WO2020094018A1 - 一种螺芳环化合物及其应用 - Google Patents

一种螺芳环化合物及其应用 Download PDF

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WO2020094018A1
WO2020094018A1 PCT/CN2019/115754 CN2019115754W WO2020094018A1 WO 2020094018 A1 WO2020094018 A1 WO 2020094018A1 CN 2019115754 W CN2019115754 W CN 2019115754W WO 2020094018 A1 WO2020094018 A1 WO 2020094018A1
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unsubstituted
alkyl
compound
substituted
formula
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PCT/CN2019/115754
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English (en)
French (fr)
Inventor
郑乾刚
许明
曾庆龙
李靖
诸葛浩
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上海奕拓医药科技有限责任公司
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First worldwide family litigation filed litigation Critical https://patents.darts-ip.com/?family=70516741&utm_source=google_patent&utm_medium=platform_link&utm_campaign=public_patent_search&patent=WO2020094018(A1) "Global patent litigation dataset” by Darts-ip is licensed under a Creative Commons Attribution 4.0 International License.
Priority to EP19881313.1A priority Critical patent/EP3712151B1/en
Priority to CN201980005853.5A priority patent/CN111566104B/zh
Priority to CA3118925A priority patent/CA3118925A1/en
Priority to CN202410530427.6A priority patent/CN118359642A/zh
Priority to EP22159530.9A priority patent/EP4074713A1/en
Priority to ES19881313T priority patent/ES2914852T3/es
Priority to SG11202104752RA priority patent/SG11202104752RA/en
Application filed by 上海奕拓医药科技有限责任公司 filed Critical 上海奕拓医药科技有限责任公司
Priority to KR1020217017152A priority patent/KR20210088639A/ko
Priority to JP2021525321A priority patent/JP7520387B2/ja
Priority to AU2019374194A priority patent/AU2019374194A1/en
Priority to BR112021008714-1A priority patent/BR112021008714A2/pt
Publication of WO2020094018A1 publication Critical patent/WO2020094018A1/zh
Priority to US16/908,604 priority patent/US10844079B2/en
Priority to US17/014,883 priority patent/US11685748B2/en
Priority to US18/196,357 priority patent/US20240116949A1/en

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    • 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
    • C07D401/02—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 two hetero rings
    • C07D401/04—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 two hetero rings directly linked by a ring-member-to-ring-member bond
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4427—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems
    • 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
    • A61K31/4965—Non-condensed pyrazines
    • A61K31/497—Non-condensed pyrazines containing further heterocyclic rings
    • 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
    • A61K31/4985—Pyrazines or piperazines ortho- or peri-condensed with heterocyclic ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • 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
    • 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/10—Spiro-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D513/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00
    • C07D513/02—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00 in which the condensed system contains two hetero rings
    • C07D513/10—Spiro-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D519/00—Heterocyclic compounds containing more than one system of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring system not provided for in groups C07D453/00 or C07D455/00

Definitions

  • the present invention relates to the field of spiroaromatic compounds, in particular, to a spiroaromatic compound that can be used as an SHP2 inhibitor, its pharmaceutically acceptable salts, or its enantiomers, diastereomers, Tautomers, solvates, isotopic substitutions, prodrugs or metabolites.
  • the invention also relates to a method for preparing such compounds, a pharmaceutical composition containing the compounds, and the use of the compounds for the preparation of a medicament for treating a disease or disorder associated with abnormal SHP2 activity.
  • Protein tyrosine phosphatase SHP2 which occupies an important position in the process of cell signal transduction, is a target for the development of major diseases such as diabetes, autoimmune diseases and cancer.
  • SHP2 is mutated or highly expressed in various diseases, such as Noonan Syndrome, Leopard Syndrome, juvenile myelomonocytic leukemia, neuroblastoma, melanoma, acute myeloid leukemia, Breast cancer, esophageal cancer, lung cancer, colon cancer, head cancer, neuroblastoma, squamous cell carcinoma of the head and neck, gastric cancer, anaplastic large cell lymphoma and glioblastoma, etc.
  • SHP2 is involved in multiple tumor cell signaling pathways, such as MAPK, JAK / STAT, and PI3K / Akt. At the same time, SHP2 is also responsible for the signaling of the PD1-PDL1 immunosuppressive pathway. Therefore, inhibition of SHP2 activity can reverse immunosuppression in the tumor microenvironment.
  • SHP2 consists of two N-terminal Src homology 2 domains (N-SH2 and C-SH2) and a protein tyrosine phosphatase catalytic domain (PTP).
  • N-SH2 combines with PTP to form a ring structure, thereby hindering the binding of PTP to the substrate, so that the enzyme catalytic activity is inhibited; when the upstream receptor protein tyrosine is phosphorylated, N-
  • SH2 When SH2 is combined with it, the PTP catalytic domain is released to exert phosphatase activity.
  • SHP2 inhibitors are mainly based on allosteric inhibitors in the non-catalytic region, such as some compounds disclosed in WO2015107493A1, WO2016203404A1, WO2016203406A1, WO2017216706A1, WO2017211303A1, CN201710062495, WO2018136265A1WO2018057884, etc.
  • This year's research shows that SHP2 as a novel drugable target has attracted more and more attention. Therefore, there is an urgent need in the art to develop SHP2 inhibitors with novel structures, good biological activity, and high druggability.
  • the object of the present invention is to provide a compound of formula I or a pharmaceutically acceptable salt thereof, a pharmaceutical composition containing the compound or a pharmaceutically acceptable salt thereof, and the compound or pharmaceutical composition thereof in the prevention and treatment of SHP2 Application in abnormally related diseases or disorders.
  • X 1 and X 2 are each independently selected from a bond, O, CR a R b or NR c ;
  • X 3 is selected from bond, CR a R b , NR c , S or O;
  • X 4 is selected from N or CR c ; and R a , R b and R c are each independently selected from H, halogen, substituted or unsubstituted C 1-6 alkyl, substituted or unsubstituted C 1-6 alkoxy base;
  • R 1 , R 2 , R 3 , R 4 and R 7 are each independently selected from H, -OH, halogen, substituted or unsubstituted amino, substituted or unsubstituted C 1-6 alkyl, substituted or unsubstituted C 1-6 alkoxy; and cannot be -OH or -NH 2 at the same time;
  • Ring A is selected from substituted or unsubstituted C 4-8 cycloalkyl, substituted or unsubstituted 4-8 membered heterocyclic group, substituted or unsubstituted C 5-10 aryl, substituted or unsubstituted 5-10 member Heteroaryl, said heterocyclyl or heteroaryl contains 1-3 heteroatoms selected from the group consisting of N, O, S or P;
  • Ring C is selected from substituted or unsubstituted C 4-8 cycloalkyl, substituted or unsubstituted 5-6 membered monocyclic heterocyclic group, substituted or unsubstituted 8-10 membered bicyclic heterocyclic group, substituted or unsubstituted C 5-10 monocyclic or bicyclic aryl group, substituted or unsubstituted 5-6 membered monocyclic heteroaryl group, substituted or unsubstituted 8-10 membered bicyclic heteroaryl group, the heterocyclic group or heteroaryl group comprising 1-4 heteroatoms selected from the group consisting of N, O, S or P;
  • R 5 and R 6 are each independently selected from H, -OH, halogen, cyano, substituted or unsubstituted amino, substituted or unsubstituted C 1-6 alkyl, substituted or unsubstituted C 1-6 alkoxy base;
  • n is any integer from 0 to 3;
  • substitution means that one or more hydrogen atoms on the group are substituted with a substituent selected from the group consisting of halogen, -OH, -NO 2 , -NH 2 , and -NH (unsubstituted or halogenated C 1- 6 alkyl), -N (unsubstituted or halogenated C 1-6 alkyl) 2 , -CN, unsubstituted or halogenated C 1-8 alkyl, unsubstituted or halogenated C 1-8 alkyl Oxygen, unsubstituted or halogenated C 1-8 alkoxy-C 1-8 alkyl, unsubstituted or halogenated C 3-8 cycloalkyl-C 1-8 alkyl, unsubstituted or halogenated C 1-6 alkylcarbonyl, unsubstituted or halogenated C 1-6 alkoxycarbonyl, hydroxamate, unsubstituted or halogenated C 1-6 al
  • one of X 1 and X 2 is CH 2 and the other is a bond.
  • X 3 is S.
  • X 4 is selected from N or CH.
  • R 1 , R 2 , R 3 , R 4 and R 7 are each independently selected from H, -OH, -F, -Cl, -Br, -NH 2 and -NHC 1-3 Alkyl, methyl, ethyl, propyl, isopropyl, butyl, methoxy, ethoxy, propoxy, or isopropoxy; halogen, -NH 2 , -OH, C 1-3 alkyl group or a C 1-3 alkoxy substituted C 1-3 alkyl; or substituted with halogen, -NH 2, -OH, C 1-3 alkyl or a C 1-3 alkoxy C 1-3 Alkoxy.
  • R 5 and R 6 are each independently selected from H, -OH, -F, -Cl, -Br, -CN, -NH 2 , -NHC 1-3 alkyl, methyl, Ethyl, propyl, isopropyl, butyl, methoxy, ethoxy, propoxy or isopropoxy; halogen, -NH 2 , -OH, C 1-3 alkyl or C 1- 3 alkoxy-substituted C 1-3 alkyl; or substituted with halogen, -NH 2, -OH, C 1-3 alkyl or a C 1-3 alkoxy group a C 1-3 alkoxy group.
  • the substituent is selected from -F, -Cl, -Br, -OH, -NO 2 , -NH 2 , -NH (C 1-6 alkyl), -N (C 1 -6 alkyl) 2 , -CN, C 1-6 alkyl, C 1-4 alkoxy, C 1-4 alkoxy-C 1-6 alkyl, C 3-8 cycloalkyl-C 1 -8 alkyl, C 1-6 alkylcarbonyl, C 1-6 alkoxycarbonyl, C 1-6 alkylmercapto, -S (O) 2 N (C 1-6 alkyl) 2 , -S ( O) 2 C 1-6 alkyl, -N (C 1-6 alkyl) S (O) 2 N (C 1-6 alkyl) 2 , -S (O) N (C 1-6 alkyl) 2 , -S (O) (C 1-6 alkyl), -N (C 1-6 alkyl) S (O) 2 N (C
  • the substituent is selected from -F, -Cl, -Br, -OH, -NO 2 , -NH 2 , -NH (C 1-3 alkyl), -N (C 1 -3 alkyl) 2 , -CN, C 1-3 alkyl, C 1-3 alkoxy, C 1-3 alkylcarbonyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, Cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cyclooctyl, pyrrolidinyl, morpholinyl, piperazinyl, homopiperazinyl, piperidinyl, thiomorpholinyl, benzene Group, naphthyl, anthracenyl, phenanthrenyl, fluorenyl, thienyl, imidazolyl, pyr
  • the substituent is selected from -F, -Cl, -Br, -OH, -NO 2 , -NH 2 , -NH (C 1-3 alkyl), -N (C 1 -3 alkyl) 2 , -CN, methyl, ethyl, propyl, isopropyl, butyl, methoxy, ethoxy, propoxy, isopropoxy or phenyl.
  • the ring C is selected from any one of the following groups:
  • X 5 , X 6 , X 7 , X 8 and X 9 are each independently selected from N or CR d ; and at most 3 are N simultaneously;
  • X 10 , X 11 , X 12 , X 13 , X 14 , X 15 , X 16 and X 17 are each independently selected from N or CR d ; and at most 5 of them are N;
  • X 18 , X 19 , X 20 and X 21 are each independently selected from N or CR d , and at most 3 are N simultaneously;
  • R 6 and R 8 are each independently selected from H, -NH 2 , -CN, -OH, -NO 2 , halogen, unsubstituted or halogenated C 1-6 alkyl, unsubstituted or halogenated C 1- 6 alkoxy; and
  • the Rd is selected from H, halogen, unsubstituted or halogenated C 1-6 alkyl, unsubstituted or halogenated C 1-6 alkoxy.
  • the ring C is selected from any one of the following:
  • X 5 , X 6 , X 7 , X 8 and X 9 are N, and the rest are CR d ;
  • R 6 is selected from H, -NH 2 , -CN, -OH, -NO 2 , -F, -Cl, -Br, methyl, ethyl, propyl, isopropyl, butyl, methoxy, ethyl , propoxy, isopropoxy, fluoro or a C 1-3 alkyl bromo, fluoro or bromo group of C 1- 3 alkoxy; and
  • the Rd is selected from H, -F, -Cl, -Br, methyl, ethyl, propyl, isopropyl, butyl, methoxy, ethoxy, propoxy, isopropoxy, Fluoro or bromo C 1-3 alkyl, fluoro or bromo C 1-3 alkoxy.
  • the ring C is selected from any one of the following groups:
  • the ring A is selected from a substituted or unsubstituted C 4-6 cyclic hydrocarbon group, a substituted or unsubstituted 4-6 membered heterocyclic group, and a substituted or unsubstituted C 5-6 aryl group.
  • the ring A is selected from any one of the following groups:
  • the ring A is selected from any one of the following groups:
  • the compound has a structure selected from the following:
  • the present invention is the isotopically substituted compounds, isotopic substituents selected from the group: hydrogen, carbon, nitrogen, oxygen, fluorine, phosphorus, chlorine and iodine; preferably 2 H, 3 H, 11 C, 13 C, 14 C, 15 N, 17 O, 18 O, 18 F, 31 P, 32 P, 35 S, 36 Cl or 125 I.
  • the disease is cancer; preferably Noonan syndrome, leopard syndrome, juvenile myeloid monocytic leukemia, neuroblastoma, melanoma, acute myeloid leukemia, breast cancer, esophageal cancer , Lung cancer, colon cancer, head cancer, neuroblastoma, squamous cell carcinoma of the head and neck, gastric cancer, anaplastic large cell lymphoma or glioblastoma.
  • a pharmaceutical composition comprising:
  • a method of inhibiting SHP2 activity comprising the steps of: administering to a subject in need thereof an effective amount of a compound of formula I of the present invention or a pharmaceutically acceptable salt thereof , Or administer an effective amount of the pharmaceutical composition of the present invention to a subject in need thereof.
  • the inventors prepared a novel allosteric inhibitor compound represented by formula I, which can bind to the non-catalytic region of SHP2 and "lock" the weak self-inhibition of SHP2 activity. State to achieve the purpose of inhibiting its activity.
  • the compound of the present invention exhibits very good biological activity and druggability, and has very good drug development prospects. It has an inhibitory effect on SHP2 at a very low concentration (which can be as low as ⁇ 100 nM / L), and the inhibitory activity is comparable Excellent, so it can be used to treat diseases or conditions associated with SHP2, such as tumors. Based on the above findings, the inventors have completed the present invention.
  • the manufacturer's instructions for the kit can be used, or the reaction and purification can be carried out in a manner well known in the art or the instructions of the present invention.
  • the above-mentioned techniques and methods can be implemented according to conventional methods well known in the art according to descriptions in a number of summary and more specific documents cited and discussed in this specification.
  • groups and their substituents can be selected by those skilled in the art to provide stable moieties and compounds.
  • substituents When a substituent is described by a general chemical formula written from left to right, the substituent also includes chemically equivalent substituents obtained when the structural formula is written from right to left. For example, -CH 2 O- is equivalent to -OCH 2- .
  • C1-C6 alkyl refers to an alkyl group having a total of 1 to 6 carbon atoms as defined below.
  • the total number of carbon atoms in the simplified symbol does not include carbons that may be present in the substituents of the group.
  • halogen refers to fluorine, chlorine, bromine or iodine.
  • Hydroxyalkyl refers to an alkyl group as defined below substituted by a hydroxy group (-OH).
  • Niro means -NO 2 .
  • Amino refers to -NH 2 .
  • Substituted amino refers to an amino group substituted with one or two alkyl groups, alkylcarbonyl groups, aralkyl groups, heteroaralkyl groups as defined below, for example, monoalkylamino, dialkylamino, alkyl Acylamino, aralkylamino, heteroaralkylamino.
  • Carboxy means -COOH.
  • alkyl means fully saturated Straight-chain or branched-chain hydrocarbon chain group consisting only of carbon atoms and hydrogen atoms, having, for example, 1 to 12 (preferably 1 to 8, more preferably 1 to 6) carbon atoms, and through a single bond with the molecule
  • the rest of the connection such as but not limited to methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl , 2,2-dimethylpropyl, n-hexyl, heptyl, 2-methylhexyl, 3-methylhexyl, octyl, nonyl and decyl and so on
  • alkenyl means consisting only of carbon atoms and hydrogen atoms, containing at least one double bond, having, for example, 2 to 20 (preferably 2 to 10) , More preferably 2 to 6) carbon atoms and a straight or branched hydrocarbon chain group connected to the rest of the molecule by a single bond, such as but not limited to vinyl, propenyl, allyl, butylene 1-alkenyl, but-2-enyl, pent-1-enyl, pent-1,4-dienyl, etc.
  • cyclic hydrocarbon group means a stable non-aromatic monocyclic or polycyclic hydrocarbon group composed of only carbon atoms and hydrogen atoms (eg, alkyl, alkenyl Or alkynyl), which may include fused ring systems, bridged ring systems, or spiro ring systems, having 3 to 15 carbon atoms, preferably 3 to 10 carbon atoms, more preferably 3 to 8 carbon atoms, such as 3 , 4, 5, 6, 7 or 8 carbon atoms, and it is saturated or unsaturated and can be connected to the rest of the molecule via a single bond via any suitable carbon atom.
  • hydrogen atoms eg, alkyl, alkenyl Or alkynyl
  • cyclic hydrocarbon groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cyclooctyl, 1H-indene Group, 2,3-dihydroindenyl, 1,2,3,4-tetrahydro-naphthyl, 5,6,7,8-tetrahydro-naphthyl, 8,9-dihydro-7H-benzo Cyclohepten-6-yl, 6,7,8,9-tetrahydro-5H-benzocycloheptenyl, 5,6,7,8,9,10-hexahydro-benzocyclooctenyl, Fluorenyl, bi
  • heterocyclic group means from 2 to 14 carbon atoms (eg, 2, 3, 4, 5, 6, 7, 8, 9, 10 , 11, 12, 13 or 14 carbon atoms) and 1 to 6 heteroatoms selected from nitrogen, phosphorus, oxygen and sulfur, a stable 3- to 20-membered non-aromatic cyclic group.
  • the heterocyclic group may be a monocyclic, bicyclic, tricyclic or more ring system, which may include a fused ring system, a bridge ring system or a spiro ring system;
  • the nitrogen, carbon or sulfur atoms of can be optionally oxidized; the nitrogen atom can be optionally quaternized; and the heterocyclic group can be partially or fully saturated.
  • the heterocyclic group may be connected to the rest of the molecule via a single bond via a carbon atom or a heteroatom.
  • one or more rings may be an aryl group or a heteroaryl group as defined below, provided that the point of attachment to the rest of the molecule is a non-aromatic ring atom.
  • the heterocyclic group is preferably a stable 4- to 11-membered non-aromatic monocyclic, bicyclic, bridged ring or spiro ring group containing 1 to 3 heteroatoms selected from nitrogen, oxygen and sulfur
  • the group is more preferably a stable 4- to 8-membered non-aromatic monocyclic, bicyclic, bridged ring or spiro ring group containing 1 to 3 heteroatoms selected from nitrogen, oxygen and sulfur.
  • heterocyclic groups include, but are not limited to: pyrrolidinyl, morpholinyl, piperazinyl, homopiperazinyl, piperidinyl, thiomorpholinyl, 2,7-diaza-spiro [3.5] non Alk-7-yl, 2-oxa-6-aza-spiro [3.3] heptane-6-yl, 2,5-diaza-bicyclo [2.2.1] heptan-2-yl, aza Cyclobutanyl, pyranyl, tetrahydropyranyl, thiopyranyl, tetrahydrofuranyl, oxazinyl, dioxolyl, tetrahydroisoquinolinyl, decahydroisoquinolinyl, imidazolinyl, Imidazolidinyl, quinazinyl, thiazolidinyl, isothiazolidinyl, isoxazolidinyl, ind
  • aryl means having 6 to 18 carbon atoms (preferably having 6 to 10 carbon atoms, such as 6, 7, 8, 9 or 10 Carbon atoms) conjugated hydrocarbon ring system group.
  • an aryl group may be a monocyclic, bicyclic, tricyclic or more ring system, or it may be fused with a cycloalkyl or heterocyclic group as defined above, provided that the aryl group passes through The atoms on the aromatic ring are connected to the rest of the molecule through a single bond.
  • aryl groups include, but are not limited to, phenyl, naphthyl, anthracenyl, phenanthrenyl, fluorenyl, 2,3-dihydro-1H-isoindolyl, 2-benzoxazolinone, 2H-1, 4-Benzoxazine-3 (4H) -one-7-yl and so on.
  • arylalkyl refers to an alkyl group as defined above substituted with an aryl group as defined above.
  • heteroaryl means having 1 to 15 carbon atoms in the ring (preferably having 1 to 10 carbon atoms, such as 1, 2, 3, 4 , 5, 6, 7, 8, 9 or 10 carbon atoms) and 1 to 6 heteroatom selected from nitrogen, oxygen and sulfur 5- to 16-membered conjugated ring system groups.
  • the heteroaryl group may be a monocyclic, bicyclic, tricyclic or more ring system, and may also be fused with a cycloalkyl or heterocyclic group as defined above, provided that The aryl group is connected to the rest of the molecule via a single bond via atoms on the aromatic ring.
  • the nitrogen, carbon or sulfur atom in the heteroaryl group may be optionally oxidized; the nitrogen atom may be optionally quaternized.
  • the heteroaryl group is preferably a stable 5- to 12-membered aromatic group containing 1 to 5 heteroatoms selected from nitrogen, oxygen, and sulfur, and more preferably 1 to 4 Stable 5- to 10-membered aromatic groups from heteroatoms of nitrogen, oxygen, and sulfur or 5- to 6-membered aromatic groups containing 1 to 3 heteroatoms selected from nitrogen, oxygen, and sulfur.
  • heteroaryl groups include, but are not limited to, thienyl, imidazolyl, pyrazolyl, thiazolyl, oxazolyl, oxadiazolyl, isoxazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, Benzimidazolyl, benzpyrazolyl, indolyl, furanyl, pyrrolyl, triazolyl, tetrazolyl, triazinyl, indolizinyl, isoindolyl, indazolyl, isoindazolyl , Purinyl, quinolinyl, isoquinolinyl, diazonaphthyl, naphthyridinyl, quinoxalinyl, pteridinyl, carbazolyl, carolinyl, phenanthridinyl, phenanthrolinyl, acrid
  • heteroarylalkyl refers to an alkyl group as defined above substituted with a heteroaryl group as defined above.
  • “optionally” or “optionally” means that the subsequently described event or condition may or may not occur, and the description includes both occurrence and non-occurrence of the event or condition.
  • “optionally substituted aryl” means that the aryl is substituted or unsubstituted, and the description includes both substituted aryl and unsubstituted aryl.
  • substituents described in the claims and description of the present invention are selected from alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, cyano, nitro , Optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cyclic hydrocarbon group, optionally substituted heterocyclic group.
  • SHP2 means "Src Homolgy-2 phosphatase", also called SH-PTP2, SH-PT3, Syp, PTP1D, PTP2C, SAP-2 or PTPN11.
  • the terms "part”, “structural part”, “chemical part”, “group”, “chemical group” refer to a specific segment or functional group in a molecule. Chemical moieties are generally considered to be chemical entities embedded or attached to molecules.
  • Stepoisomer refers to a compound composed of the same atoms, bonded by the same bond, but having different three-dimensional structures.
  • the present invention will cover various stereoisomers and mixtures thereof.
  • the compound of the present invention contains an olefinic double bond, unless otherwise specified, the compound of the present invention is intended to include E- and Z-geometric isomers.
  • Tautomer refers to an isomer formed by the transfer of protons from one atom of a molecule to another atom of the same molecule. All tautomeric forms of the compounds of the invention will also be included within the scope of the invention.
  • the compounds of the present invention or pharmaceutically acceptable salts thereof may contain one or more chiral carbon atoms, and thus may produce enantiomers, diastereomers, and other stereoisomeric forms.
  • Each chiral carbon atom can be defined as (R)-or (S)-based on stereochemistry.
  • the present invention is intended to include all possible isomers, as well as their racemates and optically pure forms.
  • racemates, diastereomers or enantiomers can be selected as starting materials or intermediates.
  • Optically active isomers can be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques, such as crystallization and chiral chromatography.
  • the present invention also includes all suitable isotopic variations of the compounds of the present invention or pharmaceutically acceptable salts thereof.
  • Isotopic variations of the compounds of the present invention or pharmaceutically acceptable salts thereof are defined as those in which at least one atom is replaced by an atom having the same atomic number but an atomic mass different from the atomic mass often found in nature.
  • Isotopes that can be incorporated into the compounds of the present invention and pharmaceutically acceptable salts include, but are not limited to, isotopes of H, C, N, and O, such as 2 H, 3 H, 11 C, 13 C, 14 C, 15 N, 17 O, 18 O, 35 S, 18 F, 36 Cl and 125 I.
  • Isotopic variants of the compounds of the present invention or pharmaceutically acceptable salts thereof can be prepared by conventional techniques using appropriate isotopic variants of suitable reagents.
  • pharmaceutically acceptable salts includes pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.
  • “Pharmaceutically acceptable acid addition salt” refers to a salt formed with an inorganic acid or an organic acid that can retain the biological effectiveness of the free base without other side effects.
  • Inorganic acid salts include but are not limited to hydrochloride, hydrobromide, sulfate, nitrate, phosphate, etc .
  • organic acid salts include but are not limited to formate, acetate, 2,2-dichloroacetate , Trifluoroacetate, propionate, hexanoate, caprylate, caprate, undecylenate, glycolate, gluconate, lactate, sebacate, adipic acid Salt, glutarate, malonate, oxalate, maleate, succinate, fumarate, tartrate, citrate, palmitate, stearate, oleate , Cinnamate, laurate, malate, glutamate, pyroglutamate, aspartate, benzoate, methanesulfonate, benzen
  • “Pharmaceutically acceptable base addition salt” refers to a salt formed with an inorganic base or an organic base that can maintain the bioavailability of the free acid without other side effects.
  • Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like.
  • Preferred inorganic salts are ammonium, sodium, potassium, calcium and magnesium salts.
  • Salts derived from organic bases include, but are not limited to, the following salts: primary amines, secondary amines, and tertiary amines, substituted amines, including natural substituted amines, cyclic amines, and basic ion exchange resins , Such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, triethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, bicyclo Hexamine, lysine, arginine, histidine, caffeine, procaine, choline, betaine, ethylenediamine, glucosamine, methylglucosamine, theobromine, purine, piperazine, piper Pyridine, N-ethylpiperidine, polyamine resin, etc.
  • Preferred organic bases include isopropylamine, diethylamine, ethanolamine, tri
  • pharmaceutical composition refers to a formulation of a compound of the present invention and a medium generally accepted in the art for delivering a biologically active compound to a mammal (eg, a human).
  • the medium includes a pharmaceutically acceptable carrier.
  • the purpose of the pharmaceutical composition is to promote the administration of living organisms, which facilitates the absorption of active ingredients and thus exerts biological activity.
  • pharmaceutically acceptable refers to a substance (such as a carrier or diluent) that does not affect the biological activity or properties of the compound of the present invention, and is relatively non-toxic, that is, the substance can be administered to an individual without causing harmful organisms React or interact with any components contained in the composition in an undesirable manner.
  • pharmaceutically acceptable excipients include, but are not limited to, any adjuvants, carriers, excipients, glidants, supplements, etc. that are approved by the relevant government regulatory agency to be acceptable for human or domestic animal use Sweeteners, diluents, preservatives, dyes / colorants, flavoring agents, surfactants, wetting agents, dispersants, suspending agents, stabilizers, isotonic agents, solvents or emulsifiers.
  • the "tumor” in the present invention includes, but is not limited to, Nunan syndrome, Leopard syndrome, juvenile myelomonocytic leukemia, neuroblastoma, sarcoma, melanoma, articular chondroma, cholangiocarcinoma, leukemia, breast cancer, stomach Intestinal stromal tumor, histiocytic lymphoma, non-small cell lung cancer, small cell lung cancer, esophageal cancer, pancreatic cancer, lung squamous cell carcinoma, lung adenocarcinoma, breast cancer, prostate cancer, liver cancer, skin cancer, epithelial cell carcinoma, cervical Cancer, ovarian cancer, intestinal cancer, nasopharyngeal cancer, brain cancer, bone cancer, kidney cancer, oral cancer / head cancer, neuroblastoma, squamous cell carcinoma of the head and neck, anaplastic large cell lymphoma or glioblastoma Tumors and other diseases.
  • preventive include causing a patient to reduce the likelihood of the occurrence or worsening of a disease or disorder.
  • treatment and other similar synonyms include the following meanings:
  • an "effective amount” for treatment is the amount of a composition comprising a compound disclosed herein required to provide a clinically significant disease relief effect. Techniques such as dose escalation tests can be used to determine the effective amount suitable for any individual case.
  • administer refers to a method capable of delivering a compound or composition to a desired site for biological action. These methods include, but are not limited to, oral route, transduodenal route, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion), local administration, and transrectal administration.
  • parenteral injection including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion
  • local administration and transrectal administration.
  • transrectal administration including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion
  • Those skilled in the art are familiar with application techniques that can be used in the compounds and methods described herein, such as in Goodman and Gilman, The Pharmacological Basis of Therapeutics, current. The ones discussed in Easton, Pa.
  • the compounds and compositions discussed herein are administered orally.
  • the terms “pharmaceutical combination”, “drug combination”, “combination drug”, “administration of other treatments”, “administration of other therapeutic agents”, etc. refer to pharmaceutical treatments obtained by mixing or combining more than one active ingredient, It includes fixed and unfixed combinations of active ingredients.
  • the term “fixed combination” refers to the simultaneous administration of at least one compound described herein and at least one synergistic agent to a patient in the form of a single entity or a single dosage form.
  • the term “unfixed combination” refers to simultaneous administration, co-administration, or sequential administration of at least one compound described herein and at least one synergistic formulation to a patient in the form of separate entities. These also apply to cocktail therapy, such as the administration of three or more active ingredients.
  • the intermediate compound functional group may need to be protected by an appropriate protecting group.
  • Such functional groups include hydroxyl, amino, mercapto and carboxylic acid.
  • Suitable hydroxy protecting groups include trialkylsilyl or diarylalkylsilyl (e.g. tert-butyldimethylsilyl, tert-butyldiphenylsilyl or trimethylsilyl) , Tetrahydropyranyl, benzyl and so on.
  • Suitable protecting groups for amino, amidino and guanidino include t-butoxycarbonyl, benzyloxycarbonyl and the like.
  • Suitable thiol protecting groups include -C (O) -R "(where R” is alkyl, aryl or aralkyl), p-methoxybenzyl, trityl and the like.
  • Suitable carboxy protecting groups include alkyl, aryl or aralkyl esters.
  • Protecting groups can be introduced and removed according to standard techniques known to those skilled in the art and as described herein. The use of protective groups is detailed in Greene, T.W. and P.G.M. Wuts, Protective Groups in Organis Synthesis, (1999), 4th Ed., Wiley.
  • the protective group may also be a polymer resin.
  • the compound of formula I provided by the present invention can be prepared by the following method: nucleophilic substitution reaction of formula Ib and formula Ic to obtain formula Id; substitution reaction of formula Id and formula Ie to obtain formula If; and formula If Acid deprotection to obtain the compound of formula I:
  • Src Hommolgy-2 phosphatase is a protein tyrosine phosphatase encoded by the PTPN11 gene, which promotes a variety of cellular functions, including proliferation, differentiation, cell cycle maintenance, and migration.
  • SHP2 is involved in signaling via the Ras-mitogen-activated protein kinase, JAK-STAT, or phosphoinositide 3-kinase-AKT pathway.
  • SHP2 mediates the activation of Erk1 and Erk2MAP kinases of receptor tyrosine kinases such as ErbB1, ErbB2 and c-Met.
  • SHP2 has two N-terminal Src homolgy2 domains (N-SH2 and C-SH2), a catalytic domain (PTP) and a C-terminal tail.
  • the two SH2 domains control the subcellular localization and functional regulation of SHP2.
  • the molecule exists in an inactive conformation, inhibiting its own activity via a binding network involving residues from N-SH2 and PTP domains.
  • SHP2 binds to specific tyrosine-phosphorylation sites on docking proteins such as Gab1 and Gab2 via its SH2 domain. This caused a conformational change, leading to SHP2 activation.
  • SHP2 is an important downstream signaling molecule for various receptor tyrosine kinases, including platelet-derived growth factor (PDGF-R), fibroblast growth factor (FGF-R) and epidermal growth factor (EGF-R) receptors.
  • PDGF-R platelet-derived growth factor
  • FGF-R fibroblast growth factor
  • EGF-R epidermal growth factor
  • SHP2 is also an important downstream signaling molecule that activates the mitogen-activated protein (MAP) kinase pathway, which can lead to cell transformation (a necessary condition for cancer development).
  • MAP mitogen-activated protein
  • Noonan Syndrome (NS) and Leopard Spot Syndrome (LS) -PTPNll mutations cause LS (Multiple Pigmented Spot Moles Syndrome), abnormal ECG conduction, excessive distance between eyes, pulmonary valve stenosis, abnormal genitalia, growth retardation Sensorineural deafness) and NS (including heart defects, craniofacial deformities, and short stature congenital abnormalities).
  • LS Multiple Pigmented Spot Moles Syndrome
  • ECG conduction abnormal ECG conduction
  • excessive distance between eyes pulmonary valve stenosis
  • abnormal genitalia abnormal genitalia, growth retardation Sensorineural deafness
  • NS including heart defects, craniofacial deformities, and short stature congenital abnormalities
  • Y62D and Y63C in the protein are the most common mutations. These two mutations affect the non-catalytically active conformation of SHP2 without interfering with the binding of phosphatase to its phosphorylated signal partner.
  • JMML Juvenile myelomonocytic leukemia
  • SHP2 Juvenile myelomonocytic leukemia
  • MPD childhood myelodysplastic disease
  • Acute myeloid leukemia-already in ⁇ 10% of pediatric acute leukemias such as myelodysplastic syndrome (MDS), ⁇ 7% of B cell acute lymphoblastic leukemia (B-ALL) and ⁇ 4% of acute myeloid leukemia (AML) PTPN11 mutation was identified.
  • MDS myelodysplastic syndrome
  • B-ALL B cell acute lymphoblastic leukemia
  • AML acute myeloid leukemia
  • NS and leukemia mutations cause amino acid changes at the interface formed by the N-SH2 and PTP domains in the self-inhibiting SHP2 conformation, disrupting inhibitory intramolecular interactions, resulting in excessive activity of the catalytic domain.
  • SHP2 acts as a positive regulator in receptor tyrosine kinase (RTK) signaling.
  • RTK receptor tyrosine kinase
  • Cancers containing changes in RTK include esophageal cancer, breast cancer, lung cancer, colon cancer, gastric cancer, glioma, Head and neck cancer.
  • Esophageal cancer is a malignant disease of the esophagus.
  • squamous cell carcinoma ⁇ 50%)
  • adenocarcinoma adenocarcinoma
  • RTK expression a high rate of RTK expression in esophageal adenocarcinoma and squamous cell carcinoma. Therefore, the SHP2 inhibitors of the present invention can be used in innovative therapeutic strategies.
  • TNBC Three-negative breast cancer
  • EGFR Epidermal growth factor receptor I
  • SHP2 can be a promising treatment for breast cancer.
  • Lung cancer-NSCLC is currently an important cause of cancer-related mortality. It accounts for about 85% of lung cancer (mainly adenocarcinoma and squamous cell carcinoma). Although cytotoxic chemotherapy is still an important part of treatment, targeted therapy based on genetic changes in tumors such as EGFR and ALK is more likely to benefit from targeted therapy.
  • Colon cancer-About 30% to 50% of colorectal tumors are known to have mutated (abnormal) KRAS, and BRAF mutations occur in 10 to 15% of colorectal cancers.
  • mutated (abnormal) KRAS mutated (abnormal) KRAS
  • BRAF mutations occur in 10 to 15% of colorectal cancers.
  • Gastric cancer is one of the most popular types of cancer. Abnormal expression of tyrosine kinase (as reflected by abnormal tyrosine phosphorylation in gastric cancer cells) is known in the art. Three receptor tyrosine kinases are often amplified in gastric cancer, namely c-met (HGF receptor), FGF receptor 2 and erbB2 / neu. Therefore, the destruction of different signaling pathways can promote the progress of different types of gastric cancer.
  • HGF receptor c-met
  • FGF receptor 2 FGF receptor 2
  • erbB2 / neu erbB2 / neu
  • Neuroblastoma is a pediatric tumor of the developing sympathetic nervous system, accounting for about 8% of childhood cancers.
  • the genomic changes of the anaplastic lymphoma kinase (ALK) gene have been proposed to promote the pathogenesis of neuroblastoma.
  • SCHN Head and neck squamous cell carcinoma
  • SCCHN head and neck squamous cell carcinoma
  • Blocking of EGFR signal leads to inhibition of receptor stimulation, cell proliferation, invasion and metastasis decline. Therefore, in SCCHN, EGFR is the best target for new anti-cancer therapies.
  • the present invention relates to compounds capable of inhibiting SHP2 activity.
  • the invention also provides a preparation method of the compound of the invention and a pharmaceutical preparation containing the compound.
  • Another aspect of the invention relates to a method of treating a disease or disorder mediated by SHP2-, which method comprises the step of administering to a patient in need thereof a therapeutically effective amount of a compound of formula I according to the invention.
  • the invention relates to the method as described above, wherein the SHP2-mediated disease or disorder is a cancer selected from but not limited to the following: JMML, AML, MDS, B-ALL, neuroblastoma , Esophageal cancer, breast cancer, lung cancer, colon cancer, stomach cancer, head and neck cancer.
  • a cancer selected from but not limited to the following: JMML, AML, MDS, B-ALL, neuroblastoma , Esophageal cancer, breast cancer, lung cancer, colon cancer, stomach cancer, head and neck cancer.
  • the present invention relates to a method for treating a disease or condition selected from NS, LS, JMML, AML, MDS, B-ALL, neuroblastoma, esophageal cancer, breast cancer, lung cancer, colon cancer , Stomach cancer, head and neck cancer.
  • the SHP2 inhibitors of the present invention can be combined with other pharmacologically active compounds or with two or more other pharmacologically active compounds, especially in the treatment of cancer.
  • the compound of formula (I) of the present invention or a pharmaceutically acceptable salt thereof may be administered simultaneously, sequentially, or separately in combination with one or more substances selected from the group consisting of chemotherapeutic agents, such as mitotic inhibitors, such as yew Alkanes, vinca alkaloids, paclitaxel, docetaxel, vinca alkaloids, vinblastine, vinorelbine or vinflunine, other anticancer agents such as cisplatin, 5-fluorouracil or 5-fluoro-2-4 (1H, 3H) -pyrimidinedione (5FU), flutamide or gemcitabine.
  • chemotherapeutic agents such as mitotic inhibitors, such as yew Alkanes, vinca alkaloids, paclitaxel, docetaxel, vinca alkaloids, vinblastine
  • Some combinations can provide significant advantages in therapy, including synergistic activity.
  • the present invention relates to the method as described above, wherein the compound is administered parenterally.
  • the invention relates to the method as described above, wherein the compound is administered intramuscularly, intravenously, subcutaneously, orally, pulmonaryly, intrathecally, topically, or intranasally.
  • the invention relates to the method as described above, wherein the compound is administered systemically.
  • the invention relates to the method as described above, wherein the patient is a mammal.
  • the invention relates to the method as described above, wherein the patient is a primate.
  • the invention relates to the method as described above, wherein the patient is a human.
  • the present invention relates to a method of treating an SHP2-mediated disease or disorder, the method comprising the steps of: administering to a patient in need thereof a therapeutically effective amount of a chemotherapeutic agent and, for example, a therapeutically effective amount of Formula I of the present invention Combination of compounds.
  • a novel SHP2 inhibitor and its preparation and application are provided.
  • the inhibitor has a high inhibitory activity against SHP2.
  • the starting materials used in the following examples can be purchased from chemical distributors such as Aldrich, TCI, Alfa Aesar, Pide, and Angie, or can be synthesized by known methods.
  • the ice bath refers to -5 ° C to 0 ° C
  • the room temperature refers to 10 ° C to 30 ° C
  • the reflux temperature generally refers to the reflux temperature of the solvent under normal pressure.
  • Overnight reaction generally refers to a time of 8-15 hours.
  • the specific operating temperature is not limited, all at room temperature.
  • the separation and purification of the intermediate and final products are performed by normal phase or reverse phase chromatography column separation or other suitable methods.
  • Normal phase flash chromatography columns use ethyl acetate and n-hexane or methanol and methylene chloride as mobile phases.
  • Reversed-phase preparative high-pressure liquid chromatography (HPLC) uses a C18 column and UV detection at 214nm and 254nm.
  • the mobile phase is A (water and 0.1% formic acid), B (acetonitrile) or mobile phase A (water and 0.1% carbonic acid). Ammonium hydrogen), B (acetonitrile).
  • LCMS instrument Pump Agilent 1260 UV detector: Agilent 1260 DAD Mass Spectrometer API 3000
  • Step 1 Add 1-oxo-1,3-dihydrospiro [indene-2,4'-piperidine] -1'-carboxylic acid tert-butyl ester (1.51g, 5mmol) in a dry 100mL single-necked bottle, Tetraethyl titanate (6.84 g, 30 mmol), (R)-(+)-tert-butylsulfinamide (2.41 g, 20 mmol) was stirred and reacted under heating and reflux for 15 hours. After cooling to room temperature, saturated brine (60 mL) was added to the reaction residue, after which the resulting mixture was stirred for 15 minutes and then filtered through celite.
  • Step 2 Add (R, Z) -1-((tert-butylsulfinyl) imino) -1,3-dihydrospiro [indene-2,4'-piper Pyridine] -1'-carboxylic acid tert-butyl ester (A1-1, 0.802 g, 2 mmol), THF (10 mL), after cooling to 0 ° C, lithium borohydride (66 mg, 3 mmol) was then added. The resulting mixture was continuously stirred and reacted for 1 hour. Methanol was slowly added to quench excess borohydride, filtered, concentrated, and volatiles were removed under reduced pressure.
  • Step 3 Add (S) -1-((((R) -tert-butylsulfinyl) amino) -1,3-dihydrospiro [indene-2,4'- Piperidine] -1'-carboxylic acid tert-butyl ester (A1-2, 0.406 g, 1 mmol), dichloromethane (5 mL), trifluoroacetic acid (1 mL) and the resulting mixture was stirred at room temperature for 1 hour. A saturated aqueous solution of Na 2 CO 3 was added until pH 7, and the aqueous mixture was extracted with DCM (3 ⁇ 30 mL).
  • Step 1 Add 4-cyanopiperidine-1-carboxylic acid tert-butyl ester (1.05 g, 5 mmol) and THF (20 mL) to a dry 100 mL flask in sequence. Under nitrogen, the mixture was cooled to -78 ° C, and then 2M LDA (3.3 mL, 6.5 mmol) was slowly added to the reaction mixture. The reaction mixture was allowed to react for 1 hour, then 3-bromo-2- (bromomethyl) pyridine (1.24 g, 5 mmol) was added thereto, and then the reaction mixture was allowed to continue to react for 2 hours.
  • 4-((3-bromopyridin-2-yl) methyl) -4-cyanopiperidine-1'-carboxylic acid tert-butyl ester A2 -1,379 mg, 1 mmol
  • triethylamine 404 mg, 4 mmol
  • Step 3 Add 5-oxo-5,7-dihydrospiro [cyclopenta [b] pyridine-6,4'-piperidine] -1'-carboxylic acid tert-butyl in a dry 100mL single-necked bottle Ester (A2-2, 0.302g, 1mmol), tetraethyl titanate (1.37g, 6mmol), (R)-(+)-tert-butylsulfinamide (0.480g, 4mmol) was stirred and heated under reflux 15 hours. After cooling to room temperature, saturated brine (15 mL) was added to the reaction residue, after which the resulting mixture was stirred for 15 minutes and then filtered through celite.
  • Step 4 Add (R, Z) -5-((tert-butylsulfinyl) imino) -5,7-dihydrospiro [cyclopentadieno [b] in a dry 100mL single-necked bottle Pyridine-6,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A2-3, 0.20g, 0.491mmol), THF (50mL), after cooling to °C, then lithium borohydride (0.018g, 0.737 mmol). The resulting mixture was continuously stirred and reacted for 1 hour. Methanol was slowly added to quench excess borohydride, filtered, concentrated, and volatiles were removed under reduced pressure.
  • Step 5 Add (S) -5-((R) -tert-butylsulfinamido) -5,7-dihydrospiro [cyclopenta [b] pyridine- in a dry 50mL single-necked bottle 6,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A2-4, 0.100g, 0.245mmol), dichloromethane (5mL), trifluoroacetic acid (1mL) The resulting mixture was stirred at room temperature for reaction 1 hour. A saturated aqueous solution of Na 2 CO 3 was added until pH 7, and the aqueous mixture was extracted with DCM (3 ⁇ 30 mL).
  • intermediate (S) -5,7-dihydrospiro [cyclopenta [b] pyridine-6,4 ' -Piperidin] -5-amine (A4).
  • LCMS m / z 204.1 [M + H] + .
  • Step a sequentially added A6-1 (11.1g, 60mmol) and NaBH 4 (2.51g, 66mmol) To a 1L flask containing 300mL THF, and in 60mL H 2 O and reacted at 20 °C 2 hours. After spot detection, the raw materials were reacted, quenched with saturated NH4Cl solution, diluted with water, and extracted with ethyl acetate. The combined organic phase was washed with saturated NaCl solution, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a white solid 2- Bromo-pyridin-3-ylmethanol (A6-2, 11.2g, yield: 100%)
  • Step 2 Add (A6-2, 6.73g, 36mmol) and Ms 2 O (6.96g, g, 40mmol) to a dry 250mL single-necked flask containing 120mL of dichloromethane sequentially, cool to 0 ° C, and then slowly add TEA (5.45g, 54mmol) was warmed to room temperature and stirred for 3 hours.
  • Step three To a dry 500 mL three-necked flask, sequentially add 4-ethoxycarbonylpiperidine-1-carboxylic acid tert-butyl ester (9.3 g, 36.2 mmol) and THF (133 mL). Under nitrogen, the mixture was cooled to -70 ° C, and then 2M LDA (21.1 mL, 42.3 mmol) was slowly added to the reaction mixture.
  • reaction mixture was allowed to react for 1 hour, then (2-bromopyridin-3-yl) methyl methanesulfonate (A6-3, 8.0 g, 30.2 mmol) dissolved in 65 mL of THF was added thereto, and then the reaction mixture was continued After 0.5 hours of reaction, the temperature was slowly raised to room temperature and stirred for 1 hour. After the reaction was completed, saturated brine was added thereto for quenching and extraction with ethyl acetate. The combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure.
  • Step 4 Add 4-ethoxycarbonyl-4- (2-bromo-3-pyridyl) methyl-piperidine-1-carboxylic acid tert-butyl ester (A6-4,7.85g, 18.4 mmol) and THF (120 mL). Under nitrogen, the mixture was cooled to -70 ° C, and then 2.5M n-butyllithium (11 mL, 27.6 mmol) was slowly added to the reaction mixture. The reaction mixture was allowed to react for 1.5 hours. After the reaction was completed, saturated aqueous ammonium chloride solution was added to quench the solution, and then saturated brine was added to dilute and separate the liquid.
  • Step 5 Refer to the synthesis method of Intermediate A2-3 and use Intermediate A6-5 instead of Intermediate A2-2 to obtain intermediate (R, Z) -7-((tert-butylsulfinyl) imino) -5,7-dihydrospiro [cyclopenta [b] pyridine-6,4′-piperidine] -1′-carboxylic acid tert-butyl ester (A6-6,1.46 g, yield: 68%).
  • Step 6 Add (R, Z) -7-((tert-butylsulfinyl) imino) -5,7-dihydrospiro [cyclopentadieno [b] in a dry 50mL three-necked bottle Pyridine-6,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A6-6,530mg, 1.31mmol), THF (10mL), cooled to -70 ° C under nitrogen, then slowly added 1.5 to the reaction mixture M Diisobutylaluminum hydride in toluene (1.3 mL, 1.95 mmol).
  • reaction mixture After allowing the reaction mixture to react for 0.5 hours, it was slowly warmed to room temperature, quenched with saturated aqueous potassium sodium tartrate solution and stirred for 0.5 hours. Extracted with ethyl acetate, the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure.
  • Step 7 Add (S) -7-((R) -tert-butylsulfinamido) -5,7-dihydrospiro [cyclopenta [b] pyridine- in a dry 25mL single-necked bottle 6,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A6-7,312mg, 0.766mmol), MeOH (5mL), 4M HCl / 1,4-dioxane solution (3.83mL, 15.3mmol), then The temperature was raised to 50 ° C for 8 hours to obtain a white suspension, which was concentrated under reduced pressure to obtain a white solid (S) -5,7-dihydrospiro [cyclopenta [b] pyridine-6,4'-piperidine] -7-amine HCl salt (A6,232 mg, yield: 97%).
  • Step 1 Refer to the synthesis method of intermediate A2 and use the raw material 4-fluorobenzyl bromide instead of the raw material 3-bromo-2- (bromomethyl) pyridine to obtain the intermediate 4-ethoxycarbonyl-4- (4-fluorobenzyl ) -Piperidine-1-carboxylic acid tert-butyl ester A7-1.
  • Step 2 To a 100 mL single-necked flask containing 20 mL of methanol and 20 mL of water, sequentially add the compound 4-ethoxycarbonyl-4- (4-fluorobenzyl) -piperidine-1-carboxylic acid tert-butyl ester (A7-1, 3.40 g , 9.30 mmol) and sodium hydroxide (1.86 g, 46.5 mmol). The reaction solution was reacted at 70 ° C for 17 hours. After cooling to room temperature, it was concentrated under reduced pressure to remove volatiles.
  • Step 4 Refer to the synthesis method of intermediate A2 and use intermediate A7-3 instead of intermediate A2-2 to obtain intermediate (R, Z) -1-((tert-butylsulfinyl) imino) -6 -Fluoro-1-oxo-1,3-dihydrospiro [indene-2,4'-piperidine] -1'-formate A7-4.
  • Step 5 Add compound (R, Z) -1-((tert-butylsulfinyl) imino) -6-fluoro-1-oxo-1,3 to the round bottom flask at -50 ° C -Dihydrospiro [indene-2,4'-piperidine] -1'-formate (A7-4, 1.59g, 3.76mmol), tetrahydrofuran / water (98: 2, 32mL) and sodium borohydride (427mg , 11.3mmol), the reaction solution was heated to 20 °C under stirring for 3 hours. TLC spotting reaction is complete.
  • reaction solution was diluted with water (30 mL) and extracted three times with ethyl acetate (30 mL). The combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure.
  • Step 6 Refer to the synthesis method of intermediate A3, use intermediate A7-5 instead of intermediate A1-2, to obtain intermediate (S) -6-fluoro-1,3-dihydrospiro [indene-2,4'- Piperidine] -1-amine A7.
  • LCMS m / z 220.1 [M + H] + .
  • Step 1 Dissolve 4-bromonicotinic acid hydrobromide (A8-1, 2.5g, 9.36mmol) in methanol (50mL), and add sodium borohydride (0.72g, 18.93mmol, batchwise) under ice bath 2.0eq), react at 0 degree for 1 hour, the reaction is completed. Under an ice bath, saturated aqueous ammonium chloride solution (50 mL) was added to quench the reaction, and ethyl acetate was added for extraction (100 mL ⁇ 2). The organic phases were combined and washed with saturated sodium chloride water.
  • Step 2 Dissolve (4-bromopyridin-3-yl) methanol (A8-2, 1.7g, 9.04mmol) in dichloromethane (100mL), and add triethylamine (2.30g, 22.7mmol) under ice bath . Methanesulfonic anhydride (1.95g, 11.2mmol) was added in batches under nitrogen, and the reaction was completed after 2 hours of reaction at 0 degrees. Under ice bath, saturated sodium chloride (50 mL) was added to quench the reaction, and the organic phase was separated and washed with saturated sodium chloride water.
  • Step 3 Dissolve 4-cyanopiperidine-1-carboxylic acid tert-butyl ester (1.61g, 7.67mmol) in anhydrous tetrahydrofuran (60mL), slowly drop into LDA under the protection of dry ice and acetone bath (-78 ° C) under nitrogen (4.6mL, 9.2mmol), the reaction was incubated for 0.5 hours, and a large amount of white solid precipitated in the reaction solution.
  • Step 4 Dissolve 4-((4-bromopyridin-3-yl) methyl) -4-cyanopiperidine-1-carboxylic acid tert-butyl ester (A8-4, 1.5g, 3.95mmol) in DMAc / H2O
  • DIPEA 1.34 g, 15.8 mmol
  • Pd (AmPhos) 2 Cl 2 142 mg, 0.20 mmol
  • Step 5 The compound 5-oxo-5,7-dihydrospiro [cyclopenta [c] pyridine-6,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A8-5,750mg, 2.48 mmol) was dissolved in THF (10 mL), and (R)-(+)-tert-butylsulfinamide (390 mg, 3.22 mmol) and tetraethyl titanate (10 mL) were added. The reaction was heated to 90 degrees under reflux with nitrogen for 18 hours. After the reaction was completed, cool to room temperature and add ethyl acetate (50 mL) to dilute it.
  • Step 6 (R, Z) -5-((tert-butylsulfinyl) imino) -5,7-dihydrospiro [cyclopenta [c] pyridine-6,4'-piper Pyridine] -1'-carboxylic acid tert-butyl ester (A8-6,800 mg, 1.98 mmol) was dissolved in THF (50 mL) and cooled to -78 degrees under nitrogen protection. 1.5 M DIBAL-H (2 mL, 3 mmol) was slowly added dropwise to the reaction solution, and the addition was completed. The reaction was completed at -78 degrees for 1 hour.
  • the reaction was quenched with water, saturated potassium sodium tartrate solution (20 mL) was added, and extracted with ethyl acetate (100 mL x 2), the organic phase was washed once with saturated aqueous sodium chloride solution, the organic phase was separated, dried over anhydrous sodium sulfate, and filtered concentrate.
  • Step 7 (S) -5-(((R) -tert-butylsulfinyl) amino) -5,7-dihydrospiro [cyclopenta [c] pyridine-6,4'-piper Pyridine] -1'-carboxylic acid tert-butyl ester (A8-7,680mg, 1.67mmol) was dissolved in MeOH (50mL), cooled to 0 degrees, 4M hydrochloric acid dioxane solution (10mL, 40mmol) was added dropwise, the reaction at room temperature Hours, the reaction is complete.
  • Step 1 Dissolve 2-bromo-6-hydroxybenzaldehyde (5g, 24.9mmol) in DMF (100mL), add anhydrous potassium carbonate (6.88g, 49.8mmol) and 4-methoxybenzyl bromide (5.26g , 26.1 mmol), and reacted at room temperature for 18 hours under nitrogen protection. The reaction was completed. The reaction solution was poured into ice water, and extracted with ethyl acetate (250 mL) twice. The organic phases were combined and washed with saturated saline. The organic phases were separated and dried over anhydrous sodium sulfate, filtered and concentrated.
  • Step 2 Dissolve 2-bromo-6-((4-methoxybenzyl) oxy) benzaldehyde (A9-1,8g, 24.9mmol) in ethanol (100mL) and cool to 0 degree under ice bath . Carefully add sodium borohydride (942 mg, 24.9 mmol) in multiple batches. The reaction was completed at 0 degrees for 0.5 hours. The reaction solution was poured into ice water and extracted twice with ethyl acetate (200 mL).
  • Step 3 Dissolve (2-bromo-6-((4-methoxybenzyl) oxy) phenyl) methanol (A9-2,7.3g, 22.6mmol) in dichloromethane (200mL), ice Cool to 0 degrees under the bath. Carbon tetrabromide (11.2g, 33.9mmol) and triphenylphosphine (8.88g, 33.9mmol) were added under nitrogen protection. The reaction was completed at 0 degrees for 5 hours. The reaction solution was poured into ice water and extracted twice with ethyl acetate (200 mL). The combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated.
  • Step 4 Dissolve tert-butyl 4-cyanopiperidine-1-carboxylate (2.6 g, 12.4 mmol) in anhydrous THF (60 mL) and cool to -78 degrees. 2M LDA (7.5 mL, 14.9 mmol) was slowly added dropwise. -78 degree reaction for 0.5 hour, continue to dropwise add 1-bromo-2- (bromomethyl) -3-((4-methoxybenzyl) oxy) benzene (A9-3, 4g, 10.4mmol) A solution of water in THF (40 mL). -78 degrees of reaction for 2 hours, slowly warming to 0 degrees. The reaction is complete.
  • Step 5 The 4- (2-bromo-6-((4-methoxybenzyl) oxy) benzyl) -4-cyanopiperidine-1-carboxylic acid tert-butyl ester (A9-4, 2.6g , 5.0 mmol) was dissolved in DMAc (100mL) was added H2O (10mL), DIPEA (3.225g , 25.0mmol) and PdCl 2 (AmPhos) 2 (354mg , 0.5mmol). The reaction was completed at 130 degrees under argon for 18 hours. After cooling to room temperature, the reaction solution was poured into ice water and extracted twice with ethyl acetate (300 mL).
  • Step 6 The 4-((4-methoxybenzyl) oxy) -1-oxo-1,3-dihydrospiro [indene-2,4'-piperidine] -1'-carboxylic acid tert-butyl
  • the ester (A9-5, 1.8g, 4.12mmol) was dissolved in THF (20mL), (R) -2-methylpropane-2-sulfinamide (748mg, 6.18mmol) and tetraethyl titanate (50mL) were added ).
  • the reaction was heated to 100 degrees under reflux with nitrogen for 18 hours. The reaction was completed.
  • Step 7 (R, Z) -1-((tert-butylsulfinyl) imino) -4-((4-methoxybenzyl) oxy) -1,3-dihydrospiro [ Indene-2,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A9-6, 1.7 g, 3.15 mmol) was dissolved in THF (100 mL) and cooled to -78 degrees under nitrogen protection. The reaction solution was slowly dropped into 1.5M DIBAL-H (3.15mL, 4.7mmol), and the dropwise addition was completed. The reaction was completed at -78 degrees for 1 hour.
  • the reaction was quenched with water, saturated potassium sodium tartrate solution (50 mL) was added, extracted with ethyl acetate (200 mL x 2), the organic phase was washed once with saturated aqueous sodium chloride solution, the organic phase was separated, dried over anhydrous sodium sulfate, and filtered concentrate.
  • Step 8 (S) -1-(((R) -tert-butylsulfinyl) amino) -4-((4-methoxybenzyl) oxy) -1,3-dihydrospiro [Indene-2,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A9-7, 1.5g, 2.77mmol) was dissolved in MeOH (50mL), cooled to 0 degrees, and 4M hydrochloric acid dioxane was added dropwise The ring solution (10 mL, 40 mmol) was reacted at room temperature for 3 hours, and the reaction was completed.
  • Step 1 Add 2-methyl-3-cyanobromobenzene (3.0g, 15.3mmol, 1.0eq) and N-bromosuccinimide (2.72g, 15.3mmol, 1.0) to a 100mL round bottom flask eq), dibenzoyl peroxide (371mg, 1.53mmol, 0.1eq) and carbon tetrachloride (40mL), stirred at 80 °C for 16 hours, the reaction solution was concentrated under reduced pressure to obtain a residue, the residue was dissolved in ethyl acetate The ester (200 mL) was washed twice with 2N aqueous NaOH solution (50 mL) and once with saturated brine (50 mL), dried over anhydrous sodium sulfate, and filtered.
  • Step 2 To a 100 mL round-bottom flask, add tetrahydrofuran (30 mL) and 4-cyanopiperidine-1-carboxylic acid tert-butyl ester (1.84 g, 8.73 mmol, 1.2 eq) sequentially, cool to -78 ° C, and then add 2.0 M LDA (5.1 mL, 10.2 mmol, 1.4 eq), stirred at -78 ° C for one hour.
  • tetrahydrofuran (30 mL) and 4-cyanopiperidine-1-carboxylic acid tert-butyl ester (1.84 g, 8.73 mmol, 1.2 eq) sequentially, cool to -78 ° C, and then add 2.0 M LDA (5.1 mL, 10.2 mmol, 1.4 eq), stirred at -78 ° C for one hour.
  • Step 3 To a dry 100mL round bottom three-necked flask, sequentially add tert-butyl-4- (2-bromo-6-cyanobenzyl) -4-cyanopiperidine-1-carboxylate (A10-2 , 1.5g, 3.71mmol, 1.0eq), Pd (AmPhos) 2 Cl 2 (262mg, 0.37mmol. 0.1eq), diisopropylethylamine (2.4g, 18.5mmol, 5.0eq), N, N- Dimethylacetamide (30 mL) and water (4 mL). In the case of stirring, it was replaced with nitrogen three times, then heated to 140 ° C. and reacted for 16 hours.
  • tert-butyl-4- (2-bromo-6-cyanobenzyl) -4-cyanopiperidine-1-carboxylate A10-2 , 1.5g, 3.71mmol, 1.0eq
  • Pd (AmPhos) 2 Cl 2 (262mg, 0.37mmol.
  • Step 4 Add tert-butyl-4-cyano-1-oxo-1,3-dihydrospiro [indene-2,4'-piperidine] -1-carboxylate to a dry 100mL single-necked bottle in sequence (A10-3, 900 mg, 2.76 mmol), titanium tetraethoxylate (3.78 g, 16.6 mmol), (R)-(+)-tert-butylsulfinamide (401 mg, 3.31 mmol) and tetrahydrofuran (20 mL) in The reaction was stirred with heating under reflux for 16 hours.
  • Step 5 Add tetrahydrofuran (15mL) and (R, Z) -1-((tert-butylsulfinyl) imine) -4-cyano-1,3-dihydrospiro to a dry 50mL three-necked flask in this order [Inden-2,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A10-4, 980 mg, 2.28 mmol, 1.0 eq).
  • diisobutylaluminum hydride (6.8 mL, 1.5 M in toluene, 10.3 mmol, 4.5 eq) was added dropwise, and stirring was continued for half an hour. Then it was raised to 0 ° C and stirring was continued for half an hour. After the reaction was completed, it was quenched with potassium sodium tartrate (4g dissolved in 20mL water), stirred for half an hour, extracted with ethyl acetate (30mL * 3), and the resulting organic phase was washed with saturated brine (30mL), anhydrous sodium sulfate Dry, filter, and concentrate under reduced pressure.
  • Step 1 Dissolve 2,2,6,6-tetramethylpiperidine (5.33g, 37.8mmol) in 100mL of tetrahydrofuran, protected by argon, cool the dry ice acetone bath to -65 degrees, add n-butyl lithium (2.5 M, 15.8mL, 39.5mmol), and then kept at 0 degrees for 30 minutes, then cooled to -65 degrees again, added dropwise a mixture of 2-bromopyrazine (5.0g, 31.4mmol) and 10mL of tetrahydrofuran, kept for 30 minutes, added DMF (5.75g, 78.6mmol) and 10mL of tetrahydrofuran mixed solution, stirring was continued for 2 hours, the reaction was complete.
  • 2-bromopyrazine 5.0g, 31.4mmol
  • DMF 5.75g, 78.6mmol
  • 10mL of tetrahydrofuran mixed solution stirring was continued for 2 hours, the reaction was complete.
  • Step 2 Dissolve (3-bromopyridin-2-yl) methanol (A11-1, 4.33g, 22.9mmol) in 80mL of diethyl ether, protect with nitrogen, add PBr 3 (6.83g, 25.2mmol) dropwise at 0 degrees, then Heated to reflux at about 40 degrees, reacted for 4 hours, the reaction was completed.
  • Step 3 Dissolve ethyl N-Boc-4-piperidinecarboxylate (6.81g, 26.5mmol) in 160mL of anhydrous tetrahydrofuran. Under the protection of argon, dry ice acetone bath to cool to -65 degrees, slowly drop LDA (2.0 M, 16.8mL, 33.6mmol), keep the reaction for 1 hour; continue dropping 2-bromo-3- (bromomethyl) pyrazine (A11-2, 5.14g, 24.1mmol) in anhydrous tetrahydrofuran (-65 degrees) 30mL) solution, continue to incubate the reaction for 2 hours, the reaction is complete.
  • LDA 2.0 M, 16.8mL, 33.6mmol
  • Step 4 Add 1- (tert-butyl) -4-ethyl 4-((3-bromopyrazin-2-yl) methyl) piperidine-1,4-dicarboxylate (A11-3, 5.8 g, 13.5mmol) dissolved in 300mL tetrahydrofuran, under the protection of argon, dry ice acetone bath was cooled to -65 degrees, n-butyllithium (2.5M, 8.2mL, 20.3mmol) was added dropwise, naturally heated to -10 degrees, keep warm After 3 hours, the reaction was complete.
  • reaction solution was poured into 300 mL of iced NH 4 Cl aqueous solution, extracted three times with ethyl acetate, and the organic phase was washed with saturated brine once.
  • the organic phases were combined, dried over anhydrous sodium sulfate, filtered, concentrated, and passed through a column to obtain a yellow solid 5-oxo -5,7-dihydrospiro [cyclopenta [b] pyrazine-6,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A11-4, 2.23g, yield: 54.4% ).
  • Step 5 The 5-oxo-5,7-dihydrospiro [cyclopenta [b] pyrazine-6,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A11-4, 2.23 g, 7.35 mmol) was dissolved in 350 mL of tetrahydrofuran, and (R) -2-methylpropane-2-sulfinamide (980 mg, 8.09 mmol) and tetraethyl titanate (21.8 g, 95.6 mmol) were added. The reaction was heated to 80 degrees under reflux with nitrogen for 10 hours. The reaction is complete.
  • Step 6 (R, Z) -5-((tert-butylsulfinyl) imino) -5,7-dihydrospiro [cyclopenta [b] pyrazine-6,4'- Piperidine] -1'-carboxylic acid tert-butyl ester (A11-5, 2.38g, 5.85mmol) was dissolved in 280mL of tetrahydrofuran, under the protection of argon, the temperature of the dry ice acetone bath was reduced to -65 degrees. 1.5M DIBAL-H (5.07mL, 7.61mmol) was slowly added dropwise, and the incubation was continued for 2 hours, and the reaction was completed.
  • DIBAL-H 5.07mL, 7.61mmol
  • Step 7 (S) -5-(((R) -tert-butylsulfinyl) amino) -5,7-dihydrospiro [cyclopenta [b] pyrazine-6,4 ' -Piperidine] -1'-carboxylic acid tert-butyl ester crude product (A11-6, 2.48g, 5.85mmol) was dissolved in 120mL methanol, under the protection of argon, the ice water was cooled to about 0 degrees, 4M hydrochloric acid dioxane was added dropwise The ring solution (18 mL, 72 mmol) was reacted at room temperature for 2 hours, and the reaction was completed.
  • Step 1 Add 2-methyl-3-methoxybromobenzene (2.0g, 9.95mmol, 1.0eq) and N-bromosuccinimide (1.77g, 9.95mmol) to a 100mL round bottom flask in sequence 1.0eq), dibenzoyl peroxide (241mg, 0.995mmol, 0.1eq) and carbon tetrachloride (40mL), stirred at 80 °C for 16 hours, the reaction solution was concentrated under reduced pressure to obtain a residue, the residue was dissolved in acetic acid Ethyl acetate (200mL), washed twice with 2N aqueous NaOH solution (50mL * 2), once with saturated brine (50mL), dried over anhydrous sodium sulfate, and filtered.
  • the filtrate was concentrated under reduced pressure to obtain a crude product.
  • the crude product was purified by silica gel chromatography to obtain a white solid 1-bromo-2- (bromomethyl) -3-methoxybenzene (A12-1, 2.0 g, yield: 71.8%).
  • Step 2 To a 100 mL round-bottom flask, add tetrahydrofuran (30 mL) and N-Boc-4-cyanopiperidine (1.8 g, 8.57 mmol, 1.2 eq) in sequence, cool to -78 ° C, and then add 2.0 M LDA ( 5mL, 10mmol, 1.4eq), stirred at -78 ° C for one hour. Then, a solution of 1-bromo-2- (bromomethyl) -3-methoxybenzene (A12-1, 2.0 g, 7.14 mmol, 1.0 eq) in tetrahydrofuran (15 mL) was added, and stirred at -78 ° C for 0.5 hour.
  • 1-bromo-2- (bromomethyl) -3-methoxybenzene A12-1, 2.0 g, 7.14 mmol, 1.0 eq
  • Step 3 Add N-Boc-4- (2-bromo-6-methoxybenzyl) -4-cyanopiperidine (A12-2, 1.5g, 3.66) to a dry 100mL round bottom three-necked flask in sequence mmol, 1.0 eq), Pd (AmPhos) 2 Cl 2 (259 mg, 0.37 mmol. 0.1 eq), diisopropylethylamine (2.37 g, 18.3 mmol, 5.0 eq), N, N-dimethylacetamide (30 mL) and water (4 mL). In the case of stirring, it was replaced with nitrogen three times, then heated to 140 ° C. and reacted for 16 hours.
  • reaction solution was cooled to room temperature, diluted with ethyl acetate (100 mL) and filtered under reduced pressure, the filter cake was washed with ethyl acetate (20 mL), and the resulting filtrate was washed with saturated brine (30 mL * 3) 3 Next, dry over anhydrous sodium sulfate and filter. The filtrate was concentrated under reduced pressure to obtain a crude product.
  • the crude product was purified by silica gel chromatography to obtain a white solid N-Boc-4-methoxy-1-oxo-1,3-dihydrospiro [indene-2,4'-piperidine] (A12-3, 0.90 g, yield: 74.1%).
  • Step 4 Add N-Boc-4-methoxy-1-oxo-1,3-dihydrospiro [indene-2,4'-piperidine] (A12-3, 900mg, 2.72mmol), tetraethoxytitanium (3.72g, 16.3mmol), (R)-(+)-tert-butylsulfinamide (395mg, 3.26mmol) and tetrahydrofuran (20mL) were stirred and reacted under heating and reflux 16 hours. After cooling to room temperature, saturated brine (60 mL) was added to the reaction residue, after which the resulting mixture was stirred for 15 minutes and then filtered through celite.
  • Step 5 Add tetrahydrofuran (15mL) and N-Boc- (R, Z) -1-((tert-butylsulfinyl) imine) -4-methoxy-1 to a dry 50mL three-necked flask in sequence 3-Dihydrospiro [indene-2,4'-piperidine] (A12-4, 800 mg, 1.84 mmol, 1.0 eq). At -78 ° C, under nitrogen protection, diisobutylaluminum hydride (5.52mL, 1.5M in toluene, 8.28mmol, 4.5eq) was added dropwise, and stirring was continued for half an hour.
  • Step 1 Dissolve 1-tert-butyl 4-ethylpiperidine-1,4-dicarboxylate (10.7g, 41.6mmol, 1.2eq) in THF (50mL), and add LDA (-78 °C) dropwise 2M, 9.8mL, 19.6mmol, 1.1eq), the reaction was stirred at this temperature for 2h, and then 2-chloro-5- (chloromethyl) thiazole (A13-1, 3g, 34.7mmol, 1.0eq) was dissolved in THF ( 10mL) was added dropwise to the system. After the completion of the reaction continued at this temperature for 1.5h. TLC detected a little unreacted.
  • Step 2 1-tert-butyl-4-ethyl 4- (2-chlorothiazol-5-yl) methyl) piperidine-1,4-dicarboxylate (A13-2, 4g, 10.3mmol) Dissolved in THF (100 mL), LDA (2M, 8.5 mL, 16.6 mmol) was added dropwise at -78 ° C. The reaction was completed in 1h. Saturated amine chloride (100 mL) was added to quench the reaction, and ethyl acetate was added for extraction (100 mL x 3).
  • Step 3 Add 2-chloro-4-oxo-4,6-dihydrospiro [cyclopentadiene [d] thiazole-5,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A13-3 , 710mg, 2.07mmol) and Ti (OEt) 4 and (R) -2-methylpropane-2-sulfinamide (276mg, 2.28mmol) were added to the reaction flask, heated to 100 °C for 5h under nitrogen protection. After the reaction was completed, it was cooled to room temperature and diluted with ethyl acetate (50 mL). Saturated brine (15 mL) was added, and a white solid precipitated.
  • Step 4 Combine tert-butyl (E) -4-((tert-butylsulfinyl) imino) -2-chloro-4,6-dihydrospiro [cyclopenta [d] thiazole-5 , 4'-piperidine] -1'-carboxylic acid tert-butyl ester (A13-4, 200 mg, 0.47 mmol) was dissolved in THF (10 mL) and cooled to -78 degrees under nitrogen. The reaction solution was slowly dropped into 1.5 M DIBAL-H (0.5 mL, 0.75 mmol). After the dropwise addition was completed, the reaction was completed at -78 degrees for 1 hour.
  • the reaction was quenched with water, saturated potassium sodium tartrate solution (20 mL) was added, extracted with ethyl acetate (50 mL x 2), the organic phase was washed once with saturated aqueous sodium chloride solution, the organic phase was separated, dried over anhydrous sodium sulfate, and filtered concentrate.
  • Step 5 Combine tert-butyl (S) -4-(((R) -tert-butylsulfinyl) amino) -2-chloro-4,6-dihydrospiro [cyclopenta [d] Thiazole-5,4'-piperidine] -1'-carboxylic acid tert-butyl ester (A13-5, 155mg, 0.35mmol) was dissolved in 4M hydrochloric acid dioxane solution (5mL, 20mmol), reacted at room temperature for 3 hours, the reaction was completed .
  • Step 1 Add 2,4-dichloro-5-iodopyrimidine (1.1 g, 4 mmol) and 20 mL of absolute ethanol to a dry 100 mL flask. Under a nitrogen condition of 0 ° C, a mixture of 80% hydrazine hydrate (601 mg, 12 mmol) was slowly added thereto, and the reaction was further stirred for 1 hour. After the reaction was completed, the mixture was filtered and washed with absolute ethanol to obtain 2-chloro-4-hydrazino-5-iodopyrimidine (B2-1, 850 mg, yield 87.7%).
  • Step 2 Add 2-chloro-4-hydrazino-5-iodopyrimidine (810 mg, 3 mmol) and trimethyl orthoformate (10 mL) to a dry 100 mL flask in sequence. Under nitrogen, the mixture was heated to 85 ° C and stirred for 5 hours.
  • Step 1 Add 2,4-dichloro-5-iodopyrimidine (1.37g, 5mmol), 2,2-dimethoxyethylamine (8.4g, 10mmol) and absolute ethanol to the dried 100mL flask 50mL). Then, at 0 ° C. under nitrogen, triethylamine (1.01 g, 10 mmol) was slowly added dropwise to the reaction mixture, and the mixture was stirred and reacted at room temperature for 10 hours.
  • Step 2 Add 2-chloro-N- (2,2-dimethoxyethyl) -5-iodopyrimidin-4-amine (B3-1, 1.03g, 3mmol) and 10mL to a dry 100mL flask in sequence Concentrated sulfuric acid. Under nitrogen, the mixture was heated to 65 ° C and stirred for 2 hours. After the reaction was completed, the reaction solution was cooled to room temperature, the mixture was slowly poured into ice water, and then the pH was adjusted to about 6-7 with 4M NaOH solution, and filtered to obtain an off-white solid 8-iodoimidazo [1,2-c] pyrimidine- 5-Alcohol (B3-2, 407 mg, 52% yield).
  • Step 3 Add 8-iodoimidazo [1,2-c] pyrimidin-5-ol (B3-2,522mg, 2mmol) and phosphorus oxychloride (8mL) to a dry 50mL single-necked flask in sequence, under nitrogen protection Next, N, N-diisopropylethylamine (1 mL) was slowly added dropwise, after which the mixture was heated to 120 ° C and stirred for 5 hours.
  • reaction solution was cooled to room temperature and concentrated in vacuo, and then quenched by adding saturated sodium bicarbonate solution, extracted with ethyl acetate (3 ⁇ 40 mL), dried over anhydrous sodium sulfate, filtered and concentrated, and the resulting residue
  • the material was purified by silica gel chromatography (0 to 30% gradient ethyl acetate: petroleum ether) to give 5-chloro-8-iodoimidazo [1,2-c] pyrimidine (B3, 360 mg, yield: 55) as a pale yellow solid %).
  • Step 1 Add 3-chloro-4-iodopyridin-2-amine (2.5g, 9.82mmol, 1.0eq), XantPhos (341mg, 0.59mmol.0.06eq), acetic acid to a dry 100mL round bottom three-necked flask Palladium (110mg, 0.49mmol, 0.05eq), DIPEA (3.25mL, 19.6mmol, 2.0q), methyl 3-mercaptopropionate (1.19mL, 10.8mmol, 1.1eq) and 1,4-dioxane ( 32.5mL). While stirring, replace with nitrogen three times, then heat to 100 ° C and react for 3 hours.
  • reaction solution was cooled to room temperature, diluted with ethyl acetate (50 mL) and filtered under reduced pressure, the filter cake was washed with ethyl acetate (25 mL), the resulting filtrate was concentrated in vacuo, and the resulting residue was subjected to silica gel chromatography Purification (0 to 30% gradient of ethyl acetate: petroleum ether) to give yellow solid methyl 3-((2-amino-3-chloropyridin-4-yl) thio) propionate (C1-1, 2.0g , Yield: 78%).
  • Step 2 In a dry 100mL round bottom three-necked flask, dissolve compound C1-1 (2g, 8.11mmol, 1.0eq) in tetrahydrofuran (28mL), under nitrogen protection, dropwise add to the reaction solution at room temperature Sodium ethoxide (2.9g, 8.51mmol, 1.05eq, 20% wt), and then stirred for one hour. After the reaction was completed, dilute with dichloromethane (60 mL) and sonicate for 5 minutes, filter under reduced pressure with suction, and dry the filter cake in vacuo to obtain a yellow solid 2-amino-3chloropyridine-4 sodium sulfide (C1, 1.4g, yield: 89%)
  • Step 1 Combine 2,3-dichloro-4-iodopyridine (1.0g, 3.65mmol, 1.0eq), methyl 3-mercaptopropionate (480mg, 4.02mmol, 1.1eq) and N, N-diisopropyl Ethyl ethylamine (950mg, 7.3mmol) was dissolved in 1,4-dioxane (15mL), replaced with argon three times, under the protection of argon was added palladium acetate (82mg, 0.37mmol, 0.1eq) and XantPhos (211mg , 0.37mmol, 0.1eq) The temperature is increased to 100 degrees after addition, and the reaction is completed for 3 hours.
  • Step 2 Dissolve methyl 3-((2,3-dichloropyridin-4-yl) thio) propionate (C2-1, 100 mg, 0.37 mmol, 1.0 eq) in tetrahydrofuran (10 mL) and ethanol (0.5 mL). Under nitrogen, sodium ethoxide (27mg, 0.39mmol, 1.05eq) was added, and the reaction was carried out at room temperature for 3 hours. The reaction was not completed. Additional sodium ethoxide (27mg, 0.39mmol, 1.05eq) was added, and the reaction was completed at room temperature for 3 hours. Purified water was added and freeze-dried to obtain crude 2,3-dichloropyridine-4-sulfur sodium (C2, 150 mg, yield: 100%) as a pale yellow solid.
  • Step 1 Combine 3-bromo-2-trifluoromethylpyridine (400mg, 1.77mmol, 1.0eq), methyl 3-mercaptopropionate (235mg, 1.95mmol, 1.1eq) and N, N-diisopropyl Ethylamine (460 mg, 3.54 mmol, 2.0 eq) was dissolved in 1,4-dioxane (15 mL), replaced with argon three times, and Pd 2 (dba) 3 (160 mg, 0.18 mmol, 0.1 eq) and XantPhos (205 mg, 0.36 mmol, 0.2 eq) were added and the temperature was raised to 110 degrees. The reaction was completed for 1 hour.
  • Step 2 Dissolve methyl 3-((2- (trifluoromethyl) pyridin-3-yl) thio) propionate (C3-1, 300 mg, 1.13 mmol, 1.0 eq) in tetrahydrofuran (10 mL) and ethanol (0.5mL). Under nitrogen protection, sodium ethoxide (84 mg, 1.23 mmol, 1.1 eq) was added, and the reaction was performed at room temperature for 3 hours, but the reaction was not completed. Additional sodium ethoxide (83 mg, 1.23 mmol, 1.1 eq) was added, and the reaction was completed at room temperature for 3 hours.
  • Step 1 Combine 4-bromo-2-trifluoromethylpyridine (1.0g, 4.4mmol, 1.0eq), methyl 3-mercaptopropionate (760mg, 6.3mmol, 1.4eq) and N, N-diisopropyl Ethyl ethylamine (2.17g, 16.8mmol, 3.8eq) was dissolved in 1,4-dioxane (25mL), replaced with argon three times, under the protection of argon was added Pd 2 (dba) 3 (200mg, 0.22mmol , 0.05eq) and XantPhos (124mg, 0.22mmol, 0.05eq) were added to warm up to 110 degrees, the reaction was completed for 1 hour.
  • dba 200mg, 0.22mmol , 0.05eq
  • XantPhos 124mg, 0.22mmol, 0.05eq
  • Step 2 Dissolve methyl 3-((2- (trifluoromethyl) pyridin-4-yl) thio) propionate (C4-1, 230 mg, 0.85 mmol, 1.0 eq) in tetrahydrofuran (10 mL) and ethanol (0.5mL). Sodium ethoxide (294mg, 0.87mmol, 1.02eq) was added under nitrogen protection, and the reaction was carried out at room temperature for 1 hour, and the reaction was completed.
  • Step 1 Dissolve 2-fluoro-4-iodopyridine (2.0 g, 8.97 mmol) in THF (30 mL). After vacuum nitrogen replacement three times, the temperature was lowered to -65 ° C, LDA (2.0M in THF, 5.4mL, 10.80mmol) was added dropwise, and the reaction solution gradually turned brown. After the drop, keep it for 1.5 hours. A solution of 1,3,2-dioxazolethiophene-2,2-dioxide (1.45 g, 11.7 mmol) in THF (30 mL) was added dropwise. After the drop, the temperature was naturally raised to room temperature and stirred overnight.
  • LDA 2.0M in THF, 5.4mL, 10.80mmol
  • reaction solution was poured into a saturated aqueous NaHCO 3 solution (50 mL), extracted with ethyl acetate (3 ⁇ 30 mL), washed sequentially with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, concentrated and dried to obtain 2- ( 2-fluoro-4-iodopyridin-3-yl) ethane-1-ol (C5-1, 2.07g, yield: 86.2%)
  • Step 3 Dissolve methyl 3-((3-chloro-2- (methylamino) pyridin-4-yl) thio) propionate (C6-2, 114 mg, 0.44 mmol) in dioxane (6 mL), Under argon protection, EtONa (20% (w / w), 150 mg, 0.44 mmol) in ethanol was added dropwise, and the reaction was stirred at room temperature for about 2.5 h. LCMS monitored the reaction to completion. The reaction solution was directly used in the next reaction (yield: 100%). LCMS: m / z 174.8 [M + H-23] +
  • Step one To a solution of 2-fluoro-3-aldehyde-4-iodopyridine (1900 mg, 7.57 mmol) in isopropanol (30 mL) was added hydrazine hydrate (3032 mg, 60.56 mmol) and stirred at 60 ° C for 3 h. The reaction solution was concentrated under reduced pressure to remove part of the solvent, then poured into water, filtered, and the filter cake was washed with water to obtain 4-iodo-1H-pyrazolo [3,4-b] pyridine (C7-1, 1.8g, yield: 97%).
  • Step 2 To a solution of 4-iodo-1H-pyrazolo [3,4-b] pyridine (C7-1,400mg, 1.63mmol) in tetrahydrofuran (10mL), p-toluenesulfonic acid (28mg, 0.16mmol) and DHP (206 mg, 2.45 mmol). Stir at 60 ° C for 16h. The reaction solution was diluted with ethyl acetate (40 mL), washed with saturated brine (2 ⁇ 40 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure.
  • Step 3 Add 4-iodo-1- (tetrahydro-2H-pyran-2-yl) -1H-pyrazolo [3,4-B] pyridine (C7-2,390mg, 1.18mmol), 3-mercapto Methyl propionate (157 mg, 1.30 mmol) and N, N-diisopropylethylamine (306 mg, 2.37 mmol) were dissolved in 1,4-dioxane (10 mL) and replaced with argon three times. Under protection, Pd (OAc) 2 (27 mg, 0.12 mmol) and Xantphos (137 mg, 0.24 mmol) were added and the temperature was raised to 110 ° C. The reaction was completed for 2 h.
  • Step 4 In a three-necked flask with a condenser, add methyl 3-((1H-pyrazolo [3,4-b] pyridin-4-yl) thio) propanoate (C7-3,280mg, 0.87mmol ) Dissolve with 1,4-dioxane (7mL). Nitrogen protection, cooling to 0 degrees, adding sodium ethoxide ethanol solution (20% (w / w), 266mg, 0.78mmol).
  • Step 1 Under the protection of nitrogen, add 5-chloro-8-iodoimidazo [1,2-c] pyrimidine (B3,80mg, 0.285mmol), ((R) -N- ((S) -1,3-dihydrospiro [indene-2,4'-piperidin] -1-yl) -2-methylpropane-2-sulfinamide (A4, 105 mg, 0.342 mmol), DIEA (55 mg, 0.428 mmol) and CH 3 CN (5 mL), and then the reaction was stirred at 95 ° C. for 3 hours.
  • Step 2 2,3-dihydrofuran [2,3-b] pyridine-4-thiocarbamic acid sodium salt (107mg, 0.61mmol) was diluted with dioxane (15mL), and then (S) -N- ( S) -1 '-(8-iodoimidazole [1,2-C] pyrimidin-5-yl) -5,7-dihydrospiro [cyclopentane [B] pyridine-6,4'-piperidine ] -5-yl) -2-methylpropane-2-sulfinamide (1-1, 200 mg, 0.36 mmol), DIPEA (141 mg, 1.09 mmol), XantPhos (63 mg, 0.11 mmol) and Pd 2 (dba) 3 (50 mg, 0.06 mmol).
  • Step 1 Under the protection of nitrogen, add 5-chloro-8-iodo- [1,2,4] triazolo [4,3-c] pyrimidine (B2,80mg, 0.285mmol ), ((R) -N-((S) -1,3-dihydrospiro [indene-2,4'-piperidin] -1-yl) -2-methylpropane-2-sulfinamide ( A1, 105 mg, 0.342 mmol), DIEA (55 mg, 0.428 mmol) and CH 3 CN (5 mL), and then the reaction was stirred at 95 ° C. for 3 hours. After the reaction was completed, the obtained residue was filtered and concentrated under reduced pressure.
  • Step 2 Add 5R microwave reaction flask to (R) -N-((S) -1 '-(8-iodine- [1,2,4] triazolo [4,3- c] pyrimidin-5-yl) -1,3-dihydrospiro [indene-2-, 4'-piperidin] -1-yl) -2-methylpropane-2-sulfinamide (2-1, 100 mg, 0.18 mmol), sodium 2-amino-3-chloropyridine-4-thiolate (49 mg, 0.27 mmol), Pd 2 (dba) 3 (16 mg, 0.018 mmol), Xantphos (21 mg, 0.036 mmol), DIPEA ( 58mg, 0.45mmol) and 1,4-dioxane solution (10mL), the mixture was heated under microwave protection at 100 ° C under nitrogen protection and stirred for 3 hours.
  • Step three Add (R) -N-((R) -1 '-(8-((2-amino-3-chloropyridin-4-yl) thio)) to a 50mL single-necked flask sequentially under nitrogen -[1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -1,3-dihydrospiro [indene-2,4'-piperidin] -1-yl) -2 -Methylpropane-2-sulfinamide (2-2, 60 mg, 0.10 mmol) and methanol (0.6 mL), and a solution of 1,4-dioxane hydrochloric acid (0.06 mL, 4M) was added dropwise at room temperature.
  • intermediate A2 is used instead of intermediate A1, and a three-step reaction is performed to obtain compound 3.
  • Step 1 2-methylamino-3chloropyridine-4 sodium sulfide (C6, 86mg) was dissolved in 10mL dioxane solution, and (R) -N-((S) -1 '-(8-iodoimidazole was added [1,2-C] pyrimidin-5-yl) -5,7-dihydrospiro [cyclopenta [b] pyridine-6,4'-piperidin] -5-yl) -2-methyl Propane-2-sulfinamide (161 mg, 0.29 mmol), Pd 2 (dba) 3 (86 mg, 0.094 mmol), Xantphos (108 mg, 0.19 mmol) and DIPEA (400 mg, 3.12 mmol).
  • Step 2 (R) -N-((S) -1 '-(8-((3-chloro-2- (methylamino) pyridin-4-yl) thio) imidazo [1,2- c] pyrimidin-5-yl) -5,7-dihydrospiro [cyclopent [b] pyridine-6,4'-piperidin] -5-yl) -2-methylpropane-2-sulfinamide ( 135 mg, 0.23 mmol) was dissolved in methanol (20 mL), protected by argon, and HCl / dioxane (4M, 1.2 mL) was slowly added dropwise.
  • Step 1 Add B3 (1.37g, 4.9mmol), A4 (1.35g, 4.9mmol), DIPEA (4.86mL, 29.41mmol) to a 25mL single-necked flask containing 3mL of acetonitrile, and then stir at 80 ° C for 2 hours . After the reaction was completed, it was cooled to room temperature, and then Boc 2 O (1.6g, 7.35mmol, 1.5eq) was added, and the temperature was raised to 50 ° C for reaction until the reaction was completed.
  • Boc 2 O 1.6g, 7.35mmol, 1.5eq
  • Step 2 Add 6-1 (1.7g, 3.11mmol), 2-amino-3-chloropyridine-4-thiol sodium (596mg, 3.27mmol), and Pd2 to a 5mL microwave reaction bottle under nitrogen.
  • dba) 3 (285 mg, 0.311 mmol), Xantphos (360 mg, 0.622 mmol), DIPEA (804 mg, 6.22 mmol) and 1,4-dioxane solution (30 mL), the mixture was stirred under microwave protection at 100 ° C under nitrogen React for 3 hours. After the reaction was completed, it was cooled to room temperature, filtered and concentrated under reduced pressure.
  • Step 3 Nitrogen protection, to (S) -tert-butyl (1 '-(8-((2-amino-3-chloropyridin-4-yl) thio) imidazole [1,2-c] Pyrimidin-5-yl) -5,7-dihydrospiro [cyclopenta [b] pyridine-6,4'-piperidinyl] -5-yl) -carbonate (6-2, 1.2g, 2.07 TFA (5 mL) was slowly added to a solution of mmol) in dichloromethane (5 mL), and the reaction was stirred at room temperature for 1 hour. TLC and LCMS showed that the reaction was complete.
  • Step 1 Dissolve (S) -5,7-dihydrospiro [cyclopenta [b] pyridine-6,4'-piperidine] -7-amine hydrochloride (A6,200mg, 1.28mmol) In acetonitrile (20mL), add DIPEA (1.6mL, 9.68mmol) and 5-chloro-8-iodo- [1,2,4] -triazolo [4,3-c] pyrimidine (B2, 180mg, 0.64 mmol), heated to 90 ° C and refluxed under nitrogen for 3 hours. The reaction was completed.
  • reaction solution was poured into a saturated aqueous sodium hydrogen carbonate solution, and extracted with dichloromethane (80 mL) twice.
  • dichloromethane 80 mL twice.
  • the organic phases were combined and washed with saturated saline.
  • the organic phases were separated and dried over anhydrous sodium sulfate, filtered and concentrated.
  • Step 2 Put (S) -1 '-(8-iodo [1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -5,7-dihydrospiro [cyclopentane Benzo [b] pyridine-6,4'-piperidin] -7-amine (21-1, 55 mg, 0.12 mmol) was dissolved in 1,4-dioxane (3 mL), and DIPEA (35 mg, 0.27) was added mmol) and sodium 2,3-dichloropyridine-4-thiolate (C2, 100 mg, crude).
  • XantPhos (30 mg, 0.05 mmol) and Pd 2 (dba) 3 (17 mg, 0.02 mmol) were added under nitrogen protection. After nitrogen substitution three times, the reaction was performed at 100 ° C for 3 hours, and the reaction was completed.
  • intermediate B2 is used instead of intermediate B3, and intermediate A6 is used instead of intermediate A4 to perform a three-step reaction to obtain compound (S) -1 '-(8-((2-amino-3-chloropyridine -4-yl) thio)-[1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -5-, 7-dihydrospiro [cyclopentadieno [b] Pyridine-6,4'-piperidine] -7-amine (11, formate, white solid).
  • intermediate A7 is used instead of intermediate A4, and a two-step reaction is performed to obtain compound (S)-(1 '-(8-((2-amino-3-chloropyridin-4-yl) thio ) Imidazo [1,2-c] pyrimidin-5-yl) -5-fluoro-1,3-dihydrospiro [inden-2,4'-piperidin] -3-yl) carboxylic acid tert-butyl ester (13 -2).
  • Step 3 Add the compound (S)-(1 '-(8-((2-amino-3-chloropyridin-4-yl) thio) imidazo [1,2-c] Pyrimidin-5-yl) -5-fluoro-1,3-dihydrospiro [indene-2,4'-piperidin] -3-yl) carboxylic acid tert-butyl ester (13-2, 60 mg, 0.101 mmol), di Chloromethane (1 mL) and trifluoroacetic acid (0.2 mL). The reaction solution was stirred at 20 ° C for 1 hour. The reaction solution was concentrated under reduced pressure.
  • the resulting residue was purified by HPLC preparation to obtain a white solid compound (S) -1 '-(8-((2-amino-3-chloropyridin-4-yl) thio) imidazo [1,2-c] pyrimidine -5-yl) -6-fluoro-1,3-dihydrospiro [indene-2,4'-piperidine] -1-amine (13,32 mg, formate, yield: 58.2%).
  • Step 1 Dissolve (S) -5,7-dihydrospiro [cyclopenta [c] pyridine-6,4'-piperidine] -5-amine hydrochloride (A8,200mg, 0.83mmol) To acetonitrile (20mL), add DIPEA (1.07g, 8.3mmol) and 5-chloro-8-iodoimidazo [1,2-c] pyrimidine (B3,208mg, 0.75mmol), and heat to 90 ° C under nitrogen protection The reaction was refluxed for 5 hours, and the reaction was completed. After cooling to room temperature, the reaction solution was poured into a saturated aqueous solution of sodium bicarbonate, and extracted with dichloromethane (50 mL) twice.
  • Step 9 Put 1 '-(8-iodoimidazo [1,2-c] pyrimidin-5-yl) -5,7-dihydrospiro [cyclopenta [c] pyridine-6,4'- Piperidine] -4-amine (14-1,50mg, 0.11mmol) was dissolved in 1,4-dioxane (10mL), added DIPEA (36mg, 0.28mmol) and 2-amino-3-chloropyridine- 4-Sodium thiolate (C1, 31 mg, 0.17 mmol). XantPhos (13 mg, 0.02 mmol) and Pd 2 (dba) 3 (10 mg, 0.01 mmol) were added under nitrogen protection.
  • intermediate B2 is used instead of intermediate B3, and a two-step reaction is performed to obtain compound (S) -1 '-(8-((2-amino-3-chloropyridin-4-yl) thio) -[1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -5-, 7-dihydrospiro [cyclopenta [c] pyridine-6,4'-piper Pyridine] -5-amine (15).
  • Step 1 Dissolve (S) -1-amino-1,3-dihydrospiro [indene-2,4'-piperidine] -4-ol hydrochloride (A9,200mg, 0.79mmol) in acetonitrile (40mL ), Add DIPEA (1.07g, 8.3mmol) and 5-chloro-8-iodo- [1,2,4] triazolo [4,3-c] pyrimidine (B2,208mg, 0.75mmol), nitrogen protection The reaction was completed at room temperature for 18 hours. After cooling to room temperature, the reaction solution was poured into a saturated aqueous solution of sodium bicarbonate, and extracted with dichloromethane (100 mL) twice.
  • Step 2 (S) -1-amino-1 '-(8-iodo- [1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -1,3-dihydro Spiro [indene-2,4 '-piperidin] -4-ol (16-1,200mg, 0.43mmol) was dissolved in 1,4-dioxane (30mL) and DIPEA (111mg, 0.86mmol) was added With sodium 2-amino-3-chloropyridine-4-thiolate (C1, 118 mg, 0.65 mmol).
  • Step 1 (S) -1-(((R) -tert-butylsulfinyl) amino) -4-cyano-1,3-dihydrospiro [indene-2,4'-piperidine] ( A10, 490mg, 1.48mmol) was dissolved in acetonitrile (20mL), added DIPEA (1.91g, 14.8mmol) and 5-chloro-8-iodo- [1,2,4] triazolo [4,3-c] Pyrimidine (B2, 415 mg, 1.48 mmol) was reacted at 85 ° C for 3 hours under nitrogen protection, and the reaction was completed.
  • Step 2 (R) -N-((S) -1 '-(8-iodine- [1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -4-cyano Yl-1,3-dihydrospiro [indene-2,4'-piperidin] -1-yl) -2-tert-butylsulfonamide (17-1, 360 mg, 0.63 mmol) dissolved in 1,4-bis To oxane (10 mL), DIPEA (202 mg, 1.56 mmol) and sodium 2-amino-3-chloropyridine-4-thiolate (C1, 171 mg, 0.94 mmol) were added.
  • DIPEA 202 mg, 1.56 mmol
  • sodium 2-amino-3-chloropyridine-4-thiolate C1, 171 mg, 0.94 mmol
  • Step 3 Add (R) -N-((S) -1 '-(8-((2-amino-3-chloropyridin-4-yl) thio)) to a 50mL single-necked flask sequentially under nitrogen -[1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -4-cyano-1,3-dihydrospiro [indene-2,4'-piperidine] -1 -Yl) -2-tert-butylsulfonamide (17-2, 130 mg, 0.21 mmol) and methanol (6 mL), 1,4-dioxane hydrochloride solution (2 mL, 4M) was added dropwise at room temperature, and the mixture The reaction was stirred at room temperature for 1 hour.
  • Step 1 (S) -5,7-dihydrospiro [cyclopenta [b] pyrazine-6,4'-piperidine] -5-amine hydrochloride (A11, 1.50g, 4.95mmol ) Dissolved in 150mL acetonitrile, added DIPEA (5.68g, 44.0mmol) and 5-chloro-8-iodoimidazo [1,2-c] pyrimidine (B3, 1.28g, 4.59mmol), heated to 95 under nitrogen protection The reaction was refluxed at °C for 3 hours, and the reaction was completed.
  • Step 2 In a 350mL glass sealed tube, place (S) -1 '-(8-iodoimidazo [1,2-c] pyrimidin-5-yl) -5,7-dihydrospiro [cyclopentadiene [B] pyrazine-6,4'-piperidine] -5-amine (18-1, 2.05g, 4.59mmol) was dissolved in 120mL 1,4-dioxane, and DIPEA (1.78g, 13.8) was added in sequence mmol), sodium 2-amino-3-chloropyridine-4-thiolate (C1, 1.26 g, 6.9 mmol), XantPhos (797 mg, 1.38 mmol) and Pd 2 (dba) 3 (630 mg, 0.69 mmol), argon Bubbling for 30 seconds, heating to 100 ° C under argon atmosphere, holding for 3 hours, the reaction was complete.
  • Step 7 (S) -1-(((R) -tert-butylsulfinyl) amino) -4-methoxy-1,3-dihydrospiro [indene-2,4'-piperidine] (A12, 520 mg, 1.55 mmol) was dissolved in acetonitrile (20 mL), and DIPEA (2.0 g, 15.5 mmol) and 5-chloro-8-iodo- [1,2,4] triazolo [4,3-c were added ] Pyrimidine (B2, 435mg, 1.55mmol), under nitrogen protection at 85 °C for 3 hours, the reaction was completed.
  • Step 8 (R) -N-((S) -1 '-(8-iodine- [1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -4-methyl Oxy-1,3-dihydrospiro [indene-2,4'-piperidin] -1-yl) -2-tert-butylsulfonamide (19-1, 420 mg, 0.72 mmol) was dissolved in 1,4- To dioxane (10 mL), DIPEA (234 mg, 1.81 mmol) and sodium 2-amino-3-chloropyridine-4-thiolate (C1, 198 mg, 1.09 mmol) were added.
  • reaction solution was concentrated and purified by silica gel chromatography to obtain a white solid compound (R) -N-((S) -1 '-(8-((2-amino-3-chloropyridin-4-yl) thio)-[ 1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -4-methoxy-1,3-dihydrospiro [indene-2,4'-piperidine] -1- Yl) -2-tert-butylsulfonamide (19-2, 310 mg, yield: 69.9%).
  • Step 9 Add (R) -N-((S) -1 '-(8-((2-amino-3-chloropyridin-4-yl) thio) to a 50mL single-necked flask in sequence under nitrogen -[1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -4-methoxy-1,3-dihydrospiro [indene-2,4'-piperidine]- 1-yl) -2-tert-butylsulfonamide (19-2, 310 mg, 0.51 mmol) and methanol (6 mL), and a solution of 1,4-dioxane hydrochloride (2 mL, 4 M) was added dropwise at room temperature.
  • Step 1 (S) -5,7-dihydrospiro [cyclopenta [b] pyrazine-6,4'-piperidine] -5-amine hydrochloride (A11, 89mg, 0.29mmol) Dissolve in 20mL of acetonitrile, add DIPEA (212mg, 1.64mmol) and 5-chloro-8-iodo- [1,2,4] triazolo [4,3-c] pyrimidine (B2, 77mg, 0.275mmol), Under nitrogen, the reaction was heated to 95 ° C and refluxed for 3 hours. The reaction was completed.
  • Step 2 In a 5mL microwave tube, place (S) -1 '-(8-iodine- [1,2,4] triazolo [4,3-c] pyrimidin-5-yl) -5,7- Dihydrospiro [cyclopenta [b] pyrazine 6,4'-piperidine] -5-amine (20-1,65mg, 0.145mmol) was dissolved in 5mL 1,4-dioxane and added sequentially DIPEA (56 mg, 0.434 mmol), sodium 2-amino-3-chloropyridine-4-thiolate (C1, 40 mg, 0.219 mmol), XantPhos (34 mg, 0.059 mmol) and Pd 2 (dba) 3 (27 mg, 0.029 mmol ), Bubbling argon for 30 seconds, heating to 100 ° C under argon atmosphere, holding for 3 hours, the reaction is complete.
  • DIPEA 56 mg, 0.434 mmol
  • Step 1 (S) -2-chloro-4,6-dihydrospiro [cyclopenta [d] thiazole-5,4'-piperidine] -4-amine hydrochloride (A13,155mg, 0.35mmol) was dissolved in acetonitrile (10mL), added DIPEA (450mg, 3.5mmol) and 5-chloro-8-iodoimidazo [1,2-c] pyrimidine (B3, 108mg, 0.39mmol), heated under the protection of nitrogen The reaction was refluxed at 100 ° C for 5 hours, and the reaction was completed.
  • Step 2 (S) -2-chloro-1 '-(8-iodoimidazo [1,2-c] pyrimidin-5-yl) -4,6-dihydrospiro [cyclopenta [d ] Thiazole-5,4'-piperidine] -4-amine (22-1,50 mg, 0.1 mmol) was dissolved in 1,4-dioxane (5 mL), and DIPEA (30 mg, 0.25 mmol) was added Sodium 2-amino-3-chloropyridine-4-thiolate (C1, 30 mg, 0.15 mmol). Under nitrogen protection, XantPhos (13 mg, 0.02 mmol) and Pd 2 (dba) 3 (20 mg, 0.02 mmol) were added.
  • Step 1 To the reaction containing 1- (tetrahydro-2H-pyran-2-yl) -1H-pyrazolo [3,4-b] pyridine-4-thiol sodium (C7,224mg, 0.8mmol) Add dioxane (20mL) to the bottle, then add (S) -N-((S) -1 '-(8-iodoimidazole [1,2-C] pyrimidin-5-yl) -5,7 -Dihydrospiro [cyclopentadiene [B] pyridine-6,4'-piperidin] -5-yl) -2-methylpropane-2-sulfinamide (1-1, 287 mg, 0.52 mmol) , DIPEA (304 mg, 2.35 mmol), XantPhos (91 mg, 0.16 mmol) and Pd 2 (dba) 3 (72 mg, 0.08 mmol).
  • Step 6 Nitrogen protection, to (R) -2-methyl-N-((5S) -1 '-(8-((1- (tetrahydro-2H-pyran-2-yl)- 1H-pyrazolo [3,4-b] pyridin-4-yl) thio) imidazo [1,2-c] pyrimidin-5-yl) -5,7-dihydrospiro [cyclopentadiene
  • b) Pyridine-6,4'-piperidin] -5-yl) propane-2-sulfinamide (23-1, 176 mg, 0.26 mmol) in dichloromethane (10 mL) was slowly added HCl / dioxane (4M , 1.3mL, 5.20mmol), the reaction was stirred at room temperature for 2h, TLC (dichloromethane / methanol 8/1) and LCMS showed the reaction was complete.
  • Control compound synthesis When applying for the present invention, the closest control to the present invention is Example 25 of WO2018136265. Refer to its synthetic route and operation steps to obtain the control compound aa.
  • the biological functions of the compounds disclosed in the present invention have been proved in tests of enzyme activity and cellular level.
  • the compounds disclosed in the present invention can achieve strong inhibitory activity (IC50 up to 1 nM).
  • the compounds disclosed in the present invention also exhibit very good activity to inhibit the proliferation of cancer cells, and the proliferation inhibitory activity on MV4-11 cell lines can reach 1 nM.
  • SHP099 6- (4-amino-4-methylpiperidin-1-yl) -3- (2,3-dichlorophenyl) pyrazine-2-amine
  • the invention patent The compounds showed superior activity both at the enzymatic level and at the cellular level.
  • Test Case 1 SHP2 enzyme activity test method
  • Compound powder is dissolved in DMSO to make mother liquor.
  • the compound storage solution was subjected to 3-fold gradient dilution with DMSO, and 10 different test concentrations of the same compound were set. Take 1 ⁇ L of compound at each concentration point into the well of the detection plate (Corning, Costar 3915), and set 2 replicates for each concentration point.
  • SHP2 E72A catalyzes its hydrolysis to produce 6,8-difluoro-4-methyl-7-hydroxyaromatic DiFMU is used to detect the fluorescence value at 455nm with PE Enspire multi-function reader with 358nm as the excitation wavelength to determine the enzyme activity of SHP2.
  • the composition is 60mmol / L Hepes, PH7.2, 75mmol / L NaCl, 75mmol / L KCl, 1mmol / L EDTA, 5mmol / L DTT.
  • the screening system consists of: SHP2 buffer, enzyme SHP2 E76A protein, substrate DiFMUP and pending Test compound.
  • Test Example 2 MV4-11 cell proliferation inhibition experiment
  • the luminescence method cell viability detection kit quantitatively measures intracellular ATP to detect the number of viable cells in the culture.
  • MV4-11 cells were seeded in 96-well plates, and cells were seeded in 96-well plates at a cell density of 2500 cells per well, with a volume of 100 ⁇ L per well. Place in a 37 ° C 5% carbon dioxide incubator overnight.
  • the second step compound treats the cells.
  • the compound to be tested is diluted 3 times, and a total of 8 concentration gradients are set; each well is added with a certain volume of DMSO or the compound to be tested, and each concentration is set to 2 replicates, and the final concentration of DMSO is controlled at 0.5%. Place in a 37 ° C 5% carbon dioxide incubator for 72 hours.
  • Luminescent Cell Viability Assay kit detects the cell viability of the control and treatment groups. Add 50ul CellTiter-Glo to each well, mix well, and incubate at room temperature for 10min. Use EnSpire (Perkin Elmer) to read the signal. The percentage of inhibition (%) is calculated by the following formula:
  • Test Example 3 Compound pharmacokinetic experiment
  • mice used in the study were administered by intragastric administration of 5-100 mg / Kg in each group. Fasting from 10 hours before administration to 4 hours after administration. Blood was collected at different time points after administration, and the plasma content of the compound (LC-MS / MS) was determined. Plasma concentration-time relationship is analyzed with professional software (winnonlin) to calculate the pharmacokinetic parameters of the compound. According to Table 3, the compounds of the present invention have excellent pharmacokinetic properties.

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Abstract

提供了一种式I化合物，或其药学上可接受的盐，或其对映异构体、非对映异构体、互变异构体、溶剂化物、同位素取代物、多晶型物、前药或代谢产物；并且还提供了制备式I化合物的方法，其中式I化合物对SHP2具有较高的抑制活性，因此可以用于预防或治疗与SHP2相关的疾病。

Description

一种螺芳环化合物及其应用 技术领域
本发明涉及螺芳环化合物领域，具体地说，涉及一种可用于SHP2抑制剂的螺芳环化合物、其药学上可接受的盐、或其对映异构体、非对映异构体、互变异构体、溶剂化物、同位素取代物、前药或代谢产物。另外，本发明还涉及该类化合物的制备方法、含有该类化合物的药物组合物以及该类化合物用于制备治疗与SHP2活性异常相关疾病或病症的药物的用途。
背景技术
蛋白酪氨酸磷酸酶SHP2，在细胞信号传导过程中占据及其重要的位置，是开发治疗糖尿病、自身免疫疾病和癌症等重大疾病的靶点。SHP2是在多种疾病中突变或者高表达，如努南综合征(Noonan Syndrome)、豹综合征(Leopard Syndrome)、青少年髓单核细胞白血病、成神经细胞瘤、黑色素瘤、急性髓性白血病、乳腺癌、食道癌、肺癌、结肠癌、头癌、成神经细胞瘤、头颈的鳞状细胞癌、胃癌、间变性大细胞淋巴瘤和成胶质细胞瘤等等。分子生物学研究表明，SHP2参与多个肿瘤细胞信号传导通路，如MAPK、JAK/STAT和PI3K/Akt等。同时，SHP2也负责PD1-PDL1免疫抑制通路的信号传导。因此，抑制SHP2的活性能够在逆转肿瘤微环境中的免疫抑制。
SHP2由两个N-末端Src同源2结构域(N-SH2和C-SH2)和一个蛋白酪氨酸磷酸酶催化结构域(PTP)构成。在自抑制状态下，N-SH2与PTP结合形成一个环状结构，从而阻碍PTP与底物的结合，使得酶催化活性被抑制；当上游受体蛋白的酪氨酸被磷酸化后，N-SH2与之相结合，PTP催化域得到释放从而发挥出磷酸酶活性。
目前针对SHP2抑制剂的开发，主要以非催化区的变构抑制剂为主，比如WO2015107493A1，WO2016203404A1，WO2016203406A1，WO2017216706A1，WO2017211303A1，CN201710062495，WO2018136265A1WO2018057884等中公开的一些化合物。今年的研究表明SHP2作为一个新颖的可成药靶点，引起了越来越多的关注。因此，本领域急需开发结构新颖、生物活性好，成药性高的SHP2抑制剂。
发明内容
本发明的目的在于提供一种式I化合物或其药学上可接受的盐，含有该化合物或其药学上可接受的盐的药物组合物，以及该化合物或其药物组合物在预防和治疗与SHP2异常相关的疾病或病症中的应用。
本发明的第一方面，提供了一种式I化合物，或其药学上可接受的盐，或其对映异构体、非对映异构体、互变异构体、溶剂化物、同位素取代物、多晶型物、前药或代谢产物，
Figure PCTCN2019115754-appb-000001
其中，
X 1和X 2各自独立地选自键、O、CR aR b或NR c；
X 3选自键、CR aR b、NR c、S或O；
X 4选自N或者CR c；且R a、R b和R c各自独立地选自H、卤素、取代或未取代的C 1-6烷基、取代或未取代的C 1-6烷氧基；
R 1、R 2、R 3、R 4和R 7各自独立地选自H、-OH、卤素、取代或未取代的氨基、取代或未取代的C 1-6烷基、取代或未取代的C 1-6烷氧基；且不能同时为-OH或-NH 2；
环A选自取代或未取代的C 4-8环烃基、取代或未取代的4-8元杂环基、取代或未取代的C 5-10芳基、取代或未取代的5-10元杂芳基，所述杂环基或杂芳基包含1-3个选自下组的杂原子：N、O、S或P；
环C选自取代或未取代的C 4-8环烃基、取代或未取代的5-6元单环杂环基、取代或未取代的8-10元双环杂环基、取代或未取代的C 5-10单环或双环芳基、取代或未取代的5-6元单环杂芳基、取代或未取代的8-10元双环杂芳基，所述杂环基或杂芳基包含1-4个选自下组的杂原子：N、O、S或P；
R 5和R 6各自独立地选自H、-OH、卤素、氰基、取代或未取代的氨基、取代或未取代的C 1-6烷基、取代或未取代的C 1-6烷氧基；
n为0至3中的任一整数；并且
所述取代是指基团上的一个或多个氢原子被选自下组的取代基取代：卤素、-OH、-NO 2、-NH 2、-NH(未取代或卤代的C 1-6烷基)、-N(未取代或卤代的C 1-6烷基) 2、-CN、未取代或卤代的C 1-8烷基、未取代或卤代的C 1-8烷氧基、未取代或卤代的C 1-8烷氧基-C 1-8烷基、未取代或卤代的C 3-8环烷基-C 1-8烷基、未取代或卤代的C 1-6烷基羰基、未取代或卤代的C 1-6烷氧基羰基、异羟肟酸基、未取代或卤代的C 1-6烷基巯基、-S(O) 2N(未取代或卤代的C 1-6烷基) 2、-S(O) 2未取代或卤代的C 1-6烷基、-N(未取代或卤代的C 1-6烷基)S(O) 2N(未取代或卤代的C 1-6烷基) 2、-S(O)N(未取代或卤代的C 1-6烷基) 2、-S(O)(未取代或卤代的C 1-6烷基)、-N(未取代或卤代的C 1-6烷基)S(O)N(未取代或卤代的C 1-6烷基) 2、-N(未取代或卤代的C 1-6烷基)S(O)(未取代或卤代的C 1-6烷基)、未取代或卤代的C 5- 10芳基、未取代或卤代的5-10元杂芳基、未取代或卤代的C 4-8环烃基、未取代或卤代的4-8元杂环基，所述杂环基和杂芳基包含1-4个选自下组的杂原子：N、O或S。
作为一种优选的实施方式，X 1和X 2中的一个为CH 2，并且另一个为键。
作为一种优选的实施方式，X 3为S。
作为一种优选的实施方式，X 4选自N或者CH。
作为一种优选的实施方式，R 1、R 2、R 3、R 4和R 7各自独立地选自H、-OH、-F、-Cl、-Br、-NH 2、-NHC 1-3烷基、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基或异丙氧基；被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷基；或被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷氧基。
作为一种优选的实施方式，R 5和R 6各自独立地选自H、-OH、-F、-Cl、-Br、-CN、-NH 2、-NHC 1-3烷基、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基或异丙氧基；被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷基；或被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷氧基。
作为一种优选的实施方式，所述取代基选自-F、-Cl、-Br、-OH、-NO 2、-NH 2、-NH(C 1-6烷基)、-N(C 1-6烷基) 2、-CN、C 1-6烷基、C 1-4烷氧基、C 1-4烷氧基-C 1-6烷基、C 3-8环烷基-C 1-8烷基、C 1-6烷基羰基、C 1-6烷氧基羰基、C 1-6烷基巯基、-S(O) 2N(C 1-6烷基) 2、-S(O) 2C 1-6烷基、-N(C 1-6烷基)S(O) 2N(C 1-6烷基) 2、-S(O)N(C 1-6烷基) 2、-S(O)(C 1-6烷基)、-N(C 1-6烷基)S(O)N(C 1-6烷基) 2、-N(C 1-6烷基)S(O)(C 1-6烷基)、取代或未取代的C 5-10芳基、取代或未取代的5-10元杂芳基、取代或未取代的C 4-8环烃基、取代或未取代的4-8元杂环基，所述杂环基和杂芳基包含1-4个选自下组的杂原子：N、O或S。
作为一种优选的实施方式，所述取代基选自-F、-Cl、-Br、-OH、-NO 2、-NH 2、-NH(C 1-3烷基)、-N(C 1-3烷基) 2、-CN、C 1-3烷基、C 1-3烷氧基、C 1-3烷基羰基、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、环辛基、吡咯烷基、吗啉基、哌嗪基、高哌嗪基、哌啶基、硫代吗啉基、苯基、萘基、蒽基、菲基、芴基、噻吩基、咪唑基、吡唑基、噻唑基、噁唑基、噁二唑基、异噁唑基、吡啶基、嘧啶基、吡嗪基、哒嗪基、苯并咪唑基、苯并吡唑基、吲哚基、呋喃基、吡咯基、三唑基、四唑基、三嗪基、吲嗪基、异吲哚基、吲唑基、异吲唑基、嘌呤基、喹啉基、异喹啉基。
作为一种优选的实施方式，所述取代基选自-F、-Cl、-Br、-OH、-NO 2、-NH 2、-NH(C 1-3烷基)、-N(C 1-3烷基) 2、-CN、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基、异丙氧基或苯基。
作为一种优选的实施方式，所述环C选自下组中的任一个：
Figure PCTCN2019115754-appb-000002
其中，
X 5、X 6、X 7、X 8和X 9各自独立地选自N或CR d；且至多同时3个为N；
X 10、X 11、X 12、X 13、X 14、X 15、X 16和X 17各自独立地选自N或CR d；且至多同时5个为N；
X 18、X 19、X 20和X 21各自独立地选自N或CR d，且至多同时3个为N；
R 6和R 8各自独立地选自H、-NH 2、-CN、-OH、-NO 2、卤素、未取代或卤代的C 1-6烷基、未取代或卤代的C 1-6烷氧基；并且
所述R d选自H、卤素、未取代或卤代的C 1-6烷基、未取代或卤代的C 1-6烷氧基。
作为一种优选的实施方式，所述环C选自以下中的任一个：
Figure PCTCN2019115754-appb-000003
其中，
X 5、X 6、X 7、X 8和X 9中的0、1或2个为N，其余为CR d；
X 18、X 19、X 20和X 21中的0、1或2个为N，其余为CR d；
R 6选自H、-NH 2、-CN、-OH、-NO 2、-F、-Cl、-Br、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基、异丙氧基、氟代或溴代的C 1-3烷基、氟代或溴代的C 1- 3烷氧基；并且
所述R d选自H、-F、-Cl、-Br、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基、异丙氧基、氟代或溴代的C 1-3烷基、氟代或溴代的C 1-3烷氧基。
作为一种优选的实施方式，所述环C选自下组中的任一个：
Figure PCTCN2019115754-appb-000004
作为一种优选的实施方式，所述环A选自取代或未取代的C 4-6环烃基、取代或未取代的4-6元杂环基、取代或未取代的C 5-6芳基、取代或未取代的5-6元杂芳基，所述杂环基或杂芳基包含1-3个N原子。
作为一种优选的实施方式，所述环A选自下组中的任一个：
Figure PCTCN2019115754-appb-000005
作为一种优选的实施方式，所述环A选自下组中的任一个：
Figure PCTCN2019115754-appb-000006
作为进一步的优选实施方式，所述化合物具有选自以下的结构：
Figure PCTCN2019115754-appb-000007
作为一种优选的实施方式，本发明所述化合物的同位素取代物，其同位素取代选自下组：氢、碳、氮、氧、氟、磷、氯和碘；优选为 2H、 3H、 11C、 13C、 14C、 15N、 17O、 18O、 18F、 31P、 32P、 35S、 36Cl或 125I。
本发明的第二方面，提供了一种制备本发明的式I化合物的方法，所述方法包括如下步骤：
(i)使式Ib与式Ic发生亲核取代反应得到式Id；
(ii)使式Id与式Ie发生取代反应得到式If；并且
(iii)将式If用酸脱除保护以得到式I化合物：
Figure PCTCN2019115754-appb-000008
本发明的第三方面，提供了一种本发明的式I化合物在以下方法中的用途：
(a)制备预防或治疗与SHP2活性异常相关的疾病或病症的药物；
(b)制备预防或治疗SHP2-介导的疾病或病症的药物
(c)制备抑制SHP2活性的抑制剂药物；
(d)体外非治疗性的抑制SHP2活性；
(e)体外非治疗性的抑制肿瘤细胞增殖；或
(f)治疗与SHP2异常相关的疾病或病症。
在一个优选的实施方式中，所述疾病为癌症；优选为努南综合症、豹综合症、青少年髓单核细胞白血病、成神经细胞瘤、黑色素瘤、急性髓性白血病、乳腺癌、食道癌、肺癌、结肠癌、头癌、成神经细胞瘤、头颈的鳞状细胞癌、胃癌、间变性大细胞淋巴瘤或成胶质细胞瘤。
本发明的第四方面，提供了一种药物组合物，所述药物组合物包含：
(i)有效量的式I化合物，或其药学上可接受的盐、对映异构体、非对映异构体、互变异构体、溶剂化物、同位素取代物、多晶型物、前药或代谢产物；和
(ii)药学上可接受的载体。
本发明的第五方面，提供了一种抑制SHP2活性的方法，所述方法包括如下步骤：对有此需要的受试者施用有效量的本发明的式I化合物或其药学上可接受的盐，或对有此需要的受试者施用有效量的本发明的药物组合物。
应理解，在本发明范围内，本发明的上述各技术特征和在下文(如实施例)中具体描述的各技术特征之间都可以互相组合，从而构成新的或优选的技术方案。限于篇幅，在此不再一一累述。
具体实施方式
本发明人经过长期而深入的研究，制备了一类具有式I所示的新颖的变构抑制剂化合物，其能够通过与SHP2非催化区域的结合并“锁”住SHP2活性很弱的自抑制状态，从而达到抑制其活性的目的。本发明的化合物表现出很好的生物活性及可成药性，具有很好的药物开发前景，其在极低浓度(可低至≤100nM/L)下，即对SHP2产生抑制作用，抑制活性相当优异，因而可以用于治疗与SHP2相关的疾病或病症，如肿瘤。基于上述发现，发明人完成了本发明。
术语
除非另有定义，否则本文所有科技术语具有的涵义与权利要求主题所属领域技术人员通常理解的涵义相同。除非另有说明，本文全文引用的所有专利、专利申请、公开材料通过引用方式整体并入本文。
应理解，上述简述和下文的详述为示例性且仅用于解释，而不对本发明主题作任何限制。在本申请中，除非另有具体说明，否则使用单数时也包括复数。必须注意，除非文中另有清楚的说明，否则在本说明书和权利要求书中所用的单数形式包括所指事物的复数形式。还应注意，除非另有说明，否则所用“或”、“或者”表示“和/或”。此外，所用术语“包括”以及其它形式，例如“包含”、“含”和“含有”并非限制性，其可以是开放式、半封闭式和封闭式的。换言之，所述术语也包括“基本上由…构成”、或“由…构成”之义。
可在参考文献(包括Carey and Sundberg"ADVANCED ORGANIC CHEMISTRY 4TH  ED."Vols.A(2000)and B(2001),Plenum Press,New York)中找到对标准化学术语的定义。除非另有说明，否则采用本领域技术范围内的常规方法，如质谱、NMR、IR和UV/VIS光谱法和药理学方法。除非提出具体定义，否则本文在分析化学、有机合成化学以及药物和药物化学的有关描述中采用的术语是本领域已知的。可在化学合成、化学分析、药物制备、制剂和递送，以及对患者的治疗中使用标准技术。例如，可利用厂商对试剂盒的使用说明，或者按照本领域公知的方式或本发明的说明来实施反应和进行纯化。通常可根据本说明书中引用和讨论的多个概要性和较具体的文献中的描述，按照本领域熟知的常规方法实施上述技术和方法。在本说明书中，可由本领域技术人员选择基团及其取代基以提供稳定的结构部分和化合物。
当通过从左向右书写的常规化学式描述取代基时，该取代基也同样包括从右向左书写结构式时所得到的在化学上等同的取代基。举例而言，-CH 2O-等同于-OCH 2-。
本文所用的章节标题仅用于组织文章的目的，而不应被解释为对所述主题的限制。本申请中引用的所有文献或文献部分包括但不限于专利、专利申请、文章、书籍、操作手册和论文，均通过引用方式整体并入本文。
在本文中定义的某些化学基团前面通过简化符号来表示该基团中存在的碳原子总数。例如，C1-C6烷基是指具有总共1至6个碳原子的如下文所定义的烷基。简化符号中的碳原子总数不包括可能存在于所述基团的取代基中的碳。
除前述以外，当用于本申请的说明书及权利要求书中时，除非另外特别指明，否则以下术语具有如下所示的含义。
在本申请中，术语“卤素”是指氟、氯、溴或碘。
“羟基”是指-OH基团。
“羟基烷基”是指被羟基(-OH)取代的如下文所定义的烷基。
“羰基”是指-C(＝O)-基团。
“硝基”是指-NO 2。
“氰基”是指-CN。
“氨基”是指-NH 2。
“取代的氨基”是指被一个或两个如下文所定义的烷基、烷基羰基、芳烷基、杂芳烷基取代的氨基，例如，单烷基氨基、二烷基氨基、烷基酰氨基、芳烷基氨基、杂芳烷基氨基。
“羧基”是指-COOH。
在本申请中，作为基团或是其它基团的一部分(例如用在卤素(例如氟、氯、溴或碘)取代的烷基等基团中)，术语“烷基”是指完全饱和的直链或支链的烃链基，仅由碳原子和氢原子组成、具有例如1至12个(优选为1至8个，更优选为1至6个)碳原子，且通过单键与分子的其余部分连接，例如包括但不限于甲基、乙基、正丙基、异丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、2-甲基丁基、2,2-二甲基丙基、正己基、庚基、2-甲基己基、3-甲基己基、辛基、壬基和癸基等。就本发明而言，术语“烷基”指含有1至8个碳原子的烷基。
在本申请中，作为基团或是其它基团的一部分，术语“烯基”意指仅由碳原子和氢原子组成、含有至少一个双键、具有例如2至20个(优选为2至10个，更优选为2至6个)碳原子且通过单键与分子的其余部分连接的直链或支链的烃链基团，例如但不限于乙烯基、丙烯基、烯丙基、丁-1-烯基、丁-2-烯基、戊-1-烯基、戊-1,4-二烯基等。
在本申请中，作为基团或是其它基团的一部分，术语“环烃基”意指仅由碳原子和氢原子组成的稳定的非芳香族单环或多环烃基(例如烷基、烯基或炔基)，其可包括稠合环体系、桥环体系或螺环体系，具有3至15个碳原子，优选具有3至10个碳原子，更优选具有3至8个碳原子，例如3、4、5、6、7或8个碳原子，且其为饱和或不饱和并可经由任何适宜的碳原子通过单键与分子的其余部分连接。除非本说明书中另外特别指明，环烃基中的碳原子可以任选地被氧化。环烃基的实例包括但不限于环丙基、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、环辛基、1H-茚基、2,3-二氢化茚基、1,2,3,4-四氢-萘基、5,6,7,8-四氢-萘基、8,9-二氢-7H-苯并环庚烯-6-基、6,7,8,9-四氢-5H-苯并环庚烯基、5,6,7,8,9,10-六氢-苯并环辛烯基、芴基、二环[2.2.1]庚基、7,7-二甲基-二环[2.2.1]庚基、二环[2.2.1]庚烯基、二环[2.2.2]辛基、二环[3.1.1]庚基、二环[3.2.1]辛基、二环[2.2.2]辛烯基、二环[3.2.1]辛烯基、金刚烷基、八氢-4,7-亚甲基-1H-茚基和八氢-2,5-亚甲基-并环戊二烯并基等。
在本申请中，作为基团或是其它基团的一部分，术语“杂环基”意指由2至14个碳原子(例如2、3、4、5、6、7、8、9、10、11、12、13或14个碳原子)以及1至6个选自氮、磷、氧和硫的杂原子组成的稳定的3元至20元非芳香族环状基团。除非本说明书中另外特别指明，否则杂环基可以为单环、双环、三环或更多环的环体系，其可包括稠合环体系、桥环体系或螺环体系；其杂环基中的氮、碳或硫原子可任选地被氧化；氮原子可任选地被季铵化；且杂环基可为部分或完全饱和。杂环基可以经由碳原子或者杂原子并通过单键与分子其余部分连接。在包含稠环的杂环基中，一个或多个环可以是下文所定义的芳基或杂芳基，条件是与分子其余部分的连接点为非芳香族环原子。就本发明的目的而言，杂环基优选为包含1至3个选自氮、氧和硫的杂原子的稳定的4元至11元非芳香性单环、双环、桥环或螺环基团，更优选为包含1至3个选自氮、氧和硫的杂原子的稳定的4元至8元非芳香性单环、双环、桥环或螺环基团。杂环基的实例包括但不限于：吡咯烷基、吗啉基、哌嗪基、高哌嗪基、哌啶基、硫代吗啉基、2,7-二氮杂-螺[3.5]壬烷-7-基、2-氧杂-6-氮杂-螺[3.3]庚烷-6-基、2,5-二氮杂-双环[2.2.1]庚烷-2-基、氮杂环丁烷基、吡喃基、四氢吡喃基、噻喃基、四氢呋喃基、噁嗪基、二氧环戊基、四氢异喹啉基、十氢异喹啉基、咪唑啉基、咪唑烷基、喹嗪基、噻唑烷基、异噻唑烷基、异噁唑烷基、二氢吲哚基、八氢吲哚基、八氢异吲哚基、吡咯烷基、吡唑烷基、邻苯二甲酰亚氨基等。
在本申请中，作为基团或是其它基团的一部分，术语“芳基”意指具有6至18个碳原子(优选具有6至10个碳原子，例如6、7、8、9或10个碳原子)的共轭烃环体系基团。就本发明的目的而言，芳基可以为单环、双环、三环或更多环的环体系，还可以与上文所定义的环烷基或杂环基稠合，条件是芳基经由芳香环上的原子通过单键与分子的其余部分连接。芳基的实例包括但不限于苯基、萘基、蒽基、菲基、芴基、2,3-二氢-1H-异吲哚基、2-苯并噁唑啉酮、2H-1,4-苯并噁嗪-3(4H)-酮-7-基等。
在本申请中，术语“芳基烷基”是指被上文所定义的芳基所取代的上文所定义的烷基。
在本申请中，作为基团或是其它基团的一部分，术语“杂芳基”意指环内具有1至15个碳原子(优选具有1至10个碳原子，例如1、2、3、4、5、6、7、8、9或10个碳原子)和1至6个选自氮、氧和硫的杂原子的5元至16元共轭环系基团。除非本说明书中另外特别指明，否则杂芳基可为单环、双环、三环或更多环的环体系，还可以与上文所定义的环烷基或杂环基稠合，条件是杂芳基经由芳香环上的原子通过单键与分子的其余部分连接。杂芳基中的氮、碳或硫原子可任选地被氧化；氮原子可任选地被季铵化。就本发明的目的而言，杂芳基优选为包含1至5个选自氮、氧和硫的杂原子的稳定的5元至12元芳香性基团，更优选为包含1至4个选自氮、氧和硫的杂原子的稳定的5元至10元芳香性基团或者包含1至3个选自氮、氧和硫的杂原子的5元至6元芳香性基团。杂芳基的实例包括但不限于噻吩基、咪唑基、吡唑基、噻唑基、噁唑基、噁二唑基、异噁唑基、吡啶基、嘧啶基、吡嗪基、哒嗪基、苯并咪唑基、苯并吡唑基、吲哚基、呋喃基、吡咯基、三唑基、四唑基、三嗪基、吲嗪基、异吲哚基、吲唑基、异吲唑基、嘌呤基、喹啉基、异喹啉基、二氮萘基、萘啶基、喹噁啉基、蝶啶基、咔唑基、咔啉基、菲啶基、菲咯啉基、吖啶基、吩嗪基、异噻唑基、苯并噻唑基、苯并噻吩基、噁三唑基、噌啉基、喹唑啉基、苯硫基、中氮茚基、邻二氮杂菲基、异噁唑基、吩噁嗪基、吩噻嗪基、4,5,6,7-四氢苯并[b]噻吩基、萘并吡啶基、[1,2,4]三唑并[4,3-b]哒嗪、[1,2,4]三唑并[4,3-a]吡嗪、[1,2,4]三唑并[4,3-c]嘧啶、[1,2,4]三唑并[4,3-a]吡啶、咪唑并[1,2-a]吡啶、咪唑并[1,2-b]哒嗪、咪唑并[1,2-a]吡嗪等。
在本申请中，术语“杂芳基烷基”是指被上文所定义的杂芳基所取代的上文所定义的烷基。
在本申请中，“任选地”或“任选地”表示随后描述的事件或状况可能发生也可能不发生，且该描述同时包括该事件或状况发生和不发生的情况。例如，“任选地被取代的芳基”表示芳基被取代或未被取代，且该描述同时包括被取代的芳基与未被取代的芳基。本发明权利要求书和说明书部分所述的“任选地”的取代基选自烷基、烯基、炔基、卤素、卤代烷基、卤代烯基、卤代炔基、氰基、硝基、任选取代的芳基、任选取代的杂芳基、任选取代的环烃基、任选取代的杂环基。
“SHP2”指“Src Homolgy-2磷酸酶”，还叫SH-PTP2、SH-PT3、Syp、PTP1D、PTP2C、SAP-2或PTPN11。
本文所用术语“部分”、“结构部分”、“化学部分”、“基团”、“化学基团”是指分子中的特定片段或官能团。化学部分通常被认为是嵌入或附加到分子上的化学实体。
“立体异构体”是指由相同原子组成，通过相同的键键合，但具有不同三维结构的化合物。本发明将涵盖各种立体异构体及其混合物。
当本发明的化合物中含有烯双键时，除非另有说明，否则本发明的化合物旨在包含E-和Z-几何异构体。
“互变异构体”是指质子从分子的一个原子转移至相同分子的另一个原子而形成的异构体。本发明的化合物的所有互变异构形式也将包含在本发明的范围内。
本发明的化合物或其药学上可接受的盐可能含有一个或多个手性碳原子，且因此可产生对映异构体、非对映异构体及其它立体异构形式。每个手性碳原子可以基于立体化学而被定义为(R)-或(S)-。本发明旨在包括所有可能的异构体，以及其外消旋体和光学纯形式。本发明的化合物的制备可以选择外消旋体、非对映异构体或对映异构体作为原料或中间体。光学活性的异构体可以使用手性合成子或手性试剂来制备，或者使用常规技术进行拆分，例如采用结晶以及手性色谱等方法。
制备/分离个别异构体的常规技术包括由合适的光学纯前体的手性合成，或者使用例如手性高效液相色谱法拆分外消旋体(或盐或衍生物的外消旋体)，例如可参见Gerald Gübitz and Martin G.Schmid(Eds.),Chiral Separations,Methods and Protocols,Methods in Molecular Biology,Vol.243,2004；A.M.Stalcup,Chiral Separations,Annu.Rev.Anal.Chem.3:341-63,2010；Fumiss et al.(eds.),VOGEL’S ENCYCLOPEDIA OF PRACTICAL ORGANIC CHEMISTRY 5.sup.TH ED.,Longman Scientific and Technical Ltd.,Essex,1991,809-816；Heller,Acc.Chem.Res.1990,23,128。
本发明还包括本发明的化合物或其药学上可接受的盐的所有适宜的同位素变体。本发明的化合物或其药学上可接受的盐的同位素变体被定义为其中至少一个原子被具有相同原子数、但原子质量与自然界经常发现的原子质量不同的原子所替换的那些。可以掺入到本发明的化合物及其药学上可接受的盐中的同位素包括但不限于H、C、N和O的同位素，例如 2H、 3H、 11C、 13C、 14C、 15N、 17O、 18O、 35S、 18F、 36Cl和 125I。本发明所述化合物或其药学上可接受的盐的同位素变体可以通过常规技术、采用适宜试剂的适当同位素变体来制备。
在本申请中，术语“药学上可接受的盐”包括药学上可接受的酸加成盐和药学上可接受的碱加成盐。
“药学上可接受的酸加成盐”是指能够保留游离碱的生物有效性而无其它副作用的，与无机酸或有机酸所形成的盐。无机酸盐包括但不限于盐酸盐、氢溴酸盐、硫酸盐、硝酸盐、磷酸盐等；有机酸盐包括但不限于甲酸盐、乙酸盐、2,2-二氯乙酸盐、三氟乙酸盐、丙酸盐、己酸盐、辛酸盐、癸酸盐、十一碳烯酸盐、乙醇酸盐、葡糖酸盐、乳酸盐、癸二酸盐、己二酸盐、戊二酸盐、丙二酸盐、草酸盐、马来酸盐、琥珀酸盐、富马酸盐、酒石酸盐、柠檬酸盐、棕榈酸盐、硬脂酸盐、油酸盐、肉桂酸盐、月桂酸盐、苹果酸盐、谷氨酸盐、焦谷氨酸盐、天冬氨酸盐、苯甲酸盐、甲磺酸盐、苯磺酸盐、对甲苯磺酸盐、海藻酸盐、抗坏血酸盐、水杨酸盐、4-氨基水杨酸盐、萘二磺酸盐等。这些盐可通过本专业已知的方法制备。
“药学上可接受的碱加成盐”是指能够保持游离酸的生物有效性而无其它副作用的、与无机碱或有机碱所形成的盐。衍生自无机碱的盐包括但不限于钠盐、钾盐、锂盐、铵盐、钙盐、镁盐、铁盐、锌盐、铜盐、锰盐、铝盐等。优选的无机盐为铵盐、钠盐、钾盐、钙盐及镁盐。衍生自有机碱的盐包括但不限于以下的盐：伯胺类、仲胺类及叔胺类，被取代的胺类，包括天然的被取代胺类、环状胺类及碱性离子交换树脂，例如氨、异丙胺、三甲胺、二乙胺、三乙胺、三丙胺、乙醇胺、二乙醇胺、三乙醇胺、二甲基乙醇胺、2-二甲氨基乙醇、2-二乙氨基乙醇、二环己胺、赖氨酸、精氨酸、组氨酸、咖啡因、普鲁卡因、胆 碱、甜菜碱、乙二胺、葡萄糖胺、甲基葡萄糖胺、可可碱、嘌呤、哌嗪、哌啶、N-乙基哌啶、聚胺树脂等。优选的有机碱包括异丙胺、二乙胺、乙醇胺、三甲胺、二环己基胺、胆碱及咖啡因。这些盐可通过本专业已知的方法制备。
在本申请中，“药物组合物”是指本发明化合物与本领域通常接受的用于将生物活性化合物输送至哺乳动物(例如人)的介质的制剂。该介质包括药学上可接受的载体。药物组合物的目的是促进生物体的给药，利于活性成分的吸收进而发挥生物活性。
本文所用术语“药学上可接受的”是指不影响本发明化合物的生物活性或性质的物质(如载体或稀释剂)，并且相对无毒，即该物质可施用于个体而不造成不良的生物反应或以不良方式与组合物中包含的任意组分相互作用。
在本申请中，“药学上可接受的赋形剂”包括但不限于任何被相关的政府管理部门许可为可接受供人类或家畜使用的佐剂、载体、赋形剂、助流剂、增甜剂、稀释剂、防腐剂、染料/着色剂、矫味剂、表面活性剂、润湿剂、分散剂、助悬剂、稳定剂、等渗剂、溶剂或乳化剂。
本发明所述“肿瘤”包括但不限于努南综合症、豹综合症、青少年髓单核细胞白血病、成神经细胞瘤、肉瘤、黑色素瘤、关节软骨瘤、胆管瘤、白血病、乳腺癌、胃肠间质瘤、组织细胞性淋巴瘤、非小细胞肺癌、小细胞肺癌、食道癌、胰腺癌、肺鳞癌、肺腺癌、乳腺癌、前列腺癌、肝癌、皮肤癌、上皮细胞癌、宫颈癌、卵巢癌、肠癌、鼻咽癌、脑癌、骨癌、肾癌、口腔癌/头癌、成神经细胞瘤、头颈的鳞状细胞癌、间变性大细胞淋巴瘤或成胶质细胞瘤等疾病。
本文所用术语“预防的”、“预防”和“防止”包括使病患减少疾病或病症的发生或恶化的可能性。
本文所用的术语“治疗”和其它类似的同义词包括以下含义：
(i)预防疾病或病症在哺乳动物中出现，特别是当这类哺乳动物易患有该疾病或病症，但尚未被诊断为已患有该疾病或病症时；
(ii)抑制疾病或病症，即遏制其发展；
(iii)缓解疾病或病症，即，使该疾病或病症的状态消退；或者
(iv)减轻该疾病或病症所造成的症状。
本文所使用术语“有效量”、“治疗有效量”或“药学有效量”是指服用后足以在某种程度上缓解所治疗的疾病或病症的一个或多个症状的至少一种药剂或化合物的量。其结果可以为迹象、症状或病因的消减和/或缓解，或生物系统的任何其它所需变化。例如，用于治疗的“有效量”是在临床上提供显著的病症缓解效果所需的包含本文公开化合物的组合物的量。可使用诸如剂量递增试验的技术测定适合于任意个体病例中的有效量。
本文所用术语“服用”、“施用”、“给药”等是指能够将化合物或组合物递送到进行生物作用的所需位点的方法。这些方法包括但不限于口服途径、经十二指肠途径、胃肠外注射(包括静脉内、皮下、腹膜内、肌内、动脉内注射或输注)、局部给药和经直肠给药。本领域技术人员熟知可用于本文所述化合物和方法的施用技术，例如在Goodman and Gilman, The Pharmacological Basis of Therapeutics,current ed.；Pergamon；and Remington’s,Pharmaceutical Sciences(current edition),Mack Publishing Co.,Easton,Pa中讨论的那些。在优选的实施方案中，本文讨论的化合物和组合物通过口服施用。
本文所使用术语“药物组合”、“药物联用”、“联合用药”、“施用其它治疗”、“施用其它治疗剂”等是指通过混合或组合不止一种活性成分而获得的药物治疗，其包括活性成分的固定和不固定组合。术语“固定组合”是指以单个实体或单个剂型的形式向患者同时施用至少一种本文所述的化合物和至少一种协同药剂。术语“不固定组合”是指以单独实体的形式向患者同时施用、合用或以可变的间隔时间顺次施用至少一种本文所述的化合物和至少一种协同制剂。这些也应用到鸡尾酒疗法中，例如施用三种或更多种活性成分。
本领域技术人员还应当理解，在下文所述的方法中，中间体化合物官能团可能需要由适当的保护基保护。这样的官能团包括羟基、氨基、巯基及羧酸。合适的羟基保护基包括三烷基甲硅烷基或二芳基烷基甲硅烷基(例如叔丁基二甲基甲硅烷基、叔丁基二苯基甲硅烷基或三甲基甲硅烷基)、四氢吡喃基、苄基等。合适的氨基、脒基及胍基的保护基包括叔丁氧羰基、苄氧羰基等。合适的巯基保护基包括-C(O)-R”(其中R”为烷基、芳基或芳烷基)、对甲氧基苄基、三苯甲基等。合适的羧基保护基包括烷基、芳基或芳烷基酯类。
保护基可根据本领域技术人员已知的和如本文所述的标准技术来引入和除去。保护基的使用详述于Greene,T.W.与P.G.M.Wuts,Protective Groups in Organi Synthesis,(1999),4th Ed.,Wiley中。保护基还可为聚合物树脂。
式I化合物的制备
本发明提供的所述式I化合物，可通过如下的方法制备：使式Ib与式Ic发生亲核取代反应得到式Id；使式Id与式Ie发生取代反应得到式If；并且将式If用酸脱除保护以得到式I化合物：
Figure PCTCN2019115754-appb-000009
式中，各基团的定义如上文中所述。
药理学和用途
Src Hommolgy-2磷酸酶(SHP2)是PTPN11基因所编码的蛋白酪氨酸磷酸酶，其促 进多种细胞功能，包括增殖、分化、细胞周期维持和迁移。SHP2牵涉在经由Ras-有丝分裂原-激活的蛋白激酶、JAK-STAT或磷酸肌醇3-激酶-AKT途径的信号传导中。SHP2介导受体酪氨酸激酶如ErbB1、ErbB2和c-Met的Erk1和Erk2MAP激酶的激活。
SHP2具有两个N-末端Src homolgy2结构域(N-SH2和C-SH2)、催化结构域(PTP)和C-末端尾。所述两个SH2结构域控制SHP2的亚细胞定位和功能调控。该分子以无活性构象存在，经由牵涉来自N-SH2和PTP结构域的残基的结合网络来抑制其自身活性。响应于生长因子的刺激，SHP2经由其SH2结构域结合停靠蛋白上的特定酪氨酸-磷酸化位点如Gab1和Gab2。这引起了构象变化，导致SHP2激活。
已经在多种人类疾病中识别到了PTPN11中的突变，所述疾病例如有努南综合征、豹斑综合征、幼年型粒-单核细胞白血病、成神经细胞瘤、黑素瘤、急性髓样白血病以及乳房、肺和结肠的癌症。SHP2是多种受体酪氨酸激酶、包括血小板衍生生长因子(PDGF-R)、成纤维细胞生长因子(FGF-R)和表皮生长因子(EGF-R)的受体的重要下游信号分子。SHP2还是激活促分裂原活化蛋白(MAP)激酶途径的重要下游信号分子，其可导致细胞转化(癌发展的必要条件)。SHP2的敲减显著抑制了具有SHP2突变或EML4/ALK易位的肺癌细胞系以及EGFR扩增乳癌和食管癌的细胞生长。SHP2还是胃癌、间变型大细胞淋巴癌和成胶质细胞瘤的癌基因的激活下游。
努南综合征(NS)和豹斑综合征(LS)-PTPNll突变引起LS(多发性色素斑样痣综合征，心电图传导异常，两眼距离过远，肺动脉瓣狭窄，异常生殖器，生长迟缓，感音神经性耳聋)和NS(包括心脏缺陷、颅面骨畸形和身材矮小的先天异常)。这两种障碍是RAS/RAF/MEK/ERK有丝分裂原活化蛋白激酶途径(正常细胞生长和分化所需)的组分中的种系突变引起的常染色体显性综合征家族的一部分。该途径的异常调控具有深远的影响，特别是对心脏发育具有深远的影响，导致多种异常，包括瓣膜中隔缺陷(valvuloseptal defects)和/或肥厚型心肌病(HCM)。已经确定MAPK信号传导途径的的扰动对于这些障碍而言是重要的，已经在人中识别出沿循该途径的一些候选基因，包括KRAS、NRAS、SOS1、RAF1、BRAF、MEKl、MEK2、SH0C2和CBL中的突变。在NS和LS中最常突变的基因是PTPN11。在～50％的NS病例中和几乎所有的具有NS的某些特征的LS患者中发现了PTPN11(SHP2)的种系突变。对于NS，蛋白质中的Y62D和Y63C是最常见的突变。这两种突变影响SHP2的无催化活性的构象，而不干扰磷酸酶与其磷酸化信号配偶体的结合。
幼年型粒-单核细胞白血病(JMML)—在约35％的JMML(—种儿童期骨髓增生病(MPD))患者中发生PTPN11(SHP2)中的体细胞突变。这些功能获得性突变通常是N-SH2结构域或磷酸酶结构域中的点突变，其阻止催化结构域和N-SH2结构域之间的自我抑制，产生SHP2活性。
急性髓样白血病——已经在～10％的儿科急性白血病如骨髓发育不良综合征(MDS)、～7％的B细胞急性淋巴母细胞白血病(B-ALL)和～4％的急性髓样白血病(AML)中识别出PTPNll突变。
NS和白血病突变引起位于由自身抑制SHP2构象中的N-SH2和PTP结构域所形成 的界面处的氨基酸的变化，破坏抑制性分子内相互作用，导致催化结构域活动过度。
SHP2在受体酪氨酸激酶(RTK)信号传导中充当正调节剂。含有RTK改变(EGFR amp,Her2 amp，FGFR amp，Met amp，易位/活化的RTK，即ALK，BCR/ABL)的癌症包括食管癌、乳癌、肺癌、结肠癌、胃癌、神经胶质瘤、头颈癌。
食管癌(或食道癌)是食管的恶性病。存在多种亚型，主要是鳞状细胞癌(<50％)和腺癌。在食管腺癌和鳞状细胞癌中有高比率的RTK表达。因此，本发明的SHP2抑制剂可用于创新的治疗策略。
乳癌是一种重要类型的癌症，并且是女性的主要死因，其中患者对现有药物出现抗性。存在四种主要的乳癌亚型，包括luminal A、luminal B、Her21ik和三阴/Basal-like。三阴乳癌(TNBC)是缺少特定靶向治疗的侵袭性乳癌。表皮生长因子受体I(EGFR)已经显现为TNBC中的有前景的靶标。Her2以及EGFR经由SHP2的抑制可以是乳癌的有前景的治疗。
肺癌——NSCLC目前是癌症相关死亡率的重要原因。占约85％的肺癌(主要是腺癌和鳞状细胞癌)。虽然细胞毒性化学治疗仍然是治疗的重要部分，但是基于肿瘤中遗传改变如EGFR和ALK的靶向治疗更可能得益于靶向治疗。
结肠癌——已知约30％至50％的结肠直肠肿瘤具有突变(异常)的KRAS，BRAF突变发生在10至15％的结肠直肠癌中。对于其结肠直肠肿瘤已经被证明过表达EGFR的患者亚群而言，这些患者呈现出对抗-EGFR疗法的有利的临床响应。
胃癌是最流行的癌症类型之一。酪氨酸激酶的异常表达(如通过胃癌细胞中的异常酪氨酸磷酸化所反映的那样)是本领域已知的。在胃癌中经常扩增三种受体酪氨酸激酶，即c-met(HGF受体)、FGF受体2和erbB2/neu。因此，不同信号途径的破坏可促进不同胃癌类型的进程。
成神经细胞瘤是发育中的交感神经系统的儿科肿瘤，占约8％的儿童癌症。间变性淋巴瘤激酶(ALK)基因的基因组改变已经被提出促进成神经细胞瘤发病机制。
头和颈鳞状细胞癌(SCCHN)——高水平的EGFR表达与多种癌症、最常见是头和颈鳞状细胞癌(SCCHN)中的预后不良和对放射治疗的抗性相关。EGFR信号阻断导致抑制受体刺激、细胞增殖、侵袭和转移下降。因此，在SCCHN中，EGFR是新抗癌疗法的最佳靶标。
本发明涉及能够抑制SHP2活性的化合物。本发明还提供了本发明的化合物的制备方法和包含该化合物的药物制剂。本发明的另一方面涉及治疗与SHP2-介导的疾病或病症的方法，该方法包括给需要其的患者施用治疗有效量的如本发明的式I化合物的步骤。
在一些实施方式中，本发明涉及如上所述的方法，其中所述SHP2-介导的疾病或病症是选自但不限于如下的癌症：JMML，AML，MDS，B-ALL，成神经细胞瘤，食管癌，乳癌，肺癌，结肠癌，胃癌，头颈癌。
本发明的化合物还可用于治疗与SHP2异常活性相关的其它疾病或病症。因此，作为优选的实施方式，本发明涉及治疗选自如下的疾病或病症的方法：NS，LS，JMML，AML， MDS，B-ALL，成神经细胞瘤，食管癌，乳癌，肺癌，结肠癌，胃癌，头颈癌。
本发明的SHP2抑制剂可以与其它具有药理活性的化合物或与两种或更多种其它具有药理活性的化合物组合，尤其是在治疗癌症中。例如，本发明的式(I)化合物或其药学上可接受的盐可以与一种或多种选自如下的物质组合地同时、依次或分别施用：化疗剂，比如有丝分裂抑制剂，如紫杉烷、长春花生物碱、紫杉醇、多西他赛、长春花新碱、长春花碱、长春瑞滨或长春氟宁，其它抗癌剂如顺铂、5-氟尿嘧啶或5-氟-2-4(1H，3H)-嘧啶二酮(5FU)、氟他胺或吉西他滨。
一些组合在疗法中可以提供显著的优点，包括协同活性。
在一些实施方式中，本发明涉及如上所述的方法，其中所述化合物经胃肠外施用。
在一些实施方式中，本发明涉及如上所述的方法，其中所述化合物经肌内、静脉内、皮下、口服、经肺、鞘内、局部或经鼻内施用。
在一些实施方式中，本发明涉及如上所述的方法，其中所述化合物经全身施用。
在一些实施方式中，本发明涉及如上所述的方法，其中所述患者是哺乳动物。
在一些实施方式中，本发明涉及如上所述的方法，其中所述患者是灵长类动物。
在一些实施方式中，本发明涉及如上所述的方法，其中所述患者是人。
在一些实施方式中，本发明涉及治疗SHP2-介导的疾病或病症的方法，该方法包括如下步骤：给需要其的患者施用治疗有效量的化疗剂与如治疗有效量的本发明的式I化合物的组合。
本发明的主要优点包括：
1、提供了一种式I化合物。
2、提供了一种结构新颖的SHP2抑制剂及其制备和应用，所述抑制剂对SHP2具有较高的抑制活性。
3、提供了一类治疗与SHP2相关疾病或病症的药物组合物。
下面结合具体实施例，进一步阐述本发明。应理解，这些实施例仅用于说明本发明而不用于限制本发明的范围。下列实施例中未注明具体条件的实验方法，通常按照常规条件，或按照制造厂商所建议的条件。除非另外说明，否则百分比和份数按重量计算。
下述实施例中所用的起始物可由化学品销售商如Aldrich、TCI、Alfa Aesar、毕得、安耐吉等处购得，或者可通过已知的方法来合成。
下述实施例中所涉及的英文简称所代表的意思如下表所述。
Figure PCTCN2019115754-appb-000010
Figure PCTCN2019115754-appb-000011
下述实施例中，冰浴是指-5℃至0℃，室温是指10℃至30℃，回流温度一般是指溶剂在常压下的回流温度。反应过夜一般是指时间为8-15小时。下述实施例中，未限定具体操作温度的，均在室温下进行。
下述实施例中，中间体和最终产物的分离提纯是通过正相或反相色谱柱分离或者其它合适的方法。正相快速色谱柱是用乙酸乙酯和正己烷或甲醇和二氯甲烷等作为流动相。反相制备性高压液相色谱(HPLC)是用C18柱并用UV 214nm和254nm来检测，其流动相为A(水和0.1％甲酸)、B(乙腈)或者流动相A(水和0.1％碳酸氢铵)、B(乙腈)。
各实施例中：LCMS仪器：Pump Agilent 1260 UV检测器：Agilent 1260 DAD Mass Spectrometer API 3000
层析柱：Waters sunfire C18,4.6×50mm,5um
流动相：A-H2O(0.1％HCOOH)；B-乙腈NMR
仪器：Bruker Ascend 400M(1H NMR:400MHz；13C NMR:100MHz)。
中间体A1的合成:(R)-N-((S)-1，3-二氢螺[茚-2,4’-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺
Figure PCTCN2019115754-appb-000012
步骤一：在干燥的100mL单口瓶中依次加入1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(1.51g,5mmol)，钛酸四乙酯(6.84g,30mmol)，(R)-(+)-叔丁基亚磺酰胺(2.41g,，20mmol)在加热回流下搅拌反应15小时。冷却到室温后，向反应残存物中加入饱和食盐水(60mL)，之后将得到的混合物搅拌15分钟然后通过硅藻土过滤。将含水的混合物用乙酸乙酯(3x80mL)萃取。将有机相用Na 2SO 4干燥，过滤，并在减压下除去挥发物。将得到的残留物通过硅胶色谱法纯化(0至30％梯度的乙酸乙酯：石油醚)得到白色固体(R,Z)-1-((叔丁基亚磺酰基)亚胺基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A1-1,1.61g,收率：80％)。
1H NMR(400MHz,CDCl 3)δ8.34(d,J＝7.2Hz,1H),7.48–7.39(m,1H),7.35–7.28(m,2H),4.21–3.92(m,2H),3.00(s,2H),2.88(t,J＝11.9Hz,2H),2.00–1.80(m,2H),1.48–1.30(m,11H),1.24(d,J＝13.1Hz,9H)；LCMS：m/z 405.1[M+H] +。
步骤二：在干燥的100mL单口瓶中依次加入(R,Z)-1-((叔丁基亚磺酰基)亚胺基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A1-1,0.802g,2mmol)，THF(10mL)，冷却到0℃后，随后加入硼氢化锂(66mg,3mmol)。将得到的混合物继续搅拌反应1小时。将甲醇缓慢加入以淬灭过量的硼氢化物，过滤，浓缩，并在减压下除去挥发物。将得到的残留物通过硅胶色谱法纯化(0至50％梯度的乙酸乙酯：石油醚)得到淡黄色固体(S)-1-(((R)-叔丁基亚磺酰基)胺基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A1-2，0.63g,收率：78％)。
1H NMR(400MHz,DMSO)δ7.30–7.15(m,4H),5.64(d,J＝10.5Hz,1H),4.38(d,J＝10.5Hz,1H),3.86(s,2H),3.05(d,J＝15.8Hz,1H),2.87(s,2H),2.62(d,J＝15.8Hz,1H),1.89(s,1H),1.61–1.35(m,12H),1.27–1.10(m,11H)；LCMS：m/z 407.1[M+H] +。
步骤三：在干燥的50mL单口瓶中依次加入(S)-1-(((R)-叔丁基亚磺酰基)胺基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A1-2，0.406g,1mmol)，二氯甲烷(5mL)，三氟乙酸(1mL)将得到的混合物在室温下搅拌反应1小时。加入Na 2CO 3饱和水溶液直到pH 7，并将含水的混合物用DCM(3x30mL)萃取。将合并的有机相用盐水洗涤，用Na 2SO 4干燥，过滤，并在减压下除去挥发物。冷却得到无色油状物(R)-N-((S)-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺(A1，0.183g,收率：70％)。
1H NMR(400MHz,CDCl 3)δ7.26–7.11(m,5H),4.45(d,J＝10.1Hz,1H),3.77(s,1H),3.18(s,2H),3.04(d,J＝15.9Hz,3H),2.67(d,J＝15.8Hz,1H),2.20(td,J＝12.7,3.5Hz,1H),1.82(t,J＝11.1Hz,1H),1.61(d,J＝12.9Hz,1H),1.34–1.11(m,10H)；
LCMS：m/z 307.1[M+H] +。
中间体A2的合成:R-N-((S)-5，7-二氢螺[环戊二烯并[b]吡啶-6,4’-哌啶]-5-基)-2-甲基丙烷-2-亚磺酰胺
Figure PCTCN2019115754-appb-000013
步骤一：向干燥的100mL烧瓶中依次加入4-氰基哌啶-1-甲酸叔丁酯(1.05g，5mmol)和THF(20mL)。在氮气条件下，将混合物降温-78℃，然后向反应混合物中缓慢加入2M LDA(3.3mL，6.5mmol)。让反应混合物反应1小时，然后向其中加入3-溴-2-(溴甲基)吡啶(1.24g，5mmol)，然后让反应混合物继续反应2小时。反应完毕后，向其中加入饱和氯化铵溶液(15mL)淬灭反应，并使用乙酸乙酯(3×30mL)萃取，混合有机层并饱和食盐水洗涤，经无水硫酸钠干燥，过滤并将减压浓缩得到的残留物通过硅胶色谱法纯化(0至30％梯度的乙酸乙酯/石油醚)，得到白色固体4-((3-溴吡啶-2-基)甲基)-4-氰基哌啶-1'-甲酸叔丁酯(A2-1，1.40g,收率：75％)
步骤二：在氮气保护下，向干燥的25mL单口烧瓶中依次加入4-((3-溴吡啶-2-基)甲基)-4-氰基哌啶-1'-甲酸叔丁酯(A2-1，379mg,1mmol)、三乙胺(404mg,4mmol)、二氯双二叔丁基-(4-二甲基氨基苯基)磷钯(II)Pd(AmPhos) 2Cl 2(71mg,0.1mmol)和DMA：H2O＝10：1(6mL)，然后在130℃下搅拌反应18小时。反应完毕后，将获得的残留物过滤并将减压浓缩得到的残留物通过硅胶色谱法纯化(0至50％梯度的乙酸乙酯：石油醚)，得到黄色固体5-氧代-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A2-2，180mg,收率：60％)LCMS：m/z 303.1[M+H] +。
步骤三：在干燥的100mL单口瓶中依次加入5-氧代-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A2-2,0.302g,1mmol)，钛酸四乙酯(1.37g,6mmol)，(R)-(+)-叔丁基亚磺酰胺(0.480g,4mmol)在加热回流下搅拌反应15小时。冷却到室温后，向反应残存物中加入饱和食盐水(15mL)，之后将得到的混合物搅拌15分钟然后通过硅藻土过滤。将含水的混合物用乙酸乙酯(3x300mL)萃取。将有机相用Na 2SO 4干燥，过滤，并在减压下除去挥发物。将得到的残留物通过硅胶色谱法纯化(0至50％梯度的乙酸乙酯：石油醚)得到黄色固体(R，Z)-5-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A2-3,0.333g,收率：82％)。LCMS：m/z 406.1[M+H] +。
步骤四：在干燥的100mL单口瓶中依次加入(R，Z)-5-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A2-3,0.20g,0.491mmol)，THF(50mL)，冷却到℃后，随后加入硼氢化锂(0.018g,0.737mmol)。将得到的混合物继续搅拌反应1小时。将甲醇缓慢加入以淬灭过量的硼氢化物，过滤，浓缩，并在减压下除去挥发物。将得到的残留物通过硅胶色谱法纯化(0至80％梯度的乙酸乙酯：石油醚)得到白色固体(S)-5-((R)-叔丁基亚磺酰胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A2-4,0.130g,收率：65％)。LCMS：m/z 408.1[M+H] +。
步骤五：在干燥的50mL单口瓶中依次加入(S)-5-((R)-叔丁基亚磺酰胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A2-4,0.100g,0.245mmol)，二氯甲烷(5mL)，三氟乙酸(1mL)将得到的混合物在室温下搅拌反应1小时。加入Na 2CO 3饱和水溶液直到pH 7，并将含水的混合物用DCM(3x30mL)萃取。将合并的有机相用盐水洗涤，用Na 2SO 4干燥，过滤，并在减压下除去挥发物。冷却得到无色油状物R-N-((S)-5，7-二氢螺[环戊二烯并[b]吡啶-6,4’-哌啶]-5-基)-2-甲基丙烷-2-亚磺酰胺(A2,0.056g,收率：75％)。LCMS：m/z 308.1[M+H] +。
中间体A3的合成:(S)-1，3-二氢螺[茚-2,4’-哌啶]-1-胺
Figure PCTCN2019115754-appb-000014
步骤：向干燥的100mL圆底烧瓶中依次加入甲醇(1mL),二氯甲烷(1mL)和A1-2(540mg,1.33mmol,1.0eq)。室温下，滴加HCl/1,4-dioxane(3.3mL,4M)，有白色固体析出，加热到50℃,搅拌两小时。反应完毕后，减压浓缩得到白色固体(S)-1，3-二氢螺[茚-2,4’-哌啶]-1-胺(A3,373mg,收率：97.1％,HCl盐)。LCMS：m/z 203.1[M+H] +。
中间体A4的合成:(S)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000015
参照中间体A3的合成方法，使用中间体A2-4代替中间体A1-2,得到中间体(S)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-胺(A4)。LCMS：m/z 204.1[M+H] +.
中间体A5的合成:(S)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000016
参考中间体A2,A3的合成方法合成A5.LCMS：m/z 204.1[M+H] +.
中间体A6的合成:(S)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000017
步骤一：向含300mL THF和60mL H 2O的1L烧瓶中依次加入A6-1(11.1g，60mmol)和NaBH 4(2.51g,66mmol)并在20℃下反应2小时。点板检测原料反应完后，用饱和NH4Cl溶液淬灭，用水稀释，并用乙酸乙酯萃取，合并有机相用饱和NaCl溶液洗涤，经无水硫酸钠干燥，过滤并减压浓缩得到白色固体2-溴-吡啶-3-基甲醇(A6-2,11.2g,收率：100％)
步骤二：向含120mL二氯甲烷的干燥的250mL单口烧瓶中依次加入(A6-2,6.73g,36mmol)和Ms 2O(6.96g,g,40mmol)后冷却至0℃，再慢慢加入TEA(5.45g，54mmol)升温至室温后搅拌3小时，反应完后反应液用水洗涤，有机相用无水硫酸钠干燥，过滤并减压浓缩，得到的残留物通过硅胶色谱法纯化(0至10％梯度的乙酸乙酯/二氯甲烷)，得到无色油状物甲基磺酸(2-溴吡啶-3-基)甲酯(A6-3,8.02g,收率：84％)。
步骤三：向干燥的500mL三口烧瓶中依次加入4-乙氧羰基哌啶-1-甲酸叔丁酯(9.3g，36.2mmol)和THF(133mL)。在氮气条件下，将混合物降温至-70℃，然后向反应混合物中缓慢加入2M LDA(21.1mL，42.3mmol)。让反应混合物反应1小时，然后向其中加入溶解于65mL THF中的甲基磺酸(2-溴吡啶-3-基)甲酯(A6-3,8.0g，30.2mmol)，然后让反应混合物继续反应0.5小时之后缓慢升温至室温并搅拌1小时。反应完毕后，向其中加入饱和食盐水淬灭，并使用乙酸乙酯萃取，合并有机相用饱和食盐水洗涤，经无水硫酸钠干燥，过滤并再减压浓缩得到的残留物通过硅胶色谱法纯化(8至12％梯度的乙酸乙酯/二氯甲烷)，得到无色油状物4-乙氧羰基-4-(2-溴-3-吡啶基)甲基-哌啶-1-甲酸叔丁酯(A6-4,10.5g,收率：81％)
步骤四：向干燥的250mL三口烧瓶中依次加入4-乙氧羰基-4-(2-溴-3-吡啶基)甲基-哌啶-1-甲酸叔丁酯(A6-4,7.85g，18.4mmol)和THF(120mL)。在氮气条件下，将混合物降温-70℃，然后向反应混合物中缓慢加入2.5M正丁基锂(11mL，27.6mmol)。让反应混合物反应1.5小时。反应完毕后，向其中加入饱和氯化铵水溶液淬灭，之后加入饱和食盐水稀释并分液，水相用乙酸乙酯萃取，合并有机相用饱和食盐水洗涤，经无水硫酸钠干燥，过滤并再减压浓缩。得到的残留物通过硅胶色谱法纯化(0至60％梯度的乙酸乙酯/石油醚)，得到浅棕色固体7-氧代-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A6-5,1.4g,收率：25％)
步骤五：参照中间体A2-3的合成方法，使用中间体A6-5代替中间体A2-2,得到中间体(R，Z)-7-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'- 甲酸叔丁酯(A6-6,1.46g,收率：68％)。
步骤六：在干燥的50mL三口瓶中依次加入(R，Z)-7-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A6-6,530mg,1.31mmol)，THF(10mL)，氮气保护下冷却至-70℃，然后向反应混合物中缓慢加入1.5M二异丁基氢化铝的甲苯溶液(1.3mL，1.95mmol)。让反应混合物反应0.5小时后缓慢升温至室温，用饱和酒石酸钾钠水溶液淬灭后并搅拌0.5小时。用乙酸乙酯萃取，合并有机相用饱和食盐水洗涤，经无水硫酸钠干燥，过滤并再减压浓缩得到的残留物通过硅胶色谱法纯化(0至5％梯度的甲醇/二氯甲烷)，得到浅黄色泡沫固体(S)-7-((R)-叔丁基亚磺酰胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A6-7,466mg,收率：87％)
步骤七：在干燥的25mL单口瓶中依次加入(S)-7-((R)-叔丁基亚磺酰胺基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A6-7,312mg,0.766mmol)，MeOH(5mL)，4M的HCl/1,4-dioxane溶液(3.83mL，15.3mmol)，然后升温至50℃反应8小时，得到白色悬浊液，减压浓缩得到白色固体(S)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺HCl盐(A6,232mg,收率：97％)。LCMS：m/z 204.1[M+H] +。
中间体A7的合成:(S)-6-氟-1,3-二氢螺[茚-2,4'-哌啶]-1-胺
Figure PCTCN2019115754-appb-000018
步骤一：参照中间体A2的合成方法，使用原料4-氟苄溴代替原料3-溴-2-(溴甲基)吡啶,得到中间体4-乙氧羰基-4-(4-氟苄基)-哌啶-1-甲酸叔丁酯A7-1。
步骤二：向含20mL甲醇和20mL水的100mL单口烧瓶中依次加入化合物4-乙氧羰基-4-(4-氟苄基)-哌啶-1-甲酸叔丁酯(A7-1,3.40g,9.30mmol)和氢氧化钠(1.86g,46.5mmol)。反应液在70℃下反应17小时。冷却至室温后，减压浓缩除去挥发物。得到的残留物加水(50mL)稀释并用稀盐酸调pH至3，用乙酸乙酯(80mL)萃取3次。合并的有机相用饱和食盐水洗涤、无水硫酸钠干燥、过滤、减压浓缩得到黄色固体1-叔丁氧羰基-4-(4-氟苄基)-哌啶-4-甲酸(A7-2,3.0g,粗品)。
步骤三：向干燥的50mL单口烧瓶中依次加入化合物1-叔丁氧羰基-4-(4-氟苄基)-哌啶-4-甲酸(A7-2,2.0g,5.93mmol)和PPA(15mL)。反应液在120℃下反应2小时。将反应液趁热倒入到冰水混合物(50mL)中，用NaOH固体调节到pH＝10。然后向得到的混合 物中加入Boc 2O(1.94g,8.90mmol)，并在20℃下搅拌1小时。反应液用乙酸乙酯(80mL)萃取3次。合并的有机相用饱和食盐水洗涤、无水硫酸钠干燥、过滤、减压浓缩。得到的残留物通过硅胶色谱法纯化(石油醚/乙酸乙酯＝5:1)，得到黄色固体6-氟-1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A7-3,1.20g,收率：63.5％)。
1HNMR(400MHz,CDCl3)δ7.46–7.38(m,2H),7.37–7.30(m,1H),4.20–4.06(m,2H),3.07–2.95(m,4H),1.96–1.85(m,2H),1.48(s,9H),1.44–1.35(m,2H).
步骤四：参照中间体A2的合成方法，使用中间体A7-3代替中间体A2-2,得到中间体(R,Z)-1-((叔丁基亚磺酰基)亚胺基)-6-氟-1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸酯A7-4。
步骤五：在-50℃下向圆底烧瓶中依次加入化合物(R,Z)-1-((叔丁基亚磺酰基)亚胺基)-6-氟-1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸酯(A7-4,1.59g,3.76mmol),四氢呋喃/水(98:2,32mL)和硼氢化钠(427mg,11.3mmol),反应液在搅拌下3小时升温至20℃。TLC点板反应完全。反应液加水(30mL)稀释，用乙酸乙酯(30mL)萃取3次。合并的有机相用饱和食盐水洗涤、无水硫酸钠干燥、过滤、减压浓缩。得到的残留物通过硅胶色谱法纯化(石油醚/乙酸乙酯＝2:1)，得到白色固体(S)-1-(((R)-叔丁基亚磺酰基)胺基)-6-氟-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A7-5,850mg,收率：53％)。
步骤六：参照中间体A3的合成方法，使用中间体A7-5代替中间体A1-2,得到中间体(S)-6-氟-1,3-二氢螺[茚-2,4'-哌啶]-1-胺A7。LCMS：m/z 220.1[M+H] +。
中间体A8:(S)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000019
步骤一：将4-溴烟醛氢溴酸盐(A8-1,，2.5g，9.36mmol)溶于甲醇(50mL)中，冰浴下分批次加入硼氢化钠(0.72g,18.93mmol,2.0eq)，0度反应1小时，反应完毕。冰浴下，加入饱和氯化铵水溶液(50mL)淬灭反应，加入乙酸乙酯萃取(100mL x 2)。有机相合并后用饱和氯化钠水洗，有机相分离后用无水硫酸钠干燥，过滤并浓缩得到白色固体(4-溴吡啶-3-基)甲醇(A8-2,1.7g，收率：100％)。LCMS：m/z 190.3[M+H]+
步骤二：将(4-溴吡啶-3-基)甲醇(A8-2，1.7g,9.04mmol)溶于二氯甲烷(100mL)中，冰浴下加入三乙胺(2.30g,22.7mmol)。氮气保护下分批次加入甲磺酸酐(1.95g，11.2mmol)，0度反应2小时后，反应完毕。冰浴下，加入饱和氯化钠(50mL)淬灭反应，分离有机相并用饱和氯化钠水洗。有机相用无水硫酸钠干燥，过滤浓缩得到红棕色固体甲基磺酸(4-溴吡啶-3-基)甲酯(A8-3,1.7g，收率：70.7％)。LCMS：m/z 266.2[M+H]+
步骤三：将4-氰基哌啶-1-甲酸叔丁酯(1.61g,7.67mmol)溶于无水四氢呋喃(60mL)中，干冰丙酮浴(-78度)氮气保护下，缓慢滴入LDA(4.6mL,9.2mmol)，保温反应0.5小时，反应液有大量白色固体析出。-78度缓慢滴入甲基磺酸(4-溴吡啶-3-基)甲酯(A8-3,1.7g，6.39mmol)的无水四氢呋喃(50mL)溶液，继续保温反应2小时，反应完毕。加入饱和氯化氨(100mL)淬灭反应，加入乙酸乙酯萃取(100mL x3)。合并有机相并用饱和氯化钠水洗，有机相用无水硫酸钠干燥，过滤浓缩残留物通过硅胶色谱法纯化(0至50％梯度的二氯甲烷/石油醚)得到白色固体4-((4-溴吡啶-3-基)甲基)-4-氰基哌啶-1-甲酸叔丁酯(A8-4,1.58g，收率：54％)。LCMS：m/z 326.0[M+H-56]+
步骤四：将4-((4-溴吡啶-3-基)甲基)-4-氰基哌啶-1-甲酸叔丁酯(A8-4，1.5g,3.95mmol)溶于DMAc/H2O(100mL/10mL)中，加入DIPEA(1.34g,15.8mmol)和Pd(AmPhos) 2Cl 2(142mg,0.20mmol),氮气置换三次,130度反应2小时，反应完毕。加入水(50mL)，用乙酸乙酯萃取(100mL x3)，合并有机相并用饱和氯化钠水洗，有机相用无水硫酸钠干燥，过滤浓缩残留物通过硅胶色谱法纯化得到固体5-氧代-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A8-5,750mg，收率：62％).LCMS：m/z 303.3[M+H]+
步骤五：将化合物5-氧代-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A8-5,750mg,2.48mmol)溶于THF(10mL)中，加入(R)-(+)-叔丁基亚磺酰胺(390mg,3.22mmol)与钛酸四乙酯(10mL)。氮气保护加热至90度回流反应18小时。反应完毕，冷却至室温加入乙酸乙酯(50mL)稀释，加入饱和食盐水(10mL)，有白色固体析出。将混合物过滤，滤饼用乙酸乙酯洗涤。滤液用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到白色固体(R，Z)-5-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A8-6,800mg,收率：80％).LCMS：m/z 406.2[M+H] +
步骤六：将(R，Z)-5-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A8-6,800mg,1.98mmol)溶于THF(50mL)中，氮气保护下冷却至-78度。向反应液缓慢滴入1.5M的DIBAL-H(2mL,3mmol)，滴加完毕，-78度反应1小时，反应完毕。加水淬灭反应，加入饱和酒石酸钾钠溶液(20mL)，用乙酸乙酯萃取(100mL x 2)，有机相用饱和氯化钠水溶液洗一次，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到白色固体(S)-5-(((R)-叔丁基亚磺酰基)胺基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A8-7,680mg，收率：84％).LCMS：m/z 408.3[M+H]+
步骤七：将(S)-5-(((R)-叔丁基亚磺酰基)氨基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-1'-甲酸叔丁酯(A8-7,680mg,1.67mmol)溶于MeOH(50mL)中，冷却至0度，滴入4M盐酸二氧六环溶液(10mL,40mmol)，室温反应3小时，反应完毕。反应液浓缩得到白色固体(S)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-7-胺盐酸盐(A8,400mg，收率：99％).LCMS：m/z 204.1[M+H] +
中间体A9:(S)-1-氨基-1,3-二氢螺[茚-2,4'-哌啶]-4-醇
Figure PCTCN2019115754-appb-000020
步骤一：将2-溴-6-羟基苯甲醛(5g,24.9mmol)溶于DMF(100mL)中，加入无水碳酸钾(6.88g,49.8mmol)与4-甲氧基苄溴(5.26g,26.1mmol)，氮气保护下室温反应18小时，反应完毕。将反应液倒入冰水，用乙酸乙酯萃取(250mL)两次。有机相合并用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到浅黄色固体2-溴-6-((4-甲氧基苄基)氧基)苯甲醛(A9-1,8g，收率100％)。LCMS：m/z 343.0[M+Na] +
步骤二：将2-溴-6-((4-甲氧基苄基)氧基)苯甲醛(A9-1,8g，24.9mmol)溶于乙醇(100mL)中，冰浴下冷却至0度。小心分多批次加入硼氢化钠(942mg,24.9mmol)。0度反应0.5小时，反应完毕。反应液倒入冰水中用乙酸乙酯萃取(200mL)两次。合并有机相用饱和食盐水洗，无水硫酸钠干燥，过滤并浓缩得到固体(2-溴-6-((4-甲氧基苄基)氧基)苯基)甲醇(A9-2,7.3g，收率：90％)。LCMS：m/z 345.0[M+Na] +
步骤三：将(2-溴-6-((4-甲氧基苄基)氧基)苯基)甲醇(A9-2,7.3g，22.6mmol)溶于二氯甲烷(200mL)中，冰浴下冷却至0度。氮气保护下加入四溴化碳(11.2g,33.9mmol)和三苯基磷(8.88g,33.9mmol)。0度反应5小时，反应完毕。反应液倒入冰水中用乙酸乙酯萃取(200mL)两次。合并有机相用饱和食盐水洗，无水硫酸钠干燥，过滤并浓缩。粗品通过硅胶色谱法纯化得到固体1-溴-2-(溴甲基)-3-((4-甲氧基苄基)氧基)苯(A9-3,4g，收率：46％)。
1H NMR(400MHz,CDCl 3)δ7.40(t,J＝4.4Hz,2H),7.17-7.20(m,1H),7.11(t,J＝8.2Hz,1H),6.92-6.95(m,2H),6.87-6.89(m,1H),5.30(s,2H),4.76(s,2H),3.83(s,3H)；
步骤四：将4-氰基哌啶-1-甲酸叔丁酯(2.6g，12.4mmol)溶于无水THF(60mL)中，冷却至-78度。缓慢滴入2M LDA(7.5mL,14.9mmol)。-78度反应0.5小时，继续滴加1-溴-2-(溴甲基)-3-((4-甲氧基苄基)氧基)苯(A9-3,4g,10.4mmol)的无水THF(40mL)溶液。-78度反应2小时，缓慢升温至0度。反应完毕。反应液倒入冰水中用乙酸乙酯萃取(200mL)两次。合并有机相用饱和食盐水洗，无水硫酸钠干燥，过滤并浓缩。粗品通过硅胶色谱法纯化得到固体4-(2-溴-6-((4-甲氧基苄基)氧基)苄基)-4-氰基哌啶-1-甲酸叔丁酯(A9-4,2.8g，收率：44％)。LCMS：m/z 537.0[M+Na] +
步骤五：将4-(2-溴-6-((4-甲氧基苄基)氧基)苄基)-4-氰基哌啶-1-甲酸叔丁酯(A9-4,2.6g，5.0mmol)溶于DMAc(100mL)中，加入H2O(10mL)，DIPEA(3.225g，25.0mmol)和PdCl 2(AmPhos) 2(354mg，0.5mmol)。氩气保护下130度反应18小时，反应完毕。冷 却至室温将反应液倒入冰水中用乙酸乙酯萃取(300mL)两次。合并有机相用水及饱和食盐水洗一次，无水硫酸钠干燥，过滤并浓缩。粗品通过硅胶色谱法纯化得到固体4-((4-甲氧基苄基)氧基)-1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A9-5,1.8g，收率：82％)。LCMS：m/z 382.2[M-56] +
步骤六：将4-((4-甲氧基苄基)氧基)-1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A9-5,1.8g,4.12mmol)溶于THF(20mL)中，加入(R)-2-甲基丙烷-2-亚磺酰胺(748mg,6.18mmol)与钛酸四乙酯(50mL)。氮气保护加热至100度回流反应18小时。反应完毕.冷却至室温加入乙酸乙酯(200mL)稀释，加入饱和食盐水(50mL)，有白色固体析出。将混合物过滤，滤饼用乙酸乙酯洗涤。滤液用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到白色固体(R，Z)-1-((叔丁基亚磺酰基)亚氨基)-4-((4-甲氧基苄基)氧基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A9-6,1.7g,收率：76％)。LCMS：m/z 541.3[M+H] +
步骤七：将(R，Z)-1-((叔丁基亚磺酰基)亚胺基)-4-((4-甲氧基苄基)氧基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A9-6,1.7g,3.15mmol)溶于THF(100mL)中，氮气保护下冷却至-78度。反应液缓慢滴入1.5M的DIBAL-H(3.15mL,4.7mmol)，滴加完毕，-78度反应1小时，反应完毕。加水淬灭反应，加入饱和酒石酸钾钠溶液(50mL)，用乙酸乙酯萃取(200mL x 2)，有机相用饱和氯化钠水溶液洗一次，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到白色固体(S)-1-(((R)-叔丁基亚磺酰基)胺基)-4-((4-甲氧基苄基)氧基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A9-7,1.5g，收率：88％).LCMS：m/z 543.3[M+H] +
步骤八：将(S)-1-(((R)-叔丁基亚磺酰基)胺基)-4-((4-甲氧基苄基)氧基)-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A9-7,1.5g,2.77mmol)溶于MeOH(50mL)中，冷却至0度，滴入4M盐酸二氧六环溶液(10mL,40mmol)，室温反应3小时，反应完毕。反应液浓缩得到白色固体(S)-1-氨基-1,3-二氢螺[茚-2,4'-哌啶]-4-醇盐酸盐(A9,600mg，收率：85％).LCMS：m/z 219.2[M+H] +
中间体A10:(S)-1-(((R)-叔丁基亚磺酰)胺基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]
Figure PCTCN2019115754-appb-000021
步骤一：向100mL的圆底烧瓶中依次加入2-甲基-3-氰基溴苯(3.0g,15.3mmol,1.0eq),N-溴代琥珀酰亚胺(2.72g,15.3mmol,1.0eq),过氧化二苯甲酰(371mg,1.53mmol,0.1eq)和 四氯化碳(40mL),80℃搅拌16小时，反应液减压浓缩后得到残留物，将残留物溶于乙酸乙酯(200mL),用2N NaOH水溶液(50mL)洗涤两次，饱和食盐水(50mL)洗涤一次，无水硫酸钠干燥，过滤。滤液减压浓缩得粗产品，粗品通过硅胶色谱法纯化得到白色固体1-溴-2-(溴甲基)-3-氰基苯(A10-1，2.0g，收率：47.5％)。
步骤二：向100mL的圆底烧瓶中依次加入四氢呋喃(30mL)和4-氰基哌啶-1-甲酸叔丁酯(1.84g,8.73mmol,1.2eq),降温到-78℃,然后加入2.0M LDA(5.1mL,10.2mmol,1.4eq),-78℃下搅拌一小时。然后加入1-溴-2-(溴甲基)-3-氰基苯(A10-1,2.0g,7.27mmol,1.0eq)的四氢呋喃(15mL)溶液,-78℃下搅拌0.5小时。然后撤去低温浴，让其自然回复至室温，继续搅拌一小时。TLC检测原料消耗完毕后，加入饱和食盐水(30mL)淬灭，然后用乙酸乙酯(100mL*2)萃取。合并的有机相用饱和食盐水(50mL)洗涤，无水硫酸钠干燥，过滤。滤液减压浓缩得粗产品，粗品通过硅胶色谱法纯化得到淡黄色油状物叔丁基-4-(2-溴-6-氰基苄基)-4-氰基哌啶-1-甲酸酯(A10-2，1.5g，收率：51％)。LC-MS：m/z 404.1，406.1[M+H] +。
步骤三：向干燥的100mL的圆底三口烧瓶中依次加入叔丁基-4-(2-溴-6-氰基苄基)-4-氰基哌啶-1-甲酸酯(A10-2，1.5g,3.71mmol,1.0eq),Pd(AmPhos) 2Cl 2(262mg,0.37mmol.0.1eq),二异丙基乙基胺(2.4g,18.5mmol,5.0eq),N,N-二甲基乙酰胺(30mL)和水(4mL)。在搅拌的情况下，用氮气置换三次，然后加热到140℃,反应16小时。反应完毕后，反应液冷却至室温,用乙酸乙酯(100mL)稀释并减压抽滤，滤饼用乙酸乙酯(20mL)洗涤,将得到的滤液用饱和食盐水(30mL)洗涤3次,无水硫酸钠干燥，过滤。滤液减压浓缩得粗产品，粗品通过硅胶色谱法纯化得到白色固体叔丁基-4-氰基-1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1-甲酸酯(A10-3，0.90g，收率：74.3％)。LC-MS：m/z 327.2[M+H] +。
步骤四：在干燥的100mL单口瓶中依次加入叔丁基-4-氰基-1-氧代-1,3-二氢螺[茚-2,4'-哌啶]-1-甲酸酯(A10-3，900mg,2.76mmol)，四乙氧基钛(3.78g,16.6mmol)，(R)-(+)-叔丁基亚磺酰胺(401mg,3.31mmol)和四氢呋喃(20mL)在加热回流下搅拌反应16小时。冷却到室温后，向反应残存物中加入饱和食盐水(60mL)，之后将得到的混合物搅拌15分钟然后通过硅藻土过滤。将含水的混合物用乙酸乙酯(3x80mL)萃取。将有机相用无水硫酸钠干燥，过滤，并在减压下浓缩。将得到的残留物通过硅胶色谱法纯化(0至30％梯度的乙酸乙酯：石油醚)得到白色固体(R,Z)-1-((叔丁基亚磺酰)亚胺)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A10-4，980mg,收率：82.7％)。LCMS：m/z 430.2[M+H] +。
步骤五：向干燥的50mL三口烧瓶中依次加入四氢呋喃(15mL)和(R,Z)-1-((叔丁基亚磺酰)亚胺)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A10-4，980mg,2.28mmol,1.0eq)。在-78℃，氮气保护条件下，滴加二异丁基氢化铝(6.8mL,1.5M in toluene,10.3mmol,4.5eq)，继续搅拌半小时。然后升至0℃，继续搅拌半小时。反应完毕后，用酒石酸钾钠(4g溶解在20mL水中)淬灭，搅拌半小时，用乙酸乙酯(30mL*3)萃取，得到的有机相用饱和食盐水(30mL)洗涤，无水硫酸钠干燥，过滤，并在减压下浓缩，残留物通过硅胶色谱法纯化(0至70％梯度的乙酸乙酯：石油醚)得到黄色固体(S)-1-(((R)-叔丁基亚磺酰)氨基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A10-5，800mg,收率：81.3％)。LC-MS：m/z 432.2[M+H] +。
步骤六：在干燥的50mL单口瓶中依次加入(S)-1-(((R)-叔丁基亚磺酰)氨基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸叔丁酯(A10-5，800mg,1.85mmol)，二氯甲烷(15mL)，三氟乙酸(5mL)将得到的混合物在室温下搅拌反应1小时。加入Na 2CO 3饱和水溶液直到pH＝7，并将含水的混合物用二氯甲烷(3x50mL)萃取。将合并的有机相用食盐水洗涤，用无水硫酸钠干燥，过滤，并在减压下浓缩，残留物通过硅胶色谱法纯化(0至70％梯度的乙酸乙酯：石油醚)得到黄色固体(R)-N-((S)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-甲基丙基-2-亚磺酰胺(A10，490mg,收率：79.8％)。LCMS：m/z 332.2[M+H] +。
中间体A11:(S)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000022
步骤一：将2,2,6,6-四甲基哌啶(5.33g,37.8mmol)溶于100mL四氢呋喃，氩气保护，干冰丙酮浴降温至-65度，滴加正丁基锂(2.5M，15.8mL,39.5mmol),然后0度保温30分钟，再次降温到-65度，滴加2-溴吡嗪(5.0g,31.4mmol)与10mL四氢呋喃混合液，保温30分钟，滴加DMF(5.75g,78.6mmol)与10mL四氢呋喃混合液，继续搅拌2小时，反应完全。继续于-65度滴加75mL甲醇，搅拌10分钟，缓慢加入NaBH 4(2.38g,62.9mmol)，缓慢升温到0度，保温1小时，反应完全。将反应液倒入500mL冰NH 4Cl水溶液中，乙酸乙酯萃取3次，有机相饱和盐水洗一次，合并有机相，无水硫酸钠干燥，过滤，浓缩，过柱，得黄色固体(3-溴吡啶-2-基)甲醇(A11-1，4.33g,收率：72.9％)。LCMS：m/z 191[M+H] +
步骤二：将(3-溴吡啶-2-基)甲醇(A11-1，4.33g,22.9mmol)溶于80mL乙醚中，氮气保护，0度滴加PBr 3(6.83g,25.2mmol),然后加热至回流约40度，反应4小时，反应完毕。将反应液倒入到冰NaHCO 3水溶液中，二氯甲烷萃取2次，饱和食盐水洗一次，合并有机相，无水硫酸钠干燥，过滤，浓缩，过柱得近无色透明油状物2-溴-3-(溴甲基)吡嗪(A11-2，5.14g,收率：89.0％)。LCMS：m/z 252.9[M+H] +
步骤三：将N-Boc-4-哌啶甲酸乙酯(6.81g,26.5mmol)溶于160mL无水四氢呋喃中，氩气保护下，干冰丙酮浴降温至-65度，缓慢滴入LDA(2.0M，16.8mL,33.6mmol)，保温反应1小时；继续在-65度滴入2-溴-3-(溴甲基)吡嗪(A11-2，5.14g,24.1mmol)的无水四氢呋喃(30mL)溶液，继续保温反应2小时，反应完毕。将反应液倒入300mL 冰NH 4Cl水溶液中，乙酸乙酯萃取3次，有机相饱和食盐水洗一次，合并有机相，无水硫酸钠干燥，过滤，浓缩，过柱，得1-(叔丁基)-4-乙基4-((3-溴吡嗪-2-基)甲基)哌啶-1,4-二羧酸酯(A11-3，5.81g,收率：56.4％)。LCMS：m/z 328.1[M+H-100] +
步骤四：将1-(叔丁基)-4-乙基4-((3-溴吡嗪-2-基)甲基)哌啶-1,4-二羧酸酯(A11-3，5.8g,13.5mmol)溶于300mL四氢呋喃中，氩气保护下，干冰丙酮浴降温至-65度，滴加正丁基锂(2.5M,8.2mL,20.3mmol),自然升温至-10度，保温3小时，反应完全。将反应液倒入300mL冰NH 4Cl水溶液中，乙酸乙酯萃取3次，有机相饱和盐水洗一次，合并有机相，无水硫酸钠干燥，过滤，浓缩，过柱，得黄色固体5-氧代-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-1'-甲酸叔丁酯(A11-4，2.23g，收率：54.4％)。
LCMS：m/z 304.2[M+H] +微弱
步骤五：将5-氧代-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-1'-甲酸叔丁酯(A11-4，2.23g，7.35mmol)溶于350mL四氢呋喃中，加入(R)-2-甲基丙烷-2-亚磺酰胺(980mg,8.09mmol)与钛酸四乙酯(21.8g,95.6mmol)。氮气保护加热至80度回流反应10小时。反应完毕。冷却至室温，倒入到400mL冰水中,加入300mL乙酸乙酯萃取3次(水相含大量白色絮状固体，有机相几乎没有固体)，有机相在用200mL水和200mL饱和食盐水各洗一次，合并有机相，无水硫酸钠干燥，过滤，浓缩，过柱，得淡橙色泡沫状固体(R，Z)-5-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-1'-甲酸叔丁酯(A11-5，2.38g，收率：79.6％)。LCMS：m/z 407.1[M+H] +
步骤六：将(R，Z)-5-((叔丁基亚磺酰基)亚胺基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-1'-甲酸叔丁酯(A11-5，2.38g，5.85mmol)溶于280mL四氢呋喃中，氩气保护下，干冰丙酮浴降温至-65度。缓慢滴入1.5M的DIBAL-H(5.07mL,7.61mmol)，继续保温2小时，反应完毕。将反应液倒至300mL饱和酒石酸钾钠水溶液中，乙酸乙酯萃取3次，合并有机相，无水硫酸钠干燥，过滤，浓缩干，得金黄色泡沫状固体(S)-5-(((R)-叔丁基亚磺酰基)胺基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-1'-甲酸叔丁酯粗品(A11-6，2.48g)。LCMS：m/z 309.3[M+H-100] +
步骤七：将(S)-5-(((R)-叔丁基亚磺酰基)胺基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-1'-甲酸叔丁酯粗品(A11-6，2.48g，5.85mmol)溶于120mL甲醇中，氩气保护下，冰水降温至0度左右，滴入4M盐酸二氧六环溶液(18mL,72mmol)，室温反应2小时，反应完毕。将反应液浓缩干，加入无水乙腈打浆，氩气保护下过滤，得墨绿色固体粉末(极易吸潮成墨绿色油滴)(S)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺盐酸盐(A11，1.77g,收率：99％)，氩气下保存。LCMS：m/z 205.3[M+H] +
中间体A12:(S)-1-(((R)-叔丁基亚磺酰)氨基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]
Figure PCTCN2019115754-appb-000023
步骤一：向100mL的圆底烧瓶中依次加入2-甲基-3-甲氧基溴苯(2.0g，9.95mmol，1.0eq),N-溴代琥珀酰亚胺(1.77g，9.95mmol，1.0eq),过氧化二苯甲酰(241mg，0.995mmol，0.1eq)和四氯化碳(40mL),80℃搅拌16小时，反应液减压浓缩后得到残留物，将残留物溶于乙酸乙酯(200mL),用2N NaOH水溶液(50mL*2)洗涤两次，饱和食盐水(50mL)洗涤一次，无水硫酸钠干燥，过滤。滤液减压浓缩得粗产品，粗品通过硅胶色谱法纯化得到白色固体1-溴-2-(溴甲基)-3-甲氧基苯(A12-1，2.0g，收率：71.8％)。
步骤二：向100mL的圆底烧瓶中依次加入四氢呋喃(30mL)和N-Boc-4-氰基哌啶(1.8g,8.57mmol,1.2eq),降温到-78℃,然后加入2.0M LDA(5mL,10mmol,1.4eq),-78℃下搅拌一小时。然后加入1-溴-2-(溴甲基)-3-甲氧基苯(A12-1，2.0g,7.14mmol,1.0eq)的四氢呋喃(15mL)溶液,-78℃下搅拌0.5小时。然后撤去低温浴，让其自然回复至室温，继续搅拌一小时。TLC检测原料消耗完毕后，加入饱和食盐水(30mL)淬灭，然后用乙酸乙酯(100mL*2)萃取。合并的有机相用饱和食盐水(50mL)洗涤，无水硫酸钠干燥，过滤。滤液减压浓缩得粗产品，粗品通过硅胶色谱法纯化得到淡黄色油状物N-Boc-4-(2-溴-6-甲氧基苄基)-4-氰基哌啶(A12-2，1.5g，收率：51.3％)。LCMS：m/z 409.1，411.1[M+H] +。
步骤三：向干燥的100mL的圆底三口烧瓶中依次加入N-Boc-4-(2-溴-6-甲氧基苄基)-4-氰基哌啶(A12-2，1.5g,3.66mmol,1.0eq),Pd(AmPhos) 2Cl 2(259mg,0.37mmol.0.1eq),二异丙基乙基胺(2.37g,18.3mmol,5.0eq),N,N-二甲基乙酰胺(30mL)和水(4mL)。在搅拌的情况下，用氮气置换三次，然后加热到140℃,反应16小时。反应完毕后，反应液冷却至室温,用乙酸乙酯(100mL)稀释并减压抽滤，滤饼用乙酸乙酯(20mL)洗涤,将得到的滤液用饱和食盐水(30mL*3)洗涤3次,无水硫酸钠干燥，过滤。滤液减压浓缩得粗产品，粗品通过硅胶色谱法纯化得到白色固体N-Boc-4-甲氧基-1-氧代-1,3-二氢螺[茚-2,4'-哌啶](A12-3，0.90g，收率：74.1％)。
LC-MS：m/z 332.2[M+H] +。
步骤四：在干燥的100mL单口瓶中依次加入N-Boc-4-甲氧基-1-氧代-1,3-二氢螺[茚-2,4'-哌啶](A12-3，900mg,2.72mmol)，四乙氧基钛(3.72g,16.3mmol)，(R)-(+)-叔丁基亚磺酰胺(395mg,3.26mmol)和四氢呋喃(20mL)在加热回流下搅拌反应16小时。冷却到室温后，向反应残存物中加入饱和食盐水(60mL)，之后将得到的混合物搅拌15分钟然后通过硅藻土过滤。将含水的混合物用乙酸乙酯(3x80mL)萃取。将有机相用无水硫酸 钠干燥，过滤，并在减压下浓缩。将得到的残留物通过硅胶色谱法纯化(0至30％梯度的乙酸乙酯：石油醚)得到白色固体N-Boc-(R,Z)-1-((叔丁基亚磺酰)亚胺)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶](A12-4，800mg,收率：67.8％)。
LC-MS：m/z 435.2[M+H] +。
步骤五：向干燥的50mL三口烧瓶中依次加入四氢呋喃(15mL)和N-Boc-(R,Z)-1-((叔丁基亚磺酰)亚胺)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶](A12-4，800mg,1.84mmol,1.0eq)。在-78℃下，氮气保护条件下，滴加二异丁基氢化铝(5.52mL,1.5M in toluene,8.28mmol,4.5eq)，继续搅拌半小时。然后升至0℃，继续搅拌半小时。反应完毕后，用酒石酸钾钠(4g溶解在20mL水中)淬灭，搅拌半小时，用乙酸乙酯(30mL*3)萃取，得到的有机相用饱和食盐水(30mL)洗涤，无水硫酸钠干燥，过滤，并在减压下浓缩，残留物通过硅胶色谱法纯化(0至70％梯度的乙酸乙酯：石油醚)得到黄色固体叔丁基(S)-1-(((R)-叔丁基亚磺酰基)胺基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸酯(A12-5，720mg,收率：89.6％)。LCMS：m/z 437.2[M+H] +。
步骤六：在干燥的50mL单口瓶中依次加入叔丁基(S)-1-(((R)-叔丁基亚磺酰基)胺基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1'-甲酸酯(A12-5，720mg,1.65mmol)，二氯甲烷(15mL)，三氟乙酸(5mL)将得到的混合物在室温下搅拌反应1小时。加入Na 2CO 3饱和水溶液直到pH＝7，并将含水的混合物用二氯甲烷(3x50mL)萃取。将合并的有机相用盐水洗涤，用无水硫酸钠干燥，过滤，并在减压下浓缩，残留物通过硅胶色谱法纯化(0至70％梯度的乙酸乙酯：石油醚)得到黄色固体(R)-N-((S)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-甲基丙基-2-亚磺酰胺(A12，520mg,收率：93.7％)。LCMS：m/z 337.2[M+H] +。
中间体A13:(S)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-4-胺
Figure PCTCN2019115754-appb-000024
步骤一：将1-叔丁基4-乙基哌啶-1,4-二甲酸酯(10.7g，41.6mmol，1.2eq)溶于THF(50mL)中，-78℃下滴加LDA(2M,9.8mL,19.6mmol,1.1eq)，此温度下搅拌反应2h，再将2-氯-5-(氯甲基)噻唑(A13-1,3g，34.7mmol,1.0eq)溶于THF(10mL)后滴加体系中。加毕继续此温度反应1.5h。TLC检测少许未反应。加水淬灭，乙酸乙酯(100mL x 2)萃取，有机相合并后用饱和氯化钠水洗，有机相分离后用无水硫酸钠干燥，过滤并浓缩，拌样过柱(PE:EA:DCM＝4:1:1)得黄色油状物1-叔丁基-4-乙基4-(2-氯噻唑-5-基)甲基)哌啶-1,4- 二羧酸酯(A13-2,2.4g，收率35％)。
步骤二：将1-叔丁基-4-乙基4-(2-氯噻唑-5-基)甲基)哌啶-1,4-二羧酸酯(A13-2，4g,10.3mmol)溶于THF(100mL)中，-78℃滴加LDA(2M,8.5mL,16.6mmol)。反应1h,反应完全。加入饱和氯化胺(100mL)淬灭反应，加入乙酸乙酯萃取(100mL x3)。合并有机相并用饱和氯化钠水洗，有机相用无水硫酸钠干燥，过滤浓缩滤液通过硅胶色谱法纯化得到黄色油状物(A13-3,2-氯-4-氧代-4,6-二氢螺[环戊二烯[d]噻唑-5,4'-哌啶]-1'-甲酸叔丁酯)(1.9g，收率54％)LCMS：m/z 343[M+H] +
步骤三：将2-氯-4-氧代-4,6-二氢螺[环戊二烯[d]噻唑-5,4'-哌啶]-1'-甲酸叔丁酯(A13-3，710mg,2.07mmol)和Ti(OEt) 4以及(R)-2-甲基丙烷-2-亚磺酰胺(276mg,2.28mmol)加到反应瓶中，氮气保护下加热到100℃反应5h。反应完毕，冷却至室温加入乙酸乙酯(50mL)稀释，加入饱和食盐水(15mL)，有白色固体析出。将混合物过滤，滤饼用乙酸乙酯洗涤。滤液用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到浅黄固体叔丁基(E)-4-((叔丁基亚磺酰基)亚胺基)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-1'-甲酸叔丁酯(A13-4,600mg,收率：68％).LCMS：m/z 426[M+H] +
步骤四：将叔丁基(E)-4-((叔丁基亚磺酰基)亚胺基)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-1'-甲酸叔丁酯(A13-4，200mg，0.47mmol)溶于THF(10mL)中，氮气保护下冷却至-78度。反应液缓慢滴入1.5M的DIBAL-H(0.5mL,0.75mmol)，滴加完毕，-78度反应1小时，反应完毕。加水淬灭反应，加入饱和酒石酸钾钠溶液(20mL)，用乙酸乙酯萃取(50mL x 2)，有机相用饱和氯化钠水溶液洗一次，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到浅黄固体叔丁基(S)-4-(((R)-叔丁基亚磺酰基)胺基)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-1'-甲酸叔丁酯A13-5(155mg，收率：76％).LCMS：m/z 448[M+H] +
步骤五：将叔丁基(S)-4-(((R)-叔丁基亚磺酰基)胺基)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-1'-甲酸叔丁酯(A13-5,155mg,0.35mmol)溶于4M盐酸二氧六环溶液(5mL,20mmol)中，室温反应3小时，反应完毕。反应液浓缩得到浅黄色油,(S)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-4-胺(A13，145mg，盐酸盐).LCMS：m/z 244[M+H] +
中间体B2的合成:5-氯-8-碘-[1,2,4]三氮唑并[4,3-c]嘧啶
Figure PCTCN2019115754-appb-000025
步骤一：向干燥的100mL烧瓶中加入2,4-二氯-5-碘嘧啶(1.1g,4mmol)和20mL无水乙醇。在0℃氮气条件下，向其中缓慢加入80％水合肼(601mg，12mmol)混合物继续搅拌反应1小时。反应完毕后，将混合物过滤并用无水乙醇洗涤得到2-氯-4-肼基-5-碘嘧啶(B2-1，850mg，产率78.7％)。
1H NMR(400MHz,CDCl 3)δ8.29(s,1H),6.67(s,1H),4.08(s,2H)；
LCMS：m/z 271.1[M+H] +。
步骤二：向干燥的100mL烧瓶中依次加入2-氯-4-肼基-5-碘嘧啶(810mg，3mmol)和原甲酸三甲酯(10mL)。在氮气条件下，将混合物加热至85℃搅拌反应5小时。反应完毕后，将获得的残留物倒入饱和NaCl溶液(50mL)，并使用乙酸乙酯(3×30mL)萃取，饱和食盐水洗涤并混合有机层，经无水硫酸钠干燥，过滤并在减压浓缩得到的残留物通过硅胶色谱法纯化(0至50％梯度的乙酸乙酯/石油醚)，得到淡黄色固体5-氯-8-碘-[1,2,4]三唑并[4,3-c]嘧啶(B2，420mg,收率：50％)。
LCMS：m/z 280.9[M+H] +。
中间体B3的合成:5-氯-8-碘咪唑并[1,2-c]嘧啶
Figure PCTCN2019115754-appb-000026
步骤一：向干燥的100mL烧瓶中依次加入2,4-二氯-5-碘嘧啶(1.37g,5mmol)和2,2-二甲氧基乙胺(8.4g,10mmol)和无水乙醇(50mL)。然后在0℃氮气条件下，向反应混合物中缓慢滴加三乙胺(1.01g,10mmol)之后将混合物在室温下搅拌反应10小时。反应完毕后，真空浓缩，得到的浓缩物加入15mL的水，并使用二氯甲烷(3×50mL)萃取，饱和食盐水洗涤并混合有机层，经无水硫酸钠干燥，过滤并浓缩得到白色固体2-氯-N-(2,2-二甲氧基乙基)-5-碘嘧啶-4-胺(B3-1，1.46g，收率:85％)。
LC-MS：m/z 344.2[M+H] +。
步骤二：向干燥的100mL烧瓶中依次加入2-氯-N-(2,2-二甲氧基乙基)-5-碘嘧啶-4-胺(B3-1,1.03g,3mmol)和10mL浓硫酸。在氮气条件下，将混合物加热至65℃搅拌反应2小时。反应完毕后，反应液冷却至室温，将混合物缓慢倒入冰水中，然后用4M的NaOH溶液调节pH约到6-7，过滤得到灰白色固体8-碘咪唑并[1,2-c]嘧啶-5-醇(B3-2，407mg，产率52％)。
1H NMR(400MHz,DMSO-d6)δ11.81(s,1H),7.93(d,J＝1.4Hz,1H),7.60(s,1H),7.40(d,J＝1.4Hz,1H)；
LC-MS：m/z 262.2[M+H] +。
步骤三：向干燥的50mL单口烧瓶中依次加入8-碘咪唑并[1,2-c]嘧啶-5-醇(B3-2,522mg,2mmol)和三氯氧磷(8mL)，在氮气的保护下，缓慢滴加N,N-二异丙基乙胺(1mL)，之后将混合物加热至120℃搅拌5小时。反应完毕后，反应液冷却至室温并将真空浓缩，然后加入饱和碳酸氢钠溶液猝灭，使用乙酸乙酯(3×40mL)萃取，经无水硫酸钠干燥，过滤并浓缩，将得到的残留物通过硅胶色谱法纯化(0至30％梯度的乙酸乙酯：石油醚)得到淡黄色固体5-氯-8-碘咪唑并[1,2-c]嘧啶(B3,360mg，收率：55％)。
1H NMR(400MHz,DMSO-d6)δ8.24(s,1H),8.20(d,J＝1.4Hz,1H),7.81(d,J＝1.4Hz,1H)；
LC-MS：m/z 280.1[M+H] +。
中间体C1的合成：2-氨基-3氯吡啶-4硫钠
Figure PCTCN2019115754-appb-000027
步骤一：向干燥的100mL的圆底三口烧瓶中依次加入3-氯-4-碘吡啶-2-胺(2.5g,9.82mmol,1.0eq),XantPhos(341mg,0.59mmol.0.06eq),醋酸钯(110mg,0.49mmol,0.05eq),DIPEA(3.25mL,19.6mmol,2.0q),3-巯基丙酸甲酯(1.19mL,10.8mmol,1.1eq)和1,4-二氧六环(32.5mL)。在搅拌的情况下，用氮气置换三次，然后加热到100℃,反应3小时。反应完毕后，反应液冷却至室温,用乙酸乙酯(50mL)稀释并减压抽滤，滤饼用乙酸乙酯(25mL)洗涤,将得到的滤液真空浓缩，将得到的残留物通过硅胶色谱法纯化(0至30％梯度的乙酸乙酯：石油醚)得到黄色固体3-((2-氨基-3-氯吡啶-4-基)硫代)丙酸甲酯(C1-1，2.0g，收率：78％)。
1H NMR(400MHz,CDCl 3)δ7.89(d,J＝5.4Hz,1H),6.53(d,J＝5.5Hz,1H),4.87(s,2H),3.74(s,3H),3.24(t,J＝7.5Hz,2H),2.75(t,J＝7.5Hz,2H)。
步骤二：在干燥的100mL的圆底三口烧瓶中，将化合物C1-1(2g,8.11mmol,1.0eq)溶解在四氢呋喃(28mL)里,氮气保护的情况下，室温下向反应液中滴加乙醇钠(2.9g,8.51mmol,1.05eq,20％wt),然后搅拌一小时。反应完毕后，用二氯甲烷(60mL)稀释并超声5分钟，减压抽滤，滤饼真空烘干得到黄色固体2-氨基-3氯吡啶-4硫钠(C1，1.4g，收率：89％)
中间体C2的合成：2，3-二氯吡啶-4-硫钠
Figure PCTCN2019115754-appb-000028
步骤一：将2,3-二氯-4-碘吡啶(1.0g，3.65mmol，1.0eq),3-巯基丙酸甲酯(480mg，4.02mmol，1.1eq)和N,N-二异丙基乙胺(950mg，7.3mmol)溶于1，4-二氧六环(15mL)中，置换三次氩气，在氩气保护下加入醋酸钯(82mg，0.37mmol，0.1eq)和XantPhos(211mg，0.37mmol，0.1eq)加完升温到100度，反应3小时，反应完毕。加入乙酸乙酯萃取(100mL)，通过硅藻土过滤并浓缩高效液相色谱制备得到类白色固体3-((2,3-二氯吡啶-4-基)硫代)丙酸甲酯(C2-1，550mg，收率：56.5％)。LCMS：m/z 266.0[M+H] +
步骤二：将3-((2,3-二氯吡啶-4-基)硫代)丙酸甲酯(C2-1，100mg,0.37mmol,1.0eq)溶于四氢呋喃(10mL)和乙醇(0.5mL)中。氮气保护下加入乙醇钠(27mg，0.39mmol,1.05eq)，室温反应3小时，反应未完成，补加乙醇钠(27mg，0.39mmol,1.05eq)，室温反应3小时，反应完毕。加入纯净水冷冻干燥得到粗品浅黄色固体2，3-二氯吡啶-4-硫钠(C2，150mg，收率：100％)。LCMS：m/z 180.0[M+H] +
中间体C3的合成：2-(三氟甲基)吡啶-3-硫钠
Figure PCTCN2019115754-appb-000029
步骤一：将3-溴-2-三氟甲基吡啶(400mg，1.77mmol，1.0eq),3-巯基丙酸甲酯(235mg，1.95mmol，1.1eq)和N,N-二异丙基乙胺(460mg，3.54mmol，2.0eq)溶于1，4-二氧六环(15mL)中，置换三次氩气，在氩气保护下加入Pd 2(dba) 3(160mg，0.18mmol，0.1eq)和XantPhos(205mg，0.36mmol，0.2eq)加完升温到110度，反应1小时，反应完毕。加入乙酸乙酯萃取(100mL)，通过硅藻土过滤并浓缩过柱(PE:EA＝8/1～6/1)得到浅黄色油状物3-((2-(三氟甲基)吡啶-3-基)硫代)丙酸甲酯(C3-1,450mg，收率：95.7％)。LCMS：m/z 266.1[M+H] +
步骤二：将3-((2-(三氟甲基)吡啶-3-基)硫代)丙酸甲酯(C3-1，300mg,1.13mmol,1.0eq)溶于四氢呋喃(10mL)和乙醇(0.5mL)中。氮气保护下加入乙醇钠(84mg，1.23mmol,1.1eq)，室温反应3小时，反应未完成，补加乙醇钠(83mg，1.23mmol,1.1eq)，室温反应3小时，反应完毕。室温减压除掉四氢呋喃后加入纯净水冷冻干燥得到粗品浅黄色固体2-(三氟甲基)吡啶-3-硫钠(C3，500mg，收率：超过100％)。LCMS：m/z 179.9[M+H] +
中间体C4的合成：2-(三氟甲基)吡啶-4-硫钠
Figure PCTCN2019115754-appb-000030
步骤一：将4-溴-2-三氟甲基吡啶(1.0g，4.4mmol，1.0eq),3-巯基丙酸甲酯(760mg，6.3mmol，1.4eq)和N,N-二异丙基乙胺(2.17g，16.8mmol，3.8eq)溶于1，4-二氧六环(25mL)中，置换三次氩气，在氩气保护下加入Pd 2(dba) 3(200mg，0.22mmol，0.05eq)和XantPhos(124mg，0.22mmol，0.05eq)加完升温到110度，反应1小时，反应完毕。加入乙酸乙酯萃取(100mL)，通过硅藻土过滤并浓缩过柱(PE:EA＝5/1)得到浅黄色油状物3-((2-(三氟甲基)吡啶-4-基)硫代)丙酸甲酯(C4-1,1.08g，收率：97％)。LCMS：m/z 266.2[M+H] +
步骤二：将3-((2-(三氟甲基)吡啶-4-基)硫代)丙酸甲酯(C4-1，230mg,0.85mmol, 1.0eq)溶于四氢呋喃(10mL)和乙醇(0.5mL)中。氮气保护下加入乙醇钠(294mg，0.87mmol,1.02eq)，室温反应1小时，反应完毕。室温减压除掉四氢呋喃后加入纯净水冷冻干燥得到粗品浅黄色固体2-(三氟甲基)吡啶-4-硫钠(C4,150mg),LCMS：m/z 180.0[M+H] +
中间体C5的合成：2-(三氟甲基)吡啶-4-硫钠
Figure PCTCN2019115754-appb-000031
步骤一：将2-氟-4-碘吡啶(2.0g，8.97mmol)溶解在THF(30mL)中。真空氮气置换3次后降温至-65℃，滴加LDA(2.0M in THF，5.4mL，10.80mmol)，反应液逐渐变成棕色。滴毕，保温1.5小时。滴加1,3，2-二噁唑噻吩-2，2-二氧化物(1.45g，11.7mmol)的THF(30mL)溶液。滴毕，自然升温至室温，搅拌过夜。TLC(石油醚/乙酸乙酯＝1/1)确认反应原料消失。重新降温到0度，滴加浓盐酸(4.48mL，40.2mmol)。滴毕，升温至室温，搅拌3小时。反应液倒至饱和NaHCO 3水溶液(50mL)中，乙酸乙酯萃取(3×30mL)，饱和盐水依次洗(50mL)，无水硫酸钠干燥，过滤，浓缩干后硅胶柱层析得2-(2-氟-4-碘吡啶-3-基)乙烷-1-醇(C5-1,2.07g,收率：86.2％)
步骤二：向2-(2-氟-4-碘吡啶-3-基)乙烷-1-醇(C5-1,2.07g,8.97mmol)的二氧六环(60mL)溶液中加入碳酸钾(4.28g，30.9mmol)。氮气保护下115℃搅拌反应48小时，LCMS和TLC(石油醚/乙酸乙酯＝1/1)显示基本为原料，仅少量产物。加入碳酸铯(7.56g，23.2mmol)，继续115℃反应6小时，LCMS和TLC(石油醚/乙酸乙酯＝1/1)显示原料基本反应完全。反应液降至室温，乙酸乙酯稀释，硅藻土过滤，乙酸乙酯淋洗。滤液用饱和盐水洗(2×50mL)，无水硫酸钠干燥，过滤，浓缩干后硅胶柱层析乙酸乙酯/石油醚/二氯甲烷＝0/5/2to 1/5/2，得白色固体4-碘-2,3-二氢呋喃并[2,3-b]吡啶(C5-2，1.21g，收率：63.2％)。LCMS:m/z 248.2[M+H] +
步骤三：向4-碘-2,3-二氢呋喃并[2,3-b]吡啶(C5-2，600mg，2.43mmol)的二氧六环(10mL)溶液中加入3-巯基丙酸甲酯(437mg，3.64mmol)，DIPEA(1.26g，9.71mmol)，XantPhos(70mg，0.12mmol)和Pd(OAc) 2(30mg，0.13mmol)。氮气保护下升温至100℃反应3小时，TLC(石油醚/乙酸乙酯＝1/1)和LCMS显示反应完全。反应液加乙酸乙酯稀释，过滤，滤液浓缩干，硅胶柱层析纯化得3-((2,3-二氢呋喃并[2,3-b]吡啶-4-基)硫代)丙酸甲酯(C5-3，565mg，收率：97.2％)。LCMS:m/z 240.3[M+H] +
步骤四：0℃下向3-((2,3-二氢呋喃并[2,3-b]吡啶-4-基)硫代)丙酸甲酯(C5-3，145mg，0.61mmol)的THF(5mL)溶液中加入乙醇钠乙醇溶液(20％(w/w)，185mg，0.54mmol)。室温反应2小时，TLC(乙酸乙酯/石油醚＝1/1)和LCMS显示反应完全。反应液不进行任何处 理，直接用于下一步反应。
中间体C6的合成：2-甲氨基-3氯吡啶-4硫钠
Figure PCTCN2019115754-appb-000032
步骤一：2,3-二氯-4-碘吡啶(1.0g,3.65mmol)与甲胺/乙醇溶液(27％，25mL)在闷罐中100℃反应12h。TLC监测反应完全(石油醚：乙酸乙酯＝5:1)。反应液减压浓缩，通过硅胶色谱法纯化(0至5％梯度的乙酸乙酯：石油醚)纯化得3-氯-4-碘-N-甲基吡啶-2-胺(C6-1,400mg，收率：41％)。LCMS:m/z 268.9[M+H] +
步骤二：向3-氯-4-碘-N-甲基吡啶-2-胺(C6-1，400mg，1.49mmol)的无水1,4-二氧六环(20mL)溶液中加入Xantphos(72mg，0.15mmol)，Pd(OAc) 2(34mg，0.15mmol)，DIPEA(770mg，5.96mmol)和3-巯基丙酸甲酯(270mg，2.23mmol)。100℃下反应5h。TLC(石油醚：乙酸乙酯＝5/1)检测反应完毕，反应降至室温后加水(50mL)和乙酸乙酯(30mL)，分液，水相乙酸乙酯再萃取(2×40mL)，合并有机相饱和盐水洗(3×50mL)，无水硫酸钠干燥，过滤浓缩，通过硅胶色谱法纯化(石油醚：乙酸乙酯＝10:1)纯化得3-((3-氯-2-(甲基氨基)吡啶-4-基)硫代)丙酸甲酯(C6-2，250mg，收率：65％)。LCMS:m/z 261.0[M+H] +
步骤三：将3-((3-氯-2-(甲基氨基)吡啶-4-基)硫代)丙酸甲酯(C6-2，114mg，0.44mmol)溶解到dioxane(6mL)中，氩气保护，滴加EtONa(20％(w/w)，150mg，0.44mmol)的乙醇溶液，室温搅拌反应约2.5h。LCMS监测反应完全。反应液直接用于下一步反应(收率：100％计)。LCMS:m/z 174.8[M+H-23] +
中间体C7的合成：1-(四氢-2H-吡喃-2-基)-1H-吡唑并[3,4-b]吡啶-4-硫醇钠
Figure PCTCN2019115754-appb-000033
步骤一：向2-氟-3-醛基-4-碘吡啶(1900mg，7.57mmol)的异丙醇(30mL)溶液中加入水合肼(3032mg,60.56mmol)，60℃搅拌3h。反应液减压浓缩除去部分溶剂后倒入水中，过滤，滤饼水洗得淡黄色固体4-碘-1H-吡唑并[3,4-b]吡啶(C7-1,1.8g，收率：97％)。 1H NMR(400MHz,CDCl 3):δ8.19(d,J＝4.4Hz,1H),7.98(s,1H),7.68(d,J＝4.8Hz,1H).
步骤二：向4-碘-1H-吡唑并[3,4-b]吡啶(C7-1,400mg，1.63mmol)的四氢呋喃(10mL)溶液中依次加入对甲苯磺酸(28mg，0.16mmol)和DHP(206mg,2.45mmol)。60℃搅拌16h。反应液加乙酸乙酯(40mL)稀释，饱和盐水洗(2×40mL)，无水硫酸钠干燥，过滤，滤液减压浓缩，将得到的残留物通过硅胶色谱法(0至50％梯度的乙酸乙酯：石油醚)纯 化得到淡黄色固体4-碘-1-(四氢-2H-吡喃-2-基)-1H-吡唑并[3,4-B]吡啶(C7-2,390mg，收率：73％)。 1H NMR(400MHz,CDCl 3):δ8.17(d,J＝4.8Hz,1H),7.93(s,1H),7.58(d,J＝5.2Hz,1H),6.10(dd,J＝10.4,2.4Hz,1H),4.14-4.10(m,1H),3.82(td,J＝11.6,2.8Hz,1H),2.69 2.59(m,1H),2.17-2.13(m,1H),2.00-1.96(m,1H),1.83-1.74(m,2H),1.64-1.62(m,1H).
步骤三：将4-碘-1-(四氢-2H-吡喃-2-基)-1H-吡唑并[3,4-B]吡啶(C7-2,390mg，1.18mmol),3-巯基丙酸甲酯(157mg，1.30mmol)和N,N-二异丙基乙胺(306mg，2.37mmol)溶于1，4-二氧六环(10mL)中，置换三次氩气，在氩气保护下加入Pd(OAc) 2(27mg，0.12mmol)和Xantphos(137mg，0.24mmol)加完升温到110℃，反应2h，反应完毕。加入乙酸乙酯萃取(30mL)，饱和盐水洗(2×30mL)。无水硫酸钠干燥，过滤，滤液减压浓缩，将得到的残留物通过硅胶色谱法(0至50％梯度的乙酸乙酯：石油醚)纯化黄褐色油状物3-((1H-吡唑并[3,4-b]吡啶-4-基)硫代)丙酸甲酯(C7-3,280mg，收率：74％)。LCMS:m/z 322.3[M+H] +
1H NMR(400MHz,CDCl 3):δ8.40(d,J＝5.2Hz,1H),8.08(s,1H),6.95(d,J＝5.2Hz,1H),6.10(dd,J＝10.8,2.4Hz,1H),4.14～4.10(m,1H),3.83(td,J＝11.2Hz,1H),3.73(s,3H),3.40(t,J＝7.2Hz,2H),2.79(t,J＝7.2Hz,2H),2.68-2.58(m,1H),2.16-2.13(m,1H),2.00-1.55(m,2H),1.63-1.60(m,1H).
步骤四：在带冷凝管的三口瓶中，将3-((1H-吡唑并[3,4-b]吡啶-4-基)硫代)丙酸甲酯(C7-3,280mg,0.87mmol)用1,4-二氧六环(7mL)溶解。氮气保护，降温至0度，加入乙醇钠乙醇溶液(20％(w/w)，266mg，0.78mmol)。室温搅拌2h，TLC(二氯甲烷/甲醇＝20/1)，显示反应完全，所得的含1-(四氢-2H-吡喃-2-基)-1H-吡唑并[3,4-b]吡啶-4-硫醇钠(C7)的反应液直接用于下一步。
实施例一：化合物1的合成
(S)-1'-(8-((2,3-二氢呋喃[2,3-b]吡啶-4-基)硫代)咪唑[1,2-c]嘧啶-5-基)-5,7-二氢螺环戊二烯并[b]吡啶-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000034
步骤一：在氮气保护下，向干燥的25mL单口烧瓶中依次加入5-氯-8-碘咪唑并[1,2-c]嘧啶(B3,80mg,0.285mmol)、((R)-N-((S)-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺(A4,105mg,0.342mmol)、DIEA(55mg,0.428mmol)和CH 3CN(5mL)，然后在95℃下搅拌反应3小时。反应完毕后，将获得的反应液过滤，减压浓缩得到的残留物通过硅胶色谱法 纯化(0至10％梯度的甲醇/乙酸乙酯)，得到(1-1,101mg,收率：65％)。LCMS：m/z551.1[M+H] +。
步骤二：2,3-二氢呋喃[2,3-b]吡啶-4-硫代甲酸钠(107mg,0.61mmol)加入二氧六环(15mL)稀释，再加入(S)-N-((S)-1'-(8-碘代咪唑[1,2-C]嘧啶-5-基)-5,7-二氢螺环[环戊烷[B]吡啶-6,4'-哌啶]-5-基)-2-甲基丙烷-2-亚磺酰胺(1-1，200mg，0.36mmol)，DIPEA(141mg，1.09mmol)，XantPhos(63mg，0.11mmol)和Pd 2(dba) 3(50mg，0.06mmol)。氮气保护下升温至100℃搅拌反应3小时，TLC(石油醚/乙酸乙酯＝1/1)和LCMS显示反应完全。反应液加乙酸乙酯稀释，过滤，滤渣乙酸乙酯淋洗，滤液浓缩干，硅胶柱层析得(S)-N-((S)-1'-(8-((2,3-二氢呋喃[2,3-b]吡啶-4-基)硫代)咪唑[1,2-c]嘧啶-5-基)-5,7-二氢螺环戊二烯并[b]吡啶-6,4'-哌啶]-5-基)-2-甲基丙烷-2-磺胺(1-2，217mg，收率：98.9％)。LCMS:m/z 576.6[M+H] +
步骤三：氮气保护，0℃下向(S)-N-((S)-1'-(8-((2,3-二氢呋喃[2,3-b]吡啶-4-基)硫代)咪唑[1,2-c]嘧啶-5-基)-5,7-二氢螺环环戊二烯并[b]吡啶-6,4'-哌啶]-5-基)-2-甲基丙烷-2-磺胺(50mg，0.29mmol)的二氯甲烷(5mL)溶液中缓慢加入HCl/dioxane(4M，0.2mL，0.80mmol)，室温搅拌反应2小时，TLC(二氯甲烷/甲醇＝8/1)和LCMS显示反应完全。反应液降温至0℃，缓慢加入氨甲醇溶液调节pH至10左右后减压浓缩，prep-HPLC纯化得(S)-1'-(8-((2,3-二氢呋喃[2,3-b]吡啶-4-基)硫代)咪唑[1,2-c]嘧啶-5-基)-5,7-二氢螺环戊二烯[b]吡啶-6,4'-哌啶]-5-胺的甲酸盐(化合物1，8.86mg，收率：20.0％)。LCMS:m/z472.5[M+H] +
1H NMR(400MHz,MeOD)δ8.47(d,J＝4.0Hz,2H),8.09(s,1H),7.92(d,J＝7.6Hz,1H),7.85(d,J＝1.6Hz,1H),7.59(d,J＝1.6Hz,1H),7.55(d,J＝6.0Hz,1H),7.37～7.34(m,1H),6.21(d,J＝5.6Hz,1H),4.72～4.67(m,2H),4.38(s,1H),4.06～4.03(m,2H),3.48～3.42(m,2H),3.28(s,2H),3.14(d,J＝16.8Hz,2H),2.11～2.078(m,2H),1.76～1.70(m,2H).
实施例二：化合物2的合成
(S)-1’-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三氮唑[4,3-c]嘧啶-5-基)-1，3-二氢螺[茚-2,4’-哌啶]-1-胺
Figure PCTCN2019115754-appb-000035
步骤一：在氮气保护下，向干燥的25mL单口烧瓶中依次加入5-氯-8-碘-[1,2,4]三唑并[4,3-c]嘧啶(B2,80mg,0.285mmol)、((R)-N-((S)-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺(A1,105mg,0.342mmol)、DIEA(55mg,0.428mmol)和CH 3CN(5mL)，然后在95℃下搅拌反应3小时。反应完毕后，将获得的残留物过滤并在减压浓缩得到的残留物通过硅胶色谱法纯化(0至10％梯度的甲醇/乙酸乙酯)，得到黄色固体(R)-N-((S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2-,4'-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺(2-1,101mg,收率：65％)。LCMS：m/z 551.1[M+H] +。
步骤二：在氮气保下向5mL的微波反应瓶中依次加入(R)-N-((S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2-,4'-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺(2-1，100mg，0.18mmol)、2-氨基-3-氯吡啶-4-硫醇钠(49mg,0.27mmol)、Pd 2(dba) 3(16mg,0.018mmol)、Xantphos(21mg,0.036mmol)、DIPEA(58mg,0.45mmol)和1,4-二氧六环溶液(10mL),该混合物在氮气保护下微波加热100℃搅拌反应3小时。反应完毕后，冷却至室温,过滤并在减压浓缩得到的残留物通过硅胶色谱法纯化(0至10％梯度的甲醇/乙酸乙酯)，得到(R)-N-((R)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺(2-2，65mg，收率：63％)。LC-MS：m/z 584.2[M+H] +。
步骤三：在氮气保下向50mL的单口烧瓶中依次加入(R)-N-((R)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-甲基丙烷-2-亚磺酰胺(2-2，60mg,0.10mmol)和甲醇(0.6mL)，在室温下滴加盐酸1，4-二氧六环溶液(0.06mL,4M)，该混合物在室温下搅拌反应1小时。反应完毕后，冷却至室温,过滤并在减压浓缩得到的残留物通过高效液相制备色谱纯化得到(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1-1,3-二氢螺[茚-2,4'-哌啶]-1-胺(化合物2,20mg，收率：42％)。
1H NMR(400MHz,DMSO-d6)δ9.38(s,1H),7.98(s,1H),7.58(d,J＝5.4Hz,1H),7.36(d,J＝4.8Hz,1H),7.22(dt,J＝8.0,4.0Hz,3H),6.33(s,2H),5.96(d,J＝5.6Hz,1H),4.22–4.07(m,2H),4.00(s,1H),3.49(dd,J＝22.5,11.2Hz,2H),3.12(d,J＝15.6Hz,1H),2.75(d,J＝15.6Hz,1H),2.03–1.86(m,2H),1.63(d,J＝13.6Hz,1H),1.39–1.29(m,1H)；
LC-MS：m/z 479.1[M+H] +。
实施例三：化合物3的合成。
(S)-1’-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三氮唑[4,3-c]嘧啶-5-基)-5，7-二氢螺[环戊二烯并[b]吡啶-6,4’-哌啶]-5-胺
Figure PCTCN2019115754-appb-000036
参照化合物2的合成方法，使用中间体A2代替中间体A1，进行三步反应得到化合物3。
1H NMR(400MHz,DMSO-d6))δ9.40(s,1H),8.35(d,J＝4.3Hz,1H),7.99(s,1H),7.71(d,J＝7.4Hz,1H),7.58(d,J＝5.4Hz,1H),7.21(dd,J＝7.4,5.0Hz,1H),6.36(s,2H),5.96(d,J＝5.4Hz,1H),4.23–4.08(m,2H),4.02(s,1H),3.51(dd,J＝23.3,11.6Hz,2H),3.16(d,J＝16.3Hz,1H),2.84(d,J＝16.3Hz,1H),2.04–1.91(m,2H),1.65(d,J＝13.4Hz,1H),1.32(d,J＝14.8Hz,1H)；
LC-MS：m/z 480.1[M+H] +。
实施例四：化合物4的合成。
(S)-1’-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑[1,2-c]嘧啶-5-基)-1，3-二氢螺[茚-2,4’-哌啶]-1-胺
Figure PCTCN2019115754-appb-000037
参照化合物2的合成方法，使用中间体B3代替中间体B2，进行三步反应得到化合物4。(甲酸盐，白色固体)。
1H NMR(400MHz,DMSO-d6)δ8.03(s,1H),7.83(s,1H),7.64–7.49(m,2H),7.43(s,1H),7.30-7.17(m,3H),6.33(s,2H),5.79(d,J＝5.2Hz,1H),4.12(s,1H),3.95(d,J＝12.8Hz,2H),3.33(dd,J＝20.0,10.8Hz,2H),3.14(d,J＝15.6Hz,1H),2.82(d,J＝15.6Hz,1H),1.98(d,J＝9.6Hz,2H),1.61(d,J＝12.8Hz,1H),1.42(d,J＝12.8Hz,1H),1.23(s,2H)；
LC-MS：m/z 478.1[M+H] +。
实施例五：化合物5的合成。
(S)-1'-(8-(((3-氯-2-(甲基氨基)吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000038
步骤一：2-甲氨基-3氯吡啶-4硫钠(C6,86mg)溶于10mL二氧六环溶液,加入(R)-N-((S)-1'-(8-碘代咪唑[1,2-C]嘧啶-5-基)-5,7-二氢螺环[环戊二烯并[b]吡啶-6,4'-哌啶]-5-基)-2-甲基丙烷-2-亚磺酰胺(161mg，0.29mmol)，Pd 2(dba) 3(86mg，0.094mmol)，Xantphos(108mg，0.19mmol)和DIPEA(400mg，3.12mmol)。氩气保护下100℃反应5h。LCMS监测反应完全。降至室温后加二氯甲烷(30mL)，过滤，母液加水(40mL)，分液。水相用二氯甲烷萃取(2×30mL)，合并有机相，饱和盐水洗(3×40mL)，无水硫酸钠干燥，过滤，通过硅胶色谱法纯化(二氯甲烷：甲醇＝15:1)纯化得浅黄色固体(R)-N-((S)-1'-(8-((3-氯-2-(甲基氨基)吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-基)-2-甲基丙烷-2-亚磺酰胺(5-1,135mg，收率：73％)。LCMS:m/z 596.9[M+H] +。
步骤二：将(R)-N-((S)-1'-(8-((3-氯-2-(甲基氨基)吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊[b]吡啶-6,4'-哌啶]-5-基)-2-甲基丙烷-2-亚磺酰胺(135mg，0.23mmol)溶解于甲醇(20mL)中,氩气保护，缓慢滴加HCl/dioxane(4M，1.2mL)，室温反应约40分钟，LCMS监测反应完全。反应液倒入冰的饱和碳酸氢钠溶液中，混合溶液冻干，加入二氯甲烷/甲醇(10:1，20mL)，过滤，滤液浓缩至干后prep-HPLC制备纯化得白色固体(化合物5，33mg，收率：30％)。LCMS:m/z 493.0[M+H] +
1H NMR(400MHz,DMSO-d6)δ8.33(d,J＝4.4Hz,1H),8.03(s,1H),7.84(d,J＝1.2Hz,1H),7.69(d,J＝7.6Hz,1H),7.65(d,J＝5.2Hz,1H),7.57(d,J＝1.2Hz,1H),7.20(dd,J＝7.2,5.2Hz,1H),6.60(q,J＝4.4Hz,1H),5.81(d,J＝5.2Hz,1H),3.98-3.89(m,3H),3.38-3.22(m,2H),3.15-3.11(d,J＝16.0Hz,1H),2.85(d,J＝4.8Hz,3H),2.81(d,J＝16.4Hz,1H),2.50-2.39(m,1H),2.05-1.90(m,3H),1.67(d,J＝13.2Hz,1H),1.28～1.25(d,J＝13.2Hz,1H).
实施例六：化合物6的合成。
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000039
步骤一：向含3mL乙腈的25mL单口烧瓶中依次加入B3(1.37g,4.9mmol)、A4(1.35g,4.9mmol)、DIPEA(4.86mL,29.41mmol)，然后在80℃下搅拌反应2小时。反应完毕后冷却至室温，然后加入Boc 2O(1.6g,7.35mmol,1.5eq),升温至50℃反应，直至反应完全，将反应液减压浓缩得到的残留物通过硅胶色谱法纯化(0至100％梯度的乙酸乙酯/石油醚)，得到黄色固体6-1(1.7g,收率：63.4％)。LC-MS：m/z＝547.0[M+H +]
步骤二：在氮气保下向5mL的微波反应瓶中依次加入6-1(1.7g，3.11mmol)、2-氨基-3-氯吡啶-4-硫醇钠(596mg,3.27mmol)、Pd2(dba)3(285mg,0.311mmol)、Xantphos(360mg,0.622mmol)、DIPEA(804mg,6.22mmol)和1,4-二氧六环溶液(30mL),该混合物在氮气保护下微波加热100℃搅拌反应3小时。反应完毕后，冷却至室温,过滤并在减压浓缩得到的残留物通过硅胶色谱法纯化(0至10％梯度的乙酸乙酯/甲醇)，得到(S)-叔丁基(1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-基)-碳酸酯(6-2，1.2g，66.7％)
步骤三：氮气保护，0℃下向(S)-叔丁基(1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-基)-碳酸酯(6-2，1.2g，2.07mmol)的二氯甲烷(5mL)溶液中缓慢加入TFA(5mL)，室温搅拌反应1小时，TLC和LCMS显示反应完全。反应液减压浓缩，再加入二氯甲烷/甲醇混合溶液溶解，用NaHCO 3调节pH至中性。过硅胶柱纯化得(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-5-胺(化合物6，400mg，收率：40.0％)。
1H NMR(400MHz,DMSO-d6)δ8.35(d,J＝4.0Hz,1H),8.03(s,1H),7.83(d,J＝1.2Hz,1H),7.72(d,J＝7.6Hz,1H),7.56(dd,J＝10.4,3.4Hz,2H),7.20(dd,J＝7.6,5.2Hz,1H),6.33(s,2H),5.80(d,J＝5.4Hz,1H),4.02(s,1H),3.95(dd,J＝11.6,7.6Hz,2H),3.31(d,J＝13.6Hz,2H),3.15(d,J＝16.4Hz,1H),2.83(d,J＝16.4Hz,1H),2.00(tt,J＝12.4,6.4Hz,2H),1.64(d,J＝13.2Hz,1H),1.48(dd,J＝13.6,6.4Hz,1H),1.34-1.29(m,2H)；
LCMS：m/z 479.0[M+H] +。
实施例七：化合物7的合成。
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5-， 7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000040
参照化合物6的合成方法，使用中间体B2代替中间体B3，使用中间体A5代替中间体A4，进行三步反应得到化合物7。
1H NMR(400MHz,DMSO)δ8.51(s,2H),8.38(d,J＝4.8Hz,1H),8.22(s,1H),7.57(d,J＝5.6Hz,1H),7.28(d,J＝4.8Hz,1H),6.36(s,2H),5.88(d,J＝5.6Hz,1H),4.92(t,J＝13.6Hz,2H),4.03(s,1H),3.61(dd,J＝26.8,12.0Hz,2H),3.16(d,J＝16.8Hz,1H),2.78(s,1H),1.92(td,J＝11.2,4.0Hz,2H),1.63(d,J＝13.2Hz,1H),1.33–1.24(m,1H).
LCMS：m/z 480.1[M+H] +。
实施例八：化合物8的合成。
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000041
参照化合物6的合成方法，使用中间体A5代替中间体A4，进行三步反应得到化合物8。
1H NMR(400MHz,DMSO)δ8.53(s,1H),8.39(d,J＝4.8Hz,1H),8.04(s,1H),7.84(d,J＝1.2Hz,1H),7.58(d,J＝1.2Hz,1H),7.56(d,J＝5.6Hz,1H),7.28(d,J＝4.8Hz,1H),6.34(s,2H),5.80(d,J＝5.6Hz,1H),4.07(s,1H),3.93(d,J＝4.0Hz,2H),3.37(dd,J＝19.6,8.0Hz,2H),3.13(d,J＝16.4Hz,1H),2.76(d,J＝16.4Hz,1H),1.98(d,J＝11.6Hz,2H),1.62(d,J＝13.6Hz,1H),1.30(s,1H).
LCMS：m/z 479.1[M+H] +。
实施例九：化合物9的合成
(S)-1'-(8-(2,3-二氯吡啶-4-基)硫代-[1,2,4]-三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000042
步骤一：将(S)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺盐酸盐(A6,200mg,1.28mmol)溶于乙腈(20mL)中，加入DIPEA(1.6mL,9.68mmol)与5-氯-8-碘-[1,2,4]-三唑并[4,3-c]嘧啶(B2,180mg,0.64mmol)，氮气保护下加热至90℃回流反应3小时，反应完毕。冷却至室温，将反应液倒入饱和碳酸氢钠水溶液，用二氯甲烷萃取(80mL)两次。有机相合并后用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化(DCM:MeOH＝20/1)得到浅黄色固体化合物(S)-1'-(8-碘[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺(9-1,260mg，收率90％)。LCMS：m/z 448.2[M+H] +
步骤二：将(S)-1'-(8-碘[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺(21-1，55mg，0.12mmol)溶于1,4-二氧六环(3mL)中，加入DIPEA(35mg,0.27mmol)与2，3-二氯吡啶-4-硫醇钠(C2,100mg,粗品)。氮气保护下加入XantPhos(30mg,0.05mmol)，Pd 2(dba) 3(17mg,0.02mmol)。氮气置换三次后100℃反应3小时，反应完毕。反应液浓缩后用高效液相色谱纯化得到浅黄色固体化合物(S)-1'-(8-(2,3-二氯吡啶-4-基)硫代-[1,2,4]-三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺(化合物9,27mg，TFA盐，收率：40％)。LCMS：m/z 499.1[M+H] +
1H NMR(400MHz,MeOD)δ9.35(s,1H),8.56(d,J＝4.8Hz,1H),8.14(s,1H),7.95(d,J＝5.2Hz,1H),7.84(d,J＝7.6Hz,1H),7.44-7.41(m,1H),6.80(d,J＝5.2Hz,1H),4.53(s,1H),4.36–4.32(m,2H),3.69-3.60(m,2H),3.40(d,J＝16.4Hz,1H),3.19(d,J＝16.4Hz,1H),2.26-2.19(m,1H),2.05-1.93(m,2H),1.73-1.70(m,1H).
实施例十：化合物10的合成。
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000043
参照化合物6的合成方法，使用中间体A6代替中间体A4，进行三步反应得到化合物10。
1H NMR(400MHz,DMSO)δ8.38(d,J＝4.3Hz,1H),8.24(s,1H),8.03(s,1H),7.84(d,J＝1.4Hz,1H),7.65(d,J＝7.4Hz,1H),7.56(dd,J＝9.1,3.4Hz,2H),7.29–7.13(m,1H),6.33(s,2H),5.80(d,J＝5.4Hz,1H),4.01(s,1H),3.95–3.86(m,2H),3.38(dd,J＝23.6,11.2Hz,3H),3.11(d,J＝16.0Hz,1H),2.75(d,J＝16.0Hz,1H),2.05–1.91(m,2H),1.69(d,J＝13.6Hz,1H),1.31(d,J＝14.0Hz,2H)。
LC-MS：m/z＝478.0[M+H +]。
实施例十一：化合物11的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5-，7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000044
参照化合物6的合成方法，使用中间体B2代替中间体B3，使用中间体A6代替中间体A4进行三步反应得到化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5-，7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺(11,甲酸盐，白色固体)。
1H NMR(400MHz,DMSO-d6)δ9.40(s,1H),8.48(d,J＝4.5Hz,1H),8.00(s,1H),7.76(d,J＝7.3Hz,1H),7.58(d,J＝5.4Hz,1H),7.33(dd,J＝7.5,5.0Hz,1H),6.35(s,2H),5.94(d,J＝5.4Hz,1H),4.31(s,1H),4.26–4.09(m,2H),3.53(dd,J＝27.1,12.0Hz,2H),3.24(d,J＝ 16.4Hz,1H),2.92(d,J＝16.3Hz,1H),2.09(s,1H),1.92(s,1H),1.76(d,J＝13.2Hz,1H),1.42(d,J＝13.2Hz,1H).
LCMS：m/z＝479.0[M+H +]。
实施例十二：化合物12的合成。
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-6-氟-1,3-二氢螺[茚-2,4'-哌啶]-1-胺
Figure PCTCN2019115754-appb-000045
参照化合物6的合成方法，使用使用中间体B2代替中间体B3,中间体A7代替中间体A4，进行三步反应得到化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-6-氟-1,3-二氢螺[茚-2,4'-哌啶]-1-胺(12,甲酸盐，白色固体)。
1H NMR(400MHz,DMSO-d6)δ9.38(s,1H),8.22(s,1H),7.98(s,1H),7.57(d,J＝5.4Hz,1H),7.31–7.20(m,1H),7.16(d,J＝7.5Hz,1H),7.01(t,J＝8.7Hz,1H),6.34(s,2H),5.95(d,J＝5.4Hz,1H),4.14(t,J＝12.0Hz,2H),4.00(s,1H),3.47(dd,J＝25.4,12.0Hz,3H),3.10(d,J＝15.5Hz,1H),2.71(d,J＝15.5Hz,1H),2.08–1.85(m,2H),1.64(d,J＝13.3Hz,1H),1.30(d,J＝13.5Hz,1H).LCMS：m/z＝495.0[M+H +]。
实施例十三：化合物13的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-6-氟-1,3-二氢螺[茚-2,4'-哌啶]-1-胺
Figure PCTCN2019115754-appb-000046
参照化合物6的合成方法，使用中间体A7代替中间体A4，进行两步反应得到化合物(S)-(1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5-氟-1,3-二氢螺[茚-2,4'-哌啶]-3-基)甲酸叔丁酯(13-2)。
步骤三：向干燥的单口烧瓶中依次加入化合物(S)-(1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5-氟-1,3-二氢螺[茚-2,4'-哌啶]-3-基)甲酸叔丁酯(13-2，60mg,0.101mmol)，二氯甲烷(1mL)和三氟乙酸(0.2mL)。反应液在20℃下搅拌1小时。反应液减压浓缩。得到的残留物反应液用饱和碳酸氢钠水溶液调至pH＝8，用DCM/MeOH(10:1)的混合溶剂(10mL)萃取3次，合并的有机相减压浓缩。得到的残留物通过HPLC制备纯化得到白色固体化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-6-氟-1,3-二氢螺[茚-2,4'-哌啶]-1-胺(13，32mg，甲酸盐，收率：58.2％)。
1H NMR(400MHz,DMSO-d6)δ8.15(s,1H),8.03(s,1H),7.83(d,J＝1.2Hz,1H),7.58(d,J＝1.6Hz,1H),7.55(d,J＝5.6Hz,1H),7.32–7.26(m,1H),7.23–7.18(m,1H),7.12–7.02(m,1H),6.32(s,2H),5.80(d,J＝5.2Hz,1H),4.14(s,1H),4.00–3.90(m,2H),3.40–3.30(m,2H),3.15–3.05(m,1H),2.85–2.75(m,1H),2.05–1.85(m,2H),1.70–1.55(m,1H),1.45–1.36(m,1H)；LC-MS：m/z 496.0[M+H]+。
实施例十四：化合物14的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000047
步骤一：将(S)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-5-胺盐酸盐(A8,200mg,0.83mmol)溶于乙腈(20mL)中，加入DIPEA(1.07g,8.3mmol)与5-氯-8-碘咪唑并[1,2-c]嘧啶(B3,208mg,0.75mmol)，氮气保护下加热至90℃回流反应5小时，反应完毕。冷却至室温，将反应液倒入饱和碳酸氢钠水溶液，用二氯甲烷萃取(50mL)两次。有机相合并后用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到浅黄色固体化合物1-(8-碘咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-4-胺(14-1,250mg，收率74％)。LCMS：m/z 447.1[M+H] +
步骤九：将1'-(8-碘咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-4-胺(14-1,50mg，0.11mmol)溶于1,4-二氧六环(10mL)中，加入DIPEA(36mg,0.28mmol)与2-氨基-3-氯吡啶-4-硫醇钠(C1,31mg,0.17mmol)。氮气保护下加入XantPhos(13mg,0.02mmol)，Pd 2(dba) 3(10mg,0.01mmol)。氮气置换三次后100℃反应3小时，反应完毕。反应液浓缩后用高效液相色谱纯化得到浅黄色固体化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-5-胺(14，12mg，收率：22.4％)。
LCMS：m/z 479.2[M+H] +
1H NMR(400MHz,MeOD)δ8.64(s,1H),8.56(d,J＝4.4Hz,1H),8.09(s,1H),7.62-7.60(m,2H),7.60-7.50(m,1H),5.95-5.94(m,1H),4.64(s,1H),4.22-4.07(m,2H),3.60(s,2H),3.6-3.49(m,2H),2.20-2.10(m,2H),2.10-2.00(m,1H),1.85-1.63(m,2H).
实施例十五：化合物15的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5-，7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000048
参照化合物14的合成方法，使用中间体B2代替中间体B3，进行两步反应得到化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5-，7-二氢螺[环戊二烯并[c]吡啶-6,4'-哌啶]-5-胺(15)。
LCMS：m/z 480.0[M+H] +
1H NMR(400MHz,MeOD)δ9.31(s,1H),8.43-8.46(m,2H),8.02(s,1H),7.49-7.54(m,2H),6.05(d,J＝5.6MHz,1H),4.17-4.29(m,3H),3.48-3.60(m,3H),2.96(d,J＝16.0MHz,1H),1.98-2.15(m,2H),1.76-1.79(m,1H),1.22-1.25(m,1H).
实施例十六：化合物16的合成
(S)-1-氨基-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'-哌啶]-4-醇
Figure PCTCN2019115754-appb-000049
步骤一：将(S)-1-氨基-1,3-二氢螺[茚-2,4'-哌啶]-4-醇盐酸盐(A9,200mg,0.79mmol)溶于乙腈(40mL)中，加入DIPEA(1.07g,8.3mmol)与5-氯-8-碘-[1,2,4]三唑并[4,3-c]嘧啶(B2,208mg,0.75mmol)，氮气保护下室温反应18小时，反应完毕。冷却至室温，将反应液倒入饱和碳酸氢钠水溶液，用二氯甲烷萃取(100mL)两次。有机相合并后用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到浅黄色固体化合物(S)-1-氨基-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'--哌啶〕-4-醇(16-1,220mg，收率63％)。LCMS：m/z 463.0[M+H] +
步骤二：将(S)-1-氨基-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'--哌啶〕-4-醇(16-1，200mg，0.43mmol)溶于1,4-二氧六环(30mL)中，加入DIPEA(111mg,0.86mmol)与2-氨基-3-氯吡啶-4-硫醇钠(C1,118mg,0.65mmol)。氮气保护下加入XantPhos(52.0mg,0.08mmol)，Pd 2(dba) 3(41mg,0.04mmol)。氮气置换三次后100℃反应3小时， 反应完毕。反应液浓缩后用高效液相色谱纯化得到白色固体化合物(S)-1-氨基-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'-哌啶]-4-醇(16,100mg，收率：47％)。LCMS：m/z 495.3[M+H] +
1H NMR(400MHz,MeOD)δ9.32(s,1H),8.01(s,1H),7.53(d,J＝5.6MHz，1H),7.04(t,J＝7.6MHz，1H),6.84(d,J＝5.6MHz，1H),6.64(d,J＝8.0MHz，1H),6.06(d,J＝5.6MHz，1H),4.20-4.24(m,2H),3.95(s,1H),3.54-3.62(m,2H),3.13(d,J＝15.6MHz，1H),2.74(d,J＝15.6MHz，1H),1.93-2.06(m,2H)，1.70(d,J＝12.8MHz，1H),1.55(d,J＝12.8MHz，1H).
实施例十七：化合物17的合成
化合物(S)-1-氨基-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'-哌啶]-4-腈
Figure PCTCN2019115754-appb-000050
步骤一：将(S)-1-(((R)-叔丁基亚磺酰)氨基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶](A10，490mg,1.48mmol)溶于乙腈(20mL)中，加入DIPEA(1.91g,14.8mmol)与5-氯-8-碘-[1,2,4]三唑并[4,3-c]嘧啶(B2，415mg,1.48mmol)，氮气保护下于85℃反应3小时，反应完毕。减压浓缩得到的残留物通过硅胶色谱法纯化(0至10％梯度的甲醇/乙酸乙酯)，得到黄色固体化合物(R)-N-((S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(17-1，360mg,收率：42.3％)。LCMS：m/z 576.1[M+H] +。
步骤二：将(R)-N-((S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(17-1，360mg，0.63mmol)溶于1,4-二氧六环(10mL)中，加入DIPEA(202mg,1.56mmol)与2-氨基-3-氯吡啶-4-硫醇钠(C1，171mg,0.94mmol)。氮气保护下加入XantPhos(73mg,0.126mmol)，Pd 2(dba) 3(57.7mg,0.063mmol)。氮气置换三次后100℃反应3小时，反应完毕。反应液浓缩后用硅胶色谱法纯化得到白色固体化合物(R)-N-((S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(17-2，130mg,收率：34.2％)。LC-MS：m/z 608.2[M+H] +。
步骤三：在氮气保下向50mL的单口烧瓶中依次加入(R)-N-((S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-氰基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(17-2，130mg,0.21mmol)和甲醇(6mL)，在室温下滴加盐酸1，4-二氧六环 溶液(2mL,4M)，该混合物在室温下搅拌反应1小时。反应完毕后，冷却至室温,过滤并减压浓缩，得到的残留物通过高效液相制备色谱纯化得到化合物(S)-1-氨基-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-1,3-二氢螺[茚-2,4'-哌啶]-4-腈(17,10mg，收率：9.3％)。
1H NMR(400MHz,CDCl 3)δ9.32(s,1H),8.02(s,1H),7.71(d,J＝7.2Hz,1H),7.59(d,J＝7.6Hz,1H),7.53(d,J＝5.6Hz,1H),7.43(d,J＝7.6Hz,1H),6.06(d,J＝5.6Hz,1H),4.23(d,J＝10.8Hz,2H),4.12(s,1H),3.60(m,2H),3.37(m,1H),3.03(d,J＝16.4Hz,1H),2.06(m,2H),1.74(t,2H),1.57(d,J＝19.2Hz,2H),1.52(s,2H)。
LCMS：m/z 504.1[M+H] +。
实施例十八：化合物18的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000051
步骤一：将(S)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺盐酸盐(A11，1.50g,4.95mmol)溶于150mL乙腈中，加入DIPEA(5.68g,44.0mmol)与5-氯-8-碘咪唑并[1,2-c]嘧啶(B3，1.28g,4.59mmol)，氮气保护下加热至95℃回流反应3小时，反应完毕。降至室温，过滤，滤液浓缩干，拌硅胶过柱，得米黄色粉末化合物(S)-1'-(8-碘咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺(18-1，2.05g，收率：92.7％)。LCMS：m/z 448.0[M+H] +
步骤二：在350mL玻璃封管中，将(S)-1'-(8-碘咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺(18-1，2.05g，4.59mmol)溶于120mL 1,4-二氧六环，依次加入DIPEA(1.78g,13.8mmol)，2-氨基-3-氯吡啶-4-硫醇钠(C1,1.26g,6.9mmol)，XantPhos(797mg,1.38mmol)和Pd 2(dba) 3(630mg,0.69mmol),氩气鼓泡30秒，氩气氛围下加热至100℃，保温3小时，反应完全。降至室温，加入二氯甲烷稀释，抽滤，滤液旋干，过柱，所得纯点旋干后，用二氯甲烷/正己烷(1/1)打浆过夜，过滤，干燥得化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺(化合物18，440mg,收率：19.9％)。LCMS：m/z 480.1[M+H] +
1H NMR(400MHz,MeOD):δ8.44-8.39(m,2H),8.05(s,1H),7.85(d,J＝1.6Hz,1H),7.56(d,J＝1.6Hz,1H),7.49(d,J＝5.6Hz,1H),5.89(d,J＝5.6Hz,1H),4.14(s,1H),4.06-4.02(m,2H),3.49-3.40(m,2H),3.33(s,1H),3.01(d,J＝16.8Hz,1H),2.24-2.10(m,2H),1.82(d,J＝13.2Hz,1H),1.50(d,J＝13.6Hz,1H).
实施例十九：化合物19的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1-胺
Figure PCTCN2019115754-appb-000052
步骤七：将(S)-1-(((R)-叔丁基亚磺酰)氨基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶](A12，520mg,1.55mmol)溶于乙腈(20mL)中，加入DIPEA(2.0g,15.5mmol)与5-氯-8-碘-[1,2,4]三唑并[4,3-c]嘧啶(B2，435mg,1.55mmol)，氮气保护下于85℃反应3小时，反应完毕。减压浓缩得到的残留物通过硅胶色谱法纯化(0至10％梯度的甲醇/乙酸乙酯)，得到黄色固体化合物(R)-N-((S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(19-1，420mg,收率：46.8％)。
LCMS：m/z 581.1[M+H] +。
步骤八：将(R)-N-((S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(19-1，420mg，0.72mmol)溶于1,4-二氧六环(10mL)中，加入DIPEA(234mg,1.81mmol)与2-氨基-3-氯吡啶-4-硫醇钠(C1，198mg,1.09mmol)。氮气保护下加入XantPhos(83mg,0.144mmol)，Pd 2(dba) 3(66mg,0.072mmol)。氮气置换三次后100℃反应3小时，反应完毕。反应液浓缩后用硅胶色谱法纯化得到白色固体化合物(R)-N-((S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(19-2，310mg,收率：69.9％)。
LCMS：m/z 613.2[M+H] +。
步骤九：在氮气保下向50mL的单口烧瓶中依次加入(R)-N-((S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1-基)-2-叔丁基磺酰胺(19-2，310mg,0.51mmol)和甲醇(6mL)，在室温下滴加盐酸1，4-二氧六环溶液(2mL,4M)，该混合物在室温下搅拌反应1小时。反应完毕后，冷却至室温,过滤并减压浓缩，得到的残留物通过高效液相制备色谱纯化得到化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-4-甲氧基-1,3-二氢螺[茚-2,4'-哌啶]-1-胺(化合物19，114mg，收率：44.3％)。
1H NMR(400MHz,CDCl 3)δ8.80(s,1H),7.95(s,1H),7.65(d,J＝5.6Hz,1H),7.26(t,2H),6.96(d,J＝7.6Hz,1H),6.77(d,J＝8.4Hz,1H),6.02(d,J＝5.6Hz,1H),4.89(s,2H),4.13-4.04(m,4H),3.86(s,3H),3.53-3.48(m,2H),3.11(d,J＝16Hz,1H),2.70(d,J＝16Hz,1H),2.08-2.01(m,3H),1.76(d,J＝14Hz,1H),1.53(d,J＝14Hz,2H)。
LCMS：m/z 509.2[M+H] +。
实施例二十：化合物20的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5-，7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺
Figure PCTCN2019115754-appb-000053
步骤一：将(S)-5,7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺盐酸盐(A11，89mg,0.29mmol)溶于20mL乙腈中，加入DIPEA(212mg,1.64mmol)与5-氯-8-碘-[1,2,4]三唑并[4,3-c]嘧啶(B2，77mg,0.275mmol)，氮气保护下加热至95℃回流反应3小时，反应完毕。降至室温，过滤，滤液浓缩干，拌硅胶过柱，得(S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡嗪6,4'-哌啶]-5-胺(20-1，65mg，收率：52.7％)。LCMS：m/z 449.2[M+H] +
步骤二：在5mL微波管中，将(S)-1'-(8-碘-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡嗪6,4'-哌啶]-5-胺(20-1，65mg，0.145mmol)溶于5mL 1,4-二氧六环，依次加入DIPEA(56mg,0.434mmol)，2-氨基-3-氯吡啶-4-硫醇钠(C1,40mg,0.219mmol)，XantPhos(34mg,0.059mmol)和Pd 2(dba) 3(27mg,0.029mmol),氩气鼓泡30秒，氩气氛围下加热至100℃，保温3小时，反应完全。降至室温，加入二氯甲烷稀释，抽滤，滤液旋干，用制备硅胶板纯化，所得纯点送制备，得纯品(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫基)-[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5-，7-二氢螺[环戊二烯并[b]吡嗪-6,4'-哌啶]-5-胺(化合物20，19.4mg,收率：27.8％)。LCMS：m/z 481.2[M+H] +
1H NMR(400MHz,MeOD)δ9.35(s,1H),8.59-8.57(m,2H),8.11(s,1H),7.53(d,J＝6.8Hz,1H),6.37(d,J＝6.4Hz,1H),4.65(s,1H),4.37-4.35(m,2H),3.69-3.62(m,2H),3.47(d,J＝17.2Hz,1H),3.26(s,1H),2.31-2.23(m,1H),2.07-1.97(m,2H),1.68(d,J＝11.6Hz,1H).
实施例二十一：化合物21的合成
(S)-1'-(8-((2-(三氟甲基)吡啶-3-基)硫代)-[1,2,4]-三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺
Figure PCTCN2019115754-appb-000054
将(S)-1'-(8-碘[1,2,4]三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺(9-1,100mg，0.22mmol)溶于1,4-二氧六环(12mL)中，加入DIPEA(300mg,2.32mmol)与2-(三氟甲基)吡啶-3-硫醇钠(C3,200mg,粗品)。氮气保护下加入XantPhos (100mg,0.17mmol)，Pd 2(dba) 3(100mg,0.11mmol)。氮气置换三次后100℃反应3小时，反应完毕。反应液浓缩过柱(EA:MeOH＝5/1,0.5％氨水)得到白色固体化合物(S)-1'-(8-((2-(三氟甲基)吡啶-3-基)硫代)-[1,2,4]-三唑并[4,3-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯并[b]吡啶-6,4'-哌啶]-7-胺(化合物21，35mg，收率：31％)。LCMS：m/z 499.1[M+H] +
1H NMR(400MHz,MeOD)δ9.32(s,1H),8.41-8.37(m,2H),8.03(s,1H),7.72(d,J＝7.6Hz,1H),7.60(d,J＝8.4Hz,1H),7.38-7.35(m,1H),7.28-7.25(m，1H)，4.18–4.14(m,2H),4.04(s，1H)，3.62-3.59(m,2H),3.23(d,J＝16.4Hz,1H),2.90(d,J＝16.4Hz,1H),2.11-2.06(m,2H),1.79-1.75(m,1H),1.56-1.53(m,1H).
实施例二十二：化合物22的合成
(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,5-c]嘧啶-5-基)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-4-胺
Figure PCTCN2019115754-appb-000055
步骤一：将(S)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-4-胺盐酸盐(A13,155mg,0.35mmol)溶于乙腈(10mL)中，加入DIPEA(450mg,3.5mmol)与5-氯-8-碘咪唑并[1,2-c]嘧啶(B3,108mg,0.39mmol)，氮气保护下加热至100℃回流反应5小时，反应完毕。冷却至室温，将反应液倒入饱和碳酸氢钠水溶液，用二氯甲烷萃取(50mL x 2)。有机相合并后用饱和食盐水洗，有机相分液后用无水硫酸钠干燥，过滤浓缩。粗品通过硅胶色谱法纯化得到浅黄色固体化合物(S)-2-氯-1'-(8-碘咪唑并[1,2-c]嘧啶-5-基)-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-4-胺(22-1,72mg，收率43％)。LCMS：m/z 487[M+H] +
步骤二：将(S)-2-氯-1'-(8-碘咪唑并[1,2-c]嘧啶-5-基)-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-4--胺(22-1,50mg，0.1mmol)溶于1,4-二氧六环(5mL)中，加入DIPEA(30mg,0.25mmol)与2-氨基-3-氯吡啶-4-硫醇钠(C1,30mg,0.15mmol)。氮气保护下加入XantPhos(13mg,0.02mmol)，Pd 2(dba) 3(20mg,0.02mmol)。氮气置换三次后100℃反应3小时。反应液浓缩后用高效液相色谱纯化得到浅黄色固体化合物(S)-1'-(8-((2-氨基-3-氯吡啶-4-基)硫代)咪唑并[1,2-c]嘧啶-5-基)-2-氯-4,6-二氢螺[环戊二烯并[d]噻唑-5,4'-哌啶]-4-胺(化合物22，3mg，收率：5％)。LCMS：m/z 519[M+H] +
1H NMR(400MHz,MeOD)δ8.06(s,1H),7.85(s,1H),7.57(s,1H),7.50-7.49(d,1H),5.89-5.87(d,1H),4.15(s,1H)，,4.07-3.99(dd,2H),3.53–3.39(m,4H),3.11-3.09(d,2H),2.21-2.04(m,4H)
实施例二十三：化合物23的合成
Figure PCTCN2019115754-appb-000056
步骤一：向含1-(四氢-2H-吡喃-2-基)-1H-吡唑并[3,4-b]吡啶-4-硫醇钠(C7,224mg，0.8mmol)的反应瓶中加入二氧六环(20mL)，再加入(S)-N-((S)-1'-(8-碘代咪唑[1,2-C]嘧啶-5-基)-5,7-二氢螺环[环戊二烯[B]吡啶-6,4'-哌啶]-5-基)-2-甲基丙烷-2-亚磺酰胺(1-1，287mg，0.52mmol)，DIPEA(304mg，2.35mmol)，XantPhos(91mg，0.16mmol)和Pd 2(dba) 3(72mg，0.08mmol)。氮气保护下升温至100℃搅拌反应3h，TLC(二氯甲烷/甲醇＝20/1)和LCMS显示反应完全。反应液加乙酸乙酯稀释，过滤，滤渣乙酸乙酯淋洗，滤液浓缩干，硅胶柱层析得(R)-2-甲基-N-((5S)-1'-(8-((1-(四氢-2H-吡喃-2-基)-1H-吡唑并[3,4-b]吡啶-4-基)硫基)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯[b]吡啶-6,4'-哌啶]-5-基)丙烷-2-亚磺酰胺(23-1,352mg，收率：97.2％)。
LCMS:m/z 658.4[M+H] +
步骤六：氮气保护，0℃下向(R)-2-甲基-N-((5S)-1'-(8-((1-(四氢-2H-吡喃-2-基)-1H-吡唑并[3,4-b]吡啶-4-基)硫基)咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯[b]吡啶-6,4'-哌啶]-5-基)丙烷-2-亚磺酰胺(23-1，176mg，0.26mmol)的二氯甲烷(10mL)溶液中缓慢加入HCl/dioxane(4M，1.3mL，5.20mmol)，室温搅拌反应2h，TLC(二氯甲烷/甲醇＝8/1)和LCMS显示反应完全。反应液降温至0℃，缓慢加入氨甲醇溶液调节PH至10左右后减压浓缩，prep-TLC纯化得(S)-1'-(8-(((1H-吡唑并[3,4-b]吡啶-4-基]硫代]咪唑并[1,2-c]嘧啶-5-基)-5,7-二氢螺[环戊二烯[b]吡啶-6,4'-哌啶]-5-胺(23，73.01mg，收率：58.1％)。LCMS:m/z 470.5[M+H] +
1H NMR(400MHz,MeOD):δ8.42(d,J＝4.0Hz,1H),8.16～8.14(m,2H),8.10(s,1H),7.91～7.87(m,2H),7.56(d,J＝1.2Hz,1H),7.34～7.31(m,1H),4.26(s,1H),4.11～4.06(m,1H),3.49～3.43(m,2H),3.28(s,1H),3.07(d,J＝16.8Hz,1H),2.15～2.07(m,2H),1.76(d,J＝12.8Hz,2H),1.64(d,J＝13.6Hz,2H).
对照化合物合成：本发明申请时，与本发明最接近的对照物为WO2018136265实施例25。参照其合成路线与操作步骤，得到对照化合物aa.
Figure PCTCN2019115754-appb-000057
本发明公开化合物的生物学功能在酶活性以及细胞水平的测试中得到了证明。比如在SHP2酶活性抑制试验中，本发明公开的化合物能够达到很强的抑制活性(IC50可达1nM)。在细胞水平上，本发明公开化合物也表现出很好的抑制癌细胞的增殖的活性,在MV4-11细胞株上增殖抑制活性可达到1nM。与SHP099(6-(4-氨基-4-甲基哌啶-1-基)-3- (2,3-二氯苯基)吡嗪-2-胺)或对照化合物aa对比，本发明专利的化合物无论在酶学水平上还是在细胞水平上，都体现出了优越的活性。
测试例一：SHP2酶活测试方法
SHP2酶活测试方法：
化合物粉末溶于DMSO中制成母液。实验时，化合物存贮液用DMSO进行3-倍梯度稀释，同一化合物设置10个不同的测试浓度。取1μL各浓度点的化合物至检测板(Corning,Costar 3915)孔内,每个浓度点设置2个平行重复。以6,8-二氟-4-甲基-7-羟基香豆素磷酸酯(DiFMUP)作为底物，SHP2 E72A催化其水解产生6,8-二氟-4-甲基-7-羟基香豆素(DiFMU)，通过PE Enspire多功能读数仪，以358nm为激发波长来检测455nm处的荧光值，确定SHP2的酶活。
反应用的SHP2缓冲液
组成为60mmol/L Hepes，PH7.2,75mmol/L NaCl，75mmol/L KCl,1mmol/L EDTA，5mmol/L DTT.筛选体系组成为：SHP2缓冲液、酶SHP2 E76A蛋白、底物DiFMUP和待测化合物。
IC50测试方法：
96孔筛选板中50ng SHP2 E76A蛋白分别与待测化合物在SHP2缓冲液中反应20min，然后与10uM DiFMUP室温共同孵育20min，用PE Enspire多功能读数仪以358nm为激发光，读取455nm处的光强度。以化合物处理组测出的荧光值比DMSO对照孔的值计算样品对酶活性的抑制率。化合物的IC 50值由Graphpad公司的Prism软件，以抑制率对抑制剂的浓度非线性拟合计算得到。通过Y＝Bottom+(Top-Bottom)/(1+10^((LogIC50-X)*HillSlope))方程拟合出酶活性随化合物浓度变化的曲线。求得各化合物的IC50值。表1显示了本发明部分化合物的IC 50值。
表1
化合物编号 酶活性(nM) 化合物编号 酶活性(nM)
SHP099 263 化合物12 2
阳性对照aa 26 化合物13 1
化合物1 11 化合物14 8
化合物2 4 化合物15 3
化合物3 11 化合物16 2
化合物4 7 化合物17 10
化合物5 5 化合物18 7
化合物6 6 化合物19 7
化合物7 3 化合物20 6
化合物8 10 化合物21 11
化合物9 7 化合物22 9
化合物10 3 化合物23 9
化合物11 1    
测试例二：MV4-11细胞增殖抑制实验
通过
Figure PCTCN2019115754-appb-000058
发光法细胞活力检测试剂盒对细胞内ATP进行定量测定来检测培养物中活细胞数目。
第一步在96孔板种接种MV4-11细胞，以每孔2500个细胞密度接种细胞到96孔板，每孔体积100μL。置于37℃5％二氧化碳培养箱培养过夜。
第二步化合物处理细胞。待测化合物进行3倍稀释，共设置8个浓度梯度；每孔分别加入一定体积的DMSO或者待测化合物，每个浓度设置2个重复，DMSO的终浓度控制在0.5％。置于37℃5％二氧化碳培养箱培养72h。
第三步
Figure PCTCN2019115754-appb-000059
Luminescent Cell Viability Assay试剂盒(Promega，G7570)检测对照组和处理组细胞活力。每孔加入50ul CellTiter-Glo，混匀，室温孵育10min。使用EnSpire(Perkin Elmer)读取信号。抑制百分率(％)通过以下公式计算获得：
抑制百分率(％)＝(1-化合物处理组信号值/DMSO处理组信号值)*100，结果示于表2中。
表2
Figure PCTCN2019115754-appb-000060
测试例三：化合物药代动力学实验
本发明化合物对药代动力学测定。本申请采用以下方法测定本申请的化合物药代动力学参数。
研究使用的健康雄性成年小鼠，每组动物单次灌胃给药5-100mg/Kg。禁食从给药前10小时至给药后4小时。给药后不同时间点后采血，并测定化合物血浆含量(LC-MS/MS)。血浆浓度----时间关系用专业软件分析(winnonlin),计算化合物的药代动力学参数。根据表3显示，本发明化合物有着优异的药代动力学性质。
表3
Figure PCTCN2019115754-appb-000061
在本发明中提及的所有文献都在本申请中引用作为参考，就如同每一篇文献被单独引用作为参考那样。以上详细描述了本发明的优选实施方式，但是，本发明并不限于上述实施方式中的具体细节，在本发明的技术构思范围内，可以对本发明的技术方案进行多种简单变型，这些简单变型均属于本发明的保护范围。
另外需要说明的是，在上述具体实施方式中所描述的各个具体技术特征，在不矛盾的情况下，可以通过任何合适的方式进行组合，为了避免不必要的重复，本发明对各种可能的组合方式不再另行说明。此外，本发明的各种不同的实施方式之间也可以进行任意组合，只要其不违背本发明的思想，其同样应当视为本发明所公开的内容。

Claims (22)

  1. 一种式I化合物，或其药学上可接受的盐，或其对映异构体、非对映异构体、互变异构体、溶剂化物、同位素取代物、多晶型物、前药或代谢产物：
    Figure PCTCN2019115754-appb-100001
    其中，
    X 1和X 2各自独立地选自键、O、CR aR b或NR c；
    X 3选自键、CR aR b、NR c、S或O；
    X 4选自N或者CR c；且R a、R b和R c各自独立地选自H、卤素、取代或未取代的C 1-6烷基、取代或未取代的C 1-6烷氧基；
    R 1、R 2、R 3、R 4和R 7各自独立地选自H、-OH、卤素、取代或未取代的氨基、取代或未取代的C 1-6烷基、取代或未取代的C 1-6烷氧基；且不能同时为-OH或-NH 2；
    环A选自取代或未取代的C 4-8环烃基、取代或未取代的4-8元杂环基、取代或未取代的C 5-10芳基、取代或未取代的5-10元杂芳基，所述杂环基或杂芳基包含1-3个选自下组的杂原子：N、O、S或P；
    环C选自取代或未取代的C 4-8环烃基、取代或未取代的5-6元单环杂环基、取代或未取代的8-10元双环杂环基、取代或未取代的C 5-10单环或双环芳基、取代或未取代的5-6元单环杂芳基、取代或未取代的8-10元双环杂芳基，所述杂环基或杂芳基包含1-4个选自下组的杂原子：N、O、S或P；
    R 5和R 6各自独立地选自H、-OH、卤素、氰基、取代或未取代的氨基、取代或未取代的C 1-6烷基、取代或未取代的C 1-6烷氧基；
    n为0至3中的任一整数；并且
    所述取代是指基团上的一个或多个氢原子被选自下组的取代基取代：卤素、-OH、-NO 2、-NH 2、-NH(未取代或卤代的C 1-6烷基)、-N(未取代或卤代的C 1-6烷基) 2、-CN、未取代或卤代的C 1-8烷基、未取代或卤代的C 1-8烷氧基、未取代或卤代的C 1-8烷氧基-C 1-8烷基、未取代或卤代的C 3-8环烷基-C 1-8烷基、未取代或卤代的C 1-6烷基羰基、未取代或卤代的C 1-6烷氧基羰基、异羟肟酸基、未取代或卤代的C 1-6烷基巯基、-S(O) 2N(未取代或卤代的C 1-6烷基) 2、-S(O) 2未取代或卤代的C 1-6烷基、-N(未取代或卤代的C 1-6烷基)S(O) 2N(未取代或卤代的C 1-6烷基) 2、-S(O)N(未取代或卤代的C 1-6烷基) 2、-S(O)(未取代或卤代的C 1-6烷基)、-N(未取代或卤代的C 1-6烷基)S(O)N(未取代或卤代的C 1-6烷基) 2、-N(未取代或卤代的C 1-6烷基)S(O)(未取代或卤代的C 1-6烷基)、未取代或卤代的C 5- 10芳基、未取代或卤代的5-10元杂芳基、未取代或卤代的C 4-8环烃基、未取代或卤代的4-8元杂环基，所述杂环基和杂芳基包含1-4个选自下组的杂原子：N、O或S。
  2. 根据权利要求1所述的式I化合物，其中，X 1和X 2中的一个为CH 2，并且另一个为键。
  3. 根据权利要求1所述的式I化合物，其中，X 3为S。
  4. 根据权利要求1所述的式I化合物，其中，X 4选自N或者CH。
  5. 根据权利要求1所述的式I化合物，其中，R 1、R 2、R 3、R 4和R 7各自独立地选自H、-OH、-F、-Cl、-Br、-NH 2、-NHC 1-3烷基、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基或异丙氧基；被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷基；或被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷氧基。
  6. 根据权利要求1所述的式I化合物，其中，R 5和R 6各自独立地选自H、-OH、-F、-Cl、-Br、-CN、-NH 2、-NHC 1-3烷基、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基或异丙氧基；被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷基；或被卤素、-NH 2、-OH、C 1-3烷基或C 1-3烷氧基取代的C 1-3烷氧基。
  7. 根据权利要求1所述的式I化合物，其中，所述取代基选自-F、-Cl、-Br、-OH、-NO 2、-NH 2、-NH(C 1-6烷基)、-N(C 1-6烷基) 2、-CN、C 1-6烷基、C 1-4烷氧基、C 1-4烷氧基-C 1-6烷基、C 3-8环烷基-C 1-8烷基、C 1-6烷基羰基、C 1-6烷氧基羰基、C 1-6烷基巯基、-S(O) 2N(C 1-6烷基) 2、-S(O) 2C 1-6烷基、-N(C 1-6烷基)S(O) 2N(C 1-6烷基) 2、-S(O)N(C 1-6烷基) 2、-S(O)(C 1-6烷基)、-N(C 1-6烷基)S(O)N(C 1-6烷基) 2、-N(C 1-6烷基)S(O)(C 1-6烷基)、取代或未取代的C 5-10芳基、取代或未取代的5-10元杂芳基、取代或未取代的C 4-8环烃基、取代或未取代的4-8元杂环基，所述杂环基和杂芳基包含1-4个选自下组的杂原子：N、O或S。
  8. 根据权利要求7所述的式I化合物，其中，所述取代基选自-F、-Cl、-Br、-OH、-NO 2、-NH 2、-NH(C 1-3烷基)、-N(C 1-3烷基) 2、-CN、C 1-3烷基、C 1-3烷氧基、C 1-3烷基羰基、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、环辛基、吡咯烷基、吗啉基、哌嗪基、高哌嗪基、哌啶基、硫代吗啉基、苯基、萘基、蒽基、菲基、芴基、噻吩基、咪唑基、吡唑基、噻唑基、噁唑基、噁二唑基、异噁唑基、吡啶基、嘧啶基、吡嗪基、哒嗪基、苯并咪唑基、苯并吡唑基、吲哚基、呋喃基、吡咯基、三唑基、四唑基、三嗪基、吲嗪基、异吲哚基、吲唑基、异吲唑基、嘌呤基、喹啉基或异喹啉基。
  9. 根据权利要求8所述的式I化合物，其中，所述取代基选自-F、-Cl、-Br、-OH、-NO 2、-NH 2、-NH(C 1-3烷基)、-N(C 1-3烷基) 2、-CN、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基、异丙氧基或苯基。
  10. 根据权利要求9所述的式I化合物，其中，所述环C选自以下中的任一个：
    Figure PCTCN2019115754-appb-100002
    其中，
    X 5、X 6、X 7、X 8和X 9各自独立地选自N或CR d；且至多同时3个为N；
    X 10、X 11、X 12、X 13、X 14、X 15、X 16和X 17各自独立地选自N或CR d；且至多同时5个为N；
    X 18、X 19、X 20和X 21各自独立地选自N或CR d，且至多同时3个为N；
    R 6和R 8各自独立地选自H、-NH 2、-CN、-OH、-NO 2、卤素、未取代或卤代的C 1-6烷基、未取代或卤代的C 1-6烷氧基；并且
    所述R d选自H、卤素、未取代或卤代的C 1-6烷基、未取代或卤代的C 1-6烷氧基。
  11. 根据权利要求10所述的式I化合物，其中，所述环C选自以下中的任一个：
    Figure PCTCN2019115754-appb-100003
    其中，
    X 5、X 6、X 7、X 8和X 9中的0、1或2个为N，其余为CR d；
    X 18、X 19、X 20和X 21中的0、1或2个为N，其余为CR d；
    R 6选自H、-NH 2、-CN、-OH、-NO 2、-F、-Cl、-Br、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基、异丙氧基、氟代或溴代的C 1-3烷基、氟代或溴代的C 1- 3烷氧基；并且
    所述R d选自H、-F、-Cl、-Br、甲基、乙基、丙基、异丙基、丁基、甲氧基、乙氧基、丙氧基、异丙氧基、氟代或溴代的C 1-3烷基、氟代或溴代的C 1-3烷氧基。
  12. 根据权利要求11所述的式I化合物，其中，所述环C选自以下中的任一个：
    Figure PCTCN2019115754-appb-100004
  13. 根据权利要求1所述的式I化合物，其中，所述环A选自取代或未取代的C 4-6环烃基、取代或未取代的4-6元杂环基、取代或未取代的C 5-6芳基、取代或未取代的5-6元杂芳基，所述杂环基或杂芳基包含1-3个N原子。
  14. 根据权利要求13所述的式I化合物，其中，所述环A选自以下中的任一个：
    Figure PCTCN2019115754-appb-100005
  15. 根据权利要求14所述的式I化合物，其中，所述环A选自以下中的任一个：
    Figure PCTCN2019115754-appb-100006
  16. 根据权利要求1所述的式I化合物，其具有选自以下的结构：
    Figure PCTCN2019115754-appb-100007
  17. 根据权利要求1所述的化合物，所述同位素的原子包括但不局限于氢、碳、氮、氧、氟、磷、氯或碘；并且优选为 2H、 3H、 11C、 13C、 14C、 15N、 17O、 18O、 18F、 31P、 32P、 35S、 36Cl或 125I。
  18. 一种制备根据权利要求1-17中任一项所述的式I化合物的方法，其包括以下步骤：
    (i)使式Ib与式Ic发生亲核取代反应得到式Id；
    (ii)使式Id与式Ie发生取代反应得到式If；并且
    (iii)将式If用酸脱除保护以得到式I化合物：
    Figure PCTCN2019115754-appb-100008
  19. 根据权利要求1-17中任一项所述的式I化合物在以下方法中的用途：
    (a)制备预防或治疗与SHP2活性异常相关的疾病或病症的药物；
    (b)制备预防或治疗SHP2-介导的疾病或病症的药物
    (c)制备抑制SHP2活性的抑制剂药物；
    (d)体外非治疗性地抑制SHP2活性；
    (e)体外非治疗性地抑制肿瘤细胞增殖；或
    (f)治疗与SHP2异常相关的疾病或病症。
  20. 根据权利要求19所述的用途，其中，所述疾病为癌症，包括但不限于努南综合症、豹综合症、青少年髓单核细胞白血病、成神经细胞瘤、黑色素瘤、急性髓性白血病、乳腺癌、食道癌、肺癌、结肠癌、头癌、成神经细胞瘤、头颈的鳞状细胞癌、胃癌、间变性大细胞淋巴瘤或成胶质细胞瘤。
  21. 一种药物组合物，其包含：
    (i)有效量的根据权利要求1-17中任一项所述的式I化合物，或其药学上可接受的盐，或其对映异构体、非对映异构体、互变异构体、溶剂化物、同位素取代物、多晶型物、前药或代谢产物；和
    (ii)药学上可接受的载体。
  22. 一种抑制SHP2活性的方法，其包括以下步骤：对有此需要的受试者施用有效量的根据权利要求1-17中任一项所述的式I化合物或其药学上可接受的盐，或对有此需要的受试者施用有效量的根据权利要求21所述的药物组合物。
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