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WO2018045956A1 - 苯并咪唑类化合物激酶抑制剂及其制备方法和应用 - Google Patents

苯并咪唑类化合物激酶抑制剂及其制备方法和应用 Download PDF

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WO2018045956A1
WO2018045956A1 PCT/CN2017/100678 CN2017100678W WO2018045956A1 WO 2018045956 A1 WO2018045956 A1 WO 2018045956A1 CN 2017100678 W CN2017100678 W CN 2017100678W WO 2018045956 A1 WO2018045956 A1 WO 2018045956A1
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cancer
amino
cycloalkyl
fluoro
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PCT/CN2017/100678
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English (en)
French (fr)
Inventor
刘世强
周远峰
吴雪松
包如迪
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江苏豪森药业集团有限公司
上海翰森生物医药科技有限公司
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Priority claimed from CN201710157847.4A external-priority patent/CN107793399A/zh
Application filed by 江苏豪森药业集团有限公司, 上海翰森生物医药科技有限公司 filed Critical 江苏豪森药业集团有限公司
Priority to CN201780050147.3A priority Critical patent/CN109963842B/zh
Priority to CN202011082470.9A priority patent/CN112225724B/zh
Publication of WO2018045956A1 publication Critical patent/WO2018045956A1/zh

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    • 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
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/517—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with carbocyclic ring systems, e.g. quinazoline, perimidine
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings
    • 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
    • A61P35/04—Antineoplastic agents specific for metastasis

Definitions

  • the invention belongs to the field of drug development, and particularly relates to a benzimidazole compound kinase inhibitor and a preparation method and application thereof.
  • Cyclin-dependent kinase is a type of serine (Ser)/threonine (Thr) kinase. This family contains 13 members, which are divided into A-L by cyclin. Different CDKs and cyclins form CDK-cyclin complexes, which catalyze the phosphorylation of different substrates through CDK kinase activity, initiate DNA synthesis, promote the promotion and transformation of different phases of the cell cycle, regulate gene transcription, and participate in Cell growth, proliferation, dormancy or entry into apoptosis. Therefore, CDKs have important functions in the regulation of proliferation and death of all cells, including tumor cells and normal cells.
  • CDK4/6-Cyclin D complex plays an important role in the transformation of cells from G1 to S phase.
  • CDK4/6 binds to cyclin D and phosphorylates a range of substrates including Retinoblastoma protein (Rb).
  • Rb phosphorylates and releases the protein bound to and inhibited by it, mainly the transcription factor E2F, etc.
  • E2F activates and transcribes some genes necessary for S phase, and promotes the transformation of G1/S cells.
  • CDK4/6-specific activation is closely related to the proliferation of some tumors, and the abnormalities of the cyclinD–CDK4/6–INK4–Rb pathway are ubiquitous.
  • the performance is as follows: (1) p16INK4a gene deletion, point mutation, or DNA methylation results in inactivation of p16INK4a; (2) CDK4 gene amplification or point mutation (R24C), loss of binding ability to p16INK4a; (3) cyclinD1 due to gene weight Excretion or gene amplification is overexpressed.
  • the change of this pathway accelerates the G1 phase process, which accelerates the proliferation of tumor cells and gains a survival advantage. Therefore, its intervention has become a therapeutic strategy, and CDK4/6 has thus become one of the targets for anti-tumor.
  • Pfizer's Palbociclib (PD0332991) is the first FDA-approved CDK4/6 small molecule inhibitor for breast cancer treatment.
  • Novartis ribociclib (LEE011) was approved in March 2017 for treatment with aromatase inhibitors.
  • Some compounds, such as Lilly Abemaciclib (LY2835219), are in clinical research and both perform well.
  • selective CDK4/6 inhibitors have multiple tumors in ovarian cancer, non-small cell lung cancer, B-cell lymphoma, liver cancer, glioma, colon cancer, multiple myeloma, and the like. Very good anti-tumor activity. Therefore, the development of new CDK4/6 small molecule inhibitors has become a new and effective method for treating these tumors, inspiring generations of scientists to make continuous efforts.
  • Inhibitors of the disclosed selective inhibition of CDK4/6 include WO2004065378, WO2012101013, WO2016192630, WO2016015604 and WO2016015604, and the like.
  • CDK4/6 inhibitor has good application prospect in cancer or tumor treatment as a medicine, the invention A novel structure of highly selective CDK4/6 inhibitor will be provided, and compounds having such a structure are found to exhibit excellent effects and effects.
  • An object of the present invention is to provide a compound represented by the formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, wherein the compound represented by the formula (I) has the following structure:
  • L is a bond, -C(O)- or -C(O)NH-;
  • Ring A is a heterocyclic group wherein the heterocyclic group is selected from the group consisting of a monocyclic heterocyclic group, a spirocyclic heterocyclic group, a fused ring heterocyclic group, and a bridged heterocyclic group;
  • R is selected from a hydrogen atom, a halogen atom or a halogen
  • R 1 is selected from the group consisting of a halogen atom, an alkyl group, a halogenated alkyl group, a halogenated alkyl group, an alkoxy group, a halogenated alkoxy group, a halogen, an amino group, a nitro group, a hydroxyl group, a cyano group, a cycloalkyl group, a heterocyclic group, an aryl group, and a hetero group.
  • aryl - (CH 2) n OR 3, - (CH 2) n SR 3, - (CH 2) n C (O) R 3, - (CH 2) n C (O) OR 3, - (CH 2 ) n S(O) m R 3 , -(CH 2 ) n NR 4 R 5 , -(CH 2 ) n C(O)NR 4 R 5 , -(CH 2 ) n C(O)NHR 4 , -(CH 2 ) n NR 4 C(O)R 5 and -(CH 2 ) n NR 4 S(O) m R 5 , wherein the alkyl group, haloalkyl group, cycloalkyl group, heterocyclic group, aryl group
  • the base and heteroaryl are optionally further selected from the group consisting of a halogen atom, an alkyl group, a halogenated alkyl group, a halogen, an amino group, a nitro group,
  • Base aryl, heteroaryl, -(CH 2 ) n OR 6 , -SR 6 , -(CH 2 ) n C(O)R 6 , -(CH 2 ) n C(O)OR 6 , -( CH 2 ) n S(O) m R 6 , -(CH 2 ) n NR 7 R 8 , -(CH 2 ) n C(O)NR 7 R 8 , -(CH 2 ) n C(O)NHR 7 Substituting one or more substituents of -(CH 2 ) n NR 7 C(O)R 8 and -(CH 2 ) n NR 7 S(O) m R 8 ;
  • R 2 is the same or different and is each independently selected from the group consisting of a hydrogen atom, a halogen atom, an alkyl group, a halogenated alkyl group, a halogenated alkyl group, an alkoxy group, an aminoalkoxy group, a halogenated alkoxy group, a halogen, an amino group, an oxo group, Nitro, hydroxy, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, -(CH 2 ) n OR 3 , -(CH 2 ) n SR 3 , -(CH 2 ) n C(O R 3 , —(CH 2 ) n C(O)OR 3 , —(CH 2 ) n S(O) m R 3 , —(CH 2 ) n NR 4 R 5 , —(CH 2 ) n C( O) NR 4 R 5 , -(CH 2 ) n
  • R 3 is selected from the group consisting of a hydrogen atom, a halogen atom, an alkyl group, a halogenated alkyl group, a halogenated alkyl group, a hydroxyl group, an amino group, an alkoxy group, a halogenated alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group and a heteroaryl group;
  • the alkyl, haloalkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl groups are optionally further selected from the group consisting of a halogen atom, an alkyl group, a halogen, an amino group, a nitro group, a cyano group, a hydroxyl group, a hydroxyalkyl group, Alkoxy, cycloalkyl, heterocyclic, aryl, heteroaryl, -(CH 2 ) n OR 6 , -(CH 2 ) n
  • R 4 and R 5 are the same or different and are each independently selected from the group consisting of a hydrogen atom, a halogen atom, an alkyl group, a halogenated alkyl group, a halogenated alkyl group, a hydroxyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group, and a heteroaryl group.
  • R 6 is selected from the group consisting of a hydrogen atom, a halogen atom, an alkyl group, a halogenated alkyl group, a halogenated alkyl group, a hydroxyl group, an amino group, an alkoxy group, a halogenated alkoxy group, a cycloalkyl group, a heterocyclic group, an aryl group and a heteroaryl group;
  • the alkyl, haloalkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl groups are optionally further selected from the group consisting of a halogen atom, an alkyl group, a halogen, an amino group, a nitro group, a cyano group, a hydroxyl group, a hydroxyalkyl group, Substituted by one or more substituents of alkoxy, cycloalkyl, heterocyclyl, aryl and heteroaryl;
  • R 7 and R 8 are the same or different and are each independently selected from the group consisting of a hydrogen atom, a halogen atom, an alkyl group, a halogenated alkyl group, a halogenated alkyl group, a hydroxyl group, an amino group, an ester group, a cycloalkyl group, a heterocyclic group, an aryl group, and a heteroaryl group, wherein said alkyl group, cycloalkyl group, heterocyclic group, aryl group and heteroaryl group are further optionally selected from the group consisting of a halogen atom, an alkyl group, a halogen group, a hydroxyl group, an amino group, a nitro group, a cyano group, and an ester group. Substituted by one or more substituents of a group, an alkoxy group, a hydroxyalkyl group, a cycloalkyl group, a heterocyclic group, an ary
  • x is an integer of 0, 1, 2, 3, 4 or 5;
  • n is an integer of 0, 1, or 2;
  • n is an integer of 0, 1, 2, 3, 4 or 5.
  • the compound of the formula (I) is a compound represented by the formula (II), a stereoisomer thereof or a pharmaceutically acceptable salt thereof:
  • B is selected from a 3-8 membered monocyclic heterocyclic group, a 6-12 membered spirocyclic heterocyclic group, a 6-12 membered fused ring heterocyclic group or a 6-12 membered bridged heterocyclic group; preferably a 3-8 membered monocyclic ring. Heterocyclic group;
  • L is a bond or -C(O)-
  • R, R 1 , R 2 and x are as defined in claim (II).
  • the compound of the formula (II) is a compound represented by the formula (III), a stereoisomer thereof or a pharmaceutically acceptable salt thereof:
  • Rings B, R, R 1 , R 2 and x are as defined in the formula (II).
  • the compound of the formula (II) is a compound represented by the formula (IV), a stereoisomer thereof or a pharmaceutically acceptable salt thereof:
  • Rings B, R, R 1 , R 2 and x are as defined in the formula (II).
  • M is CR 2 R 2 , NR 2 or O;
  • R is a hydrogen atom or a halogen, wherein the halogen is preferably a fluorine atom;
  • R 1 is alkyl or halogen, wherein the alkyl group is a C 1-6 alkyl group, preferably a C 1-3 alkyl group;
  • R 2 is the same or different and is each independently selected from a hydrogen atom, a C 1-6 alkyl group, a C 1-6 haloalkyl group, a C 1-6 alkoxy group, a C 1-6 amino alkoxy group, a C 1-6 Haloalkoxy, halogen, amino, oxo, nitro, hydroxy, cyano, C 3-8 cycloalkyl, 3-8 membered heterocyclyl, -(CH 2 ) n OR 3 and -(CH 2 ) n NR 4 R 5 , wherein the C 1-6 alkyl group, the C 1-6 haloalkyl group, the C 1-6 amino alkoxy group, the C 3-8 cycloalkyl group and the 3-8 membered heterocyclic group are optional Further selected from C 1-6 alkyl, C 1-6 haloalkyl, halogen, amino, cyano, hydroxy, alkenyl, alkynyl, C 1-6 alkoxy, C 1-6
  • the two R 2 are bonded to each other to form a 3-10 membered cycloalkyl or heterocyclic group, wherein the 3-8 membered cycloalkyl or heterocyclic group is optionally further substituted by one or more C 1 - 6 alkyl, C 1-6 haloalkyl, C 1-6 alkoxy, C 1-6 amino alkoxy, C 1-6 haloalkoxy, halogen, amino, oxo, hydroxy, cyano and C a substituent of a 3-8 cycloalkyl group; preferably a 5-8 membered cycloalkyl group or a heterocyclic group;
  • Y is an integer of 0, 1, 2 or 3;
  • R 3 to R 8 , n and x are as defined in the formula (I).
  • M is selected from CHR 2 or NR 2 ;
  • R 2 is selected from the group consisting of a hydrogen atom, a C 1-6 alkyl group, a C 1-6 haloalkyl group, a C 1-6 alkoxy group, a C 1-6 amino alkoxy group, a C 1-6 haloalkoxy group, a halogen, an amino group, Oxo, hydroxy, cyano, C 3-8 cycloalkyl, 3-8 membered heterocyclyl, -(CH 2 ) n OR 3 and -(CH 2 ) n NR 4 R 5 , wherein said C The 1-6 alkyl group, the C 1-6 haloalkyl group, the C 1-6 amino alkoxy group, the C 3-8 cycloalkyl group and the 3-8 membered heterocyclic group are optionally further selected from a C 1-6 alkyl group, C 1-6 haloalkyl, halogen, amino, cyano, hydroxy, C 2-6 alkenyl, C 2-6 alkynyl, C
  • R, R 1 , R 3 to R 8 , x, n and y are as defined in the formula (V).
  • R 4 and R 5 are each independently selected from a hydrogen atom, a C 1-6 alkyl group, a C 1-6 haloalkyl group, a C 3-8 cycloalkyl group, —(CH 2 ) n OR 6 , —(CH 2 ) n C(O)R 6 wherein said C 1-6 alkyl, C 1-6 haloalkyl, C 3-8 cycloalkyl is optionally further selected from C 1-6 alkyl, halo, hydroxy, amino Substituted with one or more substituents of cyano, C 1-6 alkoxy, C 1-6 hydroxyalkyl and C 1-6 cycloalkyl;
  • R 4 and R 5 form a 3-8 membered heterocyclic group, wherein said 3-8 membered heterocyclic group is further selected from C 1-6 alkyl, -(CH 2 )n-, Substituting one or more substituents of halogen, hydroxy, amino, cyano, C 1-6 alkoxy, C 1-6 hydroxyalkyl and C 1-6 cycloalkyl; preferably R 4 and R 5 are formed
  • the heterocyclic group is 4-6 yuan;
  • R, R 1 and n are as defined in the formula (V).
  • R 1 is selected from the group consisting of C 1-8 alkyl and halogen, wherein The C 1-8 alkyl group, preferably a C 1-6 alkyl group, more preferably a C 1-3 alkyl group; most preferably a methyl group, wherein the halogen is preferably fluorine.
  • R 2 is selected from the group consisting of a hydrogen atom, a C 1-8 alkyl group, C 1-8 haloalkyl, C 1-8 alkoxy, C 3-8 cycloalkyl, C 2-6 alkenyl, halogen, oxo, -(CH 2 ) n NR 4 R 5 and 3-10 a cyclic group wherein the C 1-8 alkyl group, C 1-8 haloalkyl group, C 1-8 alkoxy group, C 3-8 cycloalkyl group and 3-10 membered heterocyclic group are optionally further selected from the group consisting of Substituted by one or more substituents of halogen, hydroxy, cyano, C 1-8 alkyl, -(CH 2 ) n OR 6 and C 1-8 alkoxy; preferably C 1-6 alkyl, C a 1-6 haloalkyl
  • R 4 and R 5 are the same or different and are each independently selected from a hydrogen atom, a C 1-8 alkyl group, a C 1-8 haloalkyl group, a C 3-8 cycloalkyl group, a —(CH 2 ) n C(O)R 6 and a C 1-8 alkoxy group, wherein said C 1-8 alkyl, C 1-8 haloalkyl, C 3-8 cycloalkyl and C 1-8 alkoxy are optionally further selected from the group consisting of halogen, hydroxy, cyano, C 1-8 alkyl, C Substituted with one or more substituents of 3-8 cycloalkyl, -(CH 2 ) n OR 6 and C 1-8 alkoxy; preferably C 1-6 alkyl and C 3-6 cycloalkyl, Wherein the C 1-6 alkyl group and the
  • the compound of the formula (I) is obtained by coupling the compound of the formula (VA) with the compound of the formula (VB), and the compound of the formula (I) is optionally further reacted or further deprotected to give a different compound of the formula (I)
  • the catalytic reagent in the coupling reaction is preferably Pd 2 (dba) 3 and Xantphos reagent;
  • X is a halogen; preferably chlorine
  • Rings A, L, R, R 1 , R 2 and x are as described for the general formula (I). .
  • the present invention also relates to a method of treating a disease preventing and/or treating a CDK4/6 mediated pathological feature comprising administering to a patient a therapeutically effective amount of a compound of the formula (I): a stereoisomer thereof Or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof.
  • the present invention further relates to a compound represented by the formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, which is prepared for the treatment and/or prevention by CDK kinase 4 and/or 6 Use in drugs that mediate cancer or tumor-related diseases.
  • the present invention further relates to a compound represented by the formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, for use in the preparation of a medicament for treating cancer or a tumor, wherein the cancer or Tumor-associated diseases are selected from brain tumors, lung cancer, liver cancer, stomach cancer, oral cancer, head and neck cancer, intestinal or rectal cancer, colon cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, squamous cell carcinoma, thyroid cancer.
  • the cancer or Tumor-associated diseases are selected from brain tumors, lung cancer, liver cancer, stomach cancer, oral cancer, head and neck cancer, intestinal or rectal cancer, colon cancer, kidney cancer, esophageal adenocarcinoma, esophageal squamous cell carcinoma, squamous cell carcinoma, thyroid cancer.
  • bone cancer bone cancer, skin cancer, non-small cell lung cancer, carcinoma in situ, lymphoma, neurofibromatosis, neuroblastoma, mast cell tumor, multiple myeloma, melanoma, glioma, sarcoma or liposarcoma, glue Progesteroma, bladder cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, uterine cancer, cervical cancer, endometrial cancer, Prostate cancer, female genital tract cancer, testicular cancer, gastrointestinal stromal tumor or prostate tumor; preferably from bladder cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, uterine cancer, cervical cancer, endometrial cancer, Prostate cancer, female genital tract cancer, testicular cancer, gastrointestinal stromal tumor or prostate tumor.
  • the present invention further relates to a compound represented by the formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, for the preparation of a method for treating a cancer or a tumor-related disease, which comprises administering a treatment to a patient
  • the breast cancer comprises: a late-stage or metastatic breast cancer that is negative for estrogen receptor-positive and/or human epidermal growth factor receptor 2 negative in postmenopausal women.
  • the present invention further relates to a compound represented by the formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, a pharmaceutical composition comprising a therapeutically effective amount of any one of the formulas (I) a compound, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients.
  • alkyl refers to a saturated aliphatic hydrocarbon group which is a straight or branched chain group containing from 1 to 20 carbon atoms, preferably an alkyl group having from 1 to 8 carbon atoms, more preferably from 1 to 6 carbon atoms.
  • the alkyl group is most preferably an alkyl group of 1 to 3 carbon atoms.
  • Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1 ,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2- Methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3 - dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2 -methylhexyl, 3-methylhexyl, 4-methylhexyl,
  • lower alkyl groups having from 1 to 6 carbon atoms, non-limiting examples including methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl Base, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethyl Butyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl Base, 2,3-dimethylbutyl and the like.
  • the alkyl group may be substituted or unsubstituted, and when substituted, the substituent may be substituted at any available point of attachment, preferably one or more of the following groups independently selected from the group consisting of an alkane Base, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, fluorenyl, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, naphthenic An oxy group, a heterocycloalkoxy group, a cycloalkylthio group, a heterocycloalkylthio group, an oxo group, a carboxyl group or a carboxylate group, and a methyl group, an ethyl group, an isopropyl group, a t-butyl group or a halogenated alkyl group is preferred in the invention.
  • alkylene means that one hydrogen atom of the alkyl group is further substituted, for example, "methylene” refers to -CH 2 -, "ethylene” refers to -(CH 2 ) 2 -, "propylene” Refers to -(CH 2 ) 3 -, "butylene” means -(CH 2 ) 4 - and the like.
  • alkenyl refers to an alkyl group as defined above consisting of at least two carbon atoms and at least one carbon-carbon double bond, such as ethenyl, 1-propenyl, 2-propenyl, 1-, 2- or -butenyl and the like.
  • the alkenyl group may be substituted or unsubstituted, and when substituted, the substituent is preferably one or more of the following groups independently selected from the group consisting of alkyl, alkenyl, alkynyl, alkoxy, alkylthio, Alkylamino, halogen, fluorenyl, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycle Alkylthio group.
  • cycloalkyl refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent containing from 3 to 20 carbon atoms, preferably from 3 to 12 carbon atoms, more preferably from 3 to 8 carbon atoms. One carbon atom, most preferably containing from 3 to 6 carbon atoms.
  • Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatriene
  • Polycyclic cycloalkyl groups include spiro, fused and bridged cycloalkyl groups, preferably cyclopropyl, cyclobutyl, cyclohexyl, cyclopentyl and cycloheptyl.
  • spirocycloalkyl refers to a polycyclic group that shares a carbon atom (referred to as a spiro atom) between 5 to 20 members of a single ring, which may contain one or more double bonds, but none of the rings have a fully conjugated ⁇ electronic system. It is preferably 6 to 14 members, more preferably 7 to 10 members.
  • the spirocycloalkyl group is classified into a monospirocycloalkyl group, a bispirocycloalkyl group or a polyspirocycloalkyl group, preferably a monospirocycloalkyl group and a bispirocycloalkyl group, depending on the number of common spiro atoms between the rings.
  • spirocycloalkyl groups include:
  • spirocycloalkyl groups in which a monospirocycloalkyl group shares a spiro atom with a heterocycloalkyl group, and non-limiting examples include:
  • fused cycloalkyl refers to 5 to 20 members, and each ring in the system shares an all-carbon polycyclic group of an adjacent pair of carbon atoms with other rings in the system, wherein one or more of the rings may contain one or Multiple double bonds, but none of the rings have a fully conjugated ⁇ -electron system. It is preferably 6 to 14 members, more preferably 7 to 10 members.
  • fused cycloalkyl groups include:
  • bridged cycloalkyl refers to an all-carbon polycyclic group of 5 to 20 members, any two rings sharing two carbon atoms which are not directly bonded, which may contain one or more double bonds, but none of the rings have complete Conjugate ⁇ -electron system. It is preferably 6 to 14 members, more preferably 7 to 10 members. Depending on the number of constituent rings, it may be classified into a bicyclic, tricyclic, tetracyclic or polycyclic bridged cycloalkyl group, preferably a bicyclic ring, a tricyclic ring or a tetracyclic ring, and more preferably a bicyclic ring or a tricyclic ring.
  • bridged cycloalkyl groups include:
  • the cycloalkyl ring may be fused to an aryl, heteroaryl or heterocycloalkyl ring, wherein the ring to which the parent structure is attached is a cycloalkyl group, non-limiting examples include indanyl, tetrahydronaphthalene Base, benzocycloheptyl and the like.
  • the cycloalkyl group may be optionally substituted or unsubstituted, and when substituted, the substituent is preferably one or more of the following groups independently selected from the group consisting of alkyl, alkenyl, alkynyl, alkoxy, alkane Thio, alkylamino, halogen, fluorenyl, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio a heterocycloalkylthio group, an oxo group, a carboxyl group or a carboxylate group.
  • heterocyclyl refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent containing from 3 to 20 ring atoms wherein one or more ring atoms are selected from nitrogen, oxygen or S(O).
  • the hetero atom of m (where m is an integer of 0 to 2), but does not include the ring moiety of -OO-, -OS- or -SS-, and the remaining ring atoms are carbon. It preferably comprises from 3 to 12 ring atoms, wherein from 1 to 4 are heteroatoms; further preferably from 3 to 10 ring atoms; more preferably from 3 to 8 ring atoms; most preferably from 6 to 10 ring atoms.
  • Non-limiting examples of monocyclic heterocyclic groups include pyrrolidinyl, imidazolidinyl, tetrahydrofuranyl, tetrahydrothiophenyl, dihydroimidazolyl, dihydrofuranyl, dihydropyrazolyl, dihydropyrrolyl, piperidine.
  • Base piperazinyl, morpholinyl, thiomorpholinyl, homopiperazinyl, pyranyl, etc., preferably piperidinyl and
  • Polycyclic heterocyclic groups include spiro, fused, and bridged heterocyclic groups.
  • spiroheterocyclyl refers to a polycyclic heterocyclic group in which one atom (called a spiro atom) is shared between 5 to 20 members of a single ring, wherein one or more ring atoms are selected from nitrogen, oxygen or S (O). ) m (where m is an integer 0 to 2) heteroatoms, and the remaining ring atoms are carbon. It may contain one or more double bonds, but none of the rings have a fully conjugated pi-electron system. It is preferably 6 to 14 members, more preferably 6 to 10 members.
  • the spiroheterocyclyl group is classified into a monospiroheterocyclic group, a dispiroheterocyclic group or a polyspirocyclic group according to the number of shared spiro atoms between the ring and the ring, and is preferably a monospiroheterocyclic group and a dispiroheterocyclic group.
  • spiroheterocyclyl groups include:
  • fused heterocyclyl refers to 5 to 20 members, and each ring in the system shares an adjacent pair of atomic polycyclic heterocyclic groups with other rings in the system, and one or more rings may contain one or more Double bond, but none of the rings have a fully conjugated ⁇ -electron system in which one or more ring atoms are heteroatoms selected from nitrogen, oxygen or S(O) m (where m is an integer from 0 to 2), and the remaining rings
  • the atom is carbon. It is preferably 6 to 14 members, more preferably 6 to 10 members.
  • fused heterocyclic groups include:
  • bridge heterocyclyl refers to a polycyclic heterocyclic group of 5 to 14 members, any two rings sharing two atoms which are not directly bonded, which may contain one or more double bonds, but none of the rings have a total ⁇ electron conjugated system in which one or more ring atoms selected from nitrogen, oxygen, or S (O) m (wherein m is an integer of 0 to 2) heteroatoms, the remaining ring atoms being carbon. It is preferably 6 to 14 members, more preferably 6 to 10 members.
  • bridge heterocyclic groups include:
  • the heterocyclyl ring may be fused to an aryl, heteroaryl or cycloalkyl ring, wherein the ring to which the parent structure is attached is a heterocyclic group, non-limiting examples of which include:
  • the heterocyclic group may be optionally substituted or unsubstituted, and when substituted, the substituent is preferably one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkane Thio, alkylamino, halogen, fluorenyl, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio a heterocycloalkylthio group, an oxo group, a carboxyl group or a carboxylate group.
  • aryl refers to a 6 to 14 membered all-carbon monocyclic or fused polycyclic ring (ie, a ring that shares a pair of adjacent carbon atoms) having a conjugated ⁇ -electron system, preferably 6 to 10 members, such as benzene. Base and naphthyl. More preferred is phenyl.
  • the aryl ring may be fused to a heteroaryl, heterocyclyl or cycloalkyl ring, wherein the ring to which the parent structure is attached is an aryl ring, non-limiting examples of which include:
  • the aryl group may be substituted or unsubstituted, and when substituted, the substituent is preferably one or more of the following groups independently selected from the group consisting of alkyl, alkenyl, alkynyl, alkoxy, alkylthio, Alkylamino, halogen, fluorenyl, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycle An alkylthio group, a carboxyl group or a carboxylate group.
  • heteroaryl refers to a heteroaromatic system containing from 1 to 4 heteroatoms, from 5 to 14 ring atoms, wherein the heteroatoms are selected from the group consisting of oxygen, sulfur and nitrogen.
  • the heteroaryl group is preferably 5 to 10 members, more preferably 5 members or 6 members, such as imidazolyl, furyl, thienyl, thiazolyl, pyrazolyl, oxazolyl, pyrrolyl, triazolyl, tetra An oxazolyl, pyridyl, pyrimidinyl, thiadiazole, pyrazinyl or the like, preferably a triazolyl, thienyl, imidazolyl, pyrazolyl or pyrimidinyl, thiazolyl; more preferably a triazolyl, pyrrolyl, Thienyl, thiazolyl and pyrimidinyl.
  • the heteroaryl ring may be fused to
  • the heteroaryl group may be optionally substituted or unsubstituted, and when substituted, the substituent is preferably one or more of the following groups independently selected from the group consisting of alkyl, alkenyl, alkynyl, alkoxy, alkane Thio, alkylamino, halogen, fluorenyl, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio a heterocycloalkylthio group, a carboxyl group or a carboxylate group.
  • alkoxy refers to -O-(alkyl) and -O-(unsubstituted cycloalkyl), wherein alkyl is as defined above.
  • alkoxy groups include: methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy, cyclopentyloxy, cyclohexyloxy.
  • the alkoxy group may be optionally substituted or unsubstituted, and when substituted, the substituent is preferably one or more of the following groups independently selected from the group consisting of alkyl, alkenyl, alkynyl, alkoxy, alkane Thio, alkylamino, halogen, fluorenyl, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio a heterocycloalkylthio group, a carboxyl group or a carboxylate group.
  • Haloalkyl means an alkyl group substituted by one or more halogens, wherein alkyl is as defined above.
  • Haloalkoxy means an alkoxy group substituted by one or more halogens, wherein alkoxy is as defined above.
  • Hydroalkyl means an alkyl group substituted by a hydroxy group, wherein alkyl is as defined above.
  • Hydrophilicity refers to an -OH group.
  • Halogen means fluoro, chloro, bromo or iodo.
  • Amino means -NH 2 .
  • Niro means -NO 2 .
  • Carboxy refers to -C(O)OH.
  • oxo refers to Such as oxopiperidinyl
  • THF tetrahydrofuran
  • EtOAc means ethyl acetate
  • MeOH refers to methanol
  • DMF N,N-dimethylformamide
  • DIPEA diisopropylethylamine
  • TFA trifluoroacetic acid
  • MeCN means ⁇ .
  • DMA N,N-dimethylacetamide
  • Et 2 O means diethyl ether
  • DCE 1,2 dichloroethane
  • DIPEA N,N-diisopropylethylamine
  • NBS N-bromosuccinimide
  • NIS N-iodosuccinimide
  • Cbz-Cl means benzyl chloroformate
  • Pd 2 (dba) 3 refers to tris(dibenzylideneacetone) dipalladium.
  • Dppf means 1,1'-bisdiphenylphosphinoferrocene.
  • HATU means 2-(7-oxobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate.
  • KHMDS means potassium hexamethyldisilazide
  • LiHMDS refers to lithium bistrimethylsilylamine.
  • MeLi means methyl lithium
  • n-BuLi means n-butyllithium
  • NaBH(OAc) 3 refers to sodium triacetoxyborohydride.
  • X is selected from A, B, or C
  • X is selected from A, B, and C
  • X is A, B, or C
  • X is A, B, and C
  • Stepoisomerization includes three types of geometric isomerism (cis-trans isomerization), optical isomerism, and conformational isomerism.
  • the hydrogen atom of the present invention may be substituted by its isotope ruthenium, and any of the hydrogen atoms in the examples of the present invention may also be substituted by a ruthenium atom.
  • heterocyclic group optionally substituted by an alkyl group means that an alkyl group may be, but not necessarily, present, and the description includes the case where the heterocyclic group is substituted with an alkyl group and the case where the heterocyclic group is not substituted with an alkyl group.
  • Substituted refers to one or more hydrogen atoms in the group, preferably up to 5, more preferably 1 to 3, hydrogen atoms, independently of each other, substituted by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and those skilled in the art will be able to determine (by experiment or theory) substitutions that may or may not be possible without undue effort. For example, an amino group or a hydroxyl group having a free hydrogen may be unstable when combined with a carbon atom having an unsaturated (e.g., olefinic) bond.
  • “Pharmaceutical composition” means a mixture comprising one or more of the compounds described herein, or a physiologically/pharmaceutically acceptable salt or prodrug thereof, and other chemical components, as well as other components such as physiological/pharmaceutically acceptable carriers. Body and excipients. The purpose of the pharmaceutical composition is to promote the administration of the organism, which facilitates the absorption of the active ingredient and thereby exerts biological activity.
  • “Pharmaceutically acceptable salt” refers to a salt of a compound of the invention which is safe and effective for use in a mammal and which possesses the desired biological activity.
  • the structure of the compound of the present invention is determined by nuclear magnetic resonance (NMR) or/and liquid chromatography-mass spectrometry (LC-MS).
  • NMR chemical shift ( ⁇ ) is given in parts per million (ppm).
  • NMR was measured using a Bruker AVANCE-400 nuclear magnetic apparatus, and the solvent was deuterated dimethyl sulfoxide (DMSO-d 6 ), deuterated methanol (CD 3 OD) and deuterated chloroform (CDCl 3 ).
  • the internal standard was four.
  • Methyl silane (TMS) Methyl silane
  • LC-MS was determined by LC-MS using an Agilent 1200 Infinity Series mass spectrometer.
  • the HPLC was measured using an Agilent 1200 DAD high pressure liquid chromatograph (Sunfire C18 150 x 4.6 mm column) and a Waters 2695-2996 high pressure liquid chromatograph (Gimini C18 150 x 4.6 mm column).
  • the thin layer chromatography silica gel plate uses Yantai Yellow Sea HSGF254 or Qingdao GF254 silica gel plate.
  • the specification for TLC is 0.15mm ⁇ 0.20mm, and the specification for separation and purification of thin layer chromatography is 0.4mm ⁇ 0.5mm.
  • Column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as a carrier.
  • 6-Chloro-2-methylnicotonic acid 0.7 g, 4.1 mmol
  • tert-butyl 1,4-diazo heptane-1-carboxylate 1.0 g, 4.9 mmol
  • TEA 1.2 g , 12.2 mmol
  • reaction solution was (30mL) was diluted with CH 2 Cl 2 after NaHCO 3 solution (30mL), saturated brine (30mL), dried over anhydrous sodium sulfate, and concentrated by column chromatography [eluent: CH 2 Cl 2 ⁇ CH 2 Cl 2 /MeOH (10:1)] product tert-butyl 4-(6-chloro-2-methyl nicotine)-1,4-diazo heptane-1-carboxylate (1.4 g , yield 96%).
  • Second step Preparation of 5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazol-6-yl)pyrimidin-2-amine
  • the third step tert-butyl 4-(6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazole-6-yl)) Preparation of pyrimidin-2-yl)amino)-2-methylnicotinyl)-1,4-diazo heptane-1-carboxylate
  • the second step (1-(6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazol-6-yl)pyrimidine-2) Of tert-butyl ester of 2-amino)-2-methylpyridin-3-yl)-2-carbonylpiperidin-4-yl)carbamate
  • the third step (1-(6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazol-6-yl)pyrimidine-2) Of tert-butyl (meth)amino-2-methylpyridin-3-yl)-2-carbonylpiperidin-4-yl)(methyl)carbamate
  • Second step Preparation of 5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazol-6-yl)pyrimidin-2-amine
  • the third step 1-(6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazol-6-yl)pyrimidin-2- Of amino)amino)-2-methylnicotinoylpiperidin-4-one
  • the fourth step (4-(cyclopropylamino)piperidin-1-yl)(6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzene) And [d]imidazole-6-yl)pyrimidin-2-yl)amino)-2-methylpyridin-3-yl)methanone
  • the third step preparation of benzyl 3-(6-chloro-2-methylnicobutyryl)-3,9-diazabicyclo[4.2.1]nonane-9-carboxylate
  • Step 4 Benzyl 3-(6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazole-6-yl)pyrimidine) Preparation of 2-yl)amino)-2-methylnicotinyl)-3,9-diazabicyclo[4.2.1]nonane-9-carboxylate
  • Step 5 (3,9-diazabicyclo[4.2.1]nonan-3-yl)(6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-) Preparation of methyl-1H-benzo[d]imidazol-6-ylpyrimidin-2-yl)amino)-2-methylpyridin-3-yl)methanone
  • Step 6 (6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazol-6-yl)pyrimidin-2-yl) Preparation of amino)-2-methylpyridin-3-yl)(9-methyl-3,9-diazabicyclo[4.2.1]nonan-3-yl)methanone
  • reaction solution was concentrated and purified by reverse-phase column chromatography to afford compound (6-((5-fluoro-4-(4-fluoro-1-isopropyl-2-methyl-1H-benzo[d]imidazole-6) -yl)pyrimidin-2-yl)amino)-2-methylpyridin-3-yl)(9-methyl-3,9-diazabicyclo[4.2.1]nonan-3-yl)- Ketone (56 mg, yield 50%).
  • Inhibition rate (%) 100 - (signal value - min) / (max - min) * 100.
  • the compounds of the examples of the present invention have a strong inhibitory activity against CDK kinase activity, and particularly have a good inhibitory activity and selectivity for CDK 4 and/or 6 kinase activity.
  • the compound's proliferative activity against colon cancer tumor cell colo205 was tested by the following method.
  • This method was used to determine the inhibitory effect of the compound of the present invention on the proliferative activity of colon cancer tumor cell colo205.
  • the method of the present study the inhibition of a test compound on the colo205 CellTiter-Glo cell proliferation, and to obtain the compound inhibited cell proliferation half maximal inhibitory concentration IC 50 activity.
  • the plate reader measures the chemiluminescence signal value of each plate.
  • the compound of the present invention was assayed for the proliferative activity of colon cancer tumor cell colo205, and the measured IC 50 values are shown in Table 2.
  • the compound of the present invention has a strong inhibitory effect on the proliferation activity of colon cancer tumor cell colo205.
  • Example Compound 3 10 g was weighed, dissolved in 1000 mL of purified water, and 2.5 mL of Tween 80 and 0.5 mL of antifoaming agent were added. Mix well to form a clear solution. 6.1 mg of Example Compound 3, 6.9 mg of Example Compound 34, 5.8 mg of Example Compound 63 were weighed and dissolved in the solution, shaken and sonicated for 15 minutes to give a colorless clear solution at a concentration of 0.5 mg/mL.
  • HEC hydroxyethyl cellulose
  • 0.2 mL of jugular vein blood was taken before administration and 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 24.0 h after administration.
  • plasma was separated by centrifugation at 6000 rpm for 6 min at 4 ° C, and stored at -80 ° C; 4 h after administration.
  • Example 34 Example 34 and Example 63 of the present invention achieved high exposure in rat plasma at a dose of 5 mg/kg orally.
  • the highest blood drug concentration and duration of action vary widely, but the AUC difference is small. Both can guarantee an effective concentration of action.
  • Colo205 tumor-bearing mice were used as test animals, and the pharmacokinetic behavior of Compound Example Compound 3, Compound 34 and Compound 63 was orally administered in mice (plasma and tumor tissue) at a dose of 50 mg/kg.
  • Example Compound 3 10 g was weighed, dissolved in 1000 mL of purified water, and 2.5 mL of Tween 80 and 0.5 mL of antifoaming agent were added. Mix well to form a clear solution. 36.6 mg of Example Compound 3, 22.6 mg of Example Compound 34, and 35.2 mg of Example Compound 63 were weighed and dissolved in the solution, shaken, and sonicated for 15 minutes to obtain a colorless clear solution at a concentration of 5.0 mg/mL.
  • HEC hydroxyethyl cellulose
  • mice were sacrificed by CO 2 , 0.2 ml of blood was collected from the heart, placed in an EDTA-2K test tube, and the plasma was separated by centrifugation at 6000 rpm for 6 min at 4 ° C, and stored at -80 ° C. After the tumor tissue was weighed, it was placed in 2 mL. Store in a centrifuge tube at -80 °C.
  • Example 3 As shown by the data in the table, the exposure of Example 3, Example 34 and Example 63 to mouse plasma and tumor reached a very high level at a dose of 50 mg/kg, Example 3 and Example 63 Exposure in the tumor is significantly higher than in the blood, and Tmax and MRT can be seen that the concentration in the tumor is a gradually increasing process, and the metabolic rate is slower, indicating that the compound will gradually accumulate in the tumor and has been Maintain a high concentration in the tumor to ensure a better anti-tumor effect.
  • the BALB/c nude mice were used as test animals, and the human colorectal cancer cell Colo205 xenograft tumor (CDX) model was used for in vivo pharmacodynamic experiments to evaluate the antitumor effect of the test compounds.
  • CDX human colorectal cancer cell Colo205 xenograft tumor
  • Vernier caliper 500-196, Mitutoyo, Japan
  • Fetal bovine serum (FBS) (10099-141, Gibco)
  • Streptomycin double antibody (SV30010, GE)
  • PBS Phosphate buffer
  • One Colo205 cell was taken out from the cell bank, and the cells were resuscitated with 1640 medium (1640+10% FBS+1%Glu +1% SP).
  • the resuscitated cells were placed in a cell culture flask (the cell type was marked on the bottle wall, The date, culture name, etc.) were placed in a CO 2 incubator (incubator temperature 37 ° C, CO 2 concentration 5%). After the cells were covered with 80-90% of the bottom of the culture flask, they were passaged, and after passage, the cells were further cultured in a CO 2 incubator. This process is repeated until the number of cells meets the in vivo efficacy requirements.
  • the cultured cells were collected, counted by a fully automatic cell counter, and the cells were resuspended in PBS according to the counting results to prepare a cell suspension (density 4 ⁇ 10 7 /ml). Place in an ice box for use.
  • mice Female, 6-8 weeks old, weigh approximately 18-22 grams. Mice were maintained in a special pathogen-free environment and in a single ventilated cage, 5 mice per cage. All cages, litter and water are sterilized prior to use. All animals are free to access a standard certified commercial laboratory diet. Nude mice were labeled with a disposable large mouse universal ear tag prior to inoculation and the skin of the inoculated site was disinfected with 75% medical alcohol. Each mouse was inoculated subcutaneously into the right flank and at a density of 4 x 10 6 cells per 0.1 ml for tumor growth. Dosing begins when the average tumor volume reaches 100-200 cubic millimeters.
  • the test compound was orally administered orally daily at a dose of 50 mg/kg.
  • the anti-tumor efficacy is determined by dividing the average tumor-increased volume of the compound-treated animals by the average tumor-increased volume of the untreated animals.
  • Tumor inhibition rate 1 - [(Vt - V0) administration group / (Vt - V0) solvent control group] * 100%. Animals were euthanized after the experiment.
  • Example 3 In the case where the tumor in the placebo group was increased to 1763 mm 3 , the tumor of the animal group administered in Example 34 was increased to 430 mm 3 , which showed an excellent effect of inhibiting tumor growth.
  • the administration of Example 3 and Example 63 was carried out. Tumor growth was slower in the animal group. The effect was more pronounced.
  • the cells used in this experiment are transfected with hERG, cDNA and CHO stably expressing hERG channels.
  • Cell line supplied by Sophion Bioscience, Denmark
  • the cells were cultured in a medium containing the following components: Ham's F12 medium, 10% (v/v) inactivated fetal bovine serum, 100 ⁇ g/ml hygromycin B, and 100 ⁇ g/ml Geneticin.
  • CHO hERG cells were grown in a Petri dish containing the above culture medium and cultured in an incubator containing 5% CO 2 at 37 °C. 24 to 48 hours before the electrophysiological experiment, CHO hERG cells were transferred to a circular glass plate placed in a petri dish and grown under the same culture medium and culture conditions as above, and the density of CHO hERG cells on each circular slide. The need to reach the vast majority of cells is an independent, single requirement.
  • This experiment used a manual patch clamp system (HEKA EPC-10 signal amplifier and digital conversion system, purchased from HEKA Electronics, Germany) for the recording of whole cell currents.
  • a circular slide with CHO hERG cells grown on it was placed in an electrophysiology recording trough under an inverted microscope.
  • the extracellular fluid was continuously perfused in the recording tank (about 1 ml per minute).
  • the experimental procedure uses conventional whole-cell patch clamp current recording techniques. Unless otherwise stated, the experiments were carried out at regular room temperature ( ⁇ 25 ° C). The cells were clamped at a voltage of -80 mV.
  • the cell clamp voltage was depolarized to +20 mV to activate the hERG potassium channel, and after 5 seconds it was clamped to -50 mV to eliminate inactivation and generate tail current.
  • the tail current peak is used as the value of the hERG current magnitude.
  • the drug to be tested can be superimposed and filled until the inhibition of the hERG current reaches a steady state.
  • the recent three consecutive current recording lines are recombined as a criterion for judging whether or not the state is stable. After reaching a steady state, rinse with extracellular fluid until the hERG current returns to the size before the drug is added.
  • Cisapride (cisapride, purchased from Sigma) was used in the experiment as a positive control to ensure that the cells used were of normal quality.
  • DMSO dimethyl methacrylate
  • concentrations 30, 10, 3, 1, 0.3 and 0.1, 3 (30, 10, 3, 1, 0.3 and 0.1 ⁇ M) for testing.
  • DMSO diluted to 10, 3, 1, 0.3, and 0.1 mM stocks in a gradient dilution, and then diluted 1000 times with extracellular fluid to a final ⁇ M test concentration, except for a final concentration of 0.3% for the 30 ⁇ M test concentration of DMSO.
  • the final concentration of DMSO in the other concentrations of the compound solution was 0.1%.
  • the positive control Cisapride West sapride was tested at a concentration of 0.1 ⁇ M. All compound solutions were sonicated and shaken for 5 to 10 minutes to ensure complete dissolution of the compound. .
  • test data was analyzed by HEKA Patchmaster (V2x73.2), Microsoft Excel and data analysis software provided by Graphpad Prism 5.0.
  • test data in the report needs to meet the following criteria:
  • 0.1 ⁇ M cisapride (C4740-10 mg, Sigma) blocked more than 50% of the hERG current as a positive control.
  • Example Compound 3 Compound 34 and Compound 63 had substantially no inhibitory effect on cardiac hERG potassium ion channels.
  • the present invention provides a series of highly active, highly selective CDK4/6 kinase inhibitors having novel structures with stronger enzymatic activity and cell viability, and better selectivity to kinases. It shows better pharmacokinetic properties in both rats and mice, and also shows better drug efficacy. Cardiac toxicity is significantly reduced. There is great potential to be developed as a drug for diseases of cell cycle proliferative disorders. Especially for HR+/Her-type breast cancer drugs.

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Abstract

提供一种通式(I)所示的化合物、其制备方法及含有该化合物的药物组合物，及其作为苯并咪唑类化合物激酶抑制剂在制备预防和/或治疗癌症或肿瘤相关疾病特别是膀胱癌、卵巢癌、腹膜癌、胰腺癌、乳腺癌、子宫癌、子宫颈癌、子宫内膜癌、前列腺癌、雌性生殖道癌、睾丸癌、胃肠道间质瘤或前列腺肿瘤等疾病的药物中的用途。

Description

苯并咪唑类化合物激酶抑制剂及其制备方法和应用 技术领域
本发明属于药物开发领域，具体涉及一种苯并咪唑类化合物激酶抑制剂及其制备方法和应用。
背景技术
细胞周期蛋白依赖性激酶(cyclin-dependent kinase,CDK)是一类丝氨酸(Ser)/苏氨酸(Thr)激酶，该家族包含13个成员，分别为cyclin分为A-L。不同的CDK和细胞周期素(cyclin)形成CDK-cyclin复合物，通过CDK激酶活性，催化不同底物磷酸化，启动DNA合成，实现对细胞周期不同时相的推进和转化；调控基因转录，参与细胞的生长、增殖、休眠或者进入凋亡。因此，CDKs在所有细胞，包括肿瘤细胞和正常细胞的增殖和死亡调控中具有重要功能。其中，CDK4/6-Cyclin D复合物在细胞从G1期到S期的转化中具有重要作用。在G1期，CDK4/6与cyclin D结合后，使包括视网膜母细胞瘤蛋白(Retinoblastoma protein,Rb)在内的一系列底物磷酸化。Rb磷酸化后释放与其结合并被其抑制的蛋白，主要是转录因子E2F等，E2F激活并转录进入S期所必须的一些基因，促进细胞G1/S的转化。研究发现，CDK4/6特异性的激活与一些肿瘤的增殖密切相关，cyclinD–CDK4/6–INK4–Rb通路的异常普遍存在。表现为：(1)p16INK4a基因缺失，点突变，或者DNA甲基化导致p16INK4a失活；(2)CDK4基因扩增或者点突变(R24C)，失去和p16INK4a结合能力；(3)cyclinD1因为基因重排或者基因扩增而过度表达。这条通路的改变，加速了G1期进程，使得肿瘤细胞增殖加快而获得生存优势。因此，对其的干预成为一种治疗策略，CDK4/6因此成为抗肿瘤的靶点之一。
辉瑞的Palbociclib(PD0332991)是第一个FDA批准上市治疗乳腺癌的CDK4/6小分子抑制剂。接着，诺华ribociclib(LEE011)于2017年3月获批与芳香化酶抑制剂联合治疗。礼来Abemaciclib(LY2835219)等在内的一些化合物处于临床研究，而且均表现不错的治疗效果。除了乳腺癌外，研究表明,选择性CDK4/6抑制剂在卵巢癌、非小细胞肺癌、B细胞淋巴瘤、肝癌、神经胶质瘤、结肠癌、多发性骨髓瘤等多种肿瘤中均具有很好的抗肿瘤活性。因此，开发新的CDK4/6的小分子抑制剂，成为治疗这些肿瘤的新的有效方法，激励着一代代科学家们为此不断的做出努力。
公开的选择性抑制CDK4/6的抑制剂专利申请包括WO2004065378、WO2012101013、WO2016192630、WO2016015604和WO2016015604等。
CDK4/6抑制剂作为药物在癌症或肿瘤治疗中具有良好的应用前景，本发明 将提供一种新型结构的高选择性的CDK4/6抑制剂，并发现具有此类结构的化合物表现出优异的效果和作用。
发明内容
本发明的目的在于提供一种通式(I)所示的化合物、其立体异构体或其药学上可接受盐，其中通式(I)所示的化合物结构如下：
Figure PCTCN2017100678-appb-000001
其中：
L为键、-C(O)-或-C(O)NH-；
环A为杂环基，其中所述的杂环基选自单环杂环基、螺环杂环基、稠环杂环基和桥环杂环基；
R选自氢原子、氘原子或卤素；
R1选自氘原子、烷基、氘代烷基、卤代烷基、烷氧基、卤代烷氧基、卤素、氨基、硝基、羟基、氰基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR3、-(CH2)nSR3、-(CH2)nC(O)R3、-(CH2)nC(O)OR3、-(CH2)nS(O)mR3、-(CH2)nNR4R5、-(CH2)nC(O)NR4R5、-(CH2)nC(O)NHR4、-(CH2)nNR4C(O)R5和-(CH2)nNR4S(O)mR5，其中所述的烷基、卤代烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤代烷基、卤素、氨基、硝基、氰基、羟基、烷氧基、卤代烷氧基、羟烷基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、-SR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8中的一个或多个取代基所取代；
R2相同或不同，且各自独立地选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、烷氧基、氨基烷氧基、卤代烷氧基、卤素、氨基、氧代基、硝基、羟基、氰基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR3、-(CH2)nSR3、-(CH2)nC(O)R3、-(CH2)nC(O)OR3、-(CH2)nS(O)mR3、-(CH2)nNR4R5、-(CH2)nC(O)NR4R5、-(CH2)nC(O)NHR4、-(CH2)nNR4C(O)R5和-(CH2)nNR4S(O)mR5，其中所述的烷基、氘代烷基、卤代烷基、氨基烷氧基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤代烷基、卤素、氨基、硝基、氰基、羟基、烯基、炔基、烷氧基、卤代烷氧基、羟烷基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、 -(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8中的一个或多个取代基所取代；
R3选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、烷氧基、卤代烷氧基、环烷基、杂环基、芳基和杂芳基；其中所述的烷基、卤代烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、氨基、硝基、氰基、羟基、羟烷基、烷氧基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、-(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8中的一个或多个取代基所取代；
R4和R5相同或不同，且各自独立地选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、-(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8，其中所述的烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、羟基、氨基、硝基、氰基、烷氧基、羟烷基、环烷基、杂环基、芳基和杂芳基、-(CH2)nOR6、-(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8中的一个或多个取代基所取代；
R6选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、烷氧基、卤代烷氧基、环烷基、杂环基、芳基和杂芳基；其中所述的烷基、卤代烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、氨基、硝基、氰基、羟基、羟烷基、烷氧基、环烷基、杂环基、芳基和杂芳基的一个或多个取代基所取代；
R7和R8相同或不同，且各自独立地选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、酯基、环烷基、杂环基、芳基和杂芳基，其中所述的烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、羟基、氨基、硝基、氰基、酯基、烷氧基、羟烷基、环烷基、杂环基、芳基和杂芳基中的一个或多个取代基所取代；
x为0、1、2、3、4或5的整数；
m为0、1或2的整数；且
n为0、1、2、3、4或5的整数。
在本发明的一个优选实施例方案中，所述的通式(I)所示的化合物，其为通式(II)所示的化合物、其立体异构体或其药学上可接受盐：
Figure PCTCN2017100678-appb-000002
其中：
B选自3-8元单环杂环基、6-12元螺环杂环基、6-12元稠环杂环基或6-12元桥环杂环基；优选3-8元单环杂环基；
L为键或-C(O)-；
R、R1、R2和x如权利要求(II)所述。
在本发明的一个优选实施例方案中，所述的通式(II)所示的化合物，其为通式(III)所示的化合物、其立体异构体或其药学上可接受盐：
Figure PCTCN2017100678-appb-000003
其中：环B、R、R1、R2和x如通式(II)所述。
在本发明的一个优选实施例方案中，所述的通式(II)所示的化合物，其为通式(IV)所示的化合物、其立体异构体或其药学上可接受盐：
Figure PCTCN2017100678-appb-000004
其中：环B、R、R1、R2和x如通式(II)所述。
在本发明的一个优选实施例方案中，其为通式(V)所示的化合物、其立体异构体或其药学上可接受盐：
Figure PCTCN2017100678-appb-000005
其中：
M为CR2R2、NR2或O；
R为氢原子或卤素，其中所述的卤素优选氟原子；
R1为烷基或卤素，其中所述的烷基为C1-6烷基，优选C1-3烷基；
R2相同或不同，且各自独立地选自氢原子、C1-6烷基、C1-6卤代烷基、C1-6烷氧基、C1-6氨基烷氧基、C1-6卤代烷氧基、卤素、氨基、氧代基、硝基、羟基、氰基、C3-8环烷基、3-8元杂环基、-(CH2)nOR3和-(CH2)nNR4R5，其中所述的C1-6烷基、C1-6卤代烷基、C1-6氨基烷氧基、C3-8环烷基和3-8元杂环基任选进一步被选自C1-6烷基、C1-6卤代烷基、卤素、氨基、氰基、羟基、烯基、炔基、C1-6烷氧基、C1-6卤代烷氧基、C1-6羟烷基、C3-8环烷基、3-8元杂环基、-(CH2)n-、-(CH2)nOR6和-(CH2)nNR7R8中的一个或多个取代基所取代；
或者两个R2相互连接，形成一个3-10元的环烷基或者杂环基，其中所述的3-8元的环烷基或者杂环基任选进一步被一个或者多个C1-6烷基、C1-6卤代烷基、C1-6烷氧基、C1-6氨基烷氧基、C1-6卤代烷氧基、卤素、氨基、氧代基、羟基、氰基和C3-8环烷基的取代基取代；优选形成5-8元环烷基或者杂环基；且
y为0、1、2或3的整数；
R3～R8、n和x如通式(I)所述。
在本发明的一个优选实施例方案中，其为通式(V)所示的化合物、其立体异构体或其药学上可接受盐：
其中：
M选自CHR2或NR2；
R2选自氢原子、C1-6烷基、C1-6卤代烷基、C1-6烷氧基、C1-6氨基烷氧基、C1-6卤代烷氧基、卤素、氨基、氧代基、羟基、氰基、C3-8环烷基、3-8元杂环基、-(CH2)nOR3和-(CH2)nNR4R5，其中所述的C1-6烷基、C1-6卤代烷基、C1-6氨基烷氧基、C3-8环烷基和3-8元杂环基任选进一步被选自C1-6烷基、C1-6卤代烷基、卤素、氨基、氰基、羟基、C2-6烯基、C2-6炔基、C1-6烷氧基、C1-6卤代烷氧基、C1-6羟烷基、C3-8环烷基、3-8元杂环基、-(CH2)n-、-(CH2)nOR6和-(CH2)nNR7R8 中的一个或多个取代基所取代；
R、R1、R3～R8、x、n和y如通式(V)所述。
在本发明的一个优选实施例方案中，其为通式(VI)所示的化合物、其立体异构体或其药学上可接受盐：
Figure PCTCN2017100678-appb-000006
其中：
R4和R5各自独立地选自氢原子、C1-6烷基、C1-6卤代烷基、C3-8环烷基、-(CH2)nOR6、-(CH2)nC(O)R6，其中所述的C1-6烷基、C1-6卤代烷基、C3-8环烷基任选进一步被选自C1-6烷基、卤素、羟基、氨基、氰基、C1-6烷氧基、C1-6羟烷基和C1-6环烷基中的一个或多个取代基所取代；
或者R4和R5形成一个3-8元的杂环基，其中所述的3-8元的杂环基任选进一步被选自C1-6烷基、-(CH2)n-、卤素、羟基、氨基、氰基、C1-6烷氧基、C1-6羟烷基和C1-6环烷基中的一个或多个取代基所取代；优选R4和R5形成的杂环基是4-6元；
R、R1和n如通式(V)所述。
在本发明的一个优选实施例方案中，所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R选自氢原子和卤素；卤素优选氟。
在本发明的一个优选实施例方案中，所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R1选自C1-8烷基和卤素，其中所述的C1-8烷基，优选C1-6烷基，更优选C1-3烷基；最优选甲基，其中所述的卤素优选氟。
在本发明的一个优选实施例方案中，所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R2选自氢原子、C1-8烷基、C1-8卤代烷基、C1-8烷氧基、C3-8环烷基、C2-6烯基、卤素、氧代基、-(CH2)nNR4R5和3-10杂环基，其中所述的C1-8烷基、C1-8卤代烷基、C1-8烷氧基、C3-8环烷基和3-10元杂环基任选进一步被选自卤素、羟基、氰基、C1-8烷基、-(CH2)nOR6和C1-8烷氧基中的一个或多个取代基所取代；优选C1-6烷基、C1-6卤代烷基、氧代基、-(CH2)nNR4R5和3-6杂环基，其中所述的C1-6烷基任选进一步被选自卤素、羟基和氰基中的一个或多个取代基所取代；所述的C1-6烷基更优选C1-3烷基。
在本发明的一个优选实施例方案中，所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R4和R5相同或不同，且各自独立地选自氢原子、C1-8烷基、C1-8卤代烷基、C3-8环烷基、-(CH2)nC(O)R6和C1-8烷氧基，其中所述的C1-8烷基、C1-8卤代烷基、C3-8环烷基和C1-8烷氧基任选进一步被选自卤素、羟基、氰基、C1-8烷基、C3-8环烷基、-(CH2)nOR6和C1-8烷氧基中的一个或多个取代基所取代；优选C1-6烷基和C3-6环烷基，其中所述的C1-6烷基和C3-6环烷基任选进一步被选自卤素、羟基、氰基、C1-6烷基、C3-6环烷基和-(CH2)nOR6中的一个或多个取代基所取代；其中所述的C1-6烷基更优选C1-3烷基；R6选自氢原子、C1-6烷基和C1-6烷氧基；优选C1-3烷基和C1-3烷氧基。
在本发明的一个优选实施例方案中，任一所示的通式(I)化合物、其立体异构体或其药学上可接受的盐的制备方法，包括如下步骤：
Figure PCTCN2017100678-appb-000007
通式化合物(V-A)和通式化合物(V-B)偶联后得到通式(I)化合物，该通式(I)化合物任选进一步反应，或进一步脱保护基得到不同的通式(I)化合物，其中偶联反应中催化试剂优选Pd2(dba)3和Xantphos试剂；
其中：
X为卤素；优选氯；
环A、L、R、R1、R2和x如通式(I)所述。。
本发明还涉及一种治疗预防和/或治疗预防CDK4/6介导的病理学特征的疾病的方法，其包括向患者施用治疗有效剂量的通式(I)所示的化合物其立体异构体或其药学上可接受的盐，或其药物组合物。
本发明进一步涉及通式(I)所示的化合物、其立体异构体或其药学上可接受的盐，或其药物组合物在制备用于治疗和/或预防由CDK激酶4和/或6介导的癌症或肿瘤相关疾病的药物中的应用。
本发明进一步涉及通式(I)所示的化合物、其立体异构体或其药学上可接受的盐，或其药物组合物在制备治疗癌症或肿瘤相关药物中的应用，其中所述癌症或肿瘤相关疾病选自脑瘤、肺癌、肝癌、胃癌、口腔癌、头颈癌、肠癌或直肠癌、结肠癌、肾癌、食管腺癌、食管鳞状细胞癌、鳞状上皮细胞癌、甲状腺癌、骨癌、皮肤癌、非小细胞肺癌、原位癌、淋巴瘤、神经纤维瘤、成神经细胞瘤、肥大细胞肿瘤、多发性骨髓瘤、黑色素瘤、胶质瘤、肉瘤或脂肪肉瘤、胶质母细胞瘤、膀胱癌、卵巢癌、腹膜癌、胰腺癌、乳腺癌、子宫癌、子宫颈癌、子宫内膜癌、 前列腺癌、雌性生殖道癌、睾丸癌、胃肠道间质瘤或前列腺肿瘤；优选自膀胱癌、卵巢癌、腹膜癌、胰腺癌、乳腺癌、子宫癌、子宫颈癌、子宫内膜癌、前列腺癌、雌性生殖道癌、睾丸癌、胃肠道间质瘤或前列腺肿瘤。
本发明进一步涉及通式(I)所示的化合物、其立体异构体或其药学上可接受的盐，或其药物组合物在制备治疗癌症或肿瘤相关疾病的方法，其包括向患者施用治疗有效剂量的通式(I)所示的化合物其立体异构体或其药学上可接受的盐，或其药物组合物，其中所述癌症或肿瘤相关疾病选自脑瘤、肺癌、肝癌、胃癌、口腔癌、头颈癌、肠癌或直肠癌、结肠癌、肾癌、食管腺癌、食管鳞状细胞癌、鳞状上皮细胞癌、甲状腺癌、骨癌、皮肤癌、非小细胞肺癌、原位癌、淋巴瘤、神经纤维瘤、成神经细胞瘤、肥大细胞肿瘤、多发性骨髓瘤、黑色素瘤、胶质瘤、肉瘤或脂肪肉瘤、胶质母细胞瘤、膀胱癌、卵巢癌、腹膜癌、胰腺癌、乳腺癌、子宫癌、子宫颈癌、子宫内膜癌、前列腺癌、雌性生殖道癌、睾丸癌、胃肠道间质瘤或前列腺肿瘤；优选自膀胱癌、卵巢癌、腹膜癌、胰腺癌、乳腺癌、子宫癌、子宫颈癌、子宫内膜癌、前列腺癌、雌性生殖道癌、睾丸癌、胃肠道间质瘤或前列腺肿瘤。
作为更进一步优选的制备治疗乳腺癌相关疾病的方法，所述乳腺癌包括：在绝经后女性雌激素受体阳性和/或人表皮生长因子受体2阴性的局部晚期或转移性乳腺癌。
本发明进一步涉及通式(I)所示的化合物、其立体异构体或其药学上可接受的盐，一种药用组合物，其包括治疗有效剂量的任一所示的通式(I)化合物、其立体异构体或其药学上可接受的盐以及一种或多种药学上可接受的载体、稀释剂或赋形剂。
发明的详细说明
除非有相反陈述，在说明书和权利要求书中使用的术语具有下述含义。
术语“烷基”指饱和脂肪族烃基团，其为包含1至20个碳原子的直链或支链基团，优选含有1至8个碳原子的烷基，更优选1至6个碳原子的烷基，最更优选1至3个碳原子的烷基。非限制性实例包括甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基、仲丁基、正戊基、1,1-二甲基丙基、1,2-二甲基丙基、2,2-二甲基丙基、1-乙基丙基、2-甲基丁基、3-甲基丁基、正己基、1-乙基-2-甲基丙基、1,1,2-三甲基丙基、1,1-二甲基丁基、1,2-二甲基丁基、2,2-二甲基丁基、1,3-二甲基丁基、2-乙基丁基、2-甲基戊基、3-甲基戊基、4-甲基戊基、2,3-二甲基丁基、正庚基、2-甲基己基、3-甲基己基、4-甲基己基、5-甲基己基、2,3-二甲基戊基、2,4-二甲基戊基、2,2-二甲基戊基、3,3-二甲基戊基、2-乙基戊基、3-乙基戊基、正辛基、2,3-二甲基己基、2,4-二甲基己基、2,5-二甲基己基、2,2-二甲基己基、3,3-二甲基己基、4,4-二甲基己基、2-乙基己基、3-乙基己基、4-乙基己基、2-甲基-2-乙基戊基、2-甲基-3-乙基戊基、正壬基、2-甲基-2-乙基己基、2-甲基-3- 乙基己基、2,2-二乙基戊基、正癸基、3,3-二乙基己基、2,2-二乙基己基，及其各种支链异构体等。更优选的是含有1至6个碳原子的低级烷基，非限制性实施例包括甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基、仲丁基、正戊基、1,1-二甲基丙基、1,2-二甲基丙基、2,2-二甲基丙基、1-乙基丙基、2-甲基丁基、3-甲基丁基、正己基、1-乙基-2-甲基丙基、1,1,2-三甲基丙基、1,1-二甲基丁基、1,2-二甲基丁基、2,2-二甲基丁基、1,3-二甲基丁基、2-乙基丁基、2-甲基戊基、3-甲基戊基、4-甲基戊基、2,3-二甲基丁基等。烷基可以是取代的或非取代的，当被取代时，取代基可以在任何可使用的连接点上被取代，所述取代基优选为一个或多个以下基团，其独立地选自烷基、烯基、炔基、烷氧基、烷硫基、烷基氨基、卤素、巯基、羟基、硝基、氰基、环烷基、杂环烷基、芳基、杂芳基、环烷氧基、杂环烷氧基、环烷硫基、杂环烷硫基、氧代基、羧基或羧酸酯基，本发明优选甲基、乙基、异丙基、叔丁基、卤代烷基、氘代烷基、烷氧基取代的烷基和羟基取代的烷基。
术语“亚烷基”是指烷基的一个氢原子进一步被取代，例如：“亚甲基”指-CH2-、“亚乙基”指-(CH2)2-、“亚丙基”指-(CH2)3-、“亚丁基”指-(CH2)4-等。术语“烯基”指由至少由两个碳原子和至少一个碳-碳双键组成的如上定义的烷基，例如乙烯基、1-丙烯基、2-丙烯基、1-、2-或3-丁烯基等。烯基可以是取代的或非取代的，当被取代时，取代基优选为一个或多个以下基团，其独立地选自烷基、烯基、炔基、烷氧基、烷硫基、烷基氨基、卤素、巯基、羟基、硝基、氰基、环烷基、杂环烷基、芳基、杂芳基、环烷氧基、杂环烷氧基、环烷硫基、杂环烷硫基。
术语“环烷基”指饱和或部分不饱和单环或多环环状烃取代基，环烷基环包含3至20个碳原子，优选包含3至12个碳原子，更优选包含3至8个碳原子，最优选包含3至6个碳原子。单环环烷基的非限制性实例包括环丙基、环丁基、环戊基、环戊烯基、环己基、环己烯基、环己二烯基、环庚基、环庚三烯基、环辛基等；多环环烷基包括螺环、稠环和桥环的环烷基，优选环丙基、环丁基、环己基、环戊基和环庚基。
术语“螺环烷基”指5至20元的单环之间共用一个碳原子(称螺原子)的多环基团，其可以含有一个或多个双键，但没有一个环具有完全共轭的π电子系统。优选为6至14元，更优选为7至10元。根据环与环之间共用螺原子的数目将螺环烷基分为单螺环烷基、双螺环烷基或多螺环烷基，优选为单螺环烷基和双螺环烷基。更优选为4元/4元、4元/5元、4元/6元、5元/5元或5元/6元单螺环烷基。螺环烷基的非限制性实例包括：
Figure PCTCN2017100678-appb-000008
也包含单螺环烷基与杂环烷基共用螺原子的螺环烷基，非限制性实例包括：
Figure PCTCN2017100678-appb-000009
术语“稠环烷基”指5至20元，系统中的每个环与体系中的其他环共享毗邻的一对碳原子的全碳多环基团，其中一个或多个环可以含有一个或多个双键，但没有一个环具有完全共轭的π电子系统。优选为6至14元，更优选为7至10元。根据组成环的数目可以分为双环、三环、四环或多环稠环烷基，优选为双环或三环，更优选为5元/5元或5元/6元双环烷基。稠环烷基的非限制性实例包括：
Figure PCTCN2017100678-appb-000010
术语“桥环烷基”指5至20元，任意两个环共用两个不直接连接的碳原子的全碳多环基团，其可以含有一个或多个双键，但没有一个环具有完全共轭的π电子系统。优选为6至14元，更优选为7至10元。根据组成环的数目可以分为双环、三环、四环或多环桥环烷基，优选为双环、三环或四环，更有选为双环或三环。桥环烷基的非限制性实例包括：
Figure PCTCN2017100678-appb-000011
所述环烷基环可以稠合于芳基、杂芳基或杂环烷基环上，其中与母体结构连接在一起的环为环烷基，非限制性实例包括茚满基、四氢萘基、苯并环庚烷基等。环烷基可以是任选取代的或非取代的，当被取代时，取代基优选为一个或多个以下基团，其独立地选自烷基、烯基、炔基、烷氧基、烷硫基、烷基氨基、卤素、巯基、羟基、硝基、氰基、环烷基、杂环烷基、芳基、杂芳基、环烷氧基、杂环烷氧基、环烷硫基、杂环烷硫基、氧代基、羧基或羧酸酯基。
术语“杂环基”指饱和或部分不饱和单环或多环环状烃取代基，其包含3至20个环原子，其中一个或多个环原子为选自氮、氧或S(O)m(其中m是整数0至 2)的杂原子，但不包括-O-O-、-O-S-或-S-S-的环部分，其余环原子为碳。优选包含3至12个环原子，其中1～4个是杂原子；进一步优选包含3至10个环原子；更优选包含3至8个环原子；最优选包含6至10个环原子。单环杂环基的非限制性实例包括吡咯烷基、咪唑烷基、四氢呋喃基、四氢噻吩基、二氢咪唑基、二氢呋喃基、二氢吡唑基、二氢吡咯基、哌啶基、哌嗪基、吗啉基、硫代吗啉基、高哌嗪基、吡喃基等，优选哌啶基和
Figure PCTCN2017100678-appb-000012
多环杂环基包括螺环、稠环和桥环的杂环基。术语“螺杂环基”指5至20元的单环之间共用一个原子(称螺原子)的多环杂环基团，其中一个或多个环原子为选自氮、氧或S(O)m(其中m是整数0至2)的杂原子，其余环原子为碳。其可以含有一个或多个双键，但没有一个环具有完全共轭的π电子系统。优选为6至14元，更优选为6至10元。根据环与环之间共用螺原子的数目将螺杂环基分为单螺杂环基、双螺杂环基或多螺杂环基，优选为单螺杂环基和双螺杂环基。更优选为3元/6元、4元/4元、4元/5元、4元/6元、5元/5元或5元/6元单螺杂环基。螺杂环基的非限制性实例包括：
Figure PCTCN2017100678-appb-000013
术语“稠杂环基”指5至20元，系统中的每个环与体系中的其他环共享毗邻的一对原子的多环杂环基团，一个或多个环可以含有一个或多个双键，但没有一个环具有完全共轭的π电子系统，其中一个或多个环原子为选自氮、氧或S(O)m(其中m是整数0至2)的杂原子，其余环原子为碳。优选为6至14元，更优选为6至10元。根据组成环的数目可以分为双环、三环、四环或多环稠杂环基，优选为双环或三环，更优选为5元/5元或5元/6元双环稠杂环基。稠杂环基的非限制性实例包括：
Figure PCTCN2017100678-appb-000014
术语“桥杂环基”指5至14元，任意两个环共用两个不直接连接的原子的多环杂环基团，其可以含有一个或多个双键，但没有一个环具有完全共轭的π电 子系统，其中一个或多个环原子为选自氮、氧或S(O)m(其中m是整数0至2)的杂原子，其余环原子为碳。优选为6至14元，更优选为6至10元。根据组成环的数目可以分为双环、三环、四环或多环桥杂环基，优选为双环、三环或四环，更有选为双环或三环。桥杂环基的非限制性实例包括：
Figure PCTCN2017100678-appb-000015
所述杂环基环可以稠合于芳基、杂芳基或环烷基环上，其中与母体结构连接在一起的环为杂环基，其非限制性实例包括：
Figure PCTCN2017100678-appb-000016
杂环基可以是任选取代的或非取代的，当被取代时，取代基优选为一个或多个以下基团，其独立地选自烷基、烯基、炔基、烷氧基、烷硫基、烷基氨基、卤素、巯基、羟基、硝基、氰基、环烷基、杂环烷基、芳基、杂芳基、环烷氧基、杂环烷氧基、环烷硫基、杂环烷硫基、氧代基、羧基或羧酸酯基。
术语“芳基”指具有共轭的π电子体系的6至14元全碳单环或稠合多环(也就是共享毗邻碳原子对的环)基团，优选为6至10元，例如苯基和萘基。更优选苯基。所述芳基环可以稠合于杂芳基、杂环基或环烷基环上，其中与母体结构连接在一起的环为芳基环，其非限制性实例包括：
Figure PCTCN2017100678-appb-000017
芳基可以是取代的或非取代的，当被取代时，取代基优选为一个或多个以下基团，其独立地选自烷基、烯基、炔基、烷氧基、烷硫基、烷基氨基、卤素、巯基、羟基、硝基、氰基、环烷基、杂环烷基、芳基、杂芳基、环烷氧基、杂环烷氧基、环烷硫基、杂环烷硫基、羧基或羧酸酯基。
术语“杂芳基”指包含1至4个杂原子、5至14个环原子的杂芳族体系，其中杂原子选自氧、硫和氮。杂芳基优选为5至10元，更优选为5元或6元，例如咪唑基、呋喃基、噻吩基、噻唑基、吡唑基、噁唑基、吡咯基、三唑基、四 唑基、吡啶基、嘧啶基、噻二唑、吡嗪基等，优选为三唑基、噻吩基、咪唑基、吡唑基或嘧啶基、噻唑基；更有选三唑基、吡咯基、噻吩基、噻唑基和嘧啶基。所述杂芳基环可以稠合于芳基、杂环基或环烷基环上，其中与母体结构连接在一起的环为杂芳基环，其非限制性实例包括：
Figure PCTCN2017100678-appb-000018
杂芳基可以是任选取代的或非取代的，当被取代时，取代基优选为一个或多个以下基团，其独立地选自烷基、烯基、炔基、烷氧基、烷硫基、烷基氨基、卤素、巯基、羟基、硝基、氰基、环烷基、杂环烷基、芳基、杂芳基、环烷氧基、杂环烷氧基、环烷硫基、杂环烷硫基、羧基或羧酸酯基。
术语“烷氧基”指-O-(烷基)和-O-(非取代的环烷基)，其中烷基的定义如上所述。烷氧基的非限制性实例包括：甲氧基、乙氧基、丙氧基、丁氧基、环丙氧基、环丁氧基、环戊氧基、环己氧基。烷氧基可以是任选取代的或非取代的，当被取代时，取代基优选为一个或多个以下基团，其独立地选自烷基、烯基、炔基、烷氧基、烷硫基、烷基氨基、卤素、巯基、羟基、硝基、氰基、环烷基、杂环烷基、芳基、杂芳基、环烷氧基、杂环烷氧基、环烷硫基、杂环烷硫基、羧基或羧酸酯基。
“卤代烷基”指被一个或多个卤素取代的烷基，其中烷基如上所定义。
“卤代烷氧基”指被一个或多个卤素取代的烷氧基，其中烷氧基如上所定义。
“羟烷基”指被羟基取代的烷基，其中烷基如上所定义。
“羟基”指-OH基团。
“卤素”指氟、氯、溴或碘。
“氨基”指-NH2。
“氰基”指-CN。
“硝基”指-NO2。
“羧基”指-C(O)OH。
“氧代基”指
Figure PCTCN2017100678-appb-000019
例如氧代哌啶基，
Figure PCTCN2017100678-appb-000020
“THF”指四氢呋喃。
“EtOAc”指乙酸乙酯。
“MeOH”指甲醇。
“DMF”指N、N-二甲基甲酰胺。
“DIPEA”指二异丙基乙胺。
“TFA”指三氟乙酸。
“MeCN”指乙晴。
“DMA”指N,N-二甲基乙酰胺。
“Et2O”指乙醚。
“DCE”指1,2二氯乙烷。
“DIPEA”指N，N-二异丙基乙胺。
“NBS”指N-溴代琥珀酰亚胺。
“NIS”指N-碘代丁二酰亚胺。
“Cbz-Cl”指氯甲酸苄酯。
“Pd2(dba)3”指三(二亚苄基丙酮)二钯。
“Dppf”指1,1’-双二苯基膦二茂铁。
“HATU”指2-(7-氧化苯并三氮唑)-N,N,N’,N’-四甲基脲六氟磷酸酯。
“KHMDS”指六甲基二硅基胺基钾。
“LiHMDS”指双三甲基硅基胺基锂。
“MeLi”指甲基锂。
“n-BuLi”指正丁基锂。
“NaBH(OAc)3”指三乙酰氧基硼氢化钠。
“X选自A、B、或C”、“X选自A、B和C”、“X为A、B或C”、“X为A、B和C”等不同用语均表达了相同的意义，即表示X可以是A、B、C中的任意一种或几种。
“立体异构”包含几何异构(顺反异构)、旋光异构、构象异构三类。
本发明所述的氢原子均可被其同位素氘所取代，本发明涉及的实施例化合物中的任一氢原子也均可被氘原子取代。
“任选”或“任选地”意味着随后所描述的事件或环境可以但不必发生，该说明包括该事件或环境发生或不发生的场合。例如，“任选被烷基取代的杂环基团”意味着烷基可以但不必须存在，该说明包括杂环基团被烷基取代的情形和杂环基团不被烷基取代的情形。
“取代的”指基团中的一个或多个氢原子，优选为最多5个，更优选为1～3个氢原子彼此独立地被相应数目的取代基取代。不言而喻，取代基仅处在它们的可能的化学位置，本领域技术人员能够在不付出过多努力的情况下确定(通过实验或理论)可能或不可能的取代。例如，具有游离氢的氨基或羟基与具有不饱和(如烯属)键的碳原子结合时可能是不稳定的。
“药物组合物”表示含有一种或多种本文所述化合物或其生理学上/可药用的盐或前体药物与其他化学组分的混合物，以及其他组分例如生理学/可药用的载 体和赋形剂。药物组合物的目的是促进对生物体的给药，利于活性成分的吸收进而发挥生物活性。
“可药用盐”是指本发明化合物的盐，这类盐用于哺乳动物体内时具有安全性和有效性，且具有应有的生物活性。
具体实施方式
以下结合实施例进一步描述本发明，但这些实施例并非限制着本发明的范围。
实施例
本发明的化合物结构是通过核磁共振(NMR)或/和液质联用色谱(LC-MS)来确定的。NMR化学位移(δ)以百万分之一(ppm)的单位给出。NMR的测定是用Bruker AVANCE-400核磁仪，测定溶剂为氘代二甲基亚砜(DMSO-d6)，氘代甲醇(CD3OD)和氘代氯仿(CDCl3)，内标为四甲基硅烷(TMS)。
液质联用色谱LC-MS的测定用Agilent 1200 Infinity Series质谱仪。HPLC的测定使用安捷伦1200DAD高压液相色谱仪(Sunfire C18 150×4.6mm色谱柱)和Waters 2695-2996高压液相色谱仪(Gimini C18 150×4.6mm色谱柱)。
薄层层析硅胶板使用烟台黄海HSGF254或青岛GF254硅胶板，TLC采用的规格是0.15mm～0.20mm，薄层层析分离纯化产品采用的规格是0.4mm～0.5mm。柱层析一般使用烟台黄海硅胶200～300目硅胶为载体。
本发明实施例中的起始原料是已知的并且可以在市场上买到，或者可以采用或按照本领域已知的方法来合成。
在无特殊说明的情况下，本发明的所有反应均在连续的磁力搅拌下，在干燥氮气或氩气氛下进行，溶剂为干燥溶剂，反应温度单位为摄氏度。
实施例1
1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000021
第一步：叔-丁基4-(6-氯-2-甲基尼古丁酰)-1,4-重氮基庚环-1-羧酸酯的制备
Figure PCTCN2017100678-appb-000022
将6-氯-2-甲基尼古丁酸(0.7g,4.1mmol)，叔-丁基1,4-重氮基庚环-1-羧酸 酯(1.0g,4.9mmol)，TEA(1.2g,12.2mmol)溶于CH2Cl2(15mL)中，加入HATU(1.87g,4.9mmol)，室温搅拌反应两小时。反应液用CH2Cl2(30mL)稀释后用NaHCO3溶液(30mL),饱和食盐水(30mL)洗涤，无水硫酸钠干燥，浓缩后柱层析[洗脱剂:CH2Cl2～CH2Cl2/MeOH(10:1)]得产品叔-丁基4-(6-氯-2-甲基尼古丁酰)-1,4-重氮基庚环-1-羧酸酯(1.4g,产率96％)。
MS m/z(ESI):354.1[M+H]+。
第二步：5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-胺的制备
Figure PCTCN2017100678-appb-000023
4-氟-1-异丙基-2-甲基-6-(4,4,5,5-四甲基-1,3,2-二噁硼戊环-2-基)-1H-苯并[d]咪唑(2.0g,6.3mmol)、4-氯-5-氟嘧啶-2-胺(0.93g,6.3mmol)、Pd(dppf)Cl2(0.4g,0.6mmol)、碳酸钾(2.61g,18.9mmol)于二氧六环/H2O(4/1，50mL)的混合溶液中氮气保护下回流5h，冷却至室温浓缩除去有机溶剂，加水稀释，CH2Cl2(30mL*3)萃取，合并有机相并用无水硫酸钠干燥，浓缩后柱层析得到产物5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-胺(1.8g,产率94.4％)。
MS m/z(ESI):304.1[M+H]+。
第三步：叔-丁基4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-1,4-重氮基庚环-1-羧酸酯的制备
Figure PCTCN2017100678-appb-000024
化合物5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-胺(1.2g，4.0mmol)和化合物叔-丁基4-(6-氯-2-甲基尼古丁酰)-1,4-重氮基庚环-1-羧酸酯(1.7g,4.8mmol)溶于二氧六环(40mL)，然后依次加入碳酸铯(3.9g,11.9mmol)，Pd2(dba)3(0.2g，0.22mmol)，Xantphos(0.24g,0.44mmol)，置换氮气三次，搅拌回流过夜。冷却过滤，浓缩，柱层析分离[DCM～DCM/MeOH(10:1)]得到化合物叔-丁基4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-1,4-重氮基庚环-1-羧酸酯(2g,产率81％)。
MS m/z(ESI):621.3[M+H]+。
第四步：1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000025
叔-丁基-4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-1,4-重氮基庚环-1-羧酸酯(2g,3.2mmol)溶于CH2Cl2(30mL)，冷却至0℃，滴加三氟乙酸(10mL)，滴加完毕后室温搅拌2h，浓缩后的粗产物(1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的三氟乙酸盐(4g,产率99％)，粗产物无需进一步纯化可直接用于下一步反应。
1H NMR(400MHz,MeOD)δ:8.98(d,J＝3.2Hz,1H),8.61(s,1H),8.36(d,J＝9.0Hz,1H),8.24(d,J＝11.1Hz,1H),7.61(d,J＝8.8Hz,1H),5.18(dt,J＝13.8,7.1Hz,1H),4.07-3.84(m,2H),3.62-3.52(m,3H),3.43(m,3H),3.00(s,3H),2.78(s,3H),2.21(d,J＝43.5Hz,2H),1.84(d,J＝6.9Hz,6H).
19F NMR(376MHz,MeOD)δ:-129.2,-143.0.
MS m/z(ESI):521.2[M+H]+.
实施例2
1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-4-(甲基氨基)哌啶-2-酮的制备
Figure PCTCN2017100678-appb-000026
第一步：(1-(6-氯-2-甲基吡啶-3-基)-2-氧代哌啶-4-基)氨基甲酸叔丁酯的制备
Figure PCTCN2017100678-appb-000027
3-溴-6-氯-2-甲基吡啶(200mg,0.969mmol)、(2-氧代哌啶-4-基)氨基甲酸叔丁酯(249mg,1.162mmol)、Pd2(dba)3(89mg,0.0972mmol)、Xantphos(112mg, 0.194mmol)、碳酸铯(947mg,2.907mmol)于二氧六环(10mL)中100℃氮气保护下搅拌过夜，冷却，浓缩后柱层析得到产物(31mg,收率9.4％)。
MS m/z(ESI):340.1,342.1[M+H]+。
第二步：(1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-2-羰基哌啶-4-基)氨基甲酸叔丁酯的制备
Figure PCTCN2017100678-appb-000028
5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-胺(28mg,0.0923mmol)、(1-(6-氯-2-甲基吡啶-3-基)-2-氧代哌啶-4-基)氨基甲酸叔丁酯(31mg,0.0912mmol)、Pd2(dba)3(8.4mg,0.00917mmol)、Xantphos(10.6mg,0.0183mmol)、碳酸铯(89mg,0.273mmol)于二氧六环(10mL)中100℃氮气保护下搅拌5h，冷却，浓缩后柱层析得到产物(19.6mg,收率35.0％)。
MS m/z(ESI):607.2[M+H]+。
第三步：(1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-2-羰基哌啶-4-基)(甲基)氨基甲酸叔丁酯的制备
Figure PCTCN2017100678-appb-000029
(1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-2-羰基哌啶-4-基)氨基甲酸叔丁酯(19.6mg,0.0323mmol)、60％NaH(1.6mg,0.0400mmol)于THF(2mL)中搅拌，加入碘甲烷(9mg,0.0634mmol)，室温搅拌过夜，浓缩后柱层析得到产物(12mg,产率59.8％)。
MS m/z(ESI):621.3[M+H]+。
第四步：1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-4-(甲基氨基)哌啶-2-酮的制备
Figure PCTCN2017100678-appb-000030
上一步产物(12mg,0.0193mmol)溶于二氯甲烷(2mL)，加入三氟乙酸(1mL)，室温搅拌3h，浓缩至干，柱层析分离得到产物(9.1mg,产率90.4％)。
1H NMR(400MHz,CD3OD)δ:8.93(s,1H),8.53(d,J＝33.8Hz,2H),8.21(s,1H),7.91(s,1H),5.19(s,1H),4.01(s,4H),3.73(s,2H),3.02(s,3H),2.77(s,1H),2.68(s,3H),2.38(d,J＝87.2Hz,3H),1.84(s,6H).
MS m/z(ESI):521.2[M+H]+。
实施例3
(4-(环丙基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000031
第一步：1-(6-氯-2-甲基烟酰)哌啶-4-酮的制备
Figure PCTCN2017100678-appb-000032
将6-氯-2-甲基烟酸(1.0g,5.8mmol)，哌啶-4-酮(866mg,8.7mmol),HATU(2.2g,17.5mmol),DIEA(2mL)依次加入二氯甲烷(50mL)中。反应在室温下搅拌4小时，LCMS显示反应结束，反应液加入二氯甲烷(50mL)和水(50mL)分液，有机相用饱和碳酸氢钠(3x20mL)洗涤，有机相分液,用无水硫酸钠干燥，过滤浓缩。剩余粗产品通过快速硅胶柱纯化(CH2Cl2:MeOH＝20:1)得到产品1-(6-氯-2-甲基烟酰)哌啶-4-酮(1.2g,81％)。
第二步：5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-胺的制备
Figure PCTCN2017100678-appb-000033
将4-氯-5-氟嘧啶-2-胺(4.5g,31.3mmol),4-氟-1-异丙基-2-甲基-6-(4,4,5,5-四甲基-1,3,2-二噁硼戊环-2-基)-1H-苯并[d]咪唑(10.0g,31.3mmol),Pd(dppf)Cl2(200mg),碳酸钾(8.6g,62.6mmol)依次加入二氧六环(50mL)和水(5mL)中。反应在氮气保护100℃下搅拌3小时，反应结束后，反应液浓缩，剩余物加入二氯甲烷(50mL)和水(50mL)。分液，有机相用无水硫酸钠干燥，过滤浓缩。剩余粗产品通过快速硅胶柱纯化得到产品5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧 啶-2-胺(5.0g,52％)。
MS m/z(ESI):304.1[M+H]+
第三步：1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基烟酰)哌啶-4-酮的制备
Figure PCTCN2017100678-appb-000034
将5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-胺(200mg,0.66mmol)，1-(6-氯-2-甲基烟酰)哌啶-4-酮(167mg,0.66mmol),Pd2(dba)3(50mg),Xan-phos(70mg),碳酸铯(430mg,1.32mmol)依次加入无水二氧六环(5mL)中。反应在氮气保护110℃下搅拌4小时，反应结束后，反应液浓缩，剩余物加入二氯甲烷(10mL)和水(10mL)。分液，有机相用无水硫酸钠干燥，过滤浓缩。剩余粗产品通过快速硅胶柱纯化得到产品1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基烟酰)哌啶-4-酮(100mg,30％).
MS m/z(ESI):520.2[M+H]+
第四步：(4-(环丙基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000035
将1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基烟酰)哌啶-4-酮(100mg,0.19mmol)，环丙胺(0.3mL)溶于二氯甲烷(5mL)中，反应搅拌在室温下20分钟后，醋酸硼氢化钠(200mg,0.95mmol)加入反应液中，反应在室温下搅拌1小时，LCMS显示反应完全，将反应液直接旋干，剩余物用快速硅胶柱纯化，再经制备HPLC纯化得到产品(4-(环丙基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮(24.0mg,22％)。
1H NMR(400MHz,MeOD)δ：8.95(d,J＝3.1Hz,1H),8.58(s,1H),8.20(d,J＝10.7Hz,2H),7.65(d,J＝8.8Hz,1H),5.24–5.09(m,2H),3.77(s,1H),3.63(s,1H),3.02(d,J＝13.7Hz,1H),2.97(s,3H),2.84(d,J＝4.2Hz,1H),2.73(s,3H),2.35(s,1H),2.31–2.13(m,2H),2.06(s,1H),1.83(d,J＝6.9Hz,6H),1.73(s,2H),1.33(d,J＝18.5Hz,2H),1.04–0.89(m,2H).
MS m/z(ESI):561.2[M+H]+
实施例4
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(9-甲基-3,9-二氮杂二环[4.2.1]壬烷-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000036
第一步：9-苯甲基3-(叔-丁基)3,9-二氮杂二环[4.2.1]壬烷-3,9-二羧酸酯的制备
Figure PCTCN2017100678-appb-000037
叔-丁基3,9-二氮杂二环[4.2.1]壬烷-3-羧酸酯(130mg,0.57mmol)与三乙胺(0.3mL)溶于二氯甲烷(10mL)，冰浴下滴加氯甲酸苄酯(0.16mL,1.14mmol)。室温搅拌过夜。然后用二氯甲烷(30mL)稀释，依次用水(20mL)和饱和食盐水(20mL)洗涤，并用无水硫酸钠干燥，浓缩后柱层析[洗脱剂:PE～PE/EtOAc(1:1)]得到化合物9-苯甲基3-(叔-丁基)3,9-二氮杂二环[4.2.1]壬烷-3,9-二羧酸酯(170mg,产率82％)。
MS m/z(ESI):305.0[M-55]+。
第二步：苯甲基-3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯的制备
Figure PCTCN2017100678-appb-000038
化合物9-苯甲基3-(叔-丁基)3,9-二氮杂二环[4.2.1]壬烷-3,9-二羧酸酯(170mg，0.47mmol)溶于二氯甲烷(3mL),滴加三氟乙酸(2mL)，室温搅拌2h。浓缩待用。
MS m/z(ESI):261.1[M+H]+。
第三步：苯甲基3-(6-氯-2-甲基尼古丁酰)-3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯的制备
Figure PCTCN2017100678-appb-000039
化合物6-氯-2-甲基尼古丁酸(85mg，0.5mmol)和化合物苯甲基3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯溶于DMF(5mL),加DIEA(0.3mL)和HATU(380mg，1mmol)，室温搅拌2h。浓缩后柱层析纯化[洗脱剂:DCM～DCM/MeOH(10:1)]得化合物苯甲基3-(6-氯-2-甲基尼古丁酰)-3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯(180m g,产率94％)。
MS m/z(ESI):414.1[M+H]+。
第四步：苯甲基3-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯的制备
Figure PCTCN2017100678-appb-000040
化合物苯甲基3-(6-氯-2-甲基尼古丁酰)-3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯(100mg，0.24mmol)和化合物5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-胺(75mg，0.25mmol)溶于二氧六环(15mL)，然后依次加入碳酸铯(324mg,1mmol)，Pd2(dba)3(23mg，0.025mmol)与Xant-phos(30mg,0.05mmol)，置换氮气三次，回流过夜。冷却过滤,浓缩后柱层析纯化[DCM～DCM/MeOH(10:1)]得到化合物苯甲基3-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯(120mg,产率73％)。
MS m/z(ESI):681.2[M+H]+。
第五步：(3,9-二氮杂二环[4.2.1]壬烷-3-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000041
化合物苯甲基3-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-3,9-二氮杂二环[4.2.1]壬烷-9-羧酸酯(120mg,0.17mmol)溶于甲醇(10mL)，氮气保护下加入氢氧化钯(30mg)，然后置换氢气三次，室温搅拌过夜。过滤，浓缩得粗产品(90mg,产率93％)
MS m/z(ESI):547.2[M+H]+。
第六步：(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(9-甲基-3,9-二氮杂二环[4.2.1]壬烷-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000042
化合物(3,9-二氮杂二环[4.2.1]壬烷-3-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮(90mg,0.16mmol)和甲醛溶液(0.1mL)溶于二氯甲烷(5mL)，室温搅拌30min，然后加入三乙酰氧基硼氢化钠(100mg)，继续搅拌3h。反应液浓缩后经反相柱层析纯化得到化合物(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(9-甲基-3,9-二氮杂二环[4.2.1]壬烷-3-基)甲酮(56mg,产率50％)。
1H NMR(400MHz,MeOD)δ：9.00(s,1H),8.64(s,1H),8.50(s,1H),8.26(d,J＝10.6Hz,1H),7.63(s,1H),5.21(m,1H),4.20(m,4H),3.76(m,2H),3.02(m,6H),2.80(d,J＝12.8Hz,3H),2.65(m,2H),2.43–2.20(m,2H),2.19–1.95(m,2H),1.85(d,J＝6.0Hz,6H).
19F NMR(376MHz,MeOD)δ-128.9,-142.6
MS m/z(ESI):561.2[M+H]+。
实施例5
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(八氢-5H-吡咯并[3,2-c]吡啶-5-基)甲酮的制备
Figure PCTCN2017100678-appb-000043
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(八氢-5H-吡咯并[3,2-c]吡啶-5-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)):δ:9.00(s,1H),8.63(s,1H),8.38-8.22(m,2H),7.58(d,J＝8.4Hz,1H),5.25-5.15(m,1H),4.30-4.10(m,1H),4.05-3.90(m,1H),3.75-3.35(m,5H),3.02(s,3H),2.78(s,3H),2.75-2.55(m,1H),2.35-1.90(m,4H),1.85(d,J＝6.7Hz,6H).
19F NMR(400MHz,MeOD):δ:-129.16(s),-142.76(d,J＝12.8Hz).
MS m/z(ESI):547.2[M+H]+。
实施例6
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-氟吡啶-3-基)(八氢-5H-吡咯并[3,2-c]吡啶-5-基)甲酮的制备
Figure PCTCN2017100678-appb-000044
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-氟吡啶-3-基)(八氢-5H-吡咯并[3,2-c]吡啶-5-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)):δ:8.72(d,J＝3.4Hz,1H),8.60(s,1H),8.37(d,J＝8.1Hz,1H),8.00-7.90(s,1H),5.20-5.10(m,1H),4.30-4.17(m,1H),4.02-3.93(m,1H),3.79-3.38(m,5H),2.99(s,3H),2.75-2.45(m,1H),2.35-1.87(m,4H),1.82(d,J＝8Hz,6H).
19F NMR(400MHz,MeOD):δ:-129.68(d,J＝4.4Hz),-148.65(s).
MS m/z(ESI):551.2[M+H]+。
实施例7
(1,4-重氮基庚环-1-基)(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000045
(1,4-重氮基庚环-1-基)(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.59(dd,J＝9.5,3.3Hz,1H),8.49(s,1H),8.25(d,J＝8.2Hz,1H),8.07(d,J＝11.3Hz,1H),7.92(t,J＝8.9Hz,1H),5.04(dd,J＝13.7,6.9Hz,1H),3.91(s,1H),3.77(d,J＝19.3Hz,1H),3.58–3.45(m,2H),3.42–3.24(m,4H),2.89(s,3H),2.10(d,J＝24.5Hz,2H),1.73(d,J＝6.9Hz,6H).
19F NMR(376MHz,MeOD)δ:-70.3,-129.5,-148.5.
MS m/z(ESI):525.2[M+H]+.
实施例8
(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)(4-甲基-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000046
(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)(4-甲基-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.72(d,J＝3.3Hz,1H),8.61(s,1H),8.36(d,J＝8.4Hz,1H),8.20(d,J＝11.3Hz,1H),8.04(t,J＝8.9Hz,1H),5.17(dt,J＝14.0,6.9Hz,1H),4.40–4.31(m,1H),4.01–3.55(m,5H),3.46–3.36(m,2H),2.99(d,J＝10.6Hz,6H),2.29(m,2H),1.85(d,J＝6.9Hz,6H).
MS m/z(ESI):539.2[M+H]+.
实施例9
(S)-(3,4-二甲基-1,4-重氮基庚环-1-基)(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000047
(S)-(3,4-二甲基-1,4-重氮基庚环-1-基)(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ：8.62(d,J＝3.4Hz,1H),8.50(d,J＝0.9Hz,1H),8.26(dd,J＝8.3,2.0Hz,1H),8.09(d,J＝11.3Hz,1H),7.92(t,J＝8.8Hz,1H),5.13–4.97(m,1H),4.07(d,J＝15.5Hz,1H),3.82(d,J＝17.3Hz,5H),2.93(s,2H),2.89(s,3H),2.79(d,J＝18.3Hz,1H),2.14(d,J＝31.4Hz,2H),1.74(d,J＝6.9Hz,6H),1.39(dd,J＝47.3,6.8Hz,3H).
MS m/z(ESI):472.2[M+H]+.
实施例10
(4,5-二甲基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000048
(4,5-二甲基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例1。
MS m/z(ESI):472.2[M+H]+.
实施例11
(4-乙基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000049
(4-乙基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.87(s,1H),8.50(s,1H),8.30(m,1H),8.13(d,J＝10.9Hz,1H),7.49(d,J＝8.2Hz,1H),5.13–5.01(m,1H),4.23(d,J＝11.8Hz,1H),3.85–3.40(m,5H),3.235-3.25(m,4H),2.89(s,3H),2.70(m,3H),2.25-2.10(m,2H),1.73(d,J＝6.7Hz,6H),1.32(m,3H).
19F NMR(376MHz,MeOD)δ:-129.1,-142.8.
MS m/z(ESI):549.2[M+H]+.
实施例12
(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)(6-甲基-3,6-二氮杂二环[3.2.1]辛烷-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000050
(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)(6-甲基-3,6-二氮杂二环[3.2.1]辛烷-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD):δ:8.62–8.54(m,1H),8.46(s,1H),8.22(d,J＝8.2Hz,1H),8.05(d,J＝11.2Hz,1H),7.94-7.78(m,1H),5.05(dt,J＝13.6,6.8Hz,1H),4.68-4.39(m,1H),4.05-3.90(m,1H),3.91-3.40(m,2H),3.35-3.05(m,2H),2.95-2.80(m,6H),2.75-2.57(m,1H),2.45-2.30(m,1H),2.23-1.98(m,2H),1.73(d,J＝6.8Hz,6H).
19F NMR(400MHz,MeOD):δ:-69.67(d,J＝282.4Hz),-129.43(d,J＝8.9Hz),-148.33(s).
MS m/z(ESI):551.2[M+H]+。
实施例13
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(1-甲基八氢-5H-吡咯并[3,2-c]吡啶-5-基)甲酮的制备
Figure PCTCN2017100678-appb-000051
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2- 甲基吡啶-3-基)(1-甲基八氢-5H-吡咯并[3,2-c]吡啶-5-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD):δ:8.88(s,1H),8.51(s,1H),8.25-8.10(m,2H),7.52-7.44(m,1H),5.20-4.96(m,1H),4.70-4.50(m,1H),4.04-3.46(m,5H),3.40-3.30(m,1H),3.20-3.01(m,1H),2.96-2.80(m,5H),2.78-2.48(m,4H),2.35-1.9(dd,m,3H),1.92-1.51(m,7H).
19F NMR(400MHz,MeOD):δ:-129.11(s),-142.77(d,J＝26.3Hz).
MS m/z(ESI):561.2[M+H]+。
实施例14
(3,4-二甲基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000052
(3,4-二甲基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ：9.00(s,1H),8.63(s,1H),8.42(s,1H),8.26(s,1H),7.60(s,1H),5.20(s,1H),4.34–3.77(m,3H),3.63(s,3H),3.02(s,5H),2.92(s,1H),2.82(s,3H),2.32(d,J＝55.7Hz,3H),1.85(s,6H),1.60(s,2H),1.31(s,3H).
MS m/z(ESI):472.2[M+H]+.
实施例15
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(1-甲基哌啶-4-基)-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000053
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(1-甲基哌啶-4-基)-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.99(s,1H),8.63(s,1H),8.60–8.46(m,1H),8.25(d,J＝10.8Hz,1H),7.60(d,J＝7.9Hz,1H),5.20(m,1H),4.31–3.93(m,2H),3.84(m,2H),3.74(m,4H),3.57(m,3H),3.23(m,2H),3.01(s,3H),2.93(s,3H),2.80(s,3H),2.57(m,3H),2.28(m,3H),1.85(d,J＝6.4Hz,6H).
19F NMR(376MHz,MeOD)δ:-129.0,-142.7.
MS m/z(ESI):518.3[M+H]+.
实施例16
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(5-甲基-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000054
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(5-甲基-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.87(d,J＝3.0Hz,1H),8.51(s,1H),8.29(d,J＝8.9Hz,1H),8.13(d,J＝11.1Hz,1H),7.50(dd,J＝8.6,4.8Hz,1H),5.07(dt,J＝13.7,6.8Hz,1H),4.05(m,1H),3.85(m,1H),3.65–3.39(m,4H),3.32(m,1H),2.90(s,3H),2.67(s,3H),2.11–1.89(m,2H),1.73(d,J＝6.9Hz,6H),1.35(dd,J＝13.7,6.5Hz,3H).
19F NMR(376MHz,MeOD)δ:-129.1,-142.8.
MS m/z(ESI):535.2[M+H]+.
实施例17
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-甲基-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000055
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-甲基-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:9.00(d,J＝3.1Hz,1H),8.63(s,1H),8.42(dd,J＝15.8,8.9Hz,1H),8.26(d,J＝11.0Hz,1H),7.59(d,J＝8.9Hz,1H),5.20(dt,J＝13.8,6.7Hz,1H),4.36(d,J＝11.3Hz,1H),3.94–3.54(m,5H),3.50–3.40(m,2H),3.01(t,J＝7.9Hz,6H),2.80(d,J＝9.7Hz,3H),2.29(m,2H),1.85(d,J＝6.9Hz,6H).
19F NMR(376MHz,MeOD)δ:-129.3,-142.9.
MS m/z(ESI):535.2[M+H]+.
实施例18
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2-甲氧基乙基)-1,4-重氮基庚环‐1‐基)甲酮的制备
Figure PCTCN2017100678-appb-000056
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2-甲氧基乙基)-1,4-重氮基庚环-1-基)甲酮的制备方法如下。
(1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮(200mg,0.38mmol)溶于DMF(6mL),依次加入碳酸钾(150mg)和1-溴-2-甲氧基乙烷(0.5mL)，并于60℃下搅拌3h。冷却，过滤，浓缩柱层析纯化[洗脱剂:CH2Cl2～CH2Cl2/MeOH(10:1)]得白色固体产物(150mg，产率68％)。
1H NMR(400MHz,MeOD)δ:9.00(d,J＝3.2Hz,1H),8.68–8.60(m,1H),8.40(dd,J＝13.2,8.9Hz,1H),8.26(d,J＝11.1Hz,1H),7.59(d,J＝8.9Hz,1H),5.20(m,1H),4.37(m,1H),3.95–3.66(m,6H),3.59–3.39(m,8H),3.01(s,3H),2.79(s,3H),2.41–2.16(m,2H),1.85(d,J＝6.9Hz,6H).
19F NMR(376MHz,MeOD)δ:-129.3,-142.9.
MS m/z(ESI):579.3[M+H]+.
实施例19
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2,2,2-三氟乙基)-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000057
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2,2,2-三氟乙基)-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例1。
MS m/z(ESI):603.2[M+H]+.
实施例20
3-(4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-1,4-重氮基庚环-1-基)丙腈的制备
Figure PCTCN2017100678-appb-000058
3-(4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)-1,4-重氮基庚环-1-基)丙腈的制备方法参照实施例18。
1H NMR(400MHz,MeOD):δ:8.99(d,J＝3.3Hz,1H),8.63(s,1H),8.49(d,J＝9.0Hz,1H),8.25(d,J＝11.1Hz,1H),7.60(d,J＝8.9Hz,1H),5.18(dq,J＝14.1,7.0Hz,1H),4.10-3.85(m,1H),3.80-3.45(m,8H),3.26-3.18(m,2H),3.02(s,3H),2.80(s,3H),2.43-2.18(m,2H),1.85(d,J＝6.9Hz,6H).
19F NMR(400MHz,MeOD):δ:-129.12(s),-142.84(d,J＝10.2Hz).
MS m/z(ESI):574.2[M+H]+。
实施例21
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2-羟基丙基)-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000059
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2-羟基丙基)-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例18。
1H NMR(400MHz,MeOD)δ:8.56(d,J＝3.3Hz,1H),8.31(m,2H),7.77(d,J＝11.8Hz,1H),7.63(d,J＝8.4Hz,1H),4.93(m,1H),4.03–3.72(m,3H),3.57–3.41(m,2H),3.04(s,1H),2.84(d,J＝27.2Hz,3H),2.73–2.57(m,4H),2.49(m,4H),2.01(s,1H),1.87(s,1H),1.73(d,J＝6.7Hz,6H),1.17(m,3H).
19F NMR(400MHz,MeOD):δ:-130.5,-149.5.
MS m/z(ESI):579.3[M+H]+.
实施例22
(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-甲基-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000060
(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-甲基-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.97(d,J＝3.3Hz,1H),8.79(s,1H),8.42(dd,J＝15.2,8.9Hz,2H),8.04(d,J＝8.7Hz,1H),7.59(d,J＝8.9Hz,1H),5.19(dt,J＝13.8,6.9Hz,1H),4.45–4.29(m,1H),3.98–3.52(m,5H),3.49–3.38(m,2H),3.01(t,J＝7.9Hz,6H),2.81(t,J＝15.9Hz,3H),2.40–2.14(m,2H),1.84(t,J＝10.5Hz,6H).
MS m/z(ESI):517.3[M+H]+.
实施例23
6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基-N-(8-甲基-8-氮杂二环[3.2.1]辛烷-3-基)烟酰胺的制备
Figure PCTCN2017100678-appb-000061
6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基-N-(8-甲基-8-氮杂二环[3.2.1]辛烷-3-基)烟酰胺的制备方法参照实施例1。
MS m/z(ESI):561.3[M+H]+.
实施例24
(4-环丙基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000062
(4-环丙基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例18。
1H NMR(400MHz,MeOD)δ:8.47(d,J＝3.8Hz,1H),8.22(d,J＝8.7Hz,2H),7.71(d,J＝12.0Hz,1H),7.54(d,J＝8.5Hz,1H),5.04(m,1H),3.63(s,2H),3.50–3.29(m,2H),2.93(m,1H),2.83(s,1H),2.63(m,8H),2.35(s,3H),1.90(s,1H),1.76(s,1H),1.62(d,J＝6.9Hz,6H),1.19(s,2H).
19F NMR(376MHz,MeOD)δ:-130.7,-149.8.
MS m/z(ESI):561.2[M+H]+.
实施例25
(4-(2,2-二氟乙基)-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000063
(4-(2,2-二氟乙基)-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基 -1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例1。
MS m/z(ESI):585.3[M+H]+.
实施例26
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2-羟基乙基)-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000064
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(2-羟基乙基)-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例18。
MS m/z(ESI):565.3[M+H]+.
实施例27
4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-1,7-二甲基-1,4-重氮基庚环-5-酮的制备
Figure PCTCN2017100678-appb-000065
4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-1,7-二甲基-1,4-重氮基庚环-5-酮的制备方法参照实施例2。
MS m/z(ESI):535.3[M+H]+.
实施例28
3-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-1,3-噁吖己环-2-酮的制备
Figure PCTCN2017100678-appb-000066
3-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨 基)-2-甲基吡啶-3-基)-1,3-噁吖己环-2-酮的制备方法参照实施例2。
MS m/z(ESI):494.2[M+H]+.
实施例29
4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-1-甲基-1,4-重氮基庚环-5-酮的制备
Figure PCTCN2017100678-appb-000067
4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-1-甲基-1,4-重氮基庚环-5-酮的制备方法参照实施例2。
MS m/z(ESI):521.2[M+H]+.
实施例30
1-乙基-4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-1,4-重氮基庚环-5-酮的制备
Figure PCTCN2017100678-appb-000068
1-乙基-4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-1,4-重氮基庚环-5-酮的制备方法参照实施例2。
MS m/z(ESI):535.3[M+H]+.
实施例31
1-乙基-4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-7-甲基-1,4-重氮基庚环-5-酮的制备
Figure PCTCN2017100678-appb-000069
1-乙基-4-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)-7-甲基-1,4-重氮基庚环-5-酮的制备方法参照实施例2。
MS m/z(ESI):549.3[M+H]+.
实施例32
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-(2-甲氧基乙氧基)乙基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000070
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-(2-甲氧基乙氧基)乙基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):623.3[M+H]+.
实施例33
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(己基氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000071
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(己基氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ：8.97(s,1H),8.64(d,J＝7.4Hz,2H),8.44(d,J＝8.0Hz,1H),8.24(d,J＝11.0Hz,1H),7.73(d,J＝8.3Hz,1H),5.19(s,1H),3.51(s,1H),3.09(d,J＝6.8Hz,3H),3.03(s,3H),2.25(s,2H),1.86(d,J＝6.4Hz,6H),1.76(s,4H),1.46(s,2H),1.43–1.30(m,5H),0.96(t,J＝6.5Hz,3H).
MS m/z(ESI):605.3[M+H]+.
实施例34
(4-((2,2-二氟乙基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000072
(4-((2,2-二氟乙基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)):δ:8.97(d,J＝3.1Hz,1H),8.60(s,1H),8.35-8.15(m,2H),7.80–7.54(m,1H),6.39(t,J＝53.7Hz,2H),5.30-5.05(m,1H),3.90-3.55(m,4H),3.15-3.05(m,1H),2.99(s,3H),2.77(s,3H),3.40-2.10(m,2H),1.90-1.65(m,8H).
19F NMR(400MHz,MeOD):δ:-124.27(s),-129.21(s),-143.14(s).
MS m/z(ESI):585.2[M+H]+。
实施例35
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2,2,2-三氟乙基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000073
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2,2,2-三氟乙基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.95(d,J＝2.8Hz,1H),8.63(m,2H),8.42(d,J＝8.7Hz,1H),8.23(d,J＝11.1Hz,1H),7.73(d,J＝8.9Hz,1H),5.25–5.10(m,1H),4.17(q,J＝9.0Hz,2H),3.67(s,1H),3.37(m,4H),3.00(s,3H),2.39–2.19(m,2H),1.83(m,J＝11.6Hz,8H).
19F NMR(376MHz,MeOD)δ:-69.8,-129.2,-142.6.
MS m/z(ESI):603.2[M+H]+.
实施例36
(4-((环丙基甲基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000074
(4-((环丙基甲基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.99(d,J＝3.1Hz,1H),8.63(s,1H),8.27(t,J＝ 8.1Hz,2H),7.59(d,J＝8.9Hz,1H),5.19(dt,J＝13.8,6.8Hz,1H),3.79(d,J＝13.6Hz,1H),3.52(s,1H),3.01(d,J＝7.0Hz,6H),2.78(s,3H),2.30(d,J＝11.0Hz,1H),2.17(d,J＝10.2Hz,1H),1.85(d,J＝6.9Hz,6H),1.74(s,2H),1.15(td,J＝7.8,3.9Hz,1H),0.76(q,J＝5.8Hz,2H),0.47(q,J＝4.8Hz,2H).
MS m/z(ESI):573.5[M+H]+.
实施例37
(4-(环丁基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000075
(4-(环丁基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.88(s,1H),8.57–8.45(m,1H),8.14(d,J＝11.0Hz,2H),7.49(d,J＝7.9Hz,1H),5.20–5.04(m,1H),4.66(d,J＝11.9Hz,1H),3.86(d,J＝7.1Hz,1H),3.65(s,1H),3.35(s,1H),2.90(d,J＝7.7Hz,4H),2.66(s,3H),2.31–2.18(m,3H),2.17–1.97(m,2H),1.88(d,J＝16.2Hz,2H),1.72(t,J＝8.8Hz,6H),1.61(s,2H).
MS m/z(ESI):573.3[M+H]+.
实施例38
(4-((2,2-二氟环丙基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000076
(4-((2,2-二氟环丙基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):597.3[M+H]+.
实施例39
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-氟环丙基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000077
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-氟环丙基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):579.3[M+H]+.
实施例40
(4-(环戊基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000078
(4-(环戊基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):589.3[M+H]+.
实施例41
(4-((4,4-二氟环己基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000079
(4-((4,4-二氟环己基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.56(d,J＝3.8Hz,1H),8.32(d,J＝10.1Hz,2H),7.79(d,J＝11.9Hz,1H),7.61(s,1H),7.31(s,1H),4.69(d,J＝12.7Hz,1H),3.60(s,1H),3.19(t,J＝12.7Hz,1H),2.97(s,2H),2.86(s,1H),2.70(s,3H),2.46(s,3H),1.93(ddd,J＝30.6,29.7,16.7Hz,7H),1.73(d,J＝6.9Hz,6H),1.49(d,J＝9.9Hz,2H),1.30(s,3H).
MS m/z(ESI):639.2[M+H]+.
实施例42
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-羟基乙基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000080
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-羟基乙基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):565.3[M+H]+.
实施例43
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-甲氧基乙基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000081
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((2-甲氧基乙基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):579.3[M+H]+.
实施例44
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-吗啉代哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000082
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-吗啉代哌啶-1-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.56(d,J＝3.8Hz,1H),8.31(m,2H),7.77(d,J＝11.9Hz,1H),7.61(s,1H),4.91(m,1H),4.73(d,J＝12.1Hz,1H),3.82–3.69(m,4H),3.65(d,J＝13.0Hz,1H),3.17(t,J＝12.3Hz,1H),2.92(dd,J＝27.2,15.4Hz,1H),2.78–2.53(m,8H),2.45(s,3H),2.10(d,J＝10.2Hz,1H),1.94(d,J＝12.0Hz,1H),1.73(d,J＝6.9Hz,6H),1.49(m,2H).
19F NMR(376MHz,MeOD)δ:-130.5,-149.5.
MS m/z(ESI):591.3[M+H]+.
实施例45
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(3,3,4-三甲基哌嗪-1-基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000083
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(3,3,4-三甲基哌嗪-1-基)哌啶-1-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):632.4[M+H]+.
实施例46
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(4-甲基-4,7-二氮杂螺[2.5]辛烷-7-基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000084
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(4-甲基-4,7-二氮杂螺[2.5]辛烷-7-基)哌啶-1-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):630.3[M+H]+.
实施例47
(4-((3,3-二氟环丁基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000085
(4-((3,3-二氟环丁基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):611.3[M+H]+.
1H NMR(400MHz,MeOD)δ:9.00(d,J＝3.1Hz,1H),8.63(s,1H),8.28(t,J＝11.1Hz,2H),7.59(d,J＝8.9Hz,1H),5.29–5.11(m,1H),4.80(d,J＝13.5Hz,1H),3.97(dd,J＝12.9,7.2Hz,1H),3.79(d,J＝12.4Hz,1H),3.56(s,1H),3.22–2.91(m,8H),2.78(s,3H),2.21(dd,J＝54.2,9.7Hz,2H),1.85(d,J＝6.9Hz,6H),1.78(d,J＝12.2Hz,2H).
实施例48
(S)-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(吗啉-2-基甲基)-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000086
(S)-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(吗啉-2-基甲基)-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例18。
MS m/z(ESI):620.3[M+H]+.
实施例49
(R)-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(吗啉-2-基甲基)-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000087
(R)-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(吗啉-2-基甲基)-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例18。
MS m/z(ESI):620.3[M+H]+.
实施例50
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(6-甲基-3,6-二氮杂二环[3.2.1]辛烷-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000088
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(6-甲基-3,6-二氮杂二环[3.2.1]辛烷-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.58(d,J＝3.8Hz,1H),8.39–8.31(m,2H),7.82(d,J＝11.9Hz,1H),7.68–7.59(m,1H),4.98–4.92(m,1H),3.65–3.50(m, 1H),3.48–3.40(m,2H),3.27–2.96(m,2H),2.84–2.68(m,4H),2.65(s,3H),2.57–2.36(m,5H),2.22–2.00(m,1H),2.00–1.80(m,1H),1.74(d,J＝6.9Hz,6H).
MS m/z(ESI):5847.2[M+H]+.
实施例51
(4-(环丙基(甲基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000089
(4-(环丙基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮(200mg，0.357mmol)、30％甲醛水溶液(2ml)于甲醇(10ml)中搅拌，加入三乙酰氧基硼氢化钠(227mg)，室温搅拌3h。加水，二氯甲烷提取，干燥，纯化得到类白色固体(201.7mg,产率98.4％)。
1H NMR(400MHz,MeOD)δ:9.00(s,1H),8.63(s,1H),8.44–8.18(m,2H),7.60(d,J＝7.9Hz,1H),5.26–5.14(m,1H),4.87–4.82(m,1H),3.80(d,J＝9.4Hz,2H),3.35(s,1H),3.02(d,J＝9.3Hz,8H),2.78(s,3H),2.36(t,J＝48.6Hz,2H),1.94(s,2H),1.85(d,J＝6.5Hz,6H),1.29(d,J＝8.6Hz,2H),1.05(d,J＝29.8Hz,3H).
MS m/z(ESI):575.3[M+H]+.
实施例52
(4-(环丙基(异丙基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000090
(4-(环丙基(异丙基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例51。
MS m/z(ESI):603.7[M+H]+.
实施例53
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(2-甲基-2,7-二氮杂螺[3.5]壬烷-7-基)甲酮的制备
Figure PCTCN2017100678-appb-000091
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(2-甲基-2,7-二氮杂螺[3.5]壬烷-7-基)甲酮的制备方法参照实施例1。
MS m/z(ESI):561.6[M+H]+.
实施例54
(4-(2-(二甲氨基)乙基)-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000092
(4-(2-(二甲氨基)乙基)-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例18。
MS m/z(ESI):592.7[M+H]+.
实施例55
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((1-甲基哌啶-4-基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000093
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((1-甲基哌啶-4-基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):561.6[M+H]+.
实施例56
(4-(3,3-二甲基哌嗪-1-基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000094
(4-(3,3-二甲基哌嗪-1-基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯 并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):618.7[M+H]+.
实施例57
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((1-甲基环丙基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000095
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((1-甲基环丙基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.58(d,J＝3.8Hz,1H),8.35(s,1H),7.82(d,J＝11.9Hz,1H),7.62(d,J＝8.3Hz,1H),7.40–7.34(m,1H),5.00–4.92(m,1H),4.80–4.66(m,1H),3.74–3.58(m,1H),3.54–3.42(m,1H),3.30–3.12(m,1H),3.09–2.91(m,1H),2.71(s,3H),2.60–2.38(m,4H),2.25–2.10(m,1H),2.10–1.89(m,1H),1.74(d,J＝6.9Hz,6H),1.56–1.42(m,1H),1.31(s,3H),0.83–0.67(m,2H),0.66–0.50(m,2H).
MS m/z(ESI):575.7[M+H]+.
实施例58
1-((1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)哌啶-4-基)氨基)环丙烷-1-甲腈的制备
Figure PCTCN2017100678-appb-000096
1-((1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)哌啶-4-基)氨基)环丙烷-1-甲腈的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.51(d,J＝3.7Hz,1H),8.24(s,1H),8.08(d,J＝8.5Hz,1H),7.98(s,1H),7.72(d,J＝11.8Hz,1H),7.58(d,J＝8.5Hz,1H),4.89–4.81(m,1H),4.47–4.27(m,1H),3.53–3.41(m,1H),3.19–3.00(m,3H),2.61(s,3H),2.38(s,3H),2.06–1.92(m,1H),1.92–1.79(m,1H),1.62(d,J＝6.9Hz,6H),1.45–1.18(m,2H),1.19–0.85(m,4H).
MS m/z(ESI):585.7[M+H]+.
实施例59
(1,4-二氮杂二环[3.2.2]壬烷-4-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000097
(1,4-二氮杂二环[3.2.2]壬烷-4-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.88(d,J＝3.2Hz,1H),8.52(s,1H),8.26(t,J＝9.8Hz,1H),8.15(d,J＝11.1Hz,1H),7.49(d,J＝8.8Hz,1H),5.09(dt,J＝13.7,6.8Hz,1H),4.87(s,1H),4.14(s,1H),3.82(s,1H),3.52(m,6H),2.91(s,3H),2.77–2.63(m,3H),2.33(d,J＝6.3Hz,2H),2.19(d,J＝4.9Hz,2H),1.73(d,J＝6.9Hz,6H).
19F NMR(376MHz,MeOD)δ:-129.1,-142.6.
MS m/z(ESI):546.3[M+H]+.
实施例60
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(3,3,3-三氟丙基)-1,4-重氮基庚环-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000098
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(3,3,3-三氟丙基)-1,4-重氮基庚环-1-基)甲酮的制备方法参照实施例1。
MS m/z(ESI):617.6[M+H]+.
实施例61
[1,4'-联哌啶]-1'-基(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000099
[1,4'-联哌啶]-1'-基(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:9.00(d,J＝3.1Hz,1H),8.64(s,1H),8.28(t,J＝13.7Hz,2H),7.60(d,J＝8.8Hz,1H),5.27–5.15(m,1H),4.85(s,1H),3.83(d,J＝12.8Hz,1H),3.59(s,3H),3.37(s,1H),3.17–2.93(m,6H),2.78(s,3H),2.34(d,J＝11.6Hz,1H),2.20(d,J＝8.9Hz,1H),2.01(d,J＝14.0Hz,2H),1.97–1.88(m,3H),1.85(d,J＝6.9Hz,8H),1.56(d,J＝12.5Hz,1H).
MS m/z(ESI):589.3[M+H]+.
实施例62
(4-(环丙基氨基)哌啶-1-基)(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000100
(4-(环丙基氨基)哌啶-1-基)(2-氟-6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)吡啶-3-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ：8.71(d,J＝3.4Hz,1H),8.58(d,J＝1.0Hz,1H),8.38(dd,J＝8.3,1.9Hz,1H),8.16(d,J＝11.4Hz,1H),8.02–7.87(m,1H),5.22–5.05(m,1H),3.83(s,1H),3.62(t,J＝11.6Hz,1H),2.97(s,3H),2.89–2.77(m,1H),2.25(dd,J＝32.2,24.6Hz,2H),1.83(d,J＝6.9Hz,6H),1.68(dd,J＝12.3,4.3Hz,2H),1.33(d,J＝19.0Hz,2H),1.01–0.90(m,4H).
MS m/z(ESI):565.6[M+H]+.
实施例63
(4-(环丙基氨基)哌啶-1-基)(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000101
(4-(环丙基氨基)哌啶-1-基)(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.96(s,1H),8.78(s,1H),8.42(d,J＝8.1Hz,1H),8.29(s,1H),8.04(d,J＝8.1Hz,1H),7.61(d,J＝7.9Hz,1H),5.19(s,1H),4.82–4.72(m,2H),3.72(d,J＝57.5Hz,2H),3.04(d,J＝15.4Hz,4H),2.80(d,J＝24.0Hz,4H),2.46–2.16(m,2H),1.85(d,J＝6.4Hz,8H),1.08–0.87(m,4H).
MS m/z(ESI):543.2[M+H]+.
实施例64
(S)-(3,4-二甲基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000102
(S)-(3,4-二甲基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例1。
1H NMR(400MHz,MeOD)δ:8.54(s,1H),8.32–8.04(m,2H),7.72–7.48(m,2H),4.86–4.73(m,1H),3.95(m,1H),3.74–3.48(m,1H),3.40(m,2H),3.05(s,2H),2.93–2.68(m,1H),2.63(s,3H),2.51(s,2H),2.41(m,4H),2.06(s,1H),1.91(s,1H),1.66(s,6H),1.26(d,J＝6.6Hz,2H),0.93(d,J＝6.3Hz,1H).
19F NMR(376MHz,MeOD)δ:-130.2,-149.0.
MS m/z(ESI):548.3[M+H]+.
实施例65
N-(3,3-二氟环丁基)-N-(1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基尼古丁酰)哌啶-4-基)乙酰胺的制备
Figure PCTCN2017100678-appb-000103
(4-((3,3-二氟环丁基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮(200mg，0.328mmol)溶于二氯甲烷(10ml)中，加入三乙胺(1ml)，滴加醋酸酐(1ml)，室温搅拌3h。加水，二氯甲烷提取，干燥，纯化得到180.3mg目标产物，收率84.3％。
1H NMR(400MHz,MeOD)δ:9.00(d,J＝3.2Hz,1H),8.63(s,1H),8.26(d,J＝11.1Hz,1H),7.58(d,J＝8.9Hz,1H),5.27–5.12(m,1H),4.79(s,1H),4.12–3.46(m,2H),3.35(s,1H),3.33(dt,J＝3.3,1.6Hz,8H),3.02(s,4H),2.78(s,3H),2.65(s,1H),2.22(s,3H),1.82(t,J＝18.9Hz,9H).
MS m/z(ESI):653.2[M+H]+.
实施例66
(4-烯丙基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000104
(4-烯丙基-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并 [d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例18。
1H NMR(400MHz,MeOD)δ:8.53(s,1H),8.12(d,J＝17.1Hz,2H),7.53(d,J＝7.9Hz,2H),6.07–5.78(m,1H),5.27(dt,J＝48.5,14.5Hz,2H),4.82(d,J＝6.7Hz,1H),3.84(d,J＝16.7Hz,2H),3.47(d,J＝15.2Hz,2H),3.20(d,J＝5.7Hz,1H),2.98(s,1H),2.81(s,2H),2.71(s,1H),2.60(d,J＝7.4Hz,3H),2.41(d,J＝4.9Hz,3H),2.02(s,1H),1.87(s,1H),1.64(s,6H).
19F NMR(376MHz,MeOD)δ:-130.0,-148.7.
MS m/z(ESI):560.2[M+H]+.
实施例67
(4-(环丙基甲基)-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000105
(4-(环丙基甲基)-1,4-重氮基庚环-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例18。
1H NMR(400MHz,MeOD)δ:8.45(d,J＝3.3Hz,1H),8.35–8.12(m,2H),7.77–7.52(m,2H),4.90–4.81(m,1H),4.16–3.63(m,3H),3.65–3.26(m,5H),3.14–2.89(m,2H),2.59(s,3H),2.36(s,3H),2.30–1.96(m,2H),1.60(d,J＝9.0Hz,6H),1.16–0.97(m,1H),0.77–0.56(m,2H),0.48–0.19(m,2H).
MS m/z(ESI):575.2[M+H]+.
实施例68
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-氮杂环丁烷代哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000106
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-氮杂环丁烷代哌啶-1-基)甲酮的制备方法如下：
Figure PCTCN2017100678-appb-000107
将1-(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基烟酰)哌啶-4-酮(350mg,0.67mmol)，氮杂环丁烷盐酸盐(126mg,1.34mmol)溶于二氯甲烷(5mL)和甲醇(15mL)混合溶剂中，加入钛酸四乙酯(458mg,2.0mmol)，反应搅拌在室温下10小时后，醋酸硼氢化钠(425mg,2.0mmol)加入反应液中，反应在室温下搅拌3小时，LCMS显示反应完全，将反应液加到饱和碳酸氢钠溶液中(5mL),搅拌30分钟，加入无水硫酸钠，过滤，直接旋干，剩余物用快速硅胶柱纯化，再经反相柱层析纯化得到浅灰色固体产品(159mg,产率42％)。
1H NMR(400MHz,DMSO-d6)δ：10.22(s,1H),8.71(s,1H),8.31(s,1H),8.14(d,J＝8.2Hz,1H),7.70(d,J＝11.9Hz,1H),7.56(d,J＝8.3Hz,1H),4.94–4.76(m,1H),4.08(s,1H),3.12(d,J＝44.4Hz,6H),2.65(s,3H),2.28(d,J＝47.8Hz,4H),1.91(s,2H),1.60(t,J＝20.1Hz,8H),1.27–0.83(m,3H).
19F NMR(376MHz,DMSO-d6)δ：-128.75,-148.44.
MS m/z(ESI):561.3[M+H]+
实施例69
(4-(环己基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000108
(4-(环己基)氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例68。
1H NMR(400MHz,DMSO-d6)δ:10.98(s,1H),9.17(s,1H),8.87(s,1H),8.41(s,1H),8.18–7.56(m,3H),4.95(s,1H),4.58(s,1H),3.49(s,1H),3.07(s,1H),2.80(s,3H),2.48(d,J＝17.7Hz,10H),2.06(t,J＝27.0Hz,3H),1.83–1.57(m,8H),1.43–1.06(m,5H).
19F NMR(376MHz,DMSO)δ-127.75，-127.75.
MS m/z(ESI):603.3[M+H]+.
实施例70
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((3,3,3-三氟丙基)氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000109
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-((3,3,3-三氟丙基)氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ：8.57(d,J＝3.8Hz,1H),8.33(d,J＝6.9Hz,2H),7.80(d,J＝11.8Hz,1H),7.62(d,J＝8.5Hz,1H),4.74(d,J＝12.6Hz,1H),3.67(d,J＝12.2Hz,1H),3.37(s,1H),3.23(t,J＝13.5Hz,1H),3.17–3.06(m,3H),3.00(t,J＝12.5Hz,1H),2.71(s,3H),2.55(dt,J＝15.7,9.2Hz,2H),2.46(s,3H),2.20(dd,J＝17.7,10.0Hz,1H),2.01(d,J＝10.2Hz,1H),1.73(d,J＝6.9Hz,6H),1.46(dd,J＝45.8,11.0Hz,1H),1.33(d,J＝19.4Hz,1H).
MS m/z(ESI):617.2[M+H]+.
实施例71
(4-(叔-丁基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000110
(4-(叔-丁基氨基)哌啶-1-基)(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):577.2[M+H]+.
实施例72
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(异丙基氨基)哌啶-1-基)甲酮的制备
Figure PCTCN2017100678-appb-000111
(6-((5-氟-4-(4-氟-1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)(4-(异丙基氨基)哌啶-1-基)甲酮的制备方法参照实施例3。
1H NMR(400MHz,MeOD)δ:8.52(d,J＝3.8Hz,1H),8.33–8.18(m,2H),7.76(d,J＝11.9Hz,1H),7.56(d,J＝8.6Hz,1H),4.87(m,1H),4.78(m,1H),3.65(s,1H),3.52(dd,J＝12.8,6.5Hz,2H),3.22(m,1H),2.90(d,J＝13.0Hz,1H),2.64(s,3H),2.40(s,3H),2.17(s,1H),2.00(s,1H),1.68(t,J＝11.1Hz,6H),1.47(d,J＝43.5Hz,2H),1.30(d,J＝6.5Hz,6H).
19F NMR(376MHz,MeOD)δ:-130.6,-149.5.
MS m/z(ESI):562.3[M+H]+.
实施例73
(4-((4,4-二氟环己基)氨基)哌啶-1-基)(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000112
(4-((4,4-二氟环己基)氨基)哌啶-1-基)(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):621.2[M+H]+.
实施例74
(4-(环丁基氨基)哌啶-1-基)(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备
Figure PCTCN2017100678-appb-000113
(4-(环丁基氨基)哌啶-1-基)(6-((5-氟-4-(1-异丙基-2-甲基-1H-苯并[d]咪唑-6-基)嘧啶-2-基)氨基)-2-甲基吡啶-3-基)甲酮的制备方法参照实施例3。
MS m/z(ESI):557.2[M+H]+.
生物学测试评价
以下结合测试例进一步描述解释本发明，但这些实施例并非意味着限制本发明的范围。
一、本发明化合物对酶学活性分析检测方法：
本实验运用LANCE(Lanthanide chelate excite)激酶检测技术，检测分析了受试化合物对CDK4/6的酶学抑制活性。方法简述如下：
1)在384孔实验板(Greiner,Cat.No.784076)加入不同浓度的化合物，每个浓度设双复孔，之后加入CDK4(Carna,Cat.No.04-105)，或者CDK6的激酶CDK6(Carna,Cat.No.04-107)，混合均匀后；
2)加入底物Ulight-MBP peptide(PerkinElmer,Cat.No.TRF0109-D)与ATP的混合液；
3)室温反应60分钟后加入EDTA终止反应，终止5分钟后加入抗体Europium-anti-phospho-Myelin BasicProtein(MBP)(PerkinElmer,Cat.No.TRF0109-D)；
4)室温孵育60分钟后上机检测(λex＝330nm,λem＝620nm andλem＝665nm)
5)根据以下公式采集，计算数据：
信号值＝OD665/OD620*10000；
抑制率(％)＝100-(信号值-min)/(max-min)*100。
并使用Graphpad 5.0软件进行数据分析及拟合计算IC50，所得数据列表如下：
表1
实施例编号 CDK4 IC50(nM) CDK6 IC50(nM)
1 0.14 2.87
3 0.23 6.91
4 0.59 4.17
5 0.45 7.26
6 0.54 9.61
7 1 18.42
8 0.4 10.32
9 1.13 11.04
10 0.47 6.48
11 0.52 5.2
12 0.94 6.97
13 0.67 1.07
14 0.74 5.16
15 1 10.9
16 0.47 6.48
17 0.58 7.33
18 0.83 7.7
20 0.73 5.16
21 0.96 10.6
22 0.42 3.6
24 0.72 5.9
32 0.63 1.36
33 0.24 3.93
34 0.8 /
35 1.28 /
36 0.57 10.69
37 0.45 3.68
41 0.55 3.26
47 0.91 11.36
50 0.6 5.12
51 1.13 9.06
57 0.38 4.92
58 2.52 19.61
59 1.04 5.88
61 0.46 5.59
62 0.93 12.2
63 0.49 9.56
64 1.13 3.81
66 0.58 3.89
67 0.63 5.71
68 1.42 12.97
69 2.44 1.59
70 2.06 10.42
72 0.97 5.91
从以上实验结果可以看出，本发明实施例化合物对CDK激酶活性具有很强的抑制活性，尤其是对CDK 4和/或6激酶活性具有很好的抑制活性及选择性。
二、本发明化合物对结肠癌肿瘤细胞colo205增殖活性的测定
化合物对结肠癌肿瘤细胞colo205增殖活性通过以下的方法进行测试。
该方法用来测定本发明中的化合物对结肠癌肿瘤细胞colo205增殖活性的抑制作用。
实验步骤：
本实验采用CellTiter-Glo的方法测试化合物对colo205细胞增殖的抑制作用，并得出化合物抑制细胞增殖活性的半数抑制浓度IC50。
1、在96孔细胞培养板中接种50～100μL的colo205细胞悬液，密度为1～5*104细胞/ml，将培养板于培养箱培养16～24小时(37℃，5％CO2)。
2、向培养板细胞中加入梯度稀释的不同浓度的待测化合物溶液，将培养板在培养箱孵育6天(37℃，5％CO2)。
3、每孔加入50～100μL CellTiter-Glo试剂，并振荡10分钟，室温静置10分钟。
4、酶标仪测定各板的化学发光信号值。
5、通过化学发光信号值计算抑制率。
6、根据不同浓度的抑制率通过曲线拟合得出化合物的IC50。
本发明中化合物对结肠癌肿瘤细胞colo205增殖活性的试验进行测定，测得的IC50值见表2。
表2本发明中化合物对结肠癌肿瘤细胞colo205增殖活性抑制IC50
实施例编号 IC50(nM)
1 190
3 73.03
4 116.3
11 106
12 153.7
14 221.3
16 112.9
17 76.09
18 180.7
20 231.9
21 197.1
22 192.2
24 219.6
32 199.7
34 298.4
36 156.3
37 73.2
41 93.3
50 71.93
51 134.3
57 150.8
58 200
59 137.7
61 112.8
63 141.2
64 44.66
66 162.9
67 174.6
72 116.3
从以上实验结果可以看出，本发明实施例化合物对结肠癌肿瘤细胞colo205增殖活性具有很强的抑制作用。
三、大鼠药代动力学测定
1.研究目的：
以SD Rats为受试动物，研究化合物3，化合物34和化合物63，口服给药在rat体内(血浆)的药代动力学行为。
2.试验方案
2.1试验药品：
本发明实施例化合物3，化合物34和化合物63自制。
2.2试验动物：
SD rat 3只，雄性，上海杰思捷实验动物有限公司，动物生产许可证号(SCXK(沪)2013-0006 N0.311620400001794)。
2.3药物配制：
称取10g羟乙基纤维素(HEC)，溶于1000mL纯净水，加入2.5mL Tween80和0.5mL防泡剂。混合均匀成澄清溶液。称取6.1mg实施例化合物3，6.9mg实施例化合物34，5.8mg实施例化合物63，分别溶于该溶液中，摇匀，超声15分钟，得到无色澄清溶液，浓度为0.5mg/mL。
2.4给药：
SD大鼠3只，雄性，禁食一夜后分别PO，剂量为5mg/kg，给药体积10mL/kg。2.5样品采集：
于给药前和给药后0.5,1.0,2.0,4.0,6.0,8.0,24.0h颈静脉采血0.2mL，置于 EDTA-2K试管中，4℃6000rpm离心6min分离血浆，于-80℃保存；给药后4h进食。
2.5测定结果：应用LCMS/MS方法得到最后测定结果，见表3
表3：本发明化合物的大鼠药代动力学参数
Figure PCTCN2017100678-appb-000114
实验结论：表中数据显示，在口服给药剂量为5mg/kg时，本发明的3个化合物实施例3，实施例34和实施例63在大鼠血浆中都达到很高的暴露量。最高血药浓度和作用时间差异较大，但AUC差异较小。都可以保证有效的作用浓度。
四、荷瘤小鼠药代动力学测定
1.研究目的：
以Colo205荷瘤小鼠为受试动物，研究化合物实施例化合物3，化合物34和化合物63在50mg/kg剂量下口服给药在小鼠体内(血浆和肿瘤组织)的药代动力学行为。
2.试验方案
2.1试验药品：
本发明实施例化合物3，化合物34和化合物63，自制。
2.2试验动物：
nude mouse 3只，雄性，上海杰思捷实验动物有限公司，动物生产许可证号(SCXK(沪)2013-0006N0.311620400001794)。
2.3药物配制：
称取10g羟乙基纤维素(HEC)，溶于1000mL纯净水，加入2.5mL Tween80和0.5mL防泡剂。混合均匀成澄清溶液。称取36.6mg实施例化合物3，22.6mg实施例化合物34，35.2mg实施例化合物63，分别溶于该溶液中，摇匀，超声15分钟，得到无色澄清溶液，浓度为5.0mg/mL。
2.4给药：
Colo205荷瘤小鼠3只，雄性；禁食一夜后分别p.o.，剂量为50mg/kg，给药体积10mL/kg。
2.5样品采集：
小鼠给药前和给药后，CO2处死，心脏采血0.2ml，置于EDTA-2K试管中，4℃6000rpm离心6min分离血浆，于-80℃保存；肿瘤组织称重后，置于2mL离心管中，于-80℃保存。
2.5测定结果：应用LCMS/MS方法得到最后测定结果，见表4。
表4：本发明化合物的小鼠药代动力学参数
Figure PCTCN2017100678-appb-000115
实验结论：如表中数据所显示，50mg/kg剂量下，实施例3，实施例34和实施例63在小鼠血浆和肿瘤里的暴露量达到很高的水平，实施例3和实施例63在肿瘤里的暴露量明显高于血液里，而从Tmax和MRT可以看出来在肿瘤里的浓度是一个逐渐增加的过程，而代谢速度更慢，说明化合物会逐渐蓄积在肿瘤里，并一直在肿瘤里保持较高的浓度，从而保证更好的抑瘤效果。
五、Colo205移植瘤模型上对肿瘤的抑制实验
5.1实验目的：
以BALB/c裸小鼠为受试动物，采用人结直肠癌细胞Colo205异种移植瘤(CDX)模型进行体内药效实验，评价受试化合物抗肿瘤作用。
5.2实验仪器与试剂：
5.2.1仪器：
超净工作台(BSC-1300II A2，上海博讯实业有限公司医疗设备厂)
CO2培养箱(Thermo)
离心机(Centrifuge 5720R,Eppendorf)
全自动细胞计数仪(Countess II,Life Technologies)
移液器(10-20μL,Eppendorf)
显微镜(TS100，尼康)
游标卡尺(500-196，日本三丰)
细胞培养瓶(T25/T75/T225,Corning)
5.2.2试剂：
RPMI培养基1640(11875-093,Gibco)
胎牛血清(FBS)(10099-141,Gibco)
0.25％胰蛋白酶(25200-056,Gibco)
青链霉素双抗(SV30010,GE)
磷酸盐缓冲液(PBS)(10010-023,Gibco)
5.3实验操作：
从细胞库中取出一株Colo205细胞，用1640培养基(1640+10％FBS+1％Glu +1％SP)复苏细胞，复苏后的细胞置细胞培养瓶中(在瓶壁标记好细胞种类、日期、培养人名字等)置于CO2培养箱中培养(培养箱温度为37℃，CO2浓度为5％)。待细胞铺满培养瓶底部80-90％后传代，传代后细胞继续置于CO2培养箱中培养。重复该过程直到细胞数满足体内药效需求。收集培养好的细胞，用全自动细胞计数仪计数，根据计数结果用PBS重悬细胞，制成细胞悬液(密度4×107/ml)。置于冰盒中待用。
BALB/c裸小鼠，雌性，6-8周龄，体重约为18-22克。将小鼠保持在一个特殊的无病原体的环境中，且在单个通风笼中，每笼5只小鼠。所有的笼子、垫料和水在使用前进行消毒。所有动物可以自由获取标准认证的商业实验室饮食。接种前用一次性大小鼠通用耳标标记裸鼠，并用75％医用酒精消毒接种部位皮肤。每只小鼠在右胁腹皮下按每0.1毫升4×106个细胞的密度接种Colo205肿瘤细胞，用于肿瘤生长。当平均肿瘤体积达到100-200立方毫米时开始给药。将受试化合物每日经口灌胃给药，给药剂量为50mg/kg。肿瘤体积用游标卡尺测量，每周两次。体积以立方毫米计量。通过以下公式计算：V＝0.5*D*d*d，其中D和d分别是肿瘤的长径和短径。抗肿瘤药效是通过化合物处理过的动物的平均肿瘤增加体积除以未处理过动物的平均肿瘤增加体积来确定。抑瘤率＝1-[(Vt-V0)给药组/(Vt-V0)溶剂对照组]*100％。实验结束后安乐死动物。
表5：本发明化合物的移植瘤小鼠药效参数
Figure PCTCN2017100678-appb-000116
实验结论：从表中数据可以看出,在50mg/kg剂量下，本发明的化合物实施例3，实施例34和实施例63显著抑制Colo205裸小鼠移植瘤生长。
在安慰剂组肿瘤增长到1763立方毫米的情况下,实施例34给药动物组的肿瘤增长到430立方毫米,显示出优异的抑制肿瘤增长的效果.而实施例3和实施例63给药的动物组肿瘤增长更慢.效果更显著。
六、hERG钾离子通道抑制活性测试
6.1细胞培养
6.1.1本试验所用的细胞为转染有hERG、cDNA与稳定表达hERG通道的CHO 细胞系(由丹麦Sophion Bioscience公司提供)，细胞代数为P14至P16。细胞培养在含有)下列成分的培养基中：Ham’s F12培养基，10％(v/v)灭活的胎牛血清，100μg/ml潮霉素B，100μg/ml Geneticin。
表6细胞培养基组成成份
Figure PCTCN2017100678-appb-000117
6.1.2CHO hERG细胞生长于含上述培养液的培养皿中，并在37℃、含5％CO2的培养箱中进行培养。电生理实验之前24到48小时，CHO hERG细胞被转移到放置于培养皿中的圆形玻璃片上，并在以上相同的培养液及培养条件下生长，每个圆形玻片上CHO hERG细胞的密度需要达到绝大多数细胞是独立、单个的要求。
6.2实验溶液
下列溶液(由Sophion推荐)用于电生理记录。
表7细胞内液和外液的组成成分
Figure PCTCN2017100678-appb-000118
6.3电生理记录系统
本实验采用手动膜片钳系统(HEKA EPC-10信号放大器及数字转换系统，购自德国HEKA Electronics)作全细胞电流的记录。表面生长有CHO hERG细胞的圆形玻片被放置于倒置显微镜下的电生理记录槽中。记录槽内以细胞外液作持续灌流(大约每分钟1毫升)。实验过程采用常规全细胞膜片钳电流记录技术。如无特殊说明，实验都是在常规室温下进行(～25℃)。细胞钳制在-80mV的电压下。细胞钳制电压去极化到+20mV以激活hERG钾通道，5秒后再钳制到-50mV以消除失活并产生尾电流。尾电流峰值用作hERG电流大小的数值。上述步 骤所记录的hERG钾电流在记录槽内持续的细胞外液灌流下达到稳定后则可以叠加灌流待测试的药物，直到药物对hERG电流的抑制作用达到稳定状态。一般以最近的连续3个电流记录线重合作为判断是否稳定状态的标准。达到稳定态势以后以细胞外液灌流冲洗直到hERG电流回复到加药物之前的大小。一个细胞上可以测试一个或多个药物，或者同一种药物的多个浓度，但是在不同药物之间需要以细胞外液冲洗。Cisapride(西沙必利，购自Sigma)被用于实验中作为阳性对照以保证所使用的细胞质量正常。
6.4化合物处理和稀释
为了取得化合物的IC50，我们选择了下列浓度(30，10，3，1，0.3和0.1，3(30，10，3，1，0.3和0.1μM)来作测试。在试验之前，首先用DMSO以梯度稀释的方式稀释成10，3，1，0.3和0.1mM的贮备液，再用细胞外液1000倍稀释成最终的μM测试浓度，除了30μM测试浓度DMSO的最终浓度都为0.3％，其它各浓度化合物溶液中DMSO的最终浓度都为0.1％。阳性对照Cisapride(西沙比利)的测试浓度为0.1μM。所有的化合物溶液都经过常规的5到10分钟超声和振荡以保证化合物完全溶解。
6.5数据分析
试验数据由HEKA Patchmaster(V2x73.2)，Microsoft Excel以及Graphpad Prism5.0提供的数据分析软件进行分析。
6.6质量控制
报告中的试验数据需要满足以下标准：
记录参数：
膜电阻Rm>500MΩ
接入电阻(Ra)<5MΩ
尾电流幅度>300pA
电流rundown(自发性减小)每分钟<2％
漏电流<200pA或者hERG电流峰值的10％(在90％的记录时间之内)
药理学参数：
0.1μM的西沙比利(C4740-10mg,Sigma)阻断超过50％的hERG电流作为阳性对照。
6.7实验结果
实施例在多浓度对hERG电流的抑制结果如下表：
化合物编号 3 34 63
hERG(uM) >30 >10 30.3
药物对于心脏hERG钾离子通道的抑制是药物导致QT延长综合症的主要原因。从实验结果可以看出，实施例化合物3，化合物34和化合物63对于心脏hERG钾离子通道基本没有抑制作用。
综上所述，本发明提供了一系列具有新颖结构的高活性，高选择性CDK4/6激酶抑制剂，酶活性和细胞活性更强，对激酶的选择性更好。在大鼠和小鼠上都显示出更好的药代动力学性质，也体现出了更优的药效。而心脏毒性显著降低。有很大的潜力被开发成针对细胞周期增殖异常类疾病的药物。尤其是针对HR+/Her-类乳腺癌的药物。

Claims (17)

  1. 一种通式(I)所示的化合物、其立体异构体或其药学上可接受盐：
    Figure PCTCN2017100678-appb-100001
    其中：
    L为键、-C(O)-或-C(O)NH-；
    环A为杂环基，其中所述的杂环基选自单环杂环基、螺环杂环基、稠环杂环基和桥环杂环基；
    R选自氢原子、氘原子或卤素；
    R1选自氘原子、烷基、氘代烷基、卤代烷基、烷氧基、卤代烷氧基、卤素、氨基、硝基、羟基、氰基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR3、-(CH2)nSR3、-(CH2)nC(O)R3、-(CH2)nC(O)OR3、-(CH2)nS(O)mR3、-(CH2)nNR4R5、-(CH2)nC(O)NR4R5、-(CH2)nC(O)NHR4、-(CH2)nNR4C(O)R5和-(CH2)nNR4S(O)mR5，其中所述的烷基、卤代烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤代烷基、卤素、氨基、硝基、氰基、羟基、烷氧基、卤代烷氧基、羟烷基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、-SR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8中的一个或多个取代基所取代；
    R2相同或不同，且各自独立地选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、烷氧基、氨基烷氧基、卤代烷氧基、卤素、氨基、氧代基、硝基、羟基、氰基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR3、-(CH2)nSR3、-(CH2)nC(O)R3、-(CH2)nC(O)OR3、-(CH2)nS(O)mR3、-(CH2)nNR4R5、-(CH2)nC(O)NR4R5、-(CH2)nC(O)NHR4、-(CH2)nNR4C(O)R5和-(CH2)nNR4S(O)mR5，其中所述的烷基、氘代烷基、卤代烷基、氨基烷氧基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤代烷基、卤素、氨基、硝基、氰基、羟基、烯基、炔基、烷氧基、卤代烷氧基、羟烷基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、-(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8 中的一个或多个取代基所取代；
    R3选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、烷氧基、卤代烷氧基、环烷基、杂环基、芳基和杂芳基；其中所述的烷基、卤代烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、氨基、硝基、氰基、羟基、羟烷基、烷氧基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、-(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8中的一个或多个取代基所取代；
    R4和R5相同或不同，且各自独立地选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、环烷基、杂环基、芳基、杂芳基、-(CH2)nOR6、-(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8，其中所述的烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、羟基、氨基、硝基、氰基、烷氧基、羟烷基、环烷基、杂环基、芳基和杂芳基、-(CH2)nOR6、-(CH2)nSR6、-(CH2)nC(O)R6、-(CH2)nC(O)OR6、-(CH2)nS(O)mR6、-(CH2)nNR7R8、-(CH2)nC(O)NR7R8、-(CH2)nC(O)NHR7、-(CH2)nNR7C(O)R8和-(CH2)nNR7S(O)mR8中的一个或多个取代基所取代；
    R6选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、烷氧基、卤代烷氧基、环烷基、杂环基、芳基和杂芳基；其中所述的烷基、卤代烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、氨基、硝基、氰基、羟基、羟烷基、烷氧基、环烷基、杂环基、芳基和杂芳基的一个或多个取代基所取代；
    R7和R8相同或不同，且各自独立地选自氢原子、氘原子、烷基、氘代烷基、卤代烷基、羟基、氨基、酯基、环烷基、杂环基、芳基和杂芳基，其中所述的烷基、环烷基、杂环基、芳基和杂芳基任选进一步被选自氘原子、烷基、卤素、羟基、氨基、硝基、氰基、酯基、烷氧基、羟烷基、环烷基、杂环基、芳基和杂芳基中的一个或多个取代基所取代；
    x为0、1、2、3、4或5的整数；
    m为0、1或2的整数；且
    n为0、1、2、3、4或5的整数。
  2. 根据权利要求1所述的通式(I)所示的化合物，其为通式(II)所示的化合物、其立体异构体或其药学上可接受盐：
    Figure PCTCN2017100678-appb-100002
    其中：
    环B选自3-8元单环杂环基、6-10元螺环杂环基、6-10元稠环杂环基或6-10元桥环杂环基；优选3-8元单环杂环基；
    L为键或-C(O)-；
    R、R1、R2和x如权利要求1所述。
  3. 根据权利要求2所述的通式(II)所示的化合物，其为通式(III)所示的化合物、其立体异构体或其药学上可接受盐：
    Figure PCTCN2017100678-appb-100003
    其中：环B、R、R1、R2和x如权利要求2所述。
  4. 根据权利要求2所述的通式(II)所示的化合物，其为通式(IV)所示的化合物、其立体异构体或其药学上可接受盐：
    Figure PCTCN2017100678-appb-100004
    其中：环B、R、R1、R2和x如权利要求2所述。
  5. 根据权利要求3所述的通式(III)所示的化合物，其为通式(V)所示的化合物、其立体异构体或其药学上可接受盐：
    Figure PCTCN2017100678-appb-100005
    其中：
    M为CR2R2、NR2或O；
    R为氢原子或卤素，其中所述的卤素优选氟原子；
    R1为烷基或卤素，其中所述的烷基为C1-6烷基，优选C1-3烷基；
    R2相同或不同，且各自独立地选自氢原子、C1-6烷基、C1-6卤代烷基、C1-6烷氧基、C1-6氨基烷氧基、C1-6卤代烷氧基、卤素、氨基、氧代基、硝基、羟基、氰基、C3-8环烷基、3-8元杂环基、-(CH2)nOR3和-(CH2)nNR4R5，其中所述的C1-6烷基、C1-6卤代烷基、C1-6氨基烷氧基、C3-8环烷基和3-8元杂环基任选进一步被选自C1-6烷基、C1-6卤代烷基、卤素、氨基、氰基、羟基、烯基、炔基、C1-6烷氧基、C1-6卤代烷氧基、C1-6羟烷基、C3-8环烷基、3-8元杂环基、-(CH2)n-、-(CH2)nOR6和-(CH2)nNR7R8中的一个或多个取代基所取代；
    或者两个R2相互连接，形成一个3-10元的环烷基或者杂环基，其中所述的3-8元的环烷基或者杂环基任选进一步被一个或者多个C1-6烷基、C1-6卤代烷基、C1-6烷氧基、C1-6氨基烷氧基、C1-6卤代烷氧基、卤素、氨基、氧代基、羟基、氰基和C3-8环烷基的取代基取代；优选形成5-8元环烷基或者杂环基；且
    y为0、1、2或3的整数；
    R3～R8、n和x如权利要求3所述。
  6. 根据权利要求5所述的通式(V)所示的化合物、其立体异构体或其药学上可接受盐：
    其中：
    M选自CHR2或NR2；
    R2选自氢原子、C1-6烷基、C1-6卤代烷基、C1-6烷氧基、C1-6氨基烷氧基、C1-6卤代烷氧基、卤素、氨基、氧代基、羟基、氰基、C3-8环烷基、3-8元杂环基、-(CH2)nOR3和-(CH2)nNR4R5，其中所述的C1-6烷基、C1-6卤代烷基、C1-6氨基烷 氧基、C3-8环烷基和3-8元杂环基任选进一步被选自C1-6烷基、C1-6卤代烷基、卤素、氨基、氰基、羟基、C2-6烯基、C2-6炔基、C1-6烷氧基、C1-6卤代烷氧基、C1-6羟烷基、C3-8环烷基、3-8元杂环基、-(CH2)n-、-(CH2)nOR6和-(CH2)nNR7R8中的一个或多个取代基所取代；
    R、R1、R3～R8、x、n和y如权利要求5所述。
  7. 根据权利要求5所述的通式(V)所示的化合物，其为通式(VI)所示的化合物、其立体异构体或其药学上可接受盐：
    Figure PCTCN2017100678-appb-100006
    其中：
    R4和R5各自独立地选自氢原子、C1-6烷基、C1-6卤代烷基、C3-8环烷基、-(CH2)nOR6、-(CH2)nC(O)R6，其中所述的C1-6烷基、C1-6卤代烷基、C3-8环烷基任选进一步被选自C1-6烷基、卤素、羟基、氨基、氰基、C1-6烷氧基、C1-6羟烷基和C1-6环烷基中的一个或多个取代基所取代；
    或者R4和R5形成一个3-8元的杂环基，其中所述的3-8元的杂环基任选进一步被选自C1-6烷基、-(CH2)n-、卤素、羟基、氨基、氰基、C1-6烷氧基、C1-6羟烷基和C1-6环烷基中的一个或多个取代基所取代；优选R4和R5形成的杂环基是4-6元；
    R、R1和n如权利要求5所述。
  8. 根据权利要求1-7所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R选自氢原子和卤素；卤素优选氟。
  9. 根据权利要求1-8所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R1选自C1-8烷基和卤素，其中所述的C1-8烷基，优选C1-6烷基，更优选C1-3烷基；最优选甲基，其中所述的卤素优选氟。
  10. 根据权利要求1-9所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R2选自氢原子、C1-8烷基、C1-8卤代烷基、C1-8烷氧基、C3-8环烷基、 C2-6烯基、卤素、氧代基、-(CH2)nNR4R5和3-10杂环基，其中所述的C1-8烷基、C1-8卤代烷基、C1-8烷氧基、C3-8环烷基和3-10元杂环基任选进一步被选自卤素、羟基、氰基、C1-8烷基、-(CH2)nOR6和C1-8烷氧基中的一个或多个取代基所取代；优选C1-6烷基、C1-6卤代烷基、氧代基、-(CH2)nNR4R5和3-6杂环基，其中所述的C1-6烷基任选进一步被选自卤素、羟基和氰基中的一个或多个取代基所取代；所述的C1-6烷基更优选C1-3烷基。
  11. 根据权利要求1-10所示各通式、其立体异构体或其药学上可接受的盐，其特征在于，R4和R5各自独立地选自氢原子、C1-8烷基、C1-8卤代烷基、C3-8环烷基、-(CH2)nC(O)R6和C1-8烷氧基，其中所述的C1-8烷基、C1-8卤代烷基、C3-8环烷基和C1-8烷氧基任选进一步被选自卤素、羟基、氰基、C1-8烷基、C3-8环烷基、-(CH2)nOR6和C1-8烷氧基中的一个或多个取代基所取代；优选C1-6烷基和C3-6环烷基，其中所述的C1-6烷基和C3-6环烷基任选进一步被选自卤素、羟基、氰基、C1-6烷基、C3-6环烷基和-(CH2)nOR6中的一个或多个取代基所取代；其中所述的C1-6烷基更优选C1-3烷基；R6选自氢原子、C1-6烷基和C1-6烷氧基；优选C1-3烷基和C1-3烷氧基。
  12. 根据权利要求1-11任一所示的通式(I)化合物、其立体异构体或其药学上可接受的盐，其特征在于，选自如下化合物：
    Figure PCTCN2017100678-appb-100007
    Figure PCTCN2017100678-appb-100008
    Figure PCTCN2017100678-appb-100009
    Figure PCTCN2017100678-appb-100010
  13. 根据权利要求1-12任一所示的通式(I)化合物、其立体异构体或其药学上可接受的盐的制备方法，包括如下步骤：
    Figure PCTCN2017100678-appb-100011
    通式化合物(V-A)和通式化合物(V-B)偶联后得到通式(I)化合物，该通式(I)化合物任选进一步反应，或进一步脱保护基得到不同的通式(I)化合物，其中偶联反应中催化试剂优选Pd2(dba)3和Xantphos试剂；
    其中：
    X为卤素；优选氯；
    环A、L、R、R1、R2和x如权利要求1所述。
  14. 一种药用组合物，其包括治疗有效剂量的权利要求1-12任一所示的通式(I)化合物、其立体异构体或其药学上可接受的盐以及一种或多种药学上可接受的载体、稀释剂或赋形剂。
  15. 根据权利要求1-12任一所示的通式(I)化合物、其立体异构体或其药学上可接受的盐，或权利要求14所述的药物组合物在在制备用于治疗和/或预防由CDK激酶4和/或6介导的癌症或肿瘤相关疾病的药物中的应用。
  16. 根据权利要求15所述的应用，其特征在于，所述癌症或肿瘤相关疾病选自脑瘤、肺癌、肝癌、胃癌、口腔癌、头颈癌、肠癌或直肠癌、结肠癌、肾癌、食管腺癌、食管鳞状细胞癌、鳞状上皮细胞癌、甲状腺癌、骨癌、皮肤癌、非小细胞肺癌、原位癌、淋巴瘤、神经纤维瘤、成神经细胞瘤、肥大细胞肿瘤、多发性骨髓瘤、黑色素瘤、胶质瘤、肉瘤或脂肪肉瘤、胶质母细胞瘤、膀胱癌、卵巢癌、腹膜癌、胰腺癌、乳腺癌、子宫癌、子宫颈癌、子宫内膜癌、前列腺癌、雌性生殖道癌、睾丸癌、胃肠道间质瘤或前列腺肿瘤；优选自膀胱癌、卵巢癌、腹膜癌、胰腺癌、乳腺癌、子宫癌、子宫颈癌、子宫内膜癌、前列腺癌、雌性生殖道癌、睾丸癌、胃肠道间质瘤或前列腺肿瘤。
  17. 根据权利要求16所述的应用，其特征在于，所述乳腺癌包括：在绝经后女性雌激素受体阳性和/或人表皮生长因子受体2阴性的局部晚期或转移性乳腺癌。
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CN112250669B (zh) 2022-02-25
TWI765908B (zh) 2022-06-01
CN109952295A (zh) 2019-06-28
CN109952295B (zh) 2021-04-06
CN112225724A (zh) 2021-01-15
TW201811771A (zh) 2018-04-01
CN109963842A (zh) 2019-07-02
CN109963842B (zh) 2020-11-03
WO2018045957A1 (zh) 2018-03-15
CN112250669A (zh) 2021-01-22

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