Oxadiazole analog derivative, its preparation method and its in application pharmaceuticallyTechnical field
The invention belongs to field of medicaments, She Ji oxadiazole analog derivative, its preparation method and its answering on medical researchWith Gong Kai oxadiazole analog derivatives of the present invention have the tryptophan metabolism of IDO mediations for treating on the way as IDO inhibitorThe disease of footpath pathological characteristicses, described disease includes cancer, Alzheimer disease, autoimmune disease, depression, anxietyDisease, cataract, mental handicape and AIDS.
Background technology
Tumor biotherapy is the new treatment that treatment and prevention of tumour is carried out using modern biotechnology and its Related product, Yin QianEntirely, effectively, the low feature of adverse reaction, the 4th kind of pattern (Clin as the oncotherapy after operation, radiotherapy, chemotherapyCancer Res,1997；3:2623-2629), it passes through to transfer the natural immunology defense of host, such as suppress the swollen of IDO mediationsKnurl Immune escaping mechanism, or give naturally-produced targeting very strong material and obtain antineoplastic effect.
IDO (Indoleamine-pyrrole-2,3-dioxygenase, IDO) is a kind of iron contentFerroheme monomeric protein, is made up of 403 amino acid residues, including two α-helixstructure domains of folding, and big structure domain includesCatalytic pocket, substrate can occur hydrophobic grade and act on (Int J Biochem Cell Biol.2007 in catalytic pocket with IDO；39(12):2167-72).In mammal, the irrelevant enzyme containing ferroheme for having two kinds of gene codes can be catalyzed colorThe oxidative degradation of propylhomoserin：IDO and tryptophan 2,3- dioxygenases (TDO).Every kind of enzymatic identical reaction：On kynurenin roadPromote the oxidation Decomposition of the 2,3- double bonds of indole ring in first rate-limiting step tryptophan catabolism in footpath.The expression master of TDOIt is limited to liver, it appears that as a homeostasis or " house keeper " gene, it is impossible to induced or reconciled by the signal of immune system(Nat Rev Immunol.2004；4(10):762-74).IDO is to be catalyzed the enzyme that tryptophan transfer turns to formylkynurenine, extensivelyIt is general to be distributed in people and other tissues of mammal (rabbit, mouse) in addition to liver, it is that can uniquely be catalyzed tryptophan beyond liverThe rate-limiting enzyme of catabolism, and tryptophan is cell maintenance activation and amino acid necessary to propagation, is also to constitute protein notImportant component (the Adv Exp Med Biol.2003 that can lack；527:455-63、Biochim Biophys Acta.2001；1527(3):167-75).IDO is bad with interferon (interferon, IFN), interleukins (interleukin, IL), tumourThe cytokine profiles such as necrosis factor are in close relations, and they can activate IDO (J Psychiatry under certain conditionNeurosci.2004；29(1):11–17、Med Hypotheses.2003；61(5-6):519-25).And the cell of T- cellsThere is the very sensitive point of adjustment of a tryptophan level in cycle, on the one hand, IDO makes local tryptophan depletion, causes T-Cells arrest in the G1 interim phases, so as to inhibit the propagation of T cell；On the other hand, what IDO catalysis tryptophan metabolism was produced is mainProduct cynruin causes Cellular Oxidation agent and antioxidant to change and induces T- Apoptosis by Mediated by Free Radicals, and this is to depositIt is the intrinsic immunosuppression mechanism of body.Current numerous studies show IDO expression higher in leukaemia, make partT cell propagation is suppressed, suppresses the cell-mediated immune responses of T-, T- cell activation signals is transduceed and is obstructed, so that mediate tumorThe attack of cell evasion immune system.It has been found that most of mankind's tumor groups express IDO (J Exp Med.2002 with becoming second nature；196(4):459-68、Nat Med.2003；9(10):1269-74、Trends Mol Med.2004；10(1):15-8).CauseThis, IDO is a target for the cancer immunotherapy of tool potentiality.
The inhibitor patent application of disclosed Selective depression IDO include WO2004094409, WO2006122150,WO2007075598、WO200409387、WO2008147283、WO2013174947、WO2008075991、WO2004093871、WO2005051321, WO2006056304, WO2010005958 and WO2014066834 etc..
IDO inhibitor has a good application prospect as medicine in pharmaceuticals industry, but not yet finds at present wellIDO inhibitor, in order to reach the purpose of more preferable oncotherapy effect, can better meet the market demand as marketed drug,Inventor wishes to the selective IDO inhibitor of the high-efficiency low-toxicity for developing a new generation.The present invention will provide a kind of new structureSelective IDO inhibitor, and find to show excellent effect and effect with the compound of this class formation, it is particularly excellentMedicine generation absorb activity.
The content of the invention
It is an object of the invention to provide the compound or its dynamic isomer shown in a kind of logical formula (I), mesomer, outerRaceme, enantiomter, diastereoisomer or its form of mixtures, or its pharmaceutically useful salt,
Wherein：
Selected from the mixture of cis-isomer, transisomer and cis-trans-isomer；
Ring A is selected from cycloalkyl, heterocyclic radical and heteroaryl；
Ring B is aryl or heteroaryl；
R1It is identical or different, and it is each independently selected from hydrogen atom, alkyl, cyano group, amino, halogen, alkenyl, alkynyl, hydroxylBase, nitro, alkoxy, hydroxyalkyl, haloalkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl, wherein described alkyl, alkeneBase, alkynyl, alkoxy, hydroxyalkyl, haloalkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl are optionally further independently of one anotherOne or many be selected from hydroxyl, halogen, amino, cyano group, alkyl, alkoxy, cycloalkyl, heterocyclic radical, aryl or heteroarylIndividual substitution base is replaced；
R2It is identical or different, and it is each independently selected from hydrogen atom, alkyl, hydroxyl, amino, alkoxy, hydroxyalkyl, haloAlkyl, halogenated alkoxy, cycloalkyl, heterocyclic radical, aryl, heteroaryl ,-OR3、-C(O)R3、-C(O)OR3、-S(O)mR3、-C(O)NR3R4、-OC(O)NR3R4、-NR3R4、-NR3C(O)R4With-NR3S(O)mR4, wherein described alkyl, alkoxy, hydroxyalkyl, halogenSubstituted alkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl are optionally selected from alkyl, halogen, amino, nitro, cyanogen independently of one anotherBase, hydroxyl, hydroxyalkyl, alkoxy, cycloalkyl, heterocyclic radical, aryl, heteroaryl ,-OR5、-C(O)R5、-C(O)OR5、-S(O)mR5、-NR5R6、-C(O)NR5R6、-NR5C(O)R6With-NR5S(O)mR6In one or more substitution bases replaced；
R3And R4It is identical or different, and be each independently selected from hydrogen atom, alkyl, hydroxyl, amino, alkoxy, hydroxyalkyl,Haloalkyl, cycloalkyl, heterocyclic radical, aryl, heteroaryl ,-OR5、-C(O)R5、-C(O)OR5、-S(O)mR5、-NR5R6、-C(O)NR5R6、-NR5C(O)R6With-NR5S(O)mR6, wherein described alkyl, alkoxy, hydroxyalkyl, haloalkyl, cycloalkyl, heterocycleBase, aryl and heteroaryl are optionally selected from alkyl, haloalkyl, halogen, hydroxyl, amino, nitro, cyano group, alkane independently of one anotherOne or more substitution bases in epoxide, hydroxyalkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl are replaced；
R5And R6It is identical or different, and it is each independently selected from hydrogen atom, alkyl, hydroxyl, amino, carboxylic acid ester groups, alcoxylBase, hydroxyalkyl, haloalkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl, wherein described alkyl, amino, alkoxy, hydroxyl alkaneBase, haloalkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl are optionally selected from alkyl, halogen, hydroxyl, ammonia independently of one anotherOne or more in base, carboxylic acid ester groups, nitro, cyano group, alkoxy, hydroxyalkyl, cycloalkyl, heterocyclic radical, aryl and heteroarylSubstitution base is replaced；
M is 0,1 or 2；
N is 0,1,2,3,4 or 5；And
X is 0,1,2,3,4 or 5.
In yet other embodiments, lead to the compound shown in formula (I) or its dynamic isomer, mesoBody, racemic modification, enantiomter, diastereoisomer or its form of mixtures or its officinal salt, its middle ring B are virtueBase, preferably phenyl.
In yet other embodiments, lead to the compound shown in formula (I) or its dynamic isomer, mesoBody, racemic modification, enantiomter, diastereoisomer or its form of mixtures or its officinal salt, wherein R1For hydrogen is formerSon or halogen.
In yet other embodiments, lead to the compound shown in formula (I) or its dynamic isomer, mesoBody, racemic modification, enantiomter, diastereoisomer or its form of mixtures or its officinal salt, wherein R2Selected from hydrogenAtom, alkyl, hydroxyl, amino ,-C (O) R3、-S(O)mR3、-OC(O)NR3R4、-NR3C(O)R4With-NR3S(O)mR4；R3、R4And mAs defined in logical formula (I).
In yet other embodiments, lead to the compound shown in formula (I) or its dynamic isomer, mesoBody, racemic modification, enantiomter, diastereoisomer or its form of mixtures or its officinal salt, wherein n are 1 or 2.
In yet other embodiments, lead to the compound shown in formula (I) or its dynamic isomer, mesoBody, racemic modification, enantiomter, diastereoisomer or its form of mixtures or its officinal salt, wherein x are 0 or 1.
In yet other embodiments, lead to the compound shown in formula (I) or its dynamic isomer, mesoBody, racemic modification, enantiomter, diastereoisomer or its form of mixtures or its officinal salt, it is logical formula (II)Shown compound：
Or its dynamic isomer, mesomer, racemic modification, enantiomter, diastereoisomer or its mixture shapeFormula, or its officinal salt,
Wherein：
Selected from the mixture of cis-isomer, transisomer and cis-trans-isomer；
G is selected from C, O, N, S (O)mAnd S；
R2It is identical or different, and it is each independently selected from hydrogen atom, alkyl, hydroxyl, amino, alkoxy, hydroxyalkyl, haloAlkyl, halogenated alkoxy, cycloalkyl, heterocyclic radical, aryl, heteroaryl ,-OR3、-C(O)R3、-C(O)OR3、-S(O)mR3、-C(O)NR3R4、-OC(O)NR3R4、-NR3R4、-NR3C(O)R4With-NR3S(O)mR4, wherein described alkyl, alkoxy, hydroxyalkyl, halogenSubstituted alkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl are optionally selected from alkyl, halogen, amino, nitro, cyanogen independently of one anotherBase, hydroxyl, hydroxyalkyl, alkoxy, cycloalkyl, heterocyclic radical, aryl, heteroaryl ,-OR5、-C(O)R5、-C(O)OR5、-S(O)mR5、-NR5R6、-C(O)NR5R6、-NR5C(O)R6With-NR5S(O)mR6In one or more substitution bases replaced；
X is 0,1 or 2；
Y is 0,1,2 or 3；
Z is 0,1,2 or 3；And
Ring B, R1、R3~R6, m and n is as defined in logical formula (I).
In yet other embodiments, lead to the compound shown in formula (I) or its dynamic isomer, mesoBody, racemic modification, enantiomter, diastereoisomer or its form of mixtures or its officinal salt, its middle ring A are selected from ringHexyl, indanyl, pyrazolyl, cyclopenta, pyranose, cyclobutyl, piperidyl, azetidinyl and pyrrolidinyl.
The typical compound of logical formula (I) is included but is not limited to：
Or its dynamic isomer, mesomer, racemic modification, enantiomter, diastereoisomer or its mixture shapeFormula, or its officinal salt.
The present invention also provides a kind of intermediate for preparing compound shown in logical formula (I), and the intermediate is shown in logical formula (III)Compound or its dynamic isomer, mesomer, racemic modification, enantiomter, diastereoisomer or its form of mixtures,Or its officinal salt,
Wherein：
Ring B is selected from aryl and heteroaryl；
R1It is identical or different, and it is each independently selected from hydrogen atom, alkyl, cyano group, amino, halogen, alkenyl, alkynyl, hydroxylBase, nitro, alkoxy, hydroxyalkyl, haloalkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl, wherein described alkyl, alkeneBase, alkynyl, alkoxy, hydroxyalkyl, haloalkyl, cycloalkyl, heterocyclic radical, aryl and heteroaryl are optionally selected from independently of one anotherOne or more substitutions in hydroxyl, halogen, amino, cyano group, alkyl, alkoxy, cycloalkyl, heterocyclic radical, aryl or heteroarylBase is replaced；
N is 0,1,2,3,4 or 5.
The present invention also provides a kind of intermediate for preparing compound shown in logical formula (I), and the intermediate is logical formula (V) shownizationCompound or its dynamic isomer, mesomer, racemic modification, enantiomter, diastereoisomer or its form of mixtures, orIts officinal salt,
Wherein：
Ring A, ring B, R1、R2, x and n is as defined in logical formula (I).
The present invention also provides one kind and prepares compound shown in logical formula (I) or its dynamic isomer, mesomer, racemicBody, enantiomter, the method for diastereoisomer or its form of mixtures or its officinal salt, the method include：
Logical formula (III) compound reacts under room temperature alkalescence condition with logical formula (VI), obtains logical formula (I) compound；
Wherein：
Ring A, ring B, R1、R2, x and n is as defined in logical formula (I).
The present invention also provides one kind and prepares compound shown in logical formula (I) or its dynamic isomer, mesomer, racemicBody, enantiomter, the method for diastereoisomer or its form of mixtures or its officinal salt, the method include：
Logical formula (V) compound open loop under room temperature alkalescence condition, obtains logical formula (I) compound；
Wherein：
Ring A, ring B, R1、R2, x and n is as defined in logical formula (I).
Another aspect of the present invention is related to a kind of pharmaceutical composition, shown in its logical formula (I) for containing treatment effective doseCompound or its dynamic isomer, mesomer, racemic modification, enantiomter, diastereoisomer or its mixture shapeFormula or pharmaceutically useful salt, and one or more pharmaceutically acceptable carrier, diluent or excipient.The invention further relates toA kind of method for preparing above-mentioned composition, it include by the compound shown in logical formula (I) or its dynamic isomer, mesomer,Racemic modification, enantiomter, diastereoisomer or its form of mixtures or its pharmaceutically useful salt with it is pharmaceutically acceptableCarrier, diluent or excipient mix.
The invention further relates to lead to formula (I) shown in compound or its dynamic isomer, mesomer, racemic modification,It is prepared by enantiomter, diastereoisomer or its form of mixtures, or its officinal salt, or the pharmaceutical composition comprising itFor prevent and/or Prevention have IDO mediate tryptophan metabolic pathway pathological characteristicses disease medicine inPurposes.IDO inhibitor can be used for the suppression of cardiac disorder and treat other tryptophan metabolic pathways that there is IDO to mediateThe disease of pathological characteristicses, the infection of these diseases virus such as including AIDS, such as AIDS, Lyme disease and streptococcus senseCell infection, myelodysplastic syndrome, Neurodegenerative conditions (such as Alzheimer disease, Huntington disease and the handkerchiefs such as dyeThe gloomy disease of gold), autoimmune disease, depression, anxiety disorder, mental handicape, cancer (including T cell leukaemia and colon cancer),Disease of eye (such as cataract and age-related yellow), wherein described cancer can be selected from breast cancer, cervical carcinoma, knotIntestinal cancer, lung cancer, stomach cancer, the carcinoma of the rectum, cancer of pancreas, the cancer of the brain, cutaneum carcinoma, carcinoma of mouth, prostate cancer, osteocarcinoma, kidney, oophoroma, wingGuang cancer, liver cancer, fallopian tube cneoplasms, ovarioncus, peritoneal tumor, IV phases melanoma, solid tumor, glioma, neuroglia are femaleCytoma, hepatocellular carcinoma, mastoid process kidney knurl, head and neck neoplasm, leukaemia, lymthoma, myeloma and non-small cell lung cancer.
The invention further relates to lead to compound or its dynamic isomer, mesomer, racemic modification, mapping shown in formula (I)Isomers, diastereoisomer or its form of mixtures, or its officinal salt, or the pharmaceutical composition comprising it, it is used for pre-Anti- and/or Prevention has the disease of the pathological characteristicses of the tryptophan metabolic pathway of IDO mediations.These diseases are included such asThe infection of the virus such as AIDS, the cell infection such as AIDS, Lyme disease and streptococcal infection, myelodysplastic syndrome,Neurodegenerative conditions (such as Alzheimer disease, Huntington disease and Parkinson's), autoimmune disease, depression, JiaoConsider disease, mental handicape, cancer (including T cell leukaemia and colon cancer), disease of eye (such as cataract and age-relatedYellow), wherein described cancer can be selected from breast cancer, cervical carcinoma, colon cancer, lung cancer, stomach cancer, the carcinoma of the rectum, cancer of pancreas, brainCancer, cutaneum carcinoma, carcinoma of mouth, prostate cancer, osteocarcinoma, kidney, oophoroma, carcinoma of urinary bladder, liver cancer, fallopian tube cneoplasms, ovarioncus, peritonaeumTumour, IV phases melanoma, solid tumor, glioma, spongioblastoma, hepatocellular carcinoma, mastoid process kidney knurl, incidenceTumour, leukaemia, lymthoma, myeloma and non-small cell lung cancer.
There is the disease of the tryptophan metabolic pathway of IDO mediations the invention further relates to a kind of Prevention and/or PreventionThe method of the disease of feature of science, it includes compound shown in from the logical formula (I) to patient therapeuticallv's effective dose or its is mutualTautomeric, mesomer, racemic modification, enantiomter, diastereoisomer or its form of mixtures, or its is pharmaceutically acceptableSalt, or the pharmaceutical composition comprising it.The disease including AIDS etc. virus infection, such as AIDS, Lyme disease andThe cell infections such as streptococcal infection, myelodysplastic syndrome, Neurodegenerative conditions (such as Alzheimer disease, the prosperous court of a feudal rulerDisease and Parkinson's), autoimmune disease, depression, anxiety disorder, mental handicape, cancer (including T cell leukaemia andColon cancer), disease of eye (such as cataract and age-related yellow), wherein described cancer can selected from breast cancer,Cervical carcinoma, colon cancer, lung cancer, stomach cancer, the carcinoma of the rectum, cancer of pancreas, the cancer of the brain, cutaneum carcinoma, carcinoma of mouth, prostate cancer, osteocarcinoma, kidney,Oophoroma, carcinoma of urinary bladder, liver cancer, fallopian tube cneoplasms, ovarioncus, peritoneal tumor, IV phases melanoma, solid tumor, glioma,Spongioblastoma, hepatocellular carcinoma, mastoid process kidney knurl, head and neck neoplasm, leukaemia, lymthoma, myeloma and non-small cellLung cancer.
Another aspect of the present invention is related to a kind of method for the treatment of cancer, and the method is including to patient therapeuticallv's effective doseOf the invention logical formula (I) described in compound or its dynamic isomer, mesomer, racemic modification, enantiomter, non-rightReflect isomers or its form of mixtures, or its officinal salt.The method shows prominent curative effect and less side effect, whereinDescribed cancer can be selected from breast cancer, cervical carcinoma, colon cancer, lung cancer, stomach cancer, the carcinoma of the rectum, cancer of pancreas, the cancer of the brain, cutaneum carcinoma, mouthChamber cancer, prostate cancer, osteocarcinoma, kidney, oophoroma, carcinoma of urinary bladder, liver cancer, fallopian tube cneoplasms, ovarioncus, peritoneal tumor, IV phases are blackMelanoma, solid tumor, glioma, spongioblastoma, hepatocellular carcinoma, mastoid process kidney knurl, head and neck neoplasm, white bloodDisease, lymthoma, myeloma and non-small cell lung cancer, preferably fallopian tube cneoplasms, peritoneal tumor, IV phases melanoma, myelomaAnd breast cancer, more preferably breast cancer.
Pharmaceutical composition containing active component can apply to oral form, such as tablet, dragee, lozenge, waterOr oil suspension, dispersible powder or particle, emulsion, hard or soft capsule, or syrup or elixir.Can be any according to this areaKnow that the method for preparing Pharmaceutical composition prepares Orally administered composition, such composition can containing one or more selected from it is following intoPoint：Sweetener, flavouring, colouring agent and preservative, to provide pleasing and good to eat pharmaceutical formulation.Tablet contain active component andFor the suitable nontoxic pharmaceutically useful excipient for preparing tablet for mixing.These excipient can be inert excipient, such as carbonSour calcium, sodium carbonate, lactose, calcium phosphate or sodium phosphate；Granulating agent and disintegrant, such as microcrystalline cellulose, cross-linked carboxymethyl fiberPlain sodium, cornstarch or alginic acid；Adhesive, such as starch, gelatin, polyvinylpyrrolidone or Arabic gum；And lubricant, exampleSuch as magnesium stearate, stearic acid or talcum powder.These tablets can not be coated or can be by covering the taste of medicine or in intestines and stomachIt is middle to postpone disintegration and absorb, thus the known technology of offer slow releasing function is coated in a long time.For example, water can be usedDissolubility taste masked material, such as hydroxypropyl methyl cellulose or hydroxypropyl cellulose, or extension time material such as ethyl is fineDimension element, acetylbutyrylcellulose.
Also can use wherein active component hard bright with what inert solid diluent such as calcium carbonate, calcium phosphate or kaolin mixedGlue capsule, or wherein active component and water-solubility carrier such as polyethylene glycol or oil soluble matchmaker such as peanut oil, atoleine or oliveThe Perle of olive oil mixing provides oral formulations.
Water slurry contains active material and the suitable excipient for preparing water slurry for mixing.Such excipient isSuspending agent, such as sodium carboxy methyl cellulose, methylcellulose, hydroxypropyl methyl cellulose, mosanom, polyvinylpyrrolidoneAnd Arabic gum；Dispersant or wetting agent can be naturally-produced phosphatide such as lecithin, or alkylene oxide and aliphatic acid contractingClose product such as Myrj 45, or oxirane and long-chain fatty alcohol condensation product, such as 17 carbon ethylideneEpoxide cetanol (heptadecaethyleneoxy cetanol), or oxirane and the portion as derived from aliphatic acid and hexitolThe condensation product of ester, such as polyoxyethylene sorbitol monoleate, or oxirane is divided to spread out with by aliphatic acid and hexitanThe condensation product of raw partial ester, such as PEO Arlacel-80.Aqueous suspension can also containing a kind of orDetermination of Preservatives such as ethylparaben or nipalgin n-propyl, one or more colouring agent, one or more flavouring andPlant or various sweeteners, such as sucrose, saccharin or aspartame.
Oil suspension can be suspended in vegetable oil such as peanut oil, olive oil, sesame oil or coconut oil by making active component, orIt is formulated in mineral oil such as atoleine.Oil suspension can contain thickener, such as beeswax, hard paraffin or cetanol.CanAbove-mentioned sweetener and flavouring is added, to provide good to eat preparation.Can by add antioxidant such as Butylated Hydroxyanisole or α-Fertility phenol preservation these compositions.
Prepare dispersible powder and particle that water is suspended also and active component is provided and is used for by adding water to make to be applied toThe dispersant or wetting agent of mixing, suspending agent or one or more preservative.Suitable dispersant or wetting agent and suspending agent canIllustrate above-mentioned example.Also other excipient such as sweetener, flavouring and colouring agent can be added.By adding antioxidant exampleAs ascorbic acid preserves these compositions.
Pharmaceutical composition of the invention can also be the form of oil in water emulsion.Oil phase can be vegetable oil such as olive oilOr peanut oil, or mineral oil such as atoleine or its mixture.Suitable emulsifying agent can be naturally-produced phosphatide, for exampleSoybean lecithin and ester or partial ester such as sorbitan monooleate as derived from aliphatic acid and hexitan, and the partial ester and ringThe condensation product of oxidative ethane, such as polyoxyethylene sorbitol monoleate.Emulsion can also contain sweetener, flavouring, preventRotten agent and antioxidant.Syrup and elixir can be prepared with Sweetening agents such as glycerine, propane diols, sorbierite or sucrose.Such preparationModerator, preservative, colouring agent and antioxidant can be contained.
Pharmaceutical composition of the invention can be sterile injectable aqueous form.The acceptable solvent or molten that can be usedAgent has water, ringer's solution and isotonic sodium chlorrde solution.Aseptic injection preparation can be that wherein active component is dissolved in the aseptic of oil phaseInjection oil-in-water microemulsion.For example active component is dissolved in the mixture of soybean oil and lecithin.Then oil solution is added into waterMicro emulsion is formed with treatment in the mixture of glycerine.Parenteral solution or micro emulsion can be injected the blood flow of patient by local a large amount of injectionsIn.Or, solution and micro emulsion are preferably given in the way of it can keep the compounds of this invention constant circulating concentration.To keep this perseveranceDetermine concentration, continuous intravenous delivery device can be used.The example of this device is that Deltec CADD-PLUS.TM.5400 types are quietArteries and veins syringe pump.
Pharmaceutical composition can be the form of aseptic injection water for intramuscular and subcutaneous administration or oil suspension.Can be byKnow technology, the suspension is prepared with the suitable dispersant of those described above or wetting agent and suspending agent.Aseptic injection preparation can alsoIt is the aseptic injectable solution or suspension prepared in the acceptable non-toxic diluent of parenteral or solvent, such as 1,3-BDOThe solution of middle preparation.In addition, it is convenient to sterile fixed oil as solvent or suspension media.For this purpose, can be used includingSynthetic glycerine list or diester are in interior any mediation fixing oil.Additionally, aliphatic acid such as oleic acid can also prepare injection.
The compounds of this invention can be given by the suppository form for rectally.Can be by by medicine and at normal temperaturesIt is for solid but in the rectum liquid, thus can dissolves in the rectum and discharge the suitable nonirritant excipient mixing of medicineTo prepare these pharmaceutical compositions.Such material includes cocoa butter, glycerin gelatine, hydrogenated vegetable oil, the poly- second of various molecular weightThe mixture of the fatty acid ester of glycol and polyethylene glycol.
As it is well known to the skilled in the art, the dosage of medicine depends on many factors, including it is but and non-limitingIn following factor：The activity of particular compound used, the age of patient, the body weight of patient, the health status of patient, the row of patientQuilt, the diet of patient, administration time, administering mode, speed, the combination of medicine of excretion etc.；In addition, optimal therapeutic modality is such asThe species of the pattern, the consumption per day of general formula compound (I) or pharmaceutically useful salt for the treatment of can be tested according to traditional therapeutic schemeCard.
Detailed description of the invention
Unless stated to the contrary, the term for using in the specification and in the claims has following implications.
Term " alkyl " refers to saturated aliphatic hydrocarbons group, its be comprising 1 to 20 straight or branched group of carbon atom, it is excellentChoosing contains 1 to 12 alkyl of carbon atom, further preferably 1 to 6 alkyl of carbon atom.Non-limiting examples include methyl,Ethyl, n-propyl, isopropyl, normal-butyl, isobutyl group, the tert-butyl group, sec-butyl, n-pentyl, 1,1- dimethyl propyls, 1,2- diformazansBase propyl group, 2,2- dimethyl propyls, 1- ethyl propyls, 2- methyl butyls, 3- methyl butyls, n-hexyl, 1- Ethyl-2-Methyls thirdBase, 1,1,2- thmethylpropyls, 1,1- dimethylbutyls, 1,2- dimethylbutyls, 2,2- dimethylbutyls, 1,3- dimethyl butyratesBase, 2- ethyl-butyls, 2- methyl amyls, 3- methyl amyls, 4- methyl amyls, 2,3- dimethylbutyls, n-heptyl, 2- methyl oneselfBase, 3- methylhexyls, 4- methylhexyls, 5- methylhexyls, 2,3- dimethyl amyl groups, 2,4- dimethyl amyl groups, 2,2- dimethylAmyl group, 3,3- dimethyl amyl groups, 2- ethyl pentyl groups, 3- ethyl pentyl groups, n-octyl, 2,3- dimethylhexanyls, 2,4- dimethyl oneselfBase, 2,5- dimethylhexanyls, 2,2- dimethylhexanyls, 3,3- dimethylhexanyls, 4,4- dimethylhexanyls, 2- ethylhexyls, 3-Ethylhexyl, 4- ethylhexyls, 2- methyl -2- ethyl pentyl groups, 2- methyl -3- ethyl pentyl groups, n-nonyl, 2- methyl -2- ethylsHexyl, 2- methyl -3- ethylhexyls, 2,2- diethyl amyl groups, positive decyl, 3,3- diethylhexyls, 2,2- diethylhexyls, andIts various branched chain isomer etc..More preferably containing 1 to 6 low alkyl group of carbon atom, non-limiting example includes firstBase, ethyl, n-propyl, isopropyl, normal-butyl, isobutyl group, the tert-butyl group, sec-butyl, n-pentyl, 1,1- dimethyl propyls, 1,2-Dimethyl propyl, 2,2- dimethyl propyls, 1- ethyl propyls, 2- methyl butyls, 3- methyl butyls, n-hexyl, 1- ethyl -2- firstBase propyl group, 1,1,2- thmethylpropyls, 1,1- dimethylbutyls, 1,2- dimethylbutyls, 2,2- dimethylbutyls, 1,3- diformazansBase butyl, 2- ethyl-butyls, 2- methyl amyls, 3- methyl amyls, 4- methyl amyls, 2,3- dimethylbutyls etc..Alkyl can be withIt is substitution or non-substituted, when substituted, substitution base can be substituted on any usable tie point, the substitutionBase is preferably one or more following groups, its independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio group, alkyl amino,Halogen, sulfydryl, hydroxyl, nitro, cyano group, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, cycloalkyloxy, heterocyclylalkoxy groups, cycloalkanesSulfenyl, heterocycle alkylthio group, oxo base, carboxyl or carboxylic acid ester groups.
Term " alkylidene " refers to that a hydrogen atom of alkyl is further substituted, for example：" methylene " refers to-CH2-, it is " sub-Ethyl " refers to-(CH2)2-, " propylidene " refer to-(CH2)3-, " butylidene " refer to-(CH2)4- etc..
Term " alkenyl " refers to the alkane as defined above by being at least made up of two carbon atoms and at least one carbon-to-carbon double bondBase, such as vinyl, 1- acrylic, 2- acrylic, 1-, 2- or 3- cyclobutenyl etc..Alkenyl can be substitution or non-substituted,When substituted, substitution base be preferably one or more following groups, its independently selected from alkyl, alkenyl, alkynyl, alkoxy,Alkylthio group, alkyl amino, halogen, sulfydryl, hydroxyl, nitro, cyano group, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, cycloalkanes oxygenBase, heterocyclylalkoxy groups, cycloalkylthio, heterocycle alkylthio group.
Term " alkynyl " turns into alkynes containing C ≡ C in referring to molecule not comprising alkynes, and row are such as：Acetylene, propine, 1- butine, 2-Butine, 3- methyl isophthalic acids-butine or valerylene etc..Alkynyl can be substitution or non-substituted, and when substituted, substitution base is preferredBe one or more following groups, its independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio group, alkyl amino, halogen,Sulfydryl, hydroxyl, nitro, cyano group, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, cycloalkyloxy, heterocyclylalkoxy groups, cycloalkylthio,Heterocycle alkylthio group.
Term " cycloalkyl " refers to saturation or part is unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent, cycloalkyl ring comprising 3 to20 carbon atoms, preferably comprise 3 to 12 carbon atoms, more preferably comprising 3 to 6 carbon atoms.Monocyclic cycloalkyl it is non-limitingExample includes cyclopropyl, cyclobutyl, cyclopenta, cyclopentenyl, cyclohexyl, cyclohexenyl group, cyclohexadienyl, suberyl, cycloheptylTrialkenyl, cyclooctyl etc.；Polycyclic naphthene base includes the cycloalkyl of volution, condensed ring and bridged ring, preferably indanyl.
Term " spiro cycloalkyl group " shares a polycyclic moiety for carbon atom (title spiro-atom) between referring to 5 to 20 yuan monocyclic,It can contain one or more double bonds, but neither one ring has the pi-electron system of total conjugated.Preferably 6 to 14 yuan, morePreferably 7 to 10 yuan.Spiro cycloalkyl group is divided into single spiro cycloalkyl group, double spirocyclanes by the number according to spiro-atom is shared between ring and ringBase or many spiro cycloalkyl groups, preferably single spiro cycloalkyl group and double spiro cycloalkyl groups.More preferably 4 yuan/4 yuan, 4 yuan/5 yuan, 4 yuan/6 yuan, 5Unit/5 yuan or 5 yuan/6 yuan single spiro cycloalkyl groups.The non-limiting examples of spiro cycloalkyl group include：
Term " cycloalkyl " refers to 5 to 20 yuan, and each ring in system shares a pair for adjoining with other rings in systemThe full carbon polycyclic moiety of carbon atom, wherein one or more rings can contain one or more double bonds, but neither one ring hasThe pi-electron system of total conjugated.Preferably 6 to 14 yuan, more preferably 7 to 10 yuan.Number according to composition ring can be divided into doubleRing, three rings, Fourth Ring or polycyclic fused ring alkyl, preferably bicyclic or three rings, more preferably 5 yuan/5 yuan or 5 yuan/6 membered bicyclic alkyl.The non-limiting examples of cycloalkyl include：
Term " bridge ring alkyl " refers to 5 to 20 yuan, and the full carbon that any two ring shares two carbon atoms being not directly connected is moreCyclic group, it can contain one or more double bonds, but neither one ring has the pi-electron system of total conjugated.Preferably 6 to14 yuan, more preferably 7 to 10 yuan.Number according to composition ring can be divided into bicyclic, three rings, Fourth Ring or polycyclic bridge ring alkyl, excellentElect bicyclic, three rings or Fourth Ring as, more elect bicyclic or three rings as.The non-limiting examples of bridge ring alkyl include：
The cycloalkyl ring can be condensed on aryl, heteroaryl or heterocycloalkyl ring, wherein being connected to precursor structureRing together is cycloalkyl, and non-limiting examples are including indenyl, tetralyl, benzocyclohepta alkyl etc..Cycloalkyl can be appointedSelection generation or non-substituted, when substituted, substitution base is preferably one or more following groups, and it is independently selected from alkaneBase, alkenyl, alkynyl, alkoxy, alkylthio group, alkyl amino, halogen, sulfydryl, hydroxyl, nitro, cyano group, cycloalkyl, Heterocyclylalkyl,Aryl, heteroaryl, cycloalkyloxy, heterocyclylalkoxy groups, cycloalkylthio, heterocycle alkylthio group, oxo base, carboxyl or carboxylic acid ester groups.
Term " heterocyclic radical " refers to the unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent of saturation or part, and it includes 3 to 20 ringsAtom, wherein one or more annular atoms are selected from nitrogen, oxygen or S (O)mThe hetero atom of (wherein m is integer 0 to 2), but do not wrapThe loop section of-O-O- ,-O-S- or-S-S- is included, remaining annular atom is carbon.3 to 12 annular atoms are preferably comprised, wherein 1~4It is hetero atom；3 to 8 annular atoms are most preferably comprised, wherein 1~3 is hetero atom；5 to 6 annular atoms are most preferably comprised, itsIn 1~2 or 1~3 be hetero atom.The non-limiting examples of monocyclic heterocycles base include pyrrolidinyl, imidazolidinyl, tetrahydrofuranBase, tetrahydro-thienyl, glyoxalidine base, dihydrofuran base, pyrazoline base, pyrrolin base, piperidyl, piperazinyl, morpholineBase, thio-morpholinyl, homopiperazine base, pyranose, azetidinyl etc., preferably 1,2,5- oxadiazolyls, pyranose, piperidinesBase, azetidinyl, pyrrolidinyl or morpholinyl.Multiring heterocyclic includes the heterocyclic radical of volution, condensed ring and bridged ring.
Term " spiro heterocyclic radical " shares a multiring heterocyclic for atom (title spiro-atom) between referring to 5 to 20 yuan monocyclicGroup, wherein one or more annular atoms are selected from nitrogen, oxygen or S (O)mThe hetero atom of (wherein m is integer 0 to 2), remaining annular atomIt is carbon.It can contain one or more double bonds, but neither one ring has the pi-electron system of total conjugated.Preferably 6 to 14Unit, more preferably 7 to 10 yuan.Spiro heterocyclic radical is divided into single spiro heterocyclic radical, double by the number according to spiro-atom is shared between ring and ringSpiro heterocyclic radical or many spiro heterocyclic radicals, preferably single spiro heterocyclic radical and double spiro heterocyclic radicals.More preferably 4 yuan/4 yuan, 4 yuan/5 yuan, 4Unit/6 yuan, 5 yuan/5 yuan or 5 yuan/6 yuan single spiro heterocyclic radicals.The non-limiting examples of spiro heterocyclic radical include：
Term " condensed hetero ring base " refers to 5 to 20 yuan, and each ring in system shares a pair for adjoining with other rings in systemThe polycyclic heterocyclic group of atom, one or more rings can contain one or more double bonds, but neither one ring with completely commonThe pi-electron system of yoke, wherein one or more annular atoms are selected from nitrogen, oxygen or S (O)mThe miscellaneous original of (wherein m is integer 0 to 2)Son, remaining annular atom is carbon.Preferably 6 to 14 yuan, more preferably 7 to 10 yuan.According to composition ring number can be divided into it is bicyclic,Three rings, Fourth Ring or polycyclic condensed hetero ring base, preferably bicyclic or three rings, more preferably 5 yuan/5 yuan or 5 yuan/6 membered bicyclic condensed hetero ringsBase.The non-limiting examples of condensed hetero ring base include：
Term " bridge heterocyclic radical " refers to 5 to 14 yuan, and any two ring shares two polycyclic heterocycles of the atom being not directly connectedGroup, it can contain one or more double bonds, but neither one ring has the pi-electron system of total conjugated, one of them orMultiple annular atoms are selected from nitrogen, oxygen or S (O)mThe hetero atom of (wherein m is integer 0 to 2), remaining annular atom is carbon.Preferably 6To 14 yuan, more preferably 7 to 10 yuan.Number according to composition ring can be divided into bicyclic, three rings, Fourth Ring or polycyclic bridge heterocyclic radical,Preferably bicyclic, three rings or Fourth Ring, more elect bicyclic or three rings as.The non-limiting examples of bridge heterocyclic radical include：
The heterocyclic ring can be condensed on aryl, heteroaryl or cycloalkyl ring, wherein being connected to one with precursor structureThe ring for rising is heterocyclic radical, and its non-limiting examples includes：
Deng.
Heterocyclic radical can be it is optionally substituted or non-substituted, when substituted, substitution base be preferably one or more withLower group, its independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio group, alkyl amino, halogen, sulfydryl, hydroxyl, nitro,Cyano group, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, cycloalkyloxy, heterocyclylalkoxy groups, cycloalkylthio, heterocycle alkylthio group, oxoBase, carboxyl or carboxylic acid ester groups.
Term " aryl " refers to 6 to 14 yuan with the pi-electron system being conjugated, and full carbon is monocyclic or fused polycycle (is namely sharedThe ring of adjacent carbon atoms pair) group, preferably 6 to 10 yuan, such as phenyl and naphthyl.More preferably phenyl.The aryl rings can be withCondense on heteroaryl, heterocyclic radical or cycloalkyl ring, wherein the ring linked together with precursor structure is aryl rings, its is unrestrictedProperty example includes：
Aryl can be substitution or non-substituted, and when substituted, substitution base is preferably one or more following groups,It is independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio group, alkyl amino, halogen, sulfydryl, hydroxyl, nitro, cyano group, ringAlkyl, Heterocyclylalkyl, aryl, heteroaryl, cycloalkyloxy, heterocyclylalkoxy groups, cycloalkylthio, heterocycle alkylthio group, carboxyl or carboxylic acidEster group.
Term " heteroaryl " refers to the heteroaromatic system comprising 1 to 4 hetero atom, 5 to 14 annular atoms, wherein hetero atom choosingFrom oxygen, sulphur and nitrogen.Heteroaryl is preferably 5 to 10 yuan, containing 1 to 3 hetero atom；More preferably 5 yuan or 6 yuan, containing 1 to 2 miscellaneous originalSon；It is preferred that such as imidazole radicals, furyl, thienyl, thiazolyl, pyrazolyl, oxazolyl, pyrrole radicals, tetrazole radical, pyridine radicals, phoneticPiperidinyl, thiadiazoles, pyrazinyl etc., preferably imidazole radicals, thiazolyl, pyrazolyl or pyrimidine radicals, thiazolyl；More select pyrazolyl orThiazolyl.The heteroaryl ring can be condensed on aryl, heterocyclic radical or cycloalkyl ring, wherein being linked together with precursor structureRing be heteroaryl ring, its non-limiting examples includes：
Heteroaryl can be it is optionally substituted or non-substituted, when substituted, substitution base be preferably one or more withLower group, its independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio group, alkyl amino, halogen, sulfydryl, hydroxyl, nitro,Cyano group, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, cycloalkyloxy, heterocyclylalkoxy groups, cycloalkylthio, heterocycle alkylthio group, carboxylOr carboxylic acid ester groups.
Term " alkoxy " refers to-O- (alkyl) and-O- (non-substituted cycloalkyl), wherein alkyl, cycloalkyl definition such asIt is upper described.The non-limiting examples of alkoxy include：Methoxyl group, ethyoxyl, propoxyl group, butoxy, ring propoxyl group, ring fourth oxygenBase, cyclopentyloxy, cyclohexyloxy.Alkoxy can be optionally substituted or non-substituted, and when substituted, substitution base is preferablyOne or more following groups, it is independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio group, alkyl amino, halogen, mercaptoIt is base, hydroxyl, nitro, cyano group, cycloalkyl, Heterocyclylalkyl, aryl, heteroaryl, cycloalkyloxy, heterocyclylalkoxy groups, cycloalkylthio, miscellaneousCycloalkylthio, carboxyl or carboxylic acid ester groups.
Term " haloalkyl " refers to the alkyl replaced by one or more halogens, wherein alkyl as defined above.
Term " halogenated alkoxy " refers to the alkoxy replaced by one or more halogens, wherein alkoxy as defined above.
Term " hydroxyalkyl " refers to the alkyl being optionally substituted by a hydroxyl group, wherein alkyl as defined above.
Term " hydroxyl " refers to-OH groups.
Term " halogen " refers to fluorine, chlorine, bromine or iodine.
Term " amino " refers to-NH2。
Term " cyano group " refers to-CN.
Term " nitro " refers to-NO2。
Term " oxo base " refers to=O.
Term " carbonyl " refers to C=O.
Term " carboxyl " refers to-C (O) OH.
Term " isocyanate group " refers to-NCO.
Term " oximido " refers to=N-OH.
Term " sulfydryl " refers to-SH.
Term " carboxylic acid ester groups " refers to-C (O) O (alkyl) or-C (O) O (cycloalkyl), and wherein alkyl, cycloalkyl is as above determinedJustice.
Term " carboxylic acid halides " refers to the compound of the group containing-C (O)-halogen.
" optional " or " optionally " mean event described later or environment can with but need not occur, the explanation includesThe occasion that the event or environment occur or do not occur.For example, " optionally by alkyl-substituted heterocyclic group " means that alkyl can be withBut necessarily exist, the explanation includes heterocyclic group by alkyl-substituted situation and heterocyclic group not by alkyl-substituted situation.
" substituted " refers to one or more hydrogen atoms in group, preferably at most 5, more preferably 1~3 hydrogen atomReplaced by the substitution base of respective number independently of one another.Self-evident, substitution base is only in their possible chemical position, thisArt personnel can determine that (by experiment or theoretical) may or impossible take in the case where excessive effort is not paidGeneration.For example, being probably unstable when amino or hydroxyl with free hydrogen are with the carbon atom combination with unsaturated (such as olefinic) keyFixed.
" pharmaceutical composition " represent containing one or more compound described herein or its physiologically/pharmaceutically useful salt orThe mixture of pro-drug and other chemical constituents, and other components such as physiology/pharmaceutically useful carrier and excipient.MedicineThe purpose of compositions is administration of the promotion to organism, absorption and then performance bioactivity beneficial to active component.
" officinal salt " refers to the salt of the compounds of this invention, and this kind of salt has security and has when being used in mammal bodyEffect property, and with due bioactivity.
" X is selected from A, B or C ", " X is selected from A, B and C ", " X is A, B or C ", " X is A, B and C " etc. are different in the present invention usesLanguage expresses identical meaning, that is, represent X can be in A, B, C any one or a few.
The synthetic method of the compounds of this invention
In order to complete the purpose of the present invention, the present invention is adopted the following technical scheme that：
Compound or its dynamic isomer, mesomer, racemic modification, enantiomter shown in the logical formula (I) of the present invention,Diastereoisomer or its form of mixtures, or its pharmaceutically useful salt preparation method, comprise the following steps：
In the case where acid condition is heated, formula (A) compound is oxidized and obtains logical formula (III) compound；The formula for obtaining(III) compound obtains logical formula (I) compound under room temperature alkalescence condition with the reaction of logical formula (VI)；Or the formula for obtaining(III) compound obtains logical formula (V) compound under room temperature alkalescence condition with the reaction of logical formula (VI)；The logical formula (V) chemical combination for obtainingOpen loop obtains logical formula (I) compound to thing in the basic conditions.
The reagent for providing alkalescence condition includes organic base and inorganic base, and described organic bases include but is not limited to three secondAmine, pyridine, DIPEA, n-BuLi, lithium diisopropylamine, potassium acetate, sodium tert-butoxide or potassium tert-butoxide, instituteThe inorganic base stated including but not limited to sodium hydride, potassium phosphate, sodium carbonate, potassium carbonate or cesium carbonate.The examination of alkalescence condition is providedAgent is preferably NaOH.
Oxidant used is included but is not limited to：Selenium dioxide, hydrogen peroxide, potassium permanganate or manganese dioxide, preferably 30%Hydrogen peroxide solution.
The reagent for providing acid condition includes but is not limited to trifluoroacetic acid, formic acid, acetic acid, hydrochloric acid, hydrochloric acid dioxane, sulphurAcid or methanesulfonic acid, preferably trifluoroacetic acid.
Above-mentioned reaction is preferably carried out in a solvent, and solvent for use is included but is not limited to：Acetic acid, methyl alcohol, ethanol, toluene, fourHydrogen furans, dichloromethane, petroleum ether, ethyl acetate, n-hexane, dimethyl sulfoxide (DMSO), 1,4- dioxane, water, N, N- dimethylFormamide and its mixture；
Wherein：
Ring A, ring B, R1、R2, x and n defines as described in logical formula (I).
Specific embodiment
The present invention is further described with reference to embodiments, but these embodiments not limit the scope of the present invention.
Embodiment
The structure of compound is determined by nuclear magnetic resonance (NMR) or mass spectrum (MS).The measure of NMR is to use BrukerAVANCE-400 nuclear magnetic resonance spectrometers, measure solvent is deuterated dimethyl sulfoxide (DMSO-d6), deuterochloroform (CDCl3), deuterated methanol(CD3OD), tetramethylsilane (TMS) is inside designated as, chemical shift is with 10-6(ppm) be given as unit.
The measure of MS is with FINNIGAN LCQAd (ESI) mass spectrograph (manufacturer：Thermo, model：Finnigan LCQadvantage MAX)。
The measure of HPLC uses Agilent 1200DAD high pressure liquid chromatographs (Sunfire C18 150 × 4.6mm chromatogramsPost) and Waters 2695-2996 high pressure liquid chromatographs (Gimini 150 × 4.6mm of C18 chromatographic columns).
Kinases average inhibition and IC50The measure of value is with NovoStar ELIASAs (German BMG companies).
Tlc silica gel plate uses Yantai Huanghai Sea HSGF254 or Qingdao GF254 silica gel plates, and thin-layered chromatography (TLC) makesThe specification that silica gel plate is used is 0.15mm~0.2mm, the specification that thin-layer chromatography isolates and purifies product use be 0.4mm~0.5mm silica gel plates.
Column chromatography generally uses the mesh silica gel of the Yantai Huanghai Sea 200~300 for carrier.
Known initiation material of the invention can be used or synthesized according to methods known in the art, or it is commercially available fromABCR GmbH＆Co.KG, Acros Organnics, Aldrich Chemical Company, splendid remote chemistry science and technology (AccelaChemBio Inc), up to companies such as auspicious chemicals.
In embodiment unless otherwise specified, reaction is carried out under argon atmospher or blanket of nitrogen.
Argon atmospher or blanket of nitrogen refer to that reaction bulb connects the argon gas or nitrogen balloon of about 1L volumes.
Nitrogen atmosphere refers to that reaction bulb connects a hydrogen balloon for about 1L volumes.
Pressure hydration reaction hydrogenates instrument and clear indigo plant QL-500 types hydrogen generator or HC2-SS using Parr 3916EKX typesType hydrogenates instrument.
Hydrogenation is generally vacuumized, and is filled with hydrogen, is operated 3 times repeatedly.
Microwave reaction uses the type microwave reactors of CEM Discover-S 908860.
In embodiment unless otherwise specified, the solution in reaction refers to the aqueous solution.
In embodiment unless otherwise specified, the temperature of reaction is room temperature.
Room temperature is optimum reaction temperature, and temperature range is 20 DEG C~30 DEG C.
The monitoring of the reaction process in embodiment uses thin-layered chromatography (TLC), reacts the system of used solventHave：A：Dichloromethane and methanol system, B：N-hexane and ethyl acetate system, C：Petroleum ether and ethyl acetate system, D：Acetone,The volume ratio of solvent is adjusted according to the polarity difference of compound.
The system of the solvent of the system and thin-layered chromatography of the eluant, eluent of the column chromatography that purifying compound is used includes：A：Dichloromethane and methanol system, B：N-hexane and ethyl acetate system, C：N-hexane, ethyl acetate and dichloromethane system, D：Petroleum ether and ethyl acetate system, E：Ethyl acetate, the volume ratio of solvent is adjusted according to the polarity difference of compound,A small amount of triethylamine and acid or alkaline reagent etc. can be added to be adjusted.
Embodiment 1
N- (3- chlorphenyls)-N'- hydroxyls -4- (((1R, 4R) -4- (sulphamoylamino) cyclohexyl) amino) -1,2,5-Oxadiazole -3- carbonamidines
The first step
4- (3- chlorphenyls) -3- (4- nitro -1,2,5- oxadiazole -3- bases) (4H) -one of -1,2,4- oxadiazoles -5 1b
By 3- (4- amino -1,2,5- oxadiazole -3- bases) -4- (3- chlorphenyls) (4H) -one of -1,2,4- oxadiazoles -5 1a(2.0g, 7.15mmol are prepared using method disclosed in patent application " WO2006122150 ") is dissolved in 5mL trifluoroacetic acidsIn, the hydrogenperoxide steam generator of 5mL 30% is added, it is warming up to 45 DEG C and stirs 12 hours.Reaction solution is cooled to 25 DEG C, adds 20mLEthyl acetate dilutes, and adds 20mL saturated sodium thiosulfate solution that reaction is quenched, and point liquid, water is mutually extracted with ethyl acetate (15mL× 2), and merging organic phase, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, residue high performance liquid chromatography is purified,Obtain title product 1b (600mg, yellow oil), yield 40%.
MS m/z(LC-MS):279.0[M-30]
Second step
((1R, 4R) -4- ((4- (4- (3- chlorphenyls) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- oxadiazole -3- bases) amino) cyclohexyl) t-butyl carbamate 1c
By 4- (3- chlorphenyls) -3- (4- nitro -1,2,5- oxadiazole -3- bases) (4H) -one of -1,2,4- oxadiazoles -5 1b(135mg, 0.433mmol) is dissolved in 3mL tetrahydrofurans, adds ((1r, 4r) -4- aminocyclohexyls) t-butyl carbamate(206mg, 0.962mmol, Shanghai Bepharm Science ＆ Technology Co., Ltd.), adds the sodium hydroxide solution of 1.5mL 2.5M, stirringReaction 30 minutes.After reaction terminates, reaction solution is extracted with ethyl acetate (30mL) with 1N salt acid for adjusting pH to 2, merged organicPhase, with anhydrous sodium sulfate drying, filtering, filtrate decompression concentration obtains crude title product 1c (50mg, yellow solid), productIt is not purified directly to carry out next step reaction.
MS m/z(LC-MS):475.2[M-1]
3rd step
3- (4- (((1R, 4R) -4- aminocyclohexyls) amino) -1,2,5- oxadiazole -3- bases) -4- (3- chlorphenyls) -1,(4H) -one of 2,4- oxadiazoles -5 1d
Will ((1R, 4R) -4- ((4- (4- (3- chlorphenyls) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- oxadiazole -3- bases) amino) cyclohexyl) t-butyl carbamate 1c (50mg, 0.1048mmol) is dissolved in 3mL dichloromethaneIn, add 0.5mL trifluoroacetic acids, stirring reaction 12 hours.After reaction terminates, reaction solution is concentrated under reduced pressure, obtains crude titleProduct 1d (60mg, yellow oil), product is not purified directly to carry out next step reaction.MS m/z(LC-MS):377.1[M+1]
4th step
N- [4- ({ 4- [4- (3- chlorphenyls) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases] -1,2,5- Evil bis-Azoles -3- bases } amino) cyclohexyl] sulfamide 1e
By crude product 3- (4- (((1R, 4R) -4- aminocyclohexyls) amino) -1,2,5- oxadiazole -3- bases) -4- (3- chlorobenzenesBase) (4H) -one 1d (60mg, 0.105mmol) of -1,2,4- oxadiazole -5 is dissolved in 2.5mL pyridines, adds sulfamide(60mg, 0.63mmol), in 130 DEG C of microwave reactions 15 minutes.After reaction terminates, reaction solution is concentrated under reduced pressure, obtains crude product markTopic product 1e (100mg, yellow solid), product is not purified directly to carry out next step reaction.
MS m/z(ESI):454.2[M-1]
5th step
N- (3- chlorphenyls)-N'- hydroxyls -4- (((1R, 4R) -4- (sulphamoylamino) cyclohexyl) amino) -1,2,5-Oxadiazole -3- carbonamidines 1
By crude product N- [4- (4- [4- (3- chlorphenyls) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases] -1,2,5- oxadiazole -3-y bases } amino) cyclohexyl] sulfamide 1e (100mg, 0.105mmol) is dissolved in 5mL tetrahydrofurans, plusEnter the sodium hydroxide solution of 1mL 2N, stirring reaction 30 minutes.After reaction terminates, reaction solution to acidity, decompression are adjusted with acetic acidConcentration, with high performance liquid chromatography purify obtained by residue, obtain title product 1 (12mg, white solid), yield 25%.
MS m/z(ESI):430.3[M+1]
1H NMR(400MHz,DMSO-d6)δ11.56(s,1H),8.94(s,1H),7.18(t,1H),6.94(dd,1H),6.83(m,1H),6.67(dd,1H),6.45-6.55(m,3H),6.01(d,1H),3.20-3.27(m,1H),3.05-3.10(m,1H),1.95-2.05(m,4H),1.23-1.32(m,4H)
Embodiment 2
N- (the bromo- 4- fluorophenyls -4- of 3- ((2,3- dihydro -1H- indenes -2- bases) amino)-N'- hydroxyl -1,2,5- oxadiazoles -3- carbonamidines
The first step
4- (the bromo- 4- fluorophenyls of 3-) -3- (4- nitro -1,2,5- oxadiazole -3- bases) -1,2,4- oxadiazoles -5 (4H) -one2b
By 3- (4- amino -1,2,5- oxadiazole -3- bases) -4- (the bromo- 4- fluorophenyls of 3-) -1,2,4- oxadiazoles -5 (4H) -Ketone 2a (13.0g, 41.1mmol are prepared using method disclosed in patent application " WO2014066834 ") adds 150mL tri-In fluoroacetic acid, 90mL hydrogen peroxide solutions (30%) is added, reacted 48 hours in 45 DEG C.After reaction terminates, cooling adds 300mLSaturated sodium thiosulfate solution and 150mL ethyl acetate, stirring reaction 20 minutes are detected without peroxide with potassium iodide starch paper.Divide liquid, water to be mutually extracted with ethyl acetate (100mL × 2), merge organic phase, with anhydrous sodium sulfate drying, filtering, filtrate decompression is denseContracting, with silica gel column chromatography with eluant, eluent system B purify obtained by residue, obtain title product 2b (4.5g, yellow-brown solid),Yield 30%.
MS m/z(LC-MS):342.9[M-30]
Second step
N- (the bromo- 4- fluorophenyls -4- of 3- ((2,3- dihydro -1H- indenes -2- bases) amino)-N'- hydroxyl -1,2,5- oxadiazoles -3- carbonamidines 2
By 4- (the bromo- 4- fluorophenyls of 3-) -3- (4- nitro -1,2,5- oxadiazole -3- bases) -1,2,4- oxadiazoles -5 (4H) -Ketone 2b (286mg, 0.77mmol), 2,3- dihydros -1H- indenes -2- amine 2c (123mg, 0.922mmol) are dissolved in 5mL tetrahydrofurans,Add the sodium hydroxide solution of 0.924mL 2.5M, stirring reaction 4 hours.After reaction terminates, 20mL ethyl acetate and 5mL are addedWater, point liquid, water is mutually extracted with ethyl acetate, and merges organic phase, and with anhydrous sodium sulfate drying, filtering, silicon is used in filtrate decompression concentrationGlue column chromatography purifies gained residue with eluant, eluent system B, obtains title product 2 (100mg, yellow solid), yield 30%.
MS m/z(ESI):432.3/434.3[M+1]
1H NMR(400MHz,DMSO-d6)δ11.44(s,1H),8.88(s,1H),7.25-7.27(m,2H),7.11-7.19(m,4H),6.77-6.80(m,1H),6.35-6.36(d,1H),4.28-4.33(m,1H),3.28-3.34(m,2H),2.9-2.95(m,2H).
Embodiment 3
N- (the bromo- 4- fluorophenyls-N'- hydroxyls -4- of 3- ((1- methyl isophthalic acid H- pyrazoles -4- bases) amino) -1,2,5- oxadiazoles -3- carbonamidines
By 4- (the bromo- 4- fluorophenyls of 3-) -3- (4- nitro -1,2,5- oxadiazole -3- bases) -1,2,4- oxadiazoles -5 (4H) -Ketone 2b (100mg, 0.27mmol), 1- methyl isophthalic acid H- pyrazoles -4- amine 3a (26.1mg, 0.27mmol) are dissolved in 5mL tetrahydrofurans,Add the sodium hydroxide solution of 32.4mL 2.5M, stirring reaction 1.5 hours.After reaction terminates, reaction solution is poured into frozen water,With 1N salt acid for adjusting pH to neutrality, it is extracted with ethyl acetate, anhydrous sodium sulfate drying, filters, thin layer color is used in filtrate decompression concentrationSpectrometry purifies gained residue with eluant, eluent system A, obtains title product 3 (10mg, yellow solid), yield 10%.
MS m/z(ESI):396.3/398.3[M+1]
1H NMR(400MHz,DMSO-d6)δ11.52(s,1H),9.02(s,1H),8.61-8.62(m,1H),7.79-7.80(m,1H),7.46-7.48(m,1H),7.17-7.19(m,1H),6.78-6.79(m,1H),5.75(s,1H),3.82(s,3H).
Embodiment 4
N- (the bromo- 4- fluorophenyls of 3-) -4- (clopentylamino)-N'- hydroxyl -1,2,5- oxadiazole -3- carbonamidines
By 4- (the bromo- 4- fluorophenyls of 3-) -3- (4- nitro -1,2,5- oxadiazole -3- bases) -1,2,4- oxadiazoles -5 (4H) -Ketone 2b (100mg, 0.27mmol), pentamethylene amine 4a (28mg, 0.32mmol) is dissolved in 20mL tetrahydrofurans, adds 0.32mLThe sodium hydroxide solution of 2.5M, stirring reaction 1 hour.After reaction terminates, reaction solution is poured into 30mL water, extracted with ethyl acetateTake (30mL × 2), organic phase is washed with saturated ammonium chloride solution successively, saturated nacl aqueous solution washing, anhydrous sodium sulfate drying,Filtering, filtrate decompression concentration, with thin-layered chromatography with eluant, eluent system B purify obtained by residue, obtain title product 4 (50mg,White solid), yield 48.5%.MS m/z(ESI):384.3/386.3[M+1]
1H NMR(400MHz,DMSO-d6)δ11.43(s,1H),8.86(s,1H),7.10-7.20(m,2H),6.75-6.78(m,1H),6.08-6.09(m,1H),3.76-3.80(m,1H),1.89-1.98(m,2H),1.23-1.64(m,6H).
Embodiment 5
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- (((1R, 4R) -4- hydroxy-cyclohexyls) amino) -1,2,5- Evil bis-Azoles -3- carbonamidines
By 4- (the bromo- 4- fluorophenyls of 3-) -3- (4- nitro -1,2,5- oxadiazole -3- bases) -1,2,4- oxadiazoles -5 (4H) -Ketone 2b (100mg, 0.27mmol), (1R, 4R) -4- aminocyclohexanols 5a (37.15mg, 0.32mmol) are dissolved in 5mL tetrahydrofuransIn, add the sodium hydroxide solution of 0.3mL 2.5M, stirring reaction 3 hours.After reaction terminates, add 20mL saturated ammonium chlorides moltenLiquid, is extracted with ethyl acetate (15mL × 3), merges organic phase, is washed with saturated nacl aqueous solution, anhydrous sodium sulfate drying, mistakeFilter, filtrate decompression concentration, with thin-layered chromatography with eluant, eluent system A purify obtained by residue, obtain title product 5 (20mg, it is shallowYellow solid), yield 14.9%.
MS m/z(ESI):414.3/416.3[M+1]
1H NMR(400MHz,DMSO-d6)δ11.48(s,1H),8.88(s,1H),7.17-7.21(m,1H),7.10-7.13(m,1H),6.76-6.79(m,1H),5.98-6.00(m,1H),4.57-4.58(m,1H),3.43-3.46(m,1H),3.25-3.27(m,1H),1.99-2.01(m,2H),1.81-1.83(m,2H),1.23-1.32(m,4H)
Embodiment 6
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((tetrahydrochysene -2H- pyrans -4- bases) amino) -1,2,5- oxadiazoles -3-Carbonamidine
By 4- (the bromo- 4- fluorophenyls of 3-) -3- (4- nitro -1,2,5- oxadiazole -3- bases) -1,2,4- oxadiazoles -5 (4H) -Ketone 2b (100mg, 0.27mmol), tetrahydrochysene -2H- pyrans -4- amine 6a (33mg, 0.32mmol) is dissolved in 10mL tetrahydrofurans, plusEnter the sodium hydroxide solution of 0.32mL 2.5M, stirring reaction 12 hours.After reaction terminates, reaction solution is poured into 30mL water, usedEthyl acetate extracts (30mL × 2), merges organic phase, is washed with saturated ammonium chloride solution successively, saturated nacl aqueous solution washing,Anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, with thin-layered chromatography with eluant, eluent system B purify obtained by residue, obtainTitle product 6 (45mg, white solid), yield 42.1%.
MS m/z(ESI):400.3/402.3[M+1]
1H NMR(400MHz,DMSO-d6)δ11.47(s,1H),8.89(s,1H),7.16-7.20(t,1H),7.10-7.12(m,1H),6.76-6.79(m,1H),6.15-6.16(m,1H),3.82-3.85(m,2H),3.48-3.54(m,1H),3.36-3.41(m,2H),1.91-1.94(m,2H),1.42-1.51(m,6H).
Embodiment 7,8
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- (((1s, 3s) -3- (sulphamoylamino) cyclobutyl) amino) -1,2,5- oxadiazole -3- carbonamidines 7
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- (((1r, 3r) -3- (sulphamoylamino) cyclobutyl) amino) -1,2,5- oxadiazole -3- carbonamidines 8
The first step
(3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- EvilDiazole -3- bases) amino) cyclobutyl) t-butyl carbamate 7a
By 2b (1.00g, 2.69mmol), the 3- amino cyclobutane formate tert-butyl esters (500mg, 2.69mmol, using knownMethod " ACS Medicinal Chemistry Letters, 2014,5 (6), 700-705 " is prepared) add 20mL tetrahydrochysenesIn furans, stirring reaction 2 hours.After reaction terminates, be concentrated under reduced pressure reaction solution, pure with eluant, eluent system B with silica gel column chromatographyChange gained residue, obtain title product 7a (1.0g, yellow solid), yield 72.7%.
Second step
3- (4- ((3- Aminocyclobutyls) amino) -1,2,5- oxadiazole -3- bases) -4- (the bromo- 4- fluorophenyls of 3-) -1,2,4-(4H) -one of oxadiazole -5 7b
7a (1.09g, 1.95mmol) is dissolved in the hydrochloric acid of 5mL 4M, stirring reaction 3 hours.After reaction terminates, useSaturated sodium bicarbonate solution is 7 to pH, and ethyl acetate extraction merges organic phase, anhydrous sodium sulfate drying, filtering, filtrate decompressionConcentration, obtains crude title product 7b (643mg, yellow solid), and product is not purified directly to carry out next step reaction.
3rd step
N- (3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5-Oxadiazole -3- bases) amino) cyclobutyl) sulphamoylamino t-butyl formate 7c
Crude product 7b (138mg, 0.34mmol) is dissolved in 5mL dichloromethane, addition triethylamine (9.2mg,0.092mmol), chlorosulfonyl t-butyl carbamate (0.1mL, the 0.36mmol, using known of prefabricated 5mL 2M is added dropwiseMethod " Bioorganic＆Medicinal Chemistry, 2014,22 (22), 6353-6359 " is prepared) dichloromethaneSolution, stirring reaction 1 hour.After reaction terminates, 5mL water and 20mL dichloromethane are added, point liquid, water is extracted with dichloromethaneTake, merge organic phase, be concentrated under reduced pressure, with silica gel column chromatography with eluant, eluent system B purify obtained by residue, obtain title product7c (150mg, yellow solid), yield 70%.
4th step
N- [3- ({ 4- [4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases] -1,2,5-Oxadiazole -3- bases } amino) cyclobutyl] sulfamide 7d
During 7c (150mg, 0.255mmol) added into the hydrochloric acid dioxane of 5mL 4N, stirring reaction 4 hours.Reaction knotShu Hou, be concentrated under reduced pressure, with silica gel column chromatography with eluant, eluent system B purify obtained by residue, obtain title product 7d (98mg,Yellow solid), yield 80%.
5th step
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- (((1S, 3S) -3- (sulphamoylamino) cyclobutyl) amino) -1,2,5- oxadiazole -3- carbonamidines 7
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- (((1R, 3R) -3- (sulphamoylamino) cyclobutyl) amino) -1,2,5- oxadiazole -3- carbonamidines 8
7d (98mg, 0.2mmol) is dissolved in 4mL tetrahydrofurans, the sodium hydroxide solution of 0.5mL 2N is added, stirring is anti-Answer 2 hours.After reaction terminates, water is added, be extracted with ethyl acetate, merge organic phase, anhydrous sodium sulfate drying, filtering, filtrateIt is concentrated under reduced pressure, with high performance liquid chromatography purify obtained by residue, obtain title product 7 (20mg, colorless viscous thing), yield43.5%, title product 8 (12mg, white solid), yield 26.1%.
Embodiment 7:
MS m/z(ESI):464.2[M+1]
1H NMR(400MHz,DMSO-d6)δ11.50(br,1H),8.93(s,1H),7.17-7.21(m,1H),7.11-7.14(m,1H),7.00-7.02(m,1H),6.76-6.78(m,1H),6.53(s,2H),6.44-6.45(m,1H),3.86-3.96(m,2H),2.27-2.33(m,4H).
Embodiment 8:
MS m/z(ESI):464.2[M+1]
1H NMR(400MHz,DMSO-d6):δ11.55(br,1H),8.91(s,1H),7.17-7.21(m,1H),7.10-7.11(m,1H),6.89-6.91(m,1H),6.76-6.79(m,1H),6.55(s,2H),6.28-6.29(m,1H),3.63-3.64(m,1H),3.45-3.47(m,1H),2.70-2.72(m,2H),1.90-1.93(m,2H).
Embodiment 9,10
N- (3- bromine 4- fluorophenyls)-N'- hydroxyls -4- (((1S, 3S) -3- hydroxycyclobutyls) amino) -1,2,5- oxadiazoles -3- carbonamidines 9
N- (3- bromine 4- fluorophenyls)-N'- hydroxyls -4- (((1R, 3R) -3- hydroxycyclobutyls) amino) -1,2,5- oxadiazoles -3- carbonamidines 10
The first step
4- (the bromo- 4- fluorophenyls of 3-) -3- (4- ((3- hydroxycyclobutyls) amino) -1,2,5- oxadiazole -3- bases) -1,2,4-(4H) -one of oxadiazole -5 9a
2b (42.7mg, 0.12mmol) is dissolved in tetrahydrofuran, 3- amino cyclobutanol (20mg, 0.23mmol) is added,Add the sodium hydroxide solution of 0.69mL 2.5M, stirring reaction 15 minutes.After reaction terminates, point liquid, water is mutually adjusted with 1N hydrochloric acidPH to 7, ethyl acetate extraction, merges organic phase, is washed with saturated ammonium chloride solution, anhydrous sodium sulfate drying, filters, and filtrate subtractsPressure concentration, with thin-layered chromatography with solvent system B purify obtained by residue, obtain title product 9a (40mg, white solid),Yield 42.3%.
Second step
N- (3- bromine 4- fluorophenyls)-N'- hydroxyls -4- (((1S, 3S) -3- hydroxycyclobutyls) amino) -1,2,5- oxadiazoles -3- carbonamidines 9
N- (3- bromine 4- fluorophenyls)-N'- hydroxyls -4- (((1R, 3R) -3- hydroxycyclobutyls) amino) -1,2,5- oxadiazoles -3- carbonamidines 10
9a (40mg, 0.1mmol) is dissolved in 5mL tetrahydrofurans, the sodium hydroxide solution of 0.08mL 2.5M is added, stirredMix reaction 30 minutes.After reaction terminates, point liquid, water mutually uses 1N salt acid for adjusting pH to 7, and ethyl acetate extraction merges organic phase, hasMachine is washed with saturated ammonium chloride solution, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, pure with high performance liquid chromatographyChange gained residue, obtain title product 9 (13mg, yellow solid), yield 69.4%, title product 10 (3mg, yellow solid),Yield 16.0%.
Embodiment 9
MS m/z(ESI):387.3[M+1]
1H NMR(400MHz,CDCl3)δ7.12-7.14(m,1H),7.07-7.03(t,1H),6.83-6.86(m,1H),4.41-4.44(t,1H),4.08-4.11(m,1H),2.29-2.32(t,4H).
Embodiment 10
MS m/z(ESI):387.3[M+1]
1H NMR(400MHz,DMSO-d6)δ7.11-7.13(m,1H),7.03-7.07(t,1H),6.83-6.85(m,1H),3.97-4.04(m,1H),3.54-362(m,1H),2.67-2.83(m,2H),1.83-1.90(m,2H).
Embodiment 11,12
4- (((1S, 3S) -3- Aminocyclobutyls) amino)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyl -1,2,5- Evil bis-Azoles -3- carbonamidines 11
4- (((1R, 3R) -3- Aminocyclobutyls) amino)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyl -1,2,5- Evil bis-Azoles -3- carbonamidines 12
7b (100mg, 0.2433mmol) is dissolved in 5mL tetrahydrofurans, the sodium hydroxide solution of 0.4mL 2M is added, stirredMix reaction 2 hours.After reaction terminates, water is added in reaction solution, be extracted with ethyl acetate, merge organic phase, organic phase saturationSodium chloride solution wash, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, with high performance liquid chromatography purify gained remainThing, obtains title product 11 (11mg, sticky solid), yield 23.5%, title product 12 (7mg, sticky solid), yield15.0%.
Embodiment 11
MS m/z(ESI):385.2[M+1]
1H NMR(400MHz,DMSO-d6)δ11.47(br,1H),8.96(s,1H),8.00-8.05(m,2H),7.17-7.21(m,1H),7.08-7.12(m,1H),6.73-6.79(m,1H),6.61-6.62(m,1H),4.18-4.24(m,1H),3.70-3.75(m,1H),2.30-2.45(m,4H).
Embodiment 12
MS m/z(ESI):385.2[M+1]
1H NMR(400MHz,DMSO-d6)δ11.68(br,1H),8.93(s,1H),8.18-8.20(m,2H),7.17-7.23(m,1H),7.09-7.11(m,1H),6.77-6.79(m,1H),6.46-6.48(m,1H),3.77-3.82(m,1H),3.05-3.11(m,1H),2.67-2.69(m,2H),2.11-2.16(m,2H).
Embodiment 13,14
(1S, 3S) -3- ((4- (N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxy formamidines base) -1,2,5- oxadiazole -3- bases) ammoniaBase) cyclobutylmethylamino formic acid esters 13
(1R, 3R) -3- ((4- (N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxy formamidines base) -1,2,5- oxadiazole -3- bases) ammoniaBase) cyclobutylmethylamino formic acid esters 14
The first step
3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- EvilDiazole -3- bases) amino) cyclobutyl (4- nitrobenzophenones) carbonic ester 13a
9a (250mg, 0.607mmol) is dissolved in 10mL dichloromethane, triethylamine (79.7mg, 0.789mmol) is added,The dichloromethane solution of the 4- chloroformate nitrophenyl esters (146mg, 0.728mmol) of prefabricated 10mL is added dropwise, stirring reaction 2 is smallWhen.After reaction terminates, add 2mL water quenchings to go out reaction, extracted with dichloromethane, it is concentrated under reduced pressure, with silica gel column chromatography elutingAgent system B purifying gained residues, obtain title product 13a (300mg, yellow solid), yield 86%.
Second step
3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- EvilDiazole -3- bases) amino) cyclobutylmethylamino formic acid esters 13b
By in 13a (300mg, 0.52mmol) addition 10mL tetrahydrofurans, 2mL methylamines, stirring reaction 2 hours are added.InsteadAfter should terminating, it is concentrated under reduced pressure, obtains crude title product 13b (300mg, yellow solid), product is not purified is directly carried out downSingle step reaction.
3rd step
(1S, 3S) -3- ((4- (N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxy formamidines base) -1,2,5- oxadiazole -3- bases) ammoniaBase) cyclobutylmethylamino formic acid esters 13
(1R, 3R) -3- ((4- (N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxy formamidines base) -1,2,5- oxadiazole -3- bases) ammoniaBase) cyclobutylmethylamino formic acid esters 14
Crude product 13b (300mg, 0.14mmol) is dissolved in 2mL tetrahydrofurans, adds the NaOH of 0.5mL 2N moltenLiquid, stirring reaction 2 hours.After reaction terminates, it is 7 to adjust pH with saturated ammonium chloride solution, and ethyl acetate extraction merges organicPhase, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, with high performance liquid chromatography purify obtained by residue, obtain title product13 (77.7mg, white solids), yield 77.7%, title product 14 (65mg, white solid), yield 45.9%.
Embodiment 13
MS m/z(ESI):443.3[M+1]
1H NMR(400MHz,DMSO-d6)δ11.40(br,1H),8.90(s,1H),7.16-7.20(m,1H),7.10-7.12(m,1H),6.98-6.99(m,1H),6.74-6.78(m,1H),6.47-6.48(m,1H),4.56-4.64(m,1H),3.58-3.68(m,1H),2.67-2.72(m,2H),2.54-2.55(m,3H),1.96-2.03(m,2H).
Embodiment 14
MS m/z(ESI):443.2[M+1]
1H NMR(400MHz,DMSO-d6)δ11.40(br,1H),8.91(s,1H),7.16-7.21(m,1H),7.10-7.13(m,1H),7.04-7.07(m,1H),6.76-6.79(m,1H),6.53-6.54(m,1H),4.96-4.99(m,1H),4.01-4.06(m,1H),2.54-2.55(m,3H),2.31-2.34(m,4H).
Embodiment 15
(R)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- sulfamoyls pyrrolidin-3-yl) amino) -1,2,5-Oxadiazole -3- carbonamidines 15
The first step
(R) -3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5-- oxadiazole -3- bases) amino) pyrrolidines -1- carboxylic acid tert-butyl esters 15b
(S) -3- amino-pyrrolidine -1- t-butyl formates 15a (150mg, 0.81mmol, Adama this) is dissolved in 10mL tetra-In hydrogen furans, 2b (200mg, 0.54mmol), 1.08mL 1N sodium hydroxide solutions, stirring reaction 1 hour are added.Reaction terminatesAfterwards, reaction solution is poured into 50mL saturated ammonium chloride solutions, is extracted with ethyl acetate, merge organic phase, it is molten with saturated sodium-chlorideLiquid wash, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, with silica gel column chromatography with eluant, eluent system B purify obtained by it is residualThing is stayed, title product 15b (230mg, white solid), yield 83.9% is obtained.
Second step
(R) -4- (the bromo- 4- fluorophenyls of 3-) -3- (4- (pyrrolidin-3-yl amino) -1,2,5- oxadiazole -3- bases) -1,2,(4H) -one of 4- oxadiazoles -5 15c
By in 15b (230mg, 0.45mmol) addition 10mL dichloromethane, 1mL trifluoroacetic acids are added, stirring reaction 1 is smallWhen.After reaction terminates, it is concentrated under reduced pressure, obtains crude title product 15c (200mg, yellow oil), product is not purified directlyCarry out next step reaction.
3rd step
(R)-((3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- oxadiazole -3- bases) amino) pyrrolidin-1-yl) sulfonyl) t-butyl formate 15d
Crude product 15c (200mg, 0.49mmol) is dissolved in 3mL dichloromethane, at 0 DEG C add triethylamine (148mg,1.47mmol), chlorosulfonyl t-butyl carbamate (210mg, 0.98mmol) is added, room temperature is slowly increased to, stirring reaction 1 is smallWhen.After reaction terminates, reaction solution is poured into 30mL dichloromethane, washed with saturated sodium bicarbonate solution successively, saturation chlorinationAmmonium salt solution is washed, saturated nacl aqueous solution washing, anhydrous sodium sulfate drying, filtering, and silica gel column chromatography is used in filtrate decompression concentrationGained residue is purified with eluant, eluent system A, title product 15d (80mg, white solid), yield 27.8% is obtained
4th step
(R) -3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- oxadiazole -3- bases) amino) pyrrolidines -1- sulfonamide 15e
By in 15d (80mg, 0.14mmol) addition 10mL dichloromethane, 0.5mL trifluoroacetic acids are added, stirring reaction 1 is smallWhen.After reaction terminates, reaction solution is concentrated under reduced pressure, obtains crude title product 15e (80mg, white solid), product is without pureChange directly carries out next step reaction.
5th step
(R)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- sulfamoyls pyrrolidin-3-yl) amino) -1,2,5-Oxadiazole -3- carbonamidines 15
Crude product 15e (80mg, 0.16mmol) is dissolved in 5mL tetrahydrofurans, adds the NaOH of 0.13mL 2.5N moltenLiquid, stirring reaction 1 hour.After reaction terminates, reaction solution is poured into 30mL water, be extracted with ethyl acetate, merge organic phase, usedSaturated ammonium chloride solution is washed, saturated nacl aqueous solution washing, anhydrous sodium sulfate drying, filtering, and thin layer is used in filtrate decompression concentrationChromatography purifies gained residue with solvent system A, obtains title product 15 (40mg, white solid), yield 52.6%.
MS m/z(ESI):365.2[M+1]
1H NMR(400MHz,CD3OD)δ7.09-7.14(m,1H),7.03-7.07(t,1H),6.82-6.87(m,1H),4.20(s,1H),3.52-3.560(m,1H),3.39-3.42(m,2H),2.28-2.30(m,1H),2.02-2.03(m,1H),1.87(s,1H).
Embodiment 16
(S)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- sulfamoyls pyrrolidin-3-yl) amino) -1,2,5-Oxadiazole -3- carbonamidines 16
Using the synthetic route of embodiment 15, raw material 15a is replaced with into (R) -3- amino-pyrrolidine -1- t-butyl formates,Title product 16 (15mg, pale solid), yield is obtained：57%.
MS m/z(ESI):466.2[M+1]
1H NMR(400MHz,DMSO-d6)δ11.51(s,1H),8.91(s,1H),7.11-7.20(m,2H),6.78-6.84(m,3H),6.35-6.37(m,1H),4.08-4.12(m,1H),3.42-3.45(m,1H),3.17-3.23(m,3H),3.09-3.10(m,1H),2.18-2.21(m,1H),1.88-1.92(m,1H).
Embodiment 17
4- ((1- Acetylpiperidin -4- bases) amino)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyl -1,2,5- oxadiazoles -3-Carbonamidine 17
1- (4- amino piperidine -1- bases) ethyl ketone 17a (46mg, 0.32mmol) is dissolved in 10mL tetrahydrofurans, 2b is added(100mg, 0.27mmol), adds the sodium hydroxide solution of 0.81mL 2.5N, stirring reaction 1 hour.After reaction terminates, will be anti-Answer liquid to pour into 30mL saturated ammonium chloride solutions, be extracted with ethyl acetate, merge organic phase, washed with saturated ammonium chloride solution,Saturated nacl aqueous solution is washed, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, with silica gel column chromatography with eluant, eluent bodyIt is A purifying gained residues, obtains title product 17 (40mg, white solid), yield 33.9%.
MS m/z(ESI):443.4[M+1]
Embodiment 18
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- methyl azetidine -3- bases) amino) -1,2,5- Evil bis-Azoles -3- carbonamidines 18
The first step
3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- EvilDiazole -3- bases) amino) azetidine -1- t-butyl formates 18b
By 2b (500mg, 1.34mmol) and 3- aminoazetidine -1- t-butyl formates 18a (277.7mg,It is 1.61mmol, splendid remote) add 20mL tetrahydrofurans, add the sodium hydroxide solution of 2.68mL 1N, stirring reaction 16 hours.Reaction solution is concentrated under reduced pressure, with silica gel column chromatography with eluant, eluent system B purify gained residue, obtain title product 18b(400mg, white solid), yield 60.0%.
Second step
3- (4- (azetidine -3- bases amino) -1,2,5- oxadiazole -3- bases) -4- (the bromo- 4- fluorophenyls of 3-) -1,2,(4H) -one of 4- oxadiazoles -5 18c
By in 18b (497mg, 1mmol) addition 2mL 1.4- dioxane, the hydrochloric acid dioxane of 8mL 4M, LC- are addedMS monitors complete to reaction.After reaction terminates, addition saturated sodium bicarbonate solution to pH is 7, ethyl acetate extraction (20mL ×3), merge organic phase, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, obtain crude title product 18c (329.7mg, in vainColor solid), product is not purified directly to carry out next step reaction.
3rd step
4- (the bromo- 4- fluorophenyls of 3-) -3- (4- ((1- methyl azetidine -3- bases) amino) -1,2,5- oxadiazoles -3-Base) (4H) -one of -1,2,4- oxadiazoles -5 18d
Crude product 18c (39.7mg, 0.1mmol) is dissolved in 3mL methyl alcohol, 0.2mL N, N- diisopropyl ethyl amines are added,0.5mL acetic acid is added, paraformaldehyde (10mg, 0.3mmol) is added, reacted 10 minutes, addition sodium cyanoborohydride (9.4mg,0.15mmol), stirring reaction 1 hour.After reaction terminates, add saturated sodium bicarbonate solution, water to be extracted with ethyl acetate, closeAnd organic phase, washed with saturated nacl aqueous solution, anhydrous sodium sulfate drying, to filter, silica gel column chromatography is used in filtrate decompression concentrationGained residue is purified with eluant, eluent system A, title product 18d (15mg, white solid), yield 36.5% is obtained
4th step
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- methyl azetidine -3- bases) amino) -1,2,5- Evil bis-Azoles -3- carbonamidines
18d (30mg, 0.07mmol) is dissolved in 15mL tetrahydrofurans, the sodium hydroxide solution of 1mL 2.5N is added, stirredMix reaction 1 hour.After reaction terminates, saturated ammonium chloride solution is added, be extracted with ethyl acetate, merge organic phase, use saturation chlorineChange sodium solution washing, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration is purified with silica gel column chromatography with eluant, eluent system AGained residue, obtains title product 18 (8mg, white solid), yield 28.5%.
MS m/z(ESI):385.1[M+1]
1H NMR(400MHz,CD3OD)δ7.12-7.14(m,1H),7.04-7.08(m,1H),6.83-6.87(m,1H),4.48-4.51(m,1H),4.20-4.25(m,2H),4.01-4.05(m,2H),2.83(s,3H).
Embodiment 19
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- sulfamoyl azetidine -3- bases) amino) -1,2,5-Oxadiazole -3- carbonamidines 19
The first step
(3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- EvilDiazole -3- bases) amino) azetidine -1- bases) sulfonylcarbamic acid tert-butyl ester 19a
18c (137.4mg, 0.35mmol) is dissolved in 5mL dichloromethane, triethylamine (0.195mL, 1.4mmol) is added,Add chlorosulfonyl t-butyl carbamate (75.5mg, 0.35mml), stirring reaction 1 hour.After reaction terminates, 2mL first is addedAlcohol, be concentrated under reduced pressure, with thin-layered chromatography with solvent system A purify obtained by residue, obtain title product 19a (100mg, in vainColor solid), yield 49.6%.
Second step
3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- EvilDiazole -3- bases) amino) azetidine -1- sulfonamide 19b
During 19a (100mg, 0.17mmol) added into the hydrochloric acid dioxane of 3mL 4N, stirring reaction 3 hours.Reaction knotShu Hou, is concentrated under reduced pressure, and obtains crude title product 19b (70mg, white solid), and product is not purified, and directly to carry out next step anti-Should.
3rd step
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- sulfamoyl azetidine -3- bases) amino) -1,2,5-Oxadiazole -3- carbonamidines 19
Crude product 19b (70mg, 0.147mmol) is dissolved in 5mL tetrahydrofurans, adds the NaOH of 1.5mL 2.5M moltenLiquid, stirring reaction 16 hours.After reaction terminates, saturated ammonium chloride solution is added, be extracted with ethyl acetate, merge organic phase, satisfiedWith sodium chloride solution washing, anhydrous sodium sulfate drying, filtering, filtrate decompression is concentrated, pure with solvent system A with thin-layered chromatographyChange gained residue, obtain title product 19 (12mg, white solid), yield 18.1%.
MS m/z(ESI):450.1[M+1]
1H NMR(400MHz,CDCl3)δ8.91(s,1H),7.43-7.45(m,1H),7.18-7.21(m,2H),7.00-7.04(m,1H),6.81-6.85(m,1H),4.68(s,2H),3.75-3.81(m,1H),3.14-3.16(m,1H),2.90-2.95(m,1H),2.66-2.72(m,1H),2.51-2.52(m,1H).
Embodiment 20
4- ((1- acetyl group azetidine -3- bases) amino)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyl -1,2,5- EvilDiazole -3- carbonamidines 20
The first step
3- (4- ((1- acetyl group azetidine -3- bases) amino) -1,2,5- oxadiazole -3- bases) -4- (bromo- 4- fluorine of 3-Phenyl) (4H) -one of -1,2,4- oxadiazoles -5 20a
18c (29.7mg, 0.1mmol) is dissolved in 5mL dichloromethane, triethylamine (30.4mg, 0.3mmol) is added, insteadAnswer system to be cooled to 0 DEG C, add the dichloromethane solution of prefabricated 1mL acetic anhydrides (10.2mg, 0.1mmol), it is anti-in 0 DEG C of stirringAnswer 30 minutes.Reaction terminate after, reaction solution is concentrated under reduced pressure, with thin-layered chromatography with solvent system A purify obtained by residue,Obtain title product 20a (22mg, white solid), yield 50.1%.
Second step
4- ((1- acetyl group azetidine -3- bases) amino)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyl -1,2,5- EvilDiazole -3- carbonamidines 20
20a (22mg, 0.05mmol) is dissolved in 2mL tetrahydrofurans, the sodium hydroxide solution of 0.08mL 2.5M is added,Stirring reaction 1 hour.After reaction terminates, saturated ammonium chloride solution is added, be extracted with ethyl acetate, merge organic phase, saturation chlorineChange sodium solution washing, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration purifies institute with thin-layered chromatography with solvent system AResidue is obtained, title product 20 (10mg, white solid), yield 50% is obtained.
MS m/z(ESI):413.3[M+1]
1H NMR(400MHz,CDCl3)δ10.48(s,1H),7.22-7.24(m,1H),7.02-7.07(m,1H),6.92-6.96(m,2H),6.51-6.53(m,1H),4.48-4.56(m,3H),3.96-4.09(m,2H),1.96(s,3H).
Embodiment 21
(R)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- (methyl sulphonyl) pyrrolidin-3-yl) amino) -1,2,5- oxadiazole -3- carbonamidines 21
The first step
(R) -3- ((4- (4- (the bromo- 4- fluorophenyls of 3-) -5- carbonyl -4,5- dihydro -1,2,4- oxadiazole -3- bases) -1,2,5- oxadiazole -3- bases) amino) pyrrolidines -1- t-butyl formates 21a
During 2b (200mg, 0.54mmol) and 15a (150mg, 0.81mmol) added into 10mL tetrahydrofurans, stirring reaction 2Hour.Reaction solution be concentrated under reduced pressure, with silica gel column chromatography with eluant, eluent system B purify obtained by residue, obtain title product 21a(230mg, colorless oil), yield 83.9%.
Second step
(R) -4- (the bromo- 4- fluorophenyls of 3-) -3- (4- (pyrrolidin-3-yl amino) -1,2,5- oxadiazole -3- bases) -1,2,(4H) -one of 4- oxadiazoles -5 21b
By in 21a (230mg, 0.45mmol) addition 10mL dichloromethane, 1mL trifluoroacetic acids are added, stirring reaction 12 is smallWhen.After reaction terminates, reaction solution is concentrated under reduced pressure, and obtains crude title product 21b (230mg, yellow oil), and product is without pureChange directly carries out next step reaction.
3rd step
(R) -4- (the bromo- 4- fluorophenyls of 3-) -3- (4- ((1- (methyl sulphonyl) pyrrolidin-3-yl) amino) -1,2,5- EvilDiazole -3- bases) (4H) -one of -1,2,4- oxadiazoles -5 21c
Crude product 21b (90mg, 0.22mmol) is dissolved in 10mL dichloromethane, 0 DEG C is cooled to, add triethylamine (44mg,0.44mmol), methylsufonyl chloride (30mg, 0.26mmol) is added, in 0 DEG C of stirring reaction 30 minutes.After reaction terminates, will reactLiquid is poured into saturated sodium bicarbonate solution, is extracted with dichloromethane, merges organic phase, successively with saturated sodium bicarbonate solution, is satisfiedAnd ammonium chloride solution, saturated nacl aqueous solution washing, anhydrous sodium sulfate drying, filtering, filtrate decompression concentration, obtain crude title productThing 21c (100mg, yellow solid), product is not purified directly to carry out next step reaction.
4th step
(R)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- (methyl sulphonyl) pyrrolidin-3-yl) amino) -1,2,5- oxadiazole -3- carbonamidines 21
Crude product 21c (100mg, 0.2mmol) is dissolved in 15mL tetrahydrofurans, the NaOH of 0.16mL 2.5N is addedSolution, stirring reaction 12 hours.After reaction terminates, saturated ammonium chloride solution is added, is extracted with ethyl acetate, merge organic phase,Washed with saturated nacl aqueous solution, anhydrous sodium sulfate drying, filtered, filtrate decompression concentration, with silica gel column chromatography with eluant, eluentSystem A purifying gained residues, obtain title product 21 (30mg, white solid), yield 31.9%.
MS m/z(ESI):463.1[M+1]
1H NMR(400MHz,DMSO-d6)δ11.49(s,1H),8.93(s,1H),7.16-7.21(m,1H),7.10-7.12(m,1H),6.75-6.78(m,1H),6.42-6.43(m,1H),4.09-4.12(m,1H),3.52-3.56(m,1H),3.33-3.36(m,2H),3.22-3.23(m,1H),2.91(s,3H),2.18-2.23(m,1H),1.95-1.98(m,1H).
Embodiment 22
(S)-N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- (methyl sulphonyl) pyrrolidin-3-yl) amino) -1,2,5- oxadiazole -3- carbonamidines 22
Using the synthetic route of embodiment 21, raw material 15a is replaced with into (S) -3- amino-pyrrolidine -1- t-butyl formates,Title product 22 (9mg, white solid), yield is obtained：63.4%.
MS m/z(ESI):464.2[M+1]
1H NMR(400MHz,CD3OD)δ11.48(s,1H),8.92(s,1H),7.17-7.21(t,1H),7.12-7.14(t,1H),6.78-6.80(m,1H),6.42(d,1H),4.11-4.13(m,1H),3.53-3.57(m,1H),3.35-3.37(m,1H),3.21-3.24(m,1H),2.91(s,3H),2.19-2.24(m,1H),1.96-1.99(m,1H),1.24(s,1H).
Embodiment 23
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- (methyl sulphonyl) piperidin-4-yl) amino) -1,2,5- EvilDiazole -3- carbonamidines 23
Using the synthetic route of embodiment 21, raw material 15a is replaced with into 4- amino piperidine -1- t-butyl formates, mark is obtainedTopic product 23 (80mg, white solid), yield：70.2%.
MS m/z(ESI):479.2[M+1]
1H NMR(400MHz,DMSO-d6)δ11.49(s,1H),8.93(s,1H),7.17-7.22(s,1H),7.10-7.12(m,1H),6.77-6.79(m,1H),6.21-6.23(m,1H),3.43-3.53(m,3H),3.28-3.29(m,5H),2.03-2.06(m,2H),1.49-1.59(m,2H).
Embodiment 24
N- (the bromo- 4- fluorophenyls of 3-)-N'- hydroxyls -4- ((1- sulfamoyls piperidin-4-yl) amino) -1,2,5- oxadiazoles -3- carbonamidines 24
Using the synthetic route of embodiment 15, raw material 15a is replaced with into 4- amino piperidine -1- t-butyl formates, mark is obtainedTopic product 24 (40mg, white solid), yield：28.2%.
MS m/z(ESI):480.2[M+1]
Biological assessment
The explanation present invention is further described below in conjunction with test case, but these embodiments are not meant as limiting of the inventionScope.
Measure of the test case 1, the compounds of this invention to people source IDO1 protease inhibiting activities
External people source IDO1 proteinase activities are tested by following method.
The method is used for determining inhibitory action of the compound in the present invention to people source IDO1 proteinase activities.
First, experiment material and instrument
1st, ELIASA (Synergy HT, BIOTEK)
2nd, tryptophan (T0254-5G, Sigma-Aldrich)
3rd, catalase derives from cattle liver (C1345-1G, Sigma-Aldrich)
4th, methylenum careuleum (M9140-25G, Sigma-Aldrich)
5th, L-AA sodium (A7631-25G, Sigma-Aldrich)
6th, 4- (dimethylamino) benzaldehyde (D2004-25G, Sigma-Aldrich)
7th, trichloroacetic acid (T9159-100G, Sigma-Aldrich)
8th, people source IDO1 genes (SC126221, Origene)
2nd, experimental procedure
The self-control of IDO1 protease：
People source IDO1 genes are transferred in PET30a plasmids by gene clone technology, the large intestine of competence is then transferred toBacillus Rosetta (DE3) competent cell (KT1003, Shenzhen HYK Gene Technology Co., Ltd.)；In liquid LB (Luria-Bertani) culture medium [according to《Molecular Cloning:A Laboratory guide》(J. Pehanorm Brooker D.W. Russells work) prepares every liter of cultureBase] in amplify culture, collects thalline, ultrasonication, by hanging column, affords the IDO1 protease of purifying.
Compound test experiments：
100 times of the enzyme (IDO1) of 24 μ l is diluted to 2400 μ l with the KPB of 50mM, concentration is the enzyme solutions of 2.6ng/ μ l,24 μ l enzyme solutions are added per hole in 96 hole reaction plates (AXYGEN, PCR-96-FLT-C) (hereinafter referred to as reaction plate).Blank well addsEnter the 24 μ l KPB [preparations (50mM) of KPB buffer solutions：KH is weighed with assay balance2PO46.805g is put into the beaker of 1000ml,Deionized water to 900ml is added with graduated cylinder, PH to 6.5 is adjusted with the KOH of 1M, be poured into the graduated cylinder of 1L, moisturizing to 1L isCan.4 DEG C of storages].The compound or DMSO of 1 μ l are added in corresponding reacting hole in reaction plate.Prepare A liquid：Take 200 μ l500mM L-AA sodium adds 1050 μ l KPB, and turbine-type mixer maximal rate is mixed 3 seconds.B liquid：100 μ l 10mM color ammoniaAcid plus the catalase of 100 μ l 100000unit/ml, plus 5 μ l 10mM methylenum careuleum, finally plus 1050 μ l KPB, turbineBlender maximal rate is mixed 3 seconds.1200 μ l A liquid and 1200 μ l B liquid are taken, maximal rate is mixed 3 seconds on turbine-type mixer.Then by this mixed liquor with the μ l of every hole 24 addition reaction plates.Reaction plate is put into board-like centrifuge maximum speed to be centrifuged 15 seconds,Reaction liquid is all converged to bottom, oscillator is mixed 30 seconds, in constant-temperature incubation case, 37 DEG C, be incubated 1h.In reaction plate,Add 10 μ l 30% (W/V) trichloroacetic acids per hole per hole, 65 DEG C are incubated 15 minutes in incubator.By reaction plate in centrifugeUpper 4700RPM centrifugations, room temperature, 5 minutes.40 μ l supernatants to 96 hole test boards of correspondence are shifted from reaction plate with the volley of rifle fireIn (Corning, #3599).4- (dimethylamino) benzaldehyde/glacial acetic acid solution of 40 μ l 2% (W/V) is added per hole, is being shakenMaximal rate on device is swung, is mixed 1 minute.After incubation at room temperature 2 minutes, read at 480nm on Synergy HT (BIOTEK)Light absorption value.
Compound is measured to people source IDO1 protease inhibiting activities by the experiment of the above in the present invention, is measuredIC50Value is shown in Table 1.
Compound suppresses IC to people source IDO1 proteinase activities in the present invention of table 150
Conclusion：The compounds of this invention has obvious inhibitory action to people source IDO1 proteinase activities.
Measure of the test case 2, the compounds of this invention to people source TDO protease inhibiting activities
External people source TDO proteinase activities are tested by following method.
The method is used for determining inhibitory action of the compound in the present invention to people source TDO proteinase activities.
First, experiment material and instrument
1st, ELIASA (Synergy HT, BIOTEK)
2nd, tryptophan (T0254-5G, Sigma-Aldrich)
3rd, catalase derives from cattle liver (C1345-1G, Sigma-Aldrich)
4th, methylenum careuleum (M9140-25G, Sigma-Aldrich)
5th, L-AA sodium (A7631-25G, Sigma-Aldrich)
6th, 4- (dimethylamino) benzaldehyde (D2004-25G, Sigma-Aldrich)
7th, trichloroacetic acid (T9159-100G, Sigma-Aldrich)
8th, people source TDO (U32989.1, Suzhou Jin Weizhi bio tech ltd)
9th, Rosseta (CW0811A, Beijing CoWin Bioscience Co., Ltd.)
10th, turbine-type mixer (6776, Corning)
11st, mini board-like centrifuge (Mini-P25, ABSON life science equipment)
2nd, experimental procedure
The self-control of TDO protease
The plasmid of the TDO genes of source containing people that will be built, is transferred to the Escherichia coli Rosseta of competence；In liquid LB(Luria-Bertani) culture medium [according to《Molecular Cloning:A Laboratory guide》(J. Pehanorm Brooker D.W. Russells work) is prepared everyRise culture medium] in amplify culture, collects thalline, ultrasonication, by hanging column, affords the TDO1 protease of purifying.
Compound test experiments：
100 times of the enzyme (TDO) of 24 μ l is diluted to 2400 μ l with the KPB of 50mM, concentration is the enzyme solutions of 2.6ng/ μ l,96 hole reaction plates (AXYGEN, PCR-96-FLT-C) (hereinafter referred to as reaction plate) add 24 μ l enzyme solutions per hole.Blank wellAdd the 24 μ l KPB buffer solutions [preparations (50mM) of KPB buffer solutions：KH is weighed with assay balance2PO46.805g is put into 1000mlBeaker, add deionized water to 900ml with graduated cylinder, adjust PH to 6.5 with the KOH of 1M, be conducted into the graduated cylinder of 1L, moisturizingTo 1L.4 DEG C of storages].The compound or DMSO of 1 μ l are added in corresponding reacting hole in reaction plate.Prepare A liquid：Take 200μ l 500mM L-AA sodium adds 1050 μ l KPB, and turbine-type mixer maximal rate is mixed 3 seconds.B liquid：100 μ l 10mM colorsThe acid of ammonia adds the catalase of 100 μ l 100000unit/ml, plus 5 μ l 10mM methylenum careuleum, finally plus 1050 μ l KPB,Turbine-type mixer maximal rate is mixed 3 seconds.1200 μ l A liquid and 1200 μ l B liquid are taken, maximal rate is mixed on turbine-type mixer3 seconds.Then by this mixed liquor with the μ l of every hole 24 addition reaction plates.Reaction plate is put into board-like centrifuge maximum speed centrifugation 15Second, reaction liquid is all converged to bottom, oscillator is mixed 30 seconds, in constant-temperature incubation case, 37 DEG C, is incubated 1h.In reaction plateIn, 10 μ l 30% (W/V) trichloroacetic acids are added per hole, 65 DEG C are incubated 15 minutes in incubator.By reaction plate on centrifuge4700RPM is centrifuged, room temperature, 5 minutes.40 μ l supernatants to 96 hole test boards of correspondence are shifted from reaction plate with the volley of rifle fireIn (Corning, #3599).4- (dimethylamino) benzaldehyde/glacial acetic acid solution of 40 μ l 2% (W/V) is added per hole, is being shakenMaximal rate on device is swung, is mixed 1 minute.After incubation at room temperature 2 minutes, read at 480nm on Synergy HT ReaderLight absorption value.
Compound is measured to people source TDO protease inhibiting activities by the experiment of the above in the present invention, the IC for measuring50Value is shown in Table 2.
Compound suppresses IC to people source TDO proteinase activities in the present invention of table 250
Conclusion：The compounds of this invention inhibitory action weaker to people source TDO proteinase activities, is selective IDO inhibitor.
The measure of test case 3, the compounds of this invention IDO protease inhibiting activities intracellular to HeLa
The intracellular IDO proteinase activities of HeLa are tested by following method.
The method is used for determining the inhibitory action of the compound IDO proteinase activities intracellular to HeLa in the present invention.(note：HeLa cell lines express indole amine 2,3-dioxygenase (IDO) under the induction of interferon gamma (INF- γ))
First, experiment material and instrument
1st, ELIASA (Synergy HT, BIOTEK)
2nd, tryptophan (T0254-5G, Sigma-Aldrich)
3rd, 4- (dimethylamino) benzaldehyde (D2004-25G, Sigma-Aldrich)
4th, trichloroacetic acid (T9159-100G, Sigma-Aldrich)
5th, HeLa cell lines (CCL-2, ATCC)
2nd, experimental procedure
HeLa cell suspensions are produced with Fresh cell culture medium, 100 μ l cultivating systems are added with 10000 cells/wellsIn 96 porocyte culture plates, 5% 37 DEG C of carbon dioxide is cultivated 24 hours.Removal supernatant, first adds 90 μ l serum-frees DMEM per holeHigh glucose medium；Then 10 μ l are separately added into per hole and prepare compound (its end in the culture medium of γ containing INF- and tryptophanConcentration is：10000,1000,100,10,1,0.1nM) 96 porocyte culture plates are taken out in, 48 hours 5% carbon dioxide, 37 DEG C of culturesIn the holes of μ l to 96 round bottom plate of middle supernatant 80, add 16 μ l 30% (W/V) trichloroacetic acids per hole per hole, incubated for 65 DEG C in incubatorEducate 25 minutes.4700RPM centrifugations, 5 minutes on centrifuge by reaction plate.Shifted from reaction plate with the volley of rifle fire 50 μ l supernatants toIn 96 hole flat bottom clear plates, 4- (dimethylamino) benzaldehyde/glacial acetic acid solution of 50 μ l 2% (W/V) is then added per hole,Mix 1 minute on the oscillator.After incubation at room temperature 2 minutes, the extinction at 480nm is read on Synergy HT ReaderValue.
Compound IDO protease inhibiting activities intracellular to HeLa are measured by the experiment of the above in the present invention, are surveyedThe IC for obtaining50Value is shown in Table 3.
Compound IDO proteinase activities intracellular to HeLa suppress IC in the present invention of table 350
Conclusion：The compounds of this invention IDO proteinase activities intracellular to HeLa have obvious inhibitory action.