WO2022083741A1 - 吡唑并吡啶类化合物或其盐及其制备方法和用途 - Google Patents

吡唑并吡啶类化合物或其盐及其制备方法和用途 Download PDF

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WO2022083741A1
WO2022083741A1 PCT/CN2021/125738 CN2021125738W WO2022083741A1 WO 2022083741 A1 WO2022083741 A1 WO 2022083741A1 CN 2021125738 W CN2021125738 W CN 2021125738W WO 2022083741 A1 WO2022083741 A1 WO 2022083741A1
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compound
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alkyl
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membered
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French (fr)
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程耀邦
王永辉
董志强
周娟
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上海辉启生物医药科技有限公司
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/415—1,2-Diazoles
    • A61K31/4162—1,2-Diazoles condensed with heterocyclic ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/437—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a five-membered ring having nitrogen as a ring hetero atom, e.g. indolizine, beta-carboline
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4427—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems
    • A61K31/444—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems containing a six-membered ring with nitrogen as a ring heteroatom, e.g. amrinone
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • A61K31/4523—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems
    • A61K31/4545—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems containing a six-membered ring with nitrogen as a ring hetero atom, e.g. pipamperone, anabasine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/55—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having seven-membered rings, e.g. azelastine, pentylenetetrazole
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
    • C07B2200/00—Indexing scheme relating to specific properties of organic compounds
    • C07B2200/07—Optical isomers

Definitions

  • the invention belongs to the technical field of chemical medicine, and in particular relates to a pyrazolopyridine compound with RET inhibitory activity, a preparation method thereof, and a pharmaceutical composition containing the compound, and also relates to the preparation of the pyrazolopyridine compound for prevention or treatment and Use in medicine for RET-related diseases.
  • RET REarranged during Transfection protein is a receptor tyrosine kinase (RTK) and a transmembrane glycoprotein, expressed by the proto-oncogene RET located on chromosome 10, in the kidney and intestine of embryonic stage. It plays an important role in the development of the nervous system and is also critical in a variety of tissues, such as neurons, neuroendocrine, hematopoietic tissues, and male germ cells. Unlike other receptor tyrosine kinases, RET does not bind directly to ligand molecules: such as artemin, glial cell-derived neurotrophic factor (GDNF), and nerve growth factor (NGF), all of which are It belongs to the GNDF family of ligands (GFLs).
  • GDNF glial cell-derived neurotrophic factor
  • NGF nerve growth factor
  • GFR ⁇ GDNF family receptor alpha
  • RET protein GDNF family receptor alpha
  • PI3K PI3K
  • JAK-STAT PKA
  • PKC PKC
  • RET oncogenic activation of RET
  • one is a new fusion protein generated by chromosomal rearrangement, usually a fusion of the kinase domain of RET and a protein containing a self-dimerization domain; the other is RET mutation directly or indirectly.
  • RET kinase activity is activated.
  • RET chromosomal rearrangements are found in 10%-20% of papillary thyroid cancer (PTC) patients; RET point mutations are found in 60% of medullary medullary thyroid carcinomas (MTC); in all non-small cell lung cancers ( About 1-2% of patients with NSCLC have RET fusion proteins, of which KIF5B is the most common.
  • Drugs currently on the market or under clinical development that are selectively designed to target RET have shown good efficacy and safety in clinical trials of non-small cell lung cancer and thyroid cancer.
  • the present invention relates to compounds useful in the prevention or treatment of RET-related diseases.
  • the compounds of the present invention exhibit satisfactory RET inhibitory activity. Therefore, the compounds of the present invention can not only achieve the purpose of preventing or treating RET-related diseases, but also the prepared medicaments are expected to have improved absorption, increased efficacy at the same dose, or provide the same efficacy and/or lower doses. or reduce possible side effects.
  • the present invention also provides the use of a compound of the present invention in the manufacture of a medicament for the prevention or treatment of a disease associated with RET, a pharmaceutical composition comprising the compound, and the prevention and/or treatment of RET associated with RET by administering the compound methods related to the disease.
  • R 1 is selected from hydrogen, halogen, cyano, nitro and C 1 -C 6 alkyl optionally substituted with halogen or cyano;
  • R 2 is selected from hydrogen, C 6 -C 10 aryl, 5-9 membered heteroaryl, C 3 -C 8 cycloalkyl, C 3 -C 8 cycloalkenyl, 3-8 membered heterocycloalkyl, 3 -8-membered heterocycloalkenyl, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio, wherein said aryl, heteroaryl, cycloalkyl, cycloalkene group, heterocycloalkyl, heterocycloalkenyl, alkyl, alkoxy and alkylthio optionally substituted with 1, 2 or 3 groups independently selected from the group consisting of halogen, cyano, nitro, hydroxy , C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio;
  • R 3 is selected from hydrogen, halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio;
  • R 4 is selected from hydrogen, C 6 -C 10 aryl, 5-9 membered heteroaryl, C 3 -C 8 cycloalkyl, C 3 -C 8 cycloalkenyl, 3-8 membered heterocycloalkyl, 3 -8-membered heterocycloalkenyl, -SO 2 -C 1 -C 6 alkyl, -SO-C 1 -C 6 alkyl, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C alkylthio , wherein the aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl, alkyl, alkoxy and alkylthio groups are optionally independently Substituted with 1, 2 or 3 groups selected from the group consisting of halogen, cyano, nitro, C1 - C6 alkyl, C1 - C6 alkoxy and C1 -
  • R 5 is selected from halogen, cyano, nitro, hydroxyl, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio;
  • Ring A is selected from C 6 -C 10 arylene, 5-9 membered heteroarylene, C 3 -C 8 cycloalkylene, C 3 -C 8 cycloalkenylene, 3-8 membered heterocycloalkane base and 3- to 8-membered heterocycloalkenyl rings;
  • Ring B is selected from C 6 -C 10 aryl, 5-9 membered heteroaryl, C 3 -C 8 cycloalkyl, C 3 -C 8 cycloalkenyl, 3-8 membered heterocycloalkyl and 3-8 A membered heterocycloalkenyl ring;
  • n 0, 1, 2, or 3;
  • n 0, 1, 2 or 3.
  • a compound of formula (I) in another aspect of the present invention there is provided a compound of formula (I), its stereoisomers, tautomers, stable isotopic variants for use in the treatment or prevention, especially for the treatment of RET-related diseases , a pharmaceutically acceptable salt or solvate.
  • a pharmaceutical composition comprising a compound of the present invention and a pharmaceutically acceptable excipient.
  • the pharmaceutical composition of the present invention is provided for preventing or treating RET-related diseases.
  • the pharmaceutical composition may additionally contain additional therapeutically active ingredients suitable for use in combination with the compounds of the present invention.
  • a pharmaceutical combination comprising a compound of the present invention and an additional active agent.
  • a method for preventing or treating a disease associated with RET in an individual comprising administering an effective amount of a compound of the invention described herein or comprising its pharmaceutical composition.
  • the RET-related disease described in the present invention is selected from tumors or irritable bowel syndrome (IBS), and tumors include but are not limited to non-small cell lung cancer, small cell lung cancer, papillary thyroid cancer, medullary thyroid cancer, Differentiated thyroid cancer, recurrent thyroid cancer, refractory differentiated thyroid cancer, polyendocrine tumor 2A or 2B, pheochromocytoma, parathyroid hyperplasia, breast cancer, colorectal cancer, papillary renal cell carcinoma, gastrointestinal mucosa Gangliomas, pancreatic duct adenocarcinoma, multiple endocrine tumors, testicular cancer, chronic monocytic leukemia, salivary gland cancer, ovarian cancer, cervical cancer, etc.
  • IBS irritable bowel syndrome
  • halo or halogen as used herein means fluorine (F), chlorine (Cl), bromine (Br) and iodine (I). Preferred halo are fluoro or chloro.
  • halogen-substituted groups as used herein is intended to include mono- or polyhalogenated groups wherein one or more (eg, 2, 3, 4, 5 or 6) of the same or different halogen substituents One or more (eg 2, 3, 4, 5 or 6) hydrogens in the group.
  • cyano as used herein means the group -CN.
  • nitro as used herein means the group -NO2 .
  • hydroxyl refers to -OH.
  • alkyl refers to a straight or branched chain saturated hydrocarbon group consisting of carbon atoms and hydrogen atoms. Specifically, the alkyl group has 1 to 10, eg, 1 to 6, 1 to 5, 1 to 4, 1 to 3, or 1 to 2 carbon atoms.
  • Ci - C6 alkyl refers to a straight or branched chain saturated hydrocarbon group having 1 to 6 carbon atoms, examples of which are methyl, ethyl, propyl (including n-propyl and isopropyl), butyl (including n-butyl, isobutyl, sec-butyl or tert-butyl), pentyl (including n-pentyl, isopentyl, neopentyl), n-hexyl, 2-methylpentyl, etc.
  • Particular alkyl groups have 1 to 3 carbon atoms.
  • alkoxy means the group -O-alkyl, wherein alkyl has the meaning set forth herein. Specifically, the term includes the groups -OC 1-6 alkyl, more specifically -OC 1-3 alkyl. Representative examples of alkoxy include, but are not limited to, methoxy, ethoxy, propoxy (including n-propoxy, isopropoxy), butoxy (including n-butoxy, isobutoxy, tert-butoxy), pentyloxy (including n-pentyloxy, isopentyloxy, neopentyloxy), hexyloxy (including n-hexyloxy, isohexyloxy) and the like. Particular alkoxy groups have 1 to 3 carbon atoms.
  • alkylthio refers to an -S-alkyl group, wherein the alkyl group is as defined above for "alkyl”. Specifically, the term includes the groups -SC 1-6 alkyl, more specifically -SC 1-3 alkyl.
  • alkylthio include, but are not limited to, methylthio, ethylthio, propylthio (including n-propylthio, isopropylthio), butylthio (including n-butylthio, isobutylthio, tert-butylthio), pentylthio (including n-pentylthio, isopentylthio, neopentylthio), hexylthio (including n-hexylthio, isohexylthio) and the like.
  • Particular alkylthio groups have 1 to 3 carbon atoms.
  • halogen-substituted C1 - C6 alkyl refers to the C1 - C6 alkyl groups described above, wherein one or more (eg 1, 2, 3, 4 or 5) ) hydrogen atoms are replaced by halogens. It will be understood by those skilled in the art that when there is more than one halogen substituent, the halogens may be the same or different, and may be located on the same or different C atoms.
  • halogen substituted C1 - C6 alkyl are eg -CH2F , -CHF2 , -CF3 , -CCl3 , -C2F5 , -C2Cl5 , -CH2CF3 , -CH 2 Cl, -CH 2 CH 2 CF 3 or -CF(CF 3 ) 2 , etc.
  • halogen-substituted C1 - C6alkoxy refers to the above-described C1 - C6alkoxy groups, wherein one or more (eg 1, 2, 3, 4 or 5) hydrogen atoms are replaced by halogens. It will be understood by those skilled in the art that when there is more than one halogen substituent, the halogens may be the same or different, and may be located on the same or different C atoms.
  • halogen substituted C 1 -C 6 alkoxy are eg -OCH 2 F, -OCHF 2 , -OCF 3 , -OCCl 3 , -OC 2 F 5 , -OC 2 Cl 5 , -OCH 2 CF 3 , -OCH 2 Cl or -OCH 2 CH 2 CF 3 , etc.
  • cycloalkyl refers to a monocyclic, fused polycyclic, bridged polycyclic or spirocyclic non-aromatic saturated monovalent hydrocarbon ring structure having the specified number of ring atoms. Cycloalkyl groups may have 3 to 12 carbon atoms (ie C3 - C12 cycloalkyl), eg 3 to 10, 3 to 8, 3 to 7, 3 to 6, 5 to 6 carbon atoms .
  • Suitable cycloalkyl groups include, but are not limited to, monocyclic structures, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, or cyclooctyl; or polycyclic (eg, bicyclic) structures, including spiro Ring, fused or bridged systems such as bicyclo[1.1.1]pentyl, bicyclo[2.2.1]heptyl, spiro[3.4]octyl, bicyclo[3.1.1]hexyl, bicyclo[3.1. 1] heptyl or bicyclo[3.2.1] octyl, etc.).
  • monocyclic structures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, or cyclooctyl
  • polycyclic (eg, bicyclic) structures including spiro Ring
  • cycloalkylene refers to a cycloalkyl group as defined above, but which is a divalent group and the two bonds are not on the same ring atom.
  • Cycloalkylene may have 3 to 12 carbon atoms (ie C3- C12cycloalkylene ), eg 3 to 10, 3 to 8, 3 to 7, 3 to 6, 5 to 6 carbon atom.
  • suitable cycloalkylene groups include, but are not limited to, monocyclic structures such as cyclopropylene, cyclobutylene, cyclopentylene (eg, cyclopent-1,2-diyl, cyclopent-1,3- diyl), cyclohexylene (e.g.
  • cycloalkenyl means a monocyclic, fused polycyclic, bridged polycyclic, or spirocyclic non-aromatic unsaturated hydrocarbon ring structure having the specified number of ring atoms, comprising at least one (eg, 1, 2, or 3) carbon-carbon double bonds.
  • Cycloalkenyl groups may have 3 to 12 carbon atoms (ie, C3 - C12 cycloalkenyl groups), such as 3 to 10, 3 to 8, 3 to 7, 3 to 6, 5 to 6 carbon atoms .
  • Suitable cycloalkenyl groups include, but are not limited to, monocyclic structures such as cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclopentadienyl, cyclohexenyl, cyclohexadienyl, cycloheptene cycloheptadienyl, cycloheptatrienyl, or cyclooctenyl.
  • cycloalkenylene refers to a cycloalkenyl group as defined above, but which is a divalent group in which the two bonds are not on the same ring atom.
  • Cycloalkenylene may have 3 to 12 carbon atoms (ie, C3 - C12 cycloalkenylene), for example 3 to 10, 3 to 8, 3 to 7, 3 to 6, 5 to 6 carbon atom.
  • Suitable cycloalkenylene groups include, but are not limited to, monocyclic structures such as cyclopropenylene, cyclobutenylene, cyclopentenylene, cyclopentadienylene, cyclohexenylene, cyclohexylene Dienyl, cycloheptenylene, cycloheptadienylene, cycloheptatrienylene or cyclooctenylene.
  • heterocycloalkyl as used herein means a monocyclic, fused, monocyclic, fused ring comprising one or more (eg 1, 2, 3 or 4) heteroatoms independently selected from O, N and S and the specified number of ring atoms
  • Heterocycloalkyl may have 3 to 12 ring members (may be referred to as 3-12 membered heterocycloalkyl), for example 3 to 10 ring members, 3 to 8 ring members, 3 to 7 ring members, 4 to 7 ring members, 5 to 6 ring members.
  • Heterocycloalkyl groups typically contain up to 4 (eg, 1, 2, 3, or 4) heteroatoms.
  • suitable heterocycloalkyl groups include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl (eg, 1-pyrrolidinyl, 2-pyrrolidinyl, and 3-pyrrolidinyl).
  • -pyrrolidinyl tetrahydrofuranyl (eg 1-tetrahydrofuranyl, 2-tetrahydrofuranyl and 3-tetrahydrofuranyl), tetrahydrothienyl (eg 1-tetrahydrothienyl, 2-tetrahydrothienyl and 3-tetrahydrothienyl) thienyl), piperidinyl (such as 1-piperidinyl, 2-piperidinyl, 3-piperidinyl and 4-piperidinyl), tetrahydropyranyl (such as 4-tetrahydropyranyl), Tetrahydrothiopyranyl (eg 4-tetrahydrothiopyranyl), morpholinyl (eg morpholino), thiomorpholinyl, dioxanyl, piperazinyl or azepanyl, diazepine Cycloheptyl groups such as 1,4-diazacycloheptyl, 3,6-
  • heterocycloalkylene as used herein means a heterocycloalkyl group as defined above, but which is a divalent group in which the two bonds are not on the same ring atom.
  • Heterocycloalkylene may have 3 to 12 ring members (may be referred to as 3-12 membered heterocycloalkylene), such as 3 to 10 ring members, 3 to 8 ring members, 3 to 7 ring members, 4 to 7 ring members, 5 to 6 ring members.
  • Heterocycloalkylenes typically contain up to 4 (eg, 1, 2, 3, or 4) heteroatoms.
  • heterocycloalkylenes examples include, but are not limited to, azetidine, oxetylene, thietanylene, pyrrolidylene (eg, pyrrolidine-1,2-di pyrrolidine-1,3-diyl, pyrrolidine-2,3-diyl), tetrahydrofuranylidene (such as tetrahydrofuran-2,4-diyl, tetrahydrofuran-2,3-diyl and tetrahydrofuran-2, 5-diyl), piperidinylene (e.g.
  • structures having asymmetric centers encompass their racemic and/or single enantiomeric forms, such as representable and / or
  • heterocycloalkenyl as used herein means “heterocycloalkyl” as defined herein containing at least one (eg, 1, 2 or 3) double bond.
  • suitable heterocycloalkenyl groups include, but are not limited to:
  • tetrahydropyranyl eg 4-tetrahydropyranyl
  • tetrahydrothiopyranyl eg 4-tetrahydrothiopyranyl
  • heterocycloalkenylene means a heterocycloalkenyl group as defined above, but which is a divalent group in which the two bonds are not on the same ring atom.
  • suitable heterocycloalkenylene groups include, but are not limited to:
  • aryl as used herein means a monovalent aromatic hydrocarbon group derived by removing one hydrogen atom from a single carbon atom in an aromatic ring system. Specifically, aryl refers to a monocyclic or fused polycyclic aromatic ring structure having the specified number of ring atoms. In particular, the term includes groups comprising 6 to 14, such as 6 to 10, preferably 6, ring members. Particular aryl groups include phenyl and naphthyl, the most particular aryl group being phenyl.
  • arylene as used herein means an aryl group as defined above, but which is a divalent group in which the two bonds are not on the same ring atom.
  • Particular arylene groups include phenylene groups such as benzene-1,2-diyl, benzene-1,3-diyl or benzene-1,4-diyl.
  • heteroaryl as used herein means a monocyclic or fused ring comprising one or more (eg 1, 2, 3 or 4) heteroatoms independently selected from O, N and S and the specified number of ring atoms
  • a heteroaryl group can be, for example, a 5-6 membered monocyclic ring, or a fused bicyclic structure formed from two 5-membered rings fused, or a fused 5-membered ring and a 4-membered ring.
  • the heteroaryl ring contains at least one ring nitrogen atom, at least one ring sulfur atom, or at least one epoxy atom.
  • a heteroaryl group can be a 5-6 membered heteroaryl group containing 1 or 2 heteroatoms independently selected from N, O, or S.
  • suitable 5-membered monocyclic heteroaryl groups include, but are not limited to, pyrrolyl, furyl, thienyl, imidazolyl, furazanyl, oxazolyl, oxadiazolyl, oxtriazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, triazolyl, and tetrazolyl;
  • suitable 6-membered monocyclic heteroaryl groups include, but are not limited to, pyridyl, pyrazinyl, pyridazinyl, pyrimidinyl, and triazine base.
  • heteroarylene as used herein means a heteroaryl group as defined above, but which is a divalent group in which the two bonds are not on the same ring atom.
  • a heteroarylene group can be a 5-6 membered heteroarylene group containing 1 or 2 heteroatoms independently selected from N, O, or S.
  • suitable 5-membered monocyclic heteroarylenes include, but are not limited to, pyrrolidine, furanylene, thienylene, imidazolylylene, furazanylidene, oxazolylylene, oxadiazolylylene, oxane Triazolyl, isoxazolylylene, thiazolylidene, isothiazolylidene, pyrazolylidene, triazolylylene, and tetrazolylylene;
  • suitable 6-membered monocyclic heteroaryl groups include, but are not limited to Pyridinyl, pyrazinylene, pyridazinylene, pyrimidinylene and triazinylene, preferably
  • Substituents described as "optionally substituted” mean that the group may be unsubstituted or substituted by one or more (eg, 0, 1, 2, 3, 4, or 5 or more, or any derivatized therein). range) is substituted with the listed substituents for that group, wherein the substituents may be the same or different.
  • an optionally substituted group is substituted with 1 substituent.
  • an optionally substituted group is substituted with 2 substituents.
  • an optionally substituted group is substituted with 3 substituents.
  • an optionally substituted group is substituted with 4 substituents.
  • heterocycles whether aromatic or non-aromatic, in which the maximum number of heteroatoms or the type of heteroatoms contained is determined by ring size, degree of unsaturation, and valence of the heteroatoms. Decide.
  • a heterocycle can have 1 to 4 heteroatoms, provided that the heterocycle or heteroaromatic ring is chemically feasible and stable.
  • pharmaceutically acceptable means approved by or by the appropriate agency in each country, or listed in a generally recognized pharmacopoeia for use in animals and more particularly in humans, or when administered in an appropriate amount to animals such as humans Molecular entities and compositions that do not produce adverse, allergic or other adverse reactions.
  • pharmaceutically acceptable salt means a salt of a compound of the present invention that is pharmaceutically acceptable and possesses the desired pharmacological activity of the parent compound.
  • such salts are nontoxic and can be inorganic acid addition salts or organic acid addition salts and base addition salts.
  • such salts include: (1) acid addition salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; or acid addition salts formed with organic acids, which Organic acids such as acetic acid, propionic acid, caproic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandel acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, glucoheptanoic acid, 3-phenylpropionic acid, trimethylacetic acid, tert-butyl Acetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid,
  • prodrug means a compound having a cleavable group that becomes a compound of the present invention pharmaceutically active in vivo by solvolysis or under physiological conditions, including derivatives of the compound of the present invention.
  • Prodrugs include acid derivatives well known in the art, such as esters prepared by reacting the parent acid with a suitable alcohol, or amides prepared by reacting the parent acid compound with a substituted or unsubstituted amine, or anhydrides or mixed anhydrides.
  • Simple aliphatic or aromatic esters, amides, and anhydrides derived from the pendant acid groups of the compounds of the present invention are particularly suitable prodrugs.
  • Particular such prodrugs are C1-8 alkyl, C2-8 alkenyl, optionally substituted C6-10 aryl, and ( C6-10 aryl)-( C1- 4 alkyl) esters.
  • the present invention also includes all pharmaceutically acceptable isotopic compounds that are identical to the compounds of the present invention, except that one or more atoms have the same atomic number but an atomic mass or mass number different from the atomic mass or mass that predominates in nature number of atomic substitutions.
  • isotopes suitable for inclusion in the compounds of the present invention include, but are not limited to, isotopes of hydrogen (eg, 2H, 3H); isotopes of carbon (eg, 11C, 13C, and 14C); isotopes of chlorine (eg, 36Cl); isotopes of fluorine isotopes of iodine (such as 123I and 125I); isotopes of nitrogen (such as 13N and 15N); isotopes of oxygen (such as 15O, 17O, and 18O); isotopes of phosphorus (such as 32P); and isotopes of sulfur ( such as 35S).
  • isotopes of hydrogen eg, 2H, 3H
  • isotopes of carbon eg, 11C, 13C, and 14C
  • isotopes of chlorine eg, 36Cl
  • isotopes of fluorine isotopes of iodine such as 123
  • stereoisomer refers to isomers formed due to at least one asymmetric center. In compounds having one or more (eg, 1, 2, 3, or 4) asymmetric centers, it may give rise to racemic mixtures, single enantiomers, diastereomeric mixtures, and individual diastereomers. Certain individual molecules may also exist as geometric isomers (cis/trans). Similarly, the compounds of the present invention may exist as mixtures of two or more different structural forms in rapid equilibrium (often referred to as tautomers). Representative examples of tautomers include keto-enol tautomers, phenol-ketone tautomers, nitroso-oxime tautomers, imine-enamine tautomers Wait. For example, a nitroso-oxime can exist in solution in equilibrium in the following tautomeric forms:
  • the compounds of the present invention are intended to be available as stereoisomers (which include cis and trans isomers, optical isomers (eg, R and S enantiomers), diastereomers, Geometric isomers, rotational isomers, conformational isomers, atropisomers and mixtures thereof).
  • stereoisomers which include cis and trans isomers, optical isomers (eg, R and S enantiomers), diastereomers, Geometric isomers, rotational isomers, conformational isomers, atropisomers and mixtures thereof).
  • the compounds of the present invention may exhibit more than one type of isomerism and consist of mixtures thereof (eg, racemic mixtures and pairs of diastereomers).
  • solvate refers to solvent addition forms containing stoichiometric or non-stoichiometric amounts of solvent, including, for example, solvates with water, such as hydrates, or solvates with organic solvents, such as Methanol, ethanol or acetonitrile, ie as methanolate, ethanolate or acetonitrile, respectively; or in the form of any polymorph. It should be understood that such solvates of the compounds of the present invention also include solvates of pharmaceutically acceptable salts of the compounds of the present invention.
  • prophylaxis means administering to an individual, such as a mammal, such as a human, the administration of an or Various compounds of the present invention result in a reduced risk of developing a defined disease.
  • prevention encompasses the use of the compounds of the present invention prior to the diagnosis or determination of any clinical and/or pathological symptoms.
  • treating refers to administering one or more compounds of the invention described herein to a subject, eg, a mammal, eg, a human, having the disease, or a symptom of the disease, for the purpose of To cure, alleviate, alleviate or affect the disease or symptoms of the disease.
  • the disease is a RET-related disease as defined herein, especially an inflammatory or autoimmune disease.
  • RET-related diseases are selected from tumors or irritable bowel syndrome (IBS), tumors including but not limited to non-small cell lung cancer, small cell lung cancer, papillary thyroid cancer, medullary thyroid cancer, Differentiated thyroid cancer, recurrent thyroid cancer, refractory differentiated thyroid cancer, polyendocrine tumor 2A or 2B, pheochromocytoma, parathyroid hyperplasia, breast cancer, colorectal cancer, papillary renal cell carcinoma, gastrointestinal mucosa Gangliomas, pancreatic duct adenocarcinoma, multiple endocrine tumors, testicular cancer, chronic monocytic leukemia, salivary gland cancer, ovarian cancer, cervical cancer, etc.
  • IBS irritable bowel syndrome
  • cancer refers to the growth and proliferation of neoplastic cells, whether malignant or benign, and all precancerous cells and cancer cells and tissues.
  • the cancer or tumor includes but is not limited to colon cancer, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma Carcinoma, adenocarcinoma, sweat gland cancer, sebaceous gland cancer, lung cancer, leukemia, bladder cancer, stomach cancer, cervical cancer, testicular cancer, skin cancer, rectal cancer, thyroid cancer, kidney cancer, uterine cancer, pemphigus cancer, liver cancer, auditory nerve tumor, oligodendroglioma, brain (meningioma), neuroblastoma, eye cancer.
  • the term "therapeutically effective amount” means an amount sufficient to reduce or completely alleviate the symptoms or other deleterious effects of the disorder; reverse, completely stop or slow the progression of the disorder; or reduce the risk of exacerbation of the disorder when administered to an individual to treat a disease.
  • the amount, "effective amount” can vary depending on the compound, the disease and its severity, and the age, weight, etc. of the individual to be treated.
  • the term "individual” as used herein includes human or non-human animals.
  • exemplary human subjects include human subjects (referred to as patients) or normal subjects with a disease (eg, a disease described herein).
  • Non-human animals in the present invention include all vertebrates such as non-mammals (eg birds, amphibians, reptiles) and mammals such as non-human primates, livestock and/or domesticated animals (eg sheep, dogs) , cats, cows, pigs, etc.).
  • compositions refers to comprising one or more compounds of formula (I) or its stereoisomers, tautomers, stable isotope derivatives, pharmaceutically acceptable salts or solvents compositions and carriers generally accepted in the art for the delivery of biologically active compounds to organisms such as humans.
  • the term "pharmaceutical combination" as used herein means that a compound of the present invention may be used in combination with other active agents for the purposes of the present invention.
  • the other active agent may be one or more additional compounds of the present invention, or may be a second or additional (eg, third) compound that is compatible with, that is, does not adversely affect each other, or has complementary activities. ) compound.
  • Such active agents are suitably combined in amounts effective to achieve the intended purpose.
  • the other active agents may be co-administered with the compound of the present invention in a single pharmaceutical composition, or administered separately from the compound of the present invention in separate discrete units, either simultaneously or sequentially when administered separately. The sequential administrations may be close or distant in time.
  • pharmaceutically acceptable excipient or carrier refers to one or more compatible solid or liquid filler or gelling substances, which are pharmacologically inactive, incompatible with the other ingredients in the composition and should be acceptable for administration to warm-blooded animals, such as humans, for use as a carrier or vehicle for the compounds of the present invention in administration forms, examples of which include, but are not limited to, cellulose and its derivatives such as carboxymethyl cellulose sodium, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as magnesium stearate), calcium sulfate, vegetable oils, polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as Tween) class), wetting agents (such as sodium lauryl sulfate), colorants, flavors, stabilizers, antioxidants, preservatives, etc.
  • cellulose and its derivatives such as carboxymethyl cellulose sodium, cellulose acetate,
  • the stereoisomers, tautomers, stable isotopic variants, pharmaceutically acceptable salts or solvates and prodrugs are as described in the definitions section above.
  • the compounds of the present invention are in free form of a compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof; most preferably a compound of formula (I) in free form or a pharmaceutically acceptable salt thereof.
  • Certain compounds of the present invention may exist in polymorphic or amorphous forms, which also fall within the scope of the present invention.
  • the compound of formula (I) may be in the form of a co-crystal with another chemical entity, and this specification includes all such co-crystals.
  • the compounds of the present invention may exist as individual enantiomers or as mixtures of enantiomers.
  • a compound of formula (I), or a pharmaceutically acceptable salt thereof which is a single enantiomer with an enantiomeric excess (%ee) of >95, >98%, or >99%.
  • a single enantiomer is present in >99% enantiomeric excess (%ee).
  • the present invention provides compounds of formula (I), stereoisomers, tautomers, stable isotopic variants, pharmaceutically acceptable salts or solvates thereof:
  • R 1 is selected from hydrogen, halogen, cyano, nitro and C 1 -C 6 alkyl optionally substituted with halogen or cyano;
  • R 2 is selected from hydrogen, C 6 -C 10 aryl, 5-9 membered heteroaryl, C 3 -C 8 cycloalkyl, C 3 -C 8 cycloalkenyl, 3-8 membered heterocycloalkyl, 3 -8-membered heterocycloalkenyl, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio, wherein said aryl, heteroaryl, cycloalkyl, cycloalkene group, heterocycloalkyl, heterocycloalkenyl, alkyl, alkoxy and alkylthio optionally substituted with 1, 2 or 3 groups independently selected from the group consisting of halogen, cyano, nitro, hydroxy , C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio;
  • R 3 is selected from hydrogen, halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio;
  • R 4 is selected from hydrogen, C 6 -C 10 aryl, 5-9 membered heteroaryl, C 3 -C 8 cycloalkyl, C 3 -C 8 cycloalkenyl, 3-8 membered heterocycloalkyl, 3 -8-membered heterocycloalkenyl, -SO 2 -C 1 -C 6 alkyl, -SO-C 1 -C 6 alkyl, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C alkylthio , wherein the aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl, alkyl, alkoxy and alkylthio groups are optionally independently Substituted with 1, 2 or 3 groups selected from the group consisting of halogen, cyano, nitro, C1 - C6 alkyl, C1 - C6 alkoxy and C1 -
  • R 5 is selected from halogen, cyano, nitro, hydroxyl, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio;
  • Ring A is selected from C 6 -C 10 arylene, 5-9 membered heteroarylene, C 3 -C 8 cycloalkylene, C 3 -C 8 cycloalkenylene, 3-8 membered heterocycloalkane base and 3- to 8-membered heterocycloalkenyl rings;
  • Ring B is selected from C 6 -C 10 aryl, 5-9 membered heteroaryl, C 3 -C 8 cycloalkyl, C 3 -C 8 cycloalkenyl, 3-8 membered heterocycloalkyl and 3-8 A membered heterocycloalkenyl ring;
  • n 0, 1, 2, or 3;
  • n 0, 1, 2 or 3.
  • R1 is halo, cyano or nitro.
  • R1 is cyano
  • R2 is selected from 5-6 membered heteroaryl, C1 - C6 containing 1, 2 or 3 heteroatoms independently selected from N, O or S Alkyl, C1 -C6alkoxy, and C1 - C6alkylthio, optionally substituted with 1, 2 , or 3 groups independently selected from halogen, hydroxy, and C1 - C6 alkyl;
  • R 2 is selected from 5-membered heteroaryl, C 1 -C 6 alkoxy containing 1, 2 or 3 heteroatoms independently selected from N, O or S and C 1 -C 6 alkylthio, optionally substituted with 1 group independently selected from the group consisting of hydroxy and C 1 -C 6 alkyl;
  • R2 is selected from 5 containing 1, 2 or 3 heteroatoms independently selected from N, O or S, optionally substituted with C1 - C6 alkyl Member heteroaryl and C 1 -C 6 alkoxy optionally substituted by hydroxy;
  • the 5-6 membered heteroaryl in R is selected from pyrrolyl, furyl, thienyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, pyridyl, pyrazinyl, pyridazinyl or pyrimidinyl;
  • the 5-membered heteroaryl in R 2 is selected from pyrrolyl, furyl, thienyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl or pyrazolyl.
  • R 2 is methyl substituted pyrazolyl or hydroxy substituted C 1 -C 6 alkoxy
  • R 2 is
  • R3 is selected from hydrogen and C1 - C6 alkyl.
  • R3 is selected from hydrogen and C1 - C3 alkyl.
  • R3 is hydrogen or methyl.
  • R3 is hydrogen
  • R4 is selected from phenyl, 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, -SO 2 -C 1 -C 6 alkyl, -SO-C 1 -C 6 alkyl, C 1 -C 6 alkyl, C 1 -C 6 alkoxy and C 1 -C 6 alkylthio, optionally is substituted with 1, 2 or 3 halogen groups; preferably, the 5-6 membered heteroaryl is selected from pyrrolyl, furyl, thienyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, pyridyl, pyrazinyl, pyridazinyl and pyrimidinyl;
  • R4 is selected from 5 -membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, -SO2 - C1- C 6 alkyl, C 1 -C 6 alkyl and C 1 -C 6 alkoxy optionally substituted with 1, 2 or 3 halogen groups; preferably, the 5-membered heteroaryl is selected from pyrrole radical, furyl, thienyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl and pyrazolyl.
  • R4 is selected from pyrazolyl optionally substituted with halogen, -SO2 - C1 - C6 alkyl, C1 - C6 alkyl and C1- C 6 alkoxy.
  • R4 is -SO 2 -CH 3 or methoxy.
  • R 5 is selected from halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, and C 1 -C 6 alkylthio;
  • R 5 is selected from C 1 -C 6 alkyl, C 1 -C 6 alkoxy, and C 1 -C 6 alkylthio.
  • R 5 is selected from C 1 -C 3 alkyl
  • R5 is methyl
  • m is 0 or 1 .
  • n is 0 or 1 .
  • m is 1 .
  • n is zero.
  • Ring A is selected from phenylene, 5-6 membered heteroarylene containing 1, 2 or 3 heteroatoms independently selected from N, O or S, C3- C8cycloalkylene , C3 - C8cycloalkenylene , 3-8 membered heterocycloalkylene containing 1, 2 or 3 heteroatoms independently selected from N, O or S, and 3-8 membered heterocycloalkenylene containing 1, 2 or 3 heteroatoms independently selected from N, O or S;
  • Ring A is selected from phenylene, 5-membered heteroarylene containing 1, 2 or 3 heteroatoms independently selected from N, O or S, C3 -C6cycloalkylene, C3 - C6cycloalkenylene , 3-6 membered heterocycloalkylene containing 1, 2 or 3 heteroatoms independently selected from N, O or S, and containing 1 , 2 or 3 heteroatoms independently selected from N, O or S 3-6 membered heterocycloalkenylene;
  • Ring A is selected from the group consisting of phenylene, pyrrolylene, furanylene, thienylene, imidazolylide, furazanylidene, oxazolylide, oxadiene azolyl, oxtriazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, thiadiazolyl, triazolyl, tetrazolyl, pyridylene, sub Pyrazinyl, pyridazinylene, pyrimidinyl, triazinylene, azetidine, pyrrolidylene, pyrrolidylene, cyclobutylene, cyclopentylene, cyclohexylene, cyclohexylene Cycloheptyl, piperazinylene, piperidinylene and azacycloheptylene.
  • Ring A is selected from the group consisting of pyrazolylidene, azetidine, pyrrolidylene, cyclohexylene, piperidylene, and azacycloheptylene .
  • Ring A is selected from
  • Ring B is selected from phenyl, 5-9 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, C3 -C8cycloalkyl , C3- C8cycloalkenyl , 3-8 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S and containing 1, 2 or 3 3-8 membered heterocycloalkenyl rings of heteroatoms independently selected from N, O or S;
  • Ring B is selected from phenyl and 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S;
  • Ring B is selected from phenyl, and 6-membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S;
  • Ring B is selected from the group consisting of phenyl, pyrrolyl, furyl, thienyl, imidazolyl, furazanyl, oxazolyl, oxadiazolyl, oxtriazolyl, Isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, thiadiazolyl, triazolyl, tetrazolyl, pyridyl, pyrazinyl, pyridazinyl, pyrimidinyl and triazinyl rings.
  • Ring B is a phenyl or pyridyl ring.
  • the compound of formula (I) of the present invention covers each of the above independent embodiments or each specific embodiment, and also covers the embodiment formed by any combination or sub-combination of each of the above-mentioned embodiments or specific embodiments, and also covers the above Any preferred or exemplified combination constitutes an embodiment.
  • the compound of formula (I) of the present invention has the structure of formula (Ia),
  • R 1 , R 2 , R 3 , R 4 , R 5 , m, n and A each have the meanings defined above for compounds of formula (I) in general or in the specific embodiment.
  • the compound of formula (I) of the present invention has the structure of formula (Ib),
  • R 1 , R 2 , R 3 , R 4 , R 5 , n and A each have the meanings defined above for compounds of formula (I) in general or in the specific embodiment.
  • the compound of formula (I) of the present invention has the structure of formula (Ic),
  • R 2 , R 3 , R 4 , R 5 , n and A each have the meanings defined above for compounds of formula (I) in general or in the specific embodiment.
  • the compounds of formula (I) of the present invention are selected from the following specific compounds or stereoisomers, tautomers, stable isotopic variants, pharmaceutically acceptable salts or solvates thereof:
  • the present invention provides a class of pyrazolopyridine compounds with the structural features of the general formula (I). It has been found through research that such compounds can effectively inhibit RET kinase, RET fusion and mutation activities, and serve as a susceptor for related diseases with abnormal RET expression. medicine.
  • High RET kinase inhibitory activity an IC50 in the range of 0.1 nM to 1 ⁇ M, preferably in the range of 0.1 nM to 0.1 ⁇ M, in a kinase RET inhibition assay; and/or
  • the present invention also provides technical solutions in the following aspects.
  • the present invention provides compounds of the present invention for use as medicaments, particularly as RET inhibitors.
  • the present invention provides compounds of the present invention for use in the treatment, especially in the treatment and/or prevention of RET-related diseases.
  • the present invention provides the invention for the treatment and/or prevention of diseases in which RET contributes to the development and progression of the disease or in which inhibition of RET will reduce the incidence of the disease, reduce or eliminate the symptoms of the disease
  • Compounds such as tumors or irritable bowel syndrome (IBS), tumors including but not limited to non-small cell lung cancer, small cell lung cancer, papillary thyroid cancer, medullary thyroid cancer, differentiated thyroid cancer, recurrent thyroid cancer , refractory differentiated thyroid cancer, multiple endocrine tumors 2A or 2B, pheochromocytoma, parathyroid hyperplasia, breast cancer, colorectal cancer, papillary renal cell carcinoma, gastrointestinal mucosal ganglionoma, pancreatic duct adenocarcinoma, Multiple endocrine tumors, testicular cancer, chronic monocytic leukemia, salivary gland cancer, ovarian cancer, cervical cancer, etc.
  • IBS irritable bowel syndrome
  • the compounds of the present invention can be formulated into pharmaceutical compositions according to standard pharmaceutical practice. Meanwhile, based on the good pharmacokinetic properties, improved AUCO-last and good druggability of the compounds of the present invention, medicines with better pharmacokinetic properties and higher bioavailability can be prepared from the compounds of the present invention.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising the above-described compound of the present invention and a pharmaceutically acceptable excipient.
  • compositions of the present invention are provided for use in the prevention or treatment of RET-related diseases, e.g., in mammals such as human subjects.
  • compositions of the present invention may additionally comprise additional therapeutically active ingredients suitable for use in combination with the compounds of the present invention.
  • compositions of the present invention can be formulated by techniques known to those skilled in the art, such as those disclosed in Remington's Pharmaceutical Sciences 20th Edition.
  • the pharmaceutical compositions of the present invention described above can be prepared by admixing a compound of the present invention with one or more pharmaceutically acceptable excipients.
  • the preparation may further include the step of admixing one or more other active ingredients with a compound of the present invention and one or more pharmaceutically acceptable excipients.
  • excipients for inclusion in a particular composition will depend on factors such as the mode of administration and the form of the composition provided. Suitable pharmaceutically acceptable excipients are well known to those skilled in the art and are described, for example, in Ansel, Howard C., et al., Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems.
  • diluents such as glucose, lactose or mannitol
  • carriers pH adjusters, buffers, sweeteners, fillers, stabilizers, surfactants, wetting agents, lubricants, emulsifiers, suspending agents, preservatives agents, antioxidants, opacifiers, glidants, processing aids, colorants, perfuming agents, flavoring agents, other known additives.
  • compositions of the present invention can be administered in a standard manner.
  • suitable modes of administration include oral, intravenous, rectal, parenteral, topical, transdermal, ocular, nasal, buccal or pulmonary (inhalation) administration, wherein parenteral infusion includes intramuscular, intravenous, intraarterial, peritoneal Intra or subcutaneous administration.
  • the compounds of the present invention may be formulated by methods known in the art, for example, as tablets, capsules, syrups, powders, granules, aqueous or oily solutions or suspensions, (lipid) emulsions, dispersible powders, suppositories, Ointments, creams, drops, aerosols, dry powder formulations and sterile injectable aqueous or oily solutions or suspensions.
  • a prophylactic or therapeutic dose of a compound of the invention will vary depending on a range of factors, including the individual being treated, the severity of the disorder or condition, the rate of administration, the disposition of the compound, and the judgment of the prescribing physician.
  • effective doses range from about 0.0001 to about 5000 mg per kg body weight per day, eg, about 0.01 to about 1000 mg/kg/day (single or divided administration). For a 70 kg person, this would add up to about 0.007 mg/day to about 7000 mg/day, eg, about 0.7 mg/day to about 1500 mg/day.
  • the content or amount of the compound of the present invention in the pharmaceutical composition may be from about 0.01 mg to about 1000 mg, suitably 0.1-500 mg, preferably 0.5-300 mg, more preferably 1-150 mg, particularly preferably 1-50 mg, For example 1.5 mg, 2 mg, 4 mg, 10 mg, 25 mg, etc.; accordingly, the pharmaceutical composition of the present invention will comprise 0.05 to 99% w/w (weight percent), such as 0.05 to 80% w/w, such as 0.10 to 70% w/w, eg, 0.10 to 50% w/w of a compound of the invention, all weight percentages are based on the total composition. It will be understood that it may be necessary in certain circumstances to use doses above these limits.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of the present invention and one or more pharmaceutically acceptable excipients, the composition being formulated for oral administration.
  • the composition may be presented in unit dosage form, eg, in the form of a tablet, capsule, or oral liquid.
  • Such unit dosage forms may contain 0.1 mg to 1 g, eg, 5 mg to 250 mg, of a compound of the present invention as the active ingredient.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of the present invention and one or more pharmaceutically acceptable excipients, the composition being formulated for topical administration.
  • Topical administration can be in the form of, for example, creams, lotions, ointments or transdermal patches.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of the present invention and one or more pharmaceutically acceptable excipients, the composition being formulated for administration by inhalation.
  • Administration by inhalation can be by oral inhalation or by intranasal administration.
  • the compounds of the present invention can be effectively used in the present invention in daily doses, eg up to 500 ⁇ g, such as 0.1-50 ⁇ g, 0.1-40 ⁇ g, 0.1-30 ⁇ g, 0.1-20 ⁇ g or 0.1-10 ⁇ g of the present invention compound.
  • compositions of the present invention for oral inhalation may be formulated as dry powders, suspensions (in liquids or gases) or solutions (in liquids), and may be in any suitable form and using any suitable inhaler device known in the art Administration includes, for example, metered dose inhalers (MDIs), dry powder inhalers (DPIs), nebulizers, and soft mist inhalers. Multi-chamber devices can be used to deliver the compounds of the present specification and one or more other active ingredients, when present.
  • MDIs metered dose inhalers
  • DPIs dry powder inhalers
  • nebulizers nebulizers
  • soft mist inhalers soft mist inhalers
  • the compounds of the present invention can be used in methods of treating various disorders in animals, especially mammals such as humans.
  • the present invention provides a method of modulating, especially inhibiting, RET activity, the method comprising contacting a cell with a compound of the invention as previously described to modulate, especially inhibit, RET activity in the cell.
  • the present invention provides a method of preventing or treating a disease associated with RET (eg, a disease treatable or preventable by RET inhibition), the method comprising administering to an individual in need thereof an effective amount of the present invention as previously described A compound of the invention or a pharmaceutical composition of the invention comprising the same.
  • a disease associated with RET eg, a disease treatable or preventable by RET inhibition
  • the present invention provides the use of a compound of the present invention as previously described, or a pharmaceutical composition comprising the same, for inhibiting RET activity, or for treating and/or preventing RET-related diseases, such as by RET inhibition Treatable or preventable disease.
  • the present invention also provides the use of the aforementioned compound of the present invention or the pharmaceutical composition comprising the same in the preparation of medicines, especially the use of medicines with RET receptor inhibitor activity.
  • the present invention provides the use of a compound of the present invention as described above, or a pharmaceutical composition comprising the same, in the manufacture of a medicament for the treatment or prevention of a disease associated with RET, such as a disease treatable or preventable by RET inhibition , wherein the compound or pharmaceutical composition is optionally combined with one or more chemotherapy or immunotherapy.
  • the present invention also provides a process for the preparation of compounds of formula (I), and general synthetic schemes for synthesizing the compounds of the present invention are exemplified below.
  • appropriate reaction conditions are known to those skilled in the art or can be routinely determined.
  • the starting materials and reagents used in the preparation of these compounds are generally commercially available unless otherwise specified, or can be prepared by the methods below, methods analogous to those given below, or methods known in the art.
  • the raw materials and intermediates in the synthetic reaction scheme can be separated and purified by conventional techniques, including but not limited to filtration, distillation, crystallization, chromatography and the like.
  • the materials can be characterized using conventional methods including physical constants and spectral data.
  • the present invention relates to a method for synthesizing a compound of formula (I), comprising the steps of:
  • Reaction of a compound of formula 1-1 with a compound of formula 1-2 provides a compound of formula (I) wherein R 1 , R 2 , R 3 , R 4 , R 5 , m, n, A and B are each as above for formula (I) ) is defined as a compound;
  • the reaction can be carried out in the presence of a condensing agent, which is a condensing agent well known in the art for the coupling of carboxylic acids and amines, including but not limited to 1-propylphosphoric anhydride (T3P), EDC, DCC , HATU, EDCI, etc.; the reaction is preferably carried out in a suitable organic solvent, and the organic solvent can be selected from dichloromethane, tetrahydrofuran, ethers (such as diethyl ether, ethylene glycol monomethyl ether, etc.), N-methyl Pyrrolidone, N,N-dimethylformamide (DMF), N,N-dimethylacetamide, 1,4-dioxane, dimethyl sulfoxide and any combination thereof; the reaction is preferably performed in a suitable Carry out in the presence of alkali, described alkali includes but not limited to sodium carbonate, potassium carbonate, cesium carbonate, N,N-diisoprop
  • R 1 , R 2 , R 5 , n and A are each as defined above for compounds of formula (I);
  • the present invention provides a method for synthesizing a compound of formula I-1', comprising the following steps:
  • Step a formula I-3 and formula I-4 obtain the product of formula I-5 through nucleophilic substitution reaction;
  • the reaction is carried out in the presence of a base;
  • Step b Formula 1-5 with an appropriate boronic acid or boronic acid ester (e.g. ) under the action of a catalyst to obtain the product of formula I-6 through a Suzuki coupling reaction; preferably, the reaction is carried out in the presence of a base; preferably, the catalyst comprises a palladium catalyst, such as Pd(dppf) Cl2.DCM or Pd( dppf)Cl 2 ; and
  • Step c formula I-6 is deprotected in the presence of an acid to obtain a product of formula I-1', preferably, the acid is selected from hydrochloric acid, sulfuric acid, nitric acid, trifluoroacetic acid and acetic acid.
  • bases described in steps a) and b) include, but are not limited to, sodium carbonate, potassium carbonate, cesium carbonate, N,N-diisopropylethylamine, triethylamine, HOBt or pyridine;
  • steps a)-c) are preferably carried out in a suitable organic solvent, which can be selected from alcohols (eg methanol, ethanol), dichloromethane, tetrahydrofuran, ethers (eg diethyl ether, ethylene glycol monomethyl ether) etc.), N-methylpyrrolidone, N,N-dimethylformamide (DMF), N,N-dimethylacetamide, 1,4-dioxane, dimethyl sulfoxide and any combination thereof
  • the reaction is preferably carried out at a suitable temperature, such as -70-200°C, 0-200°C, 10-100°C, 100-150°C, 70-100°C, 20-50°C, room temperature (20-25°C) ) or -70 to 0°C.
  • the above-mentioned synthetic scheme merely exemplifies the preparation methods of some compounds in the present invention.
  • the compounds of the present invention, or stereoisomers, tautomers, stable isotopic derivatives, pharmaceutically acceptable salts or solvates thereof, can be prepared by a variety of methods, including the methods given above, in the Examples
  • the given method or a similar method can be prepared by those of ordinary skill in the art on the basis of the above-mentioned synthetic scheme and in combination with conventional techniques in the art.
  • experimental materials and reagents used in the following examples can be obtained from commercial sources, prepared according to methods in the prior art, or prepared according to methods similar to those disclosed in this application.
  • HATU 2-(7-Azobenzotriazole)-N,N,N',N',-tetramethylurea hexafluorophosphate
  • PE petroleum ether
  • silica gel 300-400 mesh
  • GF254 (0.25 mm) is used for thin-layer chromatography
  • Varian- 400 nuclear magnetic resonance instrument liquid chromatography mass spectrometry (LC/MS) using Agilent TechnologiESI 6120 liquid mass spectrometer.
  • the raw materials used in the present invention are all commercially available raw materials, which can be used directly without further purification, and the temperatures used in the present invention are all in degrees Celsius.
  • Step 1 Synthesis of tert-butyl (1-(5-bromopyridin-3-yl)piperidin-4-yl)carbamate
  • Step 2 1-Amino-3-bromo-5-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)pyridine-1-onium 2,4,6-trimethylbenzenesulfonic acid salt synthesis
  • Step 3 Synthesis of tert-butyl (1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)piperidin-4-yl)carbamate
  • Step 4 (1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)piperidine-4- Synthesis of tert-butyl carbamate
  • Step 6 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)piperidine- Synthesis of 4-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide and its hydrochloride
  • Step 1 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)piperidine- Synthesis of 4-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)propionamide and its hydrochloride
  • Step 1 Synthesis of tert-butyl (1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)piperidin-4-yl)carbamate
  • Step 2 (1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)piperidine-4- Synthesis of tert-butyl carbamate
  • Step 1 (1-(3-cyano-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborol-2-yl)pyrazolo[1 Synthesis of tert-butyl ,5-a]pyridin-4-yl)piperidin-4-yl)carbamate
  • Step 2 Synthesis of tert-butyl (1-(3-cyano-6-hydroxypyrazolo[1,5-a]pyridin-4-yl)piperidin-4-yl)carbamate
  • Step 3 (1-(3-cyano-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridin-4-yl)piperidin-4-yl) Synthesis of tert-butyl carbamate
  • Step 4 4-(4-Aminopiperidin-1-yl)-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridine-3-carbonitrile hydrochloride salt synthesis
  • Step 5 N-(1-(3-cyano-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridin-4-yl)piperidine-4- Synthesis of carboxylate)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide formate
  • Step 1 Synthesis of 1-amino-3-bromo-5-fluoropyridine-1-onium 2,4,6-trimethylbenzenesulfonate
  • Step 3 tert-butyl (1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)-4-methylpiperidin-4-yl)carbamate synthesis
  • Step 4 (1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)-4-methyl Synthesis of tert-butyl piperidin-4-yl)carbamate
  • Step 5 4-(4-Amino-4-methylpiperidin-1-yl)-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridine Synthesis of -3-carbonitrile hydrochloride
  • Step 6 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)-4- Synthesis of Methylpiperidin-4-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide
  • Step 1 Synthesis of tert-butyl (1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)azepan-4-yl)carbamate
  • Step 3 4-(4-Aminoazepan-1-yl)-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridine-3 - Synthesis of formonitrile hydrochloride
  • Step 4 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)azacycle Synthesis of heptan-4-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide
  • Step 1 Synthesis of tert-butyl (1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)pyrrolidin-3-yl)carbamate
  • Step 2 (1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)pyrrolidine-3- Synthesis of tert-butyl carbamate
  • Step 4 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)pyrrolidine- Synthesis of 3-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide
  • Step 1 Synthesis of tert-butyl (R)-(1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)pyrrolidin-3-yl)carbamate
  • Step 4 (R)-N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl ) Synthesis of pyrrolidin-3-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide
  • Step 1 (R)-N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl ) Preparation of pyrrolidin-3-yl)-2-(6-methoxypyridin-3-yl)acetamide hydrochloride
  • Step 1 (R)-N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl ) Preparation of pyrrolidin-3-yl)-2-(4-(methylsulfonyl)phenyl)acetamide
  • Step 1 Preparation of tert-butyl (S)-(1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)pyrrolidin-3-yl)carbamate
  • Step 2 (S)-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)pyrrole Preparation of tert-butyl alkane-3-yl)carbamate
  • Step 4 (S)-N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl ) pyrrolidin-3-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide preparation
  • Step 1 (S)-N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl ) Preparation of pyrrolidin-3-yl)-2-(4-methoxyphenyl)acetamide
  • Step 1 Synthesis of tert-butyl (1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)azetidin-3-yl)carbamate
  • Step 2 (1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)azetidine Synthesis of tert-butyl -3-yl)carbamate
  • Step 3 4-(3-Aminoazetidine-1-yl)-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridine-3 - Synthesis of formonitrile
  • Step 4 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)azacycle Synthesis of butan-3-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide
  • Step 1 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)azacycle Preparation of butan-3-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)propionamide
  • Step 1 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)azacycle Preparation of butan-3-yl)-2-(4-methoxyphenyl)acetamide
  • Step 1 N-(1-(3-Cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)azacycle Preparation of butan-3-yl)-2-(6-methoxypyridin-3-yl)acetamide
  • Step 4 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)azacycle Preparation of butan-3-yl)-2-(6-methoxypyridin-3-yl)propionamide carboxate
  • Step 2 Preparation of tert-butyl (1-(6-bromo-3-cyanopyrazolo[1,5-a]pyridin-4-yl)-1H-pyrazol-4-yl)carbamate
  • Step 3 (1-(3-cyano-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborol-2-yl)pyrazolo[1 Preparation of tert-butyl ,5-a]pyridin-4-yl)-1H-pyrazol-4-yl)carbamate
  • Step 4 Preparation of tert-butyl (1-(3-cyano-6-hydroxypyrazolo[1,5-a]pyridin-4-yl)-1H-pyrazol-4-yl)carbamate
  • Step 5 (1-(3-cyano-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridin-4-yl)-1H-pyrazol-4 -Preparation of tert-butyl carbamate
  • Step 6 4-(4-Amino-1H-pyrazol-1-yl)-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridine-3-methyl
  • Step 7 N-(1-(3-cyano-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridin-4-yl)-1H-pyrazole Preparation of -4-yl)-2-(6-methoxypyridin-3-yl)acetamide
  • Example 21 N-(1-(3-Cyano-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridin-4-yl)-1H-pyridine Azol-4 -yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide (compound 21) and its hydrochloride (compound 21-1) synthesis
  • Step 1 N-(1-(3-cyano-6-(2-hydroxy-2-methylpropoxy)pyrazolo[1,5-a]pyridin-4-yl)-1H-pyrazole Preparation of -4-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide hydrochloride
  • Step 1 (1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)-1H-pyrazole Preparation of tert-butyl-4-yl)carbamate
  • Step 2 4-(4-Amino-1H-pyrazol-1-yl)-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridine-3 - Preparation of formonitrile hydrochloride
  • Step 3 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)-1H- Preparation of pyrazol-4-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)acetamide
  • Step 1 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)-1H- Preparation of Pyrazol-4-yl)-2-(6-methoxypyridin-3-yl)acetamide
  • Example 24 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)-1H -Pyrazol-4 -yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)propanamide (Compound 24) and its hydrochloride (Compound 24-1 )Synthesis
  • Step 1 N-(1-(3-cyano-6-(1-methyl-1H-pyrazol-4-yl)pyrazolo[1,5-a]pyridin-4-yl)-1H- Preparation of pyrazol-4-yl)-2-(6-(4-fluoro-1H-pyrazol-1-yl)pyridin-3-yl)propionamide hydrochloride
  • Kinase RET inhibition assays were performed using methods similar to those described in the literature (Vivek Subbiah, J.F.G., Precision Targeted Therapy with BLU-667 for RET-Driven Cancers. Cancer Discov, 2018.8(7):836-849).
  • the inhibitory effect of the compound on the kinase RET was detected by the Caliper Mobility Shift Assay method.
  • the final concentration of the compound tested was 1000nM, 3-fold serial dilution, a total of 10 concentrations, and repeated well detection.
  • the compounds to be tested were dissolved in 100% DMSO (Sigma, D8418-1L, SHBG3288V) to prepare a 10 mM stock solution, which was stored in a nitrogen cabinet away from light.
  • DMSO 100% DMSO
  • 1 ⁇ Kinase buffer 1 ⁇ Kinase buffer
  • the initial test concentration of the compound to be tested is 1000nM
  • a 384-well plate 384-well plate (Corning, 3573, 12619003) 3-fold serial dilution with DMSO, a total of 10 concentrations, repeated well test
  • the final concentrations are 1000, 333, 111, 37, 12.3, 4.12, 1.37, 0.457, 0.152, 0.0508nM, respectively, and then use Echo550 (Labcyte, model: Echo 550) to transfer 250nl to the 384 reaction plate for use.
  • Negative control wells and 250nl of 100% DMSO were added to the positive control wells.
  • a kinase solution of RET (Carna, Cat. No. 08-159, Lot No. 13CBS-0134E) was prepared at 2.5x final concentration (1 nM final concentration) in 1 ⁇ Kinase buffer. Add 10 ⁇ L of 2.5 times the final concentration of kinase solution to compound wells and positive control wells respectively; add 10 ⁇ L of 1 ⁇ Kinase buffer to negative control wells.
  • the reaction plate was centrifuged at 1000 rpm (Eppendorf, model: 5430) for 30 seconds, the reaction plate was shaken and mixed, and incubated at room temperature for 10 minutes.
  • Conversion %_sample is the conversion rate reading of the sample
  • Conversion %_min is the mean value of the negative control wells, representing the conversion rate reading of the wells without enzymatic activity
  • Conversion %_max is the average value of the positive control wells, representing no Conversion readings for wells inhibited by compounds.
  • the kit uses luciferase as the detection substance. Luciferase needs the participation of ATP in the process of luminescence.
  • CellTiter-Glo TM reagent is added to the cell culture medium to measure the luminescence value. The light signal is proportional to the amount of ATP in the system. ATP is positively correlated with the number of viable cells, thereby determining the proliferative activity of cells.
  • DMSO dilute it 100 times with PBS to prepare a solution with a final concentration of 10 times, the highest concentration is 100 ⁇ M, and add 10 ⁇ L of the test compound to each well of the 96-well plate seeded with cells.
  • the solution ie, diluted 10-fold, reached a final concentration of 10 ⁇ M.
  • the final concentration of the compound to be tested starts from 10 ⁇ M, and the serial dilution is 3-fold, with a total of 9 concentrations, each with 3 replicate wells.
  • the 96-well plate to which the compounds to be tested and cells have been added was incubated at 37° C., 5% CO 2 , and 95% humidity for an additional 72 hours, followed by CellTiter-Glo analysis.
  • Luminescent Cell Viability Assay Promega, G7572
  • equilibrate the cell plate to room temperature for 30 minutes.
  • An equal volume of CellTiter-Glo solution was added to each well and the cells were lysed by shaking on an orbital shaker for 5 minutes.
  • the cell plate was placed at room temperature for 20 minutes to stabilize the luminescence signal, and the luminescence value was read with a SpectraMax multi-label microplate reader (MD, M3).
  • Liver microsomes (protein concentration of 0.56 mg/mL) were added to 1 ⁇ M compound working solution (diluted to 100 ⁇ M from 10 mM DMSO stock solution with 100% acetonitrile, organic phase content: 99% ACN, 1% DMSO), pre-incubated at 37°C for 10 min Afterwards, a cofactor (NADPH) (prepared from magnesium chloride solution) was added to initiate the reaction. After incubating for an appropriate time (eg 5, 10, 20, 30 and 60 minutes), take a sample and add an appropriate stop solution (ice acetonitrile containing 200ng/mL tolbutamide and 200ng/mL labetalol (ie acetonitrile at 4°C) ) to stop the reaction.
  • NADPH cofactor

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Abstract

一种吡唑并吡啶类化合物或其盐及其制备方法和用途，尤其涉及式(I)化合物或者其立体异构体、互变异构体、稳定的同位素衍生物、药学上可接受的盐或溶剂合物，及其制备方法和含有它们的药物组合物，以及所述化合物在制备治疗或预防与RET有关的疾病的药物中的用途。

Description

吡唑并吡啶类化合物或其盐及其制备方法和用途 技术领域
本发明属于化学医药技术领域，具体涉及具有RET抑制活性的吡唑并吡啶类化合物及其制备方法和含有所述化合物的药物组合物，还涉及该吡唑并吡啶类化合物在制备预防或治疗与RET有关的疾病的药物中的用途。
背景技术
RET(REarranged during Transfection)蛋白是一种受体酪氨酸激酶(RTK)，同时也是一种跨膜的糖蛋白，由位于10号染色体上的原癌基因RET表达，在胚胎阶段的肾脏和肠神经系统的发育中起着重要作用，另外在多种组织内也很关键，如神经元、神经内分泌、造血组织和男性生殖细胞等。和其他的受体酪氨酸激酶不同，RET并不是直接结合到配体分子：如神经导向素(artemin)、胶质细胞源性神经营养因子(GDNF)、神经生长因子(NGF)，这些都是属于GNDF家族配体(GFLs)。这些配体通常结合到GDNF家族受体α(GFRα)，形成的GFLs-GFRα复合物介导了RET蛋白的自二聚化，引起胞内结构域上酪氨酸的反式自磷酸化反应，招募相关接头蛋白，激活细胞增殖等信号传导的级联反应，从而导致细胞的过度增殖和癌症的产生，相关的信号通路包括MAPK、PI3K、JAK-STAT、PKA、PKC等等。
RET的致癌激活机制主要有两个：一是染色体的重排产生的新的融合蛋白，通常是RET的激酶结构域和包含自二聚化结构域的蛋白融合；二是RET突变直接或间接地激活了RET激酶活性。这些体细胞或生殖细胞水平地改变涉及多种癌症的发病机制。10％-20％的乳头状甲状腺癌(PTC)患者存在RET染色体重排；而在髓样性甲状腺髓样癌(MTC)中发现有60％存在RET点突变；在所有的非小细胞肺癌(NSCLC)患者中，大概有1-2％的具有RET融合蛋白，其中KIF5B最为常见。
目前主要采取的具有RET抑制活性的多重激酶抑制剂来治疗RET融合的癌症病人。但是在此条件下，由于脱靶效应及药物毒性，药物的剂量不足以达到足够抑制RET的表达水平。此外，在治疗癌症的过程中，癌细胞会通过变异产生耐药性。而一旦耐药性产生，病人的治疗选择将会变得非常有限。因此选择性地抑制异常的RET表达，RET融合及突变的药物非常急需。
目前上市或临床在研的选择性地针对RET靶点设计的药物，在非小细胞肺癌和甲状腺癌的临床实验上表现出了好的疗效及安全性。
目前，仍然非常需要发现和开发新的RET抑制剂化合物以用于预防和/或治疗与RET有关的疾病。这样的化合物除了应具有令人满意的RET抑制活性外，还期望基于结构的优化而具备良好的、甚至改进的成药性，以便为相关疾病患者提供更多用药选择的同时还能提供更 好的治疗效果。
发明简述
本发明涉及可用于预防或治疗与RET有关的疾病的化合物。特别地，已经鉴定，本发明的化合物显示令人满意的RET抑制活性。因此，本发明化合物不仅可实现用于预防或治疗与RET有关的疾病的目的，而且所制备的药物有望具有改善的吸收、在同等剂量下疗效提高、或以更低的剂量提供相同疗效和/或降低可能的副作用。由此，本发明还提供了本发明化合物在制备用于预防或治疗与RET有关的疾病的药物中的用途、包含所述化合物的药物组合物和通过施用所述化合物预防和/或治疗与RET有关的疾病的方法。
因此，在本发明的一方面，提供式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物：
Figure PCTCN2021125738-appb-000001
其中：
R 1选自氢、卤素、氰基、硝基和任选被卤素或氰基取代的C 1-C 6烷基；
R 2选自氢、C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基、3-8元杂环烯基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其中所述芳基、杂芳基、环烷基、环烯基、杂环烷基、杂环烯基、烷基、烷氧基和烷硫基任选被独立地选自以下的1、2或3个基团取代：卤素、氰基、硝基、羟基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
R 3选自氢、卤素、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
R 4选自氢、C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基、3-8元杂环烯基、-SO 2-C 1-C 6烷基、-SO-C 1-C 6烷基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其中所述芳基、杂芳基、环烷基、环烯基、杂环烷基、杂环烯基、烷基、烷氧基和烷硫基任选被独立地选自以下的1、2或3个基团取代：卤素、氰基、硝基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
R 5选自卤素、氰基、硝基、羟基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
环A选自C 6-C 10亚芳基、5-9元亚杂芳基、C 3-C 8亚环烷基、C 3-C 8亚环烯基、3-8元亚杂环烷基和3-8元亚杂环烯基环；
环B选自C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基和3-8元杂环烯基环；
m是0、1、2或3；且
n是0、1、2或3。
在本发明的另一方面，提供了具有RET抑制活性、用作药物、尤其是用作RET抑制剂的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物。
在本发明的另一方面，提供了用于治疗或预防、尤其是用于治疗与RET有关的疾病的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物。
在本发明的另一方面，提供了包含本发明化合物和药学上可接受的赋形剂的药物组合物。在一个具体的方面，提供了所述本发明的药物组合物，用于预防或治疗与RET有关的疾病。在一个具体的方面，药物组合物可以另外包含适合与本发明化合物组合使用的另外的治疗活性成分。
在本发明的另一方面，提供了包含本发明化合物和另外的活性剂的药物组合(或药物组合产品)。
在本发明的另一方面，提供了用于在个体，例如哺乳动物、特别是人中预防或治疗与RET有关的疾病的方法，该方法包括施用有效量的本文所述的本发明化合物或包含其的药物组合物。
在本发明的另一方面，提供了上述本发明的化合物或药物组合物用于预防或治疗与RET有关的疾病的用途。
在本发明的另一方面，提供了上述本发明的化合物或药物组合物在制备用于预防或治疗与RET有关的疾病的药物中的用途。
优选地，本发明所述的与RET有关的疾病选自肿瘤或肠易激综合症(IBS)，肿瘤包括但不限于非小细胞肺癌、小细胞肺癌、乳头状甲状腺癌、甲状腺髓质癌、分化型甲状腺癌、复发性甲状腺癌、顽固性分化型甲状腺癌、多内分泌肿瘤2A或2B、嗜铬细胞瘤、甲状旁腺增生、乳腺癌、结肠直肠癌、乳头状肾细胞癌、胃肠黏膜神经节瘤、胰管腺癌、多发性内分泌瘤、睾丸癌、慢性单核细胞性白血病、唾腺癌、卵巢癌、子宫颈癌等。
在另外的方面，提供了用于合成本发明化合物的方法，其中代表性的合成方案和途径在下文描述。
通过阅读随后的详细描述，本发明的其它目的和优点对于本领域技术人员将变得显而易见。
对于上述用作药物的本发明化合物、本发明的预防或治疗方法、药物组合物、药物组合或用途而言，优选本文所定义的式(I)化合物的各个优选实施方式，更优选文中所列的具体化合物。
定义
除非在下文中另有定义，本文中所用的所有技术术语和科学术语的含义意图与本领域技术人员通常所理解的相同。提及本文中使用的技术意图指在本领域中通常所理解的技术，包括那些对本领域技术人员显而易见的技术的变化或等效技术的替换。虽然相信以下术语对于本领域技术人员很好理解，但仍然阐述以下定义以更好地解释本发明。
本文所用的术语“卤代”或“卤素”意指氟(F)、氯(Cl)、溴(Br)及碘(I)。优选的卤代为氟或氯。本文所用的术语“被卤素取代的”基团旨在包括单卤代或多卤代基团，其中一个或多个(例如2、3、4、5或6个)相同或不同的卤素取代基团中的一个或多个(例如2、3、4、5或6个)氢。
本文所用的术语“氰基”意指基团-CN。
本文所用的术语“硝基”意指基团-NO 2。
如本文中所使用的术语“羟基”指-OH。
如本文中所使用的术语“烷基”指由碳原子和氢原子组成的直链或支链的饱和烃基团。具体地，烷基具有1-10个，例如1至6个、1至5个、1至4个、1至3个或1至2个碳原子。例如，如本文中所使用，术语“C 1-C 6烷基”指具有1至6个碳原子的直链或支链的饱和烃基团，其实例例如甲基、乙基、丙基(包括正丙基和异丙基)、丁基(包括正丁基、异丁基、仲丁基或叔丁基)、戊基(包括正戊基、异戊基、新戊基)、正己基、2-甲基戊基等。特定的烷基具有1至3个碳原子。
本文所用的术语“烷氧基”意指基团-O-烷基，其中烷基具有本文所述的含义。具体地，该术语包括基团-O-C 1-6烷基，更具体的-O-C 1-3烷基。烷氧基的代表性实例包括但不限于甲氧基、乙氧基、丙氧基(包括正丙氧基、异丙氧基)、丁氧基(包括正丁氧基、异丁氧基、叔丁氧基)、戊氧基(包括正戊氧基、异戊氧基、新戊氧基)、己氧基(包括正己氧基、异己氧基)等。特定的烷氧基具有1至3个碳原子。
如本文中所使用的术语“烷硫基”指-S-烷基，其中所述烷基如以上对于“烷基”所定义。具体地，该术语包括基团-S-C 1-6烷基，更具体的-S-C 1-3烷基。烷硫基的代表性实例包括但不限于甲硫基、乙硫基、丙硫基(包括正丙硫基、异丙硫基)、丁硫基(包括正丁硫基、异丁硫基、叔丁硫基)、戊硫基(包括正戊硫基、异戊硫基、新戊硫基)、己硫基(包括正己硫基、异己硫基)等。特定的烷硫基具有1至3个碳原子。如本文中所使用的术语“卤素取代的C 1-C 6烷基”指上文所述的C 1-C 6烷基，其中一个或多个(例如1、2、3、4或5个)氢原子被卤素代替。本领域 技术人员应当理解，当卤素取代基多于一个时，卤素可以相同也可以不同，并且可以位于相同或不同的C原子上。“卤素取代的C 1-C 6烷基”的实例有例如-CH 2F、-CHF 2、-CF 3、-CCl 3、-C 2F 5、-C 2Cl 5、-CH 2CF 3、-CH 2Cl、-CH 2CH 2CF 3或-CF(CF 3) 2等。
如本文中所使用的术语“卤素取代的C 1-C 6烷氧基”指上文所述的C 1-C 6烷氧基，其中一个或多个(例如1、2、3、4或5个)氢原子被卤素代替。本领域技术人员应当理解，当卤素取代基多于一个时，卤素可以相同也可以不同，并且可以位于相同或不同的C原子上。“卤素取代的C 1-C 6烷氧基”的实例有例如-OCH 2F、-OCHF 2、-OCF 3、-OCCl 3、-OC 2F 5、-OC 2Cl 5、-OCH 2CF 3、-OCH 2Cl或-OCH 2CH 2CF 3等。
如本文中所使用的术语“环烷基”指具有指定环原子数的单环、稠合多环、桥接多环或螺环非芳族饱和单价烃环结构。环烷基可具有3至12个碳原子(即C 3-C 12环烷基)，例如3至10个，3至8个，3至7个，3至6个，5至6个碳原子。适合的环烷基的实例包括但不限于单环结构，如环丙基、环丁基、环戊基、环己基、环庚基或环辛基；或多环(例如双环)结构，包括螺环、稠合或桥连系统，诸如双环[1.1.1]戊基、双环[2.2.1]庚基、螺[3.4]辛烷基、双环[3.1.1]己烷基、双环[3.1.1]庚基或双环[3.2.1]辛基等)。
如本文中所使用的术语“亚环烷基”指如上所定义的环烷基，但其为二价基团，该两个价键不在同一环原子上。亚环烷基可具有3至12个碳原子(即C 3-C 12亚环烷基)，例如3至10个，3至8个，3至7个，3至6个，5至6个碳原子。适合的亚环烷基的实例包括但不限于单环结构，如亚环丙基、亚环丁基、亚环戊基(例如环戊-1,2-二基、环戊-1,3-二基)、亚环己基(例如环己-1,2-二基、环己-1,3-二基、环己-1,4-二基)、亚环庚基或亚环辛基；或多环(例如双环)结构，包括螺环、稠合或桥连系统，诸如双环[1.1.1]亚戊基、双环[2.2.1]亚庚基、螺[3.4]亚辛烷基、双环[3.1.1]亚己烷基、双环[3.1.1]亚庚基或双环[3.2.1]亚辛基等)。
本文所用的术语“环烯基”意指具有指定环原子数的单环、稠合多环、桥接多环或螺环非芳族不饱和烃环结构，包含至少一个(例如1、2、或3个)碳碳双键。环烯基可具有3至12个碳原子(即C 3-C 12环烯基)，例如3至10个，3至8个，3至7个，3至6个，5至6个碳原子。适合的环烯基的实例包括但不限于单环结构，如环丙烯基、环丁烯基、环戊烯基、环戊二烯基、环己烯基、环己二烯基、环庚烯基、环庚二烯基、环庚三烯基或环辛烯基。
如本文中所使用的术语“亚环烯基”指如上所定义的环烯基，但其为二价基团，该两个价键不在同一环原子上。亚环烯基可具有3至12个碳原子(即C 3-C 12亚环烯基)，例如3至10个，3至8个，3至7个，3至6个，5至6个碳原子。适合的亚环烯基的实例包括但不限于单环结构，如亚环丙烯基、亚环丁烯基、亚环戊烯基、亚环戊二烯基、亚环己烯基、亚环己二烯基、亚环庚烯基、亚环庚二烯基、亚环庚三烯基或亚环辛烯基。
本文所用的术语“杂环烷基”意指包括一或多个(例如1、2、3或4个)独立地选自O、N及S的杂原子及指定环原子数的单环、稠合多环、螺环或桥接多环非芳族饱和环结构，或其N-氧化物，或其S-氧化物或S-二氧化物。杂环烷基可具有3至12个环成员(可称为3-12元杂环烷 基)，例如3至10个环成员，3至8个环成员，3至7个环成员，4至7个环成员，5至6个环成员。杂环烷基通常含有至多4个(例如1个、2个、3个或4个)杂原子。适合的杂环烷基的实例包括但不限于氮杂环丁烷基、氧杂环丁烷基、硫杂环丁基、吡咯烷基(例如1-吡咯烷基、2-吡咯烷基及3-吡咯烷基)、四氢呋喃基(例如1-四氢呋喃基、2-四氢呋喃基及3-四氢呋喃基)、四氢噻吩基(例如1-四氢噻吩基、2-四氢噻吩基及3-四氢噻吩基)、哌啶基(例如1-哌啶基、2-哌啶基、3-哌啶基及4-哌啶基)、四氢吡喃基(例如4-四氢吡喃基)、四氢噻喃基(例如4-四氢噻喃基)、吗啉基(例如吗啉代)、硫吗啉基、二噁烷基、哌嗪基或氮杂环庚烷基、二氮杂环庚烷基例如1,4-二氮杂环庚基、3,6-二氮杂-双环[3.1.1]庚基或3-氮杂-双环[3.2.1]辛基。
本文所用的术语“亚杂环烷基”意指如上所定义的杂环烷基，但其为二价基团，该两个价键不在同一环原子上。亚杂环烷基可具有3至12个环成员(可称为3-12元亚杂环烷基)，例如3至10个环成员，3至8个环成员，3至7个环成员，4至7个环成员，5至6个环成员。亚杂环烷基通常含有至多4个(例如1个、2个、3个或4个)杂原子。适合的亚杂环烷基的实例包括但不限于氮杂亚环丁烷基、氧杂亚环丁烷基、硫杂亚环丁基、亚吡咯烷基(例如吡咯烷-1,2-二基、吡咯烷-1,3-二基、吡咯烷-2,3-二基)、亚四氢呋喃基(例如四氢呋喃-2,4-二基、四氢呋喃-2,3-二基及四氢呋喃-2,5-二基)、亚哌啶基(例如哌啶-1,2-二基、哌啶-1,3-二基、哌啶-1,4-二基、哌啶-2,4-二基)、亚四氢噻吩基、四氢吡喃基、四氢噻喃基、亚吗啉基、亚硫吗啉基、亚二噁烷基、亚哌嗪基或氮杂亚环庚烷基、二氮杂亚环庚烷基例如1,4-二氮杂亚环庚基、3,6-二氮杂-双环[3.1.1]亚庚基或3-氮杂-双环[3.2.1]亚辛基，优选
Figure PCTCN2021125738-appb-000002
Figure PCTCN2021125738-appb-000003
应当理解的是，具有不对称中心的结构涵盖其外消旋的和/或单一的对映异构形式，例如
Figure PCTCN2021125738-appb-000004
可代表
Figure PCTCN2021125738-appb-000005
和/或
Figure PCTCN2021125738-appb-000006
本文所用的术语“杂环烯基”意指包含至少一个(例如1、2或3个)双键的本文所定义的“杂环烷基”。适合的杂环烯基的实例包括但不限于：
Figure PCTCN2021125738-appb-000007
其中各W选自CH 2、NH、O及S；各Y选自NH、O、C(＝O)、SO 2及S；且各Z选自N及CH，条件是各个环中至少包含一个选自N、O或S的原子；例如吡咯啉基(例如1-吡咯啉基、2-吡咯烷基、3-吡咯啉基、4-吡咯啉基或5-吡咯啉基)、二氢呋喃基(例如1-二氢呋喃基、2-二氢呋喃基、 3-二氢呋喃基、4-二氢呋喃基或5-二氢呋喃基)、二氢噻吩基(例如1-二氢噻吩基、2-二氢噻吩基、3-二氢噻吩基或4-二氢噻吩基)、四氢吡啶基(例如1-、2-、3-、4-、5-或6-四氢吡啶基)、四氢吡喃基(例如4-四氢吡喃基)或四氢噻喃基(例如4-四氢噻喃基)。
本文所用的术语“亚杂环烯基”意指如上所定义的杂环烯基，但其为二价基团，该两个价键不在同一环原子上。适合的亚杂环烯基的实例包括但不限于：
Figure PCTCN2021125738-appb-000008
其中各W选自CH 2、NH、O及S；各Y选自NH、O、C(＝O)、SO 2及S；且各Z选自N及CH，条件是各个环中至少包含一个选自N、O或S的原子；例如亚吡咯啉基、亚二氢呋喃基、亚二氢噻吩基、亚四氢吡啶基、亚四氢吡喃基或亚四氢噻喃基，优选
Figure PCTCN2021125738-appb-000009
更优选
Figure PCTCN2021125738-appb-000010
本文所用的术语“芳基”意指通过自芳族环系统中的单个碳原子移除一个氢原子而衍生的单价芳族烃基。具体地，芳基系指具有指定环原子数的单环或稠合多环芳族环结构。具体地，该术语包括包含6至14个、例如6至10个、优选6个环成员的基团。特定的芳基包括苯基及萘基，最具体的芳基为苯基。
本文所用的术语“亚芳基”意指如上所定义的芳基，但其为二价基团，该两个价键不在同一环原子上。特定的亚芳基包括亚苯基，例如苯-1,2-二基、苯-1,3-二基或苯-1,4-二基。
本文所用的术语“杂芳基”意指包括一或多个(例如1、2、3或4个)独立地选自O、N及S的杂原子及指定环原子数的单环或稠合多环芳族环结构，或其N-氧化物，或其S-氧化物或S-二氧化物。具体地，该芳族环结构可具有5至9个环成员。杂芳基可为例如5-6元单环、或由稠合的两个5元环或稠合的5元环和4元环形成的稠合双环结构。杂芳基环通常将含有至多4个杂原子、更通常至多3个杂原子、更通常至多2个、例如单个独立地选自O、N及S的杂原子，其中N和S可以是氧化状态如N氧化物、S＝O或S(O) 2。在一个实施方案中，杂芳基环含有至少一个环氮原子、至少一个环硫原子或至少一个环氧原子。例如，杂芳基可以是包含1或2个独立地选自N、O或S的杂原子的5-6元杂芳基。适合的5元单环杂芳基的实例包括但不限于吡咯基、呋喃基、噻吩基、咪唑基、呋咱基、噁唑基、噁二唑基、噁三唑基、异噁唑基、噻唑基、异噻唑基、吡唑基、三唑基及四唑基；适合的6元单环杂芳基的实例包括但不限于吡啶基、吡嗪基、哒嗪基、嘧啶基及三嗪基。
本文所用的术语“亚杂芳基”意指如上所定义的杂芳基，但其为二价基团，该两个价键不在同一环原子上。亚杂芳基可以是包含1或2个独立地选自N、O或S的杂原子的5-6元亚杂芳基。适合的5元单环亚杂芳基的实例包括但不限于亚吡咯基、亚呋喃基、亚噻吩基、亚咪唑基、亚呋咱基、亚噁唑基、亚噁二唑基、亚噁三唑基、亚异噁唑基、亚噻唑基、亚异噻唑基、亚吡唑基、亚三唑基及亚四唑基；适合的6元单环杂芳基的实例包括但不限于亚吡啶基、亚吡嗪基、亚哒嗪基、亚嘧啶基及亚三嗪基，优选
Figure PCTCN2021125738-appb-000011
取代基被描述为“任选取代的”意指基团可以是未取代的或被一个或多个(例如0、1、2、3、4或5或更多个，或其中可衍生的任何范围)对该基团所列的取代基取代，其中所述取代基可以相同或不同。在一个实施方案中，任选取代的基团被1个取代基取代。在另一个实施方案中，任选取代的基团被2个取代基取代。在另一个实施方案中，任选取代的基团被3个取代基取代。在另一个实施方案中，任选取代的基团被4个取代基取代。
有机合成领域普通技术人员均理解，稳定的化学可行的杂环，无论芳族还是非芳族，其中最大杂原子数或所含杂原子的类型由环大小、不饱和度及杂原子的价数决定。一般而言，杂环可具有1至4个杂原子，前提是杂环或杂芳环在化学可行及稳定。
本文所用的术语“本发明的化合物”意欲涵盖如本文所定义的通式(I)的化合物或其任一优选或具体的实施方案(包括式(Ia)和式(Ib)的化合物)、它们的立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，以及前药。类似地，本文中对“中间体”的称谓，无论其本身是否被请求保护，若上下文允许，则均意欲涵盖其游离形式以及上述各衍生形式。
本文所用的术语“药学可接受的”意指由各个国家的相应机构批准的或可由其批准，或列于用于动物且更具体地人类的一般公认药典中，或当向动物例如人类适量施用时不会产生不利、过敏或其它不良反应的分子实体和组合物。
本文所用的术语“药学可接受的盐”意指药学上可接受且具有母体化合物所需药理学活性的本发明化合物的盐。具体地，此类盐无毒，可为无机酸加成盐或有机酸加成盐及碱加成盐。具体地，此类盐包括：(1)与无机酸形成的酸加成盐，该无机酸如盐酸、氢溴酸、硫酸、硝酸、磷酸等；或与有机酸形成的酸加成盐，该有机酸如乙酸、丙酸、己酸、乙醇酸、丙酮酸、乳酸、丙二酸、丁二酸、苹果酸、马来酸、富马酸、酒石酸、柠檬酸、苯甲酸、肉桂酸、扁桃酸、甲磺酸、乙磺酸、苯磺酸、2-萘磺酸、4-甲苯磺酸、樟脑磺酸、葡糖庚酸、3-苯基丙酸、三甲基乙酸、叔丁基乙酸、月桂基硫酸、葡萄糖酸、谷氨酸、羟基萘甲酸、水杨酸、硬脂酸、黏康酸等；或(2)当存在于母体化合物中的酸性质子经金属离子如碱金属离子、碱土金属离子或铝离子置换时、或与有机碱如乙醇胺、二乙醇胺、三乙醇胺、N-甲基葡糖胺等配位 时形成的盐。本领域技术人员将知道制备药用盐的一般原理和技术，例如Berge等，Pharm ScL，66，1-19.(1977)中所述的那些。
本文所用的术语“前药”意指具有可裂解基团且通过溶剂分解或在生理条件下变成在体内具有药学活性的本发明化合物的化合物，包括本发明化合物的衍生物。前药包括本领域熟知的酸衍生物，如通过母体酸与适合醇反应制备的酯，或通过母体酸化合物与取代或未取代的胺反应制备的酰胺，或酸酐或混合酸酐。衍生自本发明化合物侧接的酸基的简单脂族或芳族酯、酰胺及酸酐为尤其适用的前药。特定的此类前药为本发明化合物的C 1-8烷基、C 2-8烯基、任选被取代的C 6-10芳基及(C 6-10芳基)-(C 1-4烷基)酯。
本发明还包括所有药学上可接受的同位素化合物，其与本发明的化合物相同，除了一个或多个原子被具有相同原子序数但原子质量或质量数不同于在自然界中占优势的原子质量或质量数的原子替代。适合包含入本发明化合物中的同位素的实例包括(但不限于)氢的同位素(例如2H、3H)；碳的同位素(例如11C、13C及14C)；氯的同位素(例如36Cl)；氟的同位素(例如18F)；碘的同位素(例如123I及125I)；氮的同位素(例如13N及15N)；氧的同位素(例如15O、17O及18O)；磷的同位素(例如32P)；及硫的同位素(例如35S)。
如本文中所使用的术语“立体异构体”表示由于至少一个不对称中心形成的异构体。在具有一个或多个(例如1个、2个、3个或4个)不对称中心的化合物中，其可产生外消旋混合物、单一对映异构体、非对映异构体混合物和单独的非对映异构体。特定个别分子也可以以几何异构体(顺式/反式)存在。类似地，本发明的化合物可以以两种或更多种处于快速平衡的不同结构形式的混合物(通常称作互变异构体)存在。互变异构体的代表性实例包括酮-烯醇互变异构体、苯酚-酮互变异构体、亚硝基-肟互变异构体、亚胺-烯胺互变异构体等。例如，亚硝基-肟在溶液中可以下列互变异构形式平衡存在：
Figure PCTCN2021125738-appb-000012
需要理解的是，本申请的范围涵盖所有这样的以任意比例(例如60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％)的异构体或其混合物。
除非另外指明，否则本发明的化合物意欲可以立体异构体(其包括顺式及反式异构体、光学异构体(例如R及S对映异构体)、非对映异构体、几何异构体、旋转异构体、构象异构体、阻转异构体及其混合物)的形式存在。本发明的化合物可表现一种以上类型的异构现象，且由其混合物(例如外消旋混合物及非对映异构体对)组成。
本文所用的术语“溶剂合物”是指包含化学计量的或非化学计量的溶剂的溶剂加成形式，包括例如与水的溶剂合物，例如水合物，或与有机溶剂的溶剂合物，例如甲醇、乙醇或乙腈，即分别作为甲醇化物、乙醇化物或乙腈化物；或为任何多晶型物的形式。应当理解的是，本发明化合物的这类溶剂合物还包括本发明化合物的药学可接受盐的溶剂合物。
本文所使用的术语“预防“意指给怀疑患上或易感于如本文所定义的与RET有关的疾病、尤其是炎症或自身免疫疾病的个体个体、例如哺乳动物、例如人施用一种或多种本发明的化合物，使得罹患所定义疾病的风险降低。术语“预防”包含在诊断或确定任何临床和/或病理症状以前使用本发明的化合物。
本文所用的术语“治疗”是指给患有所述疾病、或者具有所述疾病的症状的受试者、例如哺乳动物、例如人施用一种或多种本文所述的本发明化合物，用以治愈、缓解、减轻或影响所述疾病或所述疾病的症状。在本发明具体的实施方案中，所述疾病是本文所定义的与RET有关的疾病、尤其是炎症或自身免疫疾病。
本文所用的术语“与RET有关的疾病”意指RET对所述疾病的发生和发展起到促进作用，或抑制RET将降低疾病的发生率、减少或消除疾病病状的疾病。对于本发明而言，“与RET有关的疾病”选自肿瘤或肠易激综合症(IBS)，肿瘤包括但不限于非小细胞肺癌、小细胞肺癌、乳头状甲状腺癌、甲状腺髓质癌、分化型甲状腺癌、复发性甲状腺癌、顽固性分化型甲状腺癌、多内分泌肿瘤2A或2B、嗜铬细胞瘤、甲状旁腺增生、乳腺癌、结肠直肠癌、乳头状肾细胞癌、胃肠黏膜神经节瘤、胰管腺癌、多发性内分泌瘤、睾丸癌、慢性单核细胞性白血病、唾腺癌、卵巢癌、子宫颈癌等。
如本文中所使用的术语“癌症”或“肿瘤”是指赘生性细胞生长和增殖，无论是恶性的还是良性的，和所有的癌前期细胞和癌细胞和组织。对本发明的化合物、方法、药物组合物、药物组合及用途而言，所述癌症或肿瘤包括但不限于结肠癌、胰腺癌、乳腺癌、卵巢癌、前列腺癌、鳞状细胞癌、基底层细胞癌、腺癌、汗腺癌、皮脂腺癌、肺癌、白血病、膀胱癌、胃癌、子宫颈癌、睾丸癌、皮肤癌、直肠癌、甲状腺癌、肾癌、子宫癌、天疱疮癌、肝癌、听神经瘤、少突神经胶质瘤、脑(脊)膜瘤、成神经细胞瘤、眼癌。
本文所使用的术语“治疗有效量“意指当向个体施用以治疗疾病时足以减轻或完全缓解病症的症状或其它有害作用；逆转、完全停止或减缓病症的进展；或降低病症恶化的风险的量，“有效量”可视化合物、疾病及其严重程度及待治疗的个体的年龄、体重等而变化。
如本文所使用的术语“个体”包括人或非人动物。示例性人个体包括患有疾病(例如本文所述的疾病)的人个体(称为患者)或正常个体。本发明中“非人动物”包括所有脊椎动物，例如非哺乳动物(例如鸟类、两栖动物、爬行动物)和哺乳动物，例如非人灵长类、家畜和/或驯化动物(例如绵羊、犬、猫、奶牛、猪等)。
本发明所述的“药物组合物”，指包含一种或多种式(I)化合物或者其立体异构体、互变异构体、稳定的同位素衍生物、药学上可接受的盐或溶剂合物和在本领域中通常接受的用于将生物活性化合物输送至有机体(例如人)的载体的组合物。
本文所用的术语“药物组合”是指本发明化合物可与其它活性剂组合用于实现本发明的目的。所述其他活性剂可以是一种或多种另外的本发明化合物，或可以是与本发明化合物相容即不会相互不利影响、或具有互补活性的第二种或另外的(例如第三种)化合物。这类活性 剂以达到预期目的的有效量适宜地组合存在。所述其他活性剂可以与本发明化合物在单一药物组合物中共同施用，或与本发明化合物处于不同的离散单元中分别施用，当分别施用时可以同时或相继进行。所述相继施用在时间上可以是接近或隔远的。
本文所用的术语“药学上可接受的赋形剂或载体”是指一种或多种相容性固体或液体填料或凝胶物质，其药理学上无活性，与组合物中的其它成分相容，并且应是对温血动物如人给药可接受的，用作本发明化合物在施用形式中的载体或介质，其实例包括但不限于纤维素及其衍生物(如羧甲基纤维素钠、醋酸纤维素等)、明胶、滑石、固体润滑剂(如硬脂酸镁)、硫酸钙、植物油、多元醇(如丙二醇、甘油、甘露醇、山梨醇等)、乳化剂(如吐温类)、润湿剂(如十二烷基硫酸钠)、着色剂、调味剂、稳定剂、抗氧化剂、防腐剂等。
应理解，当本文描述本发明化合物、包含其的药物组合物、药物组合以及相关的用途和方法时所涉及的剂量，是基于游离形式的重量，而非基于其任何盐、水合物或溶剂化物等，除非说明书中另外定义。
本发明化合物
本申请通篇使用的术语“发明的化合物”和“本发明的化合物”等，除非另外指出，涵盖本文各个实施方案及其具体或优选实施方式中定义的式(I)化合物、它们的立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，以及前药。所述立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物以及前药如上文定义部分所描述。优选地，本发明化合物为式(I)化合物的游离形式或其药学上可接受的盐或溶剂合物；最优选为式(I)化合物的游离形式或其药学上可接受的盐。
本发明的某些化合物可以以多晶型或无定形形式存在，它们也落入本发明的范围内。当为固体结晶形式时，式(I)化合物可以是与另一种化学实体的共晶体形式，并且本说明书包括所有这些共晶体。
在存在手性中心时，本发明的化合物可以以单独的对映异构体或对映体混合物形式存在。根据一个实施方案，提供了式(I)化合物或其药学上可接受的盐，其是对映体过量(％ee)＞95、＞98％或＞99％的单一对映体。优选地，单一对映异构体以＞99％的对映异构体过量(％ee)存在。
具体地，一方面，本发明提供式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物：
Figure PCTCN2021125738-appb-000013
其中：
R 1选自氢、卤素、氰基、硝基和任选被卤素或氰基取代的C 1-C 6烷基；
R 2选自氢、C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基、3-8元杂环烯基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其中所述芳基、杂芳基、环烷基、环烯基、杂环烷基、杂环烯基、烷基、烷氧基和烷硫基任选被独立地选自以下的1、2或3个基团取代：卤素、氰基、硝基、羟基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
R 3选自氢、卤素、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
R 4选自氢、C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基、3-8元杂环烯基、-SO 2-C 1-C 6烷基、-SO-C 1-C 6烷基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其中所述芳基、杂芳基、环烷基、环烯基、杂环烷基、杂环烯基、烷基、烷氧基和烷硫基任选被独立地选自以下的1、2或3个基团取代：卤素、氰基、硝基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
R 5选自卤素、氰基、硝基、羟基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
环A选自C 6-C 10亚芳基、5-9元亚杂芳基、C 3-C 8亚环烷基、C 3-C 8亚环烯基、3-8元亚杂环烷基和3-8元亚杂环烯基环；
环B选自C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基和3-8元杂环烯基环；
m是0、1、2或3；且
n是0、1、2或3。
在一种式(I)化合物的实施方式中，R 1是卤素、氰基或硝基。
在一种式(I)化合物的实施方式中，R 1是氰基。
在一种式(I)化合物的实施方式中，R 2选自包含1、2或3个独立地选自N、O或S的杂原子的5-6元杂芳基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其任选被独立地选自以下的1、2或3个基团取代：卤素、羟基和C 1-C 6烷基；
在一种式(I)化合物的实施方式中，R 2选自包含1、2或3个独立地选自N、O或S的杂原子的5元杂芳基、C 1-C 6烷氧基和C 1-C 6烷硫基，其任选被独立地选自以下的1个基团取代：羟基和C 1-C 6烷基；
在一种式(I)化合物的实施方式中，R 2选自任选被C 1-C 6烷基取代的包含1、2或3个独立地选自N、O或S的杂原子的5元杂芳基和任选被羟基取代的C 1-C 6烷氧基；
在一种式(I)化合物的实施方式中，R 2中的5-6元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基、吡唑基、吡啶基、吡嗪基、哒嗪基或嘧啶基；
优选地，R 2中的5元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基或吡唑基。
在一种式(I)化合物的实施方式中，R 2是甲基取代的吡唑基或羟基取代的C 1-C 6烷氧基；
在一种式(I)化合物的实施方式中，R 2是
Figure PCTCN2021125738-appb-000014
在一种式(I)化合物的实施方式中，R 3选自氢和C 1-C 6烷基。
在一种式(I)化合物的实施方式中，R 3选自氢和C 1-C 3烷基。
在一种式(I)化合物的实施方式中，R 3是氢或甲基。
在一种式(I)化合物的实施方式中，R 3是氢。
在一种式(I)化合物的实施方式中，R 4选自苯基、包含1、2或3个独立地选自N、O或S的杂原子的5-6元杂芳基、-SO 2-C 1-C 6烷基、-SO-C 1-C 6烷基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其任选被1、2或3个卤素基团取代；优选地，所述5-6元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基、吡唑基、吡啶基、吡嗪基、哒嗪基和嘧啶基；
在一种式(I)化合物的实施方式中，R 4选自包含1、2或3个独立地选自N、O或S的杂原子的5元杂芳基、-SO 2-C 1-C 6烷基、C 1-C 6烷基和C 1-C 6烷氧基，其任选被1、2或3个卤素基团取代；优选地，所述5元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基和吡唑基。
在一种式(I)化合物的实施方式中，R 4选自任选被卤素取代的吡唑基、-SO 2-C 1-C 6烷基、C 1-C 6烷基和C 1-C 6烷氧基。
在一种式(I)化合物的实施方式中，R 4是
Figure PCTCN2021125738-appb-000015
-SO 2-CH 3或甲氧基。
在一种式(I)化合物的实施方式中，R 5选自卤素、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
在一种式(I)化合物的实施方式中，R 5选自C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基。
在一种式(I)化合物的实施方式中，R 5选自C 1-C 3烷基；
在一种式(I)化合物的实施方式中，R 5是甲基；
在一种式(I)化合物的实施方式中，m是0或1。
在一种式(I)化合物的实施方式中，n是0或1。
在一种式(I)化合物的实施方式中，m是1。
在一种式(I)化合物的实施方式中，n是0。
在一种式(I)化合物的实施方式中，环A选自亚苯基、包含1、2或3个独立地选自N、O或S的杂原子的5-6元亚杂芳基、C 3-C 8亚环烷基、C 3-C 8亚环烯基、包含1、2或3个独立地选自N、O或S的杂原子的3-8元亚杂环烷基和包含1、2或3个独立地选自N、O或S的杂原子的3-8元亚杂环烯基；
在一种式(I)化合物的实施方式中，环A选自亚苯基、包含1、2或3个独立地选自N、O或S的杂原子的5元亚杂芳基、C 3-C 6亚环烷基、C 3-C 6亚环烯基、包含1、2或3个独立地选自N、O或S的杂原子的3-6元亚杂环烷基和包含1、2或3个独立地选自N、O或S的杂原子的3-6元亚杂环烯基；
在一种式(I)化合物的实施方式中，环A选自亚苯基、亚吡咯基、亚呋喃基、亚噻吩基、亚咪唑基、亚呋咱基、亚噁唑基、亚噁二唑基、亚噁三唑基、亚异噁唑基、亚噻唑基、亚异噻唑基、亚吡唑基、亚噻二唑基、亚三唑基、亚四唑基、亚吡啶基、亚吡嗪基、亚哒嗪基、嘧啶基、亚三嗪基、亚氮杂环丁烷基、亚吡咯烷基、亚吡咯啉基、亚环丁基、亚环戊基、亚环己基、亚环庚基、亚哌嗪基、亚哌啶基和氮杂亚环庚基。
在一种式(I)化合物的实施方式中，环A选自亚吡唑基、亚氮杂环丁烷基、亚吡咯烷基、亚环己基、亚哌啶基和氮杂亚环庚基。
在一种式(I)化合物的实施方式中，环A选自
Figure PCTCN2021125738-appb-000016
在一种式(I)化合物的实施方式中，环B选自苯基、包含1、2或3个独立地选自N、O或S的杂原子的5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、包含1、2或3个独立地选自N、O或S的杂原子的3-8元杂环烷基和包含1、2或3个独立地选自N、O或S的杂原子的3-8元杂环烯基环；
在一种式(I)化合物的实施方式中，环B选自苯基和包含1、2或3个独立地选自N、O或S的杂原子的5-6元杂芳基；
在一种式(I)化合物的实施方式中，环B选自苯基、和包含1、2或3个独立地选自N、O或S的杂原子的6元杂芳基；
在一种式(I)化合物的实施方式中，环B选自苯基、吡咯基、呋喃基、噻吩基、咪唑基、呋咱基、噁唑基、噁二唑基、噁三唑基、异噁唑基、噻唑基、异噻唑基、吡唑基、噻二唑基、三唑基、四唑基、吡啶基、吡嗪基、哒嗪基、嘧啶基和三嗪基环。
在一种式(I)化合物的实施方式中，环B是苯基或吡啶基环。
在一种式(I)化合物的实施方式中，基团
Figure PCTCN2021125738-appb-000017
选自
Figure PCTCN2021125738-appb-000018
需要说明的是，本发明的式(I)化合物涵盖以上各个独立的实施方式或各个具体实施方式，还涵盖上述各个实施方式或具体实施方式的任何组合或亚组合构成的实施方式，也涵盖以上任何优选或例举的任何组合所构成的实施方式。
应当理解的是，通式化合物中基团的组合和/或取代应当以符合价键规则为前提。
优选地，本发明的式(I)化合物具有式(Ia)结构，
Figure PCTCN2021125738-appb-000019
其中X是N或CH；R 1、R 2、R 3、R 4、R 5、m、n和A各自具有上文对式(I)化合物的一般或具体实施方式中所定义的含义。
更优选地，本发明的式(I)化合物具有式(Ib)结构，
Figure PCTCN2021125738-appb-000020
其中X是N或CH；R 1、R 2、R 3、R 4、R 5、n和A各自具有上文对式(I)化合物的一般或具体实施方式中所定义的含义。
更优选地，本发明的式(I)化合物具有式(Ic)结构，
Figure PCTCN2021125738-appb-000021
其中X是N或CH；R 2、R 3、R 4、R 5、n和A各自具有上文对式(I)化合物的一般或具体实施方式中所定义的含义。
最优选地，本发明式(I)化合物选自以下具体化合物或其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物：
Figure PCTCN2021125738-appb-000022
发明的有益效果
本发明提供了一类具有通式(I)结构特征的吡唑并吡啶类化合物，经研究发现，该类化合物可有效抑制RET激酶、RET融合及突变活性，作为RET表达异常的相关类疾病的治疗药物。
本发明的化合物具有下列有益效果：
高的RET激酶抑制活性；激酶RET抑制测定实验中显示IC50在0.1nM～1μM范围，优选在0.1nM～0.1μM范围；和/或
高的RET融合及突变活性；和/或
具有良好的药物代谢动力学性质，例如具有更长的t 1/2，从而例如可以加大给药间隔，更长的半衰期，使患者具有更好的依从性；和/或
具有改善的AUC0-t数据，具有更好的成药性，更高的生物利用度；和/或
良好的安全性，如透膜性、P450(减少的药物相互作用风险)、溶解性等优异的性质。
基于以上本发明化合物的有益效果，本发明还提供以下各个方面的技术方案。
用于治疗或用作药物的本发明化合物
一方面，本发明提供用作药物、尤其是用作RET抑制剂的本发明化合物。
另一方面，本发明提供用于治疗、尤其是用于治疗和/或预防与RET有关疾病的本发明化合物。
在具体的实施方式中，本发明提供用于治疗和/或预防RET对所述疾病的发生和发展起到促进作用或抑制RET将降低疾病的发生率、减少或消除疾病病状的疾病的本发明化合物，所述疾病例如肿瘤或肠易激综合症(IBS)，肿瘤包括但不限于非小细胞肺癌、小细胞肺癌、乳头状甲状腺癌、甲状腺髓质癌、分化型甲状腺癌、复发性甲状腺癌、顽固性分化型甲状腺癌、多内分泌肿瘤2A或2B、嗜铬细胞瘤、甲状旁腺增生、乳腺癌、结肠直肠癌、乳头状肾细胞癌、胃肠黏膜神经节瘤、胰管腺癌、多发性内分泌瘤、睾丸癌、慢性单核细胞性白血病、唾腺癌、卵巢癌、子宫颈癌等。
药物组合物及其施用
另一方面，为了使用本说明书的化合物用于治疗或预防目的，可以将本发明化合物根据标准药学实践配制为药物组合物。同时，基于本发明化合物良好的药物代谢动力学性质、改善的AUC0-last、良好的成药性，由本发明化合物可制备具有更好的药动学性质、更高生物利用度的药物。
因此，本发明提供一种药物组合物，其包含上述本发明化合物和药学可接受的赋形剂。
在一个具体的实施方式中，提供了所述本发明的药物组合物，用于在例如哺乳动物如人 个体中预防或治疗与RET有关的疾病。
在一个具体的实施方式中，本发明的药物组合物可以另外包含适合与本发明化合物组合使用的另外的治疗活性成分。
本发明的药物组合物可以通过本领域技术人员已知的技术来配制，如在Remington’s Pharmaceutical Sciences第20版中公开的技术。例如，上述本发明的药物组合物，可以通过将本发明化合物与一种或多种药学可接受的赋形剂混合来制备。制备可进一步包括将一种或多种其它活性成分与本发明化合物和一种或多种药学可接受的赋形剂混合的步骤。
选择包含在特定组合物中的赋形剂将取决于多种因素、例如给药方式和所提供的组合物的形式。合适的药学可接受的赋形剂是本领域技术人员熟知的且描述于例如Ansel,Howard C.,等,Ansel’s Pharmaceutical Dosage Forms and Drug Delivery Systems.Philadelphia:Lippincott,Williams&Wilkins,2004中，包括例如佐剂、稀释剂(例如葡萄糖、乳糖或甘露醇)、载体、pH调节剂、缓冲剂、甜味剂、填充剂、稳定剂、表面活性剂、润湿剂、润滑剂、乳化剂、悬浮剂、防腐剂、抗氧化剂、遮光剂、助流剂、加工助剂、着色剂、加香剂、调味剂、其它已知添加剂。
本发明的药物组合物可以以标准方式施用。例如，合适的施用方式包括口服、静脉内、直肠、肠胃外、局部、经皮、眼、鼻、颊或肺(吸入)给药，其中肠胃外输注包括肌肉、静脉内、动脉内、腹膜内或皮下施用。为了这些目的，本发明的化合物可以通过本领域已知的方法配制成例如片剂、胶囊、糖浆、粉末、颗粒、水性或油性溶液或悬浮液、(脂质)乳剂、可分散粉末、栓剂、软膏、乳膏、滴剂、气溶胶、干粉制剂和无菌可注射水性或油性溶液或悬浮液的形式。
本发明化合物的预防或治疗剂量的大小将根据一系列因素而变化，包括所治疗的个体、病症或病况的严重性、给药的速率、化合物的处置及处方医师的判断。一般而言，有效剂量在每日每kg体重约0.0001至约5000mg，例如约0.01至约1000mg/kg/日(单次或分次给药)。对70kg的人而言，这会合计为约0.007mg/日至约7000mg/日，例如约0.7mg/日至约1500mg/日。根据给药模式，本发明化合物在药物组合物中的含量或用量可以是约0.01mg至约1000mg，适合地是0.1-500mg，优选0.5-300mg，更优选1-150mg，特别优选1-50mg，例如1.5mg、2mg、4mg、10mg、25mg等；相应地，本发明的药物组合物将包含0.05至99％w/w(重量百分比)，例如0.05至80％w/w，例如0.10至70％w/w，例如0.10至50％w/w的本发明化合物，所有重量百分比均基于总组合物。应当理解，可能有必要在某些情况下使用超出这些限制的剂量。
在一种具体实施方式中，本发明提供了一种药物组合物，其包含本发明化合物和一种或多种药学可接受的赋形剂，该组合物被配制用于口服施用。该组合物可以以单位剂型提供，例如以片剂、胶囊或口服液体制剂的形式。这样的单位剂型可以含有0.1mg至1g，例如5mg至250mg的本发明化合物作为活性成分。
在一种具体实施方式中，本发明提供了一种药物组合物，其包含本发明化合物和一种或 多种药学可接受的赋形剂，该组合物被配制用于局部施用。局部施用可以是以例如乳膏剂、洗剂、软膏剂或透皮贴剂的形式。
在一种具体实施方式中，本发明提供了一种药物组合物，其包含本发明化合物和一种或多种药学可接受的赋形剂，该组合物被配制用于吸入施用。吸入施用可以通过口服吸入，也可以通过鼻内施用。当通过口服吸入施用时，本发明的化合物可以以每日剂量有效地用于本发明，例如至多500μg，如0.1-50μg、0.1-40μg、0.1-30μg、0.1-20μg或0.1-10μg的本发明化合物。口服吸入的本发明药物组合物可以配制成干粉、悬浮液(在液体或气体中)或溶液(在液体中)，且可以以任何合适的形式和使用任何本领域已知的合适的吸入器装置施用，包括例如定量吸入器(MDI)、干粉吸入器(DPI)、喷雾器和软雾吸入器。多室装置可用于递送本说明书的化合物和一种或多种其它活性成分(当存在时)。
治疗方法和用途
基于上述本发明化合物具有的有益效果，本发明化合物可用于治疗动物，特别是哺乳动物例如人的各种病症的方法中。
因此，另一方面，本发明提供了调节、尤其是抑制RET活性的方法，所述方法包括使细胞与如前所述的本发明化合物相接触以调节、尤其是抑制细胞中RET的活性。
另一方面，本发明提供了预防或治疗与RET相关的疾病(例如通过RET抑制可治疗或预防的疾病)的方法，所述方法包括向需要其的个体施用有效量的如前所述的本发明化合物或包含其的本发明药物组合物。
另一方面，本发明提供了如前所述的本发明化合物或包含其的药物组合物的用途，用于抑制RET活性，或者用于治疗和/或预防与RET相关的疾病、例如通过RET抑制可治疗或预防的疾病。
另一方面，本发明还提供了如前所述的本发明化合物或包含其的药物组合物在制备药物中的用途，尤其是具有RET受体抑制剂活性的药物中的应用。
另一方面，本发明提供如前所述的本发明化合物或包含其的药物组合物在制备用于治疗或预防与RET相关的疾病、例如通过RET抑制可治疗或预防的疾病的药物中的用途，其中所述化合物或药物组合物任选地与一种或多种化学治疗或免疫治疗联合。
本发明化合物的合成
本发明还提供了一种制备式(I)化合物的方法，下文举例说明了合成本发明化合物的通用合成方案。对于各反应步骤而言，适当的反应条件是本领域技术人员已知的或可以常规确定的。如果没有特别说明，在制备这些化合物中使用的原料和试剂通常可商购获得，或者可以通过下文的方法、与下文给出的方法类似的方法或本领域已知的方法制得。如果需要，合成反应流程中的原料和中间体可以采用常规技术进行分离和纯化，所述技术包括但不限于过滤、蒸馏、结晶、色谱法等。所述材料可以采用包括物理常数和波谱数据在内的常规方法表征。
合成方案1
Figure PCTCN2021125738-appb-000023
一方面，本发明涉及合成式(I)化合物的方法，其包括以下步骤：
使式I-1化合物与式I-2化合物反应得到式(I)化合物，其中R 1、R 2、R 3、R 4、R 5、m、n、A和B各自如上文对式(I)化合物所定义；
其中所述反应可在缩合剂存在下进行，所述缩合剂是本领域众所周知的用于羧酸与胺偶联的缩合剂，包括但不限于1-丙基磷酸酐(T3P)、EDC、DCC、HATU、EDCI等；所述反应优选在适合的有机溶剂中进行，所述有机溶剂可选自二氯甲烷、四氢呋喃、醚类(例如乙醚、乙二醇单甲醚等)、N-甲基吡咯烷酮、N,N-二甲基甲酰胺(DMF)、N,N-二甲基乙酰胺、1,4-二氧六环、二甲基亚砜及其任意组合；所述反应优选在适合的碱存在下进行，所述的碱包括但不限于碳酸钠、碳酸钾、碳酸铯、N,N-二异丙基乙胺、三乙胺、HOBt或吡啶，优选地，所述的碱是N,N-二异丙基乙胺；所述反应优选在适合的温度下进行，例如0-200℃、10-100℃、20-50℃或室温(20-25℃)。
当式I-1化合物的A环通过N原子与吡唑并[1,5-a]吡啶环连接(即为式I-1’化合物)时，其可如合成方案2所示合成。
合成方案2
Figure PCTCN2021125738-appb-000024
其中R 1、R 2、R 5、n和A各自如上文对式(I)化合物所定义；
本发明提供了合成式I-1’化合物的方法，其包括以下步骤：
步骤a:式I-3与式I-4通过亲核取代反应得到式I-5的产物；优选地，反应在在碱存在下进行；
步骤b:式I-5与适当的硼酸或硼酸酯(例如
Figure PCTCN2021125738-appb-000025
)在催化剂作用下通过Suzuki偶联反应得到式I-6的产物；优选地，反应在碱存在下进行；优选地，所述催化剂包括钯催化剂，例如Pd(dppf)Cl 2.DCM或Pd(dppf)Cl 2；和
步骤c:式I-6在酸存在下脱保护得到式I-1’的产物，优选地，所述酸选自盐酸、硫酸、硝酸、三氟乙酸和乙酸。
步骤a)和b)中所述的碱包括但不限于碳酸钠、碳酸钾、碳酸铯、N,N-二异丙基乙胺、三乙胺、HOBt或吡啶；
步骤a)-c)的反应优选在适合的有机溶剂中进行，所述有机溶剂可选自醇(例如甲醇、乙醇)、二氯甲烷、四氢呋喃、醚类(例如乙醚、乙二醇单甲醚等)、N-甲基吡咯烷酮、N,N-二甲基甲酰胺(DMF)、N,N-二甲基乙酰胺、1,4-二氧六环、二甲基亚砜及其任意组合；所述反应优选在适合的温度下进行，例如-70-200℃、0-200℃、10-100℃、100-150℃、70-100℃、20-50℃、室温(20-25℃)或-70至0℃。
上述合成方案只是列举了本发明中部分化合物的制备方法。本发明的化合物或者其立体异构体、互变异构体、稳定的同位素衍生物、药学上可接受的盐或溶剂合物可以通过多种方法、包括上文给出的方法、实施例中给出的方法或与之类似的方法、由本领域普通技术人员在上述合成方案的基础上、结合本领域的常规技术而制备得到。
具体实施方式
以下结合具体实施例，对本发明的技术方案做进一步的描述，但是本发明的保护范围并不限于这些实施例。凡是不背离本发明构思的改变或等同替代均包括在本发明的保护范围之内。
下列实施例中未注明具体条件的实验方法，通常按照这类反应的常规条件，或按照制造厂商所建议的条件。除非另外说明，否则百分比和份数是重量百分比和重量份数。除非另外说明，否则液体的比为体积比。
以下实施例中所用的实验材料和试剂如无特别说明均可从市售渠道获得、依据现有技术的方法制得或根据与本申请公开的类似的方法制得。
本申请使用的缩写具有本领域通常理解的含义，除非说明书中另外清楚定义。在下面列出说明书中使用的缩写的含义：
Boc：叔丁基氧基羰基
Pd(dppf)Cl 2：[1,1'-双(二苯基膦基)二茂铁]二氯化钯
Pd(dppf)Cl 2.DCM:[1,1'-双(二苯基膦)二茂铁]二氯化钯二氯甲烷络合物
HATU：2-(7-偶氮苯并三氮唑)-N,N,N’,N’,-四甲基脲六氟磷酸盐
DIEA：N,N-二异丙基乙胺
DCM:二氯甲烷
EA:乙酸乙酯
PE:石油醚
DMF：N,N-二甲基甲酰胺
LC-MS：液相色谱质谱联用
ESI电喷雾离子化
m/z：质荷比
EDCI：1-乙基-(3-二甲基氨基丙基)碳酰二亚胺盐酸盐
HOBt：1-羟基苯并三唑
TLC:薄层色谱
HPLC:高效液相色谱
合成实施例
本发明提供的目标化合物制备方法中，柱层析色谱采用乳山太阳干燥剂有限公司生产的硅胶(300-400目)；薄层色谱采用GF254(0.25毫米)；核磁共振色谱(NMR)使用Varian-400核磁共振仪测定；液质联用(LC/MS)使用Agilent TechnologiESI 6120液质联用仪。
此外，凡涉及易氧化或易水解的原料的所有操作都在氮气保护下进行。除非另有说明， 本发明使用的原料都是市售原料、无需进一步纯化可以直接使用，本发明使用的温度均为摄氏度℃。
当本发明化合物结构与化合物名称不一致时，通常以结构式所示为准，除非通过上下文可以确定化合物名称正确。
实施例1： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(6-(4- 氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物1)及其盐酸盐(化合物1-1)的合成
Figure PCTCN2021125738-appb-000026
步骤1:(1-(5-溴吡啶-3-基)哌啶-4-基)氨基甲酸叔丁酯的合成
向3,5-二溴吡啶(1.00g,4.22mmol)的N-甲基吡咯烷酮(1.00mL)溶液中加入哌啶-4-氨基甲酸叔丁酯(1.69g,8.44mmol)和DIEA(0.500mL,3.02mmol)。封管，反应液加热至140℃，搅拌反应48小时。将反应混合物冷却至室温，小心倒入水(50.0mL)中得到悬浊液，过滤，滤饼用水(100mL)洗涤后得到粗产物。经柱层析分离纯化(PE：EA＝5：1-3：1)得到目标化合物(480mg，收率32.0％，白色固体)。LC-MS(ESI)m/z 356.0，358.0[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.27(d,J＝2.6Hz,1H),7.99(d,J＝1.8Hz,1H),7.52–7.49(m,1H),6.85(d,J＝7.5Hz,1H),3.82–3.70(m,2H),3.51–3.38(m,1H),2.93–2.77(m,2H),1.84–1.71(m,2H),1.48–1.40(m,2H),1.39(s,9H)。
步骤2：1-氨基-3-溴-5-(4-((叔丁氧基羰基)氨基)哌啶-1-基)吡啶-1-鎓2,4,6-三甲基苯磺酸盐的合成
(1-(5-溴吡啶-3-基)哌啶-4-基)氨基叔丁酯(480mg,1.35mmol)和邻(均三磺酰)羟胺(870mg,4.04mmol)的DCM(15.0mL)溶液在0℃下搅拌反应16小时后，升至室温继续搅拌16小时。反应液减压浓缩得到产物(700mg，粗品，黄色固体)。LC-MS(ESI)m/z 371.1[M] +。
步骤3:(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯的合成
在室温下，向1-氨基-3-溴-5-(4-((叔丁氧基羰基)氨基)哌啶-1-基)吡啶-1-胺2,4,6-三甲基苯磺酸盐(700mg,1.22mmol)的1,4-二氧六环(15.0mL)溶液中加入丙烯腈(149mg,2.82mmol)及DIEA(206mg,1.59mmol)。反应液在室温下搅拌反应2小时后，加入2,3-二氯-5,6-二氰基-1,4-苯醌(584mg,2.57mmol)。反应液在室温继续搅拌反应2小时。将反应混合物倒入水中(50.0mL)中，用EA(250mL)萃取。有机相用饱和食盐水(40.0mL×3)洗涤、无水 硫酸钠干燥及过滤，滤液减压浓缩得粗品。经柱层析分离纯化(PE：EA＝10：1-5：1)得到目标化合物(260mg，两步收率45.9％，黄色固体)。LC-MS(ESI)m/z 442.0，444.0[M+Na] +。 1H NMR(400MHz,MeOH-d 4)δ8.65(d,J＝1.4Hz,1H),8.36(s,1H),7.13(d,J＝1.4Hz,1H),6.80(d,J＝7.3Hz,1H),3.62–3.53(m,1H),3.53–3.44(m,2H),2.89(t,J＝10.7Hz,2H),2.09–2.03(m,1H),2.02–1.97(m,1H),1.93–1.82(m,2H),1.47(s,9H)。
步骤4:(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯的合成
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯(200mg,0.476mmol)和1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(119mg,0.571mmol)的水(2.00mL)和1,4-二氧六环(10.0mL)混合液中加入碳酸钠(101mg,0.952mmol)和Pd(dppf)Cl 2(15.5mg,0.024mmol)。反应液置换氩气三次后，在80℃下搅拌反应16小时。将反应液冷却至室温后，倒入水(50.0mL)中，用EA(250mL)萃取。有机相用饱和食盐水(40.0mL×3)洗、无水硫酸钠干燥并过滤，滤液减压浓缩得粗品。经柱层析分离纯化(PE：EA＝3：1-1：1)得到目标化合物(200mg，收率90.5％，类白色固体)。LC-MS(ESI)m/z 422.1[M+H] +。 1H NMR(400MHz,MeOH-d 4)δ8.64(d,J＝1.2Hz,1H),8.34(s,1H),8.13(s,1H),7.95(d,J＝0.5Hz,1H),7.23(d,J＝1.1Hz,1H),6.81(d,J＝6.9Hz,1H),3.97(s,3H),3.63–3.56(m,1H),3.56–3.48(m,2H),2.93(t,J＝10.9Hz,2H),2.09–2.01(m,2H),1.95–1.83(m,2H),1.48(s,9H)。
步骤5:4-(4-氨基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的合成
(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯(200mg,0.474mmol)的盐酸的1,4-二氧六环溶液(10.0mL,40.0mmol,4.0M)在室温下搅拌反应2小时。将反应混合物减压浓缩、干燥后得到目标化合物(160mg，收率86.0％，黄色固体)。LC-MS(ESI)m/z 322.2[M+H] +。
步骤6:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺及其盐酸盐的合成
在室温下，向2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(44.4mg,0.201mmol)和4-(4-氨基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(60.0mg,0.152mmol)的DMF(3.00mL)溶液中加入EDCI(48.3mg,0.252mmol)、HOBt(33.9mg,0.252mmol)及DIEA(65.1mg,0.504mmol)，反应混合物室温下搅拌反应16小时后，将反应加热至50℃，继续搅拌16小时。将反应液缓缓倾入水(50.0mL)中，用EA(150mL)萃取。有机相用饱和食盐水(40.0mL×3)洗涤、无水硫酸钠干燥及过滤。滤液减压浓缩得粗品(化合物1)。经制 备HPLC(乙腈/含0.05％盐酸的水)纯化得到盐酸盐形式目标化合物(化合物1-1)(28.2mg，收率33.2％，白色固体)。LC-MS(ESI)m/z 525.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.88(s,1H),8.67(d,J＝4.3Hz,1H),8.56(s,1H),8.34(s,2H),8.31(d,J＝7.3Hz,1H),8.05(s,1H),7.93–7.85(m,3H),7.25(s,1H),3.88(s,3H),3.83–3.70(m,1H),3.53(s,2H),3.44(d,J＝12.1Hz,2H),2.87(t,J＝11.2Hz,2H),1.96–1.90(m,2H),1.86-1.78(m,2H)。
实施例2： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(6-(4-氟 -1H-吡唑-1-基)吡啶-3-基)丙酰胺(化合物2)及其盐酸盐(化合物2-1)的合成
Figure PCTCN2021125738-appb-000027
步骤1:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酰胺及其盐酸盐的合成
在室温下，将4-(4-氨基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(80mg,0.203mmol)、2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酸(47.4mg,0.203mmol)、HATU(116mg,0.305mmol)和DIEA(78.7mg,0.609mmol)加入到DMF(1.50mL)中。反应混合物在室温下搅拌反应1小时。向反应混合物中加入水(10.0mL)，用EA(15.0mL×3)萃取。合并有机相用水(10.0mL)、饱和食盐水(10.0mL×1)、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品(化合物2)。经制备HPLC(乙腈/含0.05％盐酸的水)纯化得到盐酸盐形式的目标化合物(化合物2-1)(26.2mg，收率22.5％，米白色固体)。LC-MS(ESI)m/z 539.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.89(d,J＝0.7Hz,1H),8.68(d,J＝4.5Hz,1H),8.56(s,1H),8.40(d,J＝1.9Hz,1H),8.35(s,1H),8.29(d,J＝7.6Hz,1H),8.06(s,1H),7.99–7.94(m,1H),7.93–7.86(m,2H),7.24(s,1H),3.88(s,3H),3.80–3.70(m,2H),3.50–3.36(m,2H),2.93–2.79(m,2H),2.00–1.91(m,1H),1.88–1.66(m,3H),1.40(d,J＝7.0Hz,3H)。
实施例3： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(4-甲氧 基苯基)乙酰胺(化合物3)的合成
Figure PCTCN2021125738-appb-000028
步骤1:(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯的合成
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(500mg,2.08mmol)和哌啶-4-基氨基甲酸叔丁酯(500mg,2.50mmol)的N-甲基吡咯烷酮(2.00mL)溶液加入碳酸钾(576mg,4.17mmol)。封管，反应混合物在100℃下搅拌反应16小时。将反应混合物冷却至室温后倒入水(20.0mL)中，用EA(100mL)萃取。有机相用饱和食盐水(20.0mL)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝10:1-5:1)得到目标化合物(400mg，收率45.7％，灰色固体)。LC-MS(ESI)m/z 419.9，421.9[M+H] +。 1H NMR(400MHz,CDCl 3)δ8.38(d,J＝1.4Hz,1H),8.18(s,1H),6.88(d,J＝1.3Hz,1H),4.64–4.48(m,1H),3.73–3.62(m,1H),3.51–3.42(m,2H),2.89–2.78(m,2H),2.19–2.09(m,2H),1.91–1.80(m,2H),1.46(s,9H)。
步骤2:(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯的合成
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯(400mg,0.952mmol)和1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(238mg,1.14mmol)的水(1.00mL)和1,4-二氧六环(5.00mL)溶液中加入碳酸钠(202mg,1.90mmol)和Pd(dppf)Cl 2.DCM(39.0mg,0.047mmol)。氮气保护下，反应混合物在80℃下搅拌反应16小时。将反应混合物冷却至室温后倒入EA(20.0mL)中得到黑色溶液。溶液用无水硫酸钠干燥、过滤，滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝10:1-5:1)得到目标化合物(300mg，收率74.8％，灰色固体)。LC-MS(ESI)m/z 422.5[M+H] +。 1H NMR(400MHz,CDCl 3)δ8.39–8.34(m,1H),8.20(s,1H),7.78(s,1H),7.68(s,1H),7.03–6.85(m,1H),4.66–4.51(m,1H),4.02(s,3H),3.77–3.64(m,1H),3.58–3.45(m,2H),3.00–2.82(m,2H),2.20–2.12(m,2H),1.96–1.85(m,2H),1.46(s,9H)。
步骤3:4-(4-氨基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的合成
向(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯(100mg,0.237mmol)的甲醇溶液(2.00mL)中加入氯化氢甲醇溶液(3.0M,4.00mL)。反应混合物在室温下搅拌反应12小时。反应混合物减压浓缩得到目标化合物(100mg，理论收 率100％，灰色固体)。LC-MS(ESI)m/z 322.0[M+H] +。
步骤4:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(4-甲氧基苯基)乙酰胺的合成
向4-(4-氨基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(80.0mg,0.224mmol)和2-(4-甲氧基苯基)乙酸(40.0mg,0.241mmol)的DMF(2.00mL)溶液中依次加入DIEA(60.0mg,0.464mmol)、EDCI(68.0mg,0.355mmol)和HOBt(48.0mg,0.355mmol)。反应混合物在室温下搅拌反应12小时。向反应混合物中加入水(10.0mL)，用EA(30.0mL×2)萃取。合并有机相用饱和食盐水(30.0mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)得到目标化合物(21.4mg，收率20.4％，白色固体)。LC-MS(ESI)m/z 470.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.89(s,1H),8.56(s,1H),8.35(s,1H),8.17(d,J＝7.4Hz,1H),8.06(s,1H),7.25(s,1H),7.19(d,J＝8.5Hz,2H),6.86(d,J＝8.5Hz,2H),3.88(s,3H),3.79–3.74(m,1H),3.73(s,3H),3.48–3.40(m,2H),3.34(s,2H),2.86(t,J＝10.6Hz,2H),1.93–1.85(m,2H),1.85–1.72(m,2H)。
实施例4： N-(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(6-(4-氟 -1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物4)的合成
Figure PCTCN2021125738-appb-000029
步骤1:(1-(3-氰基-6-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯的合成
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯(2.21g,5.26mmol)和联硼酸频哪醇酯(6.68g,26.3mmol)的干燥1,4-二氧六环(40.0mL)溶液中加入乙酸钾(1.03g,10.5mmol)和Pd(dppf)Cl 2(227mg,0.263mmol)。氩气保护下，反应混合物在70℃下搅拌3小时。将反应液冷却至室温后，过滤，滤渣用EA(30.0mL)洗，滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝10：1-4：1)得到目标化合物(1.80g，收率73.2％,淡黄色油状物)。LC-MS(ESI)m/z 468.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.65(s,1H),8.55(s,1H),7.96(s,1H),7.02(s,1H),3.97–3.94(m,2H),3.66–3.64(m,1H),2.83–2.75(m,2H),1.91–1.84(m,2H),1.80–1.69(m,2H),1.40(s,9H),1.33(s,12H)。
步骤2:(1-(3-氰基-6-羟基吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯的合成
在0℃下，向(1-(3-氰基-6-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)吡唑并[1,5-a]吡啶 -4-基)哌啶-4-基)氨基甲酸叔丁酯(1.20g,2.57mmol)的四氢呋喃(50.0mL)溶液中加入氢氧化钠固体(514mg,12.8mmol)和30％过氧化氢水溶液(1.80mL,20.5mmol)。反应混合液在室温下搅拌反应1小时后，停止反应。向反应混合液中加入饱和氯化铵(30.0mL)水溶液，然后室温下搅拌30分钟。静置分层，分出有机相。水相用EA(60.0mL)萃取。合并有机相用无水硫酸钠干燥、过滤，滤液减压浓缩得粗产物。经柱层析分离纯化(PE：EA＝4:1-2:1)得到目标化合物(295mg，收率32.1％，黄色固体)。LC-MS(ESI)m/z 358.1[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ10.03(s,1H),8.42(s,1H),7.97(d,J＝1.6Hz,1H),6.98(d,J＝7.5Hz,1H),6.73(d,J＝1.6Hz,1H),3.47–3.41(m,1H),3.38–3.33(m,2H),2.78–2.70(m,2H),1.90–1.83(m,2H),1.79–1.68(m,2H),1.40(s,9H)。
步骤3:(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯的合成
向(1-(3-氰基-6-羟基吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯(290mg,0.811mmol)的四氢呋喃(3.50mL)溶液中加入2.0N氢氧化钠水溶液(0.81mL,1.62mmol)。反应混合液在室温下搅拌反应15分钟加入2,2-二甲基环氧乙烷(293mg,4.06mmol)。封管，反应混合液在60℃下搅拌反应16小时后，补加2,2-二甲基环氧乙烷(293mg,4.06mmol)。混合液继续在60℃下搅拌反应16小时。将反应混合物倒入水(15.0mL)中，用EA(25.0mL×2)萃取。合并有机相用无水硫酸钠干燥、过滤，滤液减压浓缩得粗产物。经反相分离纯化(乙腈/含0.05％甲酸的水)得到目标化合物(240mg，收率68.9％，黄色固体)。LC-MS(ESI)m/z430.3[M+H] +。
步骤4:4-(4-氨基哌啶-1-基)-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的合成
将(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)氨基甲酸叔丁酯(240mg,0.559mmol)加入到氯化氢甲醇溶液(3.0M,15.0mL,45.0mmol)中。反应混合物在室温下搅拌反应16小时。减压浓缩得到目标化合物(180mg，收率88.2％，黄色固体)。LC-MS(ESI)m/z 330.2[M+H] +。
步骤5:N-(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)哌啶-4-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺甲酸盐的合成
在室温下，向4-(4-氨基哌啶-1-基)-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(90.0mg,0.246mmol)和2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(54.4mg,0.246mmol)的DMF(2.00mL)溶液中加入EDCI(70.7mg,0.369mmol)、HOBt(49.9mg,0.369mmol)及DIEA(63.6mg,0.492mmol)。反应混合物在室温下搅拌反应1小时。将反应混合物倒入水中 (10.0mL)中，用EA(30.0mL)萃取。有机相用饱和食盐水(10.0mL×3)洗涤、无水硫酸钠干燥、过滤，滤液减压浓缩得到粗产品。经制备(乙腈/水含0.05％甲酸)纯化得到目标化合物(42.9mg,收率30.1％,白色固体)。LC-MS(ESI)m/z 533.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.69–8.66(m,1H),8.49(s,1H),8.35–8.29(m,3H),7.91(d,J＝3.9Hz,1H),7.89–7.85(m,2H),6.81(d,J＝1.6Hz,1H),4.68(s,1H),3.78(s,2H),3.77-3.73(m,1H),3.52(s,2H),3.41–3.37(m,2H),2.84–2.77(m,2H),1.93–1.86(m,2H),1.83–1.73(m,2H),1.21(s,6H)。
实施例5： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-4-甲基哌啶-4- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物5)及其盐酸盐(化合物5-1)的合成
Figure PCTCN2021125738-appb-000030
步骤1:1-氨基-3-溴-5-氟吡啶-1-鎓2,4,6-三甲基苯磺酸盐的合成
将3-溴-5-氟吡啶(10.0g,56.8mmol)和O-2,4,6-三甲基苯磺酰羟胺(14.7g,68.2mmol)加入到DCM(150mL)中。反应混合物在0℃下搅拌反应1小时。在0℃下向反应液中缓慢加入PE(150mL)，搅拌15分钟得到白色悬浊液。过滤后得到产物(15.5g，白色固体)和粗产物(5.50g，白色固体)。LC-MS(ESI)m/z 191.1,193.1[M] +。
步骤2:6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈的合成
在0℃下，向1-氨基-3-溴-5-氟吡啶-1-鎓2,4,6-三甲基苯磺酸盐(20.0g,51.1mmol)的1,4-二氧六环(200mL)溶液中加入丙烯腈(7.74mL,118mmol)和DIEA(11.0mL,66.4mmol)。反应液在0℃下搅拌反应2小时后，加入2,3-二氯-5,6-二氰基-1,4-苯醌(23.2g,102mmol)。反应液在0℃下继续搅拌反应1小时后，升至室温继续反应16小时。将反应混合物倒入冰水(300mL)中得到棕色悬浊液。悬浊液室温搅拌20分钟后过滤，滤饼用冰水(100mL)洗得到固体粗品。经柱层析分离纯化(PE：EA＝100：1-10：1)得到目标化合物(5.70g,两步收率41.9％,黄色固体)。
步骤3:(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)-4-甲基哌啶-4-基)氨基甲酸叔丁基酯的合成
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(300mg,1.25mmol)和(4-甲基哌啶-4-基)氨基甲酸叔丁基酯(295mg,1.37mmol)的N-甲基吡咯烷酮(2.00mL)溶液中加入碳酸钾(346mg,2.50mmol)。反应混合液在100℃下搅拌反应16小时。将反应混合物冷却至室温后倒入水中(15.0mL)中。用EA(50.0mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干 燥、过滤，滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝10:1)得到目标化合物(400mg，收率63.2％，类白色固体)。LC-MS(ESI)m/z 434.0,436.0[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.95(s,1H),8.62(s,1H),7.13(s,1H),6.55(s,1H),3.20–3.07(m,2H),3.04–2.92(m,2H),2.26–2.14(m,2H),1.82–1.68(m,2H),1.41(s,9H),1.31(s,3H)。
步骤4:(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-4-甲基哌啶-4-基)氨基甲酸叔丁基酯的合成
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)-4-甲基哌啶-4-基)氨基甲酸叔丁基酯(400mg,0.921mmol)和1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(230mg,1.11mmol)的水(0.80mL)和1,4-二氧六环(4.00mL)溶液中加入碳酸钠(195mg,1.84mmol)和Pd(dppf)Cl 2.DCM(39.7mg,0.046mmol)。置换氩气三次。反应混合物在80℃下搅拌反应16小时。将反应混合物冷却至室温后倒入水(20.0mL)中，用EA(60mL×2)萃取。合并有机相用无水硫酸钠干燥、过滤，滤液减压浓缩得到粗品。经柱层析分离纯化(PE：EA＝3：1-1：1)得到目标化合物(360mg，收率89.6％,淡黄色固体)。LC-MS(ESI)m/z 436.1[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.88(d,J＝1.0Hz,1H),8.56(s,1H),8.39(s,1H),8.08(s,1H),7.24(s,1H),6.53(s,1H),3.88(s,3H),3.21–3.11(m,2H),3.08–2.95(m,2H),2.31–2.17(m,2H),1.85–1.72(m,2H),1.42(s,9H),1.32(s,3H)。
步骤5:4-(4-氨基-4-甲基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的合成
(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-4-甲基哌啶-4-基)氨基甲酸叔丁基酯(180mg,0.413mmol)的氯化氢甲醇溶液(3.0M,5.00mL)在室温下搅拌反应16小时。反应混合物减压浓缩得到目标化合物(150mg，收率98.0％，黄色固体)。LC-MS(ESI)m/z336.1[M+H] +。
步骤6:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-4-甲基哌啶-4-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺的合成
在室温下，向2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(65.5mg,0.296mmol)的DMF(2.00mL)溶液中加入EDCI(77.3mg,0.403mmol)、HOBt(54.5mg,0.403mmol)、DIEA(69.5mg,0.538mmol)和4-(4-氨基-4-甲基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(100mg,0.269mmol)。反应混合物在室温下搅拌反应2小时。将反应混合物倒入水中(15.0mL)中，用EA(60.0mL)萃取。有机相用饱和食盐水(10.0mL×3)洗、无水硫酸钠干燥、过滤，滤液减压浓缩得到粗产品(化合物5)。经制备HPLC(乙腈/含0.05％盐酸的水)纯化得到盐酸盐形式的目标化合物(化合物5-1)(35.5mg,收率22.9％,淡黄色固体)。LC-MS (ESI)m/z:539.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.88(s,1H),8.64(d,J＝4.3Hz,1H),8.56(s,1H),8.37-8.33(m,2H),8.05(s,1H),7.94–7.87(m,3H),7.85(s,1H),7.19(s,1H),3.87(s,3H),3.56(s,2H),3.22–3.15(m,2H),3.06–2.98(m,2H),2.32–2.25(m,2H),1.89–1.80(m,2H),1.36(s,3H)。
实施例6： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-4-甲基哌啶-4- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酰胺(化合物6)及其盐酸盐(化合物6-1)的合成
Figure PCTCN2021125738-appb-000031
在室温下，向2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酸(48.7mg,0.207mmol)的DMF(2.00mL)溶液中加入HATU(107mg,0.282mmol)。反应混合物在室温搅拌反应15分钟后，加入DIEA(72.9mg,0.564mmol)和4-(4-氨基-4-甲基哌啶-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(70.0mg,0.188mmol)。反应混合物继续在室温下搅拌反应16.5小时。将反应混合物倒入水中(20.0mL)中，用EA(60.0mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤，滤液减压浓缩得到粗产品(化合物6)。经制备HPLC(乙腈/含0.05％盐酸的水)纯化得到盐酸盐形式的目标化合物(化合物6-1)(26.1mg,收率23.6％,淡黄色固体)。LC-MS(ESI)m/z 553.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.86(s,1H),8.59(d,J＝4.4Hz,1H),8.55(s,1H),8.41(d,J＝2.0Hz,1H),8.31(s,1H),8.02(s,1H),7.98–7.95(m,1H),7.89–7.86(m,2H),7.74(s,1H),7.12(s,1H),3.88(s,3H),3.85–3.83(m,1H),3.20–3.13(m,2H),3.01–2.95(m,1H),2.86–2.80(m,1H),2.33–2.29(m,1H),2.25–2.20(m,1H),1.87–1.78(m,2H),1.39(d,J＝7.0Hz,3H),1.32(s,3H)。
实施例7： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环庚烷-4- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物7)的合成
Figure PCTCN2021125738-appb-000032
步骤1:(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)氮杂环庚烷-4-基)氨基甲酸叔丁酯的合成
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(540mg,2.25mmol)和氮杂环庚烷-4-基氨基甲酸叔丁酯(579mg,2.70mmol)的N-甲基吡咯烷酮(4.00mL)溶液中加入DIEA(582mg,4.50mmol)。封管，反应混合液在100℃下搅拌反应16小时。将反应混合物冷却至室温，慢慢倾入水中(15.0mL)中。用EA(20.0mL×2)萃取。合并有机相用水(10.0mL×2)和饱和食盐水(10.0mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到目标化合物(300mg，粗品，灰色固体)，直接用于下一步。LC-MS(ESI)m/z 434.0,436.0[M+H] +。
步骤2:1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环庚烷-4-基)氨基甲酸叔丁酯的合成
将(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)氮杂环庚烷-4-基)氨基甲酸叔丁酯(300mg,0.691mmol)、1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(216mg,1.04mmol)、碳酸钾(191mg,1.38mmol)和Pd(dppf)Cl 2.DCM(28.2mg,0.034mmol)加入到1,4-二氧六环(2.00mL)和水(0.500mL)的混合溶剂中。封管，氩气置换3次。反应混合物在100℃下搅拌反应2小时。将反应混合物冷却至室温后，分出有机相。减压浓缩得到粗品。经柱层析分离纯化(PE：EA＝1:1)，再经反相纯化(乙腈/含0.05％甲酸的水)得到目标化合物(120mg，两步收率12.2％，黄色固体)。LC-MS(ESI)m/z 436.0[M+H] +。
步骤3:4-(4-氨基氮杂环庚烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的合成
将1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环庚烷-4-基)氨基甲酸叔丁酯(120mg,0.276mmol)加入到盐酸1,4-二氧六环溶液(4.0M,2.00mL)中。反应混合物在室温下搅拌反应2小时。反应混合物减压浓缩得到目标化合物(90.0mg，收率87.4％，灰色固体)。LC-MS(ESI)m/z 336.5[M+H] +。
步骤4:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环庚烷-4- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺的合成
将4-(4-氨基氮杂环庚烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(90.0mg,0.242mmol)、2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(53.5mg,0.242mmol)、HOBt(49.0mg,0.363mmol)、EDCI(92.8mg,0.484mmol)和DIEA(93.8mg,0.726mmol)依次加入到DMF(2.00mL)中。反应混合物在室温下搅拌反应16小时。加入水(10.0mL)，用EA(15.0mL×2)萃取。合并有机相用水(10.0mL×2)和饱和食盐水(10.0mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备板分离纯化(PE：EA＝1:4)得到目标化合物(19.5mg，收率14.9％，黄色固体)。LC-MS(ESI)m/z 539.2[M+H] +。 1H NMR(400MHz,MeOH-d 4)δ8.60(d,J＝1.2Hz,1H),8.49-8.46(m,1H),8.35-8.33(m,1H),8.32(s,1H),8.10(s,1H),7.93(s,1H),7.91–7.84(m,2H),7.69-7.65(m,1H),7.28(d,J＝1.2Hz,1H),4.14–4.04(m,1H),3.94(s,3H),3.57(s,2H),3.54–3.46(m,2H),3.45–3.33(m,2H),2.17–2.07(m,2H),2.05–1.97(m,2H),1.92–1.80(m,2H)。
实施例8： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)-2-(6-(4- 氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物8)的合成
Figure PCTCN2021125738-appb-000033
步骤1:(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯的合成
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(300mg,1.25mmol)和吡咯烷-3-基氨基甲酸叔丁酯(244mg,1.31mmol)的N-甲基吡咯烷酮(2.00mL)溶液加入碳酸钾(345mg,2.50mmol)。封管，反应混合物在100℃下搅拌反应16小时。将反应混合物冷却至室温后倒入水(15.0mL)中，用EA(60.0mL)萃取。有机相用饱和食盐水(15.0mL)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝3:1-1:1)得到目标化合物(290mg，收率57.2％，灰色固体)。LC-MS(ESI)m/z 350.0,351.9[M-56+H] +。 1H NMR(400MHz,CDCl 3)δ8.31(d,J＝1.3Hz,1H),8.18(s,1H),6.71–6.69(m,1H),5.02–4.91(m,1H),4.47–4.34(m,1H),3.82–3.69(m,1H),3.60–3.48(m,1H),3.46–3.36(m,1H),3.34–3.22(m,1H),2.47–2.36(m,1H),2.03–1.93(m,1H),1.46(s,9H)。
步骤2:(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯的合成
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯(160mg,0.394mmol)和1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(99.0mg,0.473mmol)的水(1.00mL)和1,4-二氧六环(5.00mL)溶液中加入碳酸钠(84.0mg,0.788mmol)和Pd(dppf)Cl 2.DCM(17.0mg,0.020mmol)。氮气保护下，反应混合物在80℃下搅拌反应16小时。将反应混合物冷却至室温后倒入EA(20.0mL)中得到黑色溶液。溶液用无水硫酸钠干燥、过滤，滤液减压浓缩得到粗品。经柱层析分离纯化(PE：EA＝9:1-5:1)得到目标化合物(110mg，收率68.8％，灰色固体)。LC-MS(ESI)m/z 408.0[M+H] +。 1H NMR(400MHz,CDCl 3)δ8.30(d,J＝1.1Hz,1H),8.19(s,1H),7.75–7.73(m,1H),7.64(s,1H),6.76(s,1H),5.12–5.02(m,1H),4.47–4.36(m,1H),3.98(s,3H),3.85–3.74(m,1H),3.53–3.37(m,2H),3.26–3.15(m,1H),2.50–2.40(m,1H),2.03–1.93(m,1H),1.46(s,9H)。
步骤3:4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的合成
向(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯(110mg,0.270mmol)的甲醇溶液(2.00mL)中加入氯化氢甲醇溶液(3.0M,2.00mL)。反应混合物在室温下搅拌反应12小时。反应混合物减压浓缩得到目标化合物(90.0mg，收率96.8％，灰色固体)。LC-MS(ESI)m/z 308.0[M+H] +。
步骤4:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺的合成
向4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(60.0mg,0.174mmol)、2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(39.0mg,0.176mmol)和DIEA(45.0mg,0.348mmol)的DMF(2.00mL)溶液中加入EDCI(51.0mg,0.266mmol)和HOBt(36.0mg,0.266mmol)。反应混合物在室温下搅拌反应4小时。向反应混合物中加入水(10.0mL)，用EA(30.0mL×3)萃取。合并有机相用饱和食盐水(30.0mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)得到目标化合物(32.0mg，收率36.0％，白色固体)。LC-MS(ESI)m/z 511.0[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.81–8.78(m,1H),8.69–8.64(m,1H),8.58(s,1H),8.54(d,J＝6.6Hz,1H),8.37(s,1H),8.32(s,1H),8.07(s,1H),7.92(d,J＝4.3Hz,1H),7.89–7.83(m,2H),7.04(s,1H),4.43–4.36(m,1H),3.87(s,3H),3.71–3.65(m,1H),3.63–3.57(m,1H),3.53(s,2H),3.45–3.41(m,1H),3.31–3.28(m,1H),2.31–2.24(m,1H),1.98–1.91(m,1H)。
实施例9： (R)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物9)的合成
Figure PCTCN2021125738-appb-000034
步骤1:(R)-(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯的合成
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(300mg,1.25mmol)和(R)-吡咯烷-3-基氨基甲酸叔丁酯(256mg,1.37mmol)的N-甲基吡咯烷酮(3.00mL)溶液加入碳酸钾(346mg,2.50mmol)。反应混合物在100℃下搅拌反应6小时。将反应混合物冷却至室温后倒入水(20.0mL)中，用EA(100mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤、滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝100:1-10:1)得到目标化合物(240mg，收率46.2％，灰色固体)。LC-MS(ESI)m/z 406.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.75(d,J＝1.4Hz,1H),8.60(s,1H),7.20(d,J＝6.2Hz,1H),6.80(d,J＝1.3Hz,1H),4.20–4.10(m,1H),3.80–3.71(m,1H),3.55–3.43(m,2H),3.30–3.26(m,1H),2.20–2.12(m,1H),1.95–1.87(m,1H),1.39(s,9H)。
步骤2:(R)-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯的合成
向(R)-(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯(240mg,0.591mmol)和1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(148mg,0.709mmol)的水(0.800mL)和1,4-二氧六环(4.00mL)溶液中加入碳酸钠(125mg,1.18mmol)和Pd(dppf)Cl 2.DCM(25.5mg,0.030mmol)。氩气保护下，反应混合物在100℃下搅拌反应8小时。将反应混合物冷却至室温后倒入EA(40.0mL)中得到黑色溶液。溶液用无水硫酸钠干燥、过滤、减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝10：1-1：1)得到目标化合物(190mg，收率79.2％,淡绿色固体)。LC-MS(ESI)m/z 408.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.77–8.75(m,1H),8.55(s,1H),8.36(s,1H),8.05(s,1H),7.23–7.17(m,1H),6.99(s,1H),4.21–4.11(m,1H),3.87(s,3H),3.77–3.69(m,1H),3.52–3.45(m,2H),3.25–3.19(m,1H),2.24–2.15(m,1H),1.95–1.86(m,1H),1.39(s,9H)。
步骤3:(R)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的合成
将(R)-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯(190mg,0.466mmol)加入到氯化氢甲醇溶液(3.0M,10.0mL)中。反应混合物在室温 下搅拌反应16小时。将反应混合物减压浓缩得到目标化合物(160mg，收率99.9％，黄色固体)。LC-MS(ESI)m/z 308.2[M+H] +。
步骤4:(R)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺的合成
在室温下，向(R)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(90.0mg,0.262mmol)和2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(69.5mg,0.314mmol)的DMF(2.00mL)溶液中加入EDCI(75.3mg,0.393mmol)、HOBt(53.1mg,0.393mmol)和DIEA(67.7mg,0.524mmol)。反应混合物室温下搅拌反应1小时。将反应混合物倒入水中(15.0mL)中，用EA(80.0mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤、减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物(55.0mg,收率41.4％,白色固体)。LC-MS(ESI)m/z 511.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.79–8.78(m,1H),8.67–8.65(m,1H),8.57(s,1H),8.52(d,J＝6.7Hz,1H),8.36(s,1H),8.33–8.31(m,1H),8.06(s,1H),7.92–7.90(m,1H),7.89–7.83(m,2H),7.03(s,1H),4.43–4.36(m,1H),3.87(s,3H),3.71–3.65(m,1H),3.64–3.57(m,1H),3.53(s,2H),3.46–3.39(m,1H),3.31–3.28(m,1H),2.34–2.24(m,1H),1.98–1.90(m,1H)。
实施例10： (R)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3- 基)-2-(4-甲氧基苯基)乙酰胺(化合物10)的合成
Figure PCTCN2021125738-appb-000035
在室温下，向(R)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(90.0mg,0.262mmol)和2-(4-甲氧基苯基)乙酸(52.2mg,0.314mmol)的DMF(2.00mL)溶液中加入EDCI(75.3mg,0.393mmol)、HOBt(53.1mg,0.393mmol)和DIEA(67.7mg,0.524mmol)。反应混合物在室温下搅拌反应1小时。将反应混合物倒入水(15.0mL)中，用EA(80.0mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤、减压浓缩得到粗产品。经制备HPLC(乙腈/水含0.05％甲酸)纯化得到目标化合物(68.3mg,收率57.1％,白色固体)。LC-MS(ESI)m/z 456.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.79–8.77(m,1H),8.57(s,1H),8.37–8.33(m,2H),8.06(s,1H),7.15(d,J＝8.6Hz,2H),7.02(s,1H),6.83 (d,J＝8.6Hz,2H),4.40–4.32(m,1H),3.87(s,3H),3.73–3.67(m,1H),3.71(s,3H),3.64–3.57(m,1H),3.47–3.39(m,1H),3.34(s,2H),3.28–3.24(m,1H),2.31–2.20(m,1H),1.97–1.87(m,1H)。
实施例11： (R)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3- 基)-2-(6-甲氧基吡啶-3-基)乙酰胺(化合物11)及其盐酸盐(化合物11-1)的合成
Figure PCTCN2021125738-appb-000036
步骤1:(R)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)-2-(6-甲氧基吡啶-3-基)乙酰胺盐酸盐的制备
向(R)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(120mg,0.349mmol)和2-(6-甲氧基吡啶-3-基)乙酸(58.3mg,0.349mmol)的DMF(2.00mL)溶液中加入EDCI(100mg,0.524mmol)、HOBt(70.7mg,0.524mmol)和DIEA(67.6mg,0.524mmol)。反应混合物在室温下搅拌反应2小时。将反应混合物倒入水(15.0mL)中，用EA(100mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。减压浓缩得到粗产品(化合物11)。经制备HPLC(乙腈/含0.05％盐酸的水)纯化得到盐酸盐形式的目标化合物(化合物11-1)(77.3mg,收率44.9％,黄色固体)。LC-MS(ESI)m/z 457.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ9.08(s,1H),8.79(s,1H),8.57(s,1H),8.49(d,J＝6.6Hz,1H),8.37(s,1H),8.06(s,1H),8.05–8.01(m,1H),7.67–7.63(m,1H),7.03(s,1H),6.80(d,J＝8.5Hz,1H),4.41–4.34(m,1H),3.88(s,3H),3.84(s,3H),3.71–3.66(m,1H),3.63–3.56(m,1H),3.47–3.42(m,1H),3.40(s,2H),3.30–3.25(m,1H),2.31–2.22(m,1H),1.97–1.88(m,1H)。
实施例12： (R)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3- 基)-2-(4-(甲磺酰基)苯基)乙酰胺(化合物12)的合成
步骤1:(R)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)-2-(4-(甲磺酰基)苯基)乙酰胺的制备
Figure PCTCN2021125738-appb-000037
在室温下，向(R)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(80.0mg,0.237mmol)和2-(4-(甲磺酰基)苯基)乙酸(64.8mg,0.302mmol)的DMF(2.00mL)溶液中加入EDCI(66.7mg,0.349mmol)、HOBt(47.2mg,0.349mmol)和DIEA(60.1mg,0.465mmol)。反应混合物在室温下搅拌反应1小时。将反应混合物倒入水(15.0mL)中，用EA(50.0mL×2)萃取。合并有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％氨水的水)纯化得到目标化合物(26.1mg,收率22.3％,类白色固体)。LC-MS(ESI)m/z 504.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.80–8.78(m,1H),8.57(s,1H),8.53(d,J＝6.5Hz,1H),8.36(s,1H),8.06(s,1H),7.87–7.82(m,2H),7.54–7.50(m,2H),7.03(s,1H),4.40–4.35(m,1H),3.87(s,3H),3.72–3.66(m,1H),3.63–3.59(m,1H),3.59-3.55(m,2H),3.44–3.40(m,1H),3.29–3.26(m,1H),3.19(s,3H),2.31–2.25(m,1H),1.96–1.90(m,1H)。
实施例13： (S)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物13)的合成
Figure PCTCN2021125738-appb-000038
步骤1:(S)-(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯的制备
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(300mg,1.25mmol)和(S)-吡咯烷-3-基氨基甲酸叔丁酯(256mg,1.37mmol)的N-甲基吡咯烷酮(2.50mL)溶液加入碳酸钾(346mg,2.50mmol)。反应混合物在100℃下搅拌反应6小时。将反应混合物冷却至室温后倒入水(15.0mL)中，用EA(60.0mL)萃取。有机相分离后用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝50:1-10:1)得到目标化合物(255mg，收率50.2％，白色固体)。LC-MS(ESI)m/z 406.1，408.1[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.75(d,J＝1.3Hz,1H),8.60(s,1H),7.20(d,J＝6.4Hz,1H),6.81(d,J＝1.2Hz, 1H),4.18–4.09(m,1H),3.79–3.72(m,1H),3.53–3.43(m,2H),3.30–3.25(m,1H),2.20–2.11(m,1H),1.95–1.86(m,1H),1.39(s,9H)。
步骤2:(S)-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯的制备
向(S)-(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯(255mg,0.628mmol)和1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(144mg,0.690mmol)的水(0.800mL)和1，4-二氧六环(4.00mL)溶液中加入碳酸钠(133mg,1.26mmol)和Pd(dppf)Cl 2.DCM(27.0mg,0.031mmol)。氮气保护下，反应混合物在100℃下搅拌反应8小时。将反应混合物冷却至室温后倒入EA(60.0mL)中得到黑色溶液。溶液用无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝3:1-1:1)得到目标化合物(190mg，收率74.2％,淡绿色固体)。LC-MS(ESI)m/z 408.3[M+H] +。
步骤3:(S)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的制备
(S)-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)氨基甲酸叔丁酯(190mg,0.466mmol)的氯化氢甲醇溶液(3.0M,10.0mL)在室温下搅拌反应16小时。将反应混合物减压浓缩得到目标化合物(150mg，收率93.6％，黄色固体)。LC-MS(ESI)m/z308.3[M+H] +。
步骤4:(S)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺的制备
在室温下，向(S)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(60.0mg,0.175mmol)和2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(46.3mg,0.209mmol)的DMF(1.50mL)溶液中加入EDCI(50.2mg,0.262mmol)、HOBt(35.4mg,0.262mmol)和DIEA(45.1mg,0.349mmol)。反应混合物在室温下搅拌反应16小时。将反应混合物倒入水(10.0mL)中，用EA(20.0mL×3)萃取。合并有机相用饱和食盐水(10.0mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备(乙腈/水含0.05％甲酸)纯化得到目标化合物(20.9mg,收率21.6％,白色固体)。LC-MS(ESI)m/z 511.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.78(s,1H),8.66(d,J＝4.5Hz,1H),8.57(s,1H),8.53(d,J＝6.6Hz,1H),8.36(s,1H),8.34-8.31(m,1H),8.06(s,1H),7.91(d,J＝4.2Hz,1H),7.88–7.83(m,2H),7.03(s,1H),4.43–4.37(m,1H),3.87(s,3H),3.71–3.65(m,1H),3.63–3.57(m,1H),3.53(s,2H),3.44–3.40(m,1H),2.34–2.24(m,2H),1.97–1.91(m,1H)。
实施例14： (S)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3- 基)-2-(4-甲氧基苯基)乙酰胺(化合物14)的合成
Figure PCTCN2021125738-appb-000039
步骤1:(S)-N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)吡咯烷-3-基)-2-(4-甲氧基苯基)乙酰胺的制备
在室温下，向(S)-4-(3-氨基吡咯烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(60mg,0.175mmol)和2-(4-甲氧基苯基)乙酸(34.8mg,0.209mmol)的DMF(1.50mL)溶液中加入EDCI(50.2mg,0.262mmol)、HOBt(35.4mg,0.262mmol)和DIEA(45.1mg,0.349mmol)。反应混合物在室温下搅拌反应16小时。将反应混合物倒入水(10.0mL)中，用EA(10.0mL×3)萃取。合并有机相用饱和食盐水(10.0mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物(37.29mg,收率46.9％,白色固体)。LC-MS(ESI)m/z 456.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.78(s,1H),8.57(s,1H),8.38–8.33(m,2H),8.06(s,1H),7.15(d,J＝8.5Hz,2H),7.02(s,1H),6.83(d,J＝8.5Hz,2H),4.41–4.32(m,1H),3.87(s,3H),3.73–3.67(m,1H),3.71(s,3H),3.64–3.56(m,1H),3.46–3.40(m,1H),3.34(s,2H),3.28–3.24(m,1H),2.30–2.21(m,1H),1.97–1.87(m,1H)。
实施例15： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物15)的合成
Figure PCTCN2021125738-appb-000040
步骤1:(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)氨基甲酸叔丁酯的合成
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(1.20g,5.00mmol)和氮杂环丁烷-3-基氨基甲酸叔丁酯(1.03g,6.00mmol)的N-甲基吡咯烷酮(6.00mL)溶液加入DIEA(2.48mL,15.0mmol)。封管，反应混合液在100℃下搅拌反应36小时。将反应混合物冷却至室温后慢慢倾入冰水(10.0mL)中，逐渐析出灰色固体。过滤，滤饼依次用水(2.00mL×2)及PE(2.00mL×2)洗， 减压干燥得到目标化合物(1.70g，粗品，灰色固体)。LC-MS(ESI)m/z 392.1,394.1[M+H] +。
步骤2:(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)氨基甲酸叔丁酯的合成
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)氨基甲酸叔丁酯(1.70g,4.33mmol)和1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(1.35g,6.50mmol)的水(2.00mL)和1,4-二氧六环(8.00mL)溶液中加入碳酸钾(1.20g,8.66mmol)和Pd(dppf)Cl 2.DCM(354mg,0.433mmol)。封管，置换氩气3次。反应混合物在100℃下搅拌反应2小时。将反应液冷却至室温后，分出有机相，减压浓缩得到粗产品。柱层析(100％EA)分离纯化得到目标化合物(1.35g，两步收率68.6％，黄色固体)。LC-MS(ESI)m/z 394.0[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.76–8.73(m,1H),8.56(s,1H),8.36(s,1H),8.06(s,1H),7.63–7.58(m,1H),6.75(s,1H),4.48–4.40(m,3H),3.99–3.93(m,2H),3.88(s,3H),1.40(s,9H)。
步骤3:4-(3-氨基氮杂环丁烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈的合成
将(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)氨基甲酸叔丁酯(200mg,0.508mmol)加入到三氟乙酸(1.00mL)中。反应混合物在室温下搅拌反应1小时。将反应混合物用冰水浴冷却，用饱和碳酸氢钠水溶液调节pH＝9，加入二甲基亚砜(5.00mL)。过滤，滤液经液相制备(乙腈/含0.05％甲酸的水)得到目标化合物(58.0mg，收率38.9％，淡黄色固体)。LC-MS(ESI)m/z 294.4[M+H] +。 1H NMR(400MHz,DMSO)δ8.73(d,J＝0.9Hz,1H),8.55(s,1H),8.36(s,1H),8.27(s,2H),8.06(s,1H),6.73(s,1H),4.42(t,J＝7.4Hz,2H),3.94–3.89(m,1H),3.88(s,3H),3.84–3.78(m,2H)。
步骤4:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺的合成
将4-(3-氨基氮杂环丁烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈(58.0mg,0.198mmol)、2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(43.7mg,0.198mmol)、HOBt(40.1mg,0.297mmol)、EDCI(75.9mg,0.396mmol)和DIEA(76.7mg,0.594mmol)依次加入到DMF(2.00mL)中。反应混合物在室温下搅拌反应16小时。将反应混合物倒入水(10.0mL)中，用EA(15.0mL×2)萃取。合并有机相用水(10.0mL)和饱和食盐水(10.0mL)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物(18.1mg，收率18.4％，米白色固体)。LC-MS(ESI)m/z 497.1[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.90(d,J＝6.7Hz,1H),8.76(d,J＝0.8Hz,1H),8.67(d,J＝4.1Hz,1H),8.57(s,1H),8.36(s,1H),8.35–8.34(m,1H),8.07(s,1H),7.93–7.90(m,1H),7.90–7.85 (m,2H),6.79(s,1H),4.67–4.58(m,1H),4.50(t,J＝7.9Hz,2H),4.06–3.99(m,2H),3.87(s,3H),3.57(s,2H)。
实施例16： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酰胺(化合物16)的合成
Figure PCTCN2021125738-appb-000041
步骤1:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酰胺的制备
在室温下，4-(3-氨基氮杂环丁烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈甲酸盐(45.0mg,0.133mmol)和2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酸(37.4mg,0.159mmol)的DMF(2.00mL)溶液中加入EDCI(38.1mg,0.199mmol)、HOBt(26.9mg,0.199mmol)和DIEA(34.2mg,0.265mmol)。反应混合物在室温下搅拌反应2小时。将反应混合物倒入水(15.0mL)中，用EA(80.0mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物(15.2mg,收率22.5％,类白色固体)。LC-MS(ESI)m/z 511.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.84(d,J＝6.5Hz,1H),8.75(s,1H),8.66(d,J＝4.2Hz,1H),8.56(s,1H),8.41–8.37(m,1H),8.35(s,1H),8.06(s,1H),7.96–7.86(m,3H),6.77(s,1H),4.65–4.58(m,1H),4.54–4.48(m,1H),4.46–4.41(m,1H),4.05–4.00(m,1H),3.96–3.90(m,1H),3.87(s,3H),3.78–3.71(m,1H),1.41(d,J＝6.9Hz,3H)。
实施例17： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3- 基)-2-(4-甲氧基苯基)乙酰胺(化合物17)的合成
Figure PCTCN2021125738-appb-000042
步骤1:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)-2-(4-甲氧基苯基)乙酰胺的制备
在室温下，向4-(3-氨基氮杂环丁烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈甲酸盐(45.0mg,0.133mmol)和2-(4-甲氧基苯基)乙酸(26.4mg,0.159mmol)的DMF(2.00mL)溶液中加入EDCI(38.1mg,0.199mmol)、HOBt(26.9mg,0.199mmol)和DIEA(34.2mg,0.266mmol)。反应混合物在室温下搅拌反应2小时。将反应混合物倒入水(15.0mL)中，用EA(80.0mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物(16.6mg,收率28.3％,白色固体)。LC-MS(ESI)m/z 442.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.76–8.72(m,2H),8.56(s,1H),8.36(s,1H),8.07(s,1H),7.18(d,J＝8.8Hz,2H),6.86(d,J＝8.8Hz,2H),6.78(s,1H),4.64–4.57(m,1H),4.51–4.45(m,2H),4.02–3.97(m,2H),3.87(s,3H),3.72(s,3H),3.37(s,2H)。
实施例18： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3- 基)-2-(6-甲氧基吡啶-3-基)乙酰胺(化合物18)的合成
Figure PCTCN2021125738-appb-000043
步骤1：N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)-2-(6-甲氧基吡啶-3-基)乙酰胺的制备
在室温下，向4-(3-氨基氮杂环丁烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈甲酸盐(84mg,0.248mmol)和2-(6-甲氧基吡啶-3-基)乙酸(62.1mg,0.371mmol)的DMF(3.0mL)溶液中加入HATU(141.2mg,0.371mmol)和DIEA(0.18mL,0.990mmol)。反应混合物室温下搅拌反应16小时。反应液直接经制备HPLC纯化(乙腈/水含0.05％甲酸)得到目标化合物(50.0mg，收率45.7％，白色固体)。LC-MS(ESI)m/z 443.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ8.81(d,J＝6.7Hz,1H),8.75(d,J＝0.9Hz,1H),8.56(s,1H),8.36(s,1H),8.07(s,1H),8.02(d,J＝2.1Hz,1H),7.60(dd,J＝8.5,2.4Hz,1H),6.79–6.74(m,2H),4.60(m,1H),4.48(t,J＝7.8Hz,2H),4.00(m,2H),3.87(s,3H),3.82(s,3H),3.40(s,2H)。
实施例19： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3- 基)-2-(6-甲氧基吡啶-3-基)丙酰胺(化合物19)的合成
Figure PCTCN2021125738-appb-000044
步骤1：2-(6-甲氧基吡啶-3-基)丙二酸二甲酯的制备
室温下，向5-溴-2-甲氧基吡啶(2.13mL,16.5mmol)、丙二酸二甲酯(3.76mL,32.9mmol)和溴化亚铜(4.72g,32.9mmol)的1,4-二氧六环(20.0mL)溶液中缓慢加入氢化钠(1.45g,36.2mmol)。氮气保护下，反应混合物在100℃下搅拌反应过夜。将反应混合物冷却至室温后过滤。滤液减压浓缩得到棕色油状物。经柱层析分离纯化(PE：EA＝10:1-2:1)得到目标化合物(490mg，收率12.8％，淡黄色固体)。LC-MS(ESI)m/z 240.3[M+H] +。
步骤2：2-(6-甲氧基吡啶-3-基)-2-甲基丙二酸二甲酯的制备
冰浴下，向2-(6-甲氧基吡啶-3-基)丙二酸二甲酯(450mg,1.88mmol)的DMF(5.00mL)溶液中缓慢加入氢化钠(150mg,3.76mmol)。反应混合物在0℃下搅拌反应30分钟后，加入碘甲烷(401mg,2.83mmol)。反应混合物继续在0℃下搅拌反应2小时。反应混合物用冰水(5.00mL)淬灭，EA(10.0mL×2)萃取。合并有机相用饱和食盐水(5.00mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝10:1-3:1)得到目标化合物(300mg，收率63.0％，黄色油状物)。LC-MS(ESI)m/z 254.3[M+H] +。
步骤3：2-(6-甲氧基吡啶-3-基)丙酸的制备
向2-(6-甲氧基吡啶-3-基)-2-甲基丙二酸二甲酯(250mg,0.987mmol)的四氢呋喃(4.00mL)溶液中加入氢氧化钠水溶液(1M,3.95mL,3.95mmol)。反应混合物在50℃下搅拌反应8小时。待反应液冷却至室温后，用稀盐酸(1M,4.50mL)调节pH～6。用EA(10.0mL×2)萃取。合并有机相用饱和食盐水(5.00mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到目标化合物(90.0mg，收率50.3％，白色固体)。LC-MS(ESI)m/z 182.1[M+H] +。
步骤4：N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)氮杂环丁烷-3-基)-2-(6-甲氧基吡啶-3-基)丙酰胺甲酸盐的制备
向4-(3-氨基氮杂环丁烷-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈甲酸盐(100mg,0.296mmol)的DMF(3.00mL)溶液中加入DIEA(84.2mg,0.652mmol)。反应混合物在室温下搅拌反应10分钟后，加入2-(6-甲氧基吡啶-3-基)丙酸(53.6mg,0.296mmol)、EDCI(93.6mg,0.488mmol)和HOBt(66.0mg,0.488mmol)。反应混合物在室温下继续搅拌反应4 小时。将反应混合物倒入水(10.0mL)中，用EA(15.0mL×3)萃取。合并有机相用饱和食盐水(10.0mL×2)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物(18.2mg,收率12.2％,白色固体)。LC-MS(ESI)m/z 457.2[M+H] +。1H NMR(400MHz,DMSO-d 6)δ8.76–8.71(m,2H),8.56(s,1H),8.36(s,1H),8.08–8.04(m,2H),7.67–7.62(m,1H),6.79–6.74(m,2H),4.62–4.57(m,1H),4.53–4.47(m,1H),4.45–4.40(m,1H),4.04–3.99(m,1H),3.94–3.89(m,1H),3.87(s,3H),3.81(s,3H),3.63–3.56(m,1H),1.33(d,J＝7.1Hz,3H)。
实施例20： N-(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)-2-(6- 甲氧基吡啶-3-基)乙酰胺(化合物20)的合成
Figure PCTCN2021125738-appb-000045
步骤1:(1H-吡唑-4-基)氨基甲酸叔丁酯的制备
向1H-吡唑-4-胺(1.58g,19.0mmol)和碳酸氢钠(3.19g,38.0mmol)的水(2.00mL)和四氢呋喃(20.0mL)溶液中加入二碳酸二叔丁酯(4.57g,20.9mmol)。反应混合液在室温搅拌反应16小时。将反应液中倒入水(50.0mL)中，用EA(100mL×3)萃取。合并有机相用无水硫酸钠干燥，过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝3：1-1:2)得到目标化合物(3.20g，收率91.8％，类白色固体)。LC-MS(ESI)m/z 184.1[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ12.44(s,1H),9.06(s,1H),7.58(s,1H),7.34(s,1H),1.44(s,9H)。
步骤2:(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯的制备
向6-溴-4-氟吡唑并[1,5-a]吡啶-3-甲腈(1.50g,6.25mmol)和(1H-吡唑-4-基)氨基甲酸叔丁酯(1.26g,6.87mmol)的N-甲基吡咯烷酮(10.0mL)溶液加入碳酸钾(1.73g,12.5mmol)。反应混合物在100℃下搅拌反应16小时。将反应混合物冷却至室温后倒入水(30.0mL)中。用EA(200mL)萃取。有机相用饱和食盐水(20.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝5:1-3:1)得到目标化合物(1.40g，收率55.6％，淡黄色固体)。LC-MS(ESI)m/z 403.0,405.0[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ9.55(s,1H),9.41–9.36(m,1H),8.74(s,1H),8.30(s,1H),8.02(s,1H),7.72(s,1H),1.48(s,9H)。
步骤3:(1-(3-氰基-6-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)吡唑并[1,5-a]吡啶-4-基)-1H- 吡唑-4-基)氨基甲酸叔丁酯的制备
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯(1.40g,3.47mmol)和联硼酸频哪醇酯(4.41g,17.4mmol)的干燥1,4-二氧六环(25.0mL)溶液中加入乙酸钾(681mg,6.94mmol)和Pd(dppf)Cl 2.DCM(150mg,0.174mmol)。氩气保护下，反应混合物在90℃下搅拌反应16小时。将反应混合物冷却至室温后倒入EA(50.0mL)中。过滤，滤饼用EA(20.0mL)洗。滤液减压浓缩得到目标化合物(7.00g，粗品,黑色固体)。LC-MS(ESI)m/z 369.1[M-82+H] +。
步骤4:(1-(3-氰基-6-羟基吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯的制备
在0℃下，向(1-(3-氰基-6-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯(7.00g,15.5mmol)的四氢呋喃(200mL)溶液中加入氢氧化钠(2.49g,62.2mmol)。随后滴加30％过氧化氢水溶液(12.7mL,124mmol)。反应混合液在室温下搅拌反应2小时。向反应混合液中加入饱和氯化铵(100mL)溶液，然后室温下搅拌30分钟。分出有机相，水相用EA(100mL×2)萃取。合并有机相合用无水硫酸钠干燥、过滤。滤液减压浓缩得粗产物。经柱层析分离纯化(PE：EA＝3:1-1:1)得到目标化合物(400mg，两步收率33.8％，黄色固体)。LC-MS(ESI)m/z 341.2[M+H] +。
步骤5:(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯的制备
向(1-(3-氰基-6-羟基吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯(400mg,1.18mmol)的四氢呋喃(6.00mL)溶液中加入氢氧化钠(2.0N,1.18mL,2.35mmol)。反应混合物在室温下搅拌反应15分钟后加入2,2-二甲基环氧乙烷(1.05mL,11.8mmol)。封管密闭条件下，反应混合物在80℃下搅拌反应16小时。将反应混合物冷却至室温后减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝2:1-1:5)得到目标化合物(200mg，收率41.2％，黄色固体)。LC-MS(ESI)m/z 413.3[M+H] +。
步骤6:4-(4-氨基-1H-吡唑-1-基)-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的制备
(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯(200mg,0.485mmol)的氯化氢甲醇溶液(3.0M,10.0mL)在室温下搅拌反应16小时。将反应混合物减压浓缩得到目标化合物(160mg，收率94.5％，黄色固体)。LC-MS(ESI)m/z313.2[M+H] +。
步骤7:N-(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)-2-(6-甲 氧基吡啶-3-基)乙酰胺的制备
向4-(4-氨基-1H-吡唑-1-基)-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(80.0mg,0.229mmol)和2-(6-甲氧基吡啶-3-基)乙酸(42.2mg,0.252mmol)的DMF(2.00mL)溶液中加入EDCI(65.9mg,0.344mmol)、HOBt(46.5mg,0.344mmol)和DIEA(59.3mg,0.459mmol)。反应混合物在室温下搅拌反应2小时。将反应混合物倒入水(15.0mL)中，用EA(100mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备(乙腈/含0.05％甲酸的水)纯化得到目标化合物(18.6mg,收率17.6％,类白色固体)。LC-MS(ESI)m/z 462.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ10.48(s,1H),8.74(d,J＝1.9Hz,1H),8.62(s,1H),8.48(s,1H),8.09(d,J＝2.1Hz,1H),7.86(s,1H),7.68–7.64(m,1H),7.62(d,J＝1.9Hz,1H),6.79(d,J＝8.5Hz,1H),4.70(s,1H),3.89(s,2H),3.83(s,3H),3.61(s,2H),1.22(s,6H)。
实施例21： N-(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物21)及其盐酸盐(化合物21-1)的合成
Figure PCTCN2021125738-appb-000046
步骤1:N-(1-(3-氰基-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺盐酸盐的制备
向4-(4-氨基-1H-吡唑-1-基)-6-(2-羟基-2-甲基丙氧基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(80.0mg,0.229mmol)和2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(60.9mg,0.275mmol)的DMF(2.00mL)溶液中加入EDCI(65.9mg,0.344mmol)、HOBt(46.5mg,0.344mmol)和DIEA(59.3mg,0.459mmol)。反应混合物在室温下搅拌反应2小时。将反应混合物倒入水(15.0mL)中，用EA(100mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品(化合物21)。经制备HPLC(乙腈/含0.05％盐酸的水)纯化得到盐酸盐形式的目标化合物(化合物21-1)(20.8mg,收率16.4％,黄色固体)。LC-MS(ESI)m/z 516.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ10.59(s,1H),8.75(d,J＝1.9Hz,1H),8.70–8.68(m,1H),8.62(s,1H),8.50(s,1H),8.41(d,J＝1.7Hz,1H),7.98–7.94(m,1H),7.93–7.88(m,2H),7.88(s,1H),7.63(d,J＝1.9Hz,1H),3.89(s,2H),3.78(s,2H),1.22(s,6H)。
实施例22： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺(化合物22)的的合成
Figure PCTCN2021125738-appb-000047
步骤1:(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯的制备
向(1-(6-溴-3-氰基吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯(190mg,0.470mmol)、1-甲基-4-(4,4,5,5-四甲基-1,3,2-二氧硼杂环戊烷-2-基)-1H-吡唑(118mg,0.570mmol)和碳酸钾(130mg,0.940mmol)的水(1.00mL)和1,4-二氧六环(5.00mL)溶液中加入Pd(dppf)Cl 2.DCM(38.4mg,0.047mmol)。氮气保护下，反应混合物在90℃下搅拌反应16小时。将反应混合物冷却至室温后倒入EA(40.0mL)中，用饱和食盐水(10.0mL)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经柱层析分离纯化(PE：EA＝10:1-1:1)得到目标化合物(130mg，收率68.4％，黄色固体)。LC-MS(ESI)m/z 405.2[M+H] +。
步骤2:4-(4-氨基-1H-吡唑-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐的制备
(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)氨基甲酸叔丁酯(101mg,0.250mmol)的HCl/EA溶液(3.0M,8.00mL)在室温下搅拌反应13小时。将反应混合物减压浓缩得到目标化合物(84.0mg，收率98.8％，白色固体)。LC-MS(ESI)m/z 305.2[M+H] +。
步骤3:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酰胺的的制备
向4-(4-氨基-1H-吡唑-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(100mg,0.293mmol)、2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)乙酸(77.9mg,0.352mmol)、EDCI(84.4mg,0.440mmol)和HOBt(59.5mg,0.440mmol)的DMF(5.00mL)溶液中加入DIEA(189mg,1.46mmol)。反应混合物室温下搅拌反应16小时。将反应混合物倒入水(15.0mL)中，用EA(30.0×2mL)萃取。合并有机相用饱和食盐水(15.0mL)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物 (35.0mg,收率21.6％,白色固体)。LC-MS(ESI)m/z 508.3[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ10.72(s,1H),9.30(s,1H),8.73–8.65(m,2H),8.57(s,1H),8.45–8.40(m,2H),8.16(s,1H),8.04(s,1H),7.99–7.95(m,1H),7.94–7.86(m,3H),3.88(s,3H),3.79(s,2H)。
实施例23： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)-2-(6- 甲氧基吡啶-3-基)乙酰胺(化合物23)的合成
Figure PCTCN2021125738-appb-000048
步骤1:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)-2-(6-甲氧基吡啶-3-基)乙酰胺的制备
向4-(4-氨基-1H-吡唑-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(65.0mg,0.191mmol)和2-(6-甲氧基吡啶-3-基)乙酸(38.3mg,0.229mmol)的DMF(2.00mL)溶液中加入EDCI(54.8mg,0.286mmol)、HOBt(38.6mg,0.286mmol)和DIEA(49.3mg,0.381mmol)。反应混合物在室温下搅拌反应16小时。将反应混合物倒入水(15.0mL)中，用EA(80.0mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品。经制备HPLC(乙腈/含0.05％甲酸的水)纯化得到目标化合物(26.5mg,收率30.6％,白色固体)。LC-MS(ESI)m/z 454.1[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ10.53(s,1H),9.30(d,J＝1.2Hz,1H),8.69(s,1H),8.55(s,1H),8.43(s,1H),8.16(s,1H),8.12–8.09(m,1H),8.05–8.02(m,1H),7.86(s,1H),7.67(dd,J＝8.5Hz,2.4Hz,1H),6.80(d,J＝8.5Hz,1H),3.88(s,3H),3.83(s,3H),3.62(s,2H)。
实施例24： N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4- 基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酰胺(化合物24)及其盐酸盐(化合物24-1)的合成
Figure PCTCN2021125738-appb-000049
步骤1:N-(1-(3-氰基-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-4-基)-1H-吡唑-4-基)-2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酰胺盐酸盐的制备
向4-(4-氨基-1H-吡唑-1-基)-6-(1-甲基-1H-吡唑-4-基)吡唑并[1,5-a]吡啶-3-甲腈盐酸盐(65.0mg,0.191mmol)和2-(6-(4-氟-1H-吡唑-1-基)吡啶-3-基)丙酸(53.8mg,0.229mmol)的DMF(2.00mL)溶液中加入EDCI(54.8mg,0.286mmol)、HOBt(38.6mg,0.286mmol)和DIEA(49.3mg,0.381mmol)。反应混合物在室温下搅拌反应16小时。将反应混合物倒入水(20.0mL)中，用EA(100mL)萃取。有机相用饱和食盐水(15.0mL×3)洗、无水硫酸钠干燥、过滤。滤液减压浓缩得到粗产品(化合物24)。经制备HPLC(乙腈/含0.05％盐酸的水)纯化得到盐酸盐形式的目标化合物(化合物24-1)(60.1mg,收率56.5％,黄色固体)。LC-MS(ESI)m/z 522.2[M+H] +。 1H NMR(400MHz,DMSO-d 6)δ10.59(s,1H),9.30(d,J＝1.2Hz,1H),8.70–8.66(m,2H),8.58(s,1H),8.48(d,J＝2.1Hz,1H),8.43(s,1H),8.16(s,1H),8.05(d,J＝1.2Hz,1H),8.03–7.99(m,1H),7.94–7.89(m,2H),7.86(s,1H),3.98(q,J＝7.0Hz,1H),3.88(s,3H),1.51(d,J＝7.0Hz,3H)。
活性实施例
如无特别说明，以下活性实施例中所用的实验材料、试剂、操作和方法均可从市售渠道获得或基于现有技术容易地获知或制备。
活性实施例1：激酶RET抑制测定
采用类似于文献(Vivek Subbiah,J.F.G.,Precision Targeted Therapy with BLU-667 for RET-Driven Cancers.Cancer Discov,2018.8(7):836-849)所述的方法进行激酶RET抑制测定。利用Caliper Mobility Shift Assay方法检测化合物对激酶RET的抑制效果，化合物测试终浓度为从1000nM起始，3倍系列稀释，共计10个浓度，复孔检测。
将待测化合物溶解在100％DMSO(Sigma,D8418-1L,SHBG3288V)中，配制成10mM储存液，于氮气柜中避光保存。配制1×Kinase buffer，同时如下配制化合物浓度梯度：待测化合物起始测试浓度为1000nM，在384孔板(Corning,3573,12619003)中用DMSO 3倍系列稀释，共10个浓度，复孔测试，终浓度分别为1000、333、111、37、12.3、4.12、1.37、0.457、0.152、0.0508nM，然后用Echo550(Labcyte，型号：Echo 550)转移250nl到384反应板中备用，阴性对照孔和阳性对照孔中分别加250nl 100％DMSO。用1×Kinase buffer配制2.5倍终浓度(终浓度为1nM)的RET(Carna,货号08-159,批号13CBS-0134E)的激酶溶液。在化合物孔和阳性对照孔分别加10μL 2.5倍终浓度的激酶溶液；在阴性对照孔中加10μL 1×Kinase buffer。
将反应板1000rpm离心(Eppendorf，型号：5430)30秒，将反应板振荡混匀后室温孵育10分钟。用1×Kinase buffer配制25倍终浓度(终浓度为16μM)的ATP和15倍终浓度(终浓度 为3μM)的Kinase substrate 2(GL,货号112394，批号P171207-MJ112394)的混合溶液，并加入15μL该混合溶液至各孔中，开始反应。将384孔板1000rpm离心30秒，振荡混匀后室温孵育60分钟。加入30μL含EDTA的终止检测液停止激酶反应，1000rpm离心30秒，振荡混匀。用酶标仪(Perkin Elmer，型号：Caliper EZ ReaderⅡ)读取转化率。
采用以下公式进行计算：
％抑制＝(转化％_max-转化％_样品)/(转化％_样品-转化％_min)*100
其中：“转化％_样品”是样品的转化率读数；“转化％_min”是阴性对照孔均值，代表没有酶活的孔的转化率读数；“转化％_max”是阳性对照孔均值，代表没有化合物抑制的孔的转化率读数。
以浓度的log值作为X轴，百分比抑制率为Y轴，采用分析软件GraphPad Prism 5的log(inhibitor)vs.response-Variable slope拟合量效曲线，
计算公式：Y＝Bottom+(Top-Bottom)/(1+10^((LogIC50-X)*HillSlope))
从而得出各个化合物对酶活性的IC 50值。
表1 RET激酶实验结果
Figure PCTCN2021125738-appb-000050
Figure PCTCN2021125738-appb-000051
活性实施例2：细胞增殖试验
采用CellTiter-Glo TM活细胞检测试剂盒，测试了本发明化合物对细胞增殖的抑制作用。该试剂盒采用萤光素酶作检测物，发光过程中萤光素酶需要ATP的参与，向细胞培养基中加入CellTiter-Glo TM试剂，测量发光值，光信号和体系中ATP量成正比，而ATP又和活细胞数正相关，由此确定细胞的增殖活性。
细胞培养和接种：收获处于对数生长期的细胞(Ba/F3 KIF5B-RET,得自康源博创，RPMI1640(Hyclone,SH30027.01)+10％FBS(GBICO,10099-141)中稳定转染)，并采用血小板计数器进行细胞计数。用台盼蓝排斥法检测细胞活力，确保细胞活力在90％以上。分别添加90μL细胞悬液至96孔透明平底黑壁板(Thermo,165305)中，调整细胞浓度到3000/孔/90ul。将96孔板中的细胞置于37℃、5％CO 2、95％湿度条件下培养过夜(Thermo，Model 3100Series)。
先用DMSO作为溶剂配制10000μM的待测化合物母液，然后用PBS稀释100倍，配制成10倍终浓度的溶液，最高浓度为100μM，在接种有细胞的96孔板中每孔加入10μL待测化合物溶液，即又稀释10倍，浓度达到最终终浓度10μM。将待测化合物终浓度从10μM开始，3倍系列稀释，共9个浓度，每个3个复孔。将已加入待测化合物和细胞的96孔板置于37℃、5％CO 2、95％湿度条件下继续培养72小时，之后进行CellTiter-Glo分析。
融化CellTiter-Glo试剂(
Figure PCTCN2021125738-appb-000052
Luminescent Cell Viability Assay,Promega,G7572)并平衡细胞板至室温30分钟。每孔加入等体积的CellTiter-Glo溶液，在定轨摇床上振动5分钟使细胞裂解。将细胞板放置于室温20分钟以稳定冷光信号，用SpectraMax多标记微孔板检测仪(MD,M3)读取冷光值。
细胞存活率(％)＝(Lum 待测药-Lum 培养液对照)/(Lum 细胞对照-Lum 培养液对照)×100％
使用GraphPad Prism 7.0软件分析数据，利用非线性S曲线回归来拟合数据得出剂量-效应曲线，并由此计算IC 50值。
表2细胞增殖实验结果
Figure PCTCN2021125738-appb-000053
活性实施例3：人和小鼠肝微粒体代谢稳定性实验
根据本领域常规的体外代谢稳定性研究的标准方法，例如(Kerns,Edward H.and Di Li(2008).Drug-like Properties:Concepts,Structure Design and Methods:from ADME to Toxicity Optimization.San Diego:Academic Press；Di,Li等人，Optimization of a Higher Throughput Microsomal Stability Screening Assay for Profiling Drug Discovery Candidates,J.Biomol.Screen.2003,8(4),453.)中所述的方法，类似地如下进行本发明化合物的肝微粒体代谢稳定性试验。
Figure PCTCN2021125738-appb-000054
将肝微粒体(蛋白浓度为0.56mg/mL)加入1μM化合物工作液(由10mM DMSO储备液用100％乙腈稀释到100μM，有机相含量：99％ACN，1％DMSO)，37℃预孵育10min后，加入辅助因子(NADPH)(由氯化镁溶液配制)启动反应。孵育适当时间(如5、10、20、30和60分钟)后取样，加入适当终止液(含有200ng/mL甲苯磺丁脲和200ng/mL拉贝洛尔的冰乙腈(即4℃的乙腈))，终止反应。
样品处理(n＝1)：各加合适样品，涡旋后高速离心，取上清液，采用HPLC-MS/MS对底物进行检测。把0min时间点峰面积作为100％。其他时间点的峰面积转换为百分剩余量，各时间点的百分剩余量的自然对数对孵育时间作图，求算出斜率(-k)，然后按固有清除率(Clint)＝(k*孵育液体积)/肝微粒体质量，计算出Clint(uL/min/mg)及化合物半衰期(T1/2，min)。结果见表3。
表3.人和小鼠肝微粒体代谢稳定性实验结果
Figure PCTCN2021125738-appb-000055
本领域技术人员将了解，上文描述本质上为示例性及说明性的，且欲说明本发明及其优选实施方案。通过常规实验，技术人员将了解可作出明显修正及变化而不悖离本发明的精神。在随附申请专利范围内的所有此类修正欲包括于其中。因此，本发明意欲并非由上述描述而是由以下权利要求范围及其等效物定义。
本说明书中所引用的所有公开出版物以引用方式并入本文中。

Claims (18)

  1. 式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物：
    Figure PCTCN2021125738-appb-100001
    其中：
    R 1选自氢、卤素、氰基、硝基和任选被卤素或氰基取代的C 1-C 6烷基；
    R 2选自氢、C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基、3-8元杂环烯基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其中所述芳基、杂芳基、环烷基、环烯基、杂环烷基、杂环烯基、烷基、烷氧基和烷硫基任选被独立地选自以下的1、2或3个基团取代：卤素、氰基、硝基、羟基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
    R 3选自氢、卤素、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
    R 4选自氢、C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基、3-8元杂环烯基、-SO 2-C 1-C 6烷基、-SO-C 1-C 6烷基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其中所述芳基、杂芳基、环烷基、环烯基、杂环烷基、杂环烯基、烷基、烷氧基和烷硫基任选被独立地选自以下的1、2或3个基团取代：卤素、氰基、硝基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
    R 5选自卤素、氰基、硝基、羟基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；
    环A选自C 6-C 10亚芳基、5-9元亚杂芳基、C 3-C 8亚环烷基、C 3-C 8亚环烯基、3-8元亚杂环烷基和3-8元亚杂环烯基环；
    环B选自C 6-C 10芳基、5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、3-8元杂环烷基和3-8元杂环烯基环；
    m是0、1、2或3；且
    n是0、1、2或3。
  2. 根据权利要求1所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中R 1是卤素、氰基或硝基；优选地，R 1是氰基。
  3. 根据权利要求1-2中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中R 2选自包含1、2或3个独立地选自N、O或S的杂原子的5-6元杂芳基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其任选被独立地选自以下的1、2或3个基团取代：卤素、羟基和C 1-C 6烷基；其中优选地，所述5-6元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基、吡唑基、吡啶基、吡嗪基、哒嗪基或嘧啶基；
    优选地，R 2选自任选被C 1-C 6烷基取代的包含1、2或3个独立地选自N、O或S的杂原子的5元杂芳基和任选被羟基取代的C 1-C 6烷氧基；其中优选地，5元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基或吡唑基。
  4. 根据权利要求3所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中R 2是甲基取代的吡唑基或羟基取代的C 1-C 6烷氧基；
    优选地，R 2是
    Figure PCTCN2021125738-appb-100002
  5. 根据权利要求1-4中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中R 3选自氢和C 1-C 6烷基；
    优选地，R 3选自氢和C 1-C 3烷基；
    更优选地，R 3是氢或甲基；
    最优选地，R 3是氢；
  6. 根据权利要求1-5中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中R 4选自苯基、包含1、2或3个独立地选自N、O或S的杂原子的5-6元杂芳基、-SO 2-C 1-C 6烷基、-SO-C 1-C 6烷基、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基，其任选被1、2或3个卤素基团取代；优选地，所述5-6元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基、吡唑基、吡啶基、吡嗪基、哒嗪基和嘧啶基；
    优选地，R 4选自包含1、2或3个独立地选自N、O或S的杂原子的5元杂芳基、-SO 2-C 1-C 6烷基、C 1-C 6烷基和C 1-C 6烷氧基，其任选被1、2或3个卤素基团取代；优选地，所述5元杂芳基选自吡咯基、呋喃基、噻吩基、咪唑基、噁唑基、异噁唑基、噻唑基、异噻唑基和吡唑基。
  7. 根据权利要求6所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中R 4选自任选被卤素取代的吡唑基、-SO 2-C 1-C 6烷基、C 1-C 6烷基和C 1-C 6烷氧基；
    优选地，R 4是
    Figure PCTCN2021125738-appb-100003
    -SO 2-CH 3或甲氧基。
  8. 根据权利要求1-7中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中R 5选自卤素、C 1-C 6烷基、C 1-C 6烷氧基和C 1-C 6烷硫基；优选地，R 5选自C 1-C 3烷基；更优选是甲基。
  9. 根据权利要求1-8中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中m是0或1；优选地m是1；和/或
    其中n是0或1；优选地，n是0。
  10. 根据权利要求1-9中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中环A选自亚苯基、包含1、2或3个独立地选自N、O或S的杂原子的5-6元亚杂芳基、C 3-C 8亚环烷基、C 3-C 8亚环烯基、包含1、2或3个独立地选自N、O或S的杂原子的3-8元亚杂环烷基和包含1、2或3个独立地选自N、O或S的杂原子的3-8元亚杂环烯基；
    优选地，环A选自亚苯基、包含1、2或3个独立地选自N、O或S的杂原子的5元亚杂芳基、C 3-C 6亚环烷基、C 3-C 6亚环烯基、包含1、2或3个独立地选自N、O或S的杂原子的3-6元亚杂环烷基和包含1、2或3个独立地选自N、O或S的杂原子的3-6元亚杂环烯基。
  11. 根据权利要求10所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中环A选自亚苯基、亚吡咯基、亚呋喃基、亚噻吩基、亚咪唑基、亚呋咱基、亚噁唑基、亚噁二唑基、亚噁三唑基、亚异噁唑基、亚噻唑基、亚异噻唑基、亚吡唑基、亚噻二唑基、亚三唑基、亚四唑基、亚吡啶基、亚吡嗪基、亚哒嗪基、嘧啶基、亚三嗪基、亚氮杂环丁烷基、亚吡咯烷基、亚吡咯啉基、亚环丁基、亚环戊基、亚环己基、亚环庚基、亚哌嗪基、亚哌啶基和氮杂亚环庚基；
    优选地，环A选自亚吡唑基、亚氮杂环丁烷基、亚吡咯烷基、亚环己基、亚哌啶基和氮杂亚环庚基；
    更优选地，环A选自
    Figure PCTCN2021125738-appb-100004
  12. 根据权利要求1-11中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳 定的同位素变体、药学上可接受的盐或溶剂合物，其中环B选自苯基、包含1、2或3个独立地选自N、O或S的杂原子的5-9元杂芳基、C 3-C 8环烷基、C 3-C 8环烯基、包含1、2或3个独立地选自N、O或S的杂原子的3-8元杂环烷基和包含1、2或3个独立地选自N、O或S的杂原子的3-8元杂环烯基环；
    优选地，环B选自苯基、和包含1、2或3个独立地选自N、O或S的杂原子的6元杂芳基；
    更优选地，环B选自苯基、吡咯基、呋喃基、噻吩基、咪唑基、呋咱基、噁唑基、噁二唑基、噁三唑基、异噁唑基、噻唑基、异噻唑基、吡唑基、噻二唑基、三唑基、四唑基、吡啶基、吡嗪基、哒嗪基、嘧啶基和三嗪基环；
    最优选地，环B是苯基或吡啶基环。
  13. 根据权利要求1-12中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中基团
    Figure PCTCN2021125738-appb-100005
    选自
    Figure PCTCN2021125738-appb-100006
  14. 根据权利要求1-13中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，其中所述的式(I)化合物具有式(Ia)结构，
    Figure PCTCN2021125738-appb-100007
    其中X是N或CH；R 1、R 2、R 3、R 4、R 5、m、n和A各自具有权利要求1-13中所定义的含义；
    更优选地，所述的式(I)化合物具有式(Ib)结构，
    Figure PCTCN2021125738-appb-100008
    其中X是N或CH；R 1、R 2、R 3、R 4、R 5、n和A各自具有权利要求1-13中所定义的含义；
    更优选地，所述的式(I)化合物具有式(Ic)结构，
    Figure PCTCN2021125738-appb-100009
    其中X是N或CH；R 2、R 3、R 4、R 5、n和A各自具有权利要求1-13中所定义的含义。
  15. 选自以下的式(I)化合物或其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物：
    Figure PCTCN2021125738-appb-100010
  16. 药物组合物，其包含根据权利要求1-15中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物，和药学上可接受的赋形剂；所述药物组合物任选地另外包含适合与根据权利要求1-15中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物组合使用的另外的治疗活性成分。
  17. 药物组合产品，其包含根据权利要求1-15中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物和另外的活性剂。
  18. 根据权利要求1-15中任何一项所述的式(I)化合物、其立体异构体、互变异构体、稳定的同位素变体、药学上可接受的盐或溶剂合物或根据权利要求16所述的药物组合物在制备用于预防或治疗与RET有关的疾病的药物中的用途；
    优选地，所述的与RET有关的疾病选自肿瘤或肠易激综合症(IBS)，肿瘤包括但不限于非小细胞肺癌、小细胞肺癌、乳头状甲状腺癌、甲状腺髓质癌、分化型甲状腺癌、复发性甲状腺癌、顽固性分化型甲状腺癌、多内分泌肿瘤2A或2B、嗜铬细胞瘤、甲状旁腺增生、乳腺癌、结肠直肠癌、乳头状肾细胞癌、胃肠黏膜神经节瘤、胰管腺癌、多发性内分泌瘤、睾丸癌、慢性单核细胞性白血病、唾腺癌、卵巢癌、子宫颈癌等。
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