之所有技術及科學術語具有與熟習此項技術者通常所理解相同之含義。出於解釋本說明書之目的，將應用以下術語闡述，且在適當時，以單數形式使用之術語亦將包括複數形式，反之亦然。所有專利、申請案、公開之申請案及其他出版物均以全文引用之方式併入。若所闡述之關於術語之任何闡述與以引用之方式併入本文中之任何文件相衝突，則以下文闡述之術語闡述為準。Unless otherwise specified, all technical and scientific terms used in this document have the same meaning as commonly understood by those skilled in the art. For the purpose of interpreting this specification, the following terminology shall apply, and where appropriate, terms used in the singular shall include the plural form and vice versa. All patents, applications, published applications, and other publications are incorporated by reference in their entirety. In the event of any conflict between any statement regarding a term and any document incorporated by reference in this document, the terminology stated below shall prevail.
如本文所使用且除非另有說明，否則術語「脂質」係指一組有機化合物，其包括但不限於脂肪酸酯，且一般以難溶於水但可溶於許多非極性有機溶劑中為特徵。儘管脂質一般具有弱水溶性，但是某些類別之脂質(例如經極性基團改質之脂質，例如DMG-PEG2000)具有有限之水溶性且在某些條件下可溶於水。已知之脂質類型包括生物分子，諸如脂肪酸、蠟、固醇、脂溶性維生素、甘油單酸酯、甘油二酸酯、甘油三酸酯及磷脂。脂質可分為至少三類：(1)「簡單脂質」，包括脂肪及油，以及蠟；(2)「化合物脂質」，包括磷脂及糖脂(例如DMPE-PEG2000)；及(3)「衍生脂質」，諸如類固醇。此外，如本文所使用，脂質亦包括類脂質化合物。術語「類脂質化合物」又簡稱為「類脂質」，係指脂質樣化合物(例如具有脂質樣物理性質之兩親性化合物)。As used herein and unless otherwise specified, the term "lipid" refers to a group of organic compounds that include, but are not limited to, fatty acid esters and are generally characterized by being poorly soluble in water but soluble in many non-polar organic solvents. Although lipids are generally poorly water soluble, certain classes of lipids (e.g., lipids modified with polar groups, such as DMG-PEG2000) have limited water solubility and are soluble in water under certain conditions. Known types of lipids include biomolecules such as fatty acids, waxes, sterols, fat-soluble vitamins, monoglycerides, diglycerides, triglycerides, and phospholipids. Lipids can be divided into at least three categories: (1) "simple lipids," including fats and oils, as well as waxes; (2) "compound lipids," including phospholipids and glycolipids (e.g., DMPE-PEG2000); and (3) "derivatized lipids," such as steroids. In addition, as used herein, lipids also include lipid-like compounds. The term "lipid-like compounds" is also referred to as "lipids" for short, and refers to lipid-like compounds (e.g., amphiphilic compounds with lipid-like physical properties).
術語「脂質奈米顆粒」或「LNP」係指具有至少一個奈米(nm)級尺寸(例如1至1,000nm)之顆粒，其含有一或多種類型之脂質分子。本文所提供之LNP可進一步含有至少一種非脂質有效負載分子(例如一或多種核酸分子)。在一些實施例中，LNP包含部分或完全包封在脂質殼內之非脂質有效負載分子。具體而言，在一些實施例中，其中有效負載係帶負電分子(例如編碼病毒蛋白之mRNA)，且LNP之脂質組分包含至少一種陽離子脂質。不受理論束縛，預期陽離子脂質可與帶負電之有效負載分子相互作用，且在LNP形成期間促進有效負載併入及/或包封至LNP中。可形成如本文所提供之LNP之一部分之其他脂質包括但不限於中性脂質及帶電脂質，諸如類固醇、聚合物結合之脂質及各種兩性離子性脂質。在某些實施例中，根據本揭示案之LNP包含如本文所述之一或多種式(I)(及其子式)之脂質。The term "lipid nanoparticle" or "LNP" refers to a particle having at least one nanometer (nm) size (e.g., 1 to 1,000 nm) that contains one or more types of lipid molecules. The LNP provided herein may further contain at least one non-lipid payload molecule (e.g., one or more nucleic acid molecules). In some embodiments, the LNP comprises a non-lipid payload molecule partially or completely encapsulated in a lipid shell. Specifically, in some embodiments, the payload is a negatively charged molecule (e.g., mRNA encoding a viral protein), and the lipid component of the LNP comprises at least one cationic lipid. Without being bound by theory, it is expected that cationic lipids can interact with negatively charged payload molecules and promote the incorporation and/or encapsulation of the payload into the LNP during LNP formation. Other lipids that may form part of the LNPs provided herein include, but are not limited to, neutral lipids and charged lipids, such as steroids, polymer-bound lipids, and various zwitterionic lipids. In certain embodiments, the LNPs according to the present disclosure comprise one or more lipids of Formula (I) (and its subformulae) as described herein.
術語「陽離子脂質」係指在其環境之任何pH值或氫離子活性下帶正電，或能夠因應於其環境(例如其預定用途之環境)之pH值或氫離子活性而帶正電之脂質。因此，術語「陽離子」涵蓋「永久性陽離子」及「可陽離子化」。在某些實施例中，陽離子脂質中之正電荷源自四級氮原子之存在。在某些實施例中，陽離子脂質包含兩性離子性脂質，該兩性離子性脂質在其預定用途之環境中(例如在生理pH值下)帶正電荷。在某些實施例中，陽離子脂質係如本文所述之一或多種式(I)(及其子式)之脂質。The term "cationic lipid" refers to a lipid that is positively charged at any pH or hydrogen ion activity of its environment, or is capable of being positively charged in response to the pH or hydrogen ion activity of its environment (e.g., the environment of its intended use). Therefore, the term "cationic" encompasses "permanently cationized" and "cationizable." In certain embodiments, the positive charge in the cationic lipid originates from the presence of a quaternary nitrogen atom. In certain embodiments, the cationic lipid comprises a zwitterionic lipid that is positively charged in the environment of its intended use (e.g., at physiological pH). In certain embodiments, the cationic lipid is one or more lipids of formula (I) (and its subformulae) as described herein.
術語「聚合物結合之脂質」係指既包含脂質部分又包含聚合物部分之分子。聚合物結合之脂質之實例係聚乙二醇化脂質(PEG-脂質)，其中聚合物部分包含聚乙二醇。The term "polymer-bound lipid" refers to a molecule that comprises both a lipid portion and a polymer portion. An example of a polymer-bound lipid is a PEGylated lipid (PEG-lipid), wherein the polymer portion comprises polyethylene glycol.
術語「中性脂質」涵蓋在所選pH值下或在所選pH值範圍內以不帶電形式或以中性兩性離子形式存在之任何脂質分子。在一些實施例中，所選之有用pH值或範圍對應於預定脂質用途之環境中之pH條件，諸如生理PH值。作為非限制性實例，可結合本揭示案使用之中性脂質包括但不限於磷脂醯膽鹼，諸如1,2-二硬脂醯基-sn-甘油-3-磷酸膽鹼(DSPC)、1,2-二棕櫚醯基-sn-甘油-3-磷酸膽鹼(DPPC)、1,2-二肉豆蔻醯基-sn-甘油-3-磷酸膽鹼(DMPC)、1-棕櫚醯基-2-油醯基-sn-甘油-3-磷酸膽鹼(POPC)、1,2-二油醯基-sn-甘油-3-磷酸膽鹼(DOPC)；磷脂醯乙醇胺，諸如1,2-二油醯基-sn-甘油-3-磷酸乙醇胺(DOPE)、2-((2,3-雙(油醯氧基)丙基))二甲基銨基)乙基磷酸氫鹽(DOCP)；鞘磷脂(SM)；神經醯胺；類固醇，諸如固醇及其衍生物。本文所提供之中性脂質可為合成的或者衍生自天然來源或化合物(自其分離或改質)。The term "neutral lipid" encompasses any lipid molecule that exists in an uncharged form or in a neutral zwitterionic form at a selected pH or within a selected pH range. In some embodiments, the selected useful pH or range corresponds to the pH conditions in the environment of the intended lipid use, such as a physiological pH. As non-limiting examples, neutral lipids that can be used in conjunction with the present disclosure include, but are not limited to, phospholipid acyl choline, such as 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1-palmitoyl-2-oleoyl-sn-glycero- 3-phosphocholine (POPC), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC); phospholipid ethanolamines, such as 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 2-((2,3-bis(oleyloxy)propyl))dimethylammonium)ethyl hydrogen phosphate (DOCP); sphingomyelin (SM); ceramides; steroids, such as sterols and their derivatives. The neutral lipids provided herein may be synthetic or derived from natural sources or compounds (isolated or modified therefrom).
術語「帶電脂質」涵蓋在所選pH值下或在所選pH值範圍內以帶正電或帶負電形式存在之任何脂質分子。在一些實施例中，所選pH值或範圍對應於預定脂質用途之環境中之pH條件，諸如生理pH值。作為非限制性實例，可與本揭示案結合使用之帶電脂質包括但不限於磷脂醯絲胺酸、磷脂酸、磷脂醯甘油、磷脂醯肌醇、固醇半琥珀酸酯、二烷基三甲基銨-丙烷(例如DOTAP、DOTMA)、二烷基二甲基胺基丙烷、乙基磷酸膽鹼、二甲基胺基乙烷胺基甲醯基固醇(例如DC-Chol)、1,2-二油醯基-sn-甘油-3-磷酸-L-絲胺酸鈉鹽(DOPS-Na)、1,2-二油醯基-sn-甘油-3-磷酸-(1’-外消旋-甘油)鈉鹽(DOPG-Na)及1,2-二油醯基-sn-甘油-3-磷酸鈉鹽(DOPA-Na)。本文所提供之帶電脂質可為合成的或者衍生自天然來源或化合物(自其分離或改質)。The term "charged lipid" encompasses any lipid molecule that exists in a positively or negatively charged form at a selected pH or within a selected pH range. In some embodiments, the selected pH or range corresponds to the pH conditions in the environment in which the lipid is intended to be used, such as a physiological pH. As non-limiting examples, charged lipids that can be used in conjunction with the present disclosure include, but are not limited to, phosphatidylserine, phosphatidic acid, phosphatidylglycerol, phosphatidyl inositol, sterol hemisuccinate, dialkyltrimethylammonium-propane (e.g., DOTAP, DOTMA), dialkyldimethylaminopropane, ethylphosphocholine, dimethylaminoethaneaminoformylsterol (e.g., DC-Chol), 1,2-dioleyl-sn-glycero-3-phospho-L-serine sodium salt (DOPS-Na), 1,2-dioleyl-sn-glycero-3-phospho-(1'-rac-glycero) sodium salt (DOPG-Na), and 1,2-dioleyl-sn-glycero-3-phosphate sodium salt (DOPA-Na). The charged lipids provided herein may be synthetic or derived from (isolated or modified from) natural sources or compounds.
如本文所使用且除非另有說明，否則術語「烷基」係指僅由碳及氫原子組成之飽和直鏈或具支鏈烴鏈基團。在一個實施例中，烷基具有例如一至二十四個碳原子(C1-C24烷基)、四至二十個碳原子(C4-C20烷基)、六至十六個碳原子(C6-C16烷基)、六至九個碳原子(C6-C9烷基)、一至十五個碳原子(C1-C15烷基)、一至十二個碳原子(C1-C12烷基)、一至八個碳原子(C1-C8烷基)或一至六個碳原子(C1-C6烷基)且其藉由單鍵連接至分子其餘部分。烷基之實例包括但不限於甲基、乙基、正丙基、1-甲基乙基(異丙基)、正丁基、正戊基、1,1-二甲基乙基(第三丁基)、3-甲基己基、2-甲基己基及諸如此類。除非另有說明，否則烷基視情況經取代。As used herein and unless otherwise indicated, the term "alkyl" refers to a saturated straight or branched hydrocarbon chain radical consisting solely of carbon and hydrogen atoms. In one embodiment, the alkyl group has, for example, one to twenty-four carbon atoms (C1 -C24 alkyl), four to twenty carbon atoms (C4 -C20 alkyl), six to sixteen carbon atoms (C6 -C16 alkyl), six to nine carbon atoms (C6 -C9 alkyl), one to fifteen carbon atoms (C1 -C15 alkyl), one to twelve carbon atoms (C1 -C12 alkyl), one to eight carbon atoms (C1 -C8 alkyl), or one to six carbon atoms (C1 -C6 alkyl) and is attached to the rest of the molecule by a single bond. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, 1-methylethyl (isopropyl), n-butyl, n-pentyl, 1,1-dimethylethyl (tert-butyl), 3-methylhexyl, 2-methylhexyl, and the like. Unless otherwise specified, alkyl groups are optionally substituted.
如本文所使用且除非另有說明，否則術語「烯基」係指僅由碳及氫原子組成之直鏈或具支鏈烴鏈基團，其含有一或多個碳-碳雙鍵。熟習此項技術者應瞭解，術語「烯基」亦包含具有「順式」及「反式」組態，或者具有「E」及「Z」組態之基團。在一個實施例中，烯基具有例如二至二十四個碳原子(C2-C24烯基)、四至二十個碳原子(C4-C20烯基)、六至十六個碳原子(C6-C16烯基)、六至九個碳原子(C6-C9烯基)、二至十五個碳原子(C2-C15烯基)、二至十二個碳原子(C2-C12烯基)、二至八個碳原子(C2-C8烯基)或二至六個碳原子(C2-C6烯基)且其藉由單鍵連接至分子其餘部分。烯基之實例包括但不限於乙烯基、丙-1-烯基、丁-1-烯基、戊-1-烯基、戊-1,4-二烯基及諸如此類。除非另有說明，否則烯基視情況經取代。As used herein and unless otherwise specified, the term "alkenyl" refers to a straight or branched hydrocarbon chain radical consisting solely of carbon and hydrogen atoms, containing one or more carbon-carbon double bonds. Those skilled in the art will appreciate that the term "alkenyl" also includes groups having "cis" and "trans" configurations, or groups having "E" and "Z" configurations. In one embodiment, the alkenyl group has, for example, two to twenty-four carbon atoms (C2 -C24 alkenyl), four to twenty carbon atoms (C4 -C20 alkenyl), six to sixteen carbon atoms (C6 -C16 alkenyl), six to nine carbon atoms (C6 -C9 alkenyl), two to fifteen carbon atoms (C2 -C15 alkenyl), two to twelve carbon atoms (C2 -C12 alkenyl), two to eight carbon atoms (C2 -C8 alkenyl), or two to six carbon atoms (C2-C6 alkenyl) and is attached to the rest of the molecule by a single bond. Examples of alkenyl groups include, but are not limited to, vinyl, prop-1-enyl, but-1-enyl, pent-1-enyl, pent-1,4-dienyl, and the like. Unless otherwise specified, alkenyl groups are optionally substituted.
如本文所使用且除非另有說明，否則術語「炔基」係指僅由碳及氫原子組成之直鏈或具支鏈烴鏈基團，其含有一或多個碳-碳三鍵。在一個實施例中，炔基具有例如二至二十四個碳原子(C2-C24炔基)、四至二十個碳原子(C4-C20炔基)、六至十六個碳原子(C6-C16炔基)、六至九個碳原子(C6-C9炔基)、二至十五個碳原子(C2-C15炔基)、二至十二個碳原子(C2-C12炔基)、二至八個碳原子(C2-C8炔基)或二至六個碳原子(C2-C6炔基)且其藉由單鍵連接至分子其餘部分。炔基之實例包括但不限於乙炔基、丙炔基、丁炔基、戊炔基及諸如此類。除非另有說明，否則炔基視情況經取代。As used herein and unless otherwise indicated, the term "alkynyl" refers to a straight or branched alkyl chain radical consisting solely of carbon and hydrogen atoms, containing one or more carbon-carbon triple bonds. In one embodiment, the alkynyl group has, for example, two to twenty-four carbon atoms (C2 -C24 alkynyl), four to twenty carbon atoms (C4 -C20 alkynyl), six to sixteen carbon atoms (C6 -C16 alkynyl), six to nine carbon atoms (C6 -C9 alkynyl), two to fifteen carbon atoms (C2 -C15 alkynyl), two to twelve carbon atoms (C2 -C12 alkynyl), two to eight carbon atoms (C2 -C8 alkynyl), or two to six carbon atoms (C2 -C6 alkynyl) and is attached to the rest of the molecule by a single bond. Examples of alkynyl groups include, but are not limited to, ethynyl, propynyl, butynyl, pentynyl, and the like. Unless otherwise specified, alkynyl groups are optionally substituted.
如本文所使用且除非另有說明，否則術語「伸烷基」或「伸烷基鏈」係指將分子其餘部分連接至一或多個基團之直鏈或具支鏈多價(例如二價或三價)烴鏈，其僅由碳及氫組成且為飽和的。在一個實施例中，伸烷基具有例如一至二十四個碳原子(C1-C24伸烷基)、一至十五個碳原子(C1-C15伸烷基)、一至十二個碳原子(C1-C12伸烷基)、一至八個碳原子(C1-C8伸烷基)、一至六個碳原子(C1-C6伸烷基)、二至四個碳原子(C2-C4伸烷基)、一至二個碳原子(C1-C2伸烷基)。伸烷基之實例包括但不限於亞甲基、伸乙基、伸丙基、伸正丁基及諸如此類。伸烷基鏈經由單鍵連接至分子其餘部分，且經由單鍵連接至基團。伸烷基鏈與分子其餘部分及與一或多個基團之連接點可經由鏈內之一個碳或任何兩個(或更多個)碳進行。除非另有說明，否則伸烷基鏈視情況經取代。As used herein and unless otherwise indicated, the term "alkylene" or "alkylene chain" refers to a linear or branched multivalent (e.g., divalent or trivalent) hydrocarbon chain that connects the rest of the molecule to one or more radicals, consisting only of carbon and hydrogen and being saturated. In one embodiment, the alkylene has, for example, one to twenty-four carbon atoms (C1 -C24 alkylene), one to fifteen carbon atoms (C1 -C15 alkylene), one to twelve carbon atoms (C1 -C12 alkylene), one to eight carbon atoms (C1 -C8 alkylene), one to six carbon atoms (C1 -C6 alkylene), two to four carbon atoms (C2 -C4 alkylene), one to two carbon atoms (C1 -C2 alkylene). Examples of alkylene groups include, but are not limited to, methylene, ethylene, propylene, n-butylene, and the like. The alkylene chain is attached to the rest of the molecule via a single bond and to the radical via a single bond. The points of attachment of the alkylene chain to the rest of the molecule and to one or more radicals may be through one carbon or any two (or more) carbons within the chain. Unless otherwise specified, alkylene chains are optionally substituted.
如本文所使用且除非另有說明，否則術語「伸烯基」係指將分子其餘部分連接至一或多個基團之直鏈或具支鏈多價(例如二價或三價)烴鏈，其僅由碳及氫組成且含有一或多個碳-碳雙鍵。在一個實施例中，伸烯基具有例如二至二十四個碳原子(C2-C24伸烯基)、二至十五個碳原子(C2-C15伸烯基)、二至十二個碳原子(C2-C12伸烯基)、二至八個碳原子(C2-C8伸烯基)、二至六個碳原子(C2-C6伸烯基)或二至四個碳原子(C2-C4伸烯基)。伸烯基之實例包括但不限於伸乙烯基、伸丙烯基、伸正丁烯基及諸如此類。伸烯基經由單鍵或雙鍵連接至分子其餘部分，且經由單鍵或雙鍵連接至基團。伸烯基與分子其餘部分及與一或多個基團之連接點可經由鏈內之一個碳或任何兩個(或更多個)碳進行。除非另有說明，否則伸烯基視情況經取代。As used herein and unless otherwise indicated, the term "alkenyl" refers to a straight or branched multivalent (e.g., divalent or trivalent) hydrocarbon chain that links the rest of the molecule to one or more radicals, consisting solely of carbon and hydrogen and containing one or more carbon-carbon double bonds. In one embodiment, the alkenyl has, for example, two to twenty-four carbon atoms (C2 -C24 alkenyl), two to fifteen carbon atoms (C2-C15 alkenyl), two to twelve carbon atoms (C2 -C12 alkenyl), two to eight carbon atoms (C2 -C8 alkenyl), two to six carbon atoms (C2 -C6 alkenyl), or two to four carbon atoms (C2 -C4 alkenyl). Examples of alkenyl groups include, but are not limited to, ethenyl, propenyl, n-butenyl, and the like. Alkenyl groups are attached to the rest of the molecule via a single or double bond and to the radical via a single or double bond. The points of attachment of an alkenyl group to the rest of the molecule and to one or more radicals may be through one carbon or any two (or more) carbons within the chain. Unless otherwise specified, alkenyl groups are optionally substituted.
如本文所使用且除非另有說明，否則術語「環烷基」係指僅由碳及氫原子組成之非芳族飽和單環或多環烴基。環烷基可包括稠合或橋連環系統。在一個實施例中，環烷基具有例如3至15個環碳原子(C3-C15環烷基)、3至10個環碳原子(C3-C10環烷基)或3至8個環碳原子(C3-C8環烷基)。環烷基藉由單鍵連接至分子其餘部分。單環環烷基之實例包括但不限於環丙基、環丁基、環戊基、環己基、環庚基及環辛基。多環環烷基之實例包括但不限於金剛烷基、降莰基、十氫萘基、7,7-二甲基-雙環[2.2.1]庚基及諸如此類。除非另有說明，否則環烷基視情況經取代。As used herein and unless otherwise specified, the term "cycloalkyl" refers to a non-aromatic saturated monocyclic or polycyclic alkyl group consisting only of carbon and hydrogen atoms. Cycloalkyl groups may include fused or bridged ring systems. In one embodiment, the cycloalkyl group has, for example, 3 to 15 ring carbon atoms (C3 -C15 cycloalkyl), 3 to 10 ring carbon atoms (C3 -C10 cycloalkyl), or 3 to 8 ring carbon atoms (C3 -C8 cycloalkyl). The cycloalkyl group is connected to the rest of the molecule by a single bond. Examples of monocyclic cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Examples of polycyclic cycloalkyl groups include, but are not limited to, adamantyl, norbornyl, decahydronaphthyl, 7,7-dimethyl-bicyclo[2.2.1]heptyl, and the like. Unless otherwise specified, cycloalkyl groups are optionally substituted.
如本文所使用且除非另有說明，否則術語「伸環烷基」係多價(例如二價或三價)環烷基。除非另有說明，否則伸環烷基視情況經取代。As used herein and unless otherwise specified, the term "cycloalkylene" refers to a polyvalent (e.g., divalent or trivalent) cycloalkylene group. Unless otherwise specified, a cycloalkylene group is optionally substituted.
如本文所使用且除非另有說明，否則術語「環烯基」係指僅由碳及氫原子組成且包括一或多個碳-碳雙鍵之非芳族單環或多環烴基。環烯基可包括稠合或橋連環系統。在一個實施例中，環烯基具有例如3至15個環碳原子(C3-C15環烯基)、3至10個環碳原子(C3-C10環烯基)或3至8個環碳原子(C3-C8環烯基)。環烯基藉由單鍵連接至分子其餘部分。單環環烯基之實例包括但不限於環丙烯基、環丁烯基、環戊烯基、環己烯基、環庚烯基、環辛烯基及諸如此類。除非另有說明，否則環烯基視情況經取代。As used herein and unless otherwise indicated, the term "cycloalkenyl" refers to a non-aromatic monocyclic or polycyclic hydrocarbon radical consisting solely of carbon and hydrogen atoms and including one or more carbon-carbon double bonds. Cycloalkenyls may include fused or bridged ring systems. In one embodiment, a cycloalkenyl has, for example, 3 to 15 ring carbon atoms (C3 -C15 cycloalkenyl), 3 to 10 ring carbon atoms (C3 -C10 cycloalkenyl), or 3 to 8 ring carbon atoms (C3 -C8 cycloalkenyl). The cycloalkenyl is attached to the rest of the molecule by a single bond. Examples of monocyclic cycloalkenyl groups include, but are not limited to, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclooctenyl, and the like. Unless otherwise specified, a cycloalkenyl group is optionally substituted.
如本文所使用且除非另有說明，否則術語「伸環烯基」係多價(例如二價或三價)環烯基。除非另有說明，否則伸環烯基視情況經取代。As used herein and unless otherwise specified, the term "cycloalkenyl" is a polyvalent (e.g., divalent or trivalent) cycloalkenyl. Unless otherwise specified, a cycloalkenyl is optionally substituted.
如本文所使用且除非另有說明，否則術語「雜環基」係指含有一或多個(例如一個、一個或兩個、一個至三個、或一個至四個)獨立地選自氮、氧、磷及硫之雜原子之非芳族基團單環或多環部分。雜環基可在任何雜原子或碳原子處連接至主結構。雜環基可為單環、雙環、三環、四環或其他多環系統，其中多環系統可為稠合、橋連或螺環系統。雜環基多環系統可在一或多個環中包含一或多個雜原子。雜環基可為飽和或部分不飽和的。飽和雜環烷基可稱為「雜環烷基」。部分不飽和之雜環烷基在雜環基含有至少一個雙鍵時可稱為「雜環烯基」，或在雜環基含有至少一個三鍵時可稱為「雜環炔基」。在一個實施例中，雜環基具有例如3至18個環原子(3至18員雜環基)、4至18個環原子(4至18員雜環基)、5至18個環原子(3至18員雜環基)、4至8個環原子(4至8員雜環基)或5至8個環原子(5至8員雜環基)。當在本文中出現時，數字範圍，諸如「3至18」係指給定範圍中之每個整數；例如，「3至18個環原子」意指雜環基可由3個環原子、4個環原子、5個環原子、6個環原子、7個環原子、8個環原子、9個環原子、10個環原子等(至多且包括18個環原子)組成。雜環基之實例包括但不限於咪唑基、咪唑啶基、
如本文所使用且除非另有說明，否則術語「伸雜環基」係多價(例如二價或三價)雜環基。除非另有說明，否則伸雜環基視情況經取代。As used herein and unless otherwise specified, the term "heterocyclic group" refers to a polyvalent (e.g., divalent or trivalent) heterocyclic group. Unless otherwise specified, a heterocyclic group is optionally substituted.
如本文所使用且除非另有說明，否則術語「芳基」係指含有至少一個芳族烴環之單環芳族基團及/或多環單價芳族基團。在某些實施例中，芳基具有6至18個環碳原子(C6-C18芳基)、6至14個環碳原子(C6-C14芳基)或6至10個環碳原子(C6-C10芳基)。芳基之實例包括但不限於苯基、萘基、茀基、薁基、蒽基、菲基、芘基、聯苯基及聯三苯基。術語「芳基」亦指雙環、三環或其他多環烴環，其中至少一個環係芳族環，且其他環可為飽和、部分不飽和或芳族環，例如二氫萘基、茚基、二氫茚基或四氫萘基(tetrahydronaphthyl/tetralinyl)。除非另有說明，否則芳基視情況經取代。As used herein and unless otherwise indicated, the term "aryl" refers to a monocyclic aromatic group and/or a polycyclic monovalent aromatic group containing at least one aromatic hydrocarbon ring. In certain embodiments, the aryl group has 6 to 18 ring carbon atoms (C6 -C18 aryl), 6 to 14 ring carbon atoms (C6 -C14 aryl), or 6 to 10 ring carbon atoms (C6 -C10 aryl). Examples of aryl groups include, but are not limited to, phenyl, naphthyl, fluorenyl, azulenyl, anthracenyl, phenanthrenyl, pyrenyl, biphenyl, and terphenyl. The term "aryl" also refers to bicyclic, tricyclic or other polycyclic hydrocarbon rings, wherein at least one ring is aromatic and the other rings may be saturated, partially unsaturated or aromatic, such as dihydronaphthyl, indenyl, dihydroindenyl or tetrahydronaphthyl (tetrahydronaphthyl/tetralinyl). Unless otherwise specified, aryl groups are optionally substituted.
如本文所使用且除非另有說明，否則術語「伸芳基」係多價(例如二價或三價)芳基。除非另有說明，否則伸芳基視情況經取代。As used herein and unless otherwise specified, the term "arylene" is a polyvalent (e.g., divalent or trivalent) aromatic group. Unless otherwise specified, an arylene group is optionally substituted.
如本文所使用且除非另有說明，否則術語「雜芳基」係指含有至少一個芳族環之單環芳族基團及/或多環芳族基團，其中至少一個芳族環含有一或多個(例如一個、一個或兩個、一個至三個、或一個至四個)獨立地選自O、S及N之雜原子。雜芳基可在任何雜原子或碳原子處連接至主結構。在某些實施例中，雜芳基具有5至20個、5至15個或5至10個環原子。術語「雜芳基」亦指雙環、三環或其他多環，其中至少一個環係芳族環，且其他環可為飽和、部分不飽和或芳族環，其中至少一個芳族環含有一或多個獨立地選自O、S及N之雜原子。單環雜芳基之實例包括但不限於吡咯基、吡唑基、吡唑啉基、咪唑基、
如本文所使用且除非另有說明，否則術語「伸雜芳基」係多價(例如二價或三價)雜芳基。除非另有說明，否則伸雜芳基視情況經取代。As used herein and unless otherwise specified, the term "heteroaryl" is a polyvalent (e.g., divalent or trivalent) heteroaryl. Unless otherwise specified, heteroaryl is optionally substituted.
當本文所述之基團被稱為「經取代」時，其可經一或多個任何適當之取代基取代。取代基之說明性實例包括但不限於在本文所提供之實例性化合物及實施例中發現之取代基，以及：鹵素原子，諸如F、Cl、Br或I；氰基；側氧基(=O)；羥基(-OH)；烷基；烯基；炔基；環烷基；芳基；-(C=O)OR’；-O(C=O)R’；-C(=O)R’；-OR’；-S(O)xR’；-S-SR’；-C(=O)SR’；-SC(=O)R’；-NR’R’；-NR’C(=O)R’；-C(=O)NR’R’；-NR’C(=O)NR’R’；-OC(=O)NR’R’；-NR’C(=O)OR’；-NR’S(O)xNR’R’；-NR’S(O)xR’；及-S(O)xNR’R’，其中：R’在每次出現時獨立地為H、C1-C15烷基或環烷基，且x係0、1或2。在一些實施例中，取代基係C1-C12烷基。在其他實施例中，取代基係環烷基。在其他實施例中，取代基係鹵基，諸如氟基。在其他實施例中，取代基係側氧基。在其他實施例中，取代基係羥基。在其他實施例中，取代基係烷氧基(-OR’)。在其他實施例中，取代基係羧基。在其他實施例中，取代基係胺基(-NR’R’)。When a group described herein is referred to as "substituted," it may be substituted with one or more any suitable substituents. Illustrative examples of substituents include, but are not limited to, those found in the exemplary compounds and Examples provided herein, as well as: halogen atoms, such as F, Cl, Br, or I; cyano; pendoxy (=O); hydroxyl (-OH); alkyl; alkenyl; alkynyl; cycloalkyl; aryl; -(C=O)OR';-O(C=O)R';-C(=O)R';-OR ';-S(O)xR';-S-SR';-C(=O)SR';-SC(=O)R';-NR'R';-NR'C(=O)R';-C(=O)NR'R';-NR'C(=O)NR'R';-OC(=O)NR'R';-NR'C(=O)OR';-NR'S(O)xNR'R';-NR'S(O)x R'; and -S(O)x NR'R', wherein: R' is independently H, C1 -C15 alkyl or cycloalkyl at each occurrence, and x is 0, 1 or 2. In some embodiments, the substituent is C1 -C12 alkyl. In other embodiments, the substituent is cycloalkyl. In other embodiments, the substituent is halogen, such as fluoro. In other embodiments, the substituent is pendooxy. In other embodiments, the substituent is hydroxyl. In other embodiments, the substituent is alkoxy (-OR'). In other embodiments, the substituent is carboxyl. In other embodiments, the substituent is amino (-NR'R').
如本文所使用且除非另有說明，否則術語「視情況選用之」或「視情況」(例如視情況經取代)意指隨後闡述之事件或情況可能會發生或可能不會發生，且該闡述包括該事件或情況發生之情況及其不發生之情況。舉例而言，「視情況經取代之烷基」意指烷基可經取代或可不經取代，且該闡述包括經取代之烷基及不具有取代之烷基二者。As used herein and unless otherwise indicated, the term "optionally" or "optionally" (e.g., optionally substituted) means that the subsequently described event or circumstance may or may not occur, and the description includes instances where the event or circumstance occurs and instances where it does not occur. For example, "optionally substituted alkyl" means that the alkyl may be substituted or unsubstituted, and the description includes both substituted alkyl and alkyl without substitution.
如本文所使用且除非另有說明，否則術語生物活性化合物之「前藥」係指可在生理條件下或藉由溶劑分解而轉化為生物活性化合物之化合物。在一個實施例中，術語「前藥」係指生物活性化合物之醫藥學上可接受之代謝前驅物。當將前藥投與有需要之個體時，前藥可能為無活性的，但在活體內轉化為生物活性化合物。前藥典型地在活體內迅速轉型以產生母體生物活性化合物，例如藉由在血液中水解而轉型。前藥化合物在哺乳動物生物體中通常提供溶解性、組織相容性或延遲釋放之優點(參見Bundgard,H.,Design of Prodrugs(1985)，第7-9頁，第21-24頁(Elsevier,Amsterdam))。關於前藥之論述提供於Higuchi,T.等人，A.C.S.Symposium Series，第14卷；以及Bioreversible Carriers in Drug Design,Edward B.Roche編輯，American Pharmaceutical Association and Pergamon Press,1987中。As used herein and unless otherwise specified, the term "prodrug" of a biologically active compound refers to a compound that can be converted to a biologically active compound under physiological conditions or by solvent decomposition. In one embodiment, the term "prodrug" refers to a pharmaceutically acceptable metabolic prodrug of a biologically active compound. When the prodrug is administered to a subject in need, the prodrug may be inactive but is converted to a biologically active compound in vivo. The prodrug is typically rapidly transformed in vivo to produce the parent biologically active compound, for example by hydrolysis in the blood. Prodrug compounds generally provide advantages of solubility, tissue compatibility or delayed release in mammalian organisms (see Bundgard, H., Design of Prodrugs (1985), pp. 7-9, pp. 21-24 (Elsevier, Amsterdam)). Discussions on prodrugs are provided in Higuchi, T. et al., A.C.S. Symposium Series, Vol. 14; and Bioreversible Carriers in Drug Design, ed. Edward B. Roche, American Pharmaceutical Association and Pergamon Press, 1987.
在一個實施例中，術語「前藥」亦意圖包括任何共價鍵合之載劑，當將此前藥投與哺乳動物個體時，其在活體內釋放活性化合物。化合物之前藥可藉由修飾化合物中存在之官能基來製備，其方式係使得修飾可在習用操作中或在活體內裂解而得到母體化合物。前藥包括羥基、胺基或巰基鍵合至任何基團之化合物，當將化合物之前藥投與哺乳動物個體時，該基團裂解而分別形成游離羥基、游離胺基或游離巰基。In one embodiment, the term "prodrug" is also intended to include any covalently bonded carrier that releases the active compound in vivo when such prodrug is administered to a mammalian subject. Prodrugs of compounds can be prepared by modifying functional groups present in the compound in such a way that the modifications are cleaved during conventional manipulations or in vivo to yield the parent compound. Prodrugs include compounds in which a hydroxyl, amine or hydroxyl group is bonded to any group that, when the prodrug of the compound is administered to a mammalian subject, is cleaved to form a free hydroxyl, free amine or free hydroxyl group, respectively.
前藥之實例包括但不限於本文所提供化合物中之醇官能基之乙酸酯、甲酸酯及苯甲酸酯衍生物或胺官能基之醯胺衍生物。Examples of prodrugs include, but are not limited to, acetate, formate, and benzoate derivatives of alcohol functional groups or amide derivatives of amine functional groups in the compounds provided herein.
如本文所使用且除非另有說明，否則術語「醫藥學上可接受之鹽」包括酸加成鹽及鹼加成鹽二者。As used herein and unless otherwise indicated, the term "pharmaceutically acceptable salt" includes both acid addition salts and base addition salts.
醫藥學上可接受之酸加成鹽之實例包括但不限於鹽酸、氫溴酸、硫酸、硝酸、磷酸及諸如此類；以及有機酸，諸如但不限於乙酸、2,2-二氯乙酸、己二酸、褐藻酸、抗壞血酸、天冬胺酸、苯磺酸、苯甲酸、4-乙醯胺基苯甲酸、樟腦酸、樟腦-10-磺酸、癸酸、己酸、辛酸、碳酸、肉桂酸、檸檬酸、環拉酸(cyclamic acid)、十二烷基硫酸、乙烷-1,2-二磺酸、乙烷磺酸、2-羥基乙烷磺酸、甲酸、富馬酸、半乳糖二酸、龍膽酸、葡庚糖酸、葡糖酸、葡糖醛酸、麩胺酸、戊二酸、2-側氧基戊二酸、甘油磷酸、乙醇酸、馬尿酸、異丁酸、乳酸、乳糖酸、月桂酸、馬來酸、蘋果酸、丙二酸、扁桃酸、甲烷磺酸、黏液酸、萘-1,5-二磺酸、萘-2-磺酸、1-羥基-2-萘甲酸、菸鹼酸、油酸、乳清酸、草酸、棕櫚酸、帕莫酸(pamoic acid)、丙酸、焦麩胺酸、丙酮酸、水楊酸、4-胺基水楊酸、癸二酸、硬脂酸、琥珀酸、酒石酸、硫氰酸、對甲苯磺酸、三氟乙酸、十一碳烯酸及諸如此類。Examples of pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like; and organic acids such as, but not limited to, acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid, ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid, 4-acetamidobenzoic acid, camphoric acid, camphor-10-sulfonic acid, capric acid, caproic acid, caprylic acid, carbonic acid, cinnamic acid, citric acid, cyclamic acid, acid), dodecyl sulfuric acid, ethane-1,2-disulfonic acid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, formic acid, fumaric acid, galactaric acid, gentianic acid, glucoheptonic acid, gluconic acid, glucuronic acid, glutaric acid, 2-hydroxyglutaric acid, glycerophosphoric acid, glycolic acid, hippuric acid, isobutyric acid, lactic acid, lactobionic acid, lauric acid, maleic acid, apple acid, malonic acid, mandelic acid, methanesulfonic acid, mucic acid, naphthalene-1,5-disulfonic acid, naphthalene-2-sulfonic acid, 1-hydroxy-2-naphthoic acid, niacin, oleic acid, orotic acid, oxalic acid, palmitic acid, pamoic acid acid), propionic acid, pyroglutamic acid, pyruvic acid, salicylic acid, 4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid, tartaric acid, thiocyanic acid, p-toluenesulfonic acid, trifluoroacetic acid, undecylenic acid and the like.
醫藥學上可接受之鹼加成鹽之實例包括但不限於藉由將無機鹼或有機鹼添加至游離酸化合物而製備之鹽。衍生自無機鹼之鹽包括但不限於鈉、鉀、鋰、銨、鈣、鎂、鐵、鋅、銅、錳、鋁鹽及諸如此類。在一個實施例中，無機鹽係銨鹽、鈉鹽、鉀鹽、鈣鹽及鎂鹽。衍生自有機鹼之鹽包括但不限於以下之鹽：一級胺、二級胺及三級胺；經取代胺，包括天然存在之經取代胺；環胺及鹼性離子交換樹脂，諸如氨、異丙胺、三甲胺、二乙胺、三乙胺、三丙胺、二乙醇胺、乙醇胺、丹醇(deanol)、2-二甲基胺基乙醇、2-二乙基胺基乙醇、二環己胺、離胺酸、精胺酸、組胺酸、咖啡因、普魯卡因(procaine)、哈胺(hydrabamine)、膽鹼、甜菜鹼、苯乙苄胺(benethamine)、苄星(benzathine)、乙二胺、葡糖胺、甲基葡糖胺、可可鹼(theobromine)、三乙醇胺、胺基丁三醇、嘌呤、六氫吡
本文所提供之化合物可含有一或多個不對稱中心，且因此可產生鏡像異構物、非鏡像異構物及其他立體異構形式，該等形式可根據絕對立體化學定義為(R)-或(S)-或對於胺基酸可定義為(D)-或(L)-。除非另有說明，否則本文所提供之化合物意圖包括所有此等可能之異構物，以及其外消旋及光學純形式。光學活性(+)與(-)、(R)-與(S)-或(D)-與(L)-異構物可使用掌性合成子或掌性試劑製備，或使用習用技術，例如層析法及分步結晶法拆分。用於製備/分離個別鏡像異構物之習用技術包括自適合之光學純前驅物掌性合成或使用例如掌性高壓液相層析法(HPLC)拆分外消旋物(或鹽或衍生物之外消旋物)。當本文所述化合物含有烯屬雙鍵或其他幾何不對稱中心時，除非另有說明，否則該等化合物意欲包括E及Z幾何異構物。同樣，亦意欲包括所有互變異構形式。The compounds provided herein may contain one or more asymmetric centers and may therefore give rise to mirror image isomers, non-mirror image isomers and other stereoisomeric forms which may be defined by absolute stereochemistry as (R)- or (S)- or, for amino acids, as (D)- or (L)-. Unless otherwise indicated, the compounds provided herein are intended to include all such possible isomers, as well as racemic and optically pure forms thereof. Optically active (+) and (-), (R)- and (S)-, or (D)- and (L)-isomers may be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques such as chromatography and fractional crystallization. Conventional techniques for preparing/isolating individual mirror image isomers include chiral synthesis from suitable optically pure precursors or resolution of racemates (or racemates of salts or derivatives) using, for example, chiral high pressure liquid chromatography (HPLC). When the compounds described herein contain olefinic double bonds or other geometric asymmetric centers, unless otherwise stated, such compounds are intended to include both E and Z geometric isomers. Likewise, all tautomeric forms are intended to be included.
如本文所使用且除非另有說明，否則術語「異構物」係指具有相同分子式之不同化合物。「立體異構物」係僅原子在空間中之排列方式不同之異構物。「阻轉異構物」係由繞單鍵之受阻旋轉得到之立體異構物。「鏡像異構物」係一對互為不可重疊之鏡像之立體異構物。一對鏡像異構物之任何比例之混合物可稱為「外消旋」混合物。「非鏡像異構物」係具有至少兩個不對稱原子但不互為鏡像之立體異構物。As used herein and unless otherwise specified, the term "isomers" refers to different compounds with the same molecular formula. "Stereoisomers" are isomers that differ only in the arrangement of their atoms in space. "Atropisomers" are stereoisomers resulting from hindered rotation about a single bond. "Mirror isomers" are a pair of stereoisomers that are non-superimposable mirror images of each other. A mixture of a pair of mirror isomers in any ratio is called a "racemic" mixture. "Non-mirror isomers" are stereoisomers that have at least two asymmetric atoms but are not mirror images of each other.
「立體異構物」亦可包括E及Z異構物或其混合物，以及順式及反式異構物或其混合物。在某些實施例中，本文所述之化合物分離為E或Z異構物。在其他實施例中，本文所述之化合物係E及Z異構物之混合物。"Stereoisomers" may also include E and Z isomers or mixtures thereof, as well as cis and trans isomers or mixtures thereof. In certain embodiments, the compounds described herein are isolated as E or Z isomers. In other embodiments, the compounds described herein are mixtures of E and Z isomers.
「互變異構物」係指化合物之彼此平衡之異構形式。異構形式之濃度將取決於發現化合物之環境，且可取決於例如化合物係固體抑或在有機溶液或水溶液中而有所不同。"Tautomers" refer to isomeric forms of a compound that are in equilibrium with each other. The concentrations of the isomeric forms will depend on the environment in which the compound is found, and may differ depending on, for example, whether the compound is a solid or in an organic or aqueous solution.
亦應注意，本文所述之化合物可在一或多個原子處含有非天然比例之原子同位素。舉例而言，化合物可用放射性同位素進行放射性標記，諸如氚(3H)、碘-125(125I)、硫-35(35S)或碳-14(14C)，或者可為同位素富集的，諸如氘(2H)、碳-13(13C)或氮-15(15N)。如本文所使用，「同位素體」係同位素富集之化合物。術語「同位素富集」係指原子之同位素組成不同於該原子之天然同位素組成。「同位素富集」亦可指化合物含有之至少一個原子之同位素組成不同於該原子之天然同位素組成。術語「同位素組成」係指給定原子存在之每種同位素之量。經放射性標記及同位素富集之化合物可用作治療劑，例如癌症治療劑；研究試劑，例如結合分析試劑；及診斷劑，例如活體內成像劑。本文所述化合物之所有同位素變異體，無論是否具有放射性，皆意欲涵蓋在本文所提供之實施例之範圍內。在一些實施例中，提供本文所述化合物之同位素體，例如同位素體係氘、碳-13及/或氮-15富集的。如本文所使用，「氘代」係指化合物中之至少一個氫(H)已經氘(以D或2H表示)置換，亦即，化合物在至少一個位置處富含氘。It should also be noted that the compounds described herein may contain unnatural proportions of atomic isotopes at one or more atoms. For example, a compound may be radiolabeled with a radioactive isotope, such as tritium (3H ), iodine-125 (125I ), sulfur-35 (35S ), or carbon-14 (14C ), or may be isotopically enriched, such as deuterium (2H ), carbon-13 (13C ), or nitrogen-15 (15N ). As used herein, an "isotopomer" is an isotopically enriched compound. The term "isotopically enriched" refers to an atom whose isotopic composition is different from the natural isotopic composition of that atom. "Isotopically enriched" may also refer to a compound containing at least one atom whose isotopic composition is different from the natural isotopic composition of that atom. The term "isotopic composition" refers to the amount of each isotope present for a given atom. Radiolabeled and isotopically enriched compounds are useful as therapeutic agents, such as cancer therapeutics; research reagents, such as binding assay reagents; and diagnostic agents, such as in vivo imaging agents. All isotopic variants of the compounds described herein, whether radioactive or not, are intended to be encompassed within the scope of the embodiments provided herein. In some embodiments, isotopologues of the compounds described herein are provided, for example, isotopologues are enriched in deuterium, carbon-13, and/or nitrogen-15. As used herein, "deuterated" means that at least one hydrogen (H) in the compound has been replaced with deuterium (represented as D or2H ), that is, the compound is enriched in deuterium at at least one position.
應注意，若所描繪之結構與該結構之名稱之間存在差異，則應以所描繪之結構為準。It should be noted that if there is a discrepancy between the structure depicted and the name of the structure, the depicted structure shall prevail.
如本文所使用且除非另有說明，否則術語「醫藥學上可接受之載劑、稀釋劑或賦形劑」包括但不限於被美國食品與藥物管理局批準可接受用於人類或家畜之任何佐劑、載劑、賦形劑、助流劑、甜味劑、稀釋劑、防腐劑、染料/著色劑、風味增強劑、表面活性劑、潤濕劑、分散劑、助懸劑、穩定劑、等滲劑、溶劑或乳化劑。As used herein and unless otherwise specified, the term "pharmaceutically acceptable carrier, diluent or excipient" includes but is not limited to any adjuvant, carrier, excipient, glidant, sweetener, diluent, preservative, dye/colorant, flavor enhancer, surfactant, wetting agent, dispersant, suspending agent, stabilizer, isotonic agent, solvent or emulsifier approved by the U.S. Food and Drug Administration for acceptable use in humans or livestock.
術語「組合物」意欲涵蓋含有視情況存在之指定量之指定成分(例如本文所提供之mRNA分子)之產品。The term "composition" is intended to encompass a product containing a specified ingredient (e.g., an mRNA molecule provided herein) in a specified amount, as appropriate.
如本文可互換使用，術語「聚核苷酸」或「核酸」係指任何長度之核苷酸聚合物，且包括例如DNA及RNA。核苷酸可為去氧核糖核苷酸、核糖核苷酸、經修飾之核苷酸或鹼基及/或其類似物，或可藉由DNA或RNA聚合酶或藉由合成反應併入聚合物中之任何受質。聚核苷酸可包含經修飾之核苷酸，諸如甲基化核苷酸及其類似物。核酸可為單股或雙股形式。如本文所使用且除非另有說明，否則「核酸」亦包括核酸模擬物，諸如鎖核酸(LNA)、肽核酸(PNA)及嗎啉核酸。如本文所使用，「寡核苷酸」係指短的合成聚核苷酸，其長度一般但未必少於約200個核苷酸。術語「寡核苷酸」與「聚核苷酸」並非互相排斥的。以上關於聚核苷酸之闡述同樣且完全適於寡核苷酸。除非另有說明，否則本文所揭示之任何單股聚核苷酸序列之左手端為5’端；雙股聚核苷酸序列之左手方向稱為5’方向。新生RNA轉錄物之5’至3’添加方向稱為轉錄方向；DNA股上具有與RNA轉錄物相同之序列且相對於RNA轉錄物之5’端而位於5’端之序列區域稱為「上游序列」；DNA股上具有與RNA轉錄物相同之序列且相對於RNA轉錄物之3’端而位於3’端之序列區域稱為「下游序列」。As used interchangeably herein, the terms "polynucleotide" or "nucleic acid" refer to polymers of nucleotides of any length, and include, for example, DNA and RNA. Nucleotides may be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases and/or analogs thereof, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase or by a synthetic reaction. Polynucleotides may include modified nucleotides, such as methylated nucleotides and their analogs. Nucleic acids may be in single-stranded or double-stranded form. As used herein and unless otherwise specified, "nucleic acid" also includes nucleic acid mimetics, such as locked nucleic acids (LNA), peptide nucleic acids (PNA), and morpholino nucleic acids. As used herein, "oligonucleotide" refers to short synthetic polynucleotides, which are generally, but not necessarily, less than about 200 nucleotides in length. The terms "oligonucleotide" and "polynucleotide" are not mutually exclusive. The above description of polynucleotides applies equally and fully to oligonucleotides. Unless otherwise specified, the left-hand end of any single-stranded polynucleotide sequence disclosed herein is the 5' end; the left-hand direction of a double-stranded polynucleotide sequence is referred to as the 5' direction. The direction of 5' to 3' addition of a nascent RNA transcript is referred to as the transcription direction; the sequence region on a DNA strand that has the same sequence as an RNA transcript and is located at the 5' end relative to the 5' end of the RNA transcript is referred to as the "upstream sequence"; the sequence region on a DNA strand that has the same sequence as an RNA transcript and is located at the 3' end relative to the 3' end of the RNA transcript is referred to as the "downstream sequence".
「分離之核酸」係指與天然地伴隨天然序列之其他基因體DNA序列以及蛋白質或複合物(諸如核糖體及聚合酶)實質上分離之核酸，例如RNA、DNA或混合核酸。「分離」之核酸分子係與存在於核酸分子之天然來源中之其他核酸分子分離之核酸分子。此外，當藉由重組技術製造時，「分離」之核酸分子，諸如mRNA分子，可實質上不含其他細胞材料或培養基，或者當化學合成時，其可實質上不含化學前驅物或其他化學品。在特定實施例中，本文所述之編碼抗原之一或多種核酸分子係分離或純化的。該術語包括已自其天然存在之環境移除之核酸序列，且包括重組或選殖之DNA或RNA分離物以及化學合成之類似物或由異源系統生物合成之類似物。實質上純之分子可包括分子之分離形式。An "isolated nucleic acid" refers to a nucleic acid, such as RNA, DNA, or mixed nucleic acid, that is substantially separated from other genomic DNA sequences and proteins or complexes (such as ribosomes and polymerases) that naturally accompany the native sequence. An "isolated" nucleic acid molecule is one that is separated from other nucleic acid molecules present in the natural source of the nucleic acid molecule. In addition, an "isolated" nucleic acid molecule, such as an mRNA molecule, may be substantially free of other cellular material or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemicals when chemically synthesized. In specific embodiments, one or more nucleic acid molecules encoding antigens described herein are isolated or purified. The term includes nucleic acid sequences that have been removed from their naturally occurring environment and includes recombinant or cloned DNA or RNA isolates as well as chemically synthesized analogs or analogs biologically synthesized by heterologous systems. A substantially pure molecule may include isolated forms of the molecule.
術語「編碼核酸」或其語法等效物當用於指核酸分子時包括：(a)處於天然狀態或藉由熟習此項技術者熟知之方法操作時可轉錄產生mRNA且接著轉譯成肽及/或多肽之核酸分子；及(b)mRNA分子本身。反義股係此一核酸分子之互補序列，且可由其推斷出編碼序列。術語「編碼區」係指編碼核酸序列中轉譯成肽或多肽之部分。術語「非轉譯區」或「UTR」係指編碼核酸中不轉譯成肽或多肽之部分。取決於UTR相對於核酸分子之編碼區之取向，UTR若位於編碼區5’端，則稱為5’-UTR，且UTR若位於編碼區3’端，則稱為3’-UTR。The term "coding nucleic acid" or its grammatical equivalents when used to refer to nucleic acid molecules include: (a) nucleic acid molecules that can be transcribed to produce mRNA and then translated into peptides and/or polypeptides when in their natural state or manipulated by methods familiar to those skilled in the art; and (b) mRNA molecules themselves. The antisense strand is the complementary sequence of such a nucleic acid molecule, and the coding sequence can be inferred from it. The term "coding region" refers to the portion of the coding nucleic acid sequence that is translated into a peptide or polypeptide. The term "untranslated region" or "UTR" refers to the portion of the coding nucleic acid that is not translated into a peptide or polypeptide. Depending on the orientation of the UTR relative to the coding region of the nucleic acid molecule, the UTR is called a 5'-UTR if it is located at the 5' end of the coding region, and a 3'-UTR if it is located at the 3' end of the coding region.
如本文所使用，術語「mRNA」係指包含一或多個開放閱讀框(ORF)之信使RNA分子，其可經具有該mRNA之細胞或生物體轉譯以產生一或多種肽或蛋白質產物。含有一或多個ORF之區域稱為mRNA分子之編碼區。在某些實施例中，mRNA分子進一步包含一或多個非轉譯區(UTR)。As used herein, the term "mRNA" refers to a messenger RNA molecule comprising one or more open reading frames (ORFs), which can be translated by a cell or organism having the mRNA to produce one or more peptide or protein products. The region containing one or more ORFs is called the coding region of the mRNA molecule. In certain embodiments, the mRNA molecule further comprises one or more untranslated regions (UTRs).
在某些實施例中，mRNA係僅包含一個ORF之單順反子mRNA。在某些實施例中，單順反子mRNA編碼包含選定抗原(例如致病性抗原或腫瘤相關抗原)之至少一個抗原決定基之肽或蛋白質。在其他實施例中，mRNA係包含兩個或更多個ORF之多順反子mRNA。在某些實施例中，多順反子mRNA編碼可彼此相同或不同之兩種或更多種肽或蛋白質。在某些實施例中，由多順反子mRNA編碼之每種肽或蛋白質包含選定抗原之至少一個抗原決定基。在某些實施例中，由多順反子mRNA編碼之不同肽或蛋白質各自包含不同抗原之至少一個抗原決定基。在本文所述實施例中之任一個中，該至少一個抗原決定基可為抗原之至少2個、至少3個、至少4個、至少5個、至少6個、至少7個、至少8個、至少9個或至少10個抗原決定基。In some embodiments, the mRNA is a monostronic mRNA comprising only one ORF. In some embodiments, the monostronic mRNA encodes a peptide or protein comprising at least one antigenic determinant of a selected antigen (e.g., a pathogenic antigen or a tumor-associated antigen). In other embodiments, the mRNA is a polystronic mRNA comprising two or more ORFs. In some embodiments, the polystronic mRNA encodes two or more peptides or proteins that may be identical or different from each other. In some embodiments, each peptide or protein encoded by the polystronic mRNA comprises at least one antigenic determinant of a selected antigen. In some embodiments, different peptides or proteins encoded by the polystronic mRNA each comprise at least one antigenic determinant of a different antigen. In any of the embodiments described herein, the at least one antigenic determinant may be at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, or at least 10 antigenic determinants of an antigen.
術語「核鹼基」涵蓋嘌呤及嘧啶，包括天然化合物腺嘌呤、胸腺嘧啶、鳥嘌呤、胞嘧啶、尿嘧啶、肌苷以及其天然或合成類似物或衍生物。The term "nucleobase" covers purines and pyrimidines, including the natural compounds adenine, thymine, guanine, cytosine, uracil, inosine, and their natural or synthetic analogs or derivatives.
如本文所使用，術語「功能性核苷酸類似物」係指經典核苷酸A、G、C、U或T之經修飾型式，該型式(a)保留對應經典核苷酸之鹼基配對特性，且(b)含有至少一種對對應天然核苷酸的(i)核鹼基、(ii)糖基、(iii)磷酸酯基或(iv)(i)至(iii)之任何組合之化學修飾。如本文所使用，鹼基配對不僅涵蓋經典沃森-克裡克(Watson-Crick)腺嘌呤-胸腺嘧啶、腺嘌呤-尿嘧啶或鳥嘌呤-胞嘧啶鹼基對，而且亦涵蓋在經典核苷酸與功能性核苷酸類似物之間或在一對功能性核苷酸類似物之間形成之鹼基對，其中氫鍵供體與氫鍵受體之佈置允許在經修飾之核鹼基與經典核鹼基之間或在兩個互補之經修飾核鹼基結構之間形成氫鍵。舉例而言，鳥苷(G)之功能性類似物保留與胞嘧啶(C)或胞嘧啶之功能性類似物鹼基配對之能力。該非經典鹼基配對之一個實例係經修飾核苷酸肌苷與腺嘌呤、胞嘧啶或尿嘧啶之間之鹼基配對。如本文所述，功能性核苷酸類似物可為天然存在或非天然存在的。因此，含有功能性核苷酸類似物之核酸分子可具有至少一個經修飾之核鹼基、糖基及/或核苷間鍵聯。本文提供對核酸分子之核鹼基、糖基或核苷間鍵聯之實例性化學修飾。As used herein, the term "functional nucleotide analog" refers to a modified form of a classical nucleotide A, G, C, U or T that (a) retains the base pairing properties of the corresponding classical nucleotide and (b) contains at least one chemical modification of the (i) nucleobase, (ii) sugar group, (iii) phosphate group, or (iv) any combination of (i) to (iii) of the corresponding natural nucleotide. As used herein, base pairing encompasses not only the classical Watson-Crick adenine-thymine, adenine-uracil or guanine-cytosine base pairs, but also base pairs formed between a classical nucleotide and a functional nucleotide analog or between a pair of functional nucleotide analogs, wherein the arrangement of hydrogen bond donors and hydrogen bond acceptors allows the formation of hydrogen bonds between a modified nucleobase and a classical nucleobase or between two complementary modified nucleobase structures. For example, a functional analog of guanosine (G) retains the ability to base pair with cytosine (C) or a functional analog of cytosine. An example of such non-classical base pairing is base pairing between modified nucleotides inosine and adenine, cytosine, or uracil. As described herein, functional nucleotide analogs can be naturally occurring or non-naturally occurring. Thus, nucleic acid molecules containing functional nucleotide analogs can have at least one modified nucleobase, sugar group, and/or internucleoside linkage. Exemplary chemical modifications of nucleobases, sugar groups, or internucleoside linkages of nucleic acid molecules are provided herein.
如本文所使用，術語「轉譯強化子元件」、「TEE」及「轉譯強化子」係指核酸分子中用於促進核酸之編碼序列轉譯成蛋白質或肽產物，諸如經由帽依賴性或非帽依賴性轉譯而轉譯成蛋白質或肽產物之區域。TEE典型地位於核酸分子(例如mRNA)之UTR區，且增強位於上游或下游之編碼序列之轉譯水準。舉例而言，核酸分子之5’-UTR中之TEE可位於核酸分子之啟動子與起始密碼子之間。各種TEE序列係此項技術中已知的(Wellensiek等人，Genome-wide profiling of human cap-independent translation-enhancing elements,Nature Methods,2013年8月；10(8)：747-750；Chappell等人，PNAS,2004年6月29日，101(26)9590-9594)。已知一些TEE在多個物種中係保守的(Pánek等人，Nucleic Acids Research，第41卷，第16期，2013年9月1日，第7625-7634頁)。As used herein, the terms "translational enhancer element", "TEE" and "translational enhancer" refer to a region in a nucleic acid molecule that is used to promote the translation of a coding sequence of the nucleic acid into a protein or peptide product, such as via cap-dependent or cap-independent translation. TEEs are typically located in the UTR region of a nucleic acid molecule (e.g., mRNA) and enhance the translation level of a coding sequence located upstream or downstream. For example, a TEE in the 5'-UTR of a nucleic acid molecule can be located between the promoter and the start codon of the nucleic acid molecule. Various TEE sequences are known in the art (Wellensiek et al., Genome-wide profiling of human cap-independent translation-enhancing elements,Nature Methods , 2013 Aug; 10(8): 747-750; Chappell et al., PNAS, 2004 Jun 29, 101(26): 9590-9594). Some TEEs are known to be conserved in multiple species (Pánek et al.,Nucleic Acids Research , Vol. 41, No. 16, 2013 Sept. 1, pp. 7625-7634).
如本文所使用，術語「莖-環序列」係指具有至少兩個區域之單股聚核苷酸序列，當以相反方向閱讀時，該兩個區域彼此互補或實質上互補，且因此能夠彼此鹼基配對形成至少一個雙螺旋及未配對之環。所得到之結構稱為莖-環結構、髮夾或髮夾環，其係在許多RNA分子中發現之二級結構。As used herein, the term "stem-loop sequence" refers to a single-stranded polynucleotide sequence having at least two regions that complement or substantially complement each other when read in opposite directions and are therefore capable of base pairing with each other to form at least one double helix and an unpaired loop. The resulting structure is called a stem-loop structure, hairpin, or hairpin loop, which is a secondary structure found in many RNA molecules.
如本文所使用，術語「肽」係指含有二至五十(2-50)個經一或多個共價肽鍵連接之胺基酸殘基之聚合物。該等術語適於天然存在之胺基酸聚合物以及一或多個胺基酸殘基係非天然存在之胺基酸(例如胺基酸類似物或非天然胺基酸)之胺基酸聚合物。As used herein, the term "peptide" refers to a polymer containing from two to fifty (2-50) amino acid residues linked by one or more covalent peptide bonds. These terms apply to naturally occurring amino acid polymers as well as amino acid polymers in which one or more amino acid residues are non-naturally occurring amino acids (e.g., amino acid analogs or non-natural amino acids).
術語「多肽」與「蛋白質」在本文中可互換使用，指具有超過五十(50)個由共價肽鍵連接之胺基酸殘基之聚合物。亦即，針對多肽之闡述同樣適於針對蛋白質之闡述，反之亦然。該等術語適於天然存在之胺基酸聚合物以及一或多個胺基酸殘基係非天然存在之胺基酸(例如胺基酸類似物)之胺基酸聚合物。如本文所使用，該等術語涵蓋任何長度之胺基酸鏈，包括全長蛋白質(例如抗原)。The terms "polypeptide" and "protein" are used interchangeably herein to refer to polymers having more than fifty (50) amino acid residues linked by covalent peptide bonds. That is, descriptions of polypeptides apply equally to descriptions of proteins, and vice versa. These terms apply to naturally occurring amino acid polymers as well as amino acid polymers in which one or more amino acid residues are non-naturally occurring amino acids (e.g., amino acid analogs). As used herein, these terms encompass amino acid chains of any length, including full-length proteins (e.g., antigens).
術語「抗原」係指能夠被個體之免疫系統(包括適應性免疫系統)識別之物質，且在個體與抗原接觸之後能夠觸發免疫反應(包括抗原特異性免疫反應)。在某些實施例中，抗原係與患病細胞，諸如感染病原體之細胞或贅生性細胞相關之蛋白質(例如腫瘤相關抗原(TAA))。The term "antigen" refers to a substance that can be recognized by an individual's immune system (including the adaptive immune system) and can trigger an immune response (including an antigen-specific immune response) after the individual comes into contact with the antigen. In certain embodiments, the antigen is a protein associated with diseased cells, such as cells infected with pathogens or proliferative cells (e.g., tumor-associated antigens (TAAs)).
在肽或多肽之情況下，如本文所使用，術語「片段」係指包含少於全長之胺基酸序列之肽或多肽。此一片段可例如來自於胺基末端之截短、羧基末端之截短及/或胺基酸序列中殘基之內部缺失。片段可例如由選擇性RNA剪接或由活體內蛋白酶活性產生。在某些實施例中，片段係指包括含多肽之胺基酸序列之至少5個連續胺基酸殘基、至少10個連續胺基酸殘基、至少15個連續胺基酸殘基、至少20個連續胺基酸殘基、至少25個連續胺基酸殘基、至少30個連續胺基酸殘基、至少40個連續胺基酸殘基、至少50個連續胺基酸殘基、至少60個連續胺基酸殘基、至少70個連續胺基酸殘基、至少80個連續胺基酸殘基、至少90個連續胺基酸殘基、至少100個連續胺基酸殘基、至少125個連續胺基酸殘基、至少150個連續胺基酸殘基、至少175個連續胺基酸殘基、至少200個連續胺基酸殘基、至少250個、至少300個、至少350個、至少400個、至少450個、至少500個、至少550個、至少600個、至少650個、至少700個、至少750個、至少800個、至少850個、至少900個或至少950個連續胺基酸殘基之胺基酸序列之多肽。在特定實施例中，多肽之片段保留該多肽之至少1種、至少2種、至少3種或更多種功能。In the case of a peptide or polypeptide, as used herein, the term "fragment" refers to a peptide or polypeptide comprising less than the full-length amino acid sequence. Such a fragment may, for example, result from truncation of the amino terminus, truncation of the carboxyl terminus, and/or internal deletion of residues in the amino acid sequence. Fragments may, for example, be produced by alternative RNA splicing or by in vivo protease activity. In certain embodiments, a fragment refers to at least 5 consecutive amino acid residues, at least 10 consecutive amino acid residues, at least 15 consecutive amino acid residues, at least 20 consecutive amino acid residues, at least 25 consecutive amino acid residues, at least 30 consecutive amino acid residues, at least 40 consecutive amino acid residues, at least 50 consecutive amino acid residues, at least 60 consecutive amino acid residues, at least 70 consecutive amino acid residues, at least 80 consecutive amino acid residues, at least 90 consecutive amino acid residues, at least 100 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 100 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 100 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 150 consecutive amino acid residues A polypeptide having an amino acid sequence of at least 125 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 175 consecutive amino acid residues, at least 200 consecutive amino acid residues, at least 250, at least 300, at least 350, at least 400, at least 450, at least 500, at least 550, at least 600, at least 650, at least 700, at least 750, at least 800, at least 850, at least 900 or at least 950 consecutive amino acid residues. In a specific embodiment, the fragment of the polypeptide retains at least 1, at least 2, at least 3 or more functions of the polypeptide.
「抗原決定基」係抗原分子表面上與單個抗體分子結合之位點，諸如抗原表面上能夠結合至抗體之一或多個抗原結合區之局部區域，且其在動物體內，諸如在哺乳動物體內(例如人類體內)具有抗原或免疫原性活性，能夠引起免疫反應。具有免疫原性活性之抗原決定基係多肽之在動物體內引起抗體反應之部分。具有抗原活性之抗原決定基係多肽之藉由此項技術中熟知之任何方法，包括例如藉由免疫分析確定之抗體所結合之部分。抗原性抗原決定基未必具有免疫原性。抗原決定基通常由分子之化學活性表面基團，諸如胺基酸或糖側鏈組成，且具有特定之三維結構特徵以及特定之電荷特徵。抗體抗原決定基可為線性抗原決定基或構形抗原決定基。線性抗原決定基係由蛋白質中之連續胺基酸序列形成的。構形抗原決定基係由在蛋白質序列中不連續但在蛋白質摺疊成其三維結構時結合在一起之胺基酸形成的。當蛋白質之三維結構呈改變之構形時，諸如在活化或結合另一種蛋白質或配位體之後，形成誘導性抗原決定基。在某些實施例中，抗原決定基係多肽之三維表面特徵。在其他實施例中，抗原決定基係多肽之線性特徵。一般而言，抗原具有若干或許多不同之抗原決定基，且可與許多不同之抗體反應。"Antigenic determinant" is a site on the surface of an antigen molecule that binds to a single antibody molecule, such as a localized area on the surface of an antigen that is capable of binding to one or more antigen binding regions of an antibody, and which has antigenic or immunogenic activity in an animal, such as a mammal (e.g., a human), and is capable of eliciting an immune response. An antigenic determinant with immunogenic activity is a portion of a polypeptide that elicits an antibody response in an animal. An antigenic determinant with antigenic activity is a portion of a polypeptide that is bound by an antibody as determined by any method known in the art, including, for example, by immunoassay. An antigenic epitope is not necessarily immunogenic. An antigenic determinant is usually composed of chemically active surface groups of a molecule, such as amino acids or sugar side chains, and has specific three-dimensional structural characteristics as well as specific charge characteristics. Antibody epitopes can be linear epitopes or conformational epitopes. Linear epitopes are formed by consecutive amino acid sequences in proteins. Conformational epitopes are formed by amino acids that are not consecutive in the protein sequence but are joined together when the protein folds into its three-dimensional structure. Inducing epitopes are formed when the three-dimensional structure of a protein is in an altered conformation, such as after activation or binding to another protein or ligand. In certain embodiments, the epitope is a three-dimensional surface feature of a polypeptide. In other embodiments, the epitope is a linear feature of a polypeptide. In general, an antigen has several or many different epitopes and can react with many different antibodies.
如本文所使用，術語「基因疫苗」係指包含至少一種核酸分子之治療性或預防性組合物，該至少一種核酸分子編碼與目標疾病(例如感染性疾病或贅生性疾病)相關之抗原。向個體投與疫苗(「疫苗接種」)允許產生編碼之肽或蛋白質，藉此在個體體內引起針對目標疾病之免疫反應。在某些實施例中，免疫反應包括適應性免疫反應，諸如產生針對編碼之抗原之抗體，及/或能夠特異性消除表現該抗原之患病細胞之免疫細胞之活化及增殖。在某些實施例中，免疫反應進一步包括先天免疫反應。根據本揭示案，疫苗可在目標疾病之臨床症狀發作之前或之後投與個體。在一些實施例中，對健康或無症狀個體進行疫苗接種使經疫苗接種之個體對目標疾病之發展具有免疫性或不太敏感。在一些實施例中，對顯示疾病症狀之個體進行疫苗接種改善經疫苗接種之個體之疾病狀況或治療該疾病。As used herein, the term "genetic vaccine" refers to a therapeutic or prophylactic composition comprising at least one nucleic acid molecule that encodes an antigen associated with a target disease (e.g., an infectious disease or a proliferative disease). Administration of a vaccine to an individual ("vaccination") allows the production of the encoded peptide or protein, thereby eliciting an immune response in the individual against the target disease. In certain embodiments, the immune response includes an adaptive immune response, such as the production of antibodies against the encoded antigen, and/or the activation and proliferation of immune cells that are capable of specifically eliminating diseased cells expressing the antigen. In certain embodiments, the immune response further includes an innate immune response. According to the present disclosure, the vaccine can be administered to an individual before or after the onset of clinical symptoms of the target disease. In some embodiments, vaccination of a healthy or asymptomatic individual renders the vaccinated individual immune or less susceptible to the development of a target disease. In some embodiments, vaccination of an individual showing symptoms of a disease improves the disease condition of the vaccinated individual or treats the disease.
術語「先天免疫反應」及「先天免疫」係此項技術中公認的，且係指身體之免疫系統在識別出病原體相關分子模式時啟動之非特異性防禦機制，其涉及不同形式之細胞活動，包括經由各種路徑之細胞介素產生及細胞死亡。如本文所使用，先天免疫反應包括但不限於發炎細胞介素之產生(例如I型干擾素或IL-10產生)增加；NFκB路徑之活化；免疫細胞之增殖、成熟、分化及/或存活增加，以及在一些情況下細胞凋亡之誘導。先天免疫之活化可使用此項技術中已知之方法偵測，諸如量測(NF)-κB活化情況。The terms "innate immune response" and "innate immunity" are recognized in the art and refer to non-specific defense mechanisms activated by the body's immune system upon recognition of pathogen-associated molecular patterns, which involve different forms of cellular activity, including interleukin production and cell death via various pathways. As used herein, innate immune responses include, but are not limited to, increased production of inflammatory interleukins (e.g., type I interferons or IL-10 production); activation of the NFκB pathway; increased proliferation, maturation, differentiation and/or survival of immune cells, and in some cases, induction of apoptosis. Activation of innate immunity can be detected using methods known in the art, such as measuring (NF)-κB activation.
術語「適應性免疫反應」及「適應性免疫」係此項技術中公認的，且係指身體之免疫系統在識別出特定抗原時啟動之抗原特異性防禦機制，包括體液反應及細胞介導之反應。如本文所使用，適應性免疫反應包括由疫苗組合物，諸如本文所述之遺傳組合物觸發及/或增強之細胞反應。在一些實施例中，疫苗組合物包含作為抗原特異性適應性免疫反應之靶標之抗原。在其他實施例中，疫苗組合物在投與後允許在經免疫之個體中產生抗原，該抗原係抗原特異性適應性免疫反應之靶標。適應性免疫反應之活化可使用此項技術中已知之方法偵測，諸如量測抗原特異性抗體之產生或抗原特異性細胞介導之細胞毒性水準。The terms "adaptive immune response" and "adaptive immunity" are recognized in the art and refer to antigen-specific defense mechanisms activated by the body's immune system when a specific antigen is recognized, including humoral responses and cell-mediated responses. As used herein, an adaptive immune response includes a cellular response triggered and/or enhanced by a vaccine composition, such as a genetic composition described herein. In some embodiments, the vaccine composition includes an antigen that is the target of an antigen-specific adaptive immune response. In other embodiments, the vaccine composition, upon administration, allows the production of an antigen in an immunized individual that is the target of an antigen-specific adaptive immune response. Activation of an adaptive immune response can be detected using methods known in the art, such as measuring the production of antigen-specific antibodies or levels of antigen-specific cell-mediated cytotoxicity.
術語「抗體」意欲包括在免疫球蛋白類多肽範圍內之B細胞之多肽產物，其能夠與特定分子抗原結合並由兩對相同之多肽鏈構成，其中每一對具有一條重鏈(約50-70kDa)及一條輕鏈(約25kDa)，每條鏈之每個胺基末端部分包括含約100至約130個或更多個胺基酸之可變區，且每條鏈之每個羧基末端部分包括恆定區。參見例如Antibody Engineering(Borrebaeck編，第2版，1995)；及Kuby,Immunology(第3版，1997)。在特定實施例中，特定分子抗原可由本文所提供之抗體結合，包括多肽、其片段或抗原決定基。抗體亦包括但不限於合成抗體、重組產生之抗體、駱駝化抗體、內抗體、抗獨特型(抗Id)抗體及以上任一種之功能片段，功能片段係指抗體重鏈或輕鏈多肽之保留作為該片段來源之抗體之部分或全部結合活性之一部分。功能片段之非限制性實例包括單鏈Fv(scFv)(例如包括單特異性、雙特異性等)、Fab片段、F(ab’)片段、F(ab)2片段、F(ab’)2片段、二硫鍵連接之Fv(dsFv)、Fd片段、Fv片段、雙功能抗體、三功能抗體、四功能抗體及微型抗體。具體而言，本文所提供之抗體包括免疫球蛋白分子及免疫球蛋白分子之免疫活性部分，例如抗原結合結構域或含有抗原結合位點之分子(例如抗體之一或多個CDR)。此等抗體片段可見於例如Harlow及Lane，Antibodies：A Laboratory Manual(1989)；Mol.Biology and Biotechnology：A Comprehensive Desk Reference(Myers編，1995)；Huston等人，1993,Cell Biophysics 22：189-224；Plückthun及Skerra,1989,Meth.Enzymol.178：497-515；及Day,Advanced Immunochemistry(第2版，1990)。本文所提供之抗體可為免疫球蛋白分子之任何類別(例如IgG、IgE、IgM、IgD及IgA)或任何亞類(例如IgG1、IgG2、IgG3、IgG4、IgA1及IgA2)。The term "antibody" is intended to include polypeptide products of B cells within the scope of immunoglobulin polypeptides, which are capable of binding to specific molecular antigens and are composed of two pairs of identical polypeptide chains, wherein each pair has a heavy chain (about 50-70 kDa) and a light chain (about 25 kDa), each amino terminal portion of each chain includes a variable region containing about 100 to about 130 or more amino acids, and each carboxyl terminal portion of each chain includes a constant region. See, for example,Antibody Engineering (Borrebaeck, ed., 2nd edition, 1995); and Kuby,Immunology (3rd edition, 1997). In specific embodiments, specific molecular antigens can be bound by antibodies provided herein, including polypeptides, fragments thereof or antigenic determinants. Antibodies also include, but are not limited to, synthetic antibodies, recombinantly produced antibodies, camelized antibodies, endoantibodies, anti-idiotype (anti-Id) antibodies, and functional fragments of any of the above, where a functional fragment refers to a portion of an antibody heavy chain or light chain polypeptide that retains some or all of the binding activity of the antibody from which the fragment originated. Non-limiting examples of functional fragments include single-chain Fv (scFv) (e.g., including monospecific, bispecific, etc.), Fab fragments, F(ab') fragments, F(ab)2 fragments, F(ab')2 fragments, disulfide-linked Fv (dsFv), Fd fragments, Fv fragments, bifunctional antibodies, trifunctional antibodies, tetrafunctional antibodies, and miniantibodies. Specifically, antibodies provided herein include immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, such as antigen binding domains or molecules containing antigen binding sites (e.g., one or more CDRs of an antibody). Such antibody fragments can be found in, for example, Harlow and Lane,Antibodies: A Laboratory Manual (1989);Mol. Biology and Biotechnology: A Comprehensive Desk Reference (Myers, 1995); Huston et al., 1993, Cell Biophysics 22: 189-224; Plückthun and Skerra, 1989, Meth. Enzymol. 178: 497-515; and Day,Advanced Immunochemistry (2nd edition, 1990). The antibodies provided herein can be of any class (eg, IgG, IgE, IgM, IgD, and IgA) or any subclass (eg, IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2) of immunoglobulin molecules.
術語「投與(administer/administration)」係指將存在於體外之物質(例如本文所述之脂質奈米顆粒組合物)注射或以其他方式物理遞送至患者體內之操作，諸如經黏膜、真皮內、靜脈內、肌肉內遞送及/或本文所述或此項技術中已知之任何其他物理遞送方法遞送。當治療疾病、病症、疾患或其症狀時，典型地在該疾病、病症、疾患或其症狀發作之後進行物質之投與。當預防疾病、病症、疾患或其症狀時，典型地在該疾病、病症、疾患或其症狀發作之前進行物質之投與。The term "administer" or "administration" refers to the act of injecting or otherwise physically delivering a substance (e.g., a lipid nanoparticle composition described herein) present outside the body into a patient's body, such as by transmucosal, intradermal, intravenous, intramuscular delivery, and/or any other physical delivery method described herein or known in the art. When treating a disease, disorder, condition, or symptom thereof, the substance is typically administered after the onset of the disease, disorder, condition, or symptom thereof. When preventing a disease, disorder, condition, or symptom thereof, the substance is typically administered before the onset of the disease, disorder, condition, or symptom thereof.
「長期」投與與急性模式相對，係指以連續模式投與一或多種劑(例如持續一段時間，諸如數天、數週、數月或數年)，藉此在較長一段時間內維持初始治療效果(活性)。「間歇性」投與係指治療並非不間斷地連續進行，而係本質上週期性的。"Long-term" administration, as opposed to acute mode, refers to the administration of one or more agents in a continuous mode (e.g., over a period of time, such as days, weeks, months, or years) to maintain the initial therapeutic effect (activity) over a longer period of time. "Intermittent" administration means that the treatment is not continuous without interruption, but is cyclical in nature.
如本文所使用，術語「靶向遞送」或動詞形式「靶向」係指相較於遞送至任何其他器官、組織、細胞或細胞內隔室(稱為非目標位置)，促進所遞送之劑(諸如本文所述之脂質奈米顆粒組合物中之治療有效負載分子)到達特定器官、組織、細胞及/或細胞內隔室(稱為目標位置)之過程。靶向遞送可使用此項技術中已知之方法偵測，例如藉由在全身投與後將所遞送之劑在目標細胞群體中之濃度與所遞送之劑在非目標細胞群體處之濃度相比較來偵測。在某些實施例中，靶向遞送使得目標位置處之濃度為非目標位置處之濃度之至少2倍高。As used herein, the term "targeted delivery" or the verb form "targeting" refers to the process of promoting the delivered agent (such as the therapeutically effective cargo molecule in the lipid nanoparticle composition described herein) to reach a specific organ, tissue, cell and/or intracellular compartment (referred to as a target location) compared to delivery to any other organ, tissue, cell or intracellular compartment (referred to as a non-target location). Targeted delivery can be detected using methods known in the art, for example, by comparing the concentration of the delivered agent in the target cell population with the concentration of the delivered agent in the non-target cell population after systemic administration. In certain embodiments, targeted delivery results in a concentration at a target location that is at least twice as high as a concentration at a non-target location.
「有效量」一般係足以降低症狀之嚴重程度及/或頻率；消除症狀及/或潛在原因；防止症狀及/或其潛在原因之發生；及/或改善或補救由疾病、病症或疾患，包括例如由感染及贅瘤形成引起或與之相關之損害之量。在一些實施例中，有效量係治療有效量或預防有效量。An "effective amount" is generally an amount sufficient to reduce the severity and/or frequency of symptoms; eliminate symptoms and/or potential causes; prevent the occurrence of symptoms and/or their potential causes; and/or ameliorate or remedy damage caused by or associated with a disease, disorder, or condition, including, for example, infection and tumor formation. In some embodiments, an effective amount is a therapeutically effective amount or a prophylactically effective amount.
如本文所使用，術語「治療有效量」係指足以減少及/或改善給定疾病、病症或疾患，及/或其相關症狀(例如感染性疾病，諸如由病毒感染引起之感染性疾病，或贅生性疾病，諸如癌症)之嚴重程度及/或持續時間之劑(例如疫苗組合物)的量。本揭示案之物質/分子/劑(例如本文所述之脂質奈米顆粒組合物)之「治療有效量」可根據諸多因素而變化，諸如個體之疾病狀態、年齡、性別及體重，以及物質/分子/劑在個體體內引起所希望反應之能力。治療有效量包含物質/分子/劑之治療有益作用勝過其任何有毒或有害作用的量。在某些實施例中，術語「治療有效量」係指有效「治療」個體或哺乳動物之疾病、病症或疾患之如本文所述之脂質奈米顆粒組合物或其中所含治療劑或預防劑(例如治療性mRNA)的量。As used herein, the term "therapeutically effective amount" refers to an amount of an agent (e.g., a vaccine composition) sufficient to reduce and/or improve the severity and/or duration of a given disease, disorder, or condition, and/or its associated symptoms (e.g., infectious diseases, such as those caused by viral infection, or proliferative diseases, such as cancer). The "therapeutically effective amount" of the substance/molecule/agent of the present disclosure (e.g., the lipid nanoparticle composition described herein) may vary according to many factors, such as the disease state, age, sex, and weight of the individual, and the ability of the substance/molecule/agent to elicit a desired response in the individual's body. A therapeutically effective amount includes an amount in which the therapeutically beneficial effects of the substance/molecule/agent outweigh any toxic or detrimental effects thereof. In certain embodiments, the term "therapeutically effective amount" refers to an amount of a lipid nanoparticle composition as described herein or a therapeutic or preventive agent (e.g., therapeutic mRNA) contained therein that is effective to "treat" a disease, disorder, or condition in an individual or mammal.
「預防有效量」係當投與個體時將具有預期預防作用，例如預防疾病、病症、疾患或相關症狀(例如感染性疾病，諸如由病毒感染引起之感染性疾病，或贅生性疾病，諸如癌症)、延遲其發作(或復發)或降低其發作(或復發)可能性之醫藥組合物的量。典型地但非必須地，由於預防劑量係在疾病、病症或疾患之前或其早期階段用於個體，因此預防有效量可能小於治療有效量。完全之治療或預防作用未必藉由投與一次劑量而發生，而可能僅在投與一系列劑量後才發生。因此，治療或預防有效量可分一或多次投與來投與。A "prophylactically effective amount" is an amount of a pharmaceutical composition that, when administered to an individual, will have the intended prophylactic effect, such as preventing a disease, disorder, condition, or related symptoms (e.g., an infectious disease, such as one caused by a viral infection, or a proliferative disease, such as cancer), delaying its onset (or recurrence), or reducing the likelihood of its onset (or recurrence). Typically, but not necessarily, since a prophylactic dose is administered to an individual before a disease, disorder, or condition or at its early stages, the prophylactic effective dose may be less than the therapeutic effective dose. A complete therapeutic or prophylactic effect may not occur by administering a single dose, but may only occur after a series of doses. Therefore, a therapeutic or prophylactic effective dose may be administered in one or more administrations.
術語「預防(prevent/preventing/prevention)」係指降低疾病、病症、疾患或相關症狀(例如感染性疾病，諸如由病毒感染引起之感染性疾病，或贅生性疾病，諸如癌症)發作(或復發)之可能性。The term "prevent/preventing/prevention" refers to reducing the likelihood of the onset (or recurrence) of a disease, illness, condition or related symptoms (e.g. infectious diseases, such as those caused by viral infections, or proliferative diseases, such as cancer).
術語「管理(manage/managing/management)」係指個體自療法(例如預防劑或治療劑)獲得之有益作用，其不會引起疾病之治癒。在某些實施例中，向個體投與一或多種療法(例如預防劑或治療劑，諸如本文所述之脂質奈米顆粒組合物)以「管理」感染性或贅生性疾病、其一或多種症狀，藉此預防疾病之進展或惡化。The term "management" refers to the beneficial effects obtained by an individual from self-treatment (e.g., a prophylactic or therapeutic agent) that does not result in the cure of the disease. In certain embodiments, one or more therapies (e.g., a prophylactic or therapeutic agent, such as the lipid nanoparticle compositions described herein) are administered to an individual to "manage" an infectious or proliferative disease, one or more symptoms thereof, thereby preventing the progression or worsening of the disease.
術語「預防劑」係指可以完全或部分地抑制個體之疾病及/或其相關症狀之發展、復發、發作或擴散之任何劑。The term "prophylactic agent" refers to any agent that can completely or partially inhibit the development, recurrence, onset or spread of a disease and/or its related symptoms in an individual.
術語「治療劑」係指可用於治療、預防或減輕疾病、病症或疾患，包括用於治療、預防或減輕疾病、病症或疾患及/或其相關症狀之一或多種症狀之任何劑。The term "therapeutic agent" means any agent useful for treating, preventing or alleviating a disease, condition or disorder, including any agent useful for treating, preventing or alleviating one or more symptoms of a disease, condition or disorder and/or its associated symptoms.
術語「療法」係指可用於預防、管理、治療及/或改善疾病、病症或疾患之任何方案、方法及/或劑。在某些實施例中，術語「療法(therapies/therapy)」係指熟習此項技術者，諸如醫務人員已知可用於預防、管理、治療及/或改善疾病、病症或疾患之生物療法、支持療法及/或其他療法。The term "therapy" refers to any regimen, method and/or agent that can be used to prevent, manage, treat and/or ameliorate a disease, disorder or condition. In certain embodiments, the term "therapies" refers to biological therapies, supportive therapies and/or other therapies known to medical professionals skilled in the art for the prevention, management, treatment and/or amelioration of a disease, disorder or condition.
如本文所使用，「預防有效血清效價」係個體(例如人類)體內完全或部分地抑制個體之疾病、病症或疾患，及/或其相關症狀之發展、復發、發作或擴散之抗體之血清效價。As used herein, "preventive effective serum titer" refers to the serum titer of antibodies in an individual (e.g., human) that completely or partially inhibits the development, recurrence, onset, or spread of a disease, disorder, or condition, and/or its associated symptoms in the individual.
在某些實施例中，「治療有效血清效價」係個體(例如人類)體內降低個體之與疾病、病症或疾患相關之嚴重程度、持續時間及/或症狀之抗體之血清效價。In certain embodiments, a "therapeutically effective serum titer" is a serum titer of an antibody in a subject (e.g., a human) that reduces the severity, duration, and/or symptoms associated with a disease, disorder, or condition in the subject.
術語「血清效價」係指一名個體中來自多個樣品(例如在多個時間點)或在至少10名、至少20名、至少40名至多達約100名、1000名或更多個體之群體中之平均血清效價。The term "serum titer" refers to the average serum titer from multiple samples (e.g., at multiple time points) in one individual or in a population of at least 10, at least 20, at least 40 up to about 100, 1000, or more individuals.
術語「副作用」涵蓋療法(例如預防劑或治療劑)之不想要及/或不良作用。不想要之作用未必為不良的。療法(例如預防劑或治療劑)之不良作用可能為有害的、令人不適的或有風險的。副作用之實例包括腹瀉、咳嗽、腸胃炎、喘鳴、噁心、嘔吐、厭食、腹部絞痛、發燒、疼痛、體重減輕、去水、脫髮、呼吸困難、失眠、頭暈、黏膜炎、神經及肌肉影響、疲勞、口乾、食欲不振、投與部位出現皮疹或腫脹、諸如發燒、發冷及疲勞之類的類似流感之症狀、消化道問題及過敏反應。患者經歷之其他不期望之作用眾多且為此項技術中所知的。有許多作用闡述於Physician’s Desk Reference(第68版，2014)中。The term "side effect" encompasses unwanted and/or adverse effects of a therapy (e.g., a preventive or therapeutic agent). Unwanted effects are not necessarily adverse. Adverse effects of a therapy (e.g., a preventive or therapeutic agent) may be harmful, uncomfortable, or risky. Examples of side effects include diarrhea, cough, gastroenteritis, wheezing, nausea, vomiting, anorexia, abdominal cramps, fever, pain, weight loss, dehydration, hair loss, difficulty breathing, insomnia, dizziness, mucositis, nerve and muscle effects, fatigue, dry mouth, loss of appetite, rash or swelling at the administration site, flu-like symptoms such as fever, chills, and fatigue, digestive problems, and allergic reactions. Other undesirable effects experienced by patients are numerous and known in the art. Many of the effects are described inthe Physician's Desk Reference (68th Edition, 2014).
術語「個體」與「患者」可互換使用。如本文所使用，在某些實施例中，個體係哺乳動物，諸如非靈長類動物(例如牛、豬、馬、貓、狗、大鼠等)或靈長類動物(例如猴及人類)。在特定實施例中，個體係人類。在一個實施例中，個體係患有感染性疾病或贅生性疾病之哺乳動物(例如人類)。在另一實施例中，個體係有發展感染性疾病或贅生性疾病之風險之哺乳動物(例如人類)。The terms "subject" and "patient" are used interchangeably. As used herein, in certain embodiments, the subject is a mammal, such as a non-primate (e.g., cows, pigs, horses, cats, dogs, rats, etc.) or a primate (e.g., monkeys and humans). In specific embodiments, the subject is a human. In one embodiment, the subject is a mammal (e.g., a human) suffering from an infectious disease or a probiotic disease. In another embodiment, the subject is a mammal (e.g., a human) at risk of developing an infectious disease or a probiotic disease.
術語「可偵測探針」係指提供可偵測信號之組合物。該術語包括但不限於經由活性提供可偵測信號之任何螢光團、發色團、放射性標記、酶、抗體或抗體片段及諸如此類。The term "detectable probe" refers to a composition that provides a detectable signal. The term includes, but is not limited to, any fluorophore, chromophore, radiolabel, enzyme, antibody or antibody fragment, and the like that provides a detectable signal through activity.
術語「可偵測劑」係指可用於確定樣品或個體中所希望分子，諸如由本文所述之mRNA分子編碼之抗原之存在(existence/presence)的物質。可偵測劑可為能夠被目測之物質或者能夠以其他方式測定及/或量測(例如藉由定量)之物質。The term "detectable agent" refers to a substance that can be used to determine the presence (existence/presence) of a desired molecule, such as an antigen encoded by an mRNA molecule described herein, in a sample or individual. A detectable agent can be a substance that can be detected visually or can be determined and/or measured (e.g., by quantitation) in other ways.
「實質上全部」係指至少約60%、至少約65%、至少約70%、至少約75%、至少約80%、至少約85%、至少約90%、至少約95%、至少約98%、至少約99%或約100%。"Substantially all" means at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 98%, at least about 99%, or about 100%.
如本文所使用且除非另有說明，否則術語「約」或「大約」意指普通熟習此項技術者所測定之特定值的可接受之誤差，其部分取決於量測或測定該值之方式。在某些實施例中，術語「約」或「大約」意指在1、2、3或4個標準偏差以內。在某些實施例中，術語「約」及「大約」意指在給定值或範圍之20%以內、15%以內、10%以內、9%以內、8%以內、7%以內、6%以內、5%以內、4%以內、3%以內、2%以內、1%以內、0.5%以內、0.05%以內或更低。As used herein and unless otherwise indicated, the term "about" or "approximately" means an acceptable error for a particular value as determined by one of ordinary skill in the art, which depends in part on the manner in which the value is measured or determined. In certain embodiments, the term "about" or "approximately" means within 1, 2, 3, or 4 standard deviations. In certain embodiments, the term "about" and "approximately" means within 20%, within 15%, within 10%, within 9%, within 8%, within 7%, within 6%, within 5%, within 4%, within 3%, within 2%, within 1%, within 0.5%, within 0.05%, or less of a given value or range.
除非上下文另外明確指出，否則本文所使用之單數形式術語「一個(種)(a/an)」及「該」包括複數個(種)參考物。Unless the context clearly indicates otherwise, the singular terms "a/an" and "the" as used herein include plural references.
本說明書中引用之所有出版物、專利申請案、寄存編號及其他參考文獻皆以全文引用之方式併入本文中，其引用程度就如同特定且單獨地指示每一個別出版物或專利申請案以引用之方式併入一般。本文所論述之出版物僅僅提供在本申請案之申請日之前的揭示內容。本文中任何內容均不應解釋為承認本發明無權憑藉在先發明而早於此等出版物。此外，提供之公開日期可能與實際公開日期有所不同，實際公開日期可能需要獨立確認。All publications, patent applications, deposit numbers, and other references cited in this specification are incorporated herein by reference in their entirety to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference. The publications discussed herein are provided only as of the date of the filing of this application. Nothing herein should be construed as an admission that the present invention is not entitled to antedate such publications by virtue of prior invention. In addition, the publication dates provided may differ from the actual publication dates, which may need to be independently confirmed.
已經闡述本發明之多個實施例。然而，應理解，在不脫離本發明之精神及範圍之情況下，可進行各種修改。因此，實驗部分及實例中之闡述旨在說明而非限制申請專利範圍中所闡述之發明範圍。Several embodiments of the present invention have been described. However, it should be understood that various modifications may be made without departing from the spirit and scope of the present invention. Therefore, the descriptions in the experimental section and examples are intended to illustrate rather than limit the scope of the invention described in the scope of the patent application.
5.3脂質化合物5.3Lipid compounds
除非另有說明，否則本文提供之闡述適用於本文提供之所有式(例如式(I)，包括其子式)，只要它們適用。Unless otherwise stated, the descriptions provided herein apply to all formulae provided herein (e.g., formula (I), including its subformulae), as long as they are applicable.
在一個實施例中，本文提供一種式(I)之化合物：
在一個實施例中，本文提供一種式(I)之化合物：
在一個實施例中，n係1。在一個實施例中，n係2。在一個實施例中，m係1。在一個實施例中，m係2。在一個實施例中，n係1，且m係1。在一個實施例中，n係1，且m係2。在一個實施例中，n係2，且m係1。在一個實施例中，n係2，且m係2。In one embodiment, n is 1. In one embodiment, n is 2. In one embodiment, m is 1. In one embodiment, m is 2. In one embodiment, n is 1 and m is 1. In one embodiment, n is 1 and m is 2. In one embodiment, n is 2 and m is 1. In one embodiment, n is 2 and m is 2.
在一個實施例中，該化合物係式(II-A)之化合物：
在一個實施例中，該化合物係式(II-B)之化合物：
在一個實施例中，該化合物係式(II-C)之化合物：
在一個實施例中，該化合物係式(II-D)之化合物：
在一個實施例中，G3係C2-C12伸烷基。在一個實施例中，G3係C2-C8伸烷基。在一個實施例中，G3係C2-C6伸烷基。在一個實施例中，G3係C2-C4伸烷基。在一個實施例中，G3係C2伸烷基。在一個實施例中，G3係C3伸烷基。在一個實施例中，G3係C4伸烷基。在一個實施例中，G3係C5伸烷基。在一個實施例中，G3係C6伸烷基。在一個實施例中，G3係-CH2CH2-。In one embodiment, G3 is C2 -C12 alkylene. In one embodiment, G3 is C2 -C8 alkylene. In one embodiment, G3 is C2 -C6 alkylene. In one embodiment, G3 is C2 -C4 alkylene. In one embodiment, G3 is C2 alkylene. In one embodiment, G3 is C3 alkylene. In one embodiment, G3 is C4 alkylene. In one embodiment, G3 is C5 alkylene. In one embodiment, G3 is C6 alkylene. In one embodiment, G3 is -CH2 CH2 -.
在一個實施例中，G3係C2-C12伸烯基。在一個實施例中，G3係C2-C8伸烯基。在一個實施例中，G3係C2-C6伸烯基。在一個實施例中，G3係C2-C4伸烯基。在一個實施例中，G3係C2伸烯基。在一個實施例中，G3係C3伸烯基。在一個實施例中，G3係C4伸烯基。在一個實施例中，G3係C5伸烯基。在一個實施例中，G3係C6伸烯基。在一個實施例中，G3係(Z)-CH2-CH=CH-CH2-。在一個實施例中，G3係(E)-CH2-CH=CH-CH2-。In one embodiment,G3 isC2 -C12 alkenyl. In one embodiment,G3 isC2 -C8 alkenyl. In one embodiment,G3 isC2 -C6 alkenyl. In one embodiment,G3 isC2 -C4 alkenyl. In one embodiment,G3 is C2 alkenyl. In one embodiment,G3 isC3 alkenyl. In one embodiment,G3 isC4 alkenyl. In one embodiment,G3 isC5alkenyl . In one embodiment,G3 isC6 alkenyl. In one embodiment,G3 is (Z)-CH2 -CH=CH-CH2- . In one embodiment, G3 is (E)-CH2 -CH=CH-CH2 -.
在一個實施例中，G3係C2-C12伸烷基或C2-C12伸烯基，其中部分或全部伸烷基或伸烯基經C3-C8伸環烷基、C3-C8伸環烯基、C3-C8伸環炔基、4至8員伸雜環基、C6-C10伸芳基或5至10員伸雜芳基置換。在一個實施例中，G3係C2-C12伸烷基或C2-C12伸烯基，其中部分或全部伸烷基或伸烯基經C3-C8伸環烷基置換。在一個實施例中，G3係C2-C12伸烷基或C2-C12伸烯基，其中所有伸烷基或伸烯基均經C3-C8伸環烷基置換，即G3係C3-C8伸環烷基。在一個實施例中，G3係伸環丙基。在一個實施例中，G3係伸環丁基。在一個實施例中，G3係伸環戊基。在一個實施例中，G3係伸環己基。在一個實施例中，G3係伸環庚基。在一個實施例中，G3係伸環辛基。In one embodiment,G3 isC2 -C12 alkylene orC2 -C12 alkenylene, wherein part or all of the alkylene or alkenylene is replaced byC3 -C8 cycloalkylene,C3 -C8 cycloalkenylene,C3 -C8 cycloalkynylene,4-8 membered heterocycloalkylene,C6 -C10 arylene or 5-10 membered heteroarylene. In one embodiment,G3 isC2 -C12 alkylene orC2 -C12 alkenylene, wherein part or all of the alkylene or alkenylene is replaced byC3-C8cycloalkylene . In one embodiment,G3 isC2 -C12 alkylene orC2 -C12 alkenylene, wherein all alkylene or alkenylene are replaced byC3 -C8 cycloalkylene, i.e., G3 isC3 -C8 cycloalkylene. In one embodiment,G3 is cyclopropylene.In one embodiment, G3 is cyclobutylene. In one embodiment, G3 is cyclopentylene. In one embodiment, G3iscyclohexylene. In one embodiment,G3 is cycloheptylene. In one embodiment,G3 is cyclooctylene.
在一個實施例中，G3係
在一個實施例中，G3未經取代。In one embodiment,G3 is unsubstituted.
在一個實施例中，該化合物係式(III-A)之化合物：
在一個實施例中，該化合物係式(III-B)之化合物：
在一個實施例中，該化合物係式(III-C)之化合物：
在一個實施例中，該化合物係式(III-D)之化合物：
在一個實施例中，s係2至12之整數。在一個實施例中，s係2至8之整數。在一個實施例中，s係2至6之整數。在一個實施例中，s係2至4之整數。在一個實施例中，s係2。在一個實施例中，s係3。在一個實施例中，s係4。在一個實施例中，s係5。在一個實施例中，s係6。In one embodiment, s is an integer from 2 to 12. In one embodiment, s is an integer from 2 to 8. In one embodiment, s is an integer from 2 to 6. In one embodiment, s is an integer from 2 to 4. In one embodiment, s is 2. In one embodiment, s is 3. In one embodiment, s is 4. In one embodiment, s is 5. In one embodiment, s is 6.
在一個實施例中，G1係鍵。在一個實施例中，G1係C2-C12伸烷基。在一個實施例中，G1係C4-C8伸烷基。在一個實施例中，G1係C5-C7伸烷基。在一個實施例中，G1係C2伸烷基。在一個實施例中，G1係C3伸烷基。在一個實施例中，G1係C4伸烷基。在一個實施例中，G1係C5伸烷基。在一個實施例中，G1係C6伸烷基。在一個實施例中，G1係C7伸烷基。在一個實施例中，G1係C2-C12伸烯基。在一個實施例中，G1係C4-C8伸烯基。在一個實施例中，G1係C5-C7伸烯基。在一個實施例中，G1係C5伸烯基。在一個實施例中，G1係C7伸烯基。在一個實施例中，G1係直鏈的。在一個實施例中，G1係具支鏈的。在一個實施例中，G1係二價的。在一個實施例中，G1係三價的。In one embodiment,G1 is a bond. In one embodiment,G1 isC2 -C12 alkylene. In one embodiment,G1 isC4 -C8 alkylene. In one embodiment,G1 isC5 -C7 alkylene. In one embodiment,G1 isC2 alkylene. In one embodiment,G1 is C3 alkylene. In one embodiment,G1 isC4 alkylene. In one embodiment,G1 isC5 alkylene. In one embodiment,G1 isC6 alkylene. In one embodiment,G1 isC7 alkylene. In one embodiment,G1 isC2 -C12 alkenylene. In one embodiment,G1 isC4 -C8 alkenylene. In one embodiment,G1 isC5 -C7 alkenyl. In one embodiment,G1 isC5 alkenyl. In one embodiment, G1 isC7 alkenyl. In one embodiment,G1 is linear. In one embodiment,G1 is branched. In one embodiment,G1 isdivalent. In one embodiment, G1is trivalent.
在一個實施例中，G2係鍵。在一個實施例中，G2係C2-C12伸烷基。在一個實施例中，G2係C4-C8伸烷基。在一個實施例中，G2係C5-C7伸烷基。在一個實施例中，G2係C2伸烷基。在一個實施例中，G2係C3伸烷基。在一個實施例中，G2係C4伸烷基。在一個實施例中，G2係C5伸烷基。在一個實施例中，G2係C6伸烷基。在一個實施例中，G2係C7伸烷基。在一個實施例中，G2係C2-C12伸烯基。在一個實施例中，G2係C4-C8伸烯基。在一個實施例中，G2係C5-C7伸烯基。在一個實施例中，G2係C5伸烯基。在一個實施例中，G2係C7伸烯基。在一個實施例中，G2係直鏈的。在一個實施例中，G2係具支鏈的。在一個實施例中，G2係二價的。在一個實施例中，G2係三價的。In one embodiment,G is a bond. In one embodiment,G is C-C alkylene. In one embodiment, G is C-C alkylene. In one embodiment,G isC -C alkylene. In one embodiment,G is C-C alkylene. In one embodiment, G is C-C alkylene. In one embodiment,G is C-C alkylene. In one embodiment,G is C-C alkylene. In one embodiment, G is C-C alkylene. In one embodiment,G is C-C alkylene. In one embodiment,G is C-C alkylene. In one embodiment,G is C-C alkylene. In one embodiment, G is C-C alkylene. In one embodiment,G is C-C alkylene. In one embodiment,G is C-C alkylene. In one embodiment, G isC -C alkenylene. In one embodiment,G isC -C alkenylene. In one embodiment,G isC5 -C7 alkenyl. In one embodiment,G isC5 alkenyl. In one embodiment,G isC7 alkenyl. In one embodiment, G is linear. In one embodiment,G is branched. In one embodiment,G isdivalent . In one embodiment,G is trivalent.
在一個實施例中，G1及G2各自獨立地為C2-C12伸烷基。在一個實施例中，G1及G2各自獨立地為C5伸烷基。在一個實施例中，G1及G2各自獨立地為C7伸烷基。In one embodiment,G1 andG2 are each independentlyC2 -C12 alkylene. In one embodiment,G1 andG2 are each independently C5 alkylene. In one embodiment,G1 andG2 areeach independentlyC7 alkylene.
在一個實施例中，G1中之一或多個-CH2-經-O-置換。在一個實施例中，G1中之一或多個非末端-CH2-經-O-置換。在一個實施例中，G1中之一個非末端-CH2-經-O-置換。在一個實施例中，G1係(C2-C5伸烷基)-O-(C2-C6伸烷基)。In one embodiment, one or more -CH2 - in G1 is replaced by -O-. In one embodiment, one or more non-terminal -CH2 - in G1 is replaced by -O-. In one embodiment, one non-terminal -CH2 - in G1 is replaced by -O-. In one embodiment, G1 is (C2 -C5 alkylene)-O-(C2 -C6 alkylene).
在一個實施例中，G1係
在一個實施例中，G2中之一或多個-CH2-經-O-置換。在一個實施例中，G2中之一或多個非末端-CH2-經-O-置換。在一個實施例中，G2中之一個非末端-CH2-經-O-置換。在一個實施例中，G2係(C2-C5伸烷基)-O-(C2-C6伸烷基)。In one embodiment, one or more -CH2 - in G2 is replaced by -O-. In one embodiment, one or more non-terminal -CH2 - in G2 is replaced by -O-. In one embodiment, one non-terminal -CH2 - in G2 is replaced by -O-. In one embodiment, G2 is (C2 -C5 alkylene)-O-(C2 -C6 alkylene).
在一個實施例中，G2係
在一個實施例中，該化合物係式(IV)之化合物：
在一個實施例中，該化合物係式(IV-A)、(IV-B)、(IV-C)、(IV-D)、(IV-E)、(IV-F)、(IV-G)或(IV-H)之化合物：
在一個實施例中，該化合物係式(V)之化合物：
在一個實施例中，該化合物係式(V-A)、(V-B)、(V-C)、(V-D)、(V-E)、(V-F)、(V-G)或(V-H)之化合物：
在一個實施例中，y及z各自獨立地為2至10之整數。在一個實施例中，y及z各自獨立地為2至6之整數。在一個實施例中，y及z各自獨立地為4至10之整數。In one embodiment, y and z are each independently an integer from 2 to 10. In one embodiment, y and z are each independently an integer from 2 to 6. In one embodiment, y and z are each independently an integer from 4 to 10.
在一個實施例中，y及z不同。在一個實施例中，y及z相同。在一個實施例中，y及z相同，且選自4、5、6、7、8及9。在一個實施例中，y係5，且z係5。In one embodiment, y and z are different. In one embodiment, y and z are the same. In one embodiment, y and z are the same and are selected from 4, 5, 6, 7, 8 and 9. In one embodiment, y is 5 and z is 5.
在一個實施例中，s係2至12之整數。在一個實施例中，s係2至8之整數。在一個實施例中，s係2至6之整數。在一個實施例中，s係2至4之整數。在一個實施例中，s係2。在一個實施例中，s係3。在一個實施例中，s係4。在一個實施例中，s係5。在一個實施例中，s係6。In one embodiment, s is an integer from 2 to 12. In one embodiment, s is an integer from 2 to 8. In one embodiment, s is an integer from 2 to 6. In one embodiment, s is an integer from 2 to 4. In one embodiment, s is 2. In one embodiment, s is 3. In one embodiment, s is 4. In one embodiment, s is 5. In one embodiment, s is 6.
在一個實施例中，y係5，z係5，且s係2。In one embodiment, y is 5, z is 5, and s is 2.
在一個實施例中，L1係R1。In one embodiment, L1 is R1 .
在一個實施例中，L1係-OC(=O)R1、-C(=O)OR1、-OC(=O)OR1、-C(=O)R1、-OR1、-S(O)xR1、-S-SR1、-C(=O)SR1、-SC(=O)R1、-NRaC(=O)R1、-C(=O)NRbRc、-NRaC(=O)NRbRc、-OC(=O)NRbRc、-NRaC(=O)OR1、-SC(=S)R1、-C(=S)SR1、-C(=S)R1、-CH(OH)R1、-P(=O)(ORb)(ORc)、-NRaP(=O)(ORb)(ORc)或-(4至8員伸雜環基)-R1。在一個實施例中，L1係-OC(=O)R1、-C(=O)OR1、-C(=O)SR1、-SC(=O)R1、-NRaC(=O)R1或-C(=O)NRbRc。在一個實施例中，L1係-OC(=O)R1、-C(=O)OR1、-NRaC(=O)R1或-C(=O)NRbRc。在一個實施例中，L1係-OC(=O)R1。在一個實施例中，L1係-C(=O)OR1。在一個實施例中，L1係-NRaC(=O)R1。在一個實施例中，L1係-C(=O)NRbRc。在一個實施例中，L1係-OR1。在一個實施例中，L1係-NRaP(=O)(ORb)(ORc)。In one embodiment, L1 is -OC(=O)R1 , -C(=O)OR1 , -OC(=O)OR1 , -C(=O)R1 , -OR1 , -S(O)xR1 , -S-SR1 , -C(=O)SR1 , -SC(=O)R1 , -NRa C(=O)R1 , -C(=O)NRb Rc , -NRa C(=O)NRb Rc , -OC(=O)NRb Rc , -NRa C(=O)OR1 , -SC(=S)R1 , -C(=S)SR1 , -C(=S)R1 , -CH(OH)R1 , -P(=O)(ORb )(ORc ), -NRa P(=O)(ORb )(ORc ) or -(4- to 8-membered heterocyclic group)-R1. In one embodiment, L1 is -OC(＝O)R1 , -C(＝O)OR1 , -C(＝O)SR1 , -SC(＝O)R1 , -NRa C(＝O)R1 , or -C(＝O)NRb Rc . In one embodiment, L1 is -OC(＝O)R1 , -C(＝O)OR1 , -NRa C(＝O)R1 , or -C(＝O)NRb Rc . In one embodiment, L1 is -OC(＝O)R1. In one embodiment, L1 is -C(＝O)OR1. In one embodiment, L1 is -NRa C(＝O)R1 . In one embodiment, L1 is -C(=O)NRb Rc . In one embodiment, L1 is -OR1 . In one embodiment, L1 is -NRa P(=O)(ORb )(ORc ).
在一個實施例中，L1係-(4至8員伸雜環基)-R1。在一個實施例中，L1係
在一個實施例中，L2係R2。In one embodiment, L2 is R2 .
在一個實施例中，L2係-OC(=O)R2、-C(=O)OR2、-OC(=O)OR2、-C(=O)R2、-OR2、-S(O)xR2、-S-SR2、-C(=O)SR2、-SC(=O)R2、-NRdC(=O)R2、-C(=O)NReRf、-NRdC(=O)NReRf、-OC(=O)NReRf、-NRdC(=O)OR2、-SC(=S)R2、-C(=S)SR2、-C(=S)R2、-CH(OH)R2、-P(=O)(ORe)(ORf)、或-NRdP(=O)(ORe)(ORf)、或-(4至8員伸雜環基)-R2。在一個實施例中，L2係-OC(=O)R2、-C(=O)OR2、-C(=O)SR2、-SC(=O)R2、-NRdC(=O)R2或-C(=O)NReRf。在一個實施例中，L2係-OC(=O)R2、-C(=O)OR2、-NRdC(=O)R2或-C(=O)NReRf。在一個實施例中，L2係-OC(=O)R2。在一個實施例中，L2係-C(=O)OR2。在一個實施例中，L2係-NRdC(=O)R2。在一個實施例中，L2係-C(=O)NReRf。在一個實施例中，L2係-OR2。在一個實施例中，L2係-NRdP(=O)(ORe)(ORf)。在一個實施例中，L2係-(4至8員伸雜環基)-R2。在一個實施例中，L2係
在一個實施例中，L1係-C(=O)OR1或-C(=O)NRbRc；且L2係-C(=O)OR2或-C(=O)NReRf。在一個實施例中，L1係-C(=O)OR1且L2係-C(=O)OR2。在一個實施例中，L1係-C(=O)OR1且L2係-C(=O)NReRf。在一個實施例中，L1係-C(=O)NRbRc且L2係-C(=O)OR2。在一個實施例中，L1係-C(=O)NRbRc且L2係-C(=O)NReRf。In one embodiment,L1 is -C(=O)OR1 or -C(=O)NRbRc; andL2 is -C(=O)OR2 or -C(=O)NReRf . In one embodiment,L1 is -C(=O)OR1andL2 is -C(=O)OR2 . In one embodiment,L1 is -C(=O)OR1 andL2 is -C(=O)NReRf. In one embodiment,L1 is -C(=O)NRbRcandL2 is -C(=O)OR2 . In one embodiment,L1 is-C (=O)NRbRc andL2 is -C(=O )NReRf .
在一個實施例中，L1係-OC(=O)R1或-NRaC(=O)R1；且L2係-OC(=O)R2或-NRdC(=O)R2。在一個實施例中，L1係-OC(=O)R1且L2係-OC(=O)R2。在一個實施例中，L1係-OC(=O)R1且L2係-NRdC(=O)R2。在一個實施例中，L1係-NRaC(=O)R1且L2係-OC(=O)R2。在一個實施例中，L1係-NRaC(=O)R1且L2係-NRdC(=O)R2。In one embodiment,L1 is -OC(=O)R1 or -NRa C(=O)R1 ; andL2 is -OC(=O)R2 or -NRd C(=O)R2 . In one embodiment,L1 is -OC(=O)R1 andL2 is -OC(=O)R2 . In one embodiment,L1 is -OC(=O)R1 andL2 is -NRd C(=O)R2 . In one embodiment,L1 is -NRa C(=O)R1 andL2 is -OC(=O)R2 . In one embodiment,L1 is -NRa C(=O)R1 andL2 is -NRd C(=O)R2 .
在一個實施例中，L1係-OR1且L2係-C(=O)OR2。在一個實施例中，L1係-OR1且L2係-C(=O)NReRf。在一個實施例中，L1係-C(=O)OR1且L2係-OR2。在一個實施例中，L1係-C(=O)NRbRc且L2係-OR2。In one embodiment,L1 is-OR1 andL2 is -C(=O)OR2 . In one embodiment,L1 is -OR1 andL2 is -C(=O)NReRf. In one embodiment,L1 is -C(=O)OR1 andL2 is-OR2 . In one embodiment,L1 is-C (=O)NRbRcandL2 is-OR2 .
在一個實施例中，該化合物係式(VI)之化合物：
在一個實施例中，z係2至10之整數。在一個實施例中，z係2至6之整數。在一個實施例中，z係4至10之整數。在一個實施例中，z選自4、5、6、7、8及9。在一個實施例中，z係5。In one embodiment, z is an integer from 2 to 10. In one embodiment, z is an integer from 2 to 6. In one embodiment, z is an integer from 4 to 10. In one embodiment, z is selected from 4, 5, 6, 7, 8 and 9. In one embodiment, z is 5.
在一個實施例中，R3係C1-C12烷基。在一個實施例中，R3係C1-C8烷基。在一個實施例中，R3係C1-C6烷基。在一個實施例中，R3係C1-C4烷基。在一個實施例中，烷基係直鏈烷基。在一個實施例中，烷基係具支鏈烷基。在一個實施例中，R3係甲基。在一個實施例中，R3係乙基。在一個實施例中，R3係正丙基。在一個實施例中，R3係異丙基。在一個實施例中，R3係正丁基。在一個實施例中，R3係正戊基。在一個實施例中，R3係正己基。在一個實施例中，R3係正辛基。在一個實施例中，R3係正壬基。In one embodiment, R3 is C1 -C12 alkyl. In one embodiment, R3 is C1 -C8 alkyl. In one embodiment, R3 is C1 -C6 alkyl. In one embodiment, R3 is C1 -C4 alkyl. In one embodiment, the alkyl group is a straight chain alkyl group. In one embodiment, the alkyl group is a branched chain alkyl group. In one embodiment, R3 is a methyl group. In one embodiment, R3 is an ethyl group. In one embodiment, R3 is a n-propyl group. In one embodiment, R3 is an isopropyl group. In one embodiment, R3 is a n-butyl group. In one embodiment, R3 is a n-pentyl group. In one embodiment, R3 is a n-hexyl group. In one embodiment, R3 is a n-octyl group. In one embodiment, R3 is n-nonyl.
在一個實施例中，R3係C2-C12烯基。在一個實施例中，R3係C2-C8烯基。在一個實施例中，R3係C2-C4烯基。在一個實施例中，烯基係直鏈烯基。在一個實施例中，烯基係具支鏈烯基。在一個實施例中，R3係乙烯基。在一個實施例中，R3係烯丙基。In one embodiment, R3 is C2 -C12 alkenyl. In one embodiment, R3 is C2 -C8 alkenyl. In one embodiment, R3 is C2 -C4 alkenyl. In one embodiment, the alkenyl is a straight chain alkenyl. In one embodiment, the alkenyl is a branched chain alkenyl. In one embodiment, R3 is vinyl. In one embodiment, R3 is allyl.
在一個實施例中，R3係C2-C12炔基。在一個實施例中，R3係C2-C8炔基。在一個實施例中，R3係C2-C4炔基。在一個實施例中，炔基係直鏈炔基。在一個實施例中，炔基係具支鏈炔基。In one embodiment, R3 is C2 -C12 alkynyl. In one embodiment, R3 is C2 -C8 alkynyl. In one embodiment, R3 is C2 -C4 alkynyl. In one embodiment, alkynyl is a straight chain alkynyl. In one embodiment, alkynyl is a branched chain alkynyl.
在一個實施例中，R3係C3-C8環烷基。在一個實施例中，R3係環丙基。在一個實施例中，R3係環丁基。在一個實施例中，R3係環戊基。在一個實施例中，R3係環己基。在一個實施例中，R3係環庚基。在一個實施例中，R3係環辛基。In one embodiment, R3 is C3 -C8 cycloalkyl. In one embodiment, R3 is cyclopropyl. In one embodiment, R3 is cyclobutyl. In one embodiment, R3 is cyclopentyl. In one embodiment, R3 is cyclohexyl. In one embodiment, R3 is cycloheptyl. In one embodiment, R3 is cyclooctyl.
在一個實施例中，R3係C3-C8環烯基。在一個實施例中，R3係環丙烯基。在一個實施例中，R3係環丁烯基。在一個實施例中，R3係環戊烯基。在一個實施例中，R3係環己烯基。在一個實施例中，R3係環庚烯基。在一個實施例中，R3係環辛烯基。In one embodiment, R3 is C3 -C8 cycloalkenyl. In one embodiment, R3 is cyclopropenyl. In one embodiment, R 3 is cyclobutenyl. In one embodiment, R3 is cyclopentenyl. In one embodiment, R3 iscyclohexenyl . In one embodiment, R3 is cycloheptenyl. In one embodiment, R3 is cyclooctenyl.
在一個實施例中，R3係4至8員雜環基。在一個實施例中，R3係4至8員雜環烷基。在一個實施例中，R3係氧雜環丁烷基。在一個實施例中，R3係四氫呋喃基。在一個實施例中，R3係四氫哌喃基。在一個實施例中，R3係四氫硫代哌喃基。In one embodiment, R3 is a 4- to 8-membered heterocyclic group. In one embodiment, R3 is a 4- to 8-membered heterocyclic alkyl group. In one embodiment, R3 is an oxacyclobutane group. In one embodiment, R3 is a tetrahydrofuranyl group. In one embodiment, R3 is a tetrahydropyranyl group. In one embodiment, R3 is a tetrahydrothiopyranyl group.
在一個實施例中，R3係C6-C10芳基。在一個實施例中，R3係苯基。In one embodiment, R3 is C6 -C10 aryl. In one embodiment, R3 is phenyl.
在一個實施例中，R3係5至10員雜芳基。在一個實施例中，R3係5員雜芳基。在一個實施例中，R3係6員雜芳基。In one embodiment, R3 is a 5- to 10-membered heteroaryl. In one embodiment, R3 is a 5-membered heteroaryl. In one embodiment, R3 is a 6-membered heteroaryl.
在一個實施例中，R3、G1或G1之一部分連同其所連接之氮一起形成環狀部分。In one embodiment, R3 , G1 or a portion of G1 together with the nitrogen to which it is attached form a cyclic moiety.
在一個實施例中，該化合物係式(VII)之化合物：
在一個實施例中，R3、G3或G3之一部分連同其所連接之氮一起形成環狀部分。In one embodiment, R3 , G3 or a portion of G3 together with the nitrogen to which it is attached form a cyclic moiety.
在一個實施例中，該化合物係式(VIII-A)、(VIII-B)、(VIII-C)、(VIII-D)、(VIII-E)、(VIII-F)或(VIII-G)之化合物：
在一個實施例中，u係1。在一個實施例中，u係2。在一個實施例中，u係3。在一個實施例中，v係1。在一個實施例中，v係2。在一個實施例中，v係3。在一個實施例中，u係1，且v係1。在一個實施例中，u係2，且v係2。在一個實施例中，u係3，且v係3。In one embodiment, u is 1. In one embodiment, u is 2. In one embodiment, u is 3. In one embodiment, v is 1. In one embodiment, v is 2. In one embodiment, v is 3. In oneembodiment, u is 1 and v is 1. In one embodiment, u is 2 and v is 2. In one embodiment, u is 3 and v is 3.
在一個實施例中，該化合物係式(IX-A)、(IX-B)、(IX-C)、(IX-D)、(IX-E)、(IX-F)、(IX-G)、(IX-H)、(IX-I)、(IX-J)、(IX-K)、(IX-L)、(IX-M)、(IX-N)、(IX-O)、(IX-P)、(IX-Q)、(IX-R)、(IX-S)、(IX-T)、(IX-U)、(IX-V)、(IX-W)、(IX-X)、(IX-Y)、(IX-Z)或(IX-AA)之化合物：
在一個實施例中，y0係1至7之整數。在一個實施例中，y0係1。在一個實施例中，y0係2。在一個實施例中，y0係3。在一個實施例中，y0係4。在一個實施例中，y0係5。在一個實施例中，y0係6。在一個實施例中，y0係7。在一個實施例中，z0係1至7之整數。在一個實施例中，z0係1。在一個實施例中，z0係2。在一個實施例中，z0係3。在一個實施例中，z0係4。在一個實施例中，z0係5。在一個實施例中，z0係6。在一個實施例中，z0係7。In one embodiment, y0 is an integer from 1 to 7. In one embodiment, y0 is 1. In one embodiment, y0 is 2. In one embodiment, y0 is 3. In one embodiment, y0 is 4. In one embodiment, y0 is 5. In one embodiment, y0 is 6. In one embodiment, y0 is 7. In one embodiment, z0 is an integer from 1 to 7. In one embodiment, z0 is 1. In one embodiment, z0 is 2. In one embodiment, z0 is 3. In one embodiment, z0 is 4. In one embodiment, z0 is 5. In one embodiment, z0 is 6. In one embodiment, z0 is 7.
在一個實施例中，y1係2至6之整數。在一個實施例中，y1係2。在一個實施例中，y1係3。在一個實施例中，y1係4。在一個實施例中，y1係5。在一個實施例中，y1係6。在一個實施例中，z1係2至6之整數。在一個實施例中，z1係2。在一個實施例中，z1係3。在一個實施例中，z1係4。在一個實施例中，z1係5。在一個實施例中，z1係6。In one embodiment, y1 is an integer from 2 to 6. In one embodiment, y1 is 2. In one embodiment, y1 is 3. In one embodiment, y1 is 4. In one embodiment, y1 is 5. In one embodiment, y1 is 6. In one embodiment, z1 is an integer from 2 to 6. In one embodiment, z1 is 2. In one embodiment, z1 is 3. In one embodiment, z1 is 4. In one embodiment, z1 is 5. In one embodiment, z1 is 6.
在一個實施例中，y2係2。在一個實施例中，y2係3。在一個實施例中，y2係4。在一個實施例中，y2係5。在一個實施例中，z2係2。在一個實施例中，z2係3。在一個實施例中，z2係4。在一個實施例中，z2係5。In one embodiment, y2 is 2. In one embodiment, y2 is 3. In one embodiment, y2 is 4. In one embodiment, y2 is 5. In one embodiment, z2 is 2. In one embodiment, z2 is 3. In one embodiment, z2 is 4. In one embodiment, z2 is 5.
在一個實施例中，y3係2。在一個實施例中，y3係3。在一個實施例中，y3係4。在一個實施例中，y3係5。在一個實施例中，y3係6。在一個實施例中，z3係2。在一個實施例中，z3係3。在一個實施例中，z3係4。在一個實施例中，z3係5。在一個實施例中，z3係6。In one embodiment, y3 is 2. In one embodiment, y3 is 3. In one embodiment, y3 is 4. In one embodiment, y3 is 5. In one embodiment, y3 is 6. In one embodiment, z3 is 2. In one embodiment, z3 is 3. In one embodiment, z3 is 4. In one embodiment, z3 is 5. In one embodiment, z3 is 6.
在一個實施例中，y4係0。在一個實施例中，y4係1。在一個實施例中，y4係2。在一個實施例中，y4係3。在一個實施例中，z4係0。在一個實施例中，z4係1。在一個實施例中，z4係2。在一個實施例中，z4係3。In one embodiment, y4 is 0. In one embodiment, y4 is 1. In one embodiment, y4 is 2. In one embodiment, y4 is 3. In one embodiment, z4 is 0. In one embodiment, z4 is 1. In one embodiment, z4 is 2. In one embodiment, z4 is 3.
在一個實施例中，y5係1。在一個實施例中，y5係2。在一個實施例中，y5係3。在一個實施例中，y5係4。在一個實施例中，y5係5。在一個實施例中，z5係1。在一個實施例中，z5係2。在一個實施例中，z5係3。在一個實施例中，z5係4。在一個實施例中，z5係5。In one embodiment, y5 is 1. In one embodiment, y5 is 2. In one embodiment, y5 is 3. In one embodiment, y5 is 4. In one embodiment, y5 is 5. In one embodiment, z5 is 1. In one embodiment, z5 is 2. In one embodiment, z5 is 3. In one embodiment, z5 is 4. In one embodiment, z5 is 5.
在一個實施例中，y2係2，且y3係2。在一個實施例中，y2係2，且y4係1。在一個實施例中，z2係2，且z3係2。在一個實施例中，z2係2，且z4係1。In one embodiment, y2 is 2 and y3 is 2. In one embodiment, y2 is 2 and y4 is 1. In one embodiment, z2 is 2 and z3 is 2. In one embodiment, z2 is 2 and z4 is 1.
在一個實施例中，s、y、z、L1及L2如別處所定義。在一個實施例中，L1係-OR1、-OC(=O)R1、-C(=O)OR1或-C(=O)NRbRc；且L2係-OR2、-OC(=O)R2、-C(=O)OR2或-C(=O)NReRf。在一個實施例中，當存在兩個L1時，每個L1獨立地為-OC(=O)R1。在一個實施例中，當存在兩個L2時，每個L2獨立地為-OC(=O)R2。在一個實施例中，當僅存在一個L1時，L1係-C(=O)OR1。在一個實施例中，當僅存在一個L1時，L1係-C(=O)NRbRc。在一個實施例中，當僅存在一個L2時，L2係-C(=O)OR2。在一個實施例中，當僅存在一個L2時，L2係-C(=O)NReRf。In one embodiment, s, y, z,L1 andL2 are as defined elsewhere. In one embodiment,L1 is-OR1 , -OC(=O)R1 , -C(=O)OR1 or-C (=O)NRbRc ; andL2 is-OR2 , -OC(=O)R2 , -C(=O)OR2 or -C(=O)NReRf. In one embodiment, when there are twoL1 , eachL1 is independently -OC(=O)R1 . In one embodiment, when there are twoL2 , eachL2 is independently -OC(=O)R2 . In one embodiment, when there is only oneL1 ,L1 is -C(=O)OR1 . In one embodiment, whenonly one L1 is present,L1 is -C(=O)NRb Rc . In one embodiment, when only oneL2 is present,L2 is -C(=O)OR2 . In one embodiment, when only oneL2 is present,L2 is -C(=O)NRe Rf .
在式(IX-A)至(IX-AA)中任一者之一個具體實施例中，當僅存在一個L1時，L1係-C(=O)OR1。在另一實施例中，L1係-C(=O)NRbRc。在一個實施例中，R1或Rc係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C4-C8烷基。In one specific embodiment of any one of formulas (IX-A) to (IX-AA), when only one L1 is present, L1 is -C(=O)OR1 . In another embodiment, L1 is -C(=O)NRb Rc . In one embodiment, R1 or Rc is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C1 alkylene, and R8 and R9 are independently C4 -C8 alkyl.
在式(IX-A)至(IX-AA)中任一者之一個具體實施例中，當僅存在一個L2時，L2係-C(=O)OR2。在另一實施例中，L2係-C(=O)NReRf。在一個實施例中，R2或Rf係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C4-C8烷基。In one specific embodiment of any one of formulas (IX-A) to (IX-AA), when only one L2 is present, L2 is -C(=O)OR2 . In another embodiment, L2 is -C(=O)NRe Rf . In one embodiment, R2 or Rf is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C1 alkylene, and R8 and R9 are independently C4 -C8 alkyl.
在式(IX-A)至(IX-AA)中任一者之一個具體實施例中，當存在
在式(IX-A)至(IX-AA)中任一者之一個具體實施例中，當存在
在式(IX-A)至(IX-AA)中任一者之一個具體實施例中，當存在
在式(IX-A)至(IX-AA)中任一者之一個具體實施例中，當存在
在一個實施例中，R3未經取代。In one embodiment, R3 is unsubstituted.
在一個實施例中，R3經一或多個選自由以下組成之群之取代基取代：C1-C6烷基、鹵基、C1-C6鹵代烷基、硝基、側氧基、-ORg、-NRgC(=O)Rh、-C(=O))NRgRh、-C(=O)Rh、-OC(=O)Rh、-C(=O)ORh及-O-Ri-OH，其中：Rg在每次出現時獨立地為H或C1-C6烷基；Rh在每次出現時獨立地為C1-C6烷基；且Ri在每次出現時獨立地為C1-C6伸烷基。In one embodiment,R3 is substituted with one or more substituents selected from the group consisting of C1-C6 alkyl, halogen,C1 -C6 haloalkyl, nitro, oxo,-ORg , -NRgC(=O)Rh ,-C (=O))NRgRh , -C(=O)Rh , -OC(=O)Rh , -C(=O )ORh , and -ORi-OH , wherein:Rg is independently H orC1 -C6 alkyl at each occurrence;Rh is independentlyC1 -C6 alkyl at each occurrence; andRi is independentlyC1 -C6 alkylene at each occurrence.
在一個實施例中，R3經一或多個C1-C6烷基(例如，甲基)取代。在一個實施例中，R3經一或多個鹵基(例如，-F)取代。在一個實施例中，R3經一或多個C1-C6鹵代烷基(例如，-CF3)取代。在一個實施例中，R3經一或多個羥基取代。在一個實施例中，R3經一個羥基取代。In one embodiment, R3 is substituted with one or more C1 -C6 alkyl (e.g., methyl). In one embodiment, R3 is substituted with one or more halogen (e.g., -F). In one embodiment, R3 is substituted with one or more C1 -C6 halogenated alkyl (e.g., -CF3 ). In one embodiment, R3 is substituted with one or more hydroxyl. In one embodiment, R3 is substituted with one hydroxyl.
在一個實施例中，R3經一或多個C3-C8環烷基、C6-C10芳基或5至10員雜芳基取代，其中每一者視情況經取代。在一個實施例中，R3係經一或多個C3-C8環烷基、C6-C10芳基或5至10員雜芳基取代之C1-C6烷基(例如，甲基)，其中每一者視情況經取代。在一個實施例中，C3-C8環烷基、C6-C10芳基或5至10員雜芳基未經取代。在一個實施例中，C3-C8環烷基、C6-C10芳基或5至10員雜芳基經一或多個C1-C6烷基、鹵基、C1-C6鹵代烷基、硝基、羥基或氰基取代。In one embodiment, R is substitutedwith one or more C3-C8cycloalkyl ,C6 -C10 aryl or 5-10 membered heteroaryl, each of which is optionally substituted. In one embodiment,R is C1-C6 alkyl (e.g., methyl) substituted with one or moreC3 -C8 cycloalkyl,C6 -C10 aryl or 5-10 membered heteroaryl, each of which is optionally substituted. In one embodiment,C3 -C8 cycloalkyl,C6 -C10 aryl or5-10 membered heteroaryl is unsubstituted. In one embodiment, the C3 -C8 cycloalkyl, C6 -C10 aryl or 5- to 10-membered heteroaryl is substituted with one or more C1 -C6 alkyl, halogen, C1 -C6 halogenated alkyl, nitro, hydroxyl or cyano groups.
在一個實施例中，R4係C1-C12烷基。在一個實施例中，R4係C1-C8烷基。在一個實施例中，R4係C1-C6烷基。在一個實施例中，R4係C1-C4烷基。在一個實施例中，R4係甲基。在一個實施例中，R4係乙基。在一個實施例中，R4係正丙基。在一個實施例中，R4係異丙基。在一個實施例中，R4係正丁基。在一個實施例中，R4係正戊基。在一個實施例中，R4係正己基。在一個實施例中，R4係正辛基。在一個實施例中，R4係正壬基。In one embodiment, R4 is C1 -C12 alkyl. In one embodiment, R4 is C1 -C8 alkyl. In one embodiment, R4 is C1 -C6 alkyl. In one embodiment, R4 is C1 -C4 alkyl. In one embodiment, R4 is methyl. In one embodiment, R4 is ethyl. In one embodiment, R4 is n-propyl. In one embodiment, R4 is isopropyl. In one embodiment, R4 is n-butyl. In one embodiment, R4 is n-pentyl. In one embodiment, R4 is n-hexyl. In one embodiment, R4 is n-octyl. In one embodiment, R4 is n-nonyl.
在一個實施例中，R4係C3-C8環烷基。在一個實施例中，R4係環丙基。在一個實施例中，R4係環丁基。在一個實施例中，R4係環戊基。在一個實施例中，R4係環己基。在一個實施例中，R4係環庚基。在一個實施例中，R4係環辛基。In one embodiment, R4 is C3 -C8 cycloalkyl. In one embodiment, R4 is cyclopropyl. In one embodiment, R4 is cyclobutyl. In one embodiment, R4 is cyclopentyl. In one embodiment, R4 is cyclohexyl. In one embodiment, R4 is cycloheptyl. In one embodiment, R4 is cyclooctyl.
在一個實施例中，R4未經取代。In one embodiment, R4 is unsubstituted.
在一個實施例中，R4經一或多個選自由以下組成之群之取代基取代：側氧基、-ORg、-NRgC(=O)Rh、-C(=O)NRgRh、-C(=O)Rh、-OC(=O)Rh、-C(=O)ORh、-O-Ri-OH及-N(R10)R11，其中：Rg在每次出現時獨立地為H或C1-C6烷基；Rh在每次出現時獨立地為C1-C6烷基；Ri在每次出現時獨立地為C1-C6伸烷基；R10係氫或C1-C6烷基；R11係C1-C6烷基、C3-C8環烷基或C3-C8環烯基；或R10及R11連同其所連接之氮一起形成環狀部分；且R11或環狀部分視情況經羥基、側氧基、-NH2、-NH(C1-C6烷基)或-N(C1-C6烷基)2中之一或多個取代。In one embodiment, R4 is substituted with one or more substituents selected from the group consisting of: oxo, -ORg , -NRg C(=O)Rh , -C(=O)NRg Rh , -C(=O)Rh , -OC(=O)Rh , -C(=O)ORh , -ORi -OH, and -N(R10 )R11 , wherein: Rg is independently H or C1 -C6 alkyl at each occurrence; Rh is independently C1 -C6 alkyl at each occurrence; Ri is independently C1 -C6 alkylene at each occurrence; R10 is hydrogen or C1 -C6 alkyl; R11 is C1 -C6 alkyl, C3 -C8 cycloalkyl or C3 -C8 cycloalkenyl; or R10 and R R11 together with the nitrogen to which it is attached forms a cyclic moiety; and R11 or the cyclic moiety is optionally substituted by one or more of a hydroxyl group, a pendoxy group, -NH2 , -NH(C1 -C6 alkyl) or -N(C1 -C6 alkyl)2 .
在一個實施例中，R4經一或多個羥基取代。在一個實施例中，R4經一個羥基取代。In one embodiment,R4 is substituted with one or more hydroxyl groups. In one embodiment,R4 is substituted with one hydroxyl group.
在一個實施例中，R4係經取代之C1-C12烷基。在一個實施例中，R4係-(CH2)pQ、-(CH2)pCHQR、-CHQR或-CQ(R)2，其中Q係C3-C8環烷基、C3-C8環烯基、C3-C8環炔基、4至8員雜環基、C6-C10芳基、5至10員雜芳基、-OR、-O(CH2)pN(R)2、-C(O)OR、-OC(O)R、-CX3、-CX2H、-CXH2、-CN、-N(R)2、-C(O)N(R)2、-N(R)C(O)R、-N(R)S(O)2R、-N(R)C(O)N(R)2、-N(R)C(S)N(R)2、-N(R)R22、-O(CH2)pOR、-N(R)C(=NR23)N(R)2、-N(R)C(=CHR23)N(R)2、-OC(O)N(R)2、-N(R)C(O)OR、-N(OR)C(O)R、-N(OR)S(O)2R、-N(OR)C(O)OR、-N(OR)C(O)N(R)2、-N(OR)C(S)N(R)2、-N(OR)C(=NR23)N(R)2、-N(OR)C(=CHR23)N(R)2、-C(=NR23)N(R)2、-C(=NR23)R、-C(O)N(R)OR或-C(R)N(R)2C(O)OR，且每個p獨立地為1、2、3、4或5；R22係C3-C8環烷基、C3-C8環烯基、C3-C8環炔基、4至8員雜環基、C6-C10芳基或5至10員雜芳基；R23係H、-CN、-NO2、C1-C6烷基、-OR、-S(O)2R、-S(O)2N(R)2、C2-C6烯基、C3-C8環烷基、C3-C8環烯基、C3-C8環炔基、4至8員雜環基、C6-C10芳基或5至10員雜芳基；每個R獨立地為H、C1-C3烷基或C2-C3烯基；或N(R)2部分中之兩個R連同其所連接之氮一起形成環狀部分；且每個X獨立地為F、CI、Br或I。In one embodiment, R4 is substituted C1 -C12 alkyl. In one embodiment, R4 is -(CH2 )p Q, -(CH2 )p CHQR, -CHQR or -CQ(R)2 , wherein Q is C3 -C8 cycloalkyl, C3 -C8 cycloalkenyl, C3 -C8 cycloalkynyl, 4-8 membered heterocyclic group, C6 -C10 aryl, 5-10 membered heteroaryl, -OR, -O(CH2 )p N(R)2 , -C(O)OR, -OC(O)R, -CX3 , -CX2 H, -CXH2 , -CN, -N(R)2 , -C(O)N(R)2 , -N(R)C(O)R, -N(R)S(O)2 R, -N(R)C(O)N(R)2 , -N(R)C(S)N(R)2 , -N(R)R22 , -O(CH2 )p OR, -N(R)C(=NR23 )N(R)2 , -N(R)C(=CHR23 )N(R)2 , -OC(O)N(R)2 , -N(R)C(O)OR, -N(OR)C(O)R, -N(OR)S(O)2 R, -N(OR)C(O)OR, -N(OR)C(O)N(R)2 , -N(OR)C(S)N(R)2 , -N(OR)C(=NR23 )N(R)2 , -N(OR)C(=CHR23 )N(R)2 , -C(=NR23 )N(R)2 , -C(=NR23 )R, -C(O)N(R)OR or -C(R)N(R)2 C(O)OR, and each p is independently 1, 2, 3, 4 or 5; R22 is C3 -C8 cycloalkyl, C3 -C8 cycloalkenyl, C3 -C8 cycloalkynyl, 4 to 8 membered heterocyclic group, C6 -C10 aryl or 5 to 10 membered heteroaryl; R23 is H, -CN, -NO2 , C1 -C6 alkyl, -OR, -S(O)2 R, -S(O)2 N(R)2 , C2 -C6 alkenyl, C3 -C8 cycloalkyl, C3 -C8 cycloalkenyl, C3 -C 88- membered cycloalkynyl, 4-8-membered heterocyclic group, C6 -C10 aryl or 5-10-membered heteroaryl; each R is independently H, C1 -C3 alkyl or C2 -C3 alkenyl; or the two Rs in the N(R)2 portion together with the nitrogen to which they are attached form a cyclic portion; and each X is independently F, CI, Br or I.
在一個實施例中，R4係-CH2CH2OH。在一個實施例中，R4係-CH2CH2CH2OH。在一個實施例中，R4係-CH2CH2CH2CH2OH。在一個實施例中，R4係-CH2CH2OCH2CH2OH。In one embodiment,R4is-CH2CH2OH . Inone embodiment,R4 is-CH2CH2CH2OH .In one embodiment,R4 is-CH2CH2CH2CH2OH .In one embodiment,R4is-CH2CH2OCH2CH2OH.
在一個實施例中，R4經一或多個-N(R10)R11取代。在一個實施例中，R4經一個-N(R10)R11取代。In one embodiment, R4 is substituted with one or more -N(R10 )R11. In one embodiment, R4 is substituted with one -N(R10 )R11 .
在一個實施例中，R10係氫。In one embodiment, R10 is hydrogen.
在一個實施例中，R11係C3-C8環烯基。在一個實施例中，R11係環丁烯基。在一個實施例中，R11經側氧基、-NH2、-NH(C1-C6烷基)或-N(C1-C6烷基)2中之一或多個取代。In one embodiment, R11 is C3 -C8 cycloalkenyl. In one embodiment, R11 is cyclobutenyl. In one embodiment, R11 is substituted by one or more of a pendoxy group, -NH2 , -NH(C1 -C6 alkyl) or -N(C1 -C6 alkyl)2 .
在一個實施例中，R10及R11連同其所連接之氮一起形成環狀部分。在一個實施例中，環狀部分係5至10員雜芳基。在一個實施例中，環狀部分係嘧啶-1-基。在一個實施例中，環狀部分係嘌呤-9-基。在一個實施例中，環狀部分經側氧基、-NH2、-NH(C1-C6烷基)或-N(C1-C6烷基)2中之一或多個取代。In one embodiment, R10 and R11 together with the nitrogen to which they are attached form a cyclic moiety. In one embodiment, the cyclic moiety is a 5-10 membered heteroaryl. In one embodiment, the cyclic moiety is a pyrimidin-1-yl. In one embodiment, the cyclic moiety is a purin-9-yl. In one embodiment, the cyclic moiety is substituted with one or more of a pendoxy group, -NH2 , -NH(C1 -C6 alkyl) or -N(C1 -C6 alkyl)2 .
在一個實施例中，R4經
在一個實施例中，R1係直鏈C6-C24烷基。在一個實施例中，R1係直鏈C7-C15烷基。在一個實施例中，R1係直鏈C7烷基。在一個實施例中，R1係直鏈C8烷基。在一個實施例中，R1係直鏈C9烷基。在一個實施例中，R1係直鏈C10烷基。在一個實施例中，R1係直鏈C11烷基。在一個實施例中，R1係直鏈C12烷基。在一個實施例中，R1係直鏈C13烷基。在一個實施例中，R1係直鏈C14烷基。在一個實施例中，R1係直鏈C15烷基。In one embodiment, R1 is a straight chain C6 -C24 alkyl. In one embodiment, R1 is a straight chain C7 -C15 alkyl. In one embodiment, R1 is a straight chain C7 alkyl. In one embodiment, R1 is a straight chain C8 alkyl. In one embodiment, R1 is a straight chain C9 alkyl. In one embodiment, R1 is a straight chain C10 alkyl. In one embodiment, R1 is a straight chain C11 alkyl. In one embodiment, R1 is a straight chain C12 alkyl. In one embodiment, R1 is a straight chain C13 alkyl. In one embodiment, R1 is a straight chain C14 alkyl. In one embodiment, R1 is a linear C15 alkyl group.
在一個實施例中，R1係直鏈C6-C24烯基。在一個實施例中，R1係直鏈C7-C17烯基。在一個實施例中，R1係直鏈C7烯基。在一個實施例中，R1係直鏈C8烯基。在一個實施例中，R1係直鏈C9烯基。在一個實施例中，R1係直鏈C10烯基。在一個實施例中，R1係直鏈C11烯基。在一個實施例中，R1係直鏈C12烯基。在一個實施例中，R1係直鏈C13烯基。在一個實施例中，R1係直鏈C14烯基。在一個實施例中，R1係直鏈C15烯基。在一個實施例中，R1係直鏈C16烯基。在一個實施例中，R1係直鏈C17烯基。In one embodiment, R1 is a straight chain C6 -C24 alkenyl. In one embodiment, R1 is a straight chain C7 -C17 alkenyl. In one embodiment, R1 is a straight chain C7 alkenyl. In one embodiment, R1 is a straight chain C8 alkenyl. In one embodiment, R1 is a straight chain C9 alkenyl. In one embodiment, R1 is a straight chain C10 alkenyl. In one embodiment, R1 is a straight chain C11 alkenyl. In one embodiment, R1 is a straight chain C12 alkenyl. In one embodiment, R1 is a straight chain C13 alkenyl. In one embodiment, R1 is a straight chain C14 alkenyl. In one embodiment, R1 is a straight chain C15 alkenyl. In one embodiment, R1 is a straight chain C16 alkenyl. In one embodiment, R1 is a straight chain C17 alkenyl.
在一個實施例中，R1係具支鏈C6-C24烷基。在一個實施例中，R1係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烷基。在一個實施例中，R1係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C4-C8烷基。In one embodiment, R1 is a branched C6 -C24 alkyl group. In one embodiment, R1 is -R7 -CH(R8 )(R9 ), wherein R7 is a C0 -C5 alkylene group, and R8 and R9 are independently C2 -C10 alkyl groups. In one embodiment, R1 is -R7 -CH(R8 )(R9 ), wherein R7 is a C0 -C1 alkylene group, and R8 and R9 are independently C4 -C8 alkyl groups.
在一個實施例中，R1係具支鏈C6-C24烯基。在一個實施例中，R1係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烯基。在一個實施例中，R1係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C6-C10烯基。In one embodiment, R1 is a branched C6 -C24 alkenyl. In one embodiment, R1 is -R7 -CH(R8 )(R9 ), wherein R7 is a C0 -C5 alkylene group, and R8 and R9 are independently C2 -C10 alkenyl. In one embodiment, R1 is -R7 -CH(R8 )(R9 ), wherein R7 is a C0 -C1 alkylene group, and R8 and R9 are independently C6 -C10 alkenyl.
在一個實施例中，R2係直鏈C6-C24烷基。在一個實施例中，R2係直鏈C7-C15烷基。在一個實施例中，R2係直鏈C7烷基。在一個實施例中，R2係直鏈C8烷基。在一個實施例中，R2係直鏈C9烷基。在一個實施例中，R2係直鏈C10烷基。在一個實施例中，R2係直鏈C11烷基。在一個實施例中，R2係直鏈C12烷基。在一個實施例中，R2係直鏈C13烷基。在一個實施例中，R2係直鏈C14烷基。在一個實施例中，R2係直鏈C15烷基。In one embodiment, R2 is a straight chain C6 -C24 alkyl. In one embodiment, R2 is a straight chain C7 -C15 alkyl. In one embodiment, R2 is a straight chain C7 alkyl. In one embodiment, R2 is a straight chain C8 alkyl. In one embodiment, R2 is a straight chain C9 alkyl. In one embodiment, R2 is a straight chain C10 alkyl. In one embodiment, R2 is a straight chain C11 alkyl. In one embodiment, R2 is a straight chain C12 alkyl. In one embodiment, R2 is a straight chain C13 alkyl. In one embodiment, R2 is a straight chain C14 alkyl. In one embodiment, R2 is a linear C15 alkyl group.
在一個實施例中，R2係直鏈C6-C24烯基。在一個實施例中，R2係直鏈C7-C17烯基。在一個實施例中，R2係直鏈C7烯基。在一個實施例中，R2係直鏈C8烯基。在一個實施例中，R2係直鏈C9烯基。在一個實施例中，R2係直鏈C10烯基。在一個實施例中，R2係直鏈C11烯基。在一個實施例中，R2係直鏈C12烯基。在一個實施例中，R2係直鏈C13烯基。在一個實施例中，R2係直鏈C14烯基。在一個實施例中，R2係直鏈C15烯基。在一個實施例中，R2係直鏈C16烯基。在一個實施例中，R2係直鏈C17烯基。In one embodiment, R2 is a straight chain C6 -C24 alkenyl. In one embodiment, R2 is a straight chain C7 -C17 alkenyl. In one embodiment, R2 is a straight chain C7 alkenyl. In one embodiment, R2 is a straight chain C8 alkenyl. In one embodiment, R2 is a straight chain C9 alkenyl. In one embodiment, R2 is a straight chain C10 alkenyl. In one embodiment, R2 is a straight chain C11 alkenyl. In one embodiment, R2 is a straight chain C12 alkenyl. In one embodiment, R2 is a straight chain C13 alkenyl. In one embodiment, R2 is a straight chain C14 alkenyl. In one embodiment, R2 is a straight chain C15 alkenyl. In one embodiment, R2 is a straight chain C16 alkenyl. In one embodiment, R2 is a straight chain C17 alkenyl.
在一個實施例中，R2係具支鏈C6-C24烷基。在一個實施例中，R2係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烷基。在一個實施例中，R2係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C4-C8烷基。In one embodiment, R2 is a branched C6 -C24 alkyl group. In one embodiment, R2 is -R7 -CH(R8 )(R9 ), wherein R7 is a C0 -C5 alkylene group, and R8 and R9 are independently C2 -C10 alkyl groups. In one embodiment, R2 is -R7 -CH(R8 )(R9 ), wherein R7 is a C0 -C1 alkylene group, and R8 and R9 are independently C4 -C8 alkyl groups.
在一個實施例中，R2係具支鏈C6-C24烯基。在一個實施例中，R2係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烯基。在一個實施例中，R2係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C6-C10烯基。In one embodiment, R2 is a branched C6 -C24 alkenyl. In one embodiment, R2 is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C5 alkylene, and R8 and R9 are independently C2 -C10 alkenyl. In one embodiment, R2 is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C1 alkylene, and R8 and R9 are independently C6 -C10 alkenyl.
在一個實施例中，Rc係直鏈C6-C24烷基。在一個實施例中，Rc係直鏈C7-C15烷基。在一個實施例中，Rc係直鏈C7烷基。在一個實施例中，Rc係直鏈C8烷基。在一個實施例中，Rc係直鏈C9烷基。在一個實施例中，Rc係直鏈C10烷基。在一個實施例中，Rc係直鏈C11烷基。在一個實施例中，Rc係直鏈C12烷基。在一個實施例中，Rc係直鏈C13烷基。在一個實施例中，Rc係直鏈C14烷基。在一個實施例中，Rc係直鏈C15烷基。In one embodiment, Rc is a straight chain C6 -C24 alkyl group. In one embodiment, Rc is a straight chain C7 -C15 alkyl group. In one embodiment, Rc is a straight chain C7 alkyl group. In one embodiment, Rc is a straight chain C8 alkyl group. In one embodiment, Rc is a straight chain C9 alkyl group. In one embodiment, Rc is a straight chain C10 alkyl group. In one embodiment, Rc is a straight chain C11 alkyl group. In one embodiment, Rc is a straight chain C12 alkyl group. In one embodiment, Rc is a straight chain C13 alkyl group. In one embodiment, Rc is a straight chain C14 alkyl group. In one embodiment, Rc is a straight chain C15 alkyl group.
在一個實施例中，Rc係直鏈C6-C24烯基。在一個實施例中，Rc係直鏈C7-C17烯基。在一個實施例中，Rc係直鏈C7烯基。在一個實施例中，Rc係直鏈C8烯基。在一個實施例中，Rc係直鏈C9烯基。在一個實施例中，Rc係直鏈C10烯基。在一個實施例中，Rc係直鏈C11烯基。在一個實施例中，Rc係直鏈C12烯基。在一個實施例中，Rc係直鏈C13烯基。在一個實施例中，Rc係直鏈C14烯基。在一個實施例中，Rc係直鏈C15烯基。在一個實施例中，Rc係直鏈C16烯基。在一個實施例中，Rc係直鏈C17烯基。In one embodiment, Rc is a straight chain C6 -C24 alkenyl. In one embodiment, Rc is a straight chain C7 -C17 alkenyl. In one embodiment, Rc is a straight chain C7 alkenyl. In one embodiment, Rc is a straight chain C8 alkenyl. In one embodiment, Rc is a straight chain C9 alkenyl. In one embodiment, Rc is a straight chain C10 alkenyl. In one embodiment, Rc is a straight chain C11 alkenyl. In one embodiment, Rc is a straight chain C12 alkenyl. In one embodiment, Rc is a straight chain C13 alkenyl. In one embodiment, Rc is a straight chain C14 alkenyl. In one embodiment, Rc is a straight chain C15 alkenyl. In one embodiment, Rc is a straight chain C16 alkenyl. In one embodiment, Rc is a straight chain C17 alkenyl.
在一個實施例中，Rc係具支鏈C6-C24烷基。在一個實施例中，Rc係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烷基。在一個實施例中，Rc係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C4-C8烷基。In one embodiment, Rc is branched C6 -C24 alkyl. In one embodiment, Rc is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C5 alkylene, and R8 and R9 are independently C2 -C10 alkyl. In one embodiment, Rc is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C1 alkylene, and R8 and R9 are independently C4 -C8 alkyl.
在一個實施例中，Rc係具支鏈C6-C24烯基。在一個實施例中，Rc係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烯基。在一個實施例中，Rc係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C6-C10烯基。In one embodiment, Rc is a branched C6 -C24 alkenyl. In one embodiment, Rc is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C5 alkylene, and R8 and R9 are independently C2 -C10 alkenyl. In one embodiment, Rc is -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C1 alkylene, and R8 and R9 are independently C6 -C10 alkenyl.
在一個實施例中，Rf係直鏈C6-C24烷基。在一個實施例中，Rf係直鏈C7-C15烷基。在一個實施例中，Rf係直鏈C7烷基。在一個實施例中，Rf係直鏈C8烷基。在一個實施例中，Rf係直鏈C9烷基。在一個實施例中，Rf係直鏈C10烷基。在一個實施例中，Rf係直鏈C11烷基。在一個實施例中，Rf係直鏈C12烷基。在一個實施例中，Rf係直鏈C13烷基。在一個實施例中，Rf係直鏈C14烷基。在一個實施例中，Rf係直鏈C15烷基。In one embodiment, Rf is a straight chain C6 -C24 alkyl group. In one embodiment, Rf is a straight chain C7 -C15 alkyl group. In one embodiment, Rf is a straight chain C7 alkyl group. In one embodiment, Rf is a straight chain C8 alkyl group. In one embodiment, Rf is a straight chain C9 alkyl group. In one embodiment, Rf is a straight chain C10 alkyl group. In one embodiment, Rf is a straight chain C11 alkyl group. In one embodiment, Rf is a straight chain C12 alkyl group. In one embodiment, Rf is a straight chain C13 alkyl group. In one embodiment, Rf is a straight chain C14 alkyl group. In one embodiment, Rf is a straight chain C15 alkyl group.
在一個實施例中，Rf係直鏈C6-C24烯基。在一個實施例中，Rf係直鏈C7-C17烯基。在一個實施例中，Rf係直鏈C7烯基。在一個實施例中，Rf係直鏈C8烯基。在一個實施例中，Rf係直鏈C9烯基。在一個實施例中，Rf係直鏈C10烯基。在一個實施例中，Rf係直鏈C11烯基。在一個實施例中，Rf係直鏈C12烯基。在一個實施例中，Rf係直鏈C13烯基。在一個實施例中，Rf係直鏈C14烯基。在一個實施例中，Rf係直鏈C15烯基。在一個實施例中，Rf係直鏈C16烯基。在一個實施例中，Rf係直鏈C17烯基。In one embodiment, Rf is a straight chain C6 -C24 alkenyl. In one embodiment, Rf is a straight chain C7 -C17 alkenyl. In one embodiment, Rf is a straight chain C7 alkenyl. In one embodiment, Rf is a straight chain C8 alkenyl. In one embodiment, Rf is a straight chain C9 alkenyl. In one embodiment, Rf is a straight chain C10 alkenyl. In one embodiment, Rf is a straight chain C11 alkenyl. In one embodiment, Rf is a straight chain C12 alkenyl. In one embodiment, Rf is a straight chain C13 alkenyl. In one embodiment, Rf is a straight chain C14 alkenyl. In one embodiment,Rf is a straight chainC15 alkenyl. In one embodiment,Rf is a straight chainC16 alkenyl. In one embodiment,Rf is a straight chainC17 alkenyl.
在一個實施例中，Rf係具支鏈C6-C24烷基。在一個實施例中，Rf係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烷基。在一個實施例中，Rf係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C4-C8烷基。In one embodiment,Rf is a branchedC6 -C24 alkyl group. In one embodiment,Rf is-R7 -CH(R8 )(R9 ), whereinR7 is aC0 -C5 alkylene group, andR8 andR9 are independentlyC2 -C10 alkyl groups. In one embodiment,Rf is-R7 -CH(R8 )(R9 ), whereinR7 is aC0 -C1 alkylene group, andR8 andR9 are independentlyC4 -C8 alkyl groups.
在一個實施例中，Rf係具支鏈C6-C24烯基。在一個實施例中，Rf係-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烯基。在一個實施例中，Rf係-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C6-C10烯基。In one embodiment,Rf is a branchedC6 -C24 alkenyl. In one embodiment,Rf is-R7 -CH(R8 )(R9 ), whereinR7 isC0 -C5 alkylene, andR8 andR9 are independentlyC2 -C10 alkenyl. In one embodiment,Rf is-R7 -CH(R8 )(R9 ), whereinR7 isC0 -C1 alkylene, andR8 andR9 are independentlyC6 -C10 alkenyl.
在一個實施例中，R1、R2、Rc及Rf各自獨立地為直鏈C6-C18烷基、直鏈C6-C18烯基或-R7-CH(R8)(R9)，其中R7係C0-C5伸烷基，且R8及R9獨立地為C2-C10烷基或C2-C10烯基。In one embodiment, R1 , R2 , Rc and Rf are each independently a linear C6 -C18 alkyl, a linear C6 -C18 alkenyl or -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C5 alkylene, and R8 and R9 are independently C2 -C10 alkyl or C2 -C10 alkenyl.
在一個實施例中，R1、R2、Rc及Rf各自獨立地為直鏈C7-C15烷基、直鏈C7-C15烯基或-R7-CH(R8)(R9)，其中R7係C0-C1伸烷基，且R8及R9獨立地為C4-C8烷基或C6-C10烯基。In one embodiment, R1 , R2 , Rc and Rf are each independently a linear C7 -C15 alkyl, a linear C7 -C15 alkenyl or -R7 -CH(R8 )(R9 ), wherein R7 is C0 -C1 alkylene, and R8 and R9 are independently C4 -C8 alkyl or C6 -C10 alkenyl.
在一個實施例中，R1、R2、Rc及Rf各自獨立地為以下結構之一：
在一個實施例中，Ra係H。在一個實施例中，Rd係H。在一個實施例中，Ra、Rb、Rd及Re各自獨立地為H。在一個實施例中，Rb係C1-C24烷基。在一個實施例中，Rb係C1-C12烷基。在一個實施例中，Rb係C2-C24烯基。在一個實施例中，Rb係C2-C12烯基。在一個實施例中，Re係C1-C24烷基。在一個實施例中，Re係C1-C12烷基。在一個實施例中，Re係C2-C24烯基。在一個實施例中，Re係C2-C12烯基。In one embodiment,Ra is H. In one embodiment,Rd is H. In one embodiment,Ra ,Rb ,Rd andRe are each independently H. In one embodiment,Rb is C1 -C24 alkyl. In one embodiment,Rb is C1 -C12 alkyl. In one embodiment,Rb is C2 -C 24 alkenyl. In one embodiment,Rb is C2 -C12 alkenyl. In one embodiment,Re is C1 -C24 alkyl. In one embodiment,Re is C1 -C12alkyl . In one embodiment,Re is C2 -C24 alkenyl. In one embodiment,Re is C2 -C12 alkenyl.
在一個實施例中，該化合物係表1中之化合物，或其醫藥學上可接受之鹽、前藥或立體異構物。In one embodiment, the compound is a compound in Table 1, or a pharmaceutically acceptable salt, prodrug or stereoisomer thereof.
在一個實施例中，該化合物係表1A中之化合物，或其醫藥學上可接受之鹽、前藥或立體異構物。In one embodiment, the compound is a compound in Table 1A, or a pharmaceutically acceptable salt, prodrug or stereoisomer thereof.
在一個實施例中，本文提供一種式(X)之化合物：
在一個實施例中，Z係-OH。在一個實施例中，Z係鹵素。在一個實施例中，Z係-Cl。In one embodiment, Z is -OH. In one embodiment, Z is a halogen. In one embodiment, Z is -Cl.
在一個實施例中，式(X)之化合物係用於製備式(I)之化合物之過程中之中間體，例如，如本文提供之實例中所舉例說明的。In one embodiment, the compound of formula (X) is an intermediate in the process of preparing a compound of formula (I), for example, as exemplified in the examples provided herein.
應理解，如上所示之本文所提供之化合物之任何實施例，以及如上所示之本文所提供之化合物之任何特定取代基及/或變量可獨立地與化合物之其他實施例及/或取代基及/或變量組合以形成以上未具體闡述之實施例。另外，在列出任何特定基團或變量之取代基及/或變量之清單之情況下，應理解，可自特定實施例及/或請求項中刪除每一個別取代基及/或變量，且其餘之取代基及/或變量清單將被認為在本文所提供之實施例之範圍內。It is understood that any embodiment of the compounds provided herein as shown above, and any specific substituents and/or variables of the compounds provided herein as shown above, can be independently combined with other embodiments and/or substituents and/or variables of the compounds to form embodiments not specifically described above. In addition, where a list of substituents and/or variables for any specific group or variable is listed, it is understood that each individual substituent and/or variable can be deleted from the specific embodiment and/or claim, and the remaining list of substituents and/or variables will be considered within the scope of the embodiments provided herein.
應理解，在本說明書中，所描繪各式之取代基及/或變量之組合僅在此等貢獻產生穩定化合物時才為容許的。It is understood that in this specification, combinations of substituents and/or variables in the various formulae depicted are permissible only if such contributions result in stable compounds.
5.4奈米顆粒組合物5.4Nanoparticle compositions
在一個態樣中，本文闡述包含本文所述之脂質化合物之奈米顆粒組合物。在特定實施例中，該奈米顆粒組合物包含如本文所述之根據式(I)(及其子式)之化合物。In one aspect, the present invention discloses a nanoparticle composition comprising a lipid compound described herein. In a specific embodiment, the nanoparticle composition comprises a compound according to formula (I) (and its subformulae) as described herein.
在一些實施例中，當例如藉由動態光散射(DLS)、透射電子顯微鏡檢查、掃描電子顯微鏡檢查或另一種方法量測時，本文提供之奈米顆粒組合物之最大尺寸係1μm或更短(例如
可結合本揭示案使用之奈米顆粒組合物包括例如脂質奈米顆粒(LNP)、奈米脂蛋白顆粒、脂質體、脂質囊泡及脂質複合物(lipoplex)。在一些實施例中，奈米顆粒組合物係包括一或多個脂質雙層之囊泡。在一些實施例中，奈米顆粒組合物包括經水性隔室隔開之兩個或更多個同心雙層。脂質雙層可經官能化及/或彼此交聯。脂質雙層可包括一或多種配位體、蛋白質或通道。Nanoparticle compositions that can be used in conjunction with the present disclosure include, for example, lipid nanoparticles (LNPs), nanolipoprotein nanoparticles, liposomes, lipid vesicles, and lipoplexes. In some embodiments, the nanoparticle composition is a vesicle comprising one or more lipid bilayers. In some embodiments, the nanoparticle composition comprises two or more concentric bilayers separated by an aqueous compartment. The lipid bilayers can be functionalized and/or cross-linked to each other. The lipid bilayer can include one or more ligands, proteins, or channels.
奈米顆粒組合物之特徵可取決於其組分。舉例而言，包含膽固醇作為結構脂質之奈米顆粒組合物可具有與包含不同結構脂質之奈米顆粒組合物不同之特徵。類似地，奈米顆粒組合物之特徵可取決於其組分之絕對或相對量。舉例而言，包含較高莫耳分率之磷脂之奈米顆粒組合物可具有與包含較低莫耳分率之磷脂之奈米顆粒組合物不同之特徵。特徵亦可取決於奈米顆粒組合物之製備方法及條件而變化。The characteristics of a nanoparticle composition may depend on its components. For example, a nanoparticle composition comprising cholesterol as a structural lipid may have different characteristics than a nanoparticle composition comprising a different structural lipid. Similarly, the characteristics of a nanoparticle composition may depend on the absolute or relative amounts of its components. For example, a nanoparticle composition comprising a higher molar fraction of phospholipids may have different characteristics than a nanoparticle composition comprising a lower molar fraction of phospholipids. Characteristics may also vary depending on the method and conditions of preparation of the nanoparticle composition.
奈米顆粒組合物可藉由多種方法表徵。舉例而言，可使用顯微鏡檢查(例如透射電子顯微鏡檢查或掃描電子顯微鏡檢查)來檢查奈米顆粒組合物之形態及大小分佈。可使用動態光散射或電位測定法(例如電位滴定法)來量測ζ電位。動態光散射亦可用於確定粒度。亦可使用儀器，諸如Zetasizer Nano ZS(Malvem Instruments Ltd,Malvem,Worcestershire,UK)來量測奈米顆粒組合物之多個特徵，諸如粒度、多分散指數及ζ電位。Nanoparticle compositions can be characterized by a variety of methods. For example, microscopic examination (e.g., transmission electron microscopy or scanning electron microscopy) can be used to examine the morphology and size distribution of nanoparticle compositions. Zeta potential can be measured using dynamic light scattering or potentiometry (e.g., potentiometric titration). Dynamic light scattering can also be used to determine particle size. Instruments such as the Zetasizer Nano ZS (Malvem Instruments Ltd, Malvem, Worcestershire, UK) can also be used to measure various characteristics of nanoparticle compositions, such as particle size, polydispersity index, and zeta potential.
Dh(大小)：奈米顆粒組合物之平均大小可在數十奈米至數百奈米之間。舉例而言，平均大小可為約40nm至約150nm，諸如約40nm、45nm、50nm、55nm、60nm、65nm、70nm、75nm、80nm、85nm、90nm、95nm、100nm、105nm、110nm、115nm、120nm、125nm、130nm、135nm、140nm、145nm或150nm。在一些實施例中，奈米顆粒組合物之平均大小可為約50nm至約100nm、約50nm至約90nm、約50nm至約80nm、約50nm至約70nm、約50nm至約60nm、約60nm至約100nm、約60nm至約90nm、約60nm至約80nm、約60nm至約70nm、約70nm至約100nm、約70nm至約90nm、約70nm至約80nm、約80nm至約100nm、約80nm至約90nm或約90nm至約100nm。在某些實施例中，奈米顆粒組合物之平均大小可為約70nm至約100nm。在一些實施例中，平均大小可為約80nm。在其他實施例中，平均大小可為約100nm。Dh (size): The average size of the nanoparticle composition can be between tens of nanometers and hundreds of nanometers. For example, the average size can be about 40 nm to about 150 nm, such as about 40 nm, 45 nm, 50 nm, 55 nm, 60 nm, 65 nm, 70 nm, 75 nm, 80 nm, 85 nm, 90 nm, 95 nm, 100 nm, 105 nm, 110 nm, 115 nm, 120 nm, 125 nm, 130 nm, 135 nm, 140 nm, 145 nm, or 150 nm. In some embodiments, the nanoparticle composition may have an average size of about 50nm to about 100nm, about 50nm to about 90nm, about 50nm to about 80nm, about 50nm to about 70nm, about 50nm to about 60nm, about 60nm to about 100nm, about 60nm to about 90nm, about 60nm to about 80nm, about 60nm to about 70nm, about 70nm to about 100nm, about 70nm to about 90nm, about 70nm to about 80nm, about 80nm to about 100nm, about 80nm to about 90nm, or about 90nm to about 100nm. In certain embodiments, the nanoparticle composition may have an average size of about 70nm to about 100nm. In some embodiments, the average size may be about 80nm. In other embodiments, the average size may be about 100nm.
PDI：奈米顆粒組合物可為相對均質的。可使用多分散指數來指示奈米顆粒組合物之均質性，例如奈米顆粒組合物之粒度分佈。較小(例如小於0.3)之多分散指數一般指示較窄之粒度分佈。奈米顆粒組合物之多分散指數可為約0至約0.25，諸如0.01、0.02、0.03、0.04、0.05、0.06、0.07、0.08、0.09、0.10、0.11、0.12、0.13、0.14、0.15、0.16、0.17、0.18、0.19、0.20、0.21、0.22、0.23、0.24或0.25。在一些實施例中，奈米顆粒組合物之多分散指數可為約0.10至約0.20。PDI: The nanoparticle composition can be relatively homogeneous. The polydispersity index can be used to indicate the homogeneity of the nanoparticle composition, such as the particle size distribution of the nanoparticle composition. A smaller polydispersity index (e.g., less than 0.3) generally indicates a narrower particle size distribution. The polydispersity index of the nanoparticle composition can be from about 0 to about 0.25, such as 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.10, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20, 0.21, 0.22, 0.23, 0.24, or 0.25. In some embodiments, the polydispersity index of the nanoparticle composition may be from about 0.10 to about 0.20.
包封效率：治療劑及/或預防劑之包封效率闡述相對於所提供之初始量，在製備後經奈米顆粒組合物包封或以其他方式與奈米顆粒組合物締合之治療劑及/或預防劑的量。包封效率期望地較高(例如接近100%)。包封效率可例如藉由比較在用一或多種有機溶劑或清潔劑破壞奈米顆粒組合物之前與之後含有奈米顆粒組合物之溶液中治療劑及/或預防劑的量來量測。螢光可用於量測溶液中游離治療劑及/或預防劑(例如RNA)的量。對於本文所述之奈米顆粒組合物，治療劑及/或預防劑之包封效率可為至少50%，例如50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%或100%。在一些實施例中，包封效率可為至少80%。在某些實施例中，包封效率可為至少90%。Encapsulation efficiency: The encapsulation efficiency of a therapeutic and/or prophylactic agent describes the amount of therapeutic and/or prophylactic agent encapsulated or otherwise associated with a nanoparticle composition after preparation relative to the initial amount provided. Encapsulation efficiency is desirably high (e.g., close to 100%). Encapsulation efficiency can be measured, for example, by comparing the amount of therapeutic and/or prophylactic agent in a solution containing a nanoparticle composition before and after the nanoparticle composition is disrupted with one or more organic solvents or detergents. Fluorescence can be used to measure the amount of free therapeutic and/or prophylactic agent (e.g., RNA) in a solution. For the nanoparticle compositions described herein, the encapsulation efficiency of the therapeutic and/or prophylactic agent may be at least 50%, for example 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%. In some embodiments, the encapsulation efficiency may be at least 80%. In certain embodiments, the encapsulation efficiency may be at least 90%.
表觀pKa：奈米顆粒組合物之ζ電位可用於指示組合物之動電位。舉例而言，ζ電位可闡述奈米顆粒組合物之表面電荷。具有相對較低正電荷或負電荷之奈米顆粒組合物一般係期望的，此乃因帶較高電荷之物質可與體內之細胞、組織及其他成分發生不期望之相互作用。在一些實施例中，奈米顆粒組合物之ζ電位可為約-10mV至約+20mV、約-10mV至約+15mV、約-10mV至約+10mV、約-10mV至約+5mV、約-10mV至約0mV、約-10mV至約-5mV、約-5mV至約+20mV、約-5mV至約+15mV、約-5mV至約+10mV、約-5mV至約+5mV、約-5mV至約0mV、約0mV至約+20mV、約0mV至約+15mV、約0mV至約+10mV、約0mV至約+5mV、約+5mV至約+20mV、約+5mV至約+15mV或約+5mV至約+10mV。Apparent pKa: The zeta potential of a nanoparticle composition can be used to indicate the zeta potential of the composition. For example, the zeta potential can describe the surface charge of the nanoparticle composition. Nanoparticle compositions with relatively low positive or negative charges are generally desirable because highly charged species can interact undesirably with cells, tissues, and other components in the body. In some embodiments, the zeta potential of the nanoparticle composition can be about -10 mV to about +20 mV, about -10 mV to about +15 mV, about -10 mV to about +10 mV, about -10 mV to about +5 mV, about -10 mV to about 0 mV, about -10 mV to about -5 mV, about -5 mV to about +20 mV, about -5 mV to about +15 mV, about -5mV to about +10mV, about -5mV to about +5mV, about -5mV to about 0mV, about 0mV to about +20mV, about 0mV to about +15mV, about 0mV to about +10mV, about 0mV to about +5mV, about +5mV to about +20mV, about +5mV to about +15mV, or about +5mV to about +10mV.
在另一實施例中，自複製型RNA可調配於脂質體中。作為非限制性實例，自複製型RNA可如以全文引用之方式併入本文中之國際公開案第WO20120067378號中所述而調配於脂質體中。在一個態樣中，脂質體可包含pKa值有利於遞送mRNA之脂質。在另一態樣中，脂質體可在生理pH值下具有實質上呈中性之表面電荷且因此可有效用於免疫(參見例如以全文引用之方式併入本文中之國際公開案第WO20120067378號中所述之脂質體)。In another embodiment, the self-replicating RNA may be formulated in liposomes. As a non-limiting example, the self-replicating RNA may be formulated in liposomes as described in International Publication No. WO20120067378, which is incorporated herein by reference in its entirety. In one aspect, the liposomes may include lipids with a pKa value that is favorable for delivering mRNA. In another aspect, the liposomes may have a substantially neutral surface charge at physiological pH and thus may be effectively used for immunization (see, for example, the liposomes described in International Publication No. WO20120067378, which is incorporated herein by reference in its entirety).
在一些實施例中，所述奈米顆粒組合物包含脂質組分，該脂質組分包括至少一種脂質，諸如本文所述之根據式(I)(及其子式)之一的化合物。舉例而言，在一些實施例中，奈米顆粒組合物可包含脂質組分，該脂質組分包括一種本文所提供之化合物。奈米顆粒組合物亦可包含一或多種如下所述之其他脂質或非脂質組分。In some embodiments, the nanoparticle composition comprises a lipid component, which includes at least one lipid, such as a compound according to one of formula (I) (and its subformulae) described herein. For example, in some embodiments, the nanoparticle composition may comprise a lipid component, which includes a compound provided herein. The nanoparticle composition may also comprise one or more other lipid or non-lipid components as described below.
在一個實施例中，包含本文提供之化合物及mRNA之奈米顆粒組合物顯示出提高之mRNA表現水準(例如，與此項技術中已知之標準陽離子脂質化合物例如MC3相比)。在一個實施例中，在向個體投與包含本文提供之化合物之奈米顆粒組合物後，該化合物顯示出快速之組織清除率(例如肝臟清除率)。In one embodiment, a nanoparticle composition comprising a compound provided herein and mRNA exhibits increased mRNA expression levels (e.g., compared to a standard cationic lipid compound known in the art, such as MC3). In one embodiment, after administering a nanoparticle composition comprising a compound provided herein to a subject, the compound exhibits rapid tissue clearance (e.g., liver clearance).
5.4.1陽離子/可電離脂質5.4.1Cationic/ionizable lipids
如本文所述，在一些實施例中，本文所提供之奈米顆粒組合物除包含根據式(I)(及其子式)之脂質以外，亦包含一或多種帶電或可電離之脂質。不受理論束縛，預期奈米顆粒組合物之某些帶電或兩性離子脂質組分類似於細胞膜中之脂質組分，藉此可改善奈米顆粒之細胞攝取。可形成本發明奈米顆粒組合物之一部分之實例性帶電或可電離之脂質包括但不限於3-(雙十二烷基胺基)-N1,N1,4-三(十二烷基)-1-六氫吡
在一些實施例中，適合陽離子脂質包括氯化N-[1-(2,3-二油基氧基)丙基]-N,N,N-三甲基銨(DOTMA)；氯化N-[1-(2,3-二油醯基氧基)丙基]-N,N,N-三甲基銨(DOTAP)；1,2-二油醯基-sn-甘油-3-乙基磷酸膽鹼(DOEPC)；1,2-二月桂醯基-sn-甘油-3-乙基磷酸膽鹼(DLEPC)；1,2-二肉豆蔻醯基-sn-甘油-3-乙基磷酸膽鹼(DMEPC)；1,2-二肉豆蔻油醯基-sn-甘油-3-乙基磷酸膽鹼(14：1)；N1-[2-((1S)-1-[(3-胺基丙基)胺基]-4-[二(3-胺基-丙基)胺基]丁基甲醯胺基)乙基]-3,4-二[油基氧基]-苯甲醯胺(MVL5)；雙十八烷基醯胺基-甘胺醯基精四胺(DOGS)；3b-[N-(N’,N’-二甲基胺基乙基)胺甲醯基]膽固醇(DC-Chol)；溴化雙十八烷基二甲基銨(DDAB)；SAINT-2,N-甲基-4-(二油基)甲基吡啶鎓；溴化1,2-二肉豆蔻基氧基丙基-3-二甲基羥乙基銨(DMRIE)；溴化1,2-二油醯基-3-二甲基-羥乙基銨(DORIE)；氯化1,2-二油醯基氧基丙基-3-二甲基羥乙基銨(DORI)；二烷基化胺基酸(DILA2)(例如C18：1-norArg-C16)；氯化二油基二甲基銨(DODAC)；1-棕櫚醯基-2-油醯基-sn-甘油-3-乙基磷酸膽鹼(POEPC)；1,2-二肉豆蔻油醯基-sn-甘油-3-乙基磷酸膽鹼(MOEPC)；二油酸(R)-5-(二甲基胺基)戊烷-1,2-二基酯鹽酸鹽(DODAPen-Cl)；二油酸(R)-5-胍基戊烷-1,2-二基酯鹽酸鹽(DOPen-G)；及氯化(R)-N,N,N-三甲基-4,5-雙(油醯基氧基)戊-1-胺鎓鹽(DOTAPen)。具有在生理pH值下帶電荷之頭基之陽離子脂質亦係適合的，諸如一級胺(例如DODAG N’,N’-雙十八烷基-N-4,8-二氮雜-10-胺基癸醯基甘胺酸醯胺)及胍鎓頭基(例如雙-胍鎓-精三胺-膽固醇(BGSC)、雙-胍鎓-tren-膽固醇(BGTC)、PONA及二油酸(R)-5-胍基戊烷-1,2-二基酯鹽酸鹽(DOPen-G))。另一適合之陽離子脂質係二油酸(R)-5-(二甲基胺基)戊烷-1,2-二基酯鹽酸鹽(DODAPen-Cl)。在某些實施例中，陽離子脂質係特定鏡像異構物或外消旋形式，且包括上述陽離子脂質之各種鹽形式(例如氯化物或硫酸鹽)。舉例而言，在一些實施例中，陽離子脂質係氯化N-[1-(2,3-二油醯基氧基)丙基]-N,N,N-三甲基銨(DOTAP-Cl)或硫酸N-[1-(2,3-二油醯基氧基)丙基]-N,N,N-三甲基銨(DOTAP-硫酸鹽)。在一些實施例中，陽離子脂質係可電離陽離子脂質，例如溴化雙十八烷基二甲基銨(DDAB)；1,2-二亞油基氧基-3-二甲基胺基丙烷(DLinDMA)；2,2-二亞油基-4-(2-二甲基胺基乙基)-[1,3]-二氧雜環戊烷(DLin-KC2-DMA)；4-(二甲基胺基)丁酸三十七碳-6,9,28,31-四烯-19-基酯(DLin-MC3-DMA)；1,2-二油醯基氧基-3-二甲基胺基丙烷(DODAP)；1,2-二油基氧基-3-二甲基胺基丙烷(DODMA)；及N-嗎啉基膽固醇(Mo-CHOL)。在某些實施例中，脂質奈米顆粒包含兩種或更多種陽離子脂質(例如兩種或更多種上述陽離子脂質)之組合。In some embodiments, suitable cationic lipids include N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA); N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTAP); 1,2-dioleyl-sn-glycero-3-ethylphosphocholine (DOEPC); ; 1,2-dilauryl-sn-glycero-3-ethylphosphocholine (DLEPC); 1,2-dimyristoyl-sn-glycero-3-ethylphosphocholine (DMEPC); 1,2-dimyristoyl-sn-glycero-3-ethylphosphocholine (14:1); N1-[2-((1S)-1-[(3-aminopropyl)amino]-4-[ 3,4-di[oleyloxy]-benzamide (MVL5); dioctadecylamido-glycine tetramine (DOGS); 3b-[N-(N',N'-dimethylaminoethyl)aminomethyl]cholesterol (DC-Chol); dioctadecyldimethylammonium bromide (DDAB); SA INT-2,N-methyl-4-(dioleyl)methylpyridinium; 1,2-dimyristyloxypropyl-3-dimethylhydroxyethylammonium bromide (DMRIE); 1,2-dioleyl-3-dimethyl-hydroxyethylammonium bromide (DORIE); 1,2-dioleyloxypropyl-3-dimethylhydroxyethylammonium chloride (DORI); dialkylated amino acids (DILA2 ) (e.g. C18: 1-norArg-C16); dioleyl dimethylammonium chloride (DODAC); 1-palmitoyl-2-oleyl-sn-glycero-3-ethylphosphocholine (POEPC); 1,2-dimyristoyl-sn-glycero-3-ethylphosphocholine (MOEPC); (R)-5-(dimethylamino)pentane-1,2-diyl dioleate hydrochloride (DODAPen-Cl); (R)-5-guanidinopentane-1,2-diyl dioleate hydrochloride (DOPen-G); and (R)-N,N,N-trimethyl-4,5-bis(oleyloxy)pentan-1-aminium chloride (DOTAPen). Cationic lipids with head groups that carry a charge at physiological pH are also suitable, such as primary amines (e.g., DODAG N',N'-dioctadecyl-N-4,8-diaza-10-aminodecanoylglycine amide) and guanidinium head groups (e.g., bis-guanidinium-spermtriamine-cholesterol (BGSC), bis-guanidinium-tren-cholesterol (BGTC), PONA, and (R)-5-guanidinopentane-1,2-diyl dioleate hydrochloride (DOPen-G)). Another suitable cationic lipid is (R)-5-(dimethylamino)pentane-1,2-diyl dioleate hydrochloride (DODAPen-Cl). In certain embodiments, the cationic lipid is a specific mirror image isomer or racemic form, and includes various salt forms (e.g., chloride or sulfate) of the above cationic lipids. For example, in some embodiments, the cationic lipid is N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTAP-Cl) or N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium sulfate (DOTAP-sulfate). In some embodiments, the cationic lipid is an ionizable cationic lipid, such as dioctadecyldimethylammonium bromide (DDAB); 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA); 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA); 4-(dimethylamino)butyric acid heptatriacontria-6,9,28,31-tetraen-19-yl ester (DLin-MC3-DMA); 1,2-dioleyloxy-3-dimethylaminopropane (DODAP); 1,2-dioleyloxy-3-dimethylaminopropane (DODMA); and N-morpholinylcholesterol (Mo-CHOL). In certain embodiments, the lipid nanoparticles comprise a combination of two or more cationic lipids (e.g., two or more of the cationic lipids described above).
另外，在一些實施例中，可形成本發明奈米顆粒組合物之一部分之帶電或可電離之脂質係包括環狀胺基之脂質。適於本文所揭示之調配物及方法之額外陽離子脂質包括WO2015199952、WO2016176330及WO2015011633中所述之彼等陽離子脂質，各案之全部內容以全文引用之方式併入本文中。In addition, in some embodiments, the charged or ionizable lipids that can form part of the nanoparticle composition of the present invention are lipids including cyclic amine groups. Additional cationic lipids suitable for the formulations and methods disclosed herein include those described in WO2015199952, WO2016176330, and WO2015011633, the entire contents of each of which are incorporated herein by reference in their entirety.
5.4.2聚合物結合之脂質5.4.2Polymer-conjugated lipids
在一些實施例中，奈米顆粒組合物之脂質組分可包括一或多種聚合物結合之脂質，諸如聚乙二醇化脂質(PEG脂質)。不受理論束縛，預期奈米顆粒組合物中之聚合物結合之脂質組分可改善膠體穩定性及/或減少奈米顆粒之蛋白質吸收。可結合本揭示案使用之實例性聚合物結合之脂質包括但不限於PEG改質之磷脂醯乙醇胺、PEG改質之磷脂酸、PEG改質之神經醯胺、PEG改質之二烷基胺、PEG改質之二醯基甘油、PEG改質之二烷基甘油及其混合物。舉例而言，PEG脂質可為PEG-c-DOMG、PEG-DMG、PEG-DLPE、PEG-DMPE、PEG-DPPC、PEG-DSPE、神經醯胺-PEG2000或Chol-PEG2000。In some embodiments, the lipid component of the nanoparticle composition may include one or more polymer-bound lipids, such as polyethylene glycolated lipids (PEG lipids). Without being bound by theory, it is expected that the polymer-bound lipid component in the nanoparticle composition can improve colloidal stability and/or reduce protein absorption of the nanoparticles. Exemplary polymer-bound lipids that can be used in conjunction with the present disclosure include, but are not limited to, PEG-modified phosphatidylethanolamine, PEG-modified phosphatidic acid, PEG-modified ceramide, PEG-modified dialkylamine, PEG-modified diacylglycerol, PEG-modified dialkylglycerol, and mixtures thereof. For example, the PEG lipid may be PEG-c-DOMG, PEG-DMG, PEG-DLPE, PEG-DMPE, PEG-DPPC, PEG-DSPE, ceramide-PEG2000 or Chol-PEG2000.
在一個實施例中，聚合物結合之脂質係聚乙二醇化脂質。舉例而言，一些實施例包括聚乙二醇化二醯基甘油(PEG-DAG)，諸如1-(單甲氧基-聚乙二醇)-2,3-二肉豆蔻醯基甘油(PEG-DMG)；聚乙二醇化磷脂醯乙醇胺(PEG-PE)；PEG琥珀酸酯二醯基甘油(PEG-S-DAG)，諸如4-O-(2’,3’-二(十四烷醯氧基)丙基-1-O-(ω-甲氧基(聚乙氧基)乙基)丁二酸酯(PEG-S-DMG)；聚乙二醇化神經醯胺(PEG-cer)；或PEG二烷氧基丙基胺基甲酸酯，諸如ω-甲氧基(聚乙氧基)乙基-N-(2,3-二(十四烷氧基)丙基)胺基甲酸酯或2,3-二(十四烷氧基)丙基-N-(ω-甲氧基)(聚乙氧基)乙基)胺基甲酸酯。In one embodiment, the polymer-bound lipid is a pegylated lipid. For example, some embodiments include pegylated diacylglycerol (PEG-DAG), such as 1-(monomethoxy-polyethylene glycol)-2,3-dimyristylglycerol (PEG-DMG); pegylated phosphatidylethanolamine (PEG-PE); PEG succinate diacylglycerol (PEG-S-DAG), such as 4-O-(2',3'-di(tetradecanoyloxy)propyl-1-O-( ω-methoxy(polyethoxy)ethyl)succinate (PEG-S-DMG); PEGylated ceramide (PEG-cer); or PEG dialkoxypropylcarbamates, such as ω-methoxy(polyethoxy)ethyl-N-(2,3-di(tetradecyloxy)propyl)carbamate or 2,3-di(tetradecyloxy)propyl-N-(ω-methoxy)(polyethoxy)ethyl)carbamate.
在一個實施例中，聚合物結合之脂質係以在1.0莫耳%至2.5莫耳%範圍內之濃度存在。在一個實施例中，聚合物結合之脂質係以約1.7莫耳%之濃度存在。在一個實施例中，聚合物結合之脂質係以約1.5莫耳%之濃度存在。In one embodiment, the polymer-bound lipid is present at a concentration ranging from 1.0 mol% to 2.5 mol%. In one embodiment, the polymer-bound lipid is present at a concentration of about 1.7 mol%. In one embodiment, the polymer-bound lipid is present at a concentration of about 1.5 mol%.
在一個實施例中，陽離子脂質與聚合物結合之脂質之莫耳比在約35：1至約25：1範圍內。在一個實施例中，陽離子脂質與聚合物結合之脂質之莫耳比在約100：1至約20：1範圍內。In one embodiment, the molar ratio of the cationic lipid to the polymer-bound lipid is in the range of about 35:1 to about 25:1. In one embodiment, the molar ratio of the cationic lipid to the polymer-bound lipid is in the range of about 100:1 to about 20:1.
在一個實施例中，聚乙二醇化脂質具有下式：
或其醫藥學上可接受之鹽、互變異構物或立體異構物，其中：R12及R13各自獨立地為含有10至30個碳原子之直鏈或具支鏈飽和或不飽和烷基鏈，其中該烷基鏈視情況間雜有一或多個酯鍵；且w具有在30至60範圍內之平均值。or a pharmaceutically acceptable salt, tautomer or stereoisomer thereof, wherein: R12 and R13 are each independently a straight or branched saturated or unsaturated alkyl chain containing 10 to 30 carbon atoms, wherein the alkyl chain is optionally doped with one or more ester bonds; and w has an average value in the range of 30 to 60.
在一個實施例中，R12及R13各自獨立地為含有12至16個碳原子之直鏈飽和烷基鏈。在其他實施例中，平均w在42至55範圍內，例如平均w係42、43、44、45、46、47、48、49、50、51、52、53、54或55。在一些特定實施例中，平均w係約49。In one embodiment, R12 and R13 are each independently a linear saturated alkyl chain containing 12 to 16 carbon atoms. In other embodiments, the average w is in the range of 42 to 55, for example, the average w is 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54 or 55. In some specific embodiments, the average w is about 49.
在一個實施例中，聚乙二醇化脂質具有下式：
其中平均w係約49。The average w is about 49.
5.4.3結構脂質5.4.3Structural lipids
在一些實施例中，奈米顆粒組合物之脂質組分可包括一或多種結構脂質。不受理論束縛，預期結構脂質可使奈米顆粒之兩親結構，諸如但不限於奈米顆粒之脂質雙層結構穩定。可結合本揭示案使用之實例性結構脂質包括但不限於膽固醇、糞固醇、穀固醇、麥角固醇、菜油固醇、豆固醇、菜子固醇、番茄鹼、番茄苷、熊果酸、α-生育酚及其混合物。在某些實施例中，結構脂質係膽固醇。在一些實施例中，結構脂質包括膽固醇及皮質類固醇(諸如普賴蘇穠(prednisolone)、地塞米松(dexamethasone)、普賴松(prednisone)及氫化可體松(hydrocortisone))或其組合。In some embodiments, the lipid component of the nanoparticle composition may include one or more structured lipids. Without being bound by theory, it is expected that the structured lipids can stabilize the amphiphilic structure of the nanoparticle, such as but not limited to the lipid bilayer structure of the nanoparticle. Exemplary structured lipids that can be used in conjunction with the present disclosure include but are not limited to cholesterol, natriuretic acid, glutathione, ergosterol, campesterol, stigmasterol, brassicasterol, tomatine, tomatin, ursolic acid, α-tocopherol, and mixtures thereof. In certain embodiments, the structured lipid is cholesterol. In some embodiments, the structured lipids include cholesterol and corticosteroids (such as prednisolone, dexamethasone, prednisone, and hydrocortisone) or a combination thereof.
在一個實施例中，本文所提供之脂質奈米顆粒包含類固醇或類固醇類似物。在一個實施例中，類固醇或類固醇類似物係膽固醇。在一個實施例中，類固醇係以在39莫耳%至49莫耳%、40莫耳%至46莫耳%、40莫耳%至44莫耳%、40莫耳%至42莫耳%、42莫耳%至44莫耳%或44莫耳%至46莫耳%範圍內之濃度存在。在一個實施例中，類固醇係以40莫耳%、41莫耳%、42莫耳%、43莫耳%、44莫耳%、45莫耳%或46莫耳%之濃度存在。In one embodiment, the lipid nanoparticles provided herein contain a steroid or a steroid analog. In one embodiment, the steroid or steroid analog is cholesterol. In one embodiment, the steroid is present at a concentration ranging from 39 mol% to 49 mol%, 40 mol% to 46 mol%, 40 mol% to 44 mol%, 40 mol% to 42 mol%, 42 mol% to 44 mol%, or 44 mol% to 46 mol%. In one embodiment, the steroid is present at a concentration of 40 mol%, 41 mol%, 42 mol%, 43 mol%, 44 mol%, 45 mol%, or 46 mol%.
在一個實施例中，陽離子脂質與類固醇之莫耳比在1.0：0.9至1.0：1.2、或1.0：1.0至1.0：1.2範圍內。在一個實施例中，陽離子脂質與膽固醇之莫耳比在約5：1至1：1範圍內。在一個實施例中，類固醇以在32莫耳%至40莫耳%類固醇範圍內之濃度存在。In one embodiment, the molar ratio of cationic lipid to steroid is in the range of 1.0:0.9 to 1.0:1.2, or 1.0:1.0 to 1.0:1.2. In one embodiment, the molar ratio of cationic lipid to cholesterol is in the range of about 5:1 to 1:1. In one embodiment, the steroid is present in a concentration in the range of 32 mol% to 40 mol% steroid.
5.4.4磷脂5.4.4Phospholipids
在一些實施例中，奈米顆粒組合物之脂質組分可包括一或多種磷脂，諸如一或多種(多)不飽和脂質。不受理論束縛，預期磷脂可組裝成一或多個脂質雙層結構。可形成本發明奈米顆粒組合物之一部分之實例性磷脂包括但不限於1,2-二硬脂醯基-sn-甘油-3-磷酸膽鹼(DSPC)、1,2-二油醯基-sn-甘油-3-磷酸乙醇胺(DOPE)、1,2-二亞油醯基-sn-甘油-3-磷酸膽鹼(DLPC)、1,2-二肉豆蔻醯基-sn-甘油磷酸膽鹼(DMPC)、1,2-二油醯基-sn-甘油-3-磷酸膽鹼(DOPC)、1,2-二棕櫚醯基-sn-甘油-3-磷酸膽鹼(DPPC)、1,2-二(十一烷醯基)-sn-甘油-磷酸膽鹼(DUPC)、1-棕櫚醯基-2-油醯基-sn-甘油-3-磷酸膽鹼(POPC)、1,2-二-O-十八碳烯基-sn-甘油-3-磷酸膽鹼(18：0 Diether PC)、1-油醯基-2-膽固醇基半琥珀醯基-sn-甘油-3-磷酸膽鹼(OChemsPC)、1-十六烷基-sn-甘油-3-磷酸膽鹼(C16 Lyso PC)、1,2-二亞油醯基-sn-甘油-3-磷酸膽鹼、1,2-二花生四烯醯基-sn-甘油-3-磷酸膽鹼、1,2-二十二碳六烯醯基-sn-甘油-3-磷酸膽鹼、1,2-二植烷醯基-sn-甘油-3-磷酸乙醇胺(ME 16.0 PE)、1,2-二硬脂醯基-sn-甘油-3-磷酸乙醇胺、1,2-二亞油醯基-sn-甘油-3-磷酸乙醇胺、1,2-二亞油醯基-sn-甘油-3-磷酸乙醇胺、1,2-二花生四烯醯基-sn-甘油-3-磷酸乙醇胺、1,2-二十二碳六烯醯基-sn-甘油-3-磷酸乙醇胺、1,2-二油醯基-sn-甘油-3-磷酸-外消旋-(1-甘油)鈉鹽(DOPG)及鞘磷脂。在某些實施例中，奈米顆粒組合物包含DSPC。在某些實施例中，奈米顆粒組合物包含DOPE。在一些實施例中，奈米顆粒組合物包含DSPC及DOPE二者。In some embodiments, the lipid component of the nanoparticle composition may include one or more phospholipids, such as one or more (poly)unsaturated lipids. Without being bound by theory, it is expected that the phospholipids can be assembled into one or more lipid bilayer structures. Exemplary phospholipids that can form part of the nanoparticle composition of the present invention include, but are not limited to, 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 1,2-dilinoleyl-sn-glycero-3-phosphocholine (DLPC), 1,2-dimyristoyl-sn-glycerophosphocholine (DMPC), 1,2 -dioleyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-di(undecanyl)-sn-glycero-phosphocholine (DUPC), 1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine (POPC), 1,2-di-O-octadecenyl-sn-glycero-3-phosphocholine (18:0 Diether PC), 1-oleyl-2-cholesterol hemisuccinyl-sn-glycero-3-phosphocholine (OChemsPC), 1-hexadecyl-sn-glycero-3-phosphocholine (C16 Lyso PC), 1,2-dilinoleyl-sn-glycero-3-phosphocholine, 1,2-diarachidonyl-sn-glycero-3-phosphocholine, 1,2-docosahexaenoyl-sn-glycero-3-phosphocholine, 1,2-diphytanyl-sn-glycero-3-phosphoethanolamine (ME 16.0 PE), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinoleyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinoleyl-sn-glycero-3-phosphoethanolamine, 1,2-diarachidonyl-sn-glycero-3-phosphoethanolamine, 1,2-docosahexaenoyl-sn-glycero-3-phosphoethanolamine, 1,2-dioleoyl-sn-glycero-3-phospho-racemic-(1-glycerol) sodium salt (DOPG) and sphingomyelin. In some embodiments, the nanoparticle composition comprises DSPC. In some embodiments, the nanoparticle composition comprises DOPE.In some embodiments, the nanoparticle composition comprises both DSPC and DOPE.
另外之實例性中性脂質包括例如二棕櫚醯基磷脂醯甘油(DPPG)、棕櫚醯基油醯基磷脂醯乙醇胺(POPE)及二油醯基磷脂醯乙醇胺4-(N-馬來醯亞胺基甲基)-環己烷-1-甲酸酯(DOPE-mal)、二棕櫚醯基磷脂醯乙醇胺(DPPE)、二肉豆蔻醯基磷酸乙醇胺(DMPE)、二硬脂醯基-磷脂醯乙醇胺(DSPE)、16-O-單甲基PE、16-O-二甲基PE、18-1-反式PE、1-硬脂醯基-2-油醯基磷脂醯乙醇胺(SOPE)及1,2-二反油醯基-sn-甘油-3-磷酸乙醇胺(反式DOPE)。在一個實施例中，中性脂質係1,2-二硬脂醯基-sn-甘油-3磷酸膽鹼(DSPC)。在一個實施例中，中性脂質選自DSPC、DPPC、DMPC、DOPC、POPC、DOPE及SM。Additional exemplary neutral lipids include, for example, dipalmitoylphosphatidylglycerol (DPPG), palmitoyloleylphosphatidylethanolamine (POPE) and dioleoylphosphatidylethanolamine 4-(N-maleimidomethyl)-cyclohexane-1-carboxylate (DOPE-mal), dipalmitoylphosphatidylethanolamine (DPPE), dimyristoylphosphoethanolamine (DMPE), distearyl-phosphatidylethanolamine (DSPE), 16-O-monomethyl PE, 16-O-dimethyl PE, 18-1-trans PE, 1-stearyl-2-oleylphosphatidylethanolamine (SOPE), and 1,2-ditransoleyl-sn-glycero-3-phosphoethanolamine (trans-DOPE). In one embodiment, the neutral lipid is 1,2-distearyl-sn-glycero-3-phosphocholine (DSPC). In one embodiment, the neutral lipid is selected from DSPC, DPPC, DMPC, DOPC, POPC, DOPE and SM.
在一個實施例中，中性脂質係磷脂醯膽鹼(PC)、磷脂醯乙醇胺(PE)、磷脂醯絲胺酸(PS)、磷脂酸(PA)或磷脂醯甘油(PG)。In one embodiment, the neutral lipid is phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidic acid (PA) or phosphatidylglycerol (PG).
可形成本發明奈米顆粒組合物之一部分之額外磷脂亦包括WO2017/112865中所闡述之彼等磷脂，其全部內容以全文引用之方式併入本文中。Additional phospholipids that may form part of the nanoparticle composition of the present invention also include those described in WO2017/112865, the entire contents of which are incorporated herein by reference in their entirety.
5.4.5治療性有效負載5.4.5Therapeutic effective load
根據本揭示案，本文所述之奈米顆粒組合物可進一步包含一或多種治療劑及/或預防劑。此等治療劑及/或預防劑在本揭示案中有時稱為「治療性有效負載」或「有效負載」。在一些實施例中，治療性有效負載可在活體內或活體外，使用奈米顆粒作為遞送媒劑投與。According to the present disclosure, the nanoparticle compositions described herein may further comprise one or more therapeutic agents and/or prophylactic agents. Such therapeutic agents and/or prophylactic agents are sometimes referred to as "therapeutic payloads" or "payloads" in the present disclosure. In some embodiments, the therapeutic payloads may be administered in vivo or in vitro using nanoparticles as a delivery vehicle.
在一些實施例中，奈米顆粒組合物包含以下作為治療性有效負載：小分子化合物(例如小分子藥物)，諸如抗癌劑(例如長春新鹼(vincristine)、多柔比星(doxorubicin)、米托蒽醌(mitoxantrone)、喜樹鹼(camptothecin)、順鉑(cisplatin)、博萊黴素(bleomycin)、環磷醯胺(cyclophosphamide)、胺甲喋呤(methotrexate)及鏈脲佐菌素(streptozotocin))、抗腫瘤劑(例如放線菌素D(actinomycin D)、長春新鹼、長春花鹼(vinblastine)、胞嘧啶阿拉伯糖苷(cytosine arabinoside)、蒽環黴素(anthracyclines)、烷基化劑、鉑類化合物、抗代謝物及核苷類似物，諸如胺甲喋呤以及嘌呤及嘧啶類似物)、抗感染劑、局麻藥(例如地布卡因(dibucaine)及氯丙嗪(chlorpromazine))、β-腎上腺素性阻斷劑(例如普萘洛爾(propranolol)、噻嗎洛爾(timolol)及拉貝洛爾(labetalol))、抗高血壓劑(例如可尼丁(clonidine)及聯胺肼(hydralazine))、抗抑鬱劑(例如伊米帕明(imipramine)、阿米替林(amitriptyline)及多塞平(doxepin))、抗痙攣劑(例如苯妥英(phenytoin))、抗組胺劑(例如苯海拉明(diphenhydramine)、氯菲安明(chlorpheniramine)及普敏太定(promethazine))、抗生素/抗細菌劑(例如慶大黴素(gentamycin)、塞普沙辛(ciprofloxacin)及頭孢西丁(cefoxitin))、抗真菌劑(例如咪康唑(miconazole)、特康唑(terconazole)、益康唑(econazole)、異康唑(isoconazole)、布康唑(butaconazole)、克氯黴唑(clotrimazole)、艾妥可那唑(itraconazole)、制黴菌素(nystatin)、萘替芬(naftifine)及兩性黴素B(amphotericin B))、抗寄生蟲劑、激素、激素拮抗劑、免疫調節劑、神經傳遞質拮抗劑、抗青光眼劑、維生素、麻醉劑及成像劑。In some embodiments, the nanoparticle composition comprises the following as a therapeutic effective cargo: small molecule compounds (e.g., small molecule drugs), such as anticancer agents (e.g., vincristine, doxorubicin, mitoxantrone, camptothecin, cisplatin, bleomycin, cyclophosphamide, methotrexate, and streptozotocin), antitumor agents (e.g., actinomycin D, vincristine, vinblastine, cytosine arabinoside, arabinoside), anthracyclines, alkylating agents, platinum compounds, anti-metabolites and nucleoside analogs such as methotrexate and purine and pyrimidine analogs), anti-infective agents, local anesthetics (such as dibucaine and chlorpromazine), beta-adrenergic blocking agents (such as propranolol, tiamoxic acid, antihypertensives (such as clonidine and hydralazine), antidepressants (such as imipramine, amitriptyline, and doxepin), anticonvulsants (such as phenytoin), antihistamines (e.g. diphenhydramine, chlorpheniramine, and promethazine), antibiotics/antibacterials (e.g. gentamycin, ciprofloxacin, and cefoxitin), antifungals (e.g. miconazole, terconazole, econazole, isoconazole, butaconazole, clotrimazole, itraconazole, nystatin, naftifine, and amphotericin B), B)), antiparasitic agents, hormones, hormone antagonists, immunomodulators, neurotransmitter antagonists, anti-glaucoma agents, vitamins, anesthetics and imaging agents.
在一些實施例中，治療性有效負載包含細胞毒素、放射性離子、化學治療劑、疫苗、引起免疫反應之化合物及/或另一治療劑及/或預防劑。細胞毒素或細胞毒性劑包括可能對細胞有害之任何劑。實例包括但不限於紫杉醇(taxol)、細胞鬆弛素B(cytochalasin B)、短桿菌肽D(gramicidin D)、溴化乙錠(ethidium bromide)、依米丁(emetine)、絲裂黴素(mitomycin)、依托泊苷(etoposide)、替尼泊苷(teniposide)、長春新鹼、長春花鹼、秋水仙鹼(colchicine)、多柔比星、道諾黴素(daunorubicin)、二羥基蒽二酮(dihydroxyanthracinedione)、米托蒽醌、米拉黴素(mithramycin)、放線菌素D、1-去氫睪酮、糖皮質激素、普魯卡因(procaine)、丁卡因(tetracaine)、利多卡因(lidocaine)、普萘洛爾、嘌呤黴素(puromycin)、美登素類化合物(maytansinoids)，例如美登醇(maytansinol)、拉奇黴素(rachelmycin)(CC-1065)以及其類似物或同系物。放射性離子包括但不限於碘(例如碘125或碘131)、鍶89、磷、鈀、銫、銥、磷酸根、鈷、釔90、釤153及鐠。In some embodiments, the therapeutic payload comprises a cytotoxin, a radioactive ion, a chemotherapeutic agent, a vaccine, an immunogenic compound, and/or another therapeutic and/or prophylactic agent. A cytotoxin or cytotoxic agent includes any agent that may be harmful to cells. Examples include, but are not limited to, taxol, cytochalasin B, gramicidin D, ethidium bromide, bromide, emetine, mitomycin, etoposide, teniposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxyanthracinedione, mitoxantrone, mithram ycin), actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, puromycin, maytansinoids, such as maytansinol, rachelmycin (CC-1065), and analogs or homologues thereof. Radioactive ions include, but are not limited to, iodine (such as iodine-125 or iodine-131), strontium-89, phosphorus, palladium, cesium, iridium, phosphate, cobalt, yttrium-90, samarium-153, and thorium.
在其他實施例中，本發明奈米顆粒組合物之治療性有效負載可包括但不限於治療劑及/或預防劑，諸如抗代謝物(例如胺甲喋呤、6-巰基嘌呤、6-硫鳥嘌呤、阿糖胞苷(cytarabine)、5-氟尿嘧啶、達卡巴嗪(dacarbazine))、烷基化劑(例如氮芥(mechlorethamine)、噻替哌(thiotepa)、苯丁酸氮芥(chlorambucil)、拉奇黴素(CC-1065)、黴法蘭(melphalan)、卡莫司汀(carmustine)(BSNU)、洛莫司汀(lomustine)(CCNU)、環磷醯胺、白消安(busulfan)、二溴甘露醇(dibromomannitol)、鏈脲佐菌素、絲裂黴素C及順二氯二胺鉑(II)(DDP)順鉑)、蒽環黴素(例如道諾黴素(先前稱為柔紅黴素(daunomycin))及多柔比星)、抗生素(例如放線菌素D(dactinomycin)(先前稱為放線菌素))、博萊黴素、米拉黴素及安曲黴素(anthramycin)(AMC))及抗有絲分裂劑(例如長春新鹼、長春花鹼、紫杉醇及美登素類化合物)。In other embodiments, the therapeutic effective load of the nanoparticle composition of the present invention may include but is not limited to therapeutic agents and/or preventive agents, such as anti-metabolites (e.g., methotrexate, 6-hydroxypurine, 6-thioguanine, cytarabine, 5-fluorouracil, dacarbazine), alkylating agents (e.g., mechlorethamine, thiotepa, chlorambucil, razumacin (CC-1065), melphalan, carmustine (BSNU), lomustine ( lomustine (CCNU), cyclophosphamide, busulfan, dibromomannitol, streptozotocin, mitomycin C and cis-dichlorodiamine platinum (II) (DDP) cis-platinum), anthracyclines (e.g. daunomycin (formerly daunomycin) and doxorubicin), antibiotics (e.g. dactinomycin (formerly actinomycin), bleomycin, milamycin and anthramycin (AMC)) and antimitotics (e.g. vincristine, vinblastine, paclitaxel and maytansine compounds).
在一些實施例中，奈米顆粒組合物包含諸如肽及多肽之類生物分子作為治療性有效負載。形成本發明奈米顆粒組合物之一部分之生物分子可為天然來源或合成的。舉例而言，在一些實施例中，本發明奈米顆粒組合物之治療性有效負載可包括但不限於慶大黴素、阿米卡星(amikacin)、胰島素、紅血球生成素(EPO)、顆粒球集落刺激因子(G-CSF)、顆粒球-巨噬細胞集落刺激因子(GM-CSF)、因子VIR、促黃體激素釋放激素(LHRH)類似物、干擾素、肝素、B型肝炎表面抗原、傷寒疫苗、霍亂疫苗以及肽及多肽。In some embodiments, the nanoparticle composition comprises biomolecules such as peptides and polypeptides as therapeutic payloads. The biomolecules forming part of the nanoparticle composition of the present invention may be of natural origin or synthetic. For example, in some embodiments, the therapeutic payload of the nanoparticle composition of the present invention may include, but is not limited to, gentamicin, amikacin, insulin, erythropoietin (EPO), granulocyte colony stimulating factor (G-CSF), granulocyte-macrophage colony stimulating factor (GM-CSF), factor VIR, luteinizing hormone-releasing hormone (LHRH) analogs, interferon, heparin, hepatitis B surface antigen, typhoid vaccine, cholera vaccine, and peptides and polypeptides.
5.4.5.1核酸5.4.5.1Nucleic Acids
在一些實施例中，本發明奈米顆粒組合物包含一或多種核酸分子(例如DNA或RNA分子)作為治療性有效負載。可包括在本發明奈米顆粒組合物中作為治療性有效負載之核酸分子之例示性形式包括但不限於以下一或多種：去氧核糖核酸(DNA)、核糖核酸(RNA)，包括信使mRNA(mRNA)、其雜交體、RNAi誘導劑、RNAi劑、siRNA、shRNA、miRNA、反義RNA、核糖酶、催化性DNA、誘導形成三螺旋之RNA、適體、載體等。在某些實施例中，治療性有效負載包含RNA。可包括在本發明奈米顆粒組合物中作為治療性有效負載之RNA分子包括但不限於短聚體(shortmer)、agomir、antagomir、反義體(antisense)、核糖酶、小干擾RNA(siRNA)、不對稱干擾RNA(aiRNA)、微小RNA(miRNA)、Dicer-受質RNA(dsRNA)、小髮夾RNA(shRNA)、轉移RNA(tRNA)、信使RNA(mRNA)及此項技術中已知之RNA分子之其他形式。在特定實施例中，RNA係mRNA。In some embodiments, the nanoparticle composition of the present invention comprises one or more nucleic acid molecules (e.g., DNA or RNA molecules) as therapeutic payloads. Exemplary forms of nucleic acid molecules that can be included in the nanoparticle composition of the present invention as therapeutic payloads include, but are not limited to, one or more of the following: deoxyribonucleic acid (DNA), ribonucleic acid (RNA), including messenger mRNA (mRNA), hybrids thereof, RNAi inducers, RNAi agents, siRNA, shRNA, miRNA, antisense RNA, ribozymes, catalytic DNA, RNA that induces triple helix formation, aptamers, carriers, etc. In certain embodiments, the therapeutic payload comprises RNA. RNA molecules that can be included in the nanoparticle composition of the present invention as therapeutic effective loads include but are not limited to shortmers, agomirs, antagomirs, antisenses, ribozymes, small interfering RNAs (siRNAs), asymmetric interfering RNAs (aiRNAs), microRNAs (miRNAs), Dicer-substrate RNAs (dsRNAs), small hairpin RNAs (shRNAs), transfer RNAs (tRNAs), messenger RNAs (mRNAs), and other forms of RNA molecules known in the art. In a specific embodiment, the RNA is mRNA.
在其他實施例中，奈米顆粒組合物包含siRNA分子作為治療性有效負載。具體而言，在一些實施例中，siRNA分子能夠選擇性干擾並下調感興趣基因之表現。舉例而言，在一些實施例中，在向有需要之個體投與包含siRNA之奈米顆粒組合物後，siRNA有效負載使與特定疾病、病症或疾患相關之基因選擇性沈默。在一些實施例中，siRNA分子包含與編碼感興趣蛋白質產物之mRNA序列互補之序列。在一些實施例中，siRNA分子係免疫調節性siRNA。In other embodiments, the nanoparticle composition comprises siRNA molecules as therapeutic payloads. Specifically, in some embodiments, the siRNA molecules are capable of selectively interfering with and downregulating the expression of a gene of interest. For example, in some embodiments, after administering a nanoparticle composition comprising siRNA to an individual in need, the siRNA payload selectively silences a gene associated with a specific disease, disorder, or condition. In some embodiments, the siRNA molecule comprises a sequence complementary to an mRNA sequence encoding a protein product of interest. In some embodiments, the siRNA molecule is an immunomodulatory siRNA.
在一些實施例中，奈米顆粒組合物包含shRNA分子或編碼shRNA分子之載體作為治療性有效負載。具體而言，在一些實施例中，治療性有效負載在投與目標細胞後，在目標細胞內產生shRNA。與shRNA相關之構築體及機制係相關技術中熟知的。In some embodiments, the nanoparticle composition comprises an shRNA molecule or a vector encoding an shRNA molecule as a therapeutic effective load. Specifically, in some embodiments, the therapeutic effective load produces shRNA in the target cell after being administered to the target cell. The constructs and mechanisms associated with shRNA are well known in the relevant technology.
在一些實施例中，奈米顆粒組合物包含mRNA分子作為治療性有效負載。具體而言，在一些實施例中，該mRNA分子編碼感興趣多肽，包括任何天然或非天然存在或以其他方式修飾之多肽。由mRNA編碼之多肽可具有任何大小且可具有任何二級結構或活性。在一些實施例中，由mRNA有效負載編碼之多肽當在細胞中表現時可具有治療作用。In some embodiments, the nanoparticle composition comprises an mRNA molecule as a therapeutic payload. Specifically, in some embodiments, the mRNA molecule encodes a polypeptide of interest, including any naturally or non-naturally occurring or otherwise modified polypeptide. The polypeptide encoded by the mRNA can be of any size and can have any secondary structure or activity. In some embodiments, the polypeptide encoded by the mRNA payload can have a therapeutic effect when expressed in a cell.
在一些實施例中，本揭示案之核酸分子包含mRNA分子。在特定實施例中，核酸分子包含至少一個編碼感興趣肽或多肽之編碼區(例如開放閱讀框(ORF))。在一些實施例中，核酸分子亦包含至少一個非轉譯區(UTR)。在特定實施例中，非轉譯區(UTR)位於編碼區之上游(5’端)，且在本文中稱為5’-UTR。在特定實施例中，非轉譯區(UTR)位於編碼區之下游(3’端)，且在本文中稱為3’-UTR。在特定實施例中，核酸分子包含5’-UTR及3’-UTR二者。在一些實施例中，5’-UTR包含5’-帽結構。在一些實施例中，核酸分子包含Kozak序列(例如在5’-UTR中)。在一些實施例中，核酸分子包含poly-A區(例如在3’-UTR中)。在一些實施例中，核酸分子包含聚腺苷酸化信號(例如在3’-UTR中)。在一些實施例中，核酸分子包含穩定區(例如在3’-UTR中)。在一些實施例中，核酸分子包含二級結構。在一些實施例中，二級結構係莖-環。在一些實施例中，核酸分子包含莖-環序列(例如在5’-UTR及/或3’-UTR中)。在一些實施例中，核酸分子包含一或多個能夠在剪接過程中切除之內含子區。在特定實施例中，核酸分子包含一或多個選自5’-UTR及編碼區之區域。在特定實施例中，核酸分子包含一或多個選自編碼區及3’-UTR之區域。在特定實施例中，核酸分子包含一或多個選自5’-UTR、編碼區及3’-UTR之區域。In some embodiments, the nucleic acid molecules of the present disclosure comprise mRNA molecules. In specific embodiments, the nucleic acid molecules comprise at least one coding region (e.g., an open reading frame (ORF)) encoding a peptide or polypeptide of interest. In some embodiments, the nucleic acid molecules also comprise at least one non-translated region (UTR). In specific embodiments, the non-translated region (UTR) is located upstream (5' end) of the coding region and is referred to herein as the 5'-UTR. In specific embodiments, the non-translated region (UTR) is located downstream (3' end) of the coding region and is referred to herein as the 3'-UTR. In specific embodiments, the nucleic acid molecules comprise both the 5'-UTR and the 3'-UTR. In some embodiments, the 5'-UTR comprises a 5'-cap structure. In some embodiments, the nucleic acid molecules comprise a Kozak sequence (e.g., in the 5'-UTR). In some embodiments, the nucleic acid molecule comprises a poly-A region (e.g., in a 3'-UTR). In some embodiments, the nucleic acid molecule comprises a polyadenylation signal (e.g., in a 3'-UTR). In some embodiments, the nucleic acid molecule comprises a stabilizing region (e.g., in a 3'-UTR). In some embodiments, the nucleic acid molecule comprises a secondary structure. In some embodiments, the secondary structure is a stem-loop. In some embodiments, the nucleic acid molecule comprises a stem-loop sequence (e.g., in a 5'-UTR and/or a 3'-UTR). In some embodiments, the nucleic acid molecule comprises one or more intron regions that can be excised during splicing. In specific embodiments, the nucleic acid molecule comprises one or more regions selected from a 5'-UTR and a coding region. In specific embodiments, the nucleic acid molecule comprises one or more regions selected from a coding region and a 3'-UTR. In certain embodiments, the nucleic acid molecule comprises one or more regions selected from 5'-UTR, coding region and 3'-UTR.
編碼區Coding area
在一些實施例中，本揭示案之核酸分子包含至少一個編碼區。在一些實施例中，編碼區係編碼單一肽或蛋白質之開放閱讀框(ORF)。在一些實施例中，編碼區包含至少兩個ORF，每個ORF編碼一種肽或蛋白質。在編碼區包含多於一個ORF之實施例中，編碼之肽及/或蛋白質可彼此相同或不同。在一些實施例中，編碼區中之多個ORF經非編碼序列隔開。在特定實施例中，隔開兩個ORF之非編碼序列包含內部核糖體進入位點(IRES)。In some embodiments, the nucleic acid molecules of the present disclosure comprise at least one coding region. In some embodiments, the coding region is an open reading frame (ORF) encoding a single peptide or protein. In some embodiments, the coding region comprises at least two ORFs, each ORF encoding a peptide or protein. In embodiments where the coding region comprises more than one ORF, the encoded peptides and/or proteins may be the same or different from each other. In some embodiments, multiple ORFs in the coding region are separated by non-coding sequences. In specific embodiments, the non-coding sequence separating two ORFs comprises an internal ribosome entry site (IRES).
不受理論束縛，預期內部核糖體進入位點(IRES)可用作唯一的核糖體結合位點，或充當mRNA之多個核糖體結合位點之一。包含多於一個功能性核糖體結合位點之mRNA分子可編碼若干肽或多肽，該等肽或多肽獨立地由核糖體轉譯(例如多順反子mRNA)。因此，在一些實施例中，本揭示案之核酸分子(例如mRNA)包含一或多個內部核糖體進入位點(IRES)。可以與本揭示案結合使用之IRES序列之實例包括但不限於來自小RNA病毒(例如FMDV)、害蟲病毒(CFFV)、脊髓灰質炎病毒(PV)、腦心肌炎病毒(ECMV)、口蹄疫病毒(FMDV)、C型肝炎病毒(HCV)、豬瘟病毒(CSFV)、鼠白血病病毒(MLV)、猴免疫缺陷病毒(SIV)或蟋蟀麻痺病毒(CrPV)之彼等IRES序列。Without being bound by theory, it is expected that an internal ribosome entry site (IRES) can function as the sole ribosome binding site, or as one of multiple ribosome binding sites for an mRNA. An mRNA molecule comprising more than one functional ribosome binding site can encode a number of peptides or polypeptides that are independently translated by the ribosome (e.g., a polycistronic mRNA). Thus, in some embodiments, a nucleic acid molecule (e.g., an mRNA) of the present disclosure comprises one or more internal ribosome entry sites (IRES). Examples of IRES sequences that can be used in conjunction with the present disclosure include, but are not limited to, those from picornaviruses (e.g., FMDV), entomoviruses (CFFV), polioviruses (PV), encephalomyocarditis virus (ECMV), foot-and-mouth disease virus (FMDV), hepatitis C virus (HCV), swine fever virus (CSFV), murine leukemia virus (MLV), simian immunodeficiency virus (SIV), or cricket paralysis virus (CrPV).
在各個實施例中，本揭示案之核酸分子編碼至少1、2、3、4、5、6、7、8、9、10種或更多種肽或蛋白質。核酸分子編碼之肽及蛋白質可相同或不同。在一些實施例中，本揭示案之核酸分子編碼二肽(例如肌肽及鵝肌肽)。在一些實施例中，核酸分子編碼三肽。在一些實施例中，核酸分子編碼四狀。在一些實施例中，核酸分子編碼五肽。在一些實施例中，核酸分子編碼六肽。在一些實施例中，核酸分子編碼七肽。在一些實施例中，核酸分子編碼八肽。在一些實施例中，核酸分子編碼九肽。在一些實施例中，核酸分子編碼十肽。在一些實施例中，核酸分子編碼具有至少約15個胺基酸之肽或多肽。在一些實施例中，核酸分子編碼具有至少約50個胺基酸之肽或多肽。在一些實施例中，核酸分子編碼具有至少約100個胺基酸之肽或多肽。在一些實施例中，核酸分子編碼具有至少約150個胺基酸之肽或多肽。在一些實施例中，核酸分子編碼具有至少約300個胺基酸之肽或多肽。在一些實施例中，核酸分子編碼具有至少約500個胺基酸之肽或多肽。在一些實施例中，核酸分子編碼具有至少約1000個胺基酸之肽或多肽。In various embodiments, the nucleic acid molecules of the present disclosure encode at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more peptides or proteins. The peptides and proteins encoded by the nucleic acid molecules may be the same or different. In some embodiments, the nucleic acid molecules of the present disclosure encode dipeptides (e.g., carnosine and goose carnosine). In some embodiments, the nucleic acid molecules encode tripeptides. In some embodiments, the nucleic acid molecules encode tetrapeptides. In some embodiments, the nucleic acid molecules encode pentapeptides. In some embodiments, the nucleic acid molecules encode hexapeptides. In some embodiments, the nucleic acid molecules encode heptapeptides. In some embodiments, the nucleic acid molecules encode octapeptides. In some embodiments, the nucleic acid molecules encode nonapeptides. In some embodiments, the nucleic acid molecules encode decapeptides. In some embodiments, the nucleic acid molecules encode peptides or polypeptides having at least about 15 amino acids. In some embodiments, the nucleic acid molecule encodes a peptide or polypeptide having at least about 50 amino acids. In some embodiments, the nucleic acid molecule encodes a peptide or polypeptide having at least about 100 amino acids. In some embodiments, the nucleic acid molecule encodes a peptide or polypeptide having at least about 150 amino acids. In some embodiments, the nucleic acid molecule encodes a peptide or polypeptide having at least about 300 amino acids. In some embodiments, the nucleic acid molecule encodes a peptide or polypeptide having at least about 500 amino acids. In some embodiments, the nucleic acid molecule encodes a peptide or polypeptide having at least about 1000 amino acids.
在一些實施例中，本揭示案之核酸分子之長度為至少約30個核苷酸(nt)。在一些實施例中，核酸分子之長度為至少約35nt。在一些實施例中，核酸分子之長度為至少約40nt。在一些實施例中，核酸分子之長度為至少約45nt。在一些實施例中，核酸分子之長度為至少約50nt。在一些實施例中，核酸分子之長度為至少約55nt。在一些實施例中，核酸分子之長度為至少約60nt。在一些實施例中，核酸分子之長度為至少約65nt。在一些實施例中，核酸分子之長度為至少約70nt。在一些實施例中，核酸分子之長度為至少約75nt。在一些實施例中，核酸分子之長度為至少約80nt。在一些實施例中，核酸分子之長度為至少約85nt。在一些實施例中，核酸分子之長度為至少約90nt。在一些實施例中，核酸分子之長度為至少約95nt。在一些實施例中，核酸分子之長度為至少約100nt。在一些實施例中，核酸分子之長度為至少約120nt。在一些實施例中，核酸分子之長度為至少約140nt。在一些實施例中，核酸分子之長度為至少約160nt。在一些實施例中，核酸分子之長度為至少約180nt。在一些實施例中，核酸分子之長度為至少約200nt。在一些實施例中，核酸分子之長度為至少約250nt。在一些實施例中，核酸分子之長度為至少約300nt。在一些實施例中，核酸分子之長度為至少約400nt。在一些實施例中，核酸分子之長度為至少約500nt。在一些實施例中，核酸分子之長度為至少約600nt。在一些實施例中，核酸分子之長度為至少約700nt。在一些實施例中，核酸分子之長度為至少約800nt。在一些實施例中，核酸分子之長度為至少約900nt。在一些實施例中，核酸分子之長度為至少約1000nt。在一些實施例中，核酸分子之長度為至少約1100nt。在一些實施例中，核酸分子之長度為至少約1200nt。在一些實施例中，核酸分子之長度為至少約1300nt。在一些實施例中，核酸分子之長度為至少約1400nt。在一些實施例中，核酸分子之長度為至少約1500nt。在一些實施例中，核酸分子之長度為至少約1600nt。在一些實施例中，核酸分子之長度為至少約1700nt。在一些實施例中，核酸分子之長度為至少約1800nt。在一些實施例中，核酸分子之長度為至少約1900nt。在一些實施例中，核酸分子之長度為至少約2000nt。在一些實施例中，核酸分子之長度為至少約2500nt。在一些實施例中，核酸分子之長度為至少約3000nt。在一些實施例中，核酸分子之長度為至少約3500nt。在一些實施例中，核酸分子之長度為至少約4000nt。在一些實施例中，核酸分子之長度為至少約4500nt。在一些實施例中，核酸分子之長度為至少約5000nt。In some embodiments, the length of the nucleic acid molecule of the present disclosure is at least about 30 nucleotides (nt). In some embodiments, the length of the nucleic acid molecule is at least about 35nt. In some embodiments, the length of the nucleic acid molecule is at least about 40nt. In some embodiments, the length of the nucleic acid molecule is at least about 45nt. In some embodiments, the length of the nucleic acid molecule is at least about 50nt. In some embodiments, the length of the nucleic acid molecule is at least about 55nt. In some embodiments, the length of the nucleic acid molecule is at least about 60nt. In some embodiments, the length of the nucleic acid molecule is at least about 65nt. In some embodiments, the length of the nucleic acid molecule is at least about 70nt. In some embodiments, the length of the nucleic acid molecule is at least about 75nt. In some embodiments, the length of the nucleic acid molecule is at least about 80nt. In some embodiments, the length of the nucleic acid molecule is at least about 85nt. In some embodiments, the length of the nucleic acid molecule is at least about 90nt. In some embodiments, the length of the nucleic acid molecule is at least about 95nt. In some embodiments, the length of the nucleic acid molecule is at least about 100nt. In some embodiments, the length of the nucleic acid molecule is at least about 120nt. In some embodiments, the length of the nucleic acid molecule is at least about 140nt. In some embodiments, the length of the nucleic acid molecule is at least about 160nt. In some embodiments, the length of the nucleic acid molecule is at least about 180nt. In some embodiments, the length of the nucleic acid molecule is at least about 200nt. In some embodiments, the length of the nucleic acid molecule is at least about 250nt. In some embodiments, the length of the nucleic acid molecule is at least about 300nt. In some embodiments, the length of the nucleic acid molecule is at least about 400nt. In some embodiments, the length of the nucleic acid molecule is at least about 500nt. In some embodiments, the length of the nucleic acid molecule is at least about 600nt. In some embodiments, the length of the nucleic acid molecule is at least about 700nt. In some embodiments, the length of the nucleic acid molecule is at least about 800nt. In some embodiments, the length of the nucleic acid molecule is at least about 900nt. In some embodiments, the length of the nucleic acid molecule is at least about 1000nt. In some embodiments, the length of the nucleic acid molecule is at least about 1100nt. In some embodiments, the length of the nucleic acid molecule is at least about 1200nt. In some embodiments, the length of the nucleic acid molecule is at least about 1300nt. In some embodiments, the length of the nucleic acid molecule is at least about 1400nt. In some embodiments, the length of the nucleic acid molecule is at least about 1500nt. In some embodiments, the length of the nucleic acid molecule is at least about 1600nt. In some embodiments, the length of the nucleic acid molecule is at least about 1700nt. In some embodiments, the length of the nucleic acid molecule is at least about 1800nt. In some embodiments, the length of the nucleic acid molecule is at least about 1900nt. In some embodiments, the length of the nucleic acid molecule is at least about 2000nt. In some embodiments, the length of the nucleic acid molecule is at least about 2500nt. In some embodiments, the length of the nucleic acid molecule is at least about 3000nt. In some embodiments, the length of the nucleic acid molecule is at least about 3500nt. In some embodiments, the length of the nucleic acid molecule is at least about 4000nt. In some embodiments, the length of the nucleic acid molecule is at least about 4500nt. In some embodiments, the length of the nucleic acid molecule is at least about 5000nt.
在特定實施例中，治療性有效負載包含如本文所述之疫苗組合物(例如基因疫苗)。在一些實施例中，治療性有效負載包含能夠引發針對一或多種目標疾患或疾病之免疫之化合物。在一些實施例中，目標疾患與病原體感染相關或由病原體感染引起，該病原體諸如為冠狀病毒(例如2019-nCoV)、流感病毒、麻疹病毒、人乳頭瘤病毒(HPV)、狂犬病病毒、腦膜炎病毒、百日咳病毒、破傷風病毒、鼠疫病毒、肝炎病毒及結核病病毒。在一些實施例中，治療性有效負載包含編碼病原體特有之致病性蛋白質或其抗原片段或抗原決定基之核酸序列(例如mRNA)。疫苗在投與經疫苗接種之個體後，允許表現編碼之致病性蛋白質(或其抗原片段或抗原決定基)，藉此在個體體內引發針對病原體之免疫。In specific embodiments, the therapeutic effective load comprises a vaccine composition as described herein (e.g., a gene vaccine). In some embodiments, the therapeutic effective load comprises a compound capable of inducing immunity against one or more target diseases or diseases. In some embodiments, the target disease is associated with or caused by a pathogen infection, such as a coronavirus (e.g., 2019-nCoV), influenza virus, measles virus, human papillomavirus (HPV), rabies virus, meningitis virus, pertussis virus, tetanus virus, plague virus, hepatitis virus, and tuberculosis virus. In some embodiments, the therapeutic effective load comprises a nucleic acid sequence (e.g., mRNA) encoding a pathogenic protein or an antigenic fragment or antigenic determinant thereof that is unique to the pathogen. When administered to a vaccinated individual, a vaccine allows the expression of the encoded pathogenic protein (or its antigenic fragment or antigenic determinant), thereby inducing immunity against the pathogen in the individual.
在一些實施例中，目標疾患與細胞之贅生性生長相關或由細胞之贅生性生長引起(諸如癌症)。在一些實施例中，治療性有效負載包含編碼癌症特有之腫瘤相關抗原(TAA)或其抗原片段或抗原決定基之核酸序列(例如mRNA)。疫苗在投與經疫苗接種之個體後，允許表現所編碼之TAA(或其抗原片段或抗原決定基)，藉此在個體體內引發針對表現TAA之贅生性細胞之免疫。In some embodiments, the target disease is associated with or caused by the mesenchymal growth of cells (such as cancer). In some embodiments, the therapeutic effective load comprises a nucleic acid sequence (such as mRNA) encoding a cancer-specific tumor-associated antigen (TAA) or an antigenic fragment or antigenic determinant thereof. After administration to the vaccinated individual, the vaccine allows the expression of the encoded TAA (or its antigenic fragment or antigenic determinant), thereby inducing immunity against mesenchymal cells expressing the TAA in the individual.
5’-帽結構5'-cap structure
不受理論束縛，預期聚核苷酸之5’-帽結構參與核輸出並增加聚核苷酸穩定性，且結合mRNA帽結合蛋白(CBP)，CBP負責細胞中之聚核苷酸穩定性，且經由CBP與poly-A結合蛋白締合形成成熟環狀mRNA物質來引起轉譯能力。5’-帽結構進一步有助於mRNA剪接期間5’-近端內含子之移除。因此，在一些實施例中，本揭示案之核酸分子包含5’-帽結構。Without being bound by theory, the 5'-cap structure of the polynucleotide is expected to participate in nuclear export and increase polynucleotide stability, and bind to the mRNA cap binding protein (CBP), which is responsible for polynucleotide stability in cells and induces translational competence through the formation of mature circular mRNA species by the binding of CBP and poly-A binding protein. The 5'-cap structure further facilitates the removal of 5'-proximal introns during mRNA splicing. Therefore, in some embodiments, the nucleic acid molecules of the present disclosure include a 5'-cap structure.
核酸分子可在5’端經細胞內源性轉錄機構戴帽，藉此在聚核苷酸之末端鳥苷帽殘基與5’末端轉錄之有義核苷酸之間產生5’-ppp-5’-三磷酸鍵聯。接著，此5’-鳥苷酸帽可經甲基化以產生N7-甲基-鳥苷酸殘基。聚核苷酸5’端之末端及/或末端前(anteterminal)轉錄之核苷酸之核糖亦可視情況經2’-O-甲基化。經由鳥苷酸帽結構水解及裂解進行之5’-脫帽可靶向核酸分子，例如mRNA分子以進行降解。Nucleic acid molecules can be capped at the 5' end by the cell's endogenous transcriptional machinery, thereby generating a 5'-ppp-5'-triphosphate linkage between the terminal guanosine cap residue of the polynucleotide and the sense nucleotide transcribed at the 5' end. This 5'-guanylate cap can then be methylated to generate an N7-methyl-guanylate residue. The ribose sugars of the terminal and/or anteterminal transcribed nucleotides at the 5' end of the polynucleotide can also be 2'-O-methylated as appropriate. 5'-Decapping via hydrolysis and cleavage of the guanylate cap structure can target nucleic acid molecules, such as mRNA molecules, for degradation.
在一些實施例中，本揭示案之核酸分子包含對由內源過程產生之天然5'-帽結構之一或多個改變。不受理論束縛，對5’-帽之修飾可增加聚核苷酸之穩定性，增加聚核苷酸之半衰期，且可增加聚核苷酸之轉譯效率。In some embodiments, the nucleic acid molecules of the present disclosure comprise one or more alterations to a native 5'-cap structure generated by an endogenous process. Without being bound by theory, modifications to the 5'-cap can increase the stability of the polynucleotide, increase the half-life of the polynucleotide, and can increase the translation efficiency of the polynucleotide.
對天然5’-帽結構之實例性改變包括產生不可水解之帽結構，以防止脫帽，且藉此增加聚核苷酸之半衰期。在一些實施例中，由於帽結構水解需要裂解5’-ppp-5’磷酸二酯鍵聯，因此在一些實施例中，可在戴帽反應期間使用經修飾之核苷酸。舉例而言，在一些實施例中，可根據製造商之說明書，將來自New England Biolabs(Ipswich,Mass.)之牛痘病毒戴帽酶(Vaccinia Capping Enzyme)用於α-硫代鳥苷核苷酸以在5’-ppp-5’帽中產生硫代磷酸酯鍵聯。可使用額外經修飾之鳥苷核苷酸，例如α-甲基膦酸及硒代磷酸核苷酸。Exemplary changes to the natural 5'-cap structure include creating a non-hydrolyzable cap structure to prevent decapping and thereby increase the half-life of the polynucleotide. In some embodiments, because hydrolysis of the cap structure requires cleavage of the 5'-ppp-5' phosphodiester linkage, in some embodiments, modified nucleotides can be used during the capping reaction. For example, in some embodiments, Vaccinia Capping Enzyme from New England Biolabs (Ipswich, Mass.) can be used with α-thioguanosine nucleotides to generate phosphorothioate linkages in the 5'-ppp-5' cap according to the manufacturer's instructions. Additional modified guanosine nucleotides can be used, such as α-methylphosphonic acid and selenophosphate nucleotides.
對於天然5’-帽結構之額外實例性改變亦包括在戴帽之鳥苷三磷酸(GTP)之2’位及/或3’位之修飾、糖環氧(產生碳環之氧)替代為亞甲基部分(CH2)、在帽結構之三磷酸橋部分處之修飾或在核鹼基(G)部分處之修飾。Additional exemplary changes to the native 5'-cap structure also include modifications at the 2' and/or 3' position of the capping guanosine triphosphate (GTP), replacement of the sugar ring oxygen (oxygen of the carbocyclic ring) with a methylene moiety (CH2 ), modifications at the triphosphate bridge portion of the cap structure, or modifications at the nucleobase (G) portion.
對於天然5’-帽結構之額外實例性改變包括但不限於聚核苷酸5’-末端及/或5’-末端前核苷酸之核糖在糖2’-羥基上之2’-O-甲基化(如上所述)。可使用多個不同之5’-帽結構產生聚核苷酸(諸如mRNA分子)之5’-帽。可與本揭示案結合使用之額外實例性5’-帽結構進一步包括國際專利公開案第WO2008127688號、第WO 2008016473號及第WO 2011015347號中闡述之彼等5’-帽結構，各案之全部內容以引用之方式併入本文中。Additional exemplary changes to the natural 5'-cap structure include, but are not limited to, 2'-O-methylation of the ribose sugar 2'-hydroxyl group of the 5'-terminus and/or the nucleotide preceding the 5'-terminus of the polynucleotide (as described above). A variety of different 5'-cap structures can be used to generate the 5'-cap of a polynucleotide (such as an mRNA molecule). Additional exemplary 5'-cap structures that can be used in conjunction with the present disclosure further include those 5'-cap structures described in International Patent Publications Nos. WO2008127688, WO 2008016473, and WO 2011015347, the entire contents of each of which are incorporated herein by reference.
在各個實施例中，5’-末端帽可包括帽類似物。帽類似物在本文中又稱為合成帽類似物、化學帽、化學帽類似物、或者結構或功能性帽類似物，其化學結構不同於天然(即，內源性、野生型或生理性)5’-帽，同時保留帽功能。帽類似物可按化學方式(即，非酶方式)或酶方式合成及/或連接至聚核苷酸。In various embodiments, the 5'-terminal cap may include a cap analog. Cap analogs are also referred to herein as synthetic cap analogs, chemical caps, chemical cap analogs, or structural or functional cap analogs, which have a chemical structure different from that of a natural (i.e., endogenous, wild-type, or physiological) 5'-cap while retaining cap function. Cap analogs can be synthesized and/or attached to polynucleotides chemically (i.e., non-enzymatically) or enzymatically.
舉例而言，抗反向帽類似物(ARCA)帽含有經5’-5’-三磷酸酯基團連接之兩個鳥苷，其中一個鳥苷含有N7-甲基以及3’-O-甲基(即，N7,3’-O-二甲基-鳥苷-5’-三磷酸-5’-鳥苷，即m7G-3’mppp-G，其可等效地稱為3’O-Me-m7G(5’)ppp(5’)G)。另一未改變之鳥苷之3’-O原子與戴帽聚核苷酸(例如mRNA)之5’-末端核苷酸連接。N7-及3’-O-甲基化之鳥苷提供戴帽聚核苷酸(例如mRNA)之末端部分。另一實例性之帽結構為mCAP，其類似於ARCA，但在鳥苷上具有2’-O-甲基(即，N7,2’-O-二甲基-鳥苷-5’-三磷酸-5’-鳥苷，即m7Gm-ppp-G)。For example, the anti-reverse cap analog (ARCA) cap contains two guanosines linked via a 5'-5'-triphosphate group, one of which contains an N7-methyl group as well as a 3'-O-methyl group (i.e., N7,3'-O-dimethyl-guanosine-5'-triphosphate-5'-guanosine, i.e.,m7G -3'mppp-G, which can be equivalently referred to as 3'O-Me-m7G(5')ppp(5')G). The 3'-O atom of the other unchanged guanosine is linked to the 5'-terminal nucleotide of the capped polynucleotide (e.g., mRNA). The N7- and 3'-O-methylated guanosine provides the terminal portion of the capped polynucleotide (e.g., mRNA). Another exemplary cap structure is mCAP, which is similar to ARCA but has a 2'-O-methyl group on guanosine (ie, N7,2'-O-dimethyl-guanosine-5'-triphosphate-5'-guanosine, ie,m7Gm -ppp-G).
在一些實施例中，帽類似物可為二核苷酸帽類似物。作為非限制性實例，二核苷酸帽類似物可在不同磷酸酯位置處用硼烷磷酸酯基(boranophosphate)或硒代磷酸酯基(phophoroselenoate)進行修飾，諸如美國專利第8,519,110號中所述之二核苷酸帽類似物，該案之全部內容以全文引用之方式併入本文中。In some embodiments, the cap analog may be a dinucleotide cap analog. As a non-limiting example, the dinucleotide cap analog may be modified with boranophosphate or phophoroselenoate at different phosphate positions, such as the dinucleotide cap analog described in U.S. Patent No. 8,519,110, the entire contents of which are incorporated herein by reference in their entirety.
在一些實施例中，帽類似物可為此項技術中已知及/或本文所述之N7-(4-氯苯氧基乙基)取代之二核苷酸帽類似物。N7-(4-氯苯氧基乙基)取代之二核苷酸帽類似物之非限制性實例包括N7-(4-氯苯氧基乙基)-G(5’)ppp(5’)G及N7-(4-氯苯氧基乙基)-m3’-OG(5’)ppp(5’)G帽類似物(參見例如Kore等人，Bioorganic & Medicinal Chemistry 2013 21：4570-4574中所述之各種帽類似物及合成帽類似物之方法；該文獻之全部內容以引用之方式併入本文中)。在其他實施例中，可與本揭示案之核酸分子結合使用之帽類似物係4-氯/溴苯氧基乙基類似物。In some embodiments, the cap analog may be a N7-(4-chlorophenoxyethyl) substituted dinucleotide cap analog known in the art and/or described herein. Non-limiting examples of N7-(4-chlorophenoxyethyl) substituted dinucleotide cap analogs include N7-(4-chlorophenoxyethyl)-G(5')ppp(5')G and N7-(4-chlorophenoxyethyl)-m3'-OG(5')ppp(5')G cap analogs (see, e.g., Kore et al., Bioorganic & Medicinal Chemistry 2013 21:4570-4574 for various cap analogs and methods for synthesizing cap analogs; the entire contents of which are incorporated herein by reference). In other embodiments, the cap analog that can be used in conjunction with the nucleic acid molecules of the present disclosure is a 4-chloro/bromophenoxyethyl analog.
在各個實施例中，帽類似物可包括鳥苷類似物。有用之鳥苷類似物包括但不限於肌苷、N1-甲基-鳥苷、2’-氟-鳥苷、7-去氮-鳥苷、8-側氧基-鳥苷、2-胺基-鳥苷、LNA-鳥苷及2-疊氮基-鳥苷。In various embodiments, the cap analog may include a guanosine analog. Useful guanosine analogs include, but are not limited to, inosine, N1-methyl-guanosine, 2'-fluoro-guanosine, 7-deaza-guanosine, 8-oxo-guanosine, 2-amino-guanosine, LNA-guanosine, and 2-azido-guanosine.
不受理論束縛，預期儘管帽類似物允許在活體外轉錄反應中同時進行聚核苷酸之戴帽，但高達20%之轉錄物仍未戴帽。此情況以及帽類似物與細胞內源轉錄機構產生之聚核苷酸之天然5’-帽結構之結構差異可能導致轉譯能力減弱及細胞穩定性降低。Without being bound by theory, it is expected that although cap analogs allow simultaneous capping of polynucleotides in in vitro transcription reactions, up to 20% of transcripts remain uncapped. This and the structural differences between the natural 5'-cap structure of polynucleotides produced by cap analogs and the endogenous transcription machinery of the cell may lead to reduced translational capacity and decreased cell stability.
因此，在一些實施例中，本揭示案之核酸分子亦可使用酶在轉錄後戴帽，以便產生更真實(authentic)之5’-帽結構。如本文所使用，片語「更真實」係指一種特徵在結構上或功能上密切反映或模仿內源或野生型特徵。亦即，與先前技術之合成特徵或類似物相比，「更真實」之特徵更好地代表內源性、野生型、天然或生理細胞功能及/或結構，或者其在一或多個態樣勝過對應內源性、野生型、天然或生理特徵。可與本揭示案之核酸分子結合使用之更真實之5’-帽結構之非限制性實例為相較於此項技術中已知之合成5’-帽結構(或相較於野生型、天然或生理性5’-帽結構)，尤其具有增強之與帽結合蛋白之結合、增加之半衰期、降低之對5’-核酸內切酶之敏感性及/或減少之5’-脫帽之結構。舉例而言，在一些實施例中，重組牛痘病毒戴帽酶及重組2’-O-甲基轉移酶可在聚核苷酸之5’-末端核苷酸與鳥苷帽核苷酸之間產生經典之5’-5’-三磷酸酯鍵聯，其中帽鳥苷含有N7-甲基化，且聚核苷酸之5’-末端核苷酸含有2’-O-甲基。此一結構稱為帽1結構。與例如此項技術中已知之其他5’帽類似物結構相比，此帽引起更高之轉譯能力、細胞穩定性及減少之細胞促炎性細胞介素之活化。其他實例性帽結構包括7mG(5’)ppp(5’)N,pN2p(帽0)、7mG(5’)ppp(5’)NlmpNp(帽1)、7mG(5’)-ppp(5’)NlmpN2mp(帽2)及m(7)Gpppm(3)(6,6,2’)Apm(2’)Apm(2’)Cpm(2)(3,2’)Up(帽4)。Therefore, in some embodiments, the nucleic acid molecules of the present disclosure may also be capped after transcription using enzymes to produce a more authentic 5'-cap structure. As used herein, the phrase "more authentic" refers to a feature that closely reflects or mimics an endogenous or wild-type feature in structure or function. That is, a "more authentic" feature better represents endogenous, wild-type, natural or physiological cellular function and/or structure than a synthetic feature or analog of the prior art, or it outperforms the corresponding endogenous, wild-type, natural or physiological feature in one or more aspects. Non-limiting examples of more realistic 5'-cap structures that can be used in conjunction with the nucleic acid molecules of the present disclosure are structures that have enhanced binding to cap-binding proteins, increased half-life, reduced sensitivity to 5'-endonucleases, and/or reduced 5'-decapping, compared to synthetic 5'-cap structures known in the art (or compared to wild-type, natural or physiological 5'-cap structures). For example, in some embodiments, recombinant vaccinia virus capping enzymes and recombinant 2'-O-methyltransferases can generate a classic 5'-5'-triphosphate linkage between the 5'-terminal nucleotide of a polynucleotide and a guanosine cap nucleotide, wherein the guanosine cap contains an N7-methylation and the 5'-terminal nucleotide of the polynucleotide contains a 2'-O-methyl group. This structure is referred to as the Cap 1 structure. This cap induces higher translational capacity, cell stability and reduced activation of cellular pro-inflammatory cytokines compared to other 5' cap analog structures known in the art, for example. Other exemplary cap structures include 7mG(5')ppp(5')N,pN2p (cap 0), 7mG(5')ppp(5')NlmpNp (cap 1), 7mG(5')-ppp(5')NlmpN2mp (cap 2) and m(7)Gpppm(3)(6,6,2')Apm(2')Apm(2')Cpm(2)(3,2')Up (cap 4).
不受理論束縛，預期本揭示案之核酸分子可在轉錄後戴帽，且由於此方法較為高效，因此幾乎100%之核酸分子可經戴帽。Without being bound by theory, it is expected that the nucleic acid molecules of the present disclosure can be capped after transcription, and since this method is relatively efficient, almost 100% of the nucleic acid molecules can be capped.
非轉譯區(UTR)Untranslated region (UTR)
在一些實施例中，本揭示案之核酸分子包含一或多個非轉譯區(UTR)。在一些實施例中，UTR位於核酸分子中編碼區之上游，且被稱為5’-UTR。在一些實施例中，UTR位於核酸分子中編碼區之下游，且被稱為3’-UTR。UTR之序列可與核酸分子中所發現之編碼區之序列同源或異源。多個UTR可包括在核酸分子中，且可具有相同或不同之序列及/或基因起源。根據本揭示案，核酸分子中UTR之任何部分(包括沒有任何部分)可經密碼子最佳化，且任何部分可在密碼子最佳化之前及/或之後獨立地含有一或多個不同之結構或化學修飾。In some embodiments, the nucleic acid molecules of the present disclosure include one or more untranslated regions (UTRs). In some embodiments, the UTR is located upstream of the coding region in the nucleic acid molecule and is referred to as the 5'-UTR. In some embodiments, the UTR is located downstream of the coding region in the nucleic acid molecule and is referred to as the 3'-UTR. The sequence of the UTR may be homologous or heterologous to the sequence of the coding region found in the nucleic acid molecule. Multiple UTRs may be included in a nucleic acid molecule and may have the same or different sequences and/or genetic origins. According to the present disclosure, any portion of the UTR in the nucleic acid molecule (including no portion) may be codon optimized, and any portion may independently contain one or more different structural or chemical modifications before and/or after codon optimization.
在一些實施例中，本揭示案之核酸分子(例如mRNA)包含相對於彼此為同源之UTR及編碼區。在其他實施例中，本揭示案之核酸分子(例如mRNA)包含相對於彼此為異源之UTR及編碼區。在一些實施例中，為了監測UTR序列之活性，可在活體外(例如細胞或組織培養物)或在活體內(例如向個體)投與包含UTR及可偵測探針之編碼序列之核酸分子，且可使用此項技術中已知之方法量測UTR序列之作用(例如調節表現水準、編碼產物之細胞定位或編碼產物之半衰期)。In some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure comprise UTRs and coding regions that are homologous to each other. In other embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure comprise UTRs and coding regions that are heterologous to each other. In some embodiments, to monitor the activity of the UTR sequence, a nucleic acid molecule comprising a UTR and a coding sequence that can be detected by a probe can be administered in vitro (e.g., in cell or tissue culture) or in vivo (e.g., to an individual), and the effect of the UTR sequence (e.g., regulating expression levels, cellular localization of the encoded product, or half-life of the encoded product) can be measured using methods known in the art.
在一些實施例中，本揭示案之核酸分子(例如mRNA)之UTR包含至少一個轉譯強化子元件(TEE)，該TEE起到增加由該核酸分子產生之多肽或蛋白質之量的作用。在一些實施例中，TEE位於核酸分子之5’-UTR中。在其他實施例中，TEE位於核酸分子之3’-UTR處。在其他實施例中，至少兩個TEE分別位於核酸分子之5’-UTR及3’-UTR處。在一些實施例中，本揭示案之核酸分子(例如mRNA)可包含一或多個拷貝之TEE序列或包含多於一個不同之TEE序列。在一些實施例中，存在於本揭示案之核酸分子中之不同TEE序列可相對於彼此為同源的或異源的。In some embodiments, the UTR of the nucleic acid molecule (e.g., mRNA) of the present disclosure comprises at least one translation enhancer element (TEE), which acts to increase the amount of polypeptide or protein produced by the nucleic acid molecule. In some embodiments, the TEE is located in the 5'-UTR of the nucleic acid molecule. In other embodiments, the TEE is located at the 3'-UTR of the nucleic acid molecule. In other embodiments, at least two TEEs are located at the 5'-UTR and 3'-UTR of the nucleic acid molecule, respectively. In some embodiments, the nucleic acid molecule (e.g., mRNA) of the present disclosure may comprise one or more copies of a TEE sequence or comprise more than one different TEE sequence. In some embodiments, different TEE sequences present in the nucleic acid molecule of the present disclosure may be homologous or heterologous to each other.
各種TEE序列係此項技術中已知的，且可與本揭示案結合使用。舉例而言，在一些實施例中，TEE可為內部核糖體進入位點(IRES)、HCV-IRES或IRES元件。Chappell等人，Proc.Natl.Acad.Sci.USA 101：9590-9594,2004；Zhou等人，Proc.Natl.Acad.Sci.102：6273-6278,2005。可與本揭示案結合使用之額外內部核糖體進入位點(IRES)包括但不限於美國專利第7,468,275號、美國專利公開案第2007/0048776號及美國專利公開案第2011/0124100號，以及國際專利公開案第WO2007/025008號及國際專利公開案第WO2001/055369號中所述之IRES，各案之內容以全文引用之方式併入本文中。在一些實施例中，TEE可為Wellensiek等人，Genome-wide profiling of human cap-independent translation-enhancing elements,Nature Methods,2013年8月；10(8)：747-750之補充表1及補充表2中所述之TEE；每篇文獻之內容以全文引用之方式併入本文中。Various TEE sequences are known in the art and can be used in conjunction with the present disclosure. For example, in some embodiments, the TEE can be an internal ribosome entry site (IRES), HCV-IRES, or an IRES element. Chappell et al.,Proc. Natl. Acad. Sci. USA 101:9590-9594, 2004; Zhou et al.,Proc. Natl. Acad. Sci. 102:6273-6278, 2005. Additional internal ribosome entry sites (IRES) that may be used in conjunction with the present disclosure include, but are not limited to, those described in U.S. Patent No. 7,468,275, U.S. Patent Publication No. 2007/0048776, and U.S. Patent Publication No. 2011/0124100, and International Patent Publication No. WO2007/025008 and International Patent Publication No. WO2001/055369, the contents of each of which are incorporated herein by reference in their entirety. In some embodiments, the TEE can be the TEE described in Supplementary Table 1 and Supplementary Table 2 of Wellensiek et al., Genome-wide profiling of human cap-independent translation-enhancing elements,Nature Methods , 2013 Aug; 10(8): 747-750; the contents of each reference are incorporated herein by reference in their entirety.
可與本揭示案結合使用之額外實例性TEE包括但不限於美國專利第6,310,197號、美國專利第6,849,405號、美國專利第7,456,273號、美國專利第7,183,395號、美國專利公開案第2009/0226470號、美國專利公開案第2013/0177581號、美國專利公開案第2007/0048776號、美國專利公開案第2011/0124100號、美國專利公開案第2009/0093049號、國際專利公開案第WO2009/075886號、國際專利公開案第WO2012/009644號及國際專利公開案第WO1999/024595號、國際專利公開案第WO2007/025008號、國際專利公開案第WO2001/055371號、歐洲專利第2610341號、歐洲專利第2610340號中所述之TEE序列，各案之內容以全文引用之方式併入本文中。Additional exemplary TEEs that may be used in conjunction with the present disclosure include, but are not limited to, U.S. Patent No. 6,310,197, U.S. Patent No. 6,849,405, U.S. Patent No. 7,456,273, U.S. Patent No. 7,183,395, U.S. Patent Publication No. 2009/0226470, U.S. Patent Publication No. 2013/0177581, U.S. Patent Publication No. 2007/0048776, U.S. Patent Publication No. 2011/0124100, U.S. Patent Publication No. 2009/0093049, International Patent Publication No. WO2009/075886, International Patent Publication No. WO2012/009644, International Patent Publication No. WO1999/024595, International Patent Publication No. WO2007/025008, International Patent Publication No. WO2001/055371, European Patent No. 2610341, European Patent No. 2610340, the contents of each case are incorporated herein by reference in their entirety.
在各個實施例中，本揭示案之核酸分子(例如mRNA)包含至少一個UTR，其包含至少1個、至少2個、至少3個、至少4個、至少5個、至少6個、至少7個、至少8個、至少9個、至少10個、至少11個、至少12個、至少13個、至少14個、至少15個、至少16個、至少17個、至少18個、至少19個、至少20個、至少21個、至少22個、至少23個、至少24個、至少25個、至少30個、至少35個、至少40個、至少45個、至少50個、至少55個或超過60個TEE序列。在一些實施例中，核酸分子UTR中之TEE序列係相同TEE序列之拷貝。在其他實施例中，核酸分子UTR中之至少兩個TEE序列具有不同之TEE序列。在一些實施例中，多個不同之TEE序列以一或多種重複模式佈置於核酸分子之UTR區中。僅出於說明之目的，重複模式可為例如ABABAB、AABBAABBAABB、ABCABCABC及諸如此類，其中在此等實例性模式中，每個大寫字母(A、B或C)代表不同之TEE序列。在一些實施例中，在核酸分子之UTR中，至少兩個TEE序列彼此為連續的(即，其間無間隔子序列)。在其他實施例中，至少兩個TEE序列藉由間隔子序列隔開。在一些實施例中，UTR可包含TEE序列-間隔子序列模組，該模組在UTR中重複至少一次、至少兩次、至少3次、至少4次、至少5次、至少6次、至少7次、至少8次、至少9次或更多次。在本段中闡述之任何實施例中，UTR可為核酸分子之5’-UTR、3’-UTR或5’-UTR及3’-UTR二者。In various embodiments, the nucleic acid molecule (e.g., mRNA) of the present disclosure comprises at least one UTR comprising at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 55, or more than 60 TEE sequences. In some embodiments, the TEE sequences in the UTR of the nucleic acid molecule are copies of the same TEE sequence. In other embodiments, at least two TEE sequences in the UTR of the nucleic acid molecule have different TEE sequences. In some embodiments, a plurality of different TEE sequences are arranged in one or more repetitive patterns in the UTR region of a nucleic acid molecule. For purposes of illustration only, the repetitive pattern may be, for example, ABABAB, AABBAABBAAB, ABCABCABC, and the like, wherein in such exemplary patterns, each capital letter (A, B, or C) represents a different TEE sequence. In some embodiments, in the UTR of a nucleic acid molecule, at least two TEE sequences are continuous with each other (i.e., there is no spacer sequence therebetween). In other embodiments, at least two TEE sequences are separated by a spacer sequence. In some embodiments, the UTR may include a TEE sequence-spacer sequence module that is repeated in the UTR at least once, at least twice, at least 3 times, at least 4 times, at least 5 times, at least 6 times, at least 7 times, at least 8 times, at least 9 times or more. In any of the embodiments described in this paragraph, the UTR may be the 5'-UTR, the 3'-UTR, or both the 5'-UTR and the 3'-UTR of the nucleic acid molecule.
在一些實施例中，本揭示案之核酸分子(例如mRNA)之UTR包含至少一個轉譯抑制元件，該元件起到減少由該核酸分子產生之多肽或蛋白質之量的作用。在一些實施例中，核酸分子之UTR包含經一或多個微小RNA識別之一或多個miR序列或其片段(例如miR種子序列(seed sequence))。在一些實施例中，核酸分子之UTR包含下調核酸分子之轉譯活性之一或多個莖-環結構。用於抑制與核酸分子相關之轉譯活性之其他機制係此項技術中已知的。在本段中闡述之任何實施例中，UTR可為核酸分子之5’-UTR、3’-UTR或5’-UTR及3’-UTR二者。In some embodiments, the UTR of a nucleic acid molecule (e.g., mRNA) of the present disclosure comprises at least one translational repressor element that acts to reduce the amount of a polypeptide or protein produced by the nucleic acid molecule. In some embodiments, the UTR of a nucleic acid molecule comprises one or more miR sequences or fragments thereof (e.g., miR seed sequences) recognized by one or more microRNAs. In some embodiments, the UTR of a nucleic acid molecule comprises one or more stem-loop structures that downregulate the translational activity of the nucleic acid molecule. Other mechanisms for inhibiting translational activity associated with nucleic acid molecules are known in the art. In any of the embodiments described in this paragraph, the UTR may be the 5'-UTR, the 3'-UTR, or both the 5'-UTR and the 3'-UTR of the nucleic acid molecule.
聚腺苷酸化(Poly-A)區Polyadenylation (Poly-A) region
在天然RNA加工過程中，通常將長鏈腺苷核苷酸(poly-A區)添加至信使RNA(mRNA)分子中以增加分子之穩定性。轉錄後，立即將轉錄物之3’-端裂解以釋放3’-羥基。接著，poly-A聚合酶將一連串腺苷核苷酸添加至RNA中。該過程稱為聚腺苷酸化，添加一個長度在100與250個殘基之間之poly-A區。不受理論束縛，預期poly-A區可賦予本揭示案之核酸分子多個優點。During natural RNA processing, long chains of adenosine nucleotides (poly-A regions) are often added to messenger RNA (mRNA) molecules to increase the stability of the molecule. Immediately after transcription, the 3'-end of the transcript is cleaved to release the 3'-hydroxyl group. Next, poly-A polymerase adds a string of adenosine nucleotides to the RNA. This process is called polyadenylation, and a poly-A region with a length of between 100 and 250 residues is added. Without being bound by theory, it is expected that the poly-A region can confer a number of advantages to the nucleic acid molecules of the present disclosure.
因此，在一些實施例中，本揭示案之核酸分子(例如mRNA)包含聚腺苷酸化信號。在一些實施例中，本揭示案之核酸分子(例如mRNA)包含一或多個聚腺苷酸化(poly-A)區。在一些實施例中，poly-A區完全由腺嘌呤核苷酸或其功能性類似物構成。在一些實施例中，核酸分子在其3’端包含至少一個poly-A區。在一些實施例中，核酸分子在其5’端包含至少一個poly-A區。在一些實施例中，核酸分子在其5’端包含至少一個poly-A區且在其3’端包含至少一個poly-A區。Thus, in some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure comprise a polyadenylation signal. In some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure comprise one or more polyadenylation (poly-A) regions. In some embodiments, the poly-A region is entirely composed of adenine nucleotides or functional analogs thereof. In some embodiments, the nucleic acid molecule comprises at least one poly-A region at its 3' end. In some embodiments, the nucleic acid molecule comprises at least one poly-A region at its 5' end. In some embodiments, the nucleic acid molecule comprises at least one poly-A region at its 5' end and at least one poly-A region at its 3' end.
根據本揭示案，在不同實施例中，poly-A區可具有不同長度。具體而言，在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少30個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少35個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少40個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少45個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少50個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少55個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少60個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少65個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少70個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少75個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少80個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少85個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少90個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少95個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少100個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少110個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少120個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少130個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少140個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少150個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少160個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少170個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少180個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少190個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少200個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少225個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少250個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少275個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少300個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少350個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少400個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少450個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少500個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少600個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少700個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少800個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少900個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1000個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1100個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1200個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1300個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1400個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1500個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1600個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1700個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1800個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少1900個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少2000個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少2250個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少2500個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少2750個核苷酸。在一些實施例中，本揭示案之核酸分子之poly-A區之長度為至少3000個核苷酸。According to the present disclosure, in different embodiments, the poly-A region may have different lengths. Specifically, in some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 30 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 35 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 40 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 45 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 50 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 55 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 60 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 65 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 70 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 75 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 80 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 85 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 90 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 95 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 100 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 110 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 120 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 130 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 140 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 150 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 160 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 170 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 180 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 190 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 200 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 225 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 250 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 275 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 300 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 350 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 400 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 450 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 500 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 600 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 700 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 800 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 900 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1000 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1100 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1200 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1300 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1400 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1500 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1600 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1700 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1800 nucleotides. In some embodiments, the length of the poly-A region of the nucleic acid molecule of the present disclosure is at least 1900 nucleotides. In some embodiments, the poly-A region of the nucleic acid molecule of the present disclosure is at least 2000 nucleotides in length. In some embodiments, the poly-A region of the nucleic acid molecule of the present disclosure is at least 2250 nucleotides in length. In some embodiments, the poly-A region of the nucleic acid molecule of the present disclosure is at least 2500 nucleotides in length. In some embodiments, the poly-A region of the nucleic acid molecule of the present disclosure is at least 2750 nucleotides in length. In some embodiments, the poly-A region of the nucleic acid molecule of the present disclosure is at least 3000 nucleotides in length.
在一些實施例中，核酸分子中poly-A區之長度可基於核酸分子之總長度或其部分(諸如核酸分子之編碼區之長度或開放閱讀框之長度等)來選擇。舉例而言，在一些實施例中，poly-A區佔含有poly-A區之核酸分子之總長度之約5%、10%、15%、20%、25%、30%、35%、40%、45%、50%、55%、60%、65%、70%、75%、80%、85%、90%、95%或更高百分比。In some embodiments, the length of the poly-A region in a nucleic acid molecule can be selected based on the total length of the nucleic acid molecule or a portion thereof (such as the length of the coding region of the nucleic acid molecule or the length of the open reading frame, etc.). For example, in some embodiments, the poly-A region accounts for about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or more of the total length of the nucleic acid molecule containing the poly-A region.
不受理論束縛，預期某些RNA結合蛋白可結合至位於mRNA分子3’端之poly-A區。此等poly-A結合蛋白(PABP)可調節mRNA表現，諸如與細胞中之轉譯起始機構相互作用及/或保護3’-poly-A尾免於降解。因此，在一些實施例中，本揭示案之核酸分子(例如mRNA)包含poly-A結合蛋白(PABP)之至少一個結合位點。在其他實施例中，在將核酸分子裝載至遞送媒劑(例如脂質奈米顆粒)中之前，使其與PABP形成結合物或複合物。Without being bound by theory, it is expected that certain RNA binding proteins can bind to the poly-A region located at the 3' end of the mRNA molecule. These poly-A binding proteins (PABPs) can regulate mRNA expression, such as interacting with the translation initiation machinery in the cell and/or protecting the 3'-poly-A tail from degradation. Therefore, in some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure include at least one binding site for a poly-A binding protein (PABP). In other embodiments, the nucleic acid molecule is bound to the PABP to form a conjugate or complex before being loaded into a delivery vehicle (e.g., lipid nanoparticles).
在一些實施例中，本揭示案之核酸分子(例如mRNA)包含poly-A-G四聯體。G四聯體係可由DNA及RNA中富含G之序列形成之氫鍵合之四個鳥苷核苷酸之環狀陣列。在此實施例中，G四聯體併入poly-A區之一端。可分析所得聚核苷酸(例如mRNA)之穩定性、蛋白質產量及其他參數，包括在不同時間點之半衰期。已發現，poly-A-G四聯體結構之蛋白質產量等於僅使用含120個核苷酸之poly-A區所觀察到之蛋白質產量之至少75%。In some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure comprise a poly-A-G quadruplex. A G-quartet is a hydrogen-bonded circular array of four guanosine nucleotides that can be formed from G-rich sequences in DNA and RNA. In this embodiment, the G-quartet is incorporated into one end of the poly-A region. The resulting polynucleotide (e.g., mRNA) can be analyzed for stability, protein yield, and other parameters, including half-life at different time points. It has been found that the protein yield of the poly-A-G quadruplex structure is at least 75% of the protein yield observed using only a poly-A region containing 120 nucleotides.
在一些實施例中，本揭示案之核酸分子(例如mRNA)可包括poly-A區且可藉由添加3’-穩定區來穩定。在一些實施例中，可用於使核酸分子(例如mRNA)穩定之3’-穩定區包括國際專利公開案第WO2013/103659號中所述之poly-A或poly-A-G四聯體結構，該公開案之內容以全文引用之方式併入本文中。In some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure may include a poly-A region and may be stabilized by adding a 3'-stabilizing region. In some embodiments, the 3'-stabilizing region that can be used to stabilize the nucleic acid molecules (e.g., mRNA) includes a poly-A or poly-A-G quadruplex structure described in International Patent Publication No. WO2013/103659, the contents of which are incorporated herein by reference in their entirety.
在其他實施例中，可與本揭示案之核酸分子結合使用之3’-穩定區包括鏈終止核苷，諸如但不限於3’-去氧腺苷(蟲草素(cordycepin))；3’-去氧尿苷；3’-去氧胞嘧啶；3’-去氧鳥苷；3’-去氧胸腺嘧啶；2’,3’-雙去氧核苷，諸如2’,3’-雙去氧腺苷、2’,3’-雙去氧尿苷、2’,3’-雙去氧胞嘧啶、2’,3’-雙去氧鳥苷、2’,3’-雙去氧胸腺嘧啶；2’-去氧核苷；或O-甲基核苷；3’-去氧核苷；2’,3’-雙去氧核苷；3’-O-甲基核苷；3’-O-乙基核苷；3’-阿拉伯糖苷，以及此項技術中已知及/或本文所述之其他替代性核苷。In other embodiments, the 3'-stabilizing region that can be used in conjunction with the nucleic acid molecules of the present disclosure includes a chain-terminating nucleoside, such as but not limited to 3'-deoxyadenosine (cordycepin); 3'-deoxyuridine; 3'-deoxycytosine; 3'-deoxyguanosine; 3'-deoxythymidine; 2',3'-dideoxynucleosides, such as 2',3'-dideoxyadenosine, 2',3' -dideoxyuridine, 2’,3’-dideoxycytosine, 2’,3’-dideoxyguanosine, 2’,3’-dideoxythymidine; 2’-deoxynucleoside; or O-methyl nucleoside; 3’-deoxynucleoside; 2’,3’-dideoxynucleoside; 3’-O-methyl nucleoside; 3’-O-ethyl nucleoside; 3’-arabinoside, and other alternative nucleosides known in the art and/or described herein.
二級結構Secondary Structure
不受理論束縛，預期莖-環結構可引導RNA摺疊，保護核酸分子(例如mRNA)之結構穩定性，提供RNA結合蛋白之識別位點，且用作酶反應之受質。舉例而言，miR序列及/或TEE序列之併入將改變莖-環區之形狀，藉此可增加及/或減少轉譯(Kedde等人，A Pumilio-induced RNA structure switch in p27-3’UTR controls miR-221 and miR-222 accessibility.Nat Cell Biol.,2010年10月；12(10)：1014-20，該文獻之內容以全文引用之方式併入本文中)。Without being bound by theory, the stem-loop structure is expected to guide RNA folding, protect the structural stability of nucleic acid molecules (e.g., mRNA), provide recognition sites for RNA binding proteins, and serve as substrates for enzymatic reactions. For example, the incorporation of miR sequences and/or TEE sequences will change the shape of the stem-loop region, thereby increasing and/or decreasing translation (Kedde et al., A Pumilio-induced RNA structure switch in p27-3'UTR controls miR-221 and miR-222 accessibility.Nat Cell Biol. , 2010 Oct;12(10):1014-20, the content of which is incorporated herein by reference in its entirety).
因此，在一些實施例中，本文所述之核酸分子(例如mRNA)或其一部分可呈莖-環結構，諸如但不限於組蛋白莖-環。在一些實施例中，莖-環結構由長度為約25個或約26個核苷酸之莖-環序列形成，諸如但不限於國際專利公開案第WO2013/103659號中所述之結構，該案內容以全文引用之方式併入本文中。莖-環序列之額外實例包括國際專利公開案第WO2012/019780號及國際專利公開案第WO201502667號中所述之序列，各案內容以引用之方式併入本文中。在一些實施例中，莖-環序列包含如本文所述之TEE。在一些實施例中，莖-環序列包含如本文所述之miR序列。在特定實施例中，莖-環序列可包括miR-122種子序列。在特定實施例中，核酸分子包含莖-環序列CAAAGGCTCTTTTCAGAGCCACCA(SEQ ID NO：1)。在其他實施例中，核酸分子包含莖-環序列CAAAGGCUCUUUUCAGAGCCACCA(SEQ ID NO：2)。Thus, in some embodiments, a nucleic acid molecule (e.g., mRNA) described herein or a portion thereof may be in a stem-loop structure, such as, but not limited to, a histone stem-loop. In some embodiments, the stem-loop structure is formed by a stem-loop sequence of about 25 or about 26 nucleotides in length, such as, but not limited to, the structure described in International Patent Publication No. WO2013/103659, the contents of which are incorporated herein by reference in their entirety. Additional examples of stem-loop sequences include sequences described in International Patent Publication No. WO2012/019780 and International Patent Publication No. WO201502667, the contents of each of which are incorporated herein by reference. In some embodiments, the stem-loop sequence comprises a TEE as described herein. In some embodiments, the stem-loop sequence comprises a miR sequence as described herein. In certain embodiments, the stem-loop sequence may include a miR-122 seed sequence. In certain embodiments, the nucleic acid molecule comprises the stem-loop sequence CAAAGGCTCTTTTCAGAGCCACCA (SEQ ID NO: 1). In other embodiments, the nucleic acid molecule comprises the stem-loop sequence CAAAGGCUCUUUUCAGAGCCACCA (SEQ ID NO: 2).
在一些實施例中，本揭示案之核酸分子(例如mRNA)包含位於核酸分子中編碼區上游(在5’端)之莖-環序列。在一些實施例中，莖-環序列位於核酸分子5’-UTR內。在一些實施例中，本揭示案之核酸分子(例如mRNA)包含位於核酸分子中編碼區下游(在3’端)之莖-環序列。在一些實施例中，莖-環序列位於核酸分子3’-UTR內。在一些情況下，核酸分子可含有多於一個莖-環序列。在一些實施例中，核酸分子在5’-UTR中包含至少一個莖-環序列且在3’-UTR中包含至少一個莖-環序列。In some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure comprise a stem-loop sequence located upstream (at the 5' end) of the coding region in the nucleic acid molecule. In some embodiments, the stem-loop sequence is located within the 5'-UTR of the nucleic acid molecule. In some embodiments, the nucleic acid molecules (e.g., mRNA) of the present disclosure comprise a stem-loop sequence located downstream (at the 3' end) of the coding region in the nucleic acid molecule. In some embodiments, the stem-loop sequence is located within the 3'-UTR of the nucleic acid molecule. In some cases, the nucleic acid molecule may contain more than one stem-loop sequence. In some embodiments, the nucleic acid molecule comprises at least one stem-loop sequence in the 5'-UTR and at least one stem-loop sequence in the 3'-UTR.
在一些實施例中，包含莖-環結構之核酸分子進一步包含穩定區。在一些實施例中，穩定區包含至少一個鏈終止核苷，其起到減慢降解並藉此增加核酸分子之半衰期之作用。可與本揭示案之核酸分子結合使用之實例性鏈終止核苷包括但不限於3’-去氧腺苷(蟲草素)；3’-去氧尿苷；3’-去氧胞嘧啶；3’-去氧鳥苷；3’-去氧胸腺嘧啶；2’,3’-雙去氧核苷，諸如2’,3’-雙去氧腺苷、2’,3’-雙去氧尿苷、2’,3’-雙去氧胞嘧啶、2’,3’-雙去氧鳥苷、2’,3’-雙去氧胸腺嘧啶；2’-去氧核苷；或O-甲基核苷；3’-去氧核苷；2’,3’-雙去氧核苷；3’-O-甲基核苷；3’-O-乙基核苷；3’-阿拉伯糖苷，以及此項技術中已知及/或本文所述之其他替代性核苷。在其他實施例中，莖-環結構可藉由改變聚核苷酸之3’-區域來穩定，該改變可防止及/或抑制寡聚(U)之添加(國際專利公開案第WO2013/103659號，其以全文引用之方式併入本文中)。In some embodiments, the nucleic acid molecule comprising a stem-ring structure further comprises a stabilizing region. In some embodiments, the stabilizing region comprises at least one chain-terminating nucleoside, which acts to slow degradation and thereby increase the half-life of the nucleic acid molecule. Exemplary chain-terminating nucleosides that can be used in conjunction with the nucleic acid molecules of the present disclosure include, but are not limited to, 3'-deoxyadenosine (cordycepin); 3'-deoxyuridine; 3'-deoxycytosine; 3'-deoxyguanosine; 3'-deoxythymidine; 2',3'-dideoxynucleosides, such as 2',3'-dideoxyadenosine, 2',3'-dideoxyuridine, 2',3'- Bis-deoxycytosine, 2’,3’-bis-deoxyguanosine, 2’,3’-bis-deoxythymine; 2’-deoxynucleoside; or O-methyl nucleoside; 3’-deoxynucleoside; 2’,3’-bis-deoxynucleoside; 3’-O-methyl nucleoside; 3’-O-ethyl nucleoside; 3’-arabinoside, and other alternative nucleosides known in the art and/or described herein. In other embodiments, the stem-ring structure can be stabilized by altering the 3’-region of the polynucleotide, which can prevent and/or inhibit the addition of oligo(U) (International Patent Publication No. WO2013/103659, which is incorporated herein by reference in its entirety).
在一些實施例中，本揭示案之核酸分子包含至少一個莖-環序列及poly-A區或聚腺苷酸化信號。包含至少一個莖-環序列及poly-A區或聚腺苷酸化信號之聚核苷酸序列之非限制性實例包括國際專利公開案第WO2013/120497號、國際專利公開案第WO2013/120629號、國際專利公開案第WO2013/120500號、國際專利公開案第WO2013/120627號、國際專利公開案第WO2013/120498號、國際專利公開案第WO2013/120626號、國際專利公開案第WO2013/120499號及國際專利公開案第WO2013/120628號中所述之序列，各案內容以全文引用之方式併入本文中。In some embodiments, the nucleic acid molecule of the present disclosure comprises at least one stem-loop sequence and a poly-A region or a polyadenylation signal. Non-limiting examples of polynucleotide sequences comprising at least one stem-loop sequence and a poly-A region or a polyadenylation signal include International Patent Publication No. WO2013/120497, International Patent Publication No. WO2013/120629, International Patent Publication No. WO2013/120500, International Patent Publication No. WO201 3/120627, International Patent Publication No. WO2013/120498, International Patent Publication No. WO2013/120626, International Patent Publication No. WO2013/120499 and International Patent Publication No. WO2013/120628, the contents of each case are incorporated herein by reference in their entirety.
在一些實施例中，包含莖-環序列及poly-A區或聚腺苷酸化信號之核酸分子可編碼病原體抗原或其片段，諸如國際專利公開案第WO2013/120499號及國際專利公開案第WO2013/120628號中所述之聚核苷酸序列，各案內容以全文引用之方式併入本文中。In some embodiments, a nucleic acid molecule comprising a stem-loop sequence and a poly-A region or a polyadenylation signal may encode a pathogen antigen or a fragment thereof, such as the polynucleotide sequences described in International Patent Publication No. WO2013/120499 and International Patent Publication No. WO2013/120628, each of which is incorporated herein by reference in its entirety.
在一些實施例中，包含莖-環序列及poly-A區或聚腺苷酸化信號之核酸分子可編碼治療性蛋白質，諸如國際專利公開案第WO2013/120497號及國際專利公開案第WO2013/120629號中所述之聚核苷酸序列，各案內容以全文引用之方式併入本文中。In some embodiments, a nucleic acid molecule comprising a stem-loop sequence and a poly-A region or a polyadenylation signal may encode a therapeutic protein, such as the polynucleotide sequences described in International Patent Publication No. WO2013/120497 and International Patent Publication No. WO2013/120629, each of which is incorporated herein by reference in its entirety.
在一些實施例中，包含莖-環序列及poly-A區或聚腺苷酸化信號之核酸分子可編碼腫瘤抗原或其片段，諸如國際專利公開案第WO2013/120500號及國際專利公開案第WO2013/120627號中所述之聚核苷酸序列，各案內容以全文引用之方式併入本文中。In some embodiments, a nucleic acid molecule comprising a stem-loop sequence and a poly-A region or a polyadenylation signal may encode a tumor antigen or a fragment thereof, such as the polynucleotide sequences described in International Patent Publication No. WO2013/120500 and International Patent Publication No. WO2013/120627, each of which is incorporated herein by reference in its entirety.
在一些實施例中，包含莖-環序列及poly-A區或聚腺苷酸化信號之核酸分子可編碼致敏性抗原或自體免疫性自體抗原，諸如國際專利公開案第WO2013/120498號及國際專利公開案第WO2013/120626號中所述之聚核苷酸序列，各案內容以全文引用之方式併入本文中。In some embodiments, a nucleic acid molecule comprising a stem-loop sequence and a poly-A region or a polyadenylation signal may encode a sensitizing antigen or an autoimmune autoantigen, such as the polynucleotide sequences described in International Patent Publication No. WO2013/120498and International Patent Publication No. WO2013/120626, each of which is incorporated herein by reference in its entirety.
功能性核苷酸類似物Functional nucleotide analogs
在一些實施例中，本文所述之有效負載核酸分子僅含有選自A(腺苷)、G(鳥苷)、C(胞嘧啶)、U(尿苷)及T(胸苷)之經典核苷酸。不受理論束縛，預期某些功能性核苷酸類似物可賦予核酸分子有用之特性。在本揭示案之上下文中，此等有用特性之實例包括但不限於核酸分子之穩定性增加、核酸分子在誘導先天免疫反應中之免疫原性降低、由核酸分子編碼之蛋白質之產量增加、核酸分子之細胞內遞送及/或保留增加，及/或核酸分子之細胞毒性降低等。In some embodiments, the effective carrier nucleic acid molecules described herein contain only classical nucleotides selected from A (adenosine), G (guanosine), C (cytosine), U (uridine) and T (thymidine). Without being bound by theory, it is expected that certain functional nucleotide analogs can confer useful properties on nucleic acid molecules. In the context of the present disclosure, examples of such useful properties include, but are not limited to, increased stability of nucleic acid molecules, reduced immunogenicity of nucleic acid molecules in inducing innate immune responses, increased production of proteins encoded by nucleic acid molecules, increased intracellular delivery and/or retention of nucleic acid molecules, and/or reduced cytotoxicity of nucleic acid molecules, etc.
因此，在一些實施例中，有效負載核酸分子包含至少一種如本文所述之功能性核苷酸類似物。在一些實施例中，功能性核苷酸類似物含有至少一個針對核鹼基、糖基及/或磷酸酯基之化學修飾。因此，包含至少一種功能性核苷酸類似物之有效負載核酸分子含有至少一個針對核鹼基、糖基及/或核苷間鍵聯之化學修飾。本文提供對核酸分子之核鹼基、糖基或核苷間鍵聯之實例性化學修飾。Thus, in some embodiments, the payload nucleic acid molecule comprises at least one functional nucleotide analog as described herein. In some embodiments, the functional nucleotide analog contains at least one chemical modification to a nucleobase, sugar group, and/or phosphate group. Thus, the payload nucleic acid molecule comprising at least one functional nucleotide analog contains at least one chemical modification to a nucleobase, sugar group, and/or internucleoside linkage. Exemplary chemical modifications to nucleobases, sugar groups, or internucleoside linkages of nucleic acid molecules are provided herein.
如本文所述，有效負載核酸分子中所有核苷酸之0%至100%範圍之核苷酸可以為如本文所述之功能性核苷酸類似物。舉例而言，在各個實施例中，核酸分子中之所有核苷酸中約1%至約20%、約1%至約25%、約1%至約50%、約1%至約60%、約1%至約70%、約1%至約80%、約1%至約90%、約1%至約95%、約10%至約20%、約10%至約25%、約10%至約50%、約10%至約60%、約10%至約70%、約10%至約80%、約10%至約90%、約10%至約95%、約10%至約100%、約20%至約25%、約20%至約50%、約20%至約60%、約20%至約70%、約20%至約80%、約20%至約90%、約20%至約95%、約20%至約100%、約50%至約60%、約50%至約70%、約50%至約80%、約50%至約90%、約50%至約95%、約50%至約100%、約70%至約80%、約70%至約90%、約70%至約95%、約70%至約100%、約80%至約90%、約80%至約95%、約80%至約100%、約90%至約95%、約90%至約100%、或約95%至約100%之核苷酸係本文所述之功能性核苷酸類似物。在此等實施例中之任一個中，功能性核苷酸類似物可存在於核酸分子之任何位置處，包括5’-末端、3’-末端及/或一或多個內部位置。在一些實施例中，單個核酸分子可含有不同之糖修飾、不同之核鹼基修飾及/或不同類型之核苷間鍵聯(例如主鏈結構)。As described herein, nucleotides ranging from 0% to 100% of all nucleotides in the effective load nucleic acid molecule can be functional nucleotide analogs as described herein. For example, in various embodiments, about 1% to about 20%, about 1% to about 25%, about 1% to about 50%, about 1% to about 60%, about 1% to about 70%, about 1% to about 80%, about 1% to about 90%, about 1% to about 95%, about 10% to about 20%, about 10% to about 25%, about 10% to about 50%, about 10% to about 60%, about 10% to about 70%, about 10% to about 80%, about 10% to about 90%, about 10% to about 95%, about 10% to about 100%, about 20% to about 25%, about 20% to about 50%, about 20% to about 60%, about 20% to about 70%, about 10% to about 80%, about 10% to about 90%, about 10% to about 95%, about 10% to about 100%, about 20% to about 25%, about 20% to about 50%, about 20% to about 60%, about 20% to about 70 %, about 20% to about 80%, about 20% to about 90%, about 20% to about 95%, about 20% to about 100%, about 50% to about 60%, about 50% to about 70%, about 50% to about 80%, about 50% to about 90%, about 50% to about 95%, about 50% to about 100%, about 70% to about 80%, about 70% to about 90%, about 70% to about 95%, about 70% to about 100%, about 80% to about 90%, about 80% to about 95%, about 80% to about 100%, about 90% to about 95%, about 90% to about 100%, or about 95% to about 100% of the nucleotides are functional nucleotide analogs described herein. In any of these embodiments, the functional nucleotide analog may be present at any position of the nucleic acid molecule, including the 5'-end, the 3'-end, and/or one or more internal positions. In some embodiments, a single nucleic acid molecule may contain different sugar modifications, different nucleobase modifications, and/or different types of internucleoside linkages (e.g., backbone structures).
如本文所述，有效負載核酸分子中一種類型之所有核苷酸(例如作為一種類型之所有含嘌呤核苷酸、或作為一種類型之所有含嘧啶核苷酸、或作為一種類型之所有A、G、C、T或U)中範圍自0%至100%之核苷酸可為本文所述之功能性核苷酸類似物。舉例而言，在各個實施例中，核酸分子中一種類型之核苷酸中約1%至約20%、約1%至約25%、約1%至約50%、約1%至約60%、約1%至約70%、約1%至約80%、約1%至約90%、約1%至約95%、約10%至約20%、約10%至約25%、約10%至約50%、約10%至約60%、約10%至約70%、約10%至約80%、約10%至約90%、約10%至約95%、約10%至約100%、約20%至約25%、約20%至約50%、約20%至約60%、約20%至約70%、約20%至約80%、約20%至約90%、約20%至約95%、約20%至約100%、約50%至約60%、約50%至約70%、約50%至約80%、約50%至約90%、約50%至約95%、約50%至約100%、約70%至約80%、約70%至約90%、約70%至約95%、約70%至約100%、約80%至約90%、約80%至約95%、約80%至約100%、約90%至約95%、約90%至約100%、或約95%至約100%之核苷酸係本文所述之功能性核苷酸類似物。在此等實施例中之任一個中，功能性核苷酸類似物可存在於核酸分子之任何位置處，包括5’-末端、3’-末端及/或一或多個內部位置。在一些實施例中，單個核酸分子可含有不同之糖修飾、不同之核鹼基修飾及/或不同類型之核苷間鍵聯(例如主鏈結構)。As described herein, ranging from 0% to 100% of all nucleotides of one type in a payload nucleic acid molecule (e.g., all purine-containing nucleotides as one type, or all pyrimidine-containing nucleotides as one type, or all A, G, C, T, or U as one type) can be functional nucleotide analogs described herein. For example, in various embodiments, about 1% to about 20%, about 1% to about 25%, about 1% to about 50%, about 1% to about 60%, about 1% to about 70%, about 1% to about 80%, about 1% to about 90%, about 1% to about 95%, about 10% to about 20%, about 10% to about 25%, about 10% to about 50%, about 10% to about 60%, about 10% to about 70%, about 10% to about 80%, about 10% to about 90%, about 10% to about 95%, about 10% to about 100%, about 20% to about 25%, about 20% to about 50%, about 20% to about 60%, about 20% to about 70%, about 10% to about 80%, about 10% to about 90%, about 10% to about 95%, about 10% to about 100%, about 20% to about 25%, about 20% to about 50%, about 20% to about 60%, about 20% to about 70%, about 10% to about 80%, about 10% to about 90%, about 10% to about 95%, about 10% to about 100%, about 20% to about 25%, about 20% to about 50%, about 20% to about 60%, about 20% to about In some embodiments, the nucleotides in the present invention are functional nucleotide analogs as described herein, such as about 0%, about 20% to about 80%, about 20% to about 90%, about 20% to about 95%, about 20% to about 100%, about 50% to about 60%, about 50% to about 70%, about 50% to about 80%, about 50% to about 90%, about 50% to about 95%, about 50% to about 100%, about 70% to about 80%, about 70% to about 90%, about 70% to about 95%, about 70% to about 100%, about 80% to about 90%, about 80% to about 95%, about 80% to about 100%, about 90% to about 95%, about 90% to about 100%, or about 95% to about 100% of the nucleotides are functional nucleotide analogs as described herein. In any of these embodiments, the functional nucleotide analog may be present at any position of the nucleic acid molecule, including the 5'-end, the 3'-end, and/or one or more internal positions. In some embodiments, a single nucleic acid molecule may contain different sugar modifications, different nucleobase modifications, and/or different types of internucleoside linkages (e.g., backbone structures).
核鹼基之修飾Modification of nucleobase
在一些實施例中，功能性核苷酸類似物含有非經典核鹼基。在一些實施例中，核苷酸中之經典核鹼基(例如腺嘌呤、鳥嘌呤、尿嘧啶、胸腺嘧啶及胞嘧啶)可經修飾或置換以提供核苷酸之一或多種功能性類似物。核鹼基之實例性修飾包括但不限於一或多個取代或修飾，包括但不限於烷基、芳基、鹵基、側氧基、羥基、烷氧基及/或硫代取代；一或多個稠環或開環、氧化及/或還原。In some embodiments, the functional nucleotide analog contains a non-classical nucleobase. In some embodiments, the classical nucleobases in the nucleotide (e.g., adenine, guanine, uracil, thymine, and cytosine) can be modified or replaced to provide one or more functional analogs of the nucleotide. Exemplary modifications of the nucleobase include, but are not limited to, one or more substitutions or modifications, including, but not limited to, alkyl, aryl, halogen, pendoxy, hydroxyl, alkoxy, and/or thio substitutions; one or more fused or opened rings, oxidations, and/or reductions.
在一些實施例中，非經典核鹼基係經修飾之尿嘧啶。具有經修飾之尿嘧啶之實例性核鹼基及核苷包括假尿苷(ψ)、吡啶-4-酮核糖核苷、5-氮雜尿嘧啶、6-氮雜尿嘧啶、2-硫代-5-氮雜尿嘧啶、2-硫代尿嘧啶(s2U)、4-硫代-尿嘧啶(s4U)、4-硫代-假尿苷、2-硫代-假尿苷、5-羥基-尿嘧啶(ho5U)、5-胺基烯丙基-尿嘧啶、5-鹵代尿嘧啶(例如5-碘尿嘧啶或5-溴尿嘧啶)、3-甲基尿嘧啶(m3U)、5-甲氧基尿嘧啶(mo5U)、尿嘧啶5-氧乙酸(cmo5U)、尿嘧啶5-氧乙酸甲酯(mcmo5U)、5-羧甲基-尿嘧啶(cm5U)、1-羧甲基-假尿苷、5-羧基羥甲基-尿嘧啶(chm5U)、5-羧基羥甲基-尿嘧啶甲酯(mchm5U)、5-甲氧基羰基甲基-尿嘧啶(mcm5U)、5-甲氧基羰基甲基-2-硫代尿嘧啶(mcm5s2U)、5-胺基甲基-2-硫代尿嘧啶(nm5s2U)、5-甲基胺基甲基尿嘧啶(mnm5U)、5-甲基胺基甲基-2-硫代尿嘧啶(mnm5s2U)、5-甲基胺基甲基-2-硒代尿嘧啶(mnm5se2U)、5-胺甲醯基甲基尿嘧啶(ncm5U)、5-羧甲基胺基甲基-尿嘧啶(cmnm5U)、5-羧甲基胺基甲基-2-硫代尿嘧啶(cmnm5s2U)、5-丙炔基-尿嘧啶、1-丙炔基-假尿嘧啶、5-牛磺酸甲基-尿嘧啶(τm5U)、1-牛磺酸甲基-假尿苷、5-牛磺酸甲基-2-硫代-尿嘧啶(τm55s2U)、1-牛磺酸甲基-4-硫代-假尿苷、5-甲基-尿嘧啶(m5U，即，具有核鹼基去氧胸腺嘧啶)、1-甲基-假尿苷(m1ψ)、1-乙基-假尿苷(Et1ψ)、5-甲基-2-硫代-尿嘧啶(m5s2U)、1-甲基-4-硫代-假尿苷(m1s4ψ)、4-硫代-1-甲基-假尿苷、3-甲基-假尿苷(m3ψ)、2-硫代-1-甲基-假尿苷、1-甲基-1-去氮-假尿苷、2-硫代-1-甲基-1-去氮-假尿苷、二氫尿嘧啶(D)、二氫假尿苷、5,6-二氫尿嘧啶、5-甲基-二氫尿嘧啶(m5D)、2-硫代-二氫尿嘧啶、2-硫代-二氫假尿苷、2-甲氧基-尿嘧啶、2-甲氧基-4-硫代-尿嘧啶、4-甲氧基-假尿苷、4-甲氧基-2-硫代-假尿苷、N1-甲基-假尿苷、3-(3-胺基-3-羧基丙基)尿嘧啶(acp3U)、1-甲基-3-(3-胺基-3-羧基丙基)假尿苷(acp3ψ)、5-(異戊烯基胺基甲基)尿嘧啶(m5U)、5-(異戊烯基胺基甲基)-2-硫代-尿嘧啶(m5s2U)、5,2’-O-二甲基-尿苷(m5Um)、2-硫代-2’-O-甲基-尿苷(s2Um)、5-甲氧基羰基甲基-2’-O-甲基-尿苷(mcm5Um)、5-胺甲醯基甲基-2’-O-甲基-尿苷(ncm5Um)、5-羧甲基胺基甲基-2’-O-甲基-尿苷(cmnm5Um)、3,2’-O-二甲基-尿苷(m3Um)及5-(異戊烯基胺基甲基)-2’-O-甲基-尿苷(inm5Um)、1-硫代-尿嘧啶、去氧胸苷、5-(2-甲氧羰基乙烯基)-尿嘧啶、5-(胺甲醯基羥甲基)-尿嘧啶、5-胺甲醯基甲基-2-硫代-尿嘧啶、5-羧甲基-2-硫代-尿嘧啶、5-氰基甲基-尿嘧啶、5-甲氧基-2-硫代-尿嘧啶及5-[3-(1-E-丙烯基胺基)]尿嘧啶。In some embodiments, the non-classical nucleobase is a modified uracil. Exemplary nucleobases and nucleosides having modified uracils include pseudouridine (ψ), pyridin-4-ketoribonucleoside, 5-azauracil, 6-azauracil, 2-thio-5-azauracil, 2-thiouracil (s2 U), 4-thio-uracil (s4 U), 4-thio-pseudouridine, 2-thio-pseudouridine, 5-hydroxy-uracil (ho5 U), 5-aminoallyl-uracil, 5-halogenated uracil (e.g., 5-iodouracil or 5-bromouracil), 3-methyluracil (m3 U), 5-methoxyuracil (mo5 U), uracil 5-oxyacetic acid (cmo5 U), uracil 5-oxyacetic acid methyl ester (mcmo5 U), 5-carboxymethyl-uracil (cm5 U), 1-carboxymethyl-pseudouridine, 5-carboxyhydroxymethyl-uracil (chm5 U), 5-carboxyhydroxymethyl-uracil methyl ester (mchm5 U), 5-methoxycarbonylmethyl-uracil (mcm5 U), 5-methoxycarbonylmethyl-2-thiouracil (mcm5 s2 U), 5-aminomethyl-2-thiouracil (nm5 s2 U), 5-methylaminomethyluracil (mnm5 U), 5-methylaminomethyl-2-thiouracil (mnm5 s2 U), 5-methylaminomethyl-2-selenouracil (mnm5 se2 U), 5-aminomethylmethyluracil (ncm5 U), 5-carboxymethylaminomethyl-uracil (cmnm5 U), 5-carboxymethylaminomethyl-2-thiouracil (cmnm5 s2 U), 5-propynyl-uracil, 1-propynyl-pseudouracil, 5-taurine methyl-uracil (τm5 U), 1-taurine methyl-pseudouridine, 5-taurine methyl-2-thio-uracil (τm5 5s2 U), 1-taurine methyl-4-thio-pseudouridine, 5-methyl-uracil (m5 U, i.e., having a nucleobase deoxythymine), 1-methyl-pseudouridine (m1 ψ), 1-ethyl-pseudouridine (Et1 ψ), 5-methyl-2-thio-uracil (m5 s2 U), 1-methyl-4-thio-pseudouridine (m1 s4 ψ), 4-thio-1-methyl-pseudouridine, 3-methyl-pseudouridine (m3 ψ), 2-thio-1-methyl-pseudouridine, 1-methyl-1-deaza-pseudouridine, 2-thio-1-methyl-1-deaza-pseudouridine, dihydrouracil (D), dihydropseudouridine, 5,6-dihydrouracil, 5-methyl-dihydrouracil (m5 D), 2-thio-dihydrouracil, 2-thio-dihydropseudouridine, 2-methoxy-uracil, 2-methoxy-4-thio-uracil, 4-methoxy-pseudouridine, 4-methoxy-2-thio-pseudouridine, N1-methyl-pseudouridine, 3-(3-amino-3-carboxypropyl)uracil (acp3 U), 1-methyl-3-(3-amino-3-carboxypropyl)pseudouridine (acp3 ψ), 5-(isopentenylaminomethyl)uracil (m5 Um), 5-(isopentenylaminomethyl)-2-thiouracil (m5 s2 Um), 5,2'-O-dimethyl-uridine (m5 Um), 2-thio-2'-O-methyl-uridine (s2 Um), 5-methoxycarbonylmethyl-2'-O-methyl-uridine (mcm5 Um), 5-aminoformylmethyl-2'-O-methyl-uridine (ncm5 Um), 5-carboxymethylaminomethyl-2'-O-methyl-uridine (cmnm5 Um), 3,2'-O-dimethyl-uridine (m3 Um) and 5-(isopentenylaminomethyl)-2'-O-methyl-uridine (inm5 Um ). Um), 1-thio-uracil, deoxythymidine, 5-(2-methoxycarbonylvinyl)-uracil, 5-(aminoformylhydroxymethyl)-uracil, 5-aminoformylmethyl-2-thio-uracil, 5-carboxymethyl-2-thio-uracil, 5-cyanomethyl-uracil, 5-methoxy-2-thio-uracil and 5-[3-(1-E-propenylamino)]uracil.
在一些實施例中，非經典核鹼基係經修飾之胞嘧啶。具有經修飾之胞嘧啶之實例性核鹼基及核苷包括5-氮雜胞嘧啶、6-氮雜胞嘧啶、假異胞苷、3-甲基胞嘧啶(m3C)、N4-乙醯基胞嘧啶(ac4C)、5-甲醯基胞嘧啶(f5C)、N4-甲基-胞嘧啶(m4C)、5-甲基-胞嘧啶(m5C)、5-鹵代-胞嘧啶(例如5-碘-胞嘧啶)、5-羥甲基-胞嘧啶(hm5C)、1-甲基-假異胞苷、吡咯并胞嘧啶、吡咯并假異胞苷、2-硫代胞嘧啶(s2C)、2-硫代-5-甲基胞嘧啶、4-硫代假異胞苷、4-硫代-1-甲基-假異胞苷、4-硫代-1-甲基-1-去氮-假異胞苷、1-甲基-1-去氮-假異胞苷、澤布拉林(zebularine)、5-氮雜-澤布拉林、5-甲基-澤布拉林、5-氮雜-2-硫代-澤布拉林、2-硫代-澤布拉林、2-甲氧基-胞嘧啶、2-甲氧基-5-甲基-胞嘧啶、4-甲氧基-假異胞苷、4-甲氧基-1-甲基-假異胞苷、賴西丁(lysidine)(k2C)、5,2’-O-二甲基-胞苷(m5Cm)、N4-乙醯基-2’-O-甲基-胞苷(ac4Cm)、N4,2’-O-二甲基-胞苷(m4Cm)、5-甲醯基-2’-O-甲基-胞苷(fSCm)、N4,N4,2’-O-三甲基-胞苷(m42Cm)、1-硫代-胞嘧啶、5-羥基-胞嘧啶、5-(3-疊氮基丙基)-胞嘧啶及5-(2-疊氮基乙基)-胞嘧啶。In some embodiments, the non-classical nucleobase is a modified cytosine. Exemplary nucleobases and nucleosides having modified cytosine include 5-azacytosine, 6-azacytosine, pseudoisocytidine, 3-methylcytosine (m3C), N4-acetylcytosine (ac4C), 5-formylcytosine (f5C), N4-methyl-cytosine (m4C), 5-methyl-cytosine (m5C), 5-halogenated-cytosine (e.g., 5-iodo-cytosine), 5-hydroxymethyl 2-thiocytosine (s2C), 2-thio-5-methylcytosine, 4-thio-pseudoisocytidine, 4-thio-1-methyl-pseudoisocytidine, 4-thio-1-methyl-pseudoisocytidine, 1-methyl-1-deaza-pseudoisocytidine, zebularine, 5-thio-1-methyl-pseudoisocytidine, 1-methyl-1-deaza ... -aza-zebularine, 5-methyl-zebularine, 5-aza-2-thio-zebularine, 2-thio-zebularine, 2-methoxy-cytosine, 2-methoxy-5-methyl-cytosine, 4-methoxy-pseudoisocytidine, 4-methoxy-1-methyl-pseudoisocytidine, lysidine (k2C), 5,2'-O-dimethyl-cytidine (m5Cm), N4-ethyl Acyl-2’-O-methyl-cytidine (ac4Cm), N4,2’-O-dimethyl-cytidine (m4Cm), 5-methyl-2’-O-methyl-cytidine (fSCm), N4,N4,2’-O-trimethyl-cytidine (m42Cm), 1-thio-cytosine, 5-hydroxy-cytosine, 5-(3-azidopropyl)-cytosine and 5-(2-azidoethyl)-cytosine.
在一些實施例中，非經典核鹼基係經修飾之腺嘌呤。具有替代性腺嘌呤之實例性核鹼基及核苷包括2-胺基-嘌呤、2,6-二胺基嘌呤、2-胺基-6-鹵代-嘌呤(例如2-胺基-6-氯-嘌呤)、6-鹵代-嘌呤(例如6-氯-嘌呤)、2-胺基-6-甲基-嘌呤、8-疊氮基-腺嘌呤、7-去氮-腺嘌呤、7-去氮-8-氮雜-腺嘌呤、7-去氮-2-胺基-嘌呤、7-去氮-8-氮雜-2-胺基-嘌呤、7-去氮-2,6-二胺基嘌呤、7-去氮-8-氮雜-2,6-二胺基嘌呤、1-甲基-腺嘌呤(m1A)、2-甲基-腺嘌呤(m2A)、N6-甲基-腺嘌呤(m6A)、2-甲硫基-N6-甲基-腺嘌呤(ms2m6A)、N6-異戊烯基-腺嘌呤(i6A)、2-甲硫基-N6-異戊烯基-腺嘌呤(ms2i6A)、N6-(順-羥基異戊烯基)腺嘌呤(io6A)、2-甲硫基-N6-(順-羥基異戊烯基)腺嘌呤(ms2io6A)、N6-甘胺醯基胺甲醯基-腺嘌呤(g6A)、N6-蘇胺醯基胺甲醯基-腺嘌呤(t6A)、N6-甲基-N6-蘇胺醯基胺甲醯基-腺嘌呤(m6t6A)、2-甲硫基-N6-蘇胺醯基胺甲醯基-腺嘌呤(ms2g6A)、N6,N6-二甲基-腺嘌呤(m62A)、N6-羥基正纈胺醯基胺甲醯基-腺嘌呤(hn6A)、2-甲硫基-N6-羥基正纈胺醯基胺甲醯基-腺嘌呤(ms2hn6A)、N6-乙醯基-腺嘌呤(ac6A)、7-甲基-腺嘌呤、2-甲硫基-腺嘌呤、2-甲氧基-腺嘌呤、N6,2’-O-二甲基-腺苷(m6Am)、N6,N6,2’-O-三甲基-腺苷(m62Am)、1,2’-O-二甲基-腺苷(m1Am)、2-胺基-N6-甲基-嘌呤、1-硫代-腺嘌呤、8-疊氮基-腺嘌呤、N6-(19-胺基-五氧雜十九烷基)-腺嘌呤、2,8-二甲基-腺嘌呤、N6-甲醯基-腺嘌呤及N6-羥甲基-腺嘌呤。In some embodiments, the non-classical nucleobase is a modified adenine. Exemplary nucleobases and nucleosides having a substituted adenine include 2-amino-purine, 2,6-diaminopurine, 2-amino-6-halogenated-purine (e.g., 2-amino-6-chloro-purine), 6-halogenated-purine (e.g., 6-chloro-purine), 2-amino-6-methyl-purine, 8-azido-adenine, 7-deaza-adenine, 7-deaza-8-aza-adenine, 7-deaza-2-amino-purine, 7-deaza-8-aza-2-amino-purine, 7-deaza-2,6-diaminopurine, 7-deaza-8-aza- 2,6-Diaminopurine, 1-methyl-adenine (m1A), 2-methyl-adenine (m2A), N6-methyl-adenine (m6A), 2-methylthio-N6-methyl-adenine (ms2m6A), N6-isopentenyl-adenine (i6A), 2-methylthio-N6-isopentenyl-adenine (ms2i6A), N6-(cis-hydroxyisopentenyl)adenine (io6A), 2-methylthio-N6-(cis-hydroxyisopentenyl)adenine (ms2io6A), N6-glycolylaminoformyl N6-methyl-N6-threonamido-methyl-adenine (m6t6A), 2-methylthio-N6-threonamido-methyl-adenine (ms2g6A), N6,N6-dimethyl-adenine (m62A), N6-hydroxy-n-valeramido-methyl-adenine (hn6A), 2-methylthio-N6-hydroxy-n-valeramido-methyl-adenine (ms2hn6A), N6-acetyl-adenine (ac6A) , 7-methyl-adenine, 2-methylthio-adenine, 2-methoxy-adenine, N6,2’-O-dimethyl-adenosine (m6Am), N6,N6,2’-O-trimethyl-adenosine (m62Am), 1,2’-O-dimethyl-adenosine (m1Am), 2-amino-N6-methyl-purine, 1-thio-adenine, 8-azido-adenine, N6-(19-amino-pentaoxonadecanyl)-adenine, 2,8-dimethyl-adenine, N6-methyl-adenine and N6-hydroxymethyl-adenine.
在一些實施例中，非經典核鹼基係經修飾之鳥嘌呤。具有經修飾之鳥嘌呤之實例性核鹼基及核苷包括肌苷(I)、1-甲基-肌苷(m1I)、懷俄苷(wyosine)(imG)、甲基懷俄苷(mimG)、4-去甲基-懷俄苷(imG-14)、異懷俄苷(imG2)、懷丁苷(wybutosine)(yW)、過氧懷丁苷(o2yW)、羥基懷丁苷(OHyW)、修飾不足(undermodified)之羥基懷丁苷(OHyW*)、7-去氮-鳥嘌呤、辮苷(queuosine)(Q)、環氧辮苷(oQ)、半乳糖基-辮苷(galQ)、甘露糖基-辮苷(manQ)、7-氰基-7-去氮-鳥嘌呤(preQO)、7-胺基甲基-7-去氮-鳥嘌呤(preQ1)、古嘌苷(archaeosine)(G+)、7-去氮-8-氮雜-鳥嘌呤、6-硫代-鳥嘌呤、6-硫代-7-去氮-鳥嘌呤、6-硫代-7-去氮-8-氮雜-鳥嘌呤、7-甲基-鳥嘌呤(m7G)、6-硫代-7-甲基-鳥嘌呤、7-甲基-肌苷、6-甲氧基-鳥嘌呤、1-甲基-鳥嘌呤(m1G)、N2-甲基-鳥嘌呤(m2G)、N2,N2-二甲基-鳥嘌呤(m22G)、N2,7-二甲基-鳥嘌呤(m2,7G)、N2,N2,7-二甲基-鳥嘌呤(m2,2,7G)、8-側氧基-鳥嘌呤、7-甲基-8-側氧基-鳥嘌呤、1-甲基-6-硫代-鳥嘌呤、N2-甲基-6-硫代-鳥嘌呤、N2,N2-二甲基-6-硫代-鳥嘌呤、N2-甲基-2’-O-甲基-鳥苷(m2Gm)、N2,N2-二甲基-2’-O-甲基-鳥苷(m22Gm)、1-甲基-2’-O-甲基-鳥苷(m1Gm)、N2,7-二甲基-2’-O-甲基-鳥苷(m2,7Gm)、2’-O-甲基-肌苷(Im)、1,2’-O-二甲基-肌苷(m1Im)、1-硫代-鳥嘌呤及O-6-甲基-鳥嘌呤。In some embodiments, the non-classical nucleobase is a modified guanine. Exemplary nucleobases and nucleosides having modified guanine include inosine (I), 1-methyl-inosine (m1I), wyosine (imG), methyl wyosine (mimG), 4-demethyl-wyosine (imG-14), iso-wyosine (imG2), wybutosine (yW), peroxy wybutosine (o2yW), hydroxy wybutosine (OHyW), undermodified hydroxy wybutosine (OHyW*), 7-deaza-guanosine (hdroxy) Purine, queuosine (Q), oxoqueuosine (oQ), galactosyl-queuosine (galQ), mannosyl-queuosine(manQ), 7-cyano-7-deaza-guanine (preQO), 7-aminomethyl-7-deaza-guanine (preQ1), archaeosine (G+), 7-deaza-8-aza-guanine, 6-thio-guanine, 6-thio-7-deaza-guanine, 6-thio-7-deaza-8-aza-guanine, 7-methyl -guanine (m7G), 6-thio-7-methyl-guanine, 7-methyl-inosine, 6-methoxy-guanine, 1-methyl-guanine (m1G), N2-methyl-guanine (m2G), N2,N2-dimethyl-guanine (m22G), N2,7-dimethyl-guanine (m2,7G), N2,N2,7-dimethyl-guanine (m2,2,7G), 8-oxo-guanine, 7-methyl-8-oxo-guanine, 1-methyl-6-thio-guanine, N2-methyl-6 -thio-guanine, N2,N2-dimethyl-6-thio-guanine, N2-methyl-2’-O-methyl-guanosine (m2Gm), N2,N2-dimethyl-2’-O-methyl-guanosine (m22Gm), 1-methyl-2’-O-methyl-guanosine (m1Gm), N2,7-dimethyl-2’-O-methyl-guanosine (m2,7Gm), 2’-O-methyl-inosine (Im), 1,2’-O-dimethyl-inosine (m1Im), 1-thio-guanine and O-6-methyl-guanine.
在一些實施例中，功能性核苷酸類似物之非經典核鹼基可獨立地為嘌呤、嘧啶、嘌呤類似物或嘧啶類似物。舉例而言，在一些實施例中，非經典核鹼基可為經修飾之腺嘌呤、胞嘧啶、鳥嘌呤、尿嘧啶或次黃嘌呤。在其他實施例中，非經典核鹼基亦可包括例如鹼基之天然存在的及合成的衍生物，包括吡唑并[3,4-d]嘧啶；5-甲基胞嘧啶(5-me-C)；5-羥甲基胞嘧啶；黃嘌呤；次黃嘌呤；2-胺基腺嘌呤；腺嘌呤及鳥嘌呤之6-甲基及其他烷基衍生物；腺嘌呤及鳥嘌呤之2-丙基及其他烷基衍生物；2-硫代尿嘧啶、2-硫代胸腺嘧啶及2-硫代胞嘧啶；5-丙炔基尿嘧啶及胞嘧啶；6-偶氮尿嘧啶、胞嘧啶及胸腺嘧啶；5-尿嘧啶(假尿嘧啶)；4-硫代尿嘧啶；8-鹵代(例如8-溴)、8-胺基、8-硫醇、8-硫代烷基、8-羥基及其他8-取代之腺嘌呤及鳥嘌呤；5-鹵代，尤其為5-溴、5-三氟甲基及其他5-取代之尿嘧啶及胞嘧啶；7-甲基鳥嘌呤及7-甲基腺嘌呤；8-氮雜鳥嘌呤及8-氮雜腺嘌呤；去氮鳥嘌呤、7-去氮鳥嘌呤、3-去氮鳥嘌呤；去氮腺嘌呤、7-去氮腺嘌呤、3-去氮腺嘌呤；吡唑并[3,4-d]嘧啶；咪唑并[1,5-a]1,3,5-三
糖之修飾Sugar Decoration
在一些實施例中，功能性核苷酸類似物含有非經典糖基。在各個實施例中，非經典糖基可為具有一或多個取代之5碳或6碳糖(諸如戊糖、核糖、阿拉伯糖、木糖、葡萄糖、半乳糖或其去氧衍生物)，該一或多個取代諸如為鹵基、羥基、硫醇基、烷基、烷氧基、烯基氧基、炔基氧基、環烷基、胺基烷氧基、烷氧基烷氧基、羥基烷氧基、胺基、疊氮基、芳基、胺基烷基、胺基烯基、胺基炔基等。In some embodiments, the functional nucleotide analog contains a non-classical sugar group. In various embodiments, the non-classical sugar group can be a 5-carbon or 6-carbon sugar (such as pentose, ribose, arabinose, xylose, glucose, galactose or its deoxy derivatives) with one or more substitutions, such as halogen, hydroxyl, thiol, alkyl, alkoxy, alkenyloxy, alkynyloxy, cycloalkyl, aminoalkoxy, alkoxyalkoxy, hydroxyalkoxy, amino, azido, aryl, aminoalkyl, aminoalkenyl, aminoalkynyl, etc.
一般而言，RNA分子含有核糖基，該核糖基係含氧5員環。實例性非限制性替代核苷酸包括核糖中之氧置換(例如用S、Se或伸烷基，諸如亞甲基或伸乙基置換)；雙鍵之添加(例如用環戊烯基或環己烯基置換核糖)；核糖之縮環(例如用於形成環丁烷或氧雜環丁烷之4員環)；核糖之擴環(例如用於形成具有額外碳或雜原子之6員或7員環，諸如對於失水己糖醇、阿卓糖醇(altritol)、甘露糖醇、環己烷基、環己烯基及N-嗎啉基(其亦具有胺基磷酸酯主鏈))；多環形式(例如三環及「未鎖定」形式，諸如二醇核酸(GNA)(例如R-GNA或S-GNA，其中核糖經連接至磷酸二酯鍵之二醇單元置換)、蘇糖核酸(TNA，其中核糖經α-L-呋喃蘇糖基-(3’→2’)置換)及肽核酸(PNA，其中2-胺基-乙基-甘胺酸鍵聯置換核糖及磷酸二酯主鏈))。In general, RNA molecules contain a ribose moiety that is an oxygen-containing 5-membered ring. Exemplary non-limiting substituted nucleotides include replacement of the oxygen in ribose (e.g., with S, Se, or an alkylene group, such as a methylene or ethylene group); addition of double bonds (e.g., replacement of ribose with a cyclopentenyl or cyclohexenyl group); Ring condensation of ribose (e.g., to form a 4-membered ring of cyclobutane or oxyheterocyclobutane); Ring expansion of ribose (e.g., to form a 6- or 7-membered ring with additional carbon or heteroatoms, such as for anhydrohexitol, altritol, mannitol, cyclohexyl, cyclohexenyl and N-morpholinyl (which also have a phosphoamidoester backbone); polycyclic forms (e.g. tricyclic and "unlocked" forms such as glycol nucleic acids (GNAs) (e.g. R-GNA or S-GNA, in which the ribose is replaced by a glycol unit linked to a phosphodiester bond), thiosyl nucleic acids (TNA, in which the ribose is replaced by an α-L-thiofuranosyl-(3'→2')) and peptide nucleic acids (PNA, in which 2-amino-ethyl-glycine bonds replace the ribose and phosphodiester backbones).
在一些實施例中，糖基含有一或多個碳，該一或多個碳具有與核糖中對應碳相反之立體化學組態。因此，核酸分子可包括含例如阿拉伯糖或L-核糖作為糖之核苷酸。在一些實施例中，核酸分子包括至少一個其中糖係L-核糖、2’-O-甲基核糖、2’-氟核糖、阿拉伯糖、己糖醇、LNA或PNA之核苷。In some embodiments, the sugar group contains one or more carbons that have a stereochemical configuration opposite to the corresponding carbon in ribose. Thus, a nucleic acid molecule can include a nucleotide containing, for example, arabinose or L-ribose as a sugar. In some embodiments, a nucleic acid molecule includes at least one nucleoside in which the sugar is L-ribose, 2'-O-methylribose, 2'-fluororibose, arabinose, hexitol, LNA, or PNA.
核苷間鍵聯之修飾Modification of internucleoside linkages
在一些實施例中，本揭示案之有效負載核酸分子可含有一或多個經修飾之核苷間鍵聯(例如磷酸酯主鏈)。主鏈磷酸酯基可藉由用不同取代基置換一或多個氧原子來改變。In some embodiments, the payload nucleic acid molecules of the present disclosure may contain one or more modified internucleoside linkages (e.g., phosphate backbones). The backbone phosphate groups may be altered by replacing one or more oxygen atoms with different substituents.
在一些實施例中，功能性核苷酸類似物可包括用本文所述之另一核苷間鍵聯置換未改變之磷酸酯部分。替代性磷酸酯基之實例包括但不限於硫代磷酸酯、硒代磷酸酯、硼烷磷酸鹽、硼烷磷酸酯、膦酸氫酯、胺基磷酸酯、二胺基磷酸酯、烷基或芳基膦酸酯及磷酸三酯。二硫代磷酸酯之兩個非連接氧均經硫置換。亦可藉由用氮(橋連之胺基磷酸酯)、硫(橋連之硫代磷酸酯)及碳(橋連之亞甲基膦酸酯)置換連接氧來改變磷酸酯連接體。In some embodiments, a functional nucleotide analog may include replacement of an unchanged phosphate moiety with another internucleoside linkage as described herein. Examples of alternative phosphate groups include, but are not limited to, phosphorothioates, phosphoroselenoates, boranophosphates, boranophosphates, hydrogen phosphonates, phosphoamidates, phosphodiamidates, alkyl or aryl phosphonates, and phosphotriesters. Both non-linking oxygens of phosphorodithioates are replaced with sulfur. Phosphate linkers may also be altered by replacement of the linking oxygen with nitrogen (bridged phosphoramidates), sulfur (bridged phosphorothioates), and carbon (bridged methylenephosphonates).
替代性核苷及核苷酸可包括一或多個非橋連氧被甲硼烷部分(BH3)、硫(硫代)、甲基、乙基及/或甲氧基置換。作為非限制性實例，在同一位置(例如alpha(α)、beta(β)或gamma(γ)位置)處之兩個非橋連氧可經硫(硫代)及甲氧基置換。置換磷酸酯部分(例如α-硫代磷酸酯)位置處之一或多個氧原子可經由非天然硫代磷酸酯主鏈鍵聯賦予RNA及DNA穩定性(諸如針對核酸外切酶及核酸內切酶之穩定性)。硫代磷酸酯DNA及RNA具有增加之核酸酶抗性，且因此在細胞環境中具有較長之半衰期。Alternative nucleosides and nucleotides may include one or more non-bridging oxygens replaced by a borane moiety (BH3 ), sulfur (thio), methyl, ethyl, and/or methoxy. As a non-limiting example, two non-bridging oxygens at the same position (e.g., alpha (α), beta (β), or gamma (γ) position) may be replaced by sulfur (thio) and methoxy. Replacing one or more oxygen atoms at a phosphate moiety (e.g., α-phosphorothioate) position may impart stability (e.g., stability against exonucleases and endonucleases) to RNA and DNA via non-natural phosphorothioate backbone linkages. Phosphorothioate DNA and RNA have increased nuclease resistance and, therefore, have a longer half-life in the cellular environment.
本文闡述可根據本揭示案使用之其他核苷間鍵聯，包括不含磷原子之核苷間鍵聯。Other internucleoside linkages that may be used in accordance with the present disclosure are described herein, including internucleoside linkages that do not contain a phosphorus atom.
可結合本揭示案使用之核酸分子(例如mRNA)、相關組合物、調配物及/或方法之額外實例進一步包括WO2002/098443、WO2003/051401、WO2008/052770、WO2009127230、WO2006122828、WO2008/083949、WO2010088927、WO2010/037539、WO2004/004743、WO2005/016376、WO2006/024518、WO2007/095976、WO2008/014979、WO2008/077592、WO2009/030481、WO2009/095226、WO2011069586、WO2011026641、WO2011/144358、WO2012019780、WO2012013326、WO2012089338、WO2012113513、WO2012116811、WO2012116810、WO2013113502、WO2013113501、WO2013113736、WO2013143698、WO2013143699、WO2013143700、WO2013/120626、WO2013120627、WO2013120628、WO2013120629、WO2013174409、WO2014127917、WO2015/024669、WO2015/024668、WO2015/024667、WO2015/024665、WO2015/024666、WO2015/024664、WO2015101415、WO2015101414、WO2015024667、WO2015062738、WO2015101416中所述者，各案之內容以全文併入本文中。Additional examples of nucleic acid molecules (e.g., mRNA), related compositions, formulations and/or methods that can be used in conjunction with the present disclosure further include WO2002/098443, WO2003/051401, WO2008/052770, WO2009127230, WO2006122828, WO2008/083949, WO2010088927, WO2010/037539, WO2004/004743, WO2005/ 016376、WO2006/024518、WO2007/095976、WO2008/014979、WO2008/077592、WO2009/030481、WO2009/095226、WO2011069586、WO2011026641, WO2011/144358, WO2012019780, WO2012013326, WO2012089338, WO20121 13513, WO2012116811, WO2012116810, WO2013113502, WO2013113501, WO2013113736, WO2013143698, WO201 3143699, WO2013143700, WO2013/120626, WO2013120627, WO2013120628, WO2013120629, WO2013174409, WO 2014127917, WO2015/024669, WO2015/024668, WO2015/024667, WO2015/024665, WO2015/024666, WO2015/024664, WO2015101415, WO2015101414, WO2015024667, WO2015062738, WO2015101416, the contents of each case are incorporated herein in their entirety.
5.5調配物5.5Preparations
根據本揭示案，本文所述之奈米顆粒組合物可包含至少一種脂質組分及一或多種額外組分，諸如治療劑及/或預防劑。奈米顆粒組合物可設計成用於一或多種特定應用或目標。奈米顆粒組合物之成分可基於特定應用或目標，及/或基於一或多種成分之功效、毒性、費用、易用性、可用性或其他特徵來選擇。類似地，奈米顆粒組合物之特定配方可根據例如每種成分之特定組合之功效及毒性，針對特定應用或目標來選擇。According to the present disclosure, the nanoparticle compositions described herein may include at least one lipid component and one or more additional components, such as therapeutic agents and/or prophylactic agents. The nanoparticle compositions may be designed for one or more specific applications or targets. The components of the nanoparticle compositions may be selected based on the specific application or target, and/or based on the efficacy, toxicity, cost, ease of use, availability or other characteristics of one or more components. Similarly, the specific formulation of the nanoparticle composition may be selected for a specific application or target based on, for example, the efficacy and toxicity of a specific combination of each component.
奈米顆粒組合物之脂質組分可包括例如本文所述之根據式(I)(及其子式)之一的脂質、磷脂(諸如不飽和脂質，例如DOPE或DSPC)、PEG脂質及結構脂質。脂質組分各成分可以特定分率提供。The lipid component of the nanoparticle composition may include, for example, lipids according to one of the formulas (I) (and its subformulae) described herein, phospholipids (such as unsaturated lipids, such as DOPE or DSPC), PEG lipids, and structured lipids. The components of the lipid component may be provided in specific fractions.
在一個實施例中，本文提供一種奈米顆粒組合物，其包含本文所提供之陽離子或可電離之脂質化合物、治療劑及一或多種賦形劑。在一個實施例中，陽離子或可電離之脂質化合物包含如本文所述之根據式(I)(及其子式)之一的化合物，以及視情況選用之一或多種額外可電離之脂質化合物。在一個實施例中，該一或多種賦形劑選自中性脂質、類固醇及聚合物結合之脂質。在一個實施例中，治療劑係包封於脂質奈米顆粒內或與脂質奈米顆粒締合。In one embodiment, a nanoparticle composition is provided herein, comprising a cationic or ionizable lipid compound provided herein, a therapeutic agent, and one or more excipients. In one embodiment, the cationic or ionizable lipid compound comprises a compound according to one of formula (I) (and its subformulae) as described herein, and one or more additional ionizable lipid compounds selected as appropriate. In one embodiment, the one or more excipients are selected from neutral lipids, steroids, and polymer-bound lipids. In one embodiment, the therapeutic agent is encapsulated in or conjugated to the lipid nanoparticles.
在一個實施例中，本文提供一種奈米顆粒組合物(脂質奈米顆粒)，其包含：i)40莫耳%至50莫耳%之陽離子脂質；ii)中性脂質；iii)類固醇；iv)聚合物結合之脂質；以及v)治療劑。In one embodiment, the present invention provides a nanoparticle composition (lipid nanoparticle) comprising: i) 40 mol% to 50 mol% of a cationic lipid; ii) a neutral lipid; iii) a steroid; iv) a polymer-bound lipid; and v) a therapeutic agent.
如本文所使用，「莫耳百分比」係指一種組分相對於LNP中所有脂質組分之總莫耳數(即，陽離子脂質、中性脂質、類固醇及聚合物結合之脂質之總莫耳數)之莫耳百分比。As used herein, "molar percentage" refers to the molar percentage of a component relative to the total molar number of all lipid components in the LNP (i.e., the total molar number of cationic lipids, neutral lipids, steroids, and polymer-bound lipids).
在一個實施例中，脂質奈米顆粒包含41莫耳%至49莫耳%、41莫耳%至48莫耳%、42莫耳%至48莫耳%、43莫耳%至48莫耳%、44莫耳%至48莫耳%、45莫耳%至48莫耳%、46莫耳%至48莫耳%或47.2莫耳%至47.8莫耳%之陽離子脂質。在一個實施例中，脂質奈米顆粒包含約47.0莫耳%、47.1莫耳%、47.2莫耳%、47.3莫耳%、47.4莫耳%、47.5莫耳%、47.6莫耳%、47.7莫耳%、47.8莫耳%、47.9莫耳%或48.0莫耳%之陽離子脂質。In one embodiment, the lipid nanoparticles contain 41 mol% to 49 mol%, 41 mol% to 48 mol%, 42 mol% to 48 mol%, 43 mol% to 48 mol%, 44 mol% to 48 mol%, 45 mol% to 48 mol%, 46 mol% to 48 mol%, or 47.2 mol% to 47.8 mol% of cationic lipids. In one embodiment, the lipid nanoparticles contain about 47.0 mol%, 47.1 mol%, 47.2 mol%, 47.3 mol%, 47.4 mol%, 47.5 mol%, 47.6 mol%, 47.7 mol%, 47.8 mol%, 47.9 mol%, or 48.0 mol% of cationic lipids.
在一個實施例中，中性脂質以在5莫耳%至15莫耳%、7莫耳%至13莫耳%或9莫耳%至11莫耳%範圍內之濃度存在。在一個實施例中，中性脂質以約9.5莫耳%、10莫耳%或10.5莫耳%之濃度存在。在一個實施例中，陽離子脂質與中性脂質之莫耳比在約4.1：1.0至約4.9：1.0、約4.5：1.0至約4.8：1.0、或約4.7：1.0至4.8：1.0範圍內。In one embodiment, the neutral lipid is present at a concentration ranging from 5 mol% to 15 mol%, 7 mol% to 13 mol%, or 9 mol% to 11 mol%. In one embodiment, the neutral lipid is present at a concentration of about 9.5 mol%, 10 mol%, or 10.5 mol%. In one embodiment, the molar ratio of cationic lipid to neutral lipid is in the range of about 4.1:1.0 to about 4.9:1.0, about 4.5:1.0 to about 4.8:1.0, or about 4.7:1.0 to 4.8:1.0.
在一個實施例中，類固醇以在39莫耳%至49莫耳%、40莫耳%至46莫耳%、40莫耳%至44莫耳%、40莫耳%至42莫耳%、42莫耳%至44莫耳%或44莫耳%至46莫耳%範圍內之濃度存在。在一個實施例中，類固醇以40莫耳%、41莫耳%、42莫耳%、43莫耳%、44莫耳%、45莫耳%或46莫耳%之濃度存在。在一個實施例中，陽離子脂質與類固醇之莫耳比在1.0：0.9至1.0：1.2、或1.0：1.0至1.0：1.2範圍內。在一個實施例中，類固醇係膽固醇。In one embodiment, the steroid is present at a concentration ranging from 39 mol% to 49 mol%, 40 mol% to 46 mol%, 40 mol% to 44 mol%, 40 mol% to 42 mol%, 42 mol% to 44 mol%, or 44 mol% to 46 mol%. In one embodiment, the steroid is present at a concentration of 40 mol%, 41 mol%, 42 mol%, 43 mol%, 44 mol%, 45 mol%, or 46 mol%. In one embodiment, the molar ratio of cationic lipid to steroid is in the range of 1.0:0.9 to 1.0:1.2, or 1.0:1.0 to 1.0:1.2. In one embodiment, the steroid is cholesterol.
在一個實施例中，LNP中治療劑與脂質之比率(即，N/P，其中N表示陽離子脂質之莫耳數，P表示作為核酸主鏈之一部分存在之磷酸酯之莫耳數)在2：1至30：1範圍內，例如在3：1至22：1範圍內。在一個實施例中，N/P在6：1至20：1或2：1至12：1範圍內。實例性N/P範圍包括約3：1、約6：1、約12：1及約22：1。In one embodiment, the ratio of therapeutic agent to lipid in the LNP (i.e., N/P, where N represents the mole number of cationic lipids and P represents the mole number of phosphates present as part of the nucleic acid backbone) is in the range of 2:1 to 30:1, such as in the range of 3:1 to 22:1. In one embodiment, N/P is in the range of 6:1 to 20:1 or 2:1 to 12:1. Exemplary N/P ranges include about 3:1, about 6:1, about 12:1, and about 22:1.
在一個實施例中，本文提供一種脂質奈米顆粒，其包含：i)有效pKa大於6.0之陽離子脂質；ii)5莫耳%至15莫耳%之中性脂質；iii)1莫耳%至15莫耳%之陰離子脂質；iv)30莫耳%至45莫耳%之類固醇；v)聚合物結合之脂質；以及vi)治療劑，或其醫藥學上可接受之鹽或前藥，其中莫耳百分比係基於脂質奈米顆粒中存在之脂質之總莫耳數確定。In one embodiment, a lipid nanoparticle is provided herein, comprising: i) a cationic lipid having an effective pKa greater than 6.0; ii) 5 mol% to 15 mol% of a neutral lipid; iii) 1 mol% to 15 mol% of an anionic lipid; iv) 30 mol% to 45 mol% of a steroid; v) a polymer-bound lipid; and vi) a therapeutic agent, or a pharmaceutically acceptable salt or prodrug thereof, wherein the molar percentage is determined based on the total molar number of lipids present in the lipid nanoparticle.
在一個實施例中，陽離子脂質可為在選定pH值，諸如生理pH值下帶有淨正電荷之多種脂質種類中之任一種。實例性陽離子脂質在下文闡述。在一個實施例中，陽離子脂質之pKa值大於6.25。在一個實施例中，陽離子脂質之pKa值大於6.5。在一個實施例中，陽離子脂質之pKa值大於6.1，大於6.2，大於6.3，大於6.35，大於6.4，大於6.45，大於6.55，大於6.6，大於6.65或大於6.7。In one embodiment, the cationic lipid can be any of a variety of lipid types that have a net positive charge at a selected pH, such as a physiological pH. Exemplary cationic lipids are described below. In one embodiment, the cationic lipid has a pKa value greater than 6.25. In one embodiment, the cationic lipid has a pKa value greater than 6.5. In one embodiment, the cationic lipid has a pKa value greater than 6.1, greater than 6.2, greater than 6.3, greater than 6.35, greater than 6.4, greater than 6.45, greater than 6.55, greater than 6.6, greater than 6.65, or greater than 6.7.
在一個實施例中，脂質奈米顆粒包含40莫耳%至45莫耳%之陽離子脂質。在一個實施例中，脂質奈米顆粒包含45莫耳%至50莫耳%之陽離子脂質。In one embodiment, the lipid nanoparticles contain 40 mol% to 45 mol% of cationic lipids. In one embodiment, the lipid nanoparticles contain 45 mol% to 50 mol% of cationic lipids.
在一個實施例中，陽離子脂質與中性脂質之莫耳比在約2：1至約8：1範圍內。在一個實施例中，脂質奈米顆粒包含5莫耳%至10莫耳%之中性脂質。In one embodiment, the molar ratio of cationic lipid to neutral lipid is in the range of about 2:1 to about 8:1. In one embodiment, the lipid nanoparticles contain 5 mol% to 10 mol% of neutral lipid.
實例性陰離子脂質包括但不限於磷脂醯甘油、二油醯基磷脂醯甘油(DOPG)、二棕櫚醯基磷脂醯甘油(DPPG)或1,2-二硬脂醯基-sn-甘油-3-磷酸-(1’-外消旋-甘油)(DSPG)。Exemplary anionic lipids include, but are not limited to, phosphatidylglycerol, dioleylphosphatidylglycerol (DOPG), dimalmitoylphosphatidylglycerol (DPPG), or 1,2-distearyl-sn-glycero-3-phospho-(1'-rac-glycerol) (DSPG).
在一個實施例中，脂質奈米顆粒包含1莫耳%至10莫耳%之陰離子脂質。在一個實施例中，脂質奈米顆粒包含1莫耳%至5莫耳%之陰離子脂質。在一個實施例中，脂質奈米顆粒包含1莫耳%至9莫耳%、1莫耳%至8莫耳%、1莫耳%至7莫耳%或1莫耳%至6莫耳%之陰離子脂質。在一個實施例中，陰離子脂質與中性脂質之莫耳比在1：1至1：10範圍內。In one embodiment, the lipid nanoparticles contain 1 mol% to 10 mol% of anionic lipids. In one embodiment, the lipid nanoparticles contain 1 mol% to 5 mol% of anionic lipids. In one embodiment, the lipid nanoparticles contain 1 mol% to 9 mol%, 1 mol% to 8 mol%, 1 mol% to 7 mol% or 1 mol% to 6 mol% of anionic lipids. In one embodiment, the molar ratio of anionic lipids to neutral lipids is in the range of 1:1 to 1:10.
在一個實施例中，類固醇係膽固醇。在一個實施例中，陽離子脂質與膽固醇之莫耳比在約5：1至1：1範圍內。在一個實施例中，脂質奈米顆粒包含32莫耳%至40莫耳%之類固醇。In one embodiment, the steroid is cholesterol. In one embodiment, the molar ratio of cationic lipid to cholesterol is in the range of about 5:1 to 1:1. In one embodiment, the lipid nanoparticles contain 32 mol% to 40 mol% of the steroid.
在一個實施例中，中性脂質之莫耳百分比與陰離子脂質之莫耳百分比之總和在5莫耳%至15莫耳%範圍內。在一個實施例中，中性脂質之莫耳百分比與陰離子脂質之莫耳百分比之總和在7莫耳%至12莫耳%範圍內。In one embodiment, the sum of the molar percentage of neutral lipids and the molar percentage of anionic lipids is in the range of 5 molar % to 15 molar %. In one embodiment, the sum of the molar percentage of neutral lipids and the molar percentage of anionic lipids is in the range of 7 molar % to 12 molar %.
在一個實施例中，陰離子脂質與中性脂質之莫耳比在1：1至1：10範圍內。在一個實施例中，中性脂質之莫耳百分比與類固醇之莫耳百分比之總和在35莫耳%至45莫耳%範圍內。In one embodiment, the molar ratio of anionic lipids to neutral lipids is in the range of 1:1 to 1:10. In one embodiment, the sum of the molar percentage of neutral lipids and the molar percentage of steroids is in the range of 35 mol% to 45 mol%.
在一個實施例中，脂質奈米顆粒包含：i)45莫耳%至55莫耳%之陽離子脂質；ii)5莫耳%至10莫耳%之中性脂質；iii)1莫耳%至5莫耳%之陰離子脂質；以及iv)32莫耳%至40莫耳%之類固醇。In one embodiment, the lipid nanoparticles contain: i) 45 mol% to 55 mol% of cationic lipids; ii) 5 mol% to 10 mol% of neutral lipids; iii) 1 mol% to 5 mol% of anionic lipids; and iv) 32 mol% to 40 mol% of steroids.
在一個實施例中，脂質奈米顆粒包含1.0莫耳%至2.5莫耳%之聚合物結合之脂質。在一個實施例中，聚合物結合之脂質係以約1.5莫耳%之濃度存在。In one embodiment, the lipid nanoparticles contain 1.0 mol% to 2.5 mol% of polymer-bound lipid. In one embodiment, the polymer-bound lipid is present at a concentration of about 1.5 mol%.
在一個實施例中，中性脂質以在5莫耳%至15莫耳%、7莫耳%至13莫耳%或9莫耳%至11莫耳%範圍內之濃度存在。在一個實施例中，中性脂質以約9.5莫耳%、10莫耳%或10.5莫耳%之濃度存在。在一個實施例中，陽離子脂質與中性脂質之莫耳比在約4.1：1.0至約4.9：1.0、約4.5：1.0至約4.8：1.0、或約4.7：1.0至4.8：1.0範圍內。In one embodiment, the neutral lipid is present at a concentration ranging from 5 mol% to 15 mol%, 7 mol% to 13 mol%, or 9 mol% to 11 mol%. In one embodiment, the neutral lipid is present at a concentration of about 9.5 mol%, 10 mol%, or 10.5 mol%. In one embodiment, the molar ratio of cationic lipid to neutral lipid is in the range of about 4.1:1.0 to about 4.9:1.0, about 4.5:1.0 to about 4.8:1.0, or about 4.7:1.0 to 4.8:1.0.
在一個實施例中，類固醇係膽固醇。在一個實施例中，類固醇以在39莫耳%至49莫耳%、40莫耳%至46莫耳%、40莫耳%至44莫耳%、40莫耳%至42莫耳%、42莫耳%至44莫耳%或44莫耳%至46莫耳%範圍內之濃度存在。在一個實施例中，類固醇以40莫耳%、41莫耳%、42莫耳%、43莫耳%、44莫耳%、45莫耳%或46莫耳%之濃度存在。在一個實施例中，陽離子脂質與類固醇之莫耳比在1.0：0.9至1.0：1.2、或1.0：1.0至1.0：1.2範圍內。In one embodiment, the steroid is cholesterol. In one embodiment, the steroid is present at a concentration ranging from 39 mol% to 49 mol%, 40 mol% to 46 mol%, 40 mol% to 44 mol%, 40 mol% to 42 mol%, 42 mol% to 44 mol%, or 44 mol% to 46 mol%. In one embodiment, the steroid is present at a concentration of 40 mol%, 41 mol%, 42 mol%, 43 mol%, 44 mol%, 45 mol%, or 46 mol%. In one embodiment, the molar ratio of cationic lipid to steroid is in the range of 1.0:0.9 to 1.0:1.2, or 1.0:1.0 to 1.0:1.2.
在一個實施例中，陽離子脂質與類固醇之莫耳比在5：1至1：1範圍內。In one embodiment, the molar ratio of cationic lipid to steroid is in the range of 5:1 to 1:1.
在一個實施例中，脂質奈米顆粒包含1.0莫耳%至2.5莫耳%之聚合物結合之脂質。在一個實施例中，聚合物結合之脂質係以約1.5莫耳%之濃度存在。In one embodiment, the lipid nanoparticles contain 1.0 mol% to 2.5 mol% of polymer-bound lipid. In one embodiment, the polymer-bound lipid is present at a concentration of about 1.5 mol%.
在一個實施例中，陽離子脂質與聚合物結合之脂質之莫耳比在約100：1至約20：1範圍內。在一個實施例中，陽離子脂質與聚合物結合之脂質之莫耳比在約35：1至約25：1範圍內。In one embodiment, the molar ratio of the cationic lipid to the polymer-bound lipid is in the range of about 100:1 to about 20:1. In one embodiment, the molar ratio of the cationic lipid to the polymer-bound lipid is in the range of about 35:1 to about 25:1.
在一個實施例中，脂質奈米顆粒之平均直徑在50nm至100nm或60nm至85nm範圍內。In one embodiment, the average diameter of the lipid nanoparticles is in the range of 50nm to 100nm or 60nm to 85nm.
在一個實施例中，組合物包含本文所提供之陽離子脂質、DSPC、膽固醇及PEG-脂質，以及mRNA。在一個實施例中，本文所提供之陽離子脂質、DSPC、膽固醇及PEG-脂質之莫耳比為約50：10：38.5：1.5。In one embodiment, the composition comprises the cationic lipids, DSPC, cholesterol and PEG-lipids provided herein, and mRNA. In one embodiment, the molar ratio of the cationic lipids, DSPC, cholesterol and PEG-lipids provided herein is about 50:10:38.5:1.5.
奈米顆粒組合物可設計成用於一或多種特定應用或目標。舉例而言，奈米顆粒組合物可設計成用於將治療劑及/或預防劑，諸如RNA遞送至哺乳動物體內之特定細胞、組織、器官或系統或其群組。奈米顆粒組合物之物理化學特性可經改變以增加對特定身體目標之選擇性。舉例而言，可基於不同器官之開窗大小來調整粒度。奈米顆粒組合物中所包含之治療劑及/或預防劑亦可基於一或多個所希望之遞送目標進行選擇。舉例而言，治療劑及/或預防劑可針對特定適應症、疾患、疾病或病症及/或針對遞送至特定細胞、組織、器官或系統或其群組(例如局部或特異性遞送)進行選擇。在某些實施例中，奈米顆粒組合物可包含編碼感興趣多肽之mRNA，其能夠在細胞內轉譯以產生感興趣多肽。此一組合物可設計成特異性遞送至特定器官。在某些實施例中，組合物可設計成特異性遞送至哺乳動物肝臟。Nanoparticle compositions can be designed for one or more specific applications or targets. For example, nanoparticle compositions can be designed to deliver therapeutic and/or prophylactic agents, such as RNA, to specific cells, tissues, organs or systems or groups thereof in mammals. The physicochemical properties of nanoparticle compositions can be altered to increase selectivity for specific body targets. For example, the particle size can be adjusted based on the size of the openings in different organs. The therapeutic and/or prophylactic agents contained in the nanoparticle compositions can also be selected based on one or more desired delivery targets. For example, therapeutic and/or prophylactic agents may be selected for specific indications, disorders, diseases or conditions and/or for delivery to specific cells, tissues, organs or systems or groups thereof (e.g., local or specific delivery). In certain embodiments, the nanoparticle composition may include mRNA encoding a polypeptide of interest, which is capable of being translated within a cell to produce the polypeptide of interest. Such a composition may be designed for specific delivery to a specific organ. In certain embodiments, the composition may be designed for specific delivery to the liver of a mammal.
奈米顆粒組合物中治療劑及/或預防劑之量可取決於奈米顆粒組合物之大小、組成、期望目標及/或應用，或其他特性，以及治療劑及/或預防劑之特性。舉例而言，可用於奈米顆粒組合物中之RNA之量可取決於RNA之大小、序列及其他特徵。奈米顆粒組合物中治療劑及/或預防劑及其他成分(例如脂質)之相對量亦可變化。在一些實施例中，奈米顆粒組合物中脂質組分與治療劑及/或預防劑之wt/wt比率可為約5：1至約60：1，諸如為5：1、6：1、7：1、8：1、9：1、10：1、11：1、12：1、13：1、14：1、15：1、16：1、17：1、18：1、19：1、20：1、25：1、30：1、35：1、40：1、45：1、50：1及60：1。舉例而言，脂質組分與治療劑及/或預防劑之wt/wt比率可為約10：1至約40：1。在某些實施例中，wt/wt比率為約20：1。奈米顆粒組合物中治療劑及/或預防劑之量可例如使用吸收光譜法(例如紫外-可見光譜法)量測。The amount of the therapeutic and/or prophylactic agent in the nanoparticle composition can depend on the size, composition, desired target and/or application, or other characteristics of the nanoparticle composition, as well as the characteristics of the therapeutic and/or prophylactic agent. For example, the amount of RNA that can be used in the nanoparticle composition can depend on the size, sequence, and other characteristics of the RNA. The relative amounts of the therapeutic and/or prophylactic agent and other components (e.g., lipids) in the nanoparticle composition can also vary. In some embodiments, the wt/wt ratio of the lipid component to the therapeutic and/or preventive agent in the nanoparticle composition may be about 5:1 to about 60:1, such as 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 11:1, 12:1, 13:1, 14:1, 15:1, 16:1, 17:1, 18:1, 19:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1 and 60:1. For example, the wt/wt ratio of the lipid component to the therapeutic and/or preventive agent may be about 10:1 to about 40:1. In certain embodiments, the wt/wt ratio is about 20:1. The amount of therapeutic and/or prophylactic agent in the nanoparticle composition can be measured, for example, using absorption spectroscopy (e.g., UV-Vis spectroscopy).
在一些實施例中，奈米顆粒組合物包含一或多種RNA，且可選擇一或多種RNA、脂質及其量來提供特定N：P比。組合物之N：P比係指一或多種脂質中之氮原子與RNA中磷酸酯基之數量之莫耳比。在一些實施例中，選擇較低之N：P比。可選擇一或多種RNA、脂質及其量以提供約2：1至約30：1，諸如2：1、3：1、4：1、5：1、6：1、7：1、8：1、9：1、10：1、12：1、14：1、16：1、18：1、20：1、22：1、24：1、26：1、28：1或30：1之N：P比。在某些實施例中，N：P比可為約2：1至約8：1。在其他實施例中，N：P比為約5：1至約8：1。舉例而言，N：P比可為約5.0：1、約5.5：1、約5.67：1、約6.0：1、約6.5：1或約7.0：1。舉例而言，N：P比可為約5.67：1。In some embodiments, the nanoparticle composition comprises one or more RNAs, and the one or more RNAs, lipids, and amounts thereof may be selected to provide a specific N:P ratio. The N:P ratio of the composition refers to the molar ratio of the number of nitrogen atoms in the one or more lipids to the number of phosphate groups in the RNA. In some embodiments, a lower N:P ratio is selected. The one or more RNAs, lipids, and amounts thereof may be selected to provide an N:P ratio of about 2:1 to about 30:1, such as 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 12:1, 14:1, 16:1, 18:1, 20:1, 22:1, 24:1, 26:1, 28:1, or 30:1. In certain embodiments, the N:P ratio may be about 2:1 to about 8:1. In other embodiments, the N:P ratio is about 5:1 to about 8:1. For example, the N:P ratio can be about 5.0:1, about 5.5:1, about 5.67:1, about 6.0:1, about 6.5:1, or about 7.0:1. For example, the N:P ratio can be about 5.67:1.
奈米顆粒組合物之物理特性可取決於其組分。舉例而言，包含膽固醇作為結構脂質之奈米顆粒組合物可具有與包含不同結構脂質之奈米顆粒組合物不同之特徵。類似地，奈米顆粒組合物之特徵可取決於其組分之絕對或相對量。舉例而言，包含較高莫耳分率之磷脂之奈米顆粒組合物可具有與包含較低莫耳分率之磷脂之奈米顆粒組合物不同之特徵。特徵亦可取決於奈米顆粒組合物之製備方法及條件而變化。The physical properties of a nanoparticle composition may depend on its components. For example, a nanoparticle composition comprising cholesterol as a structural lipid may have different characteristics than a nanoparticle composition comprising a different structural lipid. Similarly, the characteristics of a nanoparticle composition may depend on the absolute or relative amounts of its components. For example, a nanoparticle composition comprising a higher molar fraction of phospholipids may have different characteristics than a nanoparticle composition comprising a lower molar fraction of phospholipids. Characteristics may also vary depending on the method and conditions of preparation of the nanoparticle composition.
奈米顆粒組合物可藉由多種方法表徵。舉例而言，可使用顯微鏡檢查(例如透射電子顯微鏡檢查或掃描電子顯微鏡檢查)來檢查奈米顆粒組合物之形態及大小分佈。可使用動態光散射或電位測定法(例如電位滴定法)來量測ζ電位。動態光散射亦可用於確定粒度。亦可使用儀器，諸如Zetasizer Nano ZS(Malvem Instruments Ltd,Malvem,Worcestershire,UK)來量測奈米顆粒組合物之多個特徵，諸如粒度、多分散指數及ζ電位。Nanoparticle compositions can be characterized by a variety of methods. For example, microscopic examination (e.g., transmission electron microscopy or scanning electron microscopy) can be used to examine the morphology and size distribution of nanoparticle compositions. Zeta potential can be measured using dynamic light scattering or potentiometry (e.g., potentiometric titration). Dynamic light scattering can also be used to determine particle size. Instruments such as the Zetasizer Nano ZS (Malvem Instruments Ltd, Malvem, Worcestershire, UK) can also be used to measure various characteristics of nanoparticle compositions, such as particle size, polydispersity index, and zeta potential.
在各個實施例中，奈米顆粒組合物之平均大小可在數十奈米至數百奈米之間。舉例而言，平均大小可為約40nm至約150nm，諸如約40nm、45nm、50nm、55nm、60nm、65nm、70nm、75nm、80nm、85nm、90nm、95nm、100um、105nm、110nm、115nm、120nm、125nm、130nm、135nm、140nm、145nm或150nm。在一些實施例中，奈米顆粒組合物之平均大小可為約50nm至約100nm、約50nm至約90nm、約50nm至約80nm、約50nm至約70nm、約50nm至約60nm、約60nm至約100nm、約60nm至約90nm、約60nm至約80nm、約60nm至約70nm、約70nm至約100nm、約70nm至約90nm、約70nm至約80nm、約80nm至約100nm、約80nm至約90nm、或約90nm至約100nm。在某些實施例中，奈米顆粒組合物之平均大小可為約70nm至約100nm。在一些實施例中，平均大小可為約80nm。在其他實施例中，平均大小可為約100nm。In various embodiments, the average size of the nanoparticle composition can be between tens of nanometers and hundreds of nanometers. For example, the average size can be about 40nm to about 150nm, such as about 40nm, 45nm, 50nm, 55nm, 60nm, 65nm, 70nm, 75nm, 80nm, 85nm, 90nm, 95nm, 100um, 105nm, 110nm, 115nm, 120nm, 125nm, 130nm, 135nm, 140nm, 145nm or 150nm. In some embodiments, the nanoparticle composition may have an average size of about 50nm to about 100nm, about 50nm to about 90nm, about 50nm to about 80nm, about 50nm to about 70nm, about 50nm to about 60nm, about 60nm to about 100nm, about 60nm to about 90nm, about 60nm to about 80nm, about 60nm to about 70nm, about 70nm to about 100nm, about 70nm to about 90nm, about 70nm to about 80nm, about 80nm to about 100nm, about 80nm to about 90nm, or about 90nm to about 100nm. In certain embodiments, the nanoparticle composition may have an average size of about 70nm to about 100nm. In some embodiments, the average size may be about 80nm. In other embodiments, the average size may be about 100nm.
奈米顆粒組合物可為相對均質的。多分散指數可用於指示奈米顆粒組合物之均勻性，例如奈米顆粒組合物之粒度分佈。較小(例如小於0.3)之多分散指數一般指示較窄之粒度分佈。奈米顆粒組合物之多分散指數可為約0至約0.25，諸如0.01、0.02、0.03、0.04、0.05、0.06、0.07、0.08、0.09、0.10、0.11、0.12、0.13、0.14、0.15、0.16、0.17、0.18、0.19、0.20、0.21、0.22、0.23、0.24或0.25。在一些實施例中，奈米顆粒組合物之多分散指數可為約0.10至約0.20。The nanoparticle composition can be relatively homogeneous. The polydispersity index can be used to indicate the uniformity of the nanoparticle composition, such as the particle size distribution of the nanoparticle composition. A smaller polydispersity index (e.g., less than 0.3) generally indicates a narrower particle size distribution. The polydispersity index of the nanoparticle composition can be about 0 to about 0.25, such as 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.10, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20, 0.21, 0.22, 0.23, 0.24, or 0.25. In some embodiments, the polydispersity index of the nanoparticle composition may be from about 0.10 to about 0.20.
奈米顆粒組合物之ζ電位可用於指示組合物之動電位。舉例而言，ζ電位可闡述奈米顆粒組合物之表面電荷。具有相對較低正或負電荷之奈米顆粒組合物一般係期望的，此乃因帶較高電荷之物質可與體內之細胞、組織及其他成分發生不期望之相互作用。在一些實施例中，奈米顆粒組合物之ζ電位可為約-10mV至約+20mV、約-10mV至約+15mV、約-10mV至約+10mV、約-10mV至約+5mV、約-10mV至約0mV、約-10mV至約-5mV、約-5mV至約+20mV、約-5mV至約+15mV、約-5mV至約+10mV、約-5mV至約+5mV、約-5mV至約0mV、約0mV至約+20mV、約0mV至約+15mV、約0mV至約+10mV、約0mV至約+5mV、約+5mV至約+20mV、約+5mV至約+15mV、或約+5mV至約+10mV。The zeta potential of a nanoparticle composition can be used to indicate the zeta potential of the composition. For example, the zeta potential can describe the surface charge of the nanoparticle composition. Nanoparticle compositions with relatively low positive or negative charges are generally desirable because substances with higher charges can interact undesirably with cells, tissues, and other components in the body. In some embodiments, the zeta potential of the nanoparticle composition can be about -10 mV to about +20 mV, about -10 mV to about +15 mV, about -10 mV to about +10 mV, about -10 mV to about +5 mV, about -10 mV to about 0 mV, about -10 mV to about -5 mV, about -5 mV to about +20 mV, about -5 mV to about +15 mV, about -10 mV to about +10 mV, about -10 mV to about +5 mV, about -10 mV to about 0 mV, about -10 mV to about -5 mV, about -5 mV to about +20 mV, about -5 mV to about +15 mV, about -10 mV to about 0 mV, about -10 mV to about -5 mV, about -5 mV to about +20 mV, about -5 mV to about +15 mV, about -10 mV to about 0 mV, about -10 mV to about -5 mV, about -5 mV to about +20 mV, about -5 mV to about +15 mV, about -10 mV to about V, about -5mV to about +10mV, about -5mV to about +5mV, about -5mV to about 0mV, about 0mV to about +20mV, about 0mV to about +15mV, about 0mV to about +10mV, about 0mV to about +5mV, about +5mV to about +20mV, about +5mV to about +15mV, or about +5mV to about +10mV.
治療劑及/或預防劑之包封效率闡述相對於所提供之初始量，在製備後經奈米顆粒組合物包封或以其他方式與奈米顆粒組合物締合之治療劑及/或預防劑的量。包封效率期望地較高(例如接近100%)。包封效率可例如藉由比較在用一或多種有機溶劑或清潔劑破壞奈米顆粒組合物之前與之後含有奈米顆粒組合物之溶液中治療劑及/或預防劑的量來量測。螢光可用於量測溶液中游離治療劑及/或預防劑(例如RNA)的量。對於本文所述之奈米顆粒組合物，治療劑及/或預防劑之包封效率可為至少50%，例如50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%或100%。在一些實施例中，包封效率可為至少80%。在某些實施例中，包封效率可為至少90%。The encapsulation efficiency of the therapeutic and/or prophylactic agent describes the amount of the therapeutic and/or prophylactic agent encapsulated or otherwise associated with the nanoparticle composition after preparation relative to the initial amount provided. The encapsulation efficiency is desirably high (e.g., close to 100%). The encapsulation efficiency can be measured, for example, by comparing the amount of the therapeutic and/or prophylactic agent in a solution containing the nanoparticle composition before and after the nanoparticle composition is destroyed with one or more organic solvents or detergents. Fluorescence can be used to measure the amount of free therapeutic and/or prophylactic agent (e.g., RNA) in a solution. For the nanoparticle compositions described herein, the encapsulation efficiency of the therapeutic and/or prophylactic agent may be at least 50%, such as 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100%. In some embodiments, the encapsulation efficiency may be at least 80%. In certain embodiments, the encapsulation efficiency may be at least 90%.
奈米顆粒組合物可視情況包含一或多種包衣。舉例而言，可將奈米顆粒組合物調配成具有包衣之膠囊、膜片或錠劑。包含本文所述組合物之膠囊、膜片或錠劑可具有任何有用之大小、抗拉強度、硬度或密度。The nanoparticle composition may optionally include one or more coatings. For example, the nanoparticle composition may be formulated into a capsule, film, or tablet having a coating. The capsule, film, or tablet containing the composition described herein may have any useful size, tensile strength, hardness, or density.
5.6醫藥組合物5.6Pharmaceutical compositions
根據本揭示案，奈米顆粒組合物可整體或部分地調配成醫藥組合物。醫藥組合物可包含一或多種奈米顆粒組合物。舉例而言，醫藥組合物可包含一或多種奈米顆粒組合物，該一或多種奈米顆粒組合物包含一或多種不同之治療劑及/或預防劑。醫藥組合物可進一步包含一或多種醫藥學上可接受之賦形劑或輔助成分，諸如本文所述者。關於醫藥組合物及劑之調配及製造之一般準則可見於例如Remington，The Science and Practice of Pharmacy，第21版，A.R.Gennaro；Lippincott,Williams & Wilkins,Baltimore,Md.,2006。習用賦形劑及輔助成分可用於任何醫藥組合物中，除非任何習用賦形劑或輔助成分與奈米顆粒組合物之一或多種組分不相容。若賦形劑或輔助成分與奈米顆粒組合物之組分之組合會導致任何不希望之生物作用或其他有害作用，則賦形劑或輔助成分與奈米顆粒組合物之組分不相容。According to the present disclosure, the nanoparticle composition can be formulated as a pharmaceutical composition in whole or in part. The pharmaceutical composition can include one or more nanoparticle compositions. For example, the pharmaceutical composition can include one or more nanoparticle compositions, and the one or more nanoparticle compositions include one or more different therapeutic agents and/or preventive agents. The pharmaceutical composition can further include one or more pharmaceutically acceptable excipients or auxiliary ingredients, such as those described herein. General guidelines for the formulation and manufacture of pharmaceutical compositions and agents can be found in, for example, Remington, The Science and Practice of Pharmacy, 21st edition, A.R.Gennaro; Lippincott, Williams & Wilkins, Baltimore, Md., 2006. Customary excipients and auxiliary ingredients may be used in any pharmaceutical composition, unless any customary excipient or auxiliary ingredient is incompatible with one or more components of the nanoparticle composition. Excipients or auxiliary ingredients are incompatible with components of the nanoparticle composition if their combination would result in any undesirable biological or other adverse effects.
在一些實施例中，該一或多種賦形劑或輔助成分可構成包含奈米顆粒組合物之醫藥組合物之總質量或體積之超過50%。舉例而言，該一或多種賦形劑或輔助成分可構成醫藥組合物之50%、60%、70%、80%、90%或更高百分比。在一些實施例中，醫藥學上可接受之賦形劑為至少95%、至少96%、至少97%、至少98%、至少99%或100%純。在一些實施例中，賦形劑經批準用於人類及獸醫用途。在一些實施例中，賦形劑得到美國食品與藥物管理局批準。在一些實施例中，賦形劑係醫藥級的。在一些實施例中，賦形劑符合美國藥典(USP)、歐洲藥典(EP)、英國藥典及/或國際藥典之標準。In some embodiments, the one or more excipients or adjuncts may constitute more than 50% of the total mass or volume of the pharmaceutical composition comprising the nanoparticle composition. For example, the one or more excipients or adjuncts may constitute 50%, 60%, 70%, 80%, 90% or more of the pharmaceutical composition. In some embodiments, the pharmaceutically acceptable excipient is at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% pure. In some embodiments, the excipient is approved for human and veterinary use. In some embodiments, the excipient is approved by the U.S. Food and Drug Administration. In some embodiments, the excipient is pharmaceutical grade. In some embodiments, the excipient complies with the standards of the United States Pharmacopoeia (USP), the European Pharmacopoeia (EP), the British Pharmacopoeia and/or the International Pharmacopoeia.
根據本揭示案之醫藥組合物中之一或多種奈米顆粒組合物、一或多種醫藥學上可接受之賦形劑及/或任何額外成分之相對量將取決於所治療個體之身分、體格及/或狀況且進一步取決於組合物之投與途徑而變化。舉例而言，醫藥組合物可包含在0.1%與100%(wt/wt)之間之一或多種奈米顆粒組合物。The relative amounts of one or more nanoparticle compositions, one or more pharmaceutically acceptable excipients, and/or any additional ingredients in a pharmaceutical composition according to the present disclosure will vary depending on the identity, size, and/or condition of the individual being treated and further on the route of administration of the composition. For example, a pharmaceutical composition may contain between 0.1% and 100% (wt/wt) of one or more nanoparticle compositions.
在某些實施例中，本揭示案之奈米顆粒組合物及/或醫藥組合物經冷藏或冷凍儲存及/或運輸(例如在4℃或更低溫度下，諸如在約-150℃與約0℃之間或在約-80℃與約-20℃之間(例如約-5℃、-10℃、-15℃、-20℃、-25℃、-30℃、-40℃、-50℃、-60℃、-70℃、-80℃、-90℃、-130℃或-150℃)之溫度下儲存)。舉例而言，包含式(I)(及其子式)中任一個之化合物之醫藥組合物係在例如約-20℃、30℃、-40℃、-50℃、-60℃、-70℃或-80℃下冷藏儲存及/或運輸之溶液。在某些實施例中，本揭示案亦係關於一種藉由在4℃或更低溫度下，諸如在約-150℃與約0℃之間或在約-80℃與約-20℃之間之溫度下，例如在約-5℃、-10℃、-15℃、-20℃、-25℃、-30℃、-40℃、-50℃、-60℃、-70℃、-80℃、-90℃、-130℃或-150℃溫度下儲存包含式(I)(及其子式)中任一個之化合物之奈米顆粒組合物及/或醫藥組合物來增加該奈米顆粒組合物及/或醫藥組合物之穩定性之方法。舉例而言，本文所揭示之奈米顆粒組合物及/或醫藥組合物在例如4℃或更低(例如約4℃與-20℃之間)之溫度下穩定保持約至少1週、至少2週、至少3週、至少4週、至少5週、至少6週、至少1個月、至少2個月、至少4個月、至少6個月、至少8個月、至少10個月、至少12個月、至少14個月、至少16個月、至少18個月、至少20個月、至少22個月或至少24個月。在一個實施例中，調配物在約4℃下穩定保持至少4週。在某些實施例中，本揭示案之醫藥組合物包含本文所揭示之奈米顆粒組合物及醫藥學上可接受之載劑，該載劑選自以下中之一或多種：Tris、乙酸鹽(例如乙酸鈉)、檸檬酸鹽(例如檸檬酸鈉)、鹽水、PBS及蔗糖。在某些實施例中，本揭示案之醫藥組合物之pH值在約7與8之間(例如6.8、6.9、7.0、7.1、7.2、7.3、7.4、7.5、7.6、7.7、7.8、7.9或8.0，或在7.5與8之間或在7與7.8之間)。舉例而言，本揭示案之醫藥組合物包含本文所揭示之奈米顆粒組合物、Tris、鹽水及蔗糖，且具有約7.5-8之pH值，其適於在例如約-20℃下儲存及/或運輸。舉例而言，本揭示案之醫藥組合物包含本文所揭示之奈米顆粒組合物及PBS，且具有約7-7.8之pH值，其適於在例如約4℃或更低溫度下儲存及/或運輸。在本揭示案之上下文中，「穩定性」、「穩定化」及「穩定的」係指本文所揭示之奈米顆粒組合物及/或醫藥組合物在給定製造、製備、轉運、儲存及/或使用條件下，例如當施加應力，諸如剪切力、冷凍/解凍應力等時，對化學或物理變化(例如降解、粒度變化、聚集、包封之變化等)具有抗性。In certain embodiments, the nanoparticle compositions and/or pharmaceutical compositions of the present disclosure are stored and/or transported by refrigeration or freezing (e.g., at 4°C or lower, such as between about -150°C and about 0°C, or between about -80°C and about -20°C (e.g., about -5°C, -10°C, -15°C, -20°C, -25°C, -30°C, -40°C, -50°C, -60°C, -70°C, -80°C, -90°C, -130°C, or -150°C)). For example, a pharmaceutical composition comprising a compound of any one of formula (I) (and its subformulae) is a solution for storage and/or transportation under refrigeration at, for example, about -20°C, 30°C, -40°C, -50°C, -60°C, -70°C or -80°C. In certain embodiments, the disclosure also relates to a method of increasing the stability of a nanoparticle composition and/or a pharmaceutical composition comprising a compound of any one of formula (I) (and subformulae thereof) by storing the nanoparticle composition and/or a pharmaceutical composition at 4°C or lower, such as between about -150°C and about 0°C or between about -80°C and about -20°C, for example, at about -5°C, -10°C, -15°C, -20°C, -25°C, -30°C, -40°C, -50°C, -60°C, -70°C, -80°C, -90°C, -130°C or -150°C. For example, the nanoparticle compositions and/or pharmaceutical compositions disclosed herein are stable at a temperature of, for example, 4° C. or lower (e.g., between about 4° C. and -20° C.) for about at least 1 week, at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 5 weeks, at least 6 weeks, at least 1 month, at least 2 months, at least 4 months, at least 6 months, at least 8 months, at least 10 months, at least 12 months, at least 14 months, at least 16 months, at least 18 months, at least 20 months, at least 22 months, or at least 24 months. In one embodiment, the formulation is stable for at least 4 weeks at about 4° C. In certain embodiments, the pharmaceutical composition of the present disclosure comprises the nanoparticle composition disclosed herein and a pharmaceutically acceptable carrier, which is selected from one or more of the following: Tris, acetate (e.g., sodium acetate), citrate (e.g., sodium citrate), saline, PBS, and sucrose. In certain embodiments, the pH of the pharmaceutical composition of the present disclosure is between about 7 and 8 (e.g., 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8.0, or between 7.5 and 8, or between 7 and 7.8). For example, the pharmaceutical composition of the present disclosure comprises the nanoparticle composition disclosed herein, Tris, saline and sucrose, and has a pH value of about 7.5-8, which is suitable for storage and/or transportation at, for example, about -20°C. For example, the pharmaceutical composition of the present disclosure comprises the nanoparticle composition disclosed herein and PBS, and has a pH value of about 7-7.8, which is suitable for storage and/or transportation at, for example, about 4°C or lower. In the context of the present disclosure, "stability", "stabilization" and "stable" refer to the nanoparticle compositions and/or pharmaceutical compositions disclosed herein being resistant to chemical or physical changes (e.g., degradation, particle size changes, aggregation, changes in encapsulation, etc.) under given manufacturing, preparation, transportation, storage and/or use conditions, such as when stresses such as shear stress, freeze/thaw stress, etc. are applied.
可將奈米顆粒組合物及/或包含一或多種奈米顆粒組合物之醫藥組合物投與任何患者或個體，包括可受益於藉由將治療劑及/或預防劑遞送至一或多種特定細胞、組織、器官或系統或其群組，諸如腎臟系統所提供之治療作用之患者或個體。儘管本文所提供之關於奈米顆粒組合物及包含奈米顆粒組合物之醫藥組合物之闡述主要針對適於投與人類之組合物，但熟習此項技術者應理解，此等組合物一般適於投與任何其他哺乳動物。為了使組合物適於投與各種動物而對適於投與人類之組合物之改進係眾所周知的，且有普通技術之獸醫藥理學家僅藉由普通實驗(若有的話)即可設計及/或進行此等改進。經考慮，投與該等組合物之個體包括但不限於人類、其他靈長類動物及其他哺乳動物，包括商業上相關之哺乳動物，諸如牛、豬、馬、綿羊、貓、狗、小鼠及/或大鼠。Nanoparticle compositions and/or pharmaceutical compositions comprising one or more nanoparticle compositions can be administered to any patient or individual, including patients or individuals who can benefit from the therapeutic effect provided by delivering therapeutic and/or prophylactic agents to one or more specific cells, tissues, organs or systems or groups thereof, such as the renal system. Although the description of nanoparticle compositions and pharmaceutical compositions comprising nanoparticle compositions provided herein is primarily directed to compositions suitable for administration to humans, those skilled in the art will understand that such compositions are generally suitable for administration to any other mammal. Modifications of compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals are well known and can be designed and/or performed by a veterinary pharmacologist of ordinary skill with no more than ordinary experimentation, if any. It is contemplated that subjects to whom such compositions may be administered include, but are not limited to, humans, other primates, and other mammals, including commercially relevant mammals such as cattle, pigs, horses, sheep, cats, dogs, mice, and/or rats.
包含一或多種奈米顆粒組合物之醫藥組合物可藉由藥理學領域中已知或以後將開發之任何方法製備。一般而言，此等製備方法包括使活性成分與賦形劑及/或一或多種其他輔助成分結合，且接著，若需要或必要，則將產物分成、成型成及/或包裝成所希望之單劑量或多劑量單元。Pharmaceutical compositions containing one or more nanoparticle compositions can be prepared by any method known in the art of pharmacology or to be developed later.Generally speaking, such preparation methods include combining the active ingredient with a formulator and/or one or more other auxiliary ingredients, and then, if necessary or desirable, dividing, shaping and/or packaging the product into the desired single dose or multi-dose units.
根據本揭示案之醫藥組合物可以散裝、作為單次單位劑量及/或作為多個單次單位劑量製備、包裝及/或出售。如本文所使用，「單位劑量」係包含預定量之活性成分(例如奈米顆粒組合物)之醫藥組合物的離散量。活性成分的量一般等於將被投與個體之活性成分之劑量及/或此等劑量之便利部分，諸如此等劑量之一半或三分之一。The pharmaceutical compositions according to the present disclosure may be prepared, packaged and/or sold in bulk, as a single unit dose and/or as multiple single unit doses. As used herein, a "unit dose" is a discrete amount of a pharmaceutical composition containing a predetermined amount of an active ingredient (e.g., a nanoparticle composition). The amount of the active ingredient is generally equal to the dose of the active ingredient to be administered to an individual and/or a convenient fraction of such a dose, such as one-half or one-third of such a dose.
醫藥組合物可製備成適合多種投與途徑及方法之多種形式。舉例而言，醫藥組合物可製備成液體劑型(例如乳液、微乳液、奈米乳液、溶液、懸浮液、糖漿及酏劑)、可注射形式、固體劑型(例如膠囊、錠劑、丸劑、粉末及顆粒劑)、用於局部及/或經皮投與之劑型(例如軟膏、糊劑、乳膏、洗劑、凝膠劑、粉末、溶液、噴霧劑、吸入劑及貼片)、懸浮液、粉末及其他形式。Pharmaceutical compositions can be prepared in a variety of forms suitable for a variety of routes and methods of administration. For example, pharmaceutical compositions can be prepared in liquid dosage forms (e.g., emulsions, microemulsions, nanoemulsions, solutions, suspensions, syrups, and elixirs), injectable forms, solid dosage forms (e.g., capsules, tablets, pills, powders, and granules), dosage forms for topical and/or transdermal administration (e.g., ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants, and patches), suspensions, powders, and other forms.
經口及非經腸投與之液體劑型包括但不限於醫藥學上可接受之乳液、微乳液、奈米乳液、溶液、懸浮液、糖漿及/或酏劑。除活性成分外，液體劑型亦可包含此項技術中常用之惰性稀釋劑，例如水或其他溶劑、增溶劑及乳化劑，諸如乙醇、異丙醇、碳酸乙酯、乙酸乙酯、苯甲醇、苯甲酸苯甲酯、丙二醇、1,3-丁二醇、二甲基甲醯胺、油類(尤其為棉籽油、花生油、玉米油、胚芽油、橄欖油、蓖麻油及芝麻油)、甘油、四氫糠醇、聚乙二醇及去水山梨醇脂肪酸酯，以及其混合物。除惰性稀釋劑外，經口組合物亦可包含額外治療劑及/或預防劑、額外劑，諸如潤濕劑、乳化劑及懸浮劑、甜味劑、調味劑及/或加香劑。在供非經腸投與之某些實施例中，將組合物與增溶劑混合，該等增溶劑諸如為CremophorTM、醇、油、改質油、二醇、聚山梨醇酯、環糊精、聚合物及/或其組合。Liquid dosage forms for oral and parenteral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, nanoemulsions, solutions, suspensions, syrups and/or elixirs. In addition to the active ingredient, the liquid dosage form may also contain inert diluents commonly used in this art, such as water or other solvents, solubilizers and emulsifiers, such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil and sesame oil), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol and sorbitan fatty acid esters, and mixtures thereof. In addition to inert diluents, oral compositions may also contain additional therapeutic and/or prophylactic agents, excipients such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents and/or perfuming agents. In certain embodiments for parenteral administration, the composition is mixed with a solubilizing agent such as Cremophor™ , alcohols, oils, modified oils, glycols, polysorbates, cyclodextrins, polymers and/or combinations thereof.
可注射製劑，例如無菌可注射水性或油性懸浮液，可根據已知技術，使用適合分散劑、潤濕劑及/或助懸劑來調配。無菌可注射製劑可為在無毒非經腸可接受之稀釋劑及/或溶劑中之無菌可注射溶液、懸浮液及/或乳液，例如在1,3-丁二醇中之溶液。可以使用之可接受之媒劑及溶劑包括水、林格氏溶液(Ringer’s solution)、USP及等張氯化鈉溶液。無菌不揮發性油通常用作溶劑或懸浮介質。為此，可以使用任何溫和的不揮發性油，包括合成甘油單酯或甘油二酯。脂肪酸諸如油酸可用於製備注射劑。Injectable preparations, such as sterile injectable aqueous or oily suspensions, can be prepared according to known techniques using suitable dispersants, wetting agents and/or suspending agents. Sterile injectable preparations can be sterile injectable solutions, suspensions and/or emulsions in non-toxic parenterally acceptable diluents and/or solvents, such as solutions in 1,3-butanediol. Acceptable vehicles and solvents that can be used include water, Ringer’s solution, USP and isotonic sodium chloride solution. Sterile, non-volatile oils are generally used as solvents or suspending media. For this purpose, any bland, non-volatile oil may be used, including synthetic mono- or diglycerides. Fatty acids such as oleic acid can be used to prepare injectables.
可注射調配物可經滅菌，例如藉由濾過細菌截留過濾器過濾，及/或藉由摻入呈無菌固體組合物形式之滅菌劑來滅菌，該滅菌劑可在使用前溶解或分散於無菌水或其他無菌可注射介質中。The injectable formulation can be sterilized, for example, by filtration through a bacteria-retaining filter, and/or by incorporating a sterilizing agent in the form of a sterile solid composition that can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.
本揭示案之特徵在於向哺乳動物細胞或器官遞送治療劑及/或預防劑，在哺乳動物細胞中產生感興趣多肽，以及治療有需要之哺乳動物之疾病或病症之方法，該等方法包括向哺乳動物投與包含治療劑及/或預防劑之奈米顆粒組合物及/或使哺乳動物細胞與該奈米顆粒組合物接觸。The present disclosure features methods for delivering therapeutic and/or preventive agents to mammalian cells or organs, producing polypeptides of interest in mammalian cells, and treating diseases or conditions in mammals in need thereof, the methods comprising administering to a mammal a nanoparticle composition comprising a therapeutic and/or preventive agent and/or contacting a mammalian cell with the nanoparticle composition.
6.實例6. Examples
本節中之實例僅作為示例提供，而並非作為限制。The examples in this section are provided by way of illustration only and not by way of limitation.
一般方法.General approach.
一般製備型HPLC方法：HPLC純化係在配備有二極體陣列偵測器(DAD)之Waters 2767上，在Inertsil Pre-C8 OBD管柱上，一般利用含0.1% TFA之水作為溶劑A且利用乙腈作為溶劑B進行。General preparative HPLC method: HPLC purification was performed on a Waters 2767 equipped with a diode array detector (DAD) on an Inertsil Pre-C8 OBD column, typically using water containing 0.1% TFA as solvent A and acetonitrile as solvent B.
一般LCMS方法：LCMS分析係在Shimadzu(LC-MS2020)系統上進行。層析係在SunFire C18上，一般利用含0.1%甲酸之水作為溶劑A且利用含0.1%甲酸之乙腈作為溶劑B執行。General LCMS method: LCMS analysis was performed on a Shimadzu (LC-MS2020) system. Chromatography was performed on a SunFire C18, typically using water containing 0.1% formic acid as solvent A and acetonitrile containing 0.1% formic acid as solvent B.
6.1 實例1：製備起始原料及中間體.6.1 Example 1: Preparation of starting materials and intermediates.
製備化合物APreparation of Compound A
向2-己基癸-1-醇(2.0g，8.33mmol，1.0eq.)及6-溴己酸(2.0g，10.0mmol，1.2eq.)在30mL二氯甲烷中之溶液中加入二異丙基乙胺(2.7g，2.08mmol，2.5eq.)及DMAP(203mg，1.67mmol，0.2eq.)。在環境溫度下攪拌5分鐘後，加入EDCI(2.4g，12.5mmol，1.5eq.)，且將反應混合物在室溫下攪拌隔夜，之後TLC顯示起始醇完全消失。用CH2Cl2(300mL)稀釋反應混合物，且用飽和NaHCO3(100mL)、水(100mL)及鹽水(100mL)洗滌。合併之有機層經Na2SO4乾燥，真空移除溶劑。蒸發溶劑，得到粗產物，藉由管柱層析法(矽膠，己烷中之0-1%乙酸乙酯(EA))純化，得到呈無色油狀之化合物A(2.0g，57%)。To a solution of 2-hexyldecan-1-ol (2.0 g, 8.33 mmol, 1.0 eq.) and 6-bromohexanoic acid (2.0 g, 10.0 mmol, 1.2 eq.) in 30 mL of dichloromethane were added diisopropylethylamine (2.7 g, 2.08 mmol, 2.5 eq.) and DMAP (203 mg, 1.67 mmol, 0.2 eq.). After stirring at ambient temperature for 5 minutes, EDCI (2.4 g, 12.5 mmol, 1.5 eq.) was added, and the reaction mixture was stirred at room temperature overnight, after which TLC showed complete disappearance of the starting alcohol. The reaction mixture was diluted with CH2 Cl2 (300 mL) and washed with saturated NaHCO3 (100 mL), water (100 mL), and brine (100 mL). The combined organic layers were dried overNa2SO4and the solvent was removed in vacuo. The solvent was evaporated to give a crude product, which was purified by column chromatography (silica gel, 0-1% ethyl acetate (EA) in hexanes) to give Compound A (2.0 g, 57%) as a colorless oil.
製備化合物BPreparation of Compound B
在室溫、氬氣下將環己酮(2.0g，20.0mmol，1.0eq.)、異丙醇鈦(IV)(7.4g，26mmol，1.3eq.)及2-胺基乙醇(3.66g，60.0mmol，3.0eq.)在甲醇(10.0mL)中之混合物攪拌5小時。接著在0℃下加入硼氫化鈉(760.0mg，20.0mmol，1.0eq.)，將所得混合物再攪拌2小時。接著加入水(10.0mL)淬滅反應。室溫下繼續攪拌20分鐘，接著用鹽酸(1M，5mL)酸化反應混合物，用矽藻土墊過濾，用水及EA洗滌。分離有機層，經Na2SO4乾燥，減壓蒸發，且藉由急速管柱層析法(FCC)(PE/EA=5/1-0/1)純化，得到呈黃色油狀之化合物B(1.5g，52%產率)。A mixture of cyclohexanone (2.0 g, 20.0 mmol, 1.0 eq.), titanium (IV) isopropoxide (7.4 g, 26 mmol, 1.3 eq.) and 2-aminoethanol (3.66 g, 60.0 mmol, 3.0 eq.) in methanol (10.0 mL) was stirred at room temperature under an atmosphere of hydrogen for 5 hours. Sodium borohydride (760.0 mg, 20.0 mmol, 1.0 eq.) was then added at 0°C and the resulting mixture was stirred for another 2 hours. Water (10.0 mL) was then added to quench the reaction. Stirring was continued at room temperature for 20 minutes, and the reaction mixture was then acidified with hydrochloric acid (1 M, 5 mL), filtered through a celite pad, and washed with water and EA. The organic layer was separated, dried over Na2 SO4 , evaporated under reduced pressure, and purified by flash column chromatography (FCC) (PE/EA=5/1-0/1) to give compound B (1.5 g, 52% yield) as a yellow oil.
製備化合物CPreparation of Compound C
在室溫下，向化合物A(446.0mg，1.0mmol，1.0eq.)及乙醇胺(180.0mg，3.0mmol，3.0eq.)在乙腈(ACN，10.0mL)中之溶液中加入Cs2CO3(97.5mg，0.3mmol，0.3eq.)、K2CO3(414.0mg，3.0mmol，3.0eq.)及NaI(14.6mg，0.1mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈黃色油狀之化合物C(0.35g，82%產率)。To a solution of compound A (446.0 mg, 1.0 mmol, 1.0 eq.) and ethanolamine (180.0 mg, 3.0 mmol, 3.0 eq.) in acetonitrile (ACN, 10.0 mL) were added Cs2 CO3 (97.5 mg, 0.3 mmol, 0.3 eq.), K2 CO3 (414.0 mg, 3.0 mmol, 3.0 eq.) and NaI (14.6 mg, 0.1 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was complete, the mixture was evaporated under reduced pressure, and purified by FCC (DCM/MeOH=1/0-20/1) to give compound C (0.35 g, 82% yield) as a yellow oil.
製備化合物DPreparation of Compound D
在氬氣、室溫下將環丁酮(8.0g，114mol，1.0eq.)及2-胺基乙醇(20.9g，342mol，3.0eq.)在甲醇(100mL)中之混合物攪拌16小時。接著在0℃下加入硼氫化鈉(4.3g，114mmol，1.0eq.)，將所得混合物再攪拌16小時。接著將反應混合物減壓濃縮。加入水(200mL)，用二氯甲烷(DCM)萃取。合併之有機層經Na2SO4乾燥，減壓蒸發，且藉由管柱層析法(矽膠，DCM中之2%-10% MeOH)純化，得到呈淺黃色油狀之化合物D(3.9g，30%產率)。A mixture of cyclobutanone (8.0 g, 114 mol, 1.0 eq.) and 2-aminoethanol (20.9 g, 342 mol, 3.0 eq.) in methanol (100 mL) was stirred for 16 hours under argon at room temperature. Then sodium borohydride (4.3 g, 114 mmol, 1.0 eq.) was added at 0°C and the resulting mixture was stirred for another 16 hours. The reaction mixture was then concentrated under reduced pressure. Water (200 mL) was added and extracted with dichloromethane (DCM). The combined organic layers were dried overNa2SO4 , evaporated under reduced pressure, and purified by column chromatography (silica gel, 2%-10% MeOH in DCM) to give Compound D (3.9 g, 30% yield) asa light yellow oil.
製備化合物EPreparation of Compound E
步驟1：製備化合物E-1Step 1: Preparation of compound E-1
向PMB-NH2(5.166g，37.66mmol，4.0eq.)在EtOH(30mL)中之溶液中加入1,2-環氧十四烷(2.0g，9.416mmol，1.0eq.)。將反應混合物在室溫下攪拌16小時。LCMS顯示反應完成。將混合物減壓蒸發，且藉由FCC純化，得到呈白色固體狀之化合物E-1(1.42g，43.09%)。LCMS：Rt：0.815min；MS m/z(ESI)：350.3[M+H]+。To a solution of PMB-NH2 (5.166 g, 37.66 mmol, 4.0 eq.) in EtOH (30 mL) was added 1,2-epoxytetradecane (2.0 g, 9.416 mmol, 1.0 eq.). The reaction mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete. The mixture was evaporated under reduced pressure and purified by FCC to give compound E-1 (1.42 g, 43.09%) as a white solid. LCMS: Rt: 0.815 min; MS m/z (ESI): 350.3 [M+H]+ .
步驟2：製備化合物E-2Step 2: Preparation of compound E-2
向化合物E-1(1.42g，4.057mmol，1.0eq.)在ACN(25mL)中之溶液中加入化合物A(5.106mg，12.17mmol，3.0eq.)、K2CO3(1.668g，12.17mmol，3.0eq.)、Cs2CO3(397mg，1.217mmol，0.3eq.)及NaI(30mg，0.2029mmol，0.05eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑並藉由FCC純化，得到呈無色油狀之化合物E-2(2.5g，89.55%)。LCMS：Rt：0.241min；MS m/z(ESI)：688.5[M+H]+。To a solution of compound E-1 (1.42 g, 4.057 mmol, 1.0 eq.) in ACN (25 mL) were added compound A (5.106 mg, 12.17 mmol, 3.0 eq.), K2 CO3 (1.668 g, 12.17 mmol, 3.0 eq.), Cs2 CO3 (397 mg, 1.217 mmol, 0.3 eq.) and NaI (30 mg, 0.2029 mmol, 0.05 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and purified by FCC to give compound E-2 (2.5 g, 89.55%) as a colorless oil. LCMS: Rt: 0.241 min; MS m/z (ESI): 688.5 [M+H]+ .
步驟3：製備化合物EStep 3: Preparation of Compound E
向化合物E-2(250mg，0.3633mmol)在MeOH(10mL)中之溶液中加入Pd/C(50mg)。將反應混合物在室溫、H2下攪拌16小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化，得到呈無色油狀之化合物E(105mg，50.88%產率)。To a solution of compound E-2 (250 mg, 0.3633 mmol) in MeOH (10 mL) was added Pd/C (50 mg). The reaction mixture was stirred at room temperature underH2 for 16 h. LCMS showed the reaction was complete. After removal of the solvent, purification by preparative HPLC gave compound E (105 mg, 50.88% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)：3.97(d,J=6Hz,2H),3.58(s,1H),2.73-2.58(m,3H),2.45-2.40(m,1H),2.33-2.29(m,2H),1.66-1.60(m,2H),1.51-1.40(m,2H),1.39-1.34(m,4H),1.26(s,46H),0.90-0.86(m,9H)。LCMS：Rt：1.083min；MS m/z(ESI)：568.5[M+H]+。1 H NMR (400MHz, CDCl3 ): 3.97(d,J=6Hz,2H),3.58(s,1H),2.73-2.58(m,3H),2.45-2.40(m,1H),2.33-2.29(m,2H) ,1.66-1.60(m,2H),1.51-1.40(m,2H),1.39-1.34(m,4H),1.26(s,46H),0.90-0.86(m,9H). LCMS: Rt: 1.083min; MS m/z (ESI): 568.5[M+H]+ .
製備化合物FPreparation of Compound F
向環丙胺(5.7g，100mmol，2.5eq.)在EtOH(50mL)中之混合物中加入2-溴乙醇(5g，40mmol，1eq.)。將反應混合物在50℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，得到呈黃色油狀之化合物F(6.6g粗品)。2-Bromoethanol (5 g, 40 mmol, 1 eq.) was added to a mixture of cyclopropylamine (5.7 g, 100 mmol, 2.5 eq.) in EtOH (50 mL). The reaction mixture was stirred at 50 °C for 16 h. LCMS showed that the reaction was complete. The solvent was removed to give Compound F (6.6 g crude product) as a yellow oil.
製備化合物GPreparation of Compound G
將環戊酮(16.8g，200mmol，1eq.)及2-胺基乙醇(13.4g，220mmol，1.1eq.)與3滴乙酸(AcOH)在MeOH(300mL)中之溶液在室溫下攪拌隔夜，接著在0℃下向混合物中加入NaBH4(8.4g，220mmol，1.1eq.)。將混合物在室溫下攪拌2小時。混合物用水(100mL)淬滅，用EA(3×100mL)萃取，乾燥，濃縮。藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之化合物G(17.8g，49.0%產率)。A solution of cyclopentanone (16.8 g, 200 mmol, 1 eq.) and 2-aminoethanol (13.4 g, 220 mmol, 1.1 eq.) in MeOH (300 mL) was stirred at room temperature overnight, and then NaBH4 (8.4 g, 220 mmol, 1.1 eq.) was added to the mixture at 0°C. The mixture was stirred at room temperature for 2 hours. The mixture was quenched with water (100 mL), extracted with EA (3×100 mL), dried, and concentrated. Purification by silica gel column chromatography (MeOH:DCM=0% to 10%) gave Compound G (17.8 g, 49.0% yield) as a yellow oil.
製備化合物HPreparation of Compound H
向2-辛基癸-1-醇(1.5g，5.545mmol，1.0eq.)在DCM(15mL)中之溶液中加入6-溴己酸(1.3g，6.654mmol，1.2eq.)、EDCI(1.6g，8.318mmol，1.5eq.)、DMAP(135mg，1.109mmol，0.2eq.)及二異丙基乙胺(DIEA，1.4g，11.09mmol，2.0eq.)。將反應混合物在50℃下攪拌16小時。TLC顯示反應完成。移除溶劑，且藉由FCC純化粗產物，得到呈黃色油狀之化合物H(1.2g，48.36%)。To a solution of 2-octyldecan-1-ol (1.5 g, 5.545 mmol, 1.0 eq.) in DCM (15 mL) were added 6-bromohexanoic acid (1.3 g, 6.654 mmol, 1.2 eq.), EDCI (1.6 g, 8.318 mmol, 1.5 eq.), DMAP (135 mg, 1.109 mmol, 0.2 eq.) and diisopropylethylamine (DIEA, 1.4 g, 11.09 mmol, 2.0 eq.). The reaction mixture was stirred at 50 °C for 16 hours. TLC showed that the reaction was complete. The solvent was removed and the crude product was purified by FCC to give Compound H (1.2 g, 48.36%) as a yellow oil.
製備化合物KPreparation of Compound K
將環庚酮(15g，134mmol，1eq.)及2-胺基乙醇(9g，147mmol，1.1eq.)與3滴AcOH在MeOH(250mL)中之混合物在室溫下攪拌隔夜，接著在0℃下向混合物中加入NaBH4(5.6g，147mmol，1.1eq.)。將混合物在室溫下攪拌2小時。混合物用水(100mL)淬滅，用EA(3×100mL)萃取，乾燥，且濃縮。藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之化合物K(10.3g，69.2%產率)。A mixture of cycloheptanone (15 g, 134 mmol, 1 eq.) and 2-aminoethanol (9 g, 147 mmol, 1.1 eq.) and 3 drops of AcOH in MeOH (250 mL) was stirred at room temperature overnight, and then NaBH4 (5.6 g, 147 mmol, 1.1 eq.) was added to the mixture at 0°C. The mixture was stirred at room temperature for 2 hours. The mixture was quenched with water (100 mL), extracted with EA (3×100 mL), dried, and concentrated. Purification by silica gel column chromatography (MeOH:DCM=0% to 10%) gave Compound K (10.3 g, 69.2% yield) as a yellow oil.
製備化合物LPreparation of Compound L
將環辛酮(2.0g，15.85mmol，1eq.)及2-胺基乙醇(1.07g，17.43mmol，1.1eq.)與3滴AcOH在MeOH(30mL)中之混合物在室溫下攪拌隔夜，接著在0℃下向混合物中加入NaBH4(660mg，17.43mmol，1.1eq.)。將混合物在室溫下攪拌2小時。將混合物用水(100mL)淬滅，經EA(3×100mL)萃取並乾燥。濃縮後，殘餘物藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之化合物L(960mg，35%產率)。A mixture of cyclooctanone (2.0 g, 15.85 mmol, 1 eq.) and 2-aminoethanol (1.07 g, 17.43 mmol, 1.1 eq.) and 3 drops of AcOH in MeOH (30 mL) was stirred at room temperature overnight, and then NaBH4 (660 mg, 17.43 mmol, 1.1 eq.) was added to the mixture at 0°C. The mixture was stirred at room temperature for 2 hours. The mixture was quenched with water (100 mL), extracted with EA (3×100 mL) and dried. After concentration, the residue was purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give compound L (960 mg, 35% yield) as a yellow oil.
製備SM2：Preparation of SM2:
將化合物26-1(250mg，0.56mmol，1.0eq.)、2-胺基乙醇(243mg，1.68mmol，3.0eq.)、K2CO3(232mg，1.68mmol，3.0eq.)、Cs2CO3(7mg，0.02mmol，0.03eq.)及碘化鈉(30mg，0.2mmol，0.3eq.)在ACN(10mL)中之混合物在100℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物SM2(1.78g，62.1%產率)。LCMS：Rt：1.427min；MS m/z(ESI)：428.5[M+H]+。A mixture of compound 26-1 (250 mg, 0.56 mmol, 1.0 eq.), 2-aminoethanol (243 mg, 1.68 mmol, 3.0 eq.), K2 CO3 (232 mg, 1.68 mmol, 3.0 eq.), Cs2 CO3 (7 mg, 0.02 mmol, 0.03 eq.) and sodium iodide (30 mg, 0.2 mmol, 0.3 eq.) in ACN (10 mL) was stirred at 100° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product SM2 (1.78 g, 62.1% yield) as a yellow oil. LCMS: Rt: 1.427min; MS m/z (ESI): 428.5[M+H]+ .
製備SM4：Preparation of SM4:
將化合物SM4-1(2.1g，4.5mmol，1.0eq.)、2-胺基乙醇(830mg，13.6mmol，3.0eq.)、K2CO3(1.9g，13.6mmol，3.0eq.)、Cs2CO3(440mg，1.4mmol，0.3eq.)、NaI(200mg，1.4mmol，0.3eq.)在ACN(15mL)中之混合物回流攪拌隔夜。將混合物用水稀釋，用EA萃取，濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物SM4(860mg，41%產率)。LCMS：Rt：1.000min；MS m/z(ESI)：442.4[M+H]+。A mixture of compound SM4-1 (2.1 g, 4.5 mmol, 1.0 eq.), 2-aminoethanol (830 mg, 13.6 mmol, 3.0 eq.), K2 CO3 (1.9 g, 13.6 mmol, 3.0 eq.), Cs2 CO3 (440 mg, 1.4 mmol, 0.3 eq.), NaI (200 mg, 1.4 mmol, 0.3 eq.) in ACN (15 mL) was stirred under reflux overnight. The mixture was diluted with water, extracted with EA, concentrated and purified by silica gel column chromatography (MeOH: DCM = 0% to 10%) to give the desired product SM4 (860 mg, 41% yield) as a yellow oil. LCMS: Rt: 1.000 min; MS m/z (ESI): 442.4 [M+H]+ .
製備SM9：Preparation of SM9:
向化合物SM9-1(1.0g，2.166mmol，1.0eq.)在ACN(15mL)中之溶液中加入化合物SM6(0.4g，6.498mmol，3.0eq.)、K2CO3(0.9g，6.498mmol，3.0eq.)、Cs2CO3(212mg，0.6498mmol，0.3eq.)、NaI(32mg，0.2166mmol，0.1eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物SM9(350mg，37.87%)。To a solution of compound SM9-1 (1.0 g, 2.166 mmol, 1.0 eq.) in ACN (15 mL) were added compound SM6 (0.4 g, 6.498 mmol, 3.0 eq.), K2 CO3 (0.9 g, 6.498 mmol, 3.0 eq.), Cs2 CO3 (212 mg, 0.6498 mmol, 0.3 eq.), NaI (32 mg, 0.2166 mmol, 0.1 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound SM9 (350 mg, 37.87%) as a yellow oil.
製備SM10：Preparation of SM10:
步驟1：製備化合物SM10-2Step 1: Preparation of compound SM10-2
向化合物SM10-1(2.0g，6.700mmol，1.0eq.)、化合物SM8(0.83g，8.040mmol，1.2eq.)、DIEA(2.6g，20.10mmol，3.0eq.)在DCM(30mL)中之混合物中加入HATU(3.8g，10.50mmol，1.5eq.)。將反應混合物在室溫下攪拌1小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈無色油狀之化合物SM10-2(2.4g，93.36%)。To a mixture of compound SM10-1 (2.0 g, 6.700 mmol, 1.0 eq.), compound SM8 (0.83 g, 8.040 mmol, 1.2 eq.), DIEA (2.6 g, 20.10 mmol, 3.0 eq.) in DCM (30 mL) was added HATU (3.8 g, 10.50 mmol, 1.5 eq.). The reaction mixture was stirred at room temperature for 1 hour. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and dried over Na2 SO4. The solvent was removed and FCC was performed to obtain compound SM10-2 (2.4 g, 93.36%) as a colorless oil.
步驟2：製備化合物SM10-3Step 2: Preparation of compound SM10-3
在0℃、N2下，向化合物SM10-2(2.4g，6.255mmol，1.0eq.)、DIEA(1.62g，12.51mmol，2.0eq.)在DCM(60mL)中之混合物中加入MsCl(0.86g，7.506mmol，1.2eq.)。將反應混合物在0℃下攪拌1小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈黃色油狀之化合物SM10-3(2.5g，86.57%)。To a mixture of compound SM10-2 (2.4 g, 6.255 mmol, 1.0 eq.), DIEA (1.62 g, 12.51 mmol, 2.0 eq.) in DCM (60 mL) at 0 °C underN2 was added MsCl (0.86 g, 7.506 mmol, 1.2 eq.). The reaction mixture was stirred at 0 °C for 1 hour. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and driedoverNa2SO4 . The solvent was removed and FCC was performed to give compound SM10-3 (2.5 g, 86.57%) as a yellow oil.
步驟3：製備化合物SM10-4Step 3: Preparation of compound SM10-4
向化合物SM10-3(1.5g，3.249mmol，1.0eq.)在ACN(30mL)中之溶液中加入化合物B(0.45g，3.899mmol，1.2eq.)、K2CO3(1.35g，9.747mmol，3.0eq.)、Cs2CO3(318mg，0.9747mmol，0.3eq.)、NaI(49mg，0.3249mmol，0.1eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物SM10-4(700mg，44.81%)。LCMS：Rt：0.830min；MS m/z(ESI)：481.4[M+H]+。To a solution of compound SM10-3 (1.5 g, 3.249 mmol, 1.0 eq.) in ACN (30 mL) were added compound B (0.45 g, 3.899 mmol, 1.2 eq.), K2 CO3 (1.35 g, 9.747 mmol, 3.0 eq.), Cs2 CO3 (318 mg, 0.9747 mmol, 0.3 eq.), NaI (49 mg, 0.3249 mmol, 0.1 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound SM10-4 (700 mg, 44.81%) as a yellow oil. LCMS: Rt: 0.830 min; MS m/z (ESI): 481.4 [M+H]+ .
步驟4：製備化合物SM10Step 4: Preparation of compound SM10
向化合物SM10-4(300mg，0.6240mmol，1.0eq.)在DCM(15mL)中之溶液中加入SOCl2(223mg，1.872mmol，3.0eq.)。將反應混合物在35℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，得到呈黃色油狀之化合物SM10(310mg，粗品)。LCMS：Rt：0.860min；MS m/z(ESI)：499.3[M+H]+。To a solution of compound SM10-4 (300 mg, 0.6240 mmol, 1.0 eq.) in DCM (15 mL) was added SOCl2 (223 mg, 1.872 mmol, 3.0 eq.). The reaction mixture was stirred at 35° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed to give compound SM10 (310 mg, crude) as a yellow oil. LCMS: Rt: 0.860 min; MS m/z (ESI): 499.3 [M+H]+ .
製備SM11：Preparation of SM11:
步驟1：製備化合物SM11-2Step 1: Preparation of compound SM11-2
將化合物SM10-1(1.5g，5.025mmol，1.0eq.)、化合物SM7(1.26g，7.538mmol，1.5eq.)、TsOH(300mg)在甲苯(20mL)中之混合物回流攪拌2小時。TLC顯示反應完成。將混合物減壓蒸發，進行FCC，得到呈黃色油狀之化合物SM11-2(1.4g，62.26%)。A mixture of compound SM10-1 (1.5 g, 5.025 mmol, 1.0 eq.), compound SM7 (1.26 g, 7.538 mmol, 1.5 eq.), and TsOH (300 mg) in toluene (20 mL) was refluxed and stirred for 2 hours. TLC showed that the reaction was complete. The mixture was evaporated under reduced pressure and subjected to FCC to obtain compound SM11-2 (1.4 g, 62.26%) as a yellow oil.
步驟2：製備化合物SM11Step 2: Preparation of compound SM11
向化合物SM11-2(1.0g，2.235mmol，1.0eq.)在ACN(15mL)中之溶液中加入化合物SM6(0.41g，6.704mmol，3.0eq.)、K2CO3(0.93g，6.704mmol，3.0eq.)、Cs2CO3(218mg，0.6704mmol，0.3eq.)、NaI(33mg，0.2235mmol，0.1eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物SM11(700mg，44.81%)。LCMS：Rt：0.890min；MS m/z(ESI)：428.3[M+H]+。To a solution of compound SM11-2 (1.0 g, 2.235 mmol, 1.0 eq.) in ACN (15 mL) were added compound SM6 (0.41 g, 6.704 mmol, 3.0 eq.), K2 CO3 (0.93 g, 6.704 mmol, 3.0 eq.), Cs2 CO3 (218 mg, 0.6704 mmol, 0.3 eq.), NaI (33 mg, 0.2235 mmol, 0.1 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound SM11 (700 mg, 44.81%) as a yellow oil. LCMS: Rt: 0.890 min; MS m/z (ESI): 428.3 [M+H]+ .
製備SM：Preparation of SM:
步驟1：製備化合物SM-2Step 1: Preparation of compound SM-2
在0℃、N2下，向NaH(12g，227.1mmol，2.5eq.)在DMF(100mL)中之混合物中加入化合物SM-1(12g，90.84mmol，1.0eq.)。將反應混合物在0℃下攪拌1小時。向其中加入在DMF(100mL)中之C8H17Br(44g，227.1mmol，2.5eq.)。將反應混合物在室溫下攪拌16小時。TLC顯示反應完成。將混合物倒入水中並用EA洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈無色油狀之化合物SM-2(17.8g，54.96%)。1H NMR(400MHz,CCl3D)：3.71(s,6H),1.88-1.84(m,4H),1.59(s,1H),1.25(s,19H),1.14-1.10(m,4H),0.89-0.86(m,6H)。To a mixture of NaH (12 g, 227.1 mmol, 2.5 eq.) in DMF (100 mL) at 0°C underN2 , compound SM-1 (12 g, 90.84 mmol, 1.0 eq.) was added. The reaction mixture was stirred at 0°C for 1 hour.C8H17Br (44 g, 227.1 mmol, 2.5 eq.) in DMF (100 mL) was added thereto. The reaction mixture was stirred at room temperature for 16 hours. TLC showed that the reaction was complete. The mixture was poured into water and washed with EA. The organics were separated and driedoverNa2SO4 . The solvent was removed and FCC was performed to obtain compound SM-2 (17.8 g, 54.96%) as a colorless oil.1 H NMR (400MHz, CCl3 D): 3.71 (s, 6H), 1.88-1.84 (m, 4H), 1.59 (s, 1H), 1.25 (s, 19H), 1.14-1.10 (m, 4H), 0.89-0.86 (m, 6H).
步驟2：製備化合物SM-3Step 2: Preparation of compound SM-3
向SM-2(17.8g，49.93mmol，1.0eq.)在DMF(260mL)中之溶液中加入LiCl(21.17g，499.3mmol，10.0eq.)。將反應混合物在120℃下攪拌12小時。TLC顯示反應完成。將混合物倒入水中並用EA洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈無色油狀之化合物SM-3(10g，67.10%)。1H NMR(400MHz,CCl3D)：0.89-0.86(m,6H),1.25(s,22H),1.45-1.40(m,2H),1.59(s,4H),2.36-2.30(m,1H),3.67(s,3H)。To a solution of SM-2 (17.8 g, 49.93 mmol, 1.0 eq.) in DMF (260 mL) was added LiCl (21.17 g, 499.3 mmol, 10.0 eq.). The reaction mixture was stirred at 120 °C for 12 h. TLC showed the reaction was complete. The mixture was poured into water and washed with EA. The organics were separated and dried over Na2 SO4. The solvent was removed and FCC was performed to give compound SM-3 (10 g, 67.10%) as a colorless oil.1 H NMR (400MHz, CCl3 D): 0.89-0.86 (m, 6H), 1.25 (s, 22H), 1.45- 1.40 (m, 2H), 1.59 (s, 4H), 2.36-2.30 (m, 1H), 3.67 (s, 3H).
步驟3：製備化合物SMStep 3: Preparation of Compound SM
在0℃下，向化合物SM-3(10g，33.50mmol，1.0eq.)在THF(100mL)中之溶液中緩慢加入LiAlH4(2.546g，67.00mmol，2.0eq.)。將反應混合物回流攪拌1小時。TLC顯示反應完成。冷卻至0℃後，連續加入水(3.4mL)、15% NaOH水溶液(3.4mL)及水(10mL)使混合物淬滅。將所得混合物用EA稀釋並藉由過濾除去沈澱物。減壓蒸發濾液，進行FCC，得到呈黃色油狀之化合物SM(8.5g，93.80%)。1H NMR(400MHz,CCl3D)：0.90-0.86(m,6H),1.27(s,27H),1.43(s,3H),3.54(d,J=5.2Hz,2H)。To a solution of compound SM-3 (10 g, 33.50 mmol, 1.0 eq.) in THF (100 mL) was slowly added LiAlH4 (2.546 g, 67.00 mmol, 2.0 eq.) at 0°C. The reaction mixture was stirred at reflux for 1 hour. TLC showed that the reaction was complete. After cooling to 0°C, water (3.4 mL), 15% aqueous NaOH solution (3.4 mL) and water (10 mL) were successively added to quench the mixture. The resulting mixture was diluted with EA and the precipitate was removed by filtration. The filtrate was evaporated under reduced pressure and subjected to FCC to obtain compound SM (8.5 g, 93.80%) as a yellow oil.1 H NMR (400MHz, CCl3 D): 0.90-0.86 (m, 6H), 1.27 (s, 27H), 1.43 (s, 3H), 3.54 (d,J =5.2Hz, 2H).
製備SM15：Preparation of SM15:
向化合物26-1(400mg，0.89mmol，1.0eq.)在ACN(30mL)中之溶液中加入化合物SM15-1(140mg，1.79mmol，2.0eq.)、K2CO3(370mg，2.68mmol，3.0eq.)、Cs2CO3(90mg，0.27mmol，0.3eq.)及NaI(40mg，0.27mmol，0.3eq.)。將反應混合物在80℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到化合物SM15(120mg，30%)。LCMS：Rt：0.900min；MS m/z(ESI)：442.3[M+H]+。To a solution of compound 26-1 (400 mg, 0.89 mmol, 1.0 eq.) in ACN (30 mL) were added compound SM15-1 (140 mg, 1.79 mmol, 2.0 eq.), K2 CO3 (370 mg, 2.68 mmol, 3.0 eq.), Cs2 CO3 (90 mg, 0.27 mmol, 0.3 eq.) and NaI (40 mg, 0.27 mmol, 0.3 eq.). The reaction mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound SM15 (120 mg, 30%). LCMS: Rt: 0.900 min; MS m/z (ESI): 442.3 [M+H]+ .
製備SM16：Preparation of SM16:
向化合物71-7(420mg，0.88mmol，1.0eq.)及化合物SM6(108mg，1.76mmol，2.0eq.)在ACN(20mL)中之溶液中加入K2CO3(365mg，2.64mmol，3.0eq.)、Cs2CO3(85mg，0.26mmol，0.3eq.)及NaI(39mg，0.26mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=10/1)純化，得到呈黃色油狀之化合物SM16(146mg，37%產率)。LCMS：Rt：0.810min；MS m/z(ESI)：444.3[M+H]+。To a solution of compound 71-7 (420 mg, 0.88 mmol, 1.0 eq.) and compound SM6 (108 mg, 1.76 mmol, 2.0 eq.) in ACN (20 mL) were added K2 CO3 (365 mg, 2.64 mmol, 3.0 eq.), Cs2 CO3 (85 mg, 0.26 mmol, 0.3 eq.) and NaI (39 mg, 0.26 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=10/1) to give compound SM16 (146 mg, 37% yield) as a yellow oil. LCMS: Rt: 0.810min; MS m/z (ESI): 444.3[M+H]+ .
製備SM18：Preparation of SM18:
將化合物SM18-1(2.0g，4.48mmol，1.0eq.)、(2-胺基乙基)胺基甲酸第三丁酯(1.0g，6.72mmol，1.5eq.)、K2CO3(1.8g，13.4mmol，3.0eq.)、Cs2CO3(440mg，1.34mmol，0.3eq.)、NaI(200mg，1.34mmol，0.3eq.)在ACN(20mL)中之混合物在90℃下攪拌隔夜。LCMS顯示目標產物。將混合物濃縮並藉由管柱層析法純化殘餘物，得到呈白色固體狀之產物SM18(860mg，36.5%產率)。LCMS：Rt：0.870min；MS m/z(ESI)：526.5[M+H]+。A mixture of compound SM18-1 (2.0 g, 4.48 mmol, 1.0 eq.), tert-butyl (2-aminoethyl)carbamate (1.0 g, 6.72 mmol, 1.5 eq.), K2 CO3 (1.8 g, 13.4 mmol, 3.0 eq.), Cs2 CO3 (440 mg, 1.34 mmol, 0.3 eq.), and NaI (200 mg, 1.34 mmol, 0.3 eq.) in ACN (20 mL) was stirred at 90° C. overnight. LCMS showed the target product. The mixture was concentrated and the residue was purified by column chromatography to give the product SM18 (860 mg, 36.5% yield) as a white solid. LCMS: Rt: 0.870min; MS m/z (ESI): 526.5[M+H]+ .
製備SM20：Preparation of SM20:
步驟1：製備化合物SM20-1Step 1: Preparation of compound SM20-1
在室溫下，向化合物26-1(1.0g，2.24mmol，1.0eq.)及化合物B(511.0mg，4.48mmol，2.0eq.)在ACN(20.0mL)中之溶液中加入Cs2CO3(218.0mg，0.67mmol，0.3eq.)、K2CO3(927.0mg，6.72mmol，3.0eq.)及NaI(33.0mg，0.22mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈棕色油狀之化合物SM20-1(0.6g，56%產率)。LCMS：Rt：0.950min；MS m/z(ESI)：482.4[M+H]+。To a solution of compound 26-1 (1.0 g, 2.24 mmol, 1.0 eq.) and compound B (511.0 mg, 4.48 mmol, 2.0 eq.) in ACN (20.0 mL) were added Cs2 CO3 (218.0 mg, 0.67 mmol, 0.3 eq.), K2 CO3 (927.0 mg, 6.72 mmol, 3.0 eq.) and NaI (33.0 mg, 0.22 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=1/0-20/1) to give compound SM20-1 (0.6 g, 56% yield) as a brown oil. LCMS: Rt: 0.950min; MS m/z (ESI): 482.4[M+H]+ .
步驟2：製備化合物SM20Step 2: Preparation of compound SM20
在室溫下，向化合物SM20-1(0.2g，0.41mmol，1.0eq.)在DCM(5.0mL)中之溶液中加入SOCl2(144.0mg，1.23mmol，3.0eq.)。將混合物攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈棕色油狀之化合物SM20(0.23g，粗品)。LCMS：Rt：1.330min；MS m/z(ESI)：500.3[M+H]+。To a solution of compound SM20-1 (0.2 g, 0.41 mmol, 1.0 eq.) in DCM (5.0 mL) was added SOCl2 (144.0 mg, 1.23 mmol, 3.0 eq.) at room temperature. The mixture was stirred for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to give compound SM20 (0.23 g, crude) as a brown oil. LCMS: Rt: 1.330 min; MS m/z (ESI): 500.3 [M+H]+ .
製備SM22：Preparation of SM22:
步驟1：製備化合物SM22-2Step 1: Preparation of compound SM22-2
向化合物SM22-1(30.0g，98.25mmol，1.0eq.)在DMF(800mL)中之溶液中加入NaCN(9.63g，196.5mmol，2.0eq.)。將反應物在60℃下攪拌10小時。將反應混合物倒入水(500mL)中並用EtOAc(3×500mL)萃取。將合併之有機層用鹽水洗滌，經無水Na2SO4乾燥並真空濃縮。藉由急速管柱層析法(EtOAc：PE=1：20)純化粗產物，得到呈黃色油狀之目標產物(18.3g，74%產率)。To a solution of compound SM22-1 (30.0 g, 98.25 mmol, 1.0 eq.) in DMF (800 mL) was added NaCN (9.63 g, 196.5 mmol, 2.0 eq.). The reactants were stirred at 60 °C for 10 h. The reaction mixture was poured into water (500 mL) and extracted with EtOAc (3×500 mL). The combined organic layers were washed with brine, dried over anhydrous Na2 SO4 and concentrated in vacuo. The crude product was purified by flash column chromatography (EtOAc: PE = 1:20) to give the target product (18.3 g, 74% yield) as a yellow oil.
步驟2：製備化合物SM22-3Step 2: Preparation of compound SM22-3
向化合物SM22-2(17.0g，67.61mmol，1.0eq.)在EtOH(200mL)中之溶液中加入H2SO4(40mL)。將反應物在90℃下攪拌48小時。將反應混合物倒入水(500mL)中並用EtOAc(3×500mL)萃取。將合併之有機層用鹽水洗滌，經無水Na2SO4乾燥並真空濃縮，得到呈黃色油狀之目標產物(15g，75%產率)。To a solution of compound SM22-2 (17.0 g, 67.61 mmol, 1.0 eq.) in EtOH (200 mL) was added H2 SO4 (40 mL). The reaction was stirred at 90 °C for 48 h. The reaction mixture was poured into water (500 mL) and extracted with EtOAc (3×500 mL). The combined organic layers were washed with brine, dried over anhydrous Na2 SO4 and concentrated in vacuo to give the target product (15 g, 75% yield) as a yellow oil.
步驟3：製備化合物SM22Step 3: Preparation of compound SM22
向化合物SM22-3(14g，46.90mmol，1.0eq.)在MeOH(240mL)及H2O(60mL)中之溶液中加入LiOH．H2O(9.84g，234.5mmol，5.0eq.)。將反應物在50℃下攪拌10小時。將反應混合物真空濃縮，得到粗目標產物。將粗產物溶解在水中。用6M HCl將殘餘物調節至pH=2，且用EtOAc(3×500mL)萃取。將合併之有機層用鹽水洗滌，經無水Na2SO4乾燥並真空濃縮，得到呈黃色油狀之化合物SM22(15g，75%產率)。1H NMR(400MHz,CCl3D)：0.87(t,J=8Hz,6H),1.22-1.46(m,24H),1.85-1.95(m,2H),2.22-2.34(m,1H)。To a solution of compound SM22-3 (14 g, 46.90 mmol, 1.0 eq.) in MeOH (240 mL) and H2 O (60 mL) was added LiOH.H2 O (9.84 g, 234.5 mmol, 5.0 eq.). The reactants were stirred at 50 °C for 10 h. The reaction mixture was concentrated in vacuo to give the crude target product. The crude product was dissolved in water. The residue was adjusted to pH = 2 with 6M HCl and extracted with EtOAc (3×500 mL). The combined organic layers were washed with brine, dried over anhydrous Na2 SO4 and concentrated in vacuo to give compound SM22 (15 g, 75% yield) as a yellow oil.1 H NMR (400MHz, CCl3 D): 0.87 (t,J =8Hz, 6H), 1.22-1.46 (m, 24H), 1.85-1.95 (m, 2H), 2.22-2.34 (m, 1H).
鑿備SM23：Chisel SM23:
步驟1：製備化合物SM23-1Step 1: Preparation of compound SM23-1
向化合物SM22(4g，14.79mmol，1.0eq.)在CH2Cl2(100mL)中之溶液中加入DIEA(5.73g，44.37mmol，3.0eq.)、化合物SM7(2.96g，17.75mmol，1.2eq.)、EDCI(4.25g，22.18mmol，1.5eq.)及DMAP(550mg，4.44mmol，0.3eq.)。將反應物在50℃下攪拌10小時。將反應混合物真空濃縮並藉由急速管柱層析法(EtOAc：PE=20：1)純化，得到呈黃色油狀之目標產物(4g，64%產率)。To a solution of compound SM22 (4 g, 14.79 mmol, 1.0 eq.) in CH2 Cl2 (100 mL) were added DIEA (5.73 g, 44.37 mmol, 3.0 eq.), compound SM7 (2.96 g, 17.75 mmol, 1.2 eq.), EDCI (4.25 g, 22.18 mmol, 1.5 eq.) and DMAP (550 mg, 4.44 mmol, 0.3 eq.). The reactants were stirred at 50° C. for 10 hours. The reaction mixture was concentrated in vacuo and purified by flash column chromatography (EtOAc:PE=20:1) to give the target product (4 g, 64% yield) as a yellow oil.
步驟2：製備化合物SM23Step 2: Preparation of compound SM23
向化合物SM23-1(1.5g，3.58mmol，1.0eq.)在CH3CN(50mL)中之溶液中加入K2CO3(1.48g，10.73mmol，3.0eq.)、Cs2CO3(0.4g，1.07mmol，0.3eq.)、NaI(0.16g，1.07mmol，0.3eq.)及化合物SM6(0.45g，7.15mmol，2.0eq.)。將反應物在80℃下攪拌10小時。將反應混合物真空濃縮。藉由急速管柱層析法(CH2Cl2：MeOH=10：1)純化粗產物，得到呈黃色油狀之目標產物(800mg，56%產率)。LCMS：Rt：0.898min；MS m/z(ESI)：400.3[M+H]+。To a solution of compound SM23-1 (1.5 g, 3.58 mmol, 1.0 eq.) in CH3 CN (50 mL) were added K2 CO3 (1.48 g, 10.73 mmol, 3.0 eq.), Cs2 CO3 (0.4 g, 1.07 mmol, 0.3 eq.), NaI (0.16 g, 1.07 mmol, 0.3 eq.) and compound SM6 (0.45 g, 7.15 mmol, 2.0 eq.). The reactants were stirred at 80° C. for 10 hours. The reaction mixture was concentrated in vacuo. The crude product was purified by flash column chromatography (CH2 Cl2 :MeOH=10:1) to give the target product (800 mg, 56% yield) as a yellow oil. LCMS: Rt: 0.898min; MS m/z (ESI): 400.3[M+H]+ .
製備SM24：Preparation of SM24:
向化合物SM24-1(20.2g，83.3mmol，1.0eq.)及化合物W(19.5g，100mol,1.2eq.)在DCM(300mL)中之溶液中加入EDCI(24.0g，125mmol，1.5eq.)、DMAP(2.0g，16.7mmol，0.2eq.)及DIEA(27.0g，208mmol，2.5eq.)。將反應混合物在室溫下攪拌16小時。TLC顯示反應完成。將反應混合物濃縮並藉由管柱層析法(矽膠，PE中之0-1% EA)純化，得到呈無色油狀之化合物SM24(17g，49%)。EDCI (24.0 g, 125 mmol, 1.5 eq.), DMAP (2.0 g, 16.7 mmol, 0.2 eq.) and DIEA (27.0 g, 208 mmol, 2.5 eq.) were added to a solution of compound SM24-1 (20.2 g, 83.3 mmol, 1.0 eq.) and compound W (19.5 g, 100 mol, 1.2 eq.) in DCM (300 mL). The reaction mixture was stirred at room temperature for 16 hours. TLC showed that the reaction was complete. The reaction mixture was concentrated and purified by column chromatography (silica gel, 0-1% EA in PE) to obtain compound SM24 (17 g, 49%) as a colorless oil.
製備SM26：Preparation of SM26:
步驟1：製備化合物SM26-2Step 1: Preparation of compound SM26-2
向化合物SM26-1(2g，7.080mmol，1.0eq.)及化合物SM7(1.42g，8.496mmol，1.2eq.)之混合物中加入在DCM(30mL)中之DIEA(1.8g，14.16mmol，2.0eq.)、EDCI(2g，10.62mmol，1.5eq.)、DMAP(0.17g，1.416mmol，0.2eq.)。將反應混合物在50℃下攪拌16小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈黃色油狀之化合物SM26-2(1.5g，49.10%)。To a mixture of compound SM26-1 (2 g, 7.080 mmol, 1.0 eq.) and compound SM7 (1.42 g, 8.496 mmol, 1.2 eq.) were added DIEA (1.8 g, 14.16 mmol, 2.0 eq.), EDCI (2 g, 10.62 mmol, 1.5 eq.), DMAP (0.17 g, 1.416 mmol, 0.2 eq.) in DCM (30 mL). The reaction mixture was stirred at 50 °C for 16 hours. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and dried over Na2 SO4. The solvent was removed and FCC was performed to give compound SM26-2 (1.5 g, 49.10%) as a yellow oil.
步驟2：製備化合物SM26Step 2: Preparation of compound SM26
向化合物SM26-2(1.5g，3.476mmol，1.0eq.)在ACN(30mL)中之溶液中加入化合物SM6(0.64g，10.43mmol，3.0eq.)、K2CO3(1.4g，10.43mmol，3.0eq.)、Cs2CO3(0.34g，1.043mmol，0.3eq.)、NaI(0.16g，1.043mmol，0.3eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物SM26(800mg，55.90%)。To a solution of compound SM26-2 (1.5 g, 3.476 mmol, 1.0 eq.) in ACN (30 mL) were added compound SM6 (0.64 g, 10.43 mmol, 3.0 eq.), K2 CO3 (1.4 g, 10.43 mmol, 3.0 eq.), Cs2 CO3 (0.34 g, 1.043 mmol, 0.3 eq.), NaI (0.16 g, 1.043 mmol, 0.3 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound SM26 (800 mg, 55.90%) as a yellow oil.
製備SM30：Preparation of SM30:
步驟1：製備化合物SM30-2Step 1: Preparation of compound SM30-2
向化合物SM30-1(6.3g，35.2mmol，1.0eq.)在DCM(150mL)中之溶液中加入TsOH.H2O(1.3g，7.0mmol，0.2eq.)及Na2SO4(15.0g，105.6mmol，3.0eq.)。將混合物在室溫下攪拌隔夜。將混合物過濾並濃縮。藉由矽膠管柱層析法(PE/EA=100/1)純化殘餘物，得到呈無色油狀之化合物SM30-2(9.7g，66%產率)。To a solution of compound SM30-1 (6.3 g, 35.2 mmol, 1.0 eq.) in DCM (150 mL) were added TsOH.H2 O (1.3 g, 7.0 mmol, 0.2 eq.) and Na2 SO4 (15.0 g, 105.6 mmol, 3.0 eq.). The mixture was stirred at room temperature overnight. The mixture was filtered and concentrated. The residue was purified by silica gel column chromatography (PE/EA=100/1) to give compound SM30-2 (9.7 g, 66% yield) as a colorless oil.
步驟2：製備化合物SM30Step 2: Preparation of compound SM30
向化合物SM30-2(4.2g，10.0mmol，1.0eq.)及乙醇胺(1.8g，30.0mmol，3.0eq.)在ACN(50mL)中之溶液中加入K2CO3(4.1g，30.0mmol，3.0eq.)、Cs2CO3(977mg，3.0mmol，0.3eq.)及NaI(450mg，3.0mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。將殘餘物濃縮並藉由矽膠管柱層析法(PE/EA=10/1-3/1-1/1-0/1)純化，得到呈無色油狀之化合物SM30(2.3g，58%產率)。LCMS：Rt：1.010min；MS m/z(ESI)：402.4[M+H]+。To a solution of compound SM30-2 (4.2 g, 10.0 mmol, 1.0 eq.) and ethanolamine (1.8 g, 30.0 mmol, 3.0 eq.) in ACN (50 mL) were added K2 CO3 (4.1 g, 30.0 mmol, 3.0 eq.), Cs2 CO3 (977 mg, 3.0 mmol, 0.3 eq.) and NaI (450 mg, 3.0 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 h. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was concentrated and purified by silica gel column chromatography (PE/EA=10/1-3/1-1/1-0/1) to give compound SM30 (2.3 g, 58% yield) as a colorless oil. LCMS: Rt: 1.010 min; MS m/z (ESI): 402.4 [M+H]+ .
製備SM34：Preparation of SM34:
步驟1：製備化合物SM34-2Step 1: Preparation of compound SM34-2
向化合物SM22-1(30g，98.2mmol，1.0eq.)在DMF(400mL)中之溶液中加入化合物SM34-1(36.4g，196.4mmol，2.0eq.)。將混合物在90℃下攪拌16小時。將反應混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(PE/EA=100/1)純化殘餘物，得到呈黃色油狀之化合物SM34-2(31.6g，86%產率)。To a solution of compound SM22-1 (30 g, 98.2 mmol, 1.0 eq.) in DMF (400 mL) was added compound SM34-1 (36.4 g, 196.4 mmol, 2.0 eq.). The mixture was stirred at 90 °C for 16 h. The reaction mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=100/1) to give compound SM34-2 (31.6 g, 86% yield) as a yellow oil.
步驟2：製備化合物SM34-3Step 2: Preparation of compound SM34-3
向化合物SM34-2(15.8g，42.5mmol，1.0eq.)在EtOH(500mL)中之溶液中加入一水合肼(5.0g，85.0mmol，2.0eq.)。將混合物回流攪拌16小時。LCMS顯示反應完成。將混合物過濾並用EtOH洗滌。將濾液濃縮並藉由矽膠管柱層析法(DCM/MeOH=20/1)純化，得到呈黃色油狀之化合物SM34-3(9.1g，88%產率)。Hydrazine monohydrate (5.0 g, 85.0 mmol, 2.0 eq.) was added to a solution of compound SM34-2 (15.8 g, 42.5 mmol, 1.0 eq.) in EtOH (500 mL). The mixture was stirred at reflux for 16 hours. LCMS showed that the reaction was complete. The mixture was filtered and washed with EtOH. The filtrate was concentrated and purified by silica gel column chromatography (DCM/MeOH=20/1) to obtain compound SM34-3 (9.1 g, 88% yield) as a yellow oil.
步驟3：製備化合物SM34-4Step 3: Preparation of compound SM34-4
向化合物SM34-3(6.5g，26.9mol,1.2eq.)在DCM(100mL)中之溶液中加入化合物W(4.4g，22.4mmol，1.0eq.)、HATU(12.8g，33.6mmol，1.5eq.)及DIPEA(8.7g，67.2mmol，3.0eq.)。將混合物在室溫下攪拌16小時。將反應混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法純化殘餘物，得到呈黃色油狀之化合物SM34-4(7.4g，65.6%產率)。To a solution of compound SM34-3 (6.5 g, 26.9 mol, 1.2 eq.) in DCM (100 mL) were added compound W (4.4 g, 22.4 mmol, 1.0 eq.), HATU (12.8 g, 33.6 mmol, 1.5 eq.) and DIPEA (8.7 g, 67.2 mmol, 3.0 eq.). The mixture was stirred at room temperature for 16 hours. The reaction mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography to give compound SM34-4 (7.4 g, 65.6% yield) as a yellow oil.
步驟4：製備化合物SM34Step 4: Preparation of compound SM34
向化合物SM34-4(7.4g，18.0mmol，1.0eq.)及化合物SM6(3.3g，54.0mmol，3.0eq.)在THF(50mL)中之溶液中加入DIPEA(6.9g，54.0mmol，3.0eq.)及NaI(800mg，5.4mmol，0.3eq.)。將混合物在70℃下攪拌10小時。LCMS顯示反應完成。將反應混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法純化殘餘物，得到呈無色油狀之化合物SM34(6.3g，88%產率)。LCMS：Rt：1.620min；MS m/z(ESI)：399.5[M+H]+。To a solution of compound SM34-4 (7.4 g, 18.0 mmol, 1.0 eq.) and compound SM6 (3.3 g, 54.0 mmol, 3.0 eq.) in THF (50 mL) were added DIPEA (6.9 g, 54.0 mmol, 3.0 eq.) and NaI (800 mg, 5.4 mmol, 0.3 eq.). The mixture was stirred at 70 ° C for 10 hours. LCMS showed that the reaction was completed. The reaction mixture was poured into water and extracted with EA. The combined organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography to give compound SM34 (6.3 g, 88% yield) as a colorless oil. LCMS: Rt: 1.620min; MS m/z (ESI): 399.5[M+H]+ .
製備SM38：Preparation of SM38:
將化合物71-7(600mg，1.25mmol，1.0eq.)、異丙胺(739mg，12.5mmol，10.0eq.)、K2CO3(519mg，3.76mmol，3.0eq.)、Cs2CO3(124mg，0.38mmol，0.3eq.)、NaI(51mg，0.38mmol，0.3eq.)在ACN(10mL)中之混合物回流攪拌隔夜。LCMS顯示產物。將混合物用EA稀釋並用水及鹽水洗滌，乾燥並濃縮。藉由FCC純化殘餘物，得到呈無色油狀之化合物SM38(320mg，58.0%產率)。A mixture of compound 71-7 (600 mg, 1.25 mmol, 1.0 eq.), isopropylamine (739 mg, 12.5 mmol, 10.0 eq.), K2 CO3 (519 mg, 3.76 mmol, 3.0 eq.), Cs2 CO3 (124 mg, 0.38 mmol, 0.3 eq.), NaI (51 mg, 0.38 mmol, 0.3 eq.) in ACN (10 mL) was stirred under reflux overnight. LCMS showed the product. The mixture was diluted with EA and washed with water and brine, dried and concentrated. The residue was purified by FCC to give compound SM38 (320 mg, 58.0% yield) as a colorless oil.
製備SM39：Preparation of SM39:
向化合物SM24(10g，23.9mmol，1.0eq.)在CH3CN(150mL)中之溶液中加入K2CO3(9.9g，71.7mmol，3.0eq.)、Cs2CO3(2.3g，7.17mmol，0.3eq.)、NaI(1.1g，7.17mmol，0.3eq.)及化合物SM6(2.9g，47.8mmol，2.0eq.)。將反應混合物在80℃下攪拌16小時。將反應混合物真空濃縮。藉由急速管柱層析法(CH2Cl2：MeOH=20：1-10：1)純化粗產物，得到呈黃色油狀之化合物SM39(5.1g，產率：53%)。LCMS：Rt：0.880min；MS m/z(ESI)：400.3[M+H]。To a solution of compound SM24 (10 g, 23.9 mmol, 1.0 eq.) in CH3 CN (150 mL) were added K2 CO3 (9.9 g, 71.7 mmol, 3.0 eq.), Cs2 CO3 (2.3 g, 7.17 mmol, 0.3 eq.), NaI (1.1 g, 7.17 mmol, 0.3 eq.) and compound SM6 (2.9 g, 47.8 mmol, 2.0 eq.). The reaction mixture was stirred at 80° C. for 16 hours. The reaction mixture was concentrated in vacuo. The crude product was purified by flash column chromatography (CH2 Cl2 :MeOH=20:1-10:1) to give compound SM39 (5.1 g, yield: 53%) as a yellow oil. LCMS: Rt: 0.880min; MS m/z (ESI): 400.3 [M+H].
6.2 實例2：製備化合物1.6.2 Example 2: Preparation of compound 1.
步驟1：製備化合物1-1Step 1: Preparation of compound 1-1
將化合物A(1.26g，3mmol，1.5eq.)、化合物B(280mg，2mmol，1eq.)、DIEA(774mg，6mmol，3eq.)及NaI(0.1eq.)在四氫呋喃(THF，6mL)中之混合物在70℃下攪拌隔夜。混合物在真空下濃縮並藉由矽膠管柱層析法(MeOH：DCM=0：1至1：80)純化，得到呈黃色油狀之期望產物化合物1-1(269mg，28.5%產率)。LCMS：Rt：1.000min；MS m/z(ESI)：482.5[M+H]+。A mixture of compound A (1.26 g, 3 mmol, 1.5 eq.), compound B (280 mg, 2 mmol, 1 eq.), DIEA (774 mg, 6 mmol, 3 eq.) and NaI (0.1 eq.) in tetrahydrofuran (THF, 6 mL) was stirred at 70°C overnight. The mixture was concentrated under vacuum and purified by silica gel column chromatography (MeOH:DCM=0:1 to 1:80) to give the desired product compound 1-1 (269 mg, 28.5% yield) as a yellow oil. LCMS: Rt: 1.000 min; MS m/z (ESI): 482.5 [M+H]+ .
步驟2：製備化合物1-2Step 2: Preparation of compound 1-2
將化合物1-1(269mg，0.56mmol，1eq.)及SOCl2(200mg，1.68mmol，3eq.)在DCM(6mL)中之混合物在35℃下攪拌隔夜。混合物在真空下濃縮，得到呈黃色油狀之期望產物化合物1-2(313mg，粗品)。LCMS：Rt：0.970min；MS m/z(ESI)：500.4[M+H]+。A mixture of compound 1-1 (269 mg, 0.56 mmol, 1 eq.) and SOCl2 (200 mg, 1.68 mmol, 3 eq.) in DCM (6 mL) was stirred at 35° C. overnight. The mixture was concentrated under vacuum to give the desired product compound 1-2 (313 mg, crude) as a yellow oil. LCMS: Rt: 0.970 min; MS m/z (ESI): 500.4 [M+H]+ .
步驟3：製備化合物1Step 3: Preparation of compound 1
將化合物1-2(313mg，0.63mmol，1.2eq.)、化合物C(211mg，0.53mmol，1eq.)、DIEA(205mg，1.59mmol，3eq.)及NaI觸媒在THF(4mL)中之混合物在70℃下攪拌隔夜。混合物在真空下濃縮，藉由製備型HPLC純化，得到呈淺棕色油狀之化合物1(79mg，14.6%產率)。A mixture of compound 1-2 (313 mg, 0.63 mmol, 1.2 eq.), compound C (211 mg, 0.53 mmol, 1 eq.), DIEA (205 mg, 1.59 mmol, 3 eq.) and NaI catalyst in THF (4 mL) was stirred at 70°C overnight. The mixture was concentrated under vacuum and purified by preparative HPLC to obtain compound 1 (79 mg, 14.6% yield) as a light brown oil.
1H NMR(400MHz,CDCl3)δ：0.83-0.93(m,12H),1.04-1.16(m,2H),1.18-1.39(m,60H),1.40-1.55(m,3H),1.56-1.74(m,9H),1.86(s,2H),2.25-2.39(m,5H),2.56(s,3H),2.70(s,3H),3.62(s,2H),3.89-4.04(m,4H)。LCMS：Rt：2.000min；MS m/z(ESI)：863.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.83-0.93(m,12H),1.04-1.16(m,2H),1.18-1.39(m,60H),1.40-1.55(m,3H),1.56-1.74(m ,9H),1.86(s,2H),2.25-2.39(m,5H),2.56(s,3H),2.70(s,3H),3.62(s,2H),3.89-4.04(m,4H). LCMS: Rt: 2.000min; MS m/z (ESI): 863.7[M+H]+ .
以下化合物係以與化合物1類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 1 using the corresponding starting materials.
6.3 實例3：製備化合物2.6.3 Example 3: Preparation of Compound 2.
步驟1：製備化合物2-1Step 1: Preparation of compound 2-1
向1-十一醇(10g，58.03mmol，1.0eq.)在DCM(120mL)中之溶液中加入1-乙基-3-(3-二甲基胺基丙基)碳二亞胺(EDCI，16.69g，87.05mmol，1.5eq.)、4-二甲基胺基吡啶(DMAP，1.42g，11.61mmol，0.2eq.)、DIEA(15g，116.06mmol，2.0eq.)及6-溴己酸(12.45g，63.84mmol，1.1eq.)。將反應混合物在55℃下攪拌16小時。TLC顯示反應完成。移除溶劑，且藉由FCC純化粗產物，得到呈無色油狀之化合物2-1(8.6g，42.43%)。To a solution of 1-undecanol (10 g, 58.03 mmol, 1.0 eq.) in DCM (120 mL) were added 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI, 16.69 g, 87.05 mmol, 1.5 eq.), 4-dimethylaminopyridine (DMAP, 1.42 g, 11.61 mmol, 0.2 eq.), DIEA (15 g, 116.06 mmol, 2.0 eq.) and 6-bromohexanoic acid (12.45 g, 63.84 mmol, 1.1 eq.). The reaction mixture was stirred at 55 °C for 16 hours. TLC showed that the reaction was complete. The solvent was removed, and the crude product was purified by FCC to give compound 2-1 (8.6 g, 42.43%) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：4.08-4.05(m,2H),3.55-3.52(m,2H),2.34-2.30(m,2H),1.83-1.76(m,2H),1.69-1.60(m,4H),1.51-1.43(m,2H),1.23(s,16H),0.89-0.86(m,3H)。1 H NMR (400MHz, CDCl3 )δ: 4.08-4.05(m,2H),3.55-3.52(m,2H),2.34-2.30(m,2H),1.83-1.76(m, 2H),1.69-1.60(m,4H),1.51-1.43(m,2H),1.23(s,16H),0.89-0.86(m,3H).
步驟2：製備化合物2-2Step 2: Preparation of compound 2-2
向化合物2-1(1g，2.863mmol，1.2eq.)在ACN(20mL)中之溶液中加入化合物D(275mg，2.386mmol，1.0eq.)、K2CO3(989mg，7.158mmol，3.0eq.)、Cs2CO3(233mg，0.7158mmol，0.3eq.)及NaI(18mg，0.1193mmol，0.05eq.)。將反應混合物在85℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，且藉由FCC純化粗產物，得到呈黃色油狀之化合物2-2(170mg，18.57%)。LCMS：Rt：0.811min；MS m/z(ESI)：384.3[M+H]+。To a solution of compound 2-1 (1 g, 2.863 mmol, 1.2 eq.) in ACN (20 mL) were added compound D (275 mg, 2.386 mmol, 1.0 eq.), K2 CO3 (989 mg, 7.158 mmol, 3.0 eq.), Cs2 CO3 (233 mg, 0.7158 mmol, 0.3 eq.) and NaI (18 mg, 0.1193 mmol, 0.05 eq.). The reaction mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed, and the crude product was purified by FCC to give compound 2-2 (170 mg, 18.57%) as a yellow oil. LCMS: Rt: 0.811 min; MS m/z (ESI): 384.3 [M+H]+ .
步驟3：製備化合物2-3Step 3: Preparation of compound 2-3
向化合物2-2(170mg，0.4432mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(158mg，1.330mmol，3.0eq.)。將反應混合物在35℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，得到呈黃色油狀之化合物2-3(180mg，粗品)。LCMS：Rt：0.860min；MS m/z(ESI)：402.3[M+H]+。To a solution of compound 2-2 (170 mg, 0.4432 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (158 mg, 1.330 mmol, 3.0 eq.). The reaction mixture was stirred at 35° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed to give compound 2-3 (180 mg, crude) as a yellow oil. LCMS: Rt: 0.860 min; MS m/z (ESI): 402.3 [M+H]+ .
步驟4：製備化合物2Step 4: Preparation of compound 2
向化合物2-3(170mg，0.4476mmol，1.0eq.)及DIEA(289mg，2.238mmol，5.0eq.)在THF(10mL)中之混合物中加入化合物E(381mg，0.6715mmol，1.5eq.)及NaI(20mg)。將反應混合物在70℃下攪拌16小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化，得到呈無色油狀之化合物2(35mg，8.37%產率)。Compound E (381 mg, 0.6715 mmol, 1.5 eq.) and NaI (20 mg) were added to a mixture of compound 2-3 (170 mg, 0.4476 mmol, 1.0 eq.) and DIEA (289 mg, 2.238 mmol, 5.0 eq.) in THF (10 mL). The reaction mixture was stirred at 70 ° C for 16 hours. LCMS showed that the reaction was complete. After removing the solvent, it was purified by preparative HPLC to obtain compound 2 (35 mg, 8.37% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：4.07-4.04(m,2H),3.9(d,J=5.6Hz,2H),3.53(m,1H),3.08-3.04(m,1H),2.49-2.37(m,9H),2.32-2.25(m,5H),1.98-1.88(m,4H),1.66-1.58(m,9H),1.49-1.38(m,7H),1.26(s,63H),0.90-0.86(m,12H)。LCMS：Rt：0.994min；MS m/z(ESI)：933.8[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 4.07-4.04 (m, 2H), 3.9 (d,J =5.6Hz, 2H), 3.53 (m, 1H), 3.08-3.04 (m, 1H),2.49-2.37(m,9H),2.32-2.25(m,5H),1.98-1.88(m,4H),1.66-1.58(m,9H),1.49-1.38(m,7H),1.26(s,63H),0.90-0.86(m,12H). LCMS: Rt: 0.994min; MS m/z (ESI): 933.8[M+H]+ .
6.4 實例4：製備化合物3.6.4 Example 4: Preparation of Compound 3.
步驟1：製備化合物3-1Step 1: Preparation of compound 3-1
向化合物2-1(1.0g，2.86mmol，2.0eq.)及化合物F(145mg，1.43mmol，1.0eq.)在ACN(30mL)中之溶液中加入K2CO3(593mg，4.29mmol，3.0eq.)、Cs2CO3(140mg，0.429mmol，0.3eq.)及NaI(64mg，0.429mmol，0.3eq.)。將混合物在80℃下攪拌48小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=50/1-25/1)純化，得到呈黃色油狀之化合物3-1(350mg，66%產率)。LCMS：Rt：0.800min；MS m/z(ESI)：370.3[M+H]+。To a solution of compound 2-1 (1.0 g, 2.86 mmol, 2.0 eq.) and compound F (145 mg, 1.43 mmol, 1.0 eq.) in ACN (30 mL) were added K2 CO3 (593 mg, 4.29 mmol, 3.0 eq.), Cs2 CO3 (140 mg, 0.429 mmol, 0.3 eq.) and NaI (64 mg, 0.429 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 48 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=50/1-25/1) to give compound 3-1 (350 mg, 66% yield) as a yellow oil. LCMS: Rt: 0.800min; MS m/z (ESI): 370.3[M+H]+ .
步驟2：製備化合物3-2Step 2: Preparation of compound 3-2
向化合物3-1(200mg，0.54mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(193mg，1.62mmol，3.0eq.)。將混合物在30℃下攪拌16小時。LCMS顯示反應完成。將混合物減壓濃縮，得到呈黃色油狀之化合物3-2(200mg，95%)。To a solution of compound 3-1 (200 mg, 0.54 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (193 mg, 1.62 mmol, 3.0 eq.). The mixture was stirred at 30° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated under reduced pressure to give compound 3-2 (200 mg, 95%) as a yellow oil.
步驟3：製備化合物3Step 3: Preparation of compound 3
向化合物3-2(200mg，0.52mmol，1.0eq.)及化合物C(416mg，1.04mmol，2.0eq.)在THF(10mL)中之溶液中加入N,N-二異丙基乙胺(DIPEA，202mg，1.56mmol，3.0eq.)及NaI(24mg，0.16mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物3(80mg，8%產率)。N,N-diisopropylethylamine (DIPEA, 202 mg, 1.56 mmol, 3.0 eq.) and NaI (24 mg, 0.16 mmol, 0.3 eq.) were added to a solution of compound 3-2 (200 mg, 0.52 mmol, 1.0 eq.) and compound C (416 mg, 1.04 mmol, 2.0 eq.) in THF (10 mL). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 3 (80 mg, 8% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.48-0.50(m,4H),0.86-0.90(m,9H),1.26-1.30(m,45H),1.49-1.66(m,11H),1.72-1.77(m,1H),2.28-2.32(m,4H),2.52-2.76(m,10H),3.52-3.58(m,2H),3.96-3.98(m,2H),4.04-4.07(m,2H)。LCMS：Rt：1.250min；MS m/z(ESI)：751.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.48-0.50(m,4H),0.86-0.90(m,9H),1.26-1.30(m,45H),1.49-1.66(m,11H),1.72-1.77(m,1H) ),2.28-2.32(m,4H),2.52-2.76(m,10H),3.52-3.58(m,2H),3.96-3.98(m,2H),4.04-4.07(m,2H). LCMS: Rt: 1.250min; MS m/z (ESI): 751.6[M+H]+ .
以下化合物係以與化合物3類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 3 using the corresponding starting materials.
6.5 實例5：製備化合物6.6.5 Example 5: Preparation of Compound 6.
步驟1：製備化合物6-1Step 1: Preparation of compound 6-1
將化合物2-1(786mg，2.24mmol，1.2eq.)、化合物B(268mg，1.87mol,1eq.)、DIEA(724mg，5.61mmol，3eq.)及NaI(0.1eq.)在THF(10mL)中之混合物在70℃下攪拌隔夜。將混合物在真空下濃縮並藉由矽膠管柱層析法(MeOH：DCM=0：1至1：20)純化，得到呈淺棕色油狀之化合物6-1(1.18g，粗品)。LCMS：Rt：0.910min；MS m/z(ESI)：412.3[M+H]+。A mixture of compound 2-1 (786 mg, 2.24 mmol, 1.2 eq.), compound B (268 mg, 1.87 mol, 1 eq.), DIEA (724 mg, 5.61 mmol, 3 eq.) and NaI (0.1 eq.) in THF (10 mL) was stirred at 70° C. overnight. The mixture was concentrated under vacuum and purified by silica gel column chromatography (MeOH:DCM=0:1 to 1:20) to give compound 6-1 (1.18 g, crude) as a light brown oil. LCMS: Rt: 0.910 min; MS m/z (ESI): 412.3 [M+H]+ .
步驟2：製備化合物6-2Step 2: Preparation of compound 6-2
將化合物6-1(412mg，1mmol，1eq.)及SOCl2(357mg，3mmol，3eq.)在DCM(6mL)中之混合物在35℃下攪拌隔夜。在真空下濃縮混合物，得到呈黃色油狀之化合物6-2(430mg，粗品)。LCMS：Rt：0.930min；MS m/z(ESI)：430.3[M+H]+。A mixture of compound 6-1 (412 mg, 1 mmol, 1 eq.) and SOCl2 (357 mg, 3 mmol, 3 eq.) in DCM (6 mL) was stirred at 35° C. overnight. The mixture was concentrated under vacuum to give compound 6-2 (430 mg, crude) as a yellow oil. LCMS: Rt: 0.930 min; MS m/z (ESI): 430.3 [M+H]+ .
步驟3：製備化合物6Step 3: Preparation of compound 6
將化合物6-2(215mg，0.5mmol，1eq.)、化合物C(150mg，0.4mmol，0.75eq.)、DIEA(195mg，1.5mmol，3eq.)及催化量之NaI在THF(3mL)中之混合物在70℃下攪拌隔夜。將混合物在真空下濃縮，藉由製備型HPLC純化，得到呈淺棕色油狀之化合物6(15mg，12.8%產率)。A mixture of compound 6-2 (215 mg, 0.5 mmol, 1 eq.), compound C (150 mg, 0.4 mmol, 0.75 eq.), DIEA (195 mg, 1.5 mmol, 3 eq.) and a catalytic amount of NaI in THF (3 mL) was stirred at 70°C overnight. The mixture was concentrated under vacuum and purified by preparative HPLC to obtain compound 6 (15 mg, 12.8% yield) as a light brown oil.
1H NMR(400MHz,CDCl3)δ：0.83-0.92(m,9H),1.18-1.36(m,40H),1.38-1.48(m,4H),1.49-1.75(m,27H),1.85-2.15(m,5H),2.16-2.27(m,1H),2.30-2.39(m,3H),3.11-3.25(m,2H),3.35-3.48(m,1H),3.93-3.99(m,2H),4.01-4.11(m,2H)。LCMS：Rt：1.720min；MS m/z(ESI)：793.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.83-0.92(m,9H),1.18-1.36(m,40H),1.38-1.48(m,4H),1.49-1.75(m,27H),1.85-2.15(m,5H),2.16- 2.27(m,1H),2.30-2.39(m,3H),3.11-3.25(m,2H),3.35-3.48(m,1H),3.93-3.99(m,2H),4.01-4.11(m,2H). LCMS: Rt: 1.720min; MS m/z (ESI): 793.6[M+H]+ .
6.6 實例6：製備化合物8.6.6 Example 6: Preparation of Compound 8.
步驟1：製備8-1Step 1: Prepare 8-1
向化合物A(0.85g，1.98mmol)在CH3CN(50mL)中之溶液中加入K2CO3(410mg，2.97mmol)、Cs2CO3(100mg，0.29mmol)、NaI(50mg，0.29mmol)及化合物G(127mg，0.99mmol)。將反應物在80℃下攪拌10小時。真空濃縮反應混合物。藉由急速管柱層析法(CH2Cl2：MeOH=10：1)純化粗產物，得到呈黃色油狀之化合物8-1(300mg，產率：65%)。LCMS：Rt：0.88min；MS m/z(ESI)：468.4[M+H]+。To a solution of compound A (0.85 g, 1.98 mmol) in CH3 CN (50 mL) were added K2 CO3 (410 mg, 2.97 mmol), Cs2 CO3 (100 mg, 0.29 mmol), NaI (50 mg, 0.29 mmol) and compound G (127 mg, 0.99 mmol). The reactants were stirred at 80° C. for 10 hours. The reaction mixture was concentrated in vacuo. The crude product was purified by flash column chromatography (CH2 Cl2 :MeOH=10:1) to give compound 8-1 (300 mg, yield: 65%) as a yellow oil. LCMS: Rt: 0.88 min; MS m/z (ESI): 468.4 [M+H]+ .
步驟2：製備化合物8-2Step 2: Preparation of compound 8-2
向化合物8-1(300mg，0.64mmol)在CH2Cl2(10mL)中之溶液中加入SOCl2(250mg，2.05mmol)。將反應物在30℃下攪拌10小時。真空濃縮反應混合物，得到呈黃色油狀之化合物8-2(310mg，產率：100%)。To a solution of compound 8-1 (300 mg, 0.64 mmol) in CH2 Cl2 (10 mL) was added SOCl2 (250 mg, 2.05 mmol). The reactants were stirred at 30° C. for 10 hours. The reaction mixture was concentrated in vacuo to give compound 8-2 (310 mg, yield: 100%) as a yellow oil.
步驟3：製備化合物8Step 3: Preparation of compound 8
向化合物8-2(300mg，0.62mmol)在THF(10mL)中之溶液中加入DIEA(240mg，1.85mmol)、NaI(100mg，0.65mmol)及化合物C(530mg，1.31mmol)。將反應物在70℃下攪拌10小時。過濾反應混合物並真空濃縮。粗產物藉由製備型HPLC純化，得到呈黃色油狀之化合物8(45mg，產率：8.5%)。To a solution of compound 8-2 (300 mg, 0.62 mmol) in THF (10 mL) were added DIEA (240 mg, 1.85 mmol), NaI (100 mg, 0.65 mmol) and compound C (530 mg, 1.31 mmol). The reactants were stirred at 70 °C for 10 hours. The reaction mixture was filtered and concentrated in vacuo. The crude product was purified by preparative HPLC to give compound 8 (45 mg, yield: 8.5%) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.87-0.90(t,J=6.8Hz,12H),1.26(m,50H),1.40-1.51(m,8H),1.60-1.66(m,8H),1.77-1.73(m,3H),2.31-2.33(m,4H),2.48-2.61(m,10H),3.05-3.09(m,1H),3.48-3.55(m,4H),3.96-3.97(m,4H)。LCMS：Rt：1.740min；MS m/z(ESI)：849.7[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.87-0.90 (t,J =6.8Hz, 12H), 1.26 (m, 50H), 1.40-1.51 (m, 8H), 1.60-1.66 (m,8H),1.77-1.73(m,3H),2.31-2.33(m,4H),2.48-2.61(m,10H),3.05-3.09(m,1H),3.48-3.55(m,4H),3.96-3.97(m,4H). LCMS: Rt: 1.740min; MS m/z (ESI): 849.7[M+H]+ .
以下化合物係以與化合物8類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 8 using the corresponding starting materials.
6.7 實例7：製備化合物10.6.7 Example 7: Preparation of Compound 10.
步驟1：製備化合物10-1Step 1: Preparation of compound 10-1
在室溫下，向化合物H(446.0mg，1.0mmol，1.0eq.)及乙醇胺(180.0mg，3.0mmol，3.0eq.)在ACN(10.0mL)中之溶液中加入Cs2CO3(97.5mg，0.3mmol，0.3eq.)、K2CO3(414.0mg，3.0mmol，3.0eq.)及NaI(14.6mg，0.1mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈黃色油狀之化合物10-1(0.35g，82%產率)。LCMS：Rt：0.942min；MS m/z(ESI)：428.3[M+H]+。To a solution of compound H (446.0 mg, 1.0 mmol, 1.0 eq.) and ethanolamine (180.0 mg, 3.0 mmol, 3.0 eq.) in ACN (10.0 mL) were added Cs2 CO3 (97.5 mg, 0.3 mmol, 0.3 eq.), K2 CO3 (414.0 mg, 3.0 mmol, 3.0 eq.) and NaI (14.6 mg, 0.1 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, the mixture was evaporated under reduced pressure, and purified by FCC (DCM/MeOH=1/0-20/1) to give compound 10-1 (0.35 g, 82% yield) as a yellow oil. LCMS: Rt: 0.942min; MS m/z (ESI): 428.3[M+H]+ .
步驟2：製備化合物10-2Step 2: Preparation of compound 10-2
在室溫下，向化合物H(1.0g，2.24mmol，1.0eq.)及化合物D(511.0mg，4.48mmol，2.0eq.)在ACN(20.0mL)中之溶液中加入Cs2CO3(218.0mg，0.67mmol，0.3eq.)、K2CO3(927.0mg，6.72mmol，3.0eq.)及NaI(33.0mg，0.22mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈棕色油狀之化合物10-2(0.6g，56%產率)。LCMS：Rt：0.950min；MS m/z(ESI)：482.4[M+H]+。To a solution of compound H (1.0 g, 2.24 mmol, 1.0 eq.) and compound D (511.0 mg, 4.48 mmol, 2.0 eq.) in ACN (20.0 mL) were added Cs2 CO3 (218.0 mg, 0.67 mmol, 0.3 eq.), K2 CO3 (927.0 mg, 6.72 mmol, 3.0 eq.) and NaI (33.0 mg, 0.22 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, the mixture was evaporated under reduced pressure, and purified by FCC (DCM/MeOH=1/0-20/1) to give compound 10-2 (0.6 g, 56% yield) as a brown oil. LCMS: Rt: 0.950min; MS m/z (ESI): 482.4[M+H]+ .
步驟3：製備化合物10-3Step 3: Preparation of compound 10-3
在室溫下，向化合物10-2(0.2g，0.41mmol，1.0eq.)在DCM(5.0mL)中之溶液中加入SOCl2(144.0mg，1.23mmol，3.0eq.)。將混合物攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈棕色油狀之化合物10-3(0.23g，粗品)。LCMS：Rt：1.330min；MS m/z(ESI)：500.3[M+H]+。To a solution of compound 10-2 (0.2 g, 0.41 mmol, 1.0 eq.) in DCM (5.0 mL) was added SOCl2 (144.0 mg, 1.23 mmol, 3.0 eq.) at room temperature. The mixture was stirred for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to give compound 10-3 (0.23 g, crude) as a brown oil. LCMS: Rt: 1.330 min; MS m/z (ESI): 500.3 [M+H]+ .
步驟4：製備化合物10Step 4: Preparation of compound 10
在0℃下，向化合物10-3(150.0mg，0.3mmol，1.0eq.)及化合物10-1(192.0mg，0.45mmol，1.5eq.)在THF(5.0mL)中之溶液中加入DIEA(193mg，1.5mmol，5.0eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且用製備型HPLC純化，得到呈棕色油狀之化合物10(80.0mg，25%產率)。At 0°C, DIEA (193 mg, 1.5 mmol, 5.0 eq.) was added to a solution of compound 10-3 (150.0 mg, 0.3 mmol, 1.0 eq.) and compound 10-1 (192.0 mg, 0.45 mmol, 1.5 eq.) in THF (5.0 mL). The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to obtain compound 10 (80.0 mg, 25% yield) as a brown oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,12H),1.26-1.32(m,61H),1.41-1.65(m,12H),1.85-2.02(m,4H),2.28-2.61(m,14H),3.00-3.12(m,1H),3.53-3.55(m,2H),3.97(d,J=5.6Hz,4H)。LCMS：Rt：2.520min；MS m/z(ESI)：891.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,12H),1.26-1.32(m,61H),1.41-1.65(m,12H),1.85-2.02 (m,4H),2.28-2.61(m,14H),3.00-3.12(m,1H),3.53-3.55(m,2H),3.97(d,J =5.6Hz,4H). LCMS: Rt: 2.520min; MS m/z (ESI): 891.7[M+H]+ .
6.8 實例8：製備化合物11.6.8 Example 8: Preparation of Compound 11.
步驟1：製備化合物11-AStep 1: Preparation of compound 11-A
在0℃下，向2-辛基-1-癸醇(2.7g，10.0mmol，1.0eq.)及DIPEA(2.6g，20.0mmol，2.0eq.)在DCM(50mL)中之溶液中滴加甲磺醯氯(MsCl，1.4g，12.0mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將反應混合物用水、鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物11-A(3.1g，91%產率)。1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,6H),1.26-1.32(m,29H),3.00(s,3H),4.11-4.13(m,2H)。To a solution of 2-octyl-1-decanol (2.7 g, 10.0 mmol, 1.0 eq.) and DIPEA (2.6 g, 20.0 mmol, 2.0 eq.) in DCM (50 mL) was added methanesulfonyl chloride (MsCl, 1.4 g, 12.0 mmol, 1.2 eq.) dropwise at 0°C. The mixture was stirred at room temperature for 2 hours. The reaction mixture was washed with water, brine, dried over Na2 SO4 and concentrated to give compound 11-A (3.1 g, 91% yield) as a yellow oil.1 H NMR (400 MHz, CDCl3 ) δ: 0.86-0.90 (m, 6H), 1.26-1.32 (m, 29H), 3.00 (s, 3H), 4.11-4.13 (m, 2H).
步驟2：製備化合物11-1Step 2: Preparation of compound 11-1
向11-A(18.0g，51.6mmol，1.0eq.)在DMF(300mL)中之溶液中加入鄰苯二甲醯亞胺鉀(19.1g，103.2mmol，2.0eq.)。將混合物在90℃下攪拌16小時。將反應混合物倒入水中，且用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(PE/EA=100/1)純化，得到呈無色油狀之化合物11-1(14.6g，71%產率)。1H NMR(400MHz,CDCl3)δ：0.85-0.88(m,6H),1.24-1.29(m,28H),1.82-1.89(m,1H),3.56-3.58(m,2H),7.72-7.72(m,2H),7.83-7.85(m,2H)。To a solution of 11-A (18.0 g, 51.6 mmol, 1.0 eq.) in DMF (300 mL) was added potassium phthalimide (19.1 g, 103.2 mmol, 2.0 eq.). The mixture was stirred at 90 °C for 16 hours. The reaction mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. Purification by silica gel column chromatography (PE/EA=100/1) gave compound 11-1 (14.6 g, 71% yield) as a colorless oil.1 H NMR (400MHz, CDCl3 ) δ: 0.85-0.88(m,6H), 1.24-1.29(m,28H), 1.82-1.89(m,1H), 3.56-3.58(m,2H), 7.72-7.72(m,2H), 7.83-7.85(m,2H).
步驟3：製備化合物11-2Step 3: Preparation of compound 11-2
向化合物11-1(14.6g，36.5mmol，1.0eq.)在EtOH(400mL)中之溶液中加入一水合肼(3.65g，73.0mmol，2.0eq.)。將混合物回流攪拌16小時。LCMS顯示反應完成。過濾混合物並用EtOH洗滌。將濾液濃縮並藉由矽膠管柱層析法(DCM/MeOH=100/1-50/1)純化，得到呈黃色油狀之化合物11-2(6.9g，70%產率)。LCMS：Rt：1.260min；MS m/z(ESI)：270.3[M+H]+。To a solution of compound 11-1 (14.6 g, 36.5 mmol, 1.0 eq.) in EtOH (400 mL) was added hydrazine monohydrate (3.65 g, 73.0 mmol, 2.0 eq.). The mixture was stirred at reflux for 16 hours. LCMS showed that the reaction was complete. The mixture was filtered and washed with EtOH. The filtrate was concentrated and purified by silica gel column chromatography (DCM/MeOH=100/1-50/1) to give compound 11-2 (6.9 g, 70% yield) as a yellow oil. LCMS: Rt: 1.260 min; MS m/z (ESI): 270.3 [M+H]+ .
步驟4：製備化合物11-3Step 4: Preparation of compound 11-3
向化合物11-2(6.9g，25.6mmol，1.0eq.)在DCM(250mL)中之溶液中加入6-溴己酸(6.0g，30.7mmol，1.2eq.)、HATU(11.7g，30.7mmol，1.2eq.)及DIPEA(9.9g，76.8mmol，3.0eq.)。將混合物在室溫下攪拌16小時。將反應混合物倒入水中，且用DCM萃取。將合併之有機層用類水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(PE/EA=10/1-8/1)純化，得到呈黃色油狀之化合物11-3(7.1g，62%產率)。To a solution of compound 11-2 (6.9 g, 25.6 mmol, 1.0 eq.) in DCM (250 mL) were added 6-bromohexanoic acid (6.0 g, 30.7 mmol, 1.2 eq.), HATU (11.7 g, 30.7 mmol, 1.2 eq.) and DIPEA (9.9 g, 76.8 mmol, 3.0 eq.). The mixture was stirred at room temperature for 16 hours. The reaction mixture was poured into water and extracted with DCM. The combined organic layers were washed with aqueous solution, dried over Na2 SO4 and concentrated. Purification by silica gel column chromatography (PE/EA=10/1-8/1) gave compound 11-3 (7.1 g, 62% yield) as a yellow oil.
步驟5：製備化合物11-4Step 5: Preparation of compound 11-4
向化合物11-3(800mg，1.79mmol，1.5eq.)及化合物D(137mg，1.19mmol，1.0eq.)在ACN(40mL)中之溶液中加入K2CO3(493mg，3.57mmol，3.0eq.)、Cs2CO3(116mg，0.357mmol，0.3eq.)及NaI(54mg，0.357mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=10/1)純化，得到呈黃色油狀之化合物11-4(400mg，70%產率)。LCMS：Rt：0.920min；MS m/z(ESI)：481.4[M+H]+。To a solution of compound 11-3 (800 mg, 1.79 mmol, 1.5 eq.) and compound D (137 mg, 1.19 mmol, 1.0 eq.) in ACN (40 mL) were added K2 CO3 (493 mg, 3.57 mmol, 3.0 eq.), Cs2 CO3 (116 mg, 0.357 mmol, 0.3 eq.) and NaI (54 mg, 0.357 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=10/1) to give compound 11-4 (400 mg, 70% yield) as a yellow oil. LCMS: Rt: 0.920min; MS m/z (ESI): 481.4[M+H]+ .
步驟6：製備化合物11-5Step 6: Preparation of compound 11-5
向化合物11-4(200mg，0.42mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(150mg，1.26mmol，3.0eq.)。將混合物在30℃下攪拌16小時。LCMS顯示反應完成。將混合物減壓濃縮，得到呈黃色油狀之化合物11-5(200mg，95%)。LCMS：Rt：0.980min；MS m/z(ESI)：499.3[M+H]+。To a solution of compound 11-4 (200 mg, 0.42 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (150 mg, 1.26 mmol, 3.0 eq.). The mixture was stirred at 30° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated under reduced pressure to give compound 11-5 (200 mg, 95%) as a yellow oil. LCMS: Rt: 0.980 min; MS m/z (ESI): 499.3 [M+H]+ .
步驟7：製備化合物11-6Step 7: Preparation of compound 11-6
向化合物11-3(610mg，1.36mmol，1.0eq.)及乙醇胺(166mg，2.72mmol，2.0eq.)在ACN(20mL)中之溶液中加入K2CO3(564mg，4.08mmol，3.0eq.)、Cs2CO3(134mg，0.41mmol，0.3eq.)及NaI(61mg，0.41mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物倒入水中，且用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化，得到呈黃色油狀之化合物11-6(320mg，55%產率)。LCMS：Rt：0.96min；MS m/z(ESI)：427.3[M+H]+。To a solution of compound 11-3 (610 mg, 1.36 mmol, 1.0 eq.) and ethanolamine (166 mg, 2.72 mmol, 2.0 eq.) in ACN (20 mL) were added K2 CO3 (564 mg, 4.08 mmol, 3.0 eq.), Cs2 CO3 (134 mg, 0.41 mmol, 0.3 eq.) and NaI (61 mg, 0.41 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. Purification by silica gel column chromatography (DCM/MeOH=10/1) gave compound 11-6 (320 mg, 55% yield) as a yellow oil. LCMS: Rt: 0.96 min; MS m/z (ESI): 427.3 [M+H]+ .
步驟8：製備化合物11Step 8: Preparation of compound 11
向化合物11-5(175mg，0.35mmol，1.0eq.)及化合物11-6(150mg，0.35mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(136mg，1.05mmol，3.0eq.)及NaI(10mg，0.07mmol，0.2eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物11(34mg，11%產率)。DIPEA (136 mg, 1.05 mmol, 3.0 eq.) and NaI (10 mg, 0.07 mmol, 0.2 eq.) were added to a solution of compound 11-5 (175 mg, 0.35 mmol, 1.0 eq.) and compound 11-6 (150 mg, 0.35 mmol, 1.0 eq.) in THF (10 mL). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 11 (34 mg, 11% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.34(m,64H),1.41-1.54(m,6H),1.59-1.77(m,6H),1.99-2.07(m,2H),2.17-2.21(m,4H),2.47-2.71(m,10H),3.15-3.18(m,4H),3.55-3.62(m,2H),5.73-5.84(m,2H)。LCMS：Rt：1.610min；MS m/z(ESI)：889.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.34(m,64H),1.41-1.54(m,6H),1.59-1.77(m,6H),1.99-2.07(m,2H) ),2.17-2.21(m,4H),2.47-2.71(m,10H),3.15-3.18(m,4H),3.55-3.62(m,2H),5.73-5.84(m,2H). LCMS: Rt: 1.610min; MS m/z (ESI): 889.8[M+H]+ .
以下化合物係以與化合物11類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 11 using the corresponding starting materials.
6.9 實例9：製備化合物15.6.9 Example 9: Preparation of Compound 15.
向化合物11-6(221mg，0.52mmol，1.0eq.)及化合物10-3(259mg，0.52mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(202mg，1.56mmol，3.0eq.)及NaI(16mg，0.104mmol，0.2eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物15(121mg，26%產率)。To a solution of compound 11-6 (221 mg, 0.52 mmol, 1.0 eq.) and compound 10-3 (259 mg, 0.52 mmol, 1.0 eq.) in THF (10 mL) was added DIPEA (202 mg, 1.56 mmol, 3.0 eq.) and NaI (16 mg, 0.104 mmol, 0.2 eq.). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 15 (121 mg, 26% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.92(m,12H),1.26-1.30(m,67H),1.46-1.72(m,12H),1.98-2.09(m,2H),2.15-2.19(m,2H),2.31-2.71(m,8H),3.16-3.23(m,2H),3.56-3.66(m,2H),3.95-4.03(m,2H),7.30(s,1H)。LCMS：Rt：1.68min；MS m/z(ESI)：890.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.92(m,12H),1.26-1.30(m,67H),1.46-1.72(m,12H),1.98-2.09(m,2H),2.15-2.19( m,2H),2.31-2.71(m,8H),3.16-3.23(m,2H),3.56-3.66(m,2H),3.95-4.03(m,2H),7.30(s,1H). LCMS: Rt: 1.68min; MS m/z (ESI): 890.7[M+H]+ .
以下化合物係以與化合物15類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 15 using the corresponding starting materials.
6.10 實例10：製備化合物18.6.10 Example 10: Preparation of Compound 18.
步驟1：製備化合物18-1Step 1: Preparation of compound 18-1
在氬氣氣氛下，在室溫下向丙二酸二甲酯(5g，38mmol，1eq.)在DMF(76mL)中之攪拌溶液中加入氫化鈉(3.8g，95mmol，2.5eq.)。0.5小時後，將(Z)-1-溴癸-4-烯(21g，95mmol，2.5eq.)加入混合物中，將混合物在室溫下攪拌隔夜。將混合物用水(130mL)淬滅，用EA(3×65mL)萃取；將合併之有機層用鹽水(2×65mL)洗滌，經無水硫酸鈉乾燥，真空濃縮。藉由矽膠管柱層析法(EA：PE=0%-5%)純化，得到呈無色油狀之化合物18-1(10.5g，68.2%產率)。To a stirred solution of dimethyl malonate (5 g, 38 mmol, 1 eq.) in DMF (76 mL) was added sodium hydride (3.8 g, 95 mmol, 2.5 eq.) at room temperature under an argon atmosphere. After 0.5 h, (Z)-1-bromodec-4-ene (21 g, 95 mmol, 2.5 eq.) was added to the mixture, and the mixture was stirred at room temperature overnight. The mixture was quenched with water (130 mL) and extracted with EA (3×65 mL); the combined organic layers were washed with brine (2×65 mL), dried over anhydrous sodium sulfate, and concentrated in vacuo. Purification by silica gel column chromatography (EA:PE=0%-5%) gave compound 18-1 (10.5 g, 68.2% yield) as a colorless oil.
步驟2：製備化合物18-2Step 2: Preparation of compound 18-2
將化合物18-1(10.5g，25.7mmol，1eq.)及LiCl(10.9g，257mmol，10eq.)在DMF(180mL)中之混合物在120℃下攪拌24小時。將混合物用水稀釋，用EA萃取，用鹽水洗滌，乾燥並濃縮。殘餘物藉由矽膠管柱層析法(EA：PE=0%-5%)純化，得到呈無色油狀之化合物18-2(7.5g，83.2%產率)。1H NMR(400MHz,CDCl3)δ：0.80-0.95(m,6H),1.18-1.37(m,16H),1.40-1.52(m,2H),1.54-1.66(m,3H),1.90-2.08(m,7H),2.24-2.41(m,1H),3.60-3.75(m,3H),5.24-5.49(m,4H)。A mixture of compound 18-1 (10.5 g, 25.7 mmol, 1 eq.) and LiCl (10.9 g, 257 mmol, 10 eq.) in DMF (180 mL) was stirred at 120 ° C. for 24 hours. The mixture was diluted with water, extracted with EA, washed with brine, dried and concentrated. The residue was purified by silica gel column chromatography (EA: PE = 0%-5%) to give compound 18-2 (7.5 g, 83.2% yield) as a colorless oil.1 H NMR (400MHz, CDCl3 )δ: 0.80-0.95(m,6H),1.18-1.37(m,16H),1.40-1.52(m,2H),1.54-1.66(m,3H) ),1.90-2.08(m,7H),2.24-2.41(m,1H),3.60-3.75(m,3H),5.24-5.49(m,4H).
步驟3：製備化合物18-3Step 3: Preparation of compound 18-3
將化合物18-2(7.5g，21.5mmol，1eq.)、LiAlH4(1.6g，43mmol，2eq.)在THF(100mL)中之混合物在80℃下攪拌隔夜。將混合物用水淬滅，過濾；將濾液濃縮，藉由矽膠管柱層析法(EA：PE=0%至5%)純化，得到呈黃色油狀之化合物18-3(6.2g，89.8%產率)。A mixture of compound 18-2 (7.5 g, 21.5 mmol, 1 eq.) and LiAlH4 (1.6 g, 43 mmol, 2 eq.) in THF (100 mL) was stirred at 80° C. overnight. The mixture was quenched with water and filtered; the filtrate was concentrated and purified by silica gel column chromatography (EA:PE=0% to 5%) to give compound 18-3 (6.2 g, 89.8% yield) as a yellow oil.
步驟4：製備化合物18-4Step 4: Preparation of compound 18-4
將化合物18-3(1.8g，5.5mmol，1eq.)、6-溴己酸(1.3g，6.6mmol，1.2eq.)、DIEA(2.14g，16.5mmol，3eq.)、DMAP(337mg，2.76mmol，0.5eq.)及EDCI(1.27g，6.6mmol，1.2mmol)在DCM(20mL)中之溶液在40℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(EA：PE=0%-2%)純化，得到呈無色油狀之化合物18-4(2.1g，75.2%產率)。A solution of compound 18-3 (1.8 g, 5.5 mmol, 1 eq.), 6-bromohexanoic acid (1.3 g, 6.6 mmol, 1.2 eq.), DIEA (2.14 g, 16.5 mmol, 3 eq.), DMAP (337 mg, 2.76 mmol, 0.5 eq.) and EDCI (1.27 g, 6.6 mmol, 1.2 mmol) in DCM (20 mL) was stirred at 40°C overnight. The mixture was concentrated and purified by silica gel column chromatography (EA:PE = 0%-2%) to obtain compound 18-4 (2.1 g, 75.2% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.83-0.93(m,6H),1.23-1.40(m,20H),1.41-1.54(m,2H),1.62-1.72(m,3H),1.83-2.10(m,10H),2.25-2.46(m,2H),3.18-3.52(m,2H),3.87-4.03(m,2H),5.18-5.58(m,4H)。1 H NMR (400MHz, CDCl3 )δ: 0.83-0.93(m,6H),1.23-1.40(m,20H),1.41-1.54(m,2H),1.62-1.72(m,3H),1.83-2 .10(m,10H),2.25-2.46(m,2H),3.18-3.52(m,2H),3.87-4.03(m,2H),5.18-5.58(m,4H).
步驟5：製備化合物18-5Step 5: Preparation of compound 18-5
將化合物18-4(300mg，0.6mmol，1eq.)、化合物B(133mg，0.9mmol，1.5eq.)、DIEA(232mg，1.8mmol，3eq.)及碘化鈉(30mg，0.2mmol，0.3eq.)在THF(6mL)中之混合物在70℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈無色油狀之化合物18-5(147mg，43.6%產率)。LCMS：Rt：0.900min；MS m/z(ESI)：562.4[M+H]+。A mixture of compound 18-4 (300 mg, 0.6 mmol, 1 eq.), compound B (133 mg, 0.9 mmol, 1.5 eq.), DIEA (232 mg, 1.8 mmol, 3 eq.) and sodium iodide (30 mg, 0.2 mmol, 0.3 eq.) in THF (6 mL) was stirred at 70° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give compound 18-5 (147 mg, 43.6% yield) as a colorless oil. LCMS: Rt: 0.900 min; MS m/z (ESI): 562.4 [M+H]+ .
步驟6：製備化合物18-6Step 6: Preparation of compound 18-6
將化合物18-5(147mg，0.26mmol，1eq.)及SOCl2(93mg，0.78mmol，3eq.)在DCM(5mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到化合物18-6(137mg，90.2%產率)。LCMS：Rt：1.210min；MS m/z(ESI)：580.4[M+H]+。A mixture of compound 18-5 (147 mg, 0.26 mmol, 1 eq.) and SOCl2 (93 mg, 0.78 mmol, 3 eq.) in DCM (5 mL) was stirred at 35° C. overnight. The mixture was concentrated to give compound 18-6 (137 mg, 90.2% yield). LCMS: Rt: 1.210 min; MS m/z (ESI): 580.4 [M+H]+ .
步驟7：製備化合物18-7Step 7: Preparation of compound 18-7
將化合物18-4(1971mg，2mmol，1eq.)、2-胺基乙醇(147mg，2.4mmol，1.2mmol)、K2CO3(828mg，6mmol，3eq.)、Cs2CO3(20mg，0.06mmol，0.03eq.)及NaI(15mg，0.1mmol，0.05eq.)在ACN(40mL)中之混合物在80℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈棕色油狀之化合物18-7(610mg，65.4%產率)。LCMS：Rt：0.910min；MS m/z(ESI)：480.4[M+H]+。A mixture of compound 18-4 (1971 mg, 2 mmol, 1 eq.), 2-aminoethanol (147 mg, 2.4 mmol, 1.2 mmol), K2 CO3 (828 mg, 6 mmol, 3 eq.), Cs2 CO3 (20 mg, 0.06 mmol, 0.03 eq.) and NaI (15 mg, 0.1 mmol, 0.05 eq.) in ACN (40 mL) was stirred at 80° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give compound 18-7 (610 mg, 65.4% yield) as a brown oil. LCMS: Rt: 0.910 min; MS m/z (ESI): 480.4 [M+H]+ .
步驟8：製備化合物18Step 8: Preparation of compound 18
將化合物18-6(137mg，0.24mmol，1eq.)、化合物18-7(138mg，0.29mmol，1.2eq.)、碘化鈉(10mg，0.07mmol，0.3eq.)及DIEA(93mg，0.72mmol，3eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化，得到呈淺棕色油狀之化合物18(21mg，8.7%產率)。A mixture of compound 18-6 (137 mg, 0.24 mmol, 1 eq.), compound 18-7 (138 mg, 0.29 mmol, 1.2 eq.), sodium iodide (10 mg, 0.07 mmol, 0.3 eq.) and DIEA (93 mg, 0.72 mmol, 3 eq.) in THF (5 mL) was stirred at 70°C overnight. The mixture was concentrated in vacuo. Purification by preparative HPLC gave compound 18 (21 mg, 8.7% yield) as a light brown oil.
1H NMR(400MHz,CDCl3)δ：0.83-0.92(m,12H),1.15-1.23(m,3H),1.24-1.36(m,47H),1.37-1.52(m,5H),1.56-1.69(m,12H),1.71-1.79(m,4H),1.95-2.05(m,14H),2.21-2.33(m,4H),2.42-2.60(m,9H),3.49-3.56(m,1H),3.95-3.99(m,3H),5.30-5.42(m,8H)。LCMS：Rt：0.640min；MS m/z(ESI)：1023.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.83-0.92(m,12H),1.15-1.23(m,3H),1.24-1.36(m,47H),1.37-1.52(m,5H),1.56-1.69(m,12H),1.71-1.79(m,4 H),1.95-2.05(m,14H),2.21-2.33(m,4H),2.42-2.60(m,9H),3.49-3.56(m,1H),3.95-3.99(m,3H),5.30-5.42(m,8H). LCMS: Rt: 0.640min; MS m/z (ESI): 1023.7[M+H]+ .
6.11 實例11：製備化合物19.6.11 Example 11: Preparation of Compound 19.
步驟1：製備化合物19-1Step 1: Preparation of compound 19-1
向順-4-癸烯-1-醇(1.56g，10.0mmol，1.0eq.)及6-溴己酸(2.9g，15.0mmol，1.5eq.)在30mL二氯甲烷中之溶液中加入DIEA(3.87g，30.0mmol，3.0eq.)及DMAP(244.0mg，2.0mmol，0.2eq.)。在環境溫度下攪拌5分鐘後，加入EDCI(2.86g，15.0mmol，1.5eq.)，且將反應混合物在室溫下攪拌隔夜，之後TLC顯示起始醇完全消失。將反應混合物用二氯甲烷(300mL)稀釋，且用飽和NaHCO3(100mL)、水(100mL)及鹽水(100mL)洗滌。合併之有機層經Na2SO4乾燥，且真空移除溶劑。蒸發溶劑，得到粗產物，將其藉由矽膠管柱層析法(PE中之0-2% EA)純化，得到呈無色油狀之化合物19-1(1.3g，39%)。To a solution of cis-4-decen-1-ol (1.56 g, 10.0 mmol, 1.0 eq.) and 6-bromohexanoic acid (2.9 g, 15.0 mmol, 1.5 eq.) in 30 mL of dichloromethane were added DIEA (3.87 g, 30.0 mmol, 3.0 eq.) and DMAP (244.0 mg, 2.0 mmol, 0.2 eq.). After stirring at ambient temperature for 5 minutes, EDCI (2.86 g, 15.0 mmol, 1.5 eq.) was added, and the reaction mixture was stirred at room temperature overnight, after which TLC showed complete disappearance of the starting alcohol. The reaction mixture was diluted with dichloromethane (300 mL), and washed with saturated NaHCO3 (100 mL), water (100 mL), and brine (100 mL). The combined organic layers were dried over Na2 SO4 and the solvent was removed in vacuo. The solvent was evaporated to give a crude product, which was purified by silica gel column chromatography (0-2% EA in PE) to give compound 19-1 (1.3 g, 39%) as a colorless oil.
步驟2：製備化合物19-2Step 2: Preparation of compound 19-2
在室溫下，向化合物19-1(664.0mg，2.0mmol，1.0eq.)及化合物B(572.0mg，4.0mmol，2.0eq.)在ACN(10.0mL)中之溶液中加入Cs2CO3(195.0mg，0.6mmol，0.3eq.)、K2CO3(828.0mg，6.0mmol，3.0eq.)及NaI(28.0mg，0.2mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈黃色油狀之化合物19-2(0.37g，47%產率)。LCMS：Rt：0.740min；MS m/z(ESI)：396.3[M+H]+。To a solution of compound 19-1 (664.0 mg, 2.0 mmol, 1.0 eq.) and compound B (572.0 mg, 4.0 mmol, 2.0 eq.) in ACN (10.0 mL) were added Cs2 CO3 (195.0 mg, 0.6 mmol, 0.3 eq.), K2 CO3 (828.0 mg, 6.0 mmol, 3.0 eq.) and NaI (28.0 mg, 0.2 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, the mixture was evaporated under reduced pressure, and purified by FCC (DCM/MeOH=1/0-20/1) to give compound 19-2 (0.37 g, 47% yield) as a yellow oil. LCMS: Rt: 0.740min; MS m/z (ESI): 396.3[M+H]+ .
步驟3：製備化合物19-3Step 3: Preparation of compound 19-3
在室溫下，向化合物19-2(170.0mg，0.43mmol，1.0eq.)在DCM(5.0mL)中之溶液中加入SOCl2(152.0mg，1.29mmol，3.0eq.)。將混合物攪拌16小時。LCMS顯示反應完成，減壓濃縮，得到呈棕色油狀之化合物19-3(0.2g，粗品)。LCMS：Rt：0.785min；MS m/z(ESI)：414.3[M+H]+。To a solution of compound 19-2 (170.0 mg, 0.43 mmol, 1.0 eq.) in DCM (5.0 mL) was added SOCl2 (152.0 mg, 1.29 mmol, 3.0 eq.) at room temperature. The mixture was stirred for 16 hours. LCMS showed that the reaction was complete, and the product was concentrated under reduced pressure to give compound 19-3 (0.2 g, crude) as a brown oil. LCMS: Rt: 0.785 min; MS m/z (ESI): 414.3 [M+H]+ .
步驟4：製備化合物19Step 4: Preparation of compound 19
在0℃下，向化合物19-3(200.0mg，0.48mmol，1.0eq.)及化合物10-1(247.0mg，0.58mmol，1.2eq.)在THF(5.0mL)中之溶液中加入DIEA(309.0mg，2.4mmol，5.0eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，減壓濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物19(80.0mg，21%產率)。At 0°C, DIEA (309.0 mg, 2.4 mmol, 5.0 eq.) was added to a solution of compound 19-3 (200.0 mg, 0.48 mmol, 1.0 eq.) and compound 10-1 (247.0 mg, 0.58 mmol, 1.2 eq.) in THF (5.0 mL). The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was concentrated under reduced pressure and purified by preparative HPLC to obtain compound 19 (80.0 mg, 21% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,9H),1.26-1.45(m,43H),1.60-1.80(m,17H),1.98-2.16(m,4H),2.28-2.61(m,15H),3.52-3.54(m,2H),3.96-4.08(m,4H),5.26-5.46(m,2H)。LCMS：Rt：1.137min；MS m/z(ESI)：805.7[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.89 (m, 9H), 1.26-1.45 (m, 43H), 1.60-1.80 (m, 17H), 1.98-2.16 (m, 4H),2.28-2.61(m,15H),3.52-3.54(m,2H),3.96-4.08(m,4H),5.26-5.46(m,2H). LCMS: Rt: 1.137min; MS m/z (ESI): 805.7[M+H]+ .
6.12 實例12：製備化合物20.6.12 Example 12: Preparation of Compound 20.
步驟1：製備化合物20-1Step 1: Preparation of compound 20-1
向肉豆蔻醇(2.1g，10.0mmol，1.0eq.)在THF(20.0mL)中之溶液中加入NaH(0.8g，20.0mmol，2.0eq.)。將混合物在室溫下攪拌2小時，接著加入1-溴-2,3-環氧丙烷(2.5g，15.0mmol，1.5eq.)，且在70℃下攪拌16小時。LCMS顯示反應完成，加入水，用EA萃取，濃縮並藉由FCC(PE/EA=20/1)純化，得到呈無色油狀之化合物20-1(2.6g，96%產率)。1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,3H),1.21-1.35(m,20H),1.58-1.67(m,2H),2.60-2.78(m,1H),2.79-2.81(m,1H),3.13-3.17(m,1H),3.36-3.50(m,3H),3.51-3.72(m,1H)。To a solution of myristyl alcohol (2.1 g, 10.0 mmol, 1.0 eq.) in THF (20.0 mL) was added NaH (0.8 g, 20.0 mmol, 2.0 eq.). The mixture was stirred at room temperature for 2 hours, then 1-bromo-2,3-epoxypropane (2.5 g, 15.0 mmol, 1.5 eq.) was added and stirred at 70° C. for 16 hours. LCMS showed that the reaction was complete, water was added, extracted with EA, concentrated and purified by FCC (PE/EA=20/1) to give compound 20-1 (2.6 g, 96% yield) as a colorless oil.1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,3H),1.21-1.35(m,20H),1.58-1.67(m,2H),2.60-2.78(m,1H ),2.79-2.81(m,1H),3.13-3.17(m,1H),3.36-3.50(m,3H),3.51-3.72(m,1H).
步驟2：製備化合物20-2Step 2: Preparation of compound 20-2
向環丁酮(840mg，12.0mmol，1.2eq.)在MeOH(10mL)中之溶液中加入2-(苄氧基)乙-1-胺(1.5g，10.0mmol，1.0eq.)。將混合物在25℃下攪拌2小時。接著向混合物中加入NaCNBH3(1.0g，15.0mmol，1.5eq.)。將混合物在25℃下攪拌16小時。LCMS顯示反應完成。將混合物減壓蒸發，且藉由FCC(DCM/MeOH=20/1)純化，得到呈黃色油狀之化合物20-2(1.0g，粗品)。To a solution of cyclobutanone (840 mg, 12.0 mmol, 1.2 eq.) in MeOH (10 mL) was added 2-(benzyloxy)ethan-1-amine (1.5 g, 10.0 mmol, 1.0 eq.). The mixture was stirred at 25°C for 2 hours. Then NaCNBH3 (1.0 g, 15.0 mmol, 1.5 eq.) was added to the mixture. The mixture was stirred at 25°C for 16 hours. LCMS showed that the reaction was complete. The mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=20/1) to give compound 20-2 (1.0 g, crude) as a yellow oil.
步驟3：製備化合物20-3Step 3: Preparation of compound 20-3
將化合物20-1(0.8g，2.96mmol，1.0eq.)及化合物20-2(1.0g，3.84mmol，1.3eq.)在EtOH(10.0mL)中之溶液在70℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由FCC(DCM/MeOH=30/1)純化，得到呈黃色油狀之化合物20-3(0.5g，35%產率)。LCMS：Rt：0.840min；MS m/z(ESI)：476.3[M+H]+。A solution of compound 20-1 (0.8 g, 2.96 mmol, 1.0 eq.) and compound 20-2 (1.0 g, 3.84 mmol, 1.3 eq.) in EtOH (10.0 mL) was stirred at 70° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by FCC (DCM/MeOH=30/1) to give compound 20-3 (0.5 g, 35% yield) as a yellow oil. LCMS: Rt: 0.840 min; MS m/z (ESI): 476.3 [M+H]+ .
步驟4：製備化合物20-4Step 4: Preparation of compound 20-4
向化合物20-3(475mg，1.0mmol，1.0eq.)在THF(10.0mL)中之溶液中加入NaH(160mg，4.0mmol，4.0eq.)。將混合物在室溫下攪拌2小時，接著加入C8H17Br(576mg，3.0mmol，3.0eq.)，且在70℃下攪拌16小時。LCMS顯示反應完成，加入水，用EA萃取，濃縮並藉由FCC(PE/EA=20/1)純化，得到呈無色油狀之化合物20-4(300mg，51%產率)。LCMS：Rt：1.280min；MS m/z(ESI)：588.4[M+H]+。To a solution of compound 20-3 (475 mg, 1.0 mmol, 1.0 eq.) in THF (10.0 mL) was added NaH (160 mg, 4.0 mmol, 4.0 eq.). The mixture was stirred at room temperature for 2 hours, then C8 H17 Br (576 mg, 3.0 mmol, 3.0 eq.) was added and stirred at 70° C. for 16 hours. LCMS showed that the reaction was complete, water was added, extracted with EA, concentrated and purified by FCC (PE/EA=20/1) to give compound 20-4 (300 mg, 51% yield) as a colorless oil. LCMS: Rt: 1.280 min; MS m/z (ESI): 588.4 [M+H]+ .
步驟5：製備化合物20-5Step 5: Preparation of compound 20-5
向化合物20-4(250mg，0.43mmol，1.0eq.)在EA(10mL)中之溶液中加入Pd/C(25.0mg)及HCl(5滴)。將混合物在室溫、H2下攪拌16小時。LCMS顯示反應完成，過濾並濃縮，得到呈黃色油狀之化合物20-5(250mg，粗品)。LCMS：Rt：1.023min；MS m/z(ESI)：498.4[M+H]+。To a solution of compound 20-4 (250 mg, 0.43 mmol, 1.0 eq.) in EA (10 mL) was added Pd/C (25.0 mg) and HCl (5 drops). The mixture was stirred at room temperature underH2 for 16 hours. LCMS showed that the reaction was complete, and the product was filtered and concentrated to give compound 20-5 (250 mg, crude) as a yellow oil. LCMS: Rt: 1.023 min; MS m/z (ESI): 498.4 [M+H]+ .
步驟6：製備化合物20-6Step 6: Preparation of compound 20-6
在室溫下，向化合物20-5(240mg，0.5mmol，1.0eq.)在DCM(5.0mL)中之溶液中加入SOCl2(177.0mg，1.5mmol，3.0eq.)。將混合物攪拌16小時。LCMS顯示反應完成，將混合物減壓濃縮，得到呈棕色油狀之化合物20-6(0.27g，粗品)。To a solution of compound 20-5 (240 mg, 0.5 mmol, 1.0 eq.) in DCM (5.0 mL) was added SOCl2 (177.0 mg, 1.5 mmol, 3.0 eq.) at room temperature. The mixture was stirred for 16 hours. LCMS showed that the reaction was complete, and the mixture was concentrated under reduced pressure to give compound 20-6 (0.27 g, crude) as a brown oil.
步驟7：製備化合物20Step 7: Preparation of compound 20
在0℃下，向化合物20-6(120.0mg，0.23mmol，1.0eq.)及化合物10-1(120.0mg，0.28mmol，1.2eq.)在THF(5.0mL)中之溶液中加入DIEA(148.0mg，1.1mmol，5.0eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且藉由製備型HPLC純化，得到呈黃色油狀之化合物20(30.0mg，14%產率)。At 0°C, DIEA (148.0 mg, 1.1 mmol, 5.0 eq.) was added to a solution of compound 20-6 (120.0 mg, 0.23 mmol, 1.0 eq.) and compound 10-1 (120.0 mg, 0.28 mmol, 1.2 eq.) in THF (5.0 mL). The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to obtain compound 20 (30.0 mg, 14% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,12H),1.21-1.35(m,65H),1.50-1.65(m,11H),1.98-2.00(m,3H),2.28-2.32(m,2H),2.53-2.62(m,9H),3.40-3.59(m,10H),3.96(d,J=5.6Hz,2H)。LCMS：Rt：4.600min；MS m/z(ESI)：907.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,12H),1.21-1.35(m,65H),1.50-1.65(m,11H),1.98-2.00 (m,3H),2.28-2.32(m,2H),2.53-2.62(m,9H),3.40-3.59(m,10H),3.96(d,J =5.6Hz,2H). LCMS: Rt: 4.600min; MS m/z (ESI): 907.8[M+H]+ .
6.13 實例13：製備化合物22.6.13 Example 13: Preparation of Compound 22.
步驟1：製備化合物22-1Step 1: Preparation of compound 22-1
在0℃、N2下，向NaH(3g，74.07mmol，2.5eq.)在DMF(30mL)中之混合物中加入丙二酸二甲酯(4g，30mmol，1.0eq.)。將反應混合物在0℃下攪拌0.5小時。添加在DMF(30mL)中之1-溴庚烷(13.4g，75mmol，2.5eq.)。將反應混合物在室溫下攪拌16小時。TLC顯示反應完成。將反應混合物用水淬滅，且用EA洗滌。分離有機層，且經Na2SO4乾燥。移除溶劑，且藉由FCC純化，得到呈無色油狀之化合物22-1(5.3g，53.78%)。1H NMR(400MHz,CDCl3)δ：3.71(s,6H),1.88-1.84(m,4H),1.31-1.26(m,16H),1.14-1.10(m,4H),0.89-0.86(m,6H)。To a mixture of NaH (3 g, 74.07 mmol, 2.5 eq.) in DMF (30 mL) at 0 °C underN2 was added dimethyl malonate (4 g, 30 mmol, 1.0 eq.). The reaction mixture was stirred at 0 °C for 0.5 h. 1-Bromoheptane (13.4 g, 75 mmol, 2.5 eq.) in DMF (30 mL) was added. The reaction mixture was stirred at room temperature for 16 h. TLC showed that the reaction was complete. The reaction mixture was quenched with water and washed with EA. The organic layerwas separated and dried overNa2SO4 . The solvent was removed and purified by FCC to give compound 22-1 (5.3 g, 53.78%) as a colorless oil.1 H NMR (400MHz, CDCl3) δ: 3.71 (s, 6H), 1.88-1.84 (m, 4H), 1.31-1.26 (m, 16H), 1.14-1.10 (m, 4H), 0.89-0.86 (m, 6H).
步驟2：製備化合物22-2Step 2: Preparation of compound 22-2
向化合物22-1(5.3g，16.13mmol，1.0eq.)在DMF(100mL)中之溶液中加入LiCl(6.8g，161.3mmol，10.0eq.)。將反應混合物在120℃下攪拌12小時。TLC顯示反應完成。將反應混合物用水淬滅，且用EA洗滌。分離有機層，且經Na2SO4乾燥。移除溶劑，且藉由FCC純化，得到呈無色油狀之化合物22-2(3.4g，78.07%)。1H NMR(400MHz,CDCl3)δ：3.67(s,3H),2.33-2.31(m,1H),1.60-1.40(m,6H),1.25(s,18H),0.89-0.86(m,6H)。To a solution of compound 22-1 (5.3 g, 16.13 mmol, 1.0 eq.) in DMF (100 mL) was added LiCl (6.8 g, 161.3 mmol, 10.0 eq.). The reaction mixture was stirred at 120 °C for 12 hours. TLC showed that the reaction was complete. The reaction mixture was quenched with water and washed with EA. The organic layer was separated and dried over Na2 SO4. The solvent was removed and purified by FCC to give compound 22-2 (3.4 g, 78.07%) as a colorless oil.1 H NMR (400MHz, CDCl3 ) δ: 3.67 (s, 3H), 2.33-2.31 (m, 1H), 1.60-1.40 (m, 6H), 1.25 (s, 18H), 0.89-0.86 (m, 6H).
步驟3：製備化合物22-3Step 3: Preparation of compound 22-3
在0℃下，向化合物22-2(3.4g，12.57mmol，1.0eq.)在THF(60mL)中之溶液中緩慢加入LiAlH4(955mg，25.14mmol，2.0eq.)。將反應混合物回流攪拌1小時。TLC顯示反應完成。冷卻至0℃後，藉由連續加入水(1.3mL)、15% NaOH水溶液(1.3mL)及水(3.9mL)使混合物淬滅。將所得混合物用EA稀釋，且藉由過濾除去沈澱物。減壓濃縮濾液，藉由FCC純化粗產物，得到呈黃色油狀之化合物22-3(2.3g，75.48%)。1H NMR(400MHz,CDCl3)δ：3.54(d,J=5.6Hz,2H),146-1.40(m,2H),1.27(s,24H),0.90-0.87(m,6H)。To a solution of compound 22-2 (3.4 g, 12.57 mmol, 1.0 eq.) in THF (60 mL) was slowly added LiAlH4 (955 mg, 25.14 mmol, 2.0 eq.) at 0°C. The reaction mixture was stirred at reflux for 1 hour. TLC showed that the reaction was complete. After cooling to 0°C, the mixture was quenched by sequentially adding water (1.3 mL), 15% aqueous NaOH solution (1.3 mL) and water (3.9 mL). The resulting mixture was diluted with EA, and the precipitate was removed by filtration. The filtrate was concentrated under reduced pressure, and the crude product was purified by FCC to give compound 22-3 (2.3 g, 75.48%) as a yellow oil.1 H NMR (400MHz, CDCl3 ) δ: 3.54 (d,J =5.6Hz, 2H), 146-1.40 (m, 2H), 1.27 (s, 24H), 0.90-0.87 (m, 6H).
步驟4：製備化合物22-4Step 4: Preparation of compound 22-4
向化合物22-3(1g，4.125mmol，10eq.)在DCM(15mL)中之溶液中加入6-溴己酸(0.966g，4.950mmol，1.2eq.)、EDCI(1.19g，6.188mmol，1.5eq.)、DMAP(101mg，0.8250mmol，0.2eq.)及DIEA(1.07g，8.250mmol，2.0eq.)。將反應混合物在50℃下攪拌16小時。TLC顯示反應完成。移除溶劑，且藉由FCC純化，得到呈黃色油狀之化合物22-4(1g，57.79%)。To a solution of compound 22-3 (1 g, 4.125 mmol, 10 eq.) in DCM (15 mL) were added 6-bromohexanoic acid (0.966 g, 4.950 mmol, 1.2 eq.), EDCI (1.19 g, 6.188 mmol, 1.5 eq.), DMAP (101 mg, 0.8250 mmol, 0.2 eq.) and DIEA (1.07 g, 8.250 mmol, 2.0 eq.). The reaction mixture was stirred at 50 °C for 16 hours. TLC showed that the reaction was complete. The solvent was removed and purified by FCC to give compound 22-4 (1 g, 57.79%) as a yellow oil.
步驟5：製備化合物22-5Step 5: Preparation of compound 22-5
向化合物22-4(0.33g，0.79mmol，1.0eq.)在ACN(15mL)中之溶液中加入乙醇胺(49mg，0.79mmol，1.0eq.)、K2CO3(329mg，2.384mmol，3.0eq.)、Cs2CO3(78mg，0.2384mmol，0.3eq.)及NaI(6mg，0.0397mmol，0.05eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，且藉由FCC純化，得到呈黃色油狀之化合物22-5(280mg，47.73%)。To a solution of compound 22-4 (0.33 g, 0.79 mmol, 1.0 eq.) in ACN (15 mL) were added ethanolamine (49 mg, 0.79 mmol, 1.0 eq.), K2 CO3 (329 mg, 2.384 mmol, 3.0 eq.), Cs2 CO3 (78 mg, 0.2384 mmol, 0.3 eq.) and NaI (6 mg, 0.0397 mmol, 0.05 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and purified by FCC to give compound 22-5 (280 mg, 47.73%) as a yellow oil.
步驟4：製備化合物22Step 4: Preparation of compound 22
在0℃下，向化合物22-5(230.0mg，0.53mmol，1.0eq.)及化合物6-2(257.0mg，0.64mmol，1.2eq.)在THF(10.0mL)中之溶液中加入DIEA(413.0mg，3.2mmol，5.0eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且藉由製備型HPLC純化，得到呈無色油狀之化合物22(100.0mg，24%產率)。At 0°C, DIEA (413.0 mg, 3.2 mmol, 5.0 eq.) was added to a solution of compound 22-5 (230.0 mg, 0.53 mmol, 1.0 eq.) and compound 6-2 (257.0 mg, 0.64 mmol, 1.2eq.) in THF (10.0 mL). The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, the mixture was evaporated under reduced pressure, and purified by preparative HPLC to obtain compound 22 (100.0 mg, 24% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,9H),1.26-1.35(m,52H),1.46-1.49(m,3H),1.60-1.65(m,8H),1.78(s,3H),2.28-2.32(m,5H),2.49-2.60(m,10H),3.54(s,2H),3.95-4.06(m,4H)。LCMS：Rt：1.250min；MS m/z(ESI)：793.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,9H),1.26-1.35(m,52H),1.46-1.49(m,3H),1.60-1.65(m,8H),1 .78(s,3H),2.28-2.32(m,5H),2.49-2.60(m,10H),3.54(s,2H),3.95-4.06(m,4H). LCMS: Rt: 1.250min; MS m/z (ESI): 793.7[M+H]+ .
6.14 實例14：製備化合物25.6.14 Example 14: Preparation of Compound 25.
步驟1：製備化合物25-2Step 1: Preparation of compound 25-2
向化合物25-1(5g，23.25mmol，1.0eq.)在CH3CN(200mL)中之混合物中加入BnNH2(5g，46.5mmol，2.0eq.)及K2CO3(9.64g，69.75mmol，3.0eq.)。將反應混合物在80℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈無色油狀之化合物25-2(3.0g，53%產率)。LCMS：Rt：0.740min；MS m/z(ESI)：242.1[M+H]+。To a mixture of compound 25-1 (5 g, 23.25 mmol, 1.0 eq.) in CH3 CN (200 mL) was added BnNH2 (5 g, 46.5 mmol, 2.0 eq.) and K2 CO3 (9.64 g, 69.75 mmol, 3.0 eq.). The reaction mixture was stirred at 80° C. for 10 hours. LCMS showed the reaction was complete. The solvent was removed and FCC was performed to give compound 25-2 (3.0 g, 53% yield) as a colorless oil. LCMS: Rt: 0.740 min; MS m/z (ESI): 242.1 [M+H]+ .
步驟2：製備化合物25-4Step 2: Preparation of compound 25-4
將化合物25-2(2.5g，10.36mmol，1.0eq.)、化合物25-3(5.56g，20.72mmol，2.0eq.)在EtOH(100mL)中之混合物在70℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物25-4(2.5g，47%產率)。LCMS：Rt：1.320min；MS m/z(ESI)：510.4[M+H]+。A mixture of compound 25-2 (2.5 g, 10.36 mmol, 1.0 eq.) and compound 25-3 (5.56 g, 20.72 mmol, 2.0 eq.) in EtOH (100 mL) was stirred at 70° C. for 10 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound 25-4 (2.5 g, 47% yield) as a yellow oil. LCMS: Rt: 1.320 min; MS m/z (ESI): 510.4 [M+H]+ .
步驟3：製備化合物25-5Step 3: Preparation of compound 25-5
在室溫、N2下，向NaH(710mg，17.65mmol，6.0eq.)在THF(60mL)中之混合物中加入化合物25-4(1.5g，2.94mmol，1.0eq.)。將反應混合物在室溫下攪拌2小時。向其中加入C8H17Br(2.27g，11.77mmol，4.0eq.)。將反應混合物在70℃下攪拌10小時。LCMS顯示反應完成。將混合物倒入水中並用EA洗滌。分離有機物，且經Na2SO4乾燥。移除溶劑，進行FCC，得到呈黃色油狀之化合物25-5(0.8g，43%產率)。LCMS：Rt：0.733min；MS m/z(ESI)：622.5[M+H]+。To a mixture of NaH (710 mg, 17.65 mmol, 6.0 eq.) in THF (60 mL) at room temperature underN2 was added compound 25-4 (1.5 g, 2.94 mmol, 1.0 eq.). The reaction mixture was stirred at room temperature for 2 hours.C8H17Br( 2.27 g, 11.77 mmol, 4.0 eq.) was added thereto. The reaction mixture was stirred at 70 °C for 10 hours. LCMS showed that the reaction was complete. The mixture was poured into water and washed with EA. The organics were separated and dried overNa2SO4 . The solvent was removed and FCC was performed to give compound 25-5 (0.8 g, 43% yield) asa yellow oil. LCMS: Rt: 0.733min; MS m/z (ESI): 622.5[M+H]+ .
步驟4：製備化合物25-6Step 4: Preparation of compound 25-6
向化合物25-5(0.8g，1.29mmol，1.0eq.)在乙酸乙酯(100mL)中之溶液中加入Pd/C(1.0g)。將反應混合物在室溫、H2下攪拌48小時。LCMS顯示反應完成。混合物藉由矽藻土過濾。移除溶劑，得到呈黃色油狀之化合物25-6(350mg，61%產率)。LCMS：Rt：1.040min；MS m/z(ESI)：442.4[M+H]+。Pd/C (1.0 g) was added to a solution of compound 25-5 (0.8 g, 1.29 mmol, 1.0 eq.) in ethyl acetate (100 mL). The reaction mixture was stirred at room temperature underH2 for 48 h. LCMS showed that the reaction was complete. The mixture was filtered through celite. The solvent was removed to give compound 25-6 (350 mg, 61% yield) as a yellow oil. LCMS: Rt: 1.040 min; MS m/z (ESI): 442.4 [M+H]+ .
步驟5：製備化合物25Step 5: Preparation of compound 25
向化合物25-6(350mg，0.8mmol，1.0eq.)、DIEA(200mg，1.6mmol，2.0eq.)在THF(20mL)中之混合物中加入化合物25-7(200mg，0.4mmol，0.5eq.)、NaI(60mg)。將反應混合物在70℃下攪拌10小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化殘餘物，得到呈黃色油狀之標題化合物(20mg，12%產率)。Compound 25-7 (200 mg, 0.4 mmol, 0.5 eq.) and NaI (60 mg) were added to a mixture of compound 25-6 (350 mg, 0.8 mmol, 1.0 eq.) and DIEA (200 mg, 1.6 mmol, 2.0 eq.) in THF (20 mL). The reaction mixture was stirred at 70 °C for 10 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to obtain the title compound (20 mg, 12% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.87(t,J=8Hz,12H),1.26-1.97(m,91H),2.19-2.64(m,10H),3.28-3.53(m,9H)。LCMS：Rt：0.627min；MS m/z(ESI)：919.8[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.87 (t,J =8Hz, 12H), 1.26-1.97 (m, 91H), 2.19-2.64 (m, 10H), 3.28-3.53 (m, 9H). LCMS: Rt: 0.627min; MS m/z (ESI): 919.8[M+H]+ .
6.15 實例15：製備化合物26.6.15 Example 15: Preparation of Compound 26.
步驟1：製備化合物26-2Step 1: Preparation of compound 26-2
在室溫下，向化合物26-1(500mg，1.12mmol，1.0eq.)及化合物SM1(170mg，2.24mmol，2.0eq.)在ACN(10mL)中之溶液中加入Cs2CO3(95mg，0.34mmol，0.3eq.)、K2CO3(465mg，3.36mmol，3.0eq.)及NaI(14mg，0.1mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈黃色油狀之化合物26-2(500mg，81%產率)。LCMS：Rt：1.680min；MS m/z(ESI)：442.4[M+H]+。To a solution of compound 26-1 (500 mg, 1.12 mmol, 1.0 eq.) and compound SM1 (170 mg, 2.24 mmol, 2.0 eq.) in ACN (10 mL) were added Cs2 CO3 (95 mg, 0.34 mmol, 0.3 eq.), K2 CO3 (465 mg, 3.36 mmol, 3.0 eq.) and NaI (14 mg, 0.1 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, the mixture was evaporated under reduced pressure, and purified by FCC (DCM/MeOH=1/0-20/1) to give compound 26-2 (500 mg, 81% yield) as a yellow oil. LCMS: Rt: 1.680min; MS m/z (ESI): 442.4[M+H]+ .
步驟2：製備化合物26-3Step 2: Preparation of compound 26-3
在室溫下，向化合物26-2(100mg，0.23mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(82mg，0.69mmol，3.0eq.)。將混合物在35℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈黃色油狀之化合物26-3(100mg，粗品)。LCMS：Rt：0.920min；MS m/z(ESI)：460.3[M+H]+。To a solution of compound 26-2 (100 mg, 0.23 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (82 mg, 0.69 mmol, 3.0 eq.) at room temperature. The mixture was stirred at 35° C. for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to give compound 26-3 (100 mg, crude) as a yellow oil. LCMS: Rt: 0.920 min; MS m/z (ESI): 460.3 [M+H]+ .
步驟3：製備化合物26Step 3: Preparation of compound 26
在0℃下，向化合物26-3(110mg，0.24mmol，1.0eq.)及化合物SM2(100mg，0.24mmol，1.0eq.)在THF(10mL)中之溶液中加入DIEA(413mg，3.2mmol，5.0eq.)及NaI(5mg，0.02mmol，0.1eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，且用製備型HPLC純化，得到呈無色油狀之化合物26(20mg，10%產率)。1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.39(m,59H),1.58-1.68(m,9H),2.29-2.34(m,4H),2.77-3.24(m,16H),3.73(s,2H),3.95-3.97(m,4H)。LCMS：Rt：1.760min；MS m/z(ESI)：851.8[M+H]+。To a solution of compound 26-3 (110 mg, 0.24 mmol, 1.0 eq.) and compound SM2 (100 mg, 0.24 mmol, 1.0 eq.) in THF (10 mL) were added DIEA (413 mg, 3.2 mmol, 5.0 eq.) and NaI (5 mg, 0.02 mmol, 0.1 eq.) at 0°C. The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, the mixture was evaporated under reduced pressure, and purified by preparative HPLC to give compound 26 (20 mg, 10% yield) as a colorless oil.1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.39(m,59H),1.58-1.68(m,9H),2.29-2.34(m,4H),2.77-3.24(m,16H),3.73(s,2H),3.95-3.97(m,4H). LCMS: Rt: 1.760min; MS m/z (ESI): 851.8[M+H]+ .
以下化合物係以與化合物26類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 26 using the corresponding starting materials.
6.16 實例16：製備化合物28.6.16 Example 16: Preparation of Compound 28.
步驟1：製備化合物28-2Step 1: Preparation of compound 28-2
將化合物28-1(1g，10mmol，1.0eq.)、SM6(0.9g，15mmol，1.5eq.)、兩滴AcOH在MeOH(20mL)中之混合物在室溫下攪拌隔夜。將NaBH3CN試劑(0.9g，15mmol，1.5eq.)加入混合物中，且攪拌1小時。將混合物用水淬滅，經乙酸乙酯萃取，濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物化合物28-2(893mg，61.6%產率)。LCMS：Rt：0.380min；MS m/z(ESI)：146.2[M+H]+。A mixture of compound 28-1 (1 g, 10 mmol, 1.0 eq.), SM6 (0.9 g, 15 mmol, 1.5 eq.), two drops of AcOH in MeOH (20 mL) was stirred at room temperature overnight. NaBH3 CN reagent (0.9 g, 15 mmol, 1.5 eq.) was added to the mixture and stirred for 1 hour. The mixture was quenched with water, extracted with ethyl acetate, concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product compound 28-2 (893 mg, 61.6% yield) as a yellow oil. LCMS: Rt: 0.380 min; MS m/z (ESI): 146.2 [M+H]+ .
步驟2：製備化合物28-3Step 2: Preparation of compound 28-3
將化合物26-1(250mg，0.56mmol，1.0eq.)、化合物28-1(243mg，1.68mmol，3.0eq.)、K2CO3(232mg，1.68mmol，3.0eq.)、Cs2CO3(7mg，0.02mmol，0.03eq.)及碘化鈉(30mg，0.2mmol，0.3eq.)在ACN(10mL)中之混合物在80℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物化合物28-3(122mg，42.7%產率)。LCMS：Rt：0.910min；MS m/z(ESI)：512.4[M+H]+。A mixture of compound 26-1 (250 mg, 0.56 mmol, 1.0 eq.), compound 28-1 (243 mg, 1.68 mmol, 3.0 eq.), K2 CO3 (232 mg, 1.68 mmol, 3.0 eq.), Cs2 CO3 (7 mg, 0.02 mmol, 0.03 eq.) and sodium iodide (30 mg, 0.2 mmol, 0.3 eq.) in ACN (10 mL) was stirred at 80° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product compound 28-3 (122 mg, 42.7% yield) as a yellow oil. LCMS: Rt: 0.910min; MS m/z (ESI): 512.4[M+H]+ .
步驟3：製備化合物28-4Step 3: Preparation of compound 28-4
將化合物28-3(122mg，0.24mmol，1.0eq.)及SOCl2(85mg，0.72mmol，3.0eq.)在DCM(5mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到呈黃色油狀之期望產物化合物28-4(125mg，粗品)。LCMS：Rt：1.350min；MS m/z(ESI)：530.4[M+H]+。A mixture of compound 28-3 (122 mg, 0.24 mmol, 1.0 eq.) and SOCl2 (85 mg, 0.72 mmol, 3.0 eq.) in DCM (5 mL) was stirred at 35° C. overnight. The mixture was concentrated to give the desired product compound 28-4 (125 mg, crude) as a yellow oil. LCMS: Rt: 1.350 min; MS m/z (ESI): 530.4 [M+H]+ .
步驟4：製備化合物28Step 4: Preparation of compound 28
將化合物28-4(125mg，0.24mmol，1.0eq.)、化合物SM2(100mg，0.23mmol，1.0eq.)、碘化鈉(20mg，0.13mmol，0.6eq.)及DIEA(155mg，1.20mmol，5.0eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物化合物28(23mg，10.6%產率)。A mixture of compound 28-4 (125 mg, 0.24 mmol, 1.0 eq.), compound SM2 (100 mg, 0.23 mmol, 1.0 eq.), sodium iodide (20 mg, 0.13 mmol, 0.6 eq.) and DIEA (155 mg, 1.20 mmol, 5.0 eq.) in THF (5 mL) was stirred at 70°C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to obtain the desired product compound 28 (23 mg, 10.6% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.82-0.94(m,12H),1.17-1.39(m,60H),1.49-1.73(m,13H),1.95-2.06(m,1H),2.17-2.37(m,6H),2.40-3.11(m,11H),3.27-3.45(m,2H),3.90-3.97(m,4H),3.99-4.09(m,2H)。LCMS：Rt：2.240min；MS m/z(ESI)：921.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.82-0.94(m,12H),1.17-1.39(m,60H),1.49-1.73(m,13H),1.95-2.06(m,1H),2.17- 2.37(m,6H),2.40-3.11(m,11H),3.27-3.45(m,2H),3.90-3.97(m,4H),3.99-4.09(m,2H). LCMS: Rt: 2.240min; MS m/z (ESI): 921.8[M+H]+ .
以下化合物係以與化合物28類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 28 using the corresponding starting materials.
6.17 實例17：製備化合物37.6.17 Example 17: Preparation of Compound 37.
步驟1：製備化合物37-1Step 1: Preparation of compound 37-1
將化合物SM2(200mg，0.47mmol，1.0eq.)、化合物SM3(154mg，0.93mmol，2.0eq.)、DIEA(300mg，2.35mmol，5.0eq.)在THF(20mL)中之混合物回流攪拌隔夜。LCMS顯示目標產物。將混合物濃縮，且用乙酸乙酯稀釋，用水及鹽水洗滌，經Na2SO4乾燥並濃縮。殘餘物藉由製備型HPLC純化，得到呈黃色油狀之化合物37-1(180mg，82%產率)。LCMS：Rt：0.940min；MS m/z(ESI)：519.4[M+H]+。A mixture of compound SM2 (200 mg, 0.47 mmol, 1.0 eq.), compound SM3 (154 mg, 0.93 mmol, 2.0 eq.), DIEA (300 mg, 2.35 mmol, 5.0 eq.) in THF (20 mL) was stirred under reflux overnight. LCMS showed the target product. The mixture was concentrated and diluted with ethyl acetate, washed with water and brine, dried over Na2 SO4 and concentrated. The residue was purified by preparative HPLC to give compound 37-1 (180 mg, 82% yield) as a yellow oil. LCMS: Rt: 0.940 min; MS m/z (ESI): 519.4 [M+H]+ .
步驟2：製備化合物37-2Step 2: Preparation of compound 37-2
將化合物37-1(180mg，0.35mmol，1.0eq.)及SOCl2(205mg，1.7mmol，5.0eq.)在DCM(5mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到呈黃色油狀之期望產物化合物37-2(210mg，粗品)。A mixture of compound 37-1 (180 mg, 0.35 mmol, 1.0 eq.) and SOCl2 (205 mg, 1.7 mmol, 5.0 eq.) in DCM (5 mL) was stirred at 35° C. overnight. The mixture was concentrated to give the desired product compound 37-2 (210 mg, crude) as a yellow oil.
步驟3：製備化合物37Step 3: Preparation of compound 37
將化合物37-2(210mg，0.35mmol，1.0eq.)、化合物SM2(180mg，0.42mmol，1.2eq.)、碘化鈉(15mg，0.1mmol，0.3eq.)及DIEA(135mg，1.1mmol，3.0eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈淡黃色油狀之期望產物化合物37(43mg，12.4%產率)。LCMS：Rt：1.740min；MS m/z(ESI)：928.7[M+H]+。A mixture of compound 37-2 (210 mg, 0.35 mmol, 1.0 eq.), compound SM2 (180 mg, 0.42 mmol, 1.2 eq.), sodium iodide (15 mg, 0.1 mmol, 0.3 eq.) and DIEA (135 mg, 1.1 mmol, 3.0 eq.) in THF (5 mL) was stirred at 70 °C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to give the desired product compound 37 (43 mg, 12.4% yield) as a light yellow oil. LCMS: Rt: 1.740 min; MS m/z (ESI): 928.7 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26(s,62H),1.38-1.43(m,5H),1.61-1.64(m,4H),2.26-2.30(m,4H),2.41-2.44(m,4H),2.51-2.57(m,6H),3.50-3.52(m,2H),3.57(s,2H),3.96(d,J=5.6Hz,4H),7.27-7.29(m,2H),8.53-8.54(m,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26(s,62H),1.38-1.43(m,5H),1.61-1.64(m,4H),2.26-2.30 (m,4H),2.41-2.44(m,4H),2.51-2.57(m,6H),3.50-3.52(m,2H),3.57(s,2H),3.96(d,J =5.6Hz,4H),7.27-7.29(m,2H),8.53-8.54(m,2H).
6.18 實例18：製備化合物43.6.18 Example 18: Preparation of Compound 43.
步驟1：製備化合物43-2Step 1: Preparation of compound 43-2
將化合物43-1(5.0g，14.3mmol，1.0eq.)、化合物D(2.5g，17.2mmol，1.2eq.)、K2CO3(5.9g，42.9mmol，3.0eq.)、Cs2CO3(1.4g，4.3mmol，0.3eq.)、NaI(645mg，0.43mmol，0.3eq.)在ACN(60mL)中之混合物回流攪拌隔夜。將混合物用水稀釋，用乙酸乙酯萃取，濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物化合物43-2(2.6g，44.2%產率)。LCMS：Rt：0.800min；MS m/z(ESI)：412.3[M+H]+。A mixture of compound 43-1 (5.0 g, 14.3 mmol, 1.0 eq.), compound D (2.5 g, 17.2 mmol, 1.2 eq.), K2 CO3 (5.9 g, 42.9 mmol, 3.0 eq.), Cs2 CO3 (1.4 g, 4.3 mmol, 0.3 eq.), and NaI (645 mg, 0.43 mmol, 0.3 eq.) in ACN (60 mL) was stirred under reflux overnight. The mixture was diluted with water, extracted with ethyl acetate, concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product compound 43-2 (2.6 g, 44.2% yield) as a yellow oil. LCMS: Rt: 0.800min; MS m/z (ESI): 412.3[M+H]+ .
步驟2：製備化合物43-3Step 2: Preparation of compound 43-3
將化合物43-2(400mg，0.97mmol，1.0eq.)及SOCl2(580mg，4.9mmol，5.0eq.)在DCM(10mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到呈黃色油狀之期望產物化合物43-3(440mg，粗品)。A mixture of compound 43-2 (400 mg, 0.97 mmol, 1.0 eq.) and SOCl2 (580 mg, 4.9 mmol, 5.0 eq.) in DCM (10 mL) was stirred at 35° C. overnight. The mixture was concentrated to give the desired product compound 43-3 (440 mg, crude) as a yellow oil.
步驟3：製備化合物43Step 3: Preparation of compound 43
將化合物SM4(210mg，0.47mmol，1.0eq.)、化合物43-3(240mg，0.57mmol，1.2eq.)、碘化鈉(21mg，0.1mmol，0.3eq.)及DIEA(182mg，1.4mmol，3.0eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈棕色油狀之期望產物化合物43(86mg，21.8%產率)。LCMS：Rt：1.190min；MS m/z(ESI)：835.7[M+H]+。A mixture of compound SM4 (210 mg, 0.47 mmol, 1.0 eq.), compound 43-3 (240 mg, 0.57 mmol, 1.2 eq.), sodium iodide (21 mg, 0.1 mmol, 0.3 eq.) and DIEA (182 mg, 1.4 mmol, 3.0 eq.) in THF (5 mL) was stirred at 70° C. overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to give the desired product compound 43 (86 mg, 21.8% yield) as a brown oil. LCMS: Rt: 1.190 min; MS m/z (ESI): 835.7 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.14-1.26(m,59H),1.44-1.46(m,4H),1.60-1.67(m,6H),1.77-1.79(m,4H),2.28-2.32(m,4H),2.42-2.50(m,8H),2.59(s,2H),3.52-3.53(m,2H),3.96(d,J=5.6Hz,2H),4.03-4.07(m,2H)。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 9H),1.14-1.26(m,59H),1.44-1.46(m,4H),1.60-1.67(m,6H),1.77-1.79(m,4H ),2.28-2.32(m,4H),2.42-2.50(m,8H),2.59(s,2H),3.52-3.53(m,2H),3.96(d,J =5.6Hz,2H),4.03-4.07(m,2H).
以下化合物係以與化合物43類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 43 using the corresponding starting materials.
6.19 實例19：製備化合物50.6.19 Example 19: Preparation of Compound 50.
步驟1：製備化合物50-2Step 1: Preparation of compound 50-2
向化合物50-1(600mg，5.60mmol，1.0eq.)在MeOH(30mL)中之溶液中加入化合物SM5(841mg，5.60mmol，1.0eq.)及AcOH(1滴)。將混合物在室溫下攪拌2小時。接著加入NaCNBH3(387mg，6.16mmol，1.1eq.)，且將所得混合物在室溫下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=30/1)純化，得到呈黃色油狀之標題化合物(810mg，60%產率)。LCMS：Rt：0.737min；MS m/z(ESI)：242.1[M+H]+。To a solution of compound 50-1 (600 mg, 5.60 mmol, 1.0 eq.) in MeOH (30 mL) were added compound SM5 (841 mg, 5.60 mmol, 1.0 eq.) and AcOH (1 drop). The mixture was stirred at room temperature for 2 hours. NaCNBH3 (387 mg, 6.16 mmol, 1.1 eq.) was then added, and the resulting mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=30/1) to give the title compound (810 mg, 60% yield) as a yellow oil. LCMS: Rt: 0.737 min; MS m/z (ESI): 242.1 [M+H]+ .
步驟2：製備化合物50-3Step 2: Preparation of compound 50-3
向化合物50-2(600mg，2.48mmol，1.0eq.)在MeOH(10mL)中之溶液中加入Pd/C(60mg)及濃HCl(3滴)。將混合物在室溫、H2下攪拌16小時。LCMS顯示反應完成。將混合物藉由矽藻土墊過濾並用MeOH洗滌。將濾液(filtration)濃縮，得到呈黃色油狀之標題化合物(345mg，91%產率)。LCMS：Rt：0.320min；MS m/z(ESI)：152.2[M+H]+。To a solution of compound 50-2 (600 mg, 2.48 mmol, 1.0 eq.) in MeOH (10 mL) was added Pd/C (60 mg) and concentrated HCl (3 drops). The mixture was stirred at room temperature underH2 for 16 h. LCMS showed the reaction was complete. The mixture was filtered through a diatomaceous earth pad and washed with MeOH. The filtration was concentrated to give the title compound (345 mg, 91% yield) as a yellow oil. LCMS: Rt: 0.320 min; MS m/z (ESI): 152.2 [M+H]+ .
步驟3：製備化合物50-4Step 3: Preparation of compound 50-4
向化合物50-3(345mg，2.28mmol，1.0eq.)及化合物43-1(797mg，2.28mmol，1.0eq.)在ACN(20mL)中之溶液中加入K2CO3(945mg，6.84mmol，3.0eq.)、Cs2CO3(223mg，0.684mmol，0.3eq.)及NaI(102mg，0.684mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=25/1)純化，得到呈黃色油狀之標題化合物(320mg，33%產率)。LCMS：Rt：0.880min；MS m/z(ESI)：420.3[M+H]+。To a solution of compound 50-3 (345 mg, 2.28 mmol, 1.0 eq.) and compound 43-1 (797 mg, 2.28 mmol, 1.0 eq.) in ACN (20 mL) were added K2 CO3 (945 mg, 6.84 mmol, 3.0 eq.), Cs2 CO3 (223 mg, 0.684 mmol, 0.3 eq.) and NaI (102 mg, 0.684 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=25/1) to give the title compound (320 mg, 33% yield) as a yellow oil. LCMS: Rt: 0.880min; MS m/z (ESI): 420.3[M+H]+ .
步驟4：製備化合物50-5Step 4: Preparation of compound 50-5
在0℃下，向化合物50-4(160mg，0.38mmol，1.0eq.)及DIPEA(98mg，0.76mmol，2.0eq.)在DCM(5mL)中之溶液中加入MsCl(52mg，0.46mmol，1.2eq.)。將混合物在室溫下攪拌2小時。LCMS顯示反應完成。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(176mg，93%產率)。其未經進一步純化即用於下一步驟。LCMS：Rt：0.800min；MS m/z(ESI)：402.3[M-OMs]+。To a solution of compound 50-4 (160 mg, 0.38 mmol, 1.0 eq.) and DIPEA (98 mg, 0.76 mmol, 2.0 eq.) in DCM (5 mL) at 0°C was added MsCl (52 mg, 0.46 mmol, 1.2 eq.). The mixture was stirred at room temperature for 2 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated to give the title compound (176 mg, 93% yield) as a yellow oil. It was used in the next step without further purification. LCMS: Rt: 0.800 min; MS m/z (ESI): 402.3 [M-OMs]+ .
步驟5：製備化合物50Step 5: Preparation of compound 50
向化合物50-5(176mg，0.35mmol，1.0eq.)及化合物SM2(150mg，0.35mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(226mg，1.75mmol，5.0eq.)及NaI(16mg，0.11mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈無色油狀之標題化合物(29mg，10%產率)。LCMS：Rt：1.430min；MS m/z(ESI)：829.6[M+H]+。To a solution of compound 50-5 (176 mg, 0.35 mmol, 1.0 eq.) and compound SM2 (150 mg, 0.35 mmol, 1.0 eq.) in THF (10 mL) was added DIPEA (226 mg, 1.75 mmol, 5.0 eq.) and NaI (16 mg, 0.11 mmol, 0.3 eq.). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound (29 mg, 10% yield) as a colorless oil. LCMS: Rt: 1.430 min; MS m/z (ESI): 829.6 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.26-1.30(m,49H),1.39-1.48(m,4H),1.58-1.68(m,8H),2.28-2.32(m,4H),2.40-2.70(m,12H),3.05-3.13(m,2H),3.49-3.65(m,2H),3.96-3.97(m,2H),4.04-4.06(m,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,9H),1.26-1.30(m,49H),1.39-1.48(m,4H),1.58-1.68(m,8H),2.28-2.32(m,4H) ),2.40-2.70(m,12H),3.05-3.13(m,2H),3.49-3.65(m,2H),3.96-3.97(m,2H),4.04-4.06(m,2H).
6.20 實例20：製備化合物56.6.20 Example 20: Preparation of Compound 56.
步驟1：製備化合物56-2Step 1: Preparation of compound 56-2
向化合物56-1(1.85g，15mmol，1.0eq.)在DCM(30mL)中之溶液中加入DIPEA(2.9g，22.5mmol，1.5eq.)及TBSCl(2.28g，15mmol，1.0eq.)。將混合物在室溫下攪拌2小時。將反應混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(PE/EA=50/1)純化殘餘物，得到呈無色油狀之標題化合物(1.2g，34%產率)。To a solution of compound 56-1 (1.85 g, 15 mmol, 1.0 eq.) in DCM (30 mL) were added DIPEA (2.9 g, 22.5 mmol, 1.5 eq.) and TBSCl (2.28 g, 15 mmol, 1.0 eq.). The mixture was stirred at room temperature for 2 hours. The reaction mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=50/1) to give the title compound (1.2 g, 34% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.15(s,6H),0.89(s,9H),6.84-6.86(m,2H),7.67-7.70(m,2H),9.79(s,1H)。1 H NMR (400MHz, CDCl3 ) δ: 0.15 (s, 6H), 0.89 (s, 9H), 6.84-6.86 (m, 2H), 7.67-7.70 (m, 2H), 9.79 (s, 1H).
步驟2：製備化合物56-3Step 2: Preparation of compound 56-3
向化合物56-2(1.2g，5.08mmol，1.0eq.)在MeOH(25mL)中之溶液中加入化合物SM6(465mg，7.62mmol，1.5eq.)及AcOH(3滴)。將混合物在室溫下攪拌2小時。接著加入NaCNBH3(383mg，6.10mmol，1.2eq.)，且將所得混合物在室溫下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=25/1)純化，得到呈無色油狀之標題化合物(629mg，45%產率)。LCMS：Rt：0.740min；MS m/z(ESI)：282.2[M+H]+。To a solution of compound 56-2 (1.2 g, 5.08 mmol, 1.0 eq.) in MeOH (25 mL) were added compound SM6 (465 mg, 7.62 mmol, 1.5 eq.) and AcOH (3 drops). The mixture was stirred at room temperature for 2 hours. NaCNBH3 (383 mg, 6.10 mmol, 1.2 eq.) was then added, and the resulting mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=25/1) to give the title compound (629 mg, 45% yield) as a colorless oil. LCMS: Rt: 0.740 min; MS m/z (ESI): 282.2 [M+H]+ .
步驟3：製備化合物56-4Step 3: Preparation of compound 56-4
向化合物56-3(629mg，2.23mmol，1.0eq.)及化合物26-1(1.0g，2.23mmol，1.0eq.)在ACN(40mL)中之溶液中加入K2CO3(924mg，6.69mmol，3.0eq.)、Cs2CO3(218mg，0.67mmol，0.3eq.)及NaI(100mg，0.67mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=40/1)純化，得到呈無色油狀之標題化合物(290mg，33%產率)。LCMS：Rt：1.030min；MS m/z(ESI)：648.4[M+H]+。To a solution of compound 56-3 (629 mg, 2.23 mmol, 1.0 eq.) and compound 26-1 (1.0 g, 2.23 mmol, 1.0 eq.) in ACN (40 mL) were added K2 CO3 (924 mg, 6.69 mmol, 3.0 eq.), Cs2 CO3 (218 mg, 0.67 mmol, 0.3 eq.) and NaI (100 mg, 0.67 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=40/1) to give the title compound (290 mg, 33% yield) as a colorless oil. LCMS: Rt: 1.030min; MS m/z (ESI): 648.4[M+H]+ .
步驟4：製備化合物56-5Step 4: Preparation of compound 56-5
向化合物56-4(290mg，0.45mmol，1.0eq.)及DIPEA(116mg，0.90mmol，2.0eq.)在DCM(6mL)中之溶液中加入MsCl(77mg，0.68mmol，1.5eq.)。將混合物在室溫下攪拌2小時。LCMS顯示反應完成。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(324mg，100%產率)。其未經進一步純化即用於下一步驟。LCMS：Rt：0.940min；MS m/z(ESI)：630.4[M-OMs]+。To a solution of compound 56-4 (290 mg, 0.45 mmol, 1.0 eq.) and DIPEA (116 mg, 0.90 mmol, 2.0 eq.) in DCM (6 mL) was added MsCl (77 mg, 0.68 mmol, 1.5 eq.). The mixture was stirred at room temperature for 2 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give the title compound (324 mg, 100% yield) as a yellow oil. It was used in the next step without further purification. LCMS: Rt: 0.940 min; MS m/z (ESI): 630.4 [M-OMs]+ .
步驟5：製備化合物56-6Step 5: Preparation of compound 56-6
向化合物56-5(324mg，0.45mmol，1.0eq.)及化合物SM2(192mg，0.45mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(174mg，1.35mmol，3.0eq.)及NaI(20mg，0.135mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=30/1)純化，得到呈黃色油狀之標題化合物(195mg，41%產率)。LCMS：Rt：0.640min；MS m/z(ESI)：1057.7[M+H]+。To a solution of compound 56-5 (324 mg, 0.45 mmol, 1.0 eq.) and compound SM2 (192 mg, 0.45 mmol, 1.0 eq.) in THF (10 mL) were added DIPEA (174 mg, 1.35 mmol, 3.0 eq.) and NaI (20 mg, 0.135 mmol, 0.3 eq.). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=30/1) to give the title compound (195 mg, 41% yield) as a yellow oil. LCMS: Rt: 0.640 min; MS m/z (ESI): 1057.7 [M+H]+ .
步驟6：製備化合物56Step 6: Preparation of compound 56
向化合物56-6(190mg，0.18mmol，1.0eq.)在DCM(8mL)中之溶液中加入在1,4-二
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.13-1.26(m,65H),1.53-1.65(m,8H),2.27-2.37(m,6H),2.47-2.69(m,7H),3.52-3.61(m,4H),3.96-4.00(m,4H),6.80-6.82(m,2H),7.18-7.20(m,2H)。LCMS：Rt：1.560min；MS m/z(ESI)：943.5[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.13-1.26(m,65H),1.53-1.65(m,8H),2.27-2.37(m,6H),2.47- 2.69(m,7H),3.52-3.61(m,4H),3.96-4.00(m,4H),6.80-6.82(m,2H),7.18-7.20(m,2H). LCMS: Rt: 1.560min; MS m/z (ESI): 943.5[M+H]+ .
6.21 實例21：製備化合物57.6.21 Example 21: Preparation of Compound 57.
步驟1：製備化合物57-2Step 1: Preparation of compound 57-2
將化合物57-1(500mg，3.0mmol，1eq.)、化合物SM7(982mg，4.5mmol，1.5eq.)及DIEA(1.16g，9.0mmol，3eq.)在DCM(10mL)中之混合物在室溫下攪拌1小時。將混合物用水淬滅，用乙酸乙酯萃取，濃縮並藉由矽膠管柱層析法(EA：PE=0%至5%)純化，得到呈黃色油狀之期望產物57-2(942mg，59.0%產率)。A mixture of compound 57-1 (500 mg, 3.0 mmol, 1 eq.), compound SM7 (982 mg, 4.5 mmol, 1.5 eq.) and DIEA (1.16 g, 9.0 mmol, 3 eq.) in DCM (10 mL) was stirred at room temperature for 1 hour. The mixture was quenched with water, extracted with ethyl acetate, concentrated and purified by silica gel column chromatography (EA:PE = 0% to 5%) to obtain the desired product 57-2 (942 mg, 59.0% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.83-0.93(m,3H),1.14-1.38(m,16H),1.45-1.55(m,2H),1.56-1.73(m,4H),1.82-1.95(m,2H),2.25-2.34(m,2H),3.37-3.48(m,2H),4.02-4.12(m,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.83-0.93(m,3H),1.14-1.38(m,16H),1.45-1.55(m,2H),1.56-1.73(m,4H) ),1.82-1.95(m,2H),2.25-2.34(m,2H),3.37-3.48(m,2H),4.02-4.12(m,2H).
步驟2：製備化合物57-3Step 2: Preparation of compound 57-3
將化合物57-2(600mg，1.68mmol，1.0eq.)、化合物D(291mg，2.01mmol，1.2eq.)、K2CO3(696mg，5.04mmol，3.0eq.)、Cs2CO3(21mg，0.05mmol，0.03eq.)及碘化鈉(90mg，0.6mmol，0.4eq.)在ACN(24mL)中之混合物在100℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至3%)純化，得到呈黃色油狀之期望產物57-3(344mg，29.1%產率)。A mixture of compound 57-2 (600 mg, 1.68 mmol, 1.0 eq.), compound D (291 mg, 2.01 mmol, 1.2 eq.), K2 CO3 (696 mg, 5.04 mmol, 3.0 eq.), Cs2 CO3 (21 mg, 0.05 mmol, 0.03 eq.) and sodium iodide (90 mg, 0.6 mmol, 0.4 eq.) in ACN (24 mL) was stirred at 100° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 3%) to give the desired product 57-3 (344 mg, 29.1% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.83-0.93(m,3H),1.14-1.44(m,26H),1.56-1.73(m,9H),2.25-2.33(m,2H),2.56-2.81(m,4H),3.36-3.45(m,2H),4.03-4.11(m,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.83-0.93(m,3H),1.14-1.44(m,26H),1.56-1.73(m,9H),2.25-2.33(m,2H),2.56-2.81(m,4H),3.36-3.45(m,2H),4.03-4.11(m,2H).
步驟3：製備化合物57-4Step 3: Preparation of compound 57-4
將化合物57-3(344mg，0.84mmol，1.0eq.)及SOCl2(298mg，2.51mmol，3.0eq.)在DCM(10mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到呈黃色油狀之期望產物57-4(364mg，粗品)。LCMS：Rt：0.840min；MS m/z(ESI)：430.3[M+H]+。A mixture of compound 57-3 (344 mg, 0.84 mmol, 1.0 eq.) and SOCl2 (298 mg, 2.51 mmol, 3.0 eq.) in DCM (10 mL) was stirred at 35° C. overnight. The mixture was concentrated to give the desired product 57-4 (364 mg, crude) as a yellow oil. LCMS: Rt: 0.840 min; MS m/z (ESI): 430.3 [M+H]+ .
步驟4：製備化合物57Step 4: Preparation of compound 57
將化合物57-4(172mg，0.4mmol，1.0eq.)、化合物SM11(150mg，0.35mmol，0.9eq.)、碘化鈉(30mg，0.2mmol，0.5eq.)及DIEA(155mg，1.2mmol，3.0eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物57(68mg，20.7%產率)。A mixture of compound 57-4 (172 mg, 0.4 mmol, 1.0 eq.), compound SM11 (150 mg, 0.35 mmol, 0.9 eq.), sodium iodide (30 mg, 0.2 mmol, 0.5 eq.) and DIEA (155 mg, 1.2 mmol, 3.0 eq.) in THF (5 mL) was stirred at 70°C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to obtain the desired product 57 (68 mg, 20.7% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.80-0.94(m,9H),1.14-1.39(m,53H),1.41-1.53(m,5H),1.56-1.69(m,8H),1.73-1.84(m,5H),2.18-2.32(m,4H),2.38-2.54(m,8H),2.56-2.62(m,2H),3.48-3.59(m,2H),3.48-3.59(m,4H)。LCMS：Rt：1.280min；MS m/z(ESI)：821.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.80-0.94(m,9H),1.14-1.39(m,53H),1.41-1.53(m,5H),1.56-1.69(m,8H),1.73-1.84(m,5H) ),2.18-2.32(m,4H),2.38-2.54(m,8H),2.56-2.62(m,2H),3.48-3.59(m,2H),3.48-3.59(m,4H). LCMS: Rt: 1.280min; MS m/z (ESI): 821.6[M+H]+ .
以下化合物係以與化合物57類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 57 using the corresponding starting materials.
6.22 實例22：製備化合物58.6.22 Example 22: Preparation of Compound 58.
步驟1：製備化合物58-2Step 1: Preparation of compound 58-2
將化合物58-1(500mg，2.5mmol，1.0eq.)、化合物SM8(283mg，2.8mmol，1.51eq.)、HATU(1.1g，2.8mmol，1.1eq.)及DIEA(483mg，3.8mmol，1.5eq.)在DCM(10mL)中之混合物在室溫下攪拌1小時。將混合物用水淬滅，用乙酸乙酯萃取，濃縮並藉由矽膠管柱層析法(EA：PE=0%至67%)純化，得到呈白色固體狀之期望產物58-2(693mg，97.2%產率)。LCMS：Rt：1.410min；MS m/z(ESI)：286.3[M+H]+。A mixture of compound 58-1 (500 mg, 2.5 mmol, 1.0 eq.), compound SM8 (283 mg, 2.8 mmol, 1.51 eq.), HATU (1.1 g, 2.8 mmol, 1.1 eq.) and DIEA (483 mg, 3.8 mmol, 1.5 eq.) in DCM (10 mL) was stirred at room temperature for 1 hour. The mixture was quenched with water, extracted with ethyl acetate, concentrated and purified by silica gel column chromatography (EA: PE = 0% to 67%) to give the desired product 58-2 (693 mg, 97.2% yield) as a white solid. LCMS: Rt: 1.410 min; MS m/z (ESI): 286.3 [M+H]+ .
步驟2：製備化合物58-3Step 2: Preparation of compound 58-3
將化合物58-2(300mg，1.05mmol，1.0eq.)、MsCl(144mg，1.26mmol，1.2eq.)及DIEA(204mg，1.58mmol，1.5eq.)在DCM(10mL)中之混合物在室溫下攪拌1小時。將混合物用水淬滅，用乙酸乙酯萃取，濃縮，得到呈黃色固體狀之期望產物58-3(382mg，粗品)。LCMS：Rt：1.110min；MS m/z(ESI)：364.2[M+H]+。A mixture of compound 58-2 (300 mg, 1.05 mmol, 1.0 eq.), MsCl (144 mg, 1.26 mmol, 1.2 eq.) and DIEA (204 mg, 1.58 mmol, 1.5 eq.) in DCM (10 mL) was stirred at room temperature for 1 hour. The mixture was quenched with water, extracted with ethyl acetate, and concentrated to give the desired product 58-3 (382 mg, crude) as a yellow solid. LCMS: Rt: 1.110 min; MS m/z (ESI): 364.2 [M+H]+ .
步驟3：製備化合物58-4Step 3: Preparation of compound 58-4
將化合物58-3(382mg，1.05mmol，1.0eq.)、化合物D(226mg，1.58mmol，1.5eq.)、K2CO3(435mg，3.15mmol，3.0eq.)、Cs2CO3(10mg，0.03mmol，0.03eq.)在ACN(10mL)中之混合物在100℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至3%)純化，得到呈黃色油狀之期望產物58-4(162mg，37.5%產率)。LCMS：Rt：0.750min；MS m/z(ESI)：411.3[M+H]+。A mixture of compound 58-3 (382 mg, 1.05 mmol, 1.0 eq.), compound D (226 mg, 1.58 mmol, 1.5 eq.), K2 CO3 (435 mg, 3.15 mmol, 3.0 eq.), Cs2 CO3 (10 mg, 0.03 mmol, 0.03 eq.) in ACN (10 mL) was stirred at 100° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 3%) to give the desired product 58-4 (162 mg, 37.5% yield) as a yellow oil. LCMS: Rt: 0.750 min; MS m/z (ESI): 411.3 [M+H]+ .
步驟4：製備化合物58-5Step 4: Preparation of compound 58-5
將化合物58-4(162mg，0.39mmol，1.0eq.)及SOCl2(140mg，1.18mmol，3.0eq.)在DCM(10mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到呈黃色油狀之期望產物58-5(187mg，粗品)。LCMS：Rt：0.780min；MS m/z(ESI)：429.3[M+H]+。A mixture of compound 58-4 (162 mg, 0.39 mmol, 1.0 eq.) and SOCl2 (140 mg, 1.18 mmol, 3.0 eq.) in DCM (10 mL) was stirred at 35° C. overnight. The mixture was concentrated to give the desired product 58-5 (187 mg, crude) as a yellow oil. LCMS: Rt: 0.780 min; MS m/z (ESI): 429.3 [M+H]+ .
步驟5：製備化合物58Step 5: Preparation of compound 58
將化合物58-5(187mg，0.4mmol，1.0eq.)、化合物SM9(100mg，0.2mmol，0.5eq.)、碘化鈉(30mg，0.2mmol，0.5eq.)及DIEA(155mg，1.2mmol，3.0eq.)在THF(5mL)中之溶液在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物58(22mg，6.2%產率)。LCMS：Rt：1.000min；MS m/z(ESI)：819.6[M+H]+。A solution of compound 58-5 (187 mg, 0.4 mmol, 1.0 eq.), compound SM9 (100 mg, 0.2 mmol, 0.5 eq.), sodium iodide (30 mg, 0.2 mmol, 0.5 eq.) and DIEA (155 mg, 1.2 mmol, 3.0 eq.) in THF (5 mL) was stirred at 70 °C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to give the desired product 58 (22 mg, 6.2% yield) as a yellow oil. LCMS: Rt: 1.000 min; MS m/z (ESI): 819.6 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.79-0.94(m,9H),1.08-1.37(m,53H),1.42-1.55(m,8H),1.57-1.65(m,5H),1.75-1.91(m,8H),2.03-2.10(m,2H),2.11-2.20(m,2H),2.43-2.64(m,9H),3.17-3.28(m,4H),3.48-3.62(m,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.79-0.94(m,9H),1.08-1.37(m,53H),1.42-1.55(m,8H),1.57-1.65(m,5H),1.75-1.91(m,8H) ),2.03-2.10(m,2H),2.11-2.20(m,2H),2.43-2.64(m,9H),3.17-3.28(m,4H),3.48-3.62(m,2H).
以下化合物係以與化合物58類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 58 using the corresponding starting materials.
6.23 實例23：製備化合物62.6.23 Example 23: Preparation of Compound 62.
步驟1：製備化合物62-2Step 1: Preparation of compound 62-2
向化合物62-1(0.4g，0.8939mmol，1.0eq.)在ACN(15mL)中之溶液中加入化合物B(124mg，1.073mmol，1.2eq.)、K2CO3(371mg，2.682mmol，3.0eq.)、Cs2CO3(87mg，0.2682mmol，0.3eq.)、NaI(13mg，0.08939mmol，0.1eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物62-2(330mg，79.28%)。LCMS：Rt：0.970min；MS m/z(ESI)：482.4[M+H]+。To a solution of compound 62-1 (0.4 g, 0.8939 mmol, 1.0 eq.) in ACN (15 mL) were added compound B (124 mg, 1.073 mmol, 1.2 eq.), K2 CO3 (371 mg, 2.682 mmol, 3.0 eq.), Cs2 CO3 (87 mg, 0.2682 mmol, 0.3 eq.), NaI (13 mg, 0.08939 mmol, 0.1 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound 62-2 (330 mg, 79.28%) as a yellow oil. LCMS: Rt: 0.970 min; MS m/z (ESI): 482.4 [M+H]+ .
步驟2：製備化合物62-3Step 2: Preparation of compound 62-3
向化合物62-2(200mg，0.4151mmol，1.0eq.)在DCM(15mL)中之溶液中加入SOCl2(148mg，1.245mmol，3.0eq.)。將反應混合物在35℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，得到呈黃色油狀之化合物62-3(200mg，粗品)。LCMS：Rt：1.140min；MS m/z(ESI)：500.4[M+H]+。To a solution of compound 62-2 (200 mg, 0.4151 mmol, 1.0 eq.) in DCM (15 mL) was added SOCl2 (148 mg, 1.245 mmol, 3.0 eq.). The reaction mixture was stirred at 35° C. for 16 hours. LCMS showed the reaction was complete. The solvent was removed to give compound 62-3 (200 mg, crude) as a yellow oil. LCMS: Rt: 1.140 min; MS m/z (ESI): 500.4 [M+H]+ .
步驟3：製備化合物62Step 3: Preparation of compound 62
向化合物62-3(200mg，0.3998mmol，1.14eq.)、DIEA(136mg，1.052mmol，3.0eq.)在THF(15mL)中之混合物中加入化合物SM11(150mg，0.3507mmol，1.0eq.)、NaI(15mg)。將反應混合物在75℃下攪拌64小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化殘餘物，得到呈黃色油狀之標題化合物(80mg，25.59%產率)。LCMS：Rt：1.790min；MS m/z(ESI)：891.7[M+H]+。To a mixture of compound 62-3 (200 mg, 0.3998 mmol, 1.14 eq.), DIEA (136 mg, 1.052 mmol, 3.0 eq.) in THF (15 mL) were added compound SM11 (150 mg, 0.3507 mmol, 1.0 eq.), NaI (15 mg). The reaction mixture was stirred at 75 °C for 64 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to give the title compound (80 mg, 25.59% yield) as a yellow oil. LCMS: Rt: 1.790 min; MS m/z (ESI): 891.7 [M+H]+ .
1H NMR(400MHz,CDCl3)：0.86-0.90(m,12H),1.26(s,59H),1.44-1.65(m,11H),1.83-2.00(m,7H),2.22(d,J=6.8Hz,4H),2.42-2.62(m,10H),3.08(s,1H),3.54-3.56(m,2H),4.03-4.07(m,4H)。1 H NMR (400MHz, CDCl3 ): 0.86-0.90(m,12H),1.26(s,59H),1.44-1.65(m,11H),1.83-2.00(m,7H),2.22(d,J =6.8Hz,4H),2.42-2.62(m,10H),3.08(s,1H),3.54-3.56(m,2H),4.03-4.07(m,4H).
6.24 實例24：製備化合物64.6.24 Example 24: Preparation of Compound 64.
步驟1：製備化合物64-2Step 1: Preparation of compound 64-2
向化合物64-1(2.0g，13.4mmol，1.0eq.)、DIEA(4.2g，32.1mmol，2.4eq.)在DCM(30mL)中之混合物中加入Boc2O(3.5g，16.0mmol，1.2eq.)。將混合物在室溫下攪拌2小時，TLC顯示反應完成。將混合物用DCM稀釋，用水及鹽水洗滌，乾燥，濃縮，且藉由管柱層析法純化殘餘物，得到呈白色固體狀之產物64-2(2.3g，80%產率)。To a mixture of compound 64-1 (2.0 g, 13.4 mmol, 1.0 eq.), DIEA (4.2 g, 32.1 mmol, 2.4 eq.) in DCM (30 mL) was added Boc2 O (3.5 g, 16.0 mmol, 1.2 eq.). The mixture was stirred at room temperature for 2 hours, and TLC showed that the reaction was complete. The mixture was diluted with DCM, washed with water and brine, dried, concentrated, and the residue was purified by column chromatography to give product 64-2 (2.3 g, 80% yield) as a white solid.
1H NMR(400MHz,CDCl3)δ：1.46(s,9H),1.62-1.72(m,2H),2.22-2.26(m,2H),2.38-2.46(m,4H),3.93(s,1H),4.50(s,1H)。1 H NMR (400MHz, CDCl3 ) δ: 1.46 (s, 9H), 1.62-1.72 (m, 2H), 2.22-2.26 (m, 2H), 2.38-2.46 (m, 4H), 3.93 (s, 1H), 4.50 (s, 1H).
步驟2：製備化合物64-3Step 2: Preparation of compound 64-3
將化合物64-2(2.3g，10.8mmol，1eq.)、化合物SM6(2.0g，32.3mmol，3.0eq.)、NaBH3CN(1.4g，21.7mmol，2.0eq.)在MeOH(30mL)中之混合物回流攪拌隔夜。LCMS顯示反應完成。混合物用乙酸乙酯稀釋，用水及鹽水洗滌，乾燥，濃縮，且藉由管柱層析法純化殘餘物，得到呈黃色油狀之產物64-3(1.6g，57%產率)。A mixture of compound 64-2 (2.3 g, 10.8 mmol, 1 eq.), compound SM6 (2.0 g, 32.3 mmol, 3.0 eq.), NaBH3 CN (1.4 g, 21.7 mmol, 2.0 eq.) in MeOH (30 mL) was stirred under reflux overnight. LCMS showed that the reaction was complete. The mixture was diluted with ethyl acetate, washed with water and brine, dried, concentrated, and the residue was purified by column chromatography to give product 64-3 (1.6 g, 57% yield) as a yellow oil.
步驟3：製備化合物64-4Step 3: Preparation of compound 64-4
在室溫下，向化合物64-3(1.6g，6.2mmol，1.0eq.)在THF(30mL)中之溶液中加入LAH(470mg，12.4mmol，2.0eq.)。將混合物回流攪拌2小時，LCMS顯示目標產物。將混合物用水淬滅，過濾，濃縮。殘餘物未經進一步純化即用於下一步驟。LCMS：Rt：0.290min；MS m/z(ESI)：173.2[M+H]+。To a solution of compound 64-3 (1.6 g, 6.2 mmol, 1.0 eq.) in THF (30 mL) was added LAH (470 mg, 12.4 mmol, 2.0 eq.) at room temperature. The mixture was stirred at reflux for 2 hours, and LCMS showed the target product. The mixture was quenched with water, filtered, and concentrated. The residue was used in the next step without further purification. LCMS: Rt: 0.290 min; MS m/z (ESI): 173.2 [M+H]+ .
步驟4：製備化合物64Step 4: Preparation of compound 64
將化合物64-4(60mg，0.35mmol，1.0eq.)、化合物26-1(390mg，0.87mmol，2.5eq.)、K2CO3(144mg，1.04mmol，3.0eq.)、Cs2CO3(33mg，0.1mmol，0.3eq.)、NaI(15mg，0.1mmol，0.3eq.)在ACN(10mL)中之混合物回流攪拌隔夜。LCMS顯示反應完成。將混合物用乙酸乙酯稀釋，用水及鹽水洗滌，乾燥，濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之產物64(14mg，4.4%產率)。LCMS：Rt：1.340min；MS m/z(ESI)：905.8[M+H]+。A mixture of compound 64-4 (60 mg, 0.35 mmol, 1.0 eq.), compound 26-1 (390 mg, 0.87 mmol, 2.5 eq.), K2 CO3 (144 mg, 1.04 mmol, 3.0 eq.), Cs2 CO3 (33 mg, 0.1 mmol, 0.3 eq.), and NaI (15 mg, 0.1 mmol, 0.3 eq.) in ACN (10 mL) was stirred under reflux overnight. LCMS showed that the reaction was complete. The mixture was diluted with ethyl acetate, washed with water and brine, dried, and concentrated. The residue was purified by preparative HPLC to give product 64 (14 mg, 4.4% yield) as a yellow oil. LCMS: Rt: 1.340min; MS m/z (ESI): 905.8[M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26(s,58H),1.42-1.47(m,6H),1.61-1.68(m,10H),1.78-1.86(m,5H),2.23(s,3H),2.29-2.32(m,5H),2.37-2.46(m,5H),2.57-2.60(m,2H),3.45-3.48(m,2H),3.97(d,J=6.0Hz,4H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26(s,58H),1.42-1.47(m,6H),1.61-1.68(m,10H),1.78-1.86(m,5H), 2.23(s,3H),2.29-2.32(m,5H),2.37-2.46(m,5H),2.57-2.60(m,2H),3.45-3.48(m,2H),3.97(d,J =6.0Hz,4H).
6.25 實例25：製備化合物65.6.25 Example 25: Preparation of Compound 65.
步驟1：製備化合物65-1Step 1: Preparation of compound 65-1
在室溫下，向化合物26-1(892.0mg，2.0mmol，1.0eq.)及化合物SM13(426.0mg，6.0mmol，3.0eq.)溶於ACN(10.0mL)中之溶液中加入Cs2CO3(195.0mg，0.6mmol，0.3eq.)、K2CO3(826.0mg，6.0mmol，3.0eq.)及NaI(29mg，0.2mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈黃色油狀之65-1(0.6g，68%產率)。LCMS：Rt：0.950min；MS m/z(ESI)：438.3[M+H]+。To a solution of compound 26-1 (892.0 mg, 2.0 mmol, 1.0 eq.) and compound SM13 (426.0 mg, 6.0 mmol, 3.0 eq.) dissolved in ACN (10.0 mL) were added Cs2 CO3 (195.0 mg, 0.6 mmol, 0.3 eq.), K2 CO3 (826.0 mg, 6.0 mmol, 3.0 eq.) and NaI (29 mg, 0.2 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=1/0-20/1) to give 65-1 (0.6 g, 68% yield) as a yellow oil. LCMS: Rt: 0.950min; MS m/z (ESI): 438.3[M+H]+ .
步驟2：製備化合物65-2Step 2: Preparation of compound 65-2
在室溫下，向化合物65-1(100.0mg，0.23mmol，1.0eq.)及化合物SM14(58.0mg，0.27mmol，1.2eq.)在THF(5.0mL)中之溶液中加入DIEA(44.0mg，0.34mmol，1.5eq.)。將混合物在50℃下攪拌16小時。LCMS顯示反應完成，蒸發混合物，得到呈棕色油狀之65-2(230.0mg，粗品)。LCMS：Rt：0.443min；MS m/z(ESI)：572.2[M+H]+。To a solution of compound 65-1 (100.0 mg, 0.23 mmol, 1.0 eq.) and compound SM14 (58.0 mg, 0.27 mmol, 1.2 eq.) in THF (5.0 mL) was added DIEA (44.0 mg, 0.34 mmol, 1.5 eq.) at room temperature. The mixture was stirred at 50°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated to give 65-2 (230.0 mg, crude) as a brown oil. LCMS: Rt: 0.443 min; MS m/z (ESI): 572.2 [M+H]+ .
步驟3：製備化合物65Step 3: Preparation of compound 65
在0℃下，向化合物65-2(230.0mg，0.4mmol，1.0eq.)及化合物SM2(207.0mg，0.48mmol，1.2eq.)在THF(5.0mL)中之溶液中加入DIEA(258mg，2.0mmol，5.0eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用製備型HPLC純化，得到呈黃色油狀之65(32.0mg，9%產率)。To a solution of compound 65-2 (230.0 mg, 0.4 mmol, 1.0 eq.) and compound SM2 (207.0 mg, 0.48 mmol, 1.2 eq.) in THF (5.0 mL) was added DIEA (258 mg, 2.0 mmol, 5.0 eq.) at 0°C. The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to obtain 65 (32.0 mg, 9% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.13-1.38(m,64H),1.41-1.70(m,13H),2.02-2.04(m,2H),2.28-2.39(m,4H),2.41-2.60(m,5H),3.09-3.13(m,4H),3.54-3.55(m,2H),3.96(d,J=0.4Hz,4H),5.62-5.66(m,2H)。LCMS：Rt：0.581min；MS m/z(ESI)：917.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.13-1.38(m,64H),1.41-1.70(m,13H),2.02-2.04(m,2H), 2.28-2.39(m,4H),2.41-2.60(m,5H),3.09-3.13(m,4H),3.54-3.55(m,2H),3.96(d,J =0.4Hz,4H),5.62-5.66(m,2H). LCMS: Rt: 0.581min; MS m/z (ESI): 917.6[M+H]+ .
6.26 實例26：製備化合物67.6.26 Example 26: Preparation of Compound 67.
步驟1：製備化合物67-2Step 1: Preparation of compound 67-2
將化合物67-1(200mg，1.45mmol，1.0eq.)、化合物SM1(196mg，1.31mmol，0.9eq.)在甲醇(5mL)中之混合物在室溫下攪拌2小時。加入NaBH3CN(190mg，3.0mmol，2.0eq.)。將混合物在室溫下攪拌隔夜。LCMS顯示目標產物。將混合物用乙酸乙酯稀釋，且用水及鹽水洗滌，乾燥，濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物67-2(240mg，69.5%產率)。LCMS：Rt：0.730min；MS m/z(ESI)：236.2[M+H]+。A mixture of compound 67-1 (200 mg, 1.45 mmol, 1.0 eq.), compound SM1 (196 mg, 1.31 mmol, 0.9 eq.) in methanol (5 mL) was stirred at room temperature for 2 hours. NaBH3 CN (190 mg, 3.0 mmol, 2.0 eq.) was added. The mixture was stirred at room temperature overnight. LCMS showed the target product. The mixture was diluted with ethyl acetate and washed with water and brine, dried, concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product 67-2 (240 mg, 69.5% yield) as a yellow oil. LCMS: Rt: 0.730 min; MS m/z (ESI): 236.2 [M+H]+ .
步驟2：製備化合物67-3Step 2: Preparation of compound 67-3
將化合物67-2(240mg，1.0mmol，1.0eq.)、化合物26-1(550mg，1.2mmol，1.2eq.)、K2CO3(420mg，3.0mmol，3.0eq.)、Cs2CO3(100mg，0.3mmol，0.3eq.)及NaI(45mg，0.3mmol，0.3eq.)在THF(10mL)中之混合物在90℃下攪拌隔夜。將混合物真空濃縮。藉由管柱層析法純化殘餘物，得到呈棕色油狀之期望產物67-3(230mg，38.3%產率)。LCMS：Rt：0.930min；MS m/z(ESI)：602.4[M+H]+。A mixture of compound 67-2 (240 mg, 1.0 mmol, 1.0 eq.), compound 26-1 (550 mg, 1.2 mmol, 1.2 eq.), K2 CO3 (420 mg, 3.0 mmol, 3.0 eq.), Cs2 CO3 (100 mg, 0.3 mmol, 0.3 eq.) and NaI (45 mg, 0.3 mmol, 0.3 eq.) in THF (10 mL) was stirred at 90° C. overnight. The mixture was concentrated in vacuo. The residue was purified by column chromatography to give the desired product 67-3 (230 mg, 38.3% yield) as a brown oil. LCMS: Rt: 0.930 min; MS m/z (ESI): 602.4 [M+H]+ .
步驟3：製備化合物67-4Step 3: Preparation of compound 67-4
將化合物67-3(230mg，0.38mmol，1.0eq.)、Pd/C(23mg)在乙酸乙酯(5mL)中之混合物在室溫、氫氣氣氛下攪拌隔夜，LCMS顯示反應完成。過濾混合物並濃縮。殘餘物未經進一步純化即用於下一步驟。LCMS：Rt：1.687min；MS m/z(ESI)：512.4[M+H]+。A mixture of compound 67-3 (230 mg, 0.38 mmol, 1.0 eq.) and Pd/C (23 mg) in ethyl acetate (5 mL) was stirred at room temperature under hydrogen atmosphere overnight. LCMS showed that the reaction was complete. The mixture was filtered and concentrated. The residue was used in the next step without further purification. LCMS: Rt: 1.687 min; MS m/z (ESI): 512.4 [M+H]+ .
步驟4：製備化合物67-5Step 4: Preparation of compound 67-5
將化合物67-4(180mg，0.35mmol，1.0eq.)、SOCl2(210mg，1.76mmol，5.0eq.)在DCM(5mL)中之混合物在40℃下攪拌4小時，LCMS顯示反應完成。將混合物濃縮，殘餘物未經進一步純化即用於下一步驟。A mixture of compound 67-4 (180 mg, 0.35 mmol, 1.0 eq.) and SOCl2 (210 mg, 1.76 mmol, 5.0 eq.) in DCM (5 mL) was stirred at 40° C. for 4 hours, and LCMS showed that the reaction was complete. The mixture was concentrated, and the residue was used in the next step without further purification.
步驟5：製備化合物67Step 5: Preparation of compound 67
將化合物67-5(220mg(粗品)，0.35mmol，1.0eq.)、化合物SM2(150mg，0.35mmol，1.0eq.)、DIEA(225mg，1.75mmol，5.0eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜，LCMS顯示反應完成。將混合物濃縮，且藉由製備型HPLC純化殘餘物，得到產物67(102mg，31.6%產率)。LCMS：Rt：1.910min；MS m/z(ESI)：921.7[M+H]+。A mixture of compound 67-5 (220 mg (crude), 0.35 mmol, 1.0 eq.), compound SM2 (150 mg, 0.35 mmol, 1.0 eq.), DIEA (225 mg, 1.75 mmol, 5.0 eq.) in THF (5 mL) was stirred at 70°C overnight. LCMS showed that the reaction was complete. The mixture was concentrated, and the residue was purified by preparative HPLC to give product 67 (102 mg, 31.6% yield). LCMS: Rt: 1.910 min; MS m/z (ESI): 921.7 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.30(m,60H),1.38-1.48(m,5H),1.59-1.67(m,7H),1.80-1.83(m,2H),2.28-2.32(m,4H),2.36-2.51(m,10H),2.53-2.60(m,3H),3.22-3.24(m,3H),3.52-3.57(m,2H),3.96(d,J=5.6Hz,4H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.30(m,60H),1.38-1.48(m,5H),1.59-1.67(m,7H),1.80-1.83(m,2H),2 .28-2.32(m,4H),2.36-2.51(m,10H),2.53-2.60(m,3H),3.22-3.24(m,3H),3.52-3.57(m,2H),3.96(d,J =5.6Hz,4H).
以下化合物係以與化合物67類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 67 using the corresponding starting materials.
6.27 實例27：製備化合物68.6.27 Example 27: Preparation of Compound 68.
步驟1：製備化合物26-1Step 1: Preparation of compound 26-1
將化合物SM(2.0g，7.394mmol，1.0eq.)、化合物W(2.2g，11.09mmol，1.5eq.)、TsOH(500mg)在甲苯(20mL)中之混合物回流攪拌2小時。TLC顯示反應完成。將混合物減壓蒸發並進行FCC，得到呈黃色油狀之化合物26-1(3g，90.90%)。A mixture of compound SM (2.0 g, 7.394 mmol, 1.0 eq.), compound W (2.2 g, 11.09 mmol, 1.5 eq.), and TsOH (500 mg) in toluene (20 mL) was stirred under reflux for 2 hours. TLC showed that the reaction was complete. The mixture was evaporated under reduced pressure and subjected to FCC to obtain compound 26-1 (3 g, 90.90%) as a yellow oil.
步驟2：製備化合物68-1Step 2: Preparation of compound 68-1
向化合物26-1(742mg，1.658mmol，1.0eq.)在ACN(15mL)中之溶液中加入化合物50-3(0.3g，1.658mmol，1.0eq.)、K2CO3(687mg，4.974mmol，3.0eq.)、Cs2CO3(162mg，0.4974mmol，0.3eq.)、NaI(25mg，0.1658mmol，0.1eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物68-1(400mg，46.59%)。LCMS：Rt：1.200min；MS m/z(ESI)：518.4[M+H]+。To a solution of compound 26-1 (742 mg, 1.658 mmol, 1.0 eq.) in ACN (15 mL) were added compound 50-3 (0.3 g, 1.658 mmol, 1.0 eq.), K2 CO3 (687 mg, 4.974 mmol, 3.0 eq.), Cs2 CO3 (162 mg, 0.4974 mmol, 0.3 eq.), NaI (25 mg, 0.1658 mmol, 0.1 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound 68-1 (400 mg, 46.59%) as a yellow oil. LCMS: Rt: 1.200 min; MS m/z (ESI): 518.4 [M+H]+ .
步驟3：製備化合物68-2Step 3: Preparation of compound 68-2
在0℃、N2下，向化合物68-1(200mg，0.3862mmol，1.0eq.)、DIEA(100mg，0.7724mmol，2.0eq.)在DCM(20mL)中之混合物中加入MsCl(53mg，0.4635mmol，1.2eq.)。將反應混合物在0℃下攪拌1小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈黃色油狀之化合物68-2(230mg，粗品)。To a mixture of compound 68-1 (200 mg, 0.3862 mmol, 1.0 eq.), DIEA (100 mg, 0.7724 mmol, 2.0 eq.) in DCM (20 mL) at 0°C underN2 was added MsCl (53 mg, 0.4635 mmol, 1.2 eq.). The reaction mixture was stirred at 0°C for 1 hour. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and driedoverNa2SO4 . The solvent was removed and FCC was performed to give compound 68-2 (230 mg, crude) as a yellow oil.
步驟4：製備化合物68Step 4: Preparation of compound 68
向化合物68-2(230mg，0.3860mmol，1.0eq.)、DIEA(125mg，0.9648mmol，3.0eq.)在THF(15mL)中之混合物中加入化合物SM2(138mg，0.3216mmol，1.0eq.)、NaI(15mg)。將反應混合物在75℃下攪拌16小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化殘餘物，得到呈黃色油狀之標題化合物(20mg，5.59%產率)。Compound SM2 (138 mg, 0.3216 mmol, 1.0 eq.) and NaI (15 mg) were added to a mixture of compound 68-2 (230 mg, 0.3860 mmol, 1.0 eq.) and DIEA (125 mg, 0.9648 mmol, 3.0 eq.) in THF (15 mL). The reaction mixture was stirred at 75 °C for 16 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to obtain the title compound (20 mg, 5.59% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)：0.86-0.88(m,12H),1.26(s,52H),1.43-1.63(m,19H),2.28-2.32(m,4H),2.45-2.59(m,14H),3.05(s,1H),3.53(s,2H),3.95-3.97(m,4H)。LCMS：Rt：1.970min；MS m/z(ESI)：927.6[M+H]+。1 H NMR (400MHz, CDCl3 ): 0.86-0.88(m,12H),1.26(s,52H),1.43-1.63(m,19H),2.28-2.32(m ,4H),2.45-2.59(m,14H),3.05(s,1H),3.53(s,2H),3.95-3.97(m,4H). LCMS: Rt: 1.970min; MS m/z (ESI): 927.6[M+H]+ .
以下化合物係以與化合物68類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 68 using the corresponding starting materials.
6.28 實例28：製備化合物71.6.28 Example 28: Preparation of Compound 71.
步驟1：製備化合物71-2Step 1: Preparation of compound 71-2
向NaOH(2.0g，50.3mmol，2.5eq.)在水(40mL)中之溶液中加入化合物71-1(2.18g，20.1mmol，1.0eq.)、1,4-二溴丁烷(10.0g，46.3mmol，2.3eq.)及四丁基硫酸氫銨(171mg，0.50mmol，0.025eq.)。將混合物在80℃下攪拌16小時。將反應混合物用EA萃取。合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。殘餘物藉由矽膠管柱層析法(PE/EA=50/1)純化，得到呈無色油狀之標題化合物(3.3g，67%產率)。To a solution of NaOH (2.0 g, 50.3 mmol, 2.5 eq.) in water (40 mL) were added compound 71-1 (2.18 g, 20.1 mmol, 1.0 eq.), 1,4-dibromobutane (10.0 g, 46.3 mmol, 2.3 eq.) and tetrabutylammonium hydrogen sulfate (171 mg, 0.50 mmol, 0.025 eq.). The mixture was stirred at 80 °C for 16 hours. The reaction mixture was extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=50/1) to give the title compound (3.3 g, 67% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：1.72-1.80(m,2H),1.94-2.01(m,2H),3.43(t,J=6.8Hz,2H),3.50(t,J=6.2Hz,2H),4.50(s,2H),7.27-7.37(m,5H)。1 H NMR (400MHz, CDCl3 ) δ: 1.72-1.80 (m, 2H), 1.94-2.01 (m, 2H), 3.43 (t,J =6.8Hz, 2H), 3.50 (t,J =6.2Hz, 2H), 4.50 (s, 2H), 7.27-7.37 (m, 5H).
步驟2：製備化合物71-3Step 2: Preparation of compound 71-3
向NaH(653mg，16.3mmol，1.2eq.)在THF(60mL)中之懸浮液中滴加丙二酸二甲酯(3.6g，27.2mmol，2.0eq.)。接著滴加化合物71-2(3.3g，13.6mmol，1.0eq.)在THF(10mL)中之溶液，且將所得混合物回流攪拌16小時。冷卻至室溫後，將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(PE/EA=20/1-5/1)純化殘餘物，得到呈無色油狀之標題化合物(3.2g，80%產率)。To a suspension of NaH (653 mg, 16.3 mmol, 1.2 eq.) in THF (60 mL) was added dimethyl malonate (3.6 g, 27.2 mmol, 2.0 eq.) dropwise. A solution of compound 71-2 (3.3 g, 13.6 mmol, 1.0 eq.) in THF (10 mL) was then added dropwise, and the resulting mixture was stirred at reflux for 16 hours. After cooling to room temperature, the mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=20/1-5/1) to give the title compound (3.2 g, 80% yield) as a colorless oil.
步驟3：製備化合物71-4Step 3: Preparation of compound 71-4
向LiAlH4(828mg，21.8mmol，2.0eq.)在THF(40mL)中之溶液中滴加化合物71-3(3.2g，10.9mmol，1.0eq.)在THF(20mL)中之溶液。將反應混合物在室溫下攪拌16小時。向反應混合物中小心地加入乙酸乙酯及水。加入6mL 2N NaOH水溶液。將混合物藉由矽藻土墊過濾並用EA洗滌。濾液經Na2SO4乾燥並藉由矽膠管柱層析法(DCM/MeOH=30/1-20/1)純化，得到呈無色油狀之標題化合物(1.6g，62%產率)。To a solution of LiAlH4 (828 mg, 21.8 mmol, 2.0 eq.) in THF (40 mL) was added dropwise a solution of compound 71-3 (3.2 g, 10.9 mmol, 1.0 eq.) in THF (20 mL). The reaction mixture was stirred at room temperature for 16 hours. Ethyl acetate and water were carefully added to the reaction mixture. 6 mL of 2N NaOH aqueous solution was added. The mixture was filtered through a celite pad and washed with EA. The filtrate was dried over Na2 SO4 and purified by silica gel column chromatography (DCM/MeOH=30/1-20/1) to give the title compound (1.6 g, 62% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：1.20-1.24(m,2H),1.36-1.44(m,2H),1.57-1.68(m,2H),1.72-1.75(m,1H),3.33(s,2H),3.45-3.49(m,2H),3.57-3.61(m,2H),3.73-3.76(m,2H),4.49(s,2H),7.27-7.34(m,5H)。1 H NMR (400MHz, CDCl3 ) δ: 1.20-1.24(m,2H), 1.36-1.44(m,2H), 1.57-1.68(m,2H), 1.72-1.75(m, 1H),3.33(s,2H),3.45-3.49(m,2H),3.57-3.61(m,2H),3.73-3.76(m,2H),4.49(s,2H),7.27-7.34(m,5H).
步驟4：製備化合物71-5Step 4: Preparation of compound 71-5
向化合物71-4(1.0g，4.2mmol，1.0eq.)及辛酸(1.8g，12.6mmol，3.0eq.)在甲苯(40mL)中之溶液中加入TsOH．H2O(36mg)。將混合物藉由Dean-Stark分水器回流攪拌4小時。將反應混合物濃縮並藉由矽膠管柱層析法(PE/EA=30/1)純化，得到呈無色油狀之標題化合物(926mg，46%產率)。To a solution of compound 71-4 (1.0 g, 4.2 mmol, 1.0 eq.) and octanoic acid (1.8 g, 12.6 mmol, 3.0 eq.) in toluene (40 mL) was added TsOH.H2 O (36 mg). The mixture was stirred under reflux for 4 hours via a Dean-Stark trap. The reaction mixture was concentrated and purified by silica gel column chromatography (PE/EA=30/1) to give the title compound (926 mg, 46% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,6H),1.27-1.29(m,13H),1.36-1.48(m,4H),1.58-1.64(m,9H),1.92-2.02(m,1H),2.29(t,J=7.6Hz,4H),3.46(t,J=6.4Hz,2H),4.00-4.10(m,4H),4.49(s,2H),7.28-7.37(m,5H)。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 6H), 1.27-1.29 (m, 13H), 1.36-1.48 (m, 4H), 1.58-1.64 (m, 9H), 1.92-2.02 (m, 1H), 2.29 (t,J =7.6Hz,4H),3.46(t,J =6.4Hz,2H),4.00-4.10(m,4H),4.49(s,2H),7.28-7.37(m,5H).
步驟5：製備71-6Step 5: Prepare 71-6
向化合物71-5(820mg，1.67mmol，1.0eq.)在MeOH(20mL)中之溶液中加入Pd/C(82mg)。將混合物在H2及35℃下攪拌36小時。將混合物藉由矽藻土墊過濾並用MeOH洗滌。將濾液濃縮，得到呈無色油狀之標題化合物(630mg，941%產率)。To a solution of compound 71-5 (820 mg, 1.67 mmol, 1.0 eq.) in MeOH (20 mL) was added Pd/C (82 mg). The mixture was stirred underH2 at 35 °C for 36 h. The mixture was filtered through a celite pad and washed with MeOH. The filtrate was concentrated to give the title compound (630 mg, 941% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,6H),1.27-1.39(m,15H),1.41-1.51(m,6H),1.58-1.65(m,6H),1.96-2.05(m,1H),2.30(t,J=7.6Hz,4H),3.65(t,J=6.4Hz,2H),4.02-4.11(m,4H)。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 6H), 1.27-1.39 (m, 15H), 1.41-1.51 (m, 6H), 1.58-1.65 (m, 6H), 1.96-2.05 (m, 1H), 2.30 (t,J =7.6Hz,4H),3.65(t,J =6.4Hz,2H),4.02-4.11(m,4H).
步驟6：製備化合物71-7Step 6: Preparation of compound 71-7
在0℃下，向化合物71-6(630mg，1.57mmol，1.0eq.)及DIPEA(406mg，3.14mmol，2.0eq.)在DCM(15mL)中之溶液中加入MsCl(216mg，1.88mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(682mg，91%產率)。其未經進一步純化即用於下一步驟。To a solution of compound 71-6 (630 mg, 1.57 mmol, 1.0 eq.) and DIPEA (406 mg, 3.14 mmol, 2.0 eq.) in DCM (15 mL) was added MsCl (216 mg, 1.88 mmol, 1.2 eq.) at 0°C. The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted withDCM . The combined organic layers were washed with brine, dried overNa2SO4 and concentrated to give the title compound (682 mg, 91% yield) as a yellow oil. It was used in the next step without further purification.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,6H),1.27-1.37(m,14H),1.41-1.46(m,4H),1.53-1.63(m,6H),1.73-1.80(m,2H),1.96-2.03(m,1H),2.30(t,J=6.2Hz,4H),3.01(s,3H),4.02-4.10(m,4H),4.23(t,J=6.4Hz,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,6H),1.27-1.37(m,14H),1.41-1.46(m,4H),1.53-1.63(m,6H),1.73-1.80(m,2H),1.96-2.03(m,1H),2.30(t,J =6.2Hz,4H),3.01(s,3H),4.02-4.10(m,4H),4.23(t,J =6.4Hz,2H).
步驟7：製備化合物71-8Step 7: Preparation of compound 71-8
向化合物71-7(260mg，0.54mmol，1.0eq.)及化合物B(62mg，0.54mmol，1.0eq.)在ACN(15mL)中之溶液中加入K2CO3(224mg，1.62mmol，3.0eq.)、Cs2CO3(52mg，0.16mmol，0.3eq.)及NaI(24mg，0.16mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=20/1)純化，得到呈黃色油狀之標題化合物(182mg，67%產率)。LCMS：Rt：0.830min；MS m/z(ESI)：498.4[M+H]+。To a solution of compound 71-7 (260 mg, 0.54 mmol, 1.0 eq.) and compound B (62 mg, 0.54 mmol, 1.0 eq.) in ACN (15 mL) were added K2 CO3 (224 mg, 1.62 mmol, 3.0 eq.), Cs2 CO3 (52 mg, 0.16 mmol, 0.3 eq.) and NaI (24 mg, 0.16 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=20/1) to give the title compound (182 mg, 67% yield) as a yellow oil. LCMS: Rt: 0.830min; MS m/z (ESI): 498.4[M+H]+ .
步驟8：製備化合物71-9Step 8: Preparation of compound 71-9
向化合物71-8(180mg，0.36mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(128mg，1.08mmol，3.0eq.)。將混合物在30℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮，得到呈黃色油狀之標題化合物(185mg，100%產率)。其未經進一步純化即用於下一步驟。LCMS：Rt：0.870min；MS m/z(ESI)：516.3[M+H]+。To a solution of compound 71-8 (180 mg, 0.36 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (128 mg, 1.08 mmol, 3.0 eq.). The mixture was stirred at 30 °C for 16 h. LCMS showed the reaction was complete. The mixture was concentrated to give the title compound (185 mg, 100% yield) as a yellow oil. It was used in the next step without further purification. LCMS: Rt: 0.870 min; MS m/z (ESI): 516.3 [M+H]+ .
步驟9：製備化合物71Step 9: Preparation of compound 71
向化合物71-9(160mg，0.31mmol，1.0eq.)及化合物SM16(138mg，0.31mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(120mg，0.93mmol，3.0eq.)及NaI(14mg，0.093mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(100mg，35%產率)。To a solution of compound 71-9 (160 mg, 0.31 mmol, 1.0 eq.) and compound SM16 (138 mg, 0.31 mmol, 1.0 eq.) in THF (10 mL) was added DIPEA (120 mg, 0.93 mmol, 3.0 eq.) and NaI (14 mg, 0.093 mmol, 0.3 eq.). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound (100 mg, 35% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.27-1.50(m,44H),1.57-1.67(m,10H),1.85-2.05(m,6H),2.28-2.36(m,8H),2.45-3.13(m,12H),3.52-3.60(m,2H),4.01-4.10(m,8H)。LCMS：Rt：1.110min；MS m/z(ESI)：923.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.27-1.50(m,44H),1.57-1.67(m,10H),1.85-2.05(m,6 H),2.28-2.36(m,8H),2.45-3.13(m,12H),3.52-3.60(m,2H),4.01-4.10(m,8H). LCMS: Rt: 1.110min; MS m/z (ESI): 923.7[M+H]+ .
以下化合物係以與化合物71類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 71 using the corresponding starting materials.
6.29 實例29：製備化合物72.6.29 Example 29: Preparation of Compound 72.
步驟1：製備化合物72-2Step 1: Preparation of compound 72-2
將化合物72-1(400mg，0.86mmol，1eq.)、化合物C(167mg，1.3mmol，1.5eq.)及K2CO3(359mg，2.6mmol，3eq.)、Cs2CO3(10mg，0.03mmol，0.03eq.)在ACN(10mL)中之混合物在80℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至3%)純化，得到呈黃色油狀之期望產物72-2(106mg，24.7%產率)。LCMS：Rt：0.810min；MS m/z(ESI)：495.4[M+H]+。A mixture of compound 72-1 (400 mg, 0.86 mmol, 1 eq.), compound C (167 mg, 1.3 mmol, 1.5 eq.), K2 CO3 (359 mg, 2.6 mmol, 3 eq.), Cs2 CO3 (10 mg, 0.03 mmol, 0.03 eq.) in ACN (10 mL) was stirred at 80° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 3%) to give the desired product 72-2 (106 mg, 24.7% yield) as a yellow oil. LCMS: Rt: 0.810 min; MS m/z (ESI): 495.4 [M+H]+ .
步驟2：製備化合物72-3Step 2: Preparation of compound 72-3
將72-2(106mg，0.2mmol，1eq.)及SOCl2(77mg，0.6mmol，3eq.)在DCM(5mL)中之混合物在35℃下攪拌隔夜。將混合物用水稀釋，用乙酸乙酯萃取，乾燥並濃縮後得到呈黃色油狀之期望產物72-3(117mg，粗品)。LCMS：Rt：0.880min；MS m/z(ESI)：513.4[M+H]+。A mixture of 72-2 (106 mg, 0.2 mmol, 1 eq.) and SOCl2 (77 mg, 0.6 mmol, 3 eq.) in DCM (5 mL) was stirred at 35° C. overnight. The mixture was diluted with water, extracted with ethyl acetate, dried and concentrated to give the desired product 72-3 (117 mg, crude) as a yellow oil. LCMS: Rt: 0.880 min; MS m/z (ESI): 513.4 [M+H]+ .
步驟3：製備化合物72Step 3: Preparation of compound 72
將化合物72-3(91mg，0.19mmol，1.0eq.)、化合物SM2(100mg，0.23mmol，1.2eq.)、K2CO3(79mg，0.57mmol，3.0eq.)、Cs2CO3(3mg，0.01mmol，0.03eq.)、NaI(15mg，0.10mmol，0.5eq.)在ACN(5mL)中之混合物在80℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物72(31mg，18.1%產率)。LCMS：Rt：1.510min；MS m/z(ESI)：904.7[M+H]+。A mixture of compound 72-3 (91 mg, 0.19 mmol, 1.0 eq.), compound SM2 (100 mg, 0.23 mmol, 1.2 eq.), K2 CO3 (79 mg, 0.57 mmol, 3.0 eq.), Cs2 CO3 (3 mg, 0.01 mmol, 0.03 eq.), NaI (15 mg, 0.10 mmol, 0.5 eq.) in ACN (5 mL) was stirred at 80° C. overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to give the desired product 72 (31 mg, 18.1% yield) as a yellow oil. LCMS: Rt: 1.510 min; MS m/z (ESI): 904.7 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.81-0.93(m,12H),1.07-1.38(m,62H),1.39-157(m,9H),1.58-1.90(m,11H),1.96-2.10(m,3H),2.16-2.26(m,2H),2.42-2.68(m,8H),3.18-3.32(m,2H),3.49-3.61(m,2H),3.99-4.12(m,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.81-0.93(m,12H),1.07-1.38(m,62H),1.39-157(m,9H),1.58-1.90(m,11H),1.96-2.10(m,3 H),2.16-2.26(m,2H),2.42-2.68(m,8H),3.18-3.32(m,2H),3.49-3.61(m,2H),3.99-4.12(m,2H).
以下化合物係以與化合物72類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 72 using the corresponding starting materials.
6.30 實例30：製備化合物76.6.30 Example 30: Preparation of Compound 76.
步驟1：製備化合物76-2Step 1: Preparation of compound 76-2
向化合物76-1(800mg，1.79mmol，1.0eq.)在ACN(50mL)中之溶液中加入化合物B(210mg，1.79mmol，1.0eq.)、K2CO3(750mg，5.37mmol，3.0eq.)、Cs2CO3(180mg，0.54mmol，0.3eq.)及NaI(80mg，0.54mmol，0.3eq.)。將反應混合物在80℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到化合物76-1(700mg，81%)。LCMS：Rt：0.870min；MS m/z(ESI)：481.4[M+H]+。To a solution of compound 76-1 (800 mg, 1.79 mmol, 1.0 eq.) in ACN (50 mL) were added compound B (210 mg, 1.79 mmol, 1.0 eq.), K2 CO3 (750 mg, 5.37 mmol, 3.0 eq.), Cs2 CO3 (180 mg, 0.54 mmol, 0.3 eq.) and NaI (80 mg, 0.54 mmol, 0.3 eq.). The reaction mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to give compound 76-1 (700 mg, 81%). LCMS: Rt: 0.870 min; MS m/z (ESI): 481.4 [M+H]+ .
步驟2：製備化合物76-3Step 2: Preparation of compound 76-3
向化合物76-2(200mg，0.41mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(150mg，1.25mmol，3.0eq.)。將反應混合物在35℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，得到呈黃色油狀之化合物76-3(207mg，100%產率)。LCMS：Rt：0.440min；MS m/z(ESI)：499.3[M+H]。To a solution of compound 76-2 (200 mg, 0.41 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (150 mg, 1.25 mmol, 3.0 eq.). The reaction mixture was stirred at 35 °C for 10 hours. LCMS showed the reaction was complete. The solvent was removed to give compound 76-3 (207 mg, 100% yield) as a yellow oil. LCMS: Rt: 0.440 min; MS m/z (ESI): 499.3 [M+H].
步驟3：製備化合物76Step 3: Preparation of compound 76
向化合物76-3(120mg，0.23mmol，1.0eq.)、DIEA(90mg，0.68mmol，3.0eq.)在THF(10mL)中之混合物中加入化合物SM15(100mg，0.23mmol，1.0eq.)、NaI(35mg)。將反應混合物在70℃下攪拌10小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化殘餘物，得到呈黃色油狀之標題化合物(35mg，17%產率)。Compound SM15 (100 mg, 0.23 mmol, 1.0 eq.) and NaI (35 mg) were added to a mixture of compound 76-3 (120 mg, 0.23 mmol, 1.0 eq.) and DIEA (90 mg, 0.68 mmol, 3.0 eq.) in THF (10 mL). The reaction mixture was stirred at 70 °C for 10 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to obtain the title compound (35 mg, 17% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)：0.87(t,J=8Hz,12H),1.26-2.00(m,79H),2.15-2.60(m,14H),3.16-3.19(m,3H),3.75-3.77(m,2H),3.95-3.97(m,2H),5.89(brs,1H)。LCMS：Rt：0.600min；MS m/z(ESI)：904.7[M+H]+。1 H NMR (400MHz, CDCl3 ): 0.87 (t,J =8Hz, 12H), 1.26-2.00 (m, 79H), 2.15-2.60 (m, 14H), 3.16-3.19 (m, 3H),3.75-3.77(m,2H),3.95-3.97(m,2H),5.89(brs,1H). LCMS: Rt: 0.600min; MS m/z (ESI): 904.7[M+H]+ .
以下化合物係以與化合物76類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 76 using the corresponding starting materials.
6.31 實例31：製備化合物78.6.31 Example 31: Preparation of Compound 78.
步驟1：製備化合物78-2Step 1: Preparation of compound 78-2
向化合物78-1(3.0g，13.44mmol，1.0eq.)在DMF(100mL)中之溶液中加入1,3-二側氧基異吲哚啉-2-化鉀(4.98g，26.89mmol，2.0eq.)。將反應混合物在100℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到化合物78-2(2.0g，53%)。LCMS：Rt：1.120min；MS m/z(ESI)：290.1[M+H]+。To a solution of compound 78-1 (3.0 g, 13.44 mmol, 1.0 eq.) in DMF (100 mL) was added 1,3-dioxoisoindoline-2-potassium (4.98 g, 26.89 mmol, 2.0 eq.). The reaction mixture was stirred at 100°C for 10 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound 78-2 (2.0 g, 53%). LCMS: Rt: 1.120 min; MS m/z (ESI): 290.1 [M+H]+ .
步驟2：製備化合物78-3Step 2: Preparation of compound 78-3
向化合物78-2(2.0g，6.91mmol，1.0eq.)在EtOH(50mL)中之溶液中加入NH2NH2.H2O(0.7g，13.82mmol，2.0eq.)。將反應混合物在90℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到化合物78-3(0.5g，45%)。LCMS：Rt：0.590min；MS m/z(ESI)：160.2[M+H]+。To a solution of compound 78-2 (2.0 g, 6.91 mmol, 1.0 eq.) in EtOH (50 mL) was added NH2 NH2 .H2 O (0.7 g, 13.82 mmol, 2.0 eq.). The reaction mixture was stirred at 90° C. for 10 hours. LCMS showed the reaction was complete. The solvent was removed and FCC was performed to give compound 78-3 (0.5 g, 45%). LCMS: Rt: 0.590 min; MS m/z (ESI): 160.2 [M+H]+ .
步驟3：製備化合物78-4Step 3: Preparation of compound 78-4
向化合物26-1(1.4g，3.14mmol，1.0eq.)在ACN(50mL)中之溶液中加入化合物78-3(500mg，3.14mmol，1.0eq.)、K2CO3(1.3g，9.42mmol，3.0eq.)、Cs2CO3(300mg，0.94mmol，0.3eq.)及NaI(140mg，0.94mmol，0.3eq.)。將反應混合物在80℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到化合物78-4(165mg，10%)。LCMS：Rt：0.920min；MS m/z(ESI)：526.4[M+H]+。To a solution of compound 26-1 (1.4 g, 3.14 mmol, 1.0 eq.) in ACN (50 mL) were added compound 78-3 (500 mg, 3.14 mmol, 1.0 eq.), K2 CO3 (1.3 g, 9.42 mmol, 3.0 eq.), Cs2 CO3 (300 mg, 0.94 mmol, 0.3 eq.) and NaI (140 mg, 0.94 mmol, 0.3 eq.). The reaction mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to give compound 78-4 (165 mg, 10%). LCMS: Rt: 0.920 min; MS m/z (ESI): 526.4 [M+H]+ .
步驟4：製備化合物78Step 4: Preparation of compound 78
向化合物78-4(140mg，0.27mmol，1.0eq.)、DIEA(100mg，0.80mmol，3.0eq.)在THF(10mL)中之混合物中加入化合物76-3(140mg，0.27mmol，1.0eq.)、NaI(40mg)。將反應混合物在70℃下攪拌10小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化殘餘物，得到呈黃色油狀之標題化合物(20mg，7%產率)。1H NMR(400MHz,CDCl3)：0.87(t,J=8Hz,12H),1.26-1.98(m,91H),2.08-2.41(m,14H),3.00-3.09(m,3H),3.55-3.58(m,2H),3.88-3.90(m,2H),5.89(brs,1H)。LCMS：Rt：0.613min；MS m/z(ESI)：988.7[M+H]+。To a mixture of compound 78-4 (140 mg, 0.27 mmol, 1.0 eq.), DIEA (100 mg, 0.80 mmol, 3.0 eq.) in THF (10 mL) were added compound 76-3 (140 mg, 0.27 mmol, 1.0 eq.), NaI (40 mg). The reaction mixture was stirred at 70 ° C for 10 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to give the title compound as a yellow oil (20 mg, 7% yield).1 H NMR (400MHz, CDCl3 ): 0.87 (t,J =8Hz,12H),1.26-1.98(m,91H),2.08-2.41(m,14H),3.00-3.09(m,3H),3.55-3.58(m,2H),3.88-3.90(m,2H),5.89(brs,1H). LCMS: Rt: 0.613min; MS m/z (ESI): 988.7[M+H]+ .
6.32 實例32：製備化合物79.6.32 Example 32: Preparation of Compound 79.
步驟1：製備化合物79-1Step 1: Preparation of compound 79-1
向化合物SM2(500mg，1.2mmol，1.0eq.)、DIEA(300mg，2.3mmol，2.0eq.)在DCM(5mL)中之混合物中加入Boc2O(306mg，1.4mmol，1.2eq.)。將混合物在室溫下攪拌30分鐘。TLC顯示反應完成。將混合物濃縮並藉由管柱層析法純化殘餘物，得到呈無色油狀之產物79-1(520mg，91%產率)。To a mixture of compound SM2 (500 mg, 1.2 mmol, 1.0 eq.), DIEA (300 mg, 2.3 mmol, 2.0 eq.) in DCM (5 mL) was added Boc2 O (306 mg, 1.4 mmol, 1.2 eq.). The mixture was stirred at room temperature for 30 minutes. TLC showed that the reaction was complete. The mixture was concentrated and the residue was purified by column chromatography to give the product 79-1 (520 mg, 91% yield) as a colorless oil.
步驟2：製備化合物79-2Step 2: Preparation of compound 79-2
向化合物79-1(520mg，1.0mmol，1.0eq.)、DIEA(260mg，2.0mmol，2.0eq.)在DCM(5mL)中之混合物中加入MsCl(140mg，1.2mmol，1.2eq.)。將混合物在室溫下攪拌30分鐘。TLC顯示反應完成。將混合物用水及鹽水稀釋，乾燥，濃縮，殘餘物未經進一步純化即用於下一步驟。MsCl (140 mg, 1.2 mmol, 1.2 eq.) was added to a mixture of compound 79-1 (520 mg, 1.0 mmol, 1.0 eq.) and DIEA (260 mg, 2.0 mmol, 2.0 eq.) in DCM (5 mL). The mixture was stirred at room temperature for 30 minutes. TLC showed that the reaction was complete. The mixture was diluted with water and brine, dried, concentrated, and the residue was used in the next step without further purification.
步驟3：製備化合物79-3Step 3: Preparation of compound 79-3
將化合物79-2(560mg(粗品)，1.0mmol，1.0eq.)、NaN3(100mg，1.5mmol，1.5eq.)在DMF(10mL)中之混合物在100℃下攪拌隔夜，TLC顯示反應完成。將混合物用乙酸乙酯稀釋，用水及鹽水洗滌，乾燥並濃縮。藉由管柱層析法純化殘餘物，得到呈無色油狀之化合物79-3(180mg，36%產率)。A mixture of compound 79-2 (560 mg (crude), 1.0 mmol, 1.0 eq.), NaN3 (100 mg, 1.5 mmol, 1.5 eq.) in DMF (10 mL) was stirred at 100° C. overnight. TLC showed that the reaction was complete. The mixture was diluted with ethyl acetate, washed with water and brine, dried and concentrated. The residue was purified by column chromatography to give compound 79-3 (180 mg, 36% yield) as a colorless oil.
步驟4：製備化合物79-4Step 4: Preparation of compound 79-4
將化合物79-3(180mg，0.33mmol，1.0eq.)、Pd/C(18mg)在乙酸乙酯(5mL)中之混合物在室溫下攪拌隔夜，LCMS顯示反應完成。將混合物過濾並濃縮。殘餘物未經進一步純化即用於下一步驟。LCMS：Rt：0.900min；MS m/z(ESI)：527.4[M+H]+。A mixture of compound 79-3 (180 mg, 0.33 mmol, 1.0 eq.) and Pd/C (18 mg) in ethyl acetate (5 mL) was stirred at room temperature overnight. LCMS showed that the reaction was complete. The mixture was filtered and concentrated. The residue was used in the next step without further purification. LCMS: Rt: 0.900 min; MS m/z (ESI): 527.4 [M+H]+ .
步驟5：製備化合物79-5Step 5: Preparation of compound 79-5
將化合物79-4(170mg，0.33mmol，1.0eq.)及化合物SM19(170mg，1.0mmol，3.0eq.)在DCM(5mL)中之混合物在室溫下攪拌隔夜，接著加入甲胺。將混合物攪拌24小時。加入TFA(5mL)。將混合物攪拌2小時並濃縮。殘餘物用乙酸乙酯稀釋並用飽和NaHCO3(水溶液)洗滌，乾燥，濃縮。殘餘物未經進一步純化即用於下一步驟。LCMS：Rt：0.900min；MS m/z(ESI)：536.4[M+H]+。A mixture of compound 79-4 (170 mg, 0.33 mmol, 1.0 eq.) and compound SM19 (170 mg, 1.0 mmol, 3.0 eq.) in DCM (5 mL) was stirred at room temperature overnight, and then methylamine was added. The mixture was stirred for 24 hours. TFA (5 mL) was added. The mixture was stirred for 2 hours and concentrated. The residue was diluted with ethyl acetate and washed with saturated NaHCO3 (aqueous solution), dried, and concentrated. The residue was used in the next step without further purification. LCMS: Rt: 0.900 min; MS m/z (ESI): 536.4 [M+H]+ .
步驟6：製備化合物79Step 6: Preparation of compound 79
將化合物79-5(90mg(粗品)，0.17mmol，1.0eq.)、化合物26-2(180mg，0.34mmol，2.0eq.)、DIEA(110mg，0.85mmol，5.0eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜，LCMS顯示反應完成。將混合物濃縮，且藉由製備型HPLC純化殘餘物，得到產物化合物79(26mg，18%產率)。A mixture of compound 79-5 (90 mg (crude), 0.17 mmol, 1.0 eq.), compound 26-2 (180 mg, 0.34 mmol, 2.0 eq.), and DIEA (110 mg, 0.85 mmol, 5.0 eq.) in THF (5 mL) was stirred at 70°C overnight, and LCMS showed that the reaction was complete. The mixture was concentrated, and the residue was purified by preparative HPLC to obtain the product compound 79 (26 mg, 18% yield).
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.30(m,59H),1.43-1.48(m,4H),1.57-1.65(m,6H),1.93(s,3H),2.21(s,3H),2.29-2.48(m,10H),2.54(s,2H),2.61-2.63(m,2H),3.30(d,J=4.2Hz,3H),3.67(d,J=5.6Hz,2H),3.94-3.97(m,4H)。LCMS：Rt：2.47min；MS m/z(ESI)：959.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.30(m,59H),1.43-1.48(m,4H),1.57-1.65(m,6H),1. 93(s,3H),2.21(s,3H),2.29-2.48(m,10H),2.54(s,2H),2.61-2.63(m,2H),3.30(d,J =4.2Hz,3H),3.67(d,J =5.6Hz,2H),3.94-3.97(m,4H). LCMS: Rt: 2.47min; MS m/z (ESI): 959.7[M+H]+ .
6.33 實例33：製備化合物80.6.33 Example 33: Preparation of Compound 80.
步驟1：製備化合物80-1Step 1: Preparation of compound 80-1
在0℃下，向化合物26-1(600mg，1.34mmol，1.0eq.)及化合物SM17(143mg，1.61mmol，1.2eq.)溶於ACN(10mL)中之溶液中加入Cs2CO3(130mg，0.40mmol，0.3eq.)、K2CO3(555mg，4.02mmol，3.0eq.)及NaI(18mg，0.13mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈黃色油狀之化合物80-1(520mg，86%產率)。LCMS：Rt：0.890min；MS m/z(ESI)：456.4[M+H]+。To a solution of compound 26-1 (600 mg, 1.34 mmol, 1.0 eq.) and compound SM17 (143 mg, 1.61 mmol, 1.2 eq.) in ACN (10 mL) were added Cs2 CO3 (130 mg, 0.40 mmol, 0.3 eq.), K2 CO3 (555 mg, 4.02 mmol, 3.0 eq.) and NaI (18 mg, 0.13 mmol, 0.1 eq.) at 0° C. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=1/0-20/1) to give compound 80-1 (520 mg, 86% yield) as a yellow oil. LCMS: Rt: 0.890min; MS m/z (ESI): 456.4[M+H]+ .
步驟2：製備化合物80-2Step 2: Preparation of compound 80-2
在室溫下，向化合物80-1(200mg，0.44mmol，1.0eq.)溶於DCM(10mL)中之溶液中加入SOCl2(264mg，2.20mmol，5.0eq.)。將混合物在35℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈黃色油狀之化合物80-2(200mg，粗品)。LCMS：Rt：0.940min；MS m/z(ESI)：474.3[M+H]+。To a solution of compound 80-1 (200 mg, 0.44 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (264 mg, 2.20 mmol, 5.0 eq.) at room temperature. The mixture was stirred at 35° C. for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to give compound 80-2 (200 mg, crude) as a yellow oil. LCMS: Rt: 0.940 min; MS m/z (ESI): 474.3 [M+H]+ .
步驟3：製備化合物80-3Step 3: Preparation of compound 80-3
在0℃下，向化合物80-2(300mg，0.63mmol，1.2eq.)及化合物SM18(280mg，0.53mmol，1.0eq.)在THF(10mL)中之溶液中加入DIEA(340mg，2.65mmol，5.0eq.)及NaI(7mg，0.05mmol，0.1eq.)。將混合物在75℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用製備型HPLC純化，得到呈無色油狀之化合物80-3(300mg，59%產率)。LCMS：Rt：1.710min；MS m/z(ESI)：963.7[M+H]+。To a solution of compound 80-2 (300 mg, 0.63 mmol, 1.2 eq.) and compound SM18 (280 mg, 0.53 mmol, 1.0 eq.) in THF (10 mL) at 0°C, DIEA (340 mg, 2.65 mmol, 5.0 eq.) and NaI (7 mg, 0.05 mmol, 0.1 eq.) were added. The mixture was stirred at 75°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to give compound 80-3 (300 mg, 59% yield) as a colorless oil. LCMS: Rt: 1.710 min; MS m/z (ESI): 963.7 [M+H]+ .
步驟4：製備化合物80-4Step 4: Preparation of compound 80-4
在室溫下，向化合物80-3(200mg，0.21mmol，1.0eq.)在DCM(4mL)中之溶液中加入TFA(1mL)。將混合物在室溫下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈無色油狀之化合物80-4(200mg，粗品)。LCMS：Rt：0.850min；MS m/z(ESI)：863.7[M+H]+。To a solution of compound 80-3 (200 mg, 0.21 mmol, 1.0 eq.) in DCM (4 mL) was added TFA (1 mL) at room temperature. The mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to give compound 80-4 (200 mg, crude) as a colorless oil. LCMS: Rt: 0.850 min; MS m/z (ESI): 863.7 [M+H]+ .
步驟5：製備化合物80-5Step 5: Preparation of compound 80-5
在室溫下，向化合物80-4(200mg，0.23mmol，1.0eq.)溶於DCM(5mL)中之溶液中加入化合物SM19(50mg，0.28mmol，1.2eq.)。將混合物在40℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈黃色油狀之化合物80-5(200mg，粗品)。LCMS：Rt：1.310min；MS m/z(ESI)：987.7[M+H]+。Compound SM19 (50 mg, 0.28 mmol, 1.2 eq.) was added to a solution of compound 80-4 (200 mg, 0.23 mmol, 1.0 eq.) in DCM (5 mL) at room temperature. The mixture was stirred at 40°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to obtain compound 80-5 (200 mg, crude) as a yellow oil. LCMS: Rt: 1.310 min; MS m/z (ESI): 987.7 [M+H]+ .
步驟6：製備化合物80Step 6: Preparation of compound 80
將CH3NH2(25mg，0.80mmol，4.0eq.)、化合物80-5(200mg，0.2mmol，1.0eq.)、DIEA(2mL)在DCM(10mL)中之混合物在室溫下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物化合物80(63mg，18%產率)。A mixture of CH3 NH2 (25 mg, 0.80 mmol, 4.0 eq.), compound 80-5 (200 mg, 0.2 mmol, 1.0 eq.), DIEA (2 mL) in DCM (10 mL) was stirred at room temperature overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to give the desired product compound 80 (63 mg, 18% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.77-0.98(m,12H),1.14-1.41(m,61H),1.42-1.57(m,6H),1.58-1.72(m,8H),2.13-2.26(m,3H),2.27-2.38(m,3H),2.39-2.56(m,6H),2.57-2.71(m,3H),3.04-3.20(m,3H),3.21-3.36(m,3H),3.67-3.83(m,2H),3.84-4.02(m,2H),5.69-5.83(m,1H)。LCMS：Rt：0.950min；MS m/z(ESI)：972.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.77-0.98(m,12H),1.14-1.41(m,61H),1.42-1.57(m,6H),1.58-1.72(m,8H),2.13-2.26(m,3H),2.27-2.38(m,3H),2.39- 2.56(m,6H),2.57-2.71(m,3H),3.04-3.20(m,3H),3.21-3.36(m,3H),3.67-3.83(m,2H),3.84-4.02(m,2H),5.69-5.83(m,1H). LCMS: Rt: 0.950min; MS m/z (ESI): 972.7[M+H]+ .
以下化合物係以與化合物80類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 80 using the corresponding starting materials.
6.34 實例34：製備化合物81.6.34 Example 34: Preparation of Compound 81.
步驟1：製備化合物81-1Step 1: Preparation of compound 81-1
將化合物26-1(2.0g，4.47mmol，1eq.)、(2-胺基乙基)胺基甲酸第三丁酯(1.1g，6.70mmol，1.5eq.)、K2CO3(1.8g，13.4mmol，3.0eq.)、Cs2CO3(440mg，1.34mmol，0.3eq.)、NaI(200mg，1.34mmol，0.3eq.)在ACN(20mL)中之混合物在90℃下攪拌隔夜。LCMS顯示目標產物。將混合物濃縮，且藉由管柱層析法純化殘餘物，得到呈黃色油狀之化合物81-1(1.4g，64%產率)。LCMS：Rt：0.960min；MS m/z(ESI)：527.4[M+H]+。A mixture of compound 26-1 (2.0 g, 4.47 mmol, 1 eq.), tert-butyl (2-aminoethyl)carbamate (1.1 g, 6.70 mmol, 1.5 eq.), K2 CO3 (1.8 g, 13.4 mmol, 3.0 eq.), Cs2 CO3 (440 mg, 1.34 mmol, 0.3 eq.), NaI (200 mg, 1.34 mmol, 0.3 eq.) in ACN (20 mL) was stirred at 90° C. overnight. LCMS showed the target product. The mixture was concentrated, and the residue was purified by column chromatography to give compound 81-1 (1.4 g, 64% yield) as a yellow oil. LCMS: Rt: 0.960min; MS m/z (ESI): 527.4[M+H]+ .
步驟2：製備化合物81-2Step 2: Preparation of compound 81-2
將化合物81-1(500mg，0.95mmol，1.0eq.)、化合物SM20(570mg，1.14mmol，1.2eq.)、DIEA(370mg，2.85mmol，3.0eq.)、NaI(44mg，0.29mmol，0.3eq.)在THF(10mL)中之混合物回流攪拌隔夜。LCMS顯示反應完成。將混合物用乙酸乙酯稀釋，用水及鹽水洗滌，乾燥，濃縮，且藉由管柱層析法純化殘餘物，得到呈黃色油狀之產物化合物81-2(610mg，72%產率)。LCMS：Rt：2.090min；MS m/z(ESI)：990.7[M+H]+。A mixture of compound 81-1 (500 mg, 0.95 mmol, 1.0 eq.), compound SM20 (570 mg, 1.14 mmol, 1.2 eq.), DIEA (370 mg, 2.85 mmol, 3.0 eq.), NaI (44 mg, 0.29 mmol, 0.3 eq.) in THF (10 mL) was stirred under reflux overnight. LCMS showed that the reaction was complete. The mixture was diluted with ethyl acetate, washed with water and brine, dried, concentrated, and the residue was purified by column chromatography to give the product compound 81-2 (610 mg, 72% yield) as a yellow oil. LCMS: Rt: 2.090 min; MS m/z (ESI): 990.7 [M+H]+ .
步驟3：製備化合物81-3Step 3: Preparation of compound 81-3
將化合物81-2(300mg，0.3mmol，1.0eq.)、TFA(345mg，3.0mmol，10.0eq.)在DCM(2mL)中之混合物在室溫下攪拌4小時，LCMS顯示反應完成。將混合物濃縮，殘餘物未經進一步純化即用於下一步驟。LCMS：Rt：1.420min；MS m/z(ESI)：890.7[M+H]+。A mixture of compound 81-2 (300 mg, 0.3 mmol, 1.0 eq.) and TFA (345 mg, 3.0 mmol, 10.0 eq.) in DCM (2 mL) was stirred at room temperature for 4 hours. LCMS showed that the reaction was complete. The mixture was concentrated and the residue was used in the next step without further purification. LCMS: Rt: 1.420 min; MS m/z (ESI): 890.7 [M+H]+ .
步驟4：製備化合物81Step 4: Preparation of compound 81
將化合物81-3(120mg，0.13mmol，1.0eq.)、DIEA(170mg，1.3mmol，10.0eq.)、化合物SM19(46mg，0.27mmol，2.0eq.)在DCM(5mL)中之混合物在室溫下攪拌隔夜，加入甲胺(0.67mmol，5.0eq.)。LCMS顯示反應完成。藉由製備型HPLC純化殘餘物，得到呈白色固體狀之產物化合物81(45mg，28%產率)。1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.46(m,66H),1.59-1.64(m,6H),2.21(s,6H),2.29-2.49(m,12H),2.61(s,2H),3.15(s,1H),3.31(d,J=5.2Hz,3H),3.66-3.75(m,2H),3.94-3.97(m,4H)。LCMS：Rt：0.093min；MS m/z(ESI)：999.7[M+H]+。A mixture of compound 81-3 (120 mg, 0.13 mmol, 1.0 eq.), DIEA (170 mg, 1.3 mmol, 10.0 eq.), and compound SM19 (46 mg, 0.27 mmol, 2.0 eq.) in DCM (5 mL) was stirred at room temperature overnight, and methylamine (0.67 mmol, 5.0 eq.) was added. LCMS showed that the reaction was complete. The residue was purified by preparative HPLC to obtain the product compound 81 (45 mg, 28% yield) as a white solid.1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.46(m,66H),1.59-1.64(m,6H),2.21(s,6H),2.29-2.49(m,12H),2.61(s,2H),3.15(s,1H),3.31(d,J =5.2Hz,3H),3.66-3.75(m,2H),3.94-3.97(m,4H). LCMS: Rt: 0.093min; MS m/z (ESI): 999.7[M+H]+ .
以下化合物係以與化合物81類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 81 using the corresponding starting materials.
6.35 實例35：製備化合物83.6.35 Example 35: Preparation of Compound 83.
步驟1：製備化合物83-2Step 1: Preparation of compound 83-2
將化合物83-1(1.4g，4.73mmol，1.0eq.)、化合物SM7(1.2g，7.10mmol，1.5eq.)、TsOH(20mg)在甲苯(70mL)中之混合物在180℃下攪拌2小時。TLC顯示反應完成。將混合物濃縮並藉由矽膠管柱層析法(EA：PE=0%至5%)純化，得到呈無色油狀之化合物83-2(1.0g，75%產率)。A mixture of compound 83-1 (1.4 g, 4.73 mmol, 1.0 eq.), compound SM7 (1.2 g, 7.10 mmol, 1.5 eq.), and TsOH (20 mg) in toluene (70 mL) was stirred at 180°C for 2 hours. TLC showed that the reaction was complete. The mixture was concentrated and purified by silica gel column chromatography (EA: PE = 0% to 5%) to obtain compound 83-2 (1.0 g, 75% yield) as a colorless oil.
步驟2：製備化合物83-3Step 2: Preparation of compound 83-3
將化合物83-2(700mg，1.58mmol，1.0eq.)及Pd/C(310mg，1.58mmol，1.0eq.)溶於MeOH(10mL)及乙酸乙酯(10mL)中之混合物在室溫、H2下攪拌16小時。TLC顯示反應完成。將混合物濃縮，得到呈無色油狀之期望產物化合物83-3(650mg，粗品)。A mixture of compound 83-2 (700 mg, 1.58 mmol, 1.0 eq.) and Pd/C (310 mg, 1.58 mmol, 1.0 eq.) dissolved in MeOH (10 mL) and ethyl acetate (10 mL) was stirred at room temperature underH2 for 16 hours. TLC showed that the reaction was complete. The mixture was concentrated to give the desired product compound 83-3 (650 mg, crude) as a colorless oil.
步驟3：製備化合物83-4Step 3: Preparation of compound 83-4
將化合物83-3(500mg，1.12mmol，1.0eq.)、化合物B(130mg，1.12mmol，1.0eq.)及K2CO3(465mg，3.36mmol，3.0eq.)、Cs2CO3(110mg，0.34mmol，0.3eq.)、NaI(15mg，0.11mmol，0.1eq.)在ACN(10mL)中之混合物在85℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物83-4(465mg，78%產率)。LCMS：Rt：0.900min；MS m/z(ESI)：482.4[M+H]+。A mixture of compound 83-3 (500 mg, 1.12 mmol, 1.0 eq.), compound B (130 mg, 1.12 mmol, 1.0 eq.), K2 CO3 (465 mg, 3.36 mmol, 3.0 eq.), Cs2 CO3 (110 mg, 0.34 mmol, 0.3 eq.), NaI (15 mg, 0.11 mmol, 0.1 eq.) in ACN (10 mL) was stirred at 85° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product 83-4 (465 mg, 78% yield) as a yellow oil. LCMS: Rt: 0.900 min; MS m/z (ESI): 482.4 [M+H]+ .
步驟4：製備化合物83-5Step 4: Preparation of compound 83-5
在室溫下，向化合物83-4(185mg，0.39mmol，1.0eq.)溶於DCM(10mL)中之溶液中加入SOCl2(230mg，2.00mmol，5.0eq.)。將混合物在35℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈黃色油狀之化合物83-5(185mg，粗品)。LCMS：Rt：0.930min；MS m/z(ESI)：500.4[M+H]+。To a solution of compound 83-4 (185 mg, 0.39 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (230 mg, 2.00 mmol, 5.0 eq.) at room temperature. The mixture was stirred at 35° C. for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to give compound 83-5 (185 mg, crude) as a yellow oil. LCMS: Rt: 0.930 min; MS m/z (ESI): 500.4 [M+H]+ .
步驟5：製備化合物83Step 5: Preparation of compound 83
將化合物SM2(205mg，0.48mmol，1.2eq.)、化合物83-5(200mg，0.4mmol，1.0eq.)、DIEA(260mg，2.0mmol，5.0eq.)、NaI(6mg，0.04mmol，0.1eq.)在THF(10mL)中之混合物在75℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物化合物83(63mg，18%產率)。A mixture of compound SM2 (205 mg, 0.48 mmol, 1.2 eq.), compound 83-5 (200 mg, 0.4 mmol, 1.0 eq.), DIEA (260 mg, 2.0 mmol, 5.0 eq.), and NaI (6 mg, 0.04 mmol, 0.1 eq.) in THF (10 mL) was stirred at 75°C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to obtain the desired product compound 83 (63 mg, 18% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.00-1.41(m,59H),1.42-1.52(m,4H),1.57-1.69(m,10H),1.71-2.12(m,4H),2.22-2.34(m,5H),2.35-2.44(m,3H),2.45-2.56(m,4H),2.57-2.61(m,2H),3.01-3.23(m,1H),3.51-3.55(m,2H),3.90-4.00(m,2H),4.01-4.11(m,2H)。LCMS：Rt：1.980min；MS m/z(ESI)：891.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.00-1.41(m,59H),1.42-1.52(m,4H),1.57-1.69(m,10H),1.71-2.12(m,4H),2.22-2.34(m,5H),2.35- 2.44(m,3H),2.45-2.56(m,4H),2.57-2.61(m,2H),3.01-3.23(m,1H),3.51-3.55(m,2H),3.90-4.00(m,2H),4.01-4.11(m,2H). LCMS: Rt: 1.980min; MS m/z (ESI): 891.7[M+H]+ .
6.36 實例36：製備化合物84.6.36 Example 36: Preparation of Compound 84.
步驟1：製備化合物84-2Step 1: Preparation of compound 84-2
將化合物84-1(1.2g，4mmol，1.0eq.)及LiOH．H2O(1.7g，40mmol，10.0eq.)與MeOH(30mL)及H2O(5mL)之混合物在70℃下攪拌隔夜。將混合物濃縮並用乙酸乙酯萃取，乾燥，濃縮，得到呈黃色油狀之化合物84-2(853mg，74.5%產率)。A mixture of compound 84-1 (1.2 g, 4 mmol, 1.0 eq.) and LiOH.H2 O (1.7 g, 40 mmol, 10.0 eq.) with MeOH (30 mL) and H2 O (5 mL) was stirred at 70° C. overnight. The mixture was concentrated and extracted with ethyl acetate, dried, and concentrated to give compound 84-2 (853 mg, 74.5% yield) as a yellow oil.
步驟2：製備化合物84-3Step 2: Preparation of compound 84-3
將化合物84-2(853mg，3.0mmol，1.0eq.)、化合物SM12(648mg，3.6mmol，1.2eq.)、TsOH(258mg，1.5mmol，0.5eq.)在甲苯中之混合物在180℃下攪拌2小時。將混合物濃縮並藉由矽膠管柱層析法(EA：PE=0%至5%)純化，得到呈無色油狀之化合物84-3(834mg，62.1%產率)。A mixture of compound 84-2 (853 mg, 3.0 mmol, 1.0 eq.), compound SM12 (648 mg, 3.6 mmol, 1.2 eq.), and TsOH (258 mg, 1.5 mmol, 0.5 eq.) in toluene was stirred at 180°C for 2 hours. The mixture was concentrated and purified by silica gel column chromatography (EA:PE = 0% to 5%) to obtain compound 84-3 (834 mg, 62.1% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.79-0.93(m,6H),1.11-1.33(m,25H),1.35-1.51(m,6H),1.59-1.70(m,3H),1.81-2.1(m,2H),2.26-2.35(m,1H),3.36-3.45(m,2H),4.02-4.11(m,2H)。1 H NMR (400MHz, CDCl3 )δ: 0.79-0.93(m,6H),1.11-1.33(m,25H),1.35-1.51(m,6H),1.59-1.70(m,3 H),1.81-2.1(m,2H),2.26-2.35(m,1H),3.36-3.45(m,2H),4.02-4.11(m,2H).
步驟3：製備化合物84-4Step 3: Preparation of compound 84-4
將化合物84-3(300mg，0.67mmol，1.0eq.)、化合物B(93mg，0.81mmol，1.2eq.)、K2CO3(278mg，2.01mmol，3eq.)、Cs2CO3(7mg，0.02mmol，0.03eq.)及碘化鈉(51mg，0.34mmol，0.5eq.)在ACN(10mL)中之混合物在80℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物84-4(314mg，97.2%產率)。LCMS：Rt：0.920min；MS m/z(ESI)：482.4[M+H]+。A mixture of compound 84-3 (300 mg, 0.67 mmol, 1.0 eq.), compound B (93 mg, 0.81 mmol, 1.2 eq.), K2 CO3 (278 mg, 2.01 mmol, 3 eq.), Cs2 CO3 (7 mg, 0.02 mmol, 0.03 eq.) and sodium iodide (51 mg, 0.34 mmol, 0.5 eq.) in ACN (10 mL) was stirred at 80° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product 84-4 (314 mg, 97.2% yield) as a yellow oil. LCMS: Rt: 0.920 min; MS m/z (ESI): 482.4 [M+H]+ .
步驟4：製備化合物84-5Step 4: Preparation of compound 84-5
將化合物84-4(314mg，0.65mmol，1.0eq.)及SOCl2(232mg，1.95mmol，3.0eq.)在DCM(10mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到期望產物84-5(343mg，粗品)。LCMS：Rt：1.080min；MS m/z(ESI)：500.3[M+H]+。A mixture of compound 84-4 (314 mg, 0.65 mmol, 1.0 eq.) and SOCl2 (232 mg, 1.95 mmol, 3.0 eq.) in DCM (10 mL) was stirred at 35° C. overnight. The mixture was concentrated to give the desired product 84-5 (343 mg, crude). LCMS: Rt: 1.080 min; MS m/z (ESI): 500.3 [M+H]+ .
步驟5：製備化合物84Step 5: Preparation of compound 84
將化合物84-5(152mg，0.3mmol，1.0eq.)、化合物SM2(130mg，0.3mmol，1.0eq.)、碘化鈉(23mg，0.15mmol，0.5eq.)及DIEA(116mg，0.9mmol，3.0eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈棕色油狀之期望產物84(54mg，19.9%產率)。1H NMR(400MHz,CDCl3)δ：0.80-0.94(m,12H),1.00-1.34(m,59H),1.36-1.50(m,6H),1.51-1.70(m,9H),1.78-2.05(m,5H),2.26-2.33(m,3H),2.36-2.45(m,3H),2.46-2.55(m,4H),2.57-2.63(m,2H),3.01-3.14(m,1H),3.49-3.59(m,2H),3.93-3.99(m,2H),4.01-4.10(m,2H)。LCMS：Rt：2.010min；MS m/z(ESI)：891.7[M+H]+。A mixture of compound 84-5 (152 mg, 0.3 mmol, 1.0 eq.), compound SM2 (130 mg, 0.3 mmol, 1.0 eq.), sodium iodide (23 mg, 0.15 mmol, 0.5 eq.) and DIEA (116 mg, 0.9 mmol, 3.0 eq.) in THF (5 mL) was stirred at 70 ° C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to give the desired product 84 (54 mg, 19.9% yield) as a brown oil.1 H NMR (400MHz, CDCl3 )δ: 0.80-0.94(m,12H),1.00-1.34(m,59H),1.36-1.50(m,6H),1.51-1.70(m,9H),1.78-2.05(m,5H),2.26-2.33(m,3H),2.36- 2.45(m,3H),2.46-2.55(m,4H),2.57-2.63(m,2H),3.01-3.14(m,1H),3.49-3.59(m,2H),3.93-3.99(m,2H),4.01-4.10(m,2H). LCMS: Rt: 2.010min; MS m/z (ESI): 891.7[M+H]+ .
以下化合物係以與化合物84類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 84 using the corresponding starting materials.
6.37 實例37：製備化合物86.6.37 Example 37: Preparation of Compound 86.
向化合物65(0.19g，0.21mmol，1.0eq.)在MeOH(5.0mL)中之溶液中加入Pd/C(30mg)，在室溫、H2下攪拌16小時。LCMS顯示反應完成，濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物86(60mg，31%產率)。To a solution of compound 65 (0.19 g, 0.21 mmol, 1.0 eq.) in MeOH (5.0 mL) was added Pd/C (30 mg) and stirred at room temperature underH2 for 16 h. LCMS showed the reaction was complete, concentrated and purified by preparative HPLC to give compound 86 (60 mg, 31% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.39(m,60H),1.43-1.62(m,9H),1.61-1.67(m,8H),1.77-2.00(m,4H),2.28-2.47(m,12H),2.56-2.58(m,2H),2.95-3.10(m,1H),3.51-3.54(m,2H),3.96(d,J=6.0Hz,4H)。LCMS：Rt：1.490min；MS m/z(ESI)：919.9[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.39(m,60H),1.43-1.62(m,9H),1.61-1.67(m,8H),1.77-2.00 (m,4H),2.28-2.47(m,12H),2.56-2.58(m,2H),2.95-3.10(m,1H),3.51-3.54(m,2H),3.96(d,J =6.0Hz,4H). LCMS: Rt: 1.490min; MS m/z (ESI): 919.9[M+H]+ .
6.38 實例38：製備化合物87.6.38 Example 38: Preparation of Compound 87.
步驟1：製備化合物87-2Step 1: Preparation of compound 87-2
在室溫下，向化合物83-2(500mg，1.12mmol，1.0eq.)及化合物B(130mg，1.12mmol，1.0eq.)溶於ACN(10mL)中之溶液中加入Cs2CO3(110mg，0.34mmol，0.3eq.)、K2CO3(465mg，3.36mmol，3.0eq.)及NaI(15mg，0.11mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並藉由FCC(DCM/MeOH=1/0-20/1)純化，得到呈黃色油狀之化合物87-2(470mg，85%產率)。LCMS：Rt：0.930min；MS m/z(ESI)：480.4[M+H]+。To a solution of compound 83-2 (500 mg, 1.12 mmol, 1.0 eq.) and compound B (130 mg, 1.12 mmol, 1.0 eq.) dissolved in ACN (10 mL) were added Cs2 CO3 (110 mg, 0.34 mmol, 0.3 eq.), K2 CO3 (465 mg, 3.36 mmol, 3.0 eq.) and NaI (15 mg, 0.11 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=1/0-20/1) to give compound 87-2 (470 mg, 85% yield) as a yellow oil. LCMS: Rt: 0.930min; MS m/z (ESI): 480.4[M+H]+ .
步驟2：製備化合物87-3Step 2: Preparation of compound 87-3
在0℃下，向化合物87-2(2.4g，5.5mmol，1.0eq.)及MsCl(55mg，0.46mmol，1.0eq.)在無水DCM(5mL)中之混合物中緩慢加入DIEA(90mg，0.70mmol，1.5eq.)。加入後，將混合物在室溫下攪拌2小時，TLC顯示反應完成。將混合物用水洗滌並濃縮。殘餘物(210mg)未經進一步純化即用於下一步驟。DIEA (90 mg, 0.70 mmol, 1.5 eq.) was slowly added to a mixture of compound 87-2 (2.4 g, 5.5 mmol, 1.0 eq.) and MsCl (55 mg, 0.46 mmol, 1.0 eq.) in anhydrous DCM (5 mL) at 0°C. After the addition, the mixture was stirred at room temperature for 2 hours, and TLC showed that the reaction was complete. The mixture was washed with water and concentrated. The residue (210 mg) was used in the next step without further purification.
步驟3：製備化合物87Step 3: Preparation of compound 87
在0℃下，向化合物87-3(150mg，0.27mmol，1.0eq.)及化合物SM2(115mg，0.27mmol，1.0eq.)在THF(5mL)中之溶液中加入DIEA(188mg，1.35mmol，5.0eq.)及NaI(5mg，0.03mmol，0.1eq.)。將混合物在75℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用製備型HPLC純化，得到呈無色油狀之化合物87(56mg，23%產率)。At 0°C, DIEA (188 mg, 1.35 mmol, 5.0 eq.) and NaI (5 mg, 0.03 mmol, 0.1 eq.) were added to a solution of compound 87-3 (150 mg, 0.27 mmol, 1.0 eq.) and compound SM2 (115 mg, 0.27 mmol, 1.0 eq.) in THF (5 mL). The mixture was stirred at 75°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to obtain compound 87 (56 mg, 23% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.39(m,49H),1.58-1.68(m,14H),1.98-2.22(m,10H),2.23-2.34(m,5H),2.38-2.72(m,9H),2.81-3.17(m,2H),3.45-3.65(m,2H),3.95-4.00(m,2H),4.01-4.05(m,2H),5.02-5.12(m,1H)。LCMS：Rt：1.680min；MS m/z(ESI)：889.7[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 12H), 1.26-1.39 (m, 49H), 1.58-1.68 (m, 14H), 1.98-2.22 (m, 10H),2.23-2.34(m,5H),2.38-2.72(m,9H),2.81-3.17(m,2H),3.45-3.65(m,2H),3.95-4.00(m,2H),4.01-4.05(m,2H),5.02-5.12(m,1H). LCMS: Rt: 1.680min; MS m/z (ESI): 889.7[M+H]+ .
6.39 實例39：製備化合物88.6.39 Example 39: Preparation of Compound 88.
步驟1：製備化合物88-2Step 1: Preparation of compound 88-2
向化合物88-1(1.34g，10.0mmol，1.0eq.)在DCE(20.0mL)中之溶液中加入化合物SM13(0.68g，10.0mmol，1.0eq.)及AcOH(0.7g，10.0mmol，1.0eq.)，在室溫下攪拌2小時，接著加入NaCNBH3(1.02g，15.0mmol，1.5eq.)，且在室溫下攪拌16小時。LCMS顯示反應完成，加入H2O，用DCM萃取，濃縮並藉由FCC(DCM/MeOH=20/1)純化，得到呈黃色油狀之化合物88-2(0.4g，21%產率)。LCMS：Rt：0.720min；MS m/z(ESI)：190.2[M+H]。To a solution of compound 88-1 (1.34 g, 10.0 mmol, 1.0 eq.) in DCE (20.0 mL) was added compound SM13 (0.68 g, 10.0 mmol, 1.0 eq.) and AcOH (0.7 g, 10.0 mmol, 1.0 eq.), stirred at room temperature for 2 hours, then added NaCNBH3 (1.02 g, 15.0 mmol, 1.5 eq.), and stirred at room temperature for 16 hours. LCMS showed that the reaction was completed, H2 O was added, extracted with DCM, concentrated and purified by FCC (DCM/MeOH=20/1) to give compound 88-2 (0.4 g, 21% yield) as a yellow oil. LCMS: Rt: 0.720min; MS m/z (ESI): 190.2 [M+H].
步驟2：製備化合物88-3Step 2: Preparation of compound 88-3
在室溫下，向化合物88-2(0.19g，1.0mmol，1.0eq.)及化合物26-1(446.0mg，1.0mmol，1.0eq.)在ACN(10.0mL)中之溶液中加入K2CO3(414mg，3.0mmol，3.0eq.)、Cs2CO3(97.5mg，0.3mmol，0.3eq.)及NaI(14.6mg，0.1mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用FCC(DCM/MeOH=30/1)純化，得到呈黃色油狀之化合物88-3(0.26g，46%產率)。LCMS：Rt：0.990min；MS m/z(ESI)：556.4[M+H]+。To a solution of compound 88-2 (0.19 g, 1.0 mmol, 1.0 eq.) and compound 26-1 (446.0 mg, 1.0 mmol, 1.0 eq.) in ACN (10.0 mL) were added K2 CO3 (414 mg, 3.0 mmol, 3.0 eq.), Cs2 CO3 (97.5 mg, 0.3 mmol, 0.3 eq.) and NaI (14.6 mg, 0.1 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=30/1) to give compound 88-3 (0.26 g, 46% yield) as a yellow oil. LCMS: Rt: 0.990min; MS m/z (ESI): 556.4[M+H]+ .
步驟3：製備化合物88Step 3: Preparation of compound 88
向化合物88-3(0.2g，0.36mmol，1.0eq.)在DCE(10.0mL)中之溶液中加入化合物SM2(0.18g，0.43mmol，1.2eq.)及AcOH(3滴)，且在室溫下攪拌2小時，接著加入NaBH(OAc)3(0.114g，0.54mmol，1.5eq.)，且在室溫下攪拌16小時。LCMS顯示反應完成，濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物88(70mg，20%產率)。1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.39(m,60H),1.43-1.62(m,13H),1.78-2.00(m,4H),2.25-2.30(m,5H),2.45-2.62(m,5H),3.10(s,1H),3.49-3.59(m,6H),3.96(d,J=5.2Hz,4H),7.16-7.22(m,4H)。LCMS：Rt：1.350min；MS m/z(ESI)：967.7[M+H]+。To a solution of compound 88-3 (0.2 g, 0.36 mmol, 1.0 eq.) in DCE (10.0 mL), compound SM2 (0.18 g, 0.43 mmol, 1.2 eq.) and AcOH (3 drops) were added, and stirred at room temperature for 2 hours, followed by the addition of NaBH(OAc)3 (0.114 g, 0.54 mmol, 1.5 eq.), and stirred at room temperature for 16 hours. LCMS showed that the reaction was complete, concentrated and purified by preparative HPLC to give compound 88 (70 mg, 20% yield) as a yellow oil.1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 12H), 1.26-1.39 (m, 60H), 1.43-1.62 (m, 13H), 1.78-2.00 (m, 4H),2.25-2.30(m,5H),2.45-2.62(m,5H),3.10(s,1H),3.49-3.59(m,6H),3.96(d,J =5.2Hz,4H),7.16-7.22(m,4H). LCMS: Rt: 1.350min; MS m/z (ESI): 967.7[M+H]+ .
6.40 實例40：製備化合物90.6.40 Example 40: Preparation of Compound 90.
步驟1：製備化合物90-2Step 1: Preparation of compound 90-2
將90-1(5g，25.9mmol，3.0eq.)、PMBNH2(1.2g，8.6mmol，1.0eq.)及K2CO3(3.6g，25.9mmol，3.0eq.)、Cs2CO3(100mg，0.3mmol，0.03eq.)及NaI(0.6g，4.3mmol，0.5eq.)在ACN(40mL)中之混合物在80℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物90-2(2.5g，粗品)。LCMS：Rt：0.810min；MS m/z(ESI)：362.3[M+H]+。A mixture of 90-1 (5 g, 25.9 mmol, 3.0 eq.), PMBNH2 (1.2 g, 8.6 mmol, 1.0 eq.), K2 CO3 (3.6 g, 25.9 mmol, 3.0 eq.), Cs2 CO3 (100 mg, 0.3 mmol, 0.03 eq.), and NaI (0.6 g, 4.3 mmol, 0.5 eq.) in ACN (40 mL) was stirred at 80° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product 90-2 (2.5 g, crude) as a yellow oil. LCMS: Rt: 0.810 min; MS m/z (ESI): 362.3 [M+H]+ .
步驟2：製備化合物90-3Step 2: Preparation of compound 90-3
將化合物90-2(2.5g，6.9mmol，1.0eq.)及Pd/C(10%，1g)溶於MeOH(100mL)及乙酸乙酯(10mL)中之混合物在50℃下攪拌5天。將混合物濃縮，得到呈黃色半固體狀之期望產物90-3(1.48g，粗品)。LCMS：Rt：0.780min；MS m/z(ESI)：242.3[M+H]+。A mixture of compound 90-2 (2.5 g, 6.9 mmol, 1.0 eq.) and Pd/C (10%, 1 g) dissolved in MeOH (100 mL) and ethyl acetate (10 mL) was stirred at 50° C. for 5 days. The mixture was concentrated to give the desired product 90-3 (1.48 g, crude) as a yellow semisolid. LCMS: Rt: 0.780 min; MS m/z (ESI): 242.3 [M+H]+ .
步驟3：製備化合物90-4Step 3: Preparation of compound 90-4
將化合物90-3(242mg，1mmol，1.0eq.)、化合物W(214mg，1.1mmol，1.1eq.)、DIEA(194mg，3mmol，3.0eq.)及HATU(418mg，1.1mmol，1.1eq.)在DCM(5mL)中之混合物在室溫下攪拌1小時。將混合物用水稀釋，經乙酸乙酯萃取，用鹽水洗滌，乾燥，濃縮，且經矽膠管柱層析法(EA：PE=0%至33%)純化，得到呈黃色油狀之期望產物90-4(298mg，71.2%產率)。LCMS：Rt：2.250min；MS m/z(ESI)：418.2,420.2[M+H]+。A mixture of compound 90-3 (242 mg, 1 mmol, 1.0 eq.), compound W (214 mg, 1.1 mmol, 1.1 eq.), DIEA (194 mg, 3 mmol, 3.0 eq.) and HATU (418 mg, 1.1 mmol, 1.1 eq.) in DCM (5 mL) was stirred at room temperature for 1 hour. The mixture was diluted with water, extracted with ethyl acetate, washed with brine, dried, concentrated, and purified by silica gel column chromatography (EA: PE = 0% to 33%) to give the desired product 90-4 (298 mg, 71.2% yield) as a yellow oil. LCMS: Rt: 2.250 min; MS m/z (ESI): 418.2, 420.2 [M+H]+ .
步驟4：製備化合物90-5Step 4: Preparation of compound 90-5
將化合物90-4(298mg，0.71mmol，1.0eq.)、化合物SM6(130mg，2.13mmol，3.0eq.)及K2CO3(295mg，2.13mmol，3.0eq.)、Cs2CO3(7mg，0.02mmol，0.03eq.)、NaI(53mg，0.35mmol，0.5eq.)在ACN(12mL)中之混合物在80℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物90-5(150mg，52.8%產率)。LCMS：Rt：0.800min；MS m/z(ESI)：399.3[M+H]+。A mixture of compound 90-4 (298 mg, 0.71 mmol, 1.0 eq.), compound SM6 (130 mg, 2.13 mmol, 3.0 eq.), K2 CO3 (295 mg, 2.13 mmol, 3.0 eq.), Cs2 CO3 (7 mg, 0.02 mmol, 0.03 eq.), NaI (53 mg, 0.35 mmol, 0.5 eq.) in ACN (12 mL) was stirred at 80° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product 90-5 (150 mg, 52.8% yield) as a yellow oil. LCMS: Rt: 0.800 min; MS m/z (ESI): 399.3 [M+H]+ .
步驟5：製備化合物90Step 5: Preparation of compound 90
將化合物43-3(170mg，0.3mmol，1.0eq.)、化合物90-5(150mg，0.4mmol，1.3eq.)、DIEA(116mg，0.9mmol，3.0eq.)、NaI(23mg，0.15mmol，0.5eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物90(82mg，34.5%產率)。A mixture of compound 43-3 (170 mg, 0.3 mmol, 1.0 eq.), compound 90-5 (150 mg, 0.4 mmol, 1.3 eq.), DIEA (116 mg, 0.9 mmol, 3.0 eq.), and NaI (23 mg, 0.15 mmol, 0.5 eq.) in THF (5 mL) was stirred at 70°C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to obtain the desired product 90 (82 mg, 34.5% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.80-0.94(m,9H),1.00-1.37(m,47H),1.39-1.56(m,8H),1.57-1.69(m,7H),1.71-1.87(m,4H),2.22-2.34(m,4H),2.36-2.71(m,10H),3.13-3.23(m,2H),3.24-3.38(m,2H),3.48-3.60(m,2H),4.00-4.11(m,2H)。LCMS：Rt：1.070min；MS m/z(ESI)：792.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.80-0.94(m,9H),1.00-1.37(m,47H),1.39-1.56(m,8H),1.57-1.69(m,7H),1.71-1.87(m,4H),2.22-2 .34(m,4H),2.36-2.71(m,10H),3.13-3.23(m,2H),3.24-3.38(m,2H),3.48-3.60(m,2H),4.00-4.11(m,2H). LCMS: Rt: 1.070min; MS m/z (ESI): 792.6[M+H]+ .
以下化合物係以與化合物90類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 90 using the corresponding starting materials.
6.41 實例41：製備化合物100.6.41 Example 41: Preparation of Compound 100.
步驟1：製備化合物100-1Step 1: Preparation of compound 100-1
在室溫下，向化合物SM8(2.8g，17.1mmol，1.0eq.)、DIEA(7.0g，54.2mmol，2.0eq.)在DCM(100mL)中之混合物中加入Boc2O(7.1g，32.6mmol，1.2eq.)。將混合物在室溫下攪拌1小時，TLC顯示反應完成。將混合物用水及鹽水洗滌，乾燥，濃縮。藉由管柱層析法純化殘餘物，得到呈無色油狀之產物100-1(4.9g，86%產率)。To a mixture of compound SM8 (2.8 g, 17.1 mmol, 1.0 eq.), DIEA (7.0 g, 54.2 mmol, 2.0 eq.) in DCM (100 mL) was added Boc2 O (7.1 g, 32.6 mmol, 1.2 eq.) at room temperature. The mixture was stirred at room temperature for 1 hour, and TLC showed that the reaction was complete. The mixture was washed with water and brine, dried, and concentrated. The residue was purified by column chromatography to give the product 100-1 (4.9 g, 86% yield) as a colorless oil.
步驟2：製備化合物100-2Step 2: Preparation of compound 100-2
在0℃下，向化合物100-1(4.8g，24.1mmol，1.0eq.)、DIEA(6.2g，48.2mmol，2.0eq.)、DCM(100mL)之混合物中加入MsCl(3.3g，28.9mmol，1.2eq.)。TLC顯示反應完成。將混合物用水及鹽水洗滌，乾燥，濃縮，且藉由管柱層析法純化殘餘物，得到呈黃色油狀之產物100-2(6.1g，90%產率)。MsCl (3.3 g, 28.9 mmol, 1.2 eq.) was added to a mixture of compound 100-1 (4.8 g, 24.1 mmol, 1.0 eq.), DIEA (6.2 g, 48.2 mmol, 2.0 eq.), and DCM (100 mL) at 0°C. TLC showed that the reaction was complete. The mixture was washed with water and brine, dried, concentrated, and the residue was purified by column chromatography to obtain product 100-2 (6.1 g, 90% yield) as a yellow oil.
步驟3：製備化合物100-3Step 3: Preparation of compound 100-3
將化合物100-2(3.0g，10.7mmol，1.0eq.)、化合物SM6(2.0g，32.0mmol，3.0eq.)、K2CO3(2.2g，16.0mmol，1.5eq.)在ACN(30mL)中之混合物回流攪拌隔夜。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化殘餘物，得到呈黃色油狀之產物100-3(2.6g，88%產率)。LCMS：Rt：0.836min；MS m/z(ESI)：247.1[M+H]+。A mixture of compound 100-2 (3.0 g, 10.7 mmol, 1.0 eq.), compound SM6 (2.0 g, 32.0 mmol, 3.0 eq.), K2 CO3 (2.2 g, 16.0 mmol, 1.5 eq.) in ACN (30 mL) was stirred under reflux overnight. LCMS showed that the reaction was complete. The mixture was concentrated and the residue was purified by preparative HPLC to give product 100-3 (2.6 g, 88% yield) as a yellow oil. LCMS: Rt: 0.836 min; MS m/z (ESI): 247.1 [M+H]+ .
步驟4：製備化合物100-4Step 4: Preparation of compound 100-4
將化合物100-3(500mg，2.0mmol，1.0eq.)、DIEA(790mg，6.1mmol，3.0eq.)、化合物43-3(1.0g，2.4mmol，1.2eq.)、NaI(90mg，0.6mmol，0.3eq.)在THF(20mL)中之混合物回流攪拌隔夜，LCMS顯示反應完成。將混合物濃縮並藉由管柱層析法純化殘餘物，得到呈黃色油狀之標題產物(920mg，70%產率)。LCMS：Rt：0.830min；MS m/z(ESI)：640.5[M+H]+。A mixture of compound 100-3 (500 mg, 2.0 mmol, 1.0 eq.), DIEA (790 mg, 6.1 mmol, 3.0 eq.), compound 43-3 (1.0 g, 2.4 mmol, 1.2 eq.), and NaI (90 mg, 0.6 mmol, 0.3 eq.) in THF (20 mL) was stirred under reflux overnight. LCMS showed that the reaction was complete. The mixture was concentrated and the residue was purified by column chromatography to give the title product (920 mg, 70% yield) as a yellow oil. LCMS: Rt: 0.830 min; MS m/z (ESI): 640.5 [M+H]+ .
步驟5：製備化合物100-5Step 5: Preparation of compound 100-5
將化合物100-4(920mg，1.4mmol，1.0eq.)、TFA(2.0mL)在DCM(5.0mL)中之混合物回流攪拌隔夜，LCMS顯示反應完成。將混合物濃縮，將殘餘物用乙酸乙酯稀釋，用飽和NaHCO3水溶液洗滌。將有機層濃縮，且未經進一步純化即用於下一步驟。A mixture of compound 100-4 (920 mg, 1.4 mmol, 1.0 eq.) and TFA (2.0 mL) in DCM (5.0 mL) was stirred under reflux overnight. LCMS showed that the reaction was complete. The mixture was concentrated, and the residue was diluted with ethyl acetate and washed with saturated aqueous NaHCO3 solution. The organic layer was concentrated and used in the next step without further purification.
步驟6：製備化合物100Step 6: Preparation of compound 100
向化合物SM21(粗品，0.05M，7.4mL，2.0eq.)在DCM中之溶液中加入化合物100-5(100mg，0.2mmol，1.0eq.)，在室溫下攪拌隔夜，LCMS顯示目標產物。將混合物濃縮，且藉由製備型HPLC純化殘餘物，得到呈黃色油狀之100(21mg，10.8%產率)。Compound 100-5 (100 mg, 0.2 mmol, 1.0 eq.) was added to a solution of compound SM21 (crude, 0.05 M, 7.4 mL, 2.0 eq.) in DCM, stirred at room temperature overnight, and LCMS showed the target product. The mixture was concentrated, and the residue was purified by preparative HPLC to obtain 100 (21 mg, 10.8% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.26-1.30(m,46H),1.41-1.52(m,4H),1.60-1.68(m,16H),1.74-1.81(m,4H),2.27-2.31(m,2H),2.39-2.50(m,9H),2.56-2.60(m,2H),2.86-2.93(m,2H),3.52-3.54(m,2H),3.93-4.00(m,4H),4.03-4.08(m,2H)。LCMS：Rt：1.020min；MS m/z(ESI)：872.7[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 9H), 1.26-1.30 (m, 46H), 1.41-1.52 (m, 4H), 1.60-1.68 (m, 16H),1.74-1.81(m,4H),2.27-2.31(m,2H),2.39-2.50(m,9H),2.56-2.60(m,2 H),2.86-2.93(m,2H),3.52-3.54(m,2H),3.93-4.00(m,4H),4.03-4.08(m,2H). LCMS: Rt: 1.020min; MS m/z (ESI): 872.7[M+H]+ .
6.42 實例42：製備化合物108.6.42 Example 42: Preparation of Compound 108.
步驟1：製備化合物108-2Step 1: Preparation of compound 108-2
向化合物108-1(758mg，4.54mmol，1.0eq.)及化合物SM7(1.4g，5.0mmol，1.1eq.)在甲苯(40mL)中之溶液中加入TsOH.H2O(20mg)。將混合物藉由Dean-Stark分水器回流攪拌2小時。將反應混合物濃縮並藉由矽膠管柱層析法(PE/EA=50/1)純化，得到呈無色油狀之標題化合物(910mg，47%產率)。LCMS：Rt：0.840min；MS m/z(ESI)：429.1/431.1[M+H]+。To a solution of compound 108-1 (758 mg, 4.54 mmol, 1.0 eq.) and compound SM7 (1.4 g, 5.0 mmol, 1.1 eq.) in toluene (40 mL) was added TsOH.H2 O (20 mg). The mixture was stirred at reflux for 2 hours via a Dean-Stark trap. The reaction mixture was concentrated and purified by silica gel column chromatography (PE/EA=50/1) to give the title compound (910 mg, 47% yield) as a colorless oil. LCMS: Rt: 0.840 min; MS m/z (ESI): 429.1/431.1 [M+H]+ .
步驟2：製備化合物108-3Step 2: Preparation of compound 108-3
向化合物108-2(910mg，2.12mmol，1.1eq.)及化合物D(276mg，1.93mmol，1.0eq.)在ACN(30mL)中之溶液中加入K2CO3(799mg，5.78mmol，3.0eq.)、Cs2CO3(188mg，0.58mmol，0.3eq.)及NaI(87mg，0.58mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=20/1)純化，得到呈黃色油狀之標題化合物(270mg，39%產率)。LCMS：Rt：0.880min；MS m/z(ESI)：492.4[M+H]+。To a solution of compound 108-2 (910 mg, 2.12 mmol, 1.1 eq.) and compound D (276 mg, 1.93 mmol, 1.0 eq.) in ACN (30 mL) were added K2 CO3 (799 mg, 5.78 mmol, 3.0 eq.), Cs2 CO3 (188 mg, 0.58 mmol, 0.3 eq.) and NaI (87 mg, 0.58 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=20/1) to give the title compound (270 mg, 39% yield) as a yellow oil. LCMS: Rt: 0.880min; MS m/z (ESI): 492.4[M+H]+ .
步驟3：製備化合物108-4Step 3: Preparation of compound 108-4
向化合物108-3(270mg，0.55mmol，1.0eq.)及DIPEA(142mg，1.10mmol，2.0eq.)在DCM(6mL)中之溶液中加入MsCl(94mg，0.52mmol，1.5eq.)。將混合物在室溫下攪拌2小時。LCMS顯示反應完成。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(313mg，100%產率)。其未經進一步純化即用於下一步驟。LCMS：Rt：0.927min；MS m/z(ESI)：474.2[M-OMs]+。To a solution of compound 108-3 (270 mg, 0.55 mmol, 1.0 eq.) and DIPEA (142 mg, 1.10 mmol, 2.0 eq.) in DCM (6 mL) was added MsCl (94 mg, 0.52 mmol, 1.5 eq.). The mixture was stirred at room temperature for 2 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give the title compound (313 mg, 100% yield) as a yellow oil. It was used in the next step without further purification. LCMS: Rt: 0.927 min; MS m/z (ESI): 474.2 [M-OMs]+ .
步驟4：製備化合物108Step 4: Preparation of compound 108
向化合物108-4(313mg，0.55mmol，1.0eq.)及化合物SM2(235mg，0.55mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(355mg，2.75mmol，5.0eq.)及NaI(25mg，0.16mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(115mg，23%產率)。LCMS：Rt：1.450min；MS m/z(ESI)：901.7[M+H]+。To a solution of compound 108-4 (313 mg, 0.55 mmol, 1.0 eq.) and compound SM2 (235 mg, 0.55 mmol, 1.0 eq.) in THF (10 mL) was added DIPEA (355 mg, 2.75 mmol, 5.0 eq.) and NaI (25 mg, 0.16 mmol, 0.3 eq.). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound (115 mg, 23% yield) as a yellow oil. LCMS: Rt: 1.450 min; MS m/z (ESI): 901.7 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.13-1.36(m,53H),1.43-1.50(m,4H),1.51-1.64(m,8H),1.68-1.79(m,4H),2.02-2.07(m,4H),2.27-2.32(m,4H),2.42-2.54(m,8H),2.62-2.70(m,2H),2.75-2.80(m,2H),3.50-3.56(m,2H),3.96-3.97(m,2H),4.04-4.07(m,2H),5.30-5.43(m,4H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,9H),1.13-1.36(m,53H),1.43-1.50(m,4H),1.51-1.64(m,8H),1.68-1.79(m,4H),2.02-2.07(m,4H),2.27-2.32(m,4H ),2.42-2.54(m,8H),2.62-2.70(m,2H),2.75-2.80(m,2H),3.50-3.56(m,2H),3.96-3.97(m,2H),4.04-4.07(m,2H),5.30-5.43(m,4H).
以下化合物係以與化合物108類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 108 using the corresponding starting materials.
6.43 實例43：製備化合物114.6.43 Example 43: Preparation of Compound 114.
步驟1：製備化合物114-2Step 1: Preparation of compound 114-2
向化合物114-1(1.3g，2.8mmol，1.0eq.)在ACN(30mL)中之溶液中加入化合物SM6(350mg，5.59mmol，2.0eq.)、K2CO3(1.16g，8.39mmol，3.0eq.)、Cs2CO3(280mg，0.84mmol，0.3eq.)及NaI(130mg，0.84mmol，0.3eq.)。將反應混合物在80℃下攪拌10小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到化合物114-2(600mg，50%)。LCMS：Rt：0.880min；MS m/z(ESI)：430.3[M+H]+。To a solution of compound 114-1 (1.3 g, 2.8 mmol, 1.0 eq.) in ACN (30 mL) were added compound SM6 (350 mg, 5.59 mmol, 2.0 eq.), K2 CO3 (1.16 g, 8.39 mmol, 3.0 eq.), Cs2 CO3 (280 mg, 0.84 mmol, 0.3 eq.) and NaI (130 mg, 0.84 mmol, 0.3 eq.). The reaction mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to give compound 114-2 (600 mg, 50%). LCMS: Rt: 0.880 min; MS m/z (ESI): 430.3 [M+H]+ .
步驟2：製備化合物114Step 2: Preparation of compound 114
向化合物114-3(180mg，0.42mmol，1.0eq.)、DIEA(150mg，1.05mmol，2.5eq.)在THF(10mL)中之混合物中加入化合物114-2(150mg，0.35mmol,0.8eq.)、NaI(50mg)。將反應混合物在70℃下攪拌10小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化殘餘物，得到呈黃色油狀之標題化合物(30mg，11%產率)。LCMS：Rt：0.600min；MS m/z(ESI)：795.5[M+H]+。To a mixture of compound 114-3 (180 mg, 0.42 mmol, 1.0 eq.), DIEA (150 mg, 1.05 mmol, 2.5 eq.) in THF (10 mL) were added compound 114-2 (150 mg, 0.35 mmol, 0.8 eq.), NaI (50 mg). The reaction mixture was stirred at 70 °C for 10 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to give the title compound (30 mg, 11% yield) as a yellow oil. LCMS: Rt: 0.600 min; MS m/z (ESI): 795.5 [M+H]+ .
1H NMR(400MHz,CDCl3)：0.87(t,J=8Hz,9H),1.26-1.99(m,60H),2.27-2.31(m,2H),2.39-2.43(m,4H),2.56-2.76(m,8H),3.05-3.09(m,1H),3.50-3.55(m,4H),3.68-3.71(m,2H),3.98-4.07(m,4H)。1 H NMR (400MHz, CDCl3 ): 0.87(t,J=8Hz,9H),1.26-1.99(m,60H),2.27-2.31(m,2H),2.39-2.43(m,4H),2.56-2 .76(m,8H),3.05-3.09(m,1H),3.50-3.55(m,4H),3.68-3.71(m,2H),3.98-4.07(m,4H).
以下化合物係以與化合物114類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 114 using the corresponding starting materials.
6.44 實例44：製備化合物118.6.44 Example 44: Preparation of Compound 118.
步驟1：製備化合物118-2Step 1: Preparation of compound 118-2
將化合物26-1(200mg，0.45mmol，1.0eq.)、化合物118-1(64mg，0.50mmol，1.1eq.)、K2CO3(186mg，1.35mmol，3.0eq.)、Cs2CO3(3mg，0.01mmol，0.03eq.)、NaI(34mg，0.23mmol，0.5eq.)在ACN(5mL)中之混合物在90℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至3%)純化，得到呈黃色油狀之期望產物118-2(168mg，粗品)。LCMS：Rt：0.890min；MS m/z(ESI)：496.4[M+H]+。A mixture of compound 26-1 (200 mg, 0.45 mmol, 1.0 eq.), compound 118-1 (64 mg, 0.50 mmol, 1.1 eq.), K2 CO3 (186 mg, 1.35 mmol, 3.0 eq.), Cs2 CO3 (3 mg, 0.01 mmol, 0.03 eq.), NaI (34 mg, 0.23 mmol, 0.5 eq.) in ACN (5 mL) was stirred at 90° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 3%) to give the desired product 118-2 (168 mg, crude) as a yellow oil. LCMS: Rt: 0.890 min; MS m/z (ESI): 496.4 [M+H]+ .
步驟2：製備化合物118-3Step 2: Preparation of compound 118-3
將化合物118-2(156mg，0.3mmol，1.0eq.)、Pd/C(15mg)在MeOH(6mL)中之混合物在40℃、氫氣氣氛下攪拌隔夜。將混合物過濾，將濾液濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至3%)純化，得到呈黃色油狀之期望產物118-3(145mg，92.5%產率)。LCMS：Rt：1.780min；MS m/z(ESI)：498.5[M+H]+。A mixture of compound 118-2 (156 mg, 0.3 mmol, 1.0 eq.) and Pd/C (15 mg) in MeOH (6 mL) was stirred at 40° C. under a hydrogen atmosphere overnight. The mixture was filtered, and the filtrate was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 3%) to give the desired product 118-3 (145 mg, 92.5% yield) as a yellow oil. LCMS: Rt: 1.780 min; MS m/z (ESI): 498.5 [M+H]+ .
步驟3：製備化合物118-4Step 3: Preparation of compound 118-4
將化合物118-3(148mg，0.3mmol，1.0eq.)及SOCl2(108mg，0.9mmol，3.0eq.)在DCM(5mL)中之混合物在35℃下攪拌隔夜。將混合物濃縮，得到呈黃色油狀之期望產物118-4(167mg，粗品)。LCMS：Rt：1.140min；MS m/z(ESI)：516.4[M+H]+。A mixture of compound 118-3 (148 mg, 0.3 mmol, 1.0 eq.) and SOCl2 (108 mg, 0.9 mmol, 3.0 eq.) in DCM (5 mL) was stirred at 35° C. overnight. The mixture was concentrated to give the desired product 118-4 (167 mg, crude) as a yellow oil. LCMS: Rt: 1.140 min; MS m/z (ESI): 516.4 [M+H]+ .
步驟4:製備化合物118Step 4: Preparation of compound 118
將化合物118-4(167mg，0.33mmol，1.0eq.)、化合物SM2(170mg，0.40mmol，1.2eq.)、DIEA(128mg，0.99mmol，3.0eq.)及NaI(24mg，0.16mmol，0.5eq.)在THF(5mL)中之混合物在70℃下攪拌隔夜。將混合物真空濃縮。將殘餘物藉由製備型HPLC純化兩次，得到呈棕色油狀之期望產物118(22mg，7.5%產率)。LCMS：Rt：2.440min；MS m/z(ESI)：907.8[M+H]+。A mixture of compound 118-4 (167 mg, 0.33 mmol, 1.0 eq.), compound SM2 (170 mg, 0.40 mmol, 1.2 eq.), DIEA (128 mg, 0.99 mmol, 3.0 eq.) and NaI (24 mg, 0.16 mmol, 0.5 eq.) in THF (5 mL) was stirred at 70 °C overnight. The mixture was concentrated in vacuo. The residue was purified twice by preparative HPLC to give the desired product 118 (22 mg, 7.5% yield) as a brown oil. LCMS: Rt: 2.440 min; MS m/z (ESI): 907.8 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.81-0.94(m,18H),1.18-1.38(m,62H),1.41-1.56(m,5H),1.58-1.70(m,7H),2.20-2.35(m,4H),2.37-2.64(m,12H),3.45-3.56(m,2H),3.91-4.01(m,4H)。1 H NMR (400MHz, CDCl3 ) δ: 0.81-0.94 (m, 18H),1.18-1.38(m,62H),1.41-1.56(m,5H),1.58-1.70(m,7H),2.20- 2.35(m,4H),2.37-2.64(m,12H),3.45-3.56(m,2H),3.91-4.01(m,4H).
6.45 實例45：製備化合物1206.45 Example 45: Preparation of Compound 120
步驟1：製備化合物120-1Step 1: Preparation of compound 120-1
將化合物114-1(300mg，0.65mmol，1.0eq.)、化合物118-1(125mg，0.97mmol，1.5eq.)及K2CO3(269mg，1.94mmol，3.0eq.)、Cs2CO3 65mg，0.20mmol，0.3eq.)、NaI(8mg，0.06mmol，0.1eq.)在ACN(10mL)中之混合物在85℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物120-1(150mg，46%產率)。LCMS：Rt：0.890min；MS m/z(ESI)：498.4[M+H]+。A mixture of compound 114-1 (300 mg, 0.65 mmol, 1.0 eq.), compound 118-1 (125 mg, 0.97 mmol, 1.5 eq.), K2 CO3 (269 mg, 1.94 mmol, 3.0 eq.), Cs2 CO3 65 mg, 0.20 mmol, 0.3 eq.), NaI (8 mg, 0.06 mmol, 0.1 eq.) in ACN (10 mL) was stirred at 85° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product 120-1 (150 mg, 46% yield) as a yellow oil. LCMS: Rt: 0.890min; MS m/z (ESI): 498.4[M+H]+ .
步驟2：製備化合物120-2Step 2: Preparation of compound 120-2
在室溫下，向化合物120-1(150mg，0.30mmol，1.0eq.)溶於DCM(10mL)中之溶液中加入DIEA(58mg，0.45mmol，1.5eq.)及MsCl(52mg，0.45mmol，1.5eq.)。將混合物在室溫下攪拌1小時。TLC顯示反應完成，將混合物減壓蒸發，得到呈黃色油狀之120-2(110mg，64%產率)。At room temperature, DIEA (58 mg, 0.45 mmol, 1.5 eq.) and MsCl (52 mg, 0.45 mmol, 1.5 eq.) were added to a solution of compound 120-1 (150 mg, 0.30 mmol, 1.0 eq.) in DCM (10 mL). The mixture was stirred at room temperature for 1 hour. TLC showed that the reaction was complete, and the mixture was evaporated under reduced pressure to obtain 120-2 (110 mg, 64% yield) as a yellow oil.
步驟2：製備化合物120Step 2: Preparation of compound 120
將化合物114-2(103mg，0.24mmol，1.2eq.)、化合物120-2(110mg，0.20mmol，1.0eq.)、DIEA(77mg，0.60mmol，3.0eq.)、NaI(8mg，0.06mmol，0.3eq.)在THF(10mL)中之混合物在75℃下攪拌隔夜。將混合物真空濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物120(15mg，8%產率)。LCMS：Rt：2.110min；MS m/z(ESI)：909.8[M+H]+。A mixture of compound 114-2 (103 mg, 0.24 mmol, 1.2 eq.), compound 120-2 (110 mg, 0.20 mmol, 1.0 eq.), DIEA (77 mg, 0.60 mmol, 3.0 eq.), NaI (8 mg, 0.06 mmol, 0.3 eq.) in THF (10 mL) was stirred at 75 °C overnight. The mixture was concentrated in vacuo. The residue was purified by preparative HPLC to give the desired product 120 (15 mg, 8% yield) as a yellow oil. LCMS: Rt: 2.110 min; MS m/z (ESI): 909.8 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.80-0.95(m,12H),1.16-1.36(m,52H),1.53-1.70(m,6H),1.71-1.85(m,5H),2.49-2.84(m,15H),3.03-3.19(m,1H),3.43-3.64(m,6H),3.77-3.90(m,5H),3.91-4.03(m,5H),5.19-5.33(m,1H)。1 H NMR (400MHz, CDCl3 )δ: 0.80-0.95(m,12H),1.16-1.36(m,52H),1.53-1.70(m,6H),1.71-1.85(m,5H),2.49-2.84(m,15 H),3.03-3.19(m,1H),3.43-3.64(m,6H),3.77-3.90(m,5H),3.91-4.03(m,5H),5.19-5.33(m,1H).
以下化合物係以與化合物120類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 120 using the corresponding starting materials.
6.46 實例46：製備化合物1276.46 Example 46: Preparation of Compound 127
步驟1：製備化合物127-2Step 1: Preparation of compound 127-2
在室溫下，向化合物127-1(300.0mg，0.7mmol，1.0eq.)及化合物D(97.0mg，0.7mmol，1.0eq.)在ACN(15.0mL)中之溶液中加入K2CO3(276.0mg，2.01mmol，3.0eq.)、Cs2CO3(65.0mg，0.2mmol，0.3eq.)及NaI(10.0mg，0.07mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用FCC(DCM/MeOH=1/0-10/1)純化，得到呈黃色油狀之127-2(0.24g，粗品)。LCMS：Rt：0.813min；MS m/z(ESI)：497.4[M+H]+。To a solution of compound 127-1 (300.0 mg, 0.7 mmol, 1.0 eq.) and compound D (97.0 mg, 0.7 mmol, 1.0 eq.) in ACN (15.0 mL) were added K2 CO3 (276.0 mg, 2.01 mmol, 3.0 eq.), Cs2 CO3 (65.0 mg, 0.2 mmol, 0.3 eq.) and NaI (10.0 mg, 0.07 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was completed, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=1/0-10/1) to give 127-2 (0.24 g, crude) as a yellow oil. LCMS: Rt: 0.813min; MS m/z (ESI): 497.4[M+H]+ .
步驟2：製備化合物127-3Step 2: Preparation of compound 127-3
在0℃下，向化合物127-2(0.24g，0.48mmol，1.0eq.)在DCM(10.0mL)中之溶液中加入DIEA(124.0mg，0.96mmol，2.0eq.)及MsCl(67.0mg，0.58mmol，1.2eq.)。將混合物攪拌1小時。TLC顯示反應完成，加入H2O並用DCM萃取，經Na2SO4乾燥。將混合物減壓蒸發，得到呈黃色油狀之127-3(0.26g，粗品)。To a solution of compound 127-2 (0.24 g, 0.48 mmol, 1.0 eq.) in DCM (10.0 mL) at 0°C, DIEA (124.0 mg, 0.96 mmol, 2.0 eq.) and MsCl (67.0 mg, 0.58 mmol, 1.2 eq.) were added. The mixture was stirred for 1 hour. TLC showed that the reaction was complete, H2 O was added and extracted with DCM, and dried over Na2 SO4. The mixture was evaporated under reduced pressure to give 127-3 (0.26 g, crude) as a yellow oil.
步驟3：製備化合物127Step 3: Preparation of compound 127
在0℃下，向化合物127-3(240.0mg，0.4mmol，1.0eq.)及化合物SM2(180.0mg，0.4mmol，1.0eq.)在THF(5.0mL)中之溶液中加入DIEA(162.0mg，1.2mmol，3.0eq.)及NaI(6.0mg，0.04mmol，0.1eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用製備型HPLC純化，得到呈黃色油狀之127(35.0mg，9%產率)。At 0°C, DIEA (162.0 mg, 1.2 mmol, 3.0 eq.) and NaI (6.0 mg, 0.04 mmol, 0.1 eq.) were added to a solution of compound 127-3 (240.0 mg, 0.4 mmol, 1.0 eq.) and compound SM2 (180.0 mg, 0.4 mmol, 1.0 eq.) in THF (5.0 mL). The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to obtain 127 (35.0 mg, 9% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.14-1.36(m,59H),1.42-1.78(m,18H),2.20-2.60(m,12H),3.17(s,4H),3.53(s,2H),3.96-4.06(m,4H)。LCMS：Rt：1.680min；MS m/z(ESI)：906.9[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 12H), 1.14-1.36 (m, 59H), 1.42-1.78 (m, 18H), 2.20-2.60 (m, 12H),3.17(s,4H),3.53(s,2H),3.96-4.06(m,4H). LCMS: Rt: 1.680min; MS m/z (ESI): 906.9[M+H]+ .
6.47 實例47：製備化合物1286.47 Example 47: Preparation of Compound 128
步驟1：製備化合物128-1Step 1: Preparation of compound 128-1
將化合物SM7(332.0mg，2.0mmol，1.0eq.)在0℃下溶於甲苯(10.0mL)中，接著加入吡啶(1.1g，16.0mmol，8.0eq.)及三光氣(1.1g，1.2mmol，0.6eq.)。將混合物在室溫下攪拌1小時，接著加入化合物SM(540.0mg，2.0mmol，1.0eq.)。將混合物在室溫下攪拌16小時。TLC顯示反應完成，將混合物倒入H2O中，用EA萃取。將混合物減壓蒸發並藉由FCC(PE/EA=100/1-10/1)純化，得到呈黃色油狀之128-1(350.0mg，粗品)。Compound SM7 (332.0 mg, 2.0 mmol, 1.0 eq.) was dissolved in toluene (10.0 mL) at 0°C, followed by the addition of pyridine (1.1 g, 16.0 mmol, 8.0 eq.) and triphosgene (1.1 g, 1.2 mmol, 0.6 eq.). The mixture was stirred at room temperature for 1 hour, followed by the addition of compound SM (540.0 mg, 2.0 mmol, 1.0 eq.). The mixture was stirred at room temperature for 16 hours. TLC showed that the reaction was complete, and the mixture was poured into H2 O and extracted with EA. The mixture was evaporated under reduced pressure and purified by FCC (PE/EA=100/1-10/1) to give 128-1 (350.0 mg, crude product) as a yellow oil.
步驟2：製備化合物128-2Step 2: Preparation of compound 128-2
在室溫下，向化合物128-1(300.0mg，0.65mmol，1.0eq.)及乙醇胺(120.0mg，2.0mmol，3.0eq.)在ACN(15.0mL)中之溶液中加入DIEA(419.0mg，3.25mmol，5eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用FCC(DCM/MeOH=1/0-10/1)純化，得到呈黃色油狀之128-2(0.2g，粗品)。LCMS：Rt：0.882min；MS m/z(ESI)：444.4[M+H]+。To a solution of compound 128-1 (300.0 mg, 0.65 mmol, 1.0 eq.) and ethanolamine (120.0 mg, 2.0 mmol, 3.0 eq.) in ACN (15.0 mL) was added DIEA (419.0 mg, 3.25 mmol, 5 eq.) at room temperature. The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=1/0-10/1) to give 128-2 (0.2 g, crude) as a yellow oil. LCMS: Rt: 0.882 min; MS m/z (ESI): 444.4 [M+H]+ .
步驟3：製備化合物128Step 3: Preparation of compound 128
在0℃下，向化合物128-2(155.0mg，0.35mmol，1.0eq.)及化合物43-3(150.0mg，0.35mmol，1.0eq.)在THF(5.0mL)中之溶液中加入DIEA(135mg，1.2mmol，5.0eq.)及NaI(5.0mg，0.035mmol，0.1eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用製備型HPLC純化，得到呈黃色油狀之128(20.0mg，7%產率)。LCMS：Rt：1.380min；MS m/z(ESI)：837.7[M+H]+。To a solution of compound 128-2 (155.0 mg, 0.35 mmol, 1.0 eq.) and compound 43-3 (150.0 mg, 0.35 mmol, 1.0 eq.) in THF (5.0 mL) at 0°C, DIEA (135 mg, 1.2 mmol, 5.0 eq.) and NaI (5.0 mg, 0.035 mmol, 0.1 eq.) were added. The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to give 128 (20.0 mg, 7% yield) as a yellow oil. LCMS: Rt: 1.380 min; MS m/z (ESI): 837.7 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.26-1.38(m,57H),1.59-1.71(m,16H),2.28-2.32(m,2H),2.52-2.70(m,8H),3.50-3.61(m,2H),4.03-4.14(m,6H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,9H),1.26-1.38(m,57H),1.59-1.71(m,16H),2.28-2.32(m,2H),2.52-2.70(m,8H),3.50-3.61(m,2H),4.03-4.14(m,6H).
6.48 實例48：製備化合物1336.48 Example 48: Preparation of Compound 133
步驟1：製備化合物133-1Step 1: Preparation of compound 133-1
在室溫下，向化合物128-1(500.0mg，1.0mmol，1.0eq.)及化合物D(158.0mg，1.0mmol，1.0eq.)在ACN(15.0mL)中之溶液中加入DIEA(418.0mg，3.0mmol，3.0eq.)及NaI(15.0mg，0.1mmol，0.1eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用FCC(DCM/MeOH=1/0-10/1)純化，得到呈黃色油狀之133-1(0.3g，粗品)。LCMS：Rt：0.945min；MS m/z(ESI)：526.5[M+H]+。To a solution of compound 128-1 (500.0 mg, 1.0 mmol, 1.0 eq.) and compound D (158.0 mg, 1.0 mmol, 1.0 eq.) in ACN (15.0 mL) was added DIEA (418.0 mg, 3.0 mmol, 3.0 eq.) and NaI (15.0 mg, 0.1 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by FCC (DCM/MeOH=1/0-10/1) to give 133-1 (0.3 g, crude) as a yellow oil. LCMS: Rt: 0.945 min; MS m/z (ESI): 526.5 [M+H]+ .
步驟2：製備化合物133-2Step 2: Preparation of compound 133-2
在0℃下，向化合物133-1(0.3g，0.57mmol，1.0eq.)在DCM(10.0mL)中之溶液中加入DIEA(147.0mg，1.14mmol，2.0eq.)及MsCl(79.0mg，0.68mmol，1.2eq.)。將混合物攪拌1小時。TLC顯示反應完成，加入H2O並用DCM萃取，經Na2SO4乾燥。將混合物減壓蒸發，得到呈黃色油狀之133-2(0.35g，粗品)。To a solution of compound 133-1 (0.3 g, 0.57 mmol, 1.0 eq.) in DCM (10.0 mL) at 0°C, DIEA (147.0 mg, 1.14 mmol, 2.0 eq.) and MsCl (79.0 mg, 0.68 mmol, 1.2 eq.) were added. The mixture was stirred for 1 hour. TLC showed that the reaction was complete, H2 O was added and extracted with DCM, and dried over Na2 SO4. The mixture was evaporated under reduced pressure to give 133-2 (0.35 g, crude) as a yellow oil.
步驟3：製備化合物133Step 3: Preparation of compound 133
在0℃下，向化合物133-2(240.0mg，0.4mmol，1.0eq.)及化合物SM2(180.0mg，0.4mmol，1.0eq.)在THF(5.0mL)中之溶液中加入DIEA(162.0mg，1.2mmol，3.0eq.)及NaI(6.0mg，0.04mmol，0.1eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用製備型HPLC純化，得到呈黃色油狀之133(25.0mg，7%產率)。LCMS：Rt：2.170min；MS m/z(ESI)：935.8[M+H]+。To a solution of compound 133-2 (240.0 mg, 0.4 mmol, 1.0 eq.) and compound SM2 (180.0 mg, 0.4 mmol, 1.0 eq.) in THF (5.0 mL) at 0°C were added DIEA (162.0 mg, 1.2 mmol, 3.0 eq.) and NaI (6.0 mg, 0.04 mmol, 0.1 eq.). The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to give 133 (25.0 mg, 7% yield) as a yellow oil. LCMS: Rt: 2.170 min; MS m/z (ESI): 935.8 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.14-1.36(m,68H),1.60-1.82(m,15H),2.28-2.32(m,2H),2.54-2.64(m,9H),3.58(s,2H),3.98-4.14(m,6H)。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.14-1.36(m,68H),1.60-1.82(m,15H),2.28-2.32(m,2H),2.54-2.64(m,9H),3.58(s,2H),3.98-4.14(m,6H).
6.49 實例49：製備化合物1346.49 Example 49: Preparation of Compound 134
步驟1：製備化合物134Step 1: Preparation of compound 134
在室溫下，向三光氣(300mg，1.0mmol，1.0eq.)在DCM(20mL)中之溶液中加入辛醇(395mg，3.0mmol，3.0eq.)及吡啶(640mg，8.0mmol，8.0eq.)。將混合物攪拌1小時。將化合物100-5(100mg，0.20mmol，0.2eq.)加入反應混合物(4mL)中。將混合物攪拌隔夜，LCMS顯示目標產物。將混合物濃縮並藉由製備型HPLC純化殘餘物，得到呈黃色油狀之產物134(16mg，4.3%產率)。Octanol (395 mg, 3.0 mmol, 3.0 eq.) and pyridine (640 mg, 8.0 mmol, 8.0 eq.) were added to a solution of triphosgene (300 mg, 1.0 mmol, 1.0 eq.) in DCM (20 mL) at room temperature. The mixture was stirred for 1 hour. Compound 100-5 (100 mg, 0.20 mmol, 0.2 eq.) was added to the reaction mixture (4 mL). The mixture was stirred overnight and LCMS showed the target product. The mixture was concentrated and the residue was purified by preparative HPLC to give product 134 (16 mg, 4.3% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.88(m,6H),1.07-1.30(m,32H),1.47-1.59(m,26H),1.78-1.79(m,4H),1.98-2.04(m,2H),2.26-2.58(m,6H),2.10-3.12(m,1H),3.49-3.53(m,1H),4.03-4.07(m,2H),4.66-5.38(m,1H)。LCMS：Rt：0.880min；MS m/z(ESI)：696.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.88(m,6H),1.07-1.30(m,32H),1.47-1.59(m,26H),1.78-1.79(m,4H),1.98-2.04(m,2 H),2.26-2.58(m,6H),2.10-3.12(m,1H),3.49-3.53(m,1H),4.03-4.07(m,2H),4.66-5.38(m,1H). LCMS: Rt: 0.880min; MS m/z (ESI): 696.6[M+H]+ .
6.50 實例50：製備化合物1476.50 Example 50: Preparation of Compound 147
步驟1：製備化合物100-1Step 1: Preparation of compound 100-1
向化合物SM8(1g，9.690mmol，1.0eq.)、DIEA(1.9g，14.54mmol，1.5eq.)在DCM(15mL)中之混合物中加入Boc2O(2.5g，11.63mmol，1.2eq.)。將反應混合物在室溫下攪拌16小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈無色油狀之化合物100-1(1.7g，86.30%)。To a mixture of compound SM8 (1 g, 9.690 mmol, 1.0 eq.), DIEA (1.9 g, 14.54 mmol, 1.5 eq.) in DCM (15 mL) was added Boc2 O (2.5 g, 11.63 mmol, 1.2 eq.). The reaction mixture was stirred at room temperature for 16 hours. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and dried over Na2 SO4. The solvent was removed and FCC was performed to give compound 100-1 (1.7 g, 86.30%) as a colorless oil.
步驟2：製備化合物147-1Step 2: Preparation of compound 147-1
在0℃下，向化合物100-1(1.7g，8.362mmol，1.0eq.)在THF(30mL)中之溶液中緩慢加入LiAlH4(0.64g，16.72mmol，2.0eq.)。將反應混合物回流攪拌2小時。TLC顯示反應完成。冷卻至0℃後，將混合物藉由連續加入水(1.3mL)、15% NaOH水溶液(1.3mL)及水(3.9mL)淬滅。將所得混合物用乙酸乙酯稀釋並藉由滴定除去沈澱物。將濾液減壓蒸發，得到呈黃色油狀之化合物147-1(0.8g，81.63%)。To a solution of compound 100-1 (1.7 g, 8.362 mmol, 1.0 eq.) in THF (30 mL) was slowly added LiAlH4 (0.64 g, 16.72 mmol, 2.0 eq.) at 0°C. The reaction mixture was stirred at reflux for 2 hours. TLC showed that the reaction was complete. After cooling to 0°C, the mixture was quenched by the continuous addition of water (1.3 mL), 15% NaOH aqueous solution (1.3 mL) and water (3.9 mL). The resulting mixture was diluted with ethyl acetate and the precipitate was removed by titration. The filtrate was evaporated under reduced pressure to give compound 147-1 (0.8 g, 81.63%) as a yellow oil.
步驟3：製備化合物147-2Step 3: Preparation of compound 147-2
向化合物147-1(300mg，2.559mmol，1.0eq.)、化合物SM22(692mg，2.559mmol，1.0eq.)、DIEA(662mg，5.122mmol，2.0eq.)在DCM(25mL)中之混合物中加入HATU(1.46g，3.839mmol，1.5eq.)。將反應混合物在室溫下攪拌2小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈無色油狀之化合物147-2(800mg，粗品)。To a mixture of compound 147-1 (300 mg, 2.559 mmol, 1.0 eq.), compound SM22 (692 mg, 2.559 mmol, 1.0 eq.), DIEA (662 mg, 5.122 mmol, 2.0 eq.) in DCM (25 mL) was added HATU (1.46 g, 3.839 mmol, 1.5 eq.). The reaction mixture was stirred at room temperature for 2 hours. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and dried over Na2 SO4. The solvent was removed and FCC was performed to give compound 147-2 (800 mg, crude) as a colorless oil.
步驟4：製備化合物147-3Step 4: Preparation of compound 147-3
在0℃、N2下，向化合物147-2(800mg，2.165mmol，1.0eq.)、DIEA(560mg，4.329mmol，2.0eq.)在DCM(15mL)中之混合物中加入MsCl(248mg，2.165mmol，1.0eq.)。將反應混合物在0℃下攪拌2小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈黃色油狀之化合物147-3(550mg，56.74%)。To a mixture of compound 147-2 (800 mg, 2.165 mmol, 1.0 eq.), DIEA (560 mg, 4.329 mmol, 2.0 eq.) in DCM (15 mL) at 0 °C underN2 was added MsCl (248 mg, 2.165 mmol, 1.0 eq.). The reaction mixture was stirred at 0 °C for 2 hours. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and driedoverNa2SO4 . The solvent was removed and FCC was performed to give compound 147-3 (550 mg, 56.74%) as a yellow oil.
步驟5：製備化合物147-4Step 5: Preparation of compound 147-4
向化合物147-3(550mg，1.229mmol，1.0eq.)在ACN(15mL)中之溶液中加入化合物E(232mg，1.474mmol，1.2eq.)、K2CO3(509mg，3.686mmol，3.0eq.)、Cs2CO3(120mg，0.3686mmol，0.3eq.)、NaI(55mg，0.3686mmol，0.3eq.)。將反應混合物在85℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物147-4(230mg，36.77%)。LCMS：Rt：0.850min；MS m/z(ESI)：509.5[M+H]+。To a solution of compound 147-3 (550 mg, 1.229 mmol, 1.0 eq.) in ACN (15 mL) were added compound E (232 mg, 1.474 mmol, 1.2 eq.), K2 CO3 (509 mg, 3.686 mmol, 3.0 eq.), Cs2 CO3 (120 mg, 0.3686 mmol, 0.3 eq.), NaI (55 mg, 0.3686 mmol, 0.3 eq.). The reaction mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound 147-4 (230 mg, 36.77%) as a yellow oil. LCMS: Rt: 0.850 min; MS m/z (ESI): 509.5 [M+H]+ .
步驟6：製備化合物147-5Step 6: Preparation of compound 147-5
向化合物147-4(230mg，0.4520mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(161mg，1.356mmol，3.0eq.)。將反應混合物在35℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，得到呈黃色油狀之化合物147-5(270mg，粗品)。LCMS：Rt：0.890min；MS m/z(ESI)：527.5[M+H]+。To a solution of compound 147-4 (230 mg, 0.4520 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (161 mg, 1.356 mmol, 3.0 eq.). The reaction mixture was stirred at 35° C. for 16 hours. LCMS showed the reaction was complete. The solvent was removed to give compound 147-5 (270 mg, crude) as a yellow oil. LCMS: Rt: 0.890 min; MS m/z (ESI): 527.5 [M+H]+ .
步驟7：製備化合物147Step 7: Preparation of compound 147
向化合物147-5(270mg，0.4520mmol，1.0eq.)、DIEA(292mg，2.260mmol，5.0eq.)在THF(15mL)中之混合物中加入化合物SM23(217mg，0.5424mmol，1.2eq.)、NaI(15mg)。將反應混合物在70℃下攪拌16小時。LCMS顯示反應完成。移除溶劑後，藉由製備型HPLC純化殘餘物，得到呈黃色油狀之標題化合物(50mg，12.42%產率)。LCMS：Rt：1.420min；MS m/z(ESI)：890.8[M+H]+。To a mixture of compound 147-5 (270 mg, 0.4520 mmol, 1.0 eq.), DIEA (292 mg, 2.260 mmol, 5.0 eq.) in THF (15 mL) were added compound SM23 (217 mg, 0.5424 mmol, 1.2 eq.), NaI (15 mg). The reaction mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to give the title compound (50 mg, 12.42% yield) as a yellow oil. LCMS: Rt: 1.420 min; MS m/z (ESI): 890.8 [M+H]+ .
1H NMR(400MHz,CDCl3)：0.86-0.89(m,12H),1.26(s,48H),1.34-1.41(m,6H),1.45-1.52(m,10H),1.62-1.91(m,11H),2.19-2.23(m,4H),2.38-2.67(m,11H),2.94(d,J=25.2Hz,3H),3.24-3.37(m,2H),3.52-3.54(m,2H),4.04-4.07(m,2H)。1 H NMR (400MHz, CDCl3 ): 0.86-0.89(m,12H),1.26(s,48H),1.34-1.41(m,6H),1.45-1.52(m,1 0H),1.62-1.91(m,11H),2.19-2.23(m,4H),2.38-2.67(m,11H),2.94(d,J =25.2Hz,3H),3.24-3.37(m,2H),3.52-3.54(m,2H),4.04-4.07(m,2H).
6.51 實例51：製備化合物1486.51 Example 51: Preparation of Compound 148
步驟1：製備化合物148-1Step 1: Preparation of compound 148-1
向化合物118-1(600mg，4.64mmol，2.0eq.)及化合物SM24(973mg，2.32mmol，1.0eq.)在ACN(40mL)中之溶液中加入K2CO3(962mg，6.96mmol，3.0eq.)、Cs2CO3(228mg，0.70mmol，0.3eq.)及NaI(105mg，0.70mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=25/1)純化，得到呈黃色油狀之標題化合物(525mg，49%產率)。LCMS：Rt：0.850min；MS m/z(ESI)：468.4[M+H]+。To a solution of compound 118-1 (600 mg, 4.64 mmol, 2.0 eq.) and compound SM24 (973 mg, 2.32 mmol, 1.0 eq.) in ACN (40 mL) were added K2 CO3 (962 mg, 6.96 mmol, 3.0 eq.), Cs2 CO3 (228 mg, 0.70 mmol, 0.3 eq.) and NaI (105 mg, 0.70 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=25/1) to give the title compound (525 mg, 49% yield) as a yellow oil. LCMS: Rt: 0.850min; MS m/z (ESI): 468.4[M+H]+ .
步驟2：製備化合物148-2Step 2: Preparation of compound 148-2
在0℃下，向化合物148-1(220mg，0.47mmol，1.0eq.)及DIPEA(121mg，0.94mmol，2.0eq.)在DCM(5mL)中之溶液中加入MsCl(65mg，0.56mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(238mg，93%產率)。其未經進一步純化即用於下一步驟。LCMS：Rt：0.940min；MS m/z(ESI)：486.4[M-OMs+Cl]+。To a solution of compound 148-1 (220 mg, 0.47 mmol, 1.0 eq.) and DIPEA (121 mg, 0.94 mmol, 2.0 eq.) in DCM (5 mL) was added MsCl (65 mg, 0.56 mmol, 1.2 eq.) at 0°C. The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated to give the title compound (238 mg, 93% yield) as a yellow oil. It was used in the next step without further purification. LCMS: Rt: 0.940 min; MS m/z (ESI): 486.4 [M-OMs+Cl]+ .
步驟3：製備化合物148Step 3: Preparation of compound 148
向化合物148-2(200mg，0.37mmol，1.0eq.)及化合物SM16(163mg，0.37mmol，1.0eq.)在ACN(10mL)中之溶液中加入K2CO3(153mg，1.11mmol，3.0eq.)、Cs2CO3(36mg，0.11mmol，0.3eq.)及NaI(16mg，0.11mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(50mg，15%產率)。To a solution of compound 148-2 (200 mg, 0.37 mmol, 1.0 eq.) and compound SM16 (163 mg, 0.37 mmol, 1.0 eq.) in ACN (10 mL) were added K2 CO3 (153 mg, 1.11 mmol, 3.0 eq.), Cs2 CO3 (36 mg, 0.11 mmol, 0.3 eq.) and NaI (16 mg, 0.11 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound (50 mg, 15% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.35(m,48H),1.41-1.52(m,4H),1.59-1.64(m,10H),1.73-1.76(m,3H),1.95-2.01(m,1H),2.28-2.32(m,6H),2.37-2.62(m,9H),3.03-3.11(m,2H),3.50-3.56(m,2H),3.95-3.97(m,2H),4.00-4.10(m,4H),5.23-5.28(m,1H)。LCMS：Rt：1.470min；MS m/z(ESI)：893.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.35(m,48H),1.41-1.52(m,4H),1.59-1.64(m,10H),1.73-1.76(m,3H),1.95-2.01(m,1H),2.28- 2.32(m,6H),2.37-2.62(m,9H),3.03-3.11(m,2H),3.50-3.56(m,2H),3.95-3.97(m,2H),4.00-4.10(m,4H),5.23-5.28(m,1H). LCMS: Rt: 1.470min; MS m/z (ESI): 893.7[M+H]+ .
以下化合物係以與化合物148類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 148 using the corresponding starting materials.
6.52 實例52：製備化合物1496.52 Example 52: Preparation of Compound 149
步驟1：製備化合物149-2Step 1: Preparation of compound 149-2
將化合物149-1(885mg，4.56mmol，1.1eq.)、化合物W(1.0g，4.15mmol，1.0eq.)、HATU(1.9g，4.98mmol，1.2eq.)及DIEA(1.6g，4.98mmol，1.2eq.)在DCM(20mL)中之混合物在室溫下攪拌16小時。TLC顯示反應完成。將混合物濃縮並藉由矽膠管柱層析法(EA：PE=0%至5%)純化，得到呈無色油狀之化合物149-2(1.2g，63%產率)。A mixture of compound 149-1 (885 mg, 4.56 mmol, 1.1 eq.), compound W (1.0 g, 4.15 mmol, 1.0 eq.), HATU (1.9 g, 4.98 mmol, 1.2 eq.) and DIEA (1.6 g, 4.98 mmol, 1.2 eq.) in DCM (20 mL) was stirred at room temperature for 16 hours. TLC showed that the reaction was complete. The mixture was concentrated and purified by silica gel column chromatography (EA:PE = 0% to 5%) to obtain compound 149-2 (1.2 g, 63% yield) as a colorless oil.
步驟2：製備化合物149-3Step 2: Preparation of compound 149-3
將化合物149-2(500mg，1.20mmol，1.0eq.)、化合物118-1(170mg，1.31mmol，1.1eq.)、K2CO3(497mg，3.60mmol，3.0eq.)、Cs2CO3(117mg，0.36mmol，0.3eq.)及NaI(17mg，0.12mmol，0.1eq.)在ACN(10mL)中之混合物在85℃下攪拌隔夜。將混合物濃縮並藉由矽膠管柱層析法(MeOH：DCM=0%至10%)純化，得到呈黃色油狀之期望產物149-3(300mg，54%產率)。LCMS：Rt：0.820min；MS m/z(ESI)：467.4[M+H]+。A mixture of compound 149-2 (500 mg, 1.20 mmol, 1.0 eq.), compound 118-1 (170 mg, 1.31 mmol, 1.1 eq.), K2 CO3 (497 mg, 3.60 mmol, 3.0 eq.), Cs2 CO3 (117 mg, 0.36 mmol, 0.3 eq.) and NaI (17 mg, 0.12 mmol, 0.1 eq.) in ACN (10 mL) was stirred at 85° C. overnight. The mixture was concentrated and purified by silica gel column chromatography (MeOH:DCM=0% to 10%) to give the desired product 149-3 (300 mg, 54% yield) as a yellow oil. LCMS: Rt: 0.820min; MS m/z (ESI): 467.4[M+H]+ .
步驟3：製備化合物149-4Step 3: Preparation of compound 149-4
在室溫下，向化合物149-3(280mg，0.60mmol，1.0eq.)溶於DCM(10mL)中之溶液中加入MsCl(82mg，0.72mmol，1.2eq.)。將混合物在室溫下攪拌1小時。LCMS顯示反應完成，將混合物減壓蒸發，得到呈黃色油狀之化合物149-4(200mg，粗品)。LCMS：Rt：0.830min；MS m/z(ESI)：449.4[M-OMs]+。To a solution of compound 149-3 (280 mg, 0.60 mmol, 1.0 eq.) in DCM (10 mL) was added MsCl (82 mg, 0.72 mmol, 1.2 eq.) at room temperature. The mixture was stirred at room temperature for 1 hour. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure to give compound 149-4 (200 mg, crude) as a yellow oil. LCMS: Rt: 0.830 min; MS m/z (ESI): 449.4 [M-OMs]+ .
步驟4：製備化合物149Step 4: Preparation of compound 149
將化合物SM25(200mg，0.41mmol，1.2eq.)、化合物149-4(182mg，0.41mmol，1.0eq.)、K2CO3(170mg，1.23mmol，3.0eq.)、Cs2CO3(40mg，0.12mmol，0.3eq.)及NaI(5.6mg，0.04mmol，0.1eq.)在ACN(10mL)中之混合物在85℃下攪拌隔夜。LCMS顯示反應完成。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之期望產物149(62mg，17%產率)。LCMS：Rt：0.980min；MS m/z(ESI)：892.8[M+H]+。A mixture of compound SM25 (200 mg, 0.41 mmol, 1.2 eq.), compound 149-4 (182 mg, 0.41 mmol, 1.0 eq.), K2 CO3 (170 mg, 1.23 mmol, 3.0 eq.), Cs2 CO3 (40 mg, 0.12 mmol, 0.3 eq.) and NaI (5.6 mg, 0.04 mmol, 0.1 eq.) in ACN (10 mL) was stirred at 85° C. overnight. LCMS showed the reaction was complete. The residue was purified by preparative HPLC to give the desired product 149 (62 mg, 17% yield) as a yellow oil. LCMS: Rt: 0.980 min; MS m/z (ESI): 892.8 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.80-0.94(m,12H),1.19-1.42(m,42H),1.43-1.55(m,4H),1.56-1.71(m,12H),1.72-1.83(m,4H),1.83-2.03(m,2H),2.04-2.24(m,4H),2.25-2.40(m,6H),2.41-2.76(m,9H),3.06-3.24(m,3H),3.47-3.65(m,2H),4.00-4.12(m,4H),5.16-5.31(m,1H)。1 H NMR (400MHz, CDCl3 )δ: 0.80-0.94(m,12H),1.19-1.42(m,42H),1.43-1.55(m,4H),1.56-1.71(m,12H),1.72-1.83(m,4H),1.83-2.03(m,2H),2.04- 2.24(m,4H),2.25-2.40(m,6H),2.41-2.76(m,9H),3.06-3.24(m,3H),3.47-3.65(m,2H),4.00-4.12(m,4H),5.16-5.31(m,1H).
以下化合物係以與化合物149類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 149 using the corresponding starting materials.
6.53 實例53：製備化合物1516.53 Example 53: Preparation of Compound 151
步驟1：製備化合物151-2Step 1: Preparation of compound 151-2
在-78℃、N2下，向DMSO(3.2g，41.2mmol，2.0eq.)在DCM(120mL)中之溶液中滴加草醯氯(2.9g，22.7mmol，1.1eq.)在DCM(20mL)中之溶液。將混合物攪拌30分鐘，接著在-78℃下滴加化合物151-1(5.0g，20.6mmol，1.0eq.)。將混合物在-78℃下攪拌60分鐘。加入TEA(6.3g，61.8mmol，3.0eq.)，且使混合物升溫至室溫。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。將殘餘物藉由矽膠管柱層析法(PE/EA=10/1)純化，得到呈無色油狀之標題化合物(3.2g，65%產率)。To a solution of DMSO (3.2 g, 41.2 mmol, 2.0 eq.) in DCM (120 mL) was added dropwise a solution of oxalyl chloride (2.9 g, 22.7 mmol, 1.1 eq.) in DCM (20 mL) at-78 °C under N2. The mixture was stirred for 30 min, and then compound 151-1 (5.0 g, 20.6 mmol, 1.0 eq.) was added dropwise at -78 °C. The mixture was stirred at -78 °C for 60 min. TEA (6.3 g, 61.8 mmol, 3.0 eq.) was added, and the mixture was allowed to warm to room temperature. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=10/1) to give the title compound (3.2 g, 65% yield) as a colorless oil.
步驟2：製備化合物151-3Step 2: Preparation of compound 151-3
向化合物151-2(878mg，3.65mmol，1.0eq.)及化合物71-4(870mg，3.65mmol，1.0eq.)在甲苯(30mL)中之溶液中加入p-TsOH(70mg，0.37mmol，0.1eq.)。將混合物在40℃下攪拌16小時。將混合物用乙酸乙酯稀釋並用飽和NaHCO3水溶液洗滌。將有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。將殘餘物藉由矽膠管柱層析法(PE/EA=100/1)純化，得到呈無色油狀之標題化合物(1.2g，71%產率)。To a solution of compound 151-2 (878 mg, 3.65 mmol, 1.0 eq.) and compound 71-4 (870 mg, 3.65 mmol, 1.0 eq.) in toluene (30 mL) was added p-TsOH (70 mg, 0.37 mmol, 0.1 eq.). The mixture was stirred at 40 °C for 16 hours. The mixture was diluted with ethyl acetate and washed with a saturated aqueous NaHCO3 solution. The organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=100/1) to give the title compound (1.2 g, 71% yield) as a colorless oil.
步驟3：製備化合物151-4Step 3: Preparation of compound 151-4
向化合物151-3(1.2g，2.60mmol，1.0eq.)在乙酸乙酯(25mL)中之溶液中加入Pd/C(120mg)。將混合物在H2及35℃下攪拌16小時。將混合物藉由矽藻土墊過濾並用EA洗滌。將濾液濃縮並藉由矽膠管柱層析法(PE/EA=5/1)純化，得到呈無色油狀之標題化合物(610mg，63%產率)。To a solution of compound 151-3 (1.2 g, 2.60 mmol, 1.0 eq.) in ethyl acetate (25 mL) was added Pd/C (120 mg). The mixture was stirred underH2 and 35°C for 16 h. The mixture was filtered through a celite pad and washed with EA. The filtrate was concentrated and purified by silica gel column chromatography (PE/EA=5/1) to give the title compound (610 mg, 63% yield) as a colorless oil.
步驟4：製備化合物151-5Step 4: Preparation of compound 151-5
在0℃下，向化合物151-4(610mg，1.65mmol，1.0eq.)及DIPEA(426mg，3.30mmol，2.0eq.)在DCM(20mL)中之溶液中加入MsCl(227mg，1.98mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(640mg，87%產率)。其未經進一步純化即用於下一步驟。To a solution of compound 151-4 (610 mg, 1.65 mmol, 1.0 eq.) and DIPEA (426 mg, 3.30 mmol, 2.0 eq.) in DCM (20 mL) was added MsCl (227 mg, 1.98 mmol, 1.2 eq.) at 0°C. The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted withDCM . The combined organic layers were washed with brine, dried overNa2SO4 and concentrated to give the title compound (640 mg, 87% yield) as a yellow oil. It was used in the next step without further purification.
步驟5：製備化合物151-6Step 5: Preparation of compound 151-6
向化合物151-5(640mg，1.43mmol，1.0eq.)及化合物SM6(178mg，2.86mmol，2.0eq.)在ACN(28mL)中之溶液中加入K2CO3(593mg，4.29mmol，3.0eq.)、Cs2CO3(140mg，0.43mmol，0.3eq.)及NaI(64mg，0.43mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化殘餘物，得到呈黃色油狀之標題化合物(362mg，61%產率)。LCMS：Rt：0.870min；MS m/z(ESI)：414.4[M+H]+。To a solution of compound 151-5 (640 mg, 1.43 mmol, 1.0 eq.) and compound SM6 (178 mg, 2.86 mmol, 2.0 eq.) in ACN (28 mL) were added K2 CO3 (593 mg, 4.29 mmol, 3.0 eq.), Cs2 CO3 (140 mg, 0.43 mmol, 0.3 eq.) and NaI (64 mg, 0.43 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=10/1) to give the title compound (362 mg, 61% yield) as a yellow oil. LCMS: Rt: 0.870 min; MS m/z (ESI): 414.4 [M+H]+ .
步驟6：製備化合物151Step 6: Preparation of compound 151
向化合物151-6(180mg，0.44mmol，1.0eq.)及化合物148-2(240mg，0.44mmol，1.0eq.)在ACN(15mL)中之溶液中加入K2CO3(182mg，1.32mmol，3.0eq.)、Cs2CO3(42mg，0.13mmol，0.3eq.)及NaI(19mg，0.13mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(36mg，10%產率)。LCMS：Rt：1.710min；MS m/z(ESI)：863.8[M+H]+。To a solution of compound 151-6 (180 mg, 0.44 mmol, 1.0 eq.) and compound 148-2 (240 mg, 0.44 mmol, 1.0 eq.) in ACN (15 mL) were added K2 CO3 (182 mg, 1.32 mmol, 3.0 eq.), Cs2 CO3 (42 mg, 0.13 mmol, 0.3 eq.) and NaI (19 mg, 0.13 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound (36 mg, 10% yield) as a yellow oil. LCMS: Rt: 1.710 min; MS m/z (ESI): 863.8 [M+H]+ .
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,12H),0.96-0.99(m,2H),1.26-1.38(m,51H),1.46-1.59(m,6H),1.62-1.86(m,10H),1.95-1.99(m,1H),2.28-2.30(m,2H),2.41-2.71(m,9H),2.96-3.15(m,2H),3.24-3.30(m,2H),3.48-3.59(m,2H),3.80-3.85(m,1H),3.91-3.97(m,2H),4.05-4.09(m,2H),4.30-4.45(m,1H),5.22-5.28(m,1H)。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.89 (m, 12H),0.96-0.99(m,2H),1.26-1.38(m,51H),1.46-1.59(m,6H),1.62-1. 86(m,10H),1.95-1.99(m,1H),2.28-2.30(m,2H),2.41-2.71(m,9H),2.96 -3.15(m,2H),3.24-3.30(m,2H),3.48-3.59(m,2H),3.80-3.85(m,1H),3. 91-3.97(m,2H),4.05-4.09(m,2H),4.30-4.45(m,1H),5.22-5.28(m,1H).
6.54 實例54：製備化合物1526.54 Example 54: Preparation of Compound 152
步驟1：製備化合物152-1Step 1: Preparation of compound 152-1
向化合物SM22(1g，3.697mmol，1.0eq.)、化合物SM8(1.2g，4.436mmol，1.2eq.)、DIEA(0.96g，7.394mmol，2.0eq.)在DCM(15mL)中之混合物中加入HATU(2.1g，5.546mmol，1.5eq.)。將反應混合物在室溫下攪拌2小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈黃色油狀之化合物152-1(1.2g，91.28%)。To a mixture of compound SM22 (1 g, 3.697 mmol, 1.0 eq.), compound SM8 (1.2 g, 4.436 mmol, 1.2 eq.), DIEA (0.96 g, 7.394 mmol, 2.0 eq.) in DCM (15 mL) was added HATU (2.1 g, 5.546 mmol, 1.5 eq.). The reaction mixture was stirred at room temperature for 2 hours. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and dried over Na2 SO4. The solvent was removed and FCC was performed to obtain compound 152-1 (1.2 g, 91.28%) as a yellow oil.
步驟2：製備化合物152-2Step 2: Preparation of compound 152-2
在0℃、N2下，向化合物152-1(1.2g，3.375mmol，1.0eq.)、DIEA(0.87g，6.750mmol，2.0eq.)在DCM(20mL)中之混合物中加入MsCl(0.46g，4.049mmol，1.2eq.)。將反應混合物在0℃下攪拌2小時。TLC顯示反應完成。將混合物倒入水中並用DCM洗滌。將有機物分離並經Na2SO4乾燥。移除溶劑，進行FCC，得到呈黃色油狀之化合物152-2(1g，64.18%)。To a mixture of compound 152-1 (1.2 g, 3.375 mmol, 1.0 eq.), DIEA (0.87 g, 6.750 mmol, 2.0 eq.) in DCM (20 mL) at 0 °C underN2 was added MsCl (0.46 g, 4.049 mmol, 1.2 eq.). The reaction mixture was stirred at 0 °C for 2 hours. TLC showed that the reaction was complete. The mixture was poured into water and washed with DCM. The organics were separated and driedoverNa2SO4 . The solvent was removed and FCC was performed to give compound 152-2 (1 g, 64.18%) as a yellow oil.
步驟3：製備化合物152-3Step 3: Preparation of compound 152-3
向化合物152-2(1g，2.166mmol，1.0eq.)在ACN(20mL)中之溶液中加入化合物E(0.4g，2.599mmol，1.2eq.)、K2CO3(0.9g，6.498mmol，3.0eq.)、Cs2CO3(0.21g，0.6498mmol，0.3eq.)、NaI(0.1g，0.6498mmol，0.3eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，進行FCC，得到呈黃色油狀之化合物152-3(600mg，55.98%)。LCMS：Rt：0.870min；MS m/z(ESI)：495.4[M+H]+。To a solution of compound 152-2 (1 g, 2.166 mmol, 1.0 eq.) in ACN (20 mL) were added compound E (0.4 g, 2.599 mmol, 1.2 eq.), K2 CO3 (0.9 g, 6.498 mmol, 3.0 eq.), Cs2 CO3 (0.21 g, 0.6498 mmol, 0.3 eq.), NaI (0.1 g, 0.6498 mmol, 0.3 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The solvent was removed and FCC was performed to obtain compound 152-3 (600 mg, 55.98%) as a yellow oil. LCMS: Rt: 0.870 min; MS m/z (ESI): 495.4 [M+H]+ .
步驟4：製備化合物152-4Step 4: Preparation of compound 152-4
向化合物152-3(600mg，1.213mmol，1.0eq.)在DCM(15mL)中之溶液中加入SOCl2(433mg，3.638mmol，3.0eq.)。將反應混合物在35℃下攪拌16小時。LCMS顯示反應完成。移除溶劑，得到呈黃色油狀之化合物152-4(650mg，粗品)。LCMS：Rt：0.910min；MS m/z(ESI)：513.4[M+H]+。To a solution of compound 152-3 (600 mg, 1.213 mmol, 1.0 eq.) in DCM (15 mL) was added SOCl2 (433 mg, 3.638 mmol, 3.0 eq.). The reaction mixture was stirred at 35° C. for 16 hours. LCMS showed the reaction was complete. The solvent was removed to give compound 152-4 (650 mg, crude) as a yellow oil. LCMS: Rt: 0.910 min; MS m/z (ESI): 513.4 [M+H]+ .
步驟5：製備化合物152Step 5: Preparation of compound 152
向化合物152-4(200mg，0.3896mmol，1.0eq.)在ACN(10mL)中之混合物中加入化合物SM26(192mg，0.4676mmol，1.2eq.)、K2CO3(162mg，1.169mmol，330eq.)、Cs2CO3(38mg，0.1169mmol，0.3eq.)、NaI(18mg，0.1169mmol，0.3eq.)。將反應混合物在80℃下攪拌16小時。LCMS顯示反應完成。移除溶劑後，將殘餘物藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(52mg，15.02%產率)。LCMS：Rt：1.440min；MS m/z(ESI)：888.8[M+H]+。To a mixture of compound 152-4 (200 mg, 0.3896 mmol, 1.0 eq.) in ACN (10 mL) were added compound SM26 (192 mg, 0.4676 mmol, 1.2 eq.), K2 CO3 (162 mg, 1.169 mmol, 330 eq.), Cs2 CO3 (38 mg, 0.1169 mmol, 0.3 eq.), NaI (18 mg, 0.1169 mmol, 0.3 eq.). The reaction mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. After removing the solvent, the residue was purified by preparative HPLC to give the title compound (52 mg, 15.02% yield) as a yellow oil. LCMS: Rt: 1.440min; MS m/z (ESI): 888.8[M+H]+ .
1H NMR(400MHz,CDCl3)：0.86-0.89(m,9H),1.28(d,J=19.6Hz,49H),1.45-1.52(m,9H),1.62-1.68(m,10H),1.78-1.86(m,4H),1.99-2.07(m,6H),2.27-2.31(m,2H),2.50-2.68(m,11H),3.22-3.26(m,2H),3.52-3.55(m,2H),4.04-4.07(m,2H),5.33-5.36(m,2H),5.85(s,1H)。1 H NMR (400MHz, CDCl3 ): 0.86-0.89 (m, 9H), 1.28 (d,J =19.6Hz,49H),1.45-1.52(m,9H),1.62-1.68(m,10H),1.78-1.86(m,4H),1.99-2. 07(m,6H),2.27-2.31(m,2H),2.50-2.68(m,11H),3.22-3.26(m,2H),3.52-3.55(m, 2H),4.04-4.07(m,2H),5.33-5.36(m,2H),5.85(s,1H).
以下化合物係以與化合物152類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 152 using the corresponding starting materials.
6.55 實例55：製備化合物1616.55 Example 55: Preparation of Compound 161
步驟1：製備化合物161-1Step 1: Preparation of compound 161-1
向化合物71-7(500mg，1.044mmol，1.0eq.)及化合物F(215mg，1.253mmol，1.2eq.)在ACN(10mL)中之溶液中加入K2CO3(433mg，3.132mmol，3.0eq.)、Cs2CO3(102mg，0.3132mmol，0.3eq.)及NaI(51mg，0.3132mmol，0.3eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=10/1)純化，得到呈黃色油狀之標題化合物(300mg，51.88%)。LCMS：Rt：0.840min；MS m/z(ESI)：554.4[M+H]+。To a solution of compound 71-7 (500 mg, 1.044 mmol, 1.0 eq.) and compound F (215 mg, 1.253 mmol, 1.2 eq.) in ACN (10 mL) were added K2 CO3 (433 mg, 3.132 mmol, 3.0 eq.), Cs2 CO3 (102 mg, 0.3132 mmol, 0.3 eq.) and NaI (51 mg, 0.3132 mmol, 0.3 eq.). The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=10/1) to give the title compound (300 mg, 51.88%) as a yellow oil. LCMS: Rt: 0.840min; MS m/z (ESI): 554.4[M+H]+ .
步驟2：製備化合物161-2Step 2: Preparation of compound 161-2
在0℃下，向化合物161-1(300mg，0.5416mmol，1.0eq.)及DIPEA(105mg，0.8124mmol，1.5eq.)在DCM(10mL)中之溶液中加入MsCl(74mg，0.6499mmol，1.2eq.)。將混合物再攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(340mg，粗品)。To a solution of compound 161-1 (300 mg, 0.5416 mmol, 1.0 eq.) and DIPEA (105 mg, 0.8124 mmol, 1.5 eq.) in DCM (10 mL) was added MsCl (74 mg, 0.6499 mmol, 1.2 eq.) at 0°C. The mixture was stirred for another 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated to give the title compound (340 mg, crude) as a yellow oil.
步驟3：製備化合物161Step 3: Preparation of compound 161
向化合物161-2(340mg，0.5380mmol，1.04eq.)及化合物SM16(230mg，0.5184mmol，1.0eq.)在THF(15mL)中之溶液中加入DIEA(335mg，2.592mmol，5.0eq.)及NaI(15mg)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(60mg，11.82%產率)。1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.29(s,31H),1.32-1.35(m,7H),1.42-1.44(m,11H),1.59-1.72(m,19H),1.95-2.00(m,2H),2.28-2.32(m,8H),2.35-2.42(m,3H),2.47-2.60(m,6H),2.80(s,1H),3.52-3.54(m,2H),4.00-4.10(m,8H)。LCMS：Rt：1.145min；MS m/z(ESI)：979.7[M+H]+。To a solution of compound 161-2 (340 mg, 0.5380 mmol, 1.04 eq.) and compound SM16 (230 mg, 0.5184 mmol, 1.0 eq.) in THF (15 mL) was added DIEA (335 mg, 2.592 mmol, 5.0 eq.) and NaI (15 mg). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound (60 mg, 11.82% yield) as a yellow oil.1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.29(s,31H),1.32-1.35(m,7H),1.42-1.44(m,11H),1.59-1.72(m,19H),1.95-2.00(m, 2H),2.28-2.32(m,8H),2.35-2.42(m,3H),2.47-2.60(m,6H),2.80(s,1H),3.52-3.54(m,2H),4.00-4.10(m,8H). LCMS: Rt: 1.145min; MS m/z (ESI): 979.7[M+H]+ .
以下化合物係以與化合物161類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 161 using the corresponding starting materials.
6.56 實例56：製備化合物1706.56 Example 56: Preparation of Compound 170
步驟1：製備化合物170-2Step 1: Preparation of compound 170-2
向化合物170-1(500mg，3.69mmol，1.0eq.)及化合物26-1(1.3g，2.95mmol，1.0eq.)在ACN(60mL)中之溶液中加入K2CO3(1.5g，11.07mmol，3.0eq.)、Cs2CO3(361mg，1.11mmol，0.3eq.)及NaI(166mg，1.11mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=50/1)純化，得到呈黃色油狀之標題化合物(630mg，37%產率)。LCMS：Rt：1.007min；MS m/z(ESI)：466.3[M+H]+。To a solution of compound 170-1 (500 mg, 3.69 mmol, 1.0 eq.) and compound 26-1 (1.3 g, 2.95 mmol, 1.0 eq.) in ACN (60 mL) were added K2 CO3 (1.5 g, 11.07 mmol, 3.0 eq.), Cs2 CO3 (361 mg, 1.11 mmol, 0.3 eq.) and NaI (166 mg, 1.11 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=50/1) to give the title compound (630 mg, 37% yield) as a yellow oil. LCMS: Rt: 1.007min; MS m/z (ESI): 466.3[M+H]+ .
步驟2：製備化合物170Step 2: Preparation of compound 170
向化合物170-2(300mg，0.64mmol，1.0eq.)在MeOH(10mL)中之溶液中加入化合物SM2(219mg，0.51mmol，0.8eq.)及AcOH(1滴)。將混合物在室溫下攪拌2小時。接著加入NaCNBH3(40mg，0.64mmol，1.0eq.)，且將所得混合物在室溫下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈無色油狀之標題化合物(54mg，12%產率)。To a solution of compound 170-2 (300 mg, 0.64 mmol, 1.0 eq.) in MeOH (10 mL) was added compound SM2 (219 mg, 0.51 mmol, 0.8 eq.) and AcOH (1 drop). The mixture was stirred at room temperature for 2 hours. NaCNBH3 (40 mg, 0.64 mmol, 1.0 eq.) was then added, and the resulting mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound (54 mg, 12% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.36(m,60H),1.42-1.54(m,5H),1.59-1.74(m,10H),1.89-1.96(m,2H),2.30-2.35(m,6H),2.48-2.57(m,3H),2.64-2.66(m,2H),3.00-3.03(m,2H),3.49-3.52(m,2H),3.96-3.97(m,4H)。LCMS：Rt：2.360min；MS m/z(ESI)：877.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.36(m,60H),1.42-1.54(m,5H),1.59-1.74(m,10H),1.89-1.96(m,2H),2.30- 2.35(m,6H),2.48-2.57(m,3H),2.64-2.66(m,2H),3.00-3.03(m,2H),3.49-3.52(m,2H),3.96-3.97(m,4H). LCMS: Rt: 2.360min; MS m/z (ESI): 877.7[M+H]+ .
以下化合物係以與化合物170類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 170 using the corresponding starting materials.
6.57 實例57：製備化合物178.6.57 Example 57: Preparation of Compound 178.
步驟1：製備化合物178-2Step 1: Preparation of compound 178-2
向化合物178-1(10.0g，68.41mmol，1.0eq.)在THF(300mL)中之懸浮液中滴加NaH(3.28g，82.09mmol，1.2eq.)。接著滴加化合物Q(22.07g，102.61mmol，1.5eq.)，且將所得混合物在50℃下攪拌10小時。冷卻至室溫後，將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(PE/EA=4/1)純化殘餘物，得到呈黃色油狀之標題化合物(6.0g，31%產率)。To a suspension of compound 178-1 (10.0 g, 68.41 mmol, 1.0 eq.) in THF (300 mL) was added dropwise NaH (3.28 g, 82.09 mmol, 1.2 eq.). Compound Q (22.07 g, 102.61 mmol, 1.5 eq.) was then added dropwise, and the resulting mixture was stirred at 50°C for 10 hours. After cooling to room temperature, the mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=4/1) to give the title compound (6.0 g, 31% yield) as a yellow oil.
步驟2：製備化合物178-3Step 2: Preparation of compound 178-3
向化合物178-2(6.0g，21.4mmol，1.0eq.)在THF(100mL)中之溶液中加入HCl水溶液(50mL，100mmol，4.7eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用EtOAc萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈無色油狀之標題化合物(4.5g，87%產率)。To a solution of compound 178-2 (6.0 g, 21.4 mmol, 1.0 eq.) in THF (100 mL) was added aqueous HCl (50 mL, 100 mmol, 4.7 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with EtOAc. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give the title compound (4.5 g, 87% yield) as a colorless oil.
步驟3：製備化合物178-4Step 3: Preparation of compound 178-4
向化合物178-3(4.5g，18.73mmol，1.0eq.)及化合物R(8.1g，56.18mmol，3.0eq.)在DCM(200mL)中之溶液中加入DIEA(12.1g，93.63mmol，5.0eq.)、EDCI(10.77g，56.18mmol，3.0eq.)及DMAP(2.29g，18.73mmol，1.0eq.)。將混合物在40℃下攪拌10小時。將反應混合物濃縮並藉由矽膠管柱層析法(PE/EA=10/1)純化，得到呈無色油狀之標題化合物(7.0g，76%產率)。To a solution of compound 178-3 (4.5 g, 18.73 mmol, 1.0 eq.) and compound R (8.1 g, 56.18 mmol, 3.0 eq.) in DCM (200 mL) were added DIEA (12.1 g, 93.63 mmol, 5.0 eq.), EDCI (10.77 g, 56.18 mmol, 3.0 eq.) and DMAP (2.29 g, 18.73 mmol, 1.0 eq.). The mixture was stirred at 40°C for 10 hours. The reaction mixture was concentrated and purified by silica gel column chromatography (PE/EA=10/1) to obtain the title compound (7.0 g, 76% yield) as a colorless oil.
步驟4：製備化合物178-5Step 4: Preparation of compound 178-5
向化合物178-4(7.0g，14.21mmol，1.0eq.)在EtOAc(150mL)中之溶液中加入Pd/C(1.0g)。將混合物在室溫、H2下攪拌10小時。將混合物藉由矽藻土墊過濾並用MeOH洗滌。將濾液濃縮，得到呈黃色油狀之標題化合物(5.1g，58%產率)。To a solution of compound 178-4 (7.0 g, 14.21 mmol, 1.0 eq.) in EtOAc (150 mL) was added Pd/C (1.0 g). The mixture was stirred at room temperature underH2 for 10 h. The mixture was filtered through a celite pad and washed with MeOH. The filtrate was concentrated to give the title compound (5.1 g, 58% yield) as a yellow oil.
步驟5：製備化合物178-6Step 5: Preparation of compound 178-6
向化合物178-5(2.0g，4.97mmol，1.0eq.)及DIPEA(1.93g，14.90mmol，3.0eq.)在DCM(50mL)中之溶液中加入MsCl(850mg，7.45mmol，1.5eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之標題化合物(2.0g，83%產率)。其未經進一步純化即用於下一步驟。To a solution of compound 178-5 (2.0 g, 4.97 mmol, 1.0 eq.) and DIPEA (1.93 g, 14.90 mmol, 3.0 eq.) in DCM (50 mL) was added MsCl (850 mg, 7.45 mmol, 1.5 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give the title compound (2.0 g, 83% yield) as a yellow oil. It was used in the next step without further purification.
步驟6：製備化合物178-7Step 6: Preparation of compound 178-7
向化合物178-6(1.0g，2.08mmol，1.0eq.)及化合物B(480mg，4.16mmol，2.0eq.)在ACN(30mL)中之溶液中加入K2CO3(860mg，6.24mmol，3.0eq.)、Cs2CO3(200mg，0.62mmol，0.3eq.)及NaI(100mg，0.62mmol，0.3eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=10/1)純化，得到呈黃色油狀之標題化合物(500mg，48%產率)。LCMS：Rt：0.800min；MS m/z(ESI)：500.3[M+H]+。To a solution of compound 178-6 (1.0 g, 2.08 mmol, 1.0 eq.) and compound B (480 mg, 4.16 mmol, 2.0 eq.) in ACN (30 mL) were added K2 CO3 (860 mg, 6.24 mmol, 3.0 eq.), Cs2 CO3 (200 mg, 0.62 mmol, 0.3 eq.) and NaI (100 mg, 0.62 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=10/1) to give the title compound (500 mg, 48% yield) as a yellow oil. LCMS: Rt: 0.800min; MS m/z (ESI): 500.3[M+H]+ .
步驟7：製備化合物178-8Step 7: Preparation of compound 178-8
向化合物178-7(300mg，0.6mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(215mg，1.8mmol，3.0eq.)。將混合物在35℃下攪拌10小時。將混合物濃縮，得到呈黃色油狀之標題化合物(311mg，100%產率)。LCMS：Rt：0.467min；MS m/z(ESI)：518.2[M+H]+。To a solution of compound 178-7 (300 mg, 0.6 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (215 mg, 1.8 mmol, 3.0 eq.). The mixture was stirred at 35° C. for 10 hours. The mixture was concentrated to give the title compound (311 mg, 100% yield) as a yellow oil. LCMS: Rt: 0.467 min; MS m/z (ESI): 518.2 [M+H]+ .
步驟8：製備化合物178-9Step 8: Preparation of compound 178-9
向化合物178-6(1.0g，2.08mmol，1.0eq.)及化合物SM6(250mg，4.16mmol，2.0eq.)在ACN(30mL)中之溶液中加入K2CO3(860mg，6.24mmol，3.0eq.)、Cs2CO3(200mg，0.62mmol，0.3eq.)及NaI(100mg，0.62mmol，0.3eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=10/1)純化，得到呈黃色油狀之標題化合物(500mg，54%產率)。LCMS：Rt：0.810min；MS m/z(ESI)：446.3[M+H]+。To a solution of compound 178-6 (1.0 g, 2.08 mmol, 1.0 eq.) and compound SM6 (250 mg, 4.16 mmol, 2.0 eq.) in ACN (30 mL) were added K2 CO3 (860 mg, 6.24 mmol, 3.0 eq.), Cs2 CO3 (200 mg, 0.62 mmol, 0.3 eq.) and NaI (100 mg, 0.62 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=10/1) to give the title compound (500 mg, 54% yield) as a yellow oil. LCMS: Rt: 0.810min; MS m/z (ESI): 446.3[M+H]+ .
步驟9：製備化合物178Step 9: Preparation of compound 178
向化合物178-8(200mg，0.38mmol，1.0eq.)及化合物178-9(180mg，0.38mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(150mg，1.16mmol，3.0eq.)及NaI(60mg，0.38mmol，1.0eq.)。將混合物在70℃下攪拌10小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(50mg，14%產率)。To a solution of compound 178-8 (200 mg, 0.38 mmol, 1.0 eq.) and compound 178-9 (180 mg, 0.38 mmol, 1.0 eq.) in THF (10 mL) was added DIPEA (150 mg, 1.16 mmol, 3.0 eq.) and NaI (60 mg, 0.38 mmol, 1.0 eq.). The mixture was stirred at 70 °C for 10 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give the title compound as a yellow oil (50 mg, 14% yield).
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.27-1.32(m,30H),1.59-1.64(m,10H),1.85-1.99(m,7H),2.28-2.32(m,10H),2.48-2.74(m,10H),3.11-3.15(m,1H),3.43-3.53(m,10H),4.09-4.14(m,8H)。LCMS：Rt：1.080min；MS m/z(ESI)：927.5[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 12H),1.27-1.32(m,30H),1.59-1.64(m,10H),1.85-1.99(m,7H),2.28-2.32(m,10 H),2.48-2.74(m,10H),3.11-3.15(m,1H),3.43-3.53(m,10H),4.09-4.14(m,8H). LCMS: Rt: 1.080min; MS m/z (ESI): 927.5[M+H]+ .
以下化合物係以與化合物178類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 178 using the corresponding starting materials.
6.58 實例58：製備化合物99.6.58 Example 58: Preparation of Compound 99.
步驟1：製備化合物99-2Step 1: Preparation of compound 99-2
將SM7(400.0mg，2.0mmol，1.0eq.)在0℃下溶於甲苯(10.0mL)中，接著加入Py(1.1g，16.0mmol，8.0eq.)及三光氣(355.0mg，1.2mmol，0.6eq.)。將混合物在室溫下攪拌1小時，接著加入化合物99-1(578.0mg，2.4mmol，1.2eq.)。將混合物在室溫下攪拌16小時。TLC顯示反應完成，將混合物倒入H2O中，用EA萃取。將混合物減壓蒸發並藉由FCC(PE/EA=100/1-10/1)純化，得到呈黃色油狀之化合物99-2(0.3g，粗品)。SM7 (400.0 mg, 2.0 mmol, 1.0 eq.) was dissolved in toluene (10.0 mL) at 0°C, followed by the addition of Py (1.1 g, 16.0 mmol, 8.0 eq.) and triphosgene (355.0 mg, 1.2 mmol, 0.6 eq.). The mixture was stirred at room temperature for 1 hour, followed by the addition of compound 99-1 (578.0 mg, 2.4 mmol, 1.2 eq.). The mixture was stirred at room temperature for 16 hours. TLC showed that the reaction was complete, and the mixture was poured into H2 O and extracted with EA. The mixture was evaporated under reduced pressure and purified by FCC (PE/EA=100/1-10/1) to give compound 99-2 (0.3 g, crude product) as a yellow oil.
步驟2：製備化合物99-3Step 2: Preparation of compound 99-3
在室溫下，向化合物99-2(300.0mg，0.7mmol，1.0eq.)及乙醇胺(126.0mg，2.01mmol，3.0eq.)在ACN(15.0mL)中之溶液中加入K2CO3(276.0mg，2.01mmol，3.0eq.)、Cs2CO3(65.0mg，0.2mmol，0.3eq.)及NaI(10.0mg，0.07mmol，0.1eq.)。將混合物在85℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用FCC(DCM/MeOH=1/0-10/1)純化，得到呈無色油狀之化合物99-3(0.16g，粗品)。LCMS：Rt：0.863min；MS m/z(ESI)：415.3[M+H]+。To a solution of compound 99-2 (300.0 mg, 0.7 mmol, 1.0 eq.) and ethanolamine (126.0 mg, 2.01 mmol, 3.0 eq.) in ACN (15.0 mL) were added K2 CO3 (276.0 mg, 2.01 mmol, 3.0 eq.), Cs2 CO3 (65.0 mg, 0.2 mmol, 0.3 eq.) and NaI (10.0 mg, 0.07 mmol, 0.1 eq.) at room temperature. The mixture was stirred at 85° C. for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified with FCC (DCM/MeOH=1/0-10/1) to give compound 99-3 (0.16 g, crude) as a colorless oil. LCMS: Rt: 0.863min; MS m/z (ESI): 415.3[M+H]+ .
步驟3：製備化合物99Step 3: Preparation of compound 99
在0℃下，向化合物99-3(160.0mg，0.4mmol，1.0eq.)及化合物43-3(170.0mg，0.4mmol，3.0eq.)在THF(5.0mL)中之溶液中加入DIEA(153mg，1.2mmol,5.0eq.)及NaI(6.0mg，0.04mmol，0.1eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成，將混合物減壓蒸發並用製備型HPLC純化，得到呈黃色油狀之化合物99(100.0mg，31%產率)。At 0°C, DIEA (153 mg, 1.2 mmol, 5.0 eq.) and NaI (6.0 mg, 0.04 mmol, 0.1 eq.) were added to a solution of compound 99-3 (160.0 mg, 0.4 mmol, 1.0 eq.) and compound 43-3 (170.0 mg, 0.4 mmol, 3.0 eq.) in THF (5.0 mL). The mixture was stirred at 70°C for 16 hours. LCMS showed that the reaction was complete, and the mixture was evaporated under reduced pressure and purified by preparative HPLC to obtain compound 99 (100.0 mg, 31% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.23-1.36(m,48H),1.45-1.50(m,7H),1.59-1.67(m,7H),1.78-1.80(m,4H),2.27-2.31(m,2H),2.49-2.60(m,10H),3.18(s,4H),3.54(s,2H),4.03-4.06(m,4H)。LCMS：Rt：1.560min；MS m/z(ESI)：808.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,9H),1.23-1.36(m,48H),1.45-1.50(m,7H),1.59-1.67(m,7H),1.78-1.80 (m,4H),2.27-2.31(m,2H),2.49-2.60(m,10H),3.18(s,4H),3.54(s,2H),4.03-4.06(m,4H). LCMS: Rt: 1.560min; MS m/z (ESI): 808.7[M+H]+ .
6.59 實例59：製備化合物180.6.59 Example 59: Preparation of Compound 180.
步驟1：製備化合物180-1Step 1: Preparation of compound 180-1
將化合物71-7(1.2g，2.5mmol，1.0eq.)、化合物170-1(500mg，3.7mmol，1.5eq.)、K2CO3(1.0g，7.5mmol，3.0eq.)、Cs2CO3(260mg，0.8mmol，0.3eq.)及NaI(120mg，0.8mmol，0.3eq.)在ACN(30mL)中之溶液在90℃下攪拌隔夜。LCMS顯示反應完成。將混合物濃縮並藉由FCC純化，得到呈黃色油狀之化合物180-1(820mg，70%產率)。A solution of compound 71-7 (1.2 g, 2.5 mmol, 1.0 eq.), compound 170-1 (500 mg, 3.7 mmol, 1.5 eq.), K2 CO3 (1.0 g, 7.5 mmol, 3.0 eq.), Cs2 CO3 (260 mg, 0.8 mmol, 0.3 eq.) and NaI (120 mg, 0.8 mmol, 0.3 eq.) in ACN (30 mL) was stirred at 90° C. overnight. LCMS showed the reaction was complete. The mixture was concentrated and purified by FCC to give compound 180-1 (820 mg, 70% yield) as a yellow oil.
步驟2：製備化合物180Step 2: Preparation of compound 180
將化合物180-1(200mg，0.42mmol，1.0eq.)、化合物SM16(221mg，0.50mmol，1.2eq.)在DCE(5mL)中之溶液在室溫下攪拌隔夜。加入NaBH(AcO)3(176mg，0.83mmol，2.0eq.)。攪拌4小時後，LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物180(28mg，7.9%產率)。A solution of compound 180-1 (200 mg, 0.42 mmol, 1.0 eq.) and compound SM16 (221 mg, 0.50 mmol, 1.2 eq.) in DCE (5 mL) was stirred at room temperature overnight. NaBH(AcO)3 (176 mg, 0.83 mmol, 2.0 eq.) was added. After stirring for 4 hours, LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 180 (28 mg, 7.9% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.42(m,48H),1.59-1.65(m,8H),1.83-1.87(m,2H),1.96-2.07(m,3H),2.28-2.32(m,10H),2.44-2.53(m,3H),2.60-2.64(m,2H),2.93-3.03(m,2H),3.47-3.49(m,2H),4.03-4.07(m,8H)。LCMS：Rt：0.960min；MS m/z(ESI)：909.0[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.42(m,48H),1.59-1.65(m,8H),1.83-1.87(m,2H),1.96-2.07(m,3H),2.28-2 .32(m,10H),2.44-2.53(m,3H),2.60-2.64(m,2H),2.93-3.03(m,2H),3.47-3.49(m,2H),4.03-4.07(m,8H). LCMS: Rt: 0.960min; MS m/z (ESI): 909.0[M+H]+ .
6.60 實例60：製備化合物181.6.60 Example 60: Preparation of Compound 181.
步驟1：製備化合物181-1Step 1: Preparation of compound 181-1
將化合物71-7(800mg，1.67mmol，1.0eq.)、環戊胺(426mg，5.01mmol，3.0eq.)、DIEA(431mg，3.34mmol，2.0eq.)在ACN(10mL)中之溶液在70℃下攪拌隔夜。LCMS顯示反應完成。將混合物濃縮並藉由FCC純化，得到呈黃色油狀之化合物181-1(410mg，52.5%產率)。A solution of compound 71-7 (800 mg, 1.67 mmol, 1.0 eq.), cyclopentylamine (426 mg, 5.01 mmol, 3.0 eq.), and DIEA (431 mg, 3.34 mmol, 2.0 eq.) in ACN (10 mL) was stirred at 70°C overnight. LCMS showed that the reaction was complete. The mixture was concentrated and purified by FCC to give compound 181-1 (410 mg, 52.5% yield) as a yellow oil.
步驟2：製備化合物181-2Step 2: Preparation of compound 181-2
將化合物181-1(410mg，0.88mmol，1.0eq.)、化合物SM27(400mg，2.63mmol，3.0eq.)、K2CO3(363mg，2.63mmol，3.0eq.)、Cs2CO3(85mg，0.26mmol，0.3eq.)及NaI(39mg，0.26mmol，0.3eq.)在ACN(10mL)中之溶液在90℃下攪拌隔夜。LCMS顯示反應完成。將混合物濃縮並藉由FCC純化，得到呈黃色油狀之化合物181-2(320mg，62.3%產率)。A solution of compound 181-1 (410 mg, 0.88 mmol, 1.0 eq.), compound SM27 (400 mg, 2.63 mmol, 3.0 eq.), K2 CO3 (363 mg, 2.63 mmol, 3.0 eq.), Cs2 CO3 (85 mg, 0.26 mmol, 0.3 eq.) and NaI (39 mg, 0.26 mmol, 0.3 eq.) in ACN (10 mL) was stirred at 90° C. overnight. LCMS showed the reaction was complete. The mixture was concentrated and purified by FCC to give compound 181-2 (320 mg, 62.3% yield) as a yellow oil.
步驟3：製備化合物181-3Step 3: Preparation of compound 181-3
將化合物181-2(320mg，0.55mmol，1.0eq.)、TFA(310mg，2.74mmol，5.0eq.)在DCM(5mL)中之混合物在室溫下攪拌隔夜。LCMS顯示反應完成。將混合物用DCM稀釋，用水及鹽水洗滌，乾燥，濃縮，得到呈黃色油狀之粗化合物181-3(230mg，77.6%產率)，其未經進一步純化即用於下一步驟。A mixture of compound 181-2 (320 mg, 0.55 mmol, 1.0 eq.), TFA (310 mg, 2.74 mmol, 5.0 eq.) in DCM (5 mL) was stirred at room temperature overnight. LCMS showed that the reaction was complete. The mixture was diluted with DCM, washed with water and brine, dried, and concentrated to give crude compound 181-3 (230 mg, 77.6% yield) as a yellow oil, which was used in the next step without further purification.
步驟4：製備化合物181Step 4: Preparation of compound 181
將化合物181-3(230mg，0.43mmol，1.0eq.)、化合物SM16(227mg，0.51mmol，1.2eq.)在DCE(5mL)中之溶液在室溫下攪拌隔夜。加入NaBH(AcO)3(186mg，0.86mmol，2.0eq.)。攪拌4小時後，LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物181(110mg，26.5%產率)。A solution of compound 181-3 (230 mg, 0.43 mmol, 1.0 eq.) and compound SM16 (227 mg, 0.51 mmol, 1.2 eq.) in DCE (5 mL) was stirred at room temperature overnight. NaBH(AcO)3 (186 mg, 0.86 mmol, 2.0 eq.) was added. After stirring for 4 hours, LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 181 (110 mg, 26.5% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.29-1.47(m,55H),1.61-1.64(m,8H),1.74-1.80(m,2H),1.94-2.03(m,2H),2.28-2.32(m,8H),2.43-2.47(m,8H),2.55-2.61(m,2H),2.90-2.31(m,1H),3.49-3.55(m,2H),4.00-4.10(m,8H)。LCMS：Rt：1.190min；MS m/z(ESI)：965.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.29-1.47(m,55H),1.61-1.64(m,8H),1.74-1.80(m,2H),1.94-2.03(m,2H),2.28- 2.32(m,8H),2.43-2.47(m,8H),2.55-2.61(m,2H),2.90-2.31(m,1H),3.49-3.55(m,2H),4.00-4.10(m,8H). LCMS: Rt: 1.190min; MS m/z (ESI): 965.7[M+H]+ .
以下化合物係以與化合物181類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 181 using the corresponding starting materials.
6.61 實例61：製備化合物182.6.61 Example 61: Preparation of Compound 182.
步驟1：製備化合物182-1Step 1: Preparation of compound 182-1
向化合物43-1(1.0g，2.86mmol，1.0eq.)及化合物D(495mg，4.29mmol，1.5eq.)在ACN(30mL)中之溶液中加入K2CO3(1.2g，8.59mmol，3.0eq.)、Cs2CO3(28mg，0.09mmol，0.03eq.)及NaI(215mg，1.43mmol，0.5eq.)。將混合物在70℃下攪拌16小時。將混合物濃縮並藉由矽膠管柱層析法(MeOH/DCM=0/1-1/40)純化，得到呈黃色油狀之化合物182-1(1.1g，粗品)。LCMS：Rt：0.800min；MS m/z(ESI)：384.4[M+H]+。To a solution of compound 43-1 (1.0 g, 2.86 mmol, 1.0 eq.) and compound D (495 mg, 4.29 mmol, 1.5 eq.) in ACN (30 mL) were added K2 CO3 (1.2 g, 8.59 mmol, 3.0 eq.), Cs2 CO3 (28 mg, 0.09 mmol, 0.03 eq.) and NaI (215 mg, 1.43 mmol, 0.5 eq.). The mixture was stirred at 70° C. for 16 hours. The mixture was concentrated and purified by silica gel column chromatography (MeOH/DCM=0/1-1/40) to give compound 182-1 (1.1 g, crude product) as a yellow oil. LCMS: Rt: 0.800min; MS m/z (ESI): 384.4[M+H]+ .
步驟2：製備化合物182-2Step 2: Preparation of compound 182-2
將化合物182-1(300mg，0.78mmol，1.0eq.)及SOCl2(279mg，2.35mmol，3.0eq.)在DCM(6mL)中之混合物在35℃下攪拌隔夜。用水淬滅，經EA萃取，用鹽水洗滌，乾燥並濃縮，藉由FCC(MeOH/DCM=0%至20%)純化，得到呈淡黃色油狀之化合物182-2(190mg，60.42%產率)。A mixture of compound 182-1 (300 mg, 0.78 mmol, 1.0 eq.) and SOCl2 (279 mg, 2.35 mmol, 3.0 eq.) in DCM (6 mL) was stirred at 35° C. overnight. It was quenched with water, extracted with EA, washed with brine, dried and concentrated, and purified by FCC (MeOH/DCM=0% to 20%) to give compound 182-2 (190 mg, 60.42% yield) as a light yellow oil.
步驟3：製備化合物182Step 3: Preparation of compound 182
在室溫下，向化合物182-2(190mg，0.47mmol，1.0eq.)在THF(4mL)中之溶液中加入化合物SM16(210mg，0.47mmol，1.0eq.)、DIEA(183mg，1.43mmol，3.0eq.)、NaI(35mg，0.24mmol，0.5eq.)。將混合物在70℃下攪拌隔夜。將混合物用水、鹽水洗滌，濃縮有機層並藉由製備型HPLC純化，得到呈無色油狀之化合物182(57mg，15.55%產率)。Compound SM16 (210 mg, 0.47 mmol, 1.0 eq.), DIEA (183 mg, 1.43 mmol, 3.0 eq.), and NaI (35 mg, 0.24 mmol, 0.5 eq.) were added to a solution of compound 182-2 (190 mg, 0.47 mmol, 1.0 eq.) in THF (4 mL) at room temperature. The mixture was stirred at 70°C overnight. The mixture was washed with water and brine, the organic layer was concentrated and purified by preparative HPLC to obtain compound 182 (57 mg, 15.55% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,9H),1.25-1.35(m,32H),1.43-1.45(m,4H),1.56-1.65(m,17H),1.85-1.99(m,5H),2.27-2.32(m,6H),2.41-2.59(m,10H),3.05-3.10(m,1H),3.53-3.55(m,2H),4.03-4.07(m,6H)。LCMS：Rt：1.230min；MS m/z(ESI)：809.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,9H),1.25-1.35(m,32H),1.43-1.45(m,4H),1.56-1.65(m,17H),1.85-1.99(m,5H) ),2.27-2.32(m,6H),2.41-2.59(m,10H),3.05-3.10(m,1H),3.53-3.55(m,2H),4.03-4.07(m,6H). LCMS: Rt: 1.230min; MS m/z (ESI): 809.6[M+H]+ .
以下化合物係以與化合物182類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 182 using the corresponding starting materials.
6.62 實例62：製備化合物186.6.62 Example 62: Preparation of Compound 186.
步驟1：製備化合物186-1Step 1: Preparation of compound 186-1
向化合物76-1(600mg，1.29mmol，1.0eq.)在ACN(20mL)中之溶液中加入化合物SM13(184mg，2.58mmol，2.0eq.)、K2CO3(537mg，3.87mmol，3.0eq.)、Cs2CO3(126mg，0.38mmol，0.3eq.)及NaI(56mg，0.38mmol，0.3eq.)。將混合物在80℃下攪拌10小時。TLC顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化殘餘物，得到呈無色油狀之化合物186-1(350mg，61.8%產率)。To a solution of compound 76-1 (600 mg, 1.29 mmol, 1.0 eq.) in ACN (20 mL) were added compound SM13 (184 mg, 2.58 mmol, 2.0 eq.), K2 CO3 (537 mg, 3.87 mmol, 3.0 eq.), Cs2 CO3 (126 mg, 0.38 mmol, 0.3 eq.) and NaI (56 mg, 0.38 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 10 hours. TLC showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=10/1) to obtain compound 186-1 (350 mg, 61.8% yield) as a colorless oil.
步驟2：製備化合物186-2Step 2: Preparation of compound 186-2
向化合物186-1(350mg，0.8mmol，1.0eq.)及化合物SM27(360mg，2.4mmol，3.0eq.)在ACN(25mL)中之溶液中加入K2CO3(332mg，2.4mmol，3.0eq.)、Cs2CO3(78mg，0.24mmol，0.3eq.)及NaI(34mg，0.24mmol，0.3eq.)。將混合物在80℃下攪拌16小時。TLC顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化殘餘物，得到呈無色油狀之化合物186-2(380mg，85.7%產率)。To a solution of compound 186-1 (350 mg, 0.8 mmol, 1.0 eq.) and compound SM27 (360 mg, 2.4 mmol, 3.0 eq.) in ACN (25 mL) were added K2 CO3 (332 mg, 2.4 mmol, 3.0 eq.), Cs2 CO3 (78 mg, 0.24 mmol, 0.3 eq.) and NaI (34 mg, 0.24 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. TLC showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=10/1) to obtain compound 186-2 (380 mg, 85.7% yield) as a colorless oil.
步驟3：製備化合物186-3Step 3: Preparation of compound 186-3
向化合物186-2(380mg，0.68mmol，1.0eq.)在DCM(20mL)中之溶液中加入TFA(1mL)。將混合物在25℃下攪拌10小時。LCMS顯示反應完成。將反應混合物用EA及Na2CO3溶液萃取。將有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物186-3(280mg，86.1%產率)。To a solution of compound 186-2 (380 mg, 0.68 mmol, 1.0 eq.) in DCM (20 mL) was added TFA (1 mL). The mixture was stirred at 25 °C for 10 h. LCMS showed the reaction was complete. The reaction mixture was extracted with EA and Na2 CO3 solution. The organic layer was washed with brine, dried over Na2 SO4 and concentrated to give compound 186-3 (280 mg, 86.1% yield) as a yellow oil.
步驟4：製備化合物186Step 4: Preparation of compound 186
向化合物186-3(280mg，0.55mmol，1.0eq.)及化合物SM16(243mg，0.55mmol，1.0eq.)在DCE(10mL)中之溶液中加入兩滴CH3COOH，且在25℃下攪拌2小時，接著在25℃下加入NaBH(OAc)3(233mg，1.1mmol，2.0eq.)並攪拌10小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由製備型HPLC純化殘餘物，得到呈無色油狀之化合物186(10mg，2.1%產率)。To a solution of compound 186-3 (280 mg, 0.55 mmol, 1.0 eq.) and compound SM16 (243 mg, 0.55 mmol, 1.0 eq.) in DCE (10 mL) were added two drops of CH3 COOH and stirred at 25° C. for 2 hours, followed by the addition of NaBH(OAc)3 (233 mg, 1.1 mmol, 2.0 eq.) at 25° C. and stirred for 10 hours. LCMS showed the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by preparative HPLC to give compound 186 (10 mg, 2.1% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,12H),1.26-1.61(m,54H),1.63-1.71(m,18H),2.22-2.31(m,8H),2.76-2.92(m,6H),2.91-3.17(m,2H),3.44-3.80(m,6H),3.81-4.07(m,4H),5.70-5.83(m,1H)。LCMS：Rt：1.150min；MS m/z(ESI)：934.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,12H),1.26-1.61(m,54H),1.63-1.71(m,18H),2.22-2.31(m,8H),2.76- 2.92(m,6H),2.91-3.17(m,2H),3.44-3.80(m,6H),3.81-4.07(m,4H),5.70-5.83(m,1H). LCMS: Rt: 1.150min; MS m/z (ESI): 934.7[M+H]+ .
6.63 實例63：製備化合物187.6.63 Example 63: Preparation of Compound 187.
步驟1：製備化合物187-1Step 1: Preparation of compound 187-1
向化合物26-1(1.0g，2.23mmol，1.0eq.)及化合物SM13(477mg，6.70mmol，3.0eq.)在ACN(30mL)中之溶液中加入K2CO3(925mg，6.70mmol，3.0eq.)、Cs2CO3(22mg，0.07mmol，0.03eq.)及NaI(168mg，1.12mmol，0.5eq.)。將混合物在80℃下攪拌16小時。將混合物濃縮並藉由矽膠管柱層析法(MeOH/DCM=0/1-1/20)純化，得到呈深棕色油狀之化合物187-1(598mg，61.12%產率)。LCMS：Rt：0.970min；MS m/z(ESI)：438.5[M+H]+。To a solution of compound 26-1 (1.0 g, 2.23 mmol, 1.0 eq.) and compound SM13 (477 mg, 6.70 mmol, 3.0 eq.) in ACN (30 mL) were added K2 CO3 (925 mg, 6.70 mmol, 3.0 eq.), Cs2 CO3 (22 mg, 0.07 mmol, 0.03 eq.) and NaI (168 mg, 1.12 mmol, 0.5 eq.). The mixture was stirred at 80° C. for 16 hours. The mixture was concentrated and purified by silica gel column chromatography (MeOH/DCM=0/1-1/20) to give compound 187-1 (598 mg, 61.12% yield) as a dark brown oil. LCMS: Rt: 0.970min; MS m/z (ESI): 438.5[M+H]+ .
步驟2：製備化合物187-2Step 2: Preparation of compound 187-2
向化合物187-1(598mg，1.37mmol，1.0eq.)及化合物SM28(247mg，1.50mmol，1.1eq.)在ACN(12mL)中之溶液中加入K2CO3(565mg，4.10mmol，3.0eq.)、Cs2CO3(13mg，0.04mmol，0.03eq.)及NaI(102mg，0.68mmol，0.5eq.)。將混合物在80℃下攪拌16小時。將混合物濃縮並藉由矽膠管柱層析法(MeOH/DCM=0/1-1/60)純化，得到呈淡黃色油狀之化合物187-2(485mg，62.74%產率)。LCMS：Rt：1.015min；MS m/z(ESI)：566.6[M+H]+。To a solution of compound 187-1 (598 mg, 1.37 mmol, 1.0 eq.) and compound SM28 (247 mg, 1.50 mmol, 1.1 eq.) in ACN (12 mL) were added K2 CO3 (565 mg, 4.10 mmol, 3.0 eq.), Cs2 CO3 (13 mg, 0.04 mmol, 0.03 eq.) and NaI (102 mg, 0.68 mmol, 0.5 eq.). The mixture was stirred at 80° C. for 16 hours. The mixture was concentrated and purified by silica gel column chromatography (MeOH/DCM = 0/1-1/60) to give compound 187-2 (485 mg, 62.74% yield) as a light yellow oil. LCMS: Rt: 1.015min; MS m/z (ESI): 566.6[M+H]+ .
步驟3：製備化合物187-3Step 3: Preparation of compound 187-3
在室溫下，向化合物187-2(264mg，0.47mmol，1.0eq.)在DCM(6mL)中之攪拌溶液中加入TFA(6mL，80.4mmol，172.5eq.)。將混合物在室溫下攪拌隔夜。用飽和碳酸氫鈉淬滅，用EA萃取，用鹽水洗滌，乾燥並濃縮，得到呈深棕色油狀之化合物187-3(221mg，產率90.77%)。LCMS：Rt：0.960min；MS m/z(ESI)：522.5[M+H]+。To a stirred solution of compound 187-2 (264 mg, 0.47 mmol, 1.0 eq.) in DCM (6 mL) was added TFA (6 mL, 80.4 mmol, 172.5 eq.) at room temperature. The mixture was stirred at room temperature overnight. It was quenched with saturated sodium bicarbonate, extracted with EA, washed with brine, dried and concentrated to give compound 187-3 (221 mg, yield 90.77%) as a dark brown oil. LCMS: Rt: 0.960 min; MS m/z (ESI): 522.5 [M+H]+ .
步驟4：製備化合物187Step 4: Preparation of compound 187
將化合物187-3(110mg，0.21mmol，1.0eq.)、化合物SM16(94mg，0.21mmol，1.0eq.)及兩滴乙酸在DCE(6mL)中之混合物在室溫下攪拌2小時。將NaBH(OAc)3(89mg，0.42mmol，2.0eq.)加入上述混合物中，在室溫下攪拌隔夜。將混合物用水淬滅，經DCM萃取，濃縮有機層並藉由製備型HPLC純化，得到呈淡黃色油狀之化合物187(32mg，15.99%產率)。A mixture of compound 187-3 (110 mg, 0.21 mmol, 1.0 eq.), compound SM16 (94 mg, 0.21 mmol, 1.0 eq.) and two drops of acetic acid in DCE (6 mL) was stirred at room temperature for 2 hours. NaBH(OAc)3 (89 mg, 0.42 mmol, 2.0 eq.) was added to the above mixture and stirred at room temperature overnight. The mixture was quenched with water, extracted with DCM, and the organic layer was concentrated and purified by preparative HPLC to give compound 187 (32 mg, 15.99% yield) as a light yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.25-1.35(m,57H),1.40-1.47(m,6H),1.61-1.63(m,11H),1.95-2.15(m,4H),2.28-2.32(m,6H),2.44-2.62(m,8H),3.50-3.60(m,2H),3.96-4.07(m,6H)。LCMS：Rt：1.190min；MS m/z(ESI)：949.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.25-1.35(m,57H),1.40-1.47(m,6H),1.61-1.63(m,11H),1.95-2.15(m,4H),2.28-2.32(m,6H),2.44-2.62(m, 8H),3.50-3.60(m,2H),3.96-4.07(m,6H). LCMS: Rt: 1.190min; MS m/z (ESI): 949.8[M+H]+ .
以下化合物係以與化合物187類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 187 using the corresponding starting materials.
6.64 實例64：製備化合物188.6.64 Example 64: Preparation of Compound 188.
步驟1：製備化合物188-2Step 1: Preparation of compound 188-2
向化合物188-1(1.0g，2.87mmol，1.0eq.)及化合物D(660mg，5.74mmol，2.0eq.)在ACN(30mL)中之溶液中加入K2CO3(1.2g，8.61mmol，3.0eq.)、Cs2CO3(28mg，0.09mmol，0.03eq.)及NaI(216mg，1.44mmol，0.5eq.)。將混合物在80℃下攪拌16小時。將混合物濃縮並藉由矽膠管柱層析法(MeOH/DCM=0/1-1/30)純化，得到呈黃色油狀之化合物188-2(632mg，57.55%產率)。LCMS：Rt：0.770min；MS m/z(ESI)：383.4[M+H]+。To a solution of compound 188-1 (1.0 g, 2.87 mmol, 1.0 eq.) and compound D (660 mg, 5.74 mmol, 2.0 eq.) in ACN (30 mL) were added K2 CO3 (1.2 g, 8.61 mmol, 3.0 eq.), Cs2 CO3 (28 mg, 0.09 mmol, 0.03 eq.) and NaI (216 mg, 1.44 mmol, 0.5 eq.). The mixture was stirred at 80° C. for 16 hours. The mixture was concentrated and purified by silica gel column chromatography (MeOH/DCM=0/1-1/30) to give compound 188-2 (632 mg, 57.55% yield) as a yellow oil. LCMS: Rt: 0.770min; MS m/z (ESI): 383.4[M+H]+ .
步驟2：製備化合物188-3Step 2: Preparation of compound 188-3
在冰浴中向化合物188-2(341mg，0.89mmol，1.0eq.)及DIEA(172mg，1.34mmol，1.5eq.)在DCM(6mL)中之攪拌溶液中滴加MsCl(112mg，0.98mmol，1.1eq.)。將混合物在室溫下攪拌1小時。用水淬滅，經EA萃取，用鹽水洗滌，乾燥並濃縮，藉由FCC(MeOH/DCM=0%至1.67%)純化，得到呈黃色油狀之化合物188-3(141mg，34.99%產率)。LCMS：Rt：0.790min；MS m/z(ESI)：401.4[M+H]+。To a stirred solution of compound 188-2 (341 mg, 0.89 mmol, 1.0 eq.) and DIEA (172 mg, 1.34 mmol, 1.5 eq.) in DCM (6 mL) was added MsCl (112 mg, 0.98 mmol, 1.1 eq.) dropwise in an ice bath. The mixture was stirred at room temperature for 1 hour. Quenched with water, extracted with EA, washed with brine, dried and concentrated, and purified by FCC (MeOH/DCM = 0% to 1.67%) to give compound 188-3 (141 mg, 34.99% yield) as a yellow oil. LCMS: Rt: 0.790 min; MS m/z (ESI): 401.4 [M+H]+ .
步驟3：製備化合物188Step 3: Preparation of compound 188
將化合物188-3(120mg，0.30mmol，1.0eq.)、K2CO3(124mg，0.90mmol，3.0eq.)、Cs2CO3(3mg，0.01mmol，0.03eq.)、化合物SM16(159mg，0.36mmol，1.2eq.)及NaI(22mg，0.15mmol，0.5eq.)在ACN(5mL)中之混合物在90℃下攪拌48小時。將混合物用水、鹽水洗滌，濃縮有機層並藉由製備型HPLC純化，得到呈無色油狀之化合物188(32mg，13.23%產率)。A mixture of compound 188-3 (120 mg, 0.30 mmol, 1.0 eq.), K2 CO3 (124 mg, 0.90 mmol, 3.0 eq.), Cs2 CO3 (3 mg, 0.01 mmol, 0.03 eq.), compound SM16 (159 mg, 0.36 mmol, 1.2 eq.) and NaI (22 mg, 0.15 mmol, 0.5 eq.) in ACN (5 mL) was stirred at 90° C. for 48 hours. The mixture was washed with water and brine, and the organic layer was concentrated and purified by preparative HPLC to give compound 188 (32 mg, 13.23% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.26-1.35(m,41H),1.47-1.49(m,6H),1.59-1.65(m,8H),1.95-2.02(m,8H),2.15-2.20(m,2H),2.28-2.32(m,5H),2.50-2.66(m,6H),3.20-3.25(m,2H),3.55-3.61(m,2H),4.00-4.10(m,4H)。LCMS：Rt：1.050min；MS m/z(ESI)：808.7[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.86-0.90 (m, 9H), 1.26-1.35 (m, 41H), 1.47-1.49 (m, 6H), 1.59-1.65 (m, 8H),1.95-2.02(m,8H),2.15-2.20(m,2H),2.28-2.32(m,5H),2.50-2.66(m,6H),3.20-3.25(m,2H),3.55-3.61(m,2H),4.00-4.10(m,4H). LCMS: Rt: 1.050min; MS m/z (ESI): 808.7[M+H]+ .
以下化合物係以與化合物188類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 188 using the corresponding starting materials.
6.65 實例65：製備化合物190.6.65 Example 65: Preparation of Compound 190.
步驟1：製備化合物190-1Step 1: Preparation of compound 190-1
向化合物SM24(1g，2.3mmol，1.0eq.)在ACN(25mL)中之溶液中加入化合物SM29(0.37g，6.9mmol，3.0eq.)、K2CO3(0.98g，6.9mmol，3.0eq.)、Cs2CO3(0.23g，0.69mmol，0.3eq.)及NaI(0.1g，0.69mmol，0.3eq.)。將混合物在80℃下攪拌10小時。TLC顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化殘餘物，得到呈無色油狀之化合物190-1(500mg，53%產率)。To a solution of compound SM24 (1 g, 2.3 mmol, 1.0 eq.) in ACN (25 mL) were added compound SM29 (0.37 g, 6.9 mmol, 3.0 eq.), K2 CO3 (0.98 g, 6.9 mmol, 3.0 eq.), Cs2 CO3 (0.23 g, 0.69 mmol, 0.3 eq.) and NaI (0.1 g, 0.69 mmol, 0.3 eq.). The mixture was stirred at 80°C for 10 hours. TLC showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=10/1) to give compound 190-1 (500 mg, 53% yield) as a colorless oil.
步驟2：製備化合物190-2Step 2: Preparation of compound 190-2
向化合物190-1(500mg，1.26mmol，1.0eq.)及化合物SM27(578mg，3.78mmol，3.0eq.)在ACN(15mL)中之溶液中加入K2CO3(523mg，3.78mmol，3.0eq.)、Cs2CO3(123mg，0.38mmol，0.3eq.)及NaI(54mg，0.38mmol，0.3eq.)。將混合物在80℃下攪拌16小時。TLC顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化殘餘物，得到呈無色油狀之化合物190-2(523mg，80.9%產率)。To a solution of compound 190-1 (500 mg, 1.26 mmol, 1.0 eq.) and compound SM27 (578 mg, 3.78 mmol, 3.0 eq.) in ACN (15 mL) were added K2 CO3 (523 mg, 3.78 mmol, 3.0 eq.), Cs2 CO3 (123 mg, 0.38 mmol, 0.3 eq.) and NaI (54 mg, 0.38 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. TLC showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=10/1) to give compound 190-2 (523 mg, 80.9% yield) as a colorless oil.
步驟3：製備化合物190-3Step 3: Preparation of compound 190-3
向化合物190-2(523mg，1.08mmol，1.0eq.)在DCM(15mL)中之溶液中加入TFA(1mL)。將混合物在25℃下攪拌10小時。LCMS顯示反應完成。將反應混合物用EA及Na2CO3溶液萃取。將有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈無色油狀之化合物190-3(380mg，79.8%產率)。To a solution of compound 190-2 (523 mg, 1.08 mmol, 1.0 eq.) in DCM (15 mL) was added TFA (1 mL). The mixture was stirred at 25 °C for 10 h. LCMS showed the reaction was complete. The reaction mixture was extracted with EA and Na2 CO3 solution. The organic layer was washed with brine, dried over Na2 SO4 and concentrated to give compound 190-3 (380 mg, 79.8% yield) as a colorless oil.
步驟4：製備化合物190Step 4: Preparation of compound 190
向化合物190-3(350mg，0.75mmol，1.0eq.)及化合物SM16(332mg，0.75mmol，1.0eq.)在DCE(10mL)中之溶液中加入兩滴CH3COOH並在25℃下攪拌2小時，接著在25℃下加入NaBH(OAc)3(310mg，1.5mmol，2.0eq.)並攪拌10小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由製備型HPLC純化殘餘物，得到呈無色油狀之化合物190(70mg，10.4%產率)。To a solution of compound 190-3 (350 mg, 0.75 mmol, 1.0 eq.) and compound SM16 (332 mg, 0.75 mmol, 1.0 eq.) in DCE (10 mL) was added two drops of CH3 COOH and stirred at 25° C. for 2 hours, followed by addition of NaBH(OAc)3 (310 mg, 1.5 mmol, 2.0 eq.) at 25° C. and stirred for 10 hours. LCMS showed the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by preparative HPLC to give compound 190 (70 mg, 10.4% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.51-0.86(m,12H),1.28-1.39(m,48H),1.60-1.68(m,24H),2.28-2.31(m,8H),2.32-2.69(m,6H),3.96-4.06(m,6H)。LCMS：Rt：1.150min；MS m/z(ESI)：893.7[M+H]+。1 H NMR (400MHz, CDCl3 ) δ: 0.51-0.86(m,12H), 1.28-1.39(m,48H), 1.60-1.68(m,24H), 2.28-2.31(m,8H), 2.32-2.69(m,6H), 3.96-4.06(m,6H). LCMS: Rt: 1.150min; MS m/z (ESI): 893.7[M+H]+ .
以下化合物係以與化合物190類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 190 using the corresponding starting materials.
6.66 實例66：製備化合物195.6.66 Example 66: Preparation of Compound 195.
步驟1：製備化合物195-1Step 1: Preparation of compound 195-1
將化合物149-2(0.6g，1.43mmol，1.0eq.)、化合物SM29(250mg，4.3mmol，3.0eq.)、K2CO3(590mg，4.3mmol，3.0eq.)、Cs2CO3(140mg，0.43mmol，0.3eq.)及NaI(65mg，0.43mmol，0.3eq.)在ACN(10mL)中之溶液在80℃下攪拌隔夜。TLC顯示反應完成。將混合物濃縮並藉由FCC純化，得到呈黃色油狀之化合物195-1(300mg，53.15%產率)。A solution of compound 149-2 (0.6 g, 1.43 mmol, 1.0 eq.), compound SM29 (250 mg, 4.3 mmol, 3.0 eq.), K2 CO3 (590 mg, 4.3 mmol, 3.0 eq.), Cs2 CO3 (140 mg, 0.43 mmol, 0.3 eq.) and NaI (65 mg, 0.43 mmol, 0.3 eq.) in ACN (10 mL) was stirred at 80° C. overnight. TLC showed the reaction was complete. The mixture was concentrated and purified by FCC to give compound 195-1 (300 mg, 53.15% yield) as a yellow oil.
步驟2：製備化合物195-2Step 2: Preparation of compound 195-2
將化合物195-1(300mg，0.76mmol，1.0eq.)、化合物SM27(350mg，2.28mmol，3.0eq.)、K2CO3(320mg，2.28mmol，3.0eq.)、Cs2CO3(75mg，0.23mmol，0.3eq.)及NaI(35mg，0.23mmol，0.3eq.)在ACN(10mL)中之溶液在80℃下攪拌40小時。TLC顯示反應完成。將混合物濃縮並藉由FCC純化，得到呈黃色油狀之化合物195-2(300mg，77.28%產率)。A solution of compound 195-1 (300 mg, 0.76 mmol, 1.0 eq.), compound SM27 (350 mg, 2.28 mmol, 3.0 eq.), K2 CO3 (320 mg, 2.28 mmol, 3.0 eq.), Cs2 CO3 (75 mg, 0.23 mmol, 0.3 eq.) and NaI (35 mg, 0.23 mmol, 0.3 eq.) in ACN (10 mL) was stirred at 80° C. for 40 hours. TLC showed that the reaction was complete. The mixture was concentrated and purified by FCC to give compound 195-2 (300 mg, 77.28% yield) as a yellow oil.
步驟3：製備化合物195-3Step 3: Preparation of compound 195-3
將化合物195-2(300mg，0.59mmol，1.0eq.)及TFA(0.5mL)在DCM(10mL)中之混合物在室溫下攪拌隔夜。TLC顯示反應完成。將混合物用DCM稀釋，用水及鹽水洗滌，乾燥，濃縮，得到呈黃色油狀之化合物195-3(280mg，粗品)，其未經進一步純化即用於下一步驟。A mixture of compound 195-2 (300 mg, 0.59 mmol, 1.0 eq.) and TFA (0.5 mL) in DCM (10 mL) was stirred at room temperature overnight. TLC showed that the reaction was complete. The mixture was diluted with DCM, washed with water and brine, dried, and concentrated to give compound 195-3 (280 mg, crude) as a yellow oil, which was used in the next step without further purification.
步驟4：製備化合物195Step 4: Preparation of compound 195
將化合物195-3(280mg，0.59mmol，1.0eq.)、化合物SM16(310mg，0.71mmol，1.2eq.)在DCE(10mL)中之溶液在室溫下攪拌隔夜。加入NaBH(AcO)3(250mg，1.2mmol，2.0eq.)。攪拌24小時後，LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物195(110mg，20.89%產率)。A solution of compound 195-3 (280 mg, 0.59 mmol, 1.0 eq.) and compound SM16 (310 mg, 0.71 mmol, 1.2 eq.) in DCE (10 mL) was stirred at room temperature overnight. NaBH(AcO)3 (250 mg, 1.2 mmol, 2.0 eq.) was added. After stirring for 24 hours, LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 195 (110 mg, 20.89% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.36-0.45(m,4H),0.86-0.90(m,12H),1.26-1.35(m,46H),1.40-1.55(m,8H),0.60-1.77(m,9H),1.97-2.00(m,1H),2.15-2.19(m,2H),2.29-2.32(m,4H),2.43-2.59(m,10H),3.16-3.19(m,2H),3.51-3.54(m,2H),4.00-4.10(m,4H),5.50(s,1H)。LCMS：Rt：0.080min；MS m/z(ESI)：892.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.36-0.45(m,4H),0.86-0.90(m,12H),1.26-1.35(m,46H),1.40-1.55(m,8H),0.60-1.77(m,9H),1.97-2.00(m,1H),2.1 5-2.19(m,2H),2.29-2.32(m,4H),2.43-2.59(m,10H),3.16-3.19(m,2H),3.51-3.54(m,2H),4.00-4.10(m,4H),5.50(s,1H). LCMS: Rt: 0.080min; MS m/z (ESI): 892.6[M+H]+ .
以下化合物係以與化合物195類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 195 using the corresponding starting materials.
6.67 實例67：製備化合物200.6.67 Example 67: Preparation of Compound 200.
步驟1：製備化合物200-1Step 1: Preparation of compound 200-1
在0℃下，向化合物182-1(650mg，1.7mmol，1.0eq.)及DIPEA(880mg，6.8mmol，4.0eq.)在DCM(10mL)中之溶液中加入MsCl(390mg，3.4mmol,3.0eq.)。將混合物再攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物200-1(600mg，76.44%產率)。To a solution of compound 182-1 (650 mg, 1.7 mmol, 1.0 eq.) and DIPEA (880 mg, 6.8 mmol, 4.0 eq.) in DCM (10 mL) at 0°C was added MsCl (390 mg, 3.4 mmol, 3.0 eq.). The mixture was stirred for another 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated to give compound 200-1 (600 mg, 76.44% yield) as a yellow oil.
步驟2：製備化合物200Step 2: Preparation of compound 200
將化合物200-1(600mg，1.3mmol，1.0eq.)、化合物SM30(630mg，1.56mmol，1.2eq.)、K2CO3(540mg，3.9mmol，3.0eq.)、Cs2CO3(130mg，0.39mmol，0.3eq.)及NaI(60mg，0.39mmol，0.3eq.)在ACN(10mL)中之溶液在80℃下攪拌隔夜。TLC顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物200(50mg，5.01%產率)。A solution of compound 200-1 (600 mg, 1.3 mmol, 1.0 eq.), compound SM30 (630 mg, 1.56 mmol, 1.2 eq.), K2 CO3 (540 mg, 3.9 mmol, 3.0 eq.), Cs2 CO3 (130 mg, 0.39 mmol, 0.3 eq.) and NaI (60 mg, 0.39 mmol, 0.3 eq.) in ACN (10 mL) was stirred at 80° C. overnight. TLC showed the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 200 (50 mg, 5.01% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,9H),1.26(s,35H),1.41-1.46(m,6H),1.54-1.66(m,14H),1.85-1.99(m,6H),2.27-2.31(m,2H),2.39-2.59(m,12H),3.04-3.08(m,1H),3.36-3.42(m,2H),3.52-3.58(m,4H),4.03-4.07(m,2H),4.43-4.46(m,1H)。LCMS：Rt：1.730min；MS m/z(ESI)：767.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,9H),1.26(s,35H),1.41-1.46(m,6H),1.54-1.66(m,14H),1.85-1.99(m,6H),2.27-2.31(m,2H), 2.39-2.59(m,12H),3.04-3.08(m,1H),3.36-3.42(m,2H),3.52-3.58(m,4H),4.03-4.07(m,2H),4.43-4.46(m,1H). LCMS: Rt: 1.730min; MS m/z (ESI): 767.6[M+H]+ .
6.68 實例68：製備化合物201.6.68 Example 68: Preparation of Compound 201.
步驟1：製備化合物201-1Step 1: Preparation of compound 201-1
向化合物SM24(1.0g，2.38mmol，1.0eq.)及化合物D(550mg，4.77mmol，2.0eq.)在ACN(50mL)中之溶液中加入K2CO3(1.0g，7.15mmol，3.0eq.)、Cs2CO3(230mg，0.71mmol，0.3eq.)及NaI(110mg，0.71mmol，0.3eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化殘餘物，得到呈黃色油狀之化合物201-1(600mg，55%產率)。LCMS：Rt：0.940min；MS m/z(ESI)：454.4[M+H]+。To a solution of compound SM24 (1.0 g, 2.38 mmol, 1.0 eq.) and compound D (550 mg, 4.77 mmol, 2.0 eq.) in ACN (50 mL) were added K2 CO3 (1.0 g, 7.15 mmol, 3.0 eq.), Cs2 CO3 (230 mg, 0.71 mmol, 0.3 eq.) and NaI (110 mg, 0.71 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 10 h. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=10/1) to obtain compound 201-1 (600 mg, 55% yield) as a yellow oil. LCMS: Rt: 0.940 min; MS m/z (ESI): 454.4 [M+H]+ .
步驟2：製備化合物201-2Step 2: Preparation of compound 201-2
向化合物201-1(300mg，0.66mmol，1.0eq.)及DIPEA(260mg，1.99mmol，3.0eq.)在DCM(20mL)中之溶液中加入MsCl(115mg，0.99mmol，1.5eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物201-2(280mg，80%產率)。To a solution of compound 201-1 (300 mg, 0.66 mmol, 1.0 eq.) and DIPEA (260 mg, 1.99 mmol, 3.0 eq.) in DCM (20 mL) was added MsCl (115 mg, 0.99 mmol, 1.5 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 201-2 (280 mg, 80% yield) as a yellow oil.
步驟3：製備化合物201Step 3: Preparation of compound 201
向化合物201-2(250mg，0.47mmol，1.0eq.)及化合物SM30(190mg，0.47mmol，1.0eq.)在ACN(10mL)中之溶液中加入K2CO3(195mg，1.41mmol，3.0eq.)、Cs2CO3(46mg，0.14mmol，0.3eq.)及NaI(21mg，0.14mmol，0.3eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物201(35mg，9%產率)。To a solution of compound 201-2 (250 mg, 0.47 mmol, 1.0 eq.) and compound SM30 (190 mg, 0.47 mmol, 1.0 eq.) in ACN (10 mL) were added K2 CO3 (195 mg, 1.41 mmol, 3.0 eq.), Cs2 CO3 (46 mg, 0.14 mmol, 0.3 eq.) and NaI (21 mg, 0.14 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The mixture was concentrated and purified by preparative HPLC to give compound 201 (35 mg, 9% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.80-0.83(m,12H),1.20-1.56(m,66H),1.82-1.95(m,4H),2.23-2.53(m,12H),2.92-3.09(m,1H),3.31-3.40(m,2H),3.47-3.51(m,4H),3.89-3.90(m,2H),4.35-4.39(m,1H)。LCMS：Rt：2.170min；MS m/z(ESI)：837.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.80-0.83(m,12H),1.20-1.56(m,66H),1.82-1.95(m,4H),2.23-2.53(m,12H),2.92- 3.09(m,1H),3.31-3.40(m,2H),3.47-3.51(m,4H),3.89-3.90(m,2H),4.35-4.39(m,1H). LCMS: Rt: 2.170min; MS m/z (ESI): 837.7[M+H]+ .
6.69 實例69：製備化合物202.6.69 Example 69: Preparation of Compound 202.
步驟1：製備化合物202-1Step 1: Preparation of compound 202-1
向化合物149-2(0.6g，1.43mmol，1.0eq.)及化合物K(450mg，2.87mmol，2.0eq.)在ACN(30mL)中之溶液中加入K2CO3(0.6g，4.30mmol，3.0eq.)、Cs2CO3(140mg，0.43mmol，0.3eq.)及NaI(65mg，0.43mmol，0.3eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=10/1)純化殘餘物，得到呈黃色油狀之化合物202-1(350mg，48%產率)。LCMS：Rt：0.840min；MS m/z(ESI)：495.5[M+H]+。To a solution of compound 149-2 (0.6 g, 1.43 mmol, 1.0 eq.) and compound K (450 mg, 2.87 mmol, 2.0 eq.) in ACN (30 mL) were added K2 CO3 (0.6 g, 4.30 mmol, 3.0 eq.), Cs2 CO3 (140 mg, 0.43 mmol, 0.3 eq.) and NaI (65 mg, 0.43 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=10/1) to obtain compound 202-1 (350 mg, 48% yield) as a yellow oil. LCMS: Rt: 0.840 min; MS m/z (ESI): 495.5 [M+H]+ .
步驟2：製備化合物202-2Step 2: Preparation of compound 202-2
向化合物202-1(350mg，0.71mmol，1.0eq.)及DIPEA(280mg，2.12mmol，3.0eq.)在DCM(20mL)中之溶液中加入MsCl(120mg，1.06mmol，1.5eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物202-2(280mg，69%產率)。To a solution of compound 202-1 (350 mg, 0.71 mmol, 1.0 eq.) and DIPEA (280 mg, 2.12 mmol, 3.0 eq.) in DCM (20 mL) was added MsCl (120 mg, 1.06 mmol, 1.5 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 202-2 (280 mg, 69% yield) as a yellow oil.
步驟3：製備化合物202Step 3: Preparation of compound 202
向化合物202-2(250mg，0.44mmol，1.0eq.)及化合物SM30(176mg，0.44mmol，1.0eq.)在ACN(10mL)中之溶液中加入K2CO3(181mg，1.31mmol，3.0eq.)、Cs2CO3(43mg，0.13mmol，0.3eq.)及NaI(20mg，0.13mmol，0.3eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之標題化合物(25mg，6%產率)。To a solution of compound 202-2 (250 mg, 0.44 mmol, 1.0 eq.) and compound SM30 (176 mg, 0.44 mmol, 1.0 eq.) in ACN (10 mL) were added K2 CO3 (181 mg, 1.31 mmol, 3.0 eq.), Cs2 CO3 (43 mg, 0.13 mmol, 0.3 eq.) and NaI (20 mg, 0.13 mmol, 0.3 eq.). The mixture was stirred at 80 °C for 10 h. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The mixture was concentrated and purified by preparative HPLC to give the title compound (25 mg, 6% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.67(m,78H),2.16-2.20(m,2H),2.47-2.59(m,10H),3.16-3.19(m,2H),3.39-3.41(m,2H),3.53-3.57(m,4H),4.44-4.46(m,1H)。LCMS：Rt：1.770min；MS m/z(ESI)：878.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.67(m,78H),2.16-2.20(m,2H),2.47-2.59(m,10 H),3.16-3.19(m,2H),3.39-3.41(m,2H),3.53-3.57(m,4H),4.44-4.46(m,1H). LCMS: Rt: 1.770min; MS m/z (ESI): 878.8[M+H]+ .
6.70 實例70：製備化合物216.6.70 Example 70: Preparation of Compound 216.
步驟1：製備化合物216-2Step 1: Preparation of compound 216-2
向化合物216-1(3g，12.6mmol，1.0eq.)及化合物SM22(3g，11.3mmol，0.9eq.)在DCM(60mL)中之溶液中加入EDCI(3.6g，18.9mmol，1.5eq.)、DMAP(0.46g，3.78mmol，0.3eq.)、DIEA(4.9g，37.8mmol，3.0eq.)。將混合物在室溫下攪拌16小時。TLC顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法純化，得到呈黃色油狀之化合物216-2(2.5g，40.43%產率)。EDCI (3.6 g, 18.9 mmol, 1.5 eq.), DMAP (0.46 g, 3.78 mmol, 0.3 eq.), and DIEA (4.9 g, 37.8 mmol, 3.0 eq.) were added to a solution of compound 216-1 (3 g, 12.6 mmol, 1.0 eq.) and compound SM22 (3 g, 11.3 mmol, 0.9 eq.) in DCM (60 mL). The mixture was stirred at room temperature for 16 hours. TLC showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography to obtain compound 216-2 (2.5 g, 40.43% yield) as a yellow oil.
步驟2：製備化合物216-3Step 2: Preparation of compound 216-3
向化合物216-2(2.5g，5.09mmol，1.0eq.)及C10H21COOH(1.1g，6.11mmol，1.2eq.)在DCM(40mL)中之溶液中加入EDCI(1.5g，7.64mmol，1.5eq.)、DMAP(0.2g，1.53mmol，0.3eq.)、DIEA(2g，15.3mmol，3.0eq.)。將混合物在55℃下攪拌16小時。TLC顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法純化，得到呈無色油狀之化合物216-3(2.5g，74.48%產率)。To a solution of compound 216-2 (2.5 g, 5.09 mmol, 1.0 eq.) and C10 H21 COOH (1.1 g, 6.11 mmol, 1.2 eq.) in DCM (40 mL) were added EDCI (1.5 g, 7.64 mmol, 1.5 eq.), DMAP (0.2 g, 1.53 mmol, 0.3 eq.), DIEA (2 g, 15.3 mmol, 3.0 eq.). The mixture was stirred at 55° C. for 16 hours. TLC showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography to give compound 216-3 (2.5 g, 74.48% yield) as a colorless oil.
步驟3：製備化合物216-4Step 3: Preparation of compound 216-4
向化合物216-3(2.5g，3.79mmol，1.0eq.)在EA(50mL)中之溶液中加入Pd/C(300mg)。將混合物在50℃、H2下攪拌16小時。將混合物藉由矽藻土墊過濾並用EA洗滌。將濾液濃縮並藉由矽膠管柱層析法(PE/EA=3/1)純化，得到呈黃色油狀之化合物216-4(2g，92.81%產率)。Pd/C (300 mg) was added to a solution of compound 216-3 (2.5 g, 3.79 mmol, 1.0 eq.) in EA (50 mL). The mixture was stirred at 50 °C underH2 for 16 h. The mixture was filtered through a celite pad and washed with EA. The filtrate was concentrated and purified by silica gel column chromatography (PE/EA = 3/1) to give compound 216-4 (2 g, 92.81% yield) as a yellow oil.
步驟4：製備化合物216-5Step 4: Preparation of compound 216-5
在0℃下，向化合物216-4(300mg，0.53mmol，1.0eq.)及DIPEA(210mg，1.59mmol，3.0eq.)在DCM(10mL)中之溶液中加入MsCl(91mg，0.79mmol，1.5eq.)。將混合物再攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物216-5(350mg，粗品)。To a solution of compound 216-4 (300 mg, 0.53 mmol, 1.0 eq.) and DIPEA (210 mg, 1.59 mmol, 3.0 eq.) in DCM (10 mL) at 0°C was added MsCl (91 mg, 0.79 mmol, 1.5 eq.). The mixture was stirred for another 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated to give compound 216-5 (350 mg, crude) as a yellow oil.
步驟5：製備化合物216Step 5: Preparation of compound 216
向化合物216-5(350mg，0.53mmol，1.0eq.)及化合物K(250mg，1.6mmol，3.0eq.)在ACN(10mL)中之溶液中加入K2CO3(220mg，1.6mmol，3.0eq.)、Cs2CO3(50mg，0.16mmol，0.3eq.)及NaI(20mg，0.16mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物216(40mg，10.66%)。To a solution of compound 216-5 (350 mg, 0.53 mmol, 1.0 eq.) and compound K (250 mg, 1.6 mmol, 3.0 eq.) in ACN (10 mL) were added K2 CO3 (220 mg, 1.6 mmol, 3.0 eq.), Cs2 CO3 (50 mg, 0.16 mmol, 0.3 eq.) and NaI (20 mg, 0.16 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by preparative HPLC to give compound 216 (40 mg, 10.66%) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,9H),1.26-1.44(m,48H),1.53-1.83(m,12H),1.95-1.99(m,1H),2.23-2.32(m,4H),2.39-2.43(m,2H),2.56-2.63(m,3H),3.46-3.48(m,2H),4.00-4.10(m,4H)。LCMS：Rt：0.093min；MS m/z(ESI)：708.5[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,9H),1.26-1.44(m,48H),1.53-1.83(m,12H),1.95-1.99(m,1H),2.23- 2.32(m,4H),2.39-2.43(m,2H),2.56-2.63(m,3H),3.46-3.48(m,2H),4.00-4.10(m,4H). LCMS: Rt: 0.093min; MS m/z (ESI): 708.5[M+H]+ .
以下化合物係以與化合物216類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 216 using the corresponding starting materials.
6.71 實例71：製備化合物218.6.71 Example 71: Preparation of Compound 218.
步驟1：製備化合物76-1Step 1: Preparation of compound 76-1
向化合物SM31(2g，7.42mmol，1.0eq.)及化合物W(1.7g，8.91mmol，1.2eq.)在DCM(40mL)中之溶液中加入DIEA(1.9g，14.8mmol，2.0eq.)及HATU(3.4g，8.91mmol，1.2eq.)。將混合物在室溫下攪拌2小時。TLC顯示反應完成。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥，濃縮並藉由矽膠管柱層析法純化，得到呈黃色油狀之化合物76-1(3.0g，90.52%產率)。To a solution of compound SM31 (2 g, 7.42 mmol, 1.0 eq.) and compound W (1.7 g, 8.91 mmol, 1.2 eq.) in DCM (40 mL) were added DIEA (1.9 g, 14.8 mmol, 2.0 eq.) and HATU (3.4 g, 8.91 mmol, 1.2 eq.). The mixture was stirred at room temperature for 2 hours. TLC showed that the reaction was complete. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 , concentrated and purified by silica gel column chromatography to give compound 76-1 (3.0 g, 90.52% yield) as a yellow oil.
步驟2：製備化合物218-1Step 2: Preparation of compound 218-1
向化合物76-1(800mg，1.79mmol，1.0eq.)及化合物SM32(270mg，1.97mmol，1.1eq.)在ACN(20mL)中之溶液中加入K2CO3(740mg，5.37mmol，3.0eq.)、Cs2CO3(180mg，0.54mmol，0.3eq.)及NaI(27mg，0.18mmol，0.1eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮，進行FCC，得到呈黃色油狀之化合物218-1(450mg，54.06%)。To a solution of compound 76-1 (800 mg, 1.79 mmol, 1.0 eq.) and compound SM32 (270 mg, 1.97 mmol, 1.1 eq.) in ACN (20 mL) were added K2 CO3 (740 mg, 5.37 mmol, 3.0 eq.), Cs2 CO3 (180 mg, 0.54 mmol, 0.3 eq.) and NaI (27 mg, 0.18 mmol, 0.1 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and subjected to FCC to give compound 218-1 (450 mg, 54.06%) as a yellow oil.
步驟3：製備化合物218Step 3: Preparation of compound 218
向化合物218-1(220mg，0.47mmol，1.0eq.)及化合物SM16(210mg，0.47mmol，1.0eq.)在DCE(10mL)中之溶液中加入AcOH(1滴)。將混合物在室溫下攪拌16小時。接著加入NaBH(OAc)3(300mg，1.42mmol，3.0eq.)。將混合物在室溫下攪拌24小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之化合物218(50mg，11.84%產率)。To a solution of compound 218-1 (220 mg, 0.47 mmol, 1.0 eq.) and compound SM16 (210 mg, 0.47 mmol, 1.0 eq.) in DCE (10 mL) was added AcOH (1 drop). The mixture was stirred at room temperature for 16 hours. Then NaBH(OAc)3 (300 mg, 1.42 mmol, 3.0 eq.) was added. The mixture was stirred at room temperature for 24 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by preparative HPLC to give compound 218 (50 mg, 11.84% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.34(m,50H),1.41-1.52(m,5H),1.59-1.67(m,10H),1.85-2.00(m,4H),2.15-2.18(m,2H),2.28-2.32(m,6H),2.45-2.49(m,3H),2.61-2.64(m,2H),2.98(d,J=11.2Hz,2H),3.16-3.19(m,2H),3.46-3.49(m,2H),4.00-4.09(m,4H),5.33-5.36(m,1H)。LCMS：Rt：0.093min；MS m/z(ESI)：892.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.34(m,50H),1.41-1.52(m,5H),1.59-1.67(m,10H),1.85-2.0 0(m,4H),2.15-2.18(m,2H),2.28-2.32(m,6H),2.45-2.49(m,3H),2.61-2.64(m,2H),2.98(d,J =11.2Hz,2H),3.16-3.19(m,2H),3.46-3.49(m,2H),4.00-4.09(m,4H),5.33-5.36(m,1H). LCMS: Rt: 0.093min; MS m/z (ESI): 892.7[M+H]+ .
6.72 實例72：製備化合物223.6.72 Example 72: Preparation of Compound 223.
步驟1：製備化合物223-2Step 1: Preparation of compound 223-2
向化合物223-1(7.0g，23.0mmol，1.0eq.)在MeOH/H2O(50mL/50mL)中之溶液中加入NaOH(7.6g，184.0mmol，8.0eq.)。將混合物在25℃下攪拌10小時。TLC顯示反應完成。將反應混合物濃縮以除去有機層。將水層用2N HCl調節pH至5，接著用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈無色油狀之化合物223-2(4.6g，72%產率)。To a solution of compound 223-1 (7.0 g, 23.0 mmol, 1.0 eq.) in MeOH/H2 O (50 mL/50 mL) was added NaOH (7.6 g, 184.0 mmol, 8.0 eq.). The mixture was stirred at 25° C. for 10 hours. TLC showed that the reaction was complete. The reaction mixture was concentrated to remove the organic layer. The aqueous layer was adjusted to pH 5 with 2N HCl, followed by extraction with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 223-2 (4.6 g, 72% yield) as a colorless oil.
步驟2：製備化合物223-3Step 2: Preparation of compound 223-3
向化合物223-2(4.6g，17.0mmol，1.0eq.)及化合物SM33(5.4g，42.0mmol，2.5eq.)在甲苯(60mL)中之溶液中加入TsOH(0.32g，1.7mmol，0.1eq.)。將混合物藉由Dean-S-tark分水器回流攪拌3小時。TLC顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法純化殘餘物，得到呈無色油狀之化合物223-3(6.3g，74%產率)。To a solution of compound 223-2 (4.6 g, 17.0 mmol, 1.0 eq.) and compound SM33 (5.4 g, 42.0 mmol, 2.5 eq.) in toluene (60 mL) was added TsOH (0.32 g, 1.7 mmol, 0.1 eq.). The mixture was stirred at reflux for 3 hours through a Dean-S-tark water separator. TLC showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography to give compound 223-3 (6.3 g, 74% yield) as a colorless oil.
步驟3：製備化合物223-4Step 3: Preparation of compound 223-4
向化合物223-3(2.4g，4.8mmol，1.0eq.)在EA(25mL)中之溶液中加入Pd/C(0.2g)及兩滴濃鹽酸。將混合物在25℃、H2下攪拌5小時。TLC顯示反應完成。將反應混合物藉由矽藻土墊過濾並用EA洗滌。將濾液濃縮，得到呈無色油狀之化合物223-4(2g，92%產率)。To a solution of compound 223-3 (2.4 g, 4.8 mmol, 1.0 eq.) in EA (25 mL) was added Pd/C (0.2 g) and two drops of concentrated hydrochloric acid. The mixture was stirred at 25 °C underH2 for 5 h. TLC showed that the reaction was complete. The reaction mixture was filtered through a celite pad and washed with EA. The filtrate was concentrated to give compound 223-4 (2 g, 92% yield) as a colorless oil.
步驟4：製備化合物223-5Step 4: Preparation of compound 223-5
向化合物223-4(2.0g，5.0mmol，1.0eq.)及TEA(1.0g，10.0mmol，2.0eq.)在DCM(50mL)中之溶液中加入MsCl(687mg，6.0mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物223-5(12.4g，100%產率)。To a solution of compound 223-4 (2.0 g, 5.0 mmol, 1.0 eq.) and TEA (1.0 g, 10.0 mmol, 2.0 eq.) in DCM (50 mL) was added MsCl (687 mg, 6.0 mmol, 1.2 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 223-5 (12.4 g, 100% yield) as a yellow oil.
步驟5：製備化合物223-6Step 5: Preparation of compound 223-6
向化合物223-5(300mg，0.63mmol，1.0eq.)及化合物D(145mg，126mmol，2.0eq.)在ACN(12mL)中之溶液中加入K2CO3(261mg，1.89mmol，3.0eq.)、Cs2CO3(62mg，0.19mmol，0.3eq.)及NaI(28mg，0.19mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=25/1)純化殘餘物，得到呈黃色油狀之化合物223-6(126mg)，其藉由製備型HPLC進一步純化，得到呈黃色油狀之化合物223-6(65mg，21%產率)。1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,6H),1.27-1.32(m,22H),1.45-1.74(m,9H),1.87-2.10(m,6H),2.44-2.65(m,4H),3.14-3.19(m,1H),3.24-3.31(m,1H),3.54-3.61(m,2H),4.08-4.17(m,4H)。LCMS：Rt：0.860min；MS m/z(ESI)：498.5[M+H]+。To a solution of compound 223-5 (300 mg, 0.63 mmol, 1.0 eq.) and compound D (145 mg, 126 mmol, 2.0 eq.) in ACN (12 mL) were added K2 CO3 (261 mg, 1.89 mmol, 3.0 eq.), Cs2 CO3 (62 mg, 0.19 mmol, 0.3 eq.) and NaI (28 mg, 0.19 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=25/1) to give compound 223-6 (126 mg) as a yellow oil, which was further purified by preparative HPLC to give compound 223-6 (65 mg, 21% yield) as a yellow oil.1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,6H),1.27-1.32(m,22H),1.45-1.74(m,9H),1.87-2.10(m,6H),2.44-2 .65(m,4H),3.14-3.19(m,1H),3.24-3.31(m,1H),3.54-3.61(m,2H),4.08-4.17(m,4H). LCMS: Rt: 0.860min; MS m/z (ESI): 498.5[M+H]+ .
步驟6：製備化合物223-7Step 6: Preparation of compound 223-7
向化合物223-6(1.0g，2.0mmol，1.0eq.)及DIPEA(517mg，4.0mmol，2.0eq.)在DCM(20mL)中之溶液中加入MsCl(275mg，2.4mmol，1.2eq.)。將混合物在0℃下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物223-7(1.1g，95%產率)。To a solution of compound 223-6 (1.0 g, 2.0 mmol, 1.0 eq.) and DIPEA (517 mg, 4.0 mmol, 2.0 eq.) in DCM (20 mL) was added MsCl (275 mg, 2.4 mmol, 1.2 eq.). The mixture was stirred at 0 °C for 2 h. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 223-7 (1.1 g, 95% yield) as a yellow oil.
步驟7：製備化合物223Step 7: Preparation of compound 223
向化合物223-7(1.0g，1.74mmol，1.0eq.)及化合物SM39(694mg，1.74mmol，1.0eq.)在ACN(30mL)中之溶液中加入K2CO3(721mg，5.22mmol，3.0eq.)、Cs2CO3(169mg，0.52mmol，0.3eq.)及NaI(78mg，0.52mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=25/1)純化殘餘物，得到呈黃色油狀之化合物223(520mg，35%產率)。將100mg產物藉由製備型HPLC進一步純化，得到呈黃色油狀之化合物223(45mg，45%產率)。To a solution of compound 223-7 (1.0 g, 1.74 mmol, 1.0 eq.) and compound SM39 (694 mg, 1.74 mmol, 1.0 eq.) in ACN (30 mL) were added K2 CO3 (721 mg, 5.22 mmol, 3.0 eq.), Cs2 CO3 (169 mg, 0.52 mmol, 0.3 eq.) and NaI (78 mg, 0.52 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=25/1) to give compound 223 (520 mg, 35% yield) as a yellow oil. 100 mg of the product was further purified by preparative HPLC to give compound 223 (45 mg, 45% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.30(m,49H),1.44-1.68(m,12H),1.87-2.14(m,5H),2.29-3.00(m,15H),3.30-3.33(m,1H),3.53-3.69(m,2H),3.96-3.97(m,2H),4.11-4.17(m,4H)。LCMS：Rt：1.620min；MS m/z(ESI)：879.7[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.30(m,49H),1.44-1.68(m,12H),1.87-2.14(m,5H),2.29- 3.00(m,15H),3.30-3.33(m,1H),3.53-3.69(m,2H),3.96-3.97(m,2H),4.11-4.17(m,4H). LCMS: Rt: 1.620min; MS m/z (ESI): 879.7[M+H]+ .
6.73 實例73：製備化合物238.6.73 Example 73: Preparation of Compound 238.
步驟1：製備化合物238-1Step 1: Preparation of compound 238-1
向化合物108-1(3.53g，12.59mmol，1.0eq.)及化合物216-1(1.0g，4.20mmol，0.3eq.)在DCM(50mL)中之溶液中加入DIEA(2.71g，20.98mmol，1.7eq.)、EDCI(2.41g，12.59mmol，1.0eq.)及DMAP(0.52g，4.20mmol，0.3eq.)。將混合物在45℃下攪拌10小時。將反應混合物濃縮並藉由矽膠管柱層析法(PE/EA=10：1)純化，得到呈無色油狀之化合物238-1(2.0g，62%產率)。DIEA (2.71 g, 20.98 mmol, 1.7 eq.), EDCI (2.41 g, 12.59 mmol, 1.0 eq.) and DMAP (0.52 g, 4.20 mmol, 0.3 eq.) were added to a solution of compound 108-1 (3.53 g, 12.59 mmol, 1.0 eq.) and compound 216-1 (1.0 g, 4.20 mmol, 0.3 eq.) in DCM (50 mL). The mixture was stirred at 45 °C for 10 hours. The reaction mixture was concentrated and purified by silica gel column chromatography (PE/EA=10:1) to obtain compound 238-1 (2.0 g, 62% yield) as a colorless oil.
步驟2：製備化合物238-2Step 2: Preparation of compound 238-2
在-78℃下，向化合物238-1(1.0g，1.31mmol，1.0eq.)在DCM(20mL)中之溶液中加入BCl3(15.6mL，15.6mmol，12.0eq.)。將混合物在-78℃下攪拌1小時。將混合物倒入NaHCO3水溶液中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥，濃縮並藉由矽膠管柱層析法(PE/EA=4：1)純化，得到呈無色油狀之化合物238-2(0.5g，57%產率)。To a solution of compound 238-1 (1.0 g, 1.31 mmol, 1.0 eq.) in DCM (20 mL) was added BCl3 (15.6 mL, 15.6 mmol, 12.0 eq.) at -78°C. The mixture was stirred at -78°C for 1 hour. The mixture was poured into an aqueous NaHCO3 solution and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 , concentrated and purified by silica gel column chromatography (PE/EA=4:1) to give compound 238-2 (0.5 g, 57% yield) as a colorless oil.
步驟3：製備化合物238-3Step 3: Preparation of compound 238-3
向化合物238-2(800mg，1.19mmol，1.0eq.)及DIPEA(310mg，2.38mmol，2.0eq.)在DCM(20mL)中之溶液中加入MsCl(165mg，1.43mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物238-3(600mg，67%產率)，其未經進一步純化即用於下一步驟。To a solution of compound 238-2 (800 mg, 1.19 mmol, 1.0 eq.) and DIPEA (310 mg, 2.38 mmol, 2.0 eq.) in DCM (20 mL) was added MsCl (165 mg, 1.43 mmol, 1.2 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 238-3 (600 mg, 67% yield) as a yellow oil, which was used in the next step without further purification.
步驟4：製備化合物238-4Step 4: Preparation of compound 238-4
向化合物238-3(600mg，0.8mmol，1.0eq.)及化合物B(230mg，1.6mmol，2.0eq.)在ACN(30mL)中之溶液中加入K2CO3(332mg，2.4mmol，3.0eq.)、Cs2CO3(79mg，0.24mmol，0.3eq.)及NaI(36mg，0.24mmol，0.2eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=20/1)純化，得到呈黃色油狀之化合物238-4(500mg，78%產率)。LCMS：Rt：1.380min；MS m/z(ESI)：798.6[M+H]+。To a solution of compound 238-3 (600 mg, 0.8 mmol, 1.0 eq.) and compound B (230 mg, 1.6 mmol, 2.0 eq.) in ACN (30 mL) were added K2 CO3 (332 mg, 2.4 mmol, 3.0 eq.), Cs2 CO3 (79 mg, 0.24 mmol, 0.3 eq.) and NaI (36 mg, 0.24 mmol, 0.2 eq.). The mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=20/1) to give compound 238-4 (500 mg, 78% yield) as a yellow oil. LCMS: Rt: 1.380min; MS m/z (ESI): 798.6[M+H]+ .
步驟5：製備化合物238-5Step 5: Preparation of compound 238-5
向化合物238-4(250mg，0.31mmol，1.0eq.)及DIPEA(122mg，0.94mmol，3.0eq.)在DCM(20mL)中之溶液中加入MsCl(44mg，0.38mmol，1.0eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物238-5(220mg，80%產率)，其未經進一步純化即用於下一步驟。To a solution of compound 238-4 (250 mg, 0.31 mmol, 1.0 eq.) and DIPEA (122 mg, 0.94 mmol, 3.0 eq.) in DCM (20 mL) was added MsCl (44 mg, 0.38 mmol, 1.0 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organiclayers were washed with brine, dried overNa2SO4 and concentrated to give compound 238-5 (220 mg, 80% yield) as a yellow oil, which was used in the next step without further purification.
步驟6：製備化合物238Step 6: Preparation of compound 238
向化合物238-5(200mg，0.23mmol，1.0eq.)及化合物SM34(91mg，0.23mmol，1.0eq.)在ACN(10mL)中之溶液中加入K2CO3(95mg，0.69mmol，3.0eq.)、Cs2CO3(23mg，0.07mmol，0.3eq.)及NaI(11mg，0.07mmol，0.3eq.)。將混合物在80℃下攪拌10小時。LCMS顯示反應完成。將反應混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物238(11mg，4%產率)。To a solution of compound 238-5 (200 mg, 0.23 mmol, 1.0 eq.) and compound SM34 (91 mg, 0.23 mmol, 1.0 eq.) in ACN (10 mL) were added K2 CO3 (95 mg, 0.69 mmol, 3.0 eq.), Cs2 CO3 (23 mg, 0.07 mmol, 0.3 eq.) and NaI (11 mg, 0.07 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 10 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by preparative HPLC to give compound 238 (11 mg, 4% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.79-0.84(m,12H),1.19-1.74(m,81H),1.95-2.72(m,30H),3.09-3.13(m,2H),3.13-3.50(m,2H),3.92-4.09(m,4H),5.27-5.30(m,8H)。LCMS：Rt：0.627min；MS m/z(ESI)：1179.0[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.79-0.84(m,12H),1.19-1.74(m,81H),1.95-2.72(m,30H),3.09-3.13(m,2H),3.13-3.50(m,2H),3.92-4.09(m,4H),5.27-5.30(m,8H). LCMS: Rt: 0.627min; MS m/z (ESI): 1179.0[M+H]+ .
6.74 實例74：製備化合物239.6.74 Example 74: Preparation of Compound 239.
步驟1：製備化合物239-1Step 1: Preparation of compound 239-1
向化合物216-5(820mg，1.27mmol，1.0eq.)及化合物B(273mg，1.91mmol，1.5eq.)在ACN(25mL)中之溶液中加入K2CO3(527mg，3.81mmol，3.0eq.)、Cs2CO3(124mg，0.38mmol，0.3eq.)及NaI(57mg，0.38mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=50/1)純化殘餘物，得到呈黃色油狀之化合物239-1(290mg，33%產率)。LCMS：Rt：1.420min；MS m/z(ESI)：694.6[M+H]+。To a solution of compound 216-5 (820 mg, 1.27 mmol, 1.0 eq.) and compound B (273 mg, 1.91 mmol, 1.5 eq.) in ACN (25 mL) were added K2 CO3 (527 mg, 3.81 mmol, 3.0 eq.), Cs2 CO3 (124 mg, 0.38 mmol, 0.3 eq.) and NaI (57 mg, 0.38 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 h. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=50/1) to obtain compound 239-1 (290 mg, 33% yield) as a yellow oil. LCMS: Rt: 1.420 min; MS m/z (ESI): 694.6 [M+H]+ .
步驟2：製備化合物239-2Step 2: Preparation of compound 239-2
向化合物239-1(290mg，0.42mmol，1.0eq.)在DCM(10mL)中之溶液中加入SOCl2(150mg，1.26mmol，3.0eq.)。將混合物在30℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮，得到呈黃色油狀之化合物239-2(298mg，100%產率)，其未經進一步純化即用於下一步驟。LCMS：Rt：1.700min；MS m/z(ESI)：712.6[M+H]+。To a solution of compound 239-1 (290 mg, 0.42 mmol, 1.0 eq.) in DCM (10 mL) was added SOCl2 (150 mg, 1.26 mmol, 3.0 eq.). The mixture was stirred at 30° C. for 16 h. LCMS showed the reaction was complete. The mixture was concentrated to give compound 239-2 (298 mg, 100% yield) as a yellow oil, which was used in the next step without further purification. LCMS: Rt: 1.700 min; MS m/z (ESI): 712.6 [M+H]+ .
步驟3：裂備化合物239Step 3: Preparation of Compound 239
向化合物239-2(270mg，0.38mmol，1.0eq.)及化合物SM34(151mg，0.38mmol，1.0eq.)在THF(10mL)中之溶液中加入DIPEA(147mg，1.14mmol，3.0eq.)及NaI(17mg，0.114mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物239(80mg，20%產率)。To a solution of compound 239-2 (270 mg, 0.38 mmol, 1.0 eq.) and compound SM34 (151 mg, 0.38 mmol, 1.0 eq.) in THF (10 mL) was added DIPEA (147 mg, 1.14 mmol, 3.0 eq.) and NaI (17 mg, 0.114 mmol, 0.3 eq.). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 239 (80 mg, 20% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,15H),1.26-1.36(m,76H),1.37-1.70(m,10H),1.74-1.94(m,4H),1.97-2.00(m,1H),2.11-2.32(m,7H),2.37-2.71(m,8H),2.93-3.07(m,2H),3.16-3.19(m,2H),3.50-3.64(m,2H),3.99-4.10(m,4H)。LCMS：Rt：0.507min；MS m/z(ESI)：1074.9[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,15H),1.26-1.36(m,76H),1.37-1.70(m,10H),1.74-1.94(m,4H),1.97-2.00(m,1H),2.11- 2.32(m,7H),2.37-2.71(m,8H),2.93-3.07(m,2H),3.16-3.19(m,2H),3.50-3.64(m,2H),3.99-4.10(m,4H). LCMS: Rt: 0.507min; MS m/z (ESI): 1074.9[M+H]+ .
6.75 實例75：製備化合物241.6.75 Example 75: Preparation of Compound 241.
步驟1：製備化合物241-1Step 1: Preparation of compound 241-1
向化合物SM24(2.0g，4.8mmol，1.0eq.)及化合物SM35(709mg，12.0mmol，2.5eq.)在ACN(50mL)中之溶液中加入K2CO3(2.0g，14.4mmol，3.0eq.)、Cs2CO3(469mg，1.44mmol，0.3eq.)及NaI(216mg，1.44mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=25/1)純化，得到呈黃色油狀之化合物241-1(830mg，44%產率)。LCMS：Rt：0.850min；MS m/z(ESI)：398.5[M+H]+。To a solution of compound SM24 (2.0 g, 4.8 mmol, 1.0 eq.) and compound SM35 (709 mg, 12.0 mmol, 2.5 eq.) in ACN (50 mL) were added K2 CO3 (2.0 g, 14.4 mmol, 3.0 eq.), Cs2 CO3 (469 mg, 1.44 mmol, 0.3 eq.) and NaI (216 mg, 1.44 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=25/1) to give compound 241-1 (830 mg, 44% yield) as a yellow oil. LCMS: Rt: 0.850min; MS m/z (ESI): 398.5[M+H]+ .
步驟2：製備化合物241-2Step 2: Preparation of compound 241-2
向化合物241-1(400mg，1.0mmol，1.0eq.)及化合物SM36(157mg，1.0mmol，1.0eq.)在DCE(10mL)中之溶液中加入AcOH(1滴)。將混合物在室溫下攪拌4小時。接著加入NaBH3CN(94mg，1.5mmol，1.5eq.)。將混合物在室溫下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=30/1)純化，得到呈黃色油狀之化合物241-2(282mg，52%產率)。LCMS：Rt：0.467min；MS m/z(ESI)：538.5[M+H]+。To a solution of compound 241-1 (400 mg, 1.0 mmol, 1.0 eq.) and compound SM36 (157 mg, 1.0 mmol, 1.0 eq.) in DCE (10 mL) was added AcOH (1 drop). The mixture was stirred at room temperature for 4 hours. Then NaBH3 CN (94 mg, 1.5 mmol, 1.5 eq.) was added. The mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=30/1) to give compound 241-2 (282 mg, 52% yield) as a yellow oil. LCMS: Rt: 0.467 min; MS m/z (ESI): 538.5 [M+H]+ .
步驟3：製備化合物241-3Step 3: Preparation of compound 241-3
向化合物241-2(250mg，0.46mmol，1.0eq.)在DCM(9mL)中之溶液中加入TFA(3.0mL)。將混合物在室溫下攪拌24小時。LCMS顯示反應完成。將混合物用飽和NaHCO3溶液調節至pH=8，接著用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈棕色油狀之化合物241-3(223mg，97%產率)。LCMS：Rt：0.920min；MS m/z(ESI)：494.5[M+H]+。To a solution of compound 241-2 (250 mg, 0.46 mmol, 1.0 eq.) in DCM (9 mL) was added TFA (3.0 mL). The mixture was stirred at room temperature for 24 hours. LCMS showed that the reaction was complete. The mixture was adjusted to pH = 8 with saturated NaHCO3 solution, and then extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 241-3 (223 mg, 97% yield) as a brown oil. LCMS: Rt: 0.920 min; MS m/z (ESI): 494.5 [M+H]+ .
步驟4：製備化合物241-4Step 4: Preparation of compound 241-4
向化合物241-3(230mg，0.47mmol，1.0eq.)及化合物SM6(57mg，0.94mmol，2.0eq.)在DCE(10mL)中之溶液中加入AcOH(1滴)。將混合物在室溫下攪拌4小時。接著加入NaBH3CN(44mg，0.71mmol，1.5eq.)。將混合物在室溫下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。將殘餘物濃縮並藉由矽膠管柱層析法(DCM/MeOH=10/1)純化，得到呈黃色油狀之化合物241-4(144mg，57%產率)。LCMS：Rt：0.160min；MS m/z(ESI)：539.5[M+H]+。To a solution of compound 241-3 (230 mg, 0.47 mmol, 1.0 eq.) and compound SM6 (57 mg, 0.94 mmol, 2.0 eq.) in DCE (10 mL) was added AcOH (1 drop). The mixture was stirred at room temperature for 4 hours. Then NaBH3 CN (44 mg, 0.71 mmol, 1.5 eq.) was added. The mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was concentrated and purified by silica gel column chromatography (DCM/MeOH=10/1) to obtain compound 241-4 (144 mg, 57% yield) as a yellow oil. LCMS: Rt: 0.160 min; MS m/z (ESI): 539.5 [M+H]+ .
步驟5：製備化合物241Step 5: Preparation of compound 241
向化合物241-4(144mg，0.27mmol，1.0eq.)及化合物SM24(227mg，0.54mmol，2.0eq.)在ACN(10mL)中之溶液中加入K2CO3(112mg，0.81mmol，3.0eq.)、Cs2CO3(26mg，0.081mmol，0.3eq.)及NaI(12mg，0.081mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將混合物濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物241(51mg，22%產率)。To a solution of compound 241-4 (144 mg, 0.27 mmol, 1.0 eq.) and compound SM24 (227 mg, 0.54 mmol, 2.0 eq.) in ACN (10 mL) were added K2 CO3 (112 mg, 0.81 mmol, 3.0 eq.), Cs2 CO3 (26 mg, 0.081 mmol, 0.3 eq.) and NaI (12 mg, 0.081 mmol, 0.3 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was concentrated and purified by preparative HPLC to give compound 241 (51 mg, 22% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,15H),1.27(s,56H),1.38-1.41(m,6H),1.51-1.63(m,7H),1.74-1.91(m,4H),2.29-2.71(m,14H),3.43-3.52(m,2H),3.96-3.97(m,4H)。LCMS：Rt：0.400min；MS m/z(ESI)：877.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,15H),1.27(s,56H),1.38-1.41(m,6H),1.51-1.63(m,7H), 1.74-1.91(m,4H),2.29-2.71(m,14H),3.43-3.52(m,2H),3.96-3.97(m,4H). LCMS: Rt: 0.400min; MS m/z (ESI): 877.8[M+H]+ .
6.76 實例76：製備化合物244.6.76 Example 76: Preparation of Compound 244.
步驟1：製備化合物244-2Step 1: Preparation of compound 244-2
將244-1(4.0g，27.7mmol，1.0eq.)、SOCl2(9.9g，83.2mmol，3.0eq.)、吡啶(6.6g，83.2mmol，3.0eq.)在DCM(50mL)中之混合物回流攪拌4小時。TLC顯示反應完成。將混合物用DCM稀釋並用水及鹽水洗滌，乾燥並濃縮。藉由FCC純化殘餘物，得到呈無色油狀之化合物244-2(4.5g，89.7%產率)。A mixture of 244-1 (4.0 g, 27.7 mmol, 1.0 eq.), SOCl2 (9.9 g, 83.2 mmol, 3.0 eq.), pyridine (6.6 g, 83.2 mmol, 3.0 eq.) in DCM (50 mL) was stirred at reflux for 4 hours. TLC showed that the reaction was complete. The mixture was diluted with DCM and washed with water and brine, dried and concentrated. The residue was purified by FCC to give compound 244-2 (4.5 g, 89.7% yield) as a colorless oil.
步驟2：製備化合物244-3Step 2: Preparation of compound 244-3
將244-2(612mg，3.38mmol，1.0eq.)、SM16(500mg，1.13mmol，3.0eq.)、K2CO3(466mg，3.38mmol，3.0eq.)、Cs2CO3(111mg，0.34mmol，3.0eq.)、NaI(51mg，0.34mmol，3.0eq.)在ACN(10mL)中之混合物回流攪拌隔夜。LCMS顯示產物。將混合物用EA稀釋並用水及鹽水洗滌，乾燥並濃縮。藉由FCC純化殘餘物，得到呈無色油狀之化合物244-3(260mg，39.1%產率)。A mixture of 244-2 (612 mg, 3.38 mmol, 1.0 eq.), SM16 (500 mg, 1.13 mmol, 3.0 eq.), K2 CO3 (466 mg, 3.38 mmol, 3.0 eq.), Cs2 CO3 (111 mg, 0.34 mmol, 3.0 eq.), NaI (51 mg, 0.34 mmol, 3.0 eq.) in ACN (10 mL) was stirred under reflux overnight. LCMS showed the product. The mixture was diluted with EA and washed with water and brine, dried and concentrated. The residue was purified by FCC to give compound 244-3 (260 mg, 39.1% yield) as a colorless oil.
步驟3：製備化合物244-4Step 3: Preparation of compound 244-4
將244-3(260mg，0.44mmol，1.0eq.)、SM38(234mg，0.53mmol，1.2eq.)、K2CO3(184mg，1.33mmol，3.0eq.)、Cs2CO3(42mg，0.13mmol，3.0eq.)、NaI(20mg，0.13mmol，3.0eq.)在ACN(5mL)中之混合物回流攪拌隔夜。LCMS顯示產物。將混合物用EA稀釋並用水及鹽水洗滌，乾燥並濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之化合物244(38mg，8.7%產率)。A mixture of 244-3 (260 mg, 0.44 mmol, 1.0 eq.), SM38 (234 mg, 0.53 mmol, 1.2 eq.), K2 CO3 (184 mg, 1.33 mmol, 3.0 eq.), Cs2 CO3 (42 mg, 0.13 mmol, 3.0 eq.), NaI (20 mg, 0.13 mmol, 3.0 eq.) in ACN (5 mL) was stirred under reflux overnight. LCMS showed the product. The mixture was diluted with EA and washed with water and brine, dried and concentrated. The residue was purified by preparative HPLC to give compound 244 (38 mg, 8.7% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-1.08(m,18H),1.28-1.36(m,52H),1.61-1.63(m,8H),1.80-2.04(m,7H),2.26-2.32(m,11H),2.44-2.76(m,4H)2.84-3.01(m,2H),3.48-3.70(m,2H),3.99-4.11(m,8H)。LCMS：Rt：1.380min；MS m/z(ESI)：993.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-1.08(m,18H),1.28-1.36(m,52H),1.61-1.63(m,8H),1.80-2.04(m,7H),2.26- 2.32(m,11H),2.44-2.76(m,4H)2.84-3.01(m,2H),3.48-3.70(m,2H),3.99-4.11(m,8H). LCMS: Rt: 1.380min; MS m/z (ESI): 993.8[M+H]+ .
6.77 實例77：製備化合物246.6.77 Example 77: Preparation of Compound 246.
步驟1：製備化合物246-2Step 1: Preparation of compound 246-2
向246-1(1.7g，7.3mmol，1.0eq.)及十一碳-10-烯酸(4.0g，22.0mmol，3.0eq.)在DCM(40mL)中之溶液中加入DIEA(4.7g，36.6mmol，5.0eq.)、EDCI(4.2g，22.0mmol，3.0eq.)及DMAP(268mg，2.2mmol，0.3eq.)。將混合物在40℃下攪拌16小時。將反應混合物濃縮並藉由矽膠管柱層析法(PE/EA=10：1)純化，得到呈無色油狀之化合物246-2(1.7g，41.1%產率)。DIEA (4.7 g, 36.6 mmol, 5.0 eq.), EDCI (4.2 g, 22.0 mmol, 3.0 eq.) and DMAP (268 mg, 2.2 mmol, 0.3 eq.) were added to a solution of 246-1 (1.7 g, 7.3 mmol, 1.0 eq.) and undec-10-enoic acid (4.0 g, 22.0 mmol, 3.0 eq.) in DCM (40 mL). The mixture was stirred at 40 °C for 16 hours. The reaction mixture was concentrated and purified by silica gel column chromatography (PE/EA=10:1) to obtain compound 246-2 (1.7 g, 41.1% yield) as a colorless oil.
步驟2：製備化合物246-3Step 2: Preparation of compound 246-3
在0℃下，向246-2(1.7g，3.01mmol，1.0eq.)在DCM(34mL)中之溶液中加入在二
步驟3：製備化合物246-4Step 3: Preparation of compound 246-4
向246-3(869mg，1.8mmol，1.0eq.)及Et3N(350mg，2.7mmol，1.5eq.)在DCM(16mL)中之溶液中加入MsCl(228mg，2.0mmol，0.1eq.)。將混合物在室溫下攪拌1小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈無色油狀之化合物246-4(1.1g，粗品)。其未經進一步純化即用於下一步驟。To a solution of 246-3 (869 mg, 1.8 mmol, 1.0 eq.) and Et3 N (350 mg, 2.7 mmol, 1.5 eq.) in DCM (16 mL) was added MsCl (228 mg, 2.0 mmol, 0.1 eq.). The mixture was stirred at room temperature for 1 hour. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 246-4 (1.1 g, crude) as a colorless oil. It was used in the next step without further purification.
步驟4：製備化合物246-5Step 4: Preparation of compound 246-5
向246-4(1.1g，2.0mmol，1.0eq.)及化合物G(756mg，5.9mmol，3.0eq.)在ACN(30mL)中之溶液中加入K2CO3(815mg，5.9mmol，3.0eq.)、Cs2CO3(19mg，0.06mmol，0.03eq.)及NaI(148mg，1.0mmol，0.5eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由矽膠管柱層析法(DCM/MeOH=30/1)純化，得到呈無色油狀之化合物246-5(899mg，77.2%產率)。LCMS：Rt：1.630min；MS m/z(ESI)：592.5[M+H]+。To a solution of 246-4 (1.1 g, 2.0 mmol, 1.0 eq.) and compound G (756 mg, 5.9 mmol, 3.0 eq.) in ACN (30 mL) were added K2 CO3 (815 mg, 5.9 mmol, 3.0 eq.), Cs2 CO3 (19 mg, 0.06 mmol, 0.03 eq.) and NaI (148 mg, 1.0 mmol, 0.5 eq.). The mixture was stirred at 80° C. for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was concentrated and purified by silica gel column chromatography (DCM/MeOH=30/1) to give compound 246-5 (899 mg, 77.2% yield) as a colorless oil. LCMS: Rt: 1.630min; MS m/z (ESI): 592.5[M+H]+ .
步驟5：製備化合物246-6Step 5: Preparation of compound 246-6
向246-5(300mg，0.5mmol，1.0eq.)及Et3N(98mg，0.8mmol，1.6eq.)在DCM(6mL)中之溶液中加入MsCl(69mg，0.6mmol，1.2eq.)。將混合物在室溫下攪拌1小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈橙色油狀之化合物246-6(362mg，粗品)。其未經進一步純化即用於下一步驟。To a solution of 246-5 (300 mg, 0.5 mmol, 1.0 eq.) and Et3 N (98 mg, 0.8 mmol, 1.6 eq.) in DCM (6 mL) was added MsCl (69 mg, 0.6 mmol, 1.2 eq.). The mixture was stirred at room temperature for 1 hour. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated to give compound 246-6 (362 mg, crude) as an orange oil. It was used in the next step without further purification.
步驟6：製備化合物246Step 6: Preparation of compound 246
向246-6(342mg，0.5mmol，1.0eq.)及SM34(210mg，0.5mmol，1.0eq.)在ACN(10mL)中之溶液中加入K2CO3(211mg，1.5mmol，3.0eq.)、Cs2CO3(5mg，0.02mmol，0.04eq.)及NaI(38mg，0.26mmol，0.52eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物濃縮並藉由製備型HPLC純化，得到呈無色油狀之化合物246(11mg，7.22%產率)。To a solution of 246-6 (342 mg, 0.5 mmol, 1.0 eq.) and SM34 (210 mg, 0.5 mmol, 1.0 eq.) in ACN (10 mL) were added K2 CO3 (211 mg, 1.5 mmol, 3.0 eq.), Cs2 CO3 (5 mg, 0.02 mmol, 0.04 eq.) and NaI (38 mg, 0.26 mmol, 0.52 eq.). The mixture was stirred at 80 °C for 16 h. LCMS showed the reaction was complete. The reaction mixture was concentrated and purified by preparative HPLC to give compound 246 (11 mg, 7.22% yield) as a colorless oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.89(m,6H),1.26-1.39(m,51H),1.49-1.68(m,14H),1.95-2.06(m,8H),2.18-2.32(m,9H),2.52-2.69(m,7H),2.95-3.19(m,6H),3.49-3.67(m,2H),4.00-4.10(m,4H),4.91-5.02(m,4H),5.76-5.86(m,2H)。LCMS：Rt：1.510min；MS m/z(ESI)：972.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.89(m,6H),1.26-1.39(m,51H),1.49-1.68(m,14H),1.95-2.06(m,8H),2.18-2.32(m,9H),2.52- 2.69(m,7H),2.95-3.19(m,6H),3.49-3.67(m,2H),4.00-4.10(m,4H),4.91-5.02(m,4H),5.76-5.86(m,2H). LCMS: Rt: 1.510min; MS m/z (ESI): 972.8[M+H]+ .
以下化合物係以與化合物246類似之方式，使用對應起始原料製備。The following compounds were prepared in a similar manner to compound 246 using the corresponding starting materials.
6.78 實例78：製備化合物247.6.78 Example 78: Preparation of Compound 247.
步驟1：製備化合物247-2Step 1: Preparation of compound 247-2
向247-1(3.5g，12.6mmol，3.0eq.)及化合物216-1(1.0g，4.2mmol，1.0eq.)在DCM(50mL)中之溶液中加入DIEA(2.7g，21.0mmol，5.0eq.)、EDCI(2.4g，12.6mmol，3.0eq.)及DMAP(1.3g，8.4mmol，2.0eq.)。將混合物在50℃下攪拌16小時。將反應混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(PE/EA=10：1)純化殘餘物，得到呈無色油狀之化合物247-2(2.8g，87.5%產率)。1H NMR(400MHz,CDCl3)δ：0.87-1.03(m,6H),1.25-1.43(m,20H),1.58-1.65(m,6H),2.02-2.11(m,9H),2.24-2.34(m,4H),2.69-2.90(m,8H),3.45-3.51(m,2H),4.02-4.10(m,4H),4.49-4.52(m,2H),4.24-4.54(m,12H),4.27-4.38(m,5H)。To a solution of 247-1 (3.5 g, 12.6 mmol, 3.0 eq.) and compound 216-1 (1.0 g, 4.2 mmol, 1.0 eq.) in DCM (50 mL) were added DIEA (2.7 g, 21.0 mmol, 5.0 eq.), EDCI (2.4 g, 12.6 mmol, 3.0 eq.) and DMAP (1.3 g, 8.4 mmol, 2.0 eq.). The mixture was stirred at 50 °C for 16 hours. The reaction mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (PE/EA=10:1) to obtain compound 247-2 (2.8 g, 87.5% yield) as a colorless oil.1 H NMR (400MHz, CDCl3 )δ: 0.87-1.03(m,6H),1.25-1.43(m,20H),1.58-1.65(m,6H),2.02-2.11(m,9H),2.24-2.34(m,4H),2.69-2 .90(m,8H),3.45-3.51(m,2H),4.02-4.10(m,4H),4.49-4.52(m,2H),4.24-4.54(m,12H),4.27-4.38(m,5H).
步驟2：製備化合物247-3Step 2: Preparation of compound 247-3
在-78℃下，向247-2(1.0g，1.3mmol，1.0eq.)在DCM(20mL)中之溶液中加入BCl3(15.6mL，15.6mmol，12.0eq.)。將混合物在-78℃下攪拌1小時。將混合物倒入NaHCO3(水溶液)中並用DCM萃取。將有機層用鹽水洗滌，經Na2SO4乾燥，濃縮並藉由矽膠管柱層析法(PE/EA=5：1)純化，得到呈無色油狀之化合物247-3(634mg，72.87%產率)。1H NMR(400MHz,CDCl3)δ：0.90-1.00(m,6H),1.26-1.37(m,20H),1.58-1.63(m,4H),1.70-1.80(m,2H),2.03-2.12(m,8H),2.26-2.34(m,5H),2.45-2.55(m,1H),2.77-2.87(m,6H),3.63-3.66(m,2H),3.78-3.94(m,1H),4.02-4.11(m,5H),5.25-5.60(m,12H)。To a solution of 247-2 (1.0 g, 1.3 mmol, 1.0 eq.) in DCM (20 mL) was added BCl3 (15.6 mL, 15.6 mmol, 12.0 eq.) at -78°C. The mixture was stirred at -78°C for 1 hour. The mixture was poured into NaHCO3 (aq.) and extracted with DCM. The organic layer was washed with brine, dried over Na2 SO4 , concentrated and purified by silica gel column chromatography (PE/EA=5:1) to give compound 247-3 (634 mg, 72.87% yield) as a colorless oil.1 H NMR (400MHz, CDCl3 )δ: 0.90-1.00(m,6H),1.26-1.37(m,20H),1.58-1.63(m,4H),1.70-1.80(m,2H),2.03-2.12(m,8H),2.26-2.34(m,5H) ),2.45-2.55(m,1H),2.77-2.87(m,6H),3.63-3.66(m,2H),3.78-3.94(m,1H),4.02-4.11(m,5H),5.25-5.60(m,12H).
步驟3：製備化合物247-4Step 3: Preparation of compound 247-4
在0℃下，向247-3(630mg，0.94mmol，1.0eq.)及DIPEA(243mg，1.88mmol，2.0eq.)在DCM(20mL)中之溶液中加入MsCl(129mg，1.13mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物247-4(652mg，92.88%產率)。其未經進一步純化即用於下一步驟。To a solution of 247-3 (630 mg, 0.94 mmol, 1.0 eq.) and DIPEA (243 mg, 1.88 mmol, 2.0 eq.) in DCM (20 mL) at 0°C was added MsCl (129 mg, 1.13 mmol, 1.2 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated to give compound 247-4 (652 mg, 92.88% yield) as a yellow oil. It was used in the next step without further purification.
步驟4：製備化合物247-5Step 4: Preparation of compound 247-5
向247-4(600mg，0.8mmol，1.0eq.)及化合物B(229mg，1.6mmol，2.0eq.)在ACN(10mL)中之溶液中加入K2CO3(332mg，2.4mmol，3.0eq.)、Cs2CO3(78mg，0.24mmol，0.23eq.)及NaI(36mg，0.24mmol，0.3eq.)。將混合物在80℃下攪拌16小時。LCMS顯示反應完成。將反應混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=20/1)純化殘餘物，得到呈黃色油狀之化合物247-5(472mg，74.33%產率)。LCMS：Rt：1.135min；MS m/z(ESI)：794.7[M+H]+。To a solution of 247-4 (600 mg, 0.8 mmol, 1.0 eq.) and compound B (229 mg, 1.6 mmol, 2.0 eq.) in ACN (10 mL) were added K2 CO3 (332 mg, 2.4 mmol, 3.0 eq.), Cs2 CO3 (78 mg, 0.24 mmol, 0.23 eq.) and NaI (36 mg, 0.24 mmol, 0.3 eq.). The mixture was stirred at 80 °C for 16 h. LCMS showed that the reaction was complete. The reaction mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=20/1) to obtain compound 247-5 (472 mg, 74.33% yield) as a yellow oil. LCMS: Rt: 1.135 min; MS m/z (ESI): 794.7 [M+H]+ .
步驟5：製備化合物247-6Step 5: Preparation of compound 247-6
在0℃下，向247-5(472mg，0.59mmol，1.0eq.)及DIPEA(152mg，1.18mmol，2.0eq.)在DCM(10mL)中之溶液中加入MsCl(81mg，0.71mmol，1.2eq.)。將混合物在室溫下攪拌2小時。將混合物倒入水中並用DCM萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮，得到呈黃色油狀之化合物247-6(500mg，97.09%產率)。其未經進一步純化即用於下一步驟。To a solution of 247-5 (472 mg, 0.59 mmol, 1.0 eq.) and DIPEA (152 mg, 1.18 mmol, 2.0 eq.) in DCM (10 mL) at 0°C was added MsCl (81 mg, 0.71 mmol, 1.2 eq.). The mixture was stirred at room temperature for 2 hours. The mixture was poured into water and extracted with DCM. The combined organic layers were washed with brine, driedoverNa2SO4 and concentrated to give compound 247-6 (500 mg, 97.09% yield) as a yellow oil. It was used in the next step without further purification.
步驟6：製備化合物247Step 6: Preparation of compound 247
向247-6(500mg，0.6mmol，1.0eq.)及化合物SM34(480mg，1.2mmol，2.0eq.)在THF(10mL)中之溶液中加入DIEA(230mg，1.8mmol，3.0eq.)及NaI(30mg，0.18mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將反應混合物倒入水中並用EA萃取。將有機層用鹽水洗滌，經Na2SO4乾燥，濃縮並藉由製備型HPLC純化，得到呈黃色油狀之化合物247(38mg，5.4%產率)。To a solution of 247-6 (500 mg, 0.6 mmol, 1.0 eq.) and compound SM34 (480 mg, 1.2 mmol, 2.0 eq.) in THF (10 mL) were added DIEA (230 mg, 1.8 mmol, 3.0 eq.) and NaI (30 mg, 0.18 mmol, 0.3 eq.). The mixture was stirred at 70 °C for 16 hours. LCMS showed that the reaction was complete. The reaction mixture was poured into water and extracted with EA. The organic layer was washed with brine, dried over Na2 SO4 , concentrated and purified by preparative HPLC to give compound 247 (38 mg, 5.4% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.91(m,6H),0.94-1.01(m,6H),0.26-1.33(m,50H),1.57-1.69(m,10H),1.76-1.84(m,4H),1.89-2.06(m,12H),2.10-2.21(m,3H),2.26-2.32(m,6H),2.53-2.67(m,6H),2.70-2.85(m,9H),3.15-3.23(m,3H),3.47-3.74(m,3H),4.00-4.15(m,5H),5.28-5.45(m,12H)。LCMS：Rt：25.165min；MS m/z(ESI)：1174.8[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.91(m,6H),0.94-1.01(m,6H),0.26-1.33(m,50H),1.57-1.69(m,10H),1.76-1.84(m,4H),1.89-2.06(m,12H),2.10-2.21(m,3 H),2.26-2.32(m,6H),2.53-2.67(m,6H),2.70-2.85(m,9H),3.15-3.23(m,3H),3.47-3.74(m,3H),4.00-4.15(m,5H),5.28-5.45(m,12H). LCMS: Rt: 25.165min; MS m/z (ESI): 1174.8[M+H]+ .
6.79 實例79：製備化合物261.6.79 Example 79: Preparation of Compound 261.
步驟1：製備化合物261-1Step 1: Preparation of compound 261-1
向化合物26-1(895mg，2.0mmol，1.0eq.)及化合物SM32(407mg，3.0mmol，1.5eq.)在ACN(20mL)中之溶液中加入K2CO3(829mg，6.0mmol，3.0eq.)、Cs2CO3(195mg，0.6mmol，0.3eq.)及NaI(90mg，0.6mmol，0.3eq.)。將混合物在70℃下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由矽膠管柱層析法(DCM/MeOH=40/1)純化殘餘物，得到呈黃色油狀之化合物261-1(530mg，57%產率)。To a solution of compound 26-1 (895 mg, 2.0 mmol, 1.0 eq.) and compound SM32 (407 mg, 3.0 mmol, 1.5 eq.) in ACN (20 mL) were added K2 CO3 (829 mg, 6.0 mmol, 3.0 eq.), Cs2 CO3 (195 mg, 0.6 mmol, 0.3 eq.) and NaI (90 mg, 0.6 mmol, 0.3 eq.). The mixture was stirred at 70° C. for 16 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layer was washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by silica gel column chromatography (DCM/MeOH=40/1) to give compound 261-1 (530 mg, 57% yield) as a yellow oil.
步驟2：製備化合物261Step 2: Preparation of compound 261
向化合物261-1(200mg，0.43mmol，1.0eq.)及化合物SM16(191mg，0.43mmol，1.0eq.)在DCE(8mL)中之溶液中加入AcOH(1滴)。將混合物在室溫下攪拌6小時。接著加入NaBH(OAc)3(137mg，0.65mmol，1.5eq.)。將混合物在室溫下攪拌16小時。LCMS顯示反應完成。將混合物倒入水中並用EA萃取。將合併之有機層用鹽水洗滌，經Na2SO4乾燥並濃縮。藉由製備型HPLC純化殘餘物，得到呈黃色油狀之化合物261(22mg，6%產率)。To a solution of compound 261-1 (200 mg, 0.43 mmol, 1.0 eq.) and compound SM16 (191 mg, 0.43 mmol, 1.0 eq.) in DCE (8 mL) was added AcOH (1 drop). The mixture was stirred at room temperature for 6 hours. Then NaBH(OAc)3 (137 mg, 0.65 mmol, 1.5 eq.) was added. The mixture was stirred at room temperature for 16 hours. LCMS showed that the reaction was complete. The mixture was poured into water and extracted with EA. The combined organic layers were washed with brine, dried over Na2 SO4 and concentrated. The residue was purified by preparative HPLC to give compound 261 (22 mg, 6% yield) as a yellow oil.
1H NMR(400MHz,CDCl3)δ：0.86-0.90(m,12H),1.26-1.41(m,53H),1.56-1.69(m,18H),1.92-2.03(m,2H),2.29-2.32(m,7H),2.52-2.90(m,6H),3.96-4.10(m,6H)。LCMS：Rt：0.520min；MS m/z(ESI)：893.6[M+H]+。1 H NMR (400MHz, CDCl3 )δ: 0.86-0.90(m,12H),1.26-1.41(m,53H),1.56-1.69(m,18H),1.92-2.03(m,2H),2.29-2.32(m,7H),2.52-2.90(m,6H),3.96-4.10(m,6H). LCMS: Rt: 0.520min; MS m/z (ESI): 893.6[M+H]+ .
6.80 實例80：脂質奈米顆粒之製備及表徵6.80 Example 80: Preparation and Characterization of Lipid Nanoparticles
簡言之，將本文所提供之陽離子脂質、DSPC、膽固醇及PEG-脂質以50：10：38.5：1.5之莫耳比溶解於乙醇中，且在10mM至50mM檸檬酸鹽緩衝液pH=4中稀釋mRNA。藉由使用微流體設備，以在9-30mL/min範圍內之總流速，以1：3之體積比混合脂質乙醇溶液與mRNA水溶液，以大約10：1至30：1之總脂質與mRNA重量比製備出LNP。使用透析移除乙醇且以DPBS替代。最後，將脂質奈米顆粒經0.2μm無菌過濾器過濾。Briefly, cationic lipids, DSPC, cholesterol, and PEG-lipids provided herein were dissolved in ethanol at a molar ratio of 50:10:38.5:1.5, and mRNA was diluted in 10mM to 50mM citrate buffer pH=4. LNPs were prepared with a total lipid to mRNA weight ratio of approximately 10:1 to 30:1 by mixing lipid ethanol solution and mRNA aqueous solution at a total flow rate in the range of 9-30mL/min using a microfluidic device at a volume ratio of 1:3. Ethanol was removed by dialysis and replaced with DPBS. Finally, the lipid nanoparticles were filtered through a 0.2μm sterile filter.
藉由動態光散射，使用Malvern Zetasizer Nano ZS(Malvern UK)，使用173°反向散射偵測模式測定脂質奈米顆粒大小。使用Quant-it Ribogreen RNA定量分析套組(Thermo Fisher Scientific,UK)，根據製造商之說明書，測定脂質奈米顆粒之包封效率。The size of lipid nanoparticles was determined by dynamic light scattering using a Malvern Zetasizer Nano ZS (Malvern UK) in 173° backscatter detection mode. The encapsulation efficiency of lipid nanoparticles was determined using the Quant-it Ribogreen RNA Quantification Kit (Thermo Fisher Scientific, UK) according to the manufacturer’s instructions.
如文獻中所報導，LNP調配物之表觀pKa與在活體內LNP對於核酸之遞送效率相關。使用基於2-(對甲苯胺基)-6-萘磺酸(TNS)之螢光之分析來測定各調配物之表觀pKa。如上文所述，製備出在PBS中包含陽離子脂質/DSPC/膽固醇/DMG-PEG(50/10/38.5/1.5mol%)之LNP調配物。製備TNS於蒸餾水中之300μM儲備液。在3mL含有50mM檸檬酸鈉、50mM磷酸鈉、50mM硼酸鈉及30mM氯化鈉之緩衝溶液(其中pH值在3至9範圍內)中將LNP調配物稀釋至0.1mg/mL總脂質。添加TNS溶液之等分試樣以得到0.1mg/mL之最終濃度且在渦旋混合後，在室溫下，在Molecular Devices Spectramax iD3光譜儀中，使用325nm之激發波長及435nm之發射波長量測螢光強度。將S形曲線最佳擬合分析應用於螢光資料，且將pKa值量測為產生半數最大螢光強度之pH值。As reported in the literature, the apparent pKa of LNP formulations is related to the delivery efficiency of LNPs for nucleic acids in vivo. The apparent pKa of each formulation was determined using a fluorescence analysis based on 2-(p-toluidinyl)-6-naphthalenesulfonic acid (TNS). As described above, LNP formulations containing cationic lipids/DSPC/cholesterol/DMG-PEG (50/10/38.5/1.5 mol%) in PBS were prepared. A 300 μM stock solution of TNS in distilled water was prepared. The LNP formulation was diluted to 0.1 mg/mL total lipid in 3 mL of a buffer solution containing 50 mM sodium citrate, 50 mM sodium phosphate, 50 mM sodium borate, and 30 mM sodium chloride, wherein the pH value was in the range of 3 to 9. Aliquots of TNS solution were added to give a final concentration of 0.1 mg/mL and after vortex mixing, the fluorescence intensity was measured at room temperature in a Molecular Devices Spectramax iD3 spectrometer using an excitation wavelength of 325 nm and an emission wavelength of 435 nm. Sigmoidal best fit analysis was applied to the fluorescence data and the pKa value was measured as the pH value that produced half-maximal fluorescence intensity.
6.81 實例81：動物研究6.81 Example 81: Animal Research
藉由尾靜脈注射將0.5mg/kg劑量之包封人紅血球生成素(hEPO)mRNA之包含下表中之化合物之脂質奈米顆粒經全身投與6-8週齡之雌性ICR小鼠(Xipuer-Bikai,Shanghai)，且在投與後之特定時間點(例如6小時)時收集小鼠血液樣品。除前述測試組外，亦將相同劑量之包封hEPO mRNA之包含二亞油基甲基-4-二甲基胺基丁酸酯(DLin-MC3-DMA，通常縮寫為MC3)之脂質奈米顆粒以類似方式投與年齡及性別相當組之小鼠作為陽性對照。Human erythropoietin (hEPO) mRNA encapsulated lipid nanoparticles containing the compounds in the table below were systemically administered to female ICR mice aged 6-8 weeks (Xipuer-Bikai, Shanghai) by tail vein injection, and mouse blood samples were collected at specific time points (e.g., 6 hours) after administration. In addition to the aforementioned test groups, the same dose of hEPO mRNA encapsulated lipid nanoparticles containing dilinoleylmethyl-4-dimethylaminobutyrate (DLin-MC3-DMA, usually abbreviated as MC3) was also administered to mice of the same age and sex in a similar manner as a positive control.
在最後一個取樣時間點後，藉由超劑量CO2對小鼠實施安樂死。藉由在4℃下以5000g離心10分鐘，自全血分離出血清，急速冷凍並在-80℃下儲存以進行分析。使用可商購套組(DEP00,R&D systems)，根據製造商之說明書，進行ELISA分析。After the last sampling time point, mice were euthanized by overdose of CO2. Serum was separated from whole blood by centrifugation at 5000 g for 10 min at 4°C, snap frozen and stored at -80°C for analysis. ELISA analysis was performed using a commercially available kit (DEP00, R&D systems) according to the manufacturer's instructions.
自測試組量測之測試脂質奈米顆粒之特徵，包括優於MC3之表現水準列於下表中。The characteristics of the tested lipid nanoparticles measured from the test set, including the performance levels that are superior to MC3, are listed in the table below.
6.82 實例82：脂質清除研究6.82 Example 82: Lipid Clearance Study
將LNP經由尾靜脈注射到小鼠體內(ICR雌性，IV，0.5mg mRNA/kg)，接著在投與後不同時間(例如6h、24h及48h)將小鼠在二氧化碳下麻醉，且經由心臟穿刺處死。立即收集肝臟組織，接著用冰冷之鹽水洗滌。稱重肝臟樣品，且在冰水浴中藉由加入預冷之20%甲醇-水(v/v)在2-8℃下以1：5(w/v)之比率均質化。在分析前，將均質化之組織樣品儲存在-90℃至-60℃之冷凍箱中。LNPs were injected into mice via the tail vein (ICR females, IV, 0.5 mg mRNA/kg), and then mice were anesthetized under carbon dioxide and sacrificed by cardiac puncture at different times after administration (e.g., 6h, 24h, and 48h). Liver tissues were collected immediately and then washed with ice-cold saline. Liver samples were weighed and homogenized in an ice-water bath at a ratio of 1:5 (w/v) by adding pre-cooled 20% methanol-water (v/v) at 2-8°C. Homogenized tissue samples were stored in a freezer at -90°C to -60°C before analysis.
樣品處理。允許所有肝臟組織勻漿樣品在室溫下解凍。向50μL樣品之等分試樣中加入50μL MgCl2(2M)，接著加入ACN，其中含有5ng．mL-1維拉帕米(Verapamil)及50ng．mL-1格列本脲(Glibenclamide)及200ng．mL-1雙氯芬酸(Diclofenac)及200ng．mL-1甲苯磺丁脲(Tolbutamide)用於蛋白質沈澱，接著以13000rpm離心8分鐘。接著向100μL上清液中加入100μL水，接著充分渦旋。將5μL混合物之等分試樣注入LC-MS/MS系統中。Sample treatment. All liver tissue homogenate samples were allowed to thaw at room temperature. To a 50 μL aliquot of the sample, 50 μL of MgCl2 (2M) was added, followed by ACN containing 5 ng. mL-1 Verapamil and 50 ng. mL-1 Glibenclamide and 200 ng. mL-1 Diclofenac and 200 ng. mL-1 Tolbutamide for protein precipitation, followed by centrifugation at 13000 rpm for 8 minutes. Then, 100 μL of water was added to 100 μL of the supernatant, followed by thorough vortexing. A 5 μL aliquot of the mixture was injected into the LC-MS/MS system.
MC3及本文提供之選定脂質化合物之結果列於下表中。The results of MC3 and the selected lipid compounds provided herein are listed in the table below.
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| WO2024131874A1 (en)* | 2022-12-21 | 2024-06-27 | Abogen Biosciences (Shanghai) Co., Ltd. | Polynucleotides encoding cd19/cd3 bispecific antibodies |
| CN119095585A (en)* | 2022-12-21 | 2024-12-06 | 苏州艾博生物科技有限公司 | Lipid nanoparticles comprising sterol-modified phospholipids |
| WO2024179559A1 (en)* | 2023-03-02 | 2024-09-06 | Suzhou Abogen Biosciences Co., Ltd. | Nucleic acid vaccines for respiratory syncytial virus (rsv) |
| CN118666726A (en)* | 2023-03-17 | 2024-09-20 | 尧唐(上海)生物科技有限公司 | Lipid compounds for delivering therapeutic agents and preparation methods and applications thereof |
| WO2024245317A1 (en)* | 2023-05-31 | 2024-12-05 | Suzhou Abogen Biosciences Co., Ltd. | Mutated fragment of vzv glycoprotein e |
| WO2025007109A1 (en)* | 2023-06-30 | 2025-01-02 | RVAC Medicines (US), Inc. | Ionizable lipids and lipid nanoparticle compositions for delivery of nucleic acids |
| CN119464283A (en) | 2023-08-11 | 2025-02-18 | 苏州艾博生物科技有限公司 | Capped mRNA and preparation method thereof |
| WO2025087266A1 (en) | 2023-10-23 | 2025-05-01 | Abogen Biosciences (Shanghai) Co., Ltd. | Immunomodulatory mrna cassettes, and uses thereof |
| CN119912351A (en)* | 2023-10-30 | 2025-05-02 | 广州市锐博生物科技有限公司 | Lipid compounds and compositions thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2006138380A2 (en)* | 2005-06-15 | 2006-12-28 | Massachusetts Institute Of Technology | Amine-containing lipids and uses thereof |
| WO2010062322A2 (en)* | 2008-10-27 | 2010-06-03 | Massachusetts Institute Of Technology | Modulation of the immune response |
Family Cites Families (73)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5635487A (en)* | 1994-12-29 | 1997-06-03 | Wolff; Jon A. | Amphipathic, micellar delivery systems for biologically active polyions |
| AU744755B2 (en) | 1997-11-12 | 2002-03-07 | Brigham And Women's Hospital | The translation enhancer element of the human amyloid precursor protein gene |
| US7468275B2 (en) | 2000-01-28 | 2008-12-23 | The Scripps Research Institute | Synthetic internal ribosome entry sites and methods of identifying same |
| BR0107943A (en) | 2000-01-28 | 2003-01-28 | Scripps Research Inst | Synthetic Internal Ribosome Entry Sites and Methods of Identifying Them |
| EP1903054A3 (en) | 2001-06-05 | 2008-07-23 | CureVac GmbH | Pharmaceutical compound containing a stabilised mRNA which is optimised for translation in its coded areas |
| DE10162480A1 (en) | 2001-12-19 | 2003-08-07 | Ingmar Hoerr | The application of mRNA for use as a therapeutic agent against tumor diseases |
| DE10229872A1 (en) | 2002-07-03 | 2004-01-29 | Curevac Gmbh | Immune stimulation through chemically modified RNA |
| DE10335833A1 (en) | 2003-08-05 | 2005-03-03 | Curevac Gmbh | Transfection of blood cells with mRNA for immune stimulation and gene therapy |
| DE102004042546A1 (en) | 2004-09-02 | 2006-03-09 | Curevac Gmbh | Combination therapy for immune stimulation |
| DE102005023170A1 (en) | 2005-05-19 | 2006-11-23 | Curevac Gmbh | Optimized formulation for mRNA |
| WO2007025008A2 (en) | 2005-08-24 | 2007-03-01 | The Scripps Research Institute | Translation enhancer-element dependent vector systems |
| DE102006007433A1 (en) | 2006-02-17 | 2007-08-23 | Curevac Gmbh | Adjuvant in the form of a lipid-modified nucleic acid |
| JP2009544754A (en) | 2006-07-28 | 2009-12-17 | アプライド バイオシステムズ， エルエルシー | Dinucleotide MRNA cap analog |
| WO2008014979A2 (en) | 2006-07-31 | 2008-02-07 | Curevac Gmbh | NUCLEIC ACID OF FORMULA (I): GIXmGn, OR (II): CIXmCn, IN PARTICULAR AS AN IMMUNE-STIMULATING AGENT/ADJUVANT |
| DE102006051516A1 (en) | 2006-10-31 | 2008-05-08 | Curevac Gmbh | (Base) modified RNA to increase the expression of a protein |
| DE102006061015A1 (en) | 2006-12-22 | 2008-06-26 | Curevac Gmbh | Process for the purification of RNA on a preparative scale by HPLC |
| DE102007001370A1 (en) | 2007-01-09 | 2008-07-10 | Curevac Gmbh | RNA-encoded antibodies |
| WO2008127688A1 (en) | 2007-04-13 | 2008-10-23 | Hart Communication Foundation | Synchronizing timeslots in a wireless communication protocol |
| WO2008137470A1 (en)* | 2007-05-01 | 2008-11-13 | Pgr-Solutions | Multi-chain lipophilic polyamines |
| WO2009030254A1 (en) | 2007-09-04 | 2009-03-12 | Curevac Gmbh | Complexes of rna and cationic peptides for transfection and for immunostimulation |
| EP2610340B1 (en) | 2007-12-11 | 2014-10-01 | The Scripps Research Institute | Compositions and methods related to mRNA translational enhancer elements |
| RU2545701C2 (en) | 2008-01-31 | 2015-04-10 | Куревак Гмбх | NUCLEIC ACIDS OF FORMULA (I) (NuGlXmGnNv)a AND DERIVATIVES THEREOF AS IMMUNOSTIMULATING AGENTS/ADJUVANTS |
| WO2009127230A1 (en) | 2008-04-16 | 2009-10-22 | Curevac Gmbh | MODIFIED (m)RNA FOR SUPPRESSING OR AVOIDING AN IMMUNOSTIMULATORY RESPONSE AND IMMUNOSUPPRESSIVE COMPOSITION |
| PL215513B1 (en) | 2008-06-06 | 2013-12-31 | Univ Warszawski | New borane phosphate analogs of dinucleotides, their application, RNA particle, method of obtaining RNA and method of obtaining peptides or protein |
| WO2010037408A1 (en) | 2008-09-30 | 2010-04-08 | Curevac Gmbh | Composition comprising a complexed (m)rna and a naked mrna for providing or enhancing an immunostimulatory response in a mammal and uses thereof |
| WO2010088927A1 (en) | 2009-02-09 | 2010-08-12 | Curevac Gmbh | Use of pei for the improvement of endosomal release and expression of transfected nucleic acids, complexed with cationic or polycationic compounds |
| EP2281579A1 (en) | 2009-08-05 | 2011-02-09 | BioNTech AG | Vaccine composition comprising 5'-Cap modified RNA |
| US20110053829A1 (en) | 2009-09-03 | 2011-03-03 | Curevac Gmbh | Disulfide-linked polyethyleneglycol/peptide conjugates for the transfection of nucleic acids |
| WO2011069529A1 (en) | 2009-12-09 | 2011-06-16 | Curevac Gmbh | Mannose-containing solution for lyophilization, transfection and/or injection of nucleic acids |
| EP2387999A1 (en) | 2010-05-21 | 2011-11-23 | CureVac GmbH | Histidine-containing solution for transfection and/or injection of nucleic acids and uses thereof |
| WO2012009644A2 (en) | 2010-07-16 | 2012-01-19 | Arizona Board Of Regents | Methods to identify synthetic and natural rna elements that enhance protein translation |
| CA2801523C (en) | 2010-07-30 | 2021-08-03 | Curevac Gmbh | Complexation of nucleic acids with disulfide-crosslinked cationic components for transfection and immunostimulation |
| WO2012019630A1 (en) | 2010-08-13 | 2012-02-16 | Curevac Gmbh | Nucleic acid comprising or coding for a histone stem-loop and a poly(a) sequence or a polyadenylation signal for increasing the expression of an encoded protein |
| KR101648986B1 (en) | 2010-11-17 | 2016-08-30 | 두산인프라코어 주식회사 | Sliding bearing having improved lubrication property |
| WO2012089225A1 (en) | 2010-12-29 | 2012-07-05 | Curevac Gmbh | Combination of vaccination and inhibition of mhc class i restricted antigen presentation |
| WO2012116715A1 (en) | 2011-03-02 | 2012-09-07 | Curevac Gmbh | Vaccination in newborns and infants |
| WO2012113413A1 (en) | 2011-02-21 | 2012-08-30 | Curevac Gmbh | Vaccine composition comprising complexed immunostimulatory nucleic acids and antigens packaged with disulfide-linked polyethyleneglycol/peptide conjugates |
| WO2012116714A1 (en) | 2011-03-02 | 2012-09-07 | Curevac Gmbh | Vaccination in elderly patients |
| US8691750B2 (en)* | 2011-05-17 | 2014-04-08 | Axolabs Gmbh | Lipids and compositions for intracellular delivery of biologically active compounds |
| WO2013073480A1 (en)* | 2011-11-18 | 2013-05-23 | 日油株式会社 | Cationic lipid having improved intracellular kinetics |
| WO2013103659A1 (en) | 2012-01-04 | 2013-07-11 | Board Of Supervisors Of Louisiana State University And Agricultural And Mechanical College | Stabilizing rna by incorporating chain-terminating nucleosides at the 3'-terminus |
| WO2013113325A1 (en) | 2012-01-31 | 2013-08-08 | Curevac Gmbh | Negatively charged nucleic acid comprising complexes for immunostimulation |
| WO2013113326A1 (en) | 2012-01-31 | 2013-08-08 | Curevac Gmbh | Pharmaceutical composition comprising a polymeric carrier cargo complex and at least one protein or peptide antigen |
| EP2623121A1 (en) | 2012-01-31 | 2013-08-07 | Bayer Innovation GmbH | Pharmaceutical composition comprising a polymeric carrier cargo complex and an antigen |
| WO2013120498A1 (en) | 2012-02-15 | 2013-08-22 | Curevac Gmbh | Nucleic acid comprising or coding for a histone stem-loop and a poly(a) sequence or a polyadenylation signal for increasing the expression of an encoded allergenic antigen or an autoimmune self-antigen |
| WO2013120497A1 (en) | 2012-02-15 | 2013-08-22 | Curevac Gmbh | Nucleic acid comprising or coding for a histone stem-loop and a poly(a) sequence or a polyadenylation signal for increasing the expression of an encoded therapeutic protein |
| WO2013120499A1 (en) | 2012-02-15 | 2013-08-22 | Curevac Gmbh | Nucleic acid comprising or coding for a histone stem-loop and a poly (a) sequence or a polyadenylation signal for increasing the expression of an encoded pathogenic antigen |
| WO2013120500A1 (en) | 2012-02-15 | 2013-08-22 | Curevac Gmbh | Nucleic acid comprising or coding for a histone stem-loop and a poly(a) sequence or a polyadenylation signal for increasing the expression of an encoded tumour antigen |
| AU2013242403B2 (en) | 2012-03-27 | 2018-10-18 | Curevac Ag | Artificial nucleic acid molecules |
| CA2859452C (en) | 2012-03-27 | 2021-12-21 | Curevac Gmbh | Artificial nucleic acid molecules for improved protein or peptide expression |
| SG10201607966UA (en) | 2012-03-27 | 2016-11-29 | Curevac Ag | Artificial nucleic acid molecules comprising a 5'top utr |
| ES2719598T3 (en) | 2012-05-25 | 2019-07-11 | Curevac Ag | Reversible immobilization and / or controlled release of nucleic acids contained in nanoparticles by polymeric coatings (biodegradable) |
| CA2884870C (en)* | 2012-08-13 | 2022-03-29 | Massachusetts Institute Of Technology | Amine-containing lipidoids and uses thereof |
| CA2897858A1 (en) | 2013-02-22 | 2014-08-28 | Curevac Gmbh | Combination of vaccination and inhibition of the pd-1 pathway |
| WO2015002667A1 (en) | 2013-07-01 | 2015-01-08 | Myq, Inc. | A location regulated point-of-sale system and enhancements |
| CA2919226C (en) | 2013-07-23 | 2024-05-14 | Protiva Biotherapeutics, Inc. | Compositions and methods for delivering messenger rna |
| WO2015024664A1 (en) | 2013-08-21 | 2015-02-26 | Curevac Gmbh | Composition and vaccine for treating prostate cancer |
| EA037217B1 (en) | 2013-08-21 | 2021-02-20 | Куревак Аг | Composition and vaccine for treating lung cancer |
| CA2915712A1 (en) | 2013-08-21 | 2015-02-26 | Margit SCHNEE | Rabies vaccine |
| AU2014310933B2 (en) | 2013-08-21 | 2020-05-14 | CureVac SE | Method for increasing expression of RNA-encoded proteins |
| SG11201510746WA (en) | 2013-08-21 | 2016-03-30 | Curevac Ag | Respiratory syncytial virus (rsv) vaccine |
| AU2014310935B2 (en) | 2013-08-21 | 2019-11-21 | CureVac SE | Combination vaccine |
| ES2806575T3 (en) | 2013-11-01 | 2021-02-18 | Curevac Ag | Modified RNA with decreased immunostimulatory properties |
| JP6584414B2 (en) | 2013-12-30 | 2019-10-02 | キュアバック アーゲー | Artificial nucleic acid molecule |
| MX372790B (en) | 2013-12-30 | 2020-07-03 | CureVac SE | ARTIFICIAL NUCLEIC ACID MOLECULES. |
| AU2014375404C1 (en) | 2013-12-30 | 2020-11-19 | CureVac Manufacturing GmbH | Methods for RNA analysis |
| IL289934B2 (en) | 2014-06-25 | 2023-04-01 | Acuitas Therapeutics Inc | Novel lipids and lipid nanoparticle formulations for delivery of nucleic acids |
| CN104876831B (en)* | 2015-04-03 | 2017-05-17 | 苏州圣诺生物医药技术有限公司 | Liposome-modified spermine derivative and liposome prepared by derivative |
| AU2016253972B2 (en) | 2015-04-27 | 2020-01-02 | Acuitas Therapeutics Inc. | Nucleoside-modified RNA for inducing an adaptive immune response |
| HUE061564T2 (en)* | 2015-10-28 | 2023-07-28 | Acuitas Therapeutics Inc | New lipids and lipid nanoparticle formulations for nucleic acid delivery |
| HRP20220268T1 (en) | 2015-12-22 | 2022-05-13 | Modernatx, Inc. | Compounds and compositions for intracellular delivery of agents |
| WO2018160690A1 (en)* | 2017-02-28 | 2018-09-07 | Guangzhou Nanotides Pharmaceuticals Co., Ltd. | Rna pharmaceutical formulations for prophylactic and therapeutic treatment of zika virus infection |
| US20190194390A1 (en)* | 2017-11-03 | 2019-06-27 | Massachusetts Institute Of Technology | Amino-polyesters for drug delivery |
- 2022
- 2022-01-11WOPCT/CN2022/071251patent/WO2022152109A2/ennot_activeCeased
- 2022-01-13TWTW111101514Apatent/TWI885236B/enactive
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2006138380A2 (en)* | 2005-06-15 | 2006-12-28 | Massachusetts Institute Of Technology | Amine-containing lipids and uses thereof |
| WO2010062322A2 (en)* | 2008-10-27 | 2010-06-03 | Massachusetts Institute Of Technology | Modulation of the immune response |
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|---|---|
| WO2022152109A2 (en) | 2022-07-21 |
| TW202229227A (en) | 2022-08-01 |
| WO2022152109A3 (en) | 2022-08-25 |
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