TW202517674A - Anti-gpc3 antibodies and radioconjugates thereof - Google Patents

Abstract

The present invention relates to novel antibodies binding to GPC3 or antigen-binding fragments thereof. These anti-GPC3 antibodies or fragments thereof preferably don't comprise lysine residues within their CDRs and are therefore especially suitable for conjugation. The present invention further relates to targeted radionuclide conjugates such as targeted actinium-225 conjugates (TACs). Preferred are TACs of formula (I) comprising the anti-GPC3 antibodies according to the present invention or fragments thereof. Said antibodies or antigen binding fragments thereof, and/or said antibody conjugates and/or said TACs are useful in diagnostics and/or therapy, preferably in therapy, more preferably in treating cancer, most preferably in treating hepatocellular carcinoma (HCC). Provided are pharmaceutical compositions and kits with instructions for use comprising these antibodies, functional fragments and conjugates. The invention further provides methods and tools to generate these antibodies, functional fragments and conjugates.

Description

Translated fromChinese

抗GPC3抗體及其放射性結合物Anti-GPC3 antibodies and radioactive conjugates thereof

本發明係關於能夠結合至磷脂醯肌醇蛋白聚醣-3 (GPC3)之新穎單株抗體或其抗原結合片段及包括該等抗GPC3抗體或其片段之結合物。本發明另外係關於靶向放射性核素結合物，例如靶向錒-225 (Ac-225)結合物(TAC)。較佳者係包括該等本發明抗GPC3抗體或其片段之式(I)之靶向錒結合物。該等抗體或其抗原結合片段及/或該等抗體結合物及/或該等靶向錒結合物可用於診斷及/或治療(較佳地治療)，更佳地用於治療過度增殖性疾病(例如癌症)，最佳地用於治療肝細胞癌(HCC)。The present invention relates to novel monoclonal antibodies or antigen-binding fragments thereof capable of binding to phosphatidylinositol proteoglycan-3 (GPC3) and conjugates comprising such anti-GPC3 antibodies or fragments thereof. The present invention further relates to targeted radionuclide conjugates, such as targeted thiabendium-225 (Ac-225) conjugates (TAC). Preferably, the targeted thiabendium conjugates of formula (I) comprising such anti-GPC3 antibodies or fragments thereof of the present invention. Such antibodies or antigen-binding fragments thereof and/or such antibody conjugates and/or such targeted thiabendium conjugates can be used for diagnosis and/or treatment (preferably treatment), more preferably for the treatment of hyperproliferative diseases (such as cancer), and most preferably for the treatment of hepatocellular carcinoma (HCC).

提供包括本發明抗體、功能片段及結合物之醫藥組合物及具有使用說明書之套組。Provided are pharmaceutical compositions comprising the antibodies, functional fragments and conjugates of the present invention and kits with instructions for use.

本發明進一步提供用以生成本發明之抗體、功能片段及結合物之方法及工具。舉例而言，提供編碼前述抗體或片段之多核苷酸、含有其之載體以及用於產生之細胞。The present invention further provides methods and tools for producing the antibodies, functional fragments and conjugates of the present invention. For example, polynucleotides encoding the above antibodies or fragments, vectors containing the same and cells for producing the same are provided.

磷脂醯肌醇蛋白聚醣-3 (GPC3)係在各種腫瘤(例如肝細胞癌(HCC)、非小細胞肺癌(NSCLC)、胃癌、卵巢癌、黑色素瘤及兒科胚胎性腫瘤)中表現之癌胚蛋白質。GPC3在HCC中表現尤其高而成人中具有受限之表現，且其在病毒肝炎、肝硬化或NASH中不表現，此使得HCC成為基於GPC3之療法之高度有吸引力靶。考慮到原發性肝癌係全世界第六常見惡性贅瘤及引起癌症相關死亡之第二常見病因(Torre等人，Global cancer statistics, 2012. CA Cancer J Clin. 2015年3月；5(2):87-108)，此極為重要。Glypican-3 (GPC3) is an oncofetal protein expressed in various tumors, such as hepatocellular carcinoma (HCC), non-small cell lung cancer (NSCLC), gastric cancer, ovarian cancer, melanoma, and pediatric embryonal tumors. GPC3 is particularly highly expressed in HCC with restricted expression in adults, and it is not expressed in viral hepatitis, cirrhosis, or NASH, making HCC a highly attractive target for GPC3-based therapies. This is extremely important considering that primary liver cancer is the sixth most common malignancy worldwide and the second most common cause of cancer-related death (Torre et al., Global cancer statistics, 2012. CA Cancer J Clin. 2015 Mar;5(2):87-108).

迄今為止已闡述一些抗GPC3抗體，例如hYP7、hYP9.1b (Zhang等人，Humanization of high-affinity antibodies targeting glypican-3 in hepatocellular carcinoma. Sci Rep. 2016年9月26日；6:33878)、VHH抗體HN3 (WO2012/145469；Feng等人，Therapeutically targeting glypican-3 via a conformation-specific single-domain antibody in hepatocellular carcinoma. Proc Natl Acad Sci U S A. 2013年3月19日; 110(12)、完全人類抗體2-F7、4-11G、7-5B、7-8B、17-4D (Yu等人，Generation of fully human anti-GPC3 antibodies with high-affinity recognition of GPC3 positive tumors. Invest New Drugs. 2021年6月；39(3):615-626)、或小鼠抗人類GPC3抗體aGPC3 (WO2023/056474；Labadie等人，Glypican-3-Targeted 227Th α-Therapy Reduces Tumor Burden in an Orthotopic Xenograft Murine Model of Hepatocellular Carcinoma. J Nucl Med. 2022年7月；63(7):1033-1038)。然而，迄今為止該等抗GPC3抗體中無一者進入臨床階段。To date, some anti-GPC3 antibodies have been described, such as hYP7, hYP9.1b (Zhang et al., Humanization of high-affinity antibodies targeting glypican-3 in hepatocellular carcinoma. Sci Rep. 2016 Sep 26; 6:33878), VHH antibody HN3 (WO2012/145469; Feng et al., Therapeutically targeting glypican-3 via a conformation-specific single-domain antibody in hepatocellular carcinoma. Proc Natl Acad Sci U S A. 2013 Mar 19; 110(12), fully human antibodies 2-F7, 4-11G, 7-5B, 7-8B, 17-4D (Yu et al., Generation of fully human anti-GPC3 antibodies with high-affinity recognition of GPC3 positive tumors. Invest New Drugs. 2021 Jun;39(3):615-626), or mouse anti-human GPC3 antibody aGPC3 (WO2023/056474; Labadie et al., Glypican-3-Targeted 227Th α-Therapy Reduces Tumor Burden in an Orthotopic Xenograft Murine Model of Hepatocellular Carcinoma. J Nucl Med. 2022 Jul;63(7):1033-1038). However, none of these anti-GPC3 antibodies have entered the clinical stage so far.

唯一在臨床上驗證之GPC3靶向抗體係考曲妥珠單抗(codrituzumab)，即以高親和力特異性結合至GPC3之近膜結構域之鼠類單株抗體GC33之人類化版(Ishiguro等人，Anti-glypican 3 antibody as a potential antitumor agent for human liver cancer.Cancer Res. 2008年12月1日；68(23):9832-8；Nakano等人，Anti-glypican 3 antibodies cause ADCC against human hepatocellular carcinoma cells. Biochem Biophys Res Commun.2009年1月9日；378(2):279-84)。考曲妥珠單抗誘導抗體依賴性細胞細胞毒性(ADCC)及/或補體依賴性細胞細胞毒性(CDC)且抑制腫瘤生長。考曲妥珠單抗在I期試驗中良好耐受且在HCC患者中展示抗腫瘤效應。然而，在隨機化II期研究中，考曲妥珠單抗與安慰劑相比不展示益處(Abou-Alfa等人，Randomized phase II placebo-controlled study of codrituzumab in previously treated patients with advanced hepatocellular carcinoma. J Hepatol. 2016年8月；65(2):289-95)。一些隨訪臨床研究展示，考曲妥珠單抗針對具有與ADCC相關之高敏感性及GPC3之高表現之HCC患者展示預後價值(Chen等人，Combining expression of GPC3 in tumors and CD16 on NK cells from peripheral blood to identify patients responding to codrituzumab. Oncotarget. 2018年1月2日；9(12):10436-10444)。目前，臨床I期研究正在募集患有實體腫瘤之兒童及年輕成人進行考曲妥珠單抗測試(Clinical Trials.gov標識符：NCT04928677)。The only clinically validated GPC3-targeting antibody is codrituzumab, a humanized version of the murine monoclonal antibody GC33 that specifically binds to the juxtamembrane domain of GPC3 with high affinity (Ishiguro et al., Anti-glypican 3 antibody as a potential antitumor agent for human liver cancer. Cancer Res. 2008 Dec 1;68(23):9832-8; Nakano et al., Anti-glypican 3 antibodies cause ADCC against human hepatocellular carcinoma cells. Biochem Biophys Res Commun. 2009 Jan 9;378(2):279-84). Codrituzumab induces antibody-dependent cellular cytotoxicity (ADCC) and/or complement-dependent cellular cytotoxicity (CDC) and inhibits tumor growth. Codrituzumab was well tolerated in phase I trials and showed antitumor effects in HCC patients. However, in a randomized phase II study, codrituzumab did not show benefit compared with placebo (Abou-Alfa et al., Randomized phase II placebo-controlled study of codrituzumab in previously treated patients with advanced hepatocellular carcinoma. J Hepatol. 2016 Aug;65(2):289-95). Some follow-up clinical studies have shown that codrituzumab has prognostic value for HCC patients with high sensitivity associated with ADCC and high expression of GPC3 (Chen et al., Combining expression of GPC3 in tumors and CD16 on NK cells from peripheral blood to identify patients responding to codrituzumab. Oncotarget. 2018 Jan 2;9(12):10436-10444). Currently, a phase I clinical study is recruiting children and young adults with solid tumors to test codrituzumab (Clinical Trials.gov identifier: NCT04928677).

然而，因GPC3靶向抗體之ADCC/CDC活性似乎不足以有效治療其中GPC3之表現發揮作用之疾病，故需要特異性靶向GPC3之新穎治療劑。GPC3係在特定癌症類型中主要表現但在正常組織中不或僅極少表現之癌胚蛋白質，此使其成為靶向放射性核素療法之適宜靶。靶向放射性核素療法係有前景且在發展中之領域，有可能將高細胞毒性放射特異性地遞送至與疾病相關之細胞類型。目前授權用於人類之放射性醫藥之最常見形式採用β-發射及/或γ-發射放射性核素。然而，因其更強效之細胞殺死之潛能，α-發射放射性核素之使用愈來愈受關注，尤其針對癌症療法。However, because the ADCC/CDC activity of GPC3-targeted antibodies appears insufficient to effectively treat diseases in which expression of GPC3 plays a role, novel therapeutic agents that specifically target GPC3 are needed. GPC3 is an oncofetal protein that is predominantly expressed in certain cancer types but not or only minimally expressed in normal tissues, making it a suitable target for targeted radionuclide therapy. Targeted radionuclide therapy is a promising and developing field that has the potential to deliver highly cytotoxic radiation specifically to disease-associated cell types. The most common forms of radiopharmaceuticals currently licensed for use in humans employ beta-emitting and/or gamma-emitting radionuclides. However, due to their more potent cytolytic potential, the use of alpha-emitting radionuclides is gaining increasing attention, particularly for cancer therapy.

靶向α療法(TAT)之獨特作用模式係經由使用腫瘤抗原靶向部分(例如小分子、肽或單株抗體)將α-粒子發射放射性核素(如錒-225)特異性遞送至腫瘤來誘導難以修復之群集DNA雙鏈斷裂。α發射放射性核素之優異細胞毒性需要特異性生物靶向抗體以避免不可接受之副效應。同位素之生物靶向很大程度上取決於螯合劑之穩定性、抗體對螯合過程之穩健性及抗體在特異性生物背景中對其特定靶之靶向性質。The unique mode of action of targeted alpha therapy (TAT) is to induce irreparable clustered DNA double-strand breaks by specific delivery of alpha-particle emitting radionuclides (such as threonine-225) to tumors using tumor antigen targeting moieties (e.g., small molecules, peptides, or monoclonal antibodies). The excellent cytotoxicity of alpha-emitting radionuclides requires specific biological targeting antibodies to avoid unacceptable side effects. The biological targeting of the isotope depends largely on the stability of the chelator, the robustness of the antibody to the chelation process, and the targeting properties of the antibody to its specific target in a specific biological background.

典型α發射體在生理環境中之放射範圍通常小於100微米，相當於僅幾個細胞直徑。此使得該等來源非常適合於治療腫瘤(包括微小轉移)，此乃因其在腫瘤內之範圍可到達鄰近細胞，但若其良好靶向，則放射能量幾乎不會超出靶細胞。因此，並非需要靶向每一細胞且同時可最小化對外圍健康組織之損害(參見Feinendegen等人，Radiat. Res.148:195-201 (1997))。與之相比，β顆粒在水中具有大於1 mm或更大之範圍(參見Wilbur D.S., Antibody Immunoconj Radiopharm. 4: 85-97 (1991))。The radiation range of a typical alpha emitter in a physiological environment is usually less than 100 microns, equivalent to only a few cell diameters. This makes these sources very suitable for treating tumors (including micrometastases) because their range within the tumor can reach neighboring cells, but if they are well targeted, the radiation energy will hardly exceed the target cell. Therefore, it is not necessary to target every cell and at the same time damage to surrounding healthy tissue can be minimized (see Feinendegen et al., Radiat. Res. 148:195-201 (1997)). In contrast, beta particles have a range of more than 1 mm or more in water (see Wilbur D.S., Antibody Immunoconj Radiopharm. 4: 85-97 (1991)).

α-粒子放射之能量與β粒子、γ射線及X射線所載能量相比較高，通常為5-8 MeV，或係β粒子之5至10倍，且係γ射線之能量之20倍或更多倍。因此，與γ及β放射相比，此大量能量在極短距離內的沈積給予α-放射極高線性能量轉移(LET)、高相對生物效能(RBE)及低氧增強比(OER) (參見Hall, 「Radiobiology for the radiologist」，第5版，Lippincott Williams & Wilkins, Philadelphia PA, USA, 2000)。此解釋了α發射放射性核素之優異細胞毒性且亦對該等同位素之生物靶向及α發射放射性核素分佈之控制及研究程度提出嚴格需求，此為避免不可接受之副作用所必需。The energy of alpha-particle radiation is high compared to the energy carried by beta particles, gamma rays, and X-rays, typically 5-8 MeV, or 5 to 10 times the energy of beta particles and 20 or more times the energy of gamma rays. Thus, the deposition of this large amount of energy over very short distances gives alpha-radiation an extremely high linear energy transfer (LET), high relative biological effectiveness (RBE), and low oxygen enhancement ratio (OER) compared to gamma and beta radiation (see Hall, "Radiobiology for the radiologist", 5th ed., Lippincott Williams & Wilkins, Philadelphia PA, USA, 2000). This explains the excellent cytotoxicity of alpha-emitting radionuclides and also places strict demands on the biological targeting of these isotopes and the degree of control and study of the distribution of alpha-emitting radionuclides, which is necessary to avoid unacceptable side effects.

迄今為止，關於α-發射核素在放射性免疫療法中之應用，主要注意力集中於²¹¹At、²¹³Bi及²²⁵Ac，且該三種核素已在臨床免疫療法試驗中加以探索。然而，通常採用大環螯合劑DOTA (2,2′,2′′,2′′′-(1,4,7,10-四氮雜環十二烷-1,4,7,10-四基)四乙酸)之當前使用之化合物缺乏穩定性，引起放射性核素自螯合大環之解離，此導致對靶向組織之選擇性及活性減小及對非靶向組織之毒性增加。血清穩定性研究已展示，[225Ac]Ac-DOTA複合物隨時間解離(Ramogida等人，Evaluation of Polydentate Picolinic Acid Chelating Ligands and an α-Melanocyte-Stimulating Hormone Derivative for Targeted Alpha Therapy Using ISOL-Produced 225Ac. EJNMMI Radiopharm. Chem. 2019, 4, 21)，使得在投與基於DOTA之結合物之後225Ac沈積在肝及股骨中。(Deal等人，Improved in Vivo Stability of Actinium-225 Macrocyclic Complexes. J. Med. Chem. 1999, 42, 2988−2992.)To date, the main focus of attention on the application of α-emitting nuclides in radioimmunotherapy has been on²¹¹ At,²¹³ Bi and²²⁵ Ac, and these three nuclides have been explored in clinical immunotherapy trials. However, currently used compounds, which usually employ the macrocyclic chelator DOTA (2,2′,2′′,2′′′-(1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrayl)tetraacetic acid), lack stability, causing the dissociation of the radionuclide from the chelating macrocycle, which results in reduced selectivity and activity for the target tissue and increased toxicity for non-target tissues. Serum stability studies have shown that the [225Ac]Ac-DOTA complex dissociates over time (Ramogida et al., Evaluation of Polydentate Picolinic Acid Chelating Ligands and an α-Melanocyte-Stimulating Hormone Derivative for Targeted Alpha Therapy Using ISOL-Produced 225Ac. EJNMMI Radiopharm. Chem. 2019, 4, 21), resulting in 225Ac deposition in the liver and femur after administration of the DOTA-based conjugate. (Deal et al., Improved in Vivo Stability of Actinium-225 Macrocyclic Complexes. J. Med. Chem. 1999, 42, 2988−2992.)

已進一步努力以鑑別且研發針對大Ac³⁺離子之適宜螯合劑(Thiele等人，Actinium-225 for Targeted α Therapy: Coordination Chemistry and Current Chelation Approaches. Cancer Biother. Radiopharm. 2018, 33, 336−348)。非環螯合劑(如乙二胺四乙酸(H4EDTA)及二伸乙基三胺五乙酸(H5DTPA))所得之複合物具有不穩定性(Davis等人，Comparison of 225Actinium Chelates: Tissue Distribution and Radiotoxicity. Nucl. Med. Biol. 1999, 26, 581−589)。諸如四氮雜-環十二烷-1,4,7,10-四乙酸(H4DOTA)之大環螯合劑經展示更具惰性。然而，使用225Ac放射性標記H4DOTA需要與溫敏性大分子生物分子(例如抗體)不相容之高溫或需要較長培育時間以產生具有相對較低比活性及低純度之構築體。Further efforts have been made to identify and develop suitable chelators for large Ac³⁺ ions (Thiele et al., Actinium-225 for Targeted α Therapy: Coordination Chemistry and Current Chelation Approaches. Cancer Biother. Radiopharm. 2018, 33, 336−348). The complexes obtained with acyclic chelators such as ethylenediaminetetraacetic acid (H4EDTA) and diethylenetriaminepentaacetic acid (H5DTPA) are unstable (Davis et al., Comparison of 225Actinium Chelates: Tissue Distribution and Radiotoxicity. Nucl. Med. Biol. 1999, 26, 581−589). Macrocyclic chelators such as tetraaza-cyclododecane-1,4,7,10-tetraacetic acid (H4DOTA) have been shown to be more inert. However, radiolabeling H4DOTA with 225Ac requires high temperatures that are incompatible with temperature-sensitive macromolecular biomolecules (e.g., antibodies) or requires long incubation times to produce constructs with relatively low specific activity and low purity.

迄今為止，僅已闡述GPC3靶向放射性核素結合物之幾個實例。WO2022115778闡述包括結合至可與放射性同位素(例如用於治療癌症之Lu177或Ac225)複合之金屬螯合劑(例如DOTA)之大環肽之靶向部分。作為相對較小分子，該等肽展示比較大分子(例如抗體)快得多之腫瘤清除及相對較低腫瘤攝取，如Li等人針對RAYZ-8009 (一種包括GPC3結合大環肽、連接體及DOTA之結合物)所闡述。(Li等人，Novel peptide binder to Glypican-3 for targeted radiopharmaceutical therapy of hepatocellular carcinoma. Journal of Clinical Oncology 2023, 41:16_增刊，e16131-e16131)。To date, only a few examples of GPC3-targeted radionuclide conjugates have been described. WO2022115778 describes targeting moieties comprising macrocyclic peptides conjugated to metal chelators (e.g., DOTA) that can be complexed with radioisotopes (e.g., Lu177 or Ac225 for the treatment of cancer). As relatively small molecules, these peptides exhibit much faster tumor clearance and relatively lower tumor uptake than large molecules (e.g., antibodies), as described by Li et al. for RAYZ-8009, a conjugate comprising a GPC3-binding macrocyclic peptide, a linker, and DOTA. (Li et al., Novel peptide binder to Glypican-3 for targeted radiopharmaceutical therapy of hepatocellular carcinoma. Journal of Clinical Oncology 2023, 41:16_Supplement, e16131-e16131).

Labadie等人闡述用於治療正位鼠類模型之HCC之²²⁷Th標記之GPC3靶向抗體結合物(²²⁷Th-octapa-αGPC3) (Labadie等人，Glypican-3-Targeted 227Th α-Therapy Reduces Tumor Burden in an Orthotopic Xenograft Murine Model of Hepatocellular Carcinoma. J Nucl Med. 2022年7月；63(7):1033-1038；WO2023/056474)，針對該治療在室溫下2小時之後觀察到少至70%²²⁷Th標記產率。另外，此放射性藥物採用具有相對較長半衰期(18.7天)之²²⁷Th放射性核素，且²²⁷Th之第一子體²²³Rathe具有較長半衰期，從而導致與²²⁵Ac比較放射性劑量之遞送較為緩慢，²²⁵Ac之子體具有較短半衰期，由此遞送放射性劑量快得多。Labadie et al. described a²²⁷ Th-labeled GPC3-targeted antibody conjugate (²²⁷ Th-octapa-αGPC3) for the treatment of HCC in an orthotopic mouse model (Labadie et al., Glypican-3-Targeted 227 Th α-Therapy Reduces Tumor Burden in an Orthotopic Xenograft Murine Model of Hepatocellular Carcinoma. J Nucl Med. 2022 Jul;63(7):1033-1038; WO2023/056474), for which as little as 70%²²⁷ Th labeling yield was observed after 2 hours at room temperature. In addition, this radiopharmaceutical uses²²⁷ Th radionuclide with a relatively long half-life (18.7 days), and the first daughter of²²⁷ Th,²²³ Rathe, has a long half-life, resulting in slower delivery of radioactive dose compared to²²⁵ Ac, whose^daughters have a shorter half-life and thus deliver radioactive dose much faster.

Bell等人闡述基於考曲妥珠單抗之錒結合物[²²⁵Ac]Ac–Macropa–GC33 (Bell等人，Glypican-3-Targeted Alpha Particle Therapy for Hepatocellular Carcinoma. Molecules. 2020年12月22日；26(1):4)，且Kadassery等人比較[²²⁵Ac]Ac-GC33-BZmacropa與類似之[²²⁵Ac]Ac-GC33-macropa結合物(Kadassery等人，H2BZmacropa-NCS: A Bifunctional Chelator for Actinium-225 Targeted Alpha Therapy. Bioconjug Chem.2022年6月15日；33(6): 1222-1231)。儘管該等結合物採用高效α-發射放射性核素錒225，但其係基於GPC3-結合抗體考曲妥珠單抗(GC33)，然而該抗體在其CDR內包括4個離胺酸殘基。同樣，WO2023/056474 (參見上文)中揭示之αGPC3在其CDR內包括4個離胺酸殘基，且Yu等人闡述之完全人類抗體2-F74-11G、7-5B、7-8B及17-4D  (Yu等人，Generation of fully human anti-GPC3 antibodies with high-affinity recognition of GPC3 positive tumors. Invest New Drugs. 2021年6月；39(3):615-626)在其CDR內亦包括一或多種離胺酸殘基(2-F7：3個離胺酸殘基；2- 4-11G、75B、7-8B及17-4D：1個離胺酸殘基)。Bell et al. described the trastuzumab-based chelator conjugate [^225Ac ]Ac–Macropa–GC33 (Bell et al., Glypican-3-Targeted Alpha Particle Therapy for Hepatocellular Carcinoma. Molecules. 2020 Dec 22;26(1):4), and Kadassery et al. compared [^225Ac ]Ac-GC33-BZmacropa with a similar [^225Ac ]Ac-GC33-macropa conjugate (Kadassery et al., H2BZmacropa-NCS: A Bifunctional Chelator for Actinium-225 Targeted Alpha Therapy. Bioconjug Chem. 2022 Jun 15;33(6):1222-1231). Although these conjugates employ the highly potent α-emitting radionuclide 225thiocyanate, they are based on the GPC3-binding antibody trastuzumab (GC33), which however includes four lysine residues within its CDRs. Similarly, αGPC3 disclosed in WO2023/056474 (see above) includes 4 lysine residues in its CDR, and the fully human antibodies 2-F74-11G, 7-5B, 7-8B and 17-4D described by Yu et al. (Yu et al., Generation of fully human anti-GPC3 antibodies with high-affinity recognition of GPC3 positive tumors. Invest New Drugs. 2021 Jun; 39(3): 615-626) also include one or more lysine residues in their CDR (2-F7: 3 lysine residues; 2-4-11G, 75B, 7-8B and 17-4D: 1 lysine residue).

螯合劑(例如macropa)之結合通常經由抗體之離胺酸殘基進行且亦可在抗體之靶結合CDR區域內發生。該等標記阻礙抗體結合至其靶之能力。CDR中之離胺酸殘基之存在由此意指增加抗體結合物之非結合部分之風險，從而導致(例如)內化比率下降或引起毒性副效應。在每個抗體大約10個螯合劑之標記程度下，Bell等人 (2020) (參見上文)觀察到其基於考曲妥珠單抗之錒結合物[²²⁵Ac]Ac–Macropa–GC33之實質性非特異性肝累積。此發現最可能係由於藉由在GC33 CDR中標記離胺酸而增加之結合物之非結合部分。將人類化抗GPC3抗體考曲妥珠單抗用於靶向α結合物之其他缺點係其可引起免疫原性相關副效應之相對較高免疫原性潛能。Binding of chelators (e.g., macropa) typically occurs via lysine residues of the antibody and can also occur within the target-binding CDR regions of the antibody. Such labeling hinders the ability of the antibody to bind to its target. The presence of lysine residues in the CDRs thus implies an increased risk of non-binding portion of the antibody conjugate, leading to, for example, decreased internalization rates or toxic side effects. At a labeling level of approximately 10 chelators per antibody, Bell et al. (2020) (see above) observed substantial non-specific liver accumulation of their trastuzumab-based thiabendazole conjugate [^225Ac ]Ac–Macropa–GC33. This finding is most likely due to the increase in the non-binding portion of the conjugate by labeling lysine in the GC33 CDRs. Another disadvantage of using the humanized anti-GPC3 antibody trastuzumab to target the alpha binder is its relatively high immunogenic potential that can cause immunogenicity-related side effects.

本發明目標鑒於先前技術，本發明目標係提供新穎治療性抗GPC3抗體及其抗原結合片段，其尤其適於用作克服先前技術中之抗GPC3抗體及GPC3靶向放射性核素結合物之缺點之靶向放射性核素療法之結合物。具體而言本發明目標係提供：(i)具有高靶親和力、對食蟹猴抗原之交叉反應性、低免疫原性、高熱穩定性及藉由重組產生之良好可製造性之新穎抗GPC3抗體及其抗原結合片段，同時CDR中之離胺酸殘基之數量及其他序列傾向性儘可能低及(ii)仍具有高結合親和力之包括該等抗GPC3抗體及其抗原結合片段之靶向α-粒子-發射放射性核素結合物(較佳地靶向錒結合物(TAC))，其高度穩定、具有低螯合劑對抗原比率(CAR)且可在溫和條件下以高產率螯合。Objective of the present invention In view of the prior art, an object of the present invention is to provide novel therapeutic anti-GPC3 antibodies and antigen-binding fragments thereof, which are particularly suitable for use as conjugates for targeted radionuclide therapy that overcome the shortcomings of the anti-GPC3 antibodies and GPC3-targeted radionuclide conjugates in the prior art. Specifically, the present invention aims to provide: (i) novel anti-GPC3 antibodies and antigen-binding fragments thereof having high target affinity, cross-reactivity to cynomolgus monkey antigens, low immunogenicity, high thermal stability and good manufacturability produced by recombination, while the number of lysine residues in the CDRs and other sequence preferences are as low as possible and (ii) targeted α-particle-emitting radionuclide conjugates (preferably targeted titanate conjugates (TACs)) comprising such anti-GPC3 antibodies and antigen-binding fragments thereof that still have high binding affinity, are highly stable, have a low chelator to antigen ratio (CAR) and can be chelated under mild conditions with high yield.

該等新穎抗GPC3抗體及其抗GPC3-TAC在過度增殖性疾病(例如癌症，例如肝細胞癌(HCC))之治療中提供主要優點。These novel anti-GPC3 antibodies and their anti-GPC3-TACs offer major advantages in the treatment of hyperproliferative diseases such as cancers, such as hepatocellular carcinoma (HCC).

另外，該等新穎抗體亦可用作成像藥劑，例如在經由具有正電子發射體之適宜螯合劑標記時用於PET成像。In addition, these novel antibodies can also be used as imaging agents, for example, for PET imaging when labeled with a suitable chelator with a positron emitter.

藉由本發明之教示內容可達成上文所提及目標及其他目標。本發明係基於尤其適於用作靶向放射性核素療法之結合物且可向個體遞送治療性益處之新穎抗體靶向GPC3之發現。The above-mentioned objects and other objects can be achieved by the teachings of the present invention. The present invention is based on the discovery of novel antibodies targeting GPC3 that are particularly suitable for use as conjugates for targeted radionuclide therapy and can deliver therapeutic benefits to individuals.

鼠類單株抗體GC33之人類化版考曲妥珠單抗係唯一在臨床上驗證之GPC3靶向抗體。然而，此抗體具有以下缺點：高免疫原性評分、大量種系偏差、經由離胺酸偶合阻礙螯合劑之結合之CDR中之離胺酸殘基的存在、以及CDR中之潛在天門冬胺酸鹽異構化位點。為最佳化該等特徵，對考曲妥珠單抗進行序列種系化，另外引入改變，例如去除CDR中之離胺酸殘基或去除潛在天門冬胺酸鹽異構化位點。Nakano等人已闡述產生考曲妥珠單抗之GC33之人類化及最佳化。其展示，即使單一胺基酸之取代亦會導致結合之完全損失(例如CDR-L1中之N33D)。因此，期望在不完全損失有益性質之情況下，可實施更多最佳化。此外，因與人類種系(32)之大量偏差，故序列空間過大而不能進行系統性地整體評價以進一步顯著減小其衍生自小鼠之免疫化之抗體所固有之免疫原性潛能。The humanized version of the murine monoclonal antibody GC33, trastuzumab, is the only GPC3-targeting antibody that has been clinically validated. However, this antibody has the following disadvantages: high immunogenicity scores, substantial germline deviations, the presence of lysine residues in the CDRs that hinder the binding of chelators via lysine coupling, and potential aspartate isomerization sites in the CDRs. To optimize these features, trastuzumab was sequence germlined and additional changes were introduced, such as removal of lysine residues in the CDRs or removal of potential aspartate isomerization sites. The humanization and optimization of GC33 that resulted in trastuzumab has been described by Nakano et al. It has been shown that even a single amino acid substitution can result in a complete loss of binding (e.g., N33D in CDR-L1). Thus, it is expected that more optimization can be performed without a complete loss of beneficial properties. In addition, the immunogenic potential inherent in antibodies derived from mouse immunization is further significantly reduced by the large size of the sequence space that cannot be systematically evaluated overall due to the large deviations from the human germline (32).

令人吃驚地，儘管與考曲妥珠單抗相比，VH及VL二者中存在極大量之突變，但申請者能夠鑑別抗體變體，該等抗體變體a)仍可以重組方式產生；且b)在結合至重組或細胞人類GPC3中展示相當或優良親和力；且c)仍展示對食蟹猴GPC3之結合；且d)仍展示Fab部分之穩定性，即Tm高於65℃。然而，種系偏差之數量顯著減小，此在大大減小之電腦免疫原性評分(IPAD, Epibase)以及活體外免疫原性評價(Epibase)中亦得以反映。IPAD免疫原性評分比上文闡述之完全人類抗體2-F7、4-11G、7-5B、7-8B、17-4D之評分甚至更有前景。Surprisingly, despite the extremely high number of mutations in both VH and VL compared to trastuzumab, the applicants were able to identify antibody variants that a) can still be produced recombinantly; and b) show comparable or excellent affinity in binding to recombinant or cellular human GPC3; and c) still show binding to cynomolgus monkey GPC3; and d) still show stability of the Fab portion, i.e., Tm above 65° C. However, the amount of germline deviation is significantly reduced, which is also reflected in the greatly reduced in silico immunogenicity scores (IPAD, Epibase) as well as in vitro immunogenicity assessments (Epibase). The IPAD immunogenicity score is even more promising than the scores of the fully human antibodies 2-F7, 4-11G, 7-5B, 7-8B, and 17-4D described above.

令人吃驚地，藉由組合種系化及改變CDR序列，申請者能夠鑑別具有有前景之結合性質及減小之免疫原性潛能之新穎抗GPC3抗體，其在CDR中不含有任何離胺酸殘基，此使其尤其適於用作經由離胺酸殘基結合之放射性藥物或其他抗體藥物結合物之靶向部分。Surprisingly, by combining germlining and altering the CDR sequences, the applicants were able to identify novel anti-GPC3 antibodies with promising binding properties and reduced immunogenic potential that do not contain any lysine residues in the CDRs, making them particularly suitable for use as targeting moieties for radiopharmaceuticals or other antibody-drug conjugates conjugated via lysine residues.

另外，本發明中之用於靶向錒結合物(TAC)之螯合劑Macropa-NCS容許在較低螯合劑對抗原(CAR)比率下之高免疫反應性分數(IRF)值以及高單體純度，由此產生高分率之具有針對GPC3之親和力之放射性核素標記之化合物。另外，本發明GPC3-TAC展示活體內有益生物分佈及在肝中之低累積，此為放射性核素自螯合部分之解離減少及由此穩定性增加之徵象。在極溫和條件及減小之溫度(亦即，室溫)下可使用放射性核素標記本發明之抗體螯合劑結合物，由此減小抗體變性及最終產物中之非-GPC3結合部分之量。因此，本發明TAC展示改良之腫瘤對肝比率且可由此更有效及選擇性地治療疾病，從而減小對非靶組織之損害同時維持針對靶向組織中之癌細胞之高功效。In addition, the chelator Macropa-NCS used in the present invention for targeted tantalum conjugates (TACs) allows high immunoreactivity fraction (IRF) values and high monomer purity at low chelator to antigen (CAR) ratios, thereby producing a high fraction of radionuclide-labeled compounds with affinity for GPC3. In addition, the GPC3-TAC of the present invention exhibits beneficial biodistribution in vivo and low accumulation in the liver, which is a sign of reduced dissociation of the radionuclide from the chelating portion and thus increased stability. The antibody chelator conjugate of the present invention can be labeled with radionuclides under extremely mild conditions and reduced temperatures (i.e., room temperature), thereby reducing antibody denaturation and the amount of non-GPC3 bound moieties in the final product. Thus, the TACs of the present invention exhibit an improved tumor-to-liver ratio and can thereby treat the disease more effectively and selectively, thereby reducing damage to non-target tissues while maintaining high efficacy against cancer cells in the targeted tissues.

除其用作靶向放射性核素療法之結合物之適宜性外，本發明之抗體或其片段亦可用於各種其他目的，例如成像、抗體藥物結合物或在不存在酬載下之治療劑。舉例而言，在結合至螯合劑DFO* (例如在WO2021051168中所闡述)及使用⁸⁹Zr標記時，本發明之抗體或其片段可用於PET成像目的。In addition to their suitability for use as conjugates for targeted radionuclide therapy, the antibodies or fragments thereof of the present invention may also be used for various other purposes, such as imaging, antibody-drug conjugates, or therapeutic agents in the absence of a payload. For example, when conjugated to the chelator DFO* (e.g., as described in WO2021051168) and labeled with⁸⁹ Zr, the antibodies or fragments thereof of the present invention may be used for PET imaging purposes.

藉由參照本發明之下列詳細說明及其中所包含之實例可更易於理解本發明。The present invention may be more readily understood by reference to the following detailed description of the present invention and the examples included therein.

定義除非另外定義，否則本文所用所有技術及科學術語皆具有熟習本發明所屬技術領域者通常所理解之含義。然而，下列參考文獻可向熟習本發明相關之技術者提供本發明中使用之許多術語之一般定義，且只要該等定義與業內通常理解之含義一致，即可參考並使用該等參考文獻。該等參考文獻包含(但不限於)Singleton等人，Dictionary of Microbiology and Molecular Biology (1994年第2版)；The Cambridge Dictionary of Science and Technology (Walker編輯，1988年)；Hale & Marham, The Harper Collins Dictionary of Biology (1991)；及Lackie等人，The Dictionary of Cell & Molecular Biology (1999年第3版)；及Cellular and Molecular Immunology， Abbas、Lichtman及Pober編輯，第二版，W.B. Saunders Company。可查閱熟習此項技術者可用之提供本文中使用之具有業內通常理解的含義之術語的定義之任何其他技術資源。出於本發明之目的，進一步定義下列術語。其他術語在說明書其他處得以定義。除非上下文另外明確規定，否則本文及隨附申請專利範圍中所用單數形式「一」及「該」包括複數個指示物。因此， 舉例而言，提及「一基因」為提及一或多種基因且包含熟習此項技術者已知之其等效物及等等。Definitions Unless otherwise defined, all technical and scientific terms used herein have the meanings commonly understood by those skilled in the art to which the present invention belongs. However, the following references may provide general definitions of many of the terms used in the present invention to those skilled in the art, and such references may be referenced and used as long as such definitions are consistent with the meanings commonly understood in the industry. Such references include, but are not limited to, Singleton et al., Dictionary of Microbiology and Molecular Biology (2nd edition, 1994); The Cambridge Dictionary of Science and Technology (Walker ed., 1988); Hale & Marham, The Harper Collins Dictionary of Biology (1991); and Lackie et al., The Dictionary of Cell & Molecular Biology (3rd edition, 1999); and Cellular and Molecular Immunology, Abbas, Lichtman and Pober eds., 2nd edition, WB Saunders Company. Any other technical resources available to one skilled in the art that provide definitions of terms used herein with meanings generally understood in the art may be consulted. For purposes of the present invention, the following terms are further defined. Other terms are defined elsewhere in the specification. As used herein and in the appended claims, the singular forms "a,""an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a gene" is a reference to one or more genes and includes equivalents thereof known to those skilled in the art, and so forth.

在本發明之上下文中，術語「包括(comprises或comprising)」意指「包含(但不限於)」。該術語意欲為開放性，以規定所陳述特徵、要素、整數、步驟或組分之存在，但並不排除一或多個其他特徵、要素、整數、步驟、組分或其群組之存在或添加。術語「包括」由此包含更據限制性之術語「由……組成」及「基本上由……組成」。在一實施例中，術語「由……組成」可代替整個申請案中且尤其申請專利範圍內使用之術語「包括」。In the context of the present invention, the term "comprises" or "comprising" means "including (but not limited to)". The term is intended to be open to specify the presence of the stated features, elements, integers, steps or components, but does not exclude the presence or addition of one or more other features, elements, integers, steps, components or groups thereof. The term "comprising" thus includes the more restrictive terms "consisting of" and "consisting essentially of". In one embodiment, the term "consisting of" can replace the term "comprising" used throughout the application and in particular in the scope of the patent application.

在此上下文中，術語「約」或「大約」意指在既定值或範圍之80%至120%內、替代地90%至110%內(包含95%至105%內)。In this context, the term "about" or "approximately" means within 80% to 120%, alternatively within 90% to 110%, including within 95% to 105%, of a stated value or range.

若在本文內將任何術語稱為「如本文中所提及」，則其意指該術語可在本文中之任何處提及。If any term is referred to herein as "as referred to herein", it means that the term may be referred to anywhere in the document.

在本發明之上下文中，術語「GPC3」係指磷脂醯肌醇蛋白聚醣。GPC3係藉由醣基磷脂醯肌醇錨附接至細胞膜之癌胚硫酸乙醯肝素醣蛋白(Filmus J, Capurro M. Glypican-3: a marker and a therapeutic target in hepatocellular carcinoma. FEBS J. 2013年五月；280(10):2471-6)。GPC3含有580個胺基酸且具有70 kDA之分子量。兩個HS鏈附接在C-末端部分附近。藉由弗林蛋白酶(furin)在Arg358-Cys359鍵處處理單鏈GPC3以生成由藉由二硫鍵連接之40-kDa N-末端亞單元及30-kDa C-末端亞單元組成之成熟GPC3 (De Cat B, Muyldermans SY, Coomans C, Degeest G, Vanderschueren B, Creemers J, Biemar F, Peers B, David G. Processing by proprotein convertases is required for glypican-3 modulation of cell survival, Wnt signaling, and gastrulation movements.J Cell Biol. 2003年11月10日；10;163(3):625-35)。特定結合片段例如Fab、Fab'、F(ab')2及單鏈特異性結合抗體通常係片段。In the context of the present invention, the term "GPC3" refers to phosphatidylinositol protein glycan. GPC3 is an oncofetal heparan sulfate glycoprotein attached to the cell membrane via glycosylphosphatidylinositol anchors (Filmus J, Capurro M. Glypican-3: a marker and a therapeutic target in hepatocellular carcinoma. FEBS J. 2013 May; 280(10):2471-6). GPC3 contains 580 amino acids and has a molecular weight of 70 kDA. Two HS chains are attached near the C-terminal portion. Single-chain GPC3 is processed by furin at the Arg358-Cys359 bond to generate mature GPC3 consisting of a 40-kDa N-terminal subunit and a 30-kDa C-terminal subunit linked by a disulfide bond (De Cat B, Muyldermans SY, Coomans C, Degeest G, Vanderschueren B, Creemers J, Biemar F, Peers B, David G. Processing by proprotein convertases is required for glypican-3 modulation of cell survival, Wnt signaling, and gastrulation movements. J Cell Biol. 2003 Nov 10;10;163(3):625-35). Specific binding fragments such as Fab, Fab', F(ab')2 and single-chain specific binding antibodies are generally fragments.

人類GPC3蛋白質由基因GPC3 (NCBI Gene ID 2719)編碼。GPC3之同義詞係SGB、DGSX、MXR7、SDYS、SGBS、OCI-5、SGBS1及GTR2-2。GPC3蛋白質包括人類、鼠類、食蟹猴及其他哺乳動物以及非哺乳類同系物。The human GPC3 protein is encoded by the gene GPC3 (NCBI Gene ID 2719). Synonyms for GPC3 are SGB, DGSX, MXR7, SDYS, SGBS, OCI-5, SGBS1, and GTR2-2. GPC3 proteins include human, mouse, cynomolgus monkey, and other mammalian and non-mammalian homologs.

人類GPC3 (SEQ ID NO: 321)： QPPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINVDDMVNELFDSLFPVIYTQLMNPGLPDSALDINECLRGARRDLKVFGNFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILSLEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNAGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKVLKVAHVEHEETLSSRRRELIQKLKSFISFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGMKNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSMPKGRVLDKNLDEEGFESGDCGDDEDECIGGSGDGMIKVKNQLRFLAELAYDLDVDDAPGNSQQATPKDNEISTFHNLGNVHSPLKLLTSMAISVVCFFFLVH 術語「抗-GPC3抗體」或「抗-磷脂醯肌醇蛋白聚醣抗體」及「結合至磷脂醯肌醇蛋白聚醣之抗體」或「結合至GPC3之抗體」在本文中同義地使用且係指能夠結合GPC3 (較佳地具有足夠親和力)之抗體，從而使得該抗體可用作靶向GPC3中之診斷劑及/或治療劑。Human GPC3 (SEQ ID NO: 321): QPPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINV DDMVNELFDSLFPVIYTQLMNPGLPDSALDINECLRGARRDLKVFGNFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILS LEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNAGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKVLKVAHVEHEETLSSRRRELIQKLKSFISFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGM KNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSMPKGRVLDKNLDEEGFESGDCGDDEDECIGGSGDGMIKVKNQLRFLAELAYDLDVDDAPGNSQQATPKDNEISTFHNLGNVHSPLKLLTSMAISVVCFFFLVHThe terms "anti-GPC3 antibody" or "anti-GPC3 antibody" and "antibody that binds to GPC3" or "antibody that binds to GPC3" are used synonymously herein and refer to an antibody that is capable of binding to GPC3, preferably with sufficient affinity, such that the antibody can be used as a diagnostic and/or therapeutic agent in targeting GPC3.

「考曲妥珠單抗」(CAS註冊號1365267-33-9)係人類化單株抗GPC3抗體。其製備、分離及純化已闡述於(例如) US7,919,086中。考曲妥珠單抗(遍及本說明書亦稱為「TPP-14890」)包括如SEQ ID NO. 57之重鏈蛋白質序列及如SEQ ID NO.58之輕鏈蛋白質序列。另外，考曲妥珠單抗包括至少如SEQ ID NO. 42、SEQ ID NO. 43及SEQ ID NO. 44之三個CDR重鏈序列以及如SEQ ID NO. 46、SEQ ID NO. 47及SEQ ID NO. 48之三個CDR輕鏈序列： 考曲妥珠單抗CDR重鏈序列 SEQ ID NO. 42: DYEMH SEQ ID NO. 43: ALDPKTGDTAYSQKFKG SEQ ID NO. 44: FYSYTY 考曲妥珠單抗CDR輕鏈序列 SEQ ID NO. 46: RSSQSLVHSNRNTYLH SEQ ID NO. 47: KVSNRFS SEQ ID NO. 48: SQNTHVPPT 術語「多肽」及「蛋白質」在本文中可互換使用以指胺基酸殘基之聚合物。該等術語適用於其中一或多個胺基酸殘基係相應天然胺基酸之人工化學模擬物之胺基酸聚合物以及天然胺基酸聚合物及非天然胺基酸聚合物。除非另外指示，否則特定多肽序列亦隱含地涵蓋其保守修飾變體。"Trastuzumab clopidogrel" (CAS registration number 1365267-33-9) is a humanized monoclonal anti-GPC3 antibody. Its preparation, isolation and purification have been described in, for example, US7,919,086. Trastuzumab clopidogrel (also referred to as "TPP-14890" throughout this specification) includes a heavy chain protein sequence as SEQ ID NO. 57 and a light chain protein sequence as SEQ ID NO. 58. In addition, trastuzumab includes at least three CDR heavy chain sequences such as SEQ ID NO. 42, SEQ ID NO. 43 and SEQ ID NO. 44 and three CDR light chain sequences such as SEQ ID NO. 46, SEQ ID NO. 47 and SEQ ID NO. 48: Trastuzumab CDR heavy chain sequence SEQ ID NO. 42: DYEMH SEQ ID NO. 43: ALDPKTGDTAYSQKFKG SEQ ID NO. 44: FYSYTY Trastuzumab CDR light chain sequence SEQ ID NO. 46: RSSQSLVHSNRNTYLH SEQ ID NO. 47: KVSNRFS SEQ ID NO. 48: SQNTHVPPT The terms "polypeptide" and "protein" are used interchangeably herein to refer to polymers of amino acid residues. These terms apply to amino acid polymers in which one or more amino acid residues are artificial chemical mimetics of the corresponding natural amino acids as well as natural amino acid polymers and non-natural amino acid polymers. Unless otherwise indicated, a particular polypeptide sequence also implicitly encompasses conservatively modified variants thereof.

胺基酸在本文中可藉由其通常習知之三個字母符號或由IUPAC-IUB生化命名委員會(IUPAC-IUB Biochemical Nomenclature Commission)推薦之一個字母符號來提及。同樣，核苷酸可藉由其通常接受之單字母代碼來提及。Amino acids may be referred to herein by either their commonly known three letter symbols or by the one letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Likewise, nucleotides may be referred to by their commonly accepted single-letter codes.

如本文中所使用，術語「抗體」意欲指包含(但不限於)全長抗體及單價抗體之免疫球蛋白分子。「全長抗體」較佳地包括通常藉由二硫鍵互相連接之四個多肽鏈，即兩個重(H)鏈及兩個輕(L)鏈。每一重鏈包括重鏈可變區(在本文中縮寫為VH)及重鏈恆定區。重鏈恆定區包括(例如)三個結構域：CH1、CH2及CH3。每一輕鏈包括輕鏈可變區(在本文中縮寫為VL)及輕鏈恆定區。輕鏈恆定區包括一個結構域(CL)。VH及VL區可進一步細分成超變區(稱為互補決定區(CDR))及更保守之區(稱為框架區(FR))，二者間雜排列。每一VH及VL通常係由三個CDR及最多四個FR構成，其自胺基末端至羧基末端按下列順序排列：FR1、CDR1、FR2、CDR2、FR3、CDR3、FR4。本文所用之「單價抗體」較佳地包括通常藉由二硫鍵互相連接之三個多肽鏈，即兩個重(H)鏈及一個輕(L)鏈。其中一個重鏈包括重鏈可變區(在本文中縮寫為VH)及重鏈恆定區。重鏈恆定區包括(例如)三個結構域：CH1、CH2及CH3。另一個重鏈僅包括重鏈恆定區。輕鏈包括輕鏈可變區(在本文中縮寫為VL)及輕鏈恆定區。輕鏈恆定區包括一個結構域(CL)。該VH及VL區可進一步細分成超變區(稱為互補決定區(CDR))及更保守之區(稱為框架區(FR))，二者間雜排列。每一VH及VL通常係由三個CDR及最多四個FR構成，其自胺基末端至羧基末端按下列順序排列：FR1、CDR1、FR2、CDR2、FR3、CDR3、FR4。As used herein, the term "antibody" is intended to refer to immunoglobulin molecules including, but not limited to, full-length antibodies and monovalent antibodies. "Full-length antibodies" preferably include four polypeptide chains, i.e., two heavy (H) chains and two light (L) chains, which are usually interconnected by disulfide bonds. Each heavy chain includes a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region includes, for example, three domains: CH1, CH2, and CH3. Each light chain includes a light chain variable region (abbreviated herein as VL) and a light chain constant region. The light chain constant region includes one domain (CL). The VH and VL regions can be further divided into hypervariable regions, called complement-determining regions (CDRs), and more conserved regions, called framework regions (FRs), which are interspersed. Each VH and VL is usually composed of three CDRs and up to four FRs, which are arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The "monovalent antibody" used herein preferably includes three polypeptide chains, i.e., two heavy (H) chains and one light (L) chain, which are usually connected to each other by disulfide bonds. One of the heavy chains includes a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region includes, for example, three domains: CH1, CH2, and CH3. The other heavy chain includes only the heavy chain constant region. The light chain includes the light chain variable region (abbreviated herein as VL) and the light chain constant region. The light chain constant region includes a structural domain (CL). The VH and VL regions can be further divided into hypervariable regions (called complementation determining regions (CDRs)) and more conserved regions (called framework regions (FRs)), which are interspersed. Each VH and VL is usually composed of three CDRs and up to four FRs, which are arranged from the amino terminus to the carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.

如本文中所使用，術語「互補決定區域」(CDR；例如CDR1、CDR2及CDR3)係指其存在對抗原結合為必要之抗體可變結構域之胺基酸殘基。每一可變結構域通常具有鑑別為CDR1、CDR2及CDR3之三個CDR區域。每一互補決定區可包括來自由Kabat定義之「互補決定區域」之胺基酸殘基(例如約輕鏈可變結構域中之殘基24-34 (L1)、50-56 (L2)及89-97 (L3)以及重鏈可變結構域中之31-35 (H1)、50-65 (H2)及95-102 (H3)；(Kabat等人，Sequences of Proteins of Immunolological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991))及/或來自「超變環」之彼等殘基(例如約輕鏈可變結構域中之殘基26-32 (L1)、50-52 (L2)及91-96 (L3)以及重鏈可變結構域中之26- 32 (H1)、53-55 (H2)及96-101 (H3) (Chothia及Lesk; J Mol Biol 196: 901-917 (1987))。在一些情況下，互補決定區可包含來自根據Kabat定義之CDR區域及超變環二者之胺基酸。As used herein, the term "complementary determining region" (CDR; e.g., CDR1, CDR2, and CDR3) refers to the amino acid residues of an antibody variable domain whose presence is essential for antigen binding. Each variable domain typically has three CDR regions identified as CDR1, CDR2, and CDR3. Each complementation determining region may include amino acid residues from the "complementation determining region" defined by Kabat (e.g., residues 24-34 (L1), 50-56 (L2), and 89-97 (L3) in the light chain variable domain and 31-35 (H1), 50-65 (H2), and 95-102 (H3) in the heavy chain variable domain; (Kabat et al., Sequences of Proteins of Immunolological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)) and/or those residues from the "hypervariable loop" (e.g., residues 26-32 (L1), 50-52 (L2), and 91-96 in the light chain variable domain). (L3) and 26-32 (H1), 53-55 (H2) and 96-101 (H3) in the heavy chain variable domain (Chothia and Lesk; J Mol Biol 196: 901-917 (1987)). In some cases, the complementarity determining region may include amino acids from both the CDR region and the hypervariable loop according to the Kabat definition.

端視完整抗體重鏈中恆定結構域之胺基酸序列，可將完整抗體歸類為不同「種類」。存在5大類完整抗體：IgA、IgD、IgE、IgG及IgM，且該等種類中之若干可進一步分成「子類」(同型)，例如IgG1、IgG2、IgG3、IgG4、IgA1及IgA2。用於本發明之免疫球蛋白之較佳種類係IgG。在特定實施例中，本發明抗體係IgG1、IgG2、IgG3或IgG4抗體，更具體而言IgG1抗體。不同同型具有不同效應子功能。人類輕鏈分類為kappa (K)及lambda (λ)輕鏈。在輕鏈及重鏈內，可變區及恆定區由約12個或更多個胺基酸之「J」區連接，且該重鏈亦包括約10或更多個胺基酸之「D」區。通常參見Fundamental Immunology，第7章(Paul, W.編輯，第2版，Raven Press, N.Y. (1989))。對應於不同種類之抗體之重鏈恆定結構域分別稱為[α]、[δ]、[ε]、[γ]及[μ]。不同種類之免疫球蛋白之亞單元結構及三維構形已眾所周知。本文中使用之抗體係習知抗體及其功能片段。Depending on the amino acid sequence of the constant structural domains in the intact antibody heavy chain, intact antibodies can be classified into different "types". There are 5 major classes of intact antibodies: IgA, IgD, IgE, IgG and IgM, and some of these classes can be further divided into "subclasses" (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2. The preferred class of immunoglobulins used in the present invention is IgG. In specific embodiments, the antibodies of the present invention are IgG1, IgG2, IgG3 or IgG4 antibodies, more specifically IgG1 antibodies. Different isotypes have different effector functions. Human light chains are classified as kappa (K) and lambda (λ) light chains. In the light chain and heavy chain, the variable region and the constant region are connected by a "J" region of about 12 or more amino acids, and the heavy chain also includes a "D" region of about 10 or more amino acids. See generally Fundamental Immunology, Chapter 7 (Paul, W. ed., 2nd edition, Raven Press, N.Y. (1989)). The heavy chain constant structural domains corresponding to different types of antibodies are called [α], [δ], [ε], [γ] and [μ], respectively. The subunit structures and three-dimensional configurations of different types of immunoglobulins are well known. The antibodies used in this article are known antibodies and their functional fragments.

抗體/免疫球蛋白之「功能片段」或「抗原-結合抗體片段」特此定義為保留抗原結合區域之抗體/免疫球蛋白之片段(例如，IgG之可變區)。抗體之「抗原-結合區域」通常存在於抗體之一或多種超變區，例如CDR1、CDR2及/或CDR3區域；然而，可變「框架」區域亦可在抗原結合中發揮重要作用，例如藉由為CDR提供架構來發揮作用。較佳地，「抗原-結合區域」包括可變輕(VL)鏈之至少4至103位胺基酸殘基及可變重(VH)鏈之至少5至109位胺基酸殘基、更佳地VL之3至107位胺基酸殘基及VH之4至111位胺基酸殘基、及尤佳完整之VL及VH鏈(VL之1至109位胺基酸及VH之1至113位胺基酸；根據WO 97/08320編號)。"Functional fragments" or "antigen-binding antibody fragments" of antibodies/immunoglobulins are hereby defined as fragments of antibodies/immunoglobulins that retain the antigen-binding region (e.g., the variable region of IgG). The "antigen-binding region" of an antibody is generally found in one or more hypervariable regions of the antibody, such as the CDR1, CDR2, and/or CDR3 regions; however, variable "framework" regions may also play an important role in antigen binding, for example by providing a framework for the CDRs. Preferably, the "antigen-binding region" includes at least amino acid residues 4 to 103 of the variable light (VL) chain and at least amino acid residues 5 to 109 of the variable heavy (VH) chain, more preferably amino acid residues 3 to 107 of VL and amino acid residues 4 to 111 of VH, and most preferably complete VL and VH chains (amino acids 1 to 109 of VL and amino acids 1 to 113 of VH; according to WO 97/08320 numbering).

「功能片段」或「抗原-結合抗體片段」之非限制性實例包含Fab、Fab'、F(ab')2、Fv片段、結構域抗體 (dAb)、互補決定區(CDR)片段、單鏈抗體(scFv)、單鏈抗體片段、二價抗體、三價抗體、四價抗體、微小抗體、線性抗體(Zapata等人，Protein Eng.,8 (10): 1057-1062 (1995))；螯合重組抗體、三價抗體或雙特異性抗體、細胞內抗體、奈米抗體、小分子免疫醫藥(SMIP)、抗原結合-結構域免疫球蛋白融合蛋白、駝峰化抗體、含VHH之抗體、或突變蛋白或其衍生物、及含有足以賦予多肽特異性抗原結合之至少一部分免疫球蛋白(例如CDR序列)之多肽(只要抗體保留期望生物活性)；及自抗體片段形成之多特異性抗體(例如雙特異性及三特異性抗體) (C. A. K Borrebaeck編輯(1995) Antibody Engineering (Breakthroughs in Molecular Biology), Oxford University Press; R. Kontermann & S. Duebel編輯(2001) Antibody Engineering (Springer Laboratory Manual), Springer Verlag)。除「雙特異性」或「雙功能」抗體外之抗體應理解為其結合位點中之每一者相同。可改造F(ab’)₂或Fab以最小化或完全去除C_H1及C_L結構域之間發生之分子間二硫化物相互作用。抗體之木瓜酶消化產生兩個相同抗原結合片段，稱為「Fab」片段，其各自具有單一抗原結合位點；及殘餘「Fc「片段，其名稱反映其容易結晶之能力。胃蛋白酶處理產生具有兩個「Fv」片段之F(ab')2片段。「Fv」片段係含有完全抗原識別及結合位點之最小抗體片段。此區域由一個重鏈可變結構域與一個輕鏈可變結構域呈緊密非共價締合形式之二聚體組成。每一可變結構域之3個CDR以此構形相互作用以界定VH-VL二聚體之表面上之抗原結合位點。六個CDR共同賦予了該抗體抗原結合特異性。然而，甚至單一可變結構域(或僅包括三個抗原特異性CDR之一半Fv)具有識別及結合抗原之能力。Non-limiting examples of "functional fragments" or "antigen-binding antibody fragments" include Fab, Fab', F(ab')2, Fv fragments, domain antibodies (dAb), complementary determining region (CDR) fragments, single chain antibodies (scFv), single chain antibody fragments, bivalent antibodies, trivalent antibodies, tetravalent antibodies, miniantibodies, linear antibodies (Zapata et al., Protein Eng., 8 (10): 1057-1062). (1995)); chelated recombinant antibodies, trivalent antibodies or bispecific antibodies, intracellular antibodies, nanobodies, small molecule immunopharmaceuticals (SMIPs), antigen binding-domain immunoglobulin fusion proteins, peaked antibodies, VHH-containing antibodies, or mutant proteins or derivatives thereof, and polypeptides containing at least a portion of an immunoglobulin (e.g., a CDR sequence) sufficient to confer specific antigen binding to the polypeptide (as long as the antibody retains the desired biological activity); and multispecific antibodies (e.g., bispecific and trispecific antibodies) formed from antibody fragments (CA K Borrebaeck, ed. (1995) Antibody Engineering (Breakthroughs in Molecular Biology), Oxford University Press; R. Kontermann & S. Duebel, eds. (2001) Antibody Engineering (Springer Laboratory Manual), Springer Verlag). Antibodies other than "bispecific" or "bifunctional" antibodies are understood to be identical in each of their binding sites. F(ab')₂ or Fab can be engineered to minimize or completely eliminate intermolecular disulfide interactions that occur between the_CH1 and_CL domains. Papain digestion of antibodies produces two identical antigen-binding fragments, called "Fab" fragments, each with a single antigen-binding site; and a residual "Fc" fragment, whose name reflects its ability to crystallize readily. Pepsin treatment produces a F(ab')2 fragment with two "Fv" fragments. The "Fv" fragment is the smallest antibody fragment that contains complete antigen recognition and binding sites. This region consists of a dimer of one heavy chain variable domain in tight, non-covalent association with one light chain variable domain. The three CDRs of each variable domain interact in this configuration to define an antigen binding site on the surface of the VH-VL dimer. The six CDRs together confer antigen binding specificity to the antibody. However, even a single variable domain (or half of an Fv comprising only three antigen-specific CDRs) has the ability to recognize and bind to an antigen.

「單鏈Fv」或「sFv」或「scFv」抗體片段包括抗體之VH及VL結構域，其中該等結構域係以單一多肽鏈存在。"Single-chain Fv" or "sFv" or "scFv" antibody fragments comprise the VH and VL domains of an antibody, wherein the domains are present in a single polypeptide chain.

較佳地，Fv多肽進一步包含VH結構域與VL結構域之間之多肽連接體，其使Fv能形成用於抗原結合之期望結構。關於Fvs之綜述，參見Pluckthun，The Pharmacology of Monoclonal Antibodies，第113卷，Rosenburg及Moore編輯，Springer-Verlag, New York，第269-315頁(1994)。Preferably, the Fv polypeptide further comprises a polypeptide linker between the VH domain and the VL domain that enables the Fv to form a desired structure for antigen binding. For a review of Fvs, see Pluckthun, The Pharmacology of Monoclonal Antibodies, Vol. 113, Rosenburg and Moore, eds., Springer-Verlag, New York, pp. 269-315 (1994).

Fab片段亦含有輕鏈恆定結構域及重鏈之第一恆定結構域(CH1)。Fab片段與Fab'片段的不同之處在於重鏈CH1結構域之羧基末端增加了數個殘基，包括一或多個來自抗體鉸鏈區之半胱胺酸殘基。在本文中，Fab'-SH係恆定結構域中之半胱胺酸殘基具有游離硫醇基之Fab'之名稱。F(ab′)2抗體片段最初係作為在其間具有鉸鏈半胱胺酸殘基之Fab′片段對產生。Fab fragments also contain the light chain homeostatic domain and the first homeostatic domain (CH1) of the heavy chain. Fab fragments differ from Fab' fragments in that the carboxyl terminus of the heavy chain CH1 domain has several additional residues, including one or more cysteine residues from the antibody hinge region. In this article, Fab'-SH is the designation for Fab' in which the cysteine residues in the homeostatic domains have free thiol groups. F(ab')2 antibody fragments were originally produced as pairs of Fab' fragments with a hinge cysteine residue between them.

術語「突變蛋白」或「變體」可互換使用且係指含有在可變區或等效於可變區之部分中之至少一種胺基酸取代、缺失或插入之抗體或抗原結合片段，前提係突變蛋白或變體保留期望結合親和力或生物活性。本發明中考慮之抗體或抗原結合抗體片段之變體係其中抗體或抗原結合抗體片段之結合活性得以維持的分子。The terms "mutant" or "variant" are used interchangeably and refer to an antibody or antigen-binding fragment containing at least one amino acid substitution, deletion or insertion in a variable region or a portion equivalent to a variable region, provided that the mutant protein or variant retains the desired binding affinity or biological activity. Variants of antibodies or antigen-binding antibody fragments contemplated in the present invention are molecules in which the binding activity of the antibody or antigen-binding antibody fragment is maintained.

「人類化抗體」通常定義為：(I)衍生自非人類來源(例如，具有異源免疫系統之轉基因小鼠)之抗體，該抗體係基於人類種系序列；或(II) CDR接枝抗體，其中可變區之CDR係來自非人類來源，而一或多種框架區及/或可變區之CDR序列之部分係來自人類來源且通常恆定區(若存在)係來自人類來源。"Humanized antibodies" are generally defined as: (I) antibodies derived from non-human sources (e.g., transgenic mice with a heterologous immune system) that are based on human germline sequences; or (II) CDR-grafted antibodies in which the CDRs of the variable regions are from non-human sources, while one or more framework regions and/or portions of the CDR sequences of the variable regions are from human sources and typically the invariant regions (if present) are from human sources.

術語「人類化抗體」由此包含藉由來自已知特異性之非人類抗體之一或多個CDR之至少部分代替以及(視需要)藉由部分框架區代替及序列變化來改變人類抗體之重鏈或輕鏈或二者中之可變區所得之抗體。換言之，將來自已知特異性之非人類抗體(例如小鼠、大鼠、兔或非人類靈長類動物抗體)之一或多種「供體」CDR接枝至人類重或輕鏈框架區所得之抗體在本文中稱為「人類化抗體」。為使一個可變結構域之抗原結合能力轉移至另一者，可能不必用來自供體可變結構域之完整CDR來代替所有CDR。相反，可僅需要轉移維持靶結合位點之活性需要之彼等殘基。The term "humanized antibody" thus includes antibodies obtained by at least partial replacement of one or more CDRs from non-human antibodies of known specificity and, if necessary, by partial framework region replacement and sequence variation to alter the variable regions of the heavy or light chain or both of human antibodies. In other words, antibodies obtained by grafting one or more "donor" CDRs from non-human antibodies of known specificity (e.g., mouse, rat, rabbit or non-human primate antibodies) to human heavy or light chain framework regions are referred to herein as "humanized antibodies". In order to transfer the antigen binding ability of one variable domain to another, it may not be necessary to replace all CDRs with complete CDRs from the donor variable domain. Instead, it may only be necessary to transfer those residues required to maintain the activity of the target binding site.

人類化抗體之重鏈或輕鏈或二者中之可變區內之框架區(FR)可僅包括具有人類來源之殘基，在此情形下人類化抗體之該等框架區稱為「完全人類框架區」。包括人類及供體框架殘基之混合物之人類框架區在本文中稱為「部分人類框架區」。此外，人類化抗體可包括受體抗體或供體抗體中不存在之殘基。進行該等修飾以進一步改良抗體性能(例如，以獲得期望親和力)。The framework regions (FR) within the variable regions of the heavy chain or light chain or both of the humanized antibodies may include only residues of human origin, in which case the framework regions of the humanized antibodies are referred to as "fully human framework regions". Human framework regions that include a mixture of human and donor framework residues are referred to herein as "partially human framework regions". In addition, humanized antibodies may include residues that are not present in the acceptor antibody or the donor antibody. Such modifications are performed to further improve antibody performance (e.g., to obtain a desired affinity).

一般而言，人類化抗體由此包括至少一個且通常兩個可變區，其中全部或部分CDR對應於非人類免疫球蛋白之彼等CDR，且全部或實質上全部之FR係人類免疫球蛋白序列之彼等FR。人類化抗體視情況亦包括免疫球蛋白恆定區(Fc) (通常為人類免疫球蛋白恆定區)之至少一部分。其他細節參見Jones等人，Nature 331:522-25(1986)；Riechmann等人，Nature 332:323-27(1988)；及Presta, Curr. Op. Struct. Biol. 2:593-96(1992)。In general, humanized antibodies thus include at least one and usually two variable regions, in which all or part of the CDRs correspond to those of a non-human immunoglobulin, and all or substantially all of the FRs are those of a human immunoglobulin sequence. Humanized antibodies may also include at least a portion of an immunoglobulin constant region (Fc) (usually a human immunoglobulin constant region). For further details, see Jones et al., Nature 331:522-25 (1986); Riechmann et al., Nature 332:323-27 (1988); and Presta, Curr. Op. Struct. Biol. 2:593-96 (1992).

「人類抗體」或「完全人類抗體」包括人類衍生之CDR，亦即具有人類來源之CDR。「人類」抗體特此定義為嵌合或「人類化」且不來自(整個或部分)非人類物種之抗體。人類抗體或功能抗體片段可衍生自人類或可為合成人類抗體。「合成人類抗體」在本文中定義為具有在電腦上(整個或部分)衍生自基於已知人類抗體序列之分析之合成序列之序列的抗體。舉例而言，藉由分析人類抗體或抗體片段序列之資料庫及利用自其獲得之數據來設計胺基酸序列可達成人類抗體序列或其片段之電腦上設計。人類抗體或功能抗體片段之另一實例係由自具有人類來源之抗體序列庫(亦即，該庫係基於自人類天然來源獲取之抗體)分離之核酸編碼所獲得者。"Human antibodies" or "fully human antibodies" include human-derived CDRs, i.e., CDRs of human origin. "Human" antibodies are hereby defined as antibodies that are chimeric or "humanized" and are not derived (in whole or in part) from a non-human species. Human antibodies or functional antibody fragments may be derived from humans or may be synthetic human antibodies. "Synthetic human antibodies" are defined herein as antibodies having a sequence that is derived in silico (in whole or in part) from a synthetic sequence based on an analysis of known human antibody sequences. For example, in silico design of human antibody sequences or fragments thereof may be achieved by analyzing a database of human antibody or antibody fragment sequences and using the data obtained therefrom to design the amino acid sequence. Another example of a human antibody or functional antibody fragment is one obtained from a nucleic acid encoding an antibody sequence isolated from a library of antibody sequences having human origin (ie, the library is based on antibodies obtained from natural sources in humans).

如本文所用，術語「單株抗體」係指自實質上同源之抗體群體獲得之抗體，即，除可能之突變(例如，極少量存在之天然突變)外，構成該群體之個別抗體相同。因此，適用於「單株」指示並非獨立抗體混合物之抗體性質。與通常包含針對不同決定子(表位)之不同抗體之多株抗體製劑相比，單株抗體製劑之每一單株抗體針對抗原上之單一決定子。除特異性外，單株抗體製劑之優勢亦在於其通常不受其他免疫球蛋白污染。術語「單株」不應視為需要藉由任何特定方法產生抗體。舉例而言，欲使用之單株抗體可藉由首次由Kohler等人，Nature, 256: 495 [1975闡述之雜交瘤方法製得，或可藉由重組DNA方法製得(參見例如美國專利第4,816,567號)。「單株抗體」亦可為(例如)重組、嵌合、人類化、人類、Human Engineered™抗體或抗體片段。As used herein, the term "monoclonal antibody" refers to an antibody obtained from a substantially homogeneous antibody population, i.e., the individual antibodies constituting the population are identical except for possible mutations (e.g., natural mutations that occur in very small amounts). Thus, the term "monoclonal" is used to indicate the nature of an antibody that is not a mixture of independent antibodies. In contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on the antigen. In addition to specificity, the advantage of monoclonal antibody preparations is that they are generally not contaminated by other immunoglobulins. The term "monoclonal" should not be construed as requiring the production of antibodies by any particular method. For example, the monoclonal antibody to be used can be made by the hybridoma method first described by Kohler et al., Nature, 256: 495 [1975], or can be made by recombinant DNA methods (see, e.g., U.S. Patent No. 4,816,567). A "monoclonal antibody" can also be, for example, a recombinant, chimeric, humanized, human, Human Engineered™ antibody, or an antibody fragment.

「經分離」抗體係自表現其之細胞之成分鑑別及分離之抗體。細胞之污染成分係將干擾抗體之診斷或治療用途之材料，且可包含酶、激素及其他蛋白質性溶質或非蛋白質性溶質。An "isolated" antibody is one that has been identified and separated from a component of the cell in which it is expressed. Contaminating components of the cell are materials that would interfere with the diagnostic or therapeutic use of the antibody and may include enzymes, hormones, and other proteinaceous or nonproteinaceous solutes.

「經分離」核酸係已自其天然環境之成分鑑別及分離之核酸。經分離核酸包括通常含有核酸分子之細胞中所含之核酸分子，但該核酸分子存在於染色體外或存在於與其天然染色體位置不同之染色體位置處。An "isolated" nucleic acid is one that has been identified and separated from a component of its natural environment. Isolated nucleic acids include nucleic acid molecules contained in cells that normally contain nucleic acid molecules, but where the nucleic acid molecule is present extrachromosomally or at a chromosomal location that is different from its natural chromosomal location.

如本文中所使用，抗體「特異性結合至」目標抗原、對/針對目標抗原「具有特異性」或「特異性識別」目標抗原(例如GPC3)係：以足夠親和力結合抗原-靶之抗原，從而使抗體可用作靶向表現該抗原之細胞或組織中之治療劑；或以足夠親和力結合抗原結合多肽靶，從而使抗體可用作逆轉藥劑以中和此抗原結合多肽(例如抗原結合抗體)之治療性活性且與其他蛋白質不顯著交叉反應或與除上文所提及靶之直向同源物及變體(例如突變體形式、剪接變體或以蛋白水解方式截短之形式)外之蛋白質不顯著交叉反應。術語「特異性識別」或「特異性結合至」特定多肽或本文所用之特定多肽靶上之表位或對/針對其「具有特異性」可(舉例而言)由抗體或其抗原結合片段展現，該抗體或其抗原結合片段具有小於約10^-4M、替代地小於約10^-5M、替代地小於約10^-6M、替代地小於約10^-7M、替代地小於約10^-8M、替代地小於約10^-9M、替代地小於約10^-10M、替代地小於約10^-11M、替代地小於約10^-12M或更小之抗原之單價解離常數(K_D)。若該抗體能夠區分該抗體與一或多種參考抗原，則抗體「特異性結合至」抗原、對/針對抗原「具有特異性」或「特異性識別」抗原。在其最一般形式中，「特異性結合」、「特異性結合至」、「對/針對……具有特異性」或「特異性識別」係指抗體區分目標抗原與無關抗原之能力，如(舉例而言)根據下列方法中之一者所測定。該等方法包括(但不限於)表面電漿共振(SPR)、西方墨點(Western blot)、ELISA測試、RIA測試、ECL測試、IRMA測試及肽掃描。舉例而言，可實施標準ELISA分析。可藉由標準顯色(例如，使用辣根過氧化物酶之二級抗體及使用過氧化氫之四甲基聯苯胺)來實施評分。藉由光學密度(舉例而言，在450 nm下)來評分某些孔中之反應。典型背景(=陰性反應)可為0.1 OD；典型陽性反應可為1 OD。此意指陽性/陰性差異大於5倍、10倍、50倍，且較佳地大於100倍。通常，藉由使用約三至五種無關抗原(例如奶粉、BSA、轉鐵蛋白或諸如此類)而非單一參考抗原來實施結合特異性之測定。As used herein, an antibody "specifically binds to,""is specific for," or "specifically recognizes" a target antigen (e.g., GPC3) by binding to the antigen-target antigen with sufficient affinity to allow the antibody to be used as a therapeutic agent targeted to cells or tissues expressing the antigen; or by binding to an antigen-binding polypeptide target with sufficient affinity to allow the antibody to be used as a reverse agent to neutralize the therapeutic activity of such an antigen-binding polypeptide (e.g., an antigen-binding antibody) without significantly cross-reacting with other proteins or with proteins other than orthologs and variants (e.g., mutant forms, splice variants, or proteolytically truncated forms) of the targets mentioned above. The term "specifically recognizes" or "specifically binds to" or is "specific for" a particular polypeptide, or an epitope on a particular polypeptide target as used herein, may be exhibited, for example, by an antibody or antigen-binding fragment thereof having a monovalent dissociation constant (KD) for^{the antigen of less than about 10-4 M, alternatively less than about 10-5 M, alternatively less than about 10-6 M, alternatively less than about 10-7M,alternativelyless} than about^10-8 M, alternatively less than about^10-9 M, alternatively less than about^10-10 M, alternatively less than about^{10-11 M, alternatively less than about 10-12}_M , or less. An antibody "specifically binds to,""is specific for," or "specifically recognizes" an antigen if the antibody is able to distinguish between the antibody and one or more reference antigens. In its most general form, "specifically binds,""specifically binds to,""is specific for," or "specifically recognizes" refers to the ability of an antibody to distinguish a target antigen from an unrelated antigen as determined, for example, according to one of the following methods. Such methods include, but are not limited to, surface plasmon resonance (SPR), Western blot, ELISA assay, RIA assay, ECL assay, IRMA assay, and peptide scanning. For example, a standard ELISA assay can be performed. Scoring can be performed by standard color development (e.g., secondary antibodies using horseradish peroxidase and tetramethylbenzidine using hydrogen peroxide). The reactions in certain wells are scored by optical density (e.g., at 450 nm). A typical background (=negative reaction) may be 0.1 OD; a typical positive reaction may be 1 OD. This means that the positive/negative difference is greater than 5-fold, 10-fold, 50-fold, and preferably greater than 100-fold. Typically, the determination of binding specificity is performed by using about three to five unrelated antigens (e.g., milk powder, BSA, transferrin or the like) instead of a single reference antigen.

「結合親和力」或「親和力「係指分子之單一結合位點與其結合配偶體之間之非共價相互作用之總強度。除非另外指明，否則如本文所使用，「結合親和力」係指反映結合對之成員(例如，抗體與抗原)之間之1:1相互作用之固有結合親和力。可使用業內已熟知之技術來測定抗體或其片段針對靶之親和力，例如藉由ELISA、等溫滴定量熱法(ITC)、表面電漿共振(SPR)、流式細胞術或螢光偏振分析。通常使用解離常數「K_D」以闡述分子(例如抗體)與其結合配偶體(例如抗原)之間之親和力，亦即配體與特定蛋白質之結合之緊密程度。配體-蛋白質親和力受兩種分子之間之非共價分子間相互作用影響。可藉由業內已知之常用方法(包含闡述於本文中者)來量測親和力。較佳地以解離常數K_D形式來提供親和力，替代地可以EC50值形式來提供親和力。在一實施例中，藉由使用表面電漿共振分析Biacore T200儀器(Cytiva)來量測本發明之「K_D」或「K_D值」。其他適宜裝置係Biacore X100、Biacore S200、Biacore 8K、Biacore 8K+、Biacore 4000 SPR (Cytiva)、Alto Digital SPR (Nicoya Lifesciences Inc.)或SIERRA SPR-32 PRO (Bruker Corporation)。"Binding affinity" or "affinity" refers to the overall strength of the non-covalent interactions between a single binding site of a molecule and its binding partner. Unless otherwise indicated, as used herein, "binding affinity" refers to the intrinsic binding affinity that reflects a 1:1 interaction between members of a binding pair (e.g., antibody and antigen). The affinity of an antibody or fragment thereof for a target can be determined using techniques well known in the art, such as by ELISA, isothermal titration calorimetry (ITC), surface plasmon resonance (SPR), flow cytometry, or fluorescence polarization analysis. The dissociation constant "_KD " is often used to describe the affinity between a molecule (e.g., an antibody) and its binding partner (e.g., an antigen), that is, how tightly a ligand binds to a particular protein. Ligand-protein affinity is affected by non-covalent intermolecular interactions between the two molecules. Affinity can be measured by conventional methods known in the art, including those described herein. Affinity is preferably provided in the form of a dissociation constant,_KD , alternatively, affinity can be provided in the form of an EC50 value. In one embodiment, the "KD_" or "KD value" of the present invention is measured by using a surface plasmon resonance analysis Biacore T200 instrument (_Cytiva ). Other suitable devices are Biacore X100, Biacore S200, Biacore 8K, Biacore 8K+, Biacore 4000 SPR (Cytiva), Alto Digital SPR (Nicoya Lifesciences Inc.), or SIERRA SPR-32 PRO (Bruker Corporation).

「半最大有效濃度」(EC50)係指藥物、調節劑、抗體、片段、結合物或分子在指定培育時間之後誘導介於基線及最大值間之半反應的濃度。在親和力之上下文中，EC50由此反映半最大結合需要之結合物(例如抗體)之濃度。若可藉由闡述施加之藥物、抗體、片段、結合物或分子濃度與信號間之關係之劑量–反應曲線之數學建模(例如，非線性回歸)來確定拐點，則可確定EC50。舉例而言，若劑量–反應曲線遵循S形曲線，則可確定EC50。若反應係抑制，則EC50稱為「半最大抑制性濃度」(IC50)。"Half-maximal effective concentration" (EC50) refers to the concentration of a drug, modulator, antibody, fragment, conjugate or molecule that induces a response halfway between baseline and maximum after a specified incubation time. In the context of affinity, the EC50 thus reflects the concentration of the binder (e.g., antibody) required for half-maximal binding. The EC50 can be determined if the inflection point can be determined by mathematical modeling (e.g., nonlinear regression) of the dose-response curve that describes the relationship between the applied drug, antibody, fragment, conjugate or molecule concentration and the signal. For example, if the dose-response curve follows a sigmoidal curve, the EC50 can be determined. If the response is inhibitory, the EC50 is called the "half-maximal inhibitory concentration" (IC50).

如本文中所使用，術語「表位」包含能夠具體結合至免疫球蛋白或T-細胞受體之任何蛋白質決定子。表位決定子通常由分子之化學活性表面基團(例如胺基酸或糖側鏈或其組合)組成，並且通常具有特定三維結構特性以及特定電荷特性。As used herein, the term "epitope" includes any protein determinant capable of specific binding to an immunoglobulin or T-cell receptor. Epitope determinants usually consist of chemically active surface groups of molecules (such as amino acids or sugar side chains or combinations thereof) and usually have specific three-dimensional structural characteristics as well as specific charge characteristics.

術語「成熟抗體」或「成熟抗原結合片段」(例如成熟Fab變體或「最佳化」變體)包含對既定抗原(例如靶蛋白質之細胞外結構域)展現較強結合(以增加之親和力結合)之抗體或抗體片段之衍生物。成熟係鑑別引起此親和力增加之抗體或抗體片段之6個CDR內之少量突變的過程。成熟方法係將突變引入抗體中且篩選以鑑別經改良結合物之分子生物學方法之組合。The term "mature antibody" or "mature antigen-binding fragment" (e.g., mature Fab variants or "optimized" variants) includes derivatives of antibodies or antibody fragments that exhibit stronger binding (bind with increased affinity) to a given antigen (e.g., the extracellular domain of a target protein). Maturation is the process of identifying small mutations within the six CDRs of an antibody or antibody fragment that result in this increased affinity. Maturation methods are a combination of molecular biology methods to introduce mutations into antibodies and screen to identify improved binders.

術語「抗體結合物」係指包括至少一種抗體部分及一或多種其他分子或部分之結合物。一或多種其他分子或部分可選自(但不限於)藥物、螯合劑、放射性元素、細胞毒性劑、其他抗體或抗原結合片段。The term "antibody conjugate" refers to a conjugate comprising at least one antibody portion and one or more other molecules or moieties. The one or more other molecules or moieties may be selected from (but not limited to) drugs, chelating agents, radioactive elements, cytotoxic agents, other antibodies or antigen-binding fragments.

術語「抗體藥物結合物」(「ADC」)係指包括至少一種藥物部分之抗體結合物。The term "antibody-drug conjugate" ("ADC") refers to an antibody conjugate that includes at least one drug moiety.

術語「種系化」係指使用預突變種系基因中存在之殘基來代替抗體之可變結構域中之殘基，以減小免疫原性之潛能。The term "germlining" refers to the use of residues present in the pre-mutated germline genes to replace residues in the variable domains of antibodies to reduce the potential for immunogenicity.

「抗體依賴性細胞介導之細胞毒性」或「ADCC」係指以下細胞毒性形式：其中結合存在於某些細胞毒性細胞(例如NK細胞、嗜中性球及巨噬球)上之Fc γ受體(FcγR)之經分泌Ig使該等細胞毒性效應細胞能夠特異性地結合帶抗原靶細胞且隨後利用細胞毒素殺滅該靶細胞。為評價所關注抗體之ADCC活性，可實施活體外ADCC分析，例如闡述於美國專利第5,500,362號或第5,821,337號或美國專利第6,737,056號(Presta)中者。用於該等分析之可用效應細胞包含PBMC及NK細胞。"Antibody-dependent cell-mediated cytotoxicity" or "ADCC" refers to a form of cytotoxicity in which secreted Ig that binds to Fc gamma receptors (FcγRs) present on certain cytotoxic cells (e.g., NK cells, neutrophils, and macrophages) enables these cytotoxic effector cells to specifically bind to antigen-bearing target cells and subsequently kill the target cells using cytotoxins. To assess the ADCC activity of an antibody of interest, an in vitro ADCC assay, such as that described in U.S. Patent Nos. 5,500,362 or 5,821,337 or U.S. Patent No. 6,737,056 (Presta) may be performed. Useful effector cells for these analyses include PBMCs and NK cells.

「補體依賴性細胞毒性」或「CDC」係指靶細胞在補體存在下之溶胞。典型補體途徑之活化係藉由補體系統之第一組分(C1q)結合至與其同源抗原結合之抗體(或適當子類)來引發。為評價補體活化，可實施CDC分析，例如Gazzano-Santoro等人，J. Immunol. Methods 202:163 (1996)中所闡述。具有經改變Fc區胺基酸序列之多肽變體(具有變體Fc區之多肽)及增加或降低之C1q結合闡述於(例如)美國專利第6,194,551 Bl號及WO 1999/51642號中。"Complement-dependent cytotoxicity" or "CDC" refers to the lysis of target cells in the presence of complement. Activation of the canonical complement pathway is initiated by binding of the first component of the complement system (C1q) to an antibody (or appropriate subclass) bound to its cognate antigen. To assess complement activation, a CDC assay, such as that described in Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996), can be performed. Polypeptide variants having altered Fc region amino acid sequences (polypeptides having variant Fc regions) and increased or decreased C1q binding are described, for example, in U.S. Patent No. 6,194,551 Bl and WO 1999/51642.

關於參考多核苷酸或多肽序列之「序列一致性百分比(%)」定義為在比對序列且(視需要)引入間隙以達成最大百分比序列一致性之後分別與參考多核苷酸或多肽序列中之核酸或胺基酸殘基分別一致之候選序列中之核酸或胺基酸殘基分別之百分比。保守取代視為序列一致性之一部分。較佳者係無間隙比對。出於確定胺基酸序列一致性百分比之目的，可以熟習此項技術者所熟知之各種方式來達成比對，例如使用可公開獲得之電腦軟體，例如BLAST、BLAST-2、ALIGN或Megalign (DNASTAR)軟體。熟習此項技術者可確定用於比對序列之適當參數，包含在所比較序列之全長範圍內達成最大比對所需要之任何算法。"Percent sequence identity (%)" with respect to a reference polynucleotide or polypeptide sequence is defined as the percentage of nucleic acid or amino acid residues, respectively, in a candidate sequence that are identical to nucleic acid or amino acid residues, respectively, in a reference polynucleotide or polypeptide sequence, after the sequences have been aligned and (if necessary) gaps have been introduced to achieve maximum percent sequence identity. Conservative substitutions are considered part of the sequence identity. Preferably, a gapless alignment is used. For the purpose of determining the percentage of amino acid sequence identity, alignment can be achieved in a variety of ways known to those skilled in the art, such as using publicly available computer software such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithm required to achieve maximum alignment over the full length of the compared sequences.

實質性序列一致性/相似性可理解為對完整序列具有至少80%之序列相似性/一致性且/或對特異性結合區域(例如，CDR區域)具有至少90%之序列相似性/一致性。較佳序列相似性或更佳地一致性可為至少92%、95%、97%、98%或99%。可使用來自University of Wisconsin之Genetics Computer Group Version 10版軟體包之「BestFit」程式來測定序列相似性及/或一致性。該程式以默認值使用Smith及Waterman之局部具有算法(local had algorithm)：空位產生罰分=8，空位擴展罰分=2，平均匹配=2.912，平均失配=2.003。Substantial sequence identity/similarity is understood to be at least 80% sequence similarity/identity for the complete sequence and/or at least 90% sequence similarity/identity for the specific binding region (e.g., CDR region). Preferably, the sequence similarity or more preferably the identity may be at least 92%, 95%, 97%, 98%, or 99%. The "BestFit" program from the Genetics Computer Group Version 10 software package from the University of Wisconsin can be used to determine sequence similarity and/or identity. The program uses the local had algorithm of Smith and Waterman with default values: gap creation penalty = 8, gap extension penalty = 2, average match = 2.912, average mismatch = 2.003.

本文所互換使用之術語「多核苷酸」或「核酸」係指任一長度之核苷酸鏈，且包含DNA及RNA。核苷酸可為去氧核糖核苷酸、核糖核苷酸、經修飾核苷酸或鹼基及/或其類似物或任一可藉由DNA或RNA聚合酶納入鏈中之受質。多核苷酸可包括經修飾核苷酸，例如甲基化核苷酸及其類似物。The terms "polynucleotide" or "nucleic acid" used interchangeably herein refer to a nucleotide chain of any length and include DNA and RNA. The nucleotides may be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases and/or their analogs or any substrate that can be incorporated into the chain by DNA or RNA polymerase. A polynucleotide may include modified nucleotides, such as methylated nucleotides and their analogs.

「序列同源性」指示相同或代表保守胺基酸取代之胺基酸之百分比。"Sequence homology" indicates the percentage of amino acids that are identical or represent conservative amino acid substitutions.

如本文所使用，術語「載體」係指能夠轉運與其連接之另一核酸之核酸分子。該術語包括呈自複製核酸結構之載體，以及納入引入其之宿主細胞基因體中的載體。某些載體能夠引導與其可操作地連接之核酸之表現。該等載體在本文中稱為「表現載體」。As used herein, the term "vector" refers to a nucleic acid molecule capable of transporting another nucleic acid to which it is linked. The term includes vectors that are self-replicating nucleic acid structures, as well as vectors that are incorporated into the genome of a host cell into which they are introduced. Certain vectors are capable of directing the expression of nucleic acids to which they are operably linked. Such vectors are referred to herein as "expression vectors."

術語「宿主細胞」、「宿主細胞系」及「宿主細胞培養物」可互換使用且係指其中已引入至少一種外源核酸之細胞，包括該等細胞之子代。宿主細胞包括「轉化體」及「經轉化細胞」、「轉染子」及「經轉染細胞」，其包括原代經轉化/轉染/轉導細胞及源自其之子代(與傳代次數無關)。子代之核酸含量可與親代細胞並不完全相同，而是可含有突變。本文包括如經篩選或選擇用於原始經轉化細胞中之具有相同功能或生物活性之突變體子代。The terms "host cell," "host cell line," and "host cell culture" are used interchangeably and refer to cells into which at least one exogenous nucleic acid has been introduced, including the progeny of such cells. Host cells include "transformants" and "transformed cells," "transfectants," and "transfected cells," which include the primary transformed/transfected/transduced cell and progeny derived therefrom (regardless of the number of passages). The nucleic acid content of the progeny may not be exactly the same as the parent cell, but may contain mutations. Mutant progeny that have the same function or biological activity as screened or selected for use in the original transformed cell are included herein.

如本文中所使用，片語「治療有效量」意欲係指，在根據期望治療方案投與時，適於誘發期望治療性或預防性效應或反應之治療性或預防性抗體之量，該效應或反應包含緩解一些或所有該等疾病之症狀或減小患該疾病之傾向。As used herein, the phrase "therapeutically effective amount" is intended to mean an amount of a therapeutic or prophylactic antibody that, when administered according to a desired treatment regimen, is suitable to induce a desired therapeutic or prophylactic effect or response, including alleviation of some or all of the symptoms of the disease or reduction of the predisposition to develop the disease.

術語「醫藥調配物」/「醫藥組合物」係指如下製劑：其所呈現形式允許其中所含活性成分之生物活性有效，且不含對將投與該調配物之個體具有不可接受毒性之其他組分。The term "pharmaceutical formulation"/"pharmaceutical composition" refers to a preparation that is in a form that permits the biological activity of the active ingredient contained therein to be effective and does not contain other components that are unacceptably toxic to the subject to which the formulation is to be administered.

「放射性核素(radionuclide)」(亦：「放射性核素(radioactive nuclide)」、「放射性同位素(radioisotope)」或「放射性同位素(radioactive isotope)」)係發生放射性衰變之原子。舉例而言(無限制)，放射性核素可為β-發射放射性核素(β發射體)、α-粒子-發射放射性核素(α發射體)或歐傑電子(Auger electron)發射放射性核素(歐傑電子發射體)。A “radionuclide” (also: “radioactive nuclide”, “radioisotope” or “radioactive isotope”) is an atom that undergoes radioactive decay. By way of example and without limitation, a radionuclide may be a beta-emitting radionuclide (beta emitter), an alpha-particle-emitting radionuclide (alpha emitter) or an Auger electron emitting radionuclide (Auger electron emitter).

「β-發射放射性核素」或「β發射體」係發射β粒子之放射性核素。β發射體之實例包含(但不限於)銅-67 (⁶⁷Cu)、鍶-89 (⁸⁹Sr)、釔-90 (⁹⁰Y)、銠-105 (¹⁰⁵Rh)、碘-131 (¹³¹I)、鉕-149 (¹⁴⁹Pm)、鈥-166 (¹⁶⁶Ho)、鎦-177 (¹⁷⁷Lu)、錸-186 (¹⁸⁶Re)、錸-188 (¹⁸⁸Re)、金-198 (¹⁹⁸Au)及金-199 (¹⁹⁹Au)。Jongho已綜述用於螯合各種該等放射性同位素之各種技術(Jeon, Jongho. "Review of therapeutic applications of radiolabeled functional nanomaterials." International journal of molecular sciences 20.9 (2019): 2323.)。"Beta-emitting radionuclides" or "beta emitters" are radionuclides that emit beta particles. Examples of beta emitters include, but are not limited to, copper-67 (⁶⁷ Cu), strontium-89 (⁸⁹ Sr), yttrium-90 (⁹⁰ Y), rhodium-105 (¹⁰⁵ Rh), iodine-131 (¹³¹ I), bismuth-149 (¹⁴⁹ Pm), ruthenium-166 (¹⁶⁶ Ho), titanium-177 (¹⁷⁷ Lu), rhodium-186 (¹⁸⁶ Re), rhodium-188 (¹⁸⁸ Re), gold-198 (¹⁹⁸ Au), and gold-199 (¹⁹⁹ Au). Jongho has reviewed various techniques for chelating various of these radioisotopes (Jeon, Jongho. "Review of therapeutic applications of radiolabeled functional nanomaterials." International journal of molecular sciences 20.9 (2019): 2323.).

「歐傑電子發射放射性核素」或「歐傑電子發射體」係發射歐傑電子之放射性核素。歐傑電子發射體之實例包含(但不限於)溴-77、銦-111、碘-123及碘-125。"Erjer electron-emitting radionuclides" or "Erjer electron emitters" are radionuclides that emit Erjer electrons. Examples of Erjer electron emitters include, but are not limited to, bromine-77, indium-111, iodine-123, and iodine-125.

「α-發射放射性核素」或「α發射體」係發射α粒子(亦即具有+2電荷之4He細胞核)之放射性核素。α發射體之非限制性實例包含藉由45.6 min之半衰期表徵之鉍-213 (²¹³Bi)、藉由9.9 d之半衰期表徵之錒-225 (²²⁵Ac)、藉由7.2 h之半衰期表徵之砹-211 (²¹¹At)、藉由11.4 d之半衰期表徵之鐳-223 (²²³Ra)、藉由3.7 d之半衰期表徵之鐳-224 (²²⁴Ra)及藉由18.7 d之半衰期表徵之釷-227 (²²⁷Th)。An "alpha-emitting radionuclide" or "alpha emitter" is a radionuclide that emits alpha particles (i.e., 4He nuclei with a charge of +2). Non-limiting examples of alpha emitters include bismuth-213 (²¹³ Bi) characterized by a half-life of 45.6 min, ruthenium-225 (²²⁵ Ac) characterized by a half-life of 9.9 d, astatine-211 (²¹¹ At) characterized by a half-life of 7.2 h, radium-223 (²²³ Ra) characterized by a half-life of 11.4 d, radium-224 (²²⁴ Ra) characterized by a half-life of 3.7 d, and thrombium-227 (²²⁷ Th) characterized by a half-life of 18.7 d.

在本發明之上下文中，術語「Ac」意指至少一種α-發射錒同位素之離子。較佳地，α-發射錒同位素係225Ac。較佳地，至少一種α-發射錒同位素之離子係具有三重正電荷(亦即，²²⁵Ac³⁺)之²²⁵Ac。因此，在本發明之上下文中，術語「Ac」較佳地，但並非排他性地，意指²²⁵Ac³⁺。In the context of the present invention, the term "Ac" means an ion of at least one α-emitting ruthenium isotope. Preferably, the α-emitting ruthenium isotope is 225Ac. Preferably, the ion of at least one α-emitting ruthenium isotope is^225Ac having a triply positive charge (i.e.,^225Ac3+ ). Therefore, in the context of the present invention, the term "Ac" preferably, but not exclusively, means^225Ac3+ .

可根據業內已知方法獲得放射性核素。舉例而言，如Poty、Sophie等人所闡述，可藉由利用²⁰⁹Bi(α,2n)²¹¹At反應以中等能量之α-粒子光束轟擊天然鉍來在回旋加速器上產生²¹¹At (「Poty, Sophie等人，α-Emitters for radiotherapy: from basic radiochemistry to clinical studies—part 1/2.」 Journal of Nuclear Medicine 59.6/59.7 (2018): 878–884 / 1020-1027)。²²⁷Th及²²³Ra二者可自其相互之親代(²²⁷Ac)分離而獲得。使用基於²²⁷Ac/^227Th之生成器來產生臨床²²³Ra。將親代同位素裝載至錒層析樹脂，且在用1M HCl或HNO3洗脫之後獲得²²³Ra氯化物溶液，後續使用陽離子交換管柱、蒸發且溶解於鹽水溶液中。Radionuclides can be obtained according to methods known in the art. For example, as described by Poty, Sophie et al.,²¹¹ At can be produced on a cyclotron by bombarding natural bismuth with a medium-energy α-particle beam using^{the 209} Bi(α,2n)²¹¹ At reaction ("Poty, Sophie et al., α-Emitters for radiotherapy: from basic radiochemistry to clinical studies—part 1/2." Journal of Nuclear Medicine 59.6/59.7 (2018): 878–884 / 1020-1027). Both²²⁷ Th and²²³ Ra can be obtained by separation from their respective parents (²²⁷ Ac).^{A 227} Ac/^227Th -based generator is used to produce clinical²²³ Ra. The parent isotope was loaded onto a chromatographic resin and the²²³ Ra chloride solution was obtained after elution with 1 M HCl or HNO3, followed by a cation exchange column, evaporation and dissolution in aqueous saline solution.

「螯合」係指配體與單一中心金屬原子之間之兩種或更多種單獨配位鍵之形成或存在。能夠形成該等配位鍵之配體稱為「螯合劑」、「螯合試劑」或「鉗合劑」。"Chelation" refers to the formation or existence of two or more separate coordination bonds between a ligand and a single central metal atom. Ligands capable of forming such coordination bonds are called "chelators", "chelating agents" or "clamping agents".

「螯合部分」係可螯合既定放射性核素或放射性核素之群(例如(但不限於) α發射體、β發射體或歐傑電子發射體)之螯合劑。各種螯合劑在業內已熟知且可用於本發明，例如，如Price, Eric W.及Chris Orvig. "Matching chelators to radiometals for radiopharmaceuticals." Chemical Society Reviews 43.1 (2014): 260-290中所闡述，其全部內容併入本文中。A "chelating moiety" is a chelating agent that can chelate a given radionuclide or group of radionuclides, such as, but not limited to, an alpha emitter, a beta emitter, or an eutectic electron emitter. Various chelating agents are well known in the art and can be used in the present invention, for example, as described in Price, Eric W. and Chris Orvig. "Matching chelators to radiometals for radiopharmaceuticals." Chemical Society Reviews 43.1 (2014): 260-290, the entire contents of which are incorporated herein.

在本發明之上下文中，螯合部分包括(例如)18員螯合部分，例如大環螯合劑「macropa」(N,N′-雙[(6-羧基-2-吡啶基)甲基]-4,13-二氮雜-18-冠-6)。大環螯合劑macropa已闡述(舉例而言)於Thiele等人，Angew. Chem. Int. Ed. 2017, 56(46), 14712及其中之參考文獻中。Macropa螯合劑及其衍生物，例如「Macropa-NCS」(6-[[16-[(6-羧基-2-吡啶基)甲基]-1,4,10,13-四氧雜-7,16-二氮雜環十八-7-基]甲基]-4-[2-(4-異硫基氰基苯基)乙氧基]吡啶-2-甲酸)，亦闡述於WO2018/183906及WO2020/106886及其中之參考文獻中。In the context of the present invention, chelating moieties include, for example, 18-membered chelating moieties, such as the macrocyclic chelator "macropa" (N,N'-bis[(6-carboxy-2-pyridyl)methyl]-4,13-diaza-18-crown-6). The macrocyclic chelator macropa has been described, for example, in Thiele et al., Angew. Chem. Int. Ed. 2017, 56(46), 14712 and references therein. Macropa chelators and their derivatives, such as "Macropa-NCS" (6-[[16-[(6-carboxy-2-pyridyl)methyl]-1,4,10,13-tetraoxazol-7,16-diazacyclooctadeca-7-yl]methyl]-4-[2-(4-isothiocyanophenyl)ethoxy]pyridine-2-carboxylic acid), are also described in WO2018/183906 and WO2020/106886 and references therein.

可在異硫氰酸酯或羧酸部分終止錒複合之螯合部分(或「錒-複合螯合部分」)以促進其與靶向部分(例如，本發明之抗GPC3抗體或其抗原結合片段)偶合。在使用macropa螯合劑時，macropa螯合劑之吡啶環中之一者通常經取代，從而其可偶合至靶向部分。此可為簡單官能基(例如，羧酸-COOH)或更複雜之化學結構，只要其能夠偶合至靶向部分(例如，本發明之抗GPC3抗體或其抗原結合片段)。舉例而言，macropa螯合劑之吡啶環上之取代基可在異硫氰酸酯或羧酸部分終止。The chelating moiety of the thiazide complex (or "thiazide-complex chelating moiety") can be terminated in an isothiocyanate or carboxylic acid moiety to facilitate its coupling to a targeting moiety (e.g., an anti-GPC3 antibody or antigen-binding fragment thereof of the present invention). When a macropa chelator is used, one of the pyridine rings of the macropa chelator is typically substituted so that it can be coupled to the targeting moiety. This can be a simple functional group (e.g., carboxylic acid -COOH) or a more complex chemical structure, as long as it is capable of coupling to the targeting moiety (e.g., an anti-GPC3 antibody or antigen-binding fragment thereof of the present invention). For example, the substituents on the pyridine ring of the macropa chelator can be terminated in an isothiocyanate or carboxylic acid moiety.

本文所用之「靶向部分」係結合至生物靶(例如GPC3或表現GPC3之細胞)之任一化學結構。靶向部分自身(例如作為較大結構之部分)定位於需要其存在之特定位點以施加預期效應，例如在TAC之情形下遞送放射性衰變。因此，與不包括靶向部分之等效複合物之濃度對比，靶向組織之基團或部分用於將分子或結合物大大定位於個體之身體中之至少一種期望位點。靶向部分可(例如)選自(但不限於)由以下組成之群：核苷酸、DNA及RNA片段、適配體、肽、蛋白質、抗體或其片段、奈米顆粒或小分子或其任何組合。根據本發明，靶向部分較佳地係抗體或其抗原結合片段(亦即，本發明之抗GPC3-抗體或其片段)。As used herein, a "targeting moiety" is any chemical structure that binds to a biological target (e.g., GPC3 or a cell expressing GPC3). The targeting moiety itself (e.g., as part of a larger structure) is localized to a specific site where its presence is desired to exert a desired effect, such as delivery of radioactive decay in the case of TACs. Thus, a group or portion of a targeted tissue is used to localize a molecule or conjugate to at least one desired site in the body of an individual, as compared to the concentration of an equivalent complex not including the targeting moiety. The targeting moiety may, for example, be selected from the group consisting of (but not limited to) nucleotides, DNA and RNA fragments, aptamers, peptides, proteins, antibodies or fragments thereof, nanoparticles or small molecules or any combination thereof. According to the present invention, the targeting moiety is preferably an antibody or an antigen-binding fragment thereof (i.e., an anti-GPC3-antibody or fragment thereof of the present invention).

「靶向錒結合物」(亦「TAC」)係包括靶向部分(亦即，本發明之抗GPC3抗體或其片段)及用於錒複合之至少一種螯合劑或螯合基團(例如macropa、較佳地Macropa-NCS)且視情況包括錒之結合物。"Targeted Taconjugate" (also "TAC") is a conjugate comprising a targeting moiety (i.e., an anti-GPC3 antibody or fragment thereof of the present invention) and at least one chelating agent or chelating group for Taconjugate complexing (e.g., macropa, preferably Macropa-NCS), and optionally Taconjugate.

術語「DOTA」係指可螯合物放射性核素之2,2’,2’’,2’’’-(1,4,7,10-四氮雜環十二烷-1,4,7,10-四基)四乙酸或1,4,7,10-四-氮雜環十二烷-N,N′,N′′,N′′′-四乙酸及其衍生物。DOTA係經配置用於複合α-粒子-發射放射性核素(例如²¹²Bi、²¹³Bi、²²⁵Ac、²²⁷Th)之螯合劑。諸如DOTA之螯合劑與螯合之以3+氧化狀態存在之As-225形成穩定複合物。為達成DOTA-偶合抗體之足夠標記，應較佳地以兩步製程或直接在升高溫度下實施複合物步驟。The term "DOTA" refers to 2,2',2'',2'''-(1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrayl)tetraacetic acid or 1,4,7,10-tetra-azacyclododecane-N,N',N'',N'''-tetraacetic acid and its derivatives which can chelate radionuclides. DOTA is a chelator configured for complexing alpha-particle-emitting radionuclides (e.g.,²¹² Bi,²¹³ Bi,²²⁵ Ac,²²⁷ Th). Chelators such as DOTA form stable complexes with chelated As-225 in the 3+ oxidation state. To achieve adequate labeling of DOTA-conjugated antibodies, the complexation step should preferably be performed in a two-step process or directly at elevated temperature.

在本發明之上下文中，術語CAR係指螯合劑對抗原比率，其為經放射性標記之化合物(例如，本發明靶向錒結合物)之比活性之量度。In the context of the present invention, the term CAR refers to the chelator to antigen ratio, which is a measure of the specific activity of a radiolabeled compound (eg, a targeted titania conjugate of the present invention).

在本發明之上下文中，術語IRF係指免疫反應性分數，亦即能夠結合至靶(亦即，GPC3)之經標記產物(亦即，本發明靶向錒結合物)之分數。Lindmo分析(Lindmo T.等人 (1984) 「Determination of the immunoreactive fraction of radiolabeled monoclonal antibodies by linear extrapolation to binding at infinite antigen excess.」J. Immunol. Methods.72, 77–89)係用於評價免疫反應性分數之最常使用之方法。In the context of the present invention, the term IRF refers to the immunoreactive fraction, i.e., the fraction of labeled products (i.e., the targeted titanate conjugate of the present invention) that are able to bind to the target (i.e., GPC3). The Lindmo analysis (Lindmo T. et al. (1984) "Determination of the immunoreactive fraction of radiolabeled monoclonal antibodies by linear extrapolation to binding at infinite antigen excess."J. Immunol. Methods. 72, 77-89) is the most commonly used method for evaluating the immunoreactive fraction.

本發明之抗-GPC3抗體根據第一態樣，本發明係關於能夠結合至GPC3之經分離抗體或其抗原結合片段。Anti-GPC3Antibodiesof the Invention According to a first aspect, the present invention relates to isolated antibodies or antigen-binding fragments thereof that are capable of binding to GPC3.

該等抗體可為(舉例而言) IgG抗體，例如人類 IgG1、IgG2、IgG3或IgG4或小鼠IgG1、IgG2a、IgG2b或IgG2c。在一些極佳實施例中，當前態樣之經分離抗體或其抗原結合片段係人類化IgG1抗體。在一些其他較佳實施例中，當前態樣之抗體或其抗原結合片段係Fab'、F(ab')2、Fab、Fv、rlgG或scFv片段。The antibodies may be, for example, IgG antibodies, such as human IgG1, IgG2, IgG3 or IgG4 or mouse IgG1, IgG2a, IgG2b or IgG2c. In some very preferred embodiments, the isolated antibody or antigen-binding fragment thereof of the present aspect is a humanized IgG1 antibody. In some other preferred embodiments, the antibody or antigen-binding fragment thereof of the present aspect is a Fab', F(ab')2, Fab, Fv, rIgG or scFv fragment.

在一些較佳實施例中，當前態樣之抗體或其抗原結合片段結合至人類GPC3。在一些該等較佳實施例中，當前態樣之抗體或其抗原結合片段與來自其他物種之GPC3 (例如猴、食蟹獼猴(macaca fascicularis、cynomolgus monkey)、恒河猴(macaca mulatta、rhesus macaque)、齧齒類動物、小鼠、大鼠、馬、牛類、豬、狗、貓及/或駱駝GPC3)、較佳與食蟹猴GPC3交叉反應。In some preferred embodiments, the antibodies or antigen-binding fragments thereof of the present aspects bind to human GPC3. In some of these preferred embodiments, the antibodies or antigen-binding fragments thereof of the present aspects cross-react with GPC3 from other species (e.g., monkey, cynomolgus monkey (macaca fascicularis, cynomolgus monkey), rhesus monkey (macaca mulatta, rhesus macaque), rodent, mouse, rat, horse, cow, pig, dog, cat and/or camel GPC3), preferably cynomolgus monkey GPC3.

根據一些較佳實施例，抗體或抗原結合片段針對SEQ ID 321之人類GPC3具有低於2E-07 M、較佳地低於1E-08 M、1E-09 M、10E-10 M或1E-10 M之結合親和力K_D。根據相同或不同之一些實施例，抗體或抗原結合片段對SEQ ID 323之食蟹猴GPC3具有低於2E-07 M、較佳地低於1E-08 M、1E-09 M、10E-10 M或1E-10 M之結合親和力K_D。According to some preferred embodiments, the antibody or antigen-binding fragment has a binding affinity K D of less than 2E-07 M, preferably less than 1E-08 M, 1E-09 M, 10E-10 M or 1E-10 M for human GPC3 of SEQ ID 321. According to some of the same or different embodiments, the antibody or antigen-binding fragment has a binding affinity K_D of less than 2E-07 M, preferably less than 1E-08 M, 1E-09 M, 10E-10 M or 1E-10 M for cynomolgus monkey_GPC3 of SEQ ID 323.

較佳地，抗體或抗原結合片段以相同數量級之K_D結合人類及食蟹猴GPC3，例如以低於2E-07 M、較佳地低於1E-08 M、1E-09 M、10E-10 M或1E-10 M之K_D。Preferably, the antibody or antigen-binding fragment binds to human and cynomolgus monkey GPC3 with a_KD of the same order, for example, with a_KD of less than 2E-07 M, preferably less than 1E-08 M, 1E-09 M, 10E-10 M or 1E-10 M.

在其他較佳實施例中，抗體或其抗原結合片段以低於1E-07 M、較佳地低於10 E-08 M、1E-08 M、10E-9 M或1E-09 M之EC50結合至SEQ ID 321之人類GPC3。根據相同或不同之一些實施例，抗體或抗原結合片段對SEQ ID 323之食蟹猴GPC3具有低於2E-07 M、較佳地低於10 E-08 M、1E-08 M、10E-9 M或1E-09 M之EC50。In other preferred embodiments, the antibody or antigen-binding fragment thereof binds to human GPC3 of SEQ ID 321 with an EC50 of less than 1E-07 M, preferably less than 10 E-08 M, 1E-08 M, 10E-9 M or 1E-09 M. According to some of the same or different embodiments, the antibody or antigen-binding fragment thereof has an EC50 of less than 2E-07 M, preferably less than 10 E-08 M, 1E-08 M, 10E-9 M or 1E-09 M for cynomolgus monkey GPC3 of SEQ ID 323.

較佳地，抗體或抗原結合片段以相同數量級之EC50結合人類及食蟹猴GPC3，例如以低於2E-07 M、較佳地低於10E-08 M、1E-08 M、10E-9 M或1E-9 M之EC50。Preferably, the antibody or antigen-binding fragment binds to human and cynomolgus monkey GPC3 with an EC50 of the same order of magnitude, for example, with an EC50 of less than 2E-07 M, preferably less than 10E-08 M, 1E-08 M, 10E-9 M or 1E-9 M.

在一些較佳實施例中，當前態樣之抗體或其抗原結合片段與最接近種系輕鏈IGKV2-28-IGKJ2 (SEQ ID NO:325)及在CDR-H3之外側鑑別之最接近種系重鏈IGHV1-46-IGHJ4 (SEQ-ID NO: 324)具有少量種系偏差，其低於32、較佳地低於30、25、20、19、18、17、16、15、14、13、12、11、10、5、4、3、2或1。In some preferred embodiments, the antibodies or antigen-binding fragments thereof of the present aspect have a small germline deviation from the closest germline light chain IGKV2-28-IGKJ2 (SEQ ID NO: 325) and the closest germline heavy chain IGHV1-46-IGHJ4 (SEQ-ID NO: 324) identified outside of CDR-H3, which is less than 32, preferably less than 30, 25, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 5, 4, 3, 2 or 1.

在其他較佳實施例中，當前態樣之抗體或其抗原結合片段針對免疫原性具有低風險，如藉由IPAD評分(如使用實例6中闡述之v4.03演算法所預測)反映低於1000、較佳地低於900、800、700、600或550。In other preferred embodiments, the antibodies or antigen-binding fragments thereof of the present aspect have a low risk for immunogenicity, as reflected by an IPAD score (as predicted using the v4.03 algorithm described in Example 6) of less than 1000, preferably less than 900, 800, 700, 600 or 550.

在其他較佳實施例中，當前態樣之抗體或其抗原結合片段在其CDR內包括少量離胺酸殘基。較佳地抗體或其抗原結合片段之6個CDR內之離胺酸殘基之數量低於4、更佳地低於3、甚至更佳地低於2且最佳地低於1。根據極佳實施例，當前態樣之抗體或其抗原結合片段在其6個CDR內不包括離胺酸殘基。In other preferred embodiments, the antibodies or antigen-binding fragments thereof of the present aspect include a small number of lysine residues in their CDRs. Preferably, the number of lysine residues in the 6 CDRs of the antibody or antigen-binding fragment thereof is less than 4, more preferably less than 3, even more preferably less than 2 and most preferably less than 1. According to a very preferred embodiment, the antibodies or antigen-binding fragments thereof of the present aspect do not include lysine residues in their 6 CDRs.

本發明之經分離抗體或抗原結合片段可展現上述特性之任何組合。The isolated antibodies or antigen-binding fragments of the present invention may exhibit any combination of the above-mentioned properties.

在一些較佳實施例中，本發明之經分離抗體或抗原結合片段包括： i) 包括含有SEQ ID NO: 82之H-CDR1、含有SEQ ID NO: 83之H-CDR2及含有SEQ ID NO: 84之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 86之L-CDR1、含有SEQ ID NO: 87之L-CDR2及含有SEQ ID NO:88之L-CDR3之輕鏈抗原結合區域(TPP-29312)，或 ii)     包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域(TPP-29537)，或 iii)    包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域(TPP-29538)，或 iv)    包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域(TPP-29636)，或 v) 包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域(TPP-29638)，或 vi)    包括含有SEQ ID NO: 182之H-CDR1、含有SEQ ID NO: 183之H-CDR2及含有SEQ ID NO: 184之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 186之L-CDR1、含有SEQ ID NO: 187之L-CDR2及含有SEQ ID NO:188之L-CDR3之輕鏈抗原結合區域(TPP-29642)。In some preferred embodiments, the isolated antibody or antigen-binding fragment of the present invention comprises:i) a heavy chain antigen-binding region comprising H-CDR1 containing SEQ ID NO: 82, H-CDR2 containing SEQ ID NO: 83, and H-CDR3 containing SEQ ID NO: 84, and a light chain antigen-binding region comprising L-CDR1 containing SEQ ID NO: 86, L-CDR2 containing SEQ ID NO: 87, and L-CDR3 containing SEQ ID NO: 88 (TPP-29312), orii)     a heavy chain antigen-binding region comprising H-CDR1 containing SEQ ID NO: 102, H-CDR2 containing SEQ ID NO: 103, and H-CDR3 containing SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 containing SEQ ID NO: 106, L-CDR2 containing SEQ ID NO: 107 and a light chain antigen binding region containing an L-CDR2 of SEQ ID NO: 108 (TPP-29537), oriii)    comprising an H-CDR1 containing SEQ ID NO: 122, an H-CDR2 containing SEQ ID NO: 123 and an H-CDR3 containing SEQ ID NO: 124, and a light chain antigen binding region containing an L-CDR1 containing SEQ ID NO: 126, an L-CDR2 containing SEQ ID NO: 127 and an L-CDR3 containing SEQ ID NO: 128 (TPP-29538), oriv)    comprising an H-CDR1 containing SEQ ID NO: 142, an H-CDR2 containing SEQ ID NO: 143 and an H-CDR3 containing SEQ ID NO: 144 and a light chain antigen binding region comprising an L-CDR1 comprising SEQ ID NO: 146, an L-CDR2 comprising SEQ ID NO: 147 and an L-CDR3 comprising SEQ ID NO: 148 (TPP-29636), orv) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 162, an H-CDR2 comprising SEQ ID NO: 163 and an H-CDR3 comprising SEQ ID NO: 164 and a light chain antigen binding region comprising an L-CDR1 comprising SEQ ID NO: 166, an L-CDR2 comprising SEQ ID NO: 167 and an L-CDR3 comprising SEQ ID NO: 168 (TPP-29638), orvi)    comprising an L-CDR1 comprising SEQ ID NO: 182, a heavy chain antigen binding region comprising H-CDR1 of SEQ ID NO: 183, and H-CDR3 of SEQ ID NO: 184, and a light chain antigen binding region comprising L-CDR1 of SEQ ID NO: 186, L-CDR2 of SEQ ID NO: 187, and L-CDR3 of SEQ ID NO: 188 (TPP-29642).

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括： i) 包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，或 ii)     包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域，或 iii)    包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域，或 iv)    包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域。In some preferred embodiments, the isolated antibody or antigen-binding fragment of the present invention comprises:i) a heavy chain antigen-binding region comprising H-CDR1 containing SEQ ID NO: 102, H-CDR2 containing SEQ ID NO: 103, and H-CDR3 containing SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 containing SEQ ID NO: 106, L-CDR2 containing SEQ ID NO: 107, and L-CDR3 containing SEQ ID NO: 108, orii)     a heavy chain antigen-binding region comprising H-CDR1 containing SEQ ID NO: 122, H-CDR2 containing SEQ ID NO: 123, and H-CDR3 containing SEQ ID NO: 124, and a light chain antigen-binding region comprising L-CDR1 containing SEQ ID NO: 126, L-CDR2 containing SEQ ID NO: 107, and L-CDR3 containing SEQ ID NO: 108. 127 and a light chain antigen binding region containing an L-CDR2 of SEQ ID NO: 128, oriii)    comprising an H-CDR1 containing SEQ ID NO: 142, an H-CDR2 containing SEQ ID NO: 143 and an H-CDR3 containing SEQ ID NO: 144, and a light chain antigen binding region containing an L-CDR1 containing SEQ ID NO: 146, an L-CDR2 containing SEQ ID NO: 147 and an L-CDR3 containing SEQ ID NO: 148, oriv)    comprising an H-CDR1 containing SEQ ID NO: 162, an H-CDR2 containing SEQ ID NO: 163 and an H-CDR3 containing SEQ ID NO: 164, and a heavy chain antigen binding region containing SEQ ID NO: 166, L-CDR1 containing SEQ ID NO: 167, and light chain antigen binding region containing L-CDR3 containing SEQ ID NO: 168.

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括：包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域(TPP-29537)。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention include: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103, and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 106, L-CDR2 comprising SEQ ID NO: 107, and L-CDR3 comprising SEQ ID NO: 108 (TPP-29537).

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括：包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域(TPP-29538)。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention include: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 122, H-CDR2 comprising SEQ ID NO: 123, and H-CDR3 comprising SEQ ID NO: 124, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 126, L-CDR2 comprising SEQ ID NO: 127, and L-CDR3 comprising SEQ ID NO: 128 (TPP-29538).

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括：包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域(TPP-29636)。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention include: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 142, H-CDR2 comprising SEQ ID NO: 143, and H-CDR3 comprising SEQ ID NO: 144, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 146, L-CDR2 comprising SEQ ID NO: 147, and L-CDR3 comprising SEQ ID NO: 148 (TPP-29636).

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括：包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域(TPP-29638)。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention include: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 162, H-CDR2 comprising SEQ ID NO: 163, and H-CDR3 comprising SEQ ID NO: 164, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 166, L-CDR2 comprising SEQ ID NO: 167, and L-CDR3 comprising SEQ ID NO: 168 (TPP-29638).

在其他較佳實施例中，本發明之經分離抗體或抗原結合片段包括： i)      與SEQ ID NO: 81具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 85具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 ii)     與SEQ ID NO: 101具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 105具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 iii)    與SEQ ID NO: 121具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 125具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 iv)    與SEQ ID NO: 141具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 145具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 v) 與SEQ ID NO: 161具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 165具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 vi)    與SEQ ID NO: 181具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 185具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域。In other preferred embodiments, the isolated antibody or antigen-binding fragment of the present invention comprises: i)     a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 81 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 85; or ii)     a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 101 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 105; or iii)     a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 121 has a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 125; oriv)    a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 141 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 145; orv) a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 161 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: ID NO: 165 has a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity; orvi)    A variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 181 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 185.

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括： i) 與SEQ ID NO: 101具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 105具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 ii)     與SEQ ID NO: 121具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 125具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 iii)    與SEQ ID NO: 141具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 145具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域；或 iv)    與SEQ ID NO: 161具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變重鏈結構域及/或與SEQ ID NO: 165具有至少90 %、95 %、98 %、99 %或100 %序列一致性之可變輕鏈結構域。In some preferred embodiments, the isolated antibody or antigen-binding fragment of the present invention comprises: i) a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 101 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 105; or ii)     a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 121 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 125; or iii)    a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 141 has a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 145; oriv)    a variable heavy chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 161 and/or a variable light chain domain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 165.

在其他較佳實施例中，本發明之經分離抗體或抗原結合片段包括： i) 包括SEQ ID NO: 81之可變重鏈結構域及包括SEQ ID NO: 85之可變輕鏈結構域；或 ii)     包括SEQ ID NO: 101之可變重鏈結構域及包括SEQ ID NO: 105之可變輕鏈結構域；或 iii)    包括SEQ ID NO: 121之可變重鏈結構域及包括SEQ ID NO: 125之可變輕鏈結構域；或 iv)    包括SEQ ID NO: 141之可變重鏈結構域及包括SEQ ID NO: 145之可變輕鏈結構域；或 v) 包括SEQ ID NO: 161之可變重鏈結構域及包括SEQ ID NO: 165之可變輕鏈結構域；或 vi)    包括SEQ ID NO: 181之可變重鏈結構域及包括SEQ ID NO: 185之可變輕鏈結構域。In other preferred embodiments, the isolated antibody or antigen-binding fragment of the present invention comprises: i) a variable heavy chain domain comprising SEQ ID NO: 81 and a variable light chain domain comprising SEQ ID NO: 85; or ii)     a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105; or iii)    a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125; or iv)    a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145; or v) a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165; orvi)    A variable heavy chain domain comprising SEQ ID NO: 181 and a variable light chain domain comprising SEQ ID NO: 185.

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括： i) 包括SEQ ID NO: 101之可變重鏈結構域及包括SEQ ID NO: 105之可變輕鏈結構域；或 ii)     包括SEQ ID NO: 121之可變重鏈結構域及包括SEQ ID NO: 125之可變輕鏈結構域；或 iii)    包括SEQ ID NO: 141之可變重鏈結構域及包括SEQ ID NO: 145之可變輕鏈結構域；或 iv)    包括SEQ ID NO: 161之可變重鏈結構域及包括SEQ ID NO: 165之可變輕鏈結構域。In some preferred embodiments, the isolated antibody or antigen-binding fragment of the present invention comprises:i) a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105; orii)     a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125; oriii)    a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145; oriv)    a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165.

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括含有SEQ ID NO: 101之可變重鏈結構域及含有SEQ ID NO: 105之可變輕鏈結構域。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention comprise a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105.

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括含有SEQ ID NO: 121之可變重鏈結構域及含有SEQ ID NO: 125之可變輕鏈結構域。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention comprise a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125.

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括含有SEQ ID NO: 141之可變重鏈結構域及含有SEQ ID NO: 145之可變輕鏈結構域。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention comprise a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145.

在一些極佳實施例中，本發明之經分離抗體或抗原結合片段包括含有SEQ ID NO: 161之可變重鏈結構域及含有SEQ ID NO: 165之可變輕鏈結構域。In some excellent embodiments, the isolated antibodies or antigen-binding fragments of the present invention comprise a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165.

在特定實施例中，本發明經分離抗體係IgG1、IgG2、IgG3或IgG4抗體。更具體而言，本發明經分離抗體係IgG1抗體。In a specific embodiment, the isolated antibody of the present invention is an IgG1, IgG2, IgG3 or IgG4 antibody. More specifically, the isolated antibody of the present invention is an IgG1 antibody.

在其他較佳實施例中，本發明之經分離抗體或抗原結合片段包括： i) 與SEQ ID NO: 97具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 98具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈。 ii)     與SEQ ID NO: 117具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 118具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈；或 iii)    與SEQ ID NO: 137具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 138具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈；或 iv)    與SEQ ID NO: 157具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 158具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈；或 v) 與SEQ ID NO: 177具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 178具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈；或 vi)    與SEQ ID NO: 197具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 198具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈。In other preferred embodiments, the isolated antibody or antigen-binding fragment of the present invention comprises:i) a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 97 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 98.ii)     a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 117 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 118; oriii)    a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 137 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 138; oriv)    a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 157 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 158 %, 95%, 98%, 99% or 100% sequence identity to SEQ ID NO: 177; orv) a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity to SEQ ID NO: 177 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity to SEQ ID NO: 178; orvi)    a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity to SEQ ID NO: 197 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity to SEQ ID NO: 198.

在一些極佳實施例中，本發明經分離抗體包括： i) 與SEQ ID NO: 117具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 118具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈；或 ii)     與SEQ ID NO: 137具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 138具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈；或 iii)    與SEQ ID NO: 157具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 158具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈；或 iv)    與SEQ ID NO: 177具有至少90 %、95 %、98 %、99 %或100 %序列一致性之重鏈及/或與SEQ ID NO: 178具有至少90 %、95 %、98 %、99 %或100 %序列一致性之輕鏈。In some preferred embodiments, the isolated antibodies of the present invention include: i) a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 117 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 118; or ii)     a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 137 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 138; or iii)    a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity with SEQ ID NO: 157 % sequence identity to SEQ ID NO: 158; oriv)    a heavy chain having at least 90%, 95%, 98%, 99% or 100% sequence identity to SEQ ID NO: 177 and/or a light chain having at least 90%, 95%, 98%, 99% or 100% sequence identity to SEQ ID NO: 178.

在一些較佳實施例中，本發明經分離抗體包括： i) 包括SEQ ID NO: 97之重鏈及包括SEQ ID NO: 98之輕鏈；或 ii)     包括SEQ ID NO: 117之重鏈及包括SEQ ID NO: 118之輕鏈；或 iii)    包括SEQ ID NO: 137之重鏈及包括SEQ ID NO: 138之輕鏈；或 iv)    包括SEQ ID NO: 157之重鏈及包括SEQ ID NO: 158之輕鏈；或 v) 包括SEQ ID NO: 177之重鏈及包括SEQ ID NO: 178之輕鏈；或 vi)    包括SEQ ID NO: 197之重鏈及包括SEQ ID NO: 198之輕鏈。In some preferred embodiments, the isolated antibody of the present invention comprises: i) a heavy chain comprising SEQ ID NO: 97 and a light chain comprising SEQ ID NO: 98; or ii)     a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118; or iii)    a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138; or iv)    a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158; or v) a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178; or vi)    a heavy chain comprising SEQ ID NO: 197 and a light chain comprising SEQ ID NO: 198.

在一些極佳實施例中，本發明經分離抗體包括： i) 包括SEQ ID NO: 117之重鏈及包括SEQ ID NO: 118之輕鏈；或 ii)     包括SEQ ID NO: 137之重鏈及包括SEQ ID NO: 138之輕鏈；或 iii)    包括SEQ ID NO: 157之重鏈及包括SEQ ID NO: 158之輕鏈；或 iv)    包括SEQ ID NO: 177之重鏈及包括SEQ ID NO: 178之輕鏈。In some preferred embodiments, the isolated antibody of the present invention comprises:i) a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118; orii)     a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138; oriii)    a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158; oriv)    a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178.

在一些極佳實施例中，本發明經分離抗體包括含有SEQ ID NO: 117之重鏈及含有SEQ ID NO: 118之輕鏈(TPP-29537)。In some excellent embodiments, the isolated antibody of the present invention comprises a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118 (TPP-29537).

在一些極佳實施例中，本發明經分離抗體包括含有SEQ ID NO: 137之重鏈及含有SEQ ID NO: 138之輕鏈(TPP-29538)。In some excellent embodiments, the isolated antibody of the present invention comprises a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138 (TPP-29538).

在一些極佳實施例中，本發明經分離抗體包括含有SEQ ID NO: 157之重鏈及含有SEQ ID NO: 158之輕鏈(TPP-29636)。In some excellent embodiments, the isolated antibody of the present invention comprises a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158 (TPP-29636).

在一些極佳實施例中，本發明經分離抗體包括含有SEQ ID NO: 177之重鏈及含有SEQ ID NO: 178之輕鏈(TPP-29638)。In some excellent embodiments, the isolated antibody of the present invention comprises a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178 (TPP-29638).

在特定實施例中，本發明抗原結合片段想scFv、Fab、Fab’片段或F(ab’)2片段。In certain embodiments, the antigen-binding fragment of the present invention is a scFv, Fab, Fab' fragment or F(ab')2 fragment.

在特定實施例中，本發明之經分離抗體或抗原結合片段係單株抗體或抗原結合片段。In certain embodiments, the isolated antibodies or antigen-binding fragments of the present invention are monoclonal antibodies or antigen-binding fragments.

在特定實施例中，本發明之經分離抗體或抗原結合片段係人類、人類化或嵌合抗體或抗原結合片段，更具體而言人類化抗體或抗原結合片段。In certain embodiments, the isolated antibodies or antigen-binding fragments of the present invention are human, humanized or chimeric antibodies or antigen-binding fragments, more specifically humanized antibodies or antigen-binding fragments.

在特定實施例中，本發明之經分離抗體或抗原結合片段係單特異性抗體。在特定其他實施例中，本發明之經分離抗體或抗原結合片段係結合至GPC3及至少一種其他抗原之多特異性抗體，例如雙特異性、三特異性或四特異性抗體。 表1中列示本發明較佳抗體之胺基酸序列。表1：本發明較佳抗體之胺基酸序列抗體/ 同型SEQ ID NO: VHSEQ ID NO: H-CDR1SEQ ID NO: H-CDR2SEQ ID NO: H-CDR3SEQ ID NO: VLSEQ ID NO: L-CDR1SEQ ID NO: L-CDR2SEQ ID NO: L-CDR3SEQ ID NO: IgG重鏈SEQ ID NO: IgG輕鏈TPP-29312 hIgG181828384858687889798TPP-29537 hIgG1101102103104105106107108117118TPP-29538 hIgG1121122123124125126127128137138TPP-29636 hIgG1141142143144145146147148157158TPP-29638 hIgG1161162163164165166167168177178TPP-29642 hIgG1181182183184185186187188197198In certain embodiments, the isolated antibodies or antigen-binding fragments of the present invention are monospecific antibodies. In certain other embodiments, the isolated antibodies or antigen-binding fragments of the present invention are multispecific antibodies that bind to GPC3 and at least one other antigen, such as bispecific, trispecific or tetraspecific antibodies. The amino acid sequences of the preferred antibodies of the present invention are listed in Table 1.Table1 : Amino acid sequences of the preferred antibodies of the present invention Antibody/ Isotype SEQ ID NO: VH SEQ ID NO: H-CDR1 SEQ ID NO: H-CDR2 SEQ ID NO: H-CDR3 SEQ ID NO: VL SEQ ID NO: L-CDR1 SEQ ID NO: L-CDR2 SEQ ID NO: L-CDR3 SEQ ID NO: IgG heavy chain SEQ ID NO: IgG light chain TPP-29312 hIgG1 81 82 83 84 85 86 87 88 97 98 TPP-29537 hIgG1 101 102 103 104 105 106 107 108 117 118 TPP-29538 hIgG1 121 122 123 124 125 126 127 128 137 138 TPP-29636 hIgG1 141 142 143 144 145 146 147 148 157 158 TPP-29638 hIgG1 161 162 163 164 165 166 167 168 177 178 TPP-29642 hIgG1 181 182 183 184 185 186 187 188 197 198

表2中列示本發明較佳抗體之核酸序列。表2：本發明較佳抗體之核酸序列抗體/ 同型SEQ ID NO: VHSEQ ID NO: H-CDR1SEQ ID NO: H-CDR2SEQ ID NO: H-CDR3SEQ ID NO: VLSEQ ID NO: L-CDR1SEQ ID NO: L-CDR2SEQ ID NO: L-CDR3SEQ ID NO: IgG重鏈SEQ ID NO: IgG輕鏈TPP-29312 hIgG1899091929394959699100TPP-29537 hIgG1109110111112113114115116119120TPP-29538 hIgG1129130131132133134135136139140TPP-29636 hIgG1149150151152153154155156159160TPP-29638 hIgG1169170171172173174175176179180TPP-29642 hIgG1189190191192193194195196199200Table 2 lists the nucleic acid sequences of the preferred antibodies of the present invention.Table2 : Nucleic acid sequences of the preferred antibodies of the present invention Antibody/ Isotype SEQ ID NO: VH SEQ ID NO: H-CDR1 SEQ ID NO: H-CDR2 SEQ ID NO: H-CDR3 SEQ ID NO: VL SEQ ID NO: L-CDR1 SEQ ID NO: L-CDR2 SEQ ID NO: L-CDR3 SEQ ID NO: IgG heavy chain SEQ ID NO: IgG light chain TPP-29312 hIgG1 89 90 91 92 93 94 95 96 99 100 TPP-29537 hIgG1 109 110 111 112 113 114 115 116 119 120 TPP-29538 hIgG1 129 130 131 132 133 134 135 136 139 140 TPP-29636 hIgG1 149 150 151 152 153 154 155 156 159 160 TPP-29638 hIgG1 169 170 171 172 173 174 175 176 179 180 TPP-29642 hIgG1 189 190 191 192 193 194 195 196 199 200

肽變體本發明之抗體或抗原結合片段不限於本文所提供之具體肽序列。相反，本發明亦包含該等多肽之變體。參考本發明及傳統上可用技術及參考文獻，熟習此項技術者能夠製備、測試及利用本文所揭示之抗體之功能變體，同時認為具有結合至GPC3之能力之該等變體屬本發明範圍。Peptide Variants The antibodies or antigen-binding fragments of the present invention are not limited to the specific peptide sequences provided herein. Rather, the present invention also encompasses variants of such polypeptides. With reference to the present invention and conventionally available techniques and references, one skilled in the art will be able to prepare, test and utilize functional variants of the antibodies disclosed herein, and such variants having the ability to bind to GPC3 are considered to be within the scope of the present invention.

與本文所揭示之肽序列相比，變體可包含(例如)具有至少一個改變互補測定區域(CDR) (超變)及/或框架(FR) (可變)結構域/位置之抗體。Variants may include, for example, antibodies having at least one altered complementarity determining region (CDR) (hypervariable) and/or framework (FR) (variable) domain/position compared to the peptide sequences disclosed herein.

藉由改變CDR或FR區域中之一或多種胺基酸殘基，熟習此項技術者通常可生成突變或多樣化抗體序列，例如，可針對抗原篩選其，以獲得新或改良之性質。By altering one or more amino acid residues in the CDR or FR regions, one skilled in the art can typically generate mutant or diversified antibody sequences, which can be screened against antigen, for example, for new or improved properties.

本發明之其他較佳實施例係如表1中所展示來選擇VH及VL序列之抗體或抗原結合片段。熟習此項技術者可使用表1中之數據以設計本發明範圍內之肽變體。較佳地，藉由改變一或多個CDR區域內之胺基酸來構造變體；變體亦可具有一或多種經改變框架區。亦可在框架區內進行改變。舉例而言，可在與種系序列比較存在殘基之偏差之處改變肽FR結構域。Other preferred embodiments of the present invention are antibodies or antigen-binding fragments that select VH and VL sequences as shown in Table 1. One skilled in the art can use the data in Table 1 to design peptide variants within the scope of the present invention. Preferably, variants are constructed by changing amino acids within one or more CDR regions; variants may also have one or more altered framework regions. Changes may also be made within the framework regions. For example, peptide FR domains may be altered where there are residual deviations compared to the germline sequence.

或者，藉由比較本文所揭示之胺基酸序列與該等抗體之相同種類之已知序列，熟習此項技術者可進行相同分析，例如，使用由Knappik A.等人，JMB 2000, 296:57-86闡述之程序。Alternatively, one skilled in the art can perform the same analysis by comparing the amino acid sequences disclosed herein with known sequences of the same class of antibodies, for example, using the procedure described by Knappik A. et al., JMB 2000, 296:57-86.

另外，可藉由以下方式獲得變體：使用一種抗體作為起始點，以藉由多樣化抗體中之一或多種胺基酸殘基(較佳地一或多個CDR中之胺基酸殘基)進一步最佳化，且藉由篩選所得抗體變體之收集物以獲得具有改良性質之變體。尤佳者係多樣化VL及/或VH之CDR3中之一或多種胺基酸殘基。可(例如)藉由使用三核苷酸誘變(TRIM)技術合成DNA分子之收集物來進行多樣化(Virnekäs B.等人，Nucl.Acids Res.) 1994, 22: 5600.)。抗體或其抗原結合片段包含具有修飾/變化之分子，該等修飾/變化包含(但不限於)例如引起半衰期改變(例如，修飾Fc部分或附接其他分子(例如PEG))、結合親和力改變或ADCC或CDC活性改變之修飾。In addition, variants can be obtained by using one antibody as a starting point, further optimizing by diversifying one or more amino acid residues in the antibody (preferably one or more CDRs), and obtaining variants with improved properties by screening the resulting collection of antibody variants. Particularly preferred is diversifying one or more amino acid residues in CDR3 of VL and/or VH. Diversification can be performed, for example, by synthesizing a collection of DNA molecules using the trinucleotide induction mutation (TRIM) technique (Virnekäs B. et al., Nucl. Acids Res. 1994, 22: 5600.). Antibodies or antigen-binding fragments thereof include molecules with modifications/alterations including, but not limited to, modifications that, for example, result in altered half-life (e.g., modification of the Fc portion or attachment of other molecules such as PEG), altered binding affinity, or altered ADCC or CDC activity.

保守胺基酸變體可製得保留本文所闡述之抗體肽序列之整體分子結構之多肽變體。鑒於個別胺基酸之性質，熟習此項技術者會認可一些合理取代。例如，胺基酸取代(即「保守取代」)可基於所涉及殘基在以下方面中之相似性來進行：極性、電荷、溶解度、疏水性、親水性及/或兩親性特性。Conservative amino acid variants can be made to retain the overall molecular structure of the antibody peptide sequence described herein. In view of the properties of individual amino acids, those skilled in the art will recognize some reasonable substitutions. For example, amino acid substitutions (i.e., "conservative substitutions") can be made based on the similarity of the residues involved in the following aspects: polarity, charge, solubility, hydrophobicity, hydrophilicity and/or amphipathic properties.

舉例而言，(a)非極性(疏水性)胺基酸包含丙胺酸、白胺酸、異白胺酸、纈胺酸、脯胺酸、苯丙胺酸、色胺酸及甲硫胺酸；(b)極性中性胺基酸包含甘胺酸、絲胺酸、蘇胺酸、半胱胺酸、酪胺酸、天門冬醯胺酸及麩醯胺酸；(c)帶正電荷(鹼性)胺基酸包含精胺酸、離胺酸及組胺酸；且(d)帶負電荷(酸性)胺基酸包含天門冬胺酸及麩胺酸。通常可在(a)-(d)群組內進行取代。另外，甘胺酸及脯胺酸基於其破壞α螺旋之能力可彼此經取代。類似地，某些胺基酸(例如丙胺酸、半胱胺酸、白胺酸、甲硫胺酸、麩胺酸、麩醯胺酸、組胺酸及離胺酸)更常見於α螺旋中，而纈胺酸、異白胺酸、苯丙胺酸、酪胺酸、色胺酸及蘇胺酸更常見於β摺疊層。甘胺酸、絲胺酸、天門冬胺酸、天門冬醯胺酸及脯胺酸常見於轉角中。可在下列基團中進行一些較佳取代：(i) S及T；(ii) P及G；及(iii) A、V、L及I。鑒於已知遺傳密碼以及重組及合成DNA技術，熟習此項技術者可易於構造編碼保守胺基酸變體之DNA。For example, (a) non-polar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, and methionine; (b) polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, aspartate, and glutamine; (c) positively charged (basic) amino acids include arginine, lysine, and histidine; and (d) negatively charged (acidic) amino acids include aspartate and glutamine. Substitutions can generally be made within groups (a)-(d). In addition, glycine and proline can be substituted for each other based on their ability to disrupt alpha helices. Similarly, certain amino acids (e.g., alanine, cysteine, leucine, methionine, glutamine, glutamine, histidine, and lysine) are more common in alpha helices, while valine, isoleucine, phenylalanine, tyrosine, tryptophan, and threonine are more common in beta sheets. Glycine, serine, aspartate, asparagine, and proline are common in turns. Some preferred substitutions can be made in the following groups: (i) S and T; (ii) P and G; and (iii) A, V, L, and I. Given the known genetic code and recombinant and synthetic DNA technology, one skilled in the art can readily construct DNA encoding conservative amino acid variants.

醣基化變體當抗體包含Fc區時，其所附接之碳水化合物可能有所變化。由哺乳動物細胞產生之天然抗體通常包含通常藉由N-連接附接至Fc區之CH2結構域的Asn297 (使用Kabat EU編號)之支鏈、二分枝寡醣(例如，參見Wright等人， Trends Biotechnol. 15: 26-32 (1997)。Glycosylation variants When an antibody comprises an Fc region, the carbohydrates attached thereto may vary. Native antibodies produced by mammalian cells typically comprise branched, bibranched oligosaccharides attached, usually by N-linkage, to Asn297 (using Kabat EU numbering) of the CH2 domain of the Fc region (see, e.g., Wright et al., Trends Biotechnol. 15: 26-32 (1997).

在某些實施例中，改變本文所提供抗體以增加或降低抗體經醣基化之程度。抗體醣基化位點之添加或缺失可藉由改變表現系統(例如，宿主細胞)及/或藉由改變胺基酸序列使得產生或去除一或多個醣基化位點來方便地完成。In certain embodiments, the antibodies provided herein are altered to increase or decrease the degree to which the antibody is glycosylated. The addition or deletion of antibody glycosylation sites can be conveniently accomplished by altering the expression system (e.g., host cells) and/or by altering the amino acid sequence to create or remove one or more glycosylation sites.

在本發明之一種實施例中，藉由在原核宿主中之表現來製備具有降低之效應子功能之醣基抗體或抗體衍生物。適宜原核宿主包含(但不限於)大腸桿菌(E. coli)、枯草芽孢桿菌(Bacillus subtilis)、鼠傷寒沙門桿菌(Salmonella typhimurium)以及假單胞菌(Pseudomonas)、鏈黴菌(Streptomyces)及葡萄球菌(Staphylococcus)屬中之各種物種。In one embodiment of the present invention, glycoantibodies or antibody derivatives with reduced effector function are prepared by expression in prokaryotic hosts. Suitable prokaryotic hosts include, but are not limited to,E. coli ,Bacillus subtilis,Salmonella typhimurium , and various species of Pseudomonas, Streptomyces, and Staphylococcus.

在一實施例中，提供之抗體變體具有降低之效應子功能，其藉由該抗體之Fc部分之CH2結構域中之保守N-連接的位點處之修飾表徵。在本發明之一種實施例中，修飾包括重鏈醣基化位點處之突變以預防該位點之醣基化。因此，在本發明之一較佳實施例中，藉由重鏈醣基化位點之突變(亦即，N297(使用Kabat EU編號))之突變且在適當宿主細胞中表現來製備醣基抗體或抗體衍生物。In one embodiment, an antibody variant is provided with reduced effector function characterized by modification at a conserved N-linked site in the CH2 domain of the Fc portion of the antibody. In one embodiment of the invention, the modification comprises a mutation at a heavy chain glycosylation site to prevent glycosylation at the site. Thus, in a preferred embodiment of the invention, a glycoantibody or antibody derivative is prepared by mutation of a heavy chain glycosylation site (i.e., N297 (using Kabat EU numbering)) and expression in an appropriate host cell.

在本發明之另一實施例中，醣基抗體或抗體衍生物具有降低之效應子功能，其中該抗體或抗體衍生物的Fc部分之CH2結構域中之保守N-連接的位點處之修飾包括CH2結構域聚醣之去除，亦即，去醣基化。可藉由習用方法且然後以酶促方式去醣基化生成該等醣基抗體。抗體之酶促去醣基化之方法在業內已眾所周知(例如，Winkelhake & Nicolson (1976), J Biol Chem. 251(4):1074-80)。In another embodiment of the present invention, the glycoantibody or antibody derivative has reduced effector function, wherein the modification at the conserved N-linked site in the CH2 domain of the Fc portion of the antibody or antibody derivative comprises the removal of CH2 domain glycans, i.e., deglycosylation. The glycoantibodies can be generated by conventional methods and then enzymatically deglycosylated. Methods for enzymatic deglycosylation of antibodies are well known in the art (e.g., Winkelhake & Nicolson (1976), J Biol Chem. 251(4):1074-80).

在本發明之另一實施例中，可使用醣基化抑制劑衣黴素(tunicamycin)來達成去醣基化(Nose & Wigzell (1983), Proc Natl Acad Sci USA, 80(21):6632-6)。亦即，該修飾係在該抗體之Fc部分之CH2結構域中之保守N-連接的位點處預防醣基化。In another embodiment of the present invention, deglycosylation can be achieved using the glycosylation inhibitor tunicamycin (Nose & Wigzell (1983), Proc Natl Acad Sci USA, 80(21):6632-6). That is, the modification prevents glycosylation at the conserved N-linked site in the CH2 domain of the Fc portion of the antibody.

在一實施例中，提供具有缺乏附接(直接或間接)至Fc區之岩藻糖之碳水化合物結構之抗體變體。舉例而言，該抗體中岩藻糖之量可為1%至80%、1%至65%、5%至65%或20%至40%。藉由計算相對於附接至Asn 297之所有糖結構(例如複合物、雜合體及高甘露糖結構)之總量Asn297處糖鏈內岩藻糖之平均量來測定岩藻糖的量，如藉由MALDI-TOF質譜所量測，如(例如)WO 2008/077546中所述。Asn297係指位於Fc區中約297位(Fc區殘基之Eu編號)之天門冬醯胺殘基；然而，由於抗體中之微小序列變化，Asn297亦可位於297位上游或下游之約±3個胺基酸處，亦即，介於294位與300位之間。該等岩藻醣基化變體可具有改良之ADCC功能。In one embodiment, an antibody variant is provided that has a carbohydrate structure lacking fucose attached (directly or indirectly) to the Fc region. For example, the amount of fucose in the antibody may be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. The amount of fucose is determined by calculating the average amount of fucose within the sugar chain at Asn 297 relative to the total amount of all sugar structures (e.g., complexes, hybrids, and high mannose structures) attached to Asn 297, as measured by MALDI-TOF mass spectrometry, as described, for example, in WO 2008/077546. Asn297 refers to the asparagine residue located at about position 297 (Eu numbering of Fc region residues) in the Fc region; however, due to minor sequence variations in antibodies, Asn297 may also be located at about ±3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300. These fucosylation variants may have improved ADCC function.

「去岩藻糖基化」或「岩藻糖缺陷」抗體變體相關公開案之實例包含Okazaki等人， J Mol. Biol. 336: 1239-1249 (2004)；Yamane-Ohnuki等人，Biotech. Bioeng. 87: 614 (2004)。Examples of publications related to "defucosylated" or "fucose-deficient" antibody variants include Okazaki et al., J Mol. Biol. 336: 1239-1249 (2004); Yamane-Ohnuki et al., Biotech. Bioeng. 87: 614 (2004).

能夠產生去岩藻醣基化抗體之細胞系之實例包括缺少蛋白質岩藻醣基化之Lec13 CHO細胞(Ripka等人，Arch. Biochem. Biophys. 249:533-545 (1986)；及WO 2004/056312)及基因敲除細胞系，例如α-1,6-岩藻糖基轉移酶基因、FUT8、基因敲除CHO細胞(例如，參見Yamane-Ohnuki等人，Biotech. Bioeng. 87: 614 (2004)；Kanda, Y.等人，Biotechnol. Bioeng., 94(4):680-688 (2006))。Examples of cell lines capable of producing defucosylated antibodies include Lec13 CHO cells lacking protein fucosylation (Ripka et al., Arch. Biochem. Biophys. 249:533-545 (1986); and WO 2004/056312) and gene knockout cell lines, such as α-1,6-fucosyltransferase gene, FUT8, gene knockout CHO cells (e.g., see Yamane-Ohnuki et al., Biotech. Bioeng. 87:614 (2004); Kanda, Y. et al., Biotechnol. Bioeng., 94(4):680-688 (2006)).

進一步提供具有二等分寡醣之抗體變體，例如，其中附接至抗體Fc區之雙觸角寡醣由GlcNAc二等分。該等抗體變體可具有降低之岩藻醣基化及/或改良之ADCC功能。該等抗體變體之實例闡述於(例如) WO 2003/011878；美國專利第6,602,684號；及US 2005/0123546中。Further provided are antibody variants having bisected oligosaccharides, for example, wherein the biantennary oligosaccharide attached to the antibody Fc region is bisected by GlcNAc. Such antibody variants may have reduced fucosylation and/or improved ADCC function. Examples of such antibody variants are described in, for example, WO 2003/011878; U.S. Patent No. 6,602,684; and US 2005/0123546.

亦提供在附接至Fc區之寡醣中具有至少一個半乳糖殘基之抗體變體。該等抗體變體可具有改良之CDC功能。該等抗體變體闡述於(例如) WO1997/30087；WO1998/58964；及WO1999/22764中。Antibody variants having at least one galactose residue in the oligosaccharide attached to the Fc region are also provided. Such antibody variants may have improved CDC function. Such antibody variants are described in, for example, WO1997/30087; WO1998/58964; and WO1999/22764.

FC區變體在某些實施例中，可將一或多種胺基酸修飾(例如取代)引入本文所提供之抗體(例如，人類IgG1、IgG2、IgG3或IgG4 Fc區)之Fc區中，由此生成Fc區變體。FcRegion Variants In certain embodiments, one or more amino acid modifications (eg, substitutions) can be introduced into the Fc region of an antibody provided herein (eg, a human IgG1, IgG2, IgG3 or IgG4 Fc region), thereby generating an Fc region variant.

在某些實施例中，本發明涵蓋具有一些(但非全部)效應子功能之抗體變體，此使其成為許多應用之合意候選物，在該等應用中抗體之活體內半衰期較為重要，但某些效應子功能(例如補體及ADCC)係不必要或有害的。可實施活體外及/或活體內細胞毒性分析來證實CDC及/或ADCC活性之降低/消耗。舉例而言，可實施Fc受體(FcR)結合分析以確保抗體缺乏FcγR結合能力(因此可能缺乏ADCC活性)，但保留FcRn結合能力。在一些實施例中，對Fc區進行變化，從而改變(亦即，改良或減弱) C1q結合及/或補體依賴性細胞毒性(CDC)。In certain embodiments, the invention encompasses antibody variants that possess some (but not all) effector functions, making them desirable candidates for many applications where the in vivo half-life of the antibody is important, but certain effector functions (e.g., complement and ADCC) are unnecessary or detrimental. In vitro and/or in vivo cytotoxicity assays can be performed to confirm reduction/depletion of CDC and/or ADCC activity. For example, Fc receptor (FcR) binding assays can be performed to ensure that the antibody lacks FcγR binding ability (and therefore may lack ADCC activity), but retains FcRn binding ability. In some embodiments, changes are made to the Fc region to alter (i.e., improve or attenuate) C1q binding and/or complement-dependent cytotoxicity (CDC).

在某些實施例中，本發明考慮擁有增加或降低之半衰期之抗體變體。US2005/0014934 (Hinton等人)中闡述具有增加之半衰期及與新生Fc受體(FcRn)之改良之結合的抗體，該受體負責將母體IgG轉移至胎中(Guyer等人，J Immunol. 117:587 (1976)及Kim等人，J Immunol. 24:249 (1994))。彼等抗體包含具有一或多個改良Fc區與FcRn之結合之取代的Fc區。In certain embodiments, the present invention contemplates antibody variants with increased or decreased half-life. US2005/0014934 (Hinton et al.) describes antibodies with increased half-life and improved binding to the neonatal Fc receptor (FcRn), which is responsible for the transfer of maternal IgG to the fetus (Guyer et al., J Immunol. 117:587 (1976) and Kim et al., J Immunol. 24:249 (1994)). These antibodies comprise an Fc region with one or more substitutions that improve the binding of the Fc region to FcRn.

本發明DNA分子本發明亦係關於編碼本發明之抗體或抗原結合片段之經分離核酸序列。例如，可藉由Sambrook等人, 1989及Ausubel等人, 1989中闡述之技術或替代地藉由以化學方式合成來產生編碼本發明之抗體或抗原結合片段之經分離核酸序列。(例如Oligonucleotide Synthesis中(1984, Gait編輯, IRL Press, Oxford)闡述之技術)。表2中給定用於表現之抗體之DNA序列。在哺乳動物表現之某些情形中最佳化該等序列。本發明DNA分子不限於本文所揭示之序列，且亦包含其變體。可藉由參照DNA變體在雜交中之物理性質來闡述本發明內之DNA變體。熟習此項技術者將認識到，可使用核酸雜交技術將DNA用於鑑別其補體及其等效物或同系物，此乃因DNA為雙鏈。亦應認識到，雜交可在小於100%之互補性下發生。然而，鑒於適當選擇之條件，雜交技術可用於基於DNA結構與特定探針之相關性來區分DNA序列。有關條件之導則參見，Sambrook等人，1989 (見上文)及Ausubel等人，1995 (Ausubel, F. M., Brent, R., Kingston, R. E., Moore, D. D., Sedman, J. G., Smith, J. A., & Struhl, K.編輯(1995). Current Protocols in Molecular Biology. New York: John Wiley及Sons)。DNAmoleculesof the invention The invention also relates to isolated nucleic acid sequences encoding antibodies or antigen binding fragments of the invention. For example, isolated nucleic acid sequences encoding antibodies or antigen binding fragments of the invention may be generated by the techniques described in Sambrook et al., 1989 and Ausubel et al., 1989, or alternatively by chemical synthesis. (For example, the techniques described in Oligonucleotide Synthesis (1984, Gait ed., IRL Press, Oxford)). DNA sequences for antibodies used for expression are given in Table 2. These sequences are optimized in certain cases of mammalian expression. The DNA molecules of the invention are not limited to the sequences disclosed herein, and also include variants thereof. DNA variants within the invention may be described by reference to the physical properties of the DNA variants in hybridization. Those skilled in the art will recognize that DNA can be used to identify its complement and its equivalents or homologs using nucleic acid hybridization techniques because DNA is double-stranded. It should also be recognized that hybridization can occur with less than 100% complementarity. However, given the appropriate choice of conditions, hybridization techniques can be used to distinguish DNA sequences based on the association of the DNA structure with a particular probe. For guidance on relevant conditions, see Sambrook et al., 1989 (supra) and Ausubel et al., 1995 (Ausubel, FM, Brent, R., Kingston, RE, Moore, DD, Sedman, JG, Smith, JA, & Struhl, K., eds. (1995). Current Protocols in Molecular Biology. New York: John Wiley and Sons).

可依據兩種序列彼此雜交之條件之「嚴格性」來表示兩個多核苷酸序列之間之結構相似性。如本文中所使用，術語「嚴格性」係指條件不利於雜交之程度。嚴格條件強烈不利於雜交，且僅結構上最相關分子會在該等條件下彼此雜交。與之相反，非嚴格條件有利於顯示較低結構相關性程度之分子之雜交。由此，雜交嚴格度與之兩種核酸序列結構關係直接相關。The structural similarity between two polynucleotide sequences can be expressed in terms of the "strictness" of the conditions under which the two sequences hybridize to each other. As used herein, the term "strictness" refers to the degree to which the conditions are unfavorable for hybridization. Strict conditions strongly disfavor hybridization, and only the most structurally related molecules will hybridize to each other under such conditions. In contrast, non-strict conditions favor hybridization of molecules that exhibit a lower degree of structural relatedness. Thus, the stringency of hybridization is directly related to the structural relationship of the two nucleic acid sequences.

雜交嚴格度係許多因素之函數，該等因素包含整體DNA濃度、離子強度、溫度、探針大小及破壞氫鍵結之試劑之存在。促進雜交之因素包含高DNA濃度、高離子強度、低溫、較長探針大小及不存在破壞氫鍵結之藥劑。雜交通常分兩個階段實施：「結合」階段及「洗滌」階段。Hybridization severity is a function of many factors, including overall DNA concentration, ionic strength, temperature, probe size, and the presence of agents that disrupt hydrogen bonds. Factors that promote hybridization include high DNA concentration, high ionic strength, low temperature, longer probe size, and the absence of agents that disrupt hydrogen bonds. Hybridization is usually performed in two phases: a "binding" phase and a "washing" phase.

功能等效DNA變體可參考其編碼之產物來闡述本發明範圍內之又一種類之DNA變體。該等功能等效多核苷酸之特徵在於其因基因代碼之簡並性而編碼相同肽序列之事實。Functionally EquivalentDNAVariants Another class of DNA variants within the scope of the present invention can be described with reference to the products they encode. These functionally equivalent polynucleotides are characterized by the fact that they encode the same peptide sequence due to the degeneracy of the genetic code.

應瞭解，可以若干不同方式構造本文所提供之DNA分子之變體。舉例而言，可將其構造為完全合成DNA。有效地合成寡核苷酸之方法係廣泛可用的。參見Ausubel等人，第2.11部，增刊21 (1993)。可藉由Khorana等人，J. Mol. Biol. 72:209‑217 (1971)中首先報告之方式來合成及組裝重疊寡核苷；亦參見Ausubel等人(見上文)第8.2部。較佳地，藉助在基因5'及3'端改造之便利限制位點設計合成DNA以促進選殖至適當載體中。It will be appreciated that variants of the DNA molecules provided herein can be constructed in several different ways. For example, they can be constructed as completely synthetic DNA. Methods for efficiently synthesizing oligonucleotides are widely available. See Ausubel et al., Section 2.11, Supplement 21 (1993). Overlapping oligonucleotides can be synthesized and assembled in a manner first reported by Khorana et al., J. Mol. Biol. 72:209-217 (1971); see also Ausubel et al. (supra) Section 8.2. Preferably, the synthetic DNA is designed with convenient restriction sites engineered at the 5' and 3' ends of the gene to facilitate cloning into an appropriate vector.

如所指示，生成變體之方法係以本文所揭示之DNA中之一者開始且然後實施定點誘變。參見Ausubel等人(見上文)，第8章，增刊37 (1997)。在一典型方法中，將靶DNA選殖至單鏈DNA噬菌體載體中。將單鏈DNA與含有期望核苷酸變化之寡核苷酸分離及雜交。合成互補鏈且將雙鏈噬菌體引入宿主中。一些所得子代將含有期望突變體，其可使用DNA定序來證實。另外，可使用增加子代噬菌體變成期望突變體之機率之各種方法。該等方法為該領域之彼等所熟知且可自市面購得套組以生成該等突變體。As indicated, the method of generating variants starts with one of the DNAs disclosed herein and then performs site-directed mutagenesis. See Ausubel et al. (supra), Chapter 8, Supplement 37 (1997). In a typical method, the target DNA is colonized into a single-stranded DNA phage vector. The single-stranded DNA is separated and hybridized with an oligonucleotide containing the desired nucleotide changes. The complementary strands are synthesized and the double-stranded phage is introduced into the host. Some of the resulting progeny will contain the desired mutants, which can be confirmed using DNA sequencing. In addition, various methods that increase the probability of progeny phage becoming the desired mutant can be used. These methods are well known to those in the field and kits can be purchased commercially to generate these mutants.

重組DNA構築體及表現本發明進一步提供包括本發明核苷酸序列中之一或多者之重組DNA構築體。本發明之重組構築體可用於連結載體，例如質體、噬菌粒、噬菌體或病毒載體，該等載體中插入有編碼本發明抗體或其抗原結合片段或其變體之DNA分子。RecombinantDNAconstructs and expression The present invention further provides recombinant DNA constructs comprising one or more of the nucleotide sequences of the present invention. The recombinant constructs of the present invention can be used to link vectors, such as plasmids, phagemids, phages or viral vectors, into which DNA molecules encoding the antibodies of the present invention or antigen-binding fragments thereof or variants thereof are inserted.

因此，在一態樣中，本發明係關於包括本發明核酸序列之載體。Therefore, in one aspect, the present invention relates to a vector comprising a nucleic acid sequence of the present invention.

可在宿主細胞中藉由編碼輕鏈及重鏈或其部分之核酸序列之重組表現來製備本文所提供之抗體、抗原結合部分或其變體。為以重組方式表現抗體、抗原結合部分或其變體，可使用攜帶編碼輕鏈及/或重鏈或其部分之DNA片段之一或多種重組表現載體來轉染宿主細胞，從而使輕鏈及重鏈在宿主細胞中表現。使用標凖重組DNA方法以製備及/或獲得編碼重鏈及輕鏈之核酸、將該等核酸納入重組表現載體及將載體引入宿主細胞，例如Sambrook, Fritsch及Maniatis (編輯), Molecular Cloning；A Laboratory Manual, Second Edition, Cold Spring Harbor, N.Y., (1989), Ausubel, F. M.等人(編輯)Current Protocols in Molecular Biology, Greene Publishing Associates, (1989)及頒與Boss等人之美國專利第4,816,397號中闡述之方法The antibodies, antigen binding portions, or variants thereof provided herein can be prepared by recombinant expression of nucleic acid sequences encoding light and heavy chains or portions thereof in host cells. To recombinantly express an antibody, antigen binding portion, or variant thereof, one or more recombinant expression vectors carrying DNA fragments encoding light and/or heavy chains or portions thereof can be used to transfect host cells, thereby causing light and heavy chains to be expressed in host cells. Standard recombinant DNA methods are used to prepare and/or obtain nucleic acids encoding heavy and light chains, incorporate such nucleic acids into recombinant expression vectors, and introduce the vectors into host cells, such as those described in Sambrook, Fritsch and Maniatis (eds.), Molecular Cloning; A Laboratory Manual, Second Edition, Cold Spring Harbor, N.Y., (1989), Ausubel, F. M. et al. (eds.), Current Protocols in Molecular Biology, Greene Publishing Associates, (1989), and U.S. Patent No. 4,816,397 to Boss et al.

另外，可將編碼重鏈及/或輕鏈之可變區之核酸序列轉化為(舉例而言)編碼全長抗體鏈之核酸序列、Fab片段或scFv。可將VL-編碼或VH-編碼DNA片段操作性連接(從而使由兩種DNA片段編碼之胺基酸序列位於框內)至編碼(舉例而言)抗體恆定區或撓性連接體之另一DNA片段。人類重鏈及輕鏈恆定區之序列為業內已知(例如，參見Kabat, E. A.等人，(1991) Sequences of Proteins of Immunological Interest，第五版，U.S. Department of Health and Human Services, NIH公開案第91-3242號)且涵蓋該等區之DNA片段可藉由標準PCR擴增來獲得。In addition, nucleic acid sequences encoding the variable regions of the heavy and/or light chains can be converted to, for example, nucleic acid sequences encoding full-length antibody chains, Fab fragments, or scFv. A VL-encoding or VH-encoding DNA fragment can be operably linked (so that the amino acid sequences encoded by both DNA fragments are in frame) to another DNA fragment encoding, for example, an antibody constant region or a flexible linker. The sequences of human heavy and light chain constant regions are known in the art (e.g., see Kabat, E.A. et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242) and DNA fragments covering these regions can be obtained by standard PCR amplification.

為產生編碼scFv之多核苷酸序列，可將VH-編碼及VL-編碼核酸操作性連接至編碼撓性連接體之另一片段，從而使VH及VL序列可表現為連續單鏈蛋白質，其中VL及VH區域藉由撓性連接體接合(例如，參見Bird等人(1988)Science242:423-426；Huston等人 (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883；McCafferty等人，Nature (1990) 348:552-554)。To generate a polynucleotide sequence encoding an scFv, the VH-encoding and VL-encoding nucleic acids can be operably linked to another segment encoding a flexible linker so that the VH and VL sequences can be expressed as a continuous single protein chain in which the VL and VH regions are joined by a flexible linker (e.g., see Bird et al. (1988)Science 242:423-426; Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883; McCafferty et al., Nature (1990) 348:552-554).

可使用標準重組DNA表現方法來表現該等抗體、其抗原結合片段或其變體(例如，參見Goeddel; Gene Expression Technology.Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990))。舉例而言，可將編碼期望多肽之DNA插入表現載體，然後將該載體轉染至適宜宿主細胞。適宜宿主細胞係原核及真核細胞。原核宿主細胞之實例係(例如)細菌，真核宿主細胞之實例係酵母、昆蟲及昆蟲細胞、植物及植物細胞、轉基因動物或哺乳動物細胞。可使用標準技術(例如磷酸鈣轉染、DEAE右旋糖酐介導轉染、電穿孔、轉導或噬菌體感染)將重組構築體引入宿主細胞中。Standard recombinant DNA expression methods can be used to express the antibodies, antigen-binding fragments thereof, or variants thereof (see, e.g., Goeddel; Gene Expression Technology. Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990)). For example, DNA encoding the desired polypeptide can be inserted into an expression vector, which is then transfected into an appropriate host cell. Suitable host cells are prokaryotic and eukaryotic cells. Examples of prokaryotic host cells are, for example, bacteria, and examples of eukaryotic host cells are yeast, insects and insect cells, plants and plant cells, transgenic animals, or mammalian cells. Recombinant constructs can be introduced into host cells using standard techniques (e.g., calcium phosphate transfection, DEAE dextran-mediated transfection, electroporation, transduction, or phage infection).

在一些實施例中，將編碼重鏈及輕鏈之DNA插入單獨載體。在其他實施例中，將編碼重鏈及輕鏈之DNA插入相同載體。應理解，表現載體之設計(包含調控序列之選擇)受下列因素影響：例如宿主細胞之選擇、期望蛋白質之表現程度及表現是否為組成型或可誘導性。In some embodiments, DNA encoding the heavy chain and light chain are inserted into separate vectors. In other embodiments, DNA encoding the heavy chain and light chain are inserted into the same vector. It should be understood that the design of the expression vector (including the choice of regulatory sequences) is affected by factors such as the choice of host cells, the degree of expression of the desired protein, and whether the expression is constitutive or inducible.

因此，在另一態樣中，本發明係關於表現本發明之抗體或抗原結合片段及/或包括本發明核酸或本發明載體之經分離細胞。Therefore, in another aspect, the present invention relates to isolated cells expressing the antibodies or antigen-binding fragments of the present invention and/or comprising the nucleic acids of the present invention or the vectors of the present invention.

經分離細胞實際上可為可用於表現載體任何細胞。經分離細胞可為(例如)高等真核宿主細胞(例如哺乳動物細胞)、低等真核宿主細胞(例如酵母細胞)及可為原核細胞(例如細菌細胞)。The isolated cell can be virtually any cell that can be used to express the vector. The isolated cell can be, for example, a higher eukaryotic host cell (such as a mammalian cell), a lower eukaryotic host cell (such as a yeast cell), and can be a prokaryotic cell (such as a bacterial cell).

在另一態樣中，本發明係關於產生本發明之經分離抗體或抗原結合片段之方法，該方法包括培養本發明細胞。在特定實施例中，在用於抗體表現之適宜條件下培養本發明細胞且回收抗體或抗原結合片段。在特定實施例中，將抗體或抗原結合片段純化至(具體而言)至少95重量%均質性。In another aspect, the invention relates to a method of producing an isolated antibody or antigen-binding fragment of the invention, the method comprising culturing the cells of the invention. In a specific embodiment, the cells of the invention are cultured under suitable conditions for antibody expression and the antibody or antigen-binding fragment is recovered. In a specific embodiment, the antibody or antigen-binding fragment is purified to (specifically) at least 95% homogeneity by weight.

細菌表現藉由插入編碼期望蛋白質之DNA序列以及可操作讀取階段中之適宜轉譯開始及終止信號以及功能啟動子來構造用於細菌用途之有用表現載體。載體包括一或多種表型可選標記物及複製起點以確保載體之維持且(若需要)提供宿主內之擴增。用於轉化之適宜原核宿主包含(但不限於)大腸桿菌、枯草芽孢桿菌、鼠傷寒沙門桿菌以及假單胞菌、鏈黴菌及葡萄球菌屬中之各種物種。Bacterial Expression Useful expression vectors for bacterial use are constructed by inserting a DNA sequence encoding the desired protein and appropriate translation start and stop signals in operable read-through phase and a functional promoter. The vector includes one or more phenotypic selectable markers and an origin of replication to ensure maintenance of the vector and, if desired, provide for amplification within the host. Suitable prokaryotic hosts for transformation include, but are not limited to, E. coli, Bacillus subtilis, Salmonella typhimurium, and various species of Pseudomonas, Streptomyces, and Staphylococcus.

細菌載體可基於(舉例而言)噬菌體、質體或噬菌粒。該等載體可含有可選標記物及通常自含有熟知選殖載體pBR322 (ATCC 37017)之元件之市售質體衍生之細菌複製起點。在轉變適宜宿主菌株且使宿主菌株生長至適當細胞密度後，藉由適當方式去抑制/誘導選擇啟動子(例如，溫度變化或化學誘導)且將細胞額外培養一段時間。通常藉由離心收穫細胞，藉由物理或化學方式破壞細胞，且保留所得粗製提取物以進一步純化。Bacterial vectors may be based on, for example, phages, plasmids or phagemids. Such vectors may contain a selectable marker and a bacterial origin of replication usually derived from a commercially available plasmid containing elements of the well-known selection vector pBR322 (ATCC 37017). After transformation of a suitable host strain and growth of the host strain to an appropriate cell density, the selection promoter is repressed/induced by appropriate means (e.g., temperature shift or chemical induction) and the cells are cultured for an additional period of time. The cells are usually harvested by centrifugation, disrupted by physical or chemical means, and the resulting crude extract is retained for further purification.

在細菌系統中，可端視所表現分子之既定用途有利地選擇多種表現載體。例如，在欲產生大量該蛋白質時，對於抗體或篩選肽文庫之生成，舉例而言，可期望引導表現大量易純化融合蛋白質產物之載體。In bacterial systems, a variety of expression vectors may be advantageously selected depending on the intended use of the expressed molecule. For example, when large quantities of the protein are to be produced, for example, for the generation of antibodies or for screening peptide libraries, it may be desirable to direct the expression of a vector that expresses large quantities of a fusion protein product that is easily purified.

因此，本發明之一實施例係包括編碼本發明新穎抗體之核酸序列之表現載體。Therefore, one embodiment of the present invention includes an expression vector of a nucleic acid sequence encoding the novel antibody of the present invention.

本發明抗體或其抗原結合片段或其變體包含天然純化產物、化學合成程序之產物及藉由重組技術自原核宿主產生之產物，該原核宿主包含(例如)大腸桿菌、枯草芽孢桿菌、鼠傷寒沙門桿菌以及假單胞菌、鏈黴菌及葡萄球菌屬中之各種物種，較佳地來自大腸桿菌細胞。The antibodies or antigen-binding fragments or variants thereof of the present invention include naturally purified products, products of chemical synthesis processes, and products produced by recombinant technology from prokaryotic hosts, such as Escherichia coli, Bacillus subtilis, Salmonella typhimurium, and various species of Pseudomonas, Streptococcus and Staphylococcus, preferably from Escherichia coli cells.

哺乳動物表現用於哺乳動物宿主細胞表現之較佳調控序列包含在哺乳動物細胞中引導高程度之蛋白表現之病毒元件，例如衍生自巨細胞病毒(CMV) (例如CMV啟動子/增強子)、猿類病毒40 (SV40) (例如SV40啟動子/增強子)、腺病毒(例如，腺病毒主要晚期啟動子(AdMLP))及多瘤病毒之啟動子及/或增強子。抗體之表現可為組成型或調控型(例如可藉由添加或去除小分子誘導物(例如聯合Tet系統之四環素(Tetracyclin))來誘導)。關於病毒調控元件及其序列之進一步闡述，例如參見頒予Stinski之美國5,168,062、頒予Bell等人之美國4,510,245及頒予Schaffner等人之美國4,968,615。重組表現載體亦可包含複製起點及可選標記物(參見，例如美國4,399,216、4,634,665及美國5,179,017)。在已引入載體之宿主細胞中，適宜可選標記物包含賦予藥物抗性之基因，例如G418、嘌呤黴素(puromycin)、潮黴素(hygromycin)、殺稻瘟素(blasticidin)、吉歐黴素(zeocin)/博來黴素(bleomycin)或胺甲喋呤(methotrexate)或採用營養缺陷型之可選標記物(例如麩醯胺酸合成酶) (Bebbington等人， Biotechnology (N Y). 1992年2月；10(2):169-75)。舉例而言，二氫葉酸還原酶(DHFR)基因賦予胺甲喋呤抗性，neo基因賦予G418抗性，來自黑麯黴(Aspergillus terreus)之bsd基因賦予殺稻瘟素(blasticidin)抗性，嘌呤黴素N-乙醯基-轉移酶賦予嘌呤黴素抗性，Sh ble基因產物賦予吉歐黴素抗性且大腸桿菌潮黴素抗性基因(hyg或hph)賦予潮黴素抗性。可選標記物(如DHFR或麩醯胺酸合成酶)亦可用於聯合MTX及MSX之擴增技術。Mammalian Expression Preferred regulatory sequences for mammalian host cell expression include viral elements that direct high levels of protein expression in mammalian cells, such as promoters and/or enhancers derived from cytomegalovirus (CMV) (e.g., CMV promoter/enhancer), simian virus 40 (SV40) (e.g., SV40 promoter/enhancer), adenovirus (e.g., adenovirus major late promoter (AdMLP)), and polyoma virus. Expression of the antibody can be constitutive or regulated (e.g., can be induced by the addition or removal of small molecule inducers (e.g., tetracyclin associated with the Tet system)). For further description of viral regulatory elements and their sequences, see, e.g., U.S. Pat. Nos. 5,168,062 to Stinski, 4,510,245 to Bell et al., and 4,968,615 to Schaffner et al. Recombinant expression vectors may also include an origin of replication and a selectable marker (see, e.g., U.S. Pat. Nos. 4,399,216, 4,634,665, and 5,179,017). Suitable selectable markers include genes that confer drug resistance, such as G418, puromycin, hygromycin, blasticidin, zeocin/bleomycin, or methotrexate, or selectable markers that confer nutritional deficiencies, such as glutamine synthetase, in host cells into which the vector has been introduced (Bebbington et al., Biotechnology (NY). 1992 Feb;10(2):169-75). For example, the dihydrofolate reductase (DHFR) gene confers methotrexate resistance, the neo gene confers G418 resistance, the bsd gene from Aspergillus terreus confers blasticidin resistance, puromycin N-acetyl-transferase confers puromycin resistance, the Sh ble gene product confers genomycin resistance and the Escherichia coli hygromycin resistance gene (hyg or hph) confers hygromycin resistance. Selectable markers (such as DHFR or glutamine synthetase) can also be used in combination with MTX and MSX amplification techniques.

可使用標準技術(例如電穿孔、核轉染、磷酸鈣沈澱、脂轉染、基於聚陽離子之轉染(例如基於聚乙烯亞胺(PEI)之轉染)及DEAE-右旋糖酐轉染)將表現載體轉染至宿主細胞。The expression vector can be transfected into host cells using standard techniques such as electroporation, nucleofection, calcium phosphate precipitation, lipofection, polycation-based transfection such as polyethyleneimine (PEI)-based transfection, and DEAE-dextran transfection.

用於表現本文所提供之抗體、其抗原結合片段或其變體之適宜哺乳動物宿主細胞包含中國倉鼠卵巢(CHO細胞) (例如CHO-K1、CHO-S、CHO-K1SV) [包含dhfr- CHO細胞，闡述於Urlaub及Chasin, (1980) Proc. Natl. Acad. Sci. USA 77:4216-4220及Urlaub等人，Cell. 1983 Jun;33(2):405-12中，其與(例如) R. J. Kaufman及P. A. Sharp (1982) Mol. Biol. 159:601-621中闡述之DHFR可選標記物一起使用；以及Fan等人，Biotechnol Bioeng. 2012年4月；109(4):1007-15中例示之其他敲除細胞]、NS0骨髓瘤細胞、COS細胞、HEK293細胞、HKB11細胞、BHK21細胞、CAP細胞、EB66細胞及SP2細胞。Suitable mammalian host cells for expressing the antibodies, antigen-binding fragments thereof, or variants thereof provided herein include Chinese hamster ovary (CHO cells) (e.g., CHO-K1, CHO-S, CHO-K1SV) [including dhfr- CHO cells, described in Urlaub and Chasin, (1980) Proc. Natl. Acad. Sci. USA 77:4216-4220 and Urlaub et al., Cell. 1983 Jun;33(2):405-12, used with a DHFR selectable marker as described, for example, in R. J. Kaufman and P. A. Sharp (1982) Mol. Biol. 159:601-621; and Fan et al., Biotechnol Bioeng. 2012 Apr;109(4):1007-15], NS0 myeloma cells, COS cells, HEK293 cells, HKB11 cells, BHK21 cells, CAP cells, EB66 cells, and SP2 cells.

在表現系統(例如HEK293、HEK293T、HEK293-EBNA、HEK293E、HEK293-6E、HEK293-Freestyle、HKB11、Expi293F、293EBNALT75、CHOFreestyle、CHO-S、CHO-K1、CHO-K1SV、CHOEBNALT85、CHOS-XE、CHO-3E7或CAP-T細胞)中表現亦可為瞬時或半穩定的(例如Durocher等人，Nucleic Acids Res. 2002年1月15日；30(2):E9)。Expression in expression systems (e.g., HEK293, HEK293T, HEK293-EBNA, HEK293E, HEK293-6E, HEK293-Freestyle, HKB11, Expi293F, 293EBNALT75, CHOFreestyle, CHO-S, CHO-K1, CHO-K1SV, CHOEBNALT85, CHOS-XE, CHO-3E7, or CAP-T cells) can also be transient or semi-stable (e.g., Durocher et al., Nucleic Acids Res. 2002 Jan 15;30(2):E9).

在一些實施例中，表現載體經設計，從而使表現蛋白質分泌至宿主細胞生長之培養基中。可使用標準蛋白質純化方法自培養基回收抗體、其抗原結合片段或其變體。In some embodiments, the expression vector is designed so that the expressed protein is secreted into the culture medium in which the host cells are grown. The antibody, antigen-binding fragment thereof, or variant thereof can be recovered from the culture medium using standard protein purification methods.

純化可藉由熟知方法自重組細胞培養物回收並純化本發明抗體或其抗原結合片段或其變體，該等方法包含(但不限於)硫酸銨或乙醇沈澱、酸提取、蛋白質A層析、蛋白質G層析、陰離子或陽離子交換層析、磷酸-纖維素層析、疏水性相互作用層析、親和力層析、羥基磷灰石層析及凝集素層析。高效液相液相層析(「HPLC」)亦可用於純化。例如參見Colligan，Current Protocols in Immunology或Current Protocols in Protein Science, John Wiley & Sons, NY, N.Y., (1997-2001)，例如，第1、4、6、8、9、10章，其每一者以引用方式完全併入本文中。Purification The antibodies of the present invention or antigen-binding fragments or variants thereof can be recovered and purified from recombinant cell cultures by well-known methods, including but not limited to ammonium sulfate or ethanol precipitation, acid extraction, protein A chromatography, protein G chromatography, anion or cation exchange chromatography, phospho-cellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxyapatite chromatography, and lectin chromatography. High performance liquid chromatography ("HPLC") can also be used for purification. See, e.g., Colligan, Current Protocols in Immunology or Current Protocols in Protein Science, John Wiley & Sons, NY, NY, (1997-2001), e.g., Chapters 1, 4, 6, 8, 9, 10, each of which is incorporated herein by reference in its entirety.

本發明抗體或其抗原結合片段或其變體包含天然純化產物、化學合成程序之產物及藉由重組技術自真核宿主(包含(例如)酵母、高等植物、昆蟲及哺乳動物細胞)產生之產物。端視重組產生程序中採用之宿主，本發明抗體可為醣基化或可為非醣基化。該等方法闡述於許多標準實驗室手冊中，例如Sambrook (見上文)，第17.37-17.42部；Ausubel (見上文)，第10、12、13、16、18及20章。The antibodies or antigen-binding fragments or variants thereof of the present invention include naturally purified products, products of chemical synthesis procedures, and products produced by recombinant techniques from eukaryotic hosts (including, for example, yeast, higher plants, insects and mammalian cells). Depending on the host employed in the recombinant production procedure, the antibodies of the present invention may be glycosylated or may be non-glycosylated. Such methods are described in many standard laboratory manuals, such as Sambrook (supra), Sections 17.37-17.42; Ausubel (supra), Chapters 10, 12, 13, 16, 18 and 20.

在較佳實施例中，將抗體純化至(1)如(例如)藉由Lowry方法、UV-Vis光譜術或藉由SDS-毛細管凝膠電泳(例如在Caliper LabChip GXII、GX 90或Biorad Bioanalyzer裝置上)測定，大於95重量%之抗體，且在其他較佳實施例中大於99重量%，(2)足以獲得至少15個N-末端或內部胺基酸序列之殘基之程度，或(3)均質性，如藉由SDS-PAGE在還原或非還原條件下使用考馬斯藍色或(較佳地)銀染色所量測。經分離天然抗體包含重組細胞內之原位抗體，此乃因抗體天然環境之至少一種組分將不存在。然而，通常藉由至少一個純化步驟來製備經分離抗體。In preferred embodiments, the antibody is purified to (1) greater than 95% by weight of the antibody as determined, for example, by the Lowry method, UV-Vis spectroscopy, or by SDS-capillary gel electrophoresis (e.g., on a Caliper LabChip GXII, GX 90, or Biorad Bioanalyzer apparatus), and in other preferred embodiments greater than 99% by weight, (2) a degree sufficient to obtain at least 15 residues of N-terminal or internal amino acid sequence, or (3) homogeneity as measured by SDS-PAGE under reducing or non-reducing conditions using Coomassie blue or (preferably) silver staining. Isolated native antibodies include antibodies in situ within recombinant cells because at least one component of the antibody's natural environment will not be present. However, isolated antibodies are usually prepared by at least one purification step.

抗體結合物除裸抗體或其抗原結合片段外，可使用本文所揭示之抗體或抗原結合片段來設計各種基於抗體或基於抗體片段之結合物。Antibody Conjugates In addition to naked antibodies or antigen-binding fragments thereof, various antibody-based or antibody-fragment-based conjugates can be designed using the antibodies or antigen-binding fragments disclosed herein.

在一態樣中，本發明係關於靶向GPC3之抗體結合物，其包括本發明之經分離抗體或抗原結合片段。該等結合物可為用於診斷、療法、研究應用及各種其他目的之結合物。可將抗體或其抗原結合片段(例如)結合至放射性核素、細胞毒性劑、有機化合物、蛋白質毒素、免疫調節劑(例如細胞介素)、螢光部分、細胞、其他抗體或其抗原結合片段。In one aspect, the present invention relates to antibody conjugates targeting GPC3, which include isolated antibodies or antigen-binding fragments of the present invention. Such conjugates can be conjugates for diagnosis, therapy, research applications, and various other purposes. Antibodies or antigen-binding fragments thereof can be conjugated, for example, to radionuclides, cytotoxic agents, organic compounds, protein toxins, immunomodulators (e.g., cytokines), fluorescent moieties, cells, other antibodies or antigen-binding fragments thereof.

在一些其他實施例中，結合物包括細胞毒性劑(例如)以形成抗體藥物結合物(ADC)。舉例而言，細胞毒性劑可選自奧裡斯他汀(auristatin)、類美登素(maytansinoid)、驅動蛋白-紡錘體蛋白質(KSP)抑制劑、菸鹼醯胺磷酸核糖轉移酶(NAMPT)抑制劑或吡咯並苯并二氮呯衍生物。In some other embodiments, the conjugate includes a cytotoxic agent, for example, to form an antibody drug conjugate (ADC). For example, the cytotoxic agent can be selected from auristatin, maytansinoid, kinesin-spindle protein (KSP) inhibitor, nicotinamide phosphoribosyltransferase (NAMPT) inhibitor, or pyrrolobenzodiazepine derivatives.

ADC之細胞毒性及/或細胞生長抑制劑可為已知抑制細胞生長及/或複製及/或殺死細胞之任何試劑。許多具有細胞毒性及/或細胞生長抑制性質之試劑可於文獻中獲知。細胞毒性及/或細胞生長抑制劑之種類之非限制性實例包含(舉例而言且不限於)細胞週期調節劑、細胞凋亡調節劑、激酶抑制劑、蛋白質合成抑制劑、烷基化劑、DNA交聯劑、嵌入劑、線粒體抑制劑、出核轉運抑制劑、拓撲異構酶I抑制劑、拓撲異構酶II抑制劑、RNA/DNA抗代謝物及抗有絲分裂劑。The cytotoxic and/or cytostatic agent of the ADC may be any agent known to inhibit cell growth and/or replication and/or kill cells. Many agents with cytotoxic and/or cytostatic properties are known in the literature. Non-limiting examples of types of cytotoxic and/or cytostatic agents include, by way of example and not limitation, cell cycle regulators, apoptosis regulators, kinase inhibitors, protein synthesis inhibitors, alkylating agents, DNA cross-linking agents, intercalating agents, mitochondrial inhibitors, nuclear export inhibitors, topoisomerase I inhibitors, topoisomerase II inhibitors, RNA/DNA anti-metabolites, and anti-mitotic agents.

連接ADC之細胞毒性及/或細胞生長抑制與靶向部分之連接體在性質上可為較長、較短、撓性、剛性、親水或疏水，或可包括具有不同特性之區段(例如撓性區段、剛性區段等)。連接體可對細胞外環境化學穩定(例如在血流中化學穩定)，或可包含在細胞外環境中不穩定且釋放細胞毒性及/或細胞生長抑制之鍵聯。業內已知可用於將藥物連接至抗原結合部分(例如在ADC背景下之抗體)之眾多種連接體。The linker conjugating the cytotoxic and/or cytostatic and targeting moieties of the ADC may be long, short, flexible, rigid, hydrophilic or hydrophobic in nature, or may include segments with different properties (e.g., flexible segments, rigid segments, etc.). The linker may be chemically stable to the extracellular environment (e.g., chemically stable in the bloodstream), or may contain linkages that are unstable in the extracellular environment and release cytotoxic and/or cytostatic activity. A wide variety of linkers are known in the art that can be used to conjugate a drug to an antigen binding moiety (e.g., an antibody in the context of an ADC).

連接至抗GPC3 ADC之靶向部分之細胞毒性及/或細胞生長抑制劑之數量(藥物對抗體比率：DAR)可不同且僅受靶向部分上之可用附接位點之數量及連接至單一連接體之試劑數量限制。通常，連接體將單一細胞毒性及/或細胞生長抑制劑連接至ADC之靶向部分。在包含多於單一細胞毒性及/或細胞生長抑制劑之ADC之實施例中，每一藥劑可相同或不同。The amount of cytotoxic and/or cytostatic agent linked to the targeting moiety of the anti-GPC3 ADC (drug to antibody ratio: DAR) can vary and is limited only by the number of available attachment sites on the targeting moiety and the number of agents linked to a single linker. Typically, a linker links a single cytotoxic and/or cytostatic agent to the targeting moiety of the ADC. In embodiments of the ADC comprising more than a single cytotoxic and/or cytostatic agent, each agent may be the same or different.

在一些其他實施例中，結合物係經改造以用於GPC3靶向之嵌合抗原受體結合物。最近，CAR T細胞因其臨床成功而受到關注且接收到FDA批准，參照WO2020/102240，其全部內容併入本文中。在CAR T細胞方式中，T細胞經改造以表現針對腫瘤細胞中存在之抗原具有特異性之CAR。然後將該等經改造T細胞再投與至相同患者。在注射之後，CAR T細胞識別靶細胞上之靶向抗原以誘導靶細胞死亡。表現嵌合抗原受體之T細胞(CAR T細胞)由此構成用於醫學用途(例如腫瘤治療)之新穎方式。嵌合抗原受體(CAR)係經設計以靶向特異性抗原(舉例而言，腫瘤抗原)之基因改造受體。此靶向可產生針對腫瘤之細胞毒性，舉例而言，從而使表現CAR之CAR T細胞可經由特異性腫瘤抗原靶向且殺死腫瘤。In some other embodiments, the conjugate is a chimeric antigen receptor conjugate engineered for GPC3 targeting. Recently, CAR T cells have received attention for their clinical success and received FDA approval, see WO2020/102240, the entire contents of which are incorporated herein. In the CAR T cell approach, T cells are engineered to express CARs that are specific for antigens present in tumor cells. The engineered T cells are then re-administered to the same patient. After injection, the CAR T cells recognize the targeted antigen on the target cells to induce target cell death. T cells expressing chimeric antigen receptors (CAR T cells) thus constitute a novel approach for medical purposes (e.g., tumor treatment). Chimeric antigen receptors (CARs) are genetically modified receptors designed to target specific antigens (e.g., tumor antigens). This targeting can produce cytotoxicity against tumors, for example, so that CAR T cells expressing the CAR can target and kill tumors via specific tumor antigens.

根據本發明，針對GPC3提供之抗體或抗原結合片段可用於改造CAR T細胞以特異性識別GPC3表現細胞。According to the present invention, antibodies or antigen-binding fragments provided against GPC3 can be used to engineer CAR T cells to specifically recognize GPC3-expressing cells.

較佳地，本發明CAR包括共刺激結構域(例如，CD137、CD28或CD134)以達成T細胞在活體內之延長活化。添加共刺激結構域可增強含有CAR之T細胞之活體內增殖及存活，且初始臨床數據已展示，該等構築體為疾病(例如癌症)之治療中之有前景治療劑。根據本發明，CAR T細胞可用於治療局部或全身性異常存在表現GPC3之細胞之任何疾病。Preferably, the CAR of the present invention includes a co-stimulatory domain (e.g., CD137, CD28, or CD134) to achieve prolonged activation of T cells in vivo. Adding a co-stimulatory domain can enhance the proliferation and survival of CAR-containing T cells in vivo, and initial clinical data have shown that these constructs are promising therapeutic agents in the treatment of diseases such as cancer. According to the present invention, CAR T cells can be used to treat any disease in which cells expressing GPC3 are abnormally present locally or systemically.

在一些其他實施例中，結合物為或包括雙特異性抗體或多特異性抗體。在一些較佳實施例中，雙特異性抗體包括至少一種Fc結構域。In some other embodiments, the conjugate is or includes a bispecific antibody or a multispecific antibody. In some preferred embodiments, the bispecific antibody includes at least one Fc domain.

在一些較佳實施例中，雙特異性抗體之第一結合部分係本發明之抗體或抗原結合片段且雙特異性抗體之第二結合部分與本發明之抗體或抗原結合片段相同或不同。In some preferred embodiments, the first binding moiety of the bispecific antibody is an antibody or antigen-binding fragment of the present invention and the second binding moiety of the bispecific antibody is the same as or different from the antibody or antigen-binding fragment of the present invention.

在一些其他實施例中，雙特異性抗體之第一結合部分係本發明之抗體或抗原結合片段且雙特異性抗體之第二結合部分係結合至細胞表面蛋白質(例如細胞類型特異性抗原)之抗體或抗原結合片段。在一些該等實施例中，雙特異性抗體之第二結合部分係靶向檢查點蛋白之抗體或抗原結合片段，例如抗PD1抗體或抗PD-L1抗體。適宜檢查點蛋白靶向抗體包含尼沃魯單抗(Nivolumab)、帕博利珠單抗(Pembrolizumab)、阿替珠單抗(Atezolizumab)、阿維魯單抗(Avelumab)、德瓦魯單抗(Durvalumab)、西米普利單抗(Cemiplimab)、多塔利單抗(Dostarlimab)或伊匹單抗(Ipilimumab)。在一些其他該等實施例中，雙特異性抗體之第二結合部分係HER2靶向抗體，例如曲妥珠單抗、帕妥珠單抗(Pertuzumab)及/或馬妥昔單抗(Margetuximab)。In some other embodiments, the first binding portion of the bispecific antibody is an antibody or antigen-binding fragment of the present invention and the second binding portion of the bispecific antibody is an antibody or antigen-binding fragment that binds to a cell surface protein (e.g., a cell type-specific antigen). In some of these embodiments, the second binding portion of the bispecific antibody is an antibody or antigen-binding fragment that targets a checkpoint protein, such as an anti-PD1 antibody or an anti-PD-L1 antibody. Suitable checkpoint protein targeting antibodies include Nivolumab, Pembrolizumab, Atezolizumab, Avelumab, Durvalumab, Cemiplimab, Dostarlimab or Ipilimumab. In some other such embodiments, the second binding portion of the bispecific antibody is a HER2 targeting antibody, such as Trastuzumab, Pertuzumab and/or Margetuximab.

製備雙特異性抗體或多特異性抗體之技術包含(但不限於)重組共表現兩個具有不同特異性之免疫球蛋白重鏈-輕鏈對(參見Milstein及Cuello, Nature 305: 537 (1983)、WO 93/08829及Traunecker等人，EMBO J. 10: 3655 (1991))及化學結合兩個不同單株抗體(參見Staerz等人 (1985) Nature 314(6012): 628-31)。亦可藉由以下方式來製備多特異性抗體：使兩個或更多個抗體或片段交聯(例如，參見美國專利第4,676,980號及Brennan等人，Science 229: 81 (1985))；使用白胺酸拉鍊產生雙特異性抗體(例如，參見Kostelny等人，J. Immunol., 148(5):15471553 (1992))；使用用於製備雙特異性抗體片段之雙鏈抗體技術(例如，參見Hollinger等人，Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993))；使用單鏈Fv (scFv)二聚體(例如，參見Gruber等人，J. Immunol. 152:5368 (1994))；製備三特異性抗體(例如，如Tutt等人，J. Immunol. 147: 60 (1991)中所述)；及控制Fab臂交換(cFAE)，如Labrijn AF等人， Proc Natl Acad Sci USA 2013；110:5145-50。Techniques for preparing bispecific or multispecific antibodies include, but are not limited to, recombining immunoglobulin heavy chain-light chain pairs that co-express two different specificities (see Milstein and Cuello, Nature 305: 537 (1983), WO 93/08829 and Traunecker et al., EMBO J. 10: 3655 (1991)) and chemically conjugating two different monoclonal antibodies (see Staerz et al. (1985) Nature 314(6012): 628-31). Multispecific antibodies can also be prepared by cross-linking two or more antibodies or fragments (e.g., see U.S. Patent No. 4,676,980 and Brennan et al., Science 229: 81 (1985)); using leucine zipper to produce bispecific antibodies (e.g., see Kostelny et al., J. Immunol., 148(5):15471553 (1992)); using bispecific antibody technology for preparing bispecific antibody fragments (e.g., see Hollinger et al., Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993)); using single-chain Fv (scFv) dimers (e.g., see Gruber et al., J. Immunol. 152:5368 (1994)); preparation of trispecific antibodies (e.g., as described in Tutt et al., J. Immunol. 147:60 (1991)); and controlled Fab arm exchange (cFAE) as described in Labrijn AF et al., Proc Natl Acad Sci USA 2013;110:5145-50.

在一些其他實施例中，結合物包括可檢測部分。可檢測部分之實例包含各種酶、輔基、螢光材料、發光材料、生物發光材料、放射性材料、正電子發射金屬、非放射性順磁性金屬離子及反應性部分。In some other embodiments, the conjugate includes a detectable moiety. Examples of detectable moieties include various enzymes, cofactors, fluorescent materials, luminescent materials, bioluminescent materials, radioactive materials, positron emitting metals, non-radioactive paramagnetic metal ions, and reactive moieties.

可使用業內已知之技術將可檢測物質直接或(例如經由業內已知之連接體或另一部分)間接偶合或結合至抗體或其片段。酶標記之實例包含螢光素酶(例如，螢火蟲螢光素酶及細菌螢光素酶；US4,737,456)、螢光素、2,3-二氫酞嗪二酮、蘋果酸鹽去氫酶、脲酶、過氧化物酶(例如辣根過氧化物酶(HRPO))、鹼性磷酸酶、β-半乳糖苷酶、乙醯膽鹼酯酶、葡萄糖澱粉酶、溶菌酶、醣氧化酶(例如，葡萄糖氧化酶、半乳糖氧化酶及葡萄糖-6-磷酸去氫酶)、雜環氧化酶(例如尿酸酶及黃嘌呤氧化酶)、乳過氧化物酶、微過氧化物酶及諸如此類。The detectable substance can be coupled or bound to the antibody or fragment thereof directly or indirectly (eg, via a linker or another moiety known in the art) using techniques known in the art. Examples of enzyme labels include luciferase (e.g., firefly luciferase and bacterial luciferase; U.S. Pat. No. 4,737,456), luciferin, 2,3-dihydrophthalazinedione, apple salt dehydrogenase, urease, peroxidase (e.g., horseradish peroxidase (HRPO)), alkaline phosphatase, β-galactosidase, acetylcholinesterase, glucoamylase, lysozyme, carbohydrate oxidase (e.g., glucose oxidase, galactose oxidase, and glucose-6-phosphate dehydrogenase), heterocyclic oxidase (e.g., uricase and xanthine oxidase), lactoperoxidase, microperoxidase, and the like.

適宜輔基複合物之實例包含鏈黴抗生物素蛋白(streptavidin)/生物素及抗生物素蛋白/生物素；適宜螢光材料之實例包含傘形酮、螢光素、異硫氰酸螢光素、羅丹明(rhodamine)、二氯三嗪基胺螢光素、丹磺醯氯或藻紅素；發光材料之實例包含發光胺；生物發光材料之實例包含螢光素酶、螢光素及水母素；且適宜放射性材料之實例包含125I、131I、111In或99mTc。Examples of suitable cofactor complexes include streptavidin/biotin and avidin/biotin; examples of suitable fluorescent materials include fluorescein, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; examples of luminescent materials include luminescent amines; examples of bioluminescent materials include luciferase, fluorescein and aequorin; and examples of suitable radioactive materials include 125I, 131I, 111In or 99mTc.

在一些實施例中，結合物可包括放射性核素，例如β顆粒、α顆粒、γ顆粒或歐傑電子發射體。舉例而言，放射性核素可選自由以下組成之群：⁴³Sc、⁴⁴Sc、⁴⁷Sc、⁸⁹Zr、⁹⁰Y、¹¹¹In、¹⁴⁹Tb、¹⁵²Tb、¹⁵⁵Tb、¹⁶¹Tb、¹⁶⁶Ho、¹⁷⁷Lu、¹⁸⁶Re、¹⁸⁸Re、²¹²Bi、²¹³Bi、²²⁵Ac、²²⁷Th及²³²Th。本發明適宜β發射體係例如⁶⁷Cu、⁸⁹Sr、⁸⁹Zr、⁹⁰Y、¹⁰⁵Rh、¹³¹I、¹⁴⁹Pm、¹⁶⁶Ho、¹⁷⁷Lu、¹⁸⁶Re、¹⁸⁸Re、¹⁹⁸Au。在本發明之一些較佳實施例中，放射性核素係⁸⁹Zr。適宜歐傑電子發射放射性核素係例如⁶⁷Ga、⁷¹Ge、⁷⁷Br、^99mTc、¹⁰³Pd、¹¹¹In、¹²³I、¹²⁵I、¹⁴⁰Nd、¹⁷⁸Ta、¹⁹³Pt、¹⁹⁵mPt、¹⁹⁷Hg。本發明適宜α發射體係例如²¹¹At、²¹²Pb、²¹³Bi、²²³Ra、²²⁴Ra、²²⁵Ac或²²⁷Th。In some embodiments, the conjugate may include a radionuclide, such as a beta particle, an alpha particle, a gamma particle, or an orger electron emitter. For example, the radionuclide may be selected from the group consisting of:⁴³ Sc,⁴⁴ Sc,⁴⁷ Sc,⁸⁹ Zr,⁹⁰ Y,¹¹¹ In,¹⁴⁹ Tb,¹⁵² Tb,¹⁵⁵ Tb,¹⁶¹ Tb,¹⁶⁶ Ho,¹⁷⁷ Lu,¹⁸⁶ Re,¹⁸⁸ Re,²¹² Bi,²¹³ Bi,²²⁵ Ac,²²⁷ Th, and²³² Th. Suitable beta emitters of the present invention are, for example,⁶⁷ Cu,⁸⁹ Sr,⁸⁹ Zr,⁹⁰ Y,¹⁰⁵ Rh,¹³¹ I,¹⁴⁹ Pm,¹⁶⁶ Ho,¹⁷⁷ Lu,¹⁸⁶ Re,¹⁸⁸ Re,¹⁹⁸ Au. In some preferred embodiments of the present invention, the radionuclide is⁸⁹ Zr. Suitable euger electron emitting radionuclides are, for example,⁶⁷ Ga,⁷¹ Ge,⁷⁷ Br,^99m Tc,¹⁰³ Pd,¹¹¹ In,¹²³ I,¹²⁵ I,¹⁴⁰ Nd,¹⁷⁸ Ta,¹⁹³ Pt,¹⁹⁵ mPt,¹⁹⁷ Hg. Suitable α-emitting bodies in the present invention are, for example,²¹¹ At,²¹² Pb,²¹³ Bi,²²³ Ra,²²⁴ Ra,²²⁵ Ac or²²⁷ Th.

在一些較佳實施例中，放射性核素係α-粒子-發射放射性核素，例如²²⁵Ac。In some preferred embodiments, the radionuclide is an alpha-particle-emitting radionuclide, such as²²⁵ Ac.

在一些尤佳實施例中，放射性核素係²²⁵Ac。In some particularly preferred embodiments, the radionuclide is²²⁵ Ac.

在一些較佳實施例中，結合物進一步包括螯合劑以固定(例如)如本文中其他處闡述之放射性核素。In some preferred embodiments, the conjugate further comprises a chelating agent to immobilize a radionuclide, for example, as described elsewhere herein.

本發明靶向放射性核素結合物根據另一態樣，本發明係關於包括本發明之抗GPC3抗體或其抗原結合片段及用於螯合放射性核素之螯合部分之靶向放射性核素結合物。Targeted Radionuclide Conjugate of the Present Invention According to another aspect, the present invention relates to a targeted radionuclide conjugate comprising the anti-GPC3 antibody or antigen-binding fragment thereof of the present invention and a chelating portion for chelating a radionuclide.

在本發明之此態樣之一些實施例中，螯合部分能夠複合α-發射放射性核素、β-發射放射性核素、γ-發射放射性核素或歐傑電子發射體。In some embodiments of this aspect of the invention, the chelating moiety is capable of complexing an alpha-emitting radionuclide, a beta-emitting radionuclide, a gamma-emitting radionuclide, or an Ogier electron emitter.

在本發明之此態樣之一些較佳實施例中，螯合部分能夠複合α-發射放射性核素。在一些實施例中，α-發射放射性核素可係(舉例而言，但不限於)錒(²²⁵Ac³⁺)、鐳(²³³Ra²⁺)、鉍(²¹³Bi³⁺、²¹²Bi³⁺或²¹¹Bi³⁺)、鉛(²¹²Pb²⁺或²¹²Pb⁴⁺)、釷(²²⁷Th⁴⁺或²²⁶Th⁴⁺)、鏑(¹⁵²Dy³⁺)或砹(²¹¹At⁺或²¹⁷At⁺)，在本發明之本態樣之一些極佳實施例中，複合至螯合部分之α-發射放射性核素係錒(²²⁵Ac)、較佳地²²⁵Ac³⁺。In some preferred embodiments of this aspect of the invention, the chelating moiety is capable of complexing an alpha-emitting radionuclide. In some embodiments, the α-emitting radionuclide can be (for example, but not limited to) thulium (²²⁵ Ac³⁺ ), radium (²³³ Ra²⁺ ), bismuth (²¹³ Bi³⁺ ,²¹² Bi³⁺ or²¹¹ Bi³⁺ ), lead (²¹² Pb²⁺ or²¹² Pb⁴⁺ ), thulium (²²⁷ Th⁴⁺ or²²⁶ Th⁴⁺ ), thulium (¹⁵² Dy³⁺ ) or astatine (²¹¹ At⁺ or²¹⁷ At⁺ ). In some very preferred embodiments of this aspect of the invention, the α-emitting radionuclide complexed to the chelating portion is thulium (²²⁵ Ac 3+ ), preferably²²⁵ Ac³⁺ .

在本發明之此態樣之一些較佳實施例中，能夠複合錒之螯合部分包括18員螯合部分。在本發明之一些其他較佳實施例中，螯合部分包括大環macropa螯合劑N,N′-雙[(6-羧基-2-吡啶基)甲基]-4,13-二氮雜-18-冠-6。在本發明之一些極佳實施例中，螯合部分包括螯合劑Macropa-NCS (6-[[16-[(6-羧基-2-吡啶基)甲基]-1,4,10,13-四氧雜-7,16-二氮雜環十八-7-基]甲基]-4-[2-(4-異硫基氰基苯基)乙氧基]吡啶-2-甲酸) (在實例中縮寫為「Macropa」)，例如WO2020/106886中所闡述。In some preferred embodiments of this aspect of the invention, the chelating moiety capable of complexing phthalide comprises an 18-membered chelating moiety. In some other preferred embodiments of the invention, the chelating moiety comprises the macrocyclic macropa chelator N,N′-bis[(6-carboxy-2-pyridyl)methyl]-4,13-diaza-18-crown-6. In some excellent embodiments of the present invention, the chelating portion includes the chelating agent Macropa-NCS (6-[[16-[(6-carboxy-2-pyridyl)methyl]-1,4,10,13-tetraoxa-7,16-diazacyclooctadeca-7-yl]methyl]-4-[2-(4-isothiocyanatophenyl)ethoxy]pyridine-2-carboxylic acid) (abbreviated as "Macropa" in the examples), such as described in WO2020/106886.

在本發明之一些較佳實施例中，能夠複合錒之螯合部分終止於異硫氰酸酯或羧酸部分以促進其與本發明之抗體或其抗原結合片段偶合。在使用macropa螯合劑時，macropa螯合劑之吡啶環中之一者通常經取代，從而使其可偶合至本發明之抗GPC3抗體或其抗原結合片段。此可為簡單官能基(例如，羧酸-COOH)或更複雜之化學結構，只要其能夠偶合至抗GPC3抗體或其抗原結合片段。在本發明之特定實施例中，macropa螯合劑之吡啶環上之取代基較佳地終止於異硫氰酸酯或羧酸部分。In some preferred embodiments of the present invention, the chelating moiety capable of complexing phthalide terminates in an isothiocyanate or carboxylic acid moiety to facilitate its coupling to the antibody or antigen-binding fragment thereof of the present invention. When a macropa chelator is used, one of the pyridine rings of the macropa chelator is typically substituted so that it can be coupled to the anti-GPC3 antibody or antigen-binding fragment thereof of the present invention. This can be a simple functional group (e.g., carboxylic acid -COOH) or a more complex chemical structure, as long as it is capable of coupling to the anti-GPC3 antibody or antigen-binding fragment thereof. In specific embodiments of the present invention, the substituents on the pyridine ring of the macropa chelator preferably terminate in an isothiocyanate or carboxylic acid moiety.

根據此態樣，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段。According to this aspect, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I) Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3.

在其他實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In other embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, wherein [Ab] is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在其他實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之本發明第一態樣之經分離抗體或抗原結合片段： i) 包括含有SEQ ID NO: 82之H-CDR1、含有SEQ ID NO: 83之H-CDR2及含有SEQ ID NO: 84之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 86之L-CDR1、含有SEQ ID NO: 87之L-CDR2及含有SEQ ID NO:88之L-CDR3之輕鏈抗原結合區域，或 ii)     包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，或 iii)    包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域，或 iv)    包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域，或 v) 包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域，或 vi)    包括含有SEQ ID NO: 182之H-CDR1、含有SEQ ID NO: 183之H-CDR2及含有SEQ ID NO: 184之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 186之L-CDR1、含有SEQ ID NO: 187之L-CDR2及含有SEQ ID NO:188之L-CDR3之輕鏈抗原結合區域。In other embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the isolated antibodies or antigen-binding fragments of the first aspect of the present invention comprising: i) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 82, H-CDR2 comprising SEQ ID NO: 83, and H-CDR3 comprising SEQ ID NO: 84, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 86, L-CDR2 comprising SEQ ID NO: 87, and L-CDR3 comprising SEQ ID NO: 88, or ii) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103, and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising SEQ ID NO: 106, an L-CDR1 comprising SEQ ID NO: 107, and an L-CDR3 comprising SEQ ID NO: 108, or iii) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 122, an H-CDR2 comprising SEQ ID NO: 123, and an H-CDR3 comprising SEQ ID NO: 124, and a light chain antigen binding region comprising an L-CDR1 comprising SEQ ID NO: 126, an L-CDR2 comprising SEQ ID NO: 127, and an L-CDR3 comprising SEQ ID NO: 128, or iv) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 142, an H-CDR2 comprising SEQ ID NO: 143, and an H-CDR3 comprising SEQ ID NO: 144, and a light chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 145, an H-CDR2 comprising SEQ ID NO: 146, and a H-CDR3 comprising SEQ ID NO: 147. 146, an L-CDR1 comprising SEQ ID NO: 147, and an L-CDR3 comprising SEQ ID NO: 148, or v) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 162, an H-CDR2 comprising SEQ ID NO: 163, and an H-CDR3 comprising SEQ ID NO: 164, and a light chain antigen binding region comprising an L-CDR1 comprising SEQ ID NO: 166, an L-CDR2 comprising SEQ ID NO: 167, and an L-CDR3 comprising SEQ ID NO: 168, or vi) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 182, an H-CDR2 comprising SEQ ID NO: 183, and an H-CDR3 comprising SEQ ID NO: 184, and a light chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 184, and a light chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 190 The light chain antigen binding region comprises an L-CDR1 of SEQ ID NO: 186, an L-CDR2 of SEQ ID NO: 187, and an L-CDR3 of SEQ ID NO: 188.

在其他實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之本發明第一態樣之經分離抗體或抗原結合片段： i) 包括含有SEQ ID NO: 82之H-CDR1、含有SEQ ID NO: 83之H-CDR2及含有SEQ ID NO: 84之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 86之L-CDR1、含有SEQ ID NO: 87之L-CDR2及含有SEQ ID NO:88之L-CDR3之輕鏈抗原結合區域，或 ii)     包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，或 iii)    包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域，或 iv)    包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域，或 v) 包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域，或 vi)    包括含有SEQ ID NO: 182之H-CDR1、含有SEQ ID NO: 183之H-CDR2及含有SEQ ID NO: 184之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 186之L-CDR1、含有SEQ ID NO: 187之L-CDR2及含有SEQ ID NO:188之L-CDR3之輕鏈抗原結合區域， 其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In other embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the isolated antibodies or antigen-binding fragments of the first aspect of the present invention comprising: i) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 82, H-CDR2 comprising SEQ ID NO: 83, and H-CDR3 comprising SEQ ID NO: 84, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 86, L-CDR2 comprising SEQ ID NO: 87, and L-CDR3 comprising SEQ ID NO: 88, or ii) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103, and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising SEQ ID NO: 106, an L-CDR1 comprising SEQ ID NO: 107, and an L-CDR3 comprising SEQ ID NO: 108, or iii) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 122, an H-CDR2 comprising SEQ ID NO: 123, and an H-CDR3 comprising SEQ ID NO: 124, and a light chain antigen binding region comprising an L-CDR1 comprising SEQ ID NO: 126, an L-CDR2 comprising SEQ ID NO: 127, and an L-CDR3 comprising SEQ ID NO: 128, or iv) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 142, an H-CDR2 comprising SEQ ID NO: 143, and an H-CDR3 comprising SEQ ID NO: 144, and a light chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 145, an H-CDR2 comprising SEQ ID NO: 146, and a H-CDR3 comprising SEQ ID NO: 147. 146, an L-CDR1 comprising SEQ ID NO: 147, and an L-CDR3 comprising SEQ ID NO: 148, or v) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 162, an H-CDR2 comprising SEQ ID NO: 163, and an H-CDR3 comprising SEQ ID NO: 164, and a light chain antigen binding region comprising an L-CDR1 comprising SEQ ID NO: 166, an L-CDR2 comprising SEQ ID NO: 167, and an L-CDR3 comprising SEQ ID NO: 168, or vi) a heavy chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 182, an H-CDR2 comprising SEQ ID NO: 183, and an H-CDR3 comprising SEQ ID NO: 184, and a light chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 184, and a light chain antigen binding region comprising an H-CDR1 comprising SEQ ID NO: 190 186, a light chain antigen binding region comprising an L-CDR1 of SEQ ID NO: 187, and an L-CDR3 comprising SEQ ID NO: 188, wherein [Ab] is bound to the remainder of the molecule via the terminal amine group of the lysine residue of the monoclonal antibody or an antigen-binding fragment thereof.

在一些較佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，或 ii)     包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域，或 iii)    包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域，或 iv)    包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103, and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 106, L-CDR2 comprising SEQ ID NO: 107, and L-CDR3 comprising SEQ ID NO: 108, or ii) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 122, H-CDR2 comprising SEQ ID NO: 123, and H-CDR3 comprising SEQ ID NO: 124, and a light chain antigen-binding region comprising SEQ ID NO: 146, L-CDR2 containing SEQ ID NO: 147, and L-CDR3 containing SEQ ID NO: 148; or iv) a heavy chain antigen-binding region comprising a H-CDR1 comprising SEQ ID NO: 162, a H-CDR2 containing SEQ ID NO: 163, and a H-CDR3 containing SEQ ID NO: 164, and a light chain antigen-binding region comprising a H-CDR1 comprising SEQ ID NO: 165, a H-CDR2 containing SEQ ID NO: 166, and a light chain antigen-binding region comprising a H-CDR3 containing SEQ ID NO: 167; or iii) a heavy chain antigen-binding region comprising a H-CDR1 comprising SEQ ID NO: 142, a H-CDR2 containing SEQ ID NO: 143, and a H-CDR3 containing SEQ ID NO: 144, and a light chain antigen-binding region comprising a L-CDR1 comprising SEQ ID NO: 146, a L-CDR2 containing SEQ ID NO: 147, and a L-CDR3 containing SEQ ID NO: 148. The light chain antigen binding region comprises an L-CDR1 of SEQ ID NO: 166, an L-CDR2 of SEQ ID NO: 167, and an L-CDR3 of SEQ ID NO: 168.

在一些較佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，或 ii)     包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域，或 iii)    包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域，或 iv)    包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域， 其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103, and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 106, L-CDR2 comprising SEQ ID NO: 107, and L-CDR3 comprising SEQ ID NO: 108, or ii) a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 122, H-CDR2 comprising SEQ ID NO: 123, and H-CDR3 comprising SEQ ID NO: 124, and a light chain antigen-binding region comprising SEQ ID NO: 146, L-CDR2 containing SEQ ID NO: 147, and L-CDR3 containing SEQ ID NO: 148; or iv) a heavy chain antigen-binding region comprising a H-CDR1 comprising SEQ ID NO: 162, a H-CDR2 containing SEQ ID NO: 163, and a H-CDR3 containing SEQ ID NO: 164, and a light chain antigen-binding region comprising a H-CDR1 comprising SEQ ID NO: 165, a H-CDR2 containing SEQ ID NO: 166, and a light chain antigen-binding region comprising a H-CDR3 containing SEQ ID NO: 167; or iii) a heavy chain antigen-binding region comprising a H-CDR1 comprising SEQ ID NO: 142, a H-CDR2 containing SEQ ID NO: 143, and a H-CDR3 containing SEQ ID NO: 144, and a light chain antigen-binding region comprising a L-CDR1 comprising SEQ ID NO: 146, a L-CDR2 containing SEQ ID NO: 147, and a L-CDR3 containing SEQ ID NO: 148. 166, a light chain antigen binding region comprising an L-CDR1 of SEQ ID NO: 167, and an L-CDR3 comprising SEQ ID NO: 168, wherein [Ab] is bound to the remainder of the molecule via the terminal amine group of the lysine residue of the monoclonal antibody or an antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, and the [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103, and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 106, L-CDR2 comprising SEQ ID NO: 107, and L-CDR3 comprising SEQ ID NO: 108.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, and the [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 122, H-CDR2 comprising SEQ ID NO: 123, and H-CDR3 comprising SEQ ID NO: 124, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 126, L-CDR2 comprising SEQ ID NO: 127, and L-CDR3 comprising SEQ ID NO: 128.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, and the [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 142, H-CDR2 comprising SEQ ID NO: 143, and H-CDR3 comprising SEQ ID NO: 144, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 146, L-CDR2 comprising SEQ ID NO: 147, and L-CDR3 comprising SEQ ID NO: 148.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, and the [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 162, H-CDR2 comprising SEQ ID NO: 163, and H-CDR3 comprising SEQ ID NO: 164, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 166, L-CDR2 comprising SEQ ID NO: 167, and L-CDR3 comprising SEQ ID NO: 168.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係單株抗體或其能夠結合至GPC3之抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, said [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103 and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 106, L-CDR2 comprising SEQ ID NO: 107 and L-CDR3 comprising SEQ ID NO: 108, wherein [Ab] is bound to the rest of the molecule via the terminal amine group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, said [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 122, H-CDR2 comprising SEQ ID NO: 123, and H-CDR3 comprising SEQ ID NO: 124, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 126, L-CDR2 comprising SEQ ID NO: 127, and L-CDR3 comprising SEQ ID NO: 128, wherein [Ab] is bound to the rest of the molecule via the terminal amine group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, said [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 142, H-CDR2 comprising SEQ ID NO: 143, and H-CDR3 comprising SEQ ID NO: 144, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 146, L-CDR2 comprising SEQ ID NO: 147, and L-CDR3 comprising SEQ ID NO: 148, wherein [Ab] is bound to the rest of the molecule via the terminal amine group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, said [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 162, H-CDR2 comprising SEQ ID NO: 163, and H-CDR3 comprising SEQ ID NO: 164, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 166, L-CDR2 comprising SEQ ID NO: 167, and L-CDR3 comprising SEQ ID NO: 168, wherein [Ab] is bound to the rest of the molecule via the terminal amine group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在其他較佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 81之可變重鏈結構域及包括SEQ ID NO: 85之可變輕鏈結構域；或 ii)     包括SEQ ID NO: 101之可變重鏈結構域及包括SEQ ID NO: 105之可變輕鏈結構域；或 iii)    包括SEQ ID NO: 121之可變重鏈結構域及包括SEQ ID NO: 125之可變輕鏈結構域；或 iv)    包括SEQ ID NO: 141之可變重鏈結構域及包括SEQ ID NO: 145之可變輕鏈結構域；或 v) 包括SEQ ID NO: 161之可變重鏈結構域及包括SEQ ID NO: 165之可變輕鏈結構域；或 vi)    包括SEQ ID NO: 181之可變重鏈結構域及包括SEQ ID NO: 185之可變輕鏈結構域。In other preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I) wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a variable heavy chain domain comprising SEQ ID NO: 81 and a variable light chain domain comprising SEQ ID NO: 85; or ii) a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105; or iii) a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125; or iv) a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145; or v) a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: or vi) a variable heavy chain domain comprising SEQ ID NO: 181 and a variable light chain domain comprising SEQ ID NO: 185.

在其他較佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 81之可變重鏈結構域及包括SEQ ID NO: 85之可變輕鏈結構域；或 ii)     包括SEQ ID NO: 101之可變重鏈結構域及包括SEQ ID NO: 105之可變輕鏈結構域；或 iii)    包括SEQ ID NO: 121之可變重鏈結構域及包括SEQ ID NO: 125之可變輕鏈結構域；或 iv)    包括SEQ ID NO: 141之可變重鏈結構域及包括SEQ ID NO: 145之可變輕鏈結構域；或 v) 包括SEQ ID NO: 161之可變重鏈結構域及包括SEQ ID NO: 165之可變輕鏈結構域；或 vi)    包括SEQ ID NO: 181之可變重鏈結構域及包括SEQ ID NO: 185之可變輕鏈結構域， 其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In other preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I) wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a variable heavy chain domain comprising SEQ ID NO: 81 and a variable light chain domain comprising SEQ ID NO: 85; or ii) a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105; or iii) a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125; or iv) a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145; or v) a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: or vi) a variable heavy chain domain comprising SEQ ID NO: 181 and a variable light chain domain comprising SEQ ID NO: 185, wherein [Ab] is bound to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 101之可變重鏈結構域及包括SEQ ID NO: 105之可變輕鏈結構域；或 ii)     包括SEQ ID NO: 121之可變重鏈結構域及包括SEQ ID NO: 125之可變輕鏈結構域；或 iii)    包括SEQ ID NO: 141之可變重鏈結構域及包括SEQ ID NO: 145之可變輕鏈結構域；或 iv)    包括SEQ ID NO: 161之可變重鏈結構域及包括SEQ ID NO: 165之可變輕鏈結構域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105; or ii) a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125; or iii) a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145; or iv) a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 101之可變重鏈結構域及包括SEQ ID NO: 105之可變輕鏈結構域；或 ii)     包括SEQ ID NO: 121之可變重鏈結構域及包括SEQ ID NO: 125之可變輕鏈結構域；或 iii)    包括SEQ ID NO: 141之可變重鏈結構域及包括SEQ ID NO: 145之可變輕鏈結構域；或 iv)    包括SEQ ID NO: 161之可變重鏈結構域及包括SEQ ID NO: 165之可變輕鏈結構域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105; or ii) a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125; or iii) a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145; or iv) a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165.

其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。wherein [Ab] is bound to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or its antigen-binding fragment.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 101之可變重鏈結構域及含有SEQ ID NO: 105之可變輕鏈結構域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 121之可變重鏈結構域及含有SEQ ID NO: 125之可變輕鏈結構域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 141之可變重鏈結構域及含有SEQ ID NO: 145之可變輕鏈結構域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 161之可變重鏈結構域及含有SEQ ID NO: 165之可變輕鏈結構域。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 101之可變重鏈結構域及含有SEQ ID NO: 105之可變輕鏈結構域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105, wherein [Ab] is bound to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 121之可變重鏈結構域及含有SEQ ID NO: 125之可變輕鏈結構域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125, wherein [Ab] is bound to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 141之可變重鏈結構域及含有SEQ ID NO: 145之可變輕鏈結構域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145, wherein [Ab] is bound to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 161之可變重鏈結構域及含有SEQ ID NO: 165之可變輕鏈結構域，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165, wherein [Ab] is bound to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在特定實施例中，本發明靶向錒結合物中包括之經分離單株抗體係IgG1、IgG2、IgG3或IgG4抗體。更具體而言，本發明經分離單株抗體係IgG1抗體。In a specific embodiment, the isolated monoclonal antibody included in the targeted tantalum conjugate of the present invention is an IgG1, IgG2, IgG3 or IgG4 antibody. More specifically, the isolated monoclonal antibody of the present invention is an IgG1 antibody.

在其他較佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 97之重鏈及包括SEQ ID NO: 98之輕鏈；或 ii)     包括SEQ ID NO: 117之重鏈及包括SEQ ID NO: 118之輕鏈；或 iii)    包括SEQ ID NO: 137之重鏈及包括SEQ ID NO: 138之輕鏈；或 iv)    包括SEQ ID NO: 157之重鏈及包括SEQ ID NO: 158之輕鏈；或 v) 包括SEQ ID NO: 177之重鏈及包括SEQ ID NO: 178之輕鏈；或 vi)    包括SEQ ID NO: 197之重鏈及包括SEQ ID NO: 198之輕鏈。In other preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I) Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a heavy chain comprising SEQ ID NO: 97 and a light chain comprising SEQ ID NO: 98; or ii) a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118; or iii) a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138; or iv) a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158; or v) a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178; or vi) a heavy chain comprising SEQ ID NO: 197 and a light chain comprising SEQ ID NO: 198.

在其他較佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 97之重鏈及包括SEQ ID NO: 98之輕鏈；或 ii)     包括SEQ ID NO: 117之重鏈及包括SEQ ID NO: 118之輕鏈；或 iii)    包括SEQ ID NO: 137之重鏈及包括SEQ ID NO: 138之輕鏈；或 iv)    包括SEQ ID NO: 157之重鏈及包括SEQ ID NO: 158之輕鏈；或 v) 包括SEQ ID NO: 177之重鏈及包括SEQ ID NO: 178之輕鏈；或 vi)    包括SEQ ID NO: 197之重鏈及包括SEQ ID NO: 198之輕鏈， 其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In other preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I) wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a heavy chain comprising SEQ ID NO: 97 and a light chain comprising SEQ ID NO: 98; or ii) a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118; or iii) a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138; or iv) a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158; or v) a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178; or vi) a heavy chain comprising SEQ ID NO: 197 and a light chain comprising SEQ ID NO: 198, wherein [Ab] is bound to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or its antigen-binding fragment.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 117之重鏈及包括SEQ ID NO: 118之輕鏈；或 ii)     包括SEQ ID NO: 137之重鏈及包括SEQ ID NO: 138之輕鏈；或 iii)    包括SEQ ID NO: 157之重鏈及包括SEQ ID NO: 158之輕鏈；或 iv)    包括SEQ ID NO: 177之重鏈及包括SEQ ID NO: 178之輕鏈。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, which is selected from the following isolated antibodies or antigen-binding fragments: i) a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118; or ii) a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138; or iii) a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158; or iv) a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其選自包括以下之經分離抗體或抗原結合片段： i) 包括SEQ ID NO: 117之重鏈及包括SEQ ID NO: 118之輕鏈；或 ii)     包括SEQ ID NO: 137之重鏈及包括SEQ ID NO: 138之輕鏈；或 iii)    包括SEQ ID NO: 157之重鏈及包括SEQ ID NO: 158之輕鏈；或 iv)    包括SEQ ID NO: 177之重鏈及包括SEQ ID NO: 178之輕鏈， 其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or antigen-binding fragment thereof capable of binding to GPC3, selected from the following isolated antibodies or antigen-binding fragments: i) a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118; or ii) a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138; or iii) a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158; or iv) a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178, wherein [Ab] is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 117之重鏈及含有SEQ ID NO: 118之輕鏈(TPP-29537)。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain containing SEQ ID NO: 117 and a light chain containing SEQ ID NO: 118 (TPP-29537).

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 137之重鏈及含有SEQ ID NO: 138之輕鏈(TPP-29538)。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain containing SEQ ID NO: 137 and a light chain containing SEQ ID NO: 138 (TPP-29538).

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 157之重鏈及含有SEQ ID NO: 158之輕鏈(TPP-29636)。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain containing SEQ ID NO: 157 and a light chain containing SEQ ID NO: 158 (TPP-29636).

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，其包括含有SEQ ID NO: 177之重鏈及含有SEQ ID NO: 178之輕鏈(TPP-29638)。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain containing SEQ ID NO: 177 and a light chain containing SEQ ID NO: 178 (TPP-29638).

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，包括含有SEQ ID NO: 117之重鏈及含有SEQ ID NO: 118之輕鏈(TPP-29537)，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118 (TPP-29537), wherein [Ab] is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，包括含有SEQ ID NO: 137之重鏈及含有SEQ ID NO: 138之輕鏈(TPP-29538)，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138 (TPP-29538), wherein [Ab] is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，包括含有SEQ ID NO: 157之重鏈及含有SEQ ID NO: 158之輕鏈(TPP-29636)，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158 (TPP-29636), wherein [Ab] is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

在一些極佳實施例中，本發明係關於式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，包括含有SEQ ID NO: 177之重鏈及含有SEQ ID NO: 178之輕鏈(TPP-29638)，其中[Ab]經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。In some preferred embodiments, the present invention relates to a targeted tantalum conjugate (TAC) of formula (I): Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, comprising a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178 (TPP-29638), wherein [Ab] is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or the antigen-binding fragment thereof.

本發明之式(I)之靶向錒結合物(TAC)包括含有經複合錒原子(Ac)之螯合部分。The targeted tantalum conjugate (TAC) of formula (I) of the present invention comprises a chelating portion containing a complexed tantalum atom (Ac).

在本發明之式(I)之靶向錒結合物(TAC)中，錒(Ac)原子經展示為複合至大環之四個氧及兩個氮原子以及兩個羧酸酯基團。儘管此為錒原子之假設複合模式，此繪圖包含其中該等鍵中之一或多者不存在之所有可能及可構想情形，例如，其中錒原子未結合至大環之所有雜原子或羧酸酯基團等In the targeted tantalum conjugate (TAC) of formula (I) of the present invention, the tantalum (Ac) atom is shown as being complexed to four oxygen and two nitrogen atoms and two carboxylate groups of the macrocycle. Although this is a hypothetical complexation pattern for the tantalum atom, this drawing includes all possible and conceivable situations in which one or more of these bonds are not present, for example, in which the tantalum atom is not bonded to all heteroatoms or carboxylate groups of the macrocycle, etc.

抗GPC3抗體或其抗原結合片段較佳地經由抗體之離胺酸殘基偶合至分子之其餘部分從而形成本發明靶向錒結合物。因此，錒複合螯合部分與抗GPC3抗體或其抗原結合片段可經由醯胺或硫脲部分偶合。在使用較佳大環螯合劑macropa時，末端異硫氰酸酯或羧酸部分與抗GPC3抗體或其抗原結合片段中之離胺酸殘基之末端胺基偶合，從而分別形成硫脲或醯胺基團。因此，包括macropa螯合劑之較佳錒複合螯合部分經由自該抗GPC3抗體或其抗原結合片段之離胺酸殘基的末端胺基衍生之硫脲或醯胺鍵附接至抗GPC3抗體或其抗原結合片段。The anti-GPC3 antibody or its antigen-binding fragment is preferably coupled to the rest of the molecule via the lysine residue of the antibody to form the targeted tantalum conjugate of the present invention. Therefore, the tantalum complex chelating portion and the anti-GPC3 antibody or its antigen-binding fragment can be coupled via an amide or thiourea portion. When the preferred macrocyclic chelator macropa is used, the terminal isothiocyanate or carboxylic acid portion is coupled to the terminal amine group of the lysine residue in the anti-GPC3 antibody or its antigen-binding fragment to form a thiourea or amide group, respectively. Therefore, the preferred tantalum complex chelating portion including the macropa chelator is attached to the anti-GPC3 antibody or its antigen-binding fragment via a thiourea or amide bond derived from the terminal amine group of the lysine residue of the anti-GPC3 antibody or its antigen-binding fragment.

本發明靶向錒結合物之合成在另一態樣中，本發明係關於製備式(I)之靶向錒結合物(TAC)之方法其中[Ab]係結合至GPC3之單株抗體或其抗原結合片段， 該方法包括 (i)    使式(II)之螯合部分與能夠結合至GPC3之單株抗體或其抗原結合片段偶合，及 (ii)   使所得產物與包含至少一種α-發射錒同位素之離子之水溶液接觸。Synthesis of the Targeted Taconjugate of the Present Invention In another aspect, the present invention relates to a method for preparing a targeted taconjugate (TAC) of formula (I) wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof that binds to GPC3, the method comprising (i) treating a chelating moiety of formula (II) with and (ii) contacting the resulting product with an aqueous solution containing ions of at least one α-emitting helium isotope.

在另一實施例中，本發明係關於製備式(I)之靶向錒結合物(TAC)之方法其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段， 該方法包括 (i)    使式(II)之螯合部分經由單株抗體或其抗原結合片段之離胺酸殘基與能夠結合至GPC3之單株抗體或其抗原結合片段偶合，及 (ii)   使所得產物與包含至少一種α-發射錒同位素之離子之水溶液接觸。In another embodiment, the present invention relates to a method for preparing a targeted tantalum conjugate (TAC) of formula (I) wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, the method comprising (i) treating a chelating moiety of formula (II) with The invention relates to a method for coupling a monoclonal antibody or an antigen-binding fragment thereof with a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3 via a lysine residue of the monoclonal antibody or an antigen-binding fragment thereof, and (ii) contacting the resulting product with an aqueous solution containing ions of at least one α-emitting helium isotope.

在另一實施例中，本發明係關於製備式(I)之靶向錒結合物(TAC)之方法其中[Ab]係選自TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642之抗GPC3抗體或其抗原結合片段， 該方法包括 (i)    使式(II)之螯合部分經由抗GPC3抗體或其抗原結合片段之離胺酸殘基與選自TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642之抗GPC3抗體或其抗原結合片段偶合，及 (ii)   使所得產物與包含至少一種α-發射錒同位素之離子之水溶液接觸。In another embodiment, the present invention relates to a method for preparing a targeted tantalum conjugate (TAC) of formula (I) wherein [Ab] is an anti-GPC3 antibody or antigen-binding fragment thereof selected from TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638 and TPP-29642, the method comprising (i) treating a chelating moiety of formula (II) with The invention relates to a method for coupling an anti-GPC3 antibody or an antigen-binding fragment thereof with an anti-GPC3 antibody or an antigen-binding fragment thereof selected from TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638 and TPP-29642 via a lysine residue of the anti-GPC3 antibody or an antigen-binding fragment thereof, and (ii) contacting the resulting product with an aqueous solution containing ions of at least one α-emitting helium isotope.

在另一實施例中，本發明係關於製備式(I)之靶向錒結合物(TAC)之方法其中[Ab]係選自TPP-29537、TPP-29538、TPP-29636或TPP-29638之抗GPC3抗體或其抗原結合片段， 該方法包括 (i)    使式(II)之螯合部分經由單株抗體或其抗原結合片段之離胺酸殘基與選自TPP-29537、TPP-29538、TPP-29636或TPP-29638之抗GPC3抗體或其抗原結合片段偶合，及 (ii)   使所得產物與包含至少一種α-發射錒同位素之離子之水溶液接觸。In another embodiment, the present invention relates to a method for preparing a targeted tantalum conjugate (TAC) of formula (I) wherein [Ab] is an anti-GPC3 antibody or antigen-binding fragment thereof selected from TPP-29537, TPP-29538, TPP-29636 or TPP-29638, the method comprising (i) treating a chelating moiety of formula (II) with The monoclonal antibody or antigen-binding fragment thereof is coupled to an anti-GPC3 antibody or antigen-binding fragment thereof selected from TPP-29537, TPP-29538, TPP-29636 or TPP-29638 via a lysine residue of the monoclonal antibody or antigen-binding fragment thereof, and (ii) contacting the resulting product with an aqueous solution containing ions of at least one α-emitting helium isotope.

在步驟(i)中，式(II)之螯合部分與能夠結合至GPC3之單株抗體或其抗原結合片段偶合。較佳地，抗GPC3抗體選自TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642或其抗原結合片段。更佳地，抗GPC3抗體係選自TPP-29537、TPP-29538、TPP-29636及TPP-29638或其抗原結合片段。最佳地，抗GPC3抗體係TPP-29537。In step (i), the chelating moiety of formula (II) Coupled to a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3. Preferably, the anti-GPC3 antibody is selected from TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638 and TPP-29642 or an antigen-binding fragment thereof. More preferably, the anti-GPC3 antibody is selected from TPP-29537, TPP-29538, TPP-29636 and TPP-29638 or an antigen-binding fragment thereof. Most preferably, the anti-GPC3 antibody is TPP-29537.

較佳地，式(II)之螯合部經由單株抗體之離胺酸殘基偶合至抗GPC3抗體或其抗原結合片段，由此形成連接螯合部分與抗GPC3抗體或其抗原結合片段之硫脲部分。Preferably, the chelating moiety of formula (II) is coupled to the anti-GPC3 antibody or its antigen-binding fragment via the lysine residue of the monomeric antibody, thereby forming a thiourea moiety linking the chelating moiety and the anti-GPC3 antibody or its antigen-binding fragment.

較佳地，在鹼性pH下(亦即，在高於7之pH下)實施步驟(i)中之偶合。更佳地，在8至10之pH、最佳地9之pH下實施步驟(i)中之偶合。Preferably, the coupling in step (i) is carried out at an alkaline pH, i.e. at a pH above 7. More preferably, the coupling in step (i) is carried out at a pH of 8 to 10, optimally at a pH of 9.

在步驟(i)之後，式(III)之抗體藥物結合物(ADC)得以生成。因此，在其他實施例中，本發明係關於式(III)之抗體藥物結合物其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段。After step (i), an antibody-drug conjugate (ADC) of formula (III) is produced. Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3.

較佳地，抗GPC3抗體或其抗原結合片段選自TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642或其抗原結合片段。更佳地，抗GPC3抗體係選自TPP-29537、TPP-29538、TPP-29636及TPP-29638或其抗原結合片段。最佳地，抗GPC3抗體係TPP-29537。Preferably, the anti-GPC3 antibody or an antigen-binding fragment thereof is selected from TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638 and TPP-29642 or an antigen-binding fragment thereof. More preferably, the anti-GPC3 antibody is selected from TPP-29537, TPP-29538, TPP-29636 and TPP-29638 or an antigen-binding fragment thereof. Most preferably, the anti-GPC3 antibody is TPP-29537.

在步驟(ii)中，將自步驟(i)中之偶合所得之產物與包括至少一種α-發射錒同位素之離子的水溶液接觸。較佳地，α-發射錒同位素係²²⁵Ac。較佳地，至少一種α-發射錒同位素之離子係具有三重正電荷(亦即，²²⁵Ac³⁺)之²²⁵Ac。本發明容許在室溫下執行步驟(ii)，亦即，在低於相應DOTA結合物之製備需要之溫度的溫度下，該製備需要較高溫度(60℃)。In step (ii), the product obtained from the coupling in step (i) is contacted with an aqueous solution comprising ions of at least one α-emitting ruthenium isotope. Preferably, the α-emitting ruthenium isotope is²²⁵ Ac. Preferably, the ions of at least one α-emitting ruthenium isotope is²²⁵ Ac having a triply positive charge (i.e.,²²⁵ Ac³⁺ ). The present invention allows step (ii) to be performed at room temperature, i.e., at a temperature lower than that required for the preparation of the corresponding DOTA conjugate, which requires a higher temperature (60°C).

治療方法治療方法涉及向有需要之個體投與治療治療有效量之本發明考慮之抗體或其抗原結合片段或其變體及/或靶向錒結合物(TAC)。Methods of Treatment The methods of treatment involve administering to a subject in need thereof a therapeutically effective amount of an antibody or antigen-binding fragment or variant thereof and/or a targeted tantalum conjugate (TAC) contemplated by the present invention.

用途在另一態樣中，本發明之抗體及/或靶向錒結合物(TAC)可用於治療人類或非人類動物個體之疾病。具體而言，本發明之抗體及/或靶向錒結合物(TAC)可用於治療增生性及/或贅瘤性疾病，例如癌症。在一實施例中，本發明之抗體及/或靶向錒結合物(TAC)具有針對GPC3之特異性及/或結合親和力。Uses In another aspect, the antibodies and/or targeted tantalum conjugates (TACs) of the present invention can be used to treat diseases in human or non-human animal subjects. Specifically, the antibodies and/or targeted tantalum conjugates (TACs) of the present invention can be used to treat proliferative and/or neoplastic diseases, such as cancer. In one embodiment, the antibodies and/or targeted tantalum conjugates (TACs) of the present invention have specificity and/or binding affinity for GPC3.

如本文所述之本發明之各態樣係關於疾病之治療，尤其用於患病組織之選擇性靶向，以及係關於可用於該等方法之複合物、結合物、藥劑、調配物、套組等。在所有態樣中，患病組織可駐留於體內單一位點處(例如在局部性實體瘤之情形中)或可駐留於複數個位點處(例如在關節炎中影響若干個關節時或在分散性或轉移性癌性疾病之情形中)。As described herein, various aspects of the invention relate to the treatment of disease, particularly for the selective targeting of diseased tissue, and to compounds, combinations, agents, formulations, kits, etc. that can be used in such methods. In all aspects, the diseased tissue may reside at a single site in the body (e.g., in the case of a localized solid tumor) or may reside at multiple sites (e.g., when several joints are affected in arthritis or in the case of dispersed or metastatic cancerous diseases).

本發明之抗體及/或靶向錒結合物(TAC)較佳地具有針對磷脂醯肌醇蛋白聚醣-3 (GPC3)之結合親和力。已在不同類型之疾病中觀察腫瘤細胞中之GPC3表現，尤其過度增殖性疾病(例如癌症) (Nishida T, Kataoka H. Glypican 3-Targeted Therapy in Hepatocellular Carcinoma. Cancers (Basel). 2019 Sep 10;11(9):1339)。適於使用本發明較佳抗體及/或靶向錒結合物(TAC)治療之疾病及/或贅瘤之實例包含肝細胞癌(HCC)、包含鱗狀非小細胞肺癌之肺癌、神經內分泌前列腺癌、默克爾細胞癌(Merkel cell carcinoma)、兒科適應症(例如兒科實體胚胎性腫瘤(包含大部分肝母細胞瘤)、維爾姆斯腫瘤(Wilms tumor)、橫紋肌樣腫瘤、某些生殖細胞腫瘤亞型(如卵黃囊+絨毛膜癌)及一些橫紋肌肉瘤)。黑色素瘤、卵巢癌、結腸直腸癌、食道癌、子宮頸癌、頭頸癌、腎細胞癌、乳癌(包含三陰性乳癌)、胃癌、肉瘤、膀胱癌。The antibodies and/or targeted conjugates (TACs) of the present invention preferably have binding affinity for glypican-3 (GPC3). GPC3 expression in tumor cells has been observed in different types of diseases, especially hyperproliferative diseases (e.g., cancer) (Nishida T, Kataoka H. Glypican 3-Targeted Therapy in Hepatocellular Carcinoma. Cancers (Basel). 2019 Sep 10;11(9):1339). Examples of diseases and/or tumors suitable for treatment using the preferred antibodies and/or targeted titanium conjugates (TACs) of the present invention include hepatocellular carcinoma (HCC), lung cancer including squamous non-small cell lung cancer, neuroendocrine prostate cancer, Merkel cell carcinoma, pediatric indications (e.g., pediatric solid embryonal tumors (including most hepatoblastomas), Wilms tumor, rhabdoid tumor, certain germ cell tumor subtypes (such as yolk sac + choriocarcinoma) and some rhabdoid myomas). Melanoma, ovarian cancer, colorectal cancer, esophageal cancer, cervical cancer, head and neck cancer, renal cell carcinoma, breast cancer (including triple-negative breast cancer), gastric cancer, sarcoma, bladder cancer.

在一些實施例中，本發明包含使用本發明之抗體及/或靶向錒結合物(TAC)以治療疾病之方法，更佳地治療過度增殖性疾病，更具體而言其中過度增殖性疾病係癌症且甚至更具體而言其中該癌症疾病係肝細胞癌(HCC)。在一些實施例中，本發明包含使用本發明之抗體及/或靶向錒結合物(TAC)以治療疾病、較佳地過度增殖性疾病、更佳地癌症之方法，該等疾病之特徵在於腫瘤細胞中之GPC3之異位表現。In some embodiments, the invention comprises methods of using the antibodies and/or targeted titanate conjugates (TACs) of the invention to treat a disease, more preferably a hyperproliferative disease, more specifically wherein the hyperproliferative disease is cancer and even more specifically wherein the cancer disease is hepatocellular carcinoma (HCC). In some embodiments, the invention comprises methods of using the antibodies and/or targeted titanate conjugates (TACs) of the invention to treat a disease, preferably a hyperproliferative disease, more preferably a cancer, characterized by ectopic expression of GPC3 in tumor cells.

在一些實施例中，本發明包含治療過度增殖性疾病、更具體而言癌症、更具體而言肝細胞癌之方法，該方法包括向有需要之個體投與有效量之至少一種本發明之抗體及/或靶向錒結合物(TAC)。在一些實施例中，本發明包含治療特徵在於腫瘤細胞中之GPC3之異位表現之過度增殖性疾病的方法、更具體而言其中過度增殖性疾病係癌症、更具體而言肝細胞癌，該方法包括向有需要之個體投與有效量之至少一種本發明之抗體及/或靶向錒結合物(TAC)及其他醫藥劑。In some embodiments, the present invention comprises a method of treating a hyperproliferative disease, more specifically cancer, more specifically hepatocellular carcinoma, comprising administering to a subject in need thereof an effective amount of at least one antibody and/or targeted titanate conjugate (TAC) of the present invention. In some embodiments, the present invention comprises a method of treating a hyperproliferative disease characterized by ectopic expression of GPC3 in tumor cells, more specifically wherein the hyperproliferative disease is cancer, more specifically hepatocellular carcinoma, comprising administering to a subject in need thereof an effective amount of at least one antibody and/or targeted titanate conjugate (TAC) of the present invention and other pharmaceutical agents.

在一些實施例中，本發明提供如上文所闡述之抗體及/或靶向錒結合物(TAC)，其用於治療及/或預防疾病、具體而言過度增殖性病症、更具體而言癌症，甚至更具體而言其中該癌症疾病係肝細胞癌。在一些實施例中，本發明包含如上文所闡述之抗體及/或靶向錒結合物(TAC)，其用於治療及/或預防特徵在於腫瘤細胞中之GPC3之異位表現之過度增殖性疾病。In some embodiments, the present invention provides antibodies and/or targeted titanate conjugates (TACs) as described above for use in treating and/or preventing a disease, in particular a hyperproliferative disorder, more particularly a cancer, even more particularly wherein the cancer disease is hepatocellular carcinoma. In some embodiments, the present invention comprises antibodies and/or targeted titanate conjugates (TACs) as described above for use in treating and/or preventing a hyperproliferative disease characterized by ectopic expression of GPC3 in tumor cells.

在一些實施例中，本發明包含本發明之抗體及/或靶向錒結合物(TAC)，以用於抑制細胞增殖及/或誘導細胞之細胞凋亡之方法，該方法包括使細胞與抗體及/或靶向錒結合物(TAC)接觸。在一些實施例中，本發明包含本發明之抗體及/或靶向錒結合物(TAC)，以用於抑制腫瘤細胞增殖及/或誘導腫瘤細胞之細胞凋亡之方法，該方法包括使腫瘤細胞與抗體及/或靶向錒結合物(TAC)接觸，其中GPC3在腫瘤細胞中過表現。In some embodiments, the present invention comprises an antibody and/or targeted titanate conjugate (TAC) of the present invention for use in a method of inhibiting cell proliferation and/or inducing apoptosis of a cell, the method comprising contacting a cell with the antibody and/or targeted titanate conjugate (TAC). In some embodiments, the present invention comprises an antibody and/or targeted titanate conjugate (TAC) of the present invention for use in a method of inhibiting tumor cell proliferation and/or inducing apoptosis of a tumor cell, the method comprising contacting a tumor cell with the antibody and/or targeted titanate conjugate (TAC), wherein GPC3 is overexpressed in the tumor cell.

在一些實施例中，本發明包含抗體及/或靶向錒結合物(TAC)，以用於治療過度增殖性疾病之方法，更具體而言其中過度增殖性疾病係癌症且甚至更具體而言其中該癌症疾病係肝細胞癌。在一些實施例中，本發明包含抗體及/或靶向錒結合物(TAC)，以用於治療過度增殖性疾病之方法，該等疾病之特徵在於腫瘤細胞中之GPC3之異位表現。In some embodiments, the invention comprises antibodies and/or targeted titanate conjugates (TACs) for use in methods of treating hyperproliferative diseases, more specifically wherein the hyperproliferative disease is cancer and even more specifically wherein the cancer disease is hepatocellular carcinoma. In some embodiments, the invention comprises antibodies and/or targeted titanate conjugates (TACs) for use in methods of treating hyperproliferative diseases characterized by ectopic expression of GPC3 in tumor cells.

在一些實施例中，本發明包含抗體及/或靶向錒結合物(TAC)之用途，其用於治療及/或預防過度增殖性疾病。在一些實施例中，本發明包含抗體及/或靶向錒結合物(TAC)之用途，其用於製造用於治療及/或預防過度增殖性疾病之藥劑，該等疾病之特徵在於腫瘤細胞中之GPC3之異位表現。In some embodiments, the present invention comprises the use of antibodies and/or targeted titanate conjugates (TACs) for the treatment and/or prevention of hyperproliferative diseases. In some embodiments, the present invention comprises the use of antibodies and/or targeted titanate conjugates (TACs) for the manufacture of a medicament for the treatment and/or prevention of hyperproliferative diseases characterized by ectopic expression of GPC3 in tumor cells.

在一些實施例中，本發明包含抗體及/或靶向錒結合物(TAC)之用途，其用於製造用於治療過度增殖性疾病、尤其癌症及更具體而言肝細胞癌之藥劑。In some embodiments, the present invention comprises the use of antibodies and/or targeted tantalum conjugates (TACs) for the manufacture of a medicament for the treatment of a hyperproliferative disease, particularly cancer, and more specifically hepatocellular carcinoma.

上文所提及之病症已在人類中經充分描述，而且亦以類似病因存在於其他動物中(包含哺乳動物)，且其可藉由投與本發明醫藥組合物來治療。The above-mentioned disorders have been well described in humans and also exist with similar etiologies in other animals, including mammals, and can be treated by administering the pharmaceutical compositions of the present invention.

本發明之抗體或其抗原結合片段或本發明靶向錒結合物(TAC)或其變體可與已知藥劑共投與，且在一些情況下抗體或其抗原結合片段本身可經修飾。舉例而言，可將抗體或其抗原結合片段或其變體或TAC結合至藥物或另一肽或蛋白質以潛在地進一步增加效能。The antibodies or antigen-binding fragments thereof of the present invention or the targeted tantalum conjugates (TACs) or variants thereof of the present invention can be co-administered with known agents, and in some cases the antibodies or antigen-binding fragments thereof themselves can be modified. For example, the antibodies or antigen-binding fragments thereof or variants thereof or TACs can be conjugated to a drug or another peptide or protein to potentially further increase efficacy.

可將本發明之抗體或其抗原結合片段或本發明靶向錒結合物(TAC)作為唯一醫藥藥劑或與一或多種其他治療劑組合投與，其中該組合不引起不可接受之不良效應。The antibodies or antigen-binding fragments thereof of the invention or the targeted tantalum conjugates (TACs) of the invention may be administered as the sole pharmaceutical agent or in combination with one or more other therapeutic agents, where the combination does not cause unacceptable adverse effects.

因此，在另一態樣中，本發明係關於本發明之抗體或其抗原結合片段或本發明靶向錒結合物(TAC)或本發明醫藥組合物，以與一或多種具有生物活性之其他治療活性化合物同時、單獨或依序組合使用以較佳地針對癌症、尤其肝細胞癌症。Therefore, in another aspect, the present invention relates to the antibody or antigen-binding fragment thereof of the present invention or the targeted tantalum conjugate (TAC) of the present invention or the pharmaceutical composition of the present invention, for use simultaneously, separately or sequentially in combination with one or more other therapeutically active compounds with biological activity to preferably target cancer, especially hepatocellular carcinoma.

下文列示欲與本發明之抗體或其抗原結合片段或本發明靶向錒結合物(TAC)組合使用之治療活性化合物之非限制性實例。該等化合物由其種類定義，由此容許熟習此項技術者鑑別屬該種類之該等化合物。Listed below are non-limiting examples of therapeutically active compounds to be used in combination with the antibodies or antigen-binding fragments thereof of the present invention or the targeted tantalum conjugates (TACs) of the present invention. The compounds are defined by their class, thereby allowing those skilled in the art to identify the compounds belonging to that class.

在本發明之上下文中，TK抑制劑係指為酪胺酸激酶之藥理學抑制劑之化合物。酪胺酸激酶係負責藉由信號轉導級聯來活化許多蛋白質之受體。其實例包含(但不限於)索拉菲尼(sorafenib)、來瓦替尼(Lenvatinib)、瑞格非尼(regorafenib)或卡博替尼(cabozantinib)。In the context of the present invention, a TK inhibitor refers to a compound that is a pharmacological inhibitor of tyrosine kinase. Tyrosine kinase is a receptor responsible for activating many proteins through signal transduction cascades. Examples include (but are not limited to) sorafenib, lenvatinib, regorafenib or cabozantinib.

在本發明之上下文中，VEGFR抑制劑係指為血管內皮生長因子受體(VEGFR)信號傳導之藥理學抑制劑之化合物，該信號傳導在腫瘤誘導之血管生成之調控中發揮作用。其實例包含(但不限於)貝伐單抗(bevacizumab)及雷莫蘆單抗(ramucirumab)。In the context of the present invention, VEGFR inhibitors refer to compounds that are pharmacological inhibitors of vascular endothelial growth factor receptor (VEGFR) signaling, which plays a role in the regulation of tumor-induced angiogenesis. Examples include, but are not limited to, bevacizumab and ramucirumab.

在本發明之上下文中，化學治療劑係指適於GPC3陽性癌症、尤其HCC之化學療法之化合物。其實例包含(但不限於)多柔比星(doxorubicin)、表柔比星(epirubicin)或順鉑(cisplatin)。In the context of the present invention, chemotherapeutic agents refer to compounds suitable for chemotherapy of GPC3-positive cancers, especially HCC. Examples include (but are not limited to) doxorubicin, epirubicin or cisplatin.

可在本發明之背景下與本發明之抗體或其抗原結合片段或本發明靶向錒結合物(TAC)組合使用之早期產品線藥物的實例包含(但不限於)替沃尼布(tivozanib) (AVEO Oncology)、納莫德諾森(namodenoson) (Can-Fite BioPharma)、GT90001 (Kintor Pharmaceutical)、ARTEMIS T-細胞療法(Eureka Therapeutics)、MTL-CEBPA (MiNA Therapeutics)、福斯特羅西他濱-布拉帕胺(fostroxacitabine bralpamide) /福斯特羅(fostrox) (Medivir)或各種TIGIT抑制劑。Examples of early pipeline drugs that can be used in combination with the antibodies or antigen-binding fragments thereof or the targeted tantalum conjugates (TACs) of the present invention in the context of the present invention include, but are not limited to, tivozanib (AVEO Oncology), namodenoson (Can-Fite BioPharma), GT90001 (Kintor Pharmaceutical), ARTEMIS T-cell therapy (Eureka Therapeutics), MTL-CEBPA (MiNA Therapeutics), fostroxacitabine bralpamide/fostrox (Medivir), or various TIGIT inhibitors.

在本發明之上下文中，經動脈化療栓塞藥劑係指適於動脈內施加至將血液特異性供應至肝內HCC病灶之動脈中之化合物。其實例包含(但不限於)多柔比星、表柔比星或順鉑、多柔比星釋放藥物洗脫珠粒、碘油、釔-90 (β-發射體)-標記玻璃或樹脂微球體或如明膠海綿[Gelfoam]或可降解澱粉微球體之栓塞藥劑。In the context of the present invention, transarterial chemoembolic agents refer to compounds suitable for intra-arterial administration to the arteries that specifically supply blood to intrahepatic HCC lesions. Examples include, but are not limited to, doxorubicin, epirubicin or cisplatin, doxorubicin-releasing drug-eluting beads, iodized oil, yttrium-90 (β-emitter)-labeled glass or resin microspheres, or embolic agents such as gelatin sponge [Gelfoam] or degradable starch microspheres.

在本發明之上下文中，外部光束放射療法係指輻射療法，其中外部來源之電離輻射被指向及/或施加至個體之身體之特定部分。In the context of the present invention, external beam radiation therapy refers to radiation therapy in which ionizing radiation from an external source is directed at and/or applied to a specific part of an individual's body.

在本發明之上下文中，術語「組合」不僅意指含有所有組分(亦即，本發明之抗GPC3抗體或其片段及/或抗GPC3-TAC)及其他醫藥藥劑(亦稱為固定劑量組合)之劑型以及含有彼此分離之組分之組合包裝，而且意指同時或依序投與之組分，只要其用於預防或治療相同疾病。In the context of the present invention, the term "combination" means not only a dosage form containing all components (i.e., the anti-GPC3 antibody or fragment thereof and/or anti-GPC3-TAC of the present invention) and other pharmaceutical agents (also called a fixed-dose combination) and a combination package containing components separated from each other, but also means components administered simultaneously or sequentially as long as they are used for the prevention or treatment of the same disease.

根據以下之該等考量投與活性成分之量可大大變化：採用之特定化合物及劑量單元、投與之模式及時間、治療之時段、治療之患者之年齡、性別及一般條件、治療之病況之性質及嚴重程度、藥物代謝及排泄之速率、潛在藥物組合及藥物-藥物相互作用及諸如此類。The amount of active ingredient administered can vary widely depending on such considerations as the particular compound and dosage unit employed, the mode and timing of administration, the duration of treatment, the age, sex and general condition of the patient being treated, the nature and severity of the condition being treated, the rate of drug metabolism and excretion, potential drug combinations and drug-drug interactions, and the like.

醫藥組合物及投與在另一態樣中，本發明係關於包括本發明之經分離抗體或抗原結合片段或本發明靶向錒結合物之醫藥組合物。為治療前述病症中之任一者，可使用一或多種生理上可接受之載劑、賦形劑或輔助劑以任何習用方式來調配用於本發明之醫藥組合物。關於調配及投與之技術之其他細節可參見Remington's Pharmaceutical Sciences (編輯Maack Publishing Co，Easton, Pa.)之最新版本。Pharmaceutical compositions and administration In another aspect, the present invention relates to pharmaceutical compositions comprising an isolated antibody or antigen-binding fragment of the present invention or a targeted tantalum conjugate of the present invention. To treat any of the aforementioned conditions, the pharmaceutical compositions for use in the present invention may be formulated in any conventional manner using one or more physiologically acceptable carriers, excipients or adjuvants. Further details on the techniques of formulation and administration may be found in the latest edition of Remington's Pharmaceutical Sciences (ed. Maack Publishing Co, Easton, Pa.).

可藉由任何適宜方式來投與本發明之經分離抗體或抗原結合片段或本發明靶向錒結合物，該等方式端視正治療之病症類型可變化。可能投與途徑包含口服、非經腸及局部投與。非經腸遞送之方法包含動脈內、肌內、皮下、髓內、鞘內、室內、靜脈內、腹膜腔內或鼻內投與。另外，可(使用(例如)遞減劑量)藉由脈衝輸注投與本發明之經分離抗體或抗原結合片段或本發明靶向錒結合物。較佳地，藉由注射、最佳地藉由靜脈內注射投與。欲投與之量取決於各種因素，例如臨床症狀、個體之體重、是否投與其他藥物及諸如此類。熟習此項技術者將認識到，投與途徑端視欲治療之病症或病況可變化。The isolated antibodies or antigen-binding fragments of the present invention or the targeted tantalum conjugates of the present invention may be administered by any suitable means, which may vary depending on the type of condition being treated. Possible routes of administration include oral, parenteral, and topical administration. Methods of parenteral delivery include intra-arterial, intramuscular, subcutaneous, intramedullary, intrathecal, intraventricular, intravenous, intraperitoneal, or intranasal administration. In addition, the isolated antibodies or antigen-binding fragments of the present invention or the targeted tantalum conjugates of the present invention may be administered by pulse infusion (using, for example, decreasing doses). Preferably, administration is by injection, most preferably by intravenous injection. The amount to be administered depends on various factors, such as the clinical symptoms, the weight of the individual, whether other medications are being administered, and the like. Those skilled in the art will recognize that the route of administration may vary depending on the disorder or condition to be treated.

本發明醫藥組合物包括單獨或與至少一種其他藥劑(例如穩定化合物)組合之本發明之抗體或抗原結合片段或本發明靶向錒結合物。可在任何無菌、生物相容醫藥載劑中投與本發明之抗體或抗原結合片段或本發明靶向錒結合物，該等載劑包含(但不限於)鹽水、緩衝鹽水、右旋糖及水。在特定實施例中，本發明醫藥組合物可包括一或多種其他醫藥活性化合物，尤其適於治療與因部分或完全血管堵塞導致之缺血性事件有關之GPC3相關病症及/或病症之一或多種其他醫藥活性化合物。在該等藥劑與賦形劑或醫藥上可接受之載劑混合之醫藥組合物中，可將該等藥劑中之任一者單獨或與其他藥劑或藥物組合投與患者。在特定實施例中，醫藥上可接受之載劑係醫藥上惰性的。The pharmaceutical composition of the present invention includes the antibody or antigen-binding fragment of the present invention or the targeted tantalum conjugate of the present invention alone or in combination with at least one other agent (e.g., a stabilizing compound). The antibody or antigen-binding fragment of the present invention or the targeted tantalum conjugate of the present invention can be administered in any sterile, biocompatible pharmaceutical carrier, including, but not limited to, saline, buffered saline, dextrose, and water. In a specific embodiment, the pharmaceutical composition of the present invention may include one or more other pharmaceutically active compounds, particularly suitable for treating GPC3-related disorders and/or disorders associated with ischemic events caused by partial or complete vascular occlusion. In pharmaceutical compositions in which the agents are mixed with excipients or pharmaceutically acceptable carriers, any of the agents may be administered to a patient alone or in combination with other agents or drugs. In specific embodiments, the pharmaceutically acceptable carrier is pharmaceutically inert.

可使用業內已熟知之醫藥上可接受之載劑以適於口服投與之劑量來調配用於口服投與之醫藥組合物。該等載劑使得能夠將醫藥組合物調配成錠劑、丸劑、糖衣藥丸、膠囊、液體、凝膠、糖漿、漿液、懸浮液及諸如此類以用於由患者攝入。Pharmaceutical compositions for oral administration can be formulated using pharmaceutically acceptable carriers well known in the art in dosages suitable for oral administration. Such carriers enable the pharmaceutical compositions to be formulated into tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like for ingestion by patients.

用於口服使用之醫藥製劑可經由以下方式來獲得：組合活性化合物與固體賦形劑，視情況研磨所得混合物，並在添加適宜輔助劑(若需要)後處理顆粒混合物，以獲得錠劑或糖衣錠核心。適宜賦形劑為碳水化合物或蛋白質填充劑，例如糖(包含乳糖、蔗糖、甘露醇或山梨醇)；來自玉米、小麥、水稻、馬鈴薯或其他植物之澱粉；纖維素，例如甲基纖維素、羥丙基甲基纖維素或鈉羧甲基纖維素；及膠，包含阿拉伯膠(arabic)及黃蓍膠；及蛋白質，例如明膠及膠原。若期望，可添加崩解劑或增溶劑，例如交聯聚乙烯基吡咯啶酮、瓊脂、海藻酸或其鹽(例如海藻酸鈉)。Pharmaceutical preparations for oral use can be obtained by combining the active compound with a solid excipient, optionally grinding the resulting mixture, and processing the granulated mixture after adding suitable adjuvants (if necessary) to obtain tablets or dragee cores. Suitable excipients are carbohydrate or protein fillers, such as sugars (including lactose, sucrose, mannitol or sorbitol); starch from corn, wheat, rice, potato or other plants; cellulose, such as methylcellulose, hydroxypropylmethylcellulose or sodium carboxymethylcellulose; and gums, including arabic and tragacanth; and proteins, such as gelatin and collagen. If desired, disintegrating or solubilizing agents may be added, for example cross-linked polyvinyl pyrrolidone, agar, alginic acid or a salt thereof (eg sodium alginate).

可提供具有適宜包衣(例如濃縮糖溶液)之糖衣錠核心，其亦可含有阿拉伯膠、滑石粉、聚乙烯基吡咯啶酮、卡波普(carbopol)凝膠、聚乙二醇及/或二氧化鈦、漆溶液及適宜有機溶劑或溶劑混合物。可將染料或顏料添加至錠劑或糖衣錠包衣中用於產品鑒定或表徵活性化合物之量。Dragee cores may be provided with suitable coatings, such as concentrated sugar solutions, which may also contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol and/or titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures. Dyes or pigments may be added to tablets or dragee coatings for product identification or to characterize the amount of active compound.

可口服使用之醫藥製劑包含由明膠製得之推入配合式(push-fit)膠囊，以及由明膠及包衣(例如甘油或山梨醇)製得之軟密封膠囊。推入配合式膠囊可含有與填充劑或黏合劑(例如乳糖或澱粉)、潤滑劑(例如滑石粉或硬脂酸鎂)及視情況穩定劑混合之活性成分。在軟質膠囊中，可將活性化合物溶解或懸浮於適宜液體(例如脂肪油、液體石蠟或具有或不具有穩定劑之液體聚乙二醇)中。Pharmaceutical preparations that can be used orally include push-fit capsules made of gelatin, and soft-sealed capsules made of gelatin and a coating such as glycerol or sorbitol. Push-fit capsules may contain the active ingredient in admixture with fillers or binders such as lactose or starches, lubricants such as talc or magnesium stearate, and optionally stabilizers. In soft capsules, the active compound may be dissolved or suspended in a suitable liquid such as fatty oils, liquid paraffin, or liquid polyethylene glycol with or without stabilizers.

醫藥非經腸投與之醫藥調配物包含活性化合物之水溶液。對於注射而言，可將本發明之醫藥組合物調配於水溶液、較佳生理上相容之緩衝液(例如漢克氏溶液(Hank's solution)、林格氏溶液(Ringer's solution)或生理緩衝鹽水)中。水性注射懸浮液可含有增加該懸浮液黏度之物質，例如羧甲基纖維素鈉、山梨醇或右旋糖酐。另外，該等活性化合物之懸浮液可製備成適當油性注射懸浮液。適宜親脂性溶劑或媒劑包括脂肪油(例如芝麻油)或合成脂肪酸酯(例如油酸乙酯或三酸甘油酯)或脂質體。視情況，該懸浮液亦可含有適宜穩定劑或增加該等化合物之溶解度之試劑以容許製備高濃度溶液。Pharmaceutical formulations for parenteral administration include aqueous solutions of the active compounds. For injection, the pharmaceutical compositions of the present invention may be formulated in aqueous solution, preferably in a physiologically compatible buffer (e.g., Hank's solution, Ringer's solution, or physiologically buffered saline). Aqueous injection suspensions may contain substances that increase the viscosity of the suspension, such as sodium carboxymethylcellulose, sorbitol, or dextran. In addition, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils (e.g., sesame oil) or synthetic fatty acid esters (e.g., ethyl oleate or triglycerides) or liposomes. Optionally, the suspension may also contain suitable stabilizers or agents that increase the solubility of the compounds to allow the preparation of highly concentrated solutions.

針對局部或經鼻投與，在調配物中使用適於透過特定障壁之滲透劑。該等滲透劑為業內眾所周知。For topical or nasal administration, penetrants suitable for permeating the particular barrier are used in the formulation. Such penetrants are well known in the art.

本發明之醫藥組合物可以業內已知之方式製造，例如藉助習用混合、溶解、造粒、製糖衣錠、粉碎、乳化、囊封、包埋或凍乾製程。The pharmaceutical composition of the present invention can be manufactured in a manner known in the art, for example, by means of conventional mixing, dissolving, granulating, dragee-making, pulverizing, emulsifying, encapsulating, embedding or lyophilizing processes.

本發明之醫藥組合物可以鹽形式提供且可使用許多酸來形成，該等酸包括但不限於鹽酸、硫酸、乙酸、乳酸、酒石酸、蘋果酸、琥珀酸等。鹽往往更可溶於水性或相應游離鹼形式之其他質子溶劑中。在其他情形下，較佳製劑可為液體調配物或視情況包括其他物質(如聚山梨醇酯)之在組胺酸或磷酸鹽或Tris緩衝液、蔗糖及/或甘露醇中在4.5至7.5之pH範圍內之其凍乾粉末，該凍乾粉末在使用之前與緩衝液組合。The pharmaceutical compositions of the present invention may be provided in salt form and may be formed using a variety of acids, including but not limited to hydrochloric acid, sulfuric acid, acetic acid, lactic acid, tartaric acid, apple acid, succinic acid, etc. Salts are often more soluble in aqueous or other protic solvents in the form of the corresponding free base. In other cases, the preferred formulation may be a liquid formulation or a lyophilized powder thereof in histidine or phosphate or Tris buffer, sucrose and/or mannitol at a pH range of 4.5 to 7.5, optionally including other substances (such as polysorbates), which is combined with the buffer prior to use.

在已製備包括調配於可接受之載劑中之本發明化合物之醫藥組合物之後，可將其置於適當容器中且標記用於治療指示之病況。針對抗GPC3抗體或其抗原結合片段或抗GPC3-TAC之投與，該標記包含量、頻率及投與方法。After a pharmaceutical composition comprising a compound of the invention formulated in an acceptable carrier has been prepared, it can be placed in an appropriate container and labeled for treatment of an indicated condition. For administration of an anti-GPC3 antibody or antigen-binding fragment thereof or anti-GPC3-TAC, the label includes the amount, frequency, and method of administration.

治療有效劑量適用於本發明之醫藥組合物包含含有有效量之活性成分之組合物以達成預期目的，例如，治療特徵在於GPC3之異位表現之特定疾病狀態。Therapeutically Effective Amounts Pharmaceutical compositions suitable for use in the present invention include compositions containing an effective amount of the active ingredients to achieve the intended purpose, for example, to treat a particular disease state characterized by heterotopic expression of GPC3.

有效劑量之確定一定在熟習此項技術者之能力範圍內。本發明新穎抗體或其抗原結合片段或其變體之治療有效量之測定主要取決於特定患者特性、投與途徑及正治療病症之性質。通用導則可參見(例如) International Conference on Harmonization之出版物及REMINGTON'S PHARMACEUTICAL SCIENCES，第27及28章，484-528頁(第18版，Alfonso R. Gennaro, Ed., Easton, Pa.: Mack Pub. Co., 1990)。更具體而言，確定治療有效量取決於諸如藥劑之毒性及效能等因素。可使用業內熟知之及前述參考文獻中發現之方法來測定毒性。The determination of the effective dose is certainly within the capabilities of those skilled in the art. The determination of the therapeutically effective dose of the novel antibodies or antigen-binding fragments thereof or variants thereof of the present invention depends primarily on the characteristics of the particular patient, the route of administration, and the nature of the condition being treated. General guidelines can be found, for example, in publications of the International Conference on Harmonization and REMINGTON'S PHARMACEUTICAL SCIENCES, Chapters 27 and 28, pp. 484-528 (18th ed., Alfonso R. Gennaro, Ed., Easton, Pa.: Mack Pub. Co., 1990). More specifically, the determination of the therapeutically effective dose depends on factors such as the toxicity and efficacy of the agent. Toxicity can be determined using methods well known in the art and found in the aforementioned references.

針對任何化合物，可最初在細胞培養分析中或在動物模型(通常為小鼠、兔、狗、豬或猴)中估計治療有效劑量。亦使用動物模型來達成期望投與濃度範圍及途徑。然後可使用該等資訊來確定人類中之可用投與劑量及途徑。For any compound, the therapeutically effective dose can be estimated initially in cell culture assays or in animal models (usually mice, rabbits, dogs, pigs, or monkeys). Animal models are also used to arrive at the desired concentration range and route of administration. This information can then be used to determine useful doses and routes of administration in humans.

治療有效劑量係指可改善症狀或病況之抗體或其抗原結合片段之量或抗體結合物之量。可藉由標準醫藥程序在細胞培養物或實驗動物中確定該等化合物之治療效能及毒性，例如ED₅₀(於50%群體中治療有效之劑量)及LD₅₀(致死50%群體之劑量)。治療效應與毒性效應之間之劑量比率係治療指數，且其可表示為比率ED₅₀/LD₅₀。以治療指數大的之化合物較佳。可使用自細胞培養分析及動物研究獲得之數據來調配供人類使用之劑量範圍。該等化合物之劑量較佳在具有較少毒性或沒有毒性之循環濃度(包括ED₅₀)範圍內。該劑量端視所用劑型、患者之敏感性、投與途徑而定，可在此範圍內變化。A therapeutically effective dose refers to the amount of antibody or antigen-binding fragment thereof or the amount of antibody conjugate that improves the symptom or condition. The therapeutic efficacy and toxicity of the compounds can be determined by standard pharmaceutical procedures in cell culture or experimental animals, such as_ED50 (the dose that is therapeutically effective in 50% of the population) and_LD50 (the dose that is lethal to 50% of the population). The dose ratio between the therapeutic effect and the toxic effect is the therapeutic index, and it can be expressed as the ratio_ED50 /_LD50 . Compounds with large therapeutic indices are preferred. Data obtained from cell culture assays and animal studies can be used to formulate a dosage range for human use. Dosages of the compounds lie preferably within a range of circulating concentrations (including the_ED50 ) with little or no toxicity. The dosage may vary within this range depending on the dosage form employed, sensitivity of the patient, and the route of administration.

確切劑量係由個別醫師根據所治療患者來選擇。可調整劑量及投與以提供足量活性部分體或維持期望效應。可考慮之其他因素包括疾病狀態之嚴重程度；患者之年齡、體重及性別；膳食、投與之時間及頻率；藥物組合；反應敏感性；及對療法之耐受性/反應。長效醫藥組合物端視特定調配物之半衰期及清除率，可(例如)每3至4天、每週一次、每兩週一次或每三週一次投與。The exact dosage is chosen by the individual physician based on the patient being treated. Dosage and administration may be adjusted to provide adequate amounts of the active moiety or to maintain the desired effect. Other factors that may be considered include the severity of the disease state; age, weight, and sex of the patient; diet, time and frequency of administration; drug combination; reaction sensitivities; and tolerance/response to therapy. Long-acting pharmaceutical compositions may be administered, for example, every 3 to 4 days, once a week, once every two weeks, or once every three weeks, depending on the half-life and clearance rate of the particular formulation.

端視投與途徑，正常劑量量可在0.1至100,000微克之間變化，總劑量最多為約10 g。文獻中提供遞送之特定劑量及方法之指導。參見美國專利第4,657,760；5,206,344或5,225,212號。Depending on the route of administration, normal dosage amounts may vary from 0.1 to 100,000 micrograms, with a total dose of up to about 10 g. Guidance on specific dosages and methods of delivery is provided in the literature. See U.S. Patent Nos. 4,657,760; 5,206,344 or 5,225,212.

套組在另一態樣中，本發明係關於包括本發明之經分離抗體或抗原結合片段或本發明靶向錒結合物及使用說明書之套組。在特定實施例中，該套組包括填充有上文所提及本發明組合物之一或多種成分之一或多個容器。該(等)容器可附帶有監管醫藥物或生物產品之製造、使用或銷售之政府機構所規定形式之公告，該公告顯示政府機構已批准用於投與人類之製造、使用或銷售。Kits In another aspect, the present invention relates to a kit comprising an isolated antibody or antigen-binding fragment of the present invention or a targeted tantalum conjugate of the present invention and instructions for use. In a particular embodiment, the kit comprises one or more containers filled with one or more components of the composition of the present invention mentioned above. The container(s) may be accompanied by a notice in a form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, indicating that the governmental agency has approved the manufacture, use or sale for administration to humans.

診斷方法根據另一態樣，本發明之抗體或抗原結合片段或本發明抗體結合物可以原樣或在組合物中用於研究及診斷，或作為分析參考標準及諸如此類。Diagnostic Methods According to another aspect, the antibodies or antigen-binding fragments of the present invention or antibody conjugates of the present invention can be used as such or in a composition for research and diagnosis, or as analytical reference standards and the like.

抗GPC3抗體或其抗原結合片段或本發明抗體結合物可用於檢測GPC3之存在。因此，在另一態樣中，本發明係關於本發明之經分離抗體或抗原結合片段或本發明抗體結合物，以用作診斷劑。Anti-GPC3 antibodies or antigen-binding fragments thereof or antibody conjugates of the invention can be used to detect the presence of GPC3. Therefore, in another aspect, the invention relates to an isolated antibody or antigen-binding fragment or antibody conjugate of the invention for use as a diagnostic agent.

GPC3之表現之檢測通常涉及使生物試樣(個體之腫瘤、細胞、組織或體液)與一或多種本發明抗體或片段接觸(視情況結合至可檢測部分)，且檢測試樣針對GPC3是否為陽性或與對照試樣相比試樣是否改變(例如，減少或增加)表現。Detection of GPC3 expression typically involves contacting a biological sample (a tumor, cell, tissue or body fluid of an individual) with one or more antibodies or fragments of the invention (optionally bound to a detectable moiety), and testing whether the sample is positive for GPC3 or whether the sample has altered (e.g., decreased or increased) expression compared to a control sample.

在某些實施例中，提供之結合物包括一或多種可檢測部分，亦即經一或多種該等部分「標記」。在一些此類實施例中，本發明結合物可用於診斷或成像應用，例如，診斷癌症或使癌症成像。眾多種可檢測部分中之任一者可用於標記本文所闡述之抗體結合物。適宜可檢測部分包括(但不限於)：各種配體、放射性核素；螢光染料(例如螢光共振能量染料(FRET)，如Cy5、Cy7)；化學發光劑(例如，舉例而言，吖啶酯(acridinum ester)、穩定二氧雜環丁烷及諸如此類)；生物發光藥劑；可光譜解析之無機螢光半導體奈米晶體(亦即，量子點)；微顆粒；金屬奈米顆粒(例如，金、銀、銅、鉑等)；奈米簇；順磁性金屬離子；酶；比色標記(例如，舉例而言，染料、膠質金及諸如此類)；生物素；地高辛(dioxigenin)；半抗原；及抗血清或單株抗體可用之蛋白質。In certain embodiments, provided conjugates include one or more detectable moieties, i.e., are "labeled" with one or more of such moieties. In some such embodiments, the conjugates of the invention may be used in diagnostic or imaging applications, e.g., diagnosing or imaging cancer. Any of a variety of detectable moieties may be used to label the antibody conjugates described herein. Suitable detectable moieties include, but are not limited to: various ligands, radionuclides; fluorescent dyes (e.g., fluorescence resonance energy dyes (FRET), such as Cy5, Cy7); chemiluminescent agents (e.g., for example, acridinium esters); esters), stable dioxetanes, and the like); bioluminescent agents; spectrally resolvable inorganic fluorescent semiconductor nanocrystals (i.e., quantum dots); microparticles; metal nanoparticles (e.g., gold, silver, copper, platinum, etc.); nanoclusters; paramagnetic metal ions; enzymes; colorimetric labels (e.g., dyes, colloidal gold, and the like); biotin; dioxigenin; haptens; and proteins for which antisera or monoclonal antibodies can be used.

根據本發明之某些較佳實施例，可檢測部分可為螢光染料(例如Alexa Flour染料(Invitrogen))或螢光共振能量染料(FRET) (例如可用於經由離胺酸殘基直接標記抗體或抗原結合片段之pH敏感染料CypHer5E-NHS-酯(Cytiva))。According to certain preferred embodiments of the present invention, the detectable moiety can be a fluorescent dye (e.g., Alexa Flour dye (Invitrogen)) or a fluorescence resonance energy dye (FRET) (e.g., a pH-sensitive dye CypHer5E-NHS-ester (Cytiva) that can be used to directly label antibodies or antigen-binding fragments via lysine residues).

PET成像根據另一態樣，本發明之抗體或其抗原結合片段及/或結合物可用作成像劑、較佳地PET成像劑。PETImaging According to another aspect, the antibodies or antigen-binding fragments and/or conjugates thereof of the present invention can be used as imaging agents, preferably PET imaging agents.

正電子發射斷層攝影術(PET)係檢測由正電子發射體產生之γ光子且重構該等信號以顯現正電子發射體之位置之醫學成像技術。在與電腦化斷層攝影術(CT)組合時，PET/CT容許精確定位身體之解剖框架內之正電子發射體。臨床中通常使用多重正電子發射體，例如C-11、F-18、Ga-68及Zr-89。較佳地，使用Zr-89作為與本發明之抗體或抗原結合片段或本發明抗體結合物組合之正電子發射體，此乃因其長半衰期(78.4小時)及在溫和條件下之有效標記反應。Positron emission tomography (PET) is a medical imaging technique that detects gamma photons produced by positron emitters and reconstructs these signals to reveal the location of the positron emitter. When combined with computerized tomography (CT), PET/CT allows the precise localization of positron emitters within the anatomical framework of the body. Multiple positron emitters are commonly used in clinical practice, such as C-11, F-18, Ga-68 and Zr-89. Preferably, Zr-89 is used as the positron emitter in combination with the antibodies or antigen-binding fragments of the present invention or the antibody conjugates of the present invention due to its long half-life (78.4 hours) and efficient labeling reaction under mild conditions.

在PET成像應用中針對Zr-89之適宜螯合劑包含(但不限於)：例如DFO (去鐵胺)、N-琥珀醯基-去鐵胺-四氟酚酯(N-suc-DFO-TFP酯)、p-異硫氰酸根合苄基-去鐵胺(DFO-Bz-NCS，亦稱為DFO-Ph-NCS)、去鐵胺-馬來醯亞胺(DFO-馬來醯亞胺)、去鐵胺-方醯胺(DFO-sq)或DFO*。所有該等螯合劑皆可與抗體或抗體片段結合以提供將成像藥劑靶向至欲成像之腫瘤之方式。根據本發明之較佳實施例，螯合劑經由單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。Suitable chelators for Zr-89 in PET imaging applications include, but are not limited to, for example, DFO (deferoxamine), N-succinyl-deferoxamine-tetrafluorophenol ester (N-suc-DFO-TFP ester), p-isothiocyanatobenzyl-deferoxamine (DFO-Bz-NCS, also known as DFO-Ph-NCS), deferoxamine-maleimide (DFO-maleimide), deferoxamine-squaramide (DFO-sq), or DFO*. All of these chelators can be conjugated to antibodies or antibody fragments to provide a means of targeting the imaging agent to the tumor to be imaged. According to a preferred embodiment of the present invention, the chelating agent is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or antigen-binding fragment thereof.

DFO*係DFO之延伸版且與DFO複合物比較⁸⁹Zr-DFO*複合物經展示可展現增加之活體外穩定性及改良之活體內性質(WO2021051168)，且由此在與本發明抗體或抗體片段結合以用於PET成像時，其為針對Zr-89之較佳螯合劑。DFO* is an extended version of DFO and the⁸⁹ Zr-DFO* complex has been shown to exhibit increased in vitro stability and improved in vivo properties compared to the DFO complex (WO2021051168), and is thus a better chelator for Zr-89 when combined with the antibody or antibody fragment of the present invention for PET imaging.

DFO*係指5,11,16,22-四氮雜二十六烷二醯胺, N¹-[5-(乙醯基羥基胺基)戊基]-N²⁶,5,16-三羥基-N²⁶-[5-[[[(4-異硫基氰基苯基)胺基]硫代甲基]胺基]戊基]-4,12,15,23-四側氧基(CAS號1810009-29-0)：且根據本發明之一些實施例可結合至抗體或其抗原結合片段[Ab]、較佳地結合至GPC3之單株抗體或其抗原結合片段。較佳地，抗GPC3抗體選自TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642或其抗原結合片段。更佳地，抗GPC3抗體係選自TPP-29537、TPP-29538、TPP-29636及TPP-29638或其抗原結合片段。DFO* refers to 5,11,16,22-tetraazahexacosandiamide, N¹-[5-(acetylhydroxyamino)pentyl]-N²⁶,5,16-trihydroxy-N²⁶-[5-[[[(4-isothiocyanophenyl)amino]thiomethyl]amino]pentyl]-4,12,15,23-tetraoxy (CAS No. 1810009-29-0): According to some embodiments of the present invention, the antibody or antigen-binding fragment thereof [Ab] can be bound to a monoclonal antibody or antigen-binding fragment thereof that preferably binds to GPC3. Preferably, the anti-GPC3 antibody is selected from TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638 and TPP-29642 or antigen-binding fragment thereof. More preferably, the anti-GPC3 antibody is selected from TPP-29537, TPP-29538, TPP-29636 and TPP-29638 or antigen-binding fragment thereof.

根據本發明之某些極佳實施例，DFO*經由單株抗體之離胺酸殘基偶合至本發明之抗GPC3抗體或其抗原結合片段，由此形成連接螯合部分與抗GPC3抗體或其抗原結合片段之硫脲部分。 熟習此項技術者使用前述資訊可最大程度利用本發明。According to certain preferred embodiments of the present invention, DFO* is coupled to the anti-GPC3 antibody or its antigen-binding fragment of the present invention via the lysine residue of the monoclonal antibody, thereby forming a thiourea portion that connects the chelating portion and the anti-GPC3 antibody or its antigen-binding fragment.Those skilled in the art can use the above information to maximize the use of the present invention.

實驗部分使用來自Dassault Systèmes之BIOVIA Draw 2019生成化學名稱。在一些情形下，使用市售試劑之公認名稱來代替BIOVIA Draw生成之名稱。The chemical names were generatedin the experimental part using BIOVIA Draw 2019 from Dassault Systèmes. In some cases, the recognized names of commercially available reagents were used instead of the names generated by BIOVIA Draw.

下表3列示此段落中及實例部分中所用之縮寫，故在正文本體內不需對其進行闡釋。其他縮寫具有其本身為熟習此項技術者習知之含義。Table 3 below lists the abbreviations used in this paragraph and the examples, so there is no need to explain them in the text. Other abbreviations have their own meanings that are known to those skilled in the art.

表3.縮寫下表列示本文中使用之縮寫。²²⁵Ac            錒-225 Ac-225          錒-225 ACC             抗體螯合劑結合物 CAR             螯合劑對抗體比率 CT               電腦化斷層攝影術 DCTA           1,2-二胺基環己烷四乙酸 DMA            N,N-二甲基乙醯胺 DMSO          二甲基亞碸 EdU             5-乙炔基-2’-去氧尿苷 ESI              電噴霧離子化 EtOH            乙醇 FPLC            快速蛋白質液相層析 FSC              前向散射 GPC3           磷脂醯肌醇蛋白聚醣-3 HPGe            高純度鍺 HPLC           高效液相層析 iTLC            即時薄層層析 IRF              免疫反應分數 mAb             單株抗體 min              分鐘 MS               質譜 NaCl             氯化鈉 nm               奈米 nmol             奈莫耳 PBS              磷酸鹽緩衝鹽水 PET              正電子發射斷層攝影術 RAC             放射性濃度 RCP             放射化學純度 RPMI            羅斯維爾帕克紀念研究所(Roswell Park Memorial Institute) 1640介質 SEC              粒徑篩析層析 SCC             側向散射 TAC             靶向錒結合物(Ac-225標記之ACC) TFA             三氟乙酸 TOF             飛行時間 UPLC           超高效液相層析 UV               紫外Table3.Abbreviations The following table lists the abbreviations used in this article.²²⁵ Ac 225 Ac-225 ACC Antibody-chelator conjugate CAR Chelator-to-antibody ratio CT Computerized tomography DCTA 1,2-Diaminocyclohexanetetraacetic acid DMA N,N-dimethylacetamide DMSO Dimethylsulfoxide EdU 5-ethynyl-2'-deoxyuridine ESI Electrospray ionization EtOH Ethanol FPLC Fast protein liquid chromatography FSC Forward scatter GPC3 Phosphatidylinositol proteoglycan-3 HPGe High purity germanium HPLC High performance liquid chromatography iTLC Instant thin layer chromatography IRF Immunoreactivity fraction mAb Monoclonal antibody min minutes MS Mass spectrometry NaCl Sodium chloride nm nanometer nmol NamoL PBS Phosphate buffered saline PET Positron emission tomography RAC Radioactivity concentration RCP Radiochemical purity RPMI Roswell Park Memorial Institute 1640 medium SEC Size-selective chromatography SCC Side scatter TAC Targeted titanate conjugate (Ac-225 labeled ACC) TFA Trifluoroacetic acid TOF Time of flight UPLC Ultra performance liquid chromatography UV Ultraviolet

所有試劑(其合成並未闡述於實驗部分中)係市面有售或係已知化合物或可自已知化合物藉由熟習此項技術者已知之方法形成。All reagents (whose synthesis is not described in the experimental part) are commercially available or are known compounds or can be formed from known compounds by methods known to those skilled in the art.

本申請案中所闡述之本發明各態樣係藉由下列實例來闡釋，該等實例並不意欲以任何方式限制本發明。The various aspects of the present invention described in this application are illustrated by the following examples, which are not intended to limit the present invention in any way.

測試本文所述實驗之實例用於說明本發明且本發明並不限於所給出之實例。Testing The experimental examples described herein are used to illustrate the present invention and the present invention is not limited to the examples given.

抗GPC3抗體及其靶向放射性核素結合物實例1：抗GPC3抗體及參照化合物之生成所有抗體皆使用標準瞬時轉染程序在HEK293細胞中表現並經由蛋白質-A及粒徑篩析層析自細胞培養上清液純化。Anti-GPC3Antibodies and Targeted Radionuclide ConjugatesExample1: Generation ofAnti-GPC3Antibodies and Reference Compounds All antibodies were expressed in HEK293 cells using standard transient transfection procedures and purified from cell culture supernatants by protein-A and size-selection analysis.

在一些實例中，隨機改變先前技術抗體序列，例如TPP-14890 (考曲妥珠單抗)之彼等。僅少數所得抗體展示改良之行為或基本不變之行為。本文僅包含具有改良或基本不變之特性之實例。In some examples, prior art antibody sequences, such as those of TPP-14890 (trastuzumab), were randomly altered. Only a few of the resulting antibodies exhibited improved behavior or essentially unchanged behavior. Only examples with improved or essentially unchanged properties are included herein.

對先前技術抗體序列(例如TPP-14890之彼等)進一步實施序列種系化且引入其他改變，例如CDR中之離胺酸殘基或潛在天門冬胺酸鹽異構化位點之去除。對所得抗體(TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638、TPP-29642)進行關於以下之表徵：如實例2中所闡述藉由表面電漿共振(SPR)在人類及食蟹猴GPC3上評價之單價結合親和力(K_D)；及如實例3中所闡述藉由流式細胞術量測之該等抗體對使用人類或食蟹猴GPC3轉染之CHO細胞之細胞結合(EC50)。另外，如實例4中所闡述藉由差示掃描量熱法(DSC)測定Fab之熔融溫度(Tm Fab)。所有該等數據皆匯總於表4中。表4：比較本發明抗體之性質對TPP-14890之性質之匯總表。TPP-14890TPP-29312TPP-29537TPP-29538TPP-29636TPP-29638TPP-29642與相TPP-14890比，突變之數量VL: 9 VH: 10VL: 9 VH: 11VL: 10 VH: 11VL: 11 VH: 11VL: 11 VH: 11VL: 11 VH: 10CDR中之離胺酸殘基之數量4100001SPR人類GPC3K_D[M]1.3E-093.6E-095.1E-095.1E-092.1E-095.1E-105.8E-10SPR食蟹猴GPC3K_D[M]1.5E-094.2E-093.8E-095.5E-092.5E-096.7E-108.2E-10流式細胞術hGPC3:CHO EC₅₀[M]9.9E-091.2E-081.0E-082.1E-085.6E-096.7E-094.1E-09流式細胞術cynGPC3:CHO EC₅₀[M]7.7E-099.0E-094.0E-096.1E-098.3E-097.6E-094.2E-09IPAD評分1126575580545563660656DSC Fab T_m[℃]78.772.671.571.170.168.369.3表現率(使用HEK293之標凖表現(標度[L] /表現率)) [mg/L]0.035 / 2402.6 / 2160.035 / 2354.6 / 1830.035 / 3070.035 / 1460.035 / 240Prior art antibody sequences, such as those of TPP-14890, were further subjected to sequence germlining and other changes were introduced, such as removal of lysine residues in the CDRs or potential aspartate isomerization sites. The resulting antibodies (TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638, TPP-29642) were characterized for monovalent binding affinity (_KD ) evaluated on human and cynomolgus monkey GPC3 by surface plasmon resonance (SPR) as described in Example 2; and cell binding (EC50) of these antibodies to CHO cells transfected with human or cynomolgus monkey GPC3 measured by flow cytometry as described in Example 3. In addition, the melting temperature (TmFab) of the Fab was determined by differential scanning calorimetry (DSC) as described in Example 4. All these data are summarized in Table 4.Table4: Summary table comparing the properties of the antibodies of the present invention to the properties of TPP-14890.TPP-14890TPP-29312TPP-29537TPP-29538TPP-29636TPP-29638TPP-29642Compared with TPP-14890, the number of mutations VL: 9 VH: 10 VL: 9 VH: 11 VL: 10 VH: 11 VL: 11 VH: 11 VL: 11 VH: 11 VL: 11 VH: 10Number of lysine residues inCDR 4 1 0 0 0 0 1SPRHumanGPC3K_D[M] 1.3E-09 3.6E-09 5.1E-09 5.1E-09 2.1E-09 5.1E-10 5.8E-10SPRcynomolgus monkey GPC3K_D [M] 1.5E-09 4.2E-09 3.8E-09 5.5E-09 2.5E-09 6.7E-10 8.2E-10Flow cytometry hGPC3:CHO EC₅₀ [M] 9.9E-09 1.2E-08 1.0E-08 2.1E-08 5.6E-09 6.7E-09 4.1E-09Flow cytometry cynGPC3:CHO EC₅₀ [M] 7.7E-09 9.0E-09 4.0E-09 6.1E-09 8.3E-09 7.6E-09 4.2E-09IPADRating 1126 575 580 545 563 660 656DSC Fab T_m [°C] 78.7 72.6 71.5 71.1 70.1 68.3 69.3Expression rate (using HEK293standard expression (scale [L] /expression rate)) [mg/L] 0.035 / 240 2.6 / 216 0.035 / 235 4.6 / 183 0.035 / 307 0.035 / 146 0.035 / 240

令人吃驚地，儘管與TPP-14890相比VH及VL二者中有極高數量之突變，但變體經鑑別：a)仍可以重組方式產生；b)在對重組或細胞人類GPC3之結合中展示相當或優良親和力；c)對食蟹猴GPC3仍展示結合；d)仍展示Fab部分之穩定性，即T_m高於65℃；及e)在標準HEK293E表現系統中展示高於145 mg/L之表現率(如表4中所列示)，暗示良好可製造性。然而，種系偏差之數量顯著減少，此亦在大大減小之電腦免疫原性評分(IPAD, Epibase)以及活體外免疫原性評價(Epibase)中得以反映。另外，CDR內之離胺酸殘基之數量自TPP-14890之CDR中之四個離胺酸殘基減少至本發明抗體之CDR中之一個或甚至無離胺酸殘基(表4)。減少CDR內之離胺酸殘基之數量可降低螯合劑在CDR區域內偶合至抗體之風險。在CDR內之螯合劑之結合會影響抗體結合至其靶GPC3之結合。針對CDR中不含有任何離胺酸殘基之變體TPP-29537、TPP-29538、TPP-29636及TPP-29638，此風險分別得以完全避免。Surprisingly, despite the extremely high number of mutations in both VH and VL compared to TPP-14890, variants were identified that: a) could still be produced recombinantly; b) showed comparable or excellent affinity in binding to recombinant or cellular human GPC3; c) still showed binding to cynomolgus monkey GPC3; d) still showed stability of the Fab portion, i.e.,_Tm above 65°C; and e) showed expression rates above 145 mg/L in the standard HEK293E expression system (as listed in Table 4), suggesting good manufacturability. However, the amount of germline deviation was significantly reduced, which was also reflected in the greatly reduced in silico immunogenicity scores (IPAD, Epibase) as well as in vitro immunogenicity assessments (Epibase). In addition, the number of lysine residues within the CDRs was reduced from four lysine residues in the CDRs of TPP-14890 to one or even no lysine residues in the CDRs of the antibodies of the present invention (Table 4). Reducing the number of lysine residues within the CDRs reduces the risk of chelators coupling to the antibody within the CDR region. Binding of chelators within the CDRs can affect the binding of the antibody to its target GPC3. This risk was completely avoided for the variants TPP-29537, TPP-29538, TPP-29636, and TPP-29638, which do not contain any lysine residues in the CDRs, respectively.

實例2：TPP-14890與本發明抗體之SPR對比為評價抗GPC3抗體之結合動力學及親和力以及其對食蟹猴GPC3之交叉反應性，使用表面電漿共振(SPR)來實施結合分析。在Biacore T200儀器(原GE Healthcare care Inc.，現Cytiva)上在25℃下使用共價偶合至CM5感測器晶片(Cytiva)之抗人類Fc捕獲抗體來運行基於SPR之實驗。Example2:SPRcomparison ofTPP-14890and the antibody of the present invention To evaluate the binding kinetics and affinity of the anti-GPC3 antibody and its cross-reactivity to cynomolgus monkey GPC3, surface plasmon resonance (SPR) was used to perform binding analysis. SPR-based experiments were run on a Biacore T200 instrument (formerly GE Healthcare care Inc., now Cytiva) at 25°C using anti-human Fc capture antibodies covalently coupled to a CM5 sensor chip (Cytiva).

使用胺偶合套組(Cytiva)遵循製造商之說明書來共價偶合抗人類Fc捕獲抗體，直至大約5.000 RU。在SPR中，捕獲程度、R_Max以及所得活性%針對所有變體皆展示相當之行為(數據未展示)。在多循環動力學模式中以0.039 nM – 200 nM之滴定系列注射被分析物(人類及食蟹猴GPC3，R&D系統，目錄號分別為2119-GP及10509-GP)。在HBS-EP+ (Cytiva)中運行實驗。締合時間設置為300 s，解離時間設置為1.200 s且流速設置為30 µl/min。使用Biacore T200評估軟體雙重參考獲得之數據(減去參照細胞及0 nM濃度信號)且將其擬合為1:1蘭格繆爾(Langmuir)結合等溫線(RI設置為恆定)。結果顯示於表5中。表5：使用SPR之TPP-14890及本發明抗體之動力學數據。配體被分析物k_a[1/Ms]k_d[1/s]K_D[M]TPP-14890人類2.1E+052.6E-041.3E-09食蟹猴1.8E+052.6E-041.5E-09TPP-29312人類2.0E+057.2E-043.6E-09食蟹猴1.6E+056.9E-044.2E-09TPP-29537人類1.8E+056.5E-043.6E-09食蟹猴1.6E+056.3E-043.8E-09TPP-29538人類1.9E+059.7E-045.1E-09食蟹猴1.6E+059.0E-045.5E-09TPP-29636人類2.0E+054.2E-042.1E-09食蟹猴1.7E+054.1E-042.5E-09TPP-29638人類2.4E+051.2E-045.1E-10食蟹猴2.0E+051.3E-046.7E-10TPP-29642人類2.1E+051.2E-045.8E-10食蟹猴1.8E+051.5E-048.2E-10Anti-human Fc capture antibody was covalently coupled using an amine coupling kit (Cytiva) following the manufacturer's instructions to approximately 5.000 RU. In SPR, the extent of capture, R_Max and the resulting % activity showed comparable behavior for all variants (data not shown). Analytes (human and cynomolgus GPC3, R&D Systems, Cat. Nos. 2119-GP and 10509-GP, respectively) were injected in a titration series from 0.039 nM – 200 nM in multi-cycle kinetic mode. Experiments were run in HBS-EP+ (Cytiva). The association time was set to 300 s, the dissociation time to 1.200 s and the flow rate to 30 µl/min. Data obtained using Biacore T200 evaluation software double reference (reference cells and 0 nM concentration signal subtracted) were fitted to a 1:1 Langmuir binding isotherm (RI set to constant). The results are shown in Table 5.Table5: Kinetic data of TPP-14890 and antibodies of the invention using SPR.LigandAnalytek_a[1/Ms]k_d[1/s]K_D[M]TPP-14890 Human 2.1E+05 2.6E-04 1.3E-09 Cynomolgus monkey 1.8E+05 2.6E-04 1.5E-09TPP-29312 Human 2.0E+05 7.2E-04 3.6E-09 Cynomolgus monkey 1.6E+05 6.9E-04 4.2E-09TPP-29537 Human 1.8E+05 6.5E-04 3.6E-09 Cynomolgus monkey 1.6E+05 6.3E-04 3.8E-09TPP-29538 Human 1.9E+05 9.7E-04 5.1E-09 Cynomolgus monkey 1.6E+05 9.0E-04 5.5E-09TPP-29636 Human 2.0E+05 4.2E-04 2.1E-09 Cynomolgus monkey 1.7E+05 4.1E-04 2.5E-09TPP-29638 Human 2.4E+05 1.2E-04 5.1E-10 Cynomolgus monkey 2.0E+05 1.3E-04 6.7E-10TPP-29642 Human 2.1E+05 1.2E-04 5.8E-10 Cynomolgus monkey 1.8E+05 1.5E-04 8.2E-10

如自動力學數據可見，TPP-29312、TPP-29537、TPP-29538及TPP-29636展示之親和力(K_D)之數量級與TPP-14890相當。TPP-29638及TPP-29642與TPP-14890及其他變體比較展示略高之親和力。As can be seen from the autodynamics data, TPP-29312, TPP-29537, TPP-29538, and TPP-29636 displayed affinities (_KD ) of comparable magnitude to TPP- 14890. TPP-29638 and TPP-29642 displayed slightly higher affinities compared to TPP-14890 and other variants.

實例3：結合至使用人類、小鼠或食蟹猴GPC3瞬時轉染之CHO細胞之GPC3抗體為評價抗GPC3抗體對結合細胞之人類、食蟹猴或鼠類GPC3之結合，藉由FACS評價TPP-14890及本發明抗體對使用人類、食蟹猴或鼠類GPC3瞬時轉染之CHO細胞之結合。Example3:GPC3Antibody Binding toCHOCellsTransiently Transfected withHuman, Mouse or CynomolgusGPC3 To evaluate the binding of anti-GPC3 antibodies to human, cynomolgus or mouse GPC3 bound to cells, the binding of TPP-14890 and the antibodies of the present invention to CHO cells transiently transfected with human, cynomolgus or mouse GPC3 was evaluated by FACS.

使用GPC3表現構築體瞬時轉染CHO細胞CHO-3E7細胞生長於Freestyle F17 (fisherscientific, #10388033)、4mM Glutamax、0.1%普羅尼克(pluronic)、5% CO₂、37℃、35ml旋轉管中75 rpm (TPP, #87050)中。將先前藉由重組DNA技術生成之人類GPC3 (SEQ ID NO:321)、小鼠GPC3 (SEQ ID NO:322)或食蟹獼猴GPC3 (SEQ ID NO:323)表現質體以3x10⁵細胞/ml之細胞密度轉染至CHO-3E7細胞，且然後在37℃、5% CO₂下培育三天。Transient transfection ofCHOcellswithGPC3 expression constructs CHO-3E7 cells were grown in Freestyle F17 (fisherscientific, #10388033), 4mM Glutamax, 0.1% pluronic, 5% CO₂ , 37°C, 75 rpm (TPP, #87050) in a 35ml rotating tube. Human GPC3 (SEQ ID NO: 321), mouse GPC3 (SEQ ID NO: 322) or cynomolgus monkey GPC3 (SEQ ID NO: 323) expression plasmids previously generated by recombinant DNA technology were transfected into CHO-3E7 cells at a cell density of 3x10⁵ cells/ml and then incubated at 37°C, 5% CO₂ for three days.

藉由FACS分析結合至GPC3轉染之CHO細胞之抗體將5 µl之於PBS pH 7.4中之測試抗體之500 µg/ml溶液添加至微量滴定板(Eppendorf, #0030601300)之孔中。然後，製備1:5稀釋系列直至低至0.0064 µg/ml。接下來，將含有2.5x10⁴GPC3轉染之CHO細胞之45 µl之溶液在冰上預培育30 min，添加至每一孔中。在冰上培育微量滴定板達45 min後，在400xg及4℃下將板離心5 min。棄除上清液且將細胞再懸浮於110 µl冰冷FACS緩衝液(PBS, 3% FCS)中，再次離心且再懸浮於110 µl冰冷FACS緩衝液中以再次洗滌。在第三輪之離心之後，將細胞再懸浮於50 µl FACS緩衝液中，該緩衝液含有1:200稀釋之抗人類Fab-二聚體特異性及Alexa Fluor 488標記之二級試劑(Dianova, #109-456-097)。在冰上培育40 min之後，在400xg、4℃下將板離心5 min。去除上清液且將細胞再懸浮於110 µl FACS緩衝液中以用於最終離心步驟。最後，將細胞再懸浮於含有碘化丙啶(1µg/ml)之FACS緩衝液中，以2500 RPM振盪5 sec且在iQue3儀器上使用4 sec sip作為儀器設置來進行FACS量測。TPP-14980及TPP-29537以及非結合同型對照TPP-5657之結果展示於圖1、圖2、圖3及圖4中。Analysis ofAntibody Binding toGPC3-TransfectedCHO CellsbyFACS 5 µl of a 500 µg/ml solution of the test antibody in PBS pH 7.4 was added to the wells of a microtiter plate (Eppendorf, #0030601300). Then, a 1:5 dilution series was prepared down to 0.0064 µg/ml. Next, 45 µl of a solution containing 2.5x10⁴ GPC3-transfected CHO cells pre-incubated on ice for 30 min was added to each well. After incubation of the microtiter plate on ice for 45 min, the plate was centrifuged at 400xg and 4°C for 5 min. The supernatant was discarded and the cells were resuspended in 110 µl ice-cold FACS buffer (PBS, 3% FCS), centrifuged again and resuspended in 110 µl ice-cold FACS buffer to wash again. After the third round of centrifugation, the cells were resuspended in 50 µl FACS buffer containing a 1:200 dilution of anti-human Fab-dimer specific and Alexa Fluor 488 labeled secondary reagent (Dianova, #109-456-097). After incubation on ice for 40 min, the plate was centrifuged at 400xg, 4°C for 5 min. The supernatant was removed and the cells were resuspended in 110 µl FACS buffer for the final centrifugation step. Finally, the cells were resuspended in FACS buffer containing propidium iodide (1 µg/ml) and FACS measurements were performed on the iQue3 instrument with 2500 RPM shaking for 5 sec and 4 sec sip as the instrument setting. The results for TPP-14980 and TPP-29537 and the non-binding isotype control TPP-5657 are shown in Figures 1, 2, 3, and 4.

使用在軟體GraphPad Prism中積分之非線性回歸分析自FACS結合曲線來計算EC50值。已測定本文闡述之所有抗體變體之結果且匯總於表6中。表6：TPP-14890及本發明抗體在使用人類、食蟹猴或鼠類GPC3瞬時轉染之CHO細胞上之EC50值抗體流式細胞術hGPC3:CHO [EC50 M]流式細胞術cynGPC3:CHO [EC50 M]TPP-148909.90E-097.70E-09TPP-293121.20E-089.00E-09TPP-295371.00E-084.00E-09TPP-295382.10E-086.10E-09TPP-296365.60E-098.30E-09TPP-296386.70E-097.60E-09TPP-296424.10E-094.20E-09EC50 values were calculated from FACS binding curves using nonlinear regression analysis integrated in the software GraphPad Prism. The results for all antibody variants described herein have been determined and are summarized in Table 6.Table6: EC50 values of TPP-14890 and antibodies of the invention on CHO cells transiently transfected with human, cynomolgus monkey or mouse GPC3antibodyFlow cytometryhGPC3:CHO [EC50 M]Flow cytometrycynGPC3:CHO [EC50 M] TPP-14890 9.90E-09 7.70E-09 TPP-29312 1.20E-08 9.00E-09 TPP-29537 1.00E-08 4.00E-09 TPP-29538 2.10E-08 6.10E-09 TPP-29636 5.60E-09 8.30E-09 TPP-29638 6.70E-09 7.60E-09 TPP-29642 4.10E-09 4.20E-09

總而言之，TPP-14890、TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642展示相當FACS結合性質：所有抗體皆展示對人類及食蟹猴GPC3之強結合，而未檢測到對鼠類GPC3之結合。同型對照抗體TPP-5657不展示對人類、鼠類或食蟹猴GPC3之任何結合。In summary, TPP-14890, TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638 and TPP-29642 showed comparable FACS binding properties: all antibodies showed strong binding to human and cynomolgus monkey GPC3, while no binding to mouse GPC3 was detected. The isotype control antibody TPP-5657 did not show any binding to human, mouse or cynomolgus monkey GPC3.

實例4：抗GPC3抗體之熱穩定性使用VP-毛細管DSC系統(Malvern Instruments GmbH，原MicroCal Inc.)以0.137 ml之細胞體積使用差示掃描量熱法(DSC)來測定各種抗GPC3抗體之熱穩定性。在PBS緩衝液中將所有試樣稀釋至之0.5 mg/mL最終濃度，且使用無蛋白質之緩衝液對照作為參照。以120℃/h之掃描速率自20℃至95℃掃描試樣。使用Origin 7.0數據分析(OriginLab Corp.)分析數據。結果顯示於表6中。表6：各種GPC3抗體之Fab熔融溫度。抗體Fab之Tm [℃]TPP-1489078.7TPP-2931272.6TPP-2953771.5TPP-2953871.1TPP-2963670.1TPP-2963868.3TPP-2964269.3Example4:Thermal stability ofanti-GPC3 antibodies Differential scanning calorimetry (DSC) was used to determine the thermal stability of various anti-GPC3 antibodies using a VP-capillary DSC system (Malvern Instruments GmbH, formerly MicroCal Inc.) with a cell volume of 0.137 ml. All samples were diluted to a final concentration of 0.5 mg/mL in PBS buffer, and a protein-free buffer control was used as a reference. Samples were scanned from 20°C to 95°C at a scan rate of 120°C/h. Data were analyzed using Origin 7.0 data analysis (OriginLab Corp.). The results are shown in Table 6.Table6: Fab melting temperatures of various GPC3 antibodies.antibody TmofFab[℃] TPP-14890 78.7 TPP-29312 72.6 TPP-29537 71.5 TPP-29538 71.1 TPP-29636 70.1 TPP-29638 68.3 TPP-29642 69.3

所有本發明抗體皆展示遠大於65℃之Fab之Tm值且由此充分適於人類療法之臨床研發。All antibodies of the invention exhibited Tm values of Fab much greater than 65°C and are thus well suited for clinical development in human therapy.

實例5：抗體序列之種系化先前技術抗體TPP-14890與如IMGT資料庫中所闡述之人類種系相比具有多個偏差(參照http://www.imgt.org/)。Example5:Germlineation of Antibody Sequences Prior Art Antibody TPP-14890 has multiple deviations compared to the human germline as described in the IMGT database (see http://www.imgt.org/).

TPP-14890與所鑑別之最接近種系輕鏈(IGKV2-28-IGKJ2, TPP-35038；連續編號；SEQ ID NO:325)相比具有15個偏差：I2V、L30V、G34R、Y35N、N36T、D39H、L55K、G56V、A60F、M94S、A96N、L97T、Q98H、T99V、Y101P。TPP-14890 has 15 deviations from the closest identified germline light chain (IGKV2-28-IGKJ2, TPP-35038; sequential numbering; SEQ ID NO: 325): I2V, L30V, G34R, Y35N, N36T, D39H, L55K, G56V, A60F, M94S, A96N, L97T, Q98H, T99V, Y101P.

TPP-14890與在CDR-H3之外側鑑別之最接近種系重鏈(IGHV1-46-IGHJ4, TPP-35037；連續編號；SEQ-ID NO: 324)相比具有17個偏差：S31D、Y33E、I50A、I51L、N52D、S54K、G55T、S57D、S59A、A61S、Q65K、M70L、R72A、T74K、V79A、R87T、A97T。TPP-14890 has 17 deviations compared to the closest germline recombinant identified outside of CDR-H3 (IGHV1-46-IGHJ4, TPP-35037; sequential numbering; SEQ-ID NO: 324): S31D, Y33E, I50A, I51L, N52D, S54K, G55T, S57D, S59A, A61S, Q65K, M70L, R72A, T74K, V79A, R87T, A97T.

抗體與其最接近人類種系序列相比之較少量種系偏差在不影響抗原結合下減小抗體之鼠類含量或噬菌體展示衍生之人類抗體，由此增加整體「人類性」且使得由彼等分子引起之免疫原性之風險減小(Hwang等人；Methods，第36卷，第1期，2005年， 35-42頁，ISSN 1046-2023)。表7中闡述細節。表7：與TPP-14890家族之人類參考序列相比，輕鏈及重鏈(排除CDR-H3)之種系偏差之數量及各別突變。TPP-ID輕鏈偏差之數量重鏈偏差之數量突變VL突變VH(Ref.序列IGKV2-28-IGKJ2, TPP-35038)(Ref.序列IGHV1-46-IGHJ4, TPP-35037)148901517I2V、L30V、G34R、Y35N、N36T、D39H、L55K、G56V、A60F、M94S、A96N、L97T、Q98H、T99V、Y101PS31D、Y33E、I50A、I51L、N52D、S54K、G55T、S57D、S59A、A61S、Q65K、M70L、R72A、T74K、V79A、R87T、A97T2953778D39H、L55R、M94S、A96N、L97T、Q98H、T99VS31D、Y33E、I50A、I51L、N52D、G55T、K63R、A97T2931279D39H、L55R、M94S、A96N、L97T、Q98H、T99VS31D、Y33E、I50A、I51L、N52D、S54K、G55T、K63R、A97T2953868D39H、L55R、M94S、A96N、L97T、Q98HS31D、Y33E、I50A、I51L、N52D、G55T、K63R、A97T2963668D39H、L55R、M94I、A96N、L97T、Q98HS31D、Y33E、I50A、I51L、N52D、G55T、K63R、A97T2963878D39H、L55R、A60L、M94I、A96N、L97T、Q98HS31D、Y33E、I50A、I51L、N52D、G55T、K63R、A97T2964279D39H、L55R、A60L、M94I、A96N、L97T、Q98HS31D、Y33E、I50A、I51L、N52D、S54K、G55T、K63R、A97TSmaller amounts of germline deviations of antibodies compared to their closest human germline sequences reduce the murine content of the antibody or phage display derived human antibodies without affecting antigen binding, thereby increasing the overall "humanity" and reducing the risk of immunogenicity caused by those molecules (Hwang et al.; Methods, Vol. 36, No. 1, 2005, pp. 35-42, ISSN 1046-2023). Details are explained in Table 7.Table7: Number and individual mutations of germline deviations in the light and heavy chains (excluding CDR-H3) compared to the human reference sequence of the TPP-14890 family.TPP-IDThe amount of light chain deviationThe amount of rechain deviationMutationVLMutantVH (Ref. Sequence IGKV2-28-IGKJ2, TPP-35038) (Ref. Sequence IGHV1-46-IGHJ4, TPP-35037) 14890 15 17 I2V, L30V, G34R, Y35N, N36T, D39H, L55K, G56V, A60F, M94S, A96N, L97T, Q98H, T99V, Y101P S31D, Y33E, I50A, I51L, N52D, S54K, G55T, S57D, S59A, A61S, Q65K, M70L, R72A, T74K, V79A, R87T, A97T 29537 7 8 D39H, L55R, M94S, A96N, L97T, Q98H, T99V S31D, Y33E, I50A, I51L, N52D, G55T, K63R, A97T 29312 7 9 D39H, L55R, M94S, A96N, L97T, Q98H, T99V S31D, Y33E, I50A, I51L, N52D, S54K, G55T, K63R, A97T 29538 6 8 D39H, L55R, M94S, A96N, L97T, Q98H S31D, Y33E, I50A, I51L, N52D, G55T, K63R, A97T 29636 6 8 D39H, L55R, M94I, A96N, L97T, Q98H S31D, Y33E, I50A, I51L, N52D, G55T, K63R, A97T 29638 7 8 D39H, L55R, A60L, M94I, A96N, L97T, Q98H S31D, Y33E, I50A, I51L, N52D, G55T, K63R, A97T 29642 7 9 D39H, L55R, A60L, M94I, A96N, L97T, Q98H S31D, Y33E, I50A, I51L, N52D, S54K, G55T, K63R, A97T

與TPP-14890抗體比較，本發明抗體攜帶減小數量之自種系之胺基酸偏差。實際上，TPP-14890與人類種系具有總計32個種系偏差。與之相比，本發明抗體與人類種系僅具有總計14-16個種系偏差，使得其在人類療法中之免疫原性之風險減小。Compared to the TPP-14890 antibody, the antibodies of the present invention carry a reduced number of amino acid deviations from the germline. In fact, TPP-14890 has a total of 32 germline deviations from the human germline. In comparison, the antibodies of the present invention only have a total of 14-16 germline deviations from the human germline, which reduces the risk of immunogenicity in human therapy.

總而言之，與已人類化之先前技術抗體TPP-14890比較，TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642具有顯著較少量種系偏差，由此進一步降低在用於人類療法時之免疫原性反應之風險。In summary, TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638, and TPP-29642 have significantly less germline bias than the humanized prior art antibody TPP-14890, thereby further reducing the risk of immunogenic reactions when used in human therapy.

實例6：電腦免疫原性評價使用兩種獨立電腦預測方法來評價激起人類療法中之免疫原性反應之潛能：IPAD評分IPAD v4.03演算法方法依賴於NetMHCIIpan演算法(Nielsen等人，Quantitative predictions of peptide binding to any HLA-DR molecule of known sequence: NetMHCIIpan. PLoS Comput Biol. 2008年7月4日；4(7):e1000107)，該演算法預測結合至HLA II類分子之肽。簡言之，經由此演算法運行VH及VL序列並篩選出非種系肽。藉由將預測之HLA結合肽之數量與HLA頻率(%)及比例因子(0.19352)相乘來計算HLA評分。然後藉由所有個體HLA評分之總和來生成IPAD評分。使用此方法，介於600與1000之間之IPAD評分相當於市售完全人類及人類化抗體，評分低於600比較有利，而評分高於1000指示免疫原性之風險較高。表8：先前技術抗體與本發明抗體之IPAD評分之對比TPP-IDIPAD評分考曲妥珠單抗TPP-148901126本發明抗體TPP-29312575TPP-29537580TPP-29538545TPP-29636563TPP-29638660TPP-29642656先前技術抗體YP7 (TPP-32713)2333hYP7 (TPP-15081)871hYP9.1b (TPP-14877)7552-F7 (TPP-32714)5974-11G (TPP-32715)7287-5B (TPP-32716)10147-8B (TPP-32717)64517-4D (TPP-32718)1047Example6: In silico immunogenicity assessment Two independent in silico prediction methods were used to assess the potential to provoke an immunogenic response in human therapy:IPADScoring The IPAD v4.03 algorithm method relies on the NetMHCIIpan algorithm (Nielsen et al., Quantitative predictions of peptide binding to any HLA-DR molecule of known sequence: NetMHCIIpan. PLoS Comput Biol. 2008 Jul 4;4(7):e1000107), which predicts peptides that bind to HLA class II molecules. Briefly, VH and VL sequences were run through this algorithm and non-germline peptides were screened out. The HLA score was calculated by multiplying the number of predicted HLA-binding peptides by the HLA frequency (%) and a scaling factor (0.19352). The IPAD score is then generated by summing all individual HLA scores. Using this method, IPAD scores between 600 and 1000 are equivalent to commercially available fully human and humanized antibodies, with scores below 600 being more favorable and scores above 1000 indicating a higher risk of immunogenicity.Table8: Comparison of IPAD scores of prior art antibodies and antibodies of the present inventionTPP-IDIPADRating Trastuzumab TPP-14890 1126Antibodies of the present invention TPP-29312 575 TPP-29537 580 TPP-29538 545 TPP-29636 563 TPP-29638 660 TPP-29642 656Prior Art Antibodies YP7 (TPP-32713) 2333 hYP7 (TPP-15081) 871 hYP9.1b (TPP-14877) 755 2-F7 (TPP-32714) 597 4-11G (TPP-32715) 728 7-5B (TPP-32716) 1014 7-8B (TPP-32717) 645 17-4D (TPP-32718) 1047

如表8中所繪示，TPP-14890具有1129之IPAD評分，與具有低得多之IPAD評分之本發明抗體TPP-29312、TPP-29537、TPP-29538、TPP-29636、TPP-29638及TPP-29642比較，此指示免疫原性之風險較高。TPP-29312、TPP-29537、TPP-29538、TPP-29636之IPAD評分甚至低於600。完全人類抗體2-F7、4-11G、7-5B、7-8B及17-4D與本發明抗體比較展示相當或較差IPAD評分。YP7係鼠類抗體且具有最高IPAD評分。人類化變體hYP7經顯著改良但與本發明抗體比較仍展示較差IPAD評分。同樣，hYP9.1b (亦為人類化抗體)與本發明抗體比較展示較差IPAD評分且與hYP7在類似範圍內。As shown in Table 8, TPP-14890 has an IPAD score of 1129, which indicates a higher risk of immunogenicity compared to the antibodies of the present invention TPP-29312, TPP-29537, TPP-29538, TPP-29636, TPP-29638 and TPP-29642, which have much lower IPAD scores. The IPAD scores of TPP-29312, TPP-29537, TPP-29538, TPP-29636 are even lower than 600. The fully human antibodies 2-F7, 4-11G, 7-5B, 7-8B and 17-4D show comparable or inferior IPAD scores compared to the antibodies of the present invention. YP7 is a mouse antibody and has the highest IPAD score. The humanized variant hYP7 was significantly improved but still showed a poor IPAD score compared to the antibody of the present invention. Similarly, hYP9.1b (also a humanized antibody) showed a poor IPAD score compared to the antibody of the present invention and was in a similar range to hYP7.

Epibase電腦免疫原性平臺(Lonza)Epibase電腦(IS)係來自Lonza之T-細胞表位篩選平臺，其分析且預測蛋白質之潛在免疫原性。EpibaseIn silico Immunogenicity Platform(Lonza) Epibase In silico (IS) is a T-cell epitope screening platform from Lonza that analyzes and predicts the immunogenicity potential of proteins.

使用Epibase平臺之「HLA II類 – Global v4.0」設置對TPP-14890及TPP-29537之序列實施免疫剖析，且比較抗體之假定HLA II類限制性表位含量。在異型層面上針對43個DRB1、8個DRB3/4/5、22個DQ及12個DP (亦即總計86個HLA II類受體)實施剖析。DRB1評分(關鍵表位計數乘以受影響異型之等位基因頻率之產物之總和)展示於表9中。表9：全域群體中測試之抗體之DRB1風險評分。全域DRB1風險評分抗體輕鏈重鏈完整抗體TPP-29537403.4382.5785.9TPP-14890388.6536.1924.7The sequences of TPP-14890 and TPP-29537 were immunoanalyzed using the "HLA Class II - Global v4.0" setting of the Epibase platform and the content of putative HLA Class II restricted epitopes of the antibodies was compared. Analysis was performed on the allotype level for 43 DRB1, 8 DRB3/4/5, 22 DQ and 12 DP (i.e., a total of 86 HLA Class II receptors). The DRB1 score (the sum of the product of the key epitope count multiplied by the allele frequency of the affected allotype) is shown in Table 9.Table9: DRB1 risk score of antibodies tested in the global population.GlobalDRB1risk score antibody Light chain Heavy Chain Complete AntibodyTPP-29537 403.4 382.5 785.9TPP-14890 388.6 536.1 924.7

TPP-29537具有較少量關鍵DRB1表位(25個表位)，TPP-14890具有較高數量之關鍵DRB1表位(30個表位)。類似地，針對DRB3/4/5家族，TPP-29537具有最低表位計數(10個表位)且TPP-14890具有較高之12個表位。抗體TPP-14890及TPP-29537兩者不具有任何關鍵DQ表位。針對DP家族，TPP-29537及TPP-14890二者具有3個DP表位。TPP-29537 has a lower number of key DRB1 epitopes (25 epitopes) and TPP-14890 has a higher number of key DRB1 epitopes (30 epitopes). Similarly, for the DRB3/4/5 family, TPP-29537 has the lowest epitope count (10 epitopes) and TPP-14890 has a higher count of 12 epitopes. Antibodies TPP-14890 and TPP-29537 both do not have any key DQ epitopes. For the DP family, both TPP-29537 and TPP-14890 have 3 DP epitopes.

自表9可顯而易見，TPP-29537具有較低DRB1風險評分，與具有924.7之DRB1風險評分之TPP-14890相比僅為785.9。As can be seen from Table 9, TPP-29537 has a lower DRB1 risk score of only 785.9 compared to TPP-14890 which has a DRB1 risk score of 924.7.

總而言之，TPP-29537之DRB1風險評分與完全人類治療性抗體相當，且表位計數位於人類抗體之上端，而TPP-14890之DRB1風險評分與人類化治療性抗體相當且表位計數位於人類化抗體範圍之下端。因此，考慮整體關鍵表位計數及受影響HLA異型之群體頻率，在人類治療性應用中TPP-14890比TPP-29537具有較大免疫原性風險。In summary, TPP-29537 has a DRB1 risk score comparable to fully human therapeutic antibodies and an epitope count at the upper end of the human antibody range, while TPP-14890 has a DRB1 risk score comparable to humanized therapeutic antibodies and an epitope count at the lower end of the humanized antibody range. Therefore, considering the overall critical epitope count and the population frequency of affected HLA allotypes, TPP-14890 has a greater immunogenicity risk than TPP-29537 in human therapeutic applications.

總而言之，二者獨立地用於電腦預測方法以激起人類療法中之免疫原性反應，IPAD及Epibase指示與TPP-14890相比TPP-29537之免疫原性潛能減小。由此，自患者安全性之角度來看，TPP-29537比TPP-14890更有益。In summary, both independently used in silico methods to provoke immunogenic responses in human therapy, IPAD and Epibase indicate that TPP-29537 has reduced immunogenic potential compared to TPP-14890. Thus, from a patient safety perspective, TPP-29537 is more beneficial than TPP-14890.

實例7：活體外免疫原性評價為進一步證實電腦免疫原性預測，亦使用活體外免疫原性評價方法來評價激起人類療法中之免疫原性反應之潛能。為此，使用螯合劑Macropa-NCS (如實例10中所闡述)製備結合物TPP-29537-Macropa及TPP-14890-Macropa，獲得TPP-14890及TPP-29537之抗體螯合劑結合物，且對其實施「Epibase在活體外PBMC增殖分析」(Lonza)。Example7: In vitro immunogenicity assessment To further confirm the in silico immunogenicity prediction, an in vitro immunogenicity assessment method was also used to assess the potential to elicit an immunogenic response in human therapy. To this end, the conjugates TPP-29537-Macropa and TPP-14890-Macropa were prepared using the chelator Macropa-NCS (as described in Example 10), antibody chelator conjugates of TPP-14890 and TPP-29537 were obtained, and the "Epibase in vitro PBMC proliferation assay" (Lonza) was performed on them.

該分析實施Lonza活體外T細胞分析平臺以使用來自健康供體之人類外周血單核細胞(PBMC)來評價T細胞增殖。在來自HLA-DRB1頻率類似全域群體頻率之31個健康人類供體之PBMC中評價由測試分子誘導之T細胞反應。利用流式細胞術進行PBMC分析以檢測使用測試分子刺激之後之增殖細胞之數量。The assay implements the Lonza in vitro T cell assay platform to assess T cell proliferation using human peripheral blood mononuclear cells (PBMCs) from healthy donors. T cell responses induced by the test molecule were assessed in PBMCs from 31 healthy human donors with HLA-DRB1 frequencies similar to the global population frequency. PBMC analysis was performed using flow cytometry to detect the number of proliferating cells after stimulation with the test molecule.

將PBMC接種於96孔板中。將在分析介質中稀釋之測試蛋白質添加至細胞。分析介質單獨用作空白且KLH (鑰孔蟲戚血蘭素(Keyhole Limpet Hemocyanin))用作天然陽性對照。在5% CO₂中且在37℃下將PBMC培育7天。在第7天，用CD3及CD4標記物標記PBMC且納入DNA之5-乙炔基-2’-去氧尿苷(EdU)用作增殖標記物。PBMCs were seeded in 96-well plates. Test proteins diluted in assay medium were added to the cells. Assay medium alone was used as a blank and KLH (Keyhole Limpet Hemocyanin) was used as a natural positive control. PBMCs were incubated for 7 days in 5%_CO2 and at 37°C. On day 7, PBMCs were labeled with CD3 and CD4 markers and 5-ethynyl-2'-deoxyuridine (EdU) incorporated into DNA was used as a proliferation marker.

KLH用作陽性對照且為CD4+ T細胞增殖之強效驅動劑，此指示分析按預計實施：與空白條件(平均SI 28.94且p＜ 0.0001)比較，31/31供體(100%)展示對KLH之顯著T細胞反應(SI ≥2.00且p＜ 0.05)。結果顯示於表10中。表10：與空白比較每一測試條件之供體反應。KLH was used as a positive control and is a potent driver of CD4+ T cell proliferation, indicating that the assay performed as expected: 31/31 donors (100%) demonstrated significant T cell responses to KLH (SI ≥2.00 and p < 0.05) compared to blank conditions (mean SI 28.94 and p < 0.0001). The results are shown in Table 10.Table10: Donor responses for each tested condition compared to blank.

SI =刺激指數。p值係關於單一試樣t-測試假設，即既定抗原之SI之頻率分佈代表平均SI值為1之分佈。測試分子測試分子特性群體分析個別供體T細胞反應螯合劑抗體比率(CAR)來自粒徑篩析層析之%單體含量平均SIp值具有反應供體之數量(亦即SI ≥2.00且p＜ 0.05之供體)KLH不適用未測試28.94＜ 0.000131 (100%)TPP-14890-Macropa0.7991.20＜ 0.10945 (16.13%)TPP-29537-Macropa (批次1)0.599.90.88＜ 0.23912 (6.45%)TPP-29537-Macropa (批次2)0.898.30.83＜ 0.05492 (6.45%)SI = stimulation index. The p-value relates to the single sample t-test hypothesis that the frequency distribution of SI for a given antigen represents the distribution with a mean SI value of 1.Test moleculesTesting molecular propertiesGroup analysisIndividual donorTcell responses Chelator-Antibody Ratio (CAR) % Monomer content from size-screening analysis Average SI p-value The number of reactive donors (i.e. donors with SI ≥ 2.00 and p < 0.05) KLH Not applicable Not tested 28.94 < 0.0001 31 (100%) TPP-14890-Macropa 0.7 99 1.20 ＜ 0.1094 5 (16.13%) TPP-29537-Macropa (Batch 1) 0.5 99.9 0.88 ＜ 0.2391 2 (6.45%) TPP-29537-Macropa (Batch 2) 0.8 98.3 0.83 ＜ 0.0549 2 (6.45%)

與電腦預測一致，甚至在測試兩種不同批次時，與對TPP-29537抗體螯合劑結合之反應(2個供體，等於6%)比較，較多供體對TPP-14890抗體螯合劑結合具有反應(5個供體，等於16%)。Consistent with computer predictions, even when two different batches were tested, more donors responded to TPP-14890 antibody chelator binding (5 donors, equal to 16%) compared to TPP-29537 antibody chelator binding (2 donors, equal to 6%).

總而言之，TPP-14890抗體螯合劑結合在16%之供體中顯著提高CD4+ T細胞反應，表明免疫原性之中等風險。與之相比，TPP-29537抗體螯合劑結合在僅6%之供體中具有反應。因此，可將TPP-29537抗體螯合劑結合排名為針對人類療法中之免疫原性僅具有低風險(＜ 10%之具有反應之供體)。因此，自患者安全性角度看來，TPP-29537比TPP-14890更有益。In summary, TPP-14890 antibody chelator conjugate significantly increased CD4+ T cell responses in 16% of donors, indicating a moderate risk of immunogenicity. In comparison, TPP-29537 antibody chelator conjugate was reactive in only 6% of donors. Therefore, TPP-29537 antibody chelator conjugate can be ranked as having only a low risk of immunogenicity in human therapy (< 10% of reactive donors). Therefore, TPP-29537 is more beneficial than TPP-14890 from a patient safety perspective.

實例8：將抗GPC3抗體內化至HepG2細胞以下部分闡述不同抗GPC3抗體關於其內化特徵之表徵。 內化係抗體在結合至其特異性腫瘤靶之後被吸收至腫瘤細胞中之過程。內化係抗體藥物結合物(ADC)之有效性之必要前體，此乃因結合至抗體之毒素僅在細胞內側釋放且發揮其細胞毒性效應。內化並非TAC之效應之前體，此乃因甚至不經內化放射性輻射即可在TAC結合至受體之後殺死靶細胞。然而，內化亦為有益的，此乃因其有助於改良腫瘤滯留。Example8:Internalization ofanti-GPC3 antibodies intoHepG2cells The following section describes the characterization of different anti-GPC3 antibodies with respect to their internalization characteristics. Internalization is the process by which an antibody is taken up into tumor cells after binding to its specific tumor target. Internalization is a necessary precursor to the effectiveness of antibody-drug conjugates (ADCs) because the toxin bound to the antibody is only released inside the cell and exerts its cytotoxic effect. Internalization is not a precursor to the effect of TACs because even without internalizing radioactive radiation, target cells can be killed after TACs bind to receptors. However, internalization is also beneficial because it helps improve tumor retention.

CypHer5E直接標記之抗GPC3抗體之內化。在第一步驟中，使用pH敏感染料CypHer5E-NHS-酯(Cytiva, PA 15401)在1 mg/ml之濃度下直接標記抗體TPP-29537及TPP-14890，根據製造商說明書該染料偶合至抗體之離胺酸殘基。Internalization of anti-GPC3 antibodiesdirectly labeled withCypHer5E . In the first step, antibodies TPP-29537 and TPP-14890 were directly labeled at a concentration of 1 mg/ml using the pH-sensitive dye CypHer5E-NHS-ester (Cytiva, PA 15401), which was coupled to the lysine residue of the antibody according to the manufacturer's instructions.

在標記之後，染料對蛋白質比率估計為針對TPP-29537之21.5及針對TPP-14890之16.4。After labeling, the dye-to-protein ratio was estimated to be 21.5 for TPP-29537 and 16.4 for TPP-14890.

將HepG2細胞(20,000個細胞/孔)在含有10% FCS之100 µl EMEM介質(ATCC, 30-2003)中平鋪在Greiner µCLEAR 96-孔微量滴定板(Greiner, #655090)上。在37℃及5% CO₂下過夜培育之後，將於10 µl介質中之10 µg Hoechst 33342 (Invitrogen, #H3570)染色溶液添加至每一孔且於暗處培育10 min。接下來，藉由90 µl新鮮介質以及各別所關注抗體去除並代替該介質(10 µL /孔，2 µg/mL之最終濃度)。然後在培育器中在37℃及5 % CO₂下培育細胞。在0.15、1、3、5、及24小時藉由高含量分析系統ImageXpress (Molecular Devices)獲取圖片。使用軟體MetaXpress (Molecular Devices)應用Transfluor模組(設置「囊泡5_10_500」及「核染色6_32_100」)來分析影像。計算個體抗體之每個細胞之囊泡面積且將在5小時所得值展示於圖5中。HepG2 cells (20,000 cells/well) were plated in 100 µl EMEM medium (ATCC, 30-2003) containing 10% FCS on Greiner µCLEAR 96-well microtiter plates (Greiner, #655090). After overnight incubation at 37°C and 5% CO₂ , a staining solution of 10 µg Hoechst 33342 (Invitrogen, #H3570) in 10 µl medium was added to each well and incubated in the dark for 10 min. Next, the medium was removed and replaced by 90 µl fresh medium and the respective antibody of interest (10 µL/well, 2 µg/mL final concentration). The cells were then incubated in an incubator at 37°C and 5% CO₂ . Images were acquired at 0.15, 1, 3, 5, and 24 hours using the high-content analysis system ImageXpress (Molecular Devices). Images were analyzed using the software MetaXpress (Molecular Devices) using the Transfluor module (settings “Vesicle 5_10_500” and “Nuclear Stain 6_32_100”). The vesicle area per cell was calculated for individual antibodies and the values obtained at 5 hours are shown in FIG5 .

Fabfluor標記之抗體之內化。將HepG2細胞(20,000個細胞/孔)在含有10% FCS之100 µl EMEM介質(ATCC, 30-2003)中平鋪在Corning BioCoat µClear 96-孔MTP (356640)上且在37℃及5% CO₂下過夜培育。在下一天，將5 µl之Incucyte^®人類Fabfluor-pH抗體標記試劑(Satorius, #4722) (即染料標記之結合至含Fc蛋白質之Fab片段)之10 µg/ml溶液與5 µl之個體測試抗體之20 µg/ml溶液在室溫下混合20 min。然後，添加90 µl EMEM介質且在振盪器中將混合物再培育5分鐘。另外，將於110 µl介質中之10 µg Hoechst 33342 (Invitrogen, #H3570)染色溶液添加至含有細胞之板之每一孔中且於暗處培育10 min。然後，去除組織培養板之介質且替換為90 µl之抗體 – Fabfluor混合物。然後在37℃及5 % CO₂下在培育器中培育細胞。在0.15、1、3、5、及24小時藉由高含量分析系統ImageXpress (Molecular Devices)獲取圖片。使用軟體MetaXpress (Molecular Devices)應用Transfluor模組(設置「囊泡5_10_500」及「核染色6_32_100」)來分析影像。計算個體抗體之每個細胞之囊泡面積且將在5小時所得值展示於圖6中。Internalization ofFabfluor -labeled antibodies. HepG2 cells (20,000 cells/well) were plated on Corning BioCoat µClear 96-well MTP (356640) in 100 µl EMEM medium (ATCC, 30-2003) containing 10% FCS and incubated overnight at 37°C and 5% CO_2. On the next day, 5 µl of a 10 µg/ml solution of Incucyte^® Human Fabfluor-pH Antibody Labeling Reagent (Satorius, #4722) (i.e., dye-labeled Fab fragments bound to Fc-containing proteins) was mixed with 5 µl of a 20 µg/ml solution of the individual test antibody at room temperature for 20 min. Then, 90 µl of EMEM medium was added and the mixture was incubated for another 5 minutes on a shaker. In addition, 10 µg of Hoechst 33342 (Invitrogen, #H3570) staining solution in 110 µl of medium was added to each well of the plate containing cells and incubated in the dark for 10 min. Then, the medium of the tissue culture plate was removed and replaced with 90 µl of the antibody-Fabfluor mixture. The cells were then incubated in_an incubator at 37°C and 5% CO2. Images were acquired at 0.15, 1, 3, 5, and 24 hours by the high-content analysis system ImageXpress (Molecular Devices). Images were analyzed using the software MetaXpress (Molecular Devices) using the Transfluor module (settings "Vesicle 5_10_500" and "Nuclear Stain 6_32_100"). The vesicle area per cell was calculated for individual antibodies and the values obtained at 5 hours are shown in FIG6 .

顯而易見，使用CypHer5E經由離胺酸殘基之共價標記直接標記抗體時，與TPP-29537比較TPP-14890展示較低內化(參見圖5)。相反，使用Fabfluor非共價標記，兩種抗GPC3抗體展示相當內化比率(參見圖6)。內化比率之該等差異可由以下事實闡釋：使用離胺酸偶合化學之直接標記亦會標記TPP-14890之CDR內之全部4個離胺酸殘基中之一或多者，此導致結合受影響且由此內化比率降低。因本發明抗體TPP-29537在其CDR內不含離胺酸殘基，故結合及內化比率不受影響。在Fabfluor標記之情形下，兩種抗GPC3抗體展示相當內化比率，此乃因此設置中不使用離胺酸標記步驟。此實例展示，與在其CDR內包括離胺酸殘基之先前技術抗體(例如，抗體TPP-14890)相比，本發明抗體更適於基於離胺酸化學結合之應用，例如ADC或靶向錒結合物。It is evident that TPP-14890 exhibited lower internalization compared to TPP-29537 when the antibody was directly labeled using CypHer5E via covalent labeling of lysine residues (see Figure 5). In contrast, both anti-GPC3 antibodies exhibited comparable internalization ratios using Fabfluor non-covalent labeling (see Figure 6). These differences in internalization ratios can be explained by the fact that direct labeling using lysine coupling chemistry would also label one or more of all four lysine residues within the CDRs of TPP-14890, which resulted in affected binding and thus reduced internalization ratios. Since the antibody TPP-29537 of the present invention does not contain lysine residues within its CDRs, binding and internalization ratios were not affected. In the case of Fabfluor labeling, the two anti-GPC3 antibodies showed comparable internalization ratios, since no lysine labeling step was used in this setting. This example shows that the antibodies of the present invention are more suitable for applications based on lysine chemical conjugation, such as ADCs or targeted titanate conjugates, than prior art antibodies (e.g., antibody TPP-14890) that include lysine residues within their CDRs.

實例9：GPC3抗體對不同細胞系之細胞表面結合使用不同培養細胞藉由流式細胞術來探究TPP-14890、TPP-29537及同型對照TPP-754之細胞表面結合。在RPMI + 10% FCS、DMEM + 15% FCS、RPMI + 10 % FCS、MEM + 10% FCS及MEM + 10% FCS中分別培養細胞系SU-DHL-1、HFF-1、HepG2、Hep3B2、PLC/PRF/5。原代人類心臟纖維母細胞(HCF)及原代人類肺纖維母細胞(HPF)購買自Promocell且根據製造商說明書培養。使用非酶促解離溶液(Gibco #13151-014)分離細胞以進行表面染色。在冰冷FACS緩衝液(具有來自Sigma #F2442之3% FCS之PBS -/-)中將細胞洗滌兩次。所有後續步驟皆在冰上實施以預防抗體內化。將於100µl中之200,000個細胞轉移至V形96孔板之每個孔中。使用Alexa Fluor 488標記之TPP-14890、Alexa Fluor 488標記之抗GPC3抗體TPP-29537或FITC標記之同型對照抗體TPP-754染色細胞。使用偶合至離胺酸殘基上存在之一級胺基之來自Invitrogen之A10235, Alexa Fluor 488直接標記抗體。遵循製造商說明書測定標記程度。TPP-14890具有6.07之標記程度(DOL)；TPP-29537 DOL=7.22且TPP-754 DOL=3.4。所有抗體皆稀釋於FACS緩衝液中以達到10 µg/mL之最終濃度。於暗處在4℃下在Titramax100上振盪染色30 min。使用PBS-/-洗滌細胞且在4℃下以800g離心2 min。隨後小心去除上清液。將細胞再懸浮於含有1:1000 Sytot Blue (Invitrogen S34857)之FACS緩衝液中。於BD Biosciences FACSCanto^TMII流式細胞術系統上量測細胞且使用FlowJo_v10.6.1軟體(BD Biosciences)分析其。使用前向散射(FSC)、側向散射(SCC)及Pacific Blue圈選活、單一細胞。同時量測細胞MESF珠粒(Quantum 488C，批號：14506)。遵循製造商說明書藉助MESF珠粒將量測螢光信號轉化為ABC值(抗體結合能力)。在計算ABC值之後，將同型對照ABC值自靶抗體之值中扣除。將結果繪圖為ABC (細胞表面之抗體結合能力)，參見圖7、圖8及表11。表11：使用TPP-14890或TPP-29537測定之各種細胞系之ABC值試樣名稱ABC值靶-同型對照TPP-14890HFF-10.0SU-DHL-100.0PLC/PRF/50.0HepG241734.3Hep3B29750.7HCF0.0HPF0.0TPP-29537HFF-10.0SU-DHL-100.0PLC/PRF/514495.1HepG2857064.2Hep3B2192253.2HCF0.0HPF0.0Example9: Cell surface binding ofGPC3antibody to different cell lines Cell surface binding of TPP-14890, TPP-29537 and isotype control TPP-754 was investigated by flow cytometry using different cultured cells. Cell lines SU-DHL-1, HFF-1, HepG2, Hep3B2, PLC/PRF/5 were cultured in RPMI + 10% FCS, DMEM + 15% FCS, RPMI + 10% FCS, MEM + 10% FCS and MEM + 10% FCS, respectively. Primary human cardiac fibroblasts (HCF) and primary human lung fibroblasts (HPF) were purchased from Promocell and cultured according to the manufacturer's instructions. Cells were detached for surface staining using non-enzymatic dissociation solution (Gibco #13151-014). Cells were washed twice in ice-cold FACS buffer (PBS -/- with 3% FCS from Sigma #F2442). All subsequent steps were performed on ice to prevent antibody internalization. 200,000 cells in 100 µl were transferred to each well of a V-shaped 96-well plate. Cells were stained with Alexa Fluor 488-labeled TPP-14890, Alexa Fluor 488-labeled anti-GPC3 antibody TPP-29537, or FITC-labeled isotype control antibody TPP-754. Antibodies were directly labeled using A10235, Alexa Fluor 488 from Invitrogen coupled to a primary amine group present on lysine residues. The degree of labeling was determined following the manufacturer's instructions. TPP-14890 had a degree of labeling (DOL) of 6.07; TPP-29537 DOL=7.22 and TPP-754 DOL=3.4. All antibodies were diluted in FACS buffer to reach a final concentration of 10 µg/mL. Staining was performed in the dark at 4°C on a Titramax100 for 30 min with shaking. Cells were washed with PBS-/- and centrifuged at 800g for 2 min at 4°C. The supernatant was then carefully removed. The cells were resuspended in FACS buffer containing 1:1000 Sytot Blue (Invitrogen S34857). The cells were measured on a BD Biosciences FACSCanto^™ II flow cytometry system and analyzed using FlowJo_v10.6.1 software (BD Biosciences). Live, single cells were selected using forward scatter (FSC), side scatter (SCC) and Pacific Blue. Cells were measured simultaneously with MESF beads (Quantum 488C, lot number: 14506). The measured fluorescence signal was converted to ABC values (antibody binding capacity) with the aid of MESF beads following the manufacturer's instructions. After calculating the ABC value, the isotype control ABC value was subtracted from the value of the target antibody. The results were plotted as ABC (antibody binding capacity on the cell surface), see Figure 7, Figure 8 and Table 11.Table11: ABC values of various cell lines measured using TPP-14890 or TPP-29537Sample nameABCvalue target-isotype controlTPP-14890 HFF-1 0.0 SU-DHL-10 0.0 PLC/PRF/5 0.0 HepG2 41734.3 Hep3B2 9750.7 HCF 0.0 HPF 0.0TPP-29537 HFF-1 0.0 SU-DHL-10 0.0 PLC/PRF/5 14495.1 HepG2 857064.2 Hep3B2 192253.2 HCF 0.0 HPF 0.0

顯而易見，在HepG2及Hep3B2之情形下，與Alexa Fluor 488標記之TPP-14890相比，Alexa Fluor 488標記之TPP-29537檢測到之ABC值大約高20倍。針對PLC/PRF/5，Alexa Fluor 488標記之TPP-29537檢測到14495,1之ABC值，而Alexa Fluor 488標記之TPP-14890在細胞表面具有較低量之GPC3之此細胞系中不能夠檢測到任何GPC3。針對ABC = 0之細胞系，GPC3抗體之經計算ABC值低於彼各別細胞系中之同型對照抗體之經計算ABC值。0之ABC值指示無或極低GPC3蛋白質表現。ABC值之該等差異可由以下事實闡釋：使用離胺酸偶合化學以Alexa Fluor 488直接標記亦會標記先前抗體TPP-14890之CDR內之全部4個離胺酸殘基中之一或多者，此導致對GPC3之結合受影響。因本發明抗GPC3抗體TPP-29537在其CDR內不含離胺酸殘基，故結合及所得ABC值不受影響。It is evident that in the case of HepG2 and Hep3B2, Alexa Fluor 488-labeled TPP-29537 detected approximately 20-fold higher ABC values than Alexa Fluor 488-labeled TPP-14890. For PLC/PRF/5, Alexa Fluor 488-labeled TPP-29537 detected an ABC value of 14495,1, while Alexa Fluor 488-labeled TPP-14890 was unable to detect any GPC3 in this cell line with a lower amount of GPC3 on the cell surface. For cell lines with ABC = 0, the calculated ABC value of the GPC3 antibody was lower than the calculated ABC value of the isotype control antibody in those respective cell lines. An ABC value of 0 indicates no or very low GPC3 protein expression. These differences in ABC values can be explained by the fact that direct labeling with Alexa Fluor 488 using lysine coupling chemistry also labels one or more of all four lysine residues within the CDRs of the previous antibody TPP-14890, which results in affected binding to GPC3. Since the anti-GPC3 antibody TPP-29537 of the present invention does not contain lysine residues within its CDRs, binding and the resulting ABC values are not affected.

實例10：靶向錒結合物(TAC)之製備Macropa對GPC3抗體之結合物(ACC)將6-[[16-[(6-羧基-2-吡啶基)甲基]-1,4,10,13-四氧雜-7,16-二氮雜環十八-7-基]甲基]-4-[2-(4-異硫基氰基苯基)乙氧基]吡啶-2-甲酸(Macropa-NCS；下文中縮寫為「Macropa」)溶於DMA中至10 mg/ml。分別將抗GPC3抗體TPP-29537及TPP-14890溶於PBS中至10 mg/mL且使用1M碳酸鹽緩衝液將pH調節至9。將螯合劑與抗體混合且在室溫下於熱混合器中以350 rpm振盪培育兩小時。端視期望螯合劑對抗體比率(CAR)來選擇螯合劑與抗體之莫耳比。藉由FPLC純化產物(管柱：HiLoad 16/600 Superdex 200 pg管柱；流速：1 mL/min；於0.1 M乙酸酯緩衝液(pH 5)中；檢測：UV 280/254 nm)。藉由SEC-UV測定濃度及單體純度(如下文所闡述及表12中所展示)。藉由使用陽性電噴霧離子化(如下文所闡述)係SEC-MS及下列方程式測定CAR：CAR=[(A*An)/A]0，A=最大包絡熵之所有信號之強度，An=含有螯合劑之最大包絡熵之信號。不同ACC之CAR之測定如表12中所展示。表12：不同抗體螯合劑結合物之螯合劑對抗體比率(CAR)、濃度及單體純度抗GPC3 mAb螯合劑結合物螯合劑對抗體比率(CAR)ACC濃度(mg/ml)ACC單體純度(%)TPP-14890MacropaTPP-14890-Macropa0.72.399.9101.9799.9TPP-29537MacropaTPP-29537-Macropa0.55.099.9111.7199.2Example10 :Preparation ofTargeted Titanium Conjugate(TAC)Macropa conjugatetoGPC3antibody(ACC) 6-[[16-[(6-carboxy-2-pyridyl)methyl]-1,4,10,13-tetraoxa-7,16-diazacyclooctadeca-7-yl]methyl]-4-[2-(4-isothiocyanatophenyl)ethoxy]pyridine-2-carboxylic acid (Macropa-NCS; hereinafter abbreviated as "Macropa") was dissolved in DMA to 10 mg/ml. Anti-GPC3 antibodies TPP-29537 and TPP-14890 were dissolved in PBS to 10 mg/mL, respectively, and the pH was adjusted to 9 using 1 M carbonate buffer. The chelator and antibody were mixed and incubated at room temperature for two hours in a thermomixer with shaking at 350 rpm. The molar ratio of chelator to antibody was selected depending on the desired chelator to antibody ratio (CAR). The product was purified by FPLC (column: HiLoad 16/600 Superdex 200 pg column; flow rate: 1 mL/min; in 0.1 M acetate buffer (pH 5); detection: UV 280/254 nm). The concentration and monomer purity were determined by SEC-UV (as explained below and shown in Table 12). CAR was determined by using positive electrospray ionization (as explained below) SEC-MS and the following equation: CAR = [(A*An)/A]0, A = intensity of all signals with maximum entropy, An = signal with maximum entropy containing chelator. The determination of CAR for different ACCs is shown in Table 12.Table12: Chelator to Antibody Ratio (CAR), Concentration and Monomer Purity of Different Antibody Chelator ConjugatesAnti-GPC3 mAbChelating agentsConjugateChelator to Antibody Ratio(CAR)ACCconcentration(mg/ml)ACCmonomer purity(%) TPP-14890 Macropa TPP-14890-Macropa 0.7 2.3 99.9 10 1.97 99.9 TPP-29537 Macropa TPP-29537-Macropa 0.5 5.0 99.9 11 1.71 99.2

藉由粒徑篩析層析及質譜之ACC表徵藉由SEC-MS (連結至Waters XEVO TOF之Water Acquity HPLC；運行緩衝液：50/50/0.1 (v/v/v) ACN/水/TFA；流速：0.06 mL/min，用於在線去鹽之管柱：Waters Acquity BEH SEC, 1.7 µm, 2.1 x 150 mm；ESI+模式)藉由使用組分mAb、mAb + 1螯合劑、mAb + 2螯合劑、mAb + 3螯合劑等之呈主要峰高度之百分比形式之MS峰高度來測定ACC之CAR。且使用式CAR = 總(n*An)/總An，其中n等於螯合劑之數量且An等於與n個螯合劑結合之抗體之強度。藉由SEC-UV (Agilent 1260 Infinity HPLC系統，運行緩衝液：10% DMSO/PBS；流速：0.3 mL/min，管柱：Waters Acquity BEH SEC, 1.7 µm, 4.6 x 300 mm，檢測：280 nm下之UV)測定ACC之純度及濃度。該方法自ACC單體分離較高聚集體、二聚體及片段。Characterization ofACCby size-selective chromatography and mass spectrometry The CAR of ACC was determined by SEC-MS (Water Acquity HPLC coupled to Waters XEVO TOF; running buffer: 50/50/0.1 (v/v/v) ACN/water/TFA; flow rate: 0.06 mL/min, column for online desalting: Waters Acquity BEH SEC, 1.7 µm, 2.1 x 150 mm; ESI+ mode) using the MS peak heights of the component mAb, mAb + 1 chelator, mAb + 2 chelators, mAb + 3 chelators, etc. as a percentage of the main peak height. The formula CAR = total (n*An)/total An was used, where n equals the number of chelators and An equals the strength of the antibody bound to n chelators. The purity and concentration of ACC were determined by SEC-UV (Agilent 1260 Infinity HPLC system, running buffer: 10% DMSO/PBS; flow rate: 0.3 mL/min, column: Waters Acquity BEH SEC, 1.7 µm, 4.6 x 300 mm, detection: UV at 280 nm). This method separates higher aggregates, dimers and fragments from ACC monomers.

靶向錒-225結合物(TAC)之放射性標記以2 MBq/mg之比活性及1-2.5 kBq/μL之放射性濃度將錒-225 (Ac-225)添加至於0.1 M乙酸酯緩衝液(pH 5)中之TPP-14890-Macropa及TPP-29537-Macropa中。在室溫下將混合物培育一小時，隨後藉由iTLC測定放射化學純度(RCP)。Radiolabeling ofTargeted Ac-225Conjugate(TAC) Ac-225 was added to TPP-14890-Macropa and TPP-29537-Macropa in 0.1 M acetate buffer (pH 5) at a specific activity of 2 MBq/mg and a radioactivity concentration of 1-2.5 kBq/μL. The mixture was incubated at room temperature for one hour and then the radiochemical purity (RCP) was determined by iTLC.

即時薄層層析(iTLC)：實施iTLC以量測Ac-225-Macropa-GPC3結合溶液之放射化學純度(RCP)。簡言之，切割iTLC.SG.板以獲得11 cm x 1 cm條帶。標誌施加點(1 cm)、切割點(4 cm)及運行終點(10 cm)且在iTLC條帶上施加2 µl之TAC。將條帶在於LSC小瓶中之3 mL移動相中展開。游離Ac-225隨著溶劑在條帶上遷移而複合Ac-225保持在施加點。在液體前端達到標誌之終點時，將條帶自小瓶移除且置於鋁箔上以乾燥。條帶在4 cm處被切割成兩部分：較短「施加」部分(A)及較長「前端」部分(F)。將每一部分小心摺疊並置於閃爍小瓶之底部。6小時後，錒-225與子體達成長期平衡。在高純度鍺檢測器(Ortec)中施加部分計數60 sec且前部分計數120 sec。針對表12中闡述之所有試樣，試樣之放射化學純度(% RCP) (定義為[(施加部分之放射性)/(條帶上之總放射性)] × 100)皆為99.9%。Instant Thin Layer Chromatography(iTLC): iTLC was performed to measure the radiochemical purity (RCP) of the Ac-225-Macropa-GPC3 conjugate solution. Briefly, an iTLC.SG. plate was cut to obtain an 11 cm x 1 cm strip. The application point (1 cm), cut point (4 cm) and end point of the run (10 cm) were marked and 2 µl of TAC was applied on the iTLC strip. The strip was developed in 3 mL of mobile phase in an LSC vial. Free Ac-225 migrated on the strip with the solvent while complexed Ac-225 remained at the application point. When the liquid front reached the marked end point, the strip was removed from the vial and placed on aluminum foil to dry. The strip was cut at 4 cm into two parts: a shorter "applied" part (A) and a longer "front" part (F). Each part was carefully folded and placed at the bottom of a scintillation vial. After 6 hours, the retinol-225 reached a long-term equilibrium with the daughter. The applied part was counted for 60 sec and the front part for 120 sec in a high purity germanium detector (Ortec). The radiochemical purity (% RCP) of the samples (defined as [(radioactivity in the applied part)/(total radioactivity on the strip)] × 100) was 99.9% for all samples described in Table 12.

實例11：免疫反應性分數分析(IRF)中之GPC3-TAC之對比藉由免疫反應性分數(IRF)分析來測定每一經放射性標記之試樣對GPC3塗佈之M-270-羧基Dynabeads (Thermo Fisher Scientific)之結合。IRF係藉由量測結合至磁抗原塗佈之珠粒之經放射性標記之抗體之級分，與仍在上清液中之未結合級分比較來評價的。在放射性標記之後直接評價結合。Example11 :Comparison ofGPC3-TACinthe Immunoreactivity Fraction Analysis(IRF) The binding of each radiolabeled sample to GPC3-coated M-270-carboxyl Dynabeads (Thermo Fisher Scientific) was determined by immunoreactivity fraction (IRF) analysis. IRF is evaluated by measuring the fraction of radiolabeled antibody bound to the magnetic antigen-coated beads compared to the unbound fraction remaining in the supernatant. Binding was evaluated directly after radiolabeling.

簡言之，將自400至0.1 µg之降低量之磁珠粒置於2 mL埃彭道夫管(Eppendorf tube)中且再懸浮於50 µL緩衝液(PBS/ 3% BSA)中。將經放射性標記之化合物於緩衝液中稀釋至5 Bq/µL且添加至每一含珠粒管中(總計50Bq)。培育持續60-90 min，同時在室溫下在熱反應器(Eppendorf)中以750 rpm振盪該等管。在結合步驟完成之後，將40 µL之緩衝液添加至每一管且使用磁體(DynaMag-2)以自上清液分離珠粒。為使容許子體核素處於長期平衡，在至少6小時之後使用高純度鍺檢測器(Ortec)測定每一試樣之上清液中及珠粒上之放射性。總結合定義為[(珠粒中之活性-上清液中之活性)/(總活性)]x100。自阻斷試樣之相同計算獲得非特異性結合，且特異性結合定義為[總結合-非特異性結合]。結果展示於圖9及表13中。表13：Ac-225-Macropa-TPP-29537及Ac-225-Macropa-TPP-14890之CAR、EC50 (µg珠粒)及MAX % IRF。CAREC50[µg珠粒]MAX % IRFAc-225-Macropa-TPP-295370.517.26951115.0294Ac-225-Macropa-TPP-148900.718.97961035.1295Briefly, decreasing amounts of magnetic beads from 400 to 0.1 µg were placed in 2 mL Eppendorf tubes and resuspended in 50 µL buffer (PBS/ 3% BSA). Radiolabeled compounds were diluted to 5 Bq/µL in buffer and added to each tube containing beads (50 Bq total). Incubation lasted 60-90 min while shaking the tubes at 750 rpm at room temperature in a thermoreactor (Eppendorf). After the binding step was complete, 40 µL of buffer was added to each tube and a magnet (DynaMag-2) was used to separate the beads from the supernatant. To allow for long-term equilibrium of daughter nuclides, the radioactivity in the supernatant and on the beads of each sample was determined after at least 6 hours using a high purity germanium detector (Ortec). Total binding was defined as [(activity in beads - activity in supernatant) / (total activity)] x 100. Nonspecific binding was obtained from the same calculations of the blocked samples, and specific binding was defined as [total binding - nonspecific binding]. The results are shown in Figure 9 and Table 13.Table13: CAR, EC50 (µg beads) and MAX % IRF of Ac-225-Macropa-TPP-29537 and Ac-225-Macropa-TPP-14890.CAREC50 [µg beads]MAX % IRF Ac-225-Macropa-TPP-29537 0.5 17.26 95 11 15.02 94 Ac-225-Macropa-TPP-14890 0.7 18.97 96 10 35.12 95

針對Ac-225-Macropa-TPP-29537，針對低及高CAR結合物二者測得相當之EC50及max % IRF，而針對包括先前技術抗GPC3抗體TPP-14890之高CAR Ac-225-Macropa-TPP-14890結合物，與預低CAR型式比較其EC50值明顯降低，此指示其對人類GPC3肽之結合功效受損。For Ac-225-Macropa-TPP-29537, comparable EC50 and max % IRF were measured for both low and high CAR conjugates, whereas for the high CAR Ac-225-Macropa-TPP-14890 conjugate including the prior art anti-GPC3 antibody TPP-14890, its EC50 value was significantly reduced compared to the low CAR version, indicating that its binding efficacy to the human GPC3 peptide was impaired.

實例12：活體外細胞毒性在表現不同含量之GPC3之人類肝細胞癌(HCC)細胞系中測定Ac-225-Macropa-TPP-14890 (TPP-14890-TAC) CAR 0.7、Ac-225-macropa-TPP-14890 CAR 10、Ac-225-Macropa-TPP-29537 (TPP-29537-TAC) CAR 0.5及Ac-225-Macropa-TPP-29537 CAR 11之活體外細胞毒性。將HepG2及Hep3B2.1-7細胞以適當細胞密度接種至細胞介質MEM + 10% FCS中。HepG2細胞比Hep3B2.1-7細胞在細胞表面具有較高GPC3表現。在接種之一天後，使用經放射性標記之同型對照平行滴定2 MBq/mg之比活性及5 kBq/ml之放射性濃度之化合物。將細胞培育6天且隨後使用Hoechst及PI染色測定細胞生存力並在Operetta CLS高含量分析系統(Perkin Elmer)上分析其。所得IC₅₀值展示於表14、圖10A及圖10B中。表15展示不同TAC之同型/靶向比率(較高同型/靶向比率指示較高比細胞毒性)。表14：具有低及高CAR之TPP-14890-TAC及TPP-29537-TAC之IC₅₀值細胞系IC₅₀值(kBq/mL)TPP-14890-TAC CAR 0.7TPP-29537-TAC CAR 0.5TPP-14890-TAC CAR 10TPP-29537-TAC CAR 11同型CAR 0.7HepG20.00650.00410.01580.00681.003Hep3B2.1-70.01780.01100.06730.01570.2998表15：具有低及高CAR之TPP-14890-TAC及TPP-29537-TAC之IC₅₀比率(同型/靶向)細胞系IC₅₀比率(IC₅₀同型/ IC₅₀靶向)TPP-14890-TAC CAR 0.7TPP-29537-TACCAR 0.5TPP-14890-TAC CAR 10TPP-29537-TACCAR 11HepG215925266151Hep3B2.1-71727419Example12:In vitro cytotoxicity The in vitro cytotoxicity of Ac-225-Macropa-TPP-14890 (TPP-14890-TAC) CAR 0.7, Ac-225-macropa-TPP-14890 CAR 10, Ac-225-Macropa-TPP-29537 (TPP-29537-TAC) CAR 0.5 and Ac-225-Macropa-TPP-29537 CAR 11 was determined in human hepatocellular carcinoma (HCC) cell lines expressing different levels of GPC3. HepG2 and Hep3B2.1-7 cells were seeded at appropriate cell densities in the cell medium MEM + 10% FCS. HepG2 cells have higher GPC3 expression on the cell surface than Hep3B2.1-7 cells. One day after inoculation, compounds with a specific activity of 2 MBq/mg and a radioactivity concentration of 5 kBq/ml were titrated in parallel using a radiolabeled isotype control. The cells were cultured for 6 days and then cell viability was determined using Hoechst and PI staining and analyzed on the Operetta CLS High Content Analysis System (Perkin Elmer). The resulting IC₅₀ values are shown in Table 14, Figure 10A and Figure 10B. Table 15 shows the isotype/targeting ratios of different TACs (higher isotype/targeting ratios indicate higher specific cytotoxicity).Table14: IC₅₀ values of TPP-14890-TAC and TPP-29537-TAC with low and high CARCell linesIC₅₀value(kBq/mL)TPP-14890-TAC CAR 0.7TPP-29537-TAC CAR 0.5TPP-14890-TAC CAR 10TPP-29537-TAC CAR 11IsotypeCAR 0.7HepG2 0.0065 0.0041 0.0158 0.0068 1.003Hep3B2.1-7 0.0178 0.0110 0.0673 0.0157 0.2998Table15: IC₅₀ ratios of TPP-14890-TAC and TPP-29537-TAC with low and high CAR (isotype/targeting)Cell linesIC₅₀ratio(IC₅₀isotype/ IC₅₀target)TPP-14890-TAC CAR 0.7TPP-29537-TACCAR 0.5TPP-14890-TAC CAR 10TPP-29537-TACCAR 11HepG2 159 252 66 151Hep3B2.1-7 17 27 4 19

在HepG2及Hep3B2.1-7細胞系中觀察到較強比細胞毒性。針對展示最高IC₅₀比率之TPP-29537-TAC CAR 0.5測得最高比細胞毒性。高CAR TPP-29537結合物顯示與低CAR TPP-14890-TAC類似之IC₅₀。TPP-14890-TAC CAR 10展示顯著較低比細胞毒性，尤其在Hep3B2.1-7細胞系中，此指示與其他測試化合物比較具有較低有效細胞毒性，並指示因標記先前技術抗體TPP-14890的CDR內之全部4個離胺酸殘基中之一或多者導致之高CAR TPP-14890-TAC之結合受損。Strong specific cytotoxicity was observed in HepG2 and Hep3B2.1-7 cell lines. The highest specific cytotoxicity was measured for TPP-29537-TAC CAR 0.5 which exhibited the highest_IC50 ratio. The high CAR TPP-29537 conjugate showed an_IC50 similar to the low CAR TPP-14890-TAC. TPP-14890-TAC CAR 10 exhibited significantly lower specific cytotoxicity, especially in the Hep3B2.1-7 cell line, indicating lower effective cytotoxicity compared to the other tested compounds and indicating that binding of the high CAR TPP-14890-TAC was impaired due to labeling of one or more of the four lysine residues within the CDRs of the prior art antibody TPP-14890.

實例13：GPC3-TAC之生物分佈研究在低GPC3表現人類肝細胞異種移植物模型Huh-7 (ABC: 22.000)中探究Ac-225-Macropa-TPP-29537 (CAR 0.7)及Ac-225-Macropa-同型-對照(CAR 0.7)之生物分佈。使用2 x 10⁶Huh-7細胞於右側腹接種雌性RJ:NMRI-Fox1nu/nu小鼠。在熱投藥之前24小時使用200ug/小鼠非特異性IgG2a抗體預治療動物。分別使用500kBq/kg Ac-225-macropa-TPP-29537及Ac-225-Macropa-同型-對照以0.75 mg/kg之總蛋白質劑量治療動物。針對Ac-225-Macropa-TPP-29537，在治療後72、168、336及504小時處死動物且在各別時間點分離腫瘤及器官。針對Ac-225-Macropa-同型-對照，排除最後時間點(504小時)，此乃因動物達到人道終點。使用鍺檢測器HPGe (GEM-F8250-LB-C, Ortec)測定血液、腫瘤、肝、腎、脾及股骨中累積之Ac-225。結果呈現於圖11A及圖11B中。Example13:Biodistribution study ofGPC3-TAC The biodistribution of Ac-225-Macropa-TPP-29537 (CAR 0.7) and Ac-225-Macropa-isotype-control (CAR 0.7) was investigated in the low GPC3 expressing human hepatocyte xenograft model Huh-7 (ABC: 22.000). Female RJ:NMRI-Fox1nu/nu mice were inoculated in the right flank with 2 x 10⁶ Huh-7 cells. Animals were pretreated with 200ug/mouse nonspecific IgG2a antibody 24 hours before hot dosing. Animals were treated with 500 kBq/kg Ac-225-macropa-TPP-29537 and Ac-225-Macropa-isotype-control at a total protein dose of 0.75 mg/kg. For Ac-225-Macropa-TPP-29537, animals were sacrificed at 72, 168, 336, and 504 hours after treatment and tumors and organs were isolated at the respective time points. For Ac-225-Macropa-isotype-control, the last time point (504 hours) was excluded because the animal reached a humane endpoint. Accumulated Ac-225 in blood, tumors, liver, kidney, spleen, and femur was measured using a germanium detector HPGe (GEM-F8250-LB-C, Ortec). The results are presented in FIG. 11A and FIG. 11B .

針對Ac-225-Macropa-TPP-29537治療之動物，數據指示特異性、強及永久性腫瘤累積而正常器官中之累積通常較低且係瞬時的，且隨時間清除。For Ac-225-Macropa-TPP-29537 treated animals, data indicated specific, robust, and permanent tumor accumulations while accumulations in normal organs were generally low and transient and cleared over time.

實例14：比較TPP-29537-TAC及同型-TAC之活體內效能研究在人類肝細胞瘤模型Huh-7中評估Ac-225-Macropa-TPP-29537之活體內效能。使用500kBq/kg之單一劑量經靜脈內以0.75mg/kg Ac-225-Macropa-TPP-29537 (CAR 0.7)及Ac-225-Macropa-同型-對照(CAR 0.7)之總抗體劑量來治療使用低GPC3表現人類肝細胞瘤模型Huh-7異種移植之小鼠。在熱投藥之前24 h使用200ug/小鼠非特異性IgG2a抗體預治療動物。初始投藥後第25天剩餘之小鼠接收250kBq/kg之重複劑量及0.75 mg/kg總蛋白質劑量。Example14:In vivo efficacy study comparingTPP-29537-TACand isotype-TAC The in vivo efficacy of Ac-225-Macropa-TPP-29537 was evaluated in the human hepatoma model Huh-7. Mice xenografted with the low GPC3 expressing human hepatoma model Huh-7 were treated with a single dose of 500 kBq/kg intravenously with a total antibody dose of 0.75 mg/kg Ac-225-Macropa-TPP-29537 (CAR 0.7) and Ac-225-Macropa-isotype-control (CAR 0.7). Animals were pretreated with 200 ug/mouse nonspecific IgG2a antibody 24 h before hot dosing. The remaining mice received repeated doses of 250 kBq/kg and 0.75 mg/kg total protein on day 25 after the initial dosing.

如圖12中所呈現，與Ac-225-Macropa-同型-對照比較，Ac-225-Macropa-TPP-29537具有抗腫瘤活性且顯著抑制Huh-7腫瘤生長(圖12A)。與媒劑及Ac-225-Macropa-同型-對照小鼠比較，Ac-225-Macropa-TPP-29537小鼠亦具有顯著較佳存活(圖12B)。在治療群組中未觀察到體重損失。As presented in Figure 12, Ac-225-Macropa-TPP-29537 had anti-tumor activity and significantly inhibited Huh-7 tumor growth compared to Ac-225-Macropa-isotype-control (Figure 12A). Ac-225-Macropa-TPP-29537 mice also had significantly better survival compared to vehicle and Ac-225-Macropa-isotype-control mice (Figure 12B). No weight loss was observed in the treatment groups.

實例15：用於成像之Zr-89-DFO*-TPP-29537之製備及表徵為實施活體內正電子發射斷層攝影術(PET)成像，將抗GPC3抗體TPP-29537結合至DFO*且使用鋯-89放射性標記。Example15 :Preparation and Characterization ofZr-89-DFO*-TPP-29537 forImaging To perform in vivo positron emission tomography (PET) imaging, the anti-GPC3 antibody TPP-29537 was conjugated to DFO* and radiolabeled with zirconium-89.

DFO*-TPP-29537之結合及純化： 將5,11,16,22-四氮雜二十六烷二醯胺,N1-[5-(乙醯基羥基胺基)戊基]-N26,5,16-三羥基-N26-[5-[[[(4-異硫基氰基苯基)胺基]硫代甲基]胺基]戊基]-4,12,15,23-四側氧基-(p-Phe-NCS-DFO* (DFO*) (ABX advanced chemical compounds, 7272)溶於DMSO中至10 mg/ml。將抗GPC3抗體TPP-29537溶於WFI中至10 mg/mL。使用1M碳酸鹽緩衝液將抗體之pH調節至9。將p-Phe-NCS-DFO*添加至抗體且在37℃下將混合物在熱混合器中振盪培育1小時。根據製造商說明書(Cytiva, 17085101)藉由PD-10管柱純化結合物。藉由SEC-UV量測DFO*-TPP-29537結合物之單體純度。藉由使用陽性電噴霧離子化之SEC-MS及下列方程式測定CAR：CAR=[(A*An)/A]0，A=最大包絡熵之所有信號之強度，An=含有螯合劑之最大包絡熵之信號。單體純度及CAR分別測定為99.2%及1.2。Conjugation and purification of DFO*-TPP-29537: 5,11,16,22-tetraazahexacosanediamide, N1-[5-(acetylhydroxyamino)pentyl]-N26,5,16-trihydroxy-N26-[5-[[[(4-isothiocyanophenyl)amino]thiomethyl]amino]pentyl]-4,12,15,23-tetraoxy-(p-Phe-NCS-DFO* (DFO*) (ABX advanced chemical compounds, 7272) was dissolved in DMSO to 10 mg/ml. Anti-GPC3 antibody TPP-29537 was dissolved in WFI to 10 mg/mL. The pH of the antibody was adjusted to 9 using 1 M carbonate buffer. p-Phe-NCS-DFO* was added to the antibody and the mixture was incubated at 37°C for 1 hour with shaking in a thermomixer. The conjugate was purified by PD-10 column according to the manufacturer's instructions (Cytiva, 17085101). The monomer purity of the DFO*-TPP-29537 conjugate was measured by SEC-UV. CAR was determined by SEC-MS using positive electrospray ionization and the following equation: CAR=[(A*An)/A]0, A=intensity of all signals of maximum envelopment entropy, An=signal of maximum envelopment entropy containing chelating agent. Monomer purity and CAR were determined to be 99.2% and 1.2, respectively.

使用鋯-89放射性標記DFO*-TPP-29537將於1M草酸中之鋯-89添加至於1 M HEPES中之DFO*-TPP-29537 (pH 7.3)，且以34 MBq/mg之比活性及103 MBq/mL之RAC標記。在37℃下培育30分鐘之後，根據製造商說明書(Cytiva, 17085101)藉由PD-10管柱純化試樣且將緩衝液交換為形成緩衝液(於注射用水中之10 mM L-組胺酸、130 mM甘胺酸、5% (m/v)蔗糖)並稀釋至24 MBq/mL之RAC。Radiolabeling ofDFO*-TPP-29537with Zirconium-89 Zirconium-89 in 1 M oxalic acid was added to DFO*-TPP-29537 in 1 M HEPES (pH 7.3) and labeled with a specific activity of 34 MBq/mg and 103 MBq/mL of RAC. After incubation at 37°C for 30 min, the sample was purified by PD-10 column according to the manufacturer's instructions (Cytiva, 17085101) and the buffer was exchanged to form buffer (10 mM L-histidine, 130 mM glycine, 5% (m/v) sucrose in water for injection) and diluted to 24 MBq/mL of RAC.

免疫反應分數分析使用免疫反應分數(IRF)測定以評估經放射性標記之免疫反應性結合物單株抗體(mAb)產物Zr-89-DFO*-TPP-29537之結合性質。在放射性標記完成之後不久，測定Zr-89-DFO*-TPP-29537對GPC3肽-塗佈之M-270-羧基Dynabead之結合。簡而言之，將0.36-800 µg珠粒於90 µL緩衝液(PBS/3% BSA)中置於2 mL埃彭道夫管中。將經放射性標記之化合物於緩衝液中稀釋至5 Bq/µL且向每一含珠粒管中添加10 µL。試樣以一式三份運行。結合持續60 min，同時在室溫下在熱反應器(Eppendorf)中以750 rpm振盪該等管。在結合步驟完成之後，使用磁鐵(DynaMag-2)以自上清液部分分離珠粒。藉由使用高純度鍺檢測器(Ortec)測定每一試樣之上清液中及珠粒上之放射活性。總結合定義為[(珠粒中之活性-上清液中之活性) / (總活性)] × 100。圖13展示經放射性標記之Zr-89-DFO*-TPP-29537對結合至珠粒之GPC3肽之結合功效。Immunoreactivity score analysis The immunoreactivity score (IRF) assay was used to evaluate the binding properties of the radiolabeled immunoreactive binder monoclonal antibody (mAb) product Zr-89-DFO*-TPP-29537. Binding of Zr-89-DFO*-TPP-29537 to GPC3 peptide-coated M-270-carboxyl Dynabeads was assayed shortly after radiolabeling was completed. Briefly, 0.36-800 µg of beads were placed in 90 µL of buffer (PBS/3% BSA) in a 2 mL Eppendorf tube. The radiolabeled compound was diluted to 5 Bq/µL in buffer and 10 µL was added to each tube containing beads. Samples were run in triplicate. Binding was continued for 60 min while shaking the tubes at 750 rpm at room temperature in a thermoreactor (Eppendorf). After the binding step was completed, a magnet (DynaMag-2) was used to separate the beads from the supernatant fraction. The radioactivity in the supernatant and on the beads of each sample was determined by using a high purity germanium detector (Ortec). Total binding was defined as [(activity in beads - activity in supernatant) / (total activity)] × 100. FIG. 13 shows the binding efficacy of radiolabeled Zr-89-DFO*-TPP-29537 to GPC3 peptide bound to beads.

使用50-100 µg GPC3-肽塗佈珠粒可達成經選擇以產生用於結合之過量表位之抗原塗覆珠粒之數量。總而言之，在單點分析中測得IRF% ~ 90%。The amount of antigen-coated beads selected to generate excess epitope for binding was achieved using 50-100 µg of GPC3-peptide-coated beads. Overall, IRF% ~ 90% was measured in a single-point assay.

實例16：Ac-225-Macropa-TPP-29537及其成像對Zr-89-DFO*-TPP-29537之生物分佈在HEP-3B HCC腫瘤模型中評估Ac-225-Macropa-TPP-29537 (CAR 0.7)及其成像對Zr-89-DFO*-TPP-29537 (CAR 5.3)之離體生物分佈。另外，實施Zr-89-DFO*-TPP-29537活體內PET成像量化以探究其作為Ac-225-Macropa-TPP-29537之成像對以用於早期研發之潛能。Example16:Biodistribution ofAc-225-Macropa-TPP-29537and its imaging pairZr-89-DFO*-TPP-29537 The ex vivo biodistribution of Ac-225-Macropa-TPP-29537 (CAR 0.7) and its imaging pair Zr-89-DFO*-TPP-29537 (CAR 5.3) was evaluated in the HEP-3B HCC tumor model. In addition, in vivo PET imaging quantification of Zr-89-DFO*-TPP-29537 was performed to explore its potential as an imaging pair of Ac-225-Macropa-TPP-29537 for early development.

使用4.1 x 10⁶HEP-3B細胞於右側腹接種雌性RJ:NMRI-Fox1nu/nu小鼠。在熱投藥之前24小時使用200ug/小鼠非特異性IgG2a抗體預處理動物。Female RJ:NMRI-Fox1nu/nu mice were inoculated with 4.1 x 10⁶ HEP-3B cells in the right flank. Animals were pretreated with 200 ug/mouse nonspecific IgG2a antibody 24 hours before hot administration.

如表16中所闡述，以緩慢濃注形式將Zr-89-DFO*-TPP-29537注射至動物之尾部靜脈。在注射後2 h (n=1)、4 h (n=3)、24 h (n=3)、72 h (n=3)及168 h (n=2)實施PET/CT成像。藉由MILabs Vector6 PET/SPECT/CT掃描儀、使用HE-UHR-RM準直儀、4小鼠床、60分鐘之總獲取時間、螺旋獲取模式及固定視域來實施成像獲取，且在第一次獲取期間進行測定。表16：Zr-89-DFO*-TPP-29537之活體內及離體生物分佈研究以及Ac-225-Macropa-TPP-29537之離體生物分佈中之群組之概述測試物品小鼠之#劑量時間點Zr-89-DFO*-TPP-295373x6200 MBq/mg0.5、2、4、24、72及168 hAc-225-Macropa-TPP-295373x6250 kBq/kg0.5、2、4、24、72及168 hZr-89-DFO*-TPP-29537 was injected into the tail vein of the animals as a slow bolus as described in Table 16. PET/CT imaging was performed at 2 h (n=1), 4 h (n=3), 24 h (n=3), 72 h (n=3), and 168 h (n=2) after injection. Imaging acquisition was performed by a MILabs Vector6 PET/SPECT/CT scanner using a HE-UHR-RM collimator, a 4-mouse bed, a total acquisition time of 60 minutes, a spiral acquisition mode, and a fixed field of view, and measurements were performed during the first acquisition.Table16: Summary of the groups in the in vivo and in vitro biodistribution studies of Zr-89-DFO*-TPP-29537 and the in vitro biodistribution of Ac-225-Macropa-TPP-29537Test ArticlesMouse#DosageTime point Zr-89-DFO*-TPP-29537 3x6 200 MBq/mg 0.5, 2, 4, 24, 72 and 168 h Ac-225-Macropa-TPP-29537 3x6 250 kBq/kg 0.5, 2, 4, 24, 72 and 168 h

分開投藥以避免掃描儀中同時有一個以上小鼠。Dosing was separated to avoid more than one mouse being in the scanner at the same time.

為評價活體內生物分佈，使用PMOD軟體實施影像分析。將所關注CT體積(球體VOI)應用於至心臟、肝及腎。在CT上勾畫腫瘤輪廓，及對PET影像應用限制性VOI (10%臨限值)。將數據衰變校正至注射時間點。圖14展示在不同時間點之使用Zr-89-DFO*-TPP-29537之經成像之具有HEP-3B腫瘤的小鼠的冠狀PET/CT MIP。To evaluate in vivo biodistribution, image analysis was performed using PMOD software. CT volumes of interest (spherical VOIs) were applied to the heart, liver, and kidneys. Tumor contours were outlined on CT, and restrictive VOIs (10% threshold) were applied to PET images. Data decay was corrected to the injection time point. Figure 14 shows coronal PET/CT MIPs of mice with HEP-3B tumors imaged with Zr-89-DFO*-TPP-29537 at different time points.

在活體內成像之後，為評價Zr-89-DFO*-TPP-29537之離體生物分佈，將成像小鼠(每個時間點3個小鼠)處死並收集血液、腫瘤、肝、腎、脾及股骨及脛骨，且使用Perkin Elmer Wizard II γ計數器測定Zr-89活性。After in vivo imaging, to evaluate the ex vivo biodistribution of Zr-89-DFO*-TPP-29537, imaged mice (3 mice per time point) were sacrificed and blood, tumors, liver, kidney, spleen, and femur and tibia were collected, and Zr-89 activity was measured using a Perkin Elmer Wizard II gamma counter.

平行地，使用250kBq/kg Ac-225-Macropa-TPP-29537在0.75 mg/kg之蛋白質劑量下靜脈內注射具有HEP-3B腫瘤之小鼠。在治療後0.5、2、4、24、72及168小時將動物處死。使用鍺檢測器HPGe (GEM-F8250-LB-C, Ortec)測定血液、腫瘤、肝、腎、脾及股骨中累積之Ac-225。In parallel, mice bearing HEP-3B tumors were injected intravenously with 250 kBq/kg Ac-225-Macropa-TPP-29537 at a protein dose of 0.75 mg/kg. Animals were sacrificed at 0.5, 2, 4, 24, 72, and 168 hours after treatment. Accumulated Ac-225 in blood, tumor, liver, kidney, spleen, and femur was measured using a germanium detector HPGe (GEM-F8250-LB-C, Ortec).

如圖15中所繪示，Ac-225-Macropa-TPP-29537及Zr-89-DFO*-TPP-29537展示相當離體生物分佈，在研究中之最後時間點(168 h)具有類似血液清除率。Ac-225-Macropa-TPP-29537及Zr-89-DFO*-TPP-29537之腫瘤累積類似，最多72 h p.i.，此後Ac-225-Macropa-TPP-29537 %ID/g較高，其最可能由Ac-225-Macropa-TPP-29537效能導致之腫瘤收縮引起。As shown in Figure 15, Ac-225-Macropa-TPP-29537 and Zr-89-DFO*-TPP-29537 exhibited comparable ex vivo biodistribution, with similar blood clearance at the final time point in the study (168 h). Tumor accumulation of Ac-225-Macropa-TPP-29537 and Zr-89-DFO*-TPP-29537 was similar up to 72 h p.i., after which Ac-225-Macropa-TPP-29537 %ID/g was higher, which was most likely caused by tumor shrinkage due to the efficacy of Ac-225-Macropa-TPP-29537.

Zr-89-DFO*-TPP-29537之離體生物分佈及活體內PET成像量化在心臟/血液、腎、肝及腫瘤中顯示相當攝取(圖15)。The in vitro biodistribution and in vivo PET imaging quantification of Zr-89-DFO*-TPP-29537 showed comparable uptake in the heart/blood, kidney, liver, and tumor ( FIG15 ).

總而言之，該等結果證實Zr-89-DFO*-TPP-29537適於用作Ac-225-Macropa-TPP-29537之PET成像對以用於早期研發。序列TPP ID序列名稱序列區域序列類型SEQ IDSEQTPP-5657種系VH3-23-同型對照hIgG1λVHPRT1EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKLTGAEYFQHWGQGTLVTVSSTPP-5657種系VH3-23-同型對照hIgG1λHCDR1PRT2SYAMSTPP-5657種系VH3-23-同型對照hIgG1λHCDR2PRT3AISGSGGSTYYADSVKGTPP-5657種系VH3-23-同型對照hIgG1λHCDR3PRT4LTGAEYFQHTPP-5657種系VH3-23-同型對照hIgG1λVLPRT5QSVLTQPPSVSGAPGQRVTISCTGSSSNIGAGYDVHWYQQLPGTAPKLLIYGNSNRPSGVPDRFSGSKSGTSASLAITGLQAEDEADYYCQSYDSSLSGVVFGGGTKLTVLTPP-5657種系VH3-23-同型對照hIgG1λLCDR1PRT6TGSSSNIGAGYDVHTPP-5657種系VH3-23-同型對照hIgG1λLCDR2PRT7GNSNRPSTPP-5657種系VH3-23-同型對照hIgG1λLCDR3PRT8QSYDSSLSGVVTPP-5657種系VH3-23-同型對照hIgG1λVHDNA9GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCAGCAGCTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGCCATCTCTGGCAGCGGCGGCAGCACCTACTACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCAGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGCGCCAAACTGACAGGCGCCGAGTACTTCCAGCACTGGGGCCAGGGAACACTCGTGACCGTGTCATCATPP-5657種系VH3-23-同型對照hIgG1λHCDR1DNA10AGCTACGCCATGAGCTPP-5657種系VH3-23-同型對照hIgG1λHCDR2DNA11GCCATCTCTGGCAGCGGCGGCAGCACCTACTACGCCGATTCTGTGAAGGGCTPP-5657種系VH3-23-同型對照hIgG1λHCDR3DNA12CTGACAGGCGCCGAGTACTTCCAGCACTPP-5657種系VH3-23-同型對照hIgG1λVLDNA13CAGTCTGTTCTGACACAGCCTCCATCTGTGTCTGGCGCCCCTGGACAGAGAGTGACCATCAGCTGTACAGGCAGCAGCTCCAATATCGGAGCCGGCTATGACGTGCACTGGTATCAGCAGCTGCCTGGCACAGCCCCTAAACTGCTGATCTACGGCAACAGCAACAGACCCAGCGGCGTGCCCGATAGATTTTCCGGCTCTAAGAGCGGCACAAGCGCCAGCCTGGCTATTACTGGACTGCAGGCCGAGGACGAGGCCGACTACTACTGTCAGAGCTACGACAGCAGCCTGAGCGGCGTTGTGTTTGGCGGAGGCACAAAGCTGACAGTGCTATPP-5657種系VH3-23-同型對照hIgG1λLCDR1DNA14ACAGGCAGCAGCTCCAATATCGGAGCCGGCTATGACGTGCACTPP-5657種系VH3-23-同型對照hIgG1λLCDR2DNA15GGCAACAGCAACAGACCCAGCTPP-5657種系VH3-23-同型對照hIgG1λLCDR3DNA16CAGAGCTACGACAGCAGCCTGAGCGGCGTTGTGTPP-5657種系VH3-23-同型對照hIgG1λ重鏈PRT17EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKLTGAEYFQHWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-5657種系VH3-23-同型對照hIgG1λ輕鏈PRT18QSVLTQPPSVSGAPGQRVTISCTGSSSNIGAGYDVHWYQQLPGTAPKLLIYGNSNRPSGVPDRFSGSKSGTSASLAITGLQAEDEADYYCQSYDSSLSGVVFGGGTKLTVLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECSTPP-5657種系VH3-23-同型對照hIgG1λ重鏈DNA19GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCAGCAGCTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGCCATCTCTGGCAGCGGCGGCAGCACCTACTACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCAGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGCGCCAAACTGACAGGCGCCGAGTACTTCCAGCACTGGGGCCAGGGAACACTCGTGACCGTGTCATCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-5657種系VH3-23-同型對照hIgG1λ輕鏈DNA20CAGTCTGTTCTGACACAGCCTCCATCTGTGTCTGGCGCCCCTGGACAGAGAGTGACCATCAGCTGTACAGGCAGCAGCTCCAATATCGGAGCCGGCTATGACGTGCACTGGTATCAGCAGCTGCCTGGCACAGCCCCTAAACTGCTGATCTACGGCAACAGCAACAGACCCAGCGGCGTGCCCGATAGATTTTCCGGCTCTAAGAGCGGCACAAGCGCCAGCCTGGCTATTACTGGACTGCAGGCCGAGGACGAGGCCGACTACTACTGTCAGAGCTACGACAGCAGCCTGAGCGGCGTTGTGTTTGGCGGAGGCACAAAGCTGACAGTGCTAGGCCAGCCTAAAGCCGCCCCTAGCGTGACCCTGTTCCCTCCAAGCAGCGAGGAACTGCAGGCCAACAAGGCCACCCTCGTGTGCCTGATCAGCGACTTCTATCCTGGCGCCGTGACCGTGGCCTGGAAGGCCGATAGCTCTCCTGTGAAGGCCGGCGTGGAAACCACCACCCCTAGCAAGCAGAGCAACAACAAATACGCCGCCAGCAGCTACCTGAGCCTGACCCCCGAGCAGTGGAAGTCCCACAGATCCTACAGCTGCCAAGTGACCCACGAGGGCAGCACCGTGGAAAAGACAGTGGCCCCTACCGAGTGCAGCTPP-14877hYP9.1b-52T-hIgG1κVHPRT21EVQLVESGGGLVQPGGSLRLSCAASGFNFNTNAMNWVRQAPGKGLEWVGRVRNKTNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGTLVTVSSTPP-14877hYP9.1b-52T-hIgG1κHCDR1PRT22TNAMNTPP-14877hYP9.1b-52T-hIgG1κHCDR2PRT23RVRNKTNNYATYYADSVKDTPP-14877hYP9.1b-52T-hIgG1κHCDR3PRT24GNSFAYTPP-14877hYP9.1b-52T-hIgG1κVLPRT25DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKLEIKTPP-14877hYP9.1b-52T-hIgG1κLCDR1PRT26KSSQSLLYSSNQKNYLATPP-14877hYP9.1b-52T-hIgG1κLCDR2PRT27WASTRESTPP-14877hYP9.1b-52T-hIgG1κLCDR3PRT28QQYYNYPLTTPP-14877hYP9.1b-52T-hIgG1κVHDNA29GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTCTCTGAGACTGTCTTGTGCCGCCAGCGGCTTCAACTTCAACACCAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGGAAGAGTGCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTGCCTACTGGGGACAGGGCACCCTGGTTACAGTTAGCTCATPP-14877hYP9.1b-52T-hIgG1κHCDR1DNA30ACCAACGCCATGAACTPP-14877hYP9.1b-52T-hIgG1κHCDR2DNA31AGAGTGCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGACTPP-14877hYP9.1b-52T-hIgG1κHCDR3DNA32GGCAATAGCTTTGCCTACTPP-14877hYP9.1b-52T-hIgG1κVLDNA33GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTAAGCTGCTGATCTACTGGGCCAGCACCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-14877hYP9.1b-52T-hIgG1κLCDR1DNA34AAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTPP-14877hYP9.1b-52T-hIgG1κLCDR2DNA35TGGGCCAGCACCAGAGAAAGCTPP-14877hYP9.1b-52T-hIgG1κLCDR3DNA36CAGCAGTACTACAACTACCCTCTGACCTPP-14877hYP9.1b-52T-hIgG1κ重鏈PRT37EVQLVESGGGLVQPGGSLRLSCAASGFNFNTNAMNWVRQAPGKGLEWVGRVRNKTNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-14877hYP9.1b-52T-hIgG1κ輕鏈PRT38DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-14877hYP9.1b-52T-hIgG1κ重鏈DNA39GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTCTCTGAGACTGTCTTGTGCCGCCAGCGGCTTCAACTTCAACACCAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGGAAGAGTGCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTGCCTACTGGGGACAGGGCACCCTGGTTACAGTTAGCTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-14877hYP9.1b-52T-hIgG1κ輕鏈DNA40GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTAAGCTGCTGATCTACTGGGCCAGCACCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-14890考曲妥珠單抗-hIgG1κVHPRT41QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGDTAYSQKFKGRVTLTADKSTSTAYMELSSLTSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-14890考曲妥珠單抗-hIgG1κHCDR1PRT42DYEMHTPP-14890考曲妥珠單抗-hIgG1κHCDR2PRT43ALDPKTGDTAYSQKFKGTPP-14890考曲妥珠單抗-hIgG1κHCDR3PRT44FYSYTYTPP-14890考曲妥珠單抗-hIgG1κVLPRT45DVVMTQSPLSLPVTPGEPASISCRSSQSLVHSNRNTYLHWYLQKPGQSPQLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPPTFGQGTKLEIKTPP-14890考曲妥珠單抗-hIgG1κLCDR1PRT46RSSQSLVHSNRNTYLHTPP-14890考曲妥珠單抗-hIgG1κLCDR2PRT47KVSNRFSTPP-14890考曲妥珠單抗-hIgG1κLCDR3PRT48SQNTHVPPTTPP-14890考曲妥珠單抗-hIgG1κVHDNA49CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCGATACAGCCTACAGCCAGAAATTCAAGGGCAGAGTGACCCTGACCGCCGACAAGAGCACAAGCACCGCCTACATGGAACTGAGCAGCCTGACCAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-14890考曲妥珠單抗-hIgG1κHCDR1DNA50GACTACGAGATGCACTPP-14890考曲妥珠單抗-hIgG1κHCDR2DNA51GCCCTGGATCCTAAGACCGGCGATACAGCCTACAGCCAGAAATTCAAGGGCTPP-14890考曲妥珠單抗-hIgG1κHCDR3DNA52TTCTACAGCTACACCTACTPP-14890考曲妥珠單抗-hIgG1κVLDNA53GACGTTGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGCACAGCAACCGGAATACCTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAAGGTGTCCAACCGGTTCAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCACCTACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-14890考曲妥珠單抗-hIgG1κLCDR1DNA54AGAAGCAGCCAGAGCCTGGTGCACAGCAACCGGAATACCTACCTGCACTPP-14890考曲妥珠單抗-hIgG1κLCDR2DNA55AAGGTGTCCAACCGGTTCAGCTPP-14890考曲妥珠單抗-hIgG1κLCDR3DNA56AGCCAGAATACCCACGTGCCACCTACCTPP-14890考曲妥珠單抗-hIgG1κ重鏈PRT57QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGDTAYSQKFKGRVTLTADKSTSTAYMELSSLTSEDTAVYYCTRFYSYTYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-14890考曲妥珠單抗-hIgG1κ輕鏈PRT58DVVMTQSPLSLPVTPGEPASISCRSSQSLVHSNRNTYLHWYLQKPGQSPQLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPPTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-14890考曲妥珠單抗-hIgG1κ重鏈DNA59CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCGATACAGCCTACAGCCAGAAATTCAAGGGCAGAGTGACCCTGACCGCCGACAAGAGCACAAGCACCGCCTACATGGAACTGAGCAGCCTGACCAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCAGCCAGCACAAAGGGCCCTAGCGTGTTCCCTCTGGCTCCTAGCAGCAAGTCTACAAGCGGAGGAACAGCCGCTCTGGGCTGCCTGGTCAAGGATTACTTTCCCGAGCCTGTGACCGTGTCCTGGAATTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCTGTGCTGCAAAGCAGCGGCCTGTACTCTCTGAGCAGCGTGGTCACAGTGCCAAGCTCTAGCCTGGGCACCCAGACCTACATCTGCAATGTGAACCACAAGCCTAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCTCCATGTCCTGCTCCAGAACTGCTCGGCGGACCTTCCGTGTTCCTGTTTCCTCCAAAGCCTAAGGACACCCTGATGATCAGCAGAACCCCTGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGATCCCGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACAGAGTGGTGTCCGTGCTGACCGTGCTGCACCAGGATTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCTCCTATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCTAGGGAACCCCAGGTTTACACACTGCCTCCAAGCAGGGACGAGCTGACCAAGAATCAGGTGTCCCTGACCTGCCTCGTGAAGGGCTTCTACCCTTCCGATATCGCCGTGGAATGGGAGAGCAATGGCCAGCCTGAGAACAACTACAAGACAACCCCTCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACAGTGGACAAGTCCAGATGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGTCTCCTGGCAAGTPP-14890考曲妥珠單抗-hIgG1κ輕鏈DNA60GACGTTGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGCACAGCAACCGGAATACCTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAAGGTGTCCAACCGGTTCAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCACCTACCTTTGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-15081YP7-hIgG1κVHPRT61EVQLVETGGGMVQPEGSLKLSCAASGFTFNKNAMNWVRQAPGKGLEWVARIRNKTNNYATYYADSVKARFTISRDDSQSMLYLQMNNLKIEDTAMYYCVAGNSFAYWGQGTLVTVSATPP-15081YP7-hIgG1κHCDR1PRT62KNAMNTPP-15081YP7-hIgG1κHCDR2PRT63RIRNKTNNYATYYADSVKATPP-15081YP7-hIgG1κHCDR3PRT64GNSFAYTPP-15081YP7-hIgG1κVLPRT65DIVMSQSPSSLVVSIGEKVTMTCKSSQSLLYSSNQKNYLAWYQQKPGQSPKLLIYWASSRESGVPDRFTGSGSGTDFTLTISSVKAEDLAVYYCQQYYNYPLTFGAGTKLELKTPP-15081YP7-hIgG1κLCDR1PRT66KSSQSLLYSSNQKNYLATPP-15081YP7-hIgG1κLCDR2PRT67WASSRESTPP-15081YP7-hIgG1κLCDR3PRT68QQYYNYPLTTPP-15081YP7-hIgG1κVHDNA69GAAGTGCAGCTGGTGGAAACAGGCGGCGGAATGGTTCAGCCTGAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACAAGAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGCCCGGATCAGAAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCAGATTCACCATCAGCAGGGACGACAGCCAGAGCATGCTGTACCTGCAGATGAACAACCTGAAGATCGAGGACACCGCCATGTACTACTGCGTGGCCGGCAATAGCTTTGCCTATTGGGGCCAGGGCACCCTGGTCACAGTTTCTGCATPP-15081YP7-hIgG1κHCDR1DNA70AAGAACGCCATGAACTPP-15081YP7-hIgG1κHCDR2DNA71CGGATCAGAAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCTPP-15081YP7-hIgG1κHCDR3DNA72GGCAATAGCTTTGCCTATTPP-15081YP7-hIgG1κVLDNA73GACATCGTGATGTCTCAGAGCCCTAGCAGCCTGGTGGTGTCCATCGGCGAGAAAGTGACCATGACCTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGACAGTCTCCCAAGCTGCTGATCTACTGGGCCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTCACAGGCAGCGGCAGCGGAACCGACTTCACCCTGACAATCAGCAGCGTGAAGGCCGAGGATCTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGAGCCGGCACCAAGCTGGAACTGAAATPP-15081YP7-hIgG1κLCDR1DNA74AAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTPP-15081YP7-hIgG1κLCDR2DNA75TGGGCCAGCAGCAGAGAAAGCTPP-15081YP7-hIgG1κLCDR3DNA76CAGCAGTACTACAACTACCCTCTGACCTPP-15081YP7-hIgG1κ重鏈PRT77EVQLVETGGGMVQPEGSLKLSCAASGFTFNKNAMNWVRQAPGKGLEWVARIRNKTNNYATYYADSVKARFTISRDDSQSMLYLQMNNLKIEDTAMYYCVAGNSFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-15081YP7-hIgG1κ輕鏈PRT78DIVMSQSPSSLVVSIGEKVTMTCKSSQSLLYSSNQKNYLAWYQQKPGQSPKLLIYWASSRESGVPDRFTGSGSGTDFTLTISSVKAEDLAVYYCQQYYNYPLTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-15081YP7-hIgG1κ重鏈DNA79GAAGTGCAGCTGGTGGAAACAGGCGGCGGAATGGTTCAGCCTGAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACAAGAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGCCCGGATCAGAAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCAGATTCACCATCAGCAGGGACGACAGCCAGAGCATGCTGTACCTGCAGATGAACAACCTGAAGATCGAGGACACCGCCATGTACTACTGCGTGGCCGGCAATAGCTTTGCCTATTGGGGCCAGGGCACCCTGGTCACAGTTTCTGCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-15081YP7-hIgG1κ輕鏈DNA80GACATCGTGATGTCTCAGAGCCCTAGCAGCCTGGTGGTGTCCATCGGCGAGAAAGTGACCATGACCTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGACAGTCTCCCAAGCTGCTGATCTACTGGGCCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTCACAGGCAGCGGCAGCGGAACCGACTTCACCCTGACAATCAGCAGCGTGAAGGCCGAGGATCTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGAGCCGGCACCAAGCTGGAACTGAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-2931214890-gl57-gl65_hIgG1κVHPRT81QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-2931214890-gl57-gl65_hIgG1κHCDR1PRT82DYEMHTPP-2931214890-gl57-gl65_hIgG1κHCDR2PRT83ALDPKTGSTSYAQRFQGTPP-2931214890-gl57-gl65_hIgG1κHCDR3PRT84FYSYTYTPP-2931214890-gl57-gl65_hIgG1κVLPRT85DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKLEIKTPP-2931214890-gl57-gl65_hIgG1κLCDR1PRT86RSSQSLLHSNGYNYLHTPP-2931214890-gl57-gl65_hIgG1κLCDR2PRT87RGSNRASTPP-2931214890-gl57-gl65_hIgG1κLCDR3PRT88SQNTHVPYTTPP-2931214890-gl57-gl65_hIgG1κVHDNA89CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-2931214890-gl57-gl65_hIgG1κHCDR1DNA90GACTACGAGATGCACTPP-2931214890-gl57-gl65_hIgG1κHCDR2DNA91GCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCTPP-2931214890-gl57-gl65_hIgG1κHCDR3DNA92TTCTACAGCTACACCTACTPP-2931214890-gl57-gl65_hIgG1κVLDNA93GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-2931214890-gl57-gl65_hIgG1κLCDR1DNA94AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-2931214890-gl57-gl65_hIgG1κLCDR2DNA95AGAGGCAGCAACAGAGCCAGCTPP-2931214890-gl57-gl65_hIgG1κLCDR3DNA96AGCCAGAATACCCACGTGCCATACACCTPP-2931214890-gl57-gl65_hIgG1κ重鏈PRT97QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-2931214890-gl57-gl65_hIgG1κ輕鏈PRT98DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-2931214890-gl57-gl65_hIgG1κ重鏈DNA99CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-2931214890-gl57-gl65_hIgG1κ輕鏈DNA100GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-2953714890-gl53-gl65_hIgG1κVHPRT101QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-2953714890-gl53-gl65_hIgG1κHCDR1PRT102DYEMHTPP-2953714890-gl53-gl65_hIgG1κHCDR2PRT103ALDPSTGSTSYAQRFQGTPP-2953714890-gl53-gl65_hIgG1κHCDR3PRT104FYSYTYTPP-2953714890-gl53-gl65_hIgG1κVLPRT105DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKLEIKTPP-2953714890-gl53-gl65_hIgG1κLCDR1PRT106RSSQSLLHSNGYNYLHTPP-2953714890-gl53-gl65_hIgG1κLCDR2PRT107RGSNRASTPP-2953714890-gl53-gl65_hIgG1κLCDR3PRT108SQNTHVPYTTPP-2953714890-gl53-gl65_hIgG1κVHDNA109CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-2953714890-gl53-gl65_hIgG1κHCDR1DNA110GACTACGAGATGCACTPP-2953714890-gl53-gl65_hIgG1κHCDR2DNA111GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-2953714890-gl53-gl65_hIgG1κHCDR3DNA112TTCTACAGCTACACCTACTPP-2953714890-gl53-gl65_hIgG1κVLDNA113GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-2953714890-gl53-gl65_hIgG1κLCDR1DNA114AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-2953714890-gl53-gl65_hIgG1κLCDR2DNA115AGAGGCAGCAACAGAGCCAGCTPP-2953714890-gl53-gl65_hIgG1κLCDR3DNA116AGCCAGAATACCCACGTGCCATACACCTPP-2953714890-gl53-gl65_hIgG1κ重鏈PRT117QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-2953714890-gl53-gl65_hIgG1κ輕鏈PRT118DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-2953714890-gl53-gl65_hIgG1κ重鏈DNA119CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-2953714890-gl53-gl65_hIgG1κ輕鏈DNA120GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-2953814890-gl53-gl73_hIgG1κVHPRT121QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-2953814890-gl53-gl73_hIgG1κHCDR1PRT122DYEMHTPP-2953814890-gl53-gl73_hIgG1κHCDR2PRT123ALDPSTGSTSYAQRFQGTPP-2953814890-gl53-gl73_hIgG1κHCDR3PRT124FYSYTYTPP-2953814890-gl53-gl73_hIgG1κVLPRT125DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHTPYTFGQGTKLEIKTPP-2953814890-gl53-gl73_hIgG1κLCDR1PRT126RSSQSLLHSNGYNYLHTPP-2953814890-gl53-gl73_hIgG1κLCDR2PRT127RGSNRASTPP-2953814890-gl53-gl73_hIgG1κLCDR3PRT128SQNTHTPYTTPP-2953814890-gl53-gl73_hIgG1κVHDNA129CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-2953814890-gl53-gl73_hIgG1κHCDR1DNA130GACTACGAGATGCACTPP-2953814890-gl53-gl73_hIgG1κHCDR2DNA131GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-2953814890-gl53-gl73_hIgG1κHCDR3DNA132TTCTACAGCTACACCTACTPP-2953814890-gl53-gl73_hIgG1κVLDNA133GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAACACCCACACACCTTACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-2953814890-gl53-gl73_hIgG1κLCDR1DNA134AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-2953814890-gl53-gl73_hIgG1κLCDR2DNA135AGAGGCAGCAACAGAGCCAGCTPP-2953814890-gl53-gl73_hIgG1κLCDR3DNA136AGCCAGAACACCCACACACCTTACACCTPP-2953814890-gl53-gl73_hIgG1κ重鏈PRT137QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-2953814890-gl53-gl73_hIgG1κ輕鏈PRT138DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHTPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-2953814890-gl53-gl73_hIgG1κ重鏈DNA139CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-2953814890-gl53-gl73_hIgG1κ輕鏈DNA140GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAACACCCACACACCTTACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-2963614890-gl53-gl93_hIgG1κVHPRT141QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-2963614890-gl53-gl93_hIgG1κHCDR1PRT142DYEMHTPP-2963614890-gl53-gl93_hIgG1κHCDR2PRT143ALDPSTGSTSYAQRFQGTPP-2963614890-gl53-gl93_hIgG1κHCDR3PRT144FYSYTYTPP-2963614890-gl53-gl93_hIgG1κVLPRT145DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKTPP-2963614890-gl53-gl93_hIgG1κLCDR1PRT146RSSQSLLHSNGYNYLHTPP-2963614890-gl53-gl93_hIgG1κLCDR2PRT147RGSNRASTPP-2963614890-gl53-gl93_hIgG1κLCDR3PRT148IQNTHTPYTTPP-2963614890-gl53-gl93_hIgG1κVHDNA149CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-2963614890-gl53-gl93_hIgG1κHCDR1DNA150GACTACGAGATGCACTPP-2963614890-gl53-gl93_hIgG1κHCDR2DNA151GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-2963614890-gl53-gl93_hIgG1κHCDR3DNA152TTCTACAGCTACACCTACTPP-2963614890-gl53-gl93_hIgG1κVLDNA153GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-2963614890-gl53-gl93_hIgG1κLCDR1DNA154AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-2963614890-gl53-gl93_hIgG1κLCDR2DNA155AGAGGCAGCAACAGAGCCAGCTPP-2963614890-gl53-gl93_hIgG1κLCDR3DNA156ATCCAGAACACCCACACACCTTACACCTPP-2963614890-gl53-gl93_hIgG1κ重鏈PRT157QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-2963614890-gl53-gl93_hIgG1κ輕鏈PRT158DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-2963614890-gl53-gl93_hIgG1κ重鏈DNA159CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-2963614890-gl53-gl93_hIgG1κ輕鏈DNA160GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-2963814890-gl53-gl95_hIgG1κVHPRT161QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-2963814890-gl53-gl95_hIgG1κHCDR1PRT162DYEMHTPP-2963814890-gl53-gl95_hIgG1κHCDR2PRT163ALDPSTGSTSYAQRFQGTPP-2963814890-gl53-gl95_hIgG1κHCDR3PRT164FYSYTYTPP-2963814890-gl53-gl95_hIgG1κVLPRT165DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKTPP-2963814890-gl53-gl95_hIgG1κLCDR1PRT166RSSQSLLHSNGYNYLHTPP-2963814890-gl53-gl95_hIgG1κLCDR2PRT167RGSNRLSTPP-2963814890-gl53-gl95_hIgG1κLCDR3PRT168IQNTHTPYTTPP-2963814890-gl53-gl95_hIgG1κVHDNA169CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-2963814890-gl53-gl95_hIgG1κHCDR1DNA170GACTACGAGATGCACTPP-2963814890-gl53-gl95_hIgG1κHCDR2DNA171GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-2963814890-gl53-gl95_hIgG1κHCDR3DNA172TTCTACAGCTACACCTACTPP-2963814890-gl53-gl95_hIgG1κVLDNA173GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-2963814890-gl53-gl95_hIgG1κLCDR1DNA174AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-2963814890-gl53-gl95_hIgG1κLCDR2DNA175AGAGGCAGCAACAGACTGAGCTPP-2963814890-gl53-gl95_hIgG1κLCDR3DNA176ATCCAGAACACCCACACACCTTACACCTPP-2963814890-gl53-gl95_hIgG1κ重鏈PRT177QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-2963814890-gl53-gl95_hIgG1κ輕鏈PRT178DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-2963814890-gl53-gl95_hIgG1κ重鏈DNA179CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-2963814890-gl53-gl95_hIgG1κ輕鏈DNA180GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-2964214890-gl57-gl95_hIgG1κVHPRT181QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-2964214890-gl57-gl95_hIgG1κHCDR1PRT182DYEMHTPP-2964214890-gl57-gl95_hIgG1κHCDR2PRT183ALDPKTGSTSYAQRFQGTPP-2964214890-gl57-gl95_hIgG1κHCDR3PRT184FYSYTYTPP-2964214890-gl57-gl95_hIgG1κVLPRT185DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKTPP-2964214890-gl57-gl95_hIgG1κLCDR1PRT186RSSQSLLHSNGYNYLHTPP-2964214890-gl57-gl95_hIgG1κLCDR2PRT187RGSNRLSTPP-2964214890-gl57-gl95_hIgG1κLCDR3PRT188IQNTHTPYTTPP-2964214890-gl57-gl95_hIgG1κVHDNA189CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-2964214890-gl57-gl95_hIgG1κHCDR1DNA190GACTACGAGATGCACTPP-2964214890-gl57-gl95_hIgG1κHCDR2DNA191GCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCTPP-2964214890-gl57-gl95_hIgG1κHCDR3DNA192TTCTACAGCTACACCTACTPP-2964214890-gl57-gl95_hIgG1κVLDNA193GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-2964214890-gl57-gl95_hIgG1κLCDR1DNA194AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-2964214890-gl57-gl95_hIgG1κLCDR2DNA195AGAGGCAGCAACAGACTGAGCTPP-2964214890-gl57-gl95_hIgG1κLCDR3DNA196ATCCAGAACACCCACACACCTTACACCTPP-2964214890-gl57-gl95_hIgG1κ重鏈PRT197QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-2964214890-gl57-gl95_hIgG1κ輕鏈PRT198DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-2964214890-gl57-gl95_hIgG1κ重鏈DNA199CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-2964214890-gl57-gl95_hIgG1κ輕鏈DNA200GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGCAGCAACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-3271332694-hIgG1κVHPRT201EVQLVESGGGLVQPGGSLRLSCAASGFTFSDHYMDWVRQAPGKGLEWVGRIRNKTNNYATYYADSVKARFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGTLVTVSATPP-3271332694-hIgG1κHCDR1PRT202DHYMDTPP-3271332694-hIgG1κHCDR2PRT203RIRNKTNNYATYYADSVKATPP-3271332694-hIgG1κHCDR3PRT204GNSFAYTPP-3271332694-hIgG1κVLPRT205DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASSRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKLEIKTPP-3271332694-hIgG1κLCDR1PRT206KSSQSLLYSSNQKNYLATPP-3271332694-hIgG1κLCDR2PRT207WASSRESTPP-3271332694-hIgG1κLCDR3PRT208QQYYNYPLTTPP-3271332694-hIgG1κVHDNA209GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTCTCTGAGACTGTCTTGTGCCGCCAGCGGCTTCACCTTCAGCGACCACTATATGGACTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGGACGGATCCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCAGGTTCACCATCAGCAGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTGCCTACTGGGGACAGGGCACCCTGGTCACAGTTTCTGCATPP-3271332694-hIgG1κHCDR1DNA210GACCACTATATGGACTPP-3271332694-hIgG1κHCDR2DNA211CGGATCCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCTPP-3271332694-hIgG1κHCDR3DNA212GGCAATAGCTTTGCCTACTPP-3271332694-hIgG1κVLDNA213GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTAAGCTGCTGATCTATTGGGCCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-3271332694-hIgG1κLCDR1DNA214AAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTPP-3271332694-hIgG1κLCDR2DNA215TGGGCCAGCAGCAGAGAAAGCTPP-3271332694-hIgG1κLCDR3DNA216CAGCAGTACTACAACTACCCTCTGACCTPP-3271332694-hIgG1κ重鏈PRT217EVQLVESGGGLVQPGGSLRLSCAASGFTFSDHYMDWVRQAPGKGLEWVGRIRNKTNNYATYYADSVKARFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-3271332694-hIgG1κ輕鏈PRT218DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASSRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-3271332694-hIgG1κ重鏈DNA219GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTCTCTGAGACTGTCTTGTGCCGCCAGCGGCTTCACCTTCAGCGACCACTATATGGACTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGGACGGATCCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCAGGTTCACCATCAGCAGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTGCCTACTGGGGACAGGGCACCCTGGTCACAGTTTCTGCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-3271332694-hIgG1κ輕鏈DNA220GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTAAGCTGCTGATCTATTGGGCCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-327142-F7-hIgG1κVHPRT221QVQLVQSGGGLVQPGGSLRLSCGASGFTFNKFWMSWVRQAPGKGLEWVANINQDGSEKYYVDSMKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCTREATYWGQGTLVTVSSTPP-327142-F7-hIgG1κHCDR1PRT222KFWMSTPP-327142-F7-hIgG1κHCDR2PRT223NINQDGSEKYYVDSMKGTPP-327142-F7-hIgG1κHCDR3PRT224EATYTPP-327142-F7-hIgG1κVLPRT225AIQLTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLNWFQQRPGQSPRRLFYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPWTFGQGTKVEIKTPP-327142-F7-hIgG1κLCDR1PRT226RSSQSLVYSDGNTYLNTPP-327142-F7-hIgG1κLCDR2PRT227KVSNRDSTPP-327142-F7-hIgG1κLCDR3PRT228MQGTHWPWTTPP-327142-F7-hIgG1κVHDNA229CAGGTTCAGCTGGTTCAGTCTGGCGGAGGACTGGTTCAACCAGGCGGAAGCCTGAGACTGTCTTGTGGCGCCAGCGGCTTCACCTTCAACAAGTTCTGGATGAGCTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGCCAACATCAACCAGGACGGCAGCGAGAAGTACTACGTGGACAGCATGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAGAGCCGAGGACACCGCCGTGTACTACTGTACCAGAGAGGCCACATATTGGGGCCAGGGCACACTGGTTACAGTGTCCTCATPP-327142-F7-hIgG1κHCDR1DNA230AAGTTCTGGATGAGCTPP-327142-F7-hIgG1κHCDR2DNA231AACATCAACCAGGACGGCAGCGAGAAGTACTACGTGGACAGCATGAAGGGCTPP-327142-F7-hIgG1κHCDR3DNA232GAGGCCACATATTPP-327142-F7-hIgG1κVLDNA233GCTATTCAGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAGCGACGGCAACACCTACCTGAACTGGTTCCAGCAGAGGCCCGGACAGTCTCCTAGACGGCTGTTCTACAAGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACACACTGGCCCTGGACATTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-327142-F7-hIgG1κLCDR1DNA234AGATCTAGCCAGAGCCTGGTGTACAGCGACGGCAACACCTACCTGAACTPP-327142-F7-hIgG1κLCDR2DNA235AAGGTGTCCAACAGAGACAGCTPP-327142-F7-hIgG1κLCDR3DNA236ATGCAGGGCACACACTGGCCCTGGACATPP-327142-F7-hIgG1κ重鏈PRT237QVQLVQSGGGLVQPGGSLRLSCGASGFTFNKFWMSWVRQAPGKGLEWVANINQDGSEKYYVDSMKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCTREATYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-327142-F7-hIgG1κ輕鏈PRT238AIQLTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLNWFQQRPGQSPRRLFYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPWTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-327142-F7-hIgG1κ重鏈DNA239CAGGTTCAGCTGGTTCAGTCTGGCGGAGGACTGGTTCAACCAGGCGGAAGCCTGAGACTGTCTTGTGGCGCCAGCGGCTTCACCTTCAACAAGTTCTGGATGAGCTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGCCAACATCAACCAGGACGGCAGCGAGAAGTACTACGTGGACAGCATGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAGAGCCGAGGACACCGCCGTGTACTACTGTACCAGAGAGGCCACATATTGGGGCCAGGGCACACTGGTTACAGTGTCCTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-327142-F7-hIgG1κ輕鏈DNA240GCTATTCAGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAGCGACGGCAACACCTACCTGAACTGGTTCCAGCAGAGGCCCGGACAGTCTCCTAGACGGCTGTTCTACAAGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACACACTGGCCCTGGACATTTGGCCAGGGCACCAAGGTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-327154-11G-hIgG1κVHPRT241EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-327154-11G-hIgG1κHCDR1PRT242NYSMNTPP-327154-11G-hIgG1κHCDR2PRT243LISSNSSYIYYADSVKGTPP-327154-11G-hIgG1κHCDR3PRT244GGFDYTPP-327154-11G-hIgG1κVLPRT245EIVLTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWSTFGQGTKVEIK＞TPP-327154-11G-hIgG1κLCDR1PRT246RSSQSLVYNNGNTYLHTPP-327154-11G-hIgG1κLCDR2PRT247KVSNRDSTPP-327154-11G-hIgG1κLCDR3PRT248MQHTHWSTTPP-327154-11G-hIgG1κVHDNA249GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-327154-11G-hIgG1κHCDR1DNA250AACTACAGCATGAACTPP-327154-11G-hIgG1κHCDR2DNA251CTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCTPP-327154-11G-hIgG1κHCDR3DNA252GGCGGCTTTGATTACTPP-327154-11G-hIgG1κVLDNA253GAGATTGTGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGTCCACCTTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-327154-11G-hIgG1κLCDR1DNA254AGATCTAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTPP-327154-11G-hIgG1κLCDR2DNA255AAGGTGTCCAACAGAGACAGCTPP-327154-11G-hIgG1κLCDR3DNA256ATGCAGCACACCCACTGGTCCACCTPP-327154-11G-hIgG1κ重鏈PRT257EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-327154-11G-hIgG1κ輕鏈PRT258EIVLTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWSTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-327154-11G-hIgG1κ重鏈DNA259GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-327154-11G-hIgG1κ輕鏈DNA260GAGATTGTGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGTCCACCTTTGGCCAGGGCACCAAGGTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-327167-5B-hIgG1κVHPRT261QVQLVQSGGGLVKPGGSLRLSCVASGFTFSGYSMNWVRQAPGKGLEWVSFINNSSSYIVYADSVKGRFTISRDNAKNSLSLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-327167-5B-hIgG1κHCDR1PRT262GYSMNTPP-327167-5B-hIgG1κHCDR2PRT263FINNSSSYIVYADSVKGTPP-327167-5B-hIgG1κHCDR3PRT264GGFDYTPP-327167-5B-hIgG1κVLPRT265DIVMTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLHWFHQRPGQSPRRLISKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPTFGQGTKVEIKTPP-327167-5B-hIgG1κLCDR1PRT266RSSQSLVYSDGNTYLHTPP-327167-5B-hIgG1κLCDR2PRT267KVSNRGSTPP-327167-5B-hIgG1κLCDR3PRT268MQGTHWPTTPP-327167-5B-hIgG1κVHDNA269CAGGTTCAGCTGGTTCAGTCTGGCGGCGGACTTGTGAAACCTGGCGGAAGCCTGAGACTGAGCTGTGTGGCCAGCGGCTTTACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATCAACAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-327167-5B-hIgG1κHCDR1DNA270GGCTACAGCATGAACTPP-327167-5B-hIgG1κHCDR2DNA271TTCATCAACAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGGCTPP-327167-5B-hIgG1κHCDR3DNA272GGCGGCTTTGATTACTPP-327167-5B-hIgG1κVLDNA273GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGCGACGGCAATACCTACCTGCACTGGTTCCACCAGCGGCCTGGACAGTCTCCTAGACGGCTGATCTCCAAGGTGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACCCACTGGCCTACCTTTGGCCAGGGAACAAAGGTGGAAATCAAATPP-327167-5B-hIgG1κLCDR1DNA274AGAAGCAGCCAGAGCCTGGTGTACAGCGACGGCAATACCTACCTGCACTPP-327167-5B-hIgG1κLCDR2DNA275AAGGTGTCCAACAGAGGCAGCTPP-327167-5B-hIgG1κLCDR3DNA276ATGCAGGGCACCCACTGGCCTACCTPP-327167-5B-hIgG1κ重鏈PRT277QVQLVQSGGGLVKPGGSLRLSCVASGFTFSGYSMNWVRQAPGKGLEWVSFINNSSSYIVYADSVKGRFTISRDNAKNSLSLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-327167-5B-hIgG1κ輕鏈PRT278DIVMTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLHWFHQRPGQSPRRLISKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-327167-5B-hIgG1κ重鏈DNA279CAGGTTCAGCTGGTTCAGTCTGGCGGCGGACTTGTGAAACCTGGCGGAAGCCTGAGACTGAGCTGTGTGGCCAGCGGCTTTACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATCAACAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-327167-5B-hIgG1κ輕鏈DNA280GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGCGACGGCAATACCTACCTGCACTGGTTCCACCAGCGGCCTGGACAGTCTCCTAGACGGCTGATCTCCAAGGTGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACCCACTGGCCTACCTTTGGCCAGGGAACAAAGGTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-327177-8B-hIgG1κVHPRT281EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-327177-8B-hIgG1κHCDR1PRT282NYSMNTPP-327177-8B-hIgG1κHCDR2PRT283LISSNSSYIYYADSVKGTPP-327177-8B-hIgG1κHCDR3PRT284GGFDYTPP-327177-8B-hIgG1κVLPRT285DIVMTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVEIKTPP-327177-8B-hIgG1κLCDR1PRT286RSSQSLVYNNGNTYLHTPP-327177-8B-hIgG1κLCDR2PRT287KVSNRDSTPP-327177-8B-hIgG1κLCDR3PRT288MQHTHWPTTPP-327177-8B-hIgG1κVHDNA289GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-327177-8B-hIgG1κHCDR1DNA290AACTACAGCATGAACTPP-327177-8B-hIgG1κHCDR2DNA291CTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCTPP-327177-8B-hIgG1κHCDR3DNA292GGCGGCTTTGATTACTPP-327177-8B-hIgG1κVLDNA293GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-327177-8B-hIgG1κLCDR1DNA294AGAAGCAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTPP-327177-8B-hIgG1κLCDR2DNA295AAGGTGTCCAACAGAGACAGCTPP-327177-8B-hIgG1κLCDR3DNA296ATGCAGCACACCCACTGGCCTACCTPP-327177-8B-hIgG1κ重鏈PRT297EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-327177-8B-hIgG1κ輕鏈PRT298DIVMTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-327177-8B-hIgG1κ重鏈DNA299GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-327177-8B-hIgG1κ輕鏈DNA300GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-3271817-4D-hIgG1κVHPRT301QVQLVESGGGLVKPGGSLRLSCVVSGFTFSGYSMNWVRQAPGKGLEWVSFMSNSSSYIVYADSVKDRFTMSRDNAKNSLSLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-3271817-4D-hIgG1κHCDR1PRT302GYSMNTPP-3271817-4D-hIgG1κHCDR2PRT303FMSNSSSYIVYADSVKDTPP-3271817-4D-hIgG1κHCDR3PRT304GGFDYTPP-3271817-4D-hIgG1κVLPRT305DIQMTQSPLSLPVTLGQPASISCRSSQSLVYRDGNTYLHWFQQRPGQSPRRLIYKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVEIKTPP-3271817-4D-hIgG1κLCDR1PRT306RSSQSLVYRDGNTYLHTPP-3271817-4D-hIgG1κLCDR2PRT307KVSNRGSTPP-3271817-4D-hIgG1κLCDR3PRT308MQHTHWPTTPP-3271817-4D-hIgG1κVHDNA309CAGGTTCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGTGGTGTCCGGCTTCACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATGAGCAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGACCGGTTCACCATGAGCAGAGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-3271817-4D-hIgG1κHCDR1DNA310GGCTACAGCATGAACTPP-3271817-4D-hIgG1κHCDR2DNA311TTCATGAGCAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGACTPP-3271817-4D-hIgG1κHCDR3DNA312GGCGGCTTTGATTACTPP-3271817-4D-hIgG1κVLDNA313GACATCCAGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGAGATGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGGTGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-3271817-4D-hIgG1κLCDR1DNA314AGAAGCAGCCAGAGCCTGGTGTACAGAGATGGCAACACCTACCTGCACTPP-3271817-4D-hIgG1κLCDR2DNA315AAGGTGTCCAACAGAGGCAGCTPP-3271817-4D-hIgG1κLCDR3DNA316ATGCAGCACACCCACTGGCCTACCTPP-3271817-4D-hIgG1κ重鏈PRT317QVQLVESGGGLVKPGGSLRLSCVVSGFTFSGYSMNWVRQAPGKGLEWVSFMSNSSSYIVYADSVKDRFTMSRDNAKNSLSLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-3271817-4D-hIgG1κ輕鏈PRT318DIQMTQSPLSLPVTLGQPASISCRSSQSLVYRDGNTYLHWFQQRPGQSPRRLIYKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-3271817-4D-hIgG1κ重鏈DNA319CAGGTTCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGTGGTGTCCGGCTTCACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATGAGCAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGACCGGTTCACCATGAGCAGAGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-3271817-4D-hIgG1κ輕鏈DNA320GACATCCAGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGAGATGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGGTGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCCAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-16451hGPC3-fl鏈1PRT321QPPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINVDDMVNELFDSLFPVIYTQLMNPGLPDSALDINECLRGARRDLKVFGNFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILSLEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNAGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKVLKVAHVEHEETLSSRRRELIQKLKSFISFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGMKNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSMPKGRVLDKNLDEEGFESGDCGDDEDECIGGSGDGMIKVKNQLRFLAELAYDLDVDDAPGNSQQATPKDNEISTFHNLGNVHSPLKLLTSMAISVVCFFFLVHTPP-16452mGPC3-fl鏈1PRT322QPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINVDDMVNELFDSLFPVIYTQMMNPGLPESVLDINECLRGARRDLKVFGSFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILSLEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNGGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKILKVAHVEHEETLSSRRRELIQKLKSFINFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGMKNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSVPKGKVLDKSLDEEGLESGDCGDDEDECIGSSGDGMVKVKNQLRFLAELAYDLDVDDAPGNKQHGNQKDNEITTSHSVGNMPSPLKILISVAIYVACFFFLVHTPP-16453macmulGPC3-fl鏈1PRT323QPPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINVDDMVNELFDSLFPVIYTQLMNPGLPDSALDINECLRGARRDLKVFGNFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILSLEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNAGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKVLKVAHVEHEETLSSRRRELIQKLKSFISFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGMKNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSVPKGRVLDKNLDEEGFESGDCGDDEDECIGGSGDGMMKVKNQLRFLAELAYDLDVDDVPGNNQQATPKDNEISTFHNLGNVHSPLKLLTSMAISVVCFFFLVHTPP-35037IGHV1-46-IGHJ4人類種系重鏈鏈1PRT324QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYYMHWVRQAPGQGLEWMGIINPSGGSTSYAQKFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARYFDYWGQGTLVTVSSTPP-35038IGKV2-28-IGKJ2人類種系輕鏈鏈1PRT325DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWYLQKPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQALQTPYTFGQGTKLEIKTPP-754M014-G07-hIgG1-λVHPRT326EVQLLESGGGLVQPGGSLRLSCAASGFTFGDYAMSWVRQAPGKGLEWVSGISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAREGETSFGLDVWGQGTLVTVSSTPP-754M014-G07-hIgG1-λHCDR1PRT327DYAMSTPP-754M014-G07-hIgG1-λHCDR2PRT328GISGSGGSTYYADSVKGTPP-754M014-G07-hIgG1-λHCDR3PRT329EGETSFGLDVTPP-754M014-G07-hIgG1-λVLPRT330QSVLTQPPSASGTPGQRVTISCSGSSSNIGSNYVYWYQQLPGTAPKLLIYDNNDRPSGVPDRFSGSKSGTSASLAISGLRSEDEADYYCVAWDDSLNGHWVFGGGTKLTVLTPP-754M014-G07-hIgG1-λLCDR1PRT331SGSSSNIGSNYVYTPP-754M014-G07-hIgG1-λLCDR2PRT332DNNDRPSTPP-754M014-G07-hIgG1-λLCDR3PRT333VAWDDSLNGHWVTPP-754M014-G07-hIgG1-λVHDNA334GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCGGCGATTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGGCATCTCTGGCAGCGGCGGCAGCACCTACTACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCAGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGTGCCAGAGAGGGCGAGACAAGCTTCGGCCTGGATGTGTGGGGCCAGGGCACACTCGTGACCGTGTCATCTTPP-754M014-G07-hIgG1-λHCDR1DNA335GATTACGCCATGAGCTPP-754M014-G07-hIgG1-λHCDR2DNA336GGCATCTCTGGCAGCGGCGGCAGCACCTACTACGCCGATTCTGTGAAGGGCTPP-754M014-G07-hIgG1-λHCDR3DNA337GAGGGCGAGACAAGCTTCGGCCTGGATGTGTPP-754M014-G07-hIgG1-λVLDNA338CAGAGCGTGCTGACACAGCCTCCTAGCGCCTCTGGCACACCTGGCCAGAGAGTGACCATCAGCTGTAGCGGCAGCAGCAGCAACATCGGCAGCAACTACGTGTACTGGTATCAGCAGCTGCCCGGCACCGCCCCCAAACTGCTGATCTACGACAACAACGACCGGCCCAGCGGCGTGCCCGATAGATTTTCTGGCAGCAAGAGCGGCACCAGCGCCAGCCTGGCTATCAGCGGACTGAGAAGCGAGGACGAGGCCGACTACTACTGCGTGGCCTGGGACGACAGCCTGAACGGCCATTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGTPP-754M014-G07-hIgG1-λLCDR1DNA339AGCGGCAGCAGCAGCAACATCGGCAGCAACTACGTGTACTPP-754M014-G07-hIgG1-λLCDR2DNA340GACAACAACGACCGGCCCAGCTPP-754M014-G07-hIgG1-λLCDR3DNA341GTGGCCTGGGACGACAGCCTGAACGGCCATTGGGTGTPP-754M014-G07-hIgG1-λ重鏈PRT342EVQLLESGGGLVQPGGSLRLSCAASGFTFGDYAMSWVRQAPGKGLEWVSGISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAREGETSFGLDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-754M014-G07-hIgG1-λ重鏈DNA343GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCGGCGATTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGGCATCTCTGGCAGCGGCGGCAGCACCTACTACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCAGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGTGCCAGAGAGGGCGAGACAAGCTTCGGCCTGGATGTGTGGGGCCAGGGCACACTCGTGACCGTGTCATCTGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCTGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-754M014-G07-hIgG1-λ輕鏈DNA344CAGAGCGTGCTGACACAGCCTCCTAGCGCCTCTGGCACACCTGGCCAGAGAGTGACCATCAGCTGTAGCGGCAGCAGCAGCAACATCGGCAGCAACTACGTGTACTGGTATCAGCAGCTGCCCGGCACCGCCCCCAAACTGCTGATCTACGACAACAACGACCGGCCCAGCGGCGTGCCCGATAGATTTTCTGGCAGCAAGAGCGGCACCAGCGCCAGCCTGGCTATCAGCGGACTGAGAAGCGAGGACGAGGCCGACTACTACTGCGTGGCCTGGGACGACAGCCTGAACGGCCATTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGGGCCAGCCTAAAGCCGCCCCTAGCGTGACCCTGTTCCCTCCAAGCAGCGAGGAACTGCAGGCCAACAAGGCCACCCTCGTGTGCCTGATCAGCGACTTCTATCCTGGCGCCGTGACCGTGGCCTGGAAGGCCGATAGCTCTCCTGTGAAGGCCGGCGTGGAAACCACCACCCCTAGCAAGCAGAGCAACAACAAATACGCCGCCAGCAGCTACCTGAGCCTGACCCCCGAGCAGTGGAAGTCCCACAGATCCTACAGCTGCCAAGTGACCCACGAGGGCAGCACCGTGGAAAAGACAGTGGCCCCTACCGAGTGCAGCIn summary, these results demonstrate that Zr-89-DFO*-TPP-29537 is suitable for use as a PET imaging counterpart to Ac-225-Macropa-TPP-29537 for early stage development.sequenceTPP IDSequence NameSequence regionSequence TypeSEQ IDSEQTPP-5657 Germline VH3-23-isotype control hIgG1λ VH PRT 1 EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKLTGAEYFQHWGQGTLVTVSSTPP-5657 Germline VH3-23-isotype control hIgG1λ HCDR1 PRT 2 SYAMSTPP-5657 Germline VH3-23-isotype control hIgG1λ HCDR2 PRT 3 AISGSGGSTYYADSVKGTPP-5657 Germline VH3-23-isotype control hIgG1λ HCDR3 PRT 4 TGAEYFTPP-5657 Germline VH3-23-isotype control hIgG1λ V L PRT 5 QSVLTQPPSVSGAPGQRVTISCTGSSSNIGAGYDVHWYQQLPGTAPKLLIYGNSNRPSGVPDRFSGSKTSASLAITGLQAEDEADYYCQSYDSSLSGVVFGGGTKLTVLTPP-5657 Germline VH3-23-isotype control hIgG1λ LCDR1 PRT 6 TGSSSNIGAGYDVHTPP-5657 Germline VH3-23-isotype control hIgG1λ LCDR2 PRT 7 GNSNRPSTPP-5657 Germline VH3-23-isotype control hIgG1λ LCDR3 PRT 8 QSYDSSLSGVVTPP-5657 Germline VH3-23-isotype control hIgG1λ VH DNA 9 GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCAGCAGCTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGCCATCTCTGGCAGCGGCGGCAGCACCTAC TACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGCGCCAAACTGACAGGCGCCGAGTACTTCCAGCACTGGGGCCAGGGAACACTCGTGACCGTGTCATCATPP-5657 Germline VH3-23-isotype control hIgG1λ HCDR1 DNA 10 AGCTACGCCATGAGCTPP-5657 Germline VH3-23-isotype control hIgG1λ HCDR2 DNA 11 GCCATCTCTGGCAGCGGGCGGCAGCACCTACTACGCCGATTCTGTGAAGGGCTPP-5657 Germline VH3-23-isotype control hIgG1λ HCDR3 DNA 12 CTGACAGGCGCCGAGTACTTCCAGCACTPP-5657 Germline VH3-23-isotype control hIgG1λ V L DNA 13 CAGTCTGTTCTGACACAGCCTCCATCTGTGTCTGGCGCCCCTGGACAGAGAGTGACCATCAGCTTGTACAGCAGCTCCAATATCGGAGCCGGCTATGACGTGCACTGGTATCAGCAGCTGCCTGGCACAGCCCCTAAACTGCTGATCTACGGCAACAGCAACA GACCCAGCGGCGTGCCCGATAGATTTTCCGGCTCTAAGAGCGGCACAAGCGCCAGCCTGGCTATTACTGGACTGCAGGCCGAGGACGAGGCCGACTACTACTGTCAGAGCTACGACAGCAGCCTGAGCGGCGTTGTGTTTGGCGGAGGCACAAAAGCTGACAGTGCTATPP-5657 Germline VH3-23-isotype control hIgG1λ LCDR1 DNA 14 ACAGGCAGCTCCAATATCGGAGCCGGCTATGACGTGCACTPP-5657 Germline VH3-23-isotype control hIgG1λ LCDR2 DNA 15 GGCAACAGCAACAGACCCAGCTPP-5657 Germline VH3-23-isotype control hIgG1λ LCDR3 DNA 16 CAGAGCTACGACAGCAGCCTGAGCGGCGTTGTGTPP-5657 Germline VH3-23-isotype control hIgG1λ Heavy Chain PRT 17 EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKLTGAEYFQHWGQG TLVTVSSASTKGPSVFPLAPSSKSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-5657 Germline VH3-23-isotype control hIgG1λ Light chain PRT 18 QSVLTQPPSVSGAPGQRVTISCTGSSSNIGAGYDVHWYQQLPGTAPKLLIYGNSNRPSGVPDRFSGSKSGTSASLAITGLQAEDEADYYCQSYDSSLSGVVFGGGTKL TVLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECSTPP-5657 Germline VH3-23-isotype control hIgG1λ Heavy Chain DNA 19 GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCAGCAGCTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGCCATCTCTGGCAGCGGCGG CAGCACCTACTACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCAGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGCGCCAAACTGACAGGCGCCGAGTACTTCCAGCACTGGGGCCAGGGAAC ACTCGTGACCGTGTCATCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCG TGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGA CCCACACCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTAC GTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAA ACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAAC TACAAGACCACCCCCCTTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-5657 Germline VH3-23-isotype control hIgG1λ Light chain DNA 20 CAGTCTGTTCTGACACAGCCTCCATCTGTGTCTGGCGCCCCTGGACAGAGAGTGACCATCAGCTGTCAGGCAGCAGTCCAATATCGGAGCCGGCTATGACGTGCACTGGTATCAGCAGCTGCCTGGCACAGCCCCTAAACTGCTGATCTACGGCAACAGC AACAGACCCAGCGGCGTGCCCGATAGATTTTCCGGCTCTAAGAGCGGCACAAGCGCCAGCCTGGCTATTACTGGACTGCAGGCCGAGGACGAGGCCGACTACTGTCAGAGCTACGACAGCAGCCTGAGCGGCGTTGTGTTTGGCGGAGGCACAAAGCTGA CAGTGCTAGGCCAGCTAAAGCCGCCCCTAGCGTGACCCTGTTCCCTCCAAGCAGCGAGGAACTGCAGGCCAACAAGGCCACCTCGTGTGCCTGATCAGCGACTTCTATCCTGGCCGTGACCGTGGCCTGGAAGGCCGATAGCTCTCCTGTGAAGGCCGG CGTGGAAACCACCACCCCTAGCAAGCAGAGCAACAACAAATACGCCGCCAGCAGCTACCTGAGCCTGACCCCCGAGCAGTGGAAGTCCCACAGATCCTACAGCTGCCAAGTGACCCACGAGGGCAGCACCGTGGAAAAGACAGTGGCCCCTACCGAGTGCAGCTPP-14877 hYP9.1b-52T-hIgG1κ VH PRT twenty one EVQLVESGGGLVQPGGSLRLSCAASGFNFNTNAMNWVRQAPGKGLEWVGRVRNKTNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGTLVTVSSTPP-14877 hYP9.1b-52T-hIgG1κ HCDR1 PRT twenty two TNAMNTPP-14877 hYP9.1b-52T-hIgG1κ HCDR2 PRT twenty three RVRNKTNNYATYYADSVKDTPP-14877 hYP9.1b-52T-hIgG1κ HCDR3 PRT twenty four GNSFAYTPP-14877 hYP9.1b-52T-hIgG1κ V L PRT 25 DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKLEIKTPP-14877 hYP9.1b-52T-hIgG1κ LCDR1 PRT 26 KSSQSLLYSSNQKNYLATPP-14877 hYP9.1b-52T-hIgG1κ LCDR2 PRT 27 WASTRESTPP-14877 hYP9.1b-52T-hIgG1κ LCDR3 PRT 28 QQYYNYPLTTPP-14877 hYP9.1b-52T-hIgG1κ VH DNA 29 GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTCTTGAGACTGTCTTGTGCCGCCAGCGGCTTCAACTTCAACACCAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGGAAGAGTGCGGAACAAGACCAACAACTACG CCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTTGCCTACTGGGGACAGGGCACCCTGGTTACAGTTAGCTCATPP-14877 hYP9.1b-52T-hIgG1κ HCDR1 DNA 30 ACCAACGCCATGAACTPP-14877 hYP9.1b-52T-hIgG1κ HCDR2 DNA 31 AGAGTGCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGACTPP-14877 hYP9.1b-52T-hIgG1κ HCDR3 DNA 32 GGCAATAGCTTTGCCTACTPP-14877 hYP9.1b-52T-hIgG1κ V L DNA 33 GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTTAAGCTGCTGATCTACTGGG CCAGCACCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCTCTGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-14877 hYP9.1b-52T-hIgG1κ LCDR1 DNA 34 AAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTPP-14877 hYP9.1b-52T-hIgG1κ LCDR2 DNA 35 TGGGCCAGCACCAGAGAAAGCTPP-14877 hYP9.1b-52T-hIgG1κ LCDR3 DNA 36 CAGCAGTACTACAACTACCCTCTGACCTPP-14877 hYP9.1b-52T-hIgG1κ Heavy Chain PRT 37 EVQLVESGGGLVQPGGSLRLSCAASGFNFNTNAMNWVRQAPGKGLEWVGRVRNKTNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGT LVTVSSASTKGPSVFPLAPSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-14877 hYP9.1b-52T-hIgG1κ Light chain PRT 38 DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-14877 hYP9.1b-52T-hIgG1κ Heavy Chain DNA 39 GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTCTTGAGACTGTCTTGTGCCGCCAGCGGCTTCAACTTCAACACCAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGGAAGAGTGCGGAACAAGACCAA CAACTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTGCCTACTGGGGACAGGGCACCC TGGTTACAGTTAGCTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTG CACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACC CACACCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGT GGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAA CCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAAC TACAAGACCACCCCCCTTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-14877 hYP9.1b-52T-hIgG1κ Light chain DNA 40 GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTAAGCTGCTGATCTAC TGGGCCAGCACCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCTCTGACCTTCGGCCAGGGCACCAAGCTG GAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-14890 Trastuzumab-hIgG1κ VH PRT 41 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGDTAYSQKFKGRVTLTADKSTSTAYMELSSLTSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-14890 Trastuzumab-hIgG1κ HCDR1 PRT 42 DYEMTPP-14890 Trastuzumab-hIgG1κ HCDR2 PRT 43 ALDPKTGDTAYSQKFKGTPP-14890 Trastuzumab-hIgG1κ HCDR3 PRT 44 FYSYTYTPP-14890 Trastuzumab-hIgG1κ V L PRT 45 DVVMTQSPLSLPVTPGEPASISCRSSQSLVHSNRNTYLHWYLQKPGQSPQLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPPTFGQGTKLEIKTPP-14890 Trastuzumab-hIgG1κ LCDR1 PRT 46 RSSQSLVHSNRNTYLHTPP-14890 Trastuzumab-hIgG1κ LCDR2 PRT 47 KVSNRFSTPP-14890 Trastuzumab-hIgG1κ LCDR3 PRT 48 SQNTHVPPTTPP-14890 Trastuzumab-hIgG1κ VH DNA 49 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCGATA CAGCCTACAGCCAGAAATTCAAGGGCAGAGTGACCCTGACCGCCGACAAGAGCACAAGCACCGCCTACATGGAACTGAGCAGCCTGACCAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-14890 Trastuzumab-hIgG1κ HCDR1 DNA 50 GACTACGAGATGCACTPP-14890 Trastuzumab-hIgG1κ HCDR2 DNA 51 GCCCTGGATCCTAAGACCGGCGATACAGCCTACAGCCAGAAATTCAAGGGCTPP-14890 Trastuzumab-hIgG1κ HCDR3 DNA 52 TTCTACAGCTACACCTACTPP-14890 Trastuzumab-hIgG1κ V L DNA 53 GACGTTGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGCACAGCAACCGGAATAACCTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAAGGTG TCCAACCGGTTCAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCACCTACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-14890 Trastuzumab-hIgG1κ LCDR1 DNA 54 AGAAGCAGCCAGAGCCTGGTGCACAGCAACCGGAATAACCTACCTGCACTPP-14890 Trastuzumab-hIgG1κ LCDR2 DNA 55 AAGGTGTCCAACCGGTTCAGCTPP-14890 Trastuzumab-hIgG1κ LCDR3 DNA 56 AGCCAGAATACCCACGTGCCACCTACCTPP-14890 Trastuzumab-hIgG1κ Heavy Chain PRT 57 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGDTAYSQKFKGRVTLTADKSTSTAYMELSSLTSEDTAVYYCTRFYSYTYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-14890 Trastuzumab-hIgG1κ Light chain PRT 58 DVVMTQSPLSLPVTPGEPASISCRSSQSLVHSNRNTYLHWYLQKPGQSPQLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPPTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-14890 Trastuzumab-hIgG1κ Heavy Chain DNA 59 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCG GCGATACAGCCTACAGCCAGAAATTCAAGGGCAGAGTGACCCTGACCGCCGACAAGAGCACAAGCACCGCCTACATGGAACTGAGCAGCCTGACCAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTC ACAGTTTCTTCAGCCAGCACAAAGGGCCCTAGCGTGTTCCCTCTGGCTCCTAGCAGCAAGTCTACAAGCGGAGGAACAGCCGCTCTGGGCTGCCTGGTCAAGGATTACTTTCCCGAGCCTGTGACCGTGTCCTGGAATTCTGGCGCTCTGACAAGCGGCGTGCACAC CTTTCCAGCTGTGCTGCAAAGCAGCGGCCTGTACTCTCTGAGCAGCGTGGTCACAGTGCCAAGCTCTAGCCTGGGCACCCAGACCTACATCTGCAATGTGAACCACAAGCCTAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACA CCTGTCCTCCATGTCCTGCTCCAGAACTGCTCGGCGGACCTTCCGTGTTCCTGTTTCCTCCAAAGCCTAAGGACACCCTGATGATCAGCAGAACCCCTGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGATCCCGAAGTGAAGTTCAATTGGTACGTGGAC GGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACAGAGTGGTGTCCGTGCTGACCGTGCTGCACCAGGATTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCTCCTATCGAGAAAACCAT CAGCAAGGCCAAGGGCCAGCCTAGGGAACCCCAGGTTTACACACTGCCTCCAAGCAGGGACGAGCTGACCAAGAATCAGGTGTCCCTGACCTGCCTCGTGAAGGGCTTCTAACCCTTCCGATATCGCCGTGGAATGGGAGAGCAATGGCCAGCCTGAGAACAACTACA AGACAACCCCTCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACAGTGGACAAGTCCAGATGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGTCTCCTGGCAAGTPP-14890 Trastuzumab-hIgG1κ Light chain DNA 60 GACGTTGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGCACAGCAACCGGAATAACCTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAA GGTGTCCAACCGGTTCAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCTGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCACCTACCTTTGGCCAGGGCACCAAGCTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-15081 YP7-hIgG1κ VH PRT 61 EVQLVETGGGMVQPEGSLKLSCAASGFTFNKNAMNWVRQAPGKGLEWVARIRNKTNNYATYYADSVKARFTISRDDSQSMLYLQMNNLKIEDTAMYYCVAGNSFAYWGQGTLVTVSATPP-15081 YP7-hIgG1κ HCDR1 PRT 62 KNAMNTPP-15081 YP7-hIgG1κ HCDR2 PRT 63 RIRNKTNNYATYYADSVKATPP-15081 YP7-hIgG1κ HCDR3 PRT 64 GNSFAYTPP-15081 YP7-hIgG1κ V L PRT 65 DIVMSQSPSSLVVSIGEKVTMTCKSSQSLLYSSNQKNYLAWYQQKPGQSPKLLIYWASSRESGVPDRFTGSGSGTDFTLTISSVKAEDLAVYYCQQYYNYPLTFGAGTKLELKTPP-15081 YP7-hIgG1κ LCDR1 PRT 66 KSSQSLLYSSNQKNYLATPP-15081 YP7-hIgG1κ LCDR2 PRT 67 WASSRESTPP-15081 YP7-hIgG1κ LCDR3 PRT 68 QQYYNYPLTTPP-15081 YP7-hIgG1κ VH DNA 69 GAAGTGCAGCTGGTGGAAACAGGCGGCGGAATGGTTCAGCCTGAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACAAGAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGCCCGGATCAGAAACAAGACCAACAACTACG CCACCTACTACGCCGACAGCGTGAAGGCCAGATTCACCATCAGCAGGGACGACAGCCAGAGCATGCTGTACCTGCAGATGAACAACCTGAAGATCGAGGACACCGCCATGTACTACTGCGTGGCCGGCAATAGCTTTTGCCTATTGGGGCCAGGGCACCCTGGTCACAGTTTCTGCATPP-15081 YP7-hIgG1κ HCDR1 DNA 70 AAGAACGCCATGAACTPP-15081 YP7-hIgG1κ HCDR2 DNA 71 CGGATCAGAAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCTPP-15081 YP7-hIgG1κ HCDR3 DNA 72 GGCAATAGCTTTGCCTATTPP-15081 YP7-hIgG1κ V L DNA 73 GACATCGTGATGTCTCAGAGCCCTAGCAGCCTGGTGGTGTCCATCGGCGAGAAAGTGACCATGACCTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGACAGTCTCCCAAGCTGCTGATCTACTGGG CCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTCACAGGCAGCGGCAGCGGAACCGACTTCACCCTGACAATCAGCAGCGTGAAGGCCGAGGATCTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCTCTGACCTTCGGAGCCGGCACCAAGCTGGAACTGAAATPP-15081 YP7-hIgG1κ LCDR1 DNA 74 AAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTPP-15081 YP7-hIgG1κ LCDR2 DNA 75 TGGGCCAGCAGCAGAGAAAGCTPP-15081 YP7-hIgG1κ LCDR3 DNA 76 CAGCAGTACTACAACTACCCTCTGACCTPP-15081 YP7-hIgG1κ Heavy Chain PRT 77 EVQLVETGGGMVQPEGSLKLSCAASGFTFNKNAMNWVRQAPGKGLEWVARIRNKTNNYATYYADSVKARFTISRDDSQSMLYLQMNNLKIEDTAMYYCVAGNSFAYWGQGT LVTVSAASTKGPSVFPLAPSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-15081 YP7-hIgG1κ Light chain PRT 78 DIVMSQSPSSLVVSIGEKVTMTCKSSQSLLYSSNQKNYLAWYQQKPGQSPKLLIYWASSRESGVPDRFTGSGSGTDFTLTISSVKAEDLAVYYCQQYYNYPLTFGAGTKL ELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-15081 YP7-hIgG1κ Heavy Chain DNA 79 GAAGTGCAGCTGGTGGAAACAGGCGGCGGAATGGTTCAGCCTGAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACAAGAACGCCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTCGCCCGGATCAGAAACAAGACCAA CAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCAGATTCACCATCAGCAGGGACGACAGCCAGAGCATGCTGTACCTGCAGATGAACAACCTGAAGATCGAGGACACCGCCATGTACTACTGCGTGGCCGGCAATAGCTTTGCCTATTGGGGCCAGGGCACCC TGGTCACAGTTTCTGCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTG CACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACC CACACCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGT GGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAA CCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAAC TACAAGACCACCCCCCTTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-15081 YP7-hIgG1κ Light chain DNA 80 GACATCGTGATGTCTCAGAGCCCTAGCAGCCTGGTGGTGTCCATCGGCGAGAAAGTGACCATGACCTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGACAGTCTCCCAAGCTGCTGATCTAC TGGGCCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTCACAGGCAGCGGCAGCGGAACCGACTTCACCCTGACAATCAGCAGCGTGAAGGCCGAGGATCTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGAGCCGGCACCAAGCTG GAACTGAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-29312 14890-gl57-gl65_hIgG1κ VH PRT 81 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-29312 14890-gl57-gl65_hIgG1κ HCDR1 PRT 82 DYEMTPP-29312 14890-gl57-gl65_hIgG1κ HCDR2 PRT 83 ALDPKTGSTSYAQRFQGTPP-29312 14890-gl57-gl65_hIgG1κ HCDR3 PRT 84 FYSYTYTPP-29312 14890-gl57-gl65_hIgG1κ V L PRT 85 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKLEIKTPP-29312 14890-gl57-gl65_hIgG1κ LCDR1 PRT 86 RSSQSLLHSNGYNYLHTPP-29312 14890-gl57-gl65_hIgG1κ LCDR2 PRT 87 RGSNRASTPP-29312 14890-gl57-gl65_hIgG1κ LCDR3 PRT 88 QTTPP-29312 14890-gl57-gl65_hIgG1κ VH DNA 89 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCAGCA CAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-29312 14890-gl57-gl65_hIgG1κ HCDR1 DNA 90 GACTACGAGATGCACTPP-29312 14890-gl57-gl65_hIgG1κ HCDR2 DNA 91 GCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCTPP-29312 14890-gl57-gl65_hIgG1κ HCDR3 DNA 92 TTCTACAGCTACACCTACTPP-29312 14890-gl57-gl65_hIgG1κ V L DNA 93 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGC AGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-29312 14890-gl57-gl65_hIgG1κ LCDR1 DNA 94 AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-29312 14890-gl57-gl65_hIgG1κ LCDR2 DNA 95 AGAGGCAGCAACAGAGCCAGCTPP-29312 14890-gl57-gl65_hIgG1κ LCDR3 DNA 96 AGCCAGAATACCCACGTGCCATACACCTPP-29312 14890-gl57-gl65_hIgG1κ Heavy Chain PRT 97 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-29312 14890-gl57-gl65_hIgG1κ Light chain PRT 98 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-29312 14890-gl57-gl65_hIgG1κ Heavy Chain DNA 99 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCG GCAGCACAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTC ACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACAC CTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACA CCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGAC GGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCAT CAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-29312 14890-gl57-gl65_hIgG1κ Light chain DNA 100 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAG AGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-29537 14890-gl53-gl65_hIgG1κ VH PRT 101 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-29537 14890-gl53-gl65_hIgG1κ HCDR1 PRT 102 DYEMTPP-29537 14890-gl53-gl65_hIgG1κ HCDR2 PRT 103 ALDPSTGSTSYAQRFQGTPP-29537 14890-gl53-gl65_hIgG1κ HCDR3 PRT 104 FYSYTYTPP-29537 14890-gl53-gl65_hIgG1κ V L PRT 105 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKLEIKTPP-29537 14890-gl53-gl65_hIgG1κ LCDR1 PRT 106 RSSQSLLHSNGYNYLHTPP-29537 14890-gl53-gl65_hIgG1κ LCDR2 PRT 107 RGSNRASTPP-29537 14890-gl53-gl65_hIgG1κ LCDR3 PRT 108 QTTPP-29537 14890-gl53-gl65_hIgG1κ VH DNA 109 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCA CATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-29537 14890-gl53-gl65_hIgG1κ HCDR1 DNA 110 GACTACGAGATGCACTPP-29537 14890-gl53-gl65_hIgG1κ HCDR2 DNA 111 GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-29537 14890-gl53-gl65_hIgG1κ HCDR3 DNA 112 TTCTACAGCTACACCTACTPP-29537 14890-gl53-gl65_hIgG1κ V L DNA 113 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGC AGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-29537 14890-gl53-gl65_hIgG1κ LCDR1 DNA 114 AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-29537 14890-gl53-gl65_hIgG1κ LCDR2 DNA 115 AGAGGCAGCAACAGAGCCAGCTPP-29537 14890-gl53-gl65_hIgG1κ LCDR3 DNA 116 AGCCAGAATACCCACGTGCCATACACCTPP-29537 14890-gl53-gl65_hIgG1κ Heavy Chain PRT 117 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-29537 14890-gl53-gl65_hIgG1κ Light chain PRT 118 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHVPYTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-29537 14890-gl53-gl65_hIgG1κ Heavy Chain DNA 119 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCG GCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTC ACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACAC CTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACA CCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGAC GGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCAT CAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-29537 14890-gl53-gl65_hIgG1κ Light chain DNA 120 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAG AGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAATACCCACGTGCCATACACCTTTGGCCAGGGCACCAAGCTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-29538 14890-gl53-gl73_hIgG1κ VH PRT 121 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-29538 14890-gl53-gl73_hIgG1κ HCDR1 PRT 122 DYEMTPP-29538 14890-gl53-gl73_hIgG1κ HCDR2 PRT 123 ALDPSTGSTSYAQRFQGTPP-29538 14890-gl53-gl73_hIgG1κ HCDR3 PRT 124 FYSYTYTPP-29538 14890-gl53-gl73_hIgG1κ V L PRT 125 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHTPYTFGQGTKLEIKTPP-29538 14890-gl53-gl73_hIgG1κ LCDR1 PRT 126 RSSQSLLHSNGYNYLHTPP-29538 14890-gl53-gl73_hIgG1κ LCDR2 PRT 127 RGSNRASTPP-29538 14890-gl53-gl73_hIgG1κ LCDR3 PRT 128 SQTTPYTTPP-29538 14890-gl53-gl73_hIgG1κ VH DNA 129 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCA CATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-29538 14890-gl53-gl73_hIgG1κ HCDR1 DNA 130 GACTACGAGATGCACTPP-29538 14890-gl53-gl73_hIgG1κ HCDR2 DNA 131 GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-29538 14890-gl53-gl73_hIgG1κ HCDR3 DNA 132 TTCTACAGCTACACCTACTPP-29538 14890-gl53-gl73_hIgG1κ V L DNA 133 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGC AGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAACACCCACACACCTTACACCTTTGGCCAGGGCACCAAGCTGGAAATCAAATPP-29538 14890-gl53-gl73_hIgG1κ LCDR1 DNA 134 AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-29538 14890-gl53-gl73_hIgG1κ LCDR2 DNA 135 AGAGGCAGCAACAGAGCCAGCTPP-29538 14890-gl53-gl73_hIgG1κ LCDR3 DNA 136 AGCCAGAACACCCACACACCTTACACCTPP-29538 14890-gl53-gl73_hIgG1κ Heavy Chain PRT 137 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-29538 14890-gl53-gl73_hIgG1κ Light chain PRT 138 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQNTHTPYTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-29538 14890-gl53-gl73_hIgG1κ Heavy Chain DNA 139 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCG GCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTC ACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACAC CTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACA CCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGAC GGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCAT CAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-29538 14890-gl53-gl73_hIgG1κ Light chain DNA 140 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAG AGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCAGCCAGAACACCCACACACCTTACACCTTTGGCCAGGGCACCAAGCTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-29636 14890-gl53-gl93_hIgG1κ VH PRT 141 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-29636 14890-gl53-gl93_hIgG1κ HCDR1 PRT 142 DYEMTPP-29636 14890-gl53-gl93_hIgG1κ HCDR2 PRT 143 ALDPSTGSTSYAQRFQGTPP-29636 14890-gl53-gl93_hIgG1κ HCDR3 PRT 144 FYSYTYTPP-29636 14890-gl53-gl93_hIgG1κ V L PRT 145 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKTPP-29636 14890-gl53-gl93_hIgG1κ LCDR1 PRT 146 RSSQSLLHSNGYNYLHTPP-29636 14890-gl53-gl93_hIgG1κ LCDR2 PRT 147 RGSNRASTPP-29636 14890-gl53-gl93_hIgG1κ LCDR3 PRT 148 IQNTHTPYTTPP-29636 14890-gl53-gl93_hIgG1κ VH DNA 149 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCA CATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-29636 14890-gl53-gl93_hIgG1κ HCDR1 DNA 150 GACTACGAGATGCACTPP-29636 14890-gl53-gl93_hIgG1κ HCDR2 DNA 151 GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-29636 14890-gl53-gl93_hIgG1κ HCDR3 DNA 152 TTCTACAGCTACACCTACTPP-29636 14890-gl53-gl93_hIgG1κ V L DNA 153 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGC AGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-29636 14890-gl53-gl93_hIgG1κ LCDR1 DNA 154 AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-29636 14890-gl53-gl93_hIgG1κ LCDR2 DNA 155 AGAGGCAGCAACAGAGCCAGCTPP-29636 14890-gl53-gl93_hIgG1κ LCDR3 DNA 156 ATCCAGAACACCCACACACCTTACACCTPP-29636 14890-gl53-gl93_hIgG1κ Heavy Chain PRT 157 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-29636 14890-gl53-gl93_hIgG1κ Light chain PRT 158 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-29636 14890-gl53-gl93_hIgG1κ Heavy Chain DNA 159 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCG GCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTC ACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACAC CTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACA CCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGAC GGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCAT CAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-29636 14890-gl53-gl93_hIgG1κ Light chain DNA 160 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAG AGGCAGCAACAGAGCCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-29638 14890-gl53-gl95_hIgG1κ VH PRT 161 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-29638 14890-gl53-gl95_hIgG1κ HCDR1 PRT 162 DYEMTPP-29638 14890-gl53-gl95_hIgG1κ HCDR2 PRT 163 ALDPSTGSTSYAQRFQGTPP-29638 14890-gl53-gl95_hIgG1κ HCDR3 PRT 164 FYSYTYTPP-29638 14890-gl53-gl95_hIgG1κ V L PRT 165 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKTPP-29638 14890-gl53-gl95_hIgG1κ LCDR1 PRT 166 RSSQSLLHSNGYNYLHTPP-29638 14890-gl53-gl95_hIgG1κ LCDR2 PRT 167 RGSNRLSTPP-29638 14890-gl53-gl95_hIgG1κ LCDR3 PRT 168 IQNTHTPYTTPP-29638 14890-gl53-gl95_hIgG1κ VH DNA 169 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCGGCAGCA CATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-29638 14890-gl53-gl95_hIgG1κ HCDR1 DNA 170 GACTACGAGATGCACTPP-29638 14890-gl53-gl95_hIgG1κ HCDR2 DNA 171 GCCCTGGATCCTAGCACCGGCAGCACATCTTACGCCCAGAGATTCCAGGGCTPP-29638 14890-gl53-gl95_hIgG1κ HCDR3 DNA 172 TTCTACAGCTACACCTACTPP-29638 14890-gl53-gl95_hIgG1κ V L DNA 173 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGC AGCAACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-29638 14890-gl53-gl95_hIgG1κ LCDR1 DNA 174 AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-29638 14890-gl53-gl95_hIgG1κ LCDR2 DNA 175 AGAGGCAGCAACAGACTGAGCTPP-29638 14890-gl53-gl95_hIgG1κ LCDR3 DNA 176 ATCCAGAACACCCACACACCTTACACCTPP-29638 14890-gl53-gl95_hIgG1κ Heavy Chain PRT 177 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPSTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-29638 14890-gl53-gl95_hIgG1κ Light chain PRT 178 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-29638 14890-gl53-gl95_hIgG1κ Heavy Chain DNA 179 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAGCACCG GCAGCACATCTTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTC ACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACAC CTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACA CCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGAC GGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCAT CAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-29638 14890-gl53-gl95_hIgG1κ Light chain DNA 180 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAG AGGCAGCACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-29642 14890-gl57-gl95_hIgG1κ VH PRT 181 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLVTVSSTPP-29642 14890-gl57-gl95_hIgG1κ HCDR1 PRT 182 DYEMTPP-29642 14890-gl57-gl95_hIgG1κ HCDR2 PRT 183 ALDPKTGSTSYAQRFQGTPP-29642 14890-gl57-gl95_hIgG1κ HCDR3 PRT 184 FYSYTYTPP-29642 14890-gl57-gl95_hIgG1κ V L PRT 185 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKLEIKTPP-29642 14890-gl57-gl95_hIgG1κ LCDR1 PRT 186 RSSQSLLHSNGYNYLHTPP-29642 14890-gl57-gl95_hIgG1κ LCDR2 PRT 187 RGSNRLSTPP-29642 14890-gl57-gl95_hIgG1κ LCDR3 PRT 188 IQNTHTPYTTPP-29642 14890-gl57-gl95_hIgG1κ VH DNA 189 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCGGCAGCA CAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTCACAGTTTCTTCATPP-29642 14890-gl57-gl95_hIgG1κ HCDR1 DNA 190 GACTACGAGATGCACTPP-29642 14890-gl57-gl95_hIgG1κ HCDR2 DNA 191 GCCCTGGATCCTAAGACCGGCAGCACAAGCTACGCCCAGAGATTCCAGGGCTPP-29642 14890-gl57-gl95_hIgG1κ HCDR3 DNA 192 TTCTACAGCTACACCTACTPP-29642 14890-gl57-gl95_hIgG1κ V L DNA 193 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAGAGGC AGCAACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-29642 14890-gl57-gl95_hIgG1κ LCDR1 DNA 194 AGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTPP-29642 14890-gl57-gl95_hIgG1κ LCDR2 DNA 195 AGAGGCAGCAACAGACTGAGCTPP-29642 14890-gl57-gl95_hIgG1κ LCDR3 DNA 196 ATCCAGAACACCCACACACCTTACACCTPP-29642 14890-gl57-gl95_hIgG1κ Heavy Chain PRT 197 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYEMHWVRQAPGQGLEWMGALDPKTGSTSYAQRFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCTRFYSYTYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-29642 14890-gl57-gl95_hIgG1κ Light chain PRT 198 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLHWYLQKPGQSPQLLIYRGSNRLSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCIQNTHTPYTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-29642 14890-gl57-gl95_hIgG1κ Heavy Chain DNA 199 CAGGTTCAGCTGGTTCAGTCTGGCGCCGAAGTGAAGAAACCTGGCGCCTCTGTGAAGGTGTCCTGCAAGGCCAGCGGCTACACCTTTACCGACTACGAGATGCACTGGGTCCGACAGGCTCCAGGACAAGGCTTGGAATGGATGGGAGCCCTGGATCCTAAGACCG GCAGCACAAGCTACGCCCAGAGATTCCAGGGCAGAGTGACCATGACCAGAGACACCAGCACCTCCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACACCGCCGTGTACTACTGCACCCGGTTCTACAGCTACACCTACTGGGGCCAGGGCACCCTGGTC ACAGTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACAC CTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACA CCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGAC GGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCAT CAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-29642 14890-gl57-gl95_hIgG1κ Light chain DNA 200 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACCTGGCGAACCTGCCAGCATCAGCTGTAGAAGCAGCCAGTCTCTGCTGCACAGCAACGGCTACAACTACCTGCACTGGTATCTGCAGAAGCCCGGCCAGTCTCCTCAGCTGCTGATCTACAG AGGCAGCACAGACTGAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCGGCACCGACTTCACCCTGAAGATCTCCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGCATCCAGAACACCCACACACCTTACACCTTCGGCCAGGGCACCAAGCTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-32713 32694-hIgG1κ VH PRT 201 EVQLVESGGGLVQPGGSLRLSCAASGFTFSDHYMDWVRQAPGKGLEWVGRIRNKTNNYATYYADSVKARFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGTLVTVSATPP-32713 32694-hIgG1κ HCDR1 PRT 202 DHYMDTPP-32713 32694-hIgG1κ HCDR2 PRT 203 RIRNKTNNYATYYADSVKATPP-32713 32694-hIgG1κ HCDR3 PRT 204 GNSFAYTPP-32713 32694-hIgG1κ V L PRT 205 DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASSRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKLEIKTPP-32713 32694-hIgG1κ LCDR1 PRT 206 KSSQSLLYSSNQKNYLATPP-32713 32694-hIgG1κ LCDR2 PRT 207 WASSRESTPP-32713 32694-hIgG1κ LCDR3 PRT 208 QQYYNYPLTTPP-32713 32694-hIgG1κ VH DNA 209 GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTCTCTGAGACTGTCTTGTGCCGCCAGCGGCTTCACCTTCACGACCACTATATGGACTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGGACGGATCCGGAACAAGACCAACAACTACG CCACCTACTACGCCGACAGCGTGAAGGCCAGGTTCACCATCAGCAGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTGCCTACTGGGGACAGGGCACCCTGGTCACAGTTTCTGCATPP-32713 32694-hIgG1κ HCDR1 DNA 210 GACCACTATATGGACTPP-32713 32694-hIgG1κ HCDR2 DNA 211 CGGATCCGGAACAAGACCAACAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCTPP-32713 32694-hIgG1κ HCDR3 DNA 212 GGCAATAGCTTTGCCTACTPP-32713 32694-hIgG1κ V L DNA 213 GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTAAGCTGCTGATCTATTGGG CCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCTCTGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAATPP-32713 32694-hIgG1κ LCDR1 DNA 214 AAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTPP-32713 32694-hIgG1κ LCDR2 DNA 215 TGGGCCAGCAGCAGAGAAAGCTPP-32713 32694-hIgG1κ LCDR3 DNA 216 CAGCAGTACTACAACTACCCTCTGACCTPP-32713 32694-hIgG1κ Heavy Chain PRT 217 EVQLVESGGGLVQPGGSLRLSCAASGFTFSDHYMDWVRQAPGKGLEWVGRIRNKTNNYATYYADSVKARFTISRDDSKNSLYLQMNSLKTEDTAVYYCVAGNSFAYWGQGT LVTVSAASTKGPSVFPLAPSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-32713 32694-hIgG1κ Light chain PRT 218 DIVMTQSPDSLAVSLGERATINCKSSQSLLYSSNQKNYLAWYQQKPGQPPKLLIYWASSRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYNYPLTFGQGTKL EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-32713 32694-hIgG1κ Heavy Chain DNA 219 GAAGTGCAGCTGGTGGAATCTGGCGGAGGACTGGTTCAACCTGGCGGCTTCTGAGACTGTCTTGTGCCGCCAGCGGCTTCACCTTCACGACCACTATATGGACTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGGACGGATCCGGAACAAGACCAA CAACTACGCCACCTACTACGCCGACAGCGTGAAGGCCAGGTTCACCATCAGCAGGGACGACAGCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAAAACCGAGGACACCGCCGTGTACTATTGCGTGGCCGGCAATAGCTTTGCCTACTGGGGACAGGGCACCCT GGTCACAGTTTCTGCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGC ACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCC ACACCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTG GACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACC ATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTAC AAGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-32713 32694-hIgG1κ Light chain DNA 220 GACATCGTGATGACACAGAGCCCTGACAGCCTGGCCGTGTCTCTGGGAGAAAGAGCCACCATCAACTGCAAGAGCAGCCAGAGCCTGCTGTACTCCAGCAACCAGAAGAACTACCTGGCCTGGTATCAGCAGAAGCCCGGCCAGCCTCCTAAGCTGCTGATCTAT TGGGCCAGCAGCAGAGAAAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGACAATTTCTAGCCTGCAAGCCGAGGACGTGGCCGTGTACTACTGCCAGCAGTACTACAACTACCCTCTGACCTTCGGCCAGGGCACCAAGCTG GAAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-32714 2-F7-hIgG1κ VH PRT 221 QVQLVQSGGGLVQPGGSLRLSCGASGFTFNKFWMSWVRQAPGKGLEWVANINQDGSEKYYVDSMKGRFTISRDDNAKNSLYLQMNSLRAEDTAVYYCTREATYWGQGTLVTVSSTPP-32714 2-F7-hIgG1κ HCDR1 PRT 222 KFWMTPP-32714 2-F7-hIgG1κ HCDR2 PRT 223 NINQDGSEKYYVDSMKGTPP-32714 2-F7-hIgG1κ HCDR3 PRT 224 EATYTPP-32714 2-F7-hIgG1κ V L PRT 225 AIQLTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLNWFQQRPGQSPRRLFYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPWTFGQGTKVEIKTPP-32714 2-F7-hIgG1κ LCDR1 PRT 226 RSSQSLVYSDGNTYLNTPP-32714 2-F7-hIgG1κ LCDR2 PRT 227 KVSNRDSTPP-32714 2-F7-hIgG1κ LCDR3 PRT 228 QGTHWPWTTPP-32714 2-F7-hIgG1κ VH DNA 229 CAGGTTCAGCTGGTTCAGTCTGGCGGAGGACTGGTTCAACCAGGCGGAAGCCTGAGACTGTCTTGTGGCGCCAGCGGCTTCACCTTCAACAAGTTCTGGATGAGCTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGCCAACATCAACCAGGACGGCAGCG AGAAGTACTACGTGGACAGCATGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGTACCTGCAGATGAACTCCCTGAGAGCCGAGGACACCGCCGTGTACTACTGTACCAGAGAGGCCACATATTGGGGCCAGGGCACACTGGTTACAGTGTCCTCATPP-32714 2-F7-hIgG1κ HCDR1 DNA 230 AAGTTCTGGATGAGCTPP-32714 2-F7-hIgG1κ HCDR2 DNA 231 AACATCAACCAGGACGGCAGCGAGAAGTACTACGTGGACAGCATGAAGGGCTPP-32714 2-F7-hIgG1κ HCDR3 DNA 232 GAGGCCACATATTPP-32714 2-F7-hIgG1κ V L DNA 233 GCTATTCAGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAGCGACGGCAACACCTACCTGAACTGGTTCCAGCAGAGGCCCGGACAGTCTCCTAGACGGCTGTTCTACAAGGTG TCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACACACTGGCCCTGGACATTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-32714 2-F7-hIgG1κ LCDR1 DNA 234 AGATCTAGCCAGAGCCTGGTGTACAGCGACGGCAACACCTACCTGAACTPP-32714 2-F7-hIgG1κ LCDR2 DNA 235 AAGGTGTCCAACAGAGACAGCTPP-32714 2-F7-hIgG1κ LCDR3 DNA 236 ATGCAGGGCACACACTGGCCCTGGACATPP-32714 2-F7-hIgG1κ Heavy Chain PRT 237 QVQLVQSGGGLVQPGGSLRLSCGASGFTFNKFWMSWVRQAPGKGLEWVANINQDGSEKYYVDSMKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCTREATYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT CPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTI SKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-32714 2-F7-hIgG1κ Light chain PRT 238 AIQLTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLNWFQQRPGQSPRRLFYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPWTFGQGTKV EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-32714 2-F7-hIgG1κ Heavy Chain DNA 239 CAGGTTCAGCTGGTTCAGTCTGGCGGAGGACTGGTTCAACCAGGCGGAAGCCTGAGACTGTCTTGTGGCGCCAGCGGCTTCACCTTCAACAAGTTCTGGATGAGCTGGGTCCGACAGGCCCCTGGCAAAGGACTTGAGTGGGTCGCCAACATCAACCAGGACGGCA GCGAGAAGTACTACGTGGACAGCATGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGTACCTGCGATGAACTCCCTGAGAGCCGAGGACACCGCCGTGTACTACTGTACCAGAGAGGCCACATATTGGGGCCAGGGCACACTGGTTACAGT GTCCCTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCTTT CCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACCT GTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACGG CGGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATC AGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-32714 2-F7-hIgG1κ Light chain DNA 240 GCTATTCAGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAGCGACGGCAACACCTACCTGAACTGGTTCCAGCAGAGGCCCGGACAGTCTCCTAGACGGCTGTTCTACAA GGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACACACTGGCCCTGGACATTTGCCAGGGCACCAAGGTGG AAATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGC CAGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-32715 4-11G-hIgG1κ VH PRT 241 EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-32715 4-11G-hIgG1κ HCDR1 PRT 242 NYSMNTPP-32715 4-11G-hIgG1κ HCDR2 PRT 243 LISSNSSYIYYADSVKGTPP-32715 4-11G-hIgG1κ HCDR3 PRT 244 GGFTPP-32715 4-11G-hIgG1κ V L PRT 245 EIVLTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWSTFGQGTKVEIK＞TPP-32715 4-11G-hIgG1κ LCDR1 PRT 246 RSSQSLVYNNGNTYLHTPP-32715 4-11G-hIgG1κ LCDR2 PRT 247 KVSNRDSTPP-32715 4-11G-hIgG1κ LCDR3 PRT 248 MQHTHWSTTPP-32715 4-11G-hIgG1κ VH DNA 249 GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCAGCTAC ATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-32715 4-11G-hIgG1κ HCDR1 DNA 250 AACTACAGCATGAACTPP-32715 4-11G-hIgG1κ HCDR2 DNA 251 CTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCTPP-32715 4-11G-hIgG1κ HCDR3 DNA 252 GGCGGCTTTGATTACTPP-32715 4-11G-hIgG1κ V L DNA 253 GAGATTGTGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGG TGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGTCCACCTTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-32715 4-11G-hIgG1κ LCDR1 DNA 254 AGATCTAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTPP-32715 4-11G-hIgG1κ LCDR2 DNA 255 AAGGTGTCCAACAGAGACAGCTPP-32715 4-11G-hIgG1κ LCDR3 DNA 256 ATGCAGCACACCCACTGGTCCACCTPP-32715 4-11G-hIgG1κ Heavy Chain PRT 257 EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT CPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTI SKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-32715 4-11G-hIgG1κ Light chain PRT 258 EIVLTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWSTFGQGTKVE IKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-32715 4-11G-hIgG1κ Heavy Chain DNA 259 GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCA GCTACATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACC GTTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCT TTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACC TGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACG GCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATC AGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-32715 4-11G-hIgG1κ Light chain DNA 260 GAGATTGTGCTGACACAGAGCCCTCTGAGCCTGCCTGTTACACTGGGACAGCCTGCCAGCATCAGCTGCAGATCTAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACA AGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGTCCACCTTTGGCCAGGGCACCAAGGTGGAA ATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCC AGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-32716 7-5B-hIgG1κ VH PRT 261 QVQLVQSGGGLVKPGGSLRLSCVASGFTFSGYSMNWVRQAPGKGLEWVSFINNSSSYIVYADSVKGRFTISRDNAKNSSLLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-32716 7-5B-hIgG1κ HCDR1 PRT 262 GYSMTPP-32716 7-5B-hIgG1κ HCDR2 PRT 263 FINNSSSYIVYADSVKGTPP-32716 7-5B-hIgG1κ HCDR3 PRT 264 GGFTPP-32716 7-5B-hIgG1κ V L PRT 265 DIVMTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLHWFHQRPGQSPRRLISKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPTFGQGTKVEIKTPP-32716 7-5B-hIgG1κ LCDR1 PRT 266 RSSQSLVYSDGNTYLHTPP-32716 7-5B-hIgG1κ LCDR2 PRT 267 KVSNRGSTPP-32716 7-5B-hIgG1κ LCDR3 PRT 268 MQGTHWPTTPP-32716 7-5B-hIgG1κ VH DNA 269 CAGGTTCAGCTGGTTCAGTCTGGCGGCGGACTTGTGAAACCTGGCGGAAGCCTGAGACTGAGCTGTGTGGCCAGCGGCTTTACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATCAACAACAGCAGCAGCTAC ATCGTGTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-32716 7-5B-hIgG1κ HCDR1 DNA 270 GGCTACAGCATGAACTPP-32716 7-5B-hIgG1κ HCDR2 DNA 271 TTCATCAACAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGGCTPP-32716 7-5B-hIgG1κ HCDR3 DNA 272 GGCGGCTTTGATTACTPP-32716 7-5B-hIgG1κ V L DNA 273 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGCGACGGCAATAACCTACCTGCACTGGTTCCACCAGCGGCCTGGACAGTCTCCTAGACGGCTGATCTCCAAGG TGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACCCACTGGCCTACCTTTGGCCAGGGAACAAAGGTGGAAATCAAATPP-32716 7-5B-hIgG1κ LCDR1 DNA 274 AGAAGCAGCCAGAGCCTGGTGTACAGCGACGGCAATAACCTACCTGCACTPP-32716 7-5B-hIgG1κ LCDR2 DNA 275 AAGGTGTCCAACAGAGGCAGCTPP-32716 7-5B-hIgG1κ LCDR3 DNA 276 ATGCAGGGCACCCACTGGCCTACCTPP-32716 7-5B-hIgG1κ Heavy Chain PRT 277 QVQLVQSGGGLVKPGGSLRLSCVASGFTFSGYSMNWVRQAPGKGLEWVSFINNSSSYIVYADSVKGRFTISRDNAKNSSLLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT CPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTI SKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-32716 7-5B-hIgG1κ Light chain PRT 278 DIVMTQSPLSLPVTLGQPASISCRSSQSLVYSDGNTYLHWFHQRPGQSPRRLISKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQGTHWPTFGQGTKVE IKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-32716 7-5B-hIgG1κ Heavy Chain DNA 279 CAGGTTCAGCTGGTTCAGTCTGGCGGCGGACTTGTGAAACCTGGCGGAAGCCTGAGACTGAGCTGTGTGGCCAGCGGCTTTACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATCAACAACAGCAGCA GCTACATCGTGTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACC GTTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCT TTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACC TGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACG GCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATC AGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-32716 7-5B-hIgG1κ Light chain DNA 280 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGCGACGGCAATAACCTACCTGCACTGGTTCCACCAGCGGCCTGGACAGTCTCCTAGACGGCTGATCTCCA AGGTGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGGGCACCCACTGGCCTACCTTTGGCCAGGGAACAAAGGTGGAA ATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCC AGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-32717 7-8B-hIgG1κ VH PRT 281 EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-32717 7-8B-hIgG1κ HCDR1 PRT 282 NYSMNTPP-32717 7-8B-hIgG1κ HCDR2 PRT 283 LISSNSSYIYYADSVKGTPP-32717 7-8B-hIgG1κ HCDR3 PRT 284 GGFTPP-32717 7-8B-hIgG1κ V L PRT 285 DIVMTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVEIKTPP-32717 7-8B-hIgG1κ LCDR1 PRT 286 RSSQSLVYNNGNTYLHTPP-32717 7-8B-hIgG1κ LCDR2 PRT 287 KVSNRDSTPP-32717 7-8B-hIgG1κ LCDR3 PRT 288 MQHTHWPTTPP-32717 7-8B-hIgG1κ VH DNA 289 GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTCTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCAGCTAC ATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-32717 7-8B-hIgG1κ HCDR1 DNA 290 AACTACAGCATGAACTPP-32717 7-8B-hIgG1κ HCDR2 DNA 291 CTGATCAGCAGCAACAGCAGCTACATCTACTACGCCGACAGCGTGAAGGGCTPP-32717 7-8B-hIgG1κ HCDR3 DNA 292 GGCGGCTTTGATTACTPP-32717 7-8B-hIgG1κ V L DNA 293 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGG TGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-32717 7-8B-hIgG1κ LCDR1 DNA 294 AGAAGCAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTPP-32717 7-8B-hIgG1κ LCDR2 DNA 295 AAGGTGTCCAACAGAGACAGCTPP-32717 7-8B-hIgG1κ LCDR3 DNA 296 ATGCAGCACACCCACTGGCCTACCTPP-32717 7-8B-hIgG1κ Heavy Chain PRT 297 EVQLVESGGGLVKPGGSLRLSCAASGFTFSNYSMNWVRQAPGKGLEWVSLISSNSSYIYYADSVKGRFTISRDNAKNSLDLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT CPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTI SKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-32717 7-8B-hIgG1κ Light chain PRT 298 DIVMTQSPLSLPVTLGQPASISCRSSQSLVYNNGNTYLHWFQQRPGQSPRRLIYKVSNRDSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVE IKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-32717 7-8B-hIgG1κ Heavy Chain DNA 299 GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTTCTGAGACTGAGCTGTGCCGCTTCCGGCTTCACCTTCAGCAACTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCCTGATCAGCAGCAACAGCA GCTACATCTACTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAACAGCCTGGACCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACC GTTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCT TTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACC TGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACG GCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATC AGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-32717 7-8B-hIgG1κ Light chain DNA 300 GACATCGTGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAACAACGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACA AGGTGTCCAACAGAGACAGCGGCGTGCCCGATAGATTTTCTGGCAGCGGCCTCTGGCACCGACTTCACCCTGAAGATCAGCAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAA ATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCC AGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-32718 17-4D-hIgG1κ VH PRT 301 QVQLVESGGGLVKPGGSLRLSCVVSGFTFSGYSMNWVRQAPGKGLEWVSFMSNSSSYIVYADSVKDRFTMSRDNAKNSSLLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVTVSSTPP-32718 17-4D-hIgG1κ HCDR1 PRT 302 GYSMTPP-32718 17-4D-hIgG1κ HCDR2 PRT 303 FMSNSSSYIVYADSVKDTPP-32718 17-4D-hIgG1κ HCDR3 PRT 304 GGFTPP-32718 17-4D-hIgG1κ V L PRT 305 DIQMTQSPLSLPVTLGQPASISCRSSQSLVYRDGNTYLHWFQQRPGQSPRRLIYKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVEIKTPP-32718 17-4D-hIgG1κ LCDR1 PRT 306 RSSQSLVYRDGNTYLHTPP-32718 17-4D-hIgG1κ LCDR2 PRT 307 KVSNRGSTPP-32718 17-4D-hIgG1κ LCDR3 PRT 308 MQHTHWPTTPP-32718 17-4D-hIgG1κ VH DNA 309 CAGGTTCAGCTGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTTCTGAGACTGAGCTGTGTGGTGTCCGGCTTCACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATGAGCAACAGCAGCAGCTAC ATCGTGTACGCCGACAGCGTGAAGGACCGGTTCACCATGAGCAGAGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACCGTTTCTTCATPP-32718 17-4D-hIgG1κ HCDR1 DNA 310 GGCTACAGCATGAACTPP-32718 17-4D-hIgG1κ HCDR2 DNA 311 TTCATGAGCAACAGCAGCAGCTACATCGTGTACGCCGACAGCGTGAAGGACTPP-32718 17-4D-hIgG1κ HCDR3 DNA 312 GGCGGCTTTGATTACTPP-32718 17-4D-hIgG1κ V L DNA 313 GACATCCAGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGATGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACAAGG TGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAAATCAAATPP-32718 17-4D-hIgG1κ LCDR1 DNA 314 AGAAGCAGCCAGAGCCTGGTGTACAGAGATGGCAACACCTACCTGCACTPP-32718 17-4D-hIgG1κ LCDR2 DNA 315 AAGGTGTCCAACAGAGGCAGCTPP-32718 17-4D-hIgG1κ LCDR3 DNA 316 ATGCAGCACACCCACTGGCCTACCTPP-32718 17-4D-hIgG1κ Heavy Chain PRT 317 QVQLVESGGGLVKPGGSLRLSCVVSGFTFSGYSMNWVRQAPGKGLEWVSFMSNSSSYIVYADSVKDRFTMSRDNAKNSSLLQMNSLRAEDTAVYYCLTGGFDYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT CPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTI SKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKTPP-32718 17-4D-hIgG1κ Light chain PRT 318 DIQMTQSPLSLPVTLGQPASISCRSSQSLVYRDGNTYLHWFQQRPGQSPRRLIYKVSNRGSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQHTHWPTFGQGTKVE IKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECTPP-32718 17-4D-hIgG1κ Heavy Chain DNA 319 CAGGTTCAGCTGTGGAATCTGGCGGCGGACTTGTGAAACCTGGCGGCTTCTGAGACTGAGCTGTGTGGTGTCCGGCTTCACCTTCAGCGGCTACAGCATGAACTGGGTCCGACAGGCCCCTGGCAAAGGCCTTGAATGGGTGTCCTTCATGAGCAACAGCAGCA GCTACATCGTGTACGCCGACAGCGTGAAGGACCGGTTCACCATGAGCAGAGACAACGCCAAGAACAGCCTGAGCCTGCAGATGAACTCCCTGAGAGCCGAGGATACCGCCGTGTACTACTGTCTGACCGGCGGCTTTGATTACTGGGGCCAGGGAACACTGGTCACC GTTTTCTTCAGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGCGTGCACACCT TTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAGACCCACACC TGTCCCCCTTGTCCTGCCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTACGTGGACG GCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAAACCATC AGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAACTACA AGACCACCCCCCCTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCAAGTPP-32718 17-4D-hIgG1κ Light chain DNA 320 GACATCCAGATGACACAGAGCCCTCTGAGCCTGCCTGTGACACTGGGACAGCCTGCCAGCATCAGCTGTAGAAGCAGCCAGAGCCTGGTGTACAGATGGCAACACCTACCTGCACTGGTTCCAGCAGAGGCCTGGCCAGTCTCCTAGACGGCTGATCTACA AGGTGTCCAACAGAGGCAGCGGCGTGCCCGATAGATTTTCTGGCTCTGGCAGCGGCACCGACTTCACCCTGAAGATCTCTAGAGTGGAAGCCGAGGACGTGGGCGTGTACTACTGTATGCAGCACACCCACTGGCCTACCTTTGGCCAGGGCACCAAGGTGGAA ATCAAACGAACCGTGGCCGCTCCCAGCGTGTTCATCTTCCCACCTAGCGACGAGCAGCTGAAGTCCGGCACAGCCTCTGTCGTGTGCCTGCTGAACAACTTCTACCCCCGCGAGGCCAAGGTGCAGTGGAAGGTGGACAATGCCCTGCAGAGCGGCAACAGCC AGGAAAGCGTGACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAGCAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACCCACCAGGGCCTGTCTAGCCCCGTGACCAAGAGCTTCAACCGGGGCGAGTGTTPP-16451 hGPC3-fl Chain 1 PRT 321 QPPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINV DDMVNELFDSLFPVIYTQLMNPGLPDSALDINECLRGARRDLKVFGNFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILS LEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNAGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKVLKVAHVEHEETLSSRRRELIQKLKSFISFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGM KNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSMPKGRVLDKNLDEEGFESGDCGDDEDECIGGSGDGMIKVKNQLRFLAELAYDLDVDDAPGNSQQATPKDNEISTFHNLGNVHSPLKLLTSMAISVVCFFFLVHTPP-16452 mGPC3-fl Chain 1 PRT 322 QPPPPPDATCHVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINV DDMVNELFDSLFPVIYTQMMNPGLPESVLDINECLRGARRDLKVFGSFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILS LEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNGGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKILKVAHVEHEETLSSRRRELIQKLKSFINFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGM KNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSVPKGKVLDKSLDEEGLESGDCGDDEDECIGSSSGDGMVKVKNQLRFLAELAYDLDVDDAPGNKQHGNQKDNEITTSHSVGNMPSPLKILISVAIYVACFFFLVHTPP-16453 macmulGPC3-fl Chain 1 PRT 323 QPPPPPPDATCHQVRSFFQRLQPGLKWVPETPVPGSDLQVCLPKGPTCCSRKMEEKYQLTARLNMEQLLQSASMELKFLIIQNAAVFQEAFEIVVRHAKNYTNAMFKNNYPSLTPQAFEFVGEFFTDVSLYILGSDINV DDMVNELFDSLFPVIYTQLMNPGLPDSALDINECLRGARRDLKVFGNFPKLIMTQVSKSLQVTRIFLQALNLGIEVINTTDHLKFSKDCGRMLTRMWYCSYCQGLMMVKPCGGYCNVVMQGCMAGVVEIDKYWREYILS LEELVNGMYRIYDMENVLLGLFSTIHDSIQYVQKNAGKLTTTIGKLCAHSQQRQYRSAYYPEDLFIDKKVLKVAHVEHEETLSSRRRELIQKLKSFISFYSALPGYICSHSPVAENDTLCWNGQELVERYSQKAARNGM KNQFNLHELKMKGPEPVVSQIIDKLKHINQLLRTMSVPKGRVLDKNLDEEGFESGDCGDDEDECIGGSGDGMMKVKNQLRFLAELAYDLDVDDVPGNNQQATPKDNEISTFHNLGNVHSPLKLLTSMAISVVCFFFLVHTPP-35037 IGHV1-46-IGHJ4 human germline heavy chain Chain 1 PRT 324 QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYYMHWVRQAPGQGLEWMGIINPSGGSTSYAQKFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARYFDYWGQGTLVTVSSTPP-35038 IGKV2-28-IGKJ2 human germline light chain Chain 1 PRT 325 DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWYLQKPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCMQALQTPYTFGQGTKLEIKTPP-754 M014-G07-hIgG1-λ VH PRT 326 EVQLLESGGGLVQPGGSLRLSCAASGFTFGDYAMSWVRQAPGKGLEWVSGISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAREGETSFGLDVWGQGTLVTVSSTPP-754 M014-G07-hIgG1-λ HCDR1 PRT 327 DYAMSTPP-754 M014-G07-hIgG1-λ HCDR2 PRT 328 GISGSGGSTYYADSVKGTPP-754 M014-G07-hIgG1-λ HCDR3 PRT 329 EGETSFGLDVTPP-754 M014-G07-hIgG1-λ V L PRT 330 QSVLTQPPSASGTPGQRVTISCSGSSSNIGSNYVYWYQQLPGTAPKLLIYDNNDRPSGVPDRFSGSKSGTSASLAISGLRSEDEADYYCVAWDDSLNGHWVFGGGTKLTVLTPP-754 M014-G07-hIgG1-λ LCDR1 PRT 331 SGSSSNIGSNYVYTPP-754 M014-G07-hIgG1-λ LCDR2 PRT 332 DNNDRPSTPP-754 M014-G07-hIgG1-λ LCDR3 PRT 333 VAWDDSLNGHWVTPP-754 M014-G07-hIgG1-λ VH DNA 334 GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCGGCGATTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGGCATCTCTGGCAGCGGCGGCAGCACCTACT ACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGTGCCAGAGAGGGCGAGACAAGCTTCGGCCTGGATGTGTGGGGCCAGGGCACACTCGTGACCGTGTCATCTTPP-754 M014-G07-hIgG1-λ HCDR1 DNA 335 GATTACGCCATGAGCTPP-754 M014-G07-hIgG1-λ HCDR2 DNA 336 GGCATCTCTGGCAGCGGCGGGCAGCACCTACTACGCCGATTCTGTGAAGGGCTPP-754 M014-G07-hIgG1-λ HCDR3 DNA 337 GAGGGCGAGACAAGCTTCGGCCTGGATGTGTPP-754 M014-G07-hIgG1-λ V L DNA 338 CAGAGCGTGCTGACACAGCCTCCTAGCGCCTCTGGCACACCTGGCCAGAGAGTGACCATCAGCTGTAGCGGCAGCAGCAGCAACATCGGCAGCAACTACGTGTACTGGTATCAGCAGCTGCCCGGCACCGCCCCCAAACTGCTGATCTACGACAACAACGACCGGC CCAGCGGCGTGCCCGATAGATTTTCTGGCAGCAAGAGCGGCACCAGCGCCAGCCTGGCTATCAGCGGACTGAGAAGCGAGGACGAGGCCGACTACTGCGTGGCCTGGGACGACAGCCTGAACGGCCATTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGTPP-754 M014-G07-hIgG1-λ LCDR1 DNA 339 AGCGGCAGCAGCAGCAACATCGGCAGCAACTACGTGTACTPP-754 M014-G07-hIgG1-λ LCDR2 DNA 340 GACAACAACGACCGGCCCAGCTPP-754 M014-G07-hIgG1-λ LCDR3 DNA 341 GTGGCCTGGGACGACAGCCTGAACGGCCATTGGGTGTPP-754 M014-G07-hIgG1-λ Heavy Chain PRT 342 EVQLLESGGGLVQPGGSLRLSCAASGFTFGDYAMSWVRQAPGKGLEWVSGISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAREGETSFGLDVWGQG TLVTVSSASTKGPSVFPLAPSSKSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGTPP-754 M014-G07-hIgG1-λ Heavy Chain DNA 343 GAAGTGCAGCTGCTGGAATCTGGCGGCGGACTGGTGCAGCCTGGCGGATCTCTGAGACTGAGCTGTGCCGCCAGCGGCTTCACCTTCGGCGATTACGCCATGAGCTGGGTGCGCCAGGCCCCTGGAAAAGGCCTGGAATGGGTGTCCGGCATCTCTGGCAGCGGCGGC AGCACCTACTACGCCGATTCTGTGAAGGGCCGGTTCACCATCAGCCGGGACAACAGCAAGAACACCCTGTACCTGCAGATGAACAGCCTGCGGGCCGAGGACACCGCCGTGTACTATTGTGCCAGAGAGGGCGAGACAAGCTTCGGCCTGGATGTGTGGGGCCAGGGC ACACTCGTGACCGTGTCATCTGCCAGCACCAAGGGCCCCAGCGTGTTCCCTCTGGCCCCTAGCAGCAAGAGCACATCTGGCGGAACAGCCGCCCTGGGCTGCCTCGTGAAGGACTACTTTCCCGAGCCCGTGACCGTGTCCTGGAACTCTGGCGCTCTGACAAGCGGC GTGCACACCTTTCCAGCCGTGCTGCAGAGCAGCGGCCTGTACTCTCTGAGCAGCGTCGTGACAGTGCCCAGCAGCTCTCTGGGCACCCAGACCTACATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAAGGTGGAACCCAAGAGCTGCGACAAG ACCCACACCTGTCCCCCTTGTCCTGCCCCGAACTGCTGGGAGGCCCTTCCGTGTTCCTGTTCCCCCCAAAGCCCAAGGACACCCTGATGATCAGCCGGACCCCCGAAGTGACCTGCGTGGTGGTGGATGTGTCCCACGAGGACCCTGAAGTGAAGTTCAATTGGTAC GTGGACGGCGTGGAAGTGCACAACGCCAAGACCAAGCCTAGAGAGGAACAGTACAGCACCTACCGGGTGGTGTCCGTGCTGACAGTGCTGCACCAGGACTGGCTGAACGGCAAAGAGTACAAGTGCAAGGTGTCCAACAAGGCCCTGCCTGCCCCCATCGAGAAA ACCATCAGCAAGGCCAAGGGCCAGCCCCGCGAACCCCAGGTGTACACACTGCCCCCAAGCAGGGACGAGCTGACCAAGAACCAGGTGTCCCTGACCTGTCTCGTGAAAGGCTTCTAACCCCTCCGATATCGCCGTGGAATGGGAGAGCAACGGCCAGCCCGAGAACAAC TACAAGACCACCCCCCTTGTGCTGGACAGCGACGGCTCATTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCCCGGTGGCAGCAGGGCAACGTGTTCAAGCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCTGGCTPP-754 M014-G07-hIgG1-λ Light chain DNA 344 CAGAGCGTGCTGACACAGCCTCCTAGCGCCTCTGGCACACCTGGCCAGAGAGTGACCATCAGCTGTAGCGGCAGCAGCAGCAACATCGGCAGCAACTACGTGTACTGGTATCAGCAGCTGCCCGGCACCGCCCCCAAACTGCTGATCTACGACAACAACGAC CGGCCCAGCGGCGTGCCCGATAGATTTTCTGGCAGCAAGAGCGGCACCAGCGCCAGCCTGGCTATCAGCGGACTGAGAAGCGAGGACGAGGCCGACTACTGCGTGGCCTGGGACGACAGCCTGAACGGCCATTGGGTGTTCGGCGGAGGCACCAAGCTGA CCGTGCTGGGCCAGCCTAAAGCCGCCCCTAGCGTGACCCTGTTCCCTCCAAGCAGCGAGGAACTGCAGGCCAACAAGGCCACCTCGTGTGCCTGATCAGCGACTTCTATCCTGGCCGTGACCGTGGCCTGGAAGGCCGATAGCTCTCCTGTGAAGGCCGG CGTGGAAACCACCACCCCTAGCAAGCAGAGCAACAACAAATACGCCGCCAGCAGCTACCTGAGCCTGACCCCCGAGCAGTGGAAGTCCCACAGATCCTACAGCTGCCAAGTGACCCACGAGGGCAGCACCGTGGAAAAGACAGTGGCCCCTACCGAGTGCAGC

圖1：抗GPC3抗體TPP-14890及TPP-29537對瞬時轉染至CHO細胞中之人類GPC3之FACS結合。TPP-5657係同型對照抗體。 圖2：抗GPC3抗體TPP-14890及TPP-29537對瞬時轉染至CHO細胞中之食蟹猴GPC3之FACS結合。TPP-5657係同型對照抗體。 圖3：抗GPC3抗體TPP-14890及TPP-29537對瞬時轉染至CHO細胞中之小鼠GPC3之FACS結合。TPP-5657係同型對照抗體。 圖4：抗GPC3抗體TPP-14890及TPP-29537在瞬時模擬轉染CHO細胞中之FACS結合。TPP-5657係同型對照抗體。 圖5：在將直接標記之抗體添加至HepG2細胞後5小時之經計算每個細胞之囊泡面積。TPP-5657係同型對照抗體。 圖6：在將Fabfluor標記之抗體添加至HepG2細胞後5小時之經計算每個細胞之囊泡面積。TPP-5657係同型對照抗體。 圖7：使用Alexa 488標記之TPP-14890在細胞系HFF-1、SU-DHL-10、PLC/PRF/5、HepG2、Hep3B2及原代纖維母細胞HCF及HPF上藉由流式細胞術測定之細胞表面之抗體結合能力。 圖8：使用Alexa 488標記之TPP-29537在細胞系HFF-1、SU-DHL-10、PLC/PRF/5、HepG2、Hep3B2及原代纖維母細胞HCF及HPF上藉由流式細胞術測定之細胞表面之抗體結合能力。 圖9：Ac-225-Macropa-TPP-29537及Ac-225-Macropa-TPP-14890對肽塗佈之GPC3磁珠粒之結合。總結合定義為[(珠粒中之活性-上清液中之活性) / (總活性)] × 100。 圖10：HepG2細胞(A)及HEP-3B細胞(B)中之Ac-225-Macropa-TPP-29537 (CAR 0.5及CAR 11)、Ac-225-Macropa-TPP-14890 (CAR 0.7及CAR 10)及Ac-225-Macropa-同型-對照(CAR 0.7)之活體外細胞毒性數據。 圖11：A) Ac-225-Macropa-TPP-29537 (CAR 0.7)及B) 225Ac-Macropa-同型-對照(CAR 0.7)在人類肝細胞瘤異種移植物模型Huh-7之不同組織中之生物分佈。使用0.75 mg/kg、500kBq/kg靜脈內單一劑量之總抗體劑量來投與動物。累積Ac-225以注射劑量% /克(%ID/g)給出。 圖12：A) Ac-225-Macropa-TPP-29537 (CAR 0.7)及Ac-225-Macropa-同型-對照(CAR 0.7)在人類肝細胞瘤異種移植物模型Huh-7中之活體內效能。使用數位卡尺量測腫瘤，且結果呈現為腫瘤面積(mm2)，計算為腫瘤之最長直徑x最短直徑。B) Huh-7腫瘤模型之活體內效能研究之存活%。 圖13：Zr-89-DFO*-TPP-29537對GPC3肽塗佈磁珠粒之結合。總結合定義為[(珠粒中之活性-上清液中之活性) / (總活性)] × 100。 圖14：使用Zr-89-DFO*-TPP-29537之具有經成像HEP-3B腫瘤之小鼠之冠狀PET/CT MIP。2 h n=1、4 h n=1、24 h n=3、72 h n=3且168 h n=2。 圖15：在HEP-3B腫瘤模型中，Zr-89-DFO*-TPP-29537之離體及活體內PET/CT量化以及Ac-225-Macropa-TPP-29537之離體生物分佈。離體生物分佈數據：在每個時間點，n=3。基於成像之分析：2 h n=1、4 h n=1、24 h n=3、72 h n=3且168 h n=2。數據表示為平均值+SD。Figure 1: FACS binding of anti-GPC3 antibodies TPP-14890 and TPP-29537 to human GPC3 transiently transfected into CHO cells. TPP-5657 is an isotype control antibody.Figure 2: FACS binding of anti-GPC3 antibodies TPP-14890 and TPP-29537 to cynomolgus monkey GPC3 transiently transfected into CHO cells. TPP-5657 is an isotype control antibody.Figure 3: FACS binding of anti-GPC3 antibodies TPP-14890 and TPP-29537 to mouse GPC3 transiently transfected into CHO cells. TPP-5657 is an isotype control antibody.Figure 4: FACS binding of anti-GPC3 antibodies TPP-14890 and TPP-29537 in transiently simulated transfected CHO cells. TPP-5657 is an isotype control antibody.Figure 5: Vesicle area per cell calculated 5 hours after adding directly labeled antibodies to HepG2 cells. TPP-5657 is an isotype control antibody.Figure 6: Vesicle area per cell calculated 5 hours after adding Fabfluor labeled antibodies to HepG2 cells. TPP-5657 is an isotype control antibody.Figure 7: Antibody binding capacity of TPP-14890 labeled with Alexa 488 on the cell lines HFF-1, SU-DHL-10, PLC/PRF/5, HepG2, Hep3B2, and primary fibroblast HCF and HPF measured by flow cytometry.Figure 8: Antibody binding capacity of TPP-29537 labeled with Alexa 488 on the cell lines HFF-1, SU-DHL-10, PLC/PRF/5, HepG2, Hep3B2, and primary fibroblast HCF and HPF measured by flow cytometry.Figure 9: Binding of Ac-225-Macropa-TPP-29537 and Ac-225-Macropa-TPP-14890 to peptide-coated GPC3 magnetic beads. Total binding was defined as [(activity in beads - activity in supernatant) / (total activity)] × 100.Figure 10: In vitro cytotoxicity data of Ac-225-Macropa-TPP-29537 (CAR 0.5 and CAR 11), Ac-225-Macropa-TPP-14890 (CAR 0.7 and CAR 10), and Ac-225-Macropa-isotype-control (CAR 0.7) in HepG2 cells (A) and HEP-3B cells (B).Figure 11: Biodistribution of A) Ac-225-Macropa-TPP-29537 (CAR 0.7) and B) 225Ac-Macropa-isotype-control (CAR 0.7) in different tissues of the human hepatoma xenograft model Huh-7. Animals were dosed with a single intravenous dose of 0.75 mg/kg, 500 kBq/kg total antibody. Cumulative Ac-225 was given as % injected dose/gram (%ID/g).Figure 12: A) In vivo efficacy of Ac-225-Macropa-TPP-29537 (CAR 0.7) and Ac-225-Macropa-isotype-control (CAR 0.7) in the human hepatoma xenograft model Huh-7. Tumors were measured using a digital caliper and the results are presented as tumor area (mm2), calculated as the longest diameter x shortest diameter of the tumor. B) Survival % in the in vivo efficacy study of the Huh-7 tumor model.Figure 13: Binding of Zr-89-DFO*-TPP-29537 to GPC3 peptide-coated magnetic beads. Total binding is defined as [(activity in beads - activity in supernatant) / (total activity)] × 100.Figure 14: Coronal PET/CT MIP of mice with imaged HEP-3B tumors using Zr-89-DFO*-TPP-29537. 2 h n=1, 4 h n=1, 24 h n=3, 72 h n=3, and 168 h n=2.Figure 15: Ex vivo and in vivo PET/CT quantification of Zr-89-DFO*-TPP-29537 and ex vivo biodistribution of Ac-225-Macropa-TPP-29537 in the HEP-3B tumor model. Ex vivo biodistribution data: at each time point, n=3. Imaging-based analysis: 2 h n=1, 4 h n=1, 24 h n=3, 72 h n=3, and 168 h n=2. Data are presented as mean + SD.

TW202517674A_113137777_SEQL.xmlTW202517674A_113137777_SEQL.xml

Claims (22)

Translated fromChinese

一種能夠結合至GPC3之經分離抗GPC3抗體或抗原結合片段，其包括： i) 包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，或 ii)     包括含有SEQ ID NO: 122之H-CDR1、含有SEQ ID NO: 123之H-CDR2及含有SEQ ID NO: 124之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 126之L-CDR1、含有SEQ ID NO: 127之L-CDR2及含有SEQ ID NO:128之L-CDR3之輕鏈抗原結合區域，或 iii)    包括含有SEQ ID NO: 142之H-CDR1、含有SEQ ID NO: 143之H-CDR2及含有SEQ ID NO: 144之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 146之L-CDR1、含有SEQ ID NO: 147之L-CDR2及含有SEQ ID NO:148之L-CDR3之輕鏈抗原結合區域，或 iv)    包括含有SEQ ID NO: 162之H-CDR1、含有SEQ ID NO: 163之H-CDR2及含有SEQ ID NO: 164之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 166之L-CDR1、含有SEQ ID NO: 167之L-CDR2及含有SEQ ID NO:168之L-CDR3之輕鏈抗原結合區域。An isolated anti-GPC3 antibody or antigen-binding fragment capable of binding to GPC3, comprising:i) a heavy chain antigen-binding region comprising H-CDR1 containing SEQ ID NO: 102, H-CDR2 containing SEQ ID NO: 103, and H-CDR3 containing SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 containing SEQ ID NO: 106, L-CDR2 containing SEQ ID NO: 107, and L-CDR3 containing SEQ ID NO: 108, orii)     a heavy chain antigen-binding region comprising H-CDR1 containing SEQ ID NO: 122, H-CDR2 containing SEQ ID NO: 123, and H-CDR3 containing SEQ ID NO: 124, and a light chain antigen-binding region comprising L-CDR1 containing SEQ ID NO: 126, L-CDR2 containing SEQ ID NO: 127 and a light chain antigen binding region containing an L-CDR2 of SEQ ID NO: 128, oriii)    comprising an H-CDR1 containing SEQ ID NO: 142, an H-CDR2 containing SEQ ID NO: 143 and an H-CDR3 containing SEQ ID NO: 144, and a light chain antigen binding region containing an L-CDR1 containing SEQ ID NO: 146, an L-CDR2 containing SEQ ID NO: 147 and an L-CDR3 containing SEQ ID NO: 148, oriv)    comprising an H-CDR1 containing SEQ ID NO: 162, an H-CDR2 containing SEQ ID NO: 163 and an H-CDR3 containing SEQ ID NO: 164, and a heavy chain antigen binding region containing SEQ ID NO: 166, L-CDR1 containing SEQ ID NO: 167, and light chain antigen binding region containing L-CDR3 containing SEQ ID NO: 168.如請求項1之經分離抗體或抗原結合片段，其包括： i) 包括SEQ ID NO: 101之可變重鏈結構域及包括SEQ ID NO: 105之可變輕鏈結構域；或 ii)     包括SEQ ID NO: 121之可變重鏈結構域及包括SEQ ID NO: 125之可變輕鏈結構域；或 iii)    包括SEQ ID NO: 141之可變重鏈結構域及包括SEQ ID NO: 145之可變輕鏈結構域；或 iv)    包括SEQ ID NO: 161之可變重鏈結構域及包括SEQ ID NO: 165之可變輕鏈結構域。An isolated antibody or antigen-binding fragment as claimed in claim 1, comprising:i) a variable heavy chain domain comprising SEQ ID NO: 101 and a variable light chain domain comprising SEQ ID NO: 105; orii)     a variable heavy chain domain comprising SEQ ID NO: 121 and a variable light chain domain comprising SEQ ID NO: 125; oriii)    a variable heavy chain domain comprising SEQ ID NO: 141 and a variable light chain domain comprising SEQ ID NO: 145; oriv)    a variable heavy chain domain comprising SEQ ID NO: 161 and a variable light chain domain comprising SEQ ID NO: 165.如前述請求項中任一項之經分離抗體或抗原結合片段，其中該單株抗體係IgG抗體。An isolated antibody or antigen-binding fragment as claimed in any preceding claim, wherein the monoclonal antibody is an IgG antibody.如前述請求項中任一項之經分離抗體或抗原結合片段，其中該單株抗體係IgG1抗體。An isolated antibody or antigen-binding fragment as claimed in any preceding claim, wherein the monoclonal antibody is an IgG1 antibody.如前述請求項中任一項之經分離抗體或抗原結合片段，其包括 i)      包括SEQ ID NO: 117之重鏈及包括SEQ ID NO: 118之輕鏈；或 ii)     包括SEQ ID NO: 137之重鏈及包括SEQ ID NO: 138之輕鏈；或 iii)    包括SEQ ID NO: 157之重鏈及包括SEQ ID NO: 158之輕鏈；或 iv)    包括SEQ ID NO: 177之重鏈及包括SEQ ID NO: 178之輕鏈。An isolated antibody or antigen-binding fragment as claimed in any of the preceding claims, comprisingi)     a heavy chain comprising SEQ ID NO: 117 and a light chain comprising SEQ ID NO: 118; orii)     a heavy chain comprising SEQ ID NO: 137 and a light chain comprising SEQ ID NO: 138; oriii)    a heavy chain comprising SEQ ID NO: 157 and a light chain comprising SEQ ID NO: 158; oriv)    a heavy chain comprising SEQ ID NO: 177 and a light chain comprising SEQ ID NO: 178.一種抗體結合物，其包括如請求項1至5中任一項之單株抗體或抗原結合片段。An antibody conjugate comprising the monoclonal antibody or antigen-binding fragment of any one of claims 1 to 5.如請求項6之抗體結合物，其包括放射性核素、細胞毒性劑、有機化合物、蛋白質毒素、免疫調節劑、其他抗體或抗原結合片段、螢光部分或嵌合抗原受體。The antibody conjugate of claim 6, which comprises a radionuclide, a cytotoxic agent, an organic compound, a protein toxin, an immunomodulator, another antibody or an antigen-binding fragment, a fluorescent moiety, or a chimeric antigen receptor.如請求項6或7中任一項之抗體結合物，其中該抗體或其抗原結合片段經由該單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。An antibody conjugate as claimed in any one of claims 6 or 7, wherein the antibody or antigen-binding fragment thereof is conjugated to the remainder of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or antigen-binding fragment thereof.一種具有式(I)之靶向錒結合物(TAC)其中[Ab]係如請求項1至5中任一項之能夠結合至GPC3之單株抗體或其抗原結合片段。A targeted tantalum conjugate (TAC) having formula (I) wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3 as described in any one of claims 1 to 5.如請求項9之靶向錒結合物，其中[Ab]經由該單株抗體或其抗原結合片段之離胺酸殘基之末端胺基結合至分子之其餘部分。The targeted titanate conjugate of claim 9, wherein [Ab] is bound to the rest of the molecule via the terminal amino group of the lysine residue of the monoclonal antibody or antigen-binding fragment thereof.一種具有式(I)之靶向錒結合物(TAC)其中[Ab]係能夠結合至GPC3之單株抗體或其抗原結合片段，該[Ab]包括：包括含有SEQ ID NO: 102之H-CDR1、含有SEQ ID NO: 103之H-CDR2及含有SEQ ID NO: 104之H-CDR3之重鏈抗原結合區域以及包括含有SEQ ID NO: 106之L-CDR1、含有SEQ ID NO: 107之L-CDR2及含有SEQ ID NO:108之L-CDR3之輕鏈抗原結合區域，其中[Ab]經由離胺酸殘基之末端胺基結合至分子之其餘部分。A targeted tantalum conjugate (TAC) having formula (I) Wherein [Ab] is a monoclonal antibody or an antigen-binding fragment thereof capable of binding to GPC3, the [Ab] comprises: a heavy chain antigen-binding region comprising H-CDR1 comprising SEQ ID NO: 102, H-CDR2 comprising SEQ ID NO: 103, and H-CDR3 comprising SEQ ID NO: 104, and a light chain antigen-binding region comprising L-CDR1 comprising SEQ ID NO: 106, L-CDR2 comprising SEQ ID NO: 107, and L-CDR3 comprising SEQ ID NO: 108, wherein [Ab] is bound to the rest of the molecule via the terminal amine group of the lysine residue.一種醫藥組合物，其包括如請求項1至5中任一項之經分離抗體或抗原結合片段、或如請求項6至8之抗體結合物、或如請求項9至11中任一項之靶向錒結合物，及視情況一或多種醫藥上可接受之賦形劑。A pharmaceutical composition comprising the isolated antibody or antigen-binding fragment of any one of claims 1 to 5, or the antibody conjugate of any one of claims 6 to 8, or the targeted titania conjugate of any one of claims 9 to 11, and optionally one or more pharmaceutically acceptable excipients.如請求項1至5中任一項之抗體或其抗原結合片段或如請求項6至8中任一項之抗體結合物或如請求項9至11中任一項之靶向錒結合物或如請求項12之醫藥組合物，其用於治療疾病。An antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, an antibody conjugate according to any one of claims 6 to 8, a targeted titanate conjugate according to any one of claims 9 to 11, or a pharmaceutical composition according to claim 12, for use in treating a disease.如請求項13所使用之如請求項1至5中任一項之抗體或其抗原結合片段或如請求項6至8中任一項之抗體結合物或如請求項9至11中任一項之靶向錒結合物或如請求項12之醫藥組合物，其中該疾病之特徵在於GPC3之異位表現。An antibody or antigen-binding fragment thereof as claimed in any one of claims 1 to 5, an antibody conjugate as claimed in any one of claims 6 to 8, a targeted titanate conjugate as claimed in any one of claims 9 to 11, or a pharmaceutical composition as claimed in claim 12, as used in claim 13, wherein the disease is characterized by heterotopic expression of GPC3.如請求項13或14中任一項所使用之如請求項1至5中任一項之抗體或其抗原結合片段或如請求項6至8中任一項之抗體結合物或如請求項9至11中任一項之靶向錒結合物或如請求項12之醫藥組合物其中該疾病係癌症、較佳地肝細胞癌症。An antibody or antigen-binding fragment thereof as claimed in any one of claims 1 to 5, an antibody conjugate as claimed in any one of claims 6 to 8, a targeted titanate conjugate as claimed in any one of claims 9 to 11, or a pharmaceutical composition as claimed in claim 12, as used in any one of claim 13 or 14, wherein the disease is cancer, preferably hepatocellular carcinoma.如請求項1至5中任一項之經分離抗體或抗原結合片段或如請求項6至8中任一項之結合物或如請求項9至11中任一項之靶向錒結合物或如請求項12之醫藥組合物，其用於與一或多種其他治療活性化合物同時、分開或依序組合使用。An isolated antibody or antigen-binding fragment of any one of claims 1 to 5, a conjugate of any one of claims 6 to 8, a targeted titanate conjugate of any one of claims 9 to 11, or a pharmaceutical composition of claim 12, for use in combination with one or more other therapeutically active compounds simultaneously, separately or sequentially.如請求項1至5中任一項之經分離抗體或抗原結合片段或如請求項6至8中任一項之抗體結合物或如請求項9至11中任一項之靶向錒結合物，其用作診斷劑或成像劑。An isolated antibody or antigen-binding fragment of any one of claims 1 to 5, an antibody conjugate of any one of claims 6 to 8, or a targeted titanate conjugate of any one of claims 9 to 11 for use as a diagnostic or imaging agent.一種經分離核酸序列，其編碼如請求項1至5中任一項之抗體或抗原結合片段。An isolated nucleic acid sequence encoding the antibody or antigen-binding fragment of any one of claims 1 to 5.一種載體，其包括如請求項18之核酸序列。A vector comprising the nucleic acid sequence of claim 18.一種經分離細胞，其經配置以產生如請求項1至5中任一項之抗體或抗原結合片段。An isolated cell configured to produce the antibody or antigen-binding fragment of any one of claims 1 to 5.一種用於產生如請求項1至5中任一項之抗體或抗原結合片段或如請求項6至8中任一項之抗體結合物或如請求項9至11中任一項之靶向錒結合物之方法，該方法包括培養如請求項20之細胞及視情況純化該抗體或抗原結合片段。A method for producing the antibody or antigen-binding fragment of any one of claims 1 to 5, the antibody conjugate of any one of claims 6 to 8, or the targeted titanate conjugate of any one of claims 9 to 11, the method comprising culturing the cell of claim 20 and purifying the antibody or antigen-binding fragment as appropriate.一種套組，其包括如請求項1至5中任一項之經分離抗體或抗原結合片段或如請求項6至8中任一項之結合物或如請求項9至11中任一項之靶向錒結合物或如請求項12之醫藥組合物，及使用說明書。A kit comprising the isolated antibody or antigen-binding fragment of any one of claims 1 to 5, the conjugate of any one of claims 6 to 8, the targeted titanate conjugate of any one of claims 9 to 11, or the pharmaceutical composition of claim 12, and instructions for use.