WO2017035753A1 - 2-芳胺基吡啶、嘧啶或三嗪衍生物及其制备方法和用途 - Google Patents

2-芳胺基吡啶、嘧啶或三嗪衍生物及其制备方法和用途 Download PDF

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WO2017035753A1
WO2017035753A1 PCT/CN2015/088643 CN2015088643W WO2017035753A1 WO 2017035753 A1 WO2017035753 A1 WO 2017035753A1 CN 2015088643 W CN2015088643 W CN 2015088643W WO 2017035753 A1 WO2017035753 A1 WO 2017035753A1
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compound
methyl
dimethylamino
ethyl
amino
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PCT/CN2015/088643
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English (en)
French (fr)
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吴家权
张海军
曹焕岩
金深霜
张帅
陆政华
董健
王赪晨
谈秋
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无锡双良生物科技有限公司
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Priority to AU2015407300A priority Critical patent/AU2015407300B2/en
Priority to KR1020187004501A priority patent/KR102051609B1/ko
Priority to JP2018511087A priority patent/JP6606278B2/ja
Priority to PCT/CN2015/088643 priority patent/WO2017035753A1/zh
Priority to EP15902560.0A priority patent/EP3345906B1/en
Priority to CN201580002762.8A priority patent/CN106132957B/zh
Priority to US15/745,187 priority patent/US10377747B2/en
Priority to TW105119857A priority patent/TWI601718B/zh
Publication of WO2017035753A1 publication Critical patent/WO2017035753A1/zh

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    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
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    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
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    • A61K31/53—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with three nitrogens as the only ring hetero atoms, e.g. chlorazanil, melamine
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    • C07D239/02—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings
    • C07D239/24—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members
    • C07D239/28—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, directly attached to ring carbon atoms
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Definitions

  • the invention belongs to the field of new medical technology, in particular to 2-arylaminopyridine, pyrimidine or triazine derivatives or pharmaceutically acceptable salts thereof, or solvates thereof, which can be used for epidermal growth of certain variant forms Treatment or prevention of a disease or condition mediated by a factor receptor (eg, L858R activating mutant, delE746_A750 mutant, Exonl9 deletion activating mutant, and T790M resistant mutant).
  • a factor receptor eg, L858R activating mutant, delE746_A750 mutant, Exonl9 deletion activating mutant, and T790M resistant mutant.
  • Such compounds or salts thereof, or solvates thereof are useful in the treatment or prevention of many different cancers.
  • the invention also relates to a process for the preparation of intermediates useful in the preparation of said compounds.
  • NSCLC non-small-cell lung cancer
  • the epidermal growth factor receptor is a member of the transmembrane protein tyrosine kinase of the erbB receptor family.
  • the receptor can homodimerize with additional EGFR molecules, or with another family member (eg erbB2 (HER2), erbB3 (HER3) , or erbB4 (HER4)) heterodimerization occurs.
  • growth factor ligands such as epidermal growth factor (EGF)
  • the receptor can homodimerize with additional EGFR molecules, or with another family member (eg erbB2 (HER2), erbB3 (HER3) , or erbB4 (HER4)) heterodimerization occurs.
  • HER2 erbB2
  • HER3 erbB3
  • HER4 erbB4
  • erbB receptor Homology dimerization and/or heterodimerization of the erbB receptor leads to phosphorylation of key tyrosine residues in the intracellular domain, which in turn leads to stimulation of many intracellular signaling pathways involved in cell proliferation and survival. .
  • Abnormalities in the body's control of erbB family signaling promote proliferation, invasion, metastasis, angiogenesis, and tumor cell production, and have been described in many human cancers, including the lungs, head and neck, and chest. Therefore, the erbB receptor family is a reasonable target for the development of anticancer drugs.
  • Non-Patent Documents 1 and 2 provide a detailed discussion of erbB receptor signaling and its involvement in tumorigenesis.
  • Non-Patent Documents 3 and 4 report that activating mutations of the epidermal growth factor receptor in non-small cell lung cancer are associated with response to gefitinib treatment.
  • the most common epidermal growth factor receptor activating mutation (L858R del E746_A750) results in small molecule tyrosine kinase inhibitors (such as gefitinib and erlotinib relative to the wild-type (WT) epidermal growth factor receptor Increased affinity and decreased affinity for ATP.
  • WT wild-type
  • resistance to gefitinib and erlotinib treatment was developed, for example due to mutations in the gate residue T790M, which was reported to be detected in 50% of clinically resistant patients.
  • This mutation is not considered to sterically hinder the binding of gefitinib or erlotinib to EGFR, and only changes the level of ATP affinity to the level of wild-type (WT) epidermal growth factor receptor.
  • EGFR eg, L858R EGFR mutant, or delE746_A750 mutant or Exonl9 deletion EGFR
  • resistant mutant forms of EGFR eg, T790M EGFR mutant
  • WT EGFR WT EGFR
  • some irreversible ATP competitive inhibitors such as PF00299804, CI-1033, HKI-272, AZD9291, etc.
  • the irreversible inhibitor contains a Michael-added receptor fragment that is capable of generating a covalent bond with the sulfhydryl group (SH) of a conserved amino acid residue (Cys797) at the binding site.
  • Non-Patent Document 5 The ability of such an inhibitor to bind to an EGFR through an irreversible covalent bond is generally stronger than that between a reversible inhibitor and EGFR. Nevertheless, the clinical trial results of these irreversible inhibitors indicate that these inhibitors still have certain limitations, such as toxic effects due to off-target effects, side effects caused by low selectivity, and insufficient drug concentration in patients. Therefore, the development of new irreversible EGFR inhibitors It has great clinical significance and application prospects.
  • Non-Patent Document 1 New England Journal of Medicine, 2008, vol. 358, 1160-1174
  • Non-Patent Document 2 Biochemical and Biophysical Research Communications, 2004, vol. 319, 1-11
  • Non-Patent Document 3 Science, 2004, vol. 304, 1497-1500
  • Non-Patent Document 4 New England Journal of Medicine, 2004, vol. 350, 2129-2139
  • Non-Patent Document 5 Journal of Medicinal Chemistry, 2009, vol. 52, 1231-1236
  • the present inventors designed and synthesized a series of 2-arylaminopyridine, pyrimidine or triazine derivatives which have not been reported in the literature and characterized them.
  • this series of compounds was tested for cell-level activity with high EGFR inhibitory activity and showed relatively low inhibitory activity against WT EGFR.
  • the 2-arylaminopyridine, pyrimidine or triazine derivative EGFR inhibitor of the present invention can block the phosphorylation process of EGFR, inhibit the growth, proliferation and differentiation of tumor cells, and thus can be developed into a new antitumor drug. .
  • the invention is as follows.
  • X is selected from carbon, Y is selected from carbon, Z is selected from CR a , R 1 is selected from methoxy; or X is selected from nitrogen, Y is selected from carbon, Z is selected from CR b , and R 1 is selected from methoxy Or difluoromethoxy; or X is selected from nitrogen, Y is selected from nitrogen, Z is selected from CR c , R 1 is selected from methoxy or difluoromethoxy; or X is selected from nitrogen, and Y is selected from nitrogen, Z Selected from CR d , R 1 is selected from the group consisting of difluoromethoxy;
  • R a is selected from the group consisting of 3-methyl-1H-carbazol-1-yl, 1-methyl-1H-pyrrolo[2,3-b]pyridin-3-yl, 1-methyl-1H-thieno[ 3,2-c]pyrazol-3-yl, 1-methyl-1H-pyrazolo[4,3-b]pyridin-3-yl, pyrazolo[1,5-a]pyrimidine-3- Or imidazo[1,2-a]pyridin-3-yl;
  • R b is selected from the group consisting of benzo[d]isoxazol-3-yl, 1-methyl-1H-pyrrolo[3,2-b]pyridin-3-yl, 2,4-dimethyl-4H-thiophene And [3,2-b]pyrrole-6-yl, 2,5,6-trimethyl-6H-thieno[2,3-b]pyrrol-4-yl or 1-methyl-1H-thiophene [3,2-c]pyrazol-3-yl;
  • R c is selected from 1H-benzo[d]imidazol-1-yl, 1H-indol-7-yl or 1-methyl-1H-indol-7-yl;
  • R d is selected from the group consisting of 1-methyl-1H-pyrrolo[2,3-b]pyridin-3-yl;
  • R 2 is selected from (2-(dimethylamino)ethyl)(methyl)amino, 4-methylpiperazin-1-yl, 3-(dimethylamino)azetidin-1-yl, 4- (Dimethylamino)piperidin-1-yl, (S)-2-((dimethylamino)methyl)pyrrolidin-1-yl or 5-methyl-2,5-diazaspiro[3.4] Oct-2-yl.
  • R a is selected from the group consisting of 3-methyl-1H-carbazol-1-yl, 1-methyl-1H-pyrrolo[2,3-b]pyridin-3-yl, 1-methyl-1H-thiophene And [3,2-c]pyrazol-3-yl, 1-methyl-1H-pyrazolo[4,3-b]pyridin-3-yl, pyrazolo[1,5-a]pyrimidine- 3-yl or imidazo[1,2-a]pyridin-3-yl;
  • R 2 is selected from (2-(dimethylamino)ethyl)(methyl)amino, 4-methylpiperazin-1-yl, 3-(dimethylamino)azetidin-1-yl, 4- (Dimethylamino)piperidin-1-yl, (S)-2-((dimethylamino)methyl)pyrrolidin-1-yl or 5-methyl-2,5-diazaspiro[3.4] Oct-2-yl.
  • R b is selected from the group consisting of benzo[d]isoxazol-3-yl, 1-methyl-1H-pyrrolo[3,2-b]pyridin-3-yl, 2,4-dimethyl-4H -thieno[3,2-b]pyrrole-6-yl, 2,5,6-trimethyl-6H-thieno[2,3-b]pyrrol-4-yl or 1-methyl-1H- Thieno[3,2-c]pyrazol-3-yl;
  • R 1 is selected from methoxy or difluoromethoxy
  • R 2 is selected from (2-(dimethylamino)ethyl)(methyl)amino, 4-methylpiperazin-1-yl, 3-(dimethylamino)azetidin-1-yl, 4- (Dimethylamino)piperidin-1-yl, (S)-2-((dimethylamino)methyl)pyrrolidin-1-yl or 5-methyl-2,5-diazaspiro[3.4] Oct-2-yl.
  • R c is selected from 1H-benzo[d]imidazol-1-yl, 1H-indol-7-yl or 1-methyl-1H-indol-7-yl;
  • R 1 is selected from methoxy or difluoromethoxy
  • R 2 is selected from (2-(dimethylamino)ethyl)(methyl)amino, 4-methylpiperazin-1-yl, 3-(dimethylamino)azetidin-1-yl, 4- (Dimethylamino)piperidin-1-yl, (S)-2-((dimethylamino)methyl)pyrrolidin-1-yl or 5-methyl-2,5-diazaspiro[3.4] Oct-2-yl.
  • R d is selected from the group consisting of 1-methyl-1H-pyrrolo[2,3-b]pyridin-3-yl;
  • R 2 is selected from (2-(dimethylamino)ethyl)(methyl)amino, 4-methylpiperazin-1-yl, 3-(dimethylamino)azetidin-1-yl, 4- (Dimethylamino)piperidin-1-yl, (S)-2-((dimethylamino)methyl)pyrrolidin-1-yl or 5-methyl-2,5-diazaspiro[3.4] Oct-2-yl.
  • R b is selected from the group consisting of benzo[d]isoxazol-3-yl, 1-methyl-1H-pyrrole, or a pharmaceutically acceptable salt thereof, or a solvate thereof And [3,2-b]pyridin-3-yl or 1-methyl-1H-thieno[3,2-c]pyrazol-3-yl.
  • R c is 1H-indol-7-yl, or a pharmaceutically acceptable salt thereof, or a solvate thereof.
  • the compound according to the item 10 or a pharmaceutically acceptable salt thereof, or a solvate thereof comprising non-small cell lung cancer, breast cancer, glioma, prostate cancer, ovarian cancer, head and neck Squamous cell carcinoma, cervical cancer, esophageal cancer, liver cancer, kidney cancer, pancreatic cancer, colon cancer, skin cancer, leukemia, lymphoma, gastric cancer, multiple myeloma or solid tumor.
  • a pharmaceutical composition comprising the compound according to any one of items 1 to 11, or a pharmaceutically acceptable salt thereof, or a solvate thereof, and a pharmaceutically acceptable diluent and/or carrier.
  • the cancer comprises non-small cell lung cancer, breast cancer, glioma, prostate cancer, ovarian cancer, head and neck squamous cell carcinoma, cervical cancer, esophageal cancer, liver cancer, Kidney cancer, pancreatic cancer, colon cancer, skin cancer, leukemia, lymphoma, gastric cancer, multiple myeloma or solid tumor.
  • a method of producing an anti-cancer effect in a warm-blooded animal, such as a human, in need of treatment comprising: administering to the animal an effective amount of a compound according to any one of items 1 to 11 or a pharmaceutically acceptable compound thereof Salt, or a solvate thereof.
  • the compound represented by the following formula (VI) or a salt thereof is reacted with a carboxylic acid or a carboxylic acid derivative in the presence of an organic solvent.
  • organic solvent comprises dichloromethane, tetrahydrofuran, N,N-dimethylformamide or N,N-dimethylacetamide.
  • a pharmaceutically acceptable salt of a compound of the formula (I), formula (II), formula (III), formula (IV) or formula (V) may be, for example, an acid addition salt.
  • an acid addition salt for example, a mineral acid or an organic acid addition salt can be used.
  • An acid addition salt can be formed using an inorganic salt selected from the group consisting of hydrochloric acid, hydrobromic acid, sulfuric acid or phosphoric acid.
  • It may be selected from the group consisting of trifluoroacetic acid, citric acid, maleic acid, oxalic acid, acetic acid, formic acid, benzoic acid, fumaric acid, succinic acid, tartaric acid, lactic acid, pyruvic acid, methanesulfonic acid, benzenesulfonic acid or p-toluenesulfonic acid.
  • Organic The acid forms an acid addition salt.
  • N-(5-((4-(benzo[d]isoxazol-3-yl)pyrimidin-2-yl)amino)-4-(difluoromethoxy)-2 is provided a methanesulfonic acid salt of ((2-(dimethylamino)ethyl)(methyl)amino)phenyl)acrylamide.
  • N-(2-((2-(dimethylamino)ethyl))(methyl)amino)-4-methoxy-5-((4-(1-methyl-1H) is provided.
  • a mesylate salt of pyrrolo[3,2-b]pyridin-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide is provided.
  • N-(5-((4-(1H-indol-7-yl)-1,3,5-triazin-2-yl)amino)-4-(difluoromethoxy) is provided Methanesulfonate of 2-((2-(dimethylamino)ethyl)(methyl)amino)phenyl)acrylamide.
  • the compound of formula (I), formula (II), formula (III), formula (IV) or formula (V), or a pharmaceutically acceptable salt thereof may be solvated and unsolvated.
  • Form exists.
  • the solvated form can be in hydrated form. It should be understood that the present invention encompasses all such solvated and unsolvated forms.
  • the compound of the formula (I), the formula (II), the formula (III), the formula (IV) or the formula (V) can be administered in the form of a prodrug which is decomposed in the human or animal body to give a formula ( I), a compound of formula (II), formula (III), formula (IV) or formula (V).
  • prodrugs include in vivo hydrolysable esters of the compounds of formula (I), formula (II), formula (III), formula (IV) or formula (V).
  • the in vivo hydrolysable ester can be formed by esterifying a hydroxyl group in a compound represented by formula (I), formula (II), formula (III), formula (IV) or formula (V).
  • Various forms of prodrugs are known in the art. Examples of such prodrug derivatives can be found in:
  • One aspect of the invention provides a compound of formula (I), formula (II), formula (III), formula (IV) or formula (V) which inhibits activation or resistance mutation of one or more EGFR,
  • EGFR for example, the L858R EGFR mutant, or the delE746_A750 mutant or the Exonl9 deletion EGFR and/or the resistant mutant form of EGFR (such as the T790M EGFR mutant).
  • the compounds are useful in the treatment of cancer in patients who have developed or are at risk of developing a degree of resistance to existing therapies based on EGFR inhibitors.
  • a compound of formula (I), formula (II), formula (III), formula (IV) or formula (V) which exhibits an EGFR ratio for an activated or resistant mutant form WT EGFR has a higher inhibitory activity. Due to the reduced toxicity associated with WT EGFR inhibition, this compound is expected to be more suitable for use as a therapeutic, especially for cancer therapy. This toxicology is known to manifest as a rash and/or diarrhea in the human body.
  • the compound represented by the formula (VI) can be produced by deprotecting the corresponding amino-protected amine compound.
  • protecting groups include protecting groups suitable for protecting nitrogen atoms and methods for formation and final deprotection, see TW Greene and PGM Wuts, "Protective Groups in Organic Synthesis", Second Edition, John Wiley & Sons New York, 1991.
  • Another aspect of the invention provides these other intermediates.
  • Another aspect of the invention provides a compound of formula (VII) or a salt thereof:
  • X is selected from carbon, Y is selected from carbon, Z is selected from CR a , R 1 is selected from methoxy; or X is selected from nitrogen, Y is selected from carbon, Z is selected from CR b , and R 1 is selected from methoxy Or difluoromethoxy; or X is selected from nitrogen, Y is selected from nitrogen, Z is selected from CR c , R 1 is selected from methoxy or difluoromethoxy; or X is selected from nitrogen, and Y is selected from nitrogen, Z Selected from CR d , R 1 is selected from the group consisting of difluoromethoxy;
  • R a is selected from the group consisting of 3-methyl-1H-carbazol-1-yl, 1-methyl-1H-pyrrolo[2,3-b]pyridin-3-yl, 1-methyl-1H-thieno[ 3,2-c]pyrazol-3-yl, 1-methyl-1H-pyrazolo[4,3-b]pyridin-3-yl, pyrazolo[1,5-a]pyrimidine-3- Or imidazo[1,2-a]pyridin-3-yl;
  • R b is selected from the group consisting of benzo[d]isoxazol-3-yl, 1-methyl-1H-pyrrolo[3,2-b]pyridin-3-yl, 2,4-dimethyl-4H-thiophene And [3,2-b]pyrrole-6-yl, 2,5,6-trimethyl-6H-thieno[2,3-b]pyrrol-4-yl or 1-methyl-1H-thiophene [3,2-c]pyrazol-3-yl;
  • R c is selected from 1H-benzo[d]imidazol-1-yl, 1H-indol-7-yl or 1-methyl-1H-indol-7-yl;
  • R d is selected from the group consisting of 1-methyl-1H-pyrrolo[2,3-b]pyridin-3-yl;
  • R 2 is selected from (2-(dimethylamino)ethyl)(methyl)amino, 4-methylpiperazin-1-yl, 3-(dimethylamino)azetidin-1-yl, 4- (Dimethylamino)piperidin-1-yl, (S)-2-((dimethylamino)methyl)pyrrolidin-1-yl or 5-methyl-2,5-diazaspiro[3.4] Oct-2-yl.
  • the intermediate compound 12a or a salt thereof, is provided.
  • N 1 -(4-(benzo[d]isoxazol-3-yl)pyrimidin-2-yl)-N 4 -(2-(dimethylamino)ethyl) is provided.
  • the intermediate compound 19a or a salt thereof, is provided.
  • intermediate compound 29a or a salt thereof, is provided.
  • N 1 -(2-(dimethylamino)ethyl)-5-methoxy-N 1 -methyl-N 4 -(4-(1-methyl-1H-) is provided.
  • intermediate compound 37a or a salt thereof, is provided.
  • intermediate compound 49a or a salt thereof, is provided.
  • N 1 -(2-(dimethylamino)ethyl)-5-methoxy-N 1 -methyl-N 4 -(4-(1-methyl-1H-) is provided.
  • intermediate compound 49b or a salt thereof, is provided.
  • intermediate compound 78a or a salt thereof, is provided.
  • N 1 -(4-(1H-indol-7-yl)-1,3,5-triazin-2-yl)-N 4 -(2-(dimethylamino) is provided.
  • intermediate compound 78b or a salt thereof, is provided.
  • N 1 -(4-(1H-indol-7-yl)-1,3,5-triazin-2-yl)-2-(difluoromethoxy)-N is provided.
  • intermediate compound 83 or a salt thereof, is provided.
  • the pharmaceutically acceptable diluent and/or carrier there is no particular limitation, and conventionally known diluents and/or carriers can be used.
  • the pharmaceutical composition of the present invention may be in a form suitable for the following administration modes: for example, a tablet, a lozenge, a hard or soft capsule, an aqueous suspension or an oil suspension, an emulsion, a dispersible powder or a granule, or Syrup, etc.); topical use (eg creams, ointments, gels, aqueous or oil solutions, or suspensions); administration by inhalation (eg fine powder or liquid aerosol); administration by insufflation ( For example, subdivided) or parenteral (for example, sterile aqueous or oily solutions for intravenous, subcutaneous, intramuscular administration, or suppositories for rectal administration).
  • a tablet, a lozenge, a hard or soft capsule, an aqueous suspension or an oil suspension, an emulsion, a dispersible powder or a granule, or Syrup, etc. topical use (eg creams, ointments
  • compositions of the present invention can be obtained by conventional procedures using conventional pharmaceutical excipients well known in the art.
  • compositions for oral use may contain, for example, one or more coloring agents, sweeteners, flavoring agents, and/or preservatives.
  • the optimal dose of each active ingredient in the product is orally administered to a mammal daily in an amount of from about 0.0025 to 50 mg/kg body weight. Preferably, however, about 0.01 to 10 mg is administered orally per kilogram of body weight. Depending on the subject being treated, the particular route of administration, and the severity of the condition being treated, it may be necessary to change the daily dose.
  • the compounds or pharmaceutical compositions of the invention are useful in the treatment of a variety of diseases mediated by epidermal growth factor receptor kinase (EGFR).
  • EGFR epidermal growth factor receptor kinase
  • the disease mediated by EGFR is various cancers.
  • the cancer includes, but is not limited to, non-small cell lung cancer, breast cancer, glioma, prostate cancer, ovarian cancer, head and neck squamous cell carcinoma, cervical cancer, esophageal cancer, liver cancer, kidney cancer, pancreatic cancer, colon cancer, Skin cancer, leukemia, lymphoma, gastric cancer, multiple myeloma or solid tumor.
  • EGFR includes its wild type as well as various variants which cause disease. These variants include, but are not limited to, variants comprising the following mutations: T790M, L858R, L861Q, or L858R/T790M.
  • the invention also encompasses truncated forms of EGFR that cause disease.
  • the compounds of the invention, or pharmaceutical compositions thereof are useful in the treatment of a variety of diseases mediated by EGFR wild-type or various variants thereof that cause disease, including various cancers as described above, or inhibition of EGFR wild-type Or its biological activity that can cause various variants of the disease.
  • the crude compound 5 (54 g, 0.15 mol) was dissolved in DMF (500 mL), and DBU (40.2 g, 0.26 mol) was added at room temperature, and the mixture was heated to 90 ° C for 3 hours.
  • the reaction solution was cooled to room temperature, poured slowly into ice water, and the solid was evaporated, filtered, and the filter cake was washed twice with water and dried to give 47 g of a yellow solid.
  • Compound 11b was prepared in the same manner as Compound 11a except that N,N,N'-trimethylethylenediamine was replaced by 4-methylpiperazine.
  • 1 H NMR 400MHz, CDCl 3 ): ⁇ 8.23 (s, 1H), 6.52 (s, 1H), 3.98 (s, 3H), 3.20-3.10 (m, 4H), 2.69-2.59 (m, 4H) , 2.40 (s, 3H); MS (ESI) (m/z): [M+H] + 330.0.
  • Compound 14b was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 13b.
  • Compound 11c was prepared in the same manner as Compound 11a except that N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylpiperidin-4-amine.
  • Compound 12c was prepared in the same manner as Compound 12a except that Compound 11a was replaced with Compound 11c.
  • Compound 14c was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 13c.
  • Compound 14d was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 13d.
  • Compound 11e was prepared in the same manner as Compound 11a except that N,N,N'-trimethylethylenediamine was replaced by 5-methyl-2,5-diazaspiro[3.4]octane.
  • Compound 14e was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 13e.
  • Compound 18a was prepared in the same manner as Compound 11a except that Compound 10 was replaced with Compound 17.
  • Compound 19a was prepared in the same manner as Compound 12a except that Compound 11a was replaced with Compound 18a.
  • Compound 20a was prepared in the same manner as Compound 13a except that Compound 12a was replaced with Compound 19a.
  • Compound 21a was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 20a.
  • Compound 18b was prepared in the same manner as Compound 11a except that Compound 10 was replaced with Compound 17, and N,N,N'-trimethylethylenediamine was replaced by 4-methylpiperazine.
  • Compound 19b was prepared in the same manner as Compound 12a except that Compound 11a was replaced with Compound 18b.
  • Compound 21b was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 20b.
  • Compound 18c was prepared in the same manner as Compound 11a except that Compound 10 was replaced with Compound 17, N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylazetidin-3-amine. .
  • Compound 21c was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 20c.
  • Compound 18d was prepared in the same manner as Compound 11a except that Compound 10 was replaced with Compound 17, and N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylpiperidin-4-amine.
  • Compound 21d was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 20d.
  • Compound 21e was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 20e.
  • Compound 21f was prepared in the same manner as Compound 14a except that Compound 13a was replaced with Compound 20f.
  • N,N-diisopropylethylamine (280 mg, 2.14 mmol) and N,N were added sequentially to a solution of Compound 28 (650 mg, 1.65 mmol) in N,N-dimethylacetamide (15 mL).
  • N'-trimethylethylenediamine (202 mg, 1.98 mmol).
  • the mixture was heated to 70 ° C for 6 hours, and after completion of the reaction, ethyl acetate and water were added. After stirring well, the mixture was filtered over Celite, and the organic layer was washed with brine, and then dried and concentrated to give a dark solid compound 29a (450 mg, 57%).
  • Compound 29b was prepared in the same manner as Compound 29a except that N,N,N'-trimethylethylenediamine was replaced by 4-methylpiperazine. MS (ESI) (m / z ): [M + H] + 475.1.
  • Compound 30b was prepared in the same manner as Compound 30a except that Compound 29a was replaced by Compound 29b.
  • Compound 31b was prepared in the same manner as Compound 31a except that Compound 30a was replaced with Compound 30b.
  • Compound 29c was prepared in the same manner as Compound 29a except that N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylazetidin-3-amine.
  • Compound 31c was prepared in the same manner as Compound 31a except that Compound 30a was replaced by Compound 30c.
  • Compound 29d was prepared in the same manner as Compound 29a except that N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylpiperidin-4-amine.
  • Compound 30d was prepared in the same manner as Compound 30a except that Compound 29a was replaced by Compound 29d.
  • Compound 31d was prepared in the same manner as Compound 31a except that Compound 30a was replaced by Compound 30d.
  • Compound 31e was prepared in the same manner as Compound 31a except that Compound 30a was replaced with Compound 30e.
  • Compound 29f was prepared in the same manner as Compound 29a except that N,N,N'-trimethylethylenediamine was replaced by 5-methyl-2,5-diazaspiro[3.4]octane.
  • Compound 30f was prepared in the same manner as Compound 30a except that Compound 29a was replaced by Compound 29f.
  • Compound 31f was prepared in the same manner as Compound 31a except that Compound 30a was replaced by Compound 30f.
  • Compound 37b was prepared in the same manner as Compound 37a except that N,N,N'-trimethylethylenediamine was replaced by 4-methylpiperazine. MS (ESI) (m / z ): [M + H] + 511.2.
  • Compound 38b was prepared in the same manner as Compound 38a except that Compound 37a was replaced with Compound 37b.
  • Compound 39b was prepared in the same manner as Compound 39a except that Compound 38a was replaced with Compound 38b.
  • Compound 37c was prepared in the same manner as Compound 37a except that N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylazetidin-3-amine.
  • Compound 37d was prepared in the same manner as Compound 37a except that N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylpiperidin-4-amine.
  • Compound 38d was prepared in the same manner as Compound 38a except that Compound 37a was replaced with Compound 37d.
  • Compound 39d was prepared in the same manner as Compound 39a except that Compound 38a was replaced with Compound 38d.
  • Compound 39e was prepared in the same manner as Compound 39a except that Compound 38a was replaced with Compound 38e.
  • Compound 37f was prepared in the same manner as Compound 37a except that N,N,N'-trimethylethylenediamine was replaced by 5-methyl-2,5-diazaspiro[3.4]octane.
  • Compound 38f was prepared in the same manner as Compound 38a except that Compound 37a was replaced by Compound 37f.
  • 1 H NMR (400 MHz, CDCl 3 ): 8.63 - 8.59 (m, 1 H), 8.43 (d, J 5.2 Hz, 1H), 8.24 - 8.16 (m, 2H), 7.93 (s, 1H), 7.69-7.
  • Compound 39f was prepared in the same manner as Compound 39a except that Compound 38a was replaced by Compound 38f.
  • Compound 48b was prepared in the same manner as Compound 48a except that Compound 27 was replaced with Compound 35. MS (ESI) (m / z ): [M + H] + 437.1.
  • the compound 51b was prepared in the same manner as the compound 51a except that the compound 50a was replaced with the compound 50b.
  • Compound 49c was prepared in the same manner as Compound 49a except that Compound 48a was replaced by Compound 48b and N,N,N'-trimethylethylenediamine was replaced by 4-methylpiperazine.
  • Compound 51c was prepared in the same manner as Compound 51a except that Compound 50a was changed to Compound 50c.
  • Compound 49d was prepared in the same manner as Compound 49a except that Compound 48a was replaced by Compound 48b, N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylazetidin-3-amine. .
  • Compound 50d was prepared in the same manner as Compound 50a except that Compound 49a was changed to Compound 49d.
  • Compound 51d was prepared in the same manner as Compound 51a except that Compound 50a was changed to Compound 50d.
  • Compound 49e was prepared in the same manner as Compound 49a except that Compound 48a was replaced by Compound 48b and N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylpiperidin-4-amine.
  • Compound 51e was prepared in the same manner as Compound 51a except that Compound 50a was changed to Compound 50e.
  • Compound 51f was prepared in the same manner as Compound 51a except that Compound 50a was changed to Compound 50f.
  • Compound 78c was prepared in the same manner as Compound 78a except that N,N,N'-trimethylethylenediamine was replaced by 4-methylpiperazine.
  • Compound 78d was prepared in the same manner as Compound 78a except that N,N,N'-trimethylethylenediamine was replaced by N,N-dimethylazetidin-3-amine.
  • Compound 79b was prepared in the same manner as Compound 79a except that Compound 78a was changed to Compound 78b.
  • Compound 80b was prepared in the same manner as Compound 80a except that Compound 79a was replaced by Compound 79b.
  • Compound 80c was prepared in the same manner as Compound 80a except that Compound 79a was changed to Compound 79c.
  • 1 H NMR (400MHz, CDCl 3 ): ⁇ 10.94 (s, 1H), 9.59 (s, 1H), 8.86-8.48 (m, 3H), 7.95-7.76 (m, 2H), 7.41-7.25 (m, 2H), 6.82 (s, 1H), 6.62 (s, 1H), 6.51-6.47 (m, 1H), 6.37-6.30 (m, 1H), 5.81 (d, J 10.3 Hz, 1H), 3.90 (s) , 3H), 3.06-3.00 (m, 4H), 2.88-2.80 (m, 4H), 2.51 (s, 3H); MS (ESI) (m/z): [M+H] + 485.2.
  • Compound 80d was prepared in the same manner as Compound 80a except that Compound 79a was changed to Compound 79d.
  • Compound 80e was prepared in the same manner as Compound 80a except that Compound 79a was changed to Compound 79e.
  • Compound 80f was prepared in the same manner as Compound 80a except that Compound 79a was changed to Compound 79f.
  • Example 38 N-(4-(Difluoromethoxy)-2-((2-(dimethylamino)ethyl)(methyl)amino)-5-((4-(1-methyl-) 1H-pyrrolo[2,3-b]pyridin-3-yl)-1,3,5-triazin-2-yl)amino)phenyl)acrylamide (Compound 85)
  • Compound 89 was prepared in the same manner as Compound 85 except that Compound 84 was replaced with Compound 88.
  • 1 H NMR (400MHz, CDCl 3 ): ⁇ 10.23 (s, 1H), 9.74 (s, 2H), 9.61 (s, 1H), 8.73 (s, 1H), 8.61 (s, 1H), 7.98 (s , 1H), 7.88-7.86 (m, 1H), 7.38 (m, 2H), 6.70-6.36 (m, 2H), 5.76 (d, J 10.7 Hz, 1H), 3.93 (s, 3H), 2.93 ( s, 2H), 2.77 (s, 3H), 2.33 (s, 8H); MS (ESI) (m/z): [M+H] + 488.2.
  • Compound 94d was prepared in the same manner as Compound 94a except that 3-bromo-1-methyl-1H-pyrrolo[2,3-b]pyridine was replaced by 3-bromopyrazolo[1,5-a]pyrimidine. .
  • Compound 96d was prepared in the same manner as Compound 96a except that Compound 95a was replaced by Compound 95d.
  • Compound 97b was prepared in the same manner as Compound 97a except that Compound 96a was replaced by Compound 96b.
  • 1 H NMR 400 MHz, CD 3 OD
  • ⁇ 8.58 (s, 1H), 8.18-8.11 (m, 1H), 7.61 (d, J 5.3 Hz, 1H), 7.21.
  • Compound 97d was prepared in the same manner as Compound 97a except that Compound 96a was replaced by Compound 96d.
  • Compound 102 was prepared in the same manner as Compound 96a except that Compound 95a was replaced by Compound 101.
  • the MTT cell proliferation assay is described below.
  • H1975 non-small cell lung cancer cells, EGFR L858K/T790M
  • ⁇ 431 human epidermal carcinoma cells, EGFR wild type cells were purchased from the ATCC cell bank.
  • the growth medium of H1975 cells was RPMI-1640 (GIBCO, A10491-065), 10% fetal bovine serum.
  • the growth medium of A431 cells was DMEM (GIBCO, 11995-065), 10% fetal bovine serum.
  • the effect of the compound on the proliferation activity of tumor cells was examined by CellTiter-Glo assay. Tumor cells were exposed to the treatment conditions for 72 h, and the cell density used in each cell line was adjusted according to the 72 h growth curve of the cells. Ten concentration gradients (0.5 nM - 10 ⁇ M) were set for the compounds to be tested, and three sets of parallel controls were used for each concentration value.
  • the cells in the logarithmic growth phase were trypsinized to prepare a cell suspension, which was counted by a Roche counter and diluted appropriately with complete medium to a final concentration of 1 to 2 x 10 3 cells/mL.
  • the cells were seeded in a 384-well plate at 22.6 ⁇ L per well, and 3 sets of parallel were set, and cultured overnight at 37 ° C in a 5% CO 2 incubator.
  • the compound was dissolved in DMSO to prepare a mother liquor at a concentration of 10 ⁇ mol/L, and then the compound was serially diluted with a BRAVO instrument, and the concentrations of the compounds were gradually diluted to 10, 3.33, 1.11, 0.37, 0.123, 0.041, 0.0137, 0.00457, 0.00152, respectively. , 0.0005 ⁇ mol/L.
  • 2 ⁇ L of the compound solution was added to 18 ⁇ L of the medium, and after thorough mixing, 2 ⁇ L of the compound and medium mixed solution was added to 18 ⁇ L of the medium, and the mixture was thoroughly mixed.
  • 2.4 ⁇ L of the mixture was added to a 384-well plate.

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Abstract

本发明涉及2-芳胺基吡啶、嘧啶或三嗪衍生物及其制备方法和用途。所述2-芳胺基吡啶、嘧啶或三嗪衍生物可以作用于某些突变形式的表皮生长因子受体，例如L858R激活突变体、delE746_A750突变体、Exonl9缺失激活突变体和T790M耐药突变体，从而用于疾病和病况的治疗和预防。所述2-芳胺基吡啶、嘧啶或三嗪衍生物可用于癌症的治疗和预防。本发明还涉及包含2-芳胺基吡啶、嘧啶或三嗪衍生物的药物组合物，可用于制备2-芳胺基吡啶、嘧啶或三嗪衍生物的中间体，以及利用2-芳胺基吡啶、嘧啶或三嗪衍生物治疗由各种不同形式EGFR所介导的疾病的方法。

Description

2-芳胺基吡啶、嘧啶或三嗪衍生物及其制备方法和用途 技术领域
本发明属于新医药技术领域，特别涉及2-芳胺基吡啶、嘧啶或三嗪衍生物或其药学上可接受的盐、或其溶剂化物，其可用于某种由某些变异形态的表皮生长因子受体(例如L858R激活突变体、delE746_A750突变体、Exonl9缺失激活突变体和T790M耐药突变体)所介导的疾病或病况的治疗或预防。此类化合物或其盐、或其溶剂化物可用于很多不同癌症的治疗或预防。本发明还涉及包含所述化合物的制备中有用的中间体的制备方法。
背景技术
癌症患者的主要治疗为放疗、化疗及手术治疗。临床上，约80％的肺癌为非小细胞肺癌(non-small-cell lung cancer，NSCLC)，根据美国癌症协会的统计数据，美国每年大约新增20万例NSCLC患者。其中65％以上确诊时已属III、IV期。除部分III期NSCLC可经诱导治疗手术切除，大部分需要采用化疗。晚期非小细胞肺癌的治疗仍具挑战性，外科手术仅可以治愈大约30％的局限期病变。而化疗药物的全身毒副作用大，给患者带来极大痛苦。因此，寻找高效低毒的靶向治疗药物成为抗肿瘤药物发展的必然方向。
表皮生长因子受体(EGFR)是erbB受体家族的跨膜蛋白酪氨酸激酶成员。当与生长因子配体(如表皮生长因子(EGF))结合时，受体可以与附加的EGFR分子发生同源性二聚，或与另一个家庭成员(如erbB2(HER2)、erbB3(HER3)、或erbB4(HER4))发生异源性二聚。
erbB受体的同源性二聚化和/或异源性二聚化导致细胞内域关键酪氨酸残基的磷酸化，进而导致对参与细胞增殖和生存的许多细胞内信号传导通路的刺激。机体对erbB家族信号传导控制的异常，促进了扩散、入侵、转移、血管生成和肿瘤细胞的生成，并且已在许多人类癌症(包括肺，头部和颈部和胸部等)中得到了描述。因此，erbB受体家族为抗癌药物开发的合理靶点。其中以EGFR为靶点的一些药物已经上市，包括吉非替尼(Gifitinib，IRESSA TM)、厄洛替尼(Erlotinib，TARCEVATM)、拉帕替尼(TYKERBTM，TYVERBTM)等。非专利文献1和2提供了对erbB受体信号传导及其参与肿瘤发生的详细论述。
非专利文献3和4报道了在非小细胞肺癌中表皮生长因子受体的激活突变与对吉非替尼治疗的反应有关。最常见的表皮生长因子受体激活突变(L858R delE746_A750)导致相对于野生型(WT)表皮生长因子受体而言，对小分子酪氨酸激酶抑制剂(如吉非替尼和厄洛替尼)的亲和力增加以及对ATP亲和力的下降。最终，产生了对吉非替尼和厄洛替尼治疗的获得耐药性，例如由于看门残基T790M的突变，据报道在50％的临床耐药性患者中检测到该突变。该突变不被认为是在空间上阻碍了吉非替尼或厄洛替尼与EGFR结合，仅将与ATP亲和力水平改变到与野生型(WT)表皮生长因子受体的水平。
鉴于这种突变在靶向EGFR的现有疗法的抗性中的重要性，我们认为可以抑制包含看门基因突变的EGFR药物在癌症的治疗中特别有用。
相对于激活突变体形式的EGFR(如L858R EGFR突变体，或delE746_A750突变体或Exonl9缺失EGFR)和/或耐药突变形式的EGFR(如T790M EGFR突变体)，对于可表现出对WT EGFR的有效特性、和/或对其他酶受体的选择性的化合物依然存在着需求，所述选择性使得这些化合物特别有希望被开发成治疗药物。就此而言，对于某些激活或耐药性突变形式的表皮生长因子受体结合显示较高的抑制效果，且与WT EGFR抑制效果不明显的化合物存在着需求。由于与WT EGFR抑制有关毒理作用小，因而预期此类化合物可以更适于用作治疗剂，特别是对于癌症的治疗。为了克服T790M突变引起的相关的耐药性，一些不可逆的ATP竞争性抑制剂(如PF00299804，CI-1033，HKI-272，AZD9291等)已经进入临床研究阶段。不可逆抑制剂含有一个迈克尔加成的受体片段，能够和结合位点的一个保守氨基酸残基(Cys797)的巯基(SH)生成共价键。这种抑制剂和EGFR之间通过不可逆的共价键结合的能力通常比可逆性抑制剂和EGFR之间的结合能力强(非专利文献5)。尽管如此，上述这些不可逆抑制剂的临床实验结果表明这些抑制剂仍然有一定的局限性，如由于脱靶效应造成的毒性作用，低选择性导致的副作用，无法实现患者体内足够的药物浓度。因此，开发新型的不可逆EGFR抑制剂 具有重大的临床意义和应用前景。
现有技术文献
非专利文献
非专利文献1：New England Journal of Medicine,2008,vol.358,1160-1174
非专利文献2：Biochemical and Biophysical Research Communications,2004,vol.319,1-11
非专利文献3：Science,2004,vol.304,1497-1500
非专利文献4：New England Journal of Medicine,2004,vol.350,2129-2139
非专利文献5：Journal of Medicinal Chemistry,2009,vol.52,1231-1236
发明内容
本发明人设计合成了一系列未见文献报道的2-芳胺基吡啶、嘧啶或三嗪衍生物，并进行了结构表征。另外，对此系列化合物进行了细胞水平的活性测试，具有高EGFR抑制活性，同时显示了对WT EGFR相对较低的抑制活性。
本发明涉及的2-芳胺基吡啶、嘧啶或三嗪衍生物类EGFR抑制剂可以阻断EGFR的磷酸化过程，抑制肿瘤细胞的生长、增殖和分化，因而，可以开发成为新的抗肿瘤药物。
本发明如下所述。
1.式(I)所示的化合物或其药学上可接受的盐、或其溶剂化物：
Figure PCTCN2015088643-appb-000001
其中，X选自碳，Y选自碳，Z选自C-Ra，R1选自甲氧基；或X选自 氮，Y选自碳，Z选自C-Rb，R1选自甲氧基或二氟甲氧基；或X选自氮，Y选自氮，Z选自C-Rc，R1选自甲氧基或二氟甲氧基；或X选自氮，Y选自氮，Z选自C-Rd，R1选自二氟甲氧基；
Ra选自3-甲基-1H-吲唑-1-基、1-甲基-1H-吡咯并[2,3-b]吡啶-3-基、1-甲基-1H-噻吩并[3,2-c]吡唑-3-基、1-甲基-1H-吡唑并[4,3-b]吡啶-3-基、吡唑并[1,5-a]嘧啶-3-基或咪唑并[1,2-a]吡啶-3-基；
Rb选自苯并[d]异恶唑-3-基、1-甲基-1H-吡咯并[3,2-b]吡啶-3-基、2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基、2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基或1-甲基-1H-噻吩并[3,2-c]吡唑-3-基；
Rc选自1H-苯并[d]咪唑-1-基、1H-吲哚-7-基或1-甲基-1H-吲哚-7-基；
Rd选自1-甲基-1H-吡咯并[2,3-b]吡啶-3-基；
R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
2.根据项1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(II)所示的结构，
Figure PCTCN2015088643-appb-000002
其中，Ra选自3-甲基-1H-吲唑-1-基、1-甲基-1H-吡咯并[2,3-b]吡啶-3-基、1-甲基-1H-噻吩并[3,2-c]吡唑-3-基、1-甲基-1H-吡唑并[4,3-b]吡啶-3-基、吡唑并[1,5-a]嘧啶-3-基或咪唑并[1,2-a]吡啶-3-基；
R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
3.根据项1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(III)所示的结构，
Figure PCTCN2015088643-appb-000003
其中，Rb选自苯并[d]异恶唑-3-基、1-甲基-1H-吡咯并[3,2-b]吡啶-3-基、2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基、2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基或1-甲基-1H-噻吩并[3,2-c]吡唑-3-基；
R1选自甲氧基或二氟甲氧基；
R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
4.根据项1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(IV)所示的结构，
Figure PCTCN2015088643-appb-000004
其中，Rc选自1H-苯并[d]咪唑-1-基、1H-吲哚-7-基或1-甲基-1H-吲哚-7-基；
R1选自甲氧基或二氟甲氧基；
R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
5.根据项1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(V)所示的结构，
Figure PCTCN2015088643-appb-000005
其中，Rd选自1-甲基-1H-吡咯并[2,3-b]吡啶-3-基；
R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
6.根据项3所述的化合物或其药学上可接受的盐、或其溶剂化物，其中，Rb选自苯并[d]异恶唑-3-基、1-甲基-1H-吡咯并[3,2-b]吡啶-3-基或1-甲基-1H-噻吩并[3,2-c]吡唑-3-基。
7.根据项4所述的化合物或其药学上可接受的盐、或其溶剂化物，其中，Rc为1H-吲哚-7-基。
8.根据项1～7中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物，其中，R2为(2-(二甲氨基)乙基)(甲基)氨基。
9.根据项1～5中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物，其中，所述式(I)所示的化合物选自以下化合物中的任意一种：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺、N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡咯-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺、N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)氨基)苯基)丙烯酰胺、N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基苯 基)丙烯酰胺、N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡咯-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、或N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基苯基)丙烯酰胺。
10.根据项1～9中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物，其用于治疗癌症。
11.根据项10所述的化合物或其药学上可接受的盐、或其溶剂化物，所述癌症包括非小细胞肺癌、乳腺癌、神经胶质细胞瘤、前列腺癌、卵巢癌、头颈部鳞癌、宫颈癌、食管癌、肝癌、肾癌、胰腺癌、结肠癌、皮肤癌、白血病、淋巴瘤、胃癌、多发性骨髓癌或实体瘤。
12.一种药物组合物，包含根据项1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物以及药学上可接受的稀释剂和/或载体。
13.根据项1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物作为药物的用途。
14.根据项1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物在制备治疗癌症的药物中的用途。
15.根据项14所述的用途，其中，所述癌症包括非小细胞肺癌、乳腺癌、神经胶质细胞瘤、前列腺癌、卵巢癌、头颈部鳞癌、宫颈癌、食管癌、肝癌、肾癌、胰腺癌、结肠癌、皮肤癌、白血病、淋巴瘤、胃癌、多发性骨髓癌或实体瘤。
16.在需要治疗的温血动物例如人中产生抗癌作用的方法，其包括：向所述动物给药有效量的根据项1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物。
17.根据项1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物和附加的抗肿瘤物质的用途，用于癌症的同时，独立或序贯治疗。
18.一种制备根据项1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物的方法，包括：
在有机溶剂的存在下，使下述式(VI)所示的化合物或其盐与羧酸或羧酸衍生物反应，
Figure PCTCN2015088643-appb-000006
式(VI)中，X、Y、Z、R1和R2与所述式(I)同义。
19.根据项18所述的方法，其中，所述有机溶剂包括二氯甲烷、四氢呋喃、N,N-二甲基甲酰胺或N,N-二甲基乙酰胺。
20.根据项18或19所述的方法，其中，所述羧酸或羧酸衍生物包括丙烯酸、丙烯酰氯或丙烯酸酯。
21.根据项18～20中任一项所述的方法，其中，所述式(VI)所示的化合物选自以下化合物中的任意一种：N4-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺、N4-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲苯-1,2,4-三胺、N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,2,4-三胺、5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,2,4-三胺、N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,2,4-三胺、5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,2,4-三胺、N4-(4-(1H-吲哚-7基)-1,3,5-三嗪-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺、N4-(4-(1H-吲哚-7基)-1,3,5-三嗪-2-基)-5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲苯-1,2,4-三胺或5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)苯-1,2,4-三胺。
在本发明中，合适的式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物的药学上可接受的盐例如可以是酸加成盐。例如，可以用无机酸或有机酸加成盐。可以使用选自盐酸、氢溴酸、硫酸或磷酸的无机盐形成酸加成盐。可以使用选自三氟乙酸、柠檬酸、马来酸、草酸、乙酸、甲酸、苯甲酸、富马酸、琥珀酸、酒石酸、乳酸、丙酮酸、甲磺酸、苯磺酸或对甲苯磺酸的有机 酸形成酸加成盐。
在一个实施方式中，提供N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺的甲磺酸盐。
在一个实施方式中，提供N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺的甲磺酸盐。
在一个实施方式中，提供N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺的甲磺酸盐。
在一个实施方式中，提供N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡咯-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺的甲磺酸盐。
在一个实施方式中，提供N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺的甲磺酸盐。
在一个实施方式中，提供N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)氨基)苯基)丙烯酰胺的甲磺酸盐。
应当理解的是，式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物或其药学上可以接受的盐可以以溶剂化形式和未溶剂化形式存在。例如，溶剂化形式可以是水合形式。应该理解的是，本发明包括所有这种溶剂化和未溶剂化的形式。
式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物可以以前药的形式给药，该前药在人体或动物体内被分解而产生式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物。前药的例子包括式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物的体内可水解酯。可通过将式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物中的羟基酯化，形成体内可水解酯。本领域已知各种形式的前药。这种前药衍生物的例子可见于：
(a)《前药的设计(Design of Prodrugs)》，由H.Bundgaard编辑(Elsevier,1985年)和Methods in Enzymology,第42卷，第309-396页，由K.Wedder等人编辑(Academic Press，1985年)；
(b)《药物设计和开发的教科书(A Textbook of Drug Design and Development)》，由Krogsgaard-Larsen和H.Bundgaard编辑，第5章：“前药的设计和应用”，由H.Bundgaard编辑，第113-191页(1991年)；
本发明的一个方面提供式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物，其抑制一种或多种EGFR的激活或抗性突变，如L858R EGFR突变体，或delE746_A750突变体或Exonl9缺失EGFR和/或耐药突变形式的EGFR(如T790M EGFR突变体)。所述化合物可用于对基于EGFR抑制剂的现有疗法已产生或者有风险产生一定程度的抗性的患者的癌症治疗。
在本发明的一个方面，提供式(I)、式(II)、式(III)、式(IV)或式(V)所示的化合物，其对激活或抗性突变体形式的EGFR显示比WT EGFR更高的抑制活性。由于与WT EGFR抑制相关的毒性降低，因而预期这种化合物可能更适于用作治疗剂，尤其实用于癌症治疗。已知这种毒理学在人体中表现为皮疹和/或腹泻。
另一方面，在本发明的式(I)所示的化合物的制备方法中，式(VI)所示的化合物可通过将相应的氨基保护的胺化合物脱保护而制备。保护基的例子包括适用于保护氮原子的保护基以及形成和最后脱保护的方法，参见T.W.Greene和P.G.M.Wuts,《有机合成中保护基(Protective Groups in Organic Synthesis)》，第二版，John Wiley&Sons，纽约，1991年。
本发明的另一方面提供这些其它的中间体。
因此，本发明的另一方面提供式(VII)所示的化合物或其盐：
Figure PCTCN2015088643-appb-000007
其中，X选自碳，Y选自碳，Z选自C-Ra，R1选自甲氧基；或X选自氮，Y选自碳，Z选自C-Rb，R1选自甲氧基或二氟甲氧基；或X选自氮，Y 选自氮，Z选自C-Rc，R1选自甲氧基或二氟甲氧基；或X选自氮，Y选自氮，Z选自C-Rd，R1选自二氟甲氧基；
Ra选自3-甲基-1H-吲唑-1-基、1-甲基-1H-吡咯并[2,3-b]吡啶-3-基、1-甲基-1H-噻吩并[3,2-c]吡唑-3-基、1-甲基-1H-吡唑并[4,3-b]吡啶-3-基、吡唑并[1,5-a]嘧啶-3-基或咪唑并[1,2-a]吡啶-3-基；
Rb选自苯并[d]异恶唑-3-基、1-甲基-1H-吡咯并[3,2-b]吡啶-3-基、2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基、2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基或1-甲基-1H-噻吩并[3,2-c]吡唑-3-基；
Rc选自1H-苯并[d]咪唑-1-基、1H-吲哚-7-基或1-甲基-1H-吲哚-7-基；
Rd选自1-甲基-1H-吡咯并[2,3-b]吡啶-3-基；
R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
下面给出上述实施方式中的仅部分实例：
在一个实施方式中，提供中间体化合物12a、或其盐。
因此，在这种情况下，提供N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-N4-(2-(二甲氨基)乙基)-2-甲氧基-N4-甲基-5-硝基苯-1,4-二胺、或其盐。
在一个实施方式中，提供中间体化合物19a、或其盐。
因此，在这种情况下，提供N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-5-硝基苯-1,4二胺、或其盐。
在一个实施方式中，提供中间体化合物29a、或其盐。
因此，在这种情况下，提供N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-2-硝基苯-1,4-二胺、或其盐。
在一个实施方式中，提供中间体化合物37a、或其盐。
因此，在这种情况下，提供2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-5-硝基苯-1,4-二胺、或其盐。
在一个实施方式中，提供中间体化合物49a、或其盐。
因此，在这种情况下，提供N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)-2-硝基苯-1,4-二胺、或其盐。
在一个实施方式中，提供中间体化合物49b、或其盐。
因此，在这种情况下，提供2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-N1-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)-5-硝基苯-1,4二胺、或其盐。
在一个实施方式中，提供中间体化合物78a、或其盐。
因此，在这种情况下，提供N1-(4-(1H-吲哚-7基)-1,3,5-三嗪-2-基)-N4-(2-(二甲氨基)乙基)-2-甲氧基-N4-甲基-5-硝基苯-1,4二胺、或其盐。
在一个实施方式中，提供中间体化合物78b、或其盐。
因此，在这种情况下，提供N1-(4-(1H-吲哚-7基)-1,3,5-三嗪-2-基)-2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-5-硝基苯-1,4-二胺、或其盐。
在一个实施方式中，提供中间体化合物83、或其盐。
因此，在这种情况下，提供2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-N1-(4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)-5-硝基苯-1,4-二胺、或其盐。
另一方面，在本发明的药物组合物中，作为药学上可接受的稀释剂和/或载体，没有特别限制，可以使用目前公知的稀释剂和/或载体。
本发明的药物组合物可采用适合于下列给药方式的剂型：口服使用(例如片剂、锭剂、硬或软胶囊、水悬浮剂或油悬浮剂、乳剂、分散性粉剂或颗粒剂、或者糖浆剂等)；局部使用(例如霜剂、软膏剂、凝胶剂、水溶液或油溶液、或者悬浮液)；通过吸入给药(例如细粉或者液体气溶胶)；通过吹入法给药(例如细分)或者胃肠外给药(例如用于静脉、皮下、肌肉内给药的无菌水溶液或油溶液，或者用于直肠给药的栓剂)。
本发明的药物组合物可通过使用本领域熟知的常规药用辅料的常规步骤而获得。因此，用于口服使用的组合物可含有例如：一种或多种着色剂、甜味剂、矫味剂和/或防腐剂。
虽然每个人的需求各不相同，本领域技术人员可确定本发明的药物组合 物中每种活性成分的的最佳剂量。一般情况下，本发明的化合物或其药学上可接受的盐或溶剂化物，对哺乳动物每天口服给药，药量按照约0.0025到50毫克/公斤体重。但最好是每公斤体重口服给药约0.01到10毫克。根据治疗的主体，具体给药途径，正在治疗的疾病的严重程度有必要改变每日剂量。
本发明的化合物或药物组合物可用于治疗各种由表皮生长因子受体激酶(EGFR)介导的疾病。本文中，由EGFR介导的疾病为各种癌症。所述癌症包括但不仅限于非小细胞肺癌、乳腺癌、神经胶质细胞瘤、前列腺癌、卵巢癌、头颈部鳞癌、宫颈癌、食管癌、肝癌、肾癌、胰腺癌、结肠癌、皮肤癌、白血病、淋巴瘤、胃癌、多发性骨髓癌或实体瘤。
应理解，本发明中，EGFR包括其野生型以及能导致疾病的各种变异体。这些变异体包括但不限于包含以下突变的变异体：T790M、L858R、L861Q、或L858R/T790M。本发明也包括EGFR的能导致疾病的截短形式。因此，本发明化合物或其药物组合物可用于治疗由EGFR野生型或其能导致疾病的各种变异体所介导的各种疾病，包括上文所述的各种癌症，或抑制EGFR野生型或其能导致疾病的各种变异体的生物学活性。
具体实施方式
EGFR抑制剂合成部分
在以下的实施例中将进一步举例说明本发明。这些实施例仅用于说明本发明，但不以任何方式限制本发明。
化合物14a-14e的合成路线如方案1所示：
Figure PCTCN2015088643-appb-000008
方案1
(2-氟苯基)(2-(甲硫基)嘧啶-4-基)甲酮(化合物4)
Figure PCTCN2015088643-appb-000009
将化合物1(51.84g，0.32mol)溶于1,4-二氧六环(600mL)中，室温搅拌下加入化合物2(48g，0.39mol)与化合物3(23.68g，0.11mol)，冰水浴冷却下慢慢加入NaH(60％)(15.36g，0.39mol)，加毕撤去冰水浴，升温至回流反应过夜。反应液冷却至室温，慢慢倒入冰水中，用乙酸乙酯萃取三次，有机相用饱和食盐水洗一次，无水硫酸钠干燥，过滤，减压旋干，柱层析得到41g白色固体化合物4。1H NMR(400MHz,CDCl3):δ8.80(d,J＝4.9Hz,1H),7.77-7.75(m,1H),7.65-7.57(m,1H),7.53(d,J＝4.9Hz,1H),7.34-7.29(m,1H),7.20-7.12(m,1H),2.47(s,3H)。
(Z)-(2-氟苯基)(2-(甲硫基)嘧啶-4-基)甲酮肟(化合物5)
Figure PCTCN2015088643-appb-000010
将盐酸羟胺(51.8g，0.75mol)加入到无水乙醇(700mL)中，室温搅拌下加入吡啶(58.9g，0.75mol)，室温下搅拌5min，再加入化合物4(37g，0.15mol)，升温至回流反应过夜。反应液冷却至室温，减压蒸去乙醇，残余物加水与乙酸乙酯搅拌，分液，水相再用乙酸乙酯萃取一次，合并有机相，用饱和食盐水洗一次，无水硫酸钠干燥，过滤，减压蒸干得到54g黄色油状液体化合物5，直接用于下一步反应。MS(ESI)(m/z):[M+H]+264.0。
3-(2-(甲硫基)嘧啶-4-基)苯并[d]异恶唑(化合物6)
Figure PCTCN2015088643-appb-000011
将化合物5粗品(54g，0.15mol)溶于DMF(500mL)中，室温下加入DBU(40.2g，0.26mol)，升温至90℃反应3小时。反应液冷却至室温，慢慢倒入冰水中，搅拌析出固体，过滤，滤饼用水洗两次，干燥得到47g黄色固体。1H NMR(400MHz,CDCl3):δ8.71(d,J＝5.1Hz,1H),8.54(d,J＝8.0Hz,1H),7.88(d,J＝5.1Hz,1H),7.71-7.63(m,2H),7.52-7.42(m,1H),2.74(s,3H)；MS(ESI)(m/z):[M+H]+244.0。
3-(2-(甲磺酰基)嘧啶-4-基)苯并[d]异恶唑(化合物7)
Figure PCTCN2015088643-appb-000012
将化合物6(24.3g，0.1mol)溶于二氯甲烷(300mL)中，冰水浴冷却下分批加入间氯过氧苯甲酸(25.9g，0.15mol)，加毕升至室温反应过夜。反应液中加入饱和碳酸氢钠溶液，搅拌10分钟后分液，有机相再用饱和食盐水洗一次，无水硫酸钠干燥，过滤，减压蒸干得到白色固体24g，直接用于下一步反应。MS(ESI)(m/z):[M+H]+276.0。
4-(苯并[d]异恶唑-3-基)嘧啶-2-胺(化合物8)
Figure PCTCN2015088643-appb-000013
将化合物7(12g，0.044mol)加入到250mL封管反应器中，再加入乙腈(100mL)与氨水(30mL)，升至70℃反应过夜。反应液冷却，过滤得到6g白色固体。1H NMR(400MHz,CDCl3):δ8.52-8.50(m,2H),7.70-7.62(m,2H),7.53(d,J＝5.1Hz,1H),7.46-7.42(m,1H),5.30(s,2H)；MS(ESI)(m/z):[M+H]+213.1。
1-溴-4-氟-2甲氧基-5-硝基苯(化合物10)
Figure PCTCN2015088643-appb-000014
将化合物9(15g，0.073mol)溶于浓硫酸(150mL)中，冰盐浴冷却下慢慢加入硝酸钾(7.4g，0.073mol)，加毕升至室温反应1h。反应液慢慢倒入冰水中，搅拌析出固体，过滤，滤饼用水洗两次，烘干得到灰色固体13.2g。MS(ESI)(m/z):[M+H]+250.9。
N1-(4-溴-5-甲氧基-2-硝基苯基)-N1，N2，N2-三甲基乙烷-1,2-二胺(化合物11a)
Figure PCTCN2015088643-appb-000015
将化合物10(15g，0.06mol)溶于乙腈(150mL)中，加入N,N,N’-三甲基乙二胺(6.74g，0.066mol)与碳酸钾(16.56g，0.12mol)，升温至回流反应过夜。反应液冷却至室温，过滤，滤饼用二氯甲烷洗两次，合并滤液，减压蒸干，柱层析得到黄色油状物15g。1H NMR(400MHz,CDCl3):δ8.10(s,1H),6.62(s,1H),3.94(s,3H),3.33(t,J＝6.9Hz,2H),2.89(s,3H),2.57(t,J＝6.9Hz,2H),2.26(s,6H)；MS(ESI)(m/z):[M+H]+332.1。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-N4-(2-(二甲氨基)乙基)-2-甲氧基-N4-甲基-5-硝基苯-1,4-二胺(化合物12a)
Figure PCTCN2015088643-appb-000016
将化合物8(100mg，0.47mmol)溶于甲苯(10mL)中，加入化合物11a(236mg，0.71mmol)，叔丁醇钾(162mg，1.44mmol)，2-二环己基磷-2,4,6-三异丙基联苯(X-Phos)(448mg，0.94mmol)与三(二亚苄基丙酮)二钯(431mg，0.47mmol)，用氮气置换三次后，升温至90℃反应过夜。反应液冷却至室温，过滤，滤饼用二氯甲烷洗两次，合并滤液，减压蒸干，柱层析得到黄色粉末60mg。1H NMR(400MHz,CDCl3):δ9.04(s,1H),8.61(d,J＝4.7Hz,1H),8.52(d,J＝7.6Hz,1H),7.70-7.63(m，4H),7.51-7.47(m,1H),6.76(s,1H),4.02(s,3H),3.37(m,2H),2.91(s,3H),2.69(m,2H),2.36(s,6H)；MS(ESI)(m/z):[M+H]+464.2。
N4-(4-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺(化合物13a)
Figure PCTCN2015088643-appb-000017
将化合物12a(60mg，0.13mmol)溶于乙醇(9mL)与水(3mL)中，加入还原铁粉(43mg，0.77mmol)，氯化铵(5mg，0.09mmol)，升温至回流反应2h。反应液冷却至室温，过滤，滤液用饱和碳酸钾调至碱性，二氯甲烷萃取两次，合并有机相，饱和食盐水洗一次，无水硫酸钠干燥，过滤，减压蒸干，刮板得到黄色粉末40mg。MS(ESI)(m/z):[M+H]+434.2。
实施例1：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺(化合物14a)
Figure PCTCN2015088643-appb-000018
将化合物13a(40mg，0.14mmol)溶于二氯甲烷(10mL)与叔丁醇(1mL) 中，冰盐浴冷却下加入1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(EDCI，35mg，0.28mmol)，三乙胺(19mg，0.28mmol)与丙烯酸(13mg，0.28mmol)，加毕升至室温反应2h。反应液中加入饱和碳酸钾溶液，搅拌10min后分液，有机相干燥，蒸干，制备色谱纯化得到淡黄色固体8mg。1H NMR(400MHz,CDCl3):δ10.23(s,1H),9.50(s,1H),8.69(d,J＝5.0Hz,1H),8.57(d,J＝8.0Hz,1H),7.68-7.57(m,4H),7.38-7.29(m,1H),6.85(s,1H),6.45-6.28(m,2H),5.71(m,1H),3.93(s,3H),2.96-2.88(m,2H),2.75(s,3H),2.39-2.28(m,8H)；MS(ESI)(m/z):[M+H]+488.2。
1-(4-溴-5-甲氧基-2-硝基苯基)-4-甲基哌嗪(化合物11b)
Figure PCTCN2015088643-appb-000019
除了将N,N,N’-三甲基乙二胺替换成4-甲基哌嗪之外，与化合物11a同样地制备化合物11b。1H NMR(400MHz,CDCl3):δ8.23(s,1H),6.52(s,1H),3.98(s,3H),3.20-3.10(m,4H),2.69-2.59(m,4H),2.40(s,3H)；MS(ESI)(m/z):[M+H]+330.0。
4-(苯并[d]异恶唑-3-基)-N-(2-甲氧基-4-(4-甲基哌嗪-1-基)-5-硝基苯基)嘧啶-2-胺(化合物12b)
Figure PCTCN2015088643-appb-000020
将化合物8(200mg，0.94mmol)溶于1,4-二氧六环(10mL)中，加入化合物11b(311mg，0.94mmol)，碳酸铯(1.23g，3.77mmol)，2-二环己基磷-2,4,6-三异丙基联苯(X-Phos)(270mg，0.57mmol)与三(二亚苄基丙酮)二钯(173mg，0.19mmol)，用氮气置换三次后，升温至90℃反应过夜。反应液冷却至室温，过滤，滤饼用二氯甲烷洗两次，合并滤液，减压蒸干，柱层析得到红色油状物180mg。MS(ESI)(m/z):[M+H]+462.2。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-6-甲氧基-4-(4-甲基哌嗪-1-基)苯 -1,3-二胺(化合物13b)
Figure PCTCN2015088643-appb-000021
除了将化合物12a替换成化合物12b之外，与化合物13a同样地制备化合物13b。MS(ESI)(m/z):[M+H]+432.2。
实施例2：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-甲氧基-2-(4-甲基哌嗪-1-基)苯基)丙烯酰胺(化合物14b)
Figure PCTCN2015088643-appb-000022
除了将化合物13a替换成化合物13b之外，与化合物14a同样地制备化合物14b。1H NMR(400MHz,CDCl3):δ9.45(s,1H),8.68(d,J＝5.0Hz,1H),8.56-8.54(m,2H),7.68-7.58(m,4H),7.39-7.35(m,1H),6.85(s,1H),6.44-6.23(m,2H),5.78(d,J＝9.1Hz,1H),3.93(s,3H),2.99-2.98(m,4H),2.69(m,4H),2.45(s,3H)；MS(ESI)(m/z):[M+H]+486.2。
1-(4-溴-5-甲氧基-2-硝基苯基)-N,N-二甲基哌啶-4-胺(化合物11c)
Figure PCTCN2015088643-appb-000023
除了将N,N,N’-三甲基乙二胺替换成N,N-二甲基哌啶-4-胺之外，与化合物11a同样地制备化合物11c。1H NMR(400MHz,CDCl3)δ8.22(s,1H),6.51(s,1H),3.97(s,3H),3.38(d,J＝12.4Hz,2H),2.93-2.87(m,2H),2.45-2.31(m,7H),1.95-1.92(m,2H),1.84-1.74(m,2H)；MS(ESI)(m/z):[M+H]+358.1。
4-(苯并[d]异恶唑-3-基)-N-(4-(4-(二甲氨基)哌啶-1-基)-2-甲氧基-5-硝基苯基)嘧啶-2-胺(化合物12c)
Figure PCTCN2015088643-appb-000024
除了将化合物11a替换成化合物11c之外，与化合物12a同样地制备化合物12c。1H NMR(400MHz,CDCl3):δ9.18(s,1H),8.66(d,J＝5.1Hz,1H),8.55(d,J＝7.7Hz,1H),7.78-7.63(m,4H),7.55-7.50(m,1H),6.67(s,1H),4.05(s,3H),3.42-3.36(m,2H),2.94-2.78(m,2H),2.37-2.35(m,7H),1.96-1.83(m,4H)；MS(ESI)(m/z):[M+H]+490.2。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-4-(4-(二甲氨基)哌啶-1-基)-6-甲氧基苯-1,3-二胺(化合物13c)
Figure PCTCN2015088643-appb-000025
除了将化合物12a替换成化合物12c之外，与化合物13a同样地制备化合物13c。MS(ESI)(m/z):[M+H]+460.2。
实施例3：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-2-(4-(二甲氨基)哌啶-1-基)-4-甲氧基苯基)丙烯酰胺(化合物14c)
Figure PCTCN2015088643-appb-000026
除了将化合物13a替换成化合物13c之外，与化合物14a同样地制备化合物14c。1H NMR(400MHz,CDCl3):δ9.39(s,1H),8.65(d,J＝5.0Hz,1H),8.52(d,J＝7.9Hz,1H),8.41(s,1H),7.75-7.53(m,4H),7.36(t,J＝7.4Hz,1H),6.76(s,1H),6.37-6.36(m,2H),5.78-5.77(m,1H),3.92(s,3H),3.18-3.15(m,2H),2.81-2.76(m,3H),2.67(s,6H),2.25-2.22(m,2H),1.98-1.96(m,2H)；MS(ESI)(m/z):[M+H]+514.3。
(S)-1-(1-(4-溴-5-甲氧基-2-硝基苯基)吡咯烷-2-基)-N,N-二甲基甲胺(化合物11d)
Figure PCTCN2015088643-appb-000027
除了将N,N,N’-三甲基乙二胺替换成(S)-N,N-二甲基-1-(吡咯烷-2-基)-甲胺之外，与化合物11a同样地制备化合物11d。1H NMR(400MHz,CDCl3):δ8.12(s,1H),6.71(s,1H),3.95(s,3H),3.62-3.55(m,1H),2.72-2.61(m,2H),2.49-2.23(m,8H),2.09-1.71(m,4H)；MS(ESI)(m/z):[M+H]+359.1。
(S)-4-(苯并[d]异恶唑-3-基)-N-(4-(2-((二甲氨基)甲基)吡咯烷-1-基)-2-甲氧基-5-硝基苯基)嘧啶-2-胺(化合物12d)
Figure PCTCN2015088643-appb-000028
除了将化合物11a替换成化合物11d之外，与化合物12a同样地制备化合物12d。MS(ESI)(m/z):[M+H]+490.2。
(S)-N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-4-(2-((二甲氨基)甲基)吡咯烷-1-基)-6-甲氧基苯-1,3-二胺(化合物13d)
Figure PCTCN2015088643-appb-000029
除了将化合物12a替换成化合物12d之外，与化合物13a同样地制备化合物13d。MS(ESI)(m/z):[M+H]+460.2。
实施例4：(S)-N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-2-(2-((二甲氨基)甲基)吡咯烷-1-基)-4-甲氧基苯基)丙烯酰胺(化合物14d)
Figure PCTCN2015088643-appb-000030
除了将化合物13a替换成化合物13d之外，与化合物14a同样地制备化 合物14d。1H NMR(400MHz,CDCl3)；δ12.02(s,1H),9.55(s,1H),9.29(s,1H),8.66(d,J＝5.0Hz,1H),8.54(d,J＝8.0Hz,1H),7.73-7.53(m,4H),7.45(m,1H),6.72(s,1H),6.37(d,J＝16.1Hz,1H),5.72(d,J＝11.4Hz,1H),3.94(s,3H),2.97(m,1H),2.83-2.20(m,8H),2.13-1.60(m,4H)；MS(ESI)(m/z):[M+H]+514.3。
2-(4-溴-5-甲氧基-2-硝基苯基)-5-甲基-2,5-二氮杂螺[3.4]辛烷(化合物11e)
Figure PCTCN2015088643-appb-000031
除了将N,N,N’-三甲基乙二胺替换成5-甲基-2,5-二氮杂螺[3.4]辛烷之外，与化合物11a同样地制备化合物11e。1H NMR(400MHz,CDCl3):δ8.16(s,1H),5.97(s,1H),4.15(d,J＝9.2Hz,2H),3.96(s,3H),3.75(d,J＝9.2Hz,2H),2.77(t,J＝7.2Hz,2H),2.50(s,3H),2.15-2.11(m,2H),1.88-1.75(m,2H)；MS(ESI)(m/z):[M+H]+357.1。
4-(苯并[d]异恶唑-3-基)-N-(2-甲氧基-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)-5-硝基苯基)嘧啶-2-胺(化合物12e)
Figure PCTCN2015088643-appb-000032
除了将化合物11a替换成化合物11e之外，与化合物12a同样地制备化合物12e。MS(ESI)(m/z):[M+H]+488.2。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-6-甲氧基-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯-1,3-二胺(化合物13e)
Figure PCTCN2015088643-appb-000033
除了将化合物12a替换成化合物12e之外，与化合物13a同样地制备化 合物13e。MS(ESI)(m/z):[M+H]+458.2。
实施例5：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-甲氧基-2-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯基)丙烯酰胺(化合物14e)
Figure PCTCN2015088643-appb-000034
除了将化合物13a替换成化合物13e之外，与化合物14a同样地制备化合物14e。1H NMR(400MHz,CDCl3):δ8.58(d,J＝5.0Hz,1H),8.55-8.23(m,2H),7.72-7.51(m,4H),7.48-7.38(m,1H),7.18-6.91(m,1H),6.53-6.14(m,3H),5.78-5.64(m,1H),4.10-4.06(m,2H),3.98-3.95(m,3H),3.77-3.71(m,2H),2.93-2.76(m,2H),2.60-2.52(m,3H),2.28-2.12(m,2H),1.89-1.84(m,2H)；MS(ESI)(m/z):[M+H]+512.2。
化合物21a-21f的合成路线如方案2所示：
Figure PCTCN2015088643-appb-000035
方案2
1-溴-2-(二氟甲氧基)-4-氟苯(化合物16)
Figure PCTCN2015088643-appb-000036
将化合物15(80g，0.42mol)溶于DMF(800mL)中，加入碳酸钠(266g， 2.5mol)，升温至90℃，滴加二氟氯乙酸(191g，1.46mol)，滴毕保持90℃反应过夜。反应液冷却至室温，慢慢倒入冰水中，用乙酸乙酯萃取两次，合并有机相，用饱和食盐水洗一次，无水硫酸钠干燥，旋干得到粗品95g，直接用于下一步反应。1H NMR(400MHz,CDCl3):δ7.62-7.58(m,1H),7.04-7.01(m,1H),6.93-6.88(m,1H),6.57(t,J＝72.8Hz,1H)；MS(ESI)(m/z):[M+H]+240.9。
1-溴-2-(二氟甲氧基)-4-氟-5-硝基苯(化合物17)
除了将化合物9替换成化合物16之外，与化合物10同样地制备化合物17。1H NMR(400MHz,CDCl3):δ8.44(d,J＝7.8Hz,1H),7.25(d,J＝11.1Hz,1H),6.71(t,J＝71.0Hz,1H)；MS(ESI)(m/z):[M+H]+286.9。
N1-(4-溴-5-(二氟甲氧基)-2-硝基苯基)-N1,N2,N2-三甲基乙烷-1,2-二胺(化合物18a)
除了将化合物10替换成化合物17之外，与化合物11a同样地制备化合物18a。1H NMR(400MHz,CDCl3):δ8.03(s,1H),7.03(s,1H),6.60(t,J＝72.6Hz,1H),3.33(t,J＝6.8Hz,2H),2.87(s,3H),2.57(t,J＝6.8Hz,2H),2.28(s,6H)；MS(ESI)(m/z):[M+H]+369.0。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-5-硝基苯-1,4-二胺(化合物19a)
Figure PCTCN2015088643-appb-000039
除了将化合物11a替换成化合物18a之外，与化合物12a同样地制备化合物19a。1H NMR(400MHz,CDCl3):δ9.00(s,1H),8.66(d,J＝4.9Hz,1H), 8.45(d,J＝7.7Hz,1H),7.69-7.66(m,3H),7.60-7.45(m,2H),7.08(s,1H),6.70(t,J＝72.3Hz,1H),3.30(t,J＝7.0Hz,2H),2.90(s,3H),2.62-2.53(m,2H),2.29(s,6H)；MS(ESI)(m/z):[M+H]+500.2。
N4-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲苯-1,2,4-三胺(化合物20a)
Figure PCTCN2015088643-appb-000040
除了将化合物12a替换成化合物19a之外，与化合物13a同样地制备化合物20a。1H NMR(400MHz,CDCl3):δ8.61(d,J＝5.0Hz,1H),8.54(d,J＝8.0Hz,1H),7.90(s,1H),7.73-7.60(m,3H),7.49-7.44(m,2H),6.94(s,1H),6.49(t,J＝74.4Hz,1H),2.97(t,J＝6.4Hz,2H),2.72(s,3H),2.46(t,J＝6.4Hz,2H),2.31(s,6H)；MS(ESI)(m/z):[M+H]+470.2。
实施例6：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺(化合物21a)
Figure PCTCN2015088643-appb-000041
除了将化合物13a替换成化合物20a之外，与化合物14a同样地制备化合物21a。1H NMR(400MHz,CDCl3):δ10.33(s,1H),9.53(s,1H),8.69(d,J＝5.0Hz,1H),8.48(d,J＝8.0Hz,1H),7.67-7.57(m,3H),7.45(s,1H),7.34(t,J＝7.5Hz,1H),7.12(s,1H),6.77-6.28(m,3H),5.75-5.72(m,1H),2.96-2.84(m,2H),2.75(s,3H),2.39-2.32(m,8H)；MS(ESI)(m/z):[M+H]+524.2。
1-(4-溴-5-(二氟甲氧基)-2-硝基苯基)-4-甲基哌嗪(化合物18b)
Figure PCTCN2015088643-appb-000042
除了将化合物10替换成化合物17，N,N,N’-三甲基乙二胺替换成4-甲基 哌嗪之外，与化合物11a同样地制备化合物18b。MS(ESI)(m/z):[M+H]+366.0。
4-(苯并[d]异恶唑-3-基)-N-(2-(二氟甲氧基)-4-(4-甲基哌嗪-1-基)-5-硝基苯基)嘧啶-2-胺(化合物19b)
Figure PCTCN2015088643-appb-000043
除了将化合物11a替换成化合物18b之外，与化合物12a同样地制备化合物19b。1H NMR(400MHz,CDCl3):δ8.63(d,J＝5.0Hz,1H),8.53(d,J＝8.0Hz,1H),8.03(s,1H),7.77-7.61(m,3H),7.55(s,1H),7.46(t,J＝7.4Hz,1H),6.97(s,1H),6.52(t,J＝74.0Hz,1H),3.21(m,4H),3.03(m,4H),2.68(s,3H)；MS(ESI)(m/z):[M+H]+498.2。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-6-(二氟甲氧基)-4-(4-甲基哌嗪-1-基)苯-1,3-二胺(化合物20b)
Figure PCTCN2015088643-appb-000044
除了将化合物12a替换成化合物19b之外，与化合物13a同样地制备化合物20b。MS(ESI)(m/z):[M+H]+468.2。
实施例7：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-(4-甲基哌嗪-1-基)苯基)丙烯酰胺(化合物21b)
Figure PCTCN2015088643-appb-000045
除了将化合物13a替换成化合物20b之外，与化合物14a同样地制备化 合物21b。1H NMR(400MHz,CDCl3):δ9.48(s,1H),8.69(d,J＝5.0Hz,1H),8.63(s,1H),8.47(d,J＝8.0Hz,1H),7.73-7.55(m,3H),7.47(s,1H),7.36(t,J＝7.4Hz,1H),7.11(s,1H),6.80-6.25(m,3H),5.82(d,J＝9.8Hz,1H),2.98(m,4H),2.71(m,4H),2.47(s,3H)；MS(ESI)(m/z):[M+H]+522.2。
1-(4-溴-5-(二氟甲氧基)-2-硝基苯基)-N,N-二甲基氮杂环丁-3-胺(化合物18c)
Figure PCTCN2015088643-appb-000046
除了将化合物10替换成化合物17，N,N,N’-三甲基乙二胺替换成N,N-二甲基氮杂环丁-3-胺之外，与化合物11a同样地制备化合物18c。MS(ESI)(m/z):[M+H]+366.1。
4-(苯并[d]异恶唑-3-基)-N-(2-(二氟甲氧基)-4-(3-(二甲氨基)氮杂环丁-1-基)-5-硝基苯基)嘧啶-2-胺(化合物19c)
Figure PCTCN2015088643-appb-000047
除了将化合物11a替换成化合物18c之外，与化合物12a同样地制备化合物19c。1H NMR(400MHz,CDCl3):δ8.98(s,1H),8.63(s,1H),8.43(d,J＝7.7Hz,1H),7.78-7.61(m,3H),7.52-7.37(m,2H),6.67(t,J＝72.3Hz,1H),6.45(s,1H),4.14(t,J＝7.7Hz,2H),3.76-3.73(m,2H),3.27-3.18(m,1H),2.23(s,6H)；MS(ESI)(m/z):[M+H]+498.2。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-6-(二氟甲氧基)-4-(3-(二甲氨基)氮杂环丁-1-基)苯-1,3-二胺(化合物20c)
Figure PCTCN2015088643-appb-000048
除了将化合物12a替换成化合物19c之外，与化合物13a同样地制备化 合物20c。MS(ESI)(m/z):[M+H]+468.2。
实施例8：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-(3-(二甲氨基)氮杂环丁-1-基)苯基)丙烯酰胺(化合物21c)
Figure PCTCN2015088643-appb-000049
除了将化合物13a替换成化合物20c之外，与化合物14a同样地制备化合物21c。1H NMR(400MHz,CDCl3):δ8.77(s,1H),8.61(d,J＝5.0Hz,1H),8.41(d,J＝7.9Hz,1H),7.72-7.49(m,4H),7.39-7.36(m,2H),6.47-6.33(m,4H),5.79(d,J＝8.4Hz,1H),3.94(t,J＝6.6Hz,2H),3.72(t,J＝6.3Hz,2H),3.26-3.14(m,1H),2.28(s,6H)；MS(ESI)(m/z):[M+H]+522.2。
1-(4-溴-5-(二氟甲氧基)-2-硝基苯基)-N,N-二甲基哌啶-4-胺(化合物18d)
Figure PCTCN2015088643-appb-000050
除了将化合物10替换成化合物17，N,N,N’-三甲基乙二胺替换成N,N-二甲基哌啶-4-胺之外，与化合物11a同样地制备化合物18d。MS(ESI)(m/z):[M+H]+394.0。
4-(苯并[d]异恶唑-3-基)-N-(2-(二氟甲氧基)-4-(4-(二甲氨基)哌啶-1-基)-5-硝基苯基)嘧啶-2-胺(化合物19d)
Figure PCTCN2015088643-appb-000051
除了将化合物11a替换成化合物18d之外，与化合物12a同样地制备化合物19d。MS(ESI)(m/z):[M+H]+526.2。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-6-(二氟甲氧基)-4-(4-(二甲氨基) 哌啶-1-基)苯-1,3-二胺(化合物20d)
Figure PCTCN2015088643-appb-000052
除了将化合物12a替换成化合物19d之外，与化合物13a同样地制备化合物20d。MS(ESI)(m/z):[M+H]+496.2。
实施例9：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-(4-(二甲氨基)哌啶-1-基)苯基)丙烯酰胺(化合物21d)
Figure PCTCN2015088643-appb-000053
除了将化合物13a替换成化合物20d之外，与化合物14a同样地制备化合物21d。1H NMR(400MHz,CDCl3):δ9.44(s,1H),8.67(d,J＝5.0Hz,1H),8.55(s,1H),8.45(d,J＝8.0Hz,1H),7.75-7.55(m,3H),7.47(s,1H),7.35(t,J＝7.4Hz,1H),7.04(s,1H),6.79-6.31(m,3H),5.83-5.80(m,1H),3.17-3.14(m,2H),2.83-2.66(m,3H),2.63(s,6H),2.24-2.21(m,2H),2.01-1.93(m,2H)；MS(ESI)(m/z):[M+H]+550.2。
(S)-1-(1-(4-溴-5-(二氟甲氧基)-2-硝基苯基)吡咯烷-2-基)-N,N-二甲基甲胺(化合物18e)
Figure PCTCN2015088643-appb-000054
除了将化合物10替换成化合物17，N,N,N’-三甲基乙二胺替换成(S)-N,N-二甲基-1-(吡咯烷-2-基)-甲胺之外，与化合物11a同样地制备化合物18e。MS(ESI)(m/z):[M+H]+394.0。
(S)-4-(苯并[d]异恶唑-3-基)-N-(2-(二氟甲氧基)-4-(2-((二甲氨基)甲基)吡咯烷-1-基)-5-硝基苯基)嘧啶-2-胺(化合物19e)
Figure PCTCN2015088643-appb-000055
除了将化合物11a替换成化合物18e之外，与化合物12a同样地制备化合物19e。MS(ESI)(m/z):[M+H]+526.2。
(S)-N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-6-(二氟甲氧基)-4-(2-((二甲氨基)甲基)吡咯烷-1-基)苯-1,3-二胺(化合物20e)
Figure PCTCN2015088643-appb-000056
除了将化合物12a替换成化合物19e之外，与化合物13a同样地制备化合物20e。MS(ESI)(m/z):[M+H]+496.2。
实施例10：(S)-N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-(2-((二甲氨基)甲基)吡咯烷-1-基)苯基)丙烯酰胺(化合物21e)
Figure PCTCN2015088643-appb-000057
除了将化合物13a替换成化合物20e之外，与化合物14a同样地制备化合物21e。1H NMR(400MHz,CDCl3):δ9.87(s,1H),9.31(s,1H),8.66(d,J＝5.1Hz,1H),8.46(d,J＝8.0Hz,1H),7.73-7.55(m,3H),7.44-7.41(m,2H),6.96(s,1H),6.85-6.32(m,3H),5.75(d,J＝11.6Hz,1H),3.83-3.52(m,2H),3.01-2.95(m,1H),2.53(s,6H),2.32-1.95(m,4H),1.88-1.78(m,2H)；MS(ESI)(m/z):[M+H]+550.2。
2-(4-溴-5-(二氟甲氧基)-2-硝基苯基)-5-甲基-2,5-二氮杂螺[3.4]辛(化合物18f)
Figure PCTCN2015088643-appb-000058
除了将化合物10替换成化合物17，N,N,N’-三甲基乙二胺替换成5-甲基-2,5-二氮杂螺[3.4]辛烷之外，与化合物11a同样地制备化合物18f。1H NMR(400MHz,CDCl3):δ8.12(s,1H),6.84-6.36(m,2H),4.15(d,J＝9.2Hz,2H),3.73(d,J＝9.2Hz,2H),2.79(t,J＝7.2Hz,2H),2.51(s,3H),2.21-2.05(m,2H),1.86-1.79(m,2H)；MS(ESI)(m/z):[M+H]+392.0。
4-(苯并[d]异恶唑-3-基)-N-(2-(二氟甲氧基)-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)-5-硝基苯基)嘧啶-2胺(化合物19f)
Figure PCTCN2015088643-appb-000059
除了将化合物11a替换成化合物18f之外，与化合物12a同样地制备化合物19f。MS(ESI)(m/z):[M+H]+524.2。
N1-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-6-(二氟甲氧基)-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯-1,3-二胺(化合物20f)
Figure PCTCN2015088643-appb-000060
除了将化合物12a替换成化合物19f之外，与化合物13a同样地制备化合物20f。MS(ESI)(m/z):[M+H]+494.2。
实施例11：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯基)丙烯酰胺(化合物21f)
Figure PCTCN2015088643-appb-000061
除了将化合物13a替换成化合物20f之外，与化合物14a同样地制备化合物21f。1H NMR(400MHz,CDCl3):δ8.67(s,1H),8.60(d,J＝5.0Hz,1H),8.41(d,J＝7.6Hz,1H),7.72-7.48(m,4H),7.48-7.36(m,2H),6.81-6.30(m,4H),5.79(m,1H),4.10(d,J＝8.0Hz,2H),3.71(d,J＝7.7Hz,2H),2.93(m,2H),2.64(s,3H),2.31-2.17(m,2H),1.91-1.90(m,2H)；MS(ESI)(m/z):[M+H]+548.2。
化合物31a-31f的合成路线如方案3所示：
Figure PCTCN2015088643-appb-000062
方案3
1-甲基-3-(三丁基锡)-1H-吡咯并[3,2-b]吡啶(化合物25)
Figure PCTCN2015088643-appb-000063
将3-溴-1-甲基-1H-吡咯并[3,2-b]吡啶(化合物24，参考文献Bioorganic and Medicinal Chemistry Letters,2014,24,3238-3242合成)(2.4g,11.4mmol)溶于干燥的THF(60mL)中,在-70℃下滴加正丁基锂(2.5M正己烷溶液，5.0mL，12.5mmol)，滴加完成后体系保持-70℃继续搅拌30min。然后在相同温度下滴加三丁基氯化锡(4.06g,12.5mmol)，加完后缓慢升到室温搅拌3h。加入500mL乙酸乙酯,依次用水和饱和食盐水洗涤有机相，干燥浓缩后得到粗品黄色油状物(5.4g，纯度70％)。该粗品不用纯化可直接用于下一步反应。
3-(2-氯嘧啶-4-基)-1-甲基-1H-吡咯并[3,2-b]吡啶(化合物26)
Figure PCTCN2015088643-appb-000064
将化合物25(5.4g，纯度70％，9.0mmol)溶于DMF(70mL)中，依次加入2,4-二氯嘧啶(1.34g,9.0mmol)，氯化锂(1.89g,45.0mmol)和双三苯基磷二氯化钯(320mg,0.45mmol)。体系用氮气置换三次，加热到100℃反应16h。冷至室温后，加入大量乙酸乙酯，依次用水和饱和食盐水洗涤有机相。有机相干燥浓缩后过柱纯化得到淡黄色固体粗品(500mg，纯度80％)，该粗品不用纯化可直接用于下一步反应。MS(ESI)(m/z):[M+H]+245.0。
N-(4-氟-2-甲氧基-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物28)
Figure PCTCN2015088643-appb-000065
将化合物26(500mg，纯度80％，1.64mmol)溶于仲戊醇(20mL)，依次加入对甲苯磺酸(420mg,2.46mmol)和化合物27(310mg,1.64mmol)。将混合物加热到110℃反应18h。反应结束后，体系降到室温再继续搅拌30min后过滤，固体沉淀用少量仲戊醇洗涤后得到未干燥的黄色粗品化合物。该粗品用适量乙腈打浆过滤得到淡黄色固体化合物(650mg)。MS(ESI)(m/z):[M+H]+395.1。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-2-硝基苯-1,4-二胺(化合物29a)
Figure PCTCN2015088643-appb-000066
室温下向化合物28(650mg，1.65mmol)的N,N-二甲基乙酰胺(15mL)的溶液中依次加入N,N-二异丙基乙胺(280mg，2.14mmol)和N,N,N'-三甲基乙二胺(202mg，1.98mmol)。混合物加热到70℃反应6小时，反应完成后，加 入乙酸乙酯和水。充分搅拌，加硅藻土过滤，有机相用饱和食盐水多次洗涤，干燥浓缩后得到深红色固体化合物29a(450mg,57％)。1H NMR(400MHz,CDCl3):δ9.77(s,1H),δ8.65(s,1H),8.64-8.60(m,1H),8.46(d,J＝5.2Hz,1H),8.26(d,J＝5.2Hz,1H),7.72-7.68(m,1H),7.56(s,1H),7.24-7.20(m,1H),6.67(s,1H),3.99(s,6H),3.30(t,J＝7.2Hz,2H),3.01(s,3H),2.58(t,J＝7.2Hz,2H),2.27(s,6H)；MS(ESI)(m/z):MS(ESI)(m/z):[M+H]+477.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,2,4-三胺(化合物30a)
Figure PCTCN2015088643-appb-000067
将化合物29a(450mg,0.95mmol)溶于乙醇(18mL)/H2O(6mL)，加入铁粉(300mg,5.70mmol)和氯化铵(36mg,0.67mmol)。将体系加热至回流反应2h，反应结束后，反应物倒入二氯甲烷:甲醇＝10:1的溶液中，再加入适量水搅拌。加硅藻土过滤，有机相干燥浓缩后柱层析纯化(二氯甲烷:甲醇＝100:1至10:1)得到淡黄色固体30a(200mg,47％)。1H NMR(400MHz,CDCl3):δ8.62(dd,J＝4.8,1.6Hz,1H),8.45(d,J＝5.2Hz,1H),8.23(s,1H),8.17(d,J＝5.2Hz,1H),8.15(s,1H),7.69-7.65(m,1H),7.58(s,1H),7.24-7.18(m,1H),6.70(s,1H),3.89(s,3H),3.84(s,3H),3.04(t,J＝7.2Hz,2H),2.68(s,3H),2.50(t,J＝7.2Hz,2H),2.34(s,6H)；MS(ESI)(m/z):[M+H]+447.2。
实施例12：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物31a)
Figure PCTCN2015088643-appb-000068
将化合物30a(120mg，0.27mmol)溶于二氯甲烷(10mL)/叔丁醇(1mL)，在0-5℃下依次加入三乙胺(50mg，0.54mmol)，EDCI(100mg，0.54mmol)和丙烯酸(40mg，0.54mmol)。室温搅拌2h，加入适量乙酸乙酯，用水洗涤 有机相。有机相干燥浓缩后柱层析纯化得到类白色固体31a(80mg，59％)。1H NMR(400MHz,CDCl3):δ10.24(bs,1H),9.94(s,1H),9.56(bs,1H),8.61-8.57(m,1H),8.42(d,J＝5.2Hz,1H),8.24(d,J＝5.2Hz,1H),7.77(s,1H),7.70-7.65(m,1H),7.21-7.15(m,1H),6.80(s,1H),6.50-6.34(m,2H),5.72(m,1H),4.04(s,3H),3.89(s,3H),2.90(t,J＝7.2Hz,2H),2.71(s,3H),2.30-2.22(m,8H)；MS(ESI)(m/z):[M+H]+501.3。
N-(2-甲氧基-4-(4-甲基哌嗪-1-基)-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物29b)
Figure PCTCN2015088643-appb-000069
除了将N,N,N’-三甲基乙二胺替换成4-甲基哌嗪之外，与化合物29a同样地制备化合物29b。MS(ESI)(m/z):[M+H]+475.1。
6-甲氧基-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-4-(4-甲基哌嗪-1-基)苯-1,3-二胺(化合物30b)
Figure PCTCN2015088643-appb-000070
除了将化合物29a替换成化合物29b之外，与化合物30a同样地制备化合物30b。1H NMR(400MHz,CDCl3):δ8.63-8.59(m,1H),8.44(d,J＝5.2Hz,1H),8.22(s,1H),8.17(d,J＝5.2Hz,1H),8.15(s,1H),7.70-7.64(m,1H),7.56(s,1H),7.24-7.18(m,1H),6.71(s,1H),3.89(s,3H),3.84(s,3H),2.94(m,4H),2.58(m,4H),2.37(s,3H)；MS(ESI)(m/z):[M+H]+445.2。
实施例13：N-(4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)-2-(4-甲基哌嗪-1-基)苯基)丙烯酰胺(化合物31b)
Figure PCTCN2015088643-appb-000071
除了将化合物30a替换成化合物30b之外，与化合物31a同样地制备化合物31b。1H NMR(400MHz,CDCl3):δ9.91(bs,1H),9.47(s,1H),8.80(bs,1H),8.59-8.55(m,1H),8.42(d,J＝5.2Hz,1H),8.24(d,J＝5.2Hz,1H),7.77(s,1H),7.67-7.63(m,1H),7.20-7.16(m,1H),6.80(s,1H),6.48-6.30(m,2H),5.78(m,1H),4.02(s,3H),3.88(s,3H),2.92(m,4H),2.62(m,4H),2.41(s,3H)；MS(ESI)(m/z):[M+H]+499.2。
N-(4-(3-(二甲氨基)氮杂环丁-1-基)-2-甲氧基-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物29c)
Figure PCTCN2015088643-appb-000072
除了将N,N,N’-三甲基乙二胺替换成N,N-二甲基氮杂环丁-3-胺之外，与化合物29a同样地制备化合物29c。1H NMR(400MHz,CDCl3):δ9.82(s,1H),8.73(s,1H),8.64(d,J＝4.4Hz,1H),8.47(d,J＝5.1Hz,1H),8.27(d,J＝5.2Hz,1H),7.73(d,J＝8.1Hz,1H),7.54(s,1H),7.26-7.23(m,1H),6.07(s,1H),4.22(t,J＝7.6Hz,2H),4.01(s,6H),3.80-3.72(m,2H),3.25-3.22(m,1H),2.25(s,6H)；MS(ESI)(m/z):[M+H]+475.2。
4-(3-(二甲氨基)氮杂环丁-1-基)-6-甲氧基-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,3-二胺(化合物30c)
Figure PCTCN2015088643-appb-000073
除了将化合物29a替换成化合物29c之外，与化合物30a同样地制备化合物30c。MS(ESI)(m/z):[M+H]+445.2。
实施例14：N-(2-(3-(二甲氨基)氮杂环丁-1-基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物31c)
Figure PCTCN2015088643-appb-000074
除了将化合物30a替换成化合物30c之外，与化合物31a同样地制备化合物31c。1H NMR(400MHz,CDCl3):δ9.40(s,1H),9.23(s,1H),8.60(d,J＝4.5Hz,1H),8.42(d,J＝5.2Hz,1H),8.21(d,J＝5.2Hz,1H),7.76(s,1H),7.67(d,J＝8.0Hz,1H),7.62(s,1H),7.21-7.18(m,1H),6.56-6.40(m,3H),5.80-5.78(m,1H),4.01(s,3H),3.92(s,3H),3.88(t,J＝6.8Hz,2H),3.64(t,J＝6.5Hz,2H),3.17-3.11(m,1H),2.24(s,6H)；MS(ESI)(m/z):[M+H]+499.2。
N-(4-(4-(二甲氨基)哌啶-1-基)-2-甲氧基-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物29d)
Figure PCTCN2015088643-appb-000075
除了将N,N,N’-三甲基乙二胺替换成N,N-二甲基哌啶-4-胺之外，与化合物29a同样地制备化合物29d。1H NMR(400MHz,CDCl3):δ9.86(s,1H),8.67(s,1H),8.63-8.58(m,1H),8.46(d,J＝5.2Hz,1H),8.26(d,J＝5.2Hz,1H),7.73-7.67(m,1H),7.56(s,1H),7.24-7.18(m,1H),6.59(s,1H),3.99(s,3H),3.98(s,3H),3.38(m,2H),2.85(m,2H),2.48-2.22(m,7H),1.98-1.74(m,4H)；MS(ESI)(m/z):[M+H]+503.2。
4-(4-(二甲氨基)哌啶-1-基)-6-甲氧基-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,3-二胺(化合物30d)
Figure PCTCN2015088643-appb-000076
除了将化合物29a替换成化合物29d之外，与化合物30a同样地制备化 合物30d。1H NMR(400MHz,CDCl3):δ8.63-8.59(m,1H),8.45(d,J＝5.2Hz,1H),8.23(s,1H),8.17(d,J＝5.2Hz,1H),8.14(s,1H),7.70-7.65(m,1H),7.54(s,1H),7.24-7.18(m,1H),6.67(s,1H),3.89(s,3H),3.84(s,3H),3.18(m,2H),2.68-2.60(m,2H),2.34(s,6H),2.28(m,1H),1.95(m,2H),1.68(m,2H)；MS(ESI)(m/z):[M+H]+473.2。
实施例15：N-(2-(4-(二甲氨基)哌啶-1-基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物31d)
Figure PCTCN2015088643-appb-000077
除了将化合物30a替换成化合物30d之外，与化合物31a同样地制备化合物31d。1H NMR(400MHz,CDCl3):δ9.89(bs,1H),9.48(s,1H),8.77(bs,1H),8.61-8.57(m,1H),8.42(d,J＝5.2Hz,1H),8.24(d,J＝5.2Hz,1H),7.74(s,1H),7.69-7.65(m,1H),7.21-7.15(m,1H),6.76(s,1H),6.44-6.30(m,2H),5.78(m,1H),4.02(s,3H),3.89(s,3H),3.03(m,2H),2.74(m,2H),2.36(s,6H),2.19(m,1H),2.04(m,2H),1.64(m,2H)；MS(ESI)(m/z):[M+H]+527.2。
(S)-N-(4-(2-((二甲氨基)甲基)吡咯烷-1-基)-2-甲氧基-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物29e)
Figure PCTCN2015088643-appb-000078
除了将N,N,N’-三甲基乙二胺替换成(S)-N,N-二甲基-1-(吡咯烷-2-基)-甲胺之外，与化合物29a同样地制备化合物29e。MS(ESI)(m/z):[M+H]+503.2。
(S)-4-(2-((二甲氨基)甲基)吡咯烷-1-基)-6-甲氧基-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,3-二胺(化合物30e)
Figure PCTCN2015088643-appb-000079
除了将化合物29a替换成化合物29e之外，与化合物30a同样地制备化合物30e。MS(ESI)(m/z):[M+H]+473.3。
实施例16：(S)-N-(2-(2-((二甲氨基)甲基)吡咯烷-1-基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物31e)
Figure PCTCN2015088643-appb-000080
除了将化合物30a替换成化合物30e之外，与化合物31a同样地制备化合物31e。1H NMR(400MHz,CDCl3):δ9.83(s,1H),9.43(s,1H),9.39(s,1H),8.65-8.56(m,1H),8.43(d,J＝5.2Hz,1H),8.27(d,J＝5.2Hz,1H),7.75(s,1H),7.73-7.67(m,1H),7.22-7.19(m,1H),7.06(d,J＝28.2Hz,1H),6.68(s,1H),6.51-6.46(m,1H),5.79-5.76(m,1H),4.02(s,3H),3.91(s,3H),3.75-3.66(m,1H),2.93-2.81(m,2H),2.65(s,6H),2.40-2.21(m,2H),2.19-1.89(m,4H)；MS(ESI)(m/z):[M+H]+527.3。
N-(2-甲氧基-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物29f)
Figure PCTCN2015088643-appb-000081
除了将N,N,N’-三甲基乙二胺替换成5-甲基-2,5-二氮杂螺[3.4]辛烷之外，与化合物29a同样地制备化合物29f。1H NMR(400MHz,CDCl3):δ9.75(s,1H),8.70(s,1H),8.65-8.58(m,1H),8.44(d,J＝5.2Hz,1H),8.24(d,J＝5.2Hz,1H),7.73-7.67(m,1H),7.50(s,1H),7.24-7.18(m,1H),6.05(s,1H),4.13(m,2H),3.98(s,6H),3.76(m,2H),2.74(m,2H),2.50(s,3H),2.12(m,2H),1.81(m,2H)；MS(ESI)(m/z):[M+H]+501.2。
6-甲氧基-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯-1,3-二胺(化合物30f)
Figure PCTCN2015088643-appb-000082
除了将化合物29a替换成化合物29f之外，与化合物30a同样地制备化合物30f。1H NMR(400MHz,CDCl3):δ8.63-8.59(m,1H),8.44(d,J＝5.2Hz,1H),8.23(s,1H),8.13(d,J＝5.2Hz,1H),8.05(s,1H),7.70-7.64(m,1H),7.43(s,1H),7.22-7.17(m,1H),6.35(s,1H),4.13-3.54(m,10H),2.76(m,2H),2.53(s,3H),2.20(m,2H),1.82(m,2H)；MS(ESI)(m/z):[M+H]+471.2。
实施例17：N-(4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)-2-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯基)丙烯酰胺(化合物31f)
Figure PCTCN2015088643-appb-000083
除了将化合物30a替换成化合物30f之外，与化合物31a同样地制备化合物31f。1H NMR(400MHz,CDCl3):δ9.22(bs,1H),9.14(s,1H),8.59-8.55(m,1H),8.40(d,J＝5.2Hz,1H),8.18(d,J＝5.2Hz,1H),7.68-7.64(m,1H),7.54(s,1H),7.41(s,1H),7.20-7.14(m,1H),6.50-6.30(m,3H),5.76(m,1H),3.99(s,3H),3.92(m,2H),3.91(s,3H),3.76-3.64(m,2H),2.78-2.74(m,2H),2.53(s,3H),2.22-2.18(m,2H),1.86-1.82(m,2H)；MS(ESI)(m/z):[M+H]+525.2。
化合物39a-39f的合成路线如方案4所示：
Figure PCTCN2015088643-appb-000084
方案4
2-(二氟甲氧基)-4-氟-1-硝基苯(化合物33)
Figure PCTCN2015088643-appb-000085
将化合物32(10g，63.7mmol)溶于DMF(100mL)中，搅拌均匀，然后加入碳酸钠(40.5g，382.1mmol)，油浴升温至90℃后再慢慢加入二氟氯乙酸(29.1g，223.0mmol)，1h后TLC监控反应完全。后处理将反应液慢慢倒入冰水(200mL)中，用乙酸乙酯(100mL，2次)萃取，有机相用饱和食盐水洗两遍，适量硫酸钠干燥，柱层析纯化，最终得到黄色油状目标产物33(11g，83.44％)。1H NMR(400MHz,CDCl3):δ8.08-8.04(m,1H),7.21-7.09(m,2H),6.88-6.48(m,1H)；MS(ESI)(m/z):[M+H]+208.0。
2-(二氟甲氧基)-4-氟-1-苯胺(化合物34)
Figure PCTCN2015088643-appb-000086
将化合物33(11g，53.11mmol)溶于乙醇(100mL)中然后加入Pd/C(2.2g)，用H2置换3次，用氢气球加氢反应，室温反应过夜。后处理用硅藻土抽滤反应液，滤液直接旋干得到较纯产物34(9.0g，95.74％)，棕色油状。1H NMR(400MHz,CDCl3)δ6.89-6.83(m,1H),6.83-6.71(m,2H),6.70-6.31(m,1H),3.59(s,2H)；MS(ESI)(m/z):[M+H]+178.0。
2-(二氟甲氧基)-4-氟-5-硝基苯胺(化合物35)
Figure PCTCN2015088643-appb-000087
将化合物34(9g，50.8mmol)溶于浓硫酸(100mL)中，冰水浴降温，慢慢加入硝酸钾(5.14g，50.8mmol)，1h反应完全，将反应液倒入冰水中淬灭，二氯甲烷萃取，硫酸钠干燥，柱层析纯化得到目标产物35(10.4g，92.15％)，橙红色固体。MS(ESI)(m/z):[M+H]+223.0。
N-(2-(二氟甲氧基)-4-氟-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物36)
Figure PCTCN2015088643-appb-000088
将化合物26(1.2g,，纯度60％，2.95mmol)溶于仲戊醇(30mL)，依次加入对甲苯磺酸(760mg，4.4mmol)和化合物35(650mg，2.95mmol)。将混合物加热到110℃反应4h。反应结束后，体系降到室温再继续搅拌30min后过滤，固体沉淀用少量仲戊醇洗涤后得到未干燥的黄色粗品化合物36。该粗品用适量乙腈打浆过滤得到淡黄色固体化合物36(800mg，63％)。MS(ESI)(m/z):[M+H]+431.1。
2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-5-硝基苯-1,4-二胺(化合物37a)
Figure PCTCN2015088643-appb-000089
室温下向化合物36(600mg，1.39mmol)的N,N-二甲基乙酰胺(15mL)的溶液中依次加入N,N-二异丙基乙胺(233mg，1.80mmol)和N,N,N'-三甲基乙二胺(170mg，1.67mmol)。混合物加热到80℃反应2h，反应完成后，加入乙酸乙酯和水。充分搅拌，加硅藻土过滤，有机相用饱和食盐水多次洗涤，干燥浓缩后得到深红色固体化合物37a(680mg，96％)。1H NMR(400MHz,CDCl3):δ9.68(s,1H),8.62-8.58(m,1H),8.48(s,1H),8.44(d,J＝5.2Hz,1H),8.28(d,J＝5.2Hz,1H),7.70-7.64(m,1H),7.36(s,1H),7.23-7.19(m,1H),7.01(s,1H),6.67(t,J＝72Hz,1H),3.95(s,3H),3.23(t,J＝7.2Hz,2H),2.88(s,3H),2.54(t,J＝7.2Hz,2H),2.26(s,6H)；MS(ESI)(m/z):[M+H]+513.2。
5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,2,4-三胺)(化合物38a)
Figure PCTCN2015088643-appb-000090
将化合物37a(700mg，1.37mmol)溶于乙醇(15mL)/H2O(5mL)，加入铁粉(460mg，8.2mmol)和氯化铵(50mg，0.96mmol)。将体系加热至回流反应2h，反应结束后，反应物倒入二氯甲烷:甲醇＝10:1的溶液中，再加入适量水搅拌。加硅藻土过滤，有机相干燥浓缩后柱层析纯化得到淡黄色固体38a(300mg，45％)。1H NMR(400MHz,CDCl3):δ8.63-8.59(m,1H),8.45(d,J＝5.2Hz,1H),8.24-8.18(m,2H),8.05(s,1H),7.70-7.65(m,1H),7.28(s,1H),7.23-7.18(m,1H),6.88(s,1H),6.44(t,J＝72Hz,1H),3.89(s,3H),2.95(t,J＝7.2Hz,2H),2.66(s,3H),2.45(t,J＝7.2Hz,2H),2.30(s,6H)；MS(ESI)(m/z):[M+H]+483.2。
实施例18：N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物39a)
Figure PCTCN2015088643-appb-000091
将化合物38a(120mg，0.25mmol)溶于二氯甲烷(10mL)/叔丁醇(1mL)，在0-5℃下依次加入三乙胺(50mg，0.50mmol)，EDCI(96mg，0.50mmol)和丙烯酸(36mg，0.50mmol)。室温搅拌2h，加入适量乙酸乙酯，用水洗涤有机相。有机相干燥浓缩后柱层析纯化得到类白色固体39a(60mg，45％)。1H NMR(400MHz,CDCl3):δ10.36(bs,1H),10.01(s,1H),9.47(bs,1H),8.62-8.58(m,1H),8.44(d,J＝5.2Hz,1H),8.30(d,J＝5.2Hz,1H),7.70-7.65(m,1H),7.51(s,1H),7.21-7.15(m,1H),7.07(s,1H),6.68-6.32(m,3H),5.76(m,1H),4.01(s,3H),2.86(t,J＝7.2Hz,2H),2.70(s,3H),2.32-2.22(m,8H)；MS(ESI)(m/z):[M+H]+537.2。
N-(2-(二氟甲氧基)-4-(4-甲基哌嗪-1-基)-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物37b)
Figure PCTCN2015088643-appb-000092
除了将N,N,N’-三甲基乙二胺替换成4-甲基哌嗪之外，与化合物37a同样地制备化合物37b。MS(ESI)(m/z):[M+H]+511.2。
6-(二氟甲氧基)-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-4-(4-甲基哌嗪-1-基)苯-1,3-二胺(化合物38b)
Figure PCTCN2015088643-appb-000093
除了将化合物37a替换成化合物37b之外，与化合物38a同样地制备化合物38b。1H NMR(400MHz,CDCl3):δ8.64-8.60(m,1H),8.45(d,J＝5.2Hz,1H),8.24-8.18(m,2H),8.09(s,1H),7.71-7.67(m,1H),7.27(s,1H),7.22-7.18(m,1H),6.88(s,1H),6.43(t,J＝72Hz,1H),4.04(bs,2H),3.91(s,3H),2.92(m,4H),2.58(m,4H),2.37(s,3H)；MS(ESI)(m/z):[M+H]+481.2。
实施例19：N-(4-(二氟甲氧基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)-2-(4-甲基哌嗪-1-基)苯基)丙烯酰胺(化合物39b)
Figure PCTCN2015088643-appb-000094
除了将化合物38a替换成化合物38b之外，与化合物39a同样地制备化合物39b。1H NMR(400MHz,CDCl3):δ9.96(bs,1H),9.38(s,1H),8.85(bs,1H),8.62-8.58(m,1H),8.44(d,J＝5.2Hz,1H),8.32(d,J＝5.2Hz,1H),7.70-7.56(m,1H),7.50(s,1H),7.20-7.16(m,1H),7.05(s,1H),6.70-6.30(m,3H),5.82(m,1H),4.00(s,3H),2.92(m,4H),2.62(m,4H),2.41(s,3H)；MS(ESI)(m/z):[M+H]+535.2。
N-(2-(二氟甲氧基)-4-(3-(二甲氨基)氮杂环丁-1-基)-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物37c)
Figure PCTCN2015088643-appb-000095
除了将N,N,N’-三甲基乙二胺替换成N,N-二甲基氮杂环丁-3-胺之外，与化合物37a同样地制备化合物37c。1H NMR(400MHz,CDCl3):δ9.81(s,1H),8.65-8.63(m,1H),8.62(s,1H),8.48(d,J＝5.2Hz,1H),8.32(d,J＝5.2Hz,1H),7.74-7.71(m,1H),7.34(s,1H),7.27-7.23(m,1H),6.66(t,J＝72.4Hz,1H),6.43(s,1H),4.16(t,J＝7.7Hz,2H),4.00(s,3H),3.76-3.72(m,2H),3.28-3.18(m,1H),2.24(s,6H)；MS(ESI)(m/z):[M+H]+511.2。
6-(二氟甲氧基)-4-(3-(二甲氨基)氮杂环丁-1-基)-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,3-二胺(化合物38c)
Figure PCTCN2015088643-appb-000096
除了将化合物37a替换成化合物37c之外，与化合物38a同样地制备化合物38c。MS(ESI)(m/z):[M+H]+481.2。
实施例20：N-(4-(二氟甲氧基)-2-(3-(二甲氨基)氮杂环丁-1-基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物39c)
Figure PCTCN2015088643-appb-000097
除了将化合物38a替换成化合物38c之外，与化合物39a同样地制备化合物39c。1H NMR(400MHz,CDCl3):δ9.56(s,1H),9.25(s,1H),8.62(d,J＝3.8Hz,1H),8.46(d,J＝5.2Hz,1H),8.30(d,J＝5.2Hz,1H),7.82(s,1H),7.71(d,J＝8.3Hz,1H),7.41(s,1H),7.23-7.21(m,1H),6.80(s,1H),6.77-6.33(m,3H),5.85(d,J＝8.9Hz,1H),4.03(s,3H),3.90(t,J＝6.5Hz,2H),3.66(t,J＝6.6Hz,2H),3.28-3.14(m,1H),2.28(s,6H)；MS(ESI)(m/z):[M+H]+535.2。
N-(2-(二氟甲氧基)-4-(4-(二甲氨基)哌啶-1-基)-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物37d)
Figure PCTCN2015088643-appb-000098
除了将N,N,N’-三甲基乙二胺替换成N,N-二甲基哌啶-4-胺之外，与化合物37a同样地制备化合物37d。1H NMR(400MHz,CDCl3):δ9.78(s,1H),8.63-8.59(m,1H),8.53(s,1H),8.46(d,J＝5.2Hz,1H),8.30(d,J＝5.2Hz,1H),7.73-7.65(m,1H),7.42(s,1H),7.25-7.19(m,1H),6.92(s,1H),6.65(t,J＝72Hz,1H),3.95(s,3H),3.32(m,2H),2.82(m,2H),2.44-2.22(m,7H),1.98-1.74(m,4H)；MS(ESI)(m/z):[M+H]+539.2。
6-(二氟甲氧基)-4-(4-(二甲氨基)哌啶-1-基)-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,3-二胺(化合物38d)
Figure PCTCN2015088643-appb-000099
除了将化合物37a替换成化合物37d之外，与化合物38a同样地制备化合物38d。1H NMR(400MHz,CDCl3):δ8.62-8.58(m,1H),8.44(d,J＝5.2Hz,1H),8.22-8.16(m,2H),8.05(s,1H),7.68-7.63(m,1H),7.28(s,1H),7.23-7.17(m,1H),6.83(s,1H),6.44(t,J＝72Hz,1H),4.05(bs,2H),3.87(s,3H),3.18(m,2H),2.64-2.54(m,2H),2.34(s,6H),2.26(m,1H),1.95(m,2H),1.68(m,2H)；MS(ESI)(m/z):[M+H]+509.2。
实施例21：N-(4-(二氟甲氧基)-2-(4-(二甲氨基)哌啶-1-基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物39d)
Figure PCTCN2015088643-appb-000100
除了将化合物38a替换成化合物38d之外，与化合物39a同样地制备化 合物39d。1H NMR(400MHz,CDCl3):δ9.95(s,1H),9.40(s,1H),8.84(s,1H),8.62-8.58(m,1H),8.44(d,J＝5.2Hz,1H),8.30(d,J＝5.2Hz,1H),7.71-7.65(m,1H),7.49(s,1H),7.20-7.16(m,1H),7.02(s,1H),6.70-6.30(m,3H),5.80(m,1H),4.01(s,3H),3.07-3.03(m,2H),2.76-2.70(m,2H),2.36(s,6H),2.19(m,1H),2.04(m,2H),1.64(m,2H)；MS(ESI)(m/z):[M+H]+563.2。
(S)-N-(2-(二氟甲氧基)-4-(2-((二甲氨基)甲基)吡咯-1-基)-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物37e)
Figure PCTCN2015088643-appb-000101
除了将N,N,N’-三甲基乙二胺替换成(S)-N,N-二甲基-1-(吡咯烷-2-基)-甲胺之外，与化合物37a同样地制备化合物37e。1H NMR(400MHz,CDCl3):δ9.75(s,1H),8.66-8.64(m,1H),8.61(s,1H),8.49(d,J＝5.2Hz,1H),8.31(d,J＝5.2Hz,1H),7.74-7.72(m,1H),7.35(s,1H),7.26-7.24(m,1H),7.07(s,1H),6.71(t,J＝72.4Hz,1H),4.01(s,3H),3.64-3.60(m,1H),2.75-2.59(m,2H),2.48-2.28(m,8H),2.09-1.74(m,4H)；MS(ESI)(m/z):[M+H]+539.2。
(S)-6-(二氟甲氧基)-4-(2-((二甲氨基)甲基)吡咯烷-1-基)-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,3-二胺(化合物38e)
Figure PCTCN2015088643-appb-000102
除了将化合物37a替换成化合物37e之外，与化合物38a同样地制备化合物38e。MS(ESI)(m/z):[M+H]+509.3。
实施例22：(S)-N-(4-(二氟甲氧基)-2-(2-((二甲氨基)甲基)吡咯烷-1-基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物39e)
Figure PCTCN2015088643-appb-000103
除了将化合物38a替换成化合物38e之外，与化合物39a同样地制备化合物39e。1H NMR(400MHz,CDCl3)δ9.82(s,1H),9.73(s,1H),9.28(s,1H),8.59-8.58(m,1H),8.43(d,J＝5.2Hz,1H),8.30(d,J＝5.2Hz,1H),7.69-7.67(m,1H),7.53(d,J＝18.2Hz,1H),7.20-7.18(m,1H),7.03(s,1H),6.93(s,1H),6.82-6.39(m,2H),5.81-5.78(m,1H),3.98(s,3H),3.81-3.59(m,2H),2.90-2.84(m,2H),2.56(s,6H),2.24-2.20(m,1H),2.10-1.72(m,4H)；MS(ESI)(m/z):[M+H]+563.3。
N-(2-(二氟甲氧基)-4-(5-甲基-2,5二氮杂螺[3.4]辛-2-基)-5-硝基苯基)-4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-胺(化合物37f)
Figure PCTCN2015088643-appb-000104
除了将N,N,N’-三甲基乙二胺替换成5-甲基-2,5-二氮杂螺[3.4]辛烷之外，与化合物37a同样地制备化合物37f。1H NMR(400MHz,CDCl3):δ9.74(s,1H),8.64-8.58(m,1H),8.58(s,1H),8.44(d,J＝5.2Hz,1H),8.28(d,J＝5.2Hz,1H),7.72-7.64(m,1H),7.28(s,1H),7.24-7.18(m,1H),6.64(t,J＝72Hz,1H),6.40(s,1H),4.10(m,2H),3.90(s,3H),3.70(m,2H),2.74(m,2H),2.50(s,3H),2.12(m,2H),1.81(m,2H)；MS(ESI)(m/z):[M+H]+537.2。
6-(二氟甲氧基)-N1-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)-4-(5-甲基-2,5二氮杂螺[3.4]辛-2-基)苯-1,3-二胺(化合物38f)
Figure PCTCN2015088643-appb-000105
除了将化合物37a替换成化合物37f之外，与化合物38a同样地制备化合物38f。1H NMR(400MHz,CDCl3):8.63-8.59(m,1H),8.43(d,J＝5.2Hz, 1H),8.24-8.16(m,2H),7.93(s,1H),7.69-7.65(m,1H),7.23-7.18(m,1H),7.15(s,1H),6.50(s,1H),6.45(t,J＝72Hz,1H),3.94-3.84(m,5H),3.60(m,2H),2.74(m,2H),2.50(s,3H),2.18(m,2H),1.81(m,2H)；MS(ESI)(m/z):[M+H]+507.2。
实施例23：N-(4-(二氟甲氧基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)-2-(5-甲基-2,5二氮杂螺[3.4]辛-2-基)苯基)丙烯酰胺(化合物39f)
Figure PCTCN2015088643-appb-000106
除了将化合物38a替换成化合物38f之外，与化合物39a同样地制备化合物39f。1H NMR(400MHz,CDCl3):δ9.40(bs,1H),9.16(s,1H),8.62-8.58(m,1H),8.42(d,J＝5.2Hz,1H),8.26(d,J＝5.2Hz,1H),7.71-7.65(m,1H),7.50(s,1H),7.32(s,1H),7.20-7.15(m,1H),6.74-6.30(m,4H),5.82(m,1H),3.98(s,3H),3.95-3.92(m,2H),3.64-3.62(m,2H),2.81-2.77(m,2H),2.53(s,3H),2.22-2.18(m,2H),1.86-1.82(m,2H)；MS(ESI)(m/z):[M+H]+561.2。
化合物51a-51f的合成路线如方案5所示：
Figure PCTCN2015088643-appb-000107
方案5
化合物43的合成参照文献synthesis(Germany),2014,第46卷，第96-100页，化合物46的合成参照专利申请WO 2013030138。
3-(2-氯嘧啶-4-基)-1-甲基-1H-噻吩并[3,2-c]吡唑(化合物47)
Figure PCTCN2015088643-appb-000108
在100mL的单口瓶中将1-甲基-(三丁基锡)-1H-噻吩并[3,2-c]吡唑46(1.5g，3.5mmol)溶于DMF(50mL)中，然后加入2,4-二氯嘧啶(0.778g，5.3mmol)，碘化亚铜(0.138g,0.7mmol)和四(三苯基膦)钯(0.202g，0.175mmol)。在80℃氮气保护下反应1小时。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得产物(0.8g，3.2mmol)。MS(ESI)(m/z):[M+H]+251.1。
N-(4-氟-2-甲氧基-5-硝基苯基)-4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-胺(化合物48a)
Figure PCTCN2015088643-appb-000109
在100mL的单口瓶中将化合物47(600mg，2.4mmol)溶于2-戊醇(50mL)中，然后加入化合物27(580mg，3.1mmol)和对甲苯磺酸(1.2g，7.2mmol)。在110℃反应3h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得产物(300mg，0.75mmol)。MS(ESI)(m/z):[M+H]+401.1。
N-(2-(二氟甲氧基)-4-氟-5-硝基苯基)-4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-胺(化合物48b)
Figure PCTCN2015088643-appb-000110
除了将化合物27换成化合物35之外，与化合物48a同样地制备化合物48b。MS(ESI)(m/z):[M+H]+437.1。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)-2-硝基苯-1,4-二胺(化合物49a)
Figure PCTCN2015088643-appb-000111
在100mL的单口瓶中将化合物48a(400mg,1.0mmol)溶于二甲基乙酰胺(50mL)中，然后加入N,N,N'-三甲基乙二胺(153mg，1.5mmol)和N,N-二异丙基乙胺(387mg，3.0mmol)。在80℃反应2h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物49a(300mg，0.622mmol)。MS(ESI)(m/z):[M+H]+483.3。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,2,4-三胺(化合物50a)
Figure PCTCN2015088643-appb-000112
在100mL的单口瓶中将化合物49a(300mg，0.622mmol)溶于乙醇(20ml)中，然后加入饱和氯化铵溶液(20ml)和铁粉(174mg，3.11mmol)。在50℃反应2h。加入50mL水，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物50a(150mg，0.33mmol)。MS(ESI)(m/z):[M+H]+453.2。
实施例24：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物51a)
Figure PCTCN2015088643-appb-000113
在100mL的单口瓶中将化合物50a(150mg，0.33mmol)溶于20mL二氯甲烷中，然后加入丙烯酸(28.7mg，0.39mmol)，EDCI(127mg,0.66mmol)和三乙胺(100mg，0.99mmol)，室温反应2h。加入水30(mL)，二氯甲烷萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物51a(30mg，0.059mmol).1H NMR(400MHz,CDCl3):δ10.20(s,1H),9.47(s,1H),8.57(d,J＝5.1Hz,1H),7.62(s,1H),7.43(d,J＝5.3Hz,1H),7.37(d,J＝5.2Hz,1H),6.93(d,J＝5.3Hz, 1H),6.81(s,1H),6.45-6.39(m,2H),5.70-5.67(m,1H),4.13(s,3H),3.90(s,3H),2.93(s,2H),2.74(s,3H),2.26-2.33(m,8H).MS(ESI)(m/z):[M+H]+507.2。
2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-N1-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)-5-硝基苯-1,4-二胺(化合物49b)
Figure PCTCN2015088643-appb-000114
除了将化合物48a换成化合物48b之外，与化合物49a同样地制备化合物49b。MS(ESI)(m/z):[M+H]+519.2。
5-(二氟甲氧基-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,2,4-三胺)(化合物50b)
Figure PCTCN2015088643-appb-000115
除了将化合物49a换成化合物49b之外，与化合物50a同样地制备化合物50b。1H NMR(400MHz,CDCl3):δ8.47(d,J＝5.1Hz,1H),7.73(s,1H),7.39(s,1H),7.35(d,J＝5.3Hz,1H),7.11(d,J＝5.3Hz,1H),6.93(d,J＝5.1Hz,1H),6.86(s,1H),6.45(t,J＝74.4Hz,1H),4.43(s,3H),3.00(t,J＝6.4Hz,2H),2.66(s,3H),2.61(t,J＝6.0Hz,2H),2.42(s,6H).MS(ESI)(m/z):[M+H]+489.2。
实施例25：N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物51b)
Figure PCTCN2015088643-appb-000116
除了将化合物50a换成化合物50b之外，与化合物51a同样地制备化合 物51b。1H NMR(400MHz,CDCl3):δ10.34(s,1H),9.40(s,1H),8.60(d,J＝5.2Hz,1H),7.39(d,J＝5.4Hz,1H),7.31(s,1H),7.17(d,J＝5.4Hz,1H),7.09(s,1H),7.03(d,J＝5.2Hz,1H),6.71-6.33(m,3H),5.76-5.73(m,1H),4.48(s,3H),2.89-2.87(m,2H),2.74(s,3H),2.36-2.34(m,2H),2.30(s,6H).MS(ESI)(m/z):[M+H]+543.3。
N-(2-(二氟甲氧基)-4-(4-甲基哌嗪-1-基)-5-硝基苯基)-4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-胺(化合物49c)
Figure PCTCN2015088643-appb-000117
除了将化合物48a换成化合物48b，N,N,N’-三甲基乙二胺替换成4-甲基哌嗪之外，与化合物49a同样地制备化合物49c。MS(ESI)(m/z):[M+H]+517.1。
6-(二氟甲氧基)-N1-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)-4-(4-甲基哌嗪-1-基)苯-1,3-二胺(化合物50c)
Figure PCTCN2015088643-appb-000118
除了将化合物49a换成化合物49c之外，与化合物50a同样地制备化合物50c。MS(ESI)(m/z):[M+H]+487.1。
实施例26：N-(4-(二氟甲氧基)-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)氨基)-2-(4-甲基哌嗪-1-基)苯基)丙烯酰胺(化合物51c)
Figure PCTCN2015088643-appb-000119
除了将化合物50a换成化合物50c之外，与化合物51a同样地制备化合物51c。1H NMR(400MHz,CDCl3):δ9.36(s,1H),8.62(s,1H),8.58(d,J＝5.2Hz,1H),7.36(d,J＝5.2Hz,1H),7.31(s,1H),7.14(d,J＝5.2Hz,1H),7.06(s, 1H),7.02(d,J＝5.2Hz,1H),6.50(t,J＝72Hz,1H),6.44-6.22(m,2H),5.82-5.78(m,1H),4.47(s,3H),2.93-2.89(m,4H),2.64-2.60(m,4H),2.39(s,3H)；MS(ESI)(m/z):[M+H]+541.2。
N-(2-(二氟甲氧基)-4-(3-(二甲氨基)氮杂环丁-1-基)-5-硝基苯基)-4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-胺(化合物49d)
Figure PCTCN2015088643-appb-000120
除了将化合物48a换成化合物48b，N,N,N’-三甲基乙二胺替换成N,N-二甲基氮杂环丁-3-胺之外，与化合物49a同样地制备化合物49d。1H NMR(400MHz,CDCl3):δ8.88(s,1H),8.54(d,J＝5.2Hz,1H),7.38(d,J＝5.2Hz,1H),7.24(s,1H),7.17(d,J＝5.2Hz,1H),7.04(d,J＝5.2Hz,1H),6.62(t,J＝72Hz,1H),6.40(s,1H),4.52(s,3H),4.20-4.05(m,2H),3.78-3.62(m,2H),3.26-3.16(m,1H),2.20(s,6H)；MS(ESI)(m/z):[M+H]+517.2。
6-(二氟甲氧基)-4-(3-(二甲氨基)氮杂环丁-1-基)-N1-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,3-二胺(化合物50d)
Figure PCTCN2015088643-appb-000121
除了将化合物49a换成化合物49d之外，与化合物50a同样地制备化合物50d。1H NMR(400MHz,CDCl3):δ8.51(d,J＝5.2Hz,1H),7.65(s,1H),δ7.38(d,J＝5.2Hz,1H),7.22(s,1H),7.16(d,J＝5.2Hz,1H),6.96(d,J＝5.2Hz,1H),6.52(s,1H),6.43(t,J＝72Hz,1H),4.46(s,3H),3.96-3.90(m,2H),3.62-3.52(m,2H),3.16-3.08(m,1H),2.20(s,6H)；MS(ESI)(m/z):[M+H]+487.2。
实施例27：N-(4-(二氟甲氧基)-2-(3-(二甲氨基)氮杂环丁-1-基)-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物51d)
Figure PCTCN2015088643-appb-000122
除了将化合物50a换成化合物50d之外，与化合物51a同样地制备化合物51d。1H NMR(400MHz,CDCl3):δ8.65(s,1H),8.51(d,J＝5.2Hz,2H),7.42(s,1H),7.36(d,J＝5.2Hz,1H),7.21(s,1H),7.14(d,J＝5.2Hz,1H),6.98(d,J＝5.2Hz,1H),6.62(s,1H),6.51(t,J＝72Hz,1H),6.48-6.26(m,2H),5.84-5.76(m,1H),4.43(s,3H),3.94-3.84(m,2H),3.66-3.58(m,2H),3.16-3.08(m,1H),2.19(s,6H)；MS(ESI)(m/z):[M+H]+541.2。
N-(2-(二氟甲氧基)-4-(4-(二甲氨基)哌啶-1-基)-5-硝基苯基)-4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-胺(化合物49e)
Figure PCTCN2015088643-appb-000123
除了将化合物48a换成化合物48b，N,N,N’-三甲基乙二胺替换成N,N-二甲基哌啶-4-胺之外，与化合物49a同样地制备化合物49e。MS(ESI)(m/z):[M+H]+545.2。
6-(二氟甲氧基)-4-(4-(二甲氨基)哌啶-1-基)-N1-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,3-二胺(化合物50e)
Figure PCTCN2015088643-appb-000124
除了将化合物49a换成化合物49e之外，与化合物50a同样地制备化合物50e。MS(ESI)(m/z):[M+H]+515.2。
实施例28：N-(4-(二氟甲氧基)-2-(4-(二甲氨基)哌啶-1-基)-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物51e)
Figure PCTCN2015088643-appb-000125
除了将化合物50a换成化合物50e之外，与化合物51a同样地制备化合物51e。1H NMR(400MHz,CDCl3):δ9.33(s,1H),8.62-8.54(m,2H),7.36(d,J＝5.2Hz,1H),7.30(s,1H),7.14(d,J＝5.2Hz,1H),7.05-6.98(m,2H),6.50(t,J＝72.0Hz,1H),6.44-6.22(m,2H),5.82-5.76(m,1H),4.47(s,3H),3.10-3.03(m,2H),2.76-2.66(m,2H),2.42(s,6H),2.41-2.39(m,1H),2.14-2.05(m,2H),1.78-1.66(m,2H)；MS(ESI)(m/z):[M+H]+569.2。
N-(2-(二氟甲氧基)-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)-5-硝基苯基)-4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-胺(化合物49f)
Figure PCTCN2015088643-appb-000126
除了将化合物48a换成化合物48b，N,N,N’-三甲基乙二胺替换成5-甲基-2,5-二氮杂螺[3.4]辛烷之外，与化合物49a同样地制备化合物49f。1H NMR(400MHz,CDCl3):δ8.88(s,1H),8.54(d,J＝5.2Hz,1H),7.38(d,J＝5.2Hz,1H),7.24(s,1H),7.17(d,J＝5.2Hz,1H),7.04(d,J＝5.2Hz,1H),6.64(t,J＝72Hz,1H),6.42(s,1H),4.52(s,3H),4.16-4.06(m,2H),3.74-3.64(m,2H),2.80-2.70(m,2H),2.49(s,3H),2.15-2.07(m,2H),1.86-1.76(m,2H)；MS(ESI)(m/z):[M+H]+543.2。
6-(二氟甲氧基)-N1-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯-1,3-二胺(化合物50f)
Figure PCTCN2015088643-appb-000127
除了将化合物49a换成化合物49f之外，与化合物50a同样地制备化合物50f。1H NMR(400MHz,CDCl3):δ8.50(d,J＝5.2Hz,1H),7.64(s,1H), 7.38(d,J＝5.2Hz,1H),7.21(s,1H),7.16(d,J＝5.2Hz,1H),6.97(d,J＝5.2Hz,1H),6.50(s,1H),6.44(t,J＝72Hz,1H),4.46(s,3H),3.94-3.88(m,2H),3.66-3.60(m,2H),3.55(bs,2H),2.80-2.72(m,2H),2.51(s,3H),2.22-2.14(m,2H),1.86-1.76(m,2H)；MS(ESI)(m/z):[M+H]+513.2。
实施例29：N-(4-(二氟甲氧基)-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)氨基)-2-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯基)丙烯酰胺(化合物51f)
Figure PCTCN2015088643-appb-000128
除了将化合物50a换成化合物50f之外，与化合物51a同样地制备化合物51f。1H NMR(400MHz,CDCl3):δ8.57(s,1H),8.50(d,J＝5.2Hz,1H),7.36(d,J＝5.2Hz,1H),7.25(m,1H),7.20(s,1H),7.14(d,J＝5.2Hz,1H),6.98(d,J＝5.2Hz,1H),6.69-6.29(m,4H),5.82-5.76(m,1H),4.43(s,3H),4.02-3.94(m,2H),3.68-3.60(m,2H),2.82-2.72(m,2H),2.52(s,3H),2.22-2.12(m,2H),1.86-1.76(m,2H)；MS(ESI)(m/z):[M+H]+567.2。
化合物63的合成路线如方案6所示：
Figure PCTCN2015088643-appb-000129
方案6
2-甲基-4H-噻吩并[3,2-b]吡咯(化合物53)
Figure PCTCN2015088643-appb-000130
将化合物52(3g，16.6mmol，参考专利申请US20080004327合成)溶于500ml四氢呋喃中，冷却后缓慢加入四氢铝锂(2.5g，66mmol)，然后加热 至70℃搅拌4小时。冷却至室温后，用1M的碳酸氢钠溶液在0℃淬灭，接着过滤旋干，柱层析纯化得到白色固体(1.8g，79％)。MS(ESI)(m/z):[M+H]+138.1。1H NMR(400MHz,CDCl3):δ8.11(br,1H)，6.91-6.89(m,1H)，6.65(s,1H)，6.39(s,1H)，2.54(s,3H)。
6-(2-氯嘧啶-4-基)-2-甲基-4H-噻吩并[3,2-b]吡咯(化合物54)
Figure PCTCN2015088643-appb-000131
将2,4-二氯嘧啶(2.44g，16.4mmol)和三氯化铁(2.66g，16.4mmol)加入200mL 1，2-二氯乙烷中，室温搅拌3小时，再加入化合物53(1.5g，11mmol)，室温搅拌4小时。过滤浓缩后，硅胶柱纯化得到褐色固体(100mg，2.4％)。MS(ESI)(m/z):[M+H]+250.1。1H NMR(400MHz,CDCl3):δ9.50(br,1H),8.38(d,J＝5.6Hz,1H),7.33(d,J＝5.2Hz,1H),7.05(d,J＝1.6Hz,1H),6.67(s,1H),2.57(d,J＝1.2Hz,3H)。
6-(2-氯嘧啶-4-基)-2,4-二甲基-4H-噻吩并[3,2-b]吡咯(化合物55)
Figure PCTCN2015088643-appb-000132
将化合物54(100mg，0.4mmol)，碘甲烷(1.42g，4mmol)和碳酸钾(276mg，2mmol)加入DMF 5mL中，室温搅拌一周后，加入乙酸乙酯稀释，依次用水和饱和食盐水洗涤，干燥后硅胶柱纯化得到黄色固体(50mg，47％)。MS(ESI)(m/z):[M+H]+264.1。1H NMR(400MHz,CDCl3):δ8.36(d,J＝5.6Hz,1H),7.36(d,J＝5.6Hz,1H),7.01(s,1H),6.66(s,1H),4.14(s,3H),2.57(d,J＝0.8Hz,3H)。
4-(2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基)-N-(4-氟-2-甲氧基-5-硝基苯基)嘧啶-2-胺(化合物56)
Figure PCTCN2015088643-appb-000133
将化合物55(50mg，0.19mmol)，2-甲氧基-4-氟-5-硝基苯胺(71mg，0.38mmol)和一水合对甲苯磺酸(108mg，0.57mmol)加入仲戊醇5mL中，100℃搅拌5小时。冷却后过滤，固体用乙酸乙酯洗涤得到黄色固体(65mg，83％)。MS(ESI)(m/z):[M+H]+414.1。1H NMR(400MHz,CDCl3):δ9.35(d,J＝8.4Hz,1H),8.33(d,J＝5.2Hz,1H),7.59(br,1H),7.05(d,J＝5.2Hz,1H),6.96(s,1H),6.75(d,J＝12Hz,1H),6.68(s,1H),4.15(s,3H),4.02(s,3H),2.57(s,3H)。
N1-(4-(2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基)嘧啶-2-基)-N4-(2-(二甲氨基)乙基)-2-甲氧基-N4-甲基-5-硝基苯-1,4-二胺(化合物57)
Figure PCTCN2015088643-appb-000134
将化合物56(65mg，0.16mmol)和N,N,N’-三甲基乙二胺(32mg，0.31mmol)溶于10ml乙腈中，加入碳酸钾(43mg，0.31mmol)，80℃搅拌4小时。然后过滤浓缩，硅胶柱纯化得到红色固体(55mg，70％)。MS(ESI)(m/z):[M+H]+496.3。1H NMR(400MHz,CDCl3):δ9.05(s,1H),8.31(d,J＝5.2Hz,1H),7.48(s,1H),7.00(d,J＝5.6Hz,1H),6.94(s,1H),6.76(s,1H),6.68(s,1H),4.15(s,3H),4.00(s,3H),3.41-3.37(m,2H),2.88(s,3H),2.80-2.77(m,2H),2.57(s,3H),2.46(s,6H)。
N4-(4-(2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基)嘧啶-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺(化合物58)
Figure PCTCN2015088643-appb-000135
将化合物57(55mg，0.11mmol)和钯碳在甲醇中的混合物在氢气氛围下 室温搅拌1h。硅藻土过滤后，滤液浓缩得到黄色固体(50mg，98％)。MS(ESI)(m/z):[M+H]+466.2。1H NMR(400MHz,CDCl3):δ8.28(d,J＝5.2Hz,1H),7.94(s,1H),7.48(s,1H),6.91-6.90(m,2H),6.70(s,1H),6.66(s,1H),4.13(s,3H),3.83(s,3H),2.97-2.94(m,2H),2.67(s,3H),2.57(d,J＝0.8Hz,3H),2.41-2.38(m,2H),2.26(s,6H)。
实施例30：N-(5-((4-(2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基)嘧啶-2-基)氨基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基苯)丙烯酰胺(化合物59)
Figure PCTCN2015088643-appb-000136
将化合物58(50mg，0.1mmol)，三乙胺(33mg，0.32mmol)和丙烯酸(23mg，0.32mmol)溶于二氯甲烷2mL，加入催化量的叔丁醇，冷却至-10℃，接着加入EDCI(41mg，0.22mmol)，室温搅拌2小时。反应液用二氯甲烷稀释，碳酸氢钠水溶液和饱和食盐水洗涤，干燥后硅胶柱和制备板纯化得到黄色固体(7.7mg，13.8％)。MS(ESI)(m/z):[M+H]+520.2。1H NMR(400MHz,CDCl3):δ9.49(s,1H),9.27(s,1H),8.32(d,J＝5.2Hz,1H),7.51(s,1H),7.06-6.93(m,3H),6.68(s,1H),6.63(s,1H),6.42(d,J＝16.8Hz,1H),5.71(d,J＝10.4Hz,1H),4.06(s,3H),3.87(s,3H),3.18(s,2H),2.93(s,2H),2.69(s,3H),2.66(s,6H),2.56(s,3H)。
化合物72的合成路线如方案7所示：
Figure PCTCN2015088643-appb-000137
方案7
(5-甲基噻吩-2-基)氨基甲酸叔丁酯(化合物61)
Figure PCTCN2015088643-appb-000138
将5-甲基-2-噻吩甲酸(10g，70.3mmol)溶于叔丁醇(100mL)中，然后依次加入三乙胺(7.83g，77.4mmol)和叠氮磷酸二苯酯(21.29g，77.4mmol)，室温搅拌30min后油浴升温至80℃回流过夜反应。后处理直接旋干反应液，柱层析得到化合物61(7g，46.7％)。1H NMR(400MHz,d6-DMSO):δ10.13(s,1H),6.45-6.44(m,1H),6.29(d,J＝3.6Hz,1H),2.32(t,J＝4.0Hz,3H),1.48(d,J＝16.5Hz,9H)；MS(ESI)(m/z):[M+H]+214.1。
(3-甲酰基-5-甲基噻吩-2-基)氨基甲酸叔丁酯(化合物62)
Figure PCTCN2015088643-appb-000139
在冰水浴条件下，慢慢将POCl3(15.1g，98.5mmol)滴加进DMF(4.8g，65.7mmol)中，搅拌15-30分钟出现白色固体。另将化合物61(7g，32.8mmol)溶于四氢呋喃(100mL)，加入反应瓶中，升至室温搅拌过夜反应。反应液慢慢加到温水(100mL)中淬灭反应，乙酸乙酯萃取，柱层析得到化合物62(3g，38.0％)。1H NMR(400MHz,CDCl3):δ10.47(s,1H),9.69(s,1H),6.76(s,1H),2.39(s,3H),1.56(s,9H)；MS(ESI)(m/z):[M+H]+242.1。
2-((叔丁氧羰基)(3-甲酰基-5-甲基噻吩-2-基)氨基)乙酸乙酯(化合物63)
Figure PCTCN2015088643-appb-000140
将化合物62(2.9g，12.0mmol)溶于DMF(30mL)中，再加入碳酸钾(2.5g，18.1mmol)和溴乙酸乙酯(2.4g，14.4mmol)，室温搅拌过夜。后处理加水(50mL)，乙酸乙酯萃取，硫酸钠干燥旋干得到粗产品63(4g，100％)，直接用作下一步投料。MS(ESI)(m/z):[M+H]+328.1。
6-叔丁基5-乙基4-羟基-2-甲基-4H-噻吩[2,3-b]吡咯-5,6(5H)-二羧酸酯(化合物64)
Figure PCTCN2015088643-appb-000141
将化合物63(4g，12.2mmol)溶于DMF(40mL)中，加入碳酸钾(2.53g，18.3mmol)，油浴升温60℃反应过夜。后处理加水(50mL)，乙酸乙酯萃取，柱层析得到化合物64(3.8g，86.2％)。
2-甲基-6H-噻吩[2,3-b]吡咯-5-甲酸乙酯(化合物65)
Figure PCTCN2015088643-appb-000142
将化合物64(3.8g，11.6mmol)溶于乙醇(15mL)中，加入3mL浓盐酸，室温搅拌，待反应完全后，直接将反应液旋干，加水(50mL)，用碳酸钾中和，乙酸乙酯(50mL)萃取，硫酸钠干燥，旋干后得到化合物65(1.8g，74％)。MS(ESI)(m/z):[M+H]+210.1。
2,5-二甲基-6H-噻吩并[2,3-b]吡咯(化合物66)
Figure PCTCN2015088643-appb-000143
将化合物65(500mg，2.4mmol)溶于无水THF(5mL)中，冰盐浴冷却，慢慢加入氢化铝锂(226.7mg，6.0mmol)，放热且大量气泡产生，加毕，油浴升温至70℃回流。待反应完全，冰水浴降温，慢慢加水，NaOH(aq)，乙酸乙酯萃取，硅藻土过滤，旋干得到化合物66(250mg，69.1％)。1H NMR(400MHz,CDCl3):δ7.88(s,1H),6.62(t,J＝6.0Hz,1H),6.08-5.94(m,1H),2.52(d,J＝1.2Hz,3H),2.39-2.37(m,3H)；MS(ESI)(m/z):[M+H]+152.0。
4-(2-氯嘧啶-4-基)-2,5-二甲基-6H-噻吩并[2,3-b]吡咯(化合物67)
Figure PCTCN2015088643-appb-000144
将化合物66(200mg，1.3mmol)溶于1,2-二氯甲烷(5mL)中，然后依次加入2,4-二氯嘧啶(295.5mg，2.0mmol)、三氯化铝(264.5mg，2.0mmol)，油浴升温至80℃反应。待反应完全后加水(10mL)淬灭反应，抽滤掉固体， 加二氯甲烷萃取，有机相干燥旋干，柱层析提纯得到化合物67(100mg，26.7％)。1H NMR(400MHz,CDCl3):δ9.73(s,1H),8.29(t,J＝7.7Hz,1H),7.18(d,J＝5.5Hz,1H),6.88(d,J＝1.1Hz,1H),2.59(d,J＝1.0Hz,3H),2.49(s,3H)；MS(ESI)(m/z):[M+H]+264.0。
4-(2-氯嘧啶-4-基)-2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯(化合物68)
Figure PCTCN2015088643-appb-000145
将化合物67(73mg，0.38mmol)溶于DMF(5mL)中，然后依次加入碘甲烷(47.14mg，0.45mmol)、碳酸钾(57.30mg，0.57mmol)室温搅拌。后处理加水(10mL)和乙酸乙酯(10mL)萃取，饱和食盐水洗两遍，干燥浓缩，得到化合物68(80mg)。MS(ESI)(m/z):[M+H]+278.0。
N-(4-氟-2-甲氧基-5-硝基苯基)-4-(2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基)嘧啶-2-胺(化合物69)
Figure PCTCN2015088643-appb-000146
将化合物68(80mg，0.288mmol)溶于仲戊醇(5mL)中，然后依次加入4-氟-2-甲氧基-5-硝基苯胺(53.6mg，0.288mmol)、对甲苯磺酸(82.18mg，0.430mmol)，油浴升温105℃反应。后处理，直接旋干反应液，制备色谱纯化得到化合物69(60mg，48.7％)。1H NMR(400MHz,CDCl3):δ9.48(d,J＝8.4Hz,1H),8.40(d,J＝5.3Hz,1H),7.65(s,1H),7.10(d,J＝5.3Hz,1H),6.96(s,1H),6.80(d,J＝4.2Hz,1H),4.05(s,3H),3.68(s,3H),2.79(s,3H),2.59(s,3H)；MS(ESI)(m/z):[M+H]+428.1。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-2-硝基-N4-(4-(2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基)嘧啶-2-基)苯-1,4-二胺(化合物70)
Figure PCTCN2015088643-appb-000147
将化合物69(60mg，0.14mmol)溶于DMF(5mL)中，然后依次加入N,N,N’-三甲基乙二胺(17.2mg，0.16mmol)、N,N-二异丙基乙胺(23.6mg，0.18mmol)，油浴升温至85℃搅拌。反应完全后加水(10mL)淬灭反应，乙酸乙酯(10mL)萃取，饱和食盐水洗两遍，干燥旋干，制备色谱纯化得到化合物70(50mg，69.9％)。1H NMR(400MHz,CDCl3):δ9.19(s,1H),8.37(d,J＝5.3Hz,1H),7.60(s,1H),7.03(d,J＝5.3Hz,1H),6.95(s,1H),6.87(s,1H),4.05(s,3H),3.65(s,3H),3.52(t,J＝6.6Hz,2H),3.49(s,3H),3.02(t,J＝6.8Hz,2H),2.88(s,3H),2.76(s,3H),2.66(s,6H)；MS(ESI)(m/z):[M+H]+510.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基)嘧啶-2-基)苯-1,2,4-三胺(化合物71)
Figure PCTCN2015088643-appb-000148
将化合物70(50mL，0.098mmol)溶于乙醇(6mL)和水(2mL)中，再依次加入铁粉(32.87mg，0.587mmol)、氯化铵(3.67mg，0.067mmol)，油浴升温80℃反应，2小时反应完全。抽滤反应液，制备色谱纯化得到化合物71(21mg，44.68％)。MS(ESI)(m/z):[M+H]+480.2。
实施例31：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(2,5,6-三甲基-噻吩并[2,3-b]吡咯-4-基)嘧啶-2-基)氨基)苯基)丙烯酰胺(化合物72)
Figure PCTCN2015088643-appb-000149
将化合物71(21mg，0.044mmol)溶于DCM(5mL)和叔丁醇(0.5mL)中，冰水浴冷却后，依次加入EDCI(16.79mg，0.088mmol)、三乙胺(8.84mg， 0.088mmol)、丙烯酸(6.31mg，0.088mmol)搅拌3h反应完全。制备色谱纯化得到化合物72(5mg，21.39％)。1H NMR(400MHz,CDCl3):δ9.64(s,1H),9.33(s,1H),8.33(d,J＝5.4Hz,1H),7.48(s,1H),6.96(d,J＝5.5Hz,1H),6.87(s,1H),6.71(s,1H),6.41(d,J＝16.8Hz,1H),5.71(d,J＝11.7Hz,1H),4.08(s,3H),3.90(s,3H),3.12(s,2H),2.77(s,2H),2.72(s,3H),2.60(s,6H),2.54(s,3H),2.41(s,3H)；MS(ESI)(m/z):[M+H]+534.3。
化合物80a-80f的合成路线如方案8所示：
Figure PCTCN2015088643-appb-000150
方案8
4-氯-N-(4-氟-2-甲氧基-5-硝基苯基)-1,3,5-三嗪-2-胺(化合物74a)
Figure PCTCN2015088643-appb-000151
将化合物73(1.77g，11.80mmol)溶于N,N-二甲基甲酰胺(20mL)中，然后依次加入2-甲氧基-4-氟-5-硝基苯胺(2g，10.74mmol)、N,N-二异丙基乙胺(1.67g，12.92mmol)，室温搅拌，3小时后反应完全，加水(50mL)，析出大量固体，抽滤得到粗品，再用乙酸乙酯洗两遍，得到纯品产物74a(1.8g，56.25％)。MS(ESI)(m/z):[M+H]+300.0。
4-氯-N-(2-(二氟甲氧基)-4-氟-5-硝基苯基)-1,3,5-三嗪-2-胺(化合物74b)
Figure PCTCN2015088643-appb-000152
在250mL的单口瓶中将2-(二氟甲氧基)-4-氟-5-硝基苯胺(30g，0.135mol)溶于DMF(100mL)中，加入碳酸氢钠(17g，0.20mol)然后在冰浴下慢慢加入化合物73(30g，0.20mol)，升到室温反应2h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物74b(50g，0.149mol)。1H NMR(400MHz,d6-DMSO):δ10.64(s,1H),8.64(s,1H),8.49(d,J＝7.7Hz,1H),7.63(d,J＝12.0Hz,1H),7.61-7.22(m,1H)；MS(ESI)(m/z):[M+H]+336.0。
7-(4,4,5,5-四甲基-1,3,2-二氧杂戊硼烷-2-基)-1H-吲哚(化合物76a)
Figure PCTCN2015088643-appb-000153
在1L的单口瓶中将7-溴吲哚(20g，0.102mol)溶于DMSO(100mL)中，加入联硼酸频哪醇酯(39g，0.153mol)，[1,1'-双(二苯基磷)二茂铁]二氯化钯(Pd(dppf)Cl2，4g，0.005mol)和醋酸钾(15g，0.153mol)，升温到85℃反应2h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析产物(25g，0.103mol)。1H NMR(400MHz,CDCl3):δ9.29(s,1H),7.82(d,J＝7.9Hz,1H),7.70(d,J＝7.0Hz,1H),7.32-7.29(m,1H),7.21-7.14(m,1H),6.62-6.57(m,1H),1.44(s,12H).MS(ESI)(m/z):[M+H]+244.1。
1-甲基-7-(4,4,5,5-四甲基-1,3,2-二氧杂戊硼烷-2-基)-1H-吲哚(化合物76b)
Figure PCTCN2015088643-appb-000154
在100mL的单口瓶中将7-溴-1-甲基-1H-吲哚(2g，0.01mol)溶于DMSO(50mL)中，加入联硼酸频哪醇酯(5g，0.02mol)，Pd(dppf)Cl2(0.8g，0.001mol)和醋酸钾(2.4g，0.024mol)。升温到85℃反应2h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物76b(2.5g，0.012mol)。1H NMR(400MHz,CDCl3):δ7.79-7.72(m,1H),7.70(d,J＝6.9Hz, 1H),7.13(t,J＝7.5Hz,1H),7.04(d,J＝3.1Hz,1H),6.52(d,J＝3.1Hz,1H),4.00(s,3H),1.43(s,12H)；MS(ESI)(m/z):[M+H]+258.1。
N-(4-氟-2-甲氧基-5-硝基苯基)-4-(1H-吲哚-7-基)-1,3,5-三嗪-2-胺(化合物77a)
Figure PCTCN2015088643-appb-000155
在100mL的单口瓶中将化合物76a(609mg，2.5mmol)溶于1,4-二氧六环(50mL)中，加入化合物74a(500mg，1.67mmol)，四(三苯基膦)钯(193mg，0.167mmol)，碳酸钠(442mg，4.17mmol)和水(15mL)升到80℃反应2h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物77a(1g，2.6mmol)。MS(ESI)(m/z):[M+H]+381.1。
N-(2-(二氟甲氧基)-4-氟-5-硝基苯基)-4-(1H-吲哚-7-基)-1,3,5-三嗪-2-胺(化合物77b)
Figure PCTCN2015088643-appb-000156
除了将化合物74a换成化合物74b之外，与化合物77a同样地制备化合物77b。MS(ESI)(m/z):[M+H]+417.1。
N-(4-氟-2-甲氧基-5-硝基苯基)-4-(1-甲基-1H-吲哚-7-基)-1,3,5-三嗪-2-胺(化合物77c)
Figure PCTCN2015088643-appb-000157
除了将化合物76a换成化合物76b之外，与化合物77a同样地制备化合物77c。MS(ESI)(m/z):[M+H]+395.1。
N1-(4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)-N4-(2-(二甲氨基)乙基)-2-甲氧基-N4-甲基-5-硝基苯-1,4-二胺(化合物78a)
Figure PCTCN2015088643-appb-000158
在100mL的单口瓶中将化合物77a(1g,2.63mmol)溶于N,N-二甲基甲酰胺(30mL)中，加入N,N,N’-三甲基乙二胺(458mg,3.95mmol)，N,N-二异丙基乙胺(1.01g，7.89mmol)。升到60℃反应2小时。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得产物78a(400mg,0.866mmol)。1H NMR(400MHz,CDCl3):δ10.91(s,1H),9.31(s,1H),8.84(s,1H),8.57(s,1H),7.91(d,J＝7.7Hz,1H),7.69(s,1H),7.40-7.30(m,2H),6.70(s,1H),6.66-6.62(m,1H),3.99(s,3H),3.35(t,J＝6.9Hz,2H),2.94(s,3H),2.62(t,J＝7.0Hz,2H),2.31(s,6H)；MS(ESI)(m/z):[M+H]+463.2。
N1-(4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)-2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-5-硝基苯-1,4-二胺(化合物78b)
Figure PCTCN2015088643-appb-000159
除了将化合物77a换成化合物77b之外，与化合物78a同样地制备化合物78b。MS(ESI)(m/z):[M+H]+499.2。
4-(1H-吲哚-7-基)-N-(2-甲氧基-4-(4-甲基哌嗪-1-基)-5-硝基苯基)-1,3,5-三嗪-2-胺(化合物78c)
Figure PCTCN2015088643-appb-000160
除了将N,N,N’-三甲基乙二胺换成4-甲基哌嗪之外，与化合物78a同样地制备化合物78c。1H NMR(400MHz,d6-DMSO):δ11.90(s,1H),9.90(s,1H),8.85(s,1H),8.74(s,1H),8.35-8.33(m,1H),7.88(d,J＝7.6Hz,1H),7.58(s,1H),7.20(t,J＝7.7Hz,1H),6.90(s,1H),6.64(s,1H),4.07(s,3H),3.27-3.23(m,4H),2.98-2.82(m,4H),2.74(s,3H)；MS(ESI)(m/z):[M+H]+461.2。
N-(4-(3-(二甲氨基)氮杂环丁-1-基)-2-甲氧基-5-硝基苯基)-4-(1H-吲哚-7-基)-1,3,5-三嗪-2-胺(化合物78d)
Figure PCTCN2015088643-appb-000161
除了将N,N,N’-三甲基乙二胺换成N,N-二甲基氮杂环丁-3-胺之外，与化合物78a同样地制备化合物78d。1H NMR(400MHz,d6-DMSO):δ11.86(s,1H),9.74(s,1H),8.81(s,1H),8.53(s,1H),8.35(s,1H),7.88(d,J＝6.9Hz,1H),7.57(s,1H),7.20(t,J＝7.3Hz,1H),6.64(s,1H),6.32(s,1H),4.08-3.89(m,5H),3.82-3.78(m,2H),3.20-3.16(m,1H),2.17(s,6H)；MS(ESI)(m/z):[M+H]+461.2。
4-(1H-吲哚-7基)-N-(2-甲氧基-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)-5-硝基苯基)-1,3,5-三嗪-2胺(化合物78e)
Figure PCTCN2015088643-appb-000162
除了将N,N,N’-三甲基乙二胺换成N,N-二甲基氮杂环丁-3-胺之外，与化合物78a同样地制备化合物78e。MS(ESI)(m/z):[M+H]+487.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吲哚-7-基)-1,3,5-三嗪-2-基)-2-硝基苯-1,4-二胺(化合物78f)
Figure PCTCN2015088643-appb-000163
除了将化合物77a换成化合物77c之外，与化合物78a同样地制备化合物78f。MS(ESI)(m/z):[M+H]+477.2。
N4-(4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺(化合物79a)
Figure PCTCN2015088643-appb-000164
在100mL的单口瓶中将化合物78a(200mg,0.433mmol)溶于乙醇(20mL)-水(20mL)中，加入铁粉(606mg,10.82mmol)，氯化铵(139mg，2.6mmol)。升到50℃反应2小时。加饱和碳酸钠水溶液淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得产物79a(150mg,0.35mmol)。MS(ESI)(m/z):[M+H]+433.2。
N4-(4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)-5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲苯-1,2,4-三胺(化合物79b)
Figure PCTCN2015088643-appb-000165
除了将化合物78a换成化合物78b之外，与化合物79a同样地制备化合物79b。1H NMR(400MHz,CDCl3):δ10.86(s,1H),8.83(s,1H),8.49-8.42(m,1H),7.90(d,J＝7.7Hz,1H),7.66(s,1H),7.60(s,1H),7.34(s,1H),7.28(m,1H),6.95(s,1H),6.68-6.27(m,2H),2.99(t,J＝6.3Hz,2H),2.74(s,3H),2.49(t,J＝6.3Hz,2H),2.33(s,6H)；MS(ESI)(m/z):[M+H]+469.2。
N1-(4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)-6-甲氧基-4-(4-甲基哌嗪-1-基)苯-1,3-二胺(化合物79c)
Figure PCTCN2015088643-appb-000166
除了将化合物78a换成化合物78c之外，与化合物79a同样地制备化合物79c。MS(ESI)(m/z):[M+H]+431.2。
N1-(4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)-4-(3-(二甲氨基)氮杂环丁-1-基)-6-甲氧基苯-1,3-二胺(化合物79d)
Figure PCTCN2015088643-appb-000167
除了将化合物78a换成化合物78d之外，与化合物79a同样地制备化合物79d。MS(ESI)(m/z):[M+H]+431.2。
N1-(4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)-6-甲氧基-4-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯-1,3-二胺(化合物79e)
Figure PCTCN2015088643-appb-000168
除了将化合物78a换成化合物78e之外，与化合物79a同样地制备化合物79e。MS(ESI)(m/z):[M+H]+457.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吲哚-7-基)-1,3,5-三嗪-2-基)苯-1,2,4-三胺(化合物79f)
Figure PCTCN2015088643-appb-000169
除了将化合物78a换成化合物78f之外，与化合物79a同样地制备化合物79f。MS(ESI)(m/z):[M+H]+447.3。
实施例32：N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基苯)丙烯酰胺(化合物80a)
Figure PCTCN2015088643-appb-000170
在100mL的单口瓶中将化合物79a(200mg,0.463mmol)溶于二氯甲烷(20mL)中，加入丙烯酸(33.3mg,0.463mmol)，EDCI(222mg，1.15mmol)。室温反应2小时。加水淬灭反应，二氯甲烷萃取3遍，硫酸钠干燥，浓缩后柱层析得产物84a(95mg,0.195mmol)。1H NMR(400MHz,CDCl3):δ10.94 (s,1H),10.20(s,1H),9.68(s,1H),8.89(s,1H),7.89(d,J＝7.7Hz,1H),7.79(s,1H),7.32(d,J＝7.2Hz,1H),6.86(s,1H),6.66-6.49(m,2H),6.41-6.32(m,1H),5.79-5.72(m,1H),3.92(s,3H),2.99-2.88(m,2H),2.77(s,3H),2.40-2.34(m,2H),2.32(s,6H)；MS(ESI)(m/z):[M+H]+487.2。
实施例33：N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺(化合物80b)
Figure PCTCN2015088643-appb-000171
除了将化合物79a换成化合物79b之外，与化合物80a同样地制备化合物80b。1H NMR(400MHz,CDCl3):δ10.90(s,1H),10.36(s,1H),9.69(s,1H),8.88(s,1H),8.72(s,1H),7.90(d,J＝7.7Hz,1H),7.57(s,1H),7.31(d,J＝7.8Hz,2H),7.12(s,1H),6.73-6.34(m,4H),5.80(d,J＝10.2Hz,1H),2.96-2.86(m,2H),2.76(s,3H),2.45-2.35(m,2H),2.33(s,6H)；MS(ESI)(m/z):[M+H]+523.2。
实施例34：N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-4-甲氧基-2-(4-甲基哌嗪-1-基)苯基)丙烯酰胺(化合物80c)
Figure PCTCN2015088643-appb-000172
除了将化合物79a换成化合物79c之外，与化合物80a同样地制备化合物80c。1H NMR(400MHz,CDCl3):δ10.94(s,1H),9.59(s,1H),8.86-8.48(m,3H),7.95-7.76(m,2H),7.41-7.25(m,2H),6.82(s,1H),6.62(s,1H),6.51-6.47(m,1H),6.37-6.30(m,1H),5.81(d,J＝10.3Hz,1H),3.90(s,3H),3.06-3.00(m,4H),2.88-2.80(m,4H),2.51(s,3H)；MS(ESI)(m/z):[M+H]+485.2。
实施例35：N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-2-(3-(二甲氨基)氮杂环丁-1-基)-4-甲氧基苯基)丙烯酰胺(化合物80d)
Figure PCTCN2015088643-appb-000173
除了将化合物79a换成化合物79d之外，与化合物80a同样地制备化合物80d。1H NMR(400MHz,CDCl3):δ10.92(s,1H),8.79(s,1H),8.59(s,1H),7.87(d,J＝7.3Hz,1H),7.68(s,1H),7.42-7.21(m,4H),6.61(s,1H),6.52-6.48(m,1H),6.38-6.31(m,2H),5.81(d,J＝10.2Hz,1H),4.13-3.81(m,5H),3.71-3.67(m,2H),3.16-3.12(m,1H),2.22(s,6H)；MS(ESI)(m/z):[M+H]+485.2。
实施例36：N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-4-甲氧基-2-(5-甲基-2,5-二氮杂螺[3.4]辛-2-基)苯基)丙烯酰胺(化合物80e)
Figure PCTCN2015088643-appb-000174
除了将化合物79a换成化合物79e之外，与化合物80a同样地制备化合物80e。1H NMR(400MHz,CDCl3):δ11.05-10.71(m,1H),8.72(s,1H),8.58-8.00(m,2H),7.85-7.56(m,3H),7.31(s,1H),7.21(t,J＝7.6Hz,1H),6.65-6.29(m,3H),6.05(s,1H),5.77(d,J＝10.7Hz,1H),4.02-4.00(m,2H),3.79(s,3H),3.65-3.63(m,2H),2.88-2.73(m,2H),2.53(s,3H),2.12-2.10(m,2H),1.81-1.77(m,2H)；MS(ESI)(m/z):[M+H]+511.3。
实施例37：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)苯基)丙烯酰胺(化合物80f)
Figure PCTCN2015088643-appb-000175
除了将化合物79a换成化合物79f之外，与化合物80a同样地制备化合物80f。1H NMR(400MHz,CDCl3):δ10.14(s,1H),9.65(s,1H),8.98(s,1H),7.87(s,1H),7.79(d,J＝7.2Hz,1H),7.24(t,J＝7.7Hz,1H),7.09(d,J＝3.1Hz, 1H),6.83(s,1H),6.60(d,J＝3.1Hz,1H),6.49(d,J＝15.7Hz,1H),6.39-6.28(m,1H),5.73(d,J＝10.6Hz,1H),3.90(s,3H),3.76(s,3H),2.99-2.86(m,2H),2.75(s,3H),2.33(m,8H)。
化合物85的合成路线如方案9所示：
Figure PCTCN2015088643-appb-000176
方案9
N-(2-(二氟甲氧基)-4-氟-5-硝基苯基)-4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-胺(化合物82)
Figure PCTCN2015088643-appb-000177
将化合物81(15g，0.058mol)溶于1,4-二氧六环(150mL)和水(50mL)中，然后加入化合物74b(13g，0.038mol)、碳酸钠(12.3g，0.116mol)，氮气置换3次后，加入四(三苯基膦)钯(9g，0.008mol)再用氮气置换3次，油浴升温90℃反应，1h后反应完全，硅藻土抽滤，柱层析纯化，得到化合物82(7g，41.90％)。1H NMR(400MHz,CDCl3):δ9.80(s,1H),8.85-8.83(m,1H),8.80(s,1H),8.47-8.45(m,2H),7.48-7.45(m,1H),7.33-7.30(m,1H),7.22(d,J＝10.7Hz,1H),6.97-6.59(m,1H),4.05(s,3H)；MS(ESI)(m/z):[M+H]+432.1。
2-(二氟甲氧基)-N4-(2-(二甲氨基)乙基)-N4-甲基-N1-(4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)-5-硝基苯-1,4-二胺(化合物83)
Figure PCTCN2015088643-appb-000178
将化合物82(7g，0.016mol)溶于DMF(100mL)中，将N,N-二异丙基乙 胺(2.73g，0.021mL),N,N,N’-三甲基乙二胺(1.99g，0.019mL)加入反应中，油浴升温至80℃搅拌，半小时反应完全，加水(100mL)淬灭反应，用二氯甲烷萃取，饱和食盐水洗两遍，柱层析纯化，得到化合物83(3.0g，36.14％)。1H NMR(400MHz,CDCl3):δ9.23(s,1H),8.82-8.80(m,1H),8.72(s,1H),8.44-8.43(m,1H),8.41(s,1H),7.43(s,1H),7.31-7.28(m,1H),7.05(s,1H),6.91-6.48(m,1H),4.03(s,3H),3.33(t,J＝7.0Hz,2H),2.93(d,J＝10.1Hz,3H),2.59(t,J＝7.0Hz,2H),2.29(s,6H)；MS(ESI)(m/z):[M+H]+514.2。
5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)苯-1,2,4-三胺(化合物84)
Figure PCTCN2015088643-appb-000179
将化合物83(3.2g，6.23mmol)溶于乙醇(30mL)中,加水(10mL)，将铁粉(2.09g，37.42mmol)，氯化铵(0.23g，4.30mmol)加入反应中油浴升温80℃搅拌，3h后反应完全，直接抽滤反应液，滤液旋干，柱层析纯化，得到化合物84(3.1g，99.7％)。MS(ESI)(m/z):[M+H]+484.2。
实施例38：N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)氨基)苯基)丙烯酰胺(化合物85)
Figure PCTCN2015088643-appb-000180
将化合物84(3.1g，6.41mmol)溶于二氯甲烷(50mL)中，加入叔丁醇(5mL)，冰水浴降温，然后加入丙烯酸(0.92g，12.77mmol)，EDCI(2.46g，12.83mmol)，三乙胺(1.30g，12.87mmol)，3h后反应完全，加入饱和碳酸钾溶液搅拌10min，二氯甲烷萃取，柱层析纯化，得到化合物85(1.1g，31.9％)。1H NMR(400MHz,CDCl3):δ10.41(s,1H),10.03(s,1H),9.39(bs,1H),8.87-8.85(m,1H),8.76(s,1H),8.43-8.41(m,1H),7.56(s,1H),7.26-7.24 (m,1H),7.11(s,1H),6.76-6.33(m,3H),5.83-5.81(m,1H),4.10(s,3H),2.93-2.85(m,2H),2.75(s,3H),2.36-2.34(m,2H),2.32(s,6H)；MS(ESI)(m/z):[M+H]+538.2。
化合物89的合成路线如方案10所示：
Figure PCTCN2015088643-appb-000181
方案10
4-(1H-苯并[d]咪唑-1-基)-N-(4-氟-2-甲氧基-5-硝基苯基)-1,3,5-三嗪-2-胺(化合物86)的合成
Figure PCTCN2015088643-appb-000182
在100mL的单口瓶中将化合物74a(500mg，1.67mmol)溶于DMF(30mL)中，加入苯并咪唑(179mg,1.52mmol)，碳酸钾(419mg，3.04mmol)。室温反应16h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物86(300mg,0.785mmol)。1H NMR(400MHz,d6-DMSO):δ10.17(s,1H),9.14-8.95(m,1H),8.81(s,1H),8.61-8.52(m,1H),7.79(d,J＝7.3Hz,1H),7.57-7.18(m,3H),3.99(s,3H)；MS(ESI)(m/z):[M+H]+382.1。
N1-(4-(1H-苯并[d]咪唑-1-基)-1,3,5-三嗪-2-基)-N4-(2-(二甲氨基)乙基)-2-甲氧基-N4-甲基-5-硝基苯-1,4-二胺(化合物87)
Figure PCTCN2015088643-appb-000183
在100mL的单口瓶中将化合物86(650mg，1.7mmol)溶于N,N-二甲基乙酰胺(50mL)中，加入N,N,N'-三甲基乙二胺(297mg，2.6mmol)，N,N-二异丙基乙胺(658mg，5.1mmol)。升到60℃反应2h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物87(300mg，0.65mmol)。1H  NMR(400MHz,CDCl3)δ9.12-8.63(m,3H),8.59-8.46(m,1H),7.84(d,J＝7.8Hz,1H),7.81-7.69(m,1H),7.47-7.38(m,2H),6.71(s,1H),3.99(s,3H),3.36(s,2H),2.94(s,3H),2.62(t,J＝6.7Hz,2H),2.30(s,6H)；MS(ESI)(m/z):[M+H]+464.2。
N4-(4-(1H-苯并[d]咪唑-1-基)-1,3,5-三嗪-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺(化合物88)
Figure PCTCN2015088643-appb-000184
在100mL的单口瓶中将化合物87(500mg，1.08mmol)溶于乙醇(20mL)-水(2mL)中，加入铁粉(604mg，10.08mmol)，氯化铵(604mg，10.08mmol)。升到50℃反应2h。加饱和碳酸钠水溶液淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物88(250mg，0.577mmol)。MS(ESI)(m/z):[M+H]+434.2。
实施例39：N-(5-((4-(1H-苯并[d]咪唑-1-基)-1,3,5-三嗪-2-基)氨基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基苯基)丙烯酰胺(化合物89)
Figure PCTCN2015088643-appb-000185
除了将化合物84换成化合物88之外，与化合物85同样地制备化合物89。1H NMR(400MHz,CDCl3):δ10.23(s,1H),9.74(s,2H),9.61(s,1H),8.73(s,1H),8.61(s,1H),7.98(s,1H),7.88-7.86(m,1H),7.38(m,2H),6.70-6.36(m,2H),5.76(d,J＝10.7Hz,1H),3.93(s,3H),2.93(s,2H),2.77(s,3H),2.33(s,8H)；MS(ESI)(m/z):[M+H]+488.2。
化合物97a-97e的合成路线如方案11所示：
Figure PCTCN2015088643-appb-000186
方案11
4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)吡啶-2-胺(化合物94a)
Figure PCTCN2015088643-appb-000187
在100mL的单口瓶中将化合物92(1.3g，6.25mmol)溶于1,4-二氧六环(50mL)中，加入3-溴-1-甲基-1H-吡咯并[2,3-b]吡啶(2g，6.25mmol)，四(三苯基膦)钯(360mg，0.31mmol)，碳酸钠(1.3g，12.5mmol)和水(15ml)升到80℃反应2h。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，旋干得粗产物93a。然后在粗产物93a中加入盐酸水溶液(50mL)加热到50℃反应2h，先用乙酸乙酯萃取2遍倒掉有机层，然后再用NaOH调水相pH到9，再用乙酸乙酯萃取3遍，干燥，浓缩，柱层析得化合物94a(300mg,1.34mmol)。MS(ESI)(m/z):[M+H]+225.1。
4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)吡啶-2-胺(化合物94b)
Figure PCTCN2015088643-appb-000188
在100mL的单口瓶中将化合物46(1.5g,3.5mmol)溶于N,N-二甲基甲酰胺(50mL)中，然后加入化合物90(223mg,1.288mmol),碘化亚铜(44mg,0.24mmol)和四三苯基膦钯(135mg,0.117mmol)。在80℃氮气保护下反应1小时。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后产物94b(300mg,1.3mmol)。MS(ESI)(m/z):[M+H]+231.1。
4-(1-甲基-1H-吡唑并[4,3-c]吡啶-3-基)吡啶-2-胺(化合物94c)
Figure PCTCN2015088643-appb-000189
在100mL的单口瓶中将92(1.3g,4.06mmol)溶于1,4-二氧六环(50mL)中，加入3-碘-1-甲基-1H-吡唑[4,3-b]吡啶(400mg,1.54mmol)，四三苯基膦钯(178mg，0.308mmol),碳酸钠(408mg,3.85mmol)和水(15ml)升到80℃反应2小时。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩得粗产物93c,然后在粗产物中加入盐酸水溶液(50ml)加热到50℃反应2h,先用乙酸乙酯萃取2遍倒掉有机层，后再用NaOH调水相pH到9，再用乙酸乙酯萃取3遍，干燥，浓缩，柱层析得产物94c(350mg,1.56mmol)。MS(ESI)(m/z):[M+H]+226.1。
4-(吡唑并[1,5-a]嘧啶-3-基)吡啶-2-胺(化合物94d)
Figure PCTCN2015088643-appb-000190
除了将3-溴-1-甲基-1H-吡咯并[2,3-b]吡啶换成3-溴吡唑并[1,5-a]嘧啶之外，与化合物94a同样地制备化合物94d。1H NMR(400MHz,d6-DMSO):δ9.23-9.19(m,1H),8.75(s,1H),8.73-8.71(m,1H),7.92(d,J＝5.2Hz,1H),7.31(s,1H),7.22-7.14(m,2H),5.90(bs,2H)；MS(ESI)(m/z):[M+H]+212.1。
4-(咪唑并[1,2-a]吡啶-3-基)吡啶-2-胺(化合物94e)
Figure PCTCN2015088643-appb-000191
除了将3-溴-1-甲基-1H-吡咯并[2,3-b]吡啶换成3-溴咪唑并[1,2-a]吡啶之外，与化合物94a同样地制备化合物94e。MS(ESI)(m/z):[M+H]+211.1。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)吡啶-2-基)-2-硝基苯-1,4-二胺(化合物95a)
Figure PCTCN2015088643-appb-000192
在100mL的单口瓶中将化合物94a(400mg，1.79mmol)溶于DMF(50mL)中，加入化合物11a(886mg，2.7mmol)，三(二亚苄基丙酮)二钯(16mg，0.0179mmol)，4,5-双(二苯基膦)-9,9-二甲基氧杂蒽(21mg，0.0358mmol)和碳酸铯(1.75g，5.37mmol)升到85℃反应12小时。加水淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得化合物95a(300mg，0.63mmol)。MS(ESI)(m/z):[M+H]+476.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)吡啶-2-基)-2-硝基苯-1,4-二胺(化合物95b)
Figure PCTCN2015088643-appb-000193
除了将化合物94a换成化合物94b之外，与化合物95a同样地制备化合物95b。MS(ESI)(m/z):[M+H]+482.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡唑并[4,3-c]吡啶-3-基)吡啶-2-基)-2-硝基苯-1,4-二胺(化合物95c)
Figure PCTCN2015088643-appb-000194
除了将化合物94a换成化合物94c之外，与化合物95a同样地制备化合物95c。MS(ESI)(m/z):[M+H]+477.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-2-硝基-N4-(4-(吡唑并[1,5-a]嘧啶-3-基)吡啶-2-基)苯-1,4-二胺(化合物95d)
Figure PCTCN2015088643-appb-000195
除了将化合物94a换成化合物94d之外，与化合物95a同样地制备化合物95d。MS(ESI)(m/z):[M+H]+463.1。
N1-(2-(二甲氨基)乙基)-N4-(4-(咪唑并[1,2-a]吡啶-3-基)吡啶-2-基)-5-甲氧基-N1-甲基-2-硝基苯-1,4-二胺(化合物95e)
Figure PCTCN2015088643-appb-000196
除了将化合物94a换成化合物94e之外，与化合物95a同样地制备化合物95e。1H NMR(400MHz,CDCl3):δ8.82(s,1H),8.53(d,J＝7.0Hz,1H),8.43-8.35(m,1H),7.85(s,1H),7.73(d,J＝9.1Hz,1H),7.35-7.26(m,1H),7.05-6.95(m,3H),6.91(s,1H),6.71(s,1H),3.99(s,3H),3.35-3.26(m,2H),2.90(s,3H),2.67-2.56(m,2H),2.30(s,6H)；MS(ESI)(m/z):[M+H]+462.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)吡啶-2-基)苯-1,2,4-三胺(化合物96a)
Figure PCTCN2015088643-appb-000197
在100mL的单口瓶中将95a(475mg,2.37mmol)溶于乙醇(20mL)-水(20mL)中，加入铁粉(1.3g,23.75mmol)，氯化铵(1.3g，23.75mmol)。升到50℃反应2小时。加饱和碳酸钠水溶液淬灭反应，乙酸乙酯萃取3遍，硫酸钠干燥，浓缩后柱层析得产物96a(100mg,0.224mmol)。MS(ESI)(m/z):[M+H]+446.3。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)吡啶-2-基)苯-1,2,4-三胺(化合物96b)
Figure PCTCN2015088643-appb-000198
除了将化合物95a换成化合物95b之外，与化合物96a同样地制备化合 物96b。MS(ESI)(m/z):[M+H]+452.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡唑并[4,3-c]吡啶-3-基)吡啶-2-基)苯-1,2,4-三胺(化合物96c)
Figure PCTCN2015088643-appb-000199
除了将化合物95a换成化合物95c之外，与化合物96a同样地制备化合物96c。MS(ESI)(m/z):[M+H]+447.3。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(吡唑并[1,5-a]嘧啶-3-基)吡啶-2-基)苯-1,2,4-三胺(化合物96d)
Figure PCTCN2015088643-appb-000200
除了将化合物95a换成化合物95d之外，与化合物96a同样地制备化合物96d。1H NMR(400MHz,d6-DMSO):δ9.25-9.21(m,1H),8.78(s,1H),8.75-8.71(m,1H),8.13-8.10(d,J＝5.2Hz,1H),7.69(s,1H),7.63(s,1H),7.49(s,1H),7.48-7.43(m,1H),7.21-7.17(m,1H),6.74(s,1H),3.75(s,3H),3.18(s,3H),2.90(t,J＝7.2Hz,2H),2.42(t,J＝7.2Hz,2H),2.23(s,6H)；MS(ESI)(m/z):[M+H]+433.2。
N1-(2-(二甲氨基)乙基)-N4-(4-(咪唑并[1,2-a]吡啶-3-基)吡啶-2-基)-5-甲氧基-N1-甲苯-1,2,4-三胺(化合物96e)
Figure PCTCN2015088643-appb-000201
除了将化合物95a换成化合物95e之外，与化合物96a同样地制备化合物96e。MS(ESI)(m/z):[M+H]+432.2。
实施例40：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)吡啶-2-基)氨基)苯基)丙烯酰胺(化合物97a)
Figure PCTCN2015088643-appb-000202
在100mL的单口瓶中将96a(445mg,0.45mmol)溶于二氯甲烷(50mL)中，加入丙烯酸(72mg，0.67mmol)，EDCI(173mg，0.9mmol)，室温反应2小时，加水淬灭反应，DCM萃取3遍，硫酸钠干燥，浓缩后柱层析得产物97a(30mg,0.06mmol)。1H NMR(400MHz,CD3OD):δ8.52(s,1H),8.45(dd,J＝8.0,1.3Hz,1H),8.38-8.32(m,1H),8.12(d,J＝5.4Hz,1H),8.06(s,1H),7.41(s,1H),7.24(dd,J＝8.0,4.8Hz,1H),7.16(dd,J＝5.5,1.4Hz,1H),7.00(s,1H),6.62-6.55(m,1H),6.48-6.41(m,1H),5.86-5.83(m,1H),3.96(d,J＝3.8Hz,6H),3.20(s,2H),2.73(s,3H),2.50(s,5H)；MS(ESI)(m/z):[M+H]+500.3。
实施例41：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)吡啶-2-基)氨基)苯基)丙烯酰胺(化合物97b)
Figure PCTCN2015088643-appb-000203
除了将化合物96a换成化合物96b之外，与化合物97a同样地制备化合物97b。1H NMR(400MHz,CD3OD)δ8.58(s,1H),8.18-8.11(m,1H),7.61(d,J＝5.3Hz,1H),7.21(d,J＝1.7Hz,2H),7.15(d,J＝5.3Hz,1H),7.00(s,1H),6.63-6.48(m,1H),6.40-6.29(m,1H),5.80(d,J＝10.2Hz,1H),4.09(s,3H),3.92(s,4H),3.13(t,J＝5.5Hz,3H),2.72(d,J＝5.9Hz,3H),2.59(s,2H),2.39(s,6H)；MS(ESI)(m/z):[M+H]+506.2。
实施例42：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-吡唑并[4,3-b]吡啶-3-基)吡啶-2-基)氨基)苯基)丙烯酰胺(化合物97c)
Figure PCTCN2015088643-appb-000204
除了将化合物96a换成化合物96c之外，与化合物97a同样地制备化合物97c。1H NMR(400MHz,CD3OD)δ8.72(d,J＝11.8Hz,1H),8.63(m,1H),8.20(d,J＝5.4Hz,1H),8.12-8.06(m,2H),7.75(m,1H),7.46(m,1H),6.99(s,1H),6.55-6.50(m,1H),6.35(m,1H),5.79(d,J＝10.2Hz,1H),4.18(s,3H),3.93(d,J＝4.8Hz,3H),3.11(d,J＝5.5Hz,2H),2.72(s,3H),2.36(m,6H)；MS(ESI)(m/z):[M+H]+501.3。
实施例43：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(吡唑并[1,5-a]嘧啶-3-基)吡啶-2-基)氨基)苯基)丙烯酰胺(化合物97d)
Figure PCTCN2015088643-appb-000205
除了将化合物96a换成化合物96d之外，与化合物97a同样地制备化合物97d。1H NMR(400MHz,CDCl3):δ10.07(bs,1H),9.06(s,1H),8.87(bs,1H),8.74-8.70(m,1H),8.63-8.59(m,1H),8.28(d,J＝5.2Hz,1H),7.82(s,1H),7.64-7.60(m,1H),7.14(s,1H),6.93-6.87(m,1H),6.78(s,1H),6.52-6.38(m,2H),5.73-5.67(m,1H),3.87(s,3H),2.94(t,J＝7.2Hz,2H),2.71(s,3H),2.41(t,J＝7.2Hz,2H),2.34(s,6H)；MS(ESI)(m/z):[M+H]+487.2。
实施例44：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(咪唑并[1,2-a]吡啶-3-基)吡啶-2-基)氨基)-4-甲氧基苯)丙烯酰胺(化合物97e)
Figure PCTCN2015088643-appb-000206
除了将化合物96a换成化合物96e之外，与化合物97a同样地制备化合物97e。1H NMR(400MHz,CDCl3):δ9.85(s,1H),9.02(s,1H),8.77(d,J＝6.9Hz,1H),8.32(d,J＝5.2Hz,1H),7.94(s,1H),7.73(d,J＝7.3Hz,1H),7.39-7.21(m,2H),7.10-6.84(m,4H),6.76(s,1H),6.50-6.45(m,1H),5.76-5.74(m,1H),3.86(s,3H),3.12-3.06(m,2H),2.75-2.72(m,5H),2.55(s,6H)；MS(ESI)(m/z):[M+H]+486.3。
化合物103的合成路线如方案12所示：
Figure PCTCN2015088643-appb-000207
方案12
1-(2-氯吡啶-4-基)-3-甲基-1H-吲唑(化合物99)
Figure PCTCN2015088643-appb-000208
在氮气保护下，在100mL反应瓶中加入化合物98(1.1g，8.3mmol)，2-氯-4-碘吡啶(4.0g，16.7mmol)，碘化亚铜(158mg，0.83mmol)，磷酸钾(3.5g，16.7mmol)，反式-N,N’-二甲基环己二胺(237mg，1.67mmol)，甲苯(30mL)，升温至80℃反应过夜。反应完毕，过滤，滤液浓缩后，柱层析得到化合物99(1.2g)。1H NMR(400MHz,CDCl3):δ8.47(d,J＝5.6Hz,1H),7.90-7.85(m,2H),7.79-7.70(m,2H),7.62-7.53(m,1H),7.35(t,J＝7.5Hz,1H),2.67(s,3H)；MS(ESI)(m/z):[M+H]+244.1。
N-(4-氟-2-甲氧基-5-硝基苯基)-4-(3-甲基-1H-吲唑-1-基)吡啶-2-胺(化合物100)
Figure PCTCN2015088643-appb-000209
在氮气保护下，在100mL反应瓶中加入化合物99(1.2g，4.9mmol)，4-氟-2-甲氧基-5-硝基苯胺(1.2g，6.3mmol)，醋酸钯(110mg，0.49mmol)，4,5-双(二苯基膦)-9,9-二甲基氧杂蒽(566mg，0.98mmol)，叔丁醇钾(1.1g，9.8mmol)，甲苯(20mL)，加热至100℃反应过夜。反应完毕，过滤，滤液浓 缩后，柱层析得到化合物100(500mg)。1H NMR(400MHz,CDCl3):δ9.37(d,J＝8.2Hz,1H),8.42(d,J＝5.2Hz,1H),7.93(d,J＝8.5Hz,1H),7.77(d,J＝8.0Hz,1H),7.58-7.54(m,1H),7.37-7.31(m,2H),7.26(s,1H),7.13(s,1H),6.77(d,J＝12.1Hz,1H),4.05(s,3H),2.68(s,3H)；MS(ESI)(m/z):[M+H]+394.1。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(3-甲基-1H-吲唑-1-基)吡啶-2-基)-2-硝基苯-1,4-二胺(化合物101)
Figure PCTCN2015088643-appb-000210
室温下向化合物100(500mg，1.4mmol)的N,N-二甲基乙酰胺(15mL)的溶液中依次加入N,N-二异丙基乙胺(0.5mL)和N,N,N'-三甲基乙二胺(0.36mL，2.8mmol)。混合物加热到80℃反应2h，反应完成后，加入乙酸乙酯和水。充分搅拌，加硅藻土过滤，有机相用饱和食盐水多次洗涤，干燥浓缩后，柱层析得到化合物101(500mg)。MS(ESI)(m/z):[M+H]+476.2。
N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(3-甲基-1H-吲唑-1-基)吡啶-2-基)苯-1,2,4-三胺(化合物102)
Figure PCTCN2015088643-appb-000211
除了将化合物95a换成化合物101之外，与化合物96a同样地制备化合物102。1H NMR(400MHz,CDCl3):δ8.32(d,J＝6.0Hz,1H),7.87(d,J＝8.3Hz,1H),7.76(d,J＝8.2Hz,1H),7.64(s,1H),7.58-7.44(m,1H),7.34-7.24(m,2H),7.20(s,1H),6.99(s,1H),6.74(s,1H),3.85(s,3H),3.01(t,J＝6.6Hz,2H),2.70(s,3H),2.67(s,3H),2.48(m,2H),2.33(s,6H)；MS(ESI)(m/z):[M+H]+446.3。
实施例45：N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(3-甲基-1H-吲唑-1-基)吡啶-2-基)氨基)苯基)丙烯酰胺(化合物103)
Figure PCTCN2015088643-appb-000212
除了将化合物96a换成化合物102之外，与化合物97a同样地制备化合物103。1H NMR(400MHz,CDCl3)δ10.16(s,1H),9.22(s,1H),8.35(d,J＝5.6Hz,1H),8.11(d,J＝8.5Hz,1H),7.71(d,J＝8.0Hz,1H),7.48(t,J＝7.7Hz,1H),7.38(s,1H),7.27-7.24(m,2H),7.03(s,1H),6.82(s,1H),6.52-6.42(m,J＝19.5Hz,2H),5.75(d,J＝11.6Hz,1H),3.88(s,3H),2.92(m,2H),2.74(s,3H),2.66(s,3H),2.31-2.25(m,8H)；MS(ESI)(m/z):[M+H]+500.3。
生物活性测试部分
以下描述了MTT细胞增殖试验。
肿瘤细胞增殖及生长抑制分析：H1975(非小细胞肺癌细胞，EGFR L858K/T790M)、Α431(人表皮癌细胞，EGFR野生型)细胞均从ATCC细胞库购买。H1975细胞的生长培养基为RPMI-1640(GIBCO，A10491-065)，10％胎牛血清。A431细胞的生长培养基为DMEM(GIBCO，11995-065)，10％胎牛血清。采用CellTiter-Glo分析法检测化合物对肿瘤细胞的增殖活性的影响。肿瘤细胞暴露在处理条件下72h，各细胞系每次实验所使用的细胞密度根据细胞72h生长曲线进行调整。为待测试化合物设置了10个浓度梯度3倍稀释(0.5nM-10μM)，每个浓度值使用3组平行对照。
将对数生长期的细胞用胰蛋白酶消化制成细胞悬液，Roche计数仪计数并用完全培养基适当稀释，使细胞终浓度为1～2×103个cell/mL。将细胞接种于384孔板中，每孔为22.6μL，设置3组平行，置于37℃，5％CO2培养箱中培养过夜。将化合物溶于DMSO，配制浓度为10μmol/L的母液，随后将化合物用BRAVO仪器进行梯度稀释，逐步稀释得到的化合物浓度分别为10、3.33、1.11、0.37、0.123、0.041、0.0137、0.00457、0.00152、0.0005μmol/L。取2μL化合物溶液加到18μL的培养基中，充分混合后，再取2μL化合物和培养基混合溶液加到18μL的培养基中，充分混合。取2.4μL的混合物加入384孔板中。2.4μL稀释后的DMSO代替化合物溶液用作0％抑制对照(DMSO终浓度要小于0.1％，以减少DMSO带来的影响)。培养72h 之后,加入24μL CellTiter-Glo试剂。在一个定轨振荡器上混合内容物2min，诱导细胞裂解。将384孔板室温孵育10min，使荧光信号值稳定。用
Figure PCTCN2015088643-appb-000213
M1000PRO(TECAN)读取数据。数据使用GraphPad Prism version 5.0进行计算，GI50值通过使用剂量反应曲线的非线性回归模型调整得到。
测试结果如下表所示。
Figure PCTCN2015088643-appb-000214
Figure PCTCN2015088643-appb-000215

Claims (21)

  1. 式(I)所示的化合物或其药学上可接受的盐、或其溶剂化物：
    Figure PCTCN2015088643-appb-100001
    其中，X选自碳，Y选自碳，Z选自C-Ra，R1选自甲氧基；或X选自氮，Y选自碳，Z选自C-Rb，R1选自甲氧基或二氟甲氧基；或X选自氮，Y选自氮，Z选自C-Rc，R1选自甲氧基或二氟甲氧基；或X选自氮，Y选自氮，Z选自C-Rd，R1选自二氟甲氧基；
    Ra选自3-甲基-1H-吲唑-1-基、1-甲基-1H-吡咯并[2,3-b]吡啶-3-基、1-甲基-1H-噻吩并[3,2-c]吡唑-3-基、1-甲基-1H-吡唑并[4,3-b]吡啶-3-基、吡唑并[1,5-a]嘧啶-3-基或咪唑并[1,2-a]吡啶-3-基；
    Rb选自苯并[d]异恶唑-3-基、1-甲基-1H-吡咯并[3,2-b]吡啶-3-基、2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基、2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基或1-甲基-1H-噻吩并[3,2-c]吡唑-3-基；
    Rc选自1H-苯并[d]咪唑-1-基、1H-吲哚-7-基或1-甲基-1H-吲哚-7-基；
    Rd选自1-甲基-1H-吡咯并[2,3-b]吡啶-3-基；
    R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
  2. 根据权利要求1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(II)所示的结构，
    Figure PCTCN2015088643-appb-100002
    其中，Ra选自3-甲基-1H-吲唑-1-基、1-甲基-1H-吡咯并[2,3-b]吡啶-3-基、 1-甲基-1H-噻吩并[3,2-c]吡唑-3-基、1-甲基-1H-吡唑并[4,3-b]吡啶-3-基、吡唑并[1,5-a]嘧啶-3-基或咪唑并[1,2-a]吡啶-3-基；
    R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
  3. 根据权利要求1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(III)所示的结构，
    Figure PCTCN2015088643-appb-100003
    其中，Rb选自苯并[d]异恶唑-3-基、1-甲基-1H-吡咯并[3,2-b]吡啶-3-基、2,4-二甲基-4H-噻吩并[3,2-b]吡咯-6-基、2,5,6-三甲基-6H-噻吩并[2,3-b]吡咯-4-基或1-甲基-1H-噻吩并[3,2-c]吡唑-3-基；
    R1选自甲氧基或二氟甲氧基；
    R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
  4. 根据权利要求1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(IV)所示的结构，
    Figure PCTCN2015088643-appb-100004
    其中，Rc选自1H-苯并[d]咪唑-1-基、1H-吲哚-7-基或1-甲基-1H-吲哚-7-基；
    R1选自甲氧基或二氟甲氧基；
    R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
  5. 根据权利要求1所述的化合物或其药学上可接受的盐、或其溶剂化物，其具有式(V)所示的结构，
    Figure PCTCN2015088643-appb-100005
    其中，Rd选自1-甲基-1H-吡咯并[2,3-b]吡啶-3-基；
    R2选自(2-(二甲氨基)乙基)(甲基)氨基、4-甲基哌嗪-1-基、3-(二甲氨基)氮杂环丁-1-基、4-(二甲氨基)哌啶-1-基、(S)-2-((二甲氨基)甲基)吡咯烷-1-基或5-甲基-2,5-二氮杂螺[3.4]辛-2-基。
  6. 根据权利要求3所述的化合物或其药学上可接受的盐、或其溶剂化物，其特征在于，Rb选自苯并[d]异恶唑-3-基、1-甲基-1H-吡咯并[3,2-b]吡啶-3-基或1-甲基-1H-噻吩并[3,2-c]吡唑-3-基。
  7. 根据权利要求4所述的化合物或其药学上可接受的盐、或其溶剂化物，其特征在于，Rc为1H-吲哚-7-基。
  8. 根据权利要求1～7中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物，其特征在于，R2为(2-(二甲氨基)乙基)(甲基)氨基。
  9. 根据权利要求1～5中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物，其特征在于，所述式(I)所示的化合物选自以下化合物中的任意一种：N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺、N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H- 噻吩并[3,2-c]吡咯-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)苯基)丙烯酰胺、N-(4-(二氟甲氧基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-5-((4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2-基)氨基)苯基)丙烯酰胺、N-(5-((4-(苯并[d]异恶唑-3-基)嘧啶-2-基)氨基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基苯基)丙烯酰胺、N-(2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基-5-((4-(1-甲基-1H-噻吩并[3,2-c]吡咯-3-基)嘧啶-2-基)氨基)苯基)丙烯酰胺、或N-(5-((4-(1H-吲哚-7-基)-1,3,5-三嗪-2-基)氨基)-2-((2-(二甲氨基)乙基)(甲基)氨基)-4-甲氧基苯基)丙烯酰胺。
  10. 根据权利要求1～9中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物，其用于治疗癌症。
  11. 根据权利要求10所述的化合物或其药学上可接受的盐、或其溶剂化物，所述癌症包括非小细胞肺癌、乳腺癌、神经胶质细胞瘤、前列腺癌、卵巢癌、头颈部鳞癌、宫颈癌、食管癌、肝癌、肾癌、胰腺癌、结肠癌、皮肤癌、白血病、淋巴瘤、胃癌、多发性骨髓癌或实体瘤。
  12. 一种药物组合物，包含根据权利要求1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物以及药学上可接受的稀释剂和/或载体。
  13. 根据权利要求1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物作为药物的用途。
  14. 根据权利要求1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物在制备治疗癌症的药物中的用途。
  15. 根据权利要求14所述的用途，其特征在于，所述癌症包括非小细胞肺癌、乳腺癌、神经胶质细胞瘤、前列腺癌、卵巢癌、头颈部鳞癌、宫颈癌、食管癌、肝癌、肾癌、胰腺癌、结肠癌、皮肤癌、白血病、淋巴瘤、胃癌、多发性骨髓癌或实体瘤。
  16. 在需要治疗的温血动物例如人中产生抗癌作用的方法，其包括：向所述动物给药有效量的根据权利要求1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物。
  17. 根据权利要求1～11中任一项所述的化合物或其药学上可接受的盐、 或其溶剂化物和附加的抗肿瘤物质的用途，用于癌症的同时，独立或序贯治疗。
  18. 一种制备根据权利要求1～11中任一项所述的化合物或其药学上可接受的盐、或其溶剂化物的方法，包括：
    在有机溶剂的存在下，使下述式(VI)所示的化合物或其盐与羧酸或羧酸衍生物反应，
    Figure PCTCN2015088643-appb-100006
    式(VI)中，X、Y、Z、R1和R2与所述式(I)同义。
  19. 根据权利要求18所述的方法，其特征在于，所述有机溶剂包括二氯甲烷、四氢呋喃、N,N-二甲基甲酰胺或N,N-二甲基乙酰胺。
  20. 根据权利要求18或19所述的方法，其特征在于，所述羧酸或羧酸衍生物包括丙烯酸、丙烯酰氯或丙烯酸酯。
  21. 根据权利要求18～20中任一项所述的方法，其特征在于，所述式(VI)所示的化合物选自以下化合物中的任意一种：N4-(4-(4-苯并[d]异恶唑-3-基)嘧啶-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺、N4-(4-(苯并[d]异恶唑-3-基)嘧啶-2-基)-5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲苯-1,2,4-三胺、N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,2,4-三胺、5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[3,2-b]吡啶-3-基)嘧啶-2-基)苯-1,2,4-三胺、N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,2,4-三胺、5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-噻吩并[3,2-c]吡唑-3-基)嘧啶-2-基)苯-1,2,4-三胺、N4-(4-(1H-吲哚-7基)-1,3,5-三嗪-2-基)-N1-(2-(二甲氨基)乙基)-5-甲氧基-N1-甲苯-1,2,4-三胺、N4-(4-(1H-吲哚-7基)-1,3,5-三嗪-2-基)-5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲苯-1,2,4-三胺或5-(二氟甲氧基)-N1-(2-(二甲氨基)乙基)-N1-甲基-N4-(4-(1-甲基-1H-吡咯并[2,3-b]吡啶-3-基)-1,3,5-三嗪-2- 基)苯-1,2,4-三胺。
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