WO2024046454A1 - Heteroaryl-substituted pyridopyrrolidone derivative, and pharmaceutical composition and use thereof - Google Patents

Heteroaryl-substituted pyridopyrrolidone derivative, and pharmaceutical composition and use thereof Download PDF

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WO2024046454A1
WO2024046454A1 PCT/CN2023/116425 CN2023116425W WO2024046454A1 WO 2024046454 A1 WO2024046454 A1 WO 2024046454A1 CN 2023116425 W CN2023116425 W CN 2023116425W WO 2024046454 A1 WO2024046454 A1 WO 2024046454A1
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alkyl
ring
group
compound
substituted
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PCT/CN2023/116425
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Chinese (zh)
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孙洪鹏
许峰
王海龙
席宝信
童忠安
马前
李鑫
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上海海雁医药科技有限公司
扬子江药业集团有限公司
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Publication of WO2024046454A1 publication Critical patent/WO2024046454A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/425—Thiazoles
    • A61K31/427—Thiazoles not condensed and 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
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • 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
    • 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

Definitions

  • the present application relates to the field of medical technology, and in particular to a heteroaryl-substituted pyridopyrrolone derivative, its pharmaceutical composition and application.
  • Phosphoinositide 3-kinase is a type of phosphatidyl kinase with the function of phosphorylating the phosphoinositide ring. It is divided into three categories: type I, type II and type III. Among them, type I PI3K is the most widely studied. This type is further divided into two subgroups (IA and IB). Class IA PI3K consists of three closely related kinases, PI3K ⁇ , PI3K ⁇ , and PI3K ⁇ , now present as a heterodimer composed of a catalytic subunit (p110 ⁇ , p110 ⁇ , or p110 ⁇ ) and one of several regulatory subunits.
  • PI3K ⁇ and PI3K ⁇ are widely expressed and play roles in cell growth, division and survival (Thomas M, et al., Curr. Opin. Pharmacol., 2008; 8:267-274).
  • the roles of these two kinases in many biological functions are enhanced by the embryonic lethality observed in mice lacking PI3K ⁇ or PI3K ⁇ . Due to their role in homeostasis, clinical evaluation of PI3K ⁇ and PI3K ⁇ has been limited to the field of oncology, and some compounds are also in various stages of clinical development.
  • the single class 1B isoform of PI3K ⁇ mainly responds to G-protein coupled receptors (GPCR) and is composed of the p110 ⁇ catalytic subunit and one of two different regulatory subunits.
  • GPCR G-protein coupled receptors
  • PI3K ⁇ isoforms are expressed in immune cells and have limited expression in normal or malignant epithelial and connective tissue cells.
  • Research results on PI3K ⁇ knockout mice indicate that PI3K ⁇ is important for cell activation and the migration of some chemokines (Sasaki T., et al., Science, 2000; 287:1040-1046; Hirsch E., et al. ,Science,2000;287:1049-1053).
  • PI3K ⁇ signaling is particularly important for myeloid cell function, and Camps et al. described that treatment with the selective PI3K ⁇ inhibitor AS-60485023 inhibited the progression of joint inflammation and damage in two different mouse models of rheumatoid arthritis (Camps M, et al., Nat. Med. 2005, 11, 936-943). Based on studies at the cellular level and efficacy observed in various disease models, PI3K ⁇ inhibitors can potentially be used to treat various diseases, such as inflammation, metabolism, and cancer (Cushing, T.D., et al., J. Med.
  • the purpose of this application is to provide a heteroaryl-substituted pyridopyrrolone derivative with good selectivity and good pharmacokinetic properties.
  • the first aspect of this application provides a compound represented by formula (I), or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof:
  • Ring A is a 5- or 6-membered heteroaryl ring
  • (R 3 ) n means that the hydrogen on ring A is replaced by n R 3s , n is 0, 1, 2, 3 or 4; each R 3 is the same or different, and each R 3 is independently deuterium, halogen, cyano, or hydroxyl , carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halo C 1-3 alkyl, halo C 1-3 alkoxy , NR a1 R b1 , -N(R a3 )-C(O)C 1-3 alkyl, -N(R a3 )-C(O)-deuterated C 1-3 alkyl, -N(R a3 )-C(O)OC 1-3 alkyl, -SO 2 C 1-3 alkyl, -SO 2 C 3-6 cycloalkyl, -C(O)NR a1 R b1 , -C(O)OC
  • R Z , RT , and RU are each independently hydrogen, halogen, NR a1 R b1 or C 1-8 alkyl; wherein, the C 1-8 alkyl is preferably C 1-6 alkyl, and more preferably C 1-3 alkyl;
  • R W and R Y are each independently hydrogen, halogen, C 1-8 alkyl, halogenated C 1-8 alkyl, halogenated C 1-8 alkoxy, NR a1 R b1 or C 3-6 cycloalkyl base; wherein, the C 1-8 alkyl group is preferably a C 1-6 alkyl group, and more preferably a C 1-3 alkyl group;
  • R W and R Y are connected to form a 9- or 10-membered heteroaryl ring, a 9- or 10-membered phenyl heterocycloalkyl ring, or a 9- or 10-membered heteroaryl heterocycloalkyl ring with the adjacent 6-membered ring.
  • the 9- or 10-membered heteroaryl ring, the 9- or 10-membered phenyl heterocycloalkyl ring, and the 9- or 10-membered heteroaryl heterocycloalkyl ring are each independently unsubstituted or substituted by 1 or 2 Or substituted by 3 substituents each independently selected from the substituent group Q;
  • R 1 and R 2 are each independently C 1-8 alkyl, halogenated C 1-8 alkyl or C 3-6 cycloalkyl; wherein, the C 3-6 cycloalkyl is unsubstituted or substituted 1 or 2 substituents each independently selected from halogen and C 1-3 alkyl are substituted; the C 1-8 alkyl is preferably C 1-6 alkyl, more preferably C 1-3 alkyl; so The halogenated C 1-8 alkyl group is preferably a halogenated C 1-6 alkyl group, and more preferably a halogenated C 1-3 alkyl group;
  • the substituent group Q is selected from halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halogenated C 1-3 Alkyl group, halogenated C 1-3 alkoxy group, NR a1 R b1 , -SO 2 C 1-3 alkyl group, -S(O)C 1-3 alkyl group, -C(O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy, 3 to 6-membered heterocycloalkyl , phenyl or 5 to 6-membered heteroaryl;
  • R a1 and R b1 are each independently hydrogen, C 1-3 alkyl or acetyl, or R a1 , R b1 and the connected nitrogen atom together form a 4- to 6-membered saturated monoheterocyclic ring; the 4- to 6-membered saturated
  • the monoheterocycle is unsubstituted or substituted by 1, 2 or 3 substituents each independently selected from the following: deuterium, halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkyl Oxygen group, C 2-4 alkenyl group, C 2-4 alkynyl group, halogenated C 1-3 alkyl group, halogenated C 1-3 alkoxy group, -SO 2 C 1-3 alkyl group, -S(O )C 1-3 alkyl, -C(O)NH 2 , -C(O)NH(C 1-3 alkyl), -C(O)N(C 1-3 alkyl) 2
  • R a3 is hydrogen or C 1-8 alkyl; wherein the C 1-8 alkyl is preferably C 1-6 alkyl, more preferably C 1-3 alkyl.
  • one of T, W, Y, and U is N.
  • T is CRT ; U is CRU ; W is N or CRW ; Y is N or CRY ; and W and Y are not N at the same time.
  • two of T, W, Y, and U are N.
  • the structure is selected from the group consisting of formula (A), formula (B) and formula (C):
  • the 9- or 10-membered heteroaryl ring is selected from: indolyl, isoindolyl, indazolyl, benzotriazolyl, benzothienyl, isobenzothiophene base, benzofuryl, benzisofuryl, benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzoxadiazolyl, benzothiazolyl, benzisothiazolyl, benzene Thiadiazolyl, indanazinyl, purinyl, pyrido[3,2-d]pyrimidinyl, pyrido[2,3-d]pyrimidinyl, pyrido[3,4-d]pyrimidinyl, pyridine And[4,3-d]pyrimidinyl, 1,8-naphthyridinyl, 1,7-naphthyridinyl, 1,6-naphthy
  • the heterocycloalkyl groups in the 9- or 10-membered phenylheterocycloalkyl ring and the 9- or 10-membered heteroarylheterocycloalkyl ring are each independently selected from tetrahydrofuranyl.
  • tetrahydrothienyl tetrahydropyrrolyl, oxazolidinyl, dioxolanyl, piperidyl, piperazinyl, morpholinyl, dioxane, thiomorpholinyl, thiomorpholine -1,1-dioxide, tetrahydropyranyl, azetidine-2-one, oxetane-2-one, dihydrofuran-2(3H)-one, pyrrole Alk-2-one group, pyrrolidine-2,5-dione group, dihydrofuran-2,5-dione group, piperidin-2-one group, tetrahydro-2H-pyran-2-one group , piperazine-2-one group, morpholin-3-one group.
  • the heteroaryl group in the 9- or 10-membered heteroaryl heterocycloalkyl ring is selected from the group consisting of pyridyl, pyridazinyl, pyrimidinyl, and pyrazinyl.
  • R T is hydrogen; R U is hydrogen; R W is hydrogen, halogen, C 1-6 alkyl, halo C 1-6 alkyl or halo C 1-6 alkoxy.
  • R T is hydrogen;
  • R U is hydrogen;
  • R W is hydrogen, fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, Trifluoromethyl or trifluoromethoxy.
  • R T is hydrogen;
  • R U is hydrogen;
  • R Y is hydrogen, halogen, C 1-6 alkyl, halo C 1-6 alkyl, halo C 1-6 alkoxy or C 3-6 cycloalkyl.
  • R T is hydrogen;
  • R U is hydrogen;
  • R Y is fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, trifluoro Methyl, trifluoromethoxy, cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl.
  • the structure Choose one of the following structures:
  • Ring A is a 5- or 6-membered heteroaryl ring selected from the following group: thiophene ring, furan ring, thiazole ring, isothiazole ring, imidazole ring, oxazole ring, pyrrole ring, pyrazole Ring, 1,2,3-triazole ring, 1,2,4-triazole ring, 1,2,5-triazole ring, 1,3,4-triazole ring, tetrazole ring, isoxazole ring , 1,2,3-oxadiazole ring, 1,2,4-oxadiazole ring, 1,2,5-oxadiazole ring, 1,3,4-oxadiazole ring, thiadiazole ring, Pyridine ring, pyridazine ring, pyrimidine ring, pyrazine ring, triazine ring or tetrazine ring.
  • Ring A is a thiazole ring.
  • n is 2.
  • each R 3 is the same or different, and each is independently C 1-3 alkyl, -NH-C(O)C 1-3 alkyl, -NH-C(O)- Deuterated C 1-3 alkyl or -NH-C(O)OC 1-3 alkyl.
  • R 01 and R 02 are each independently deuterium, halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halogen Substituted C 1-3 alkyl, halogenated C 1-3 alkoxy, NR a1 R b1 , -N(R a3 )-C(O)C 1-3 alkyl, -N(R a3 )-C( O)- Deuterated C 1-3 alkyl, -N(R a3 )-C(O)OC 1-3 alkyl, -SO 2 C 1-3 alkyl, -SO 2 C 3-6 cycloalkyl, -C (O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy,
  • R 01 is halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, halo C 1-3 alkyl or halo C 1- 3 alkoxy;
  • R 02 is -NH-C(O)C 1-3 alkyl, -NH-C(O)-deuterated C 1-3 alkyl or -NH-C(O)OC 1-3 alkyl.
  • R 01 is methyl, ethyl, n-propyl or isopropyl. Further preferably, R 01 is methyl.
  • R 02 is -NH-C(O)C 1-3 alkyl, -NH-C(O)-deuterated C 1-3 alkyl or -NH-C(O)OC 1-3 alkyl . Further preferably, R 02 is -NH-C(O)CH 3 , -NH-C(O)-CD 3 or -NH-C(O)OCH 3 .
  • R 1 is monochloromethyl, dichloromethyl, trichloromethyl, monochloroethyl, 1,2-dichloroethyl, trichloroethyl, monobromoethyl , monofluoromethyl, difluoromethyl, trifluoromethyl, monofluoroethyl, difluoroethyl, trifluoroethyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, monofluoro-substituted Cyclopropyl or monofluoro-substituted cyclobutyl;
  • R 2 is methyl, ethyl, n-propyl, isopropyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, monofluoro-substituted cyclopropyl group or a fluorine-substi
  • R1 is cyclopropyl, cyclobutyl or trifluoromethyl.
  • R2 is methyl, ethyl, cyclopropyl or cyclobutyl.
  • Z is N or CH.
  • the 5- or 6-membered heteroaryl (ring) described in the group R 3 and substituent group Q in each of the above structural formulas are each independently selected from: thienyl, furyl, thiazole base, isothiazolyl, imidazolyl, oxazolyl, pyrrolyl, pyrazolyl, triazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,5- Triazolyl, 1,3,4-triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl or tetrazinyl.
  • the 5- or 6-membered heteroaryl (ring) is each independently selected from the following structures:
  • the 3- to 6-membered heterocycloalkyl groups described in the above structural formulas, the group R 3 and the substituent group Q are 4 to 6-membered heterocycloalkyl groups, each independently selected from : Azetidinyl, oxetanyl, tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyrrolyl, oxazolidinyl, dioxolanyl, piperidinyl, piperazinyl, morpholinyl , dioxanyl, thiomorpholinyl, thiomorpholine-1,1-dioxide, tetrahydropyranyl, pyrrolidin-2-one, dihydrofuran-2(3H)-one base, morpholin-3-one group, piperazin-2-one group and piperidin-2-one group.
  • the compound of formula (I) is selected from any one of the following structures:
  • the second aspect of the present application provides a pharmaceutical composition, which includes the compound described in the first aspect of the present application, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, and a pharmaceutically acceptable carrier.
  • the third aspect of this application provides the compounds described in the first aspect of this application, or pharmaceutically acceptable salts thereof, or their stereoisomers, and the pharmaceutical compositions described in the second aspect of this application in the preparation of treatments and/or Use in drugs to prevent diseases associated with or mediated by PI3K ⁇ activity.
  • the disease related to or mediated by PI3K ⁇ activity is inflammation, metabolic disease or cancer.
  • Figure 1 shows the tumor growth curve of the mixed inoculation model of MC38 and macrophages in C57BL/6 mice
  • Figure 2 shows the tumor weight on day 27 of C57BL/6 mouse MC38 and macrophage mixed inoculation model.
  • Alkyl refers to straight and branched chain saturated aliphatic hydrocarbon groups.
  • C 1-8 alkyl refers to an alkyl group having 1 to 8 carbon atoms, preferably C 1-6 alkyl, more preferably C 1-3 alkyl; non-limiting examples of alkyl include : Methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-di Methylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl , 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethyl But
  • Alkynyl refers to straight-chain and branched unsaturated aliphatic hydrocarbon groups with one or more carbon-carbon triple bonds
  • C 2-8 alkynyl refers to alkynyl groups with 2 to 8 carbon atoms, preferably C 2-6 alkynyl, more preferably C 2-4 alkynyl, is similarly defined; non-limiting examples of alkynyl include ethynyl, propynyl, n-butynyl, isobutynyl, pentynyl, hexynyl Alkynyl etc.
  • Cycloalkyl and “cycloalkyl ring” are used interchangeably and both refer to a saturated monocyclic, bicyclic or polycyclic cyclic hydrocarbon group which may be fused to an aryl or heteroaryl group. Cycloalkyl rings may be optionally substituted. In certain embodiments, cycloalkyl rings contain one or more carbonyl groups, such as oxo groups. “C 3-8 cycloalkyl” refers to a monocyclic cycloalkyl group having 3 to 8 carbon atoms.
  • Non-limiting examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, Cycloheptyl, cyclooctyl, cyclobutanone, cyclopentanone, cyclopentane-1,3-dione, etc.
  • Preferred is C 3-6 cycloalkyl, including cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.
  • C 8-10 cycloalkyl refers to a fused bicyclic cyclic hydrocarbon group having 8 to 10 ring atoms.
  • Non-limiting examples of C 8-10 cycloalkyl include
  • Heterocycloalkyl and “heterocycloalkyl ring” are used interchangeably and both refer to a cycloalkyl group containing at least one heteroatom selected from nitrogen, oxygen, and sulfur, which group may be associated with an aryl or heteroaryl group. Condensation. Heterocycloalkyl rings may be optionally substituted. In certain embodiments, heterocycloalkyl rings contain one or more carbonyl or thiocarbonyl groups, such as groups containing oxo and thio.
  • 3- to 8-membered heterocycloalkyl refers to a monocyclic cyclic hydrocarbon group having 3 to 8 ring atoms, wherein 1, 2 or 3 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur, preferably 4 to 8-membered heterocycloalkyl. More preferred is a 3- to 6-membered heterocycloalkyl group, which has 3 to 6 ring atoms, of which 1 or 2 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. Further preferred is a 4- to 6-membered heterocycloalkyl group having 4 to 6 ring atoms, of which 1 or 2 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur.
  • Non-limiting examples include aziridyl, oxiranyl, azetidinyl, oxetanyl, tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyrrolyl, oxazolidinyl, di Oxopentyl, piperidinyl, piperazinyl, morpholinyl, dioxanyl, thiomorpholinyl, thiomorpholine-1,1-dioxide, tetrahydropyranyl, aza cyclobutane-2-one group, oxetan-2-one group, dihydrofuran-2(3H)-one group, pyrrolidine-2-one group, pyrrolidine-2,5-dione group , dihydrofuran-2,5-dione group, piperidin-2-one group, tetrahydro-2H-pyran-2-one group, piperazine-2-one group, morpholin-3-one group, etc
  • 6- to 12-membered heterocycloalkyl and “6- to 12-membered fused heterocycloalkyl” are used interchangeably and refer to having 6 to 12 ring atoms, of which 1, 2 or 3 ring atoms are selected from A fused bicyclic cyclic hydrocarbon radical of heteroatoms of nitrogen, oxygen and sulfur.
  • 8 to 10 membered heterocycloalkyl and “8 to 10 membered fused heterocycloalkyl” can Used interchangeably, it refers to a fused bicyclic cyclic hydrocarbon group having 8 to 10 ring atoms, of which 1, 2 or 3 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur.
  • Non-limiting examples include hexahydro-1H-furan[3,4-c]pyrrole, octahydro-1H-cyclopenta[c]pyridine, hexahydro-1H-pyrrolo[2,1-c][1, 4]oxazine, octahydropyrrolo[1,2-a]pyrazine, hexahydropyrro[1,2-a]pyrazin-4(1H)-one, octahydrocyclopenta[c]pyrrole, etc.
  • the point of attachment may be a carbon or nitrogen atom as long as the valency permits.
  • Bicyclic heterocycloalkyl systems may include one or more heteroatoms in one or both rings.
  • Heteroaryl and “heteroaryl ring” are used interchangeably and both refer to a monocyclic, bicyclic or polycyclic 4n+2 aromatic ring system having ring carbon atoms and ring heteroatoms (e.g., having a Arrange shared 6 or 10 ⁇ electrons) groups in which each heteroatom is independently selected from nitrogen, oxygen and sulfur. Heteroaryl rings may be optionally substituted.
  • "5- to 10-membered heteroaryl” refers to a monocyclic or bicyclic heteroaryl group having 5 to 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms.
  • 5- to 6-membered heteroaryl refers to a monocyclic heteroaryl group with 5 to 6 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms.
  • Non-limiting examples include thienyl, furan base, thiazolyl, isothiazolyl, imidazolyl, oxazolyl, pyrrolyl, pyrazolyl, triazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2 ,5-triazolyl, 1,3,4-triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl Azolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl
  • 9- or 10-membered heteroaryl refers to a bicyclic heteroaryl group with 9 or 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms.
  • Non-limiting examples include indolyl, iso- Indolyl, indazolyl, benzotriazolyl, benzothienyl, isobenzothienyl, benzofuranyl, benzisofuranyl, benzimidazolyl, benzoxazolyl, benzoiso Oxazolyl, benzoxadiazolyl, benzothiazolyl, benzisothiazolyl, benzothiadiazolyl, indanazinyl, purinyl, pyrido[3,2-d]pyrimidinyl, pyrido [2,3-d]pyrimidinyl, pyrido[3,4-d]pyrimidinyl, pyrido[4,3-d]pyrimidinyl, 1,8-naph
  • Heteroatom means nitrogen, oxygen or sulfur. In heteroaryl groups containing one or more nitrogen atoms, the point of attachment may be a carbon or nitrogen atom as long as the valency permits. Heteroaryl bicyclic systems may include one or more heteroatoms in one or both rings.
  • “Fused” refers to a structure in which two or more rings share one or more bonds.
  • Phenylheterocycloalkyl refers to a group in which a benzene ring is fused with a heterocycloalkyl ring to form a bicyclic, tricyclic or polycyclic system, wherein the heterocycloalkyl ring is as defined above.
  • "9- or 10-membered phenylheterocycloalkyl” refers to a bicyclic cyclic group having 9 or 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. .
  • Non-limiting examples include indoline, benzo[d][1,3]dioxazole, 1,2,3,4-tetrahydroisoquinoline, 3,4-dihydro-2H-benzo[ b][1,4]oxazine, indole-2-one, isoindolin-1-one, 1,4-dihydroisoquinolin-3(2H)-one, etc.
  • Heteroarylheterocycloalkyl refers to a group in which a heteroaryl ring is fused with a heterocycloalkyl ring to form a bicyclic, tricyclic or polycyclic system, wherein the heterocycloalkyl ring is as defined above.
  • “9- or 10-membered heteroarylheterocycloalkyl” refers to a bicyclic cyclic group having 9 or 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. group.
  • Non-limiting examples include 2,3-dihydro-1H-pyrrolo[2,3-b]pyridine, [1,3]dioxola[4,5-b]pyridine, 2,3-dihydro -1H-pyrido[3,4-b][1,4]oxazine, 2,3,4,6-tetrahydropyrrolo[3,4-b][1,4]oxazine, 2,4 ,5,6-tetrahydropyrano[2,3-c]pyrazole, 5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine, etc.
  • Alkoxy refers to -O-alkyl, where alkyl is as defined above. It is preferably a C 1-8 alkoxy group, more preferably a C 1-6 alkoxy group, and most preferably a C 1-3 alkoxy group. Non-limiting examples of alkoxy include methoxy, ethoxy, n-propoxy, isopropoxy, butoxy, tert-butoxy, isobutoxy, pentyloxy, and the like.
  • Cycloalkyloxy refers to -O-cycloalkyl, where cycloalkyl is as defined above. A C 3-8 cycloalkyloxy group is preferred, and a C 3-6 cycloalkyloxy group is more preferred. Non-limiting examples of cycloalkyloxy include cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy, and the like.
  • a bond means that the two groups connected thereto are connected by a covalent bond.
  • Halogen means fluorine, chlorine, bromine or iodine.
  • Halo means that one or more (eg, 1, 2, 3, 4 or 5) hydrogens in a group are replaced by halogen.
  • haloalkyl refers to an alkyl group in which the hydrogens are replaced by one or more (eg, 1, 2, 3, 4 or 5) halogens, where alkyl is as defined above.
  • a halogenated C 1-8 alkyl group is preferred, a halogenated C 1-6 alkyl group is more preferred, and a halogenated C 1-3 alkyl group is even more preferred.
  • haloalkyl groups include, but are not limited to, monochloromethyl, dichloromethyl, trichloromethyl, monochloroethyl, 1,2-dichloroethyl, trichloroethyl, monobromoethyl, monofluoromethyl base, difluoromethyl, trifluoromethyl, monofluoroethyl, difluoroethyl, trifluoroethyl, etc.
  • haloalkoxy means that the hydrogen on the alkoxy group is substituted by one or more (such as 1, 2, 3, 4 or 5) halogens, where the alkoxy group is as defined above.
  • a halogenated C 1-8 alkoxy group is preferred, a halogenated C 1-6 alkoxy group is more preferred, and a halogenated C 1-3 alkoxy group is even more preferred.
  • Haloalkoxy groups include, but are not limited to, trifluoromethoxy, trifluoroethoxy, monofluoromethoxy, monofluoroethoxy, difluoromethoxy, difluoroethoxy, and the like.
  • halogenated cycloalkyl means that the hydrogen on the cycloalkyl group is substituted by one or more (such as 1, 2, 3, 4 or 5) halogens, wherein the cycloalkyl group is as defined above.
  • a halogenated C 3-8 cycloalkyl group is preferred, and a halogenated C 3-6 cycloalkyl group is more preferred.
  • Halogenated cycloalkyl groups include, but are not limited to, trifluorocyclopropyl, monofluorocyclopropyl, monofluorocyclohexyl, difluorocyclopropyl, difluorocyclohexyl, etc.
  • Deuterated alkyl means that the hydrogen on the alkyl group is replaced by one or more (eg, 1, 2, 3, 4 or 5) deuterium atoms, where alkyl is as defined above. It is preferably a deuterated C 1-8 alkyl group, more preferably a deuterated C 1-6 alkyl group, and even more preferably a deuterated C 1-3 alkyl group. Examples of deuterated alkyl groups include, but are not limited to, monodeuterated methyl, monodeuterated ethyl, dideuterated methyl, dideuterated ethyl, trideuterated methyl, trideuterated ethyl, and the like.
  • Amino refers to NH 2
  • cyano refers to CN
  • nitro refers to NO 2
  • benzyl refers to -CH 2 -phenyl
  • oxo O
  • carboxyl refers to -C (O)OH
  • acetyl refers to -C(O)CH 3
  • hydroxymethyl refers to -CH 2 OH
  • hydroxyethyl refers to -CH 2 CH 2 OH or -CHOHCH 3
  • hydroxyl refers to -OH
  • mercapto refers to SH
  • the structure of "cyclopropylene” is:
  • Substituted refers to one or more hydrogen atoms in the group, preferably 1 to 5 hydrogen atoms are each independently substituted by a corresponding number of substituents, and more preferably 1 to 3 hydrogen atoms are each independently substituted by a corresponding number of substituents. Substituted with substituents. It goes without saying that the substituents are only in their possible chemical positions, and the person skilled in the art is able to determine (either experimentally or theoretically) possible or impossible substitutions without undue effort. For example, an amino or hydroxyl group with a free hydrogen may be unstable when combined with a carbon atom with an unsaturated (eg, olefinic) bond.
  • L is (CR 01 R 02 ) s .
  • L is (CR 01 R 02 )-(CR 01 R 02 ).
  • the two R 01 can be the same or different, and the two R 02 can be Identical or different, they are independent species.
  • L can be C(CH 3 )(CN)-C(CH 2 CH 3 )(OH), C(CH 3 )(CN)-C(CH 3 )(OH ) or C(CN)(CH 2 CH 3 )-C(OH)(CH 2 CH 3 ).
  • any group in this application may be substituted or unsubstituted.
  • the substituents are preferably 1 to 5 or less groups, independently selected from cyano, halogen (preferably fluorine or chlorine), C 1-8 alkyl (preferably C 1-6 alkyl) , more preferably C 1-3 alkyl), C 1-8 alkoxy (preferably C 1-6 alkoxy, more preferably C 1-3 alkoxy), halogenated C 1-8 alkyl (preferably halogenated C 1-6 alkyl, more preferably halogenated C 1-3 alkyl), C 3-8 cycloalkyl (preferably C 3-6 cycloalkyl), halogenated C 1-8 Alkoxy (preferably halogenated C 1-6 alkoxy, more preferably halogenated C 1-3 alkoxy), C 1-8 alkyl substituted amino, halogenated C 1-8 alkyl substituted Amino, acetyl, hydroxyl, hydroxymethyl, hydroxyethyl
  • any two “preferences” may be independent of each other.
  • any two substituents may be the same or different.
  • it can be substituted by two identical or different halogens, or it can be substituted by one halogen and one hydroxyl group.
  • Sub represents various substituents described in this application; Represents the connection position to other atoms.
  • the compounds of the present application or their pharmaceutically acceptable salts or their stereoisomers can be combined with one or more pharmaceutical carriers to form a suitable dosage form for administration.
  • dosage forms are suitable for oral, rectal, topical, intraoral, and other parenteral administration (e.g., subcutaneous, intramuscular, intravenous, etc.).
  • dosage forms suitable for oral administration include capsules, tablets, granules, and and syrup, etc.
  • the compounds of the present application contained in these preparations can be solid powders or granules; solutions or suspensions in aqueous or non-aqueous liquids; water-in-oil or oil-in-water emulsions, etc.
  • the above dosage forms can be prepared from the active compound and one or more carriers or excipients through common pharmaceutical methods.
  • the above carriers need to be compatible with the active compound or other excipients.
  • the active compounds can form solutions or suspensions with carriers.
  • “Pharmaceutically acceptable carrier” means a non-toxic, inert, solid, semi-solid substance or liquid filling machine, diluent, encapsulating material or auxiliary preparation or excipient of any type that is compatible with the patient, preferably a mammal , more preferably human, which is suitable for delivering the active agent to the intended target site without terminating the activity of the agent.
  • Compounds of the present application refer to compounds of formula (I) of the present application, or pharmaceutically acceptable salts thereof, or stereoisomers thereof, which have PI3K ⁇ selective inhibitory activity, and preparation of compounds of formula (I), or pharmaceutically acceptable salts thereof. Acceptable salts, or intermediate compounds of its stereoisomers.
  • compositions of the present application are formulated, dosed, and administered in a manner consistent with good medical practice.
  • the "therapeutically effective amount" of a compound administered is determined by factors such as the specific condition to be treated, the individual being treated, the cause of the condition, the target of the drug, and the mode of administration.
  • “Therapeutically effective amount” refers to the amount of a compound of the present application that will induce a biological or medical response in an individual, such as reducing or inhibiting enzyme or protein activity or improving symptoms, alleviating disease, slowing or delaying disease progression, or preventing disease, etc.
  • the pharmaceutical composition of the present application or the therapeutically effective amount of the compound of the present application or its pharmaceutically acceptable salt or its stereoisomer contained in the pharmaceutical composition is preferably 0.1 mg/kg-5g/kg ( weight).
  • Patient means an animal, preferably a mammal, more preferably a human being.
  • mammal refers to warm-blooded vertebrate mammals, including, for example, cats, dogs, rabbits, bears, foxes, wolves, monkeys, deer, rats, pigs, and humans.
  • Treatment means to alleviate, delay the progression, attenuate, prevent, or maintain an existing disease or condition (eg, cancer). Treatment also includes curing, preventing the progression of, or alleviating to some degree one or more symptoms of a disease or condition.
  • compositions include pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.
  • Pharmaceutically acceptable acid addition salts refer to salts formed with inorganic or organic acids that retain the biological effectiveness of the free base without other side effects. These salts can be prepared by methods known in the art.
  • “Pharmaceutically acceptable base addition salts” include, but are not limited to, salts of inorganic bases such as sodium salts, potassium salts, calcium salts and magnesium salts. Including but not limited to salts of organic bases, such as ammonium salts, triethylamine salts, lysine salts, arginine salts, etc. These salts can be prepared by methods known in the art.
  • the compound represented by formula (I) of the present application may exist in different optically active forms.
  • a compound of formula (I) contains a chiral center, the compound contains a pair of enantiomers.
  • Two enantiomers of the compound as well as mixtures of the pair of enantiomers, such as racemic mixtures, are also within the scope of the present application.
  • Enantiomers can be resolved by methods known in the art, such as crystallization and chiral chromatography.
  • the compound contains more than one chiral center, the compound contains enantiomers and diastereomers.
  • Enantiomers and diastereomers of this compound can be resolved by methods known in the art, such as crystallization and preparative chromatography.
  • the compounds described in this application can be synthesized using methods similar to those described below or the exemplary methods described in the Examples, or relevant publications used by those skilled in the art, by using appropriate alternative starting materials.
  • Starting materials for the compounds described herein may be synthesized or may be obtained from commercial sources.
  • the compounds described herein and other related compounds having various substituents can be synthesized using techniques and starting materials known to those skilled in the art.
  • General methods for preparing the compounds disclosed herein can be derived from reactions known in the art, and the reactions can be modified by reagents and conditions deemed appropriate by those skilled in the art to introduce various moieties in the molecules provided herein.
  • the IC 50 value of the inhibitory activity against PI3K ⁇ kinase is less than 100 nM. , in some embodiments having an IC 50 value of less than 50 nM, in some embodiments having an IC 50 value of less than 10 nM, and therefore can be used as a therapeutic agent for the treatment and/or prevention of diseases associated with or mediated by PI3K ⁇ activity. drug.
  • Known starting materials can be synthesized by methods known in the art, or can be purchased from ABCR GmbH&Co.KG, Acros Organics, Aldrich Chemical Company, Shaoyuan Chemical Technology (Accela ChemBio Inc) and Dari Chemicals, etc. company.
  • DCM dichloromethane
  • DMF dimethylformamide
  • THF tetrahydrofuran
  • Pd(dppf)Cl 2 [1,1'-bis(diphenylphosphorus)ferrocene]palladium dichloride
  • dppf 1 ,1'-bis(diphenylphosphine)ferrocene
  • Pin 2 B 2 pinacol diboronate
  • SEM-Cl 2-(trimethylsilyl)ethoxymethyl chloride
  • ACN acetonitrile
  • TFA trifluoroacetic acid
  • DCM dichloromethane
  • MeOH methanol
  • n-BuLi n-butyllithium
  • KOAc potassium acetate
  • 1,4-Dioxane 1,4-dioxane
  • CombiFlash fast liquid phase Preparative chromatograph
  • EA ethyl acetate
  • PE petroleum ether
  • DMSO dimethyl sulfoxide
  • the percentage content involved in this application refers to mass percentage for solid-liquid mixing and solid-solid phase mixing, and refers to volume percentage for liquid-liquid phase mixing.
  • the final product of the present application can be obtained through preparation and purification, and the purification and separation conditions can be obtained by those skilled in the art based on the characteristics of the compound through tests with common knowledge in the art.
  • room temperature refers to about 20-30°C.
  • Step 1 Dissolve 2,6-dichloro-4-methylnicotinic acid (40.00g, 194.15mmol) and potassium carbonate (67.08g, 485.38mmol) in DMF (200mL), add methyl iodide (41.34g) dropwise at room temperature ,291.23mmol), then react at 24°C for 1 hour, Pour the reaction solution into water, extract with ethyl acetate, wash the organic phase twice with saturated brine, dry over anhydrous sodium sulfate, filter, and concentrate the filtrate to obtain a brown crude product, 2,6-dichloro-4-methylnicotinic acid methyl. Ester (39.00g, yield: 92.86%). MS(ESI)220.0[M+H] + .
  • Step 2 Combine the crude product 2,6-dichloro-4-methylnicotinic acid methyl ester (39.00g, 177.23mmol) and N-bromosuccinimide (47.32g, 265.85mmol) and benzyl peroxide
  • the acyl 64.40 g, 265.85 mmol
  • Step 3 Combine the crude product 2,6-dichloro-4-methylnicotinic acid-4-(bromomethyl)-2,6-dichloronicotinic acid methyl ester (25.00g, 84.20mmol) and 2,6- Dichloro-4-(dibromomethyl)nicotinic acid methyl ester (25.00g, 66.17mmol) was dissolved in ACN (300mL), and the temperature was lowered to 5°C under N 2 protection. At this temperature, N, N-dioic acid was added dropwise.
  • Step 4 Combine 2,6-dichloro-4-methylnicotinic acid-4-(bromomethyl)-2,6-dichloronicotinic acid methyl ester (28.00g, 93.66mmol) and (S)-1- Cyclopropylethylamine hydrochloride (11.39g, 93.66mmol) was dissolved in acetonitrile (200mL), and then boric acid (5.79g, 93.66mmol) and potassium carbonate (25.89g, 187.32mmol) were added. The reaction was stirred at 24°C for 12 hours. The suspension was filtered, and the filtrate was spun to dryness to obtain crude product.
  • Step 1 Dissolve 6-chloropyrazolopyridine (1g, 6.51mmol, 1eq) in DMF (10mL), protect it with argon, add NaH (188mg, 7.81mmol, 1.2eq, 60wt%) at -5°C, React for 10 minutes, add SEM-Cl (1.3g, 7.81mmol, 1.2eq) dropwise at -5°C, remove the ice bath after adding, return to room temperature (rt), and react for 3 hours and 10 minutes. Send LCMS to detect the success of the reaction. Pour the reaction solution into an appropriate amount of ice water, extract three times with ethyl acetate (30 mL ⁇ 3), and wash once with saturated aqueous sodium chloride solution (30 mL).
  • Step 2 Combine 6-chloro-1-(2-trimethylsilylethoxy)methylpyrazolopyridine (1.3g, 4.58mmol, 1eq), Pin 2 B 2 (1.4g, 5.50mmol, 1.2 eq), KOAc (899mg, 9.16mmol, 2eq) were dissolved in DMF (20mL), replaced with argon, added Pd(dppf)Cl 2 (335mg, 0.4mmol, 0.1eq), replaced with argon, under argon protection. , stirring reaction at 100°C for 5 hours. Sent LCMS for testing and the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL), and extracted three times with dichloromethane (30 mL ⁇ 3). The organic phase was washed once with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, and spin-dried to obtain intermediate 5 (1.9 g, purity: 77%, yield: 99%, black-gray solid).
  • Step 1 Dissolve 6-chloropyrrole (4.5g, 29.49mmol, 1eq) in DMF (40mL), replace with argon, protect with argon, and add NaH (60wt%) (849mg, 35.39mmol, 1.2) at -5°C. eq), react for 10 minutes, add SEM-Cl (5.9g, 35.39mmol, 1.2eq) dropwise at -5°C, remove the ice bath after adding, return to room temperature (RT), and react for 3 hours. Send LCMS to detect the success of the reaction.
  • reaction solution was poured into ice water, extracted three times with ethyl acetate, washed once with sodium chloride, dried over anhydrous sodium sulfate, spin-dried and purified through column to obtain the product 6-chloro-1-(2-trimethylsilylethane) Oxy)methylpyrrolidine (8g, purity: 80.08%, yield: 76.80%, light yellow liquid).
  • Step 2 Combine 6-chloro-1-(2-trimethylsilylethoxy)methylpyrrolidine (3g, 10.61mmol, 1eq), Pin 2 B 2 (3.2g, 12.73mmol, 1.2eq), KOAc (2.1g, 21.22mmol, 2eq) was dissolved in 1,4-dioxane (30mL), replaced with argon, added Pd(dppf)Cl 2 (776mg, 1.06mmol, 0.1eq), replaced with argon, Under argon protection, the reaction was stirred at 90°C for 5 hours. Sent LCMS for testing and the reaction solution was successful.
  • Step 1 Dissolve 6-bromo-1H-pyrazolopyridine (4.5g, 22.73mmol, 1eq) in DMF (40mL), replace with argon, protect with argon, and add NaH (60wt%) at -5°C ( 654mg, 27.27mmol, 1.2eq), react for 10 minutes, add SEM-Cl (4.5g, 27.27mmol, 1.2eq) dropwise at -5°C, remove the ice bath after adding, return to room temperature, and react for 3 hours. Send LCMS to detect the success of the reaction.
  • reaction solution was poured into ice water, extracted three times with ethyl acetate, washed once with sodium chloride, dried over anhydrous sodium sulfate, spin-dried and purified through column to obtain the product 6-bromo-1-(2-trimethylsilylethane) Oxy)methyl-1H-pyrazolo[4,3-b]pyridine (4.3g, purity: 97.50%, yield: 56.20%, light yellow liquid).
  • Step 2 Combine 6-bromo-1-(2-trimethylsilylethoxy)methyl-1H-pyrazolo[4,3-b]pyridine (4.3g, 13.098mmol, 1eq), Pin 2 B 2 (3.991g, 15.718mmol, 1.2eq) and KOAc (2.571g, 26.196mmol, 2eq) were dissolved in 1,4-dioxane (100mL), replaced with argon, and Pd(dppf)Cl 2 ( 479 mg, 0.655 mmol, 0.05 eq), replaced with argon, stirred and reacted at 120°C for 12 hours under argon protection. Sent LCMS for testing and the reaction solution was successful.
  • Step 1 Dissolve 6-chloro-1H-pyrazole[3,4-b]pyrazine (2g, 13mmol, 1eq) in DMF (50mL), put it in an ice-salt bath (-5°C), and add NaH ( 0.67g, 16.9mmol, 1.3eq, 60wt%), stir for 30 minutes, add SEM-Cl (2.6g, 15.6mmol, 1.2eq) dropwise, protect with argon, return to room temperature, react for 3 hours, add ice water (30mL ), quenched, extracted three times with ethyl acetate (50mL )Methyl)-1h-pyrazoline[3,4-b]pyrazine (2.3g, crude product), brown oily product. The product was directly used in the next reaction.
  • Step 2 Add 6-chloro-1-((2-(trimethylsilyl)ethoxy)methyl)-1h-pyrazoline[3,4-b]pyrazine (1.5g, 5.3mmol, 1eq) was dissolved in 1,4-dioxane (50mL), add Pin 2 B 2 (1.74g, 6.84mmol, 1.3eq), Pd(dppf)Cl 2 ) (384mg, 0.53mmol, 0.1eq), KOAc (1.56g, 15.9mmol, 3eq), replaced with argon three times, heated the oil bath to 120°C and reacted for 12 hours.
  • Step 1 Place the raw material 4-bromopyridine (2g, 12.6mmol, 1eq) in a 250mL single-mouth bottle, add 100mL THF to dissolve, protect with argon, cool to -78°C, and start adding cyclopropylmagnesium bromide ( 40mL, 25.2mmol, 2eq), keep the temperature at -78°C, stir for 5 minutes after the dropwise addition, start to dropwise add phenyl chloroformate (2.4mL, 12.6mmol, 1eq), complete the dropwise addition, react for 10 minutes, return to room temperature, react The solution was diluted with ethyl acetate (150 mL), washed with water (30 mL), 0.1 M (mol/L) hydrochloric acid (30 mL), and brine (30 mL) respectively, and the organic phase was dried and spin-dried to obtain 2.1 g of crude product.
  • cyclopropylmagnesium bromide 40mL, 25.2mmol, 2
  • Step 2 Combine the raw materials 2-cyclopropyl-4-bromopyridine (1.5g, 7.5mmol, 1eq), Pin 2 B 2 (2.3g, 9mmol, 1.2eq), potassium acetate (1.5g, 15mmol, 2eq) ), place it in a 250mL three-necked flask, add 75mL of 1,4-dioxane, replace with argon, add Pd(dppf)Cl 2 (280mg, 0.39mmol, 0.05eq), replace with argon, stir and heat to 120°C. After 12 hours of reaction, LCMS sampling was performed. LCMS showed that the raw material was completely reacted, but LCMS of the product did not show it.
  • Step 1 Dissolve intermediate 1 (400mg, 1.02mmol, 1eq), intermediate 5 (900mg, 3.07mmol, 3eq), K 2 CO 3 (283mg, 2.05mmol, 2eq) in 1,4-dioxane (20 mL) and water (4 mL), replace with argon, add Pd(dppf)Cl 2 (75 mg, 0.10 mmol, 0.1 eq), replace with argon, stir and react at 100°C for 2 hours under argon protection. LCMS detection showed that the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL) and extracted with dichloromethane (20 mL ⁇ 3). The combined organic phases were washed once with saturated sodium chloride solution (20 mL) and dried over anhydrous sodium sulfate. Spin to dryness to obtain 1.2g of crude compound S6-1 (1.2g, purity: 73%).
  • Step 2 Dissolve compound S6-1 (1.2g, 1.45mmol, 1eq) in dichloromethane (50mL), add trifluoroacetic acid (9.4g, 82.7mmol, 57eq), protect with argon, and react at room temperature for 12 hours. Add K 2 CO 3 (1.0g, 7.25mmol, 5eq), methanol (50mL), water (10mL), and stir for 12 hours. Extract with dichloromethane (80mL ⁇ 3). The combined organic phases were washed once with saturated sodium chloride solution (20 mL) and dried over anhydrous sodium sulfate. Spin to dryness and send to preparation and purification to obtain compound S6 (16.31 mg, 0.034 mmol, yield: 2.37%).
  • Step 1 Dissolve intermediate 1 (300mg, 0.77mmol, 1eq), intermediate 6 (673mg, 2.30mmol, 3eq), K 2 CO 3 (212mg, 1.53mmol, 2eq) in 1,4-dioxane (20 mL), add Troubles distilled water (4 mL), replace with argon, add Pd(dppf)Cl 2 (56 mg, 0.077 mmol, 0.1 eq), replace with argon, stir the reaction at 100°C under argon protection 2 Hour. Sent LCMS for testing and the reaction solution was successful.
  • Step 2 Dissolve compound S7-1 (453mg, purity: 76.59%, 0.58mmol, 1eq) in dichloromethane (50mL), add trifluoroacetic acid (3.7g, 32.81mmol, 57eq), protect with argon, room temperature Reaction takes 10 hours. At the end of the reaction, adjust the pH to neutral, spin to dryness, add K 2 CO 3 (398 mg, 2.88 mmol, 5 eq), 50 mL of methanol, and 10 mL of Treconite distilled water, and react for 2 hours.
  • Step 1 Dissolve intermediate 1 (300mg, 0.77mmol, 1eq), intermediate 7 (432mg, 1.15mmol, 1.5eq), K 2 CO 3 (212mg, 1.53mmol, 2eq) in 1,4-dioxane ring (20 mL), add Tribune distilled water (4 mL), replace with argon, add Pd(dppf)Cl 2 (56 mg, 0.077 mmol, 0.1 eq), replace with argon, and stir the reaction at 100°C under argon protection. 2 hours. Sent LCMS for testing and the reaction solution was successful.
  • Step 2 Dissolve compound S8-1 (500mg, purity: 74.49%, 0.617mmol, 1eq) in dichloromethane (50mL), add trifluoroacetic acid (4.0g, 35.16mmol, 57eq), protect with argon, room temperature Reaction takes 10 hours. After the reaction is completed, adjust the pH to neutral, spin to dryness, add K 2 CO 3 (426 mg, 3.08 mmol, 5 eq), 50 mL of methanol, and 10 mL of Trecroft distilled water, and react for 2 hours.
  • Step 1 Dissolve intermediate 1 (500mg, 1.28mmol, 1eq) in 1,4-dioxane (25mL), water (5mL), add intermediate 8 (1.5g, 5.1mmol, 4eq), Pd (dppf)Cl 2 (94.2mg, 0.13mmol, 0.1eq), sodium carbonate (1.25g, 0.77mmol, 3eq), protected by argon, stirred and reacted at 100°C for 2 hours.
  • LCMS monitored product formation. After cooling, the reaction solution was diluted with water (30 mL), extracted with dichloromethane three times (30 mL ⁇ 3), and separated. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude compound S9-1 (1 g).
  • Step 2 Dissolve compound S9-1 (1g, crude product) in DCM (35mL), add trifluoroacetic acid (5mL), and react at room temperature for 10 hours.
  • the reaction solution was concentrated to dryness, the crude product was dissolved in 25 mL of methanol and 5 mL of water, potassium carbonate (1.37 g, 10 mmol, 5 eq) was added, stirred at room temperature for 2 hours, filtered, the filter cake was washed with a small amount of methanol, and the solvent was concentrated under reduced pressure.
  • Step 1 Dissolve compound 1 (100mg, 0.256mmol) in N,N-dimethylformamide (3mL), add zinc cyanide (150mg, 1.28mmol) and 1,1'-bis(diphenyl) respectively.
  • Phosphine) ferrocene (56.7 mg, 0.1 mmol)
  • activated zinc powder (16.6 mg, 0.256 mmol)
  • blow argon gas into the reaction solution for 30 seconds
  • tris(dibenzylideneacetone) dipalladium 46.6 mg, 0.05 mmol
  • Microwave at 125°C for one hour.
  • the reaction solution was brought to room temperature, filtered, and the filter cake was washed twice with dichloromethane.
  • Step 2 Dissolve compound S-34-a (30 mg, 0.079 mmol) and azidotributylstannane (261 mg, 0.79 mmol) in toluene (2 mL), replace the argon gas three times, and react in an oil bath at 120°C for 36 hours.
  • the reaction solution was brought to room temperature, filtered, and the filter cake was washed twice with dichloromethane.
  • the filter cake was dissolved in dimethyl sulfoxide (5 mL) and prepared using alkaline method [NH 4 HCO 3 ] conditions to obtain compound S-34 (1.21 mg, white solid, purity 99%).
  • Test Example 1 Inhibition of Akt S473 phosphorylation level in Raw264.7 cells
  • This experiment uses ELISA method for detection.
  • Raw264.7 cells were purchased from ATCC; mC5a protein was purchased from R&D; DMEM serum-free medium was purchased from Gibco; fetal calf serum (BSA) was purchased from Gibco; RIPA buffer was purchased from CST; protease phosphatase inhibitor was purchased from CST; phosphate
  • the ELISA detection kit for Akt S473 was purchased from R&D; the 96-well plate was purchased from Falcon; the high binding 96-well plate was purchased from Costar; the Infinite M1000 fluorescence microplate reader was purchased from Tecan.
  • Capture Prepare the capture antibody (Capture) solution according to the instructions of the ELISA detection kit for phosphorylated Akt S473 (R&D DYC887BE). Add 100 ⁇ L of the Capture solution per well into a high binding 96-well plate (Costar 42592) and incubate at room temperature overnight. . Wash three times with 300 ⁇ L PBST, add blocking solution (PBS solution containing 1% (w/v) BSA) (Genview FA016-100G), incubate at room temperature for 2 hours, then wash three times with 300 ⁇ L PBST; add 90 ⁇ L control group and blank group respectively.
  • PBS solution containing 1% (w/v) BSA Block FA016-100G
  • Adding DMSO without mC5a protein was the blank group, and adding DMSO and mC5a protein was the control group.
  • Inhibition% [1-(Total mean(experimental group)-total mean(blank group))/(Total mean(control group)-total mean(blank group))] ⁇ 100%; among them, Inhibition%: compound’s effect on Raw264 .7% inhibition of Akt S473 phosphorylation level in cells; total mean: total mean.
  • Test Example 2 Inhibition of Akt S473 phosphorylation levels in T47D and PC3 cells
  • T47D cells were purchased from ATCC; PC3 cells were purchased from SIBS; RPMI 1640 and F12K culture media were purchased from Gibco; phosphorylated Akt S473 (Ser473-p-Akt) antibody was purchased from CST; 96-well plates were purchased from Falcon; Infinite M1000 fluorescent enzyme label The instrument was purchased from Tecan; YZJ-0673 is compound S-37 in patent WO2016095833.
  • T47D (PI3K ⁇ ) (culture conditions: RPMI 1640+10% FBS (v/v)) and PC3 (PI3K ⁇ ) (culture conditions: F12K + 10% FBS (v/v)) cells in the logarithmic growth phase and incubate them at 2.2 ⁇ 90 ⁇ L of a concentration of 10 5 cells/mL (PC3) or 4.4 ⁇ 10 5 cells/mL (T47D) was plated in a 96-well plate (Falcon 353072), cultured at 37°C, 5% CO 2 , and left overnight until the cells were attached.
  • PC3 10 5 cells/mL
  • the control group, background group and experimental group respectively took out the Triton X-100 solution, washed it twice with 200 ⁇ L PBS (300 rpm, 1 minute each time), and added blocking solution (PBS solution containing 1% (w/v) BSA) (Genview FA016-100G), incubate at room temperature for 2 hours, take it out, wash once with 200 ⁇ L PBS (300 rpm, 1 minute), add 30 ⁇ L Ser473-p-Akt antibody 0.1% (w/v) BSA dilution (cell signaling 4060L), 4°C Incubate overnight.
  • PBS solution containing 1% (w/v) BSA Genview FA016-100G
  • the compound of the present application has high inhibitory activity on the phosphorylation level of Akt S473 in Raw264.7 cells and low inhibitory activity on T47D and PC3 cells, indicating that the compound of the present application has high inhibitory activity on PI3K ⁇ . Select inhibitory activity.
  • Test Example 3 Determination of PI3K kinase inhibitory activity
  • Human PI3K ⁇ and PI3K ⁇ kinase were purchased from Millipore, human PI3K ⁇ kinase was purchased from Invitrogen, human PI3K ⁇ kinase was purchased from Eurofins, ADP-Glo kinase detection kit was purchased from Promega, and PIP2 lipid substrate was purchased from Life Technologies, 384-well plate Purchased from PE.
  • This experiment uses the ADP-Glo method to detect the inhibition of the activities of PI3K ⁇ , PI3K ⁇ , PI3K ⁇ and PI3K ⁇ by compounds.
  • the reagents used are PI3K ⁇ (Millipore 14-558), PI3K ⁇ (Invitrogen PV4788), PI3K ⁇ (Eurofins 14-603M), PI3K ⁇ (Millipore 14-604M), PIP2 (Life Technologies PR8982B), ADP-Glo (Promega V9102/3).
  • the inhibitory effect of the test compounds on PI3K kinase activity in Table 3 was determined by the following method.
  • the test compounds were prepared with 100% DMSO into a 100 ⁇ DMSO solution with the maximum concentration required for the experiment, and then 3-fold gradient dilution was performed with 100% DMSO. There are 10 concentration points in total. Add 50nL of the prepared test compound solution to the 384-well plate (PE 6008289), and add DMSO to the blank wells and control wells.
  • Inhibition rate % [1-(average value (experimental group)-average value (blank group))/(average value (control group)-average value (blank group))] ⁇ 100%
  • Table 3 IC 50 value of compounds inhibiting PI3K ⁇ kinase activity
  • Table 4 IC 50 values of compounds inhibiting PI3K ⁇ , PI3K ⁇ and PI3K ⁇ kinase activity
  • the LC/MS/MS method was used to determine the drug concentration in the plasma of mice at different times after intravenous injection and oral gavage administration of the compound of the present application.
  • the known compound D1 and the compounds of the present application S11, S5 and S18 were For example, study the pharmacokinetic behavior of the compound of the present application in mice and evaluate its pharmacokinetic characteristics.
  • compound D1 (CAS No. 2504036-13-7) is as follows, which can be prepared by referring to existing literature or purchased commercially.
  • mice in the intravenous injection group can drink water and eat freely; mice in the intragastric administration group are fasted overnight, and can drink water and eat freely 4 hours after administration) ;
  • Dosing method and dosage Select the drug that meets the experimental requirements before administration (qualifies for quarantine, passes the laboratory adaptation period, and has a weight that meets the requirements of the experiment) Animals that meet the requirements are weighed and marked. ICR mice were administered via tail vein, the dosage was 2 mg/kg, the solvent was 5% DMSO + 30% (10% Solutol HS15) + 65% (20% Captisol, pH 7.4), v/v/v; administered by gavage The dosage is 10 mg/kg, the solvent is 5% DMSO + 30% (10% Solutol HS15) + 65% (20% Captisol, pH 7.4), v/v/v.
  • Blood sample collection Before collecting blood samples, bind the mice, and each administered mouse will collect blood at predetermined blood collection time points (intravenous administration: respectively at 0.083, 0.25, 0.5, 1, 2, 4, Collect blood at 7 and 24 hours, a total of 8 time points; intragastric administration: Collect blood at 0.083, 0.25, 0.5, 1, 2, 4, 7, and 24 hours after administration, a total of 8 time points), through the canthus vein Collect approximately 100 ⁇ L of blood. Transfer the whole blood to a 1.5 mL test tube pre-added with the anticoagulant EDTA-K 2 , centrifuge for 6 minutes (8000 rpm, 4°C), and separate the plasma. The entire process is completed within 15 minutes after blood collection. All samples need to be stored in a -20°C refrigerator until sample analysis.
  • the chromatographic column is: Waters XBridge-C18 (2.1 ⁇ 50mm, 3.5 ⁇ m), mobile phase A: containing Aqueous solution of 0.05% formic acid and 5mmol/L ammonium acetate (v/v), mobile phase B: methanol solution (v/v) containing 0.05% formic acid and 5mmol/L ammonium acetate, the flow rate is 0.6mL/min, and the column temperature is 40°C, the injection volume is 2 ⁇ L, and the liquid phase gradient is:
  • Mass spectrometry conditions are: scan mode: multiple reaction ion monitoring (positive ion mode);
  • Ion source turbine spray
  • ionization mode electrospray ionization.
  • C max in Table 5 is the peak concentration
  • AUC 0-t is the area under the plasma concentration-time curve from time 0 to the final quantifiable time point
  • F is the oral bioavailability
  • Test Example 5 Pharmacodynamic test in a tumor model in which mouse colon cancer cells MC38 and in vitro differentiated M2 macrophages were mixed subcutaneously and transplanted
  • the anti-tumor effect of the compound of the present application was evaluated in a tumor model in which mouse colon cancer cell MC38 cells and macrophages cultured in vitro were mixed subcutaneously and transplanted.
  • mice C57BL/6 mice, female, 8-9 weeks old (the age of mice when tumor cells were inoculated), weight 18.0-20.0g, 8 mice in each group. Purchased from Shanghai Jihui Experimental Animal Breeding Co., Ltd.
  • Mouse colon cancer cell MC38 (purchased from Shanghai Jiao Tong University) was cultured in a medium containing 10% (v/v) fetal calf serum (Gibco; 10099-141C), 100 U/mL penicillin and 100 pg/mL streptomycin (Gibco; 15140122). in RPMI-1640 culture medium (Gibco; 61870-036). MC38 cells in the exponential growth phase were collected and resuspended in PBS to a suitable concentration for subcutaneous tumor inoculation in mice.
  • Bone marrow cells were isolated from the leg bones of C57BL/6 mice, prepared into single cell suspension, and added with 50ng/mL M-CSF (Peprotech; 315-02) to induce bone marrow-derived M0 macrophages; in M0 macrophage induction During the process, 20ng/mL interleukin 4 (IL4) (Peprotech; 214-14) was added and cultured for 48 hours to obtain polarized M2 macrophages; M0 macrophages and M2 macrophages cultured in vitro were closely related to mouse colon cancer.
  • the cell line MC38 was mixed at a ratio of 1:1 (number) and inoculated subcutaneously, with a total of 10 6 cells/mouse.
  • mice in the control group were administered with vehicle as the vehicle group, and the administration volume was 10 ⁇ L/g; the mice in the M0 group were administered with vehicle as the vehicle group mixed with M0 macrophages.
  • the drug volume is 10 ⁇ L/g; 8 mice in the M2 group are randomly selected to be gavaged with the vehicle as the vehicle group mixed with M2 macrophages, and the dosage volume is 10 ⁇ L/g; 8 mice in the M2 group are randomly selected to be gavaged with 1 mg/mL of the drug.
  • the compound S11 administration preparation was used as the compound S11 group mixed with M2 macrophages, and the dosage was 10 mg/kg; the mice in the remaining M2 group were administered 1 mg/mL compound D1 administration preparation as the compound D1 mixed with M2 macrophages.
  • the dosage was 10 mg/kg.
  • Routine monitoring includes observing the impact of tumor growth and treatment on the normal behavior of animals, including the activity of experimental animals, food and water intake, weight gain or loss, eyes, coat and other abnormalities. Weigh and measure the tumor volume twice a week. The dosing period is 27 days. On the 27th day, the body weight is measured and the tumor volume is measured. The mice are then killed and the tumor block is taken out and weighed (TW). The tumor volume (TV) and relative tumor weight are calculated. Growth rate (T/C) and tumor growth inhibition rate (tumor weight) (TGI) were analyzed statistically.
  • T/C (%) T/C ⁇ 100%
  • TGI (%) (1-T/C) ⁇ 100%
  • T and C are the treatment groups respectively (compound S11 group mixed with M2 macrophages or The relative tumor volume or weight at a specific time point between the compound D1 group mixed with M2 macrophages) and the control group (vehicle group).
  • the mouse tumor growth curve is shown in Figure 1.
  • vehicle vehicle group
  • M0+ vehicle vehicle group mixed with M0 macrophages
  • M2+ vehicle vehicle group mixed with M2 macrophages
  • M2+ compound D1 compound mixed with M2 macrophages D1 group, 10 mg/kg (mpk), administered by gavage, once a day
  • M2+ compound S11 compound S11 group mixed with M2 macrophages, 10 mg/kg, administered by gavage, once a day.
  • **, P ⁇ 0.01; ***, P ⁇ 0.001; ns, P>0.05. (n 8, mean ⁇ standard error).
  • mice The tumor weight analysis of mice in each group of the model on the 27th day after administration is shown in Figure 2.
  • vehicle vehicle group
  • M0+ vehicle vehicle group mixed with M0 macrophages
  • M2+ vehicle vehicle group mixed with M2 macrophages
  • M2+ compound D1 compound mixed with M2 macrophages D1 group, 10 mg/kg, administered by gavage, once a day
  • M2+ compound S11 compound S11 group mixed with M2 macrophages, 10 mg/kg, administered by gavage, once a day.
  • **, P ⁇ 0.01; ns, P>0.05. (n 8, mean ⁇ standard error).
  • T/C in Table 6 indicates relative tumor weight growth rate
  • TGI tumor growth inhibition rate (tumor weight)
  • ⁇ indicates no corresponding parameter

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Abstract

The present application relates to a heteroaryl-substituted pyridopyrrolidone derivative, and a pharmaceutical composition and the use thereof. The heteroaryl-substituted pyridopyrrolidone derivative has a structure as shown in formula (I). The compound of the present application has a relatively high selective inhibitory activity on PI3Kγ and an improved in-vivo pharmacokinetic activity, and has practical value.

Description

杂芳基取代的吡啶并吡咯酮衍生物、其药物组合物及应用Heteroaryl-substituted pyridopyrrolone derivatives, pharmaceutical compositions and applications thereof
本申请要求于2022年09月01日提交中国专利局、申请号为CN202211062585.0发明名称为“杂芳基取代的吡啶并吡咯酮衍生物、其药物组合物及应用”的中国专利申请的优先权，其全部内容通过引用结合在本申请中。This application requires priority for the Chinese patent application with the application number CN202211062585.0 and the invention name is "Heteroaryl-substituted pyridopyrrolone derivatives, pharmaceutical compositions and applications thereof" submitted to the China Patent Office on September 1, 2022. rights, the entire contents of which are incorporated herein by reference.
技术领域Technical field
本申请涉及医药技术领域，特别涉及一种杂芳基取代的吡啶并吡咯酮衍生物、其药物组合物及应用。The present application relates to the field of medical technology, and in particular to a heteroaryl-substituted pyridopyrrolone derivative, its pharmaceutical composition and application.
背景技术Background technique
磷酸肌醇3-激酶(PI3K)是一类具有磷酸肌醇肌醇环磷酸化功能的磷脂酰激酶，其被分成I型、II型和III型三类，其中I类PI3K的研究最为广泛。该类型进一步被分为两个亚组(IA和IB)。IA类PI3K由三个密切相关的激酶PI3Kα、PI3Kβ及PI3Kδ，现存为由催化次单元(p110α、p110β、或p110δ)所组成之异二聚体)及数个调节次单元之一者所组成。PI3Kα和PI3Kβ被广泛表达并在细胞生长、分裂和存活中发挥作用(Thomas M,et al.,Curr.Opin.Pharmacol.,2008；8:267-274)。这两种激酶在许多生物学功能中的作用在缺乏PI3Kα或PI3Kβ的小鼠中观察到的胚胎致死性被增强。由于它们在体内平衡中的作用，PI3Kα和PI3Kβ的临床评价受限于肿瘤学领域，并且一些化合物也处于临床开发的不同阶段。PI3Kγ单一类别1B异构体，主要响应于G蛋白偶联受体(G-protein coupled receptors,GPCR)，且由p110γ催化次单元及两个不同的调节次单元之一者所组成。PI3Kγ亚型在免疫细胞中表达，并且在正常或恶性的上皮细胞和结缔组织细胞中具有有限的表达。PI3Kγ敲除小鼠的研究结果表明，PI3Kγ对于细胞活化和一些趋化因子的迁移是重要的(Sasaki T.,et al.,Science,2000；287:1040-1046；Hirsch E.,et al.,Science,2000；287:1049-1053)。PI3Kγ信号传导对骨髓细胞功能特别重要，Camps等人描述了用选择性PI3Kγ抑制剂AS-60485023治疗可在两种相异的类风湿性关节炎小鼠模型中抑制关节发炎及损伤之进展(Camps M,et al.,Nat.Med.2005,11,936-943)。基于在各种疾病模型中观察到的细胞水平及功效的研究，可潜在地使用PI3Kγ抑制剂来治疗各种疾病，例如发炎、代谢、癌症(Cushing,T.D.,et al.，J.Med.Chem.2012,55,8559-8581；Ruckle,T.,et al.，Nat.Rev.Drug Discovery 2006,5,903-918；Stark,A.K.,et al.，Curr.Opin.Pharmacol.2015,23,82-91)。Phosphoinositide 3-kinase (PI3K) is a type of phosphatidyl kinase with the function of phosphorylating the phosphoinositide ring. It is divided into three categories: type I, type II and type III. Among them, type I PI3K is the most widely studied. This type is further divided into two subgroups (IA and IB). Class IA PI3K consists of three closely related kinases, PI3Kα, PI3Kβ, and PI3Kδ, now present as a heterodimer composed of a catalytic subunit (p110α, p110β, or p110δ) and one of several regulatory subunits. PI3Kα and PI3Kβ are widely expressed and play roles in cell growth, division and survival (Thomas M, et al., Curr. Opin. Pharmacol., 2008; 8:267-274). The roles of these two kinases in many biological functions are enhanced by the embryonic lethality observed in mice lacking PI3Kα or PI3Kβ. Due to their role in homeostasis, clinical evaluation of PI3Kα and PI3Kβ has been limited to the field of oncology, and some compounds are also in various stages of clinical development. The single class 1B isoform of PI3Kγ mainly responds to G-protein coupled receptors (GPCR) and is composed of the p110γ catalytic subunit and one of two different regulatory subunits. PI3Kγ isoforms are expressed in immune cells and have limited expression in normal or malignant epithelial and connective tissue cells. Research results on PI3Kγ knockout mice indicate that PI3Kγ is important for cell activation and the migration of some chemokines (Sasaki T., et al., Science, 2000; 287:1040-1046; Hirsch E., et al. ,Science,2000;287:1049-1053). PI3Kγ signaling is particularly important for myeloid cell function, and Camps et al. described that treatment with the selective PI3Kγ inhibitor AS-60485023 inhibited the progression of joint inflammation and damage in two different mouse models of rheumatoid arthritis (Camps M, et al., Nat. Med. 2005, 11, 936-943). Based on studies at the cellular level and efficacy observed in various disease models, PI3Kγ inhibitors can potentially be used to treat various diseases, such as inflammation, metabolism, and cancer (Cushing, T.D., et al., J. Med. Chem .2012,55,8559-8581; Ruckle,T.,et al., Nat.Rev.Drug Discovery 2006,5,903-918; Stark,A.K.,et al., Curr.Opin.Pharmacol.2015,23,82- 91).
尽管目前已有多篇文献报道了选择性PI3Kγ抑制剂，如WO2017153527、WO2020210379，然而这些PI3Kγ抑制剂的选择性、药代动力学参数等性质还有待进一步提高，因此本技术领域还需要开发具有更加优异的选择性以及药代动力学性质等适合后期开发的PI3Kγ抑制剂以更好地用于临床需求。Although there have been many reports on selective PI3Kγ inhibitors, such as WO2017153527 and WO2020210379, the selectivity, pharmacokinetic parameters and other properties of these PI3Kγ inhibitors need to be further improved. Therefore, this technical field still needs to develop more effective PI3Kγ inhibitors. The excellent selectivity and pharmacokinetic properties are suitable for late-stage development of PI3Kγ inhibitors to better meet clinical needs.
发明内容Contents of the invention
本申请的目的是提供一种选择性好、药代动力学性质好的杂芳基取代的吡啶并吡咯酮衍生物。The purpose of this application is to provide a heteroaryl-substituted pyridopyrrolone derivative with good selectivity and good pharmacokinetic properties.
本申请第一方面提供了一种式(I)所示的化合物、或其药学上可接受的盐、或其立体异构体：
The first aspect of this application provides a compound represented by formula (I), or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof:
其中，in,
环A为5或6元杂芳基环；Ring A is a 5- or 6-membered heteroaryl ring;
(R3)n表示环A上的氢被n个R3取代，n为0、1、2、3或4；每个R3相同或不同，各自独立地为氘、卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、NRa1Rb1、-N(Ra3)-C(O)C1-3烷基、-N(Ra3)-C(O)-氘代C1-3烷基、-N(Ra3)-C(O)OC1-3烷基、-SO2C1-3烷基、-SO2C3-6环烷基、-C(O)NRa1Rb1、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基、3至6元杂环烷基、苯基或5至6元杂芳基；其中所述3至6元杂环烷基、苯基、5至6元杂芳基各自独立地为未取代的或被1、2或3个各自独立地选自取代基组Q的取代基取代；(R 3 ) n means that the hydrogen on ring A is replaced by n R 3s , n is 0, 1, 2, 3 or 4; each R 3 is the same or different, and each R 3 is independently deuterium, halogen, cyano, or hydroxyl , carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halo C 1-3 alkyl, halo C 1-3 alkoxy , NR a1 R b1 , -N(R a3 )-C(O)C 1-3 alkyl, -N(R a3 )-C(O)-deuterated C 1-3 alkyl, -N(R a3 )-C(O)OC 1-3 alkyl, -SO 2 C 1-3 alkyl, -SO 2 C 3-6 cycloalkyl, -C(O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy, 3 to 6 membered heterocycloalkyl, phenyl or 5 to 6-membered heteroaryl; wherein the 3- to 6-membered heterocycloalkyl, phenyl, and 5- to 6-membered heteroaryl groups are each independently unsubstituted or 1, 2, or 3 are independently selected from substituents Substituent substitution of group Q;
Z为N或CRZ；T为N或CRT；U为N或CRU；W为N或CRW；Y为N或CRY；Z is N or CR Z ; T is N or CR T ; U is N or CR U ; W is N or CR W ; Y is N or CR Y ;
RZ、RT、RU各自独立地为氢、卤素、NRa1Rb1或C1-8烷基；其中，所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基；R Z , RT , and RU are each independently hydrogen, halogen, NR a1 R b1 or C 1-8 alkyl; wherein, the C 1-8 alkyl is preferably C 1-6 alkyl, and more preferably C 1-3 alkyl;
RW、RY各自独立地为氢、卤素、C1-8烷基、卤代C1-8烷基、卤代C1-8烷氧基、NRa1Rb1或C3-6环烷基；其中，所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基；R W and R Y are each independently hydrogen, halogen, C 1-8 alkyl, halogenated C 1-8 alkyl, halogenated C 1-8 alkoxy, NR a1 R b1 or C 3-6 cycloalkyl base; wherein, the C 1-8 alkyl group is preferably a C 1-6 alkyl group, and more preferably a C 1-3 alkyl group;
或者RW、RY相连，与相邻6元环形成9或10元杂芳基环、9或10元苯基并杂环烷基环或9或10元杂芳基并杂环烷基环；所述9或10元杂芳基环、9或10元苯基并杂环烷基环、9或10元杂芳基并杂环烷基环各自独立地为未取代的或被1、2或3个各自独立地选自取代基组Q的取代基取代；Or R W and R Y are connected to form a 9- or 10-membered heteroaryl ring, a 9- or 10-membered phenyl heterocycloalkyl ring, or a 9- or 10-membered heteroaryl heterocycloalkyl ring with the adjacent 6-membered ring. ; The 9- or 10-membered heteroaryl ring, the 9- or 10-membered phenyl heterocycloalkyl ring, and the 9- or 10-membered heteroaryl heterocycloalkyl ring are each independently unsubstituted or substituted by 1 or 2 Or substituted by 3 substituents each independently selected from the substituent group Q;
R1、R2各自独立地为C1-8烷基、卤代C1-8烷基或C3-6环烷基；其中，所述C3-6环烷基为未取代的或被1或2个各自独立地选自卤素和C1-3烷基的取代基取代；所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基；所述卤代C1-8烷基优选为卤代C1-6烷基，更优选为卤代C1-3烷基；R 1 and R 2 are each independently C 1-8 alkyl, halogenated C 1-8 alkyl or C 3-6 cycloalkyl; wherein, the C 3-6 cycloalkyl is unsubstituted or substituted 1 or 2 substituents each independently selected from halogen and C 1-3 alkyl are substituted; the C 1-8 alkyl is preferably C 1-6 alkyl, more preferably C 1-3 alkyl; so The halogenated C 1-8 alkyl group is preferably a halogenated C 1-6 alkyl group, and more preferably a halogenated C 1-3 alkyl group;
取代基组Q选自卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、NRa1Rb1、-SO2C1-3烷基、-S(O)C1-3烷基、-C(O)NRa1Rb1、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基、3至6元杂环烷基、苯基或5至6元杂芳基；The substituent group Q is selected from halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halogenated C 1-3 Alkyl group, halogenated C 1-3 alkoxy group, NR a1 R b1 , -SO 2 C 1-3 alkyl group, -S(O)C 1-3 alkyl group, -C(O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy, 3 to 6-membered heterocycloalkyl , phenyl or 5 to 6-membered heteroaryl;
Ra1、Rb1各自独立地为氢、C1-3烷基或乙酰基，或Ra1、Rb1与相连的氮原子共同形成4至6元饱和单杂环；所述4至6元饱和单杂环为未取代的或被1、2或3个各自独立地选自下述的取代基取代：氘、卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、-SO2C1-3烷基、-S(O)C1-3烷基、-C(O)NH2、-C(O)NH(C1-3烷基)、-C(O)N(C1-3烷基)2、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基或3至6元杂环烷基；R a1 and R b1 are each independently hydrogen, C 1-3 alkyl or acetyl, or R a1 , R b1 and the connected nitrogen atom together form a 4- to 6-membered saturated monoheterocyclic ring; the 4- to 6-membered saturated The monoheterocycle is unsubstituted or substituted by 1, 2 or 3 substituents each independently selected from the following: deuterium, halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkyl Oxygen group, C 2-4 alkenyl group, C 2-4 alkynyl group, halogenated C 1-3 alkyl group, halogenated C 1-3 alkoxy group, -SO 2 C 1-3 alkyl group, -S(O )C 1-3 alkyl, -C(O)NH 2 , -C(O)NH(C 1-3 alkyl), -C(O)N(C 1-3 alkyl) 2 , -C( O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy or 3 to 6-membered heterocycloalkyl;
Ra3为氢或C1-8烷基；其中所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基。R a3 is hydrogen or C 1-8 alkyl; wherein the C 1-8 alkyl is preferably C 1-6 alkyl, more preferably C 1-3 alkyl.
本申请的一种实施方案中，T、W、Y、U中的一个为N。In one embodiment of the present application, one of T, W, Y, and U is N.
本申请的一种实施方案中，T为CRT；U为CRU；W为N或CRW；Y为N或CRY；且W、Y不同时为N。 In one embodiment of the present application, T is CRT ; U is CRU ; W is N or CRW ; Y is N or CRY ; and W and Y are not N at the same time.
本申请的一种实施方案中，T、W、Y、U中的两个为N。In one embodiment of the present application, two of T, W, Y, and U are N.
本申请的一种实施方案中，T为N；U为CRU；W为CRW；Y为N。In one embodiment of the present application, T is N; U is CRU ; W is CRW ; Y is N.
本申请的一种实施方案中，为选自式(A)、式(B)和式(C)所示结构：
In one embodiment of the present application, The structure is selected from the group consisting of formula (A), formula (B) and formula (C):
本申请的一种实施方案中，所述9或10元杂芳基环选自：吲哚基、异吲哚基、吲唑基、苯并三唑基、苯并噻吩基、异苯并噻吩基、苯并呋喃基、苯并异呋喃基、苯并咪唑基、苯并噁唑基、苯并异噁唑基、苯并噁二唑基、苯并噻唑基、苯并异噻唑基、苯并噻二唑基、茚嗪基、嘌呤基、吡啶并[3,2-d]嘧啶基、吡啶并[2,3-d]嘧啶基、吡啶并[3,4-d]嘧啶基、吡啶并[4,3-d]嘧啶基、1,8-萘啶基、1,7-萘啶基、1,6-萘啶基、1,5-萘啶基、喋啶基、喹啉基、异喹啉基、噌琳基、喹喔啉基、酞嗪基或喹唑啉基。In one embodiment of the present application, the 9- or 10-membered heteroaryl ring is selected from: indolyl, isoindolyl, indazolyl, benzotriazolyl, benzothienyl, isobenzothiophene base, benzofuryl, benzisofuryl, benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzoxadiazolyl, benzothiazolyl, benzisothiazolyl, benzene Thiadiazolyl, indanazinyl, purinyl, pyrido[3,2-d]pyrimidinyl, pyrido[2,3-d]pyrimidinyl, pyrido[3,4-d]pyrimidinyl, pyridine And[4,3-d]pyrimidinyl, 1,8-naphthyridinyl, 1,7-naphthyridinyl, 1,6-naphthyridinyl, 1,5-naphthyridinyl, pyridinyl, quinolinyl , isoquinolinyl, quinolinyl, quinoxalinyl, phthalazinyl or quinazolinyl.
本申请的一种实施方案中，所述9或10元苯基并杂环烷基环、9或10元杂芳基并杂环烷基环中的杂环烷基各自独立地选自四氢呋喃基、四氢噻吩基、四氢吡咯基、噁唑烷基、二氧戊环基、哌啶基、哌嗪基、吗啉基、二氧六环基、硫代吗啉基、硫代吗啉-1,1-二氧化物、四氢吡喃基、氮杂环丁烷-2-酮基、氧杂环丁烷-2-酮基、二氢呋喃-2(3H)-酮基、吡咯烷-2-酮基、吡咯烷-2,5-二酮基、二氢呋喃-2,5-二酮基、哌啶-2-酮基、四氢-2H-吡喃-2-酮基、哌嗪-2-酮基、吗啉-3-酮基。In one embodiment of the present application, the heterocycloalkyl groups in the 9- or 10-membered phenylheterocycloalkyl ring and the 9- or 10-membered heteroarylheterocycloalkyl ring are each independently selected from tetrahydrofuranyl. , tetrahydrothienyl, tetrahydropyrrolyl, oxazolidinyl, dioxolanyl, piperidyl, piperazinyl, morpholinyl, dioxane, thiomorpholinyl, thiomorpholine -1,1-dioxide, tetrahydropyranyl, azetidine-2-one, oxetane-2-one, dihydrofuran-2(3H)-one, pyrrole Alk-2-one group, pyrrolidine-2,5-dione group, dihydrofuran-2,5-dione group, piperidin-2-one group, tetrahydro-2H-pyran-2-one group , piperazine-2-one group, morpholin-3-one group.
本申请的一种实施方案中，所述9或10元杂芳基并杂环烷基环中的杂芳基选自吡啶基、哒嗪基、嘧啶基、吡嗪基。In one embodiment of the present application, the heteroaryl group in the 9- or 10-membered heteroaryl heterocycloalkyl ring is selected from the group consisting of pyridyl, pyridazinyl, pyrimidinyl, and pyrazinyl.
本申请的一种实施方案中，式(A)中，RT为氢；RU为氢；RW为氢、卤素、C1-6烷基、卤代C1-6烷基或卤代C1-6烷氧基。In one embodiment of the present application, in formula (A), R T is hydrogen; R U is hydrogen; R W is hydrogen, halogen, C 1-6 alkyl, halo C 1-6 alkyl or halo C 1-6 alkoxy.
本申请的一种实施方案中，式(A)中，RT为氢；RU为氢；RW为氢、氟、氯、溴、甲基、乙基、正丙基、异丙基、三氟甲基或三氟甲氧基。In one embodiment of the present application, in formula (A), R T is hydrogen; R U is hydrogen; R W is hydrogen, fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, Trifluoromethyl or trifluoromethoxy.
本申请的一种实施方案中，式(B)中，RT为氢；RU为氢；RY为氢、卤素、C1-6烷基、卤代C1-6烷基、卤代C1-6烷氧基或C3-6环烷基。In one embodiment of the present application, in formula (B), R T is hydrogen; R U is hydrogen; R Y is hydrogen, halogen, C 1-6 alkyl, halo C 1-6 alkyl, halo C 1-6 alkoxy or C 3-6 cycloalkyl.
本申请的一种实施方案中，式(B)中，RT为氢；RU为氢；RY为氟、氯、溴、甲基、乙基、正丙基、异丙基、三氟甲基、三氟甲氧基、环丙基、环丁基、环戊基或环己基。In one embodiment of the present application, in formula (B), R T is hydrogen; R U is hydrogen; R Y is fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, trifluoro Methyl, trifluoromethoxy, cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl.
本申请的一种实施方案中，结构选自以下结构中的一种：
In one embodiment of the present application, the structure Choose one of the following structures:
本申请的一种实施方案中，环A选自下组的5或6元杂芳基环：噻吩环、呋喃环、噻唑环、异噻唑环、咪唑环、噁唑环、吡咯环、吡唑环、1,2,3-三唑环、1,2,4-三唑环、1,2,5-三唑环、1,3,4-三唑环、四唑环、异噁唑环、1,2,3-噁二唑环、1,2,4-噁二唑环、1,2,5-噁二唑环、1,3,4-噁二唑环、噻二唑环、吡啶环、哒嗪环、嘧啶环、吡嗪环、三嗪环或四嗪环。In one embodiment of the present application, Ring A is a 5- or 6-membered heteroaryl ring selected from the following group: thiophene ring, furan ring, thiazole ring, isothiazole ring, imidazole ring, oxazole ring, pyrrole ring, pyrazole Ring, 1,2,3-triazole ring, 1,2,4-triazole ring, 1,2,5-triazole ring, 1,3,4-triazole ring, tetrazole ring, isoxazole ring , 1,2,3-oxadiazole ring, 1,2,4-oxadiazole ring, 1,2,5-oxadiazole ring, 1,3,4-oxadiazole ring, thiadiazole ring, Pyridine ring, pyridazine ring, pyrimidine ring, pyrazine ring, triazine ring or tetrazine ring.
优选地，环A为噻唑环。Preferably, Ring A is a thiazole ring.
优选地，n为2。Preferably, n is 2.
本申请的一种实施方案中，每个R3相同或不同，各自独立地为C1-3烷基、-NH-C(O)C1-3烷基、-NH-C(O)-氘代C1-3烷基或-NH-C(O)OC1-3烷基。In one embodiment of the application, each R 3 is the same or different, and each is independently C 1-3 alkyl, -NH-C(O)C 1-3 alkyl, -NH-C(O)- Deuterated C 1-3 alkyl or -NH-C(O)OC 1-3 alkyl.
本申请的一种实施方案中，式(I)中，为式(a)所示结构：
In one embodiment of the present application, in formula (I), It is the structure shown in formula (a):
R01、R02各自独立地为氘、卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、NRa1Rb1、-N(Ra3)-C(O)C1-3烷基、-N(Ra3)-C(O)- 氘代C1-3烷基、-N(Ra3)-C(O)OC1-3烷基、-SO2C1-3烷基、-SO2C3-6环烷基、-C(O)NRa1Rb1、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基、3至6元杂环烷基、苯基或5至6元杂芳基；其中所述3至6元杂环烷基、苯基、5至6元杂芳基各自独立地为未取代的或被1、2或3个各自独立地选自取代基组Q的取代基取代。R 01 and R 02 are each independently deuterium, halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halogen Substituted C 1-3 alkyl, halogenated C 1-3 alkoxy, NR a1 R b1 , -N(R a3 )-C(O)C 1-3 alkyl, -N(R a3 )-C( O)- Deuterated C 1-3 alkyl, -N(R a3 )-C(O)OC 1-3 alkyl, -SO 2 C 1-3 alkyl, -SO 2 C 3-6 cycloalkyl, -C (O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy, 3 to 6 membered heterocycloalkyl, phenyl or 5 to 6 membered heteroaryl; wherein the 3 to 6 membered heterocycloalkyl, phenyl, 5 to 6 membered heteroaryl are each independently unsubstituted or Substituted by 1, 2 or 3 substituents each independently selected from the substituent group Q.
本申请的一种实施方案中，R01为卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、卤代C1-3烷基或卤代C1-3烷氧基；R02为-NH-C(O)C1-3烷基、-NH-C(O)-氘代C1-3烷基或-NH-C(O)OC1-3烷基。In one embodiment of the present application, R 01 is halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, halo C 1-3 alkyl or halo C 1- 3 alkoxy; R 02 is -NH-C(O)C 1-3 alkyl, -NH-C(O)-deuterated C 1-3 alkyl or -NH-C(O)OC 1-3 alkyl.
优选地，R01为甲基、乙基、正丙基或异丙基。进一步优选地，R01为甲基。Preferably, R 01 is methyl, ethyl, n-propyl or isopropyl. Further preferably, R 01 is methyl.
优选地，R02为-NH-C(O)C1-3烷基、-NH-C(O)-氘代C1-3烷基或-NH-C(O)OC1-3烷基。进一步优选地，R02为-NH-C(O)CH3、-NH-C(O)-CD3或-NH-C(O)OCH3。Preferably, R 02 is -NH-C(O)C 1-3 alkyl, -NH-C(O)-deuterated C 1-3 alkyl or -NH-C(O)OC 1-3 alkyl . Further preferably, R 02 is -NH-C(O)CH 3 , -NH-C(O)-CD 3 or -NH-C(O)OCH 3 .
本申请的一种实施方案中，R1为一氯甲基、二氯甲基、三氯甲基、一氯乙基、1,2-二氯乙基、三氯乙基、一溴乙基、一氟甲基、二氟甲基、三氟甲基、一氟乙基、二氟乙基、三氟乙基、环丙基、环丁基、环戊基、环己基、一氟取代的环丙基或一氟取代的环丁基；R2为甲基、乙基、正丙基、异丙基、环丙基、环丁基、环戊基、环己基、一氟取代的环丙基或一氟取代的环丁基。In one embodiment of the application, R 1 is monochloromethyl, dichloromethyl, trichloromethyl, monochloroethyl, 1,2-dichloroethyl, trichloroethyl, monobromoethyl , monofluoromethyl, difluoromethyl, trifluoromethyl, monofluoroethyl, difluoroethyl, trifluoroethyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, monofluoro-substituted Cyclopropyl or monofluoro-substituted cyclobutyl; R 2 is methyl, ethyl, n-propyl, isopropyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, monofluoro-substituted cyclopropyl group or a fluorine-substituted cyclobutyl group.
优选地，R1为环丙基、环丁基或三氟甲基。Preferably, R1 is cyclopropyl, cyclobutyl or trifluoromethyl.
优选地，R2为甲基、乙基、环丙基或环丁基。Preferably, R2 is methyl, ethyl, cyclopropyl or cyclobutyl.
本申请的一种实施方案中，式(I)中，选自以下结构中的一种：
In one embodiment of the present application, in formula (I), Choose one of the following structures:
本申请的一种实施方案中，式(I)中，选自以下结构中的一种：
In one embodiment of the present application, in formula (I), Choose one of the following structures:
本申请的一种实施方案中，Z为N或CH。In one embodiment of the present application, Z is N or CH.
本申请的一种实施方案中，上述各结构式中、基团R3和取代基组Q中所述的5或6元杂芳基(环)各自独立地选自：噻吩基、呋喃基、噻唑基、异噻唑基、咪唑基、噁唑基、吡咯基、吡唑基、三唑基、1,2,3-三唑基、1,2,4-三唑基、1,2,5-三唑基、1,3,4-三唑基、四唑基、异噁唑基、噁二唑基、1,2,3-噁二唑基、1,2,4-噁二唑基、1,2,5-噁二唑基、1,3,4-噁二唑基、噻二唑基、吡啶基、哒嗪基、嘧啶基、吡嗪基、三嗪基或四嗪基。 In one embodiment of the present application, the 5- or 6-membered heteroaryl (ring) described in the group R 3 and substituent group Q in each of the above structural formulas are each independently selected from: thienyl, furyl, thiazole base, isothiazolyl, imidazolyl, oxazolyl, pyrrolyl, pyrazolyl, triazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,5- Triazolyl, 1,3,4-triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl or tetrazinyl.
本申请的一种实施方案中，所述5或6元杂芳基(环)各自独立地选自如下结构：
In one embodiment of the present application, the 5- or 6-membered heteroaryl (ring) is each independently selected from the following structures:
本申请的一种实施方案中，上述各结构式中、基团R3和取代基组Q中所述的3至6元杂环烷基为4至6元杂环烷基，各自独立地选自：氮杂环丁烷基、氧杂环丁烷基、四氢呋喃基、四氢噻吩基、四氢吡咯基、噁唑烷基、二氧戊环基、哌啶基、哌嗪基、吗啉基、二氧六环基、硫代吗啉基、硫代吗啉-1,1-二氧化物、四氢吡喃基、吡咯烷-2-酮基、二氢呋喃-2(3H)-酮基、吗啉-3-酮基、哌嗪-2-酮基和哌啶-2-酮基。In one embodiment of the present application, the 3- to 6-membered heterocycloalkyl groups described in the above structural formulas, the group R 3 and the substituent group Q are 4 to 6-membered heterocycloalkyl groups, each independently selected from : Azetidinyl, oxetanyl, tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyrrolyl, oxazolidinyl, dioxolanyl, piperidinyl, piperazinyl, morpholinyl , dioxanyl, thiomorpholinyl, thiomorpholine-1,1-dioxide, tetrahydropyranyl, pyrrolidin-2-one, dihydrofuran-2(3H)-one base, morpholin-3-one group, piperazin-2-one group and piperidin-2-one group.
本申请的一种实施方案中，式(I)化合物选自以下结构中的任意一种：

In one embodiment of the present application, the compound of formula (I) is selected from any one of the following structures:

本申请第二方面提供了一种药物组合物，其包括本申请第一方面所述的化合物、或其药学上可接受的盐、或其立体异构体，以及药学可接受的载体。The second aspect of the present application provides a pharmaceutical composition, which includes the compound described in the first aspect of the present application, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, and a pharmaceutically acceptable carrier.
本申请第三方面提供了本申请第一方面所述的化合物、或其药学上可接受的盐、或其立体异构体以及本申请第二方面所述的药物组合物在制备治疗和/或预防与PI3Kγ活性相关的或由PI3Kγ活性介导的疾病的药物中的应用。The third aspect of this application provides the compounds described in the first aspect of this application, or pharmaceutically acceptable salts thereof, or their stereoisomers, and the pharmaceutical compositions described in the second aspect of this application in the preparation of treatments and/or Use in drugs to prevent diseases associated with or mediated by PI3Kγ activity.
本申请的一种实施方案中，所述与PI3Kγ活性相关的或由PI3Kγ活性介导的疾病为炎症、代谢性疾病或癌症。In one embodiment of the present application, the disease related to or mediated by PI3Kγ activity is inflammation, metabolic disease or cancer.
应理解，在本申请范围内中，本申请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。It should be understood that within the scope of the present application, the above-mentioned technical features of the present application and the technical features specifically described below (such as embodiments) can be combined with each other to constitute a new or preferred technical solution. Due to space limitations, they will not be described one by one here.
附图说明Description of drawings
此处所说明的附图用来提供对本申请的进一步理解，构成本申请的一部分，本申请的示意性实施例及其说明用于解释本申请，并不构成对本申请的不当限定。The drawings described here are used to provide a further understanding of the present application and constitute a part of the present application. The illustrative embodiments of the present application and their descriptions are used to explain the present application and do not constitute an improper limitation of the present application.
图1为C57BL/6小鼠MC38和巨噬细胞混合接种模型肿瘤生长曲线；Figure 1 shows the tumor growth curve of the mixed inoculation model of MC38 and macrophages in C57BL/6 mice;
图2为C57BL/6小鼠MC38和巨噬细胞混合接种模型给药第27天的肿瘤重量。Figure 2 shows the tumor weight on day 27 of C57BL/6 mouse MC38 and macrophage mixed inoculation model.
具体实施方式Detailed ways
为使本申请的目的、技术方案、及优点更加清楚明白，以下参照附图并举实施例，对本申请进一步详细说明。显然，所描述的实施例仅仅是本申请一部分实施例，而不是全部的实施例。本领域技术人员基于本申请所获得的所有其他实施例，都属于本申请保护的范围。In order to make the purpose, technical solutions, and advantages of the present application clearer, the present application will be further described in detail below with reference to the accompanying drawings and examples. Obviously, the described embodiments are only some of the embodiments of the present application, but not all of the embodiments. All other embodiments obtained by those skilled in the art based on this application fall within the scope of protection of this application.
本申请人经过广泛而深入的研究，意外地发现了式(I)所示的杂芳基取代的吡啶并吡咯酮衍生物，其具有优异的PI3Kγ激酶和细胞选择抑制活性、更优异的体内药代参数因此该系列化合物有望开发成为PI3Kγ选择性抑制剂，并用于治疗和/或预防与PI3Kγ活性相关的疾病。在此基础上，申请人完成了本申请。After extensive and in-depth research, the applicant unexpectedly discovered a heteroaryl-substituted pyridopyrrolone derivative represented by formula (I), which has excellent PI3Kγ kinase and cell-selective inhibitory activities and better in vivo drug properties. Therefore, this series of compounds is expected to be developed into PI3Kγ selective inhibitors and used to treat and/or prevent diseases related to PI3Kγ activity. On this basis, the applicant completed this application.
术语定义Definition of Terms
为了能够更清楚地理解本申请的技术内容，下面对本申请的术语作进一步说明。 In order to understand the technical content of this application more clearly, the terminology of this application is further explained below.
“烷基”指直链和支链的饱和的脂族烃基。“C1-8烷基”是指具有1至8个碳原子的烷基，优选为C1-6烷基，更优选为C1-3烷基；烷基的非限制性的实施例包括：甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基、仲丁基、正戊基、1,1-二甲基丙基、1,2-二甲基丙基、2,2-二甲基丙基、1-乙基丙基、2-甲基丁基、3-甲基丁基、正己基、1-乙基-2-甲基丙基、1,1,2-三甲基丙基、1,1-二甲基丁基、1,2-二甲基丁基、2,2-二甲基丁基、1,3-二甲基丁基、2-乙基丁基、2-甲基戊基、3-甲基戊基、4-甲基戊基、2,3-二甲基丁基、正庚基、2-甲基己基、3-甲基己基、4-甲基己基、5-甲基己基、2,3-二甲基戊基、2,4-二甲基戊基、2,2-二甲基戊基、3,3-二甲基戊基、2-乙基戊基、3-乙基戊基、正辛基、2,3-二甲基己基、2,4-二甲基己基、2,5-二甲基己基、2,2-二甲基己基、3,3-二甲基己基、4,4-二甲基己基、2-乙基己基、3-乙基己基、4-乙基己基、2-甲基-2-乙基戊基、2-甲基-3-乙基戊基、正壬基、2-甲基-2-乙基己基、2-甲基-3-乙基己基、2,2-二乙基戊基、正癸基、3,3-二乙基己基、2,2-二乙基己基，及其各种支链异构体。"Alkyl" refers to straight and branched chain saturated aliphatic hydrocarbon groups. "C 1-8 alkyl" refers to an alkyl group having 1 to 8 carbon atoms, preferably C 1-6 alkyl, more preferably C 1-3 alkyl; non-limiting examples of alkyl include : Methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-di Methylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl , 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethyl Butyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2-methylhexyl , 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3 ,3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2,3-dimethylhexyl, 2,4-dimethylhexyl, 2,5-di Methylhexyl, 2,2-dimethylhexyl, 3,3-dimethylhexyl, 4,4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2 -Methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, n-nonyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2 , 2-diethylpentyl, n-decyl, 3,3-diethylhexyl, 2,2-diethylhexyl, and various branched isomers.
“烯基”指直链或支链的具有一个或多个碳碳双键(C＝C)的不饱和脂族烃基，“C2-8烯基”指具有2至8个碳原子的烯基，优选为C2-6烯基，更优选为C2-4烯基，定义类似；烯基的非限制性实施例包括乙烯基、丙烯基、异丙烯基、正丁烯基、异丁烯基、戊烯基、己烯基等。"Alkenyl" refers to a straight-chain or branched unsaturated aliphatic hydrocarbon group with one or more carbon-carbon double bonds (C=C), and "C 2-8 alkenyl" refers to an alkenyl group with 2 to 8 carbon atoms. group, preferably C 2-6 alkenyl, more preferably C 2-4 alkenyl, similarly defined; non-limiting examples of alkenyl include vinyl, propenyl, isopropenyl, n-butenyl, isobutenyl , pentenyl, hexenyl, etc.
“炔基”指直链和支链的具有一个或多个碳碳三键的不饱和脂族烃基，“C2-8炔基”指具有2至8个碳原子的炔基，优选为C2-6炔基，更优选为C2-4炔基，定义类似；炔基的非限制性实施例包括乙炔基、丙炔基、正丁炔基、异丁炔基、戊炔基、己炔基等。"Alkynyl" refers to straight-chain and branched unsaturated aliphatic hydrocarbon groups with one or more carbon-carbon triple bonds, and "C 2-8 alkynyl" refers to alkynyl groups with 2 to 8 carbon atoms, preferably C 2-6 alkynyl, more preferably C 2-4 alkynyl, is similarly defined; non-limiting examples of alkynyl include ethynyl, propynyl, n-butynyl, isobutynyl, pentynyl, hexynyl Alkynyl etc.
“环烷基”和“环烷基环”可互换使用，均指饱和单环、双环或多环环状烃基，该基团可以与芳基或杂芳基稠合。环烷基环可以任选地被取代。在某些实施方案中，环烷基环含有一个或多个羰基，例如氧代的基团。“C3-8环烷基”是指具有3至8个碳原子的单环环烷基，环烷基的非限制性实施例包括环丙基、环丁基、环戊基、环己基、环庚基、环辛基、环丁酮、环戊酮、环戊烷-1,3-二酮等。优选为C3-6环烷基，包括环丙基、环丁基、环戊基和环己基。“C8-10环烷基”是指具有8至10个环原子的稠合双环环状烃基，C8-10环烷基的非限制性实施例包括 "Cycloalkyl" and "cycloalkyl ring" are used interchangeably and both refer to a saturated monocyclic, bicyclic or polycyclic cyclic hydrocarbon group which may be fused to an aryl or heteroaryl group. Cycloalkyl rings may be optionally substituted. In certain embodiments, cycloalkyl rings contain one or more carbonyl groups, such as oxo groups. "C 3-8 cycloalkyl" refers to a monocyclic cycloalkyl group having 3 to 8 carbon atoms. Non-limiting examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, Cycloheptyl, cyclooctyl, cyclobutanone, cyclopentanone, cyclopentane-1,3-dione, etc. Preferred is C 3-6 cycloalkyl, including cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. "C 8-10 cycloalkyl" refers to a fused bicyclic cyclic hydrocarbon group having 8 to 10 ring atoms. Non-limiting examples of C 8-10 cycloalkyl include
“杂环烷基”和“杂环烷基环”可互换使用，均指包含至少一个选自氮、氧和硫的杂原子的环烷基，该基团可以与芳基或杂芳基稠合。杂环烷基环可以任选地被取代。在某些实施方案中，杂环烷基环含有一个或多个羰基或硫代羰基，例如包含氧代和硫代的基团。“3至8元杂环烷基”是指具有3至8个环原子，其中1、2或3个环原子为选自氮、氧和硫的杂原子的单环环状烃基，优选为4至8元杂环烷基。更优选为3至6元杂环烷基，其具有3至6个环原子，其中1或2个环原子为选自氮、氧和硫的杂原子。进一步优选为4至6元杂环烷基，其具有4至6个环原子，其中1或2个环原子为选自氮、氧和硫的杂原子。非限制性实施例包括氮丙环基、环氧乙烷基、氮杂环丁烷基、氧杂环丁烷基、四氢呋喃基、四氢噻吩基、四氢吡咯基、噁唑烷基、二氧戊环基、哌啶基、哌嗪基、吗啉基、二氧六环基、硫代吗啉基、硫代吗啉-1,1-二氧化物、四氢吡喃基、氮杂环丁烷-2-酮基、氧杂环丁烷-2-酮基、二氢呋喃-2(3H)-酮基、吡咯烷-2-酮基、吡咯烷-2,5-二酮基、二氢呋喃-2,5-二酮基、哌啶-2-酮基、四氢-2H-吡喃-2-酮基、哌嗪-2-酮基、吗啉-3-酮基等。“6至12元杂环烷基”和“6至12元稠合杂环烷基”可互换使用，是指具有6至12个环原子，其中1、2或3个环原子为选自氮、氧和硫的杂原子的稠合双环环状烃基。“8至10元杂环烷基”和“8至10元稠合杂环烷基”可 互换使用，是指具有8至10个环原子，其中1、2或3个环原子为选自氮、氧和硫的杂原子的稠合双环环状烃基。非限制性实施例包括六氢-1H-呋喃[3,4-c]吡咯、八氢-1H-环戊[c]吡啶、六氢-1H-吡咯并[2,1-c][1,4]噁嗪、八氢吡咯并[1,2-a]吡嗪、六氢吡咯并[1,2-a]吡嗪-4(1H)-酮、八氢环戊[c]吡咯等。在含有一个或多个氮原子的稠合双环杂环烷基中，只要化合价允许，连接点可以是碳或氮原子。双环杂环烷基系统在一个或两个环中可以包括一个或多个杂原子。"Heterocycloalkyl" and "heterocycloalkyl ring" are used interchangeably and both refer to a cycloalkyl group containing at least one heteroatom selected from nitrogen, oxygen, and sulfur, which group may be associated with an aryl or heteroaryl group. Condensation. Heterocycloalkyl rings may be optionally substituted. In certain embodiments, heterocycloalkyl rings contain one or more carbonyl or thiocarbonyl groups, such as groups containing oxo and thio. "3- to 8-membered heterocycloalkyl" refers to a monocyclic cyclic hydrocarbon group having 3 to 8 ring atoms, wherein 1, 2 or 3 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur, preferably 4 to 8-membered heterocycloalkyl. More preferred is a 3- to 6-membered heterocycloalkyl group, which has 3 to 6 ring atoms, of which 1 or 2 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. Further preferred is a 4- to 6-membered heterocycloalkyl group having 4 to 6 ring atoms, of which 1 or 2 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. Non-limiting examples include aziridyl, oxiranyl, azetidinyl, oxetanyl, tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyrrolyl, oxazolidinyl, di Oxopentyl, piperidinyl, piperazinyl, morpholinyl, dioxanyl, thiomorpholinyl, thiomorpholine-1,1-dioxide, tetrahydropyranyl, aza cyclobutane-2-one group, oxetan-2-one group, dihydrofuran-2(3H)-one group, pyrrolidine-2-one group, pyrrolidine-2,5-dione group , dihydrofuran-2,5-dione group, piperidin-2-one group, tetrahydro-2H-pyran-2-one group, piperazine-2-one group, morpholin-3-one group, etc. . "6- to 12-membered heterocycloalkyl" and "6- to 12-membered fused heterocycloalkyl" are used interchangeably and refer to having 6 to 12 ring atoms, of which 1, 2 or 3 ring atoms are selected from A fused bicyclic cyclic hydrocarbon radical of heteroatoms of nitrogen, oxygen and sulfur. "8 to 10 membered heterocycloalkyl" and "8 to 10 membered fused heterocycloalkyl" can Used interchangeably, it refers to a fused bicyclic cyclic hydrocarbon group having 8 to 10 ring atoms, of which 1, 2 or 3 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. Non-limiting examples include hexahydro-1H-furan[3,4-c]pyrrole, octahydro-1H-cyclopenta[c]pyridine, hexahydro-1H-pyrrolo[2,1-c][1, 4]oxazine, octahydropyrrolo[1,2-a]pyrazine, hexahydropyrro[1,2-a]pyrazin-4(1H)-one, octahydrocyclopenta[c]pyrrole, etc. In fused bicyclic heterocycloalkyl groups containing one or more nitrogen atoms, the point of attachment may be a carbon or nitrogen atom as long as the valency permits. Bicyclic heterocycloalkyl systems may include one or more heteroatoms in one or both rings.
“杂芳基”和“杂芳基环”可互换使用，均指具有环碳原子和环杂原子的单环、双环或多环的4n+2芳族环体系(例如，具有以环状排列共享的6或10个π电子)的基团，其中每个杂原子独立地选自氮、氧和硫。杂芳基环可以任选地被取代。“5至10元杂芳基”是指具有5至10个环原子，其中1、2、3或4个环原子为杂原子的单环或双环杂芳基。“5至6元杂芳基”是指具有5至6个环原子，其中1、2、3或4个环原子为杂原子的单环杂芳基，非限制性实施例包括噻吩基、呋喃基、噻唑基、异噻唑基、咪唑基、噁唑基、吡咯基、吡唑基、三唑基、1,2,3-三唑基、1,2,4-三唑基、1,2,5-三唑基、1,3,4-三唑基、四唑基、异噁唑基、噁二唑基、1,2,3-噁二唑基、1,2,4-噁二唑基、1,2,5-噁二唑基、1,3,4-噁二唑基、噻二唑基、吡啶基、哒嗪基、嘧啶基、吡嗪基、三嗪基、四嗪基。“9或10元杂芳基”是指具有9或10个环原子，其中1、2、3或4个环原子为杂原子的双环杂芳基，非限制性实施例包括吲哚基、异吲哚基、吲唑基、苯并三唑基、苯并噻吩基、异苯并噻吩基、苯并呋喃基、苯并异呋喃基、苯并咪唑基、苯并噁唑基、苯并异噁唑基、苯并噁二唑基、苯并噻唑基、苯并异噻唑基、苯并噻二唑基、茚嗪基、嘌呤基、吡啶并[3,2-d]嘧啶基、吡啶并[2,3-d]嘧啶基、吡啶并[3,4-d]嘧啶基、吡啶并[4,3-d]嘧啶基、1,8-萘啶基、1,7-萘啶基、1,6-萘啶基、1,5-萘啶基、喋啶基、喹啉基、异喹啉基、噌琳基、喹喔啉基、酞嗪基和喹唑啉基。“杂原子”是指氮、氧或硫。在含有一个或多个氮原子的杂芳基中，只要化合价允许，连接点可以是碳或氮原子。杂芳基双环系统在一个或两个环中可以包括一个或多个杂原子。"Heteroaryl" and "heteroaryl ring" are used interchangeably and both refer to a monocyclic, bicyclic or polycyclic 4n+2 aromatic ring system having ring carbon atoms and ring heteroatoms (e.g., having a Arrange shared 6 or 10 π electrons) groups in which each heteroatom is independently selected from nitrogen, oxygen and sulfur. Heteroaryl rings may be optionally substituted. "5- to 10-membered heteroaryl" refers to a monocyclic or bicyclic heteroaryl group having 5 to 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms. "5- to 6-membered heteroaryl" refers to a monocyclic heteroaryl group with 5 to 6 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms. Non-limiting examples include thienyl, furan base, thiazolyl, isothiazolyl, imidazolyl, oxazolyl, pyrrolyl, pyrazolyl, triazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2 ,5-triazolyl, 1,3,4-triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl Azolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, thiadiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, tetrazine base. "9- or 10-membered heteroaryl" refers to a bicyclic heteroaryl group with 9 or 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms. Non-limiting examples include indolyl, iso- Indolyl, indazolyl, benzotriazolyl, benzothienyl, isobenzothienyl, benzofuranyl, benzisofuranyl, benzimidazolyl, benzoxazolyl, benzoiso Oxazolyl, benzoxadiazolyl, benzothiazolyl, benzisothiazolyl, benzothiadiazolyl, indanazinyl, purinyl, pyrido[3,2-d]pyrimidinyl, pyrido [2,3-d]pyrimidinyl, pyrido[3,4-d]pyrimidinyl, pyrido[4,3-d]pyrimidinyl, 1,8-naphthyridinyl, 1,7-naphthyridinyl, 1,6-naphthyridinyl, 1,5-naphthyridinyl, pteridinyl, quinolinyl, isoquinolinyl, quinolinyl, quinoxalinyl, phthalazinyl and quinazolinyl. "Heteroatom" means nitrogen, oxygen or sulfur. In heteroaryl groups containing one or more nitrogen atoms, the point of attachment may be a carbon or nitrogen atom as long as the valency permits. Heteroaryl bicyclic systems may include one or more heteroatoms in one or both rings.
“稠合”是指两个或多个环共用一个或多个键的结构。"Fused" refers to a structure in which two or more rings share one or more bonds.
“苯基并杂环烷基”是指苯环与杂环烷基环稠合形成双环、三环或多环体系的基团，其中杂环烷基环如上述所定义。“9或10元苯基并杂环烷基”指具有9或10个环原子，其中1、2、3或4个环原子为选自氮、氧和硫的杂原子的双环环状基团。非限制性实施例包含吲哚啉、苯并[d][1,3]二噁唑、1,2,3,4-四氢异喹啉、3,4-二氢-2H-苯并[b][1,4]噁嗪、吲哚-2-酮、异吲哚林-1-酮、1,4-二氢异喹啉-3(2H)-酮等。"Phenylheterocycloalkyl" refers to a group in which a benzene ring is fused with a heterocycloalkyl ring to form a bicyclic, tricyclic or polycyclic system, wherein the heterocycloalkyl ring is as defined above. "9- or 10-membered phenylheterocycloalkyl" refers to a bicyclic cyclic group having 9 or 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. . Non-limiting examples include indoline, benzo[d][1,3]dioxazole, 1,2,3,4-tetrahydroisoquinoline, 3,4-dihydro-2H-benzo[ b][1,4]oxazine, indole-2-one, isoindolin-1-one, 1,4-dihydroisoquinolin-3(2H)-one, etc.
“杂芳基并杂环烷基”是指杂芳基环与杂环烷基环稠合形成双环、三环或多环体系的基团，其中杂环烷基环如上述所定义。“9或10元杂芳基并杂环烷基”指具有9或10个环原子，其中1、2、3或4个环原子为选自氮、氧和硫的杂原子的双环环状基团。非限制性实施例包含2,3-二氢-1H-吡咯并[2,3-b]吡啶、[1,3]二氧戊环[4,5-b]吡啶、2,3-二氢-1H-吡啶并[3,4-b][1,4]噁嗪、2,3,4,6-四氢吡咯并[3,4-b][1,4]噁嗪、2,4,5,6-四氢吡喃并[2,3-c]吡唑、5,6,7,8-四氢吡啶并[3,4-d]嘧啶等。"Heteroarylheterocycloalkyl" refers to a group in which a heteroaryl ring is fused with a heterocycloalkyl ring to form a bicyclic, tricyclic or polycyclic system, wherein the heterocycloalkyl ring is as defined above. "9- or 10-membered heteroarylheterocycloalkyl" refers to a bicyclic cyclic group having 9 or 10 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms selected from nitrogen, oxygen and sulfur. group. Non-limiting examples include 2,3-dihydro-1H-pyrrolo[2,3-b]pyridine, [1,3]dioxola[4,5-b]pyridine, 2,3-dihydro -1H-pyrido[3,4-b][1,4]oxazine, 2,3,4,6-tetrahydropyrrolo[3,4-b][1,4]oxazine, 2,4 ,5,6-tetrahydropyrano[2,3-c]pyrazole, 5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine, etc.
“烷氧基”指-O-烷基，其中烷基的定义如上所述。优选为C1-8烷氧基，更优选为C1-6烷氧基，最优选为C1-3烷氧基。烷氧基的非限制性实施例包含甲氧基、乙氧基、正丙氧基、异丙氧基、丁氧基、叔丁氧基、异丁氧基、戊氧基等。"Alkoxy" refers to -O-alkyl, where alkyl is as defined above. It is preferably a C 1-8 alkoxy group, more preferably a C 1-6 alkoxy group, and most preferably a C 1-3 alkoxy group. Non-limiting examples of alkoxy include methoxy, ethoxy, n-propoxy, isopropoxy, butoxy, tert-butoxy, isobutoxy, pentyloxy, and the like.
“环烷基氧基”指-O-环烷基，其中环烷基的定义如上所述。优选为C3-8环烷基氧基，更优选为C3-6环烷基氧基。环烷基氧基的非限制性实施例包含环丙基氧基、环丁基氧基、环戊基氧基、环己基氧基等。"Cycloalkyloxy" refers to -O-cycloalkyl, where cycloalkyl is as defined above. A C 3-8 cycloalkyloxy group is preferred, and a C 3-6 cycloalkyloxy group is more preferred. Non-limiting examples of cycloalkyloxy include cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy, and the like.
“一个键”指由其连接的两个基团通过一个共价键连接。"A bond" means that the two groups connected thereto are connected by a covalent bond.
“卤素”指氟、氯、溴或碘。"Halogen" means fluorine, chlorine, bromine or iodine.
“卤代”指基团中一个或多个(如1、2、3、4或5个)氢被卤素所取代。 "Halo" means that one or more (eg, 1, 2, 3, 4 or 5) hydrogens in a group are replaced by halogen.
例如，“卤代烷基”指烷基上的氢被一个或多个(如1、2、3、4或5个)卤素取代，其中烷基的定义如上所述。优选为卤代C1-8烷基，更优选为卤代C1-6烷基，进一步优选为卤代C1-3烷基。卤代烷基的例子包括但不限于一氯甲基、二氯甲基、三氯甲基、一氯乙基、1,2-二氯乙基、三氯乙基、一溴乙基、一氟甲基、二氟甲基、三氟甲基、一氟乙基、二氟乙基、三氟乙基等。For example, "haloalkyl" refers to an alkyl group in which the hydrogens are replaced by one or more (eg, 1, 2, 3, 4 or 5) halogens, where alkyl is as defined above. A halogenated C 1-8 alkyl group is preferred, a halogenated C 1-6 alkyl group is more preferred, and a halogenated C 1-3 alkyl group is even more preferred. Examples of haloalkyl groups include, but are not limited to, monochloromethyl, dichloromethyl, trichloromethyl, monochloroethyl, 1,2-dichloroethyl, trichloroethyl, monobromoethyl, monofluoromethyl base, difluoromethyl, trifluoromethyl, monofluoroethyl, difluoroethyl, trifluoroethyl, etc.
又例如，“卤代烷氧基”指烷氧基上的氢被一个或多个(如1、2、3、4或5个)卤素取代，其中烷氧基的定义如上所述。优选为卤代C1-8烷氧基，更优选为卤代C1-6烷氧基，进一步优选为卤代C1-3烷氧基。卤代烷氧基包括但不限于三氟甲氧基、三氟乙氧基、一氟甲氧基、一氟乙氧基、二氟甲氧基、二氟乙氧基等。As another example, "haloalkoxy" means that the hydrogen on the alkoxy group is substituted by one or more (such as 1, 2, 3, 4 or 5) halogens, where the alkoxy group is as defined above. A halogenated C 1-8 alkoxy group is preferred, a halogenated C 1-6 alkoxy group is more preferred, and a halogenated C 1-3 alkoxy group is even more preferred. Haloalkoxy groups include, but are not limited to, trifluoromethoxy, trifluoroethoxy, monofluoromethoxy, monofluoroethoxy, difluoromethoxy, difluoroethoxy, and the like.
又例如，“卤代环烷基”指环烷基上的氢被一个或多个(如1、2、3、4或5个)卤素取代，其中环烷基的定义如上所述。优选为卤代C3-8环烷基，更优选为卤代C3-6环烷基。卤代环烷基包括但不限于三氟环丙基、一氟环丙基、一氟环己基、二氟环丙基、二氟环己基等。As another example, "halogenated cycloalkyl" means that the hydrogen on the cycloalkyl group is substituted by one or more (such as 1, 2, 3, 4 or 5) halogens, wherein the cycloalkyl group is as defined above. A halogenated C 3-8 cycloalkyl group is preferred, and a halogenated C 3-6 cycloalkyl group is more preferred. Halogenated cycloalkyl groups include, but are not limited to, trifluorocyclopropyl, monofluorocyclopropyl, monofluorocyclohexyl, difluorocyclopropyl, difluorocyclohexyl, etc.
“氘代烷基”指烷基上的氢被一个或多个(如1、2、3、4或5个)氘原子取代，其中烷基的定义如上所述。优选为氘代C1-8烷基，更优选为氘代C1-6烷基，更优选为氘代C1-3烷基。氘代烷基的例子包括但不限于单氘代甲基、单氘代乙基、二氘代甲基、二氘代乙基、三氘代甲基、三氘代乙基等。"Deuterated alkyl" means that the hydrogen on the alkyl group is replaced by one or more (eg, 1, 2, 3, 4 or 5) deuterium atoms, where alkyl is as defined above. It is preferably a deuterated C 1-8 alkyl group, more preferably a deuterated C 1-6 alkyl group, and even more preferably a deuterated C 1-3 alkyl group. Examples of deuterated alkyl groups include, but are not limited to, monodeuterated methyl, monodeuterated ethyl, dideuterated methyl, dideuterated ethyl, trideuterated methyl, trideuterated ethyl, and the like.
“氨基”指NH2，“氰基”指CN，“硝基”指NO2，“苯甲基”指-CH2-苯基，“氧代基”指＝O，“羧基”指-C(O)OH，“乙酰基”指-C(O)CH3，“羟甲基”指-CH2OH，“羟乙基”指-CH2CH2OH或-CHOHCH3，“羟基”指-OH，“巯基”指SH，“亚环丙基”结构为： "Amino" refers to NH 2 , "cyano" refers to CN, "nitro" refers to NO 2 , "benzyl" refers to -CH 2 -phenyl, "oxo" refers to =O, and "carboxyl" refers to -C (O)OH, "acetyl" refers to -C(O)CH 3 , "hydroxymethyl" refers to -CH 2 OH, "hydroxyethyl" refers to -CH 2 CH 2 OH or -CHOHCH 3 , "hydroxyl" refers to -OH, "mercapto" refers to SH, and the structure of "cyclopropylene" is:
“取代的”指基团中的一个或多个氢原子，优选为1～5个氢原子各自独立地被相应数目的取代基取代，更优选为1～3个氢原子各自独立地被相应数目的取代基取代。不言而喻，取代基仅处在它们的可能的化学位置，本领域技术人员能够在不付出过多努力的情况下确定(通过实验或理论)可能或不可能的取代。例如，具有游离氢的氨基或羟基与具有不饱和(如烯属)键的碳原子结合时可能是不稳定的。"Substituted" refers to one or more hydrogen atoms in the group, preferably 1 to 5 hydrogen atoms are each independently substituted by a corresponding number of substituents, and more preferably 1 to 3 hydrogen atoms are each independently substituted by a corresponding number of substituents. Substituted with substituents. It goes without saying that the substituents are only in their possible chemical positions, and the person skilled in the art is able to determine (either experimentally or theoretically) possible or impossible substitutions without undue effort. For example, an amino or hydroxyl group with a free hydrogen may be unstable when combined with a carbon atom with an unsaturated (eg, olefinic) bond.
除非另有定义，本申请所述“各自独立地选自……的取代基”是指当基团上的一个以上的氢被取代基取代时，所述的取代基种类可以相同或不同，所选自的取代基为各自独立的种类。Unless otherwise defined, the "substituents each independently selected from..." mentioned in this application mean that when more than one hydrogen on the group is replaced by a substituent, the types of the substituents may be the same or different, so The substituents selected from are of their own independent species.
除非另有定义，本申请所述“……相同或不同，且各自独立地为……”是指当通式中存在一个以上的相同取代基团时，该基团可以相同或不同，为各自独立的种类。例如L为(CR01R02)s，当s为2时，即L为(CR01R02)-(CR01R02)，其中的两个R01可以相同或不同，两个R02可以相同或不同，为各自独立的种类，例如L可以为C(CH3)(CN)-C(CH2CH3)(OH)，C(CH3)(CN)-C(CH3)(OH)或C(CN)(CH2CH3)-C(OH)(CH2CH3)。Unless otherwise defined, "...the same or different, and each independently is..." mentioned in this application means that when there is more than one same substituent group in the general formula, the group can be the same or different, and is each independently independent kind. For example, L is (CR 01 R 02 ) s . When s is 2, L is (CR 01 R 02 )-(CR 01 R 02 ). The two R 01 can be the same or different, and the two R 02 can be Identical or different, they are independent species. For example, L can be C(CH 3 )(CN)-C(CH 2 CH 3 )(OH), C(CH 3 )(CN)-C(CH 3 )(OH ) or C(CN)(CH 2 CH 3 )-C(OH)(CH 2 CH 3 ).
除非另有定义，本申请中任一基团可以是取代的或未取代的。上述基团被取代时，取代基优选为1至5个以下基团，独立地选自氰基、卤素(优选为氟或氯)、C1-8烷基(优选为C1-6烷基，更优选为C1-3烷基)、C1-8烷氧基(优选为C1-6烷氧基，更优选为C1-3烷氧基)、卤代C1-8烷基(优选为卤代C1-6烷基，更优选为卤代C1-3烷基)、C3-8环烷基(优选为C3-6环烷基)、卤代C1-8烷氧基(优选为卤代C1-6烷氧基，更优选为卤代C1-3烷氧基)、C1-8烷基取代的氨基、卤代C1-8烷基取代的氨基、乙酰基、羟基、羟甲基、羟乙基、羧基、硝基、C6-10芳基(优选为苯基)、C3-8环烷基氧基(优选为C3-6环烷基氧基)、C2-8烯基(优选为C2-6烯基，更优选为C2-4烯基)、C2-8炔基(优选为C2-6炔基，更优选为C2-4炔基)、-CONRa0Rb0、-C(O)OC1-10烷基(优选为-C(O)OC1-6烷基，更优选为-C(O)OC1-3烷基)、-CHO、-OC(O)C1-10烷基(优选为-OC(O)C1-6烷基，更优选为-OC(O)C1-3烷基)、-SO2C1-10烷基(优选为-SO2C1-6烷基，更优选 为-SO2C1-3烷基)、-SO2C6-10芳基(优选为-SO2C6芳基，如-SO2-苯基)、-COC6-10芳基(优选为-COC6芳基，如-CO-苯基)、4至6元饱和或不饱和单杂环、4至6元饱和或不饱和单环、5至6元单环杂芳基环、8至10元双环杂芳基环、螺环、螺杂环、桥环或桥杂环，其中Ra0、Rb0各自独立地为氢或C1-3烷基。Unless otherwise defined, any group in this application may be substituted or unsubstituted. When the above groups are substituted, the substituents are preferably 1 to 5 or less groups, independently selected from cyano, halogen (preferably fluorine or chlorine), C 1-8 alkyl (preferably C 1-6 alkyl) , more preferably C 1-3 alkyl), C 1-8 alkoxy (preferably C 1-6 alkoxy, more preferably C 1-3 alkoxy), halogenated C 1-8 alkyl (preferably halogenated C 1-6 alkyl, more preferably halogenated C 1-3 alkyl), C 3-8 cycloalkyl (preferably C 3-6 cycloalkyl), halogenated C 1-8 Alkoxy (preferably halogenated C 1-6 alkoxy, more preferably halogenated C 1-3 alkoxy), C 1-8 alkyl substituted amino, halogenated C 1-8 alkyl substituted Amino, acetyl, hydroxyl, hydroxymethyl, hydroxyethyl, carboxyl, nitro, C 6-10 aryl (preferably phenyl), C 3-8 cycloalkyloxy (preferably C 3-6 cyclic Alkyloxy), C 2-8 alkenyl (preferably C 2-6 alkenyl, more preferably C 2-4 alkenyl), C 2-8 alkynyl (preferably C 2-6 alkynyl, more preferably Preferably C 2-4 alkynyl), -CONR a0 R b0 , -C(O)OC 1-10 alkyl (preferably -C(O)OC 1-6 alkyl, more preferably -C(O) OC 1-3 alkyl), -CHO, -OC(O)C 1-10 alkyl (preferably -OC(O)C 1-6 alkyl, more preferably -OC(O)C 1-3 alkyl base), -SO 2 C 1-10 alkyl (preferably -SO 2 C 1-6 alkyl, more preferably is -SO 2 C 1-3 alkyl), -SO 2 C 6-10 aryl (preferably -SO 2 C 6 aryl, such as -SO 2 -phenyl), -COC 6-10 aryl (preferably is -COC 6 aryl, such as -CO-phenyl), 4 to 6 membered saturated or unsaturated monoheterocyclic ring, 4 to 6 membered saturated or unsaturated monocyclic ring, 5 to 6 membered monocyclic heteroaryl ring, 8 to a 10-membered bicyclic heteroaryl ring, spiro ring, spiro heterocyclic ring, bridged ring or bridged heterocyclic ring, wherein R a0 and R b0 are each independently hydrogen or C 1-3 alkyl.
本申请中，一个方案中出现两个或更多个“优选”时，任意的两个“优选”可以是彼此独立的。In this application, when two or more "preferences" appear in a solution, any two "preferences" may be independent of each other.
本申请中，当取代基的数量大于1时，任意的两个取代基可以相同或不同。如，可以为两个相同或不同的卤素取代，可以为一个卤素和一个羟基取代。In this application, when the number of substituents is greater than 1, any two substituents may be the same or different. For example, it can be substituted by two identical or different halogens, or it can be substituted by one halogen and one hydroxyl group.
本申请以上所述的各类取代基团其自身也是可以被本申请所描述的基团取代。The various substituent groups described above in this application can themselves be substituted by the groups described in this application.
本申请所述的4至6元饱和单杂环被取代时，取代基的位置可处在它们可能的化学位置，示例性的单杂环的代表性的取代情况如下所示：
When the 4- to 6-membered saturated monoheterocycle described in this application is substituted, the substituents can be positioned at their possible chemical positions. The representative substitution situations of the exemplary monoheterocycle are as follows:
其中“Sub”表示本申请所述的各类取代基；表示与其他原子的连接位置。“Sub” represents various substituents described in this application; Represents the connection position to other atoms.
药物组合物pharmaceutical composition
通常本申请化合物或其药学可接受的盐或其立体异构体可以与一种或多种药用载体形成适合的剂型施用。这些剂型适用于口服、直肠给药、局部给药、口内给药以及其他非胃肠道施用(例如，皮下、肌肉、静脉等)。例如，适合口服给药的剂型包括胶囊、片剂、颗粒剂以 及糖浆等。这些制剂中包含的本申请的化合物可以是固体粉末或颗粒；水性或非水性液体中的溶液或是混悬液；油包水或水包油的乳剂等。上述剂型可由活性化合物与一种或多种载体或辅料经由通用的药剂学方法制成。上述的载体需要与活性化合物或其他辅料兼容。活性化合物可与载体形成溶液或是混悬液。Generally, the compounds of the present application or their pharmaceutically acceptable salts or their stereoisomers can be combined with one or more pharmaceutical carriers to form a suitable dosage form for administration. These dosage forms are suitable for oral, rectal, topical, intraoral, and other parenteral administration (e.g., subcutaneous, intramuscular, intravenous, etc.). For example, dosage forms suitable for oral administration include capsules, tablets, granules, and and syrup, etc. The compounds of the present application contained in these preparations can be solid powders or granules; solutions or suspensions in aqueous or non-aqueous liquids; water-in-oil or oil-in-water emulsions, etc. The above dosage forms can be prepared from the active compound and one or more carriers or excipients through common pharmaceutical methods. The above carriers need to be compatible with the active compound or other excipients. The active compounds can form solutions or suspensions with carriers.
“药学可接受的载体”是指无毒、惰性、固态、半固态的物质或液体灌装机、稀释剂、封装材料或辅助制剂或任何类型辅料，其与患者相兼容，最好为哺乳动物，更优选为人，其适合将活性试剂输送到目标靶点而不终止试剂的活性。"Pharmaceutically acceptable carrier" means a non-toxic, inert, solid, semi-solid substance or liquid filling machine, diluent, encapsulating material or auxiliary preparation or excipient of any type that is compatible with the patient, preferably a mammal , more preferably human, which is suitable for delivering the active agent to the intended target site without terminating the activity of the agent.
“本申请的化合物”是指本申请式(I)化合物、或其药学上可接受的盐、或其立体异构体，其具有PI3Kγ选择抑制活性，以及制备式(I)化合物、或其药学上可接受的盐、或其立体异构体的中间体化合物。"Compounds of the present application" refer to compounds of formula (I) of the present application, or pharmaceutically acceptable salts thereof, or stereoisomers thereof, which have PI3Kγ selective inhibitory activity, and preparation of compounds of formula (I), or pharmaceutically acceptable salts thereof. Acceptable salts, or intermediate compounds of its stereoisomers.
本申请的组合物以符合医学实践规范的方式配制，定量和给药。给予化合物的“治疗有效量”由要治疗的具体病症、治疗的个体、病症的起因、药物的靶点以及给药方式等因素决定。The compositions of the present application are formulated, dosed, and administered in a manner consistent with good medical practice. The "therapeutically effective amount" of a compound administered is determined by factors such as the specific condition to be treated, the individual being treated, the cause of the condition, the target of the drug, and the mode of administration.
“治疗有效量”是指将引起个体的生物学或医学响应，例如降低或抑制酶或蛋白质活性或改善症状、缓解病症、减缓或延迟疾病进程或预防疾病等的本申请化合物的量。"Therapeutically effective amount" refers to the amount of a compound of the present application that will induce a biological or medical response in an individual, such as reducing or inhibiting enzyme or protein activity or improving symptoms, alleviating disease, slowing or delaying disease progression, or preventing disease, etc.
本申请的所述药物组合物或所述药用组合物中含有的本申请化合物或其药学上可接受的盐或其立体异构体的治疗有效量优选为0.1mg/kg-5g/kg(体重)。The pharmaceutical composition of the present application or the therapeutically effective amount of the compound of the present application or its pharmaceutically acceptable salt or its stereoisomer contained in the pharmaceutical composition is preferably 0.1 mg/kg-5g/kg ( weight).
“患者”是指一种动物，最好为哺乳动物，更好的为人。术语“哺乳动物”是指温血脊椎类哺乳动物，包括如猫、狗、兔、熊、狐狸、狼、猴子、鹿、鼠、猪和人类。"Patient" means an animal, preferably a mammal, more preferably a human being. The term "mammal" refers to warm-blooded vertebrate mammals, including, for example, cats, dogs, rabbits, bears, foxes, wolves, monkeys, deer, rats, pigs, and humans.
“治疗”是指减轻、延缓进展、衰减、预防，或维持现有疾病或病症(例如癌症)。治疗还包括将疾病或病症的一个或多个症状治愈、预防其发展或减轻到某种程度。"Treat" means to alleviate, delay the progression, attenuate, prevent, or maintain an existing disease or condition (eg, cancer). Treatment also includes curing, preventing the progression of, or alleviating to some degree one or more symptoms of a disease or condition.
所述“药学上可接受的盐”包括药学可接受的酸加成盐和药学可接受的碱加成盐。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他副作用的，与无机酸或有机酸所形成的盐。这些盐可通过本专业已知的方法制备。The "pharmaceutically acceptable salts" include pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts. Pharmaceutically acceptable acid addition salts refer to salts formed with inorganic or organic acids that retain the biological effectiveness of the free base without other side effects. These salts can be prepared by methods known in the art.
“药学可接受的碱加成盐”，包括但不限于无机碱的盐如钠盐，钾盐，钙盐和镁盐等。包括但不限于有机碱的盐，比如铵盐，三乙胺盐，赖氨酸盐，精氨酸盐等。这些盐可通过本专业已知的方法制备。"Pharmaceutically acceptable base addition salts" include, but are not limited to, salts of inorganic bases such as sodium salts, potassium salts, calcium salts and magnesium salts. Including but not limited to salts of organic bases, such as ammonium salts, triethylamine salts, lysine salts, arginine salts, etc. These salts can be prepared by methods known in the art.
当本申请式(I)所示的化合物含有一个或多个手性中心时，可以以不同的光学活性形式存在。当式(I)化合物含有一个手性中心时，该化合物包含一对对映异构体。该化合物的两个对映异构体以及该一对对映异构体的混合物，如外消旋混合物也在本申请的保护范围内。对映异构体可以通过本专业已知的方法拆分，例如结晶以及手性色谱等方法。当式(I)化合物含有多于一个手性中心时，该化合物包含对映异构体和非对映异构体。该化合物的所有对映异构体和非对映异构体，以及对映异构体的混合物，非对映异构体的混合物，以及对映异构体和非对映异构体的混合物也在本申请的保护范围内。对映异构体、非对映异构体可由本专业已知方法拆分，比如结晶以及制备色谱。When the compound represented by formula (I) of the present application contains one or more chiral centers, it may exist in different optically active forms. When a compound of formula (I) contains a chiral center, the compound contains a pair of enantiomers. Two enantiomers of the compound as well as mixtures of the pair of enantiomers, such as racemic mixtures, are also within the scope of the present application. Enantiomers can be resolved by methods known in the art, such as crystallization and chiral chromatography. When a compound of formula (I) contains more than one chiral center, the compound contains enantiomers and diastereomers. All enantiomers and diastereomers of this compound, as well as mixtures of enantiomers, mixtures of diastereomers, and mixtures of enantiomers and diastereomers It is also within the protection scope of this application. Enantiomers and diastereomers can be resolved by methods known in the art, such as crystallization and preparative chromatography.
制备方法Preparation
本申请提供了式(I)化合物的制备方法，使用本领域技术人员已知的标准合成技术或使用本领域已知的方法与本申请描述的方法组合可以合成式(I)化合物。本申请给出的溶剂、温度和其它反应条件可以根据本领域技术而改变。所述反应可以按顺序使用，以提供本申请的化合物，或者它们可以用于合成片段，所述片段通过本申请所描述的方法和/或本领域已知的方法随后加入。This application provides methods for the preparation of compounds of formula (I), which can be synthesized using standard synthesis techniques known to those skilled in the art or using methods known in the art in combination with the methods described in this application. Solvents, temperatures and other reaction conditions given herein may vary according to skill in the art. The reactions can be used sequentially to provide compounds of the present application, or they can be used to synthesize fragments that are added subsequently by methods described herein and/or methods known in the art.
本申请描述的化合物可以使用与下述类似的方法或实施例中所述的示例性方法，或本领域技术人员所用的相关公开文献，通过使用适当的可选择的起始原料合成化合物。用于合成 本申请所描述的化合物的起始原料可以被合成或可以从商业来源获得。本申请描述的化合物和其它相关具有不同取代基的化合物可以使用本领域技术人员已知的技术和原料合成。制备本申请公开的化合物的一般方法可以来自本领域已知的反应，并且该反应可以通过由本领域技术人员所认为适当的试剂和条件修改，以引入本申请提供的分子中的各种部分。The compounds described in this application can be synthesized using methods similar to those described below or the exemplary methods described in the Examples, or relevant publications used by those skilled in the art, by using appropriate alternative starting materials. for synthesis Starting materials for the compounds described herein may be synthesized or may be obtained from commercial sources. The compounds described herein and other related compounds having various substituents can be synthesized using techniques and starting materials known to those skilled in the art. General methods for preparing the compounds disclosed herein can be derived from reactions known in the art, and the reactions can be modified by reagents and conditions deemed appropriate by those skilled in the art to introduce various moieties in the molecules provided herein.
本申请的有益效果：Beneficial effects of this application:
提供了一系列杂芳基取代的吡啶并吡咯酮衍生物，其对PI3Kγ酶和细胞具有较高的选择抑制活性以及具有更优异的体内药代参数，对PI3Kγ激酶的抑制活性IC50值小于100nM，在一些实施方案中具有小于50nM的IC50值，在一些实施方案中具有小于10nM的IC50值，因此可用作治疗和/或预防与PI3Kγ活性相关的或由PI3Kγ活性介导的疾病的药物。Provides a series of heteroaryl-substituted pyridopyrrolone derivatives, which have higher selective inhibitory activity against PI3Kγ enzymes and cells and better in vivo pharmacokinetic parameters. The IC 50 value of the inhibitory activity against PI3Kγ kinase is less than 100 nM. , in some embodiments having an IC 50 value of less than 50 nM, in some embodiments having an IC 50 value of less than 10 nM, and therefore can be used as a therapeutic agent for the treatment and/or prevention of diseases associated with or mediated by PI3Kγ activity. drug.
下面结合具体实施例，进一步阐述本申请。应理解，这些实施例仅用于说明本申请而不用于限制本申请的范围。下列实施例中未注明具体条件的实验方法，通常按照常规条件如Sambrook等人，分子克隆：实验室手册(New York:Cold Spring Harbor Laboratory Press，1989)中所述的条件，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。除非另行定义，本申请所用的术语与本领域熟练人员所熟悉的意义相同。此外，任何与所记载内容相似或同等的方法及材料皆可应用于本申请中。The present application will be further described below in conjunction with specific embodiments. It should be understood that these examples are only used to illustrate the present application and are not intended to limit the scope of the present application. Experimental methods without specifying specific conditions in the following examples usually follow conventional conditions such as those described in Sambrook et al., Molecular Cloning: Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or according to the manufacturer Suggested conditions. Unless otherwise stated, percentages and parts are by weight. Unless otherwise defined, terms used herein have the same meaning as is familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to those described can be used in this application.
已知的起始原料可以采用或按照本领域已知的方法来合成，或可以购自ABCR GmbH&Co.KG，Acros Organics，Aldrich Chemical Company，韶远化学科技(Accela ChemBio Inc)和达瑞化学品等公司。Known starting materials can be synthesized by methods known in the art, or can be purchased from ABCR GmbH&Co.KG, Acros Organics, Aldrich Chemical Company, Shaoyuan Chemical Technology (Accela ChemBio Inc) and Dari Chemicals, etc. company.
若无特殊说明，实施例中的反应均在氮气氛或氩气氛下进行。Unless otherwise specified, the reactions in the examples were all carried out under a nitrogen atmosphere or an argon atmosphere.
DCM：二氯甲烷；DMF：二甲基甲酰胺；THF：四氢呋喃；Pd(dppf)Cl2：[1,1'-双(二苯基磷)二茂铁]二氯化钯；dppf：1,1'-双(二苯基膦)二茂铁；Pin2B2：联硼酸频那醇酯；SEM-Cl：2-(三甲基硅烷基)乙氧甲基氯；ACN：乙腈；TFA：三氟乙酸；DCM：二氯甲烷；MeOH：甲醇；n-BuLi：正丁基锂；KOAc：乙酸钾；1,4-Dioxane：1,4-二氧六环；CombiFlash：快速液相制备色谱仪；EA：乙酸乙酯；PE：石油醚；DMSO：二甲基亚砜；BSA：牛血清白蛋白；FA：甲酸；DDQ：2,3-二氯-5,6-二氰对苯醌。DCM: dichloromethane; DMF: dimethylformamide; THF: tetrahydrofuran; Pd(dppf)Cl 2 : [1,1'-bis(diphenylphosphorus)ferrocene]palladium dichloride; dppf: 1 ,1'-bis(diphenylphosphine)ferrocene; Pin 2 B 2 : pinacol diboronate; SEM-Cl: 2-(trimethylsilyl)ethoxymethyl chloride; ACN: acetonitrile; TFA: trifluoroacetic acid; DCM: dichloromethane; MeOH: methanol; n-BuLi: n-butyllithium; KOAc: potassium acetate; 1,4-Dioxane: 1,4-dioxane; CombiFlash: fast liquid phase Preparative chromatograph; EA: ethyl acetate; PE: petroleum ether; DMSO: dimethyl sulfoxide; BSA: bovine serum albumin; FA: formic acid; DDQ: 2,3-dichloro-5,6-dicyanide Benzoquinone.
本申请中涉及的百分比含量，如无特别说明，对于固液混合和固相-固相混合均指质量百分比，对于液相-液相混合指体积百分比。The percentage content involved in this application, unless otherwise specified, refers to mass percentage for solid-liquid mixing and solid-solid phase mixing, and refers to volume percentage for liquid-liquid phase mixing.
本申请的终产物可通过制备纯化得到，纯化分离条件是本领域技术人员依据化合物特性通过本领域常规知识试验得到。The final product of the present application can be obtained through preparation and purification, and the purification and separation conditions can be obtained by those skilled in the art based on the characteristics of the compound through tests with common knowledge in the art.
如本申请所用，室温是指约20-30℃。As used herein, room temperature refers to about 20-30°C.
收率＝实际合成产物质量/理论合成产物质量×100％。Yield=actual synthesis product mass/theoretical synthesis product mass×100%.
中间体a的制备
Preparation of intermediate a
步骤1:将2,6-二氯-4-甲基烟酸(40.00g,194.15mmol)和碳酸钾(67.08g,485.38mmol)溶于DMF(200mL)中，室温滴加碘甲烷(41.34g,291.23mmol)，然后24℃反应1小时， 将反应液倒入水中，用乙酸乙酯萃取，有机相用饱和食盐水洗两次，无水硫酸钠干燥，过滤，滤液浓缩得到棕色粗产物2,6-二氯-4-甲基烟酸甲酯(39.00g,收率：92.86％)。MS(ESI)220.0[M+H]+.Step 1: Dissolve 2,6-dichloro-4-methylnicotinic acid (40.00g, 194.15mmol) and potassium carbonate (67.08g, 485.38mmol) in DMF (200mL), add methyl iodide (41.34g) dropwise at room temperature ,291.23mmol), then react at 24°C for 1 hour, Pour the reaction solution into water, extract with ethyl acetate, wash the organic phase twice with saturated brine, dry over anhydrous sodium sulfate, filter, and concentrate the filtrate to obtain a brown crude product, 2,6-dichloro-4-methylnicotinic acid methyl. Ester (39.00g, yield: 92.86%). MS(ESI)220.0[M+H] + .
步骤2:将粗产物2,6-二氯-4-甲基烟酸甲酯(39.00g,177.23mmol)和N-溴代丁二酰亚胺(47.32g,265.85mmol)和过氧化苯甲酰(64.40g,265.85mmol)溶于四氯化碳CCl4(300mL)中，然后反应升到90℃，该反应在90℃下搅拌48小时。加入水，用二氯甲烷萃取，无水硫酸钠干燥，过滤，滤液浓缩得到粗产物。采用CombiFlash(0～30％，乙酸乙酯/石油醚)分离，成功得到黄色油状混合产物2,6-二氯-4-甲基烟酸-4-(溴甲基)-2,6-二氯烟酸甲酯(25.00g,收率：48.08％)和2,6-二氯-4-(二溴甲基)烟酸甲酯(25.00g,收率：37.88％)。MS(ESI)299.8[M+H]+，MS(ESI)379.7[M+H]+.Step 2: Combine the crude product 2,6-dichloro-4-methylnicotinic acid methyl ester (39.00g, 177.23mmol) and N-bromosuccinimide (47.32g, 265.85mmol) and benzyl peroxide The acyl (64.40 g, 265.85 mmol) was dissolved in carbon tetrachloride CCl 4 (300 mL) and the reaction was warmed to 90°C. The reaction was stirred at 90°C for 48 hours. Add water, extract with dichloromethane, dry over anhydrous sodium sulfate, filter, and concentrate the filtrate to obtain crude product. CombiFlash (0~30%, ethyl acetate/petroleum ether) was used to separate and successfully obtain the yellow oily mixed product 2,6-dichloro-4-methylnicotinic acid-4-(bromomethyl)-2,6-di Chloronicotinic acid methyl ester (25.00g, yield: 48.08%) and 2,6-dichloro-4-(dibromomethyl)nicotinic acid methyl ester (25.00g, yield: 37.88%). MS(ESI)299.8[M+H]+, MS(ESI)379.7[M+H] + .
步骤3:将粗产物2,6-二氯-4-甲基烟酸-4-(溴甲基)-2,6-二氯烟酸甲酯(25.00g,84.20mmol)和2,6-二氯-4-(二溴甲基)烟酸甲酯(25.00g,66.17mmol)溶于ACN(300mL)中，N2保护下降温至5℃，在此温度下滴加N,N-二异丙基乙胺(34.20g,264.66mmol)，然后滴加亚磷酸二乙酯(18.14g,132.33mmol)，5℃反应0.5小时，原料反应完全，加入饱和碳酸氢钠水溶液，用乙酸乙酯萃取，饱和食盐水洗两次，无水硫酸钠干燥，过滤，滤液浓缩，采用CombiFlash(0～20％，乙酸乙酯/石油醚)分离。成功得到黄色的油状产物2,6-二氯-4-甲基烟酸-4-(溴甲基)-2,6-二氯烟酸甲酯(28.00g,收率：70.78％)。MS(ESI)299.9[M+H]+.Step 3: Combine the crude product 2,6-dichloro-4-methylnicotinic acid-4-(bromomethyl)-2,6-dichloronicotinic acid methyl ester (25.00g, 84.20mmol) and 2,6- Dichloro-4-(dibromomethyl)nicotinic acid methyl ester (25.00g, 66.17mmol) was dissolved in ACN (300mL), and the temperature was lowered to 5°C under N 2 protection. At this temperature, N, N-dioic acid was added dropwise. Isopropylethylamine (34.20g, 264.66mmol), then dropwise add diethyl phosphite (18.14g, 132.33mmol), react at 5°C for 0.5 hours, the raw material reaction is complete, add saturated sodium bicarbonate aqueous solution, and use ethyl acetate Extract, wash twice with saturated brine, dry over anhydrous sodium sulfate, filter, concentrate the filtrate, and separate using CombiFlash (0-20%, ethyl acetate/petroleum ether). The yellow oily product 2,6-dichloro-4-methylnicotinic acid-4-(bromomethyl)-2,6-dichloronicotinic acid methyl ester (28.00g, yield: 70.78%) was successfully obtained. MS(ESI)299.9[M+H] + .
步骤4:将2,6-二氯-4-甲基烟酸-4-(溴甲基)-2,6-二氯烟酸甲酯(28.00g,93.66mmol)和(S)-1-环丙基乙胺盐酸盐(11.39g,93.66mmol)溶于乙腈(200mL)中，然后将硼酸(5.79g,93.66mmol)和碳酸钾(25.89g,187.32mmol)加入其中。该反应在24℃条件下搅拌12小时。将悬浊液过滤，滤液旋干得到粗产物。采用CombiFlash(0～50％，乙酸乙酯/石油醚)分离，得到白色固体产物中间体a(22.60g,收率：88.99％)。MS(ESI)271.0[M+H]+.Step 4: Combine 2,6-dichloro-4-methylnicotinic acid-4-(bromomethyl)-2,6-dichloronicotinic acid methyl ester (28.00g, 93.66mmol) and (S)-1- Cyclopropylethylamine hydrochloride (11.39g, 93.66mmol) was dissolved in acetonitrile (200mL), and then boric acid (5.79g, 93.66mmol) and potassium carbonate (25.89g, 187.32mmol) were added. The reaction was stirred at 24°C for 12 hours. The suspension was filtered, and the filtrate was spun to dryness to obtain crude product. Use CombiFlash (0-50%, ethyl acetate/petroleum ether) to separate, and obtain the white solid product intermediate a (22.60g, yield: 88.99%). MS(ESI)271.0[M+H] + .
中间体b的制备
Preparation of intermediate b
取n-BuLi(2.5M in THF,5.12mL,12.8mmol),在0℃下滴加入二异丙基氨(1.29g,12.8mmol)的THF(20mL)溶液中,保持0℃反应30分钟。在-78℃下，将上述溶液缓慢滴加入N-(4-甲基噻唑-2-基)乙酰胺(500mg,3.2mmol)的THF(5mL)溶液中，继续反应30分钟。将2-异丙基-4,4,5,5-四甲基-1,3,2-二氧硼烷(1.19g,6.4mmol)缓慢滴入上述反应液中，保持-78℃反应两个小时。用饱和氯化铵溶液淬灭，乙酸乙酯萃取，食盐水洗涤有机相两遍，硫酸钠干燥，过滤浓缩，得到淡黄色固体产物中间体b(410mg,收率：45％)。1HNMR(400MHz,CDCl3)δ2.53(s,3H),2.25(d,J＝2.4Hz,3H),1.25(s,12H).Take n-BuLi (2.5M in THF, 5.12mL, 12.8mmol), add it dropwise to a solution of diisopropylamine (1.29g, 12.8mmol) in THF (20mL) at 0°C, and maintain the reaction at 0°C for 30 minutes. At -78°C, the above solution was slowly added dropwise to a solution of N-(4-methylthiazol-2-yl)acetamide (500 mg, 3.2 mmol) in THF (5 mL), and the reaction was continued for 30 minutes. Slowly drop 2-isopropyl-4,4,5,5-tetramethyl-1,3,2-dioxaborane (1.19g, 6.4mmol) into the above reaction solution and keep it at -78°C for two seconds. Hours. Quench with saturated ammonium chloride solution, extract with ethyl acetate, wash the organic phase twice with brine, dry with sodium sulfate, filter and concentrate to obtain the light yellow solid product intermediate b (410 mg, yield: 45%). 1 HNMR (400MHz, CDCl 3 ) δ2.53 (s, 3H), 2.25 (d, J = 2.4Hz, 3H), 1.25 (s, 12H).
中间体1的制备
Preparation of intermediate 1
中间体a(1.200g,4.43mmol)和中间体b(3.75g,13.28mmol)溶解在N,N-二甲基甲酰胺(20mL)中，氩气保护下加入双(2-二苯基膦基环戊烷-2,4-二烯-1-基)铁二氯钯(323.83mg,442.57μmol)，碳酸钠(1.41g,13.28mmol)，升温至100℃搅拌10小时。LC-MS检测发现目标产物。反应液倒入水中，二氯甲烷(50ml×2)萃取，无水硫酸钠干燥，减压浓缩干，硅胶柱分离纯化(EA：PE＝0～100％)得到黄色固体中间体1(380mg,972.15μmol,收率：21.97％)。MS(ESI)391[M+H]+.Intermediate a (1.200g, 4.43mmol) and intermediate b (3.75g, 13.28mmol) were dissolved in N,N-dimethylformamide (20mL), and bis(2-diphenylphosphine) was added under argon protection. Cyclopentane-2,4-dien-1-yl)iron palladium dichloride (323.83 mg, 442.57 μmol) and sodium carbonate (1.41 g, 13.28 mmol) were heated to 100°C and stirred for 10 hours. LC-MS detection found the target product. The reaction solution was poured into water, extracted with dichloromethane (50ml×2), dried over anhydrous sodium sulfate, concentrated to dryness under reduced pressure, and separated and purified on a silica gel column (EA: PE=0~100%) to obtain yellow solid intermediate 1 (380mg, 972.15 μmol, yield: 21.97%). MS(ESI)391[M+H] + .
中间体2的制备
Preparation of intermediate 2
将6-溴吲哚啉-2-酮(500mg，2.36mmol,1eq)、Pin2B2(719mg，2.83mmol,1.2eq)、KOAc(463mg，4.72mmol,2eq)溶于1,4-二氧六环(20mL)中，氩气置换，加入Pd(dppf)Cl2(176mg，0.24mmol,0.1eq)，氩气置换，氩气保护状态下，120℃搅拌反应6小时。LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(30mL×3)萃取三次。有机相用饱和氯化钠水溶液洗涤一次，无水硫酸钠干燥，旋干得黑灰色固体中间体2(1.1g，粗品)。可直接投下一步。Dissolve 6-bromoindolin-2-one (500mg, 2.36mmol, 1eq), Pin 2 B 2 (719mg, 2.83mmol, 1.2eq), KOAc (463mg, 4.72mmol, 2eq) in 1,4-bis In oxygen hexacyclic ring (20 mL), replace with argon, add Pd(dppf)Cl 2 (176 mg, 0.24 mmol, 0.1eq), replace with argon, and stir and react at 120°C for 6 hours under argon protection. LCMS detection showed that the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL), and extracted three times with dichloromethane (30 mL × 3). The organic phase was washed once with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, and spin-dried to obtain black-gray solid intermediate 2 (1.1 g, crude product). You can directly vote for the next step.
中间体3的制备
Preparation of intermediate 3
将6-溴异吲哚啉-1-酮(500mg，2.36mmol,1eq)、Pin2B2(719mg，2.83mmol,1.2eq)、KOAc(463mg，4.72mmol,2eq)溶于1,4-二氧六环(20mL)中，氩气置换，加入Pd(dppf)Cl2(176mg，0.24mmol,0.1eq)，氩气置换，氩气保护状态下，120℃搅拌反应6小时。LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(30mL×3)萃取三次。有机相用饱和氯化钠水溶液洗涤一次，无水硫酸钠干燥，旋干得1g黑灰色粗品固体中间体3(1g，粗品)。直接投下一步。Dissolve 6-bromoisoindolin-1-one (500mg, 2.36mmol, 1eq), Pin 2 B 2 (719mg, 2.83mmol, 1.2eq), KOAc (463mg, 4.72mmol, 2eq) in 1,4- In dioxane (20 mL), replace with argon, add Pd(dppf)Cl 2 (176 mg, 0.24 mmol, 0.1eq), replace with argon, stir and react at 120°C for 6 hours under argon protection. LCMS detection showed that the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL), and extracted three times with dichloromethane (30 mL × 3). The organic phase was washed once with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, and spin-dried to obtain 1 g of black-gray crude solid intermediate 3 (1 g, crude product). Directly vote for the next step.
中间体4的制备
Preparation of intermediate 4
将4-溴-2-氟吡啶(2g，11.36mmol,1eq)、Pin2B2(3.463g,13.64mmol,1.2eq)、KOAc(2.23g，22.72mmol,2eq)溶于1,4-二氧六环(20mL)中，氩气置换，加入Pd(dppf)Cl2(416mg，0.568mmol，0.1eq)，氩气置换，氩气保护状态下，120℃下搅拌5小时。送LCMS检测反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(30mL×3)萃取三次。有机相用饱和氯化钠水溶液洗涤一次，无水硫酸钠干燥，旋干得粗产物中间体4(2.4g，粗品)。Dissolve 4-bromo-2-fluoropyridine (2g, 11.36mmol, 1eq), Pin 2 B 2 (3.463g, 13.64mmol, 1.2eq), and KOAc (2.23g, 22.72mmol, 2eq) in 1,4-bis In oxyhexanes (20 mL), replace with argon, add Pd(dppf)Cl 2 (416 mg, 0.568 mmol, 0.1 eq), replace with argon, and stir at 120°C for 5 hours under argon protection. Send LCMS to detect the success of the reaction. After the reaction solution was cooled, it was diluted with water (20 mL), and extracted three times with dichloromethane (30 mL × 3). The organic phase was washed once with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, and spin-dried to obtain crude product Intermediate 4 (2.4 g, crude product).
中间体5的制备
Preparation of intermediate 5
步骤1：将6-氯吡唑并吡啶(1g，6.51mmol,1eq)溶于DMF(10mL)中，氩气保护，-5℃下加NaH(188mg，7.81mmol,1.2eq，60wt％)，反应10分钟，-5℃下滴加SEM-Cl(1.3g，7.81mmol,1.2eq)，加完撤走冰浴，恢复至室温(rt)，反应3小时10分钟。送LCMS检测反应成功。将反应液倒入适量冰水中，乙酸乙酯(30mL×3)萃取3次，饱和氯化钠水溶液(30mL)洗涤1次。合并有机相用无水硫酸钠干燥，旋干，过柱纯化得产物6-氯-1-(2-三甲基硅基乙氧基)甲基吡唑并吡啶(1.3g,纯度:88％，收率:62％，呈淡黄色液体)。Step 1: Dissolve 6-chloropyrazolopyridine (1g, 6.51mmol, 1eq) in DMF (10mL), protect it with argon, add NaH (188mg, 7.81mmol, 1.2eq, 60wt%) at -5°C, React for 10 minutes, add SEM-Cl (1.3g, 7.81mmol, 1.2eq) dropwise at -5°C, remove the ice bath after adding, return to room temperature (rt), and react for 3 hours and 10 minutes. Send LCMS to detect the success of the reaction. Pour the reaction solution into an appropriate amount of ice water, extract three times with ethyl acetate (30 mL × 3), and wash once with saturated aqueous sodium chloride solution (30 mL). The combined organic phases were dried over anhydrous sodium sulfate, spun to dryness, and purified through column to obtain the product 6-chloro-1-(2-trimethylsilylethoxy)methylpyrazopyridine (1.3g, purity: 88% , Yield: 62%, light yellow liquid).
步骤2：将6-氯-1-(2-三甲基硅基乙氧基)甲基吡唑并吡啶(1.3g，4.58mmol,1eq)、Pin2B2(1.4g，5.50mmol,1.2eq)、KOAc(899mg，9.16mmol,2eq)溶于DMF(20mL)中，氩气置换，加入Pd(dppf)Cl2(335mg，0.4mmol,0.1eq)，氩气置换，氩气保护状态下，100℃搅拌反应5小时。送LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(30mL×3)萃取三次。有机相用饱和氯化钠水溶液洗涤一次，无水硫酸钠干燥，旋干得中间体5(1.9g，纯度:77％，收率：99％，呈黑灰色固体)。Step 2: Combine 6-chloro-1-(2-trimethylsilylethoxy)methylpyrazolopyridine (1.3g, 4.58mmol, 1eq), Pin 2 B 2 (1.4g, 5.50mmol, 1.2 eq), KOAc (899mg, 9.16mmol, 2eq) were dissolved in DMF (20mL), replaced with argon, added Pd(dppf)Cl 2 (335mg, 0.4mmol, 0.1eq), replaced with argon, under argon protection. , stirring reaction at 100°C for 5 hours. Sent LCMS for testing and the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL), and extracted three times with dichloromethane (30 mL × 3). The organic phase was washed once with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, and spin-dried to obtain intermediate 5 (1.9 g, purity: 77%, yield: 99%, black-gray solid).
中间体6的制备
Preparation of intermediate 6
步骤1：将6-氯吡咯(4.5g，29.49mmol,1eq)溶于DMF(40mL)中,氩气置换，氩气保护，-5℃下加NaH(60wt％)(849mg，35.39mmol,1.2eq)，反应10分钟，-5℃下滴加SEM-Cl(5.9g，35.39mmol,1.2eq)，加完撤走冰浴，恢复至室温(RT)，反应3小时。送LCMS检测反应成功。将反应液倒入冰水中，乙酸乙酯萃取3次，氯化钠水洗1次，无水硫酸钠干燥，旋干过柱纯化得产物6-氯-1-(2-三甲基硅基乙氧基)甲基吡咯烷(8g，纯度:80.08％，收率:76.80％，呈淡黄色液体)。Step 1: Dissolve 6-chloropyrrole (4.5g, 29.49mmol, 1eq) in DMF (40mL), replace with argon, protect with argon, and add NaH (60wt%) (849mg, 35.39mmol, 1.2) at -5°C. eq), react for 10 minutes, add SEM-Cl (5.9g, 35.39mmol, 1.2eq) dropwise at -5°C, remove the ice bath after adding, return to room temperature (RT), and react for 3 hours. Send LCMS to detect the success of the reaction. The reaction solution was poured into ice water, extracted three times with ethyl acetate, washed once with sodium chloride, dried over anhydrous sodium sulfate, spin-dried and purified through column to obtain the product 6-chloro-1-(2-trimethylsilylethane) Oxy)methylpyrrolidine (8g, purity: 80.08%, yield: 76.80%, light yellow liquid).
步骤2：将6-氯-1-(2-三甲基硅基乙氧基)甲基吡咯烷(3g,10.61mmol，1eq)、Pin2B2(3.2g，12.73mmol,1.2eq)、KOAc(2.1g，21.22mmol,2eq)溶于1,4-二氧六环(30mL)中，氩气置换，加入Pd(dppf)Cl2(776mg，1.06mmol,0.1eq)，氩气置换，氩气保护状态下，90℃搅拌反应5小时。送LCMS检测，反应液成功。反应液冷却后，二氯甲烷与水萃取，饱和氯化钠水洗，无水硫酸钠干燥，旋干得中间体6(3.5g，纯度:76.53％，收率:86.42％，呈黑灰 色固体)。Step 2: Combine 6-chloro-1-(2-trimethylsilylethoxy)methylpyrrolidine (3g, 10.61mmol, 1eq), Pin 2 B 2 (3.2g, 12.73mmol, 1.2eq), KOAc (2.1g, 21.22mmol, 2eq) was dissolved in 1,4-dioxane (30mL), replaced with argon, added Pd(dppf)Cl 2 (776mg, 1.06mmol, 0.1eq), replaced with argon, Under argon protection, the reaction was stirred at 90°C for 5 hours. Sent LCMS for testing and the reaction solution was successful. After the reaction solution was cooled, extracted with dichloromethane and water, washed with saturated sodium chloride, dried with anhydrous sodium sulfate, and spin-dried to obtain intermediate 6 (3.5g, purity: 76.53%, yield: 86.42%, in the form of black gray color solid).
中间体7的制备
Preparation of intermediate 7
步骤1：将6-溴-1H-吡唑并吡啶(4.5g，22.73mmol,1eq)溶于DMF(40mL)中,氩气置换，氩气保护，-5℃下加NaH(60wt％)(654mg，27.27mmol,1.2eq)，反应10分钟，-5℃下滴加SEM-Cl(4.5g，27.27mmol,1.2eq)，加完撤走冰浴，恢复至室温，反应3小时。送LCMS检测反应成功。将反应液倒入冰水中，乙酸乙酯萃取3次，氯化钠水洗1次，无水硫酸钠干燥，旋干过柱纯化得产物6-溴-1-(2-三甲基硅基乙氧基)甲基-1H-吡唑并[4,3-b]吡啶(4.3g，纯度:97.50％，收率:56.20％，呈淡黄色液体)。Step 1: Dissolve 6-bromo-1H-pyrazolopyridine (4.5g, 22.73mmol, 1eq) in DMF (40mL), replace with argon, protect with argon, and add NaH (60wt%) at -5°C ( 654mg, 27.27mmol, 1.2eq), react for 10 minutes, add SEM-Cl (4.5g, 27.27mmol, 1.2eq) dropwise at -5°C, remove the ice bath after adding, return to room temperature, and react for 3 hours. Send LCMS to detect the success of the reaction. The reaction solution was poured into ice water, extracted three times with ethyl acetate, washed once with sodium chloride, dried over anhydrous sodium sulfate, spin-dried and purified through column to obtain the product 6-bromo-1-(2-trimethylsilylethane) Oxy)methyl-1H-pyrazolo[4,3-b]pyridine (4.3g, purity: 97.50%, yield: 56.20%, light yellow liquid).
步骤2：将6-溴-1-(2-三甲基硅基乙氧基)甲基-1H-吡唑并[4,3-b]吡啶(4.3g,13.098mmol，1eq)、Pin2B2(3.991g，15.718mmol,1.2eq)、KOAc(2.571g，26.196mmol,2eq)溶于1,4-二氧六环(100mL)中，氩气置换，加入Pd(dppf)Cl2(479mg，0.655mmol,0.05eq)，氩气置换，氩气保护状态下，120℃搅拌反应12小时。送LCMS检测，反应液成功。反应液冷却后，二氯甲烷与水萃取，饱和氯化钠水洗，无水硫酸钠干燥，旋干得中间体7(5.9g，纯度:62.90％，收率:98.68％)。Step 2: Combine 6-bromo-1-(2-trimethylsilylethoxy)methyl-1H-pyrazolo[4,3-b]pyridine (4.3g, 13.098mmol, 1eq), Pin 2 B 2 (3.991g, 15.718mmol, 1.2eq) and KOAc (2.571g, 26.196mmol, 2eq) were dissolved in 1,4-dioxane (100mL), replaced with argon, and Pd(dppf)Cl 2 ( 479 mg, 0.655 mmol, 0.05 eq), replaced with argon, stirred and reacted at 120°C for 12 hours under argon protection. Sent LCMS for testing and the reaction solution was successful. After the reaction solution was cooled, it was extracted with dichloromethane and water, washed with saturated sodium chloride, dried with anhydrous sodium sulfate, and spin-dried to obtain Intermediate 7 (5.9g, purity: 62.90%, yield: 98.68%).
中间体8的制备
Preparation of intermediate 8
步骤1：将6-氯-1H-吡唑[3,4-b]吡嗪(2g，13mmol,1eq)溶于DMF(50mL)中,冰盐浴(-5℃)，分批加入NaH(0.67g,16.9mmol,1.3eq,60wt％),搅拌30分钟，滴加SEM-Cl(2.6g,15.6mmol,1.2eq),氩气保护，恢复至室温，反应3小时，加冰水(30mL)淬灭，乙酸乙酯(50mL×3)萃取三次，分液，无水硫酸钠干燥，过滤旋干，得产物6-氯-1-((2-(三甲基硅基)乙氧基)甲基)-1h-吡唑啉[3,4-b]吡嗪(2.3g,粗品)，棕色油状产物。产物直接用于下一步反应。Step 1: Dissolve 6-chloro-1H-pyrazole[3,4-b]pyrazine (2g, 13mmol, 1eq) in DMF (50mL), put it in an ice-salt bath (-5°C), and add NaH ( 0.67g, 16.9mmol, 1.3eq, 60wt%), stir for 30 minutes, add SEM-Cl (2.6g, 15.6mmol, 1.2eq) dropwise, protect with argon, return to room temperature, react for 3 hours, add ice water (30mL ), quenched, extracted three times with ethyl acetate (50mL )Methyl)-1h-pyrazoline[3,4-b]pyrazine (2.3g, crude product), brown oily product. The product was directly used in the next reaction.
步骤2：将6-氯-1-((2-(三甲基硅基)乙氧基)甲基)-1h-吡唑啉[3,4-b]吡嗪(1.5g,5.3mmol,1eq)溶于1,4-二氧六环(50mL)中，加入Pin2B2(1.74g,6.84mmol,1.3eq)，Pd(dppf)Cl2)(384mg,0.53mmol,0.1eq)，KOAc(1.56g,15.9mmol,3eq),氩气置换三次，油浴升温至120℃反应12小时。反应液冷却后加水(30mL)稀释，用乙酸乙酯(50mL×3)萃取三次，分液，无水硫酸钠干燥，过滤旋干，得中间体8(1.8g,粗品)，棕色油状产物。直接用于下一步反应。1H NMR(400MHz,DMSO-d6)δ8.49(s,1H),8.45(s,1H),5.78(s,2H),3.95–3.90(m,2H),1.03(dd,J＝14.1,6.0Hz,3H),0.08(d,J＝6.4Hz,12H),-0.00(s,9H).Step 2: Add 6-chloro-1-((2-(trimethylsilyl)ethoxy)methyl)-1h-pyrazoline[3,4-b]pyrazine (1.5g, 5.3mmol, 1eq) was dissolved in 1,4-dioxane (50mL), add Pin 2 B 2 (1.74g, 6.84mmol, 1.3eq), Pd(dppf)Cl 2 ) (384mg, 0.53mmol, 0.1eq), KOAc (1.56g, 15.9mmol, 3eq), replaced with argon three times, heated the oil bath to 120°C and reacted for 12 hours. After cooling, the reaction solution was diluted with water (30 mL), extracted three times with ethyl acetate (50 mL × 3), separated, dried over anhydrous sodium sulfate, filtered and spin-dried to obtain Intermediate 8 (1.8 g, crude product), a brown oily product. used directly for the next reaction. 1 H NMR (400MHz, DMSO-d 6 ) δ8.49 (s, 1H), 8.45 (s, 1H), 5.78 (s, 2H), 3.95–3.90 (m, 2H), 1.03 (dd, J = 14.1 ,6.0Hz,3H),0.08(d,J＝6.4Hz,12H),-0.00(s,9H).
中间体9的制备
Preparation of intermediate 9
将5-溴-2-甲基吡啶(1g，5.81mmol,1eq)、Pin2B2(1.771g,6.98mmol,1.2eq)、KOAc(1.140g，11.63mmol,2eq)溶于1,4-二氧六环(20mL)中，氩气置换，加入Pd(dppf)Cl2(425mg，0.581mmol，0.05eq)，氩气置换，氩气保护状态下，120℃下搅拌12小时。送LCMS检测反应成功。反应液冷却后，将反应液转移水中(100mL)，用二氯甲烷(200mL)萃取，萃取3次。将收集的有机相用无水硫酸钠干燥，旋干得中间体9(2g，粗品)。Dissolve 5-bromo-2-methylpyridine (1g, 5.81mmol, 1eq), Pin 2 B 2 (1.771g, 6.98mmol, 1.2eq), KOAc (1.140g, 11.63mmol, 2eq) in 1,4- In dioxane (20 mL), replace with argon, add Pd(dppf)Cl 2 (425 mg, 0.581 mmol, 0.05 eq), replace with argon, and stir at 120°C for 12 hours under argon protection. Send LCMS to detect the success of the reaction. After the reaction solution was cooled, the reaction solution was transferred to water (100 mL), and extracted with dichloromethane (200 mL) three times. The collected organic phase was dried over anhydrous sodium sulfate and spun to dryness to obtain intermediate 9 (2g, crude product).
中间体10的制备
Preparation of intermediate 10
将4-溴-2-甲基吡啶(1.5g，8.7mmol,1eq)溶于1,4-二氧六环(20mL)中,加入Pin2B2(2.88g,11.3mmol,1.3eq),Pd(dppf)Cl2(631mg,0.87mmol)，KOAc(2.13g,21.7mmol)，氩气置换三次，油浴升温至120℃反应12小时。反应液冷却后加水(30mL)稀释，用乙酸乙酯(50mL×3)萃取三次，分液，无水硫酸钠干燥，过滤旋干，得中间体10(3.2g,粗品)，棕色油状产物。产物直接用于下一步反应。LCMS：MS m/z(ESI):138.1[M+H]+.Dissolve 4-bromo-2-methylpyridine (1.5g, 8.7mmol, 1eq) in 1,4-dioxane (20mL), add Pin 2 B 2 (2.88g, 11.3mmol, 1.3eq), Pd(dppf)Cl 2 (631 mg, 0.87 mmol), KOAc (2.13 g, 21.7 mmol), replaced with argon three times, heated the oil bath to 120°C and reacted for 12 hours. After cooling, the reaction solution was diluted with water (30 mL), extracted three times with ethyl acetate (50 mL × 3), separated, dried over anhydrous sodium sulfate, filtered and spin-dried to obtain Intermediate 10 (3.2 g, crude product), a brown oily product. The product was directly used in the next reaction. LCMS: MS m/z(ESI):138.1[M+H] + .
中间体11a和11b的制备
Preparation of Intermediates 11a and 11b
将5-溴-2-环丙基吡啶(1g，5.049mmol,1eq)、Pin2B2(1.539g,6.059mmol,1.2eq)、KOAc(991mg，10.098mmol,2eq)溶于1,4-二氧六环(20mL)中，氩气置换。再加入Pd(dppf)Cl2(369mg，0.505mmol，0.1eq)，氩气置换，氩气保护状态下，120℃下搅拌12小时。送LCMS检测反应成功。反应液冷却后，将反应液转移至200mL水中，用二氯甲烷(200mL)萃取，萃取3次。将收集的有机相用无水硫酸钠干燥，旋干，得中间体11a和11b的混合物(2g，粗品)。Dissolve 5-bromo-2-cyclopropylpyridine (1g, 5.049mmol, 1eq), Pin 2 B 2 (1.539g, 6.059mmol, 1.2eq), KOAc (991mg, 10.098mmol, 2eq) in 1,4- Dioxane (20 mL) was replaced with argon. Then add Pd(dppf)Cl 2 (369 mg, 0.505 mmol, 0.1 eq), replace with argon, and stir at 120°C for 12 hours under argon protection. Send LCMS to detect the success of the reaction. After the reaction liquid was cooled, the reaction liquid was transferred to 200 mL of water, and extracted with dichloromethane (200 mL) three times. The collected organic phase was dried over anhydrous sodium sulfate and spun to dryness to obtain a mixture of intermediates 11a and 11b (2g, crude product).
中间体12的制备
Preparation of intermediate 12
将5-氯-2-三氟甲基吡啶(1g，5.508mmol,1eq)、Pin2B2(1.678g,6.610mmol,1.2eq)、KOAc(1.081g，11.016mmol,2eq)溶于1,4-二氧六环(20mL)中，氩气置换。再加入Pd(dppf)Cl2(403mg，0.5508mmol，0.1eq)，氩气置换，氩气保护状态下，120℃下搅拌5小时。送LCMS检测反应成功。反应液冷却后，将反应液转移至200mL水中，用二氯甲烷(200mL)萃取，萃取3次。将收集的有机相用无水硫酸钠干燥，旋干，过反相柱纯化得中间体12(480mg，2.51mmol，收率:45.64％)。Dissolve 5-chloro-2-trifluoromethylpyridine (1g, 5.508mmol, 1eq), Pin 2 B 2 (1.678g, 6.610mmol, 1.2eq), KOAc (1.081g, 11.016mmol, 2eq) in 1, 4-Dioxane (20 mL) was replaced with argon. Then add Pd(dppf)Cl 2 (403 mg, 0.5508 mmol, 0.1 eq), replace with argon, and stir at 120°C for 5 hours under argon protection. Send LCMS to detect the success of the reaction. After the reaction liquid was cooled, the reaction liquid was transferred to 200 mL of water, and extracted with dichloromethane (200 mL) three times. The collected organic phase was dried over anhydrous sodium sulfate, spun to dryness, and purified through a reversed-phase column to obtain intermediate 12 (480 mg, 2.51 mmol, yield: 45.64%).
中间体13的制备
Preparation of intermediate 13
将7-溴-1,4-二氢异喹啉-3(2H)-酮(500mg，2.2mmol)溶于1,4-二氧六环(25mL)中，加入Pin2B2(620mg,2.4mmol)，Pd(dppf)Cl2(80mg,0.11mmol)，KOAc(755mg,7.7mmol)，dppf(60mg,0.11mmol)，氩气保护，120℃反应12小时。反应液放至室温，过滤，减压浓缩，得700mg产物中间体13，棕色油(0.7g，粗品)。可直接投下一步。LCMS：MS m/z(ESI):274.2[M+H]+.Dissolve 7-bromo-1,4-dihydroisoquinolin-3(2H)-one (500mg, 2.2mmol) in 1,4-dioxane (25mL), and add Pin 2 B 2 (620mg, 2.4mmol), Pd(dppf)Cl 2 (80mg, 0.11mmol), KOAc (755mg, 7.7mmol), dppf (60mg, 0.11mmol), argon protection, reaction at 120°C for 12 hours. The reaction solution was brought to room temperature, filtered, and concentrated under reduced pressure to obtain 700 mg of product intermediate 13, a brown oil (0.7 g, crude product). You can directly vote for the next step. LCMS: MS m/z(ESI):274.2[M+H] + .
中间体14的制备
Preparation of intermediate 14
将4-氯-2-三氟甲基吡啶(1g，5.52mmol,1eq)置于100mL的洁净三口烧瓶中,室温下依次加入Pin2B2(2.8g,11.00mmol,2eq)，KOAc(1.6g,16.56mmol,3eq)和Pd(dppf)Cl2(0.6g,0.83mmoL,0.15eq)，加入1,4-二氧六环(20mL)溶解，120℃搅拌12小时。冷却至室温，向反应液中加入20mL水，乙酸乙酯(30mL×3)萃取三次，有机相中加入氯化钠饱和溶液洗涤，分液，有机相用无水硫酸钠干燥，过滤浓缩。得到粗产品中间体14(2.1g,粗品)，呈黑色。产物直接用于下一步反应。LCMS：MS m/z(ESI):274.0[M+H]+.Place 4-chloro-2-trifluoromethylpyridine (1g, 5.52mmol, 1eq) into a 100mL clean three-necked flask, and add Pin 2 B 2 (2.8g, 11.00mmol, 2eq) and KOAc (1.6) at room temperature. g, 16.56mmol, 3eq) and Pd(dppf)Cl 2 (0.6g, 0.83mmoL, 0.15eq), add 1,4-dioxane (20mL) to dissolve, and stir at 120°C for 12 hours. Cool to room temperature, add 20 mL of water to the reaction solution, and extract three times with ethyl acetate (30 mL × 3). Add a saturated sodium chloride solution to the organic phase to wash and separate the layers. The organic phase is dried over anhydrous sodium sulfate, filtered and concentrated. The crude product intermediate 14 (2.1 g, crude product) was obtained, which was black. The product was directly used in the next reaction. LCMS: MS m/z(ESI):274.0[M+H] + .
中间体15的制备
Preparation of intermediate 15
将5-氯-2-氟吡啶(1.0g，7.63mmol,1eq)置于100mL的洁净三口烧瓶中，室温下依次加入Pin2B2(3.88g,15.3mmol,2eq)，KOAc(2.24g,22.9mmol,3eq)和Pd(dppf)Cl2(0.83g,1.14mmoL,0.15eq)，加1,4-二氧六环(20mL)溶解，120℃搅拌12小时。冷却至室温，向反应液中加入20mL水，加乙酸乙酯(30mL×3)萃取三次，有机相中加入氯化钠饱和溶液洗涤，分液，有机相用无水硫酸钠干燥，过滤浓缩。得到中间体15(2.3g,粗品)，呈黑色油。产物直接用于下一步反应。LCMS：MS m/z(ESI):224.1[M+H]+.Place 5-chloro-2-fluoropyridine (1.0g, 7.63mmol, 1eq) into a 100mL clean three-necked flask, and add Pin 2 B 2 (3.88g, 15.3mmol, 2eq) and KOAc (2.24g, 2.24g, 22.9mmol, 3eq) and Pd(dppf)Cl 2 (0.83g, 1.14mmoL, 0.15eq), add 1,4-dioxane (20mL) to dissolve, and stir at 120°C for 12 hours. Cool to room temperature, add 20 mL of water to the reaction solution, add ethyl acetate (30 mL × 3) for extraction three times, add saturated sodium chloride solution to the organic phase to wash, separate the layers, dry the organic phase over anhydrous sodium sulfate, filter and concentrate. Intermediate 15 (2.3 g, crude product) was obtained as black oil. The product was directly used in the next reaction. LCMS: MS m/z(ESI):224.1[M+H] + .
中间体16的制备
Preparation of intermediate 16
步骤1：将原料4-溴吡啶(2g，12.6mmol，1eq)，置于250mL单口瓶中，加入100mL THF溶解，氩气保护，降温至-78℃，开始滴加环丙基溴化镁(40mL，25.2mmol，2eq)，温度保持-78℃，滴加完搅拌5分钟，开始滴加氯甲酸苯酯(2.4mL，12.6mmol，1eq)，滴加完毕，反应10分钟，恢复室温，反应液用乙酸乙酯(150mL)稀释，分别用水(30mL)，0.1M(mol/L)盐酸(30mL)，盐水(30mL)分别洗涤，有机相干燥旋干，得粗品2.1g。将上述粗品放入250mL单口瓶中，加入80mL甲苯溶解，加入DDQ(2.9g，12.6mmol,1eq)，室温搅拌过夜。LCMS取样检测原料反应完全，用1M NaOH(40mL)淬灭，调节pH＝7，分层，水相乙酸乙酯(50×3)萃取三次，合并有机相，盐水洗涤，分液，有机相无水硫酸钠干燥，旋干，过正相柱(PE：EA＝7:1)纯化得2-环丙基-4-溴吡啶(1.5g，收率：86％)。LCMS：MS m/z(ESI):198.0[M+H]+。Step 1: Place the raw material 4-bromopyridine (2g, 12.6mmol, 1eq) in a 250mL single-mouth bottle, add 100mL THF to dissolve, protect with argon, cool to -78°C, and start adding cyclopropylmagnesium bromide ( 40mL, 25.2mmol, 2eq), keep the temperature at -78°C, stir for 5 minutes after the dropwise addition, start to dropwise add phenyl chloroformate (2.4mL, 12.6mmol, 1eq), complete the dropwise addition, react for 10 minutes, return to room temperature, react The solution was diluted with ethyl acetate (150 mL), washed with water (30 mL), 0.1 M (mol/L) hydrochloric acid (30 mL), and brine (30 mL) respectively, and the organic phase was dried and spin-dried to obtain 2.1 g of crude product. Put the above crude product into a 250 mL single-neck bottle, add 80 mL of toluene to dissolve, add DDQ (2.9 g, 12.6 mmol, 1 eq), and stir at room temperature overnight. LCMS sampling was used to detect the complete reaction of the raw materials, quenched with 1M NaOH (40 mL), adjusted to pH = 7, separated into layers, and the aqueous phase was extracted three times with ethyl acetate (50×3). The organic phases were combined, washed with brine, and separated. The organic phase was Dry over sodium sulfate, spin to dryness, and purify through normal phase column (PE:EA=7:1) to obtain 2-cyclopropyl-4-bromopyridine (1.5g, yield: 86%). LCMS: MS m/z(ESI):198.0[M+H] + .
步骤2：将原料2-环丙基-4-溴吡啶(1.5g，7.5mmol,，1eq)，Pin2B2(2.3g，9mmol,，1.2eq)，乙酸钾(1.5g，15mmol，2eq)，置于250mL三口瓶中，加入75mL1,4-二氧六环，氩气置换，加入Pd(dppf)Cl2(280mg，0.39mmol，0.05eq)，氩气置换，搅拌升温至120℃，反应12小时，LCMS取样检测，LCMS显示原料反应完全，产物LCMS不显示。反应液加入50mL水，100mL乙酸乙酯，分层，水相EA萃取三次，合并有机相，盐水洗涤，分层，有机相干燥脱溶。将产物直接用于下一步投料。Step 2: Combine the raw materials 2-cyclopropyl-4-bromopyridine (1.5g, 7.5mmol, 1eq), Pin 2 B 2 (2.3g, 9mmol, 1.2eq), potassium acetate (1.5g, 15mmol, 2eq) ), place it in a 250mL three-necked flask, add 75mL of 1,4-dioxane, replace with argon, add Pd(dppf)Cl 2 (280mg, 0.39mmol, 0.05eq), replace with argon, stir and heat to 120°C. After 12 hours of reaction, LCMS sampling was performed. LCMS showed that the raw material was completely reacted, but LCMS of the product did not show it. Add 50 mL of water and 100 mL of ethyl acetate to the reaction solution, separate the layers, extract the aqueous phase with EA three times, combine the organic phases, wash with brine, separate the layers, and dry and remove the organic phase. The product is directly used for the next step of feeding.
实施例1化合物S1的制备
Example 1 Preparation of Compound S1
将中间体1(200mg，0.51mmol)和3-吡啶硼酸(76mg，0.61mmol)溶于1,4-二氧六环(20mL)和水(4mL)中，然后加入[1,1'-双(二苯基膦)二茂铁]二氯化钯二氯甲烷络合物(42mg，0.05mmol)和碳酸钾(142mg，1.02mmol)，反应液在氮气保护下100℃搅拌2小时。反应液冷却至室温，过滤，滤液用乙酸乙酯和水萃取三次，合并有机相，用饱和食盐水洗涤，用无水硫酸钠干燥，过滤，滤液浓缩后经制备纯化得到化合物S1(38.03mg，收率：17.1％)。LCMS：MS m/z(ESI):434.1[M+H]+.1H NMR(400MHz,DMSO-d6)δ12.23(s,1H),9.07(s,1H),8.66(d,J＝4.8Hz,1H),8.32(d,J＝7.9Hz,1H),7.94(s,1H),7.54(dd,J＝7.9,4.8Hz,1H),4.70(s,2H),3.62–3.56(m,1H),2.66(s,3H),2.17(s,3H),1.29(d,J＝6.8Hz,3H),1.17–1.11(m,1H),0.62–0.55(m,1H),0.44–0.35(m,2H),0.24(dd,J＝8.7,3.5Hz,1H).Intermediate 1 (200 mg, 0.51 mmol) and 3-pyridine boronic acid (76 mg, 0.61 mmol) were dissolved in 1,4-dioxane (20 mL) and water (4 mL), and then [1,1'-bis (Diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex (42 mg, 0.05 mmol) and potassium carbonate (142 mg, 1.02 mmol), the reaction solution was stirred at 100°C for 2 hours under nitrogen protection. The reaction solution was cooled to room temperature and filtered. The filtrate was extracted three times with ethyl acetate and water. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and purified to obtain compound S1 (38.03 mg, Yield: 17.1%). LCMS: MS m/z(ESI):434.1[M+H] + . 1 H NMR(400MHz, DMSO-d 6 )δ12.23(s,1H),9.07(s,1H),8.66(d,J ＝4.8Hz,1H),8.32(d,J＝7.9Hz,1H),7.94(s,1H),7.54(dd,J＝7.9,4.8Hz,1H),4.70(s,2H),3.62–3.56 (m,1H),2.66(s,3H),2.17(s,3H),1.29(d,J＝6.8Hz,3H),1.17–1.11(m,1H),0.62–0.55(m,1H), 0.44–0.35(m,2H),0.24(dd,J＝8.7,3.5Hz,1H).
实施例2化合物S2的制备
Example 2 Preparation of Compound S2
将中间体1(120mg，0.30mmol)和5-硼酸嘧啶(77mg，0.36mmol)溶于1,4-二氧六环(10mL)和水(2mL)中，然后加入[1,1'-双(二苯基膦)二茂铁]二氯化钯二氯甲烷络合物(26mg，0.03mmol)和碳酸钾(85mg，0.61mmol)，反应液在氮气保护下100℃搅拌2小时。反应液冷却至室温，过滤，滤液用乙酸乙酯和水萃取三次，合并有机相，用饱和食盐水洗涤，用无水硫酸钠干燥，过滤，滤液浓缩后经制备纯化得到化合物S2(31.48mg，收率：23.4％)。LCMS：MS m/z(ESI):435.2[M+H]+.1H NMR(400MHz,DMSO-d6)δ12.25(s,1H),9.28(d,J＝1.8Hz,3H),7.99(s,1H),4.72(s,2H),3.63–3.57(m,1H),2.66(s,3H),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.15(s,1H),0.58(s,1H),0.45–0.36(m,2H),0.25(d,J＝5.4Hz,1H).Intermediate 1 (120 mg, 0.30 mmol) and 5-boronic acid pyrimidine (77 mg, 0.36 mmol) were dissolved in 1,4-dioxane (10 mL) and water (2 mL), and then [1,1'-bis (Diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex (26 mg, 0.03 mmol) and potassium carbonate (85 mg, 0.61 mmol), the reaction solution was stirred at 100°C for 2 hours under nitrogen protection. The reaction solution was cooled to room temperature and filtered. The filtrate was extracted three times with ethyl acetate and water. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated and purified to obtain compound S2 (31.48 mg, Yield: 23.4%). LCMS: MS m/z (ESI): 435.2[M+H] + . 1 H NMR (400MHz, DMSO-d 6 ) δ 12.25 (s, 1H), 9.28 (d, J = 1.8Hz, 3H), 7.99(s,1H),4.72(s,2H),3.63–3.57(m,1H),2.66(s,3H),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.15 (s,1H),0.58(s,1H),0.45–0.36(m,2H),0.25(d,J＝5.4Hz,1H).
实施例3化合物S3的制备
Example 3 Preparation of Compound S3
将中间体1(100mg，0.26mmol,1eq)、中间体2(199mg，0.77mmol,3eq)、K2CO3(71mg，0.51mmol,2eq)溶于1,4-二氧六环(20mL)和水(4mL)中，氩气置换，加入Pd(dppf)Cl2(19mg，0.03mmol,0.1eq)，氩气置换，氩气保护状态下，100℃搅拌反应2小时。LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(30mL×3) 萃取，合并有机相用饱和氯化钠水洗涤，无水硫酸钠干燥。旋干送制备纯化，得化合物S3(18.11mg，0.037mmol，收率:14％)。LCMS：MS m/z(ESI):488.1[M+H]+.1H NMR(400MHz,DMSO-d6)δ12.18(s,1H),10.48(s,1H),7.85(s,1H),7.51(d,J＝7.7Hz,1H),7.41(s,1H),7.30(d,J＝7.7Hz,1H),4.65(s,2H),3.59(dd,J＝15.7,6.9Hz,1H),2.64(s,3H),2.16(s,3H),1.26(t,J＝9.3Hz,3H),1.13(dd,J＝8.3,4.4Hz,1H),0.57(dd,J＝8.2,4.5Hz,1H),0.47–0.31(m,2H),0.28–0.16(m,1H).Dissolve Intermediate 1 (100mg, 0.26mmol, 1eq), Intermediate 2 (199mg, 0.77mmol, 3eq), K 2 CO 3 (71mg, 0.51mmol, 2eq) in 1,4-dioxane (20mL) and water (4 mL), replace with argon, add Pd(dppf)Cl 2 (19 mg, 0.03 mmol, 0.1eq), replace with argon, stir and react at 100°C for 2 hours under argon protection. LCMS detection showed that the reaction was successful. After the reaction solution is cooled, add water (20mL) to dilute, dichloromethane (30mL×3) Extract, combine the organic phases, wash with saturated sodium chloride water, and dry over anhydrous sodium sulfate. Spin dry and send for preparation and purification to obtain compound S3 (18.11 mg, 0.037 mmol, yield: 14%). LCMS: MS m/z(ESI):488.1[M+H] + . 1 H NMR(400MHz, DMSO-d6)δ12.18(s,1H),10.48(s,1H),7.85(s,1H) ,7.51(d,J＝7.7Hz,1H),7.41(s,1H),7.30(d,J＝7.7Hz,1H),4.65(s,2H),3.59(dd,J＝15.7,6.9Hz, 1H),2.64(s,3H),2.16(s,3H),1.26(t,J＝9.3Hz,3H),1.13(dd,J＝8.3,4.4Hz,1H),0.57(dd,J＝8.2 ,4.5Hz,1H),0.47–0.31(m,2H),0.28–0.16(m,1H).
实施例4化合物S4的制备
Example 4 Preparation of Compound S4
将中间体1(300mg，0.77mmol,1eq)、中间体3(597mg，2.30mmol,3eq)、K2CO3(212mg，1.53mmol,2eq)溶于1,4-二氧六环(20mL)和水(4mL)中，氩气置换，加入Pd(dppf)Cl2(56mg，0.077mmol,0.1eq)，氩气置换，氩气保护状态下，100℃搅拌反应2小时。送LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(20mL×3)萃取。合并有机相用饱和氯化钠溶液(20mL)洗涤一次，无水硫酸钠干燥。旋干送制备纯化，得化合物S4(3.85mg，0.0079mmol，收率:1.03％)。LCMS：MS m/z(ESI):488.2[M+H]+.1H NMR(400MHz,DMSO-d6))δ12.22(s,1H),8.63(s,1H),8.21(s,1H),8.14(dd,J＝7.9,1.5Hz,1H),7.91(s,1H),7.68(d,J＝7.9Hz,1H),4.69(s,2H),4.46(s,2H),3.64–3.49(m,1H),2.66(s,3H),2.16(s,3H),1.29(d,J＝6.8Hz,3H),1.18–1.05(m,1H),0.59(d,J＝8.5Hz,1H),0.47–0.33(m,2H),0.24(d,J＝5.1Hz,1H).Dissolve Intermediate 1 (300mg, 0.77mmol, 1eq), Intermediate 3 (597mg, 2.30mmol, 3eq), K 2 CO 3 (212mg, 1.53mmol, 2eq) in 1,4-dioxane (20mL) and water (4 mL), replaced with argon, added Pd(dppf)Cl 2 (56 mg, 0.077 mmol, 0.1eq), replaced with argon, and stirred at 100°C for 2 hours under argon protection. Sent LCMS for testing and the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL) and extracted with dichloromethane (20 mL × 3). The combined organic phases were washed once with saturated sodium chloride solution (20 mL) and dried over anhydrous sodium sulfate. Spin dry and send for preparation and purification to obtain compound S4 (3.85 mg, 0.0079 mmol, yield: 1.03%). LCMS: MS m/z(ESI):488.2[M+H] + . 1 H NMR(400MHz, DMSO-d6))δ12.22(s,1H),8.63(s,1H),8.21(s,1H ),8.14(dd,J＝7.9,1.5Hz,1H),7.91(s,1H),7.68(d,J＝7.9Hz,1H),4.69(s,2H),4.46(s,2H),3.64 –3.49(m,1H),2.66(s,3H),2.16(s,3H),1.29(d,J＝6.8Hz,3H),1.18–1.05(m,1H),0.59(d,J＝8.5 Hz,1H),0.47–0.33(m,2H),0.24(d,J＝5.1Hz,1H).
实施例5化合物S5的制备
Example 5 Preparation of Compound S5
将中间体1(1.4g,3.58mmol，1eq)、中间体4(1.261g,8.95mmol，2.5eq)、K2CO3(990mg,7.16mmol，2eq)、溶于1,4-二氧六环(30mL)和水(6mL)中，氩气置换，加入Pd(dppf)Cl2(131mg,0.179mmol，0.05eq)氩气置换，氩气保护状态下，100℃搅拌2小时。LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(20mL×3)萃取。合并有机相用饱和氯化钠溶液(20mL)洗涤一次，无水硫酸钠干燥。旋干，送制备纯化，得化合物S5(1175mg，2.60mmol，收率:73％)。LCMS：MS m/z(ESI):452.2[M+H]+.1H NMR(400MHz,DMSO-d6))δ12.23(s,1H),8.37(d,J＝5.1Hz,1H),7.96(s,1H),7.89(d,J＝4.6Hz,1H),7.74(s,1H),4.69(s,2H),3.65–3.54(m,1H),2.63(s,3H),2.16(s,3H),1.28(d,J＝6.7Hz,3H),1.17–1.08(m,1H),0.58(d,J＝3.9Hz,1H),0.40(dd,J＝14.4,7.5Hz,2H),0.25(d,J＝5.0Hz,1H).Dissolve Intermediate 1 (1.4g, 3.58mmol, 1eq), Intermediate 4 (1.261g, 8.95mmol, 2.5eq), K 2 CO 3 (990mg, 7.16mmol, 2eq) in 1,4-dioxane Ring (30 mL) and water (6 mL) were replaced with argon, and Pd(dppf)Cl 2 (131 mg, 0.179 mmol, 0.05 eq) was added to replace with argon. Under argon protection, stir at 100°C for 2 hours. LCMS detection showed that the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL) and extracted with dichloromethane (20 mL × 3). The combined organic phases were washed once with saturated sodium chloride solution (20 mL) and dried over anhydrous sodium sulfate. Spin to dryness and send to preparation and purification to obtain compound S5 (1175 mg, 2.60 mmol, yield: 73%). LCMS: MS m/z (ESI): 452.2[M+H] + . 1 H NMR (400MHz, DMSO-d6)) δ12.23 (s, 1H), 8.37 (d, J = 5.1Hz, 1H), 7.96(s,1H),7.89(d,J＝4.6Hz,1H),7.74(s,1H),4.69(s,2H),3.65–3.54(m,1H),2.63(s,3H),2.16 (s,3H),1.28(d,J＝6.7Hz,3H),1.17–1.08(m,1H),0.58(d,J＝3.9Hz,1H),0.40(dd,J＝14.4,7.5Hz, 2H),0.25(d,J=5.0Hz,1H).
实施例6化合物S6的制备
Example 6 Preparation of Compound S6
步骤1：将中间体1(400mg，1.02mmol,1eq)、中间体5(900mg，3.07mmol,3eq)、K2CO3(283mg，2.05mmol,2eq)溶于1,4-二氧六环(20mL)和水(4mL)中，氩气置换，加入Pd(dppf)Cl2(75mg，0.10mmol,0.1eq)，氩气置换，氩气保护状态下，100℃搅拌反应2小时。LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(20mL×3)萃取。合并有机相用饱和氯化钠溶液(20mL)洗涤一次，无水硫酸钠干燥。旋干得1.2g粗品化合物S6-1(1.2g，纯度:73％)。Step 1: Dissolve intermediate 1 (400mg, 1.02mmol, 1eq), intermediate 5 (900mg, 3.07mmol, 3eq), K 2 CO 3 (283mg, 2.05mmol, 2eq) in 1,4-dioxane (20 mL) and water (4 mL), replace with argon, add Pd(dppf)Cl 2 (75 mg, 0.10 mmol, 0.1 eq), replace with argon, stir and react at 100°C for 2 hours under argon protection. LCMS detection showed that the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL) and extracted with dichloromethane (20 mL × 3). The combined organic phases were washed once with saturated sodium chloride solution (20 mL) and dried over anhydrous sodium sulfate. Spin to dryness to obtain 1.2g of crude compound S6-1 (1.2g, purity: 73%).
步骤2：将化合物S6-1(1.2g，1.45mmol，1eq)溶于二氯甲烷(50mL)中，加入三氟乙酸(9.4g,82.7mmol，57eq)，氩气保护，室温反应12小时。加K2CO3(1.0g，7.25mmol，5eq)、甲醇(50mL)，水(10mL)，搅拌12小时。二氯甲烷(80mL×3)萃取。合并有机相用饱和氯化钠溶液(20mL)洗涤一次，无水硫酸钠干燥。旋干，送制备纯化，得化合物S6(16.31mg，0.034mmol,收率:2.37％)。LCMS：MS m/z(ESI):474.2[M+H]+.1H NMR(400MHz,DMSO-d6))δ13.78(s,1H),12.21(s,1H),8.33(d,J＝8.2Hz,1H),8.23(s,1H),8.00(s,1H),7.51(d,J＝8.2Hz,1H),4.71(s,2H),3.56–3.47(m,1H),2.65(s,3H),2.15(s,3H),1.27(d,J＝6.8Hz,3H),1.13(d,J＝8.6Hz,1H),0.62–0.14(m,4H).Step 2: Dissolve compound S6-1 (1.2g, 1.45mmol, 1eq) in dichloromethane (50mL), add trifluoroacetic acid (9.4g, 82.7mmol, 57eq), protect with argon, and react at room temperature for 12 hours. Add K 2 CO 3 (1.0g, 7.25mmol, 5eq), methanol (50mL), water (10mL), and stir for 12 hours. Extract with dichloromethane (80mL×3). The combined organic phases were washed once with saturated sodium chloride solution (20 mL) and dried over anhydrous sodium sulfate. Spin to dryness and send to preparation and purification to obtain compound S6 (16.31 mg, 0.034 mmol, yield: 2.37%). LCMS: MS m/z(ESI):474.2[M+H] + . 1 H NMR(400MHz, DMSO-d6))δ13.78(s,1H),12.21(s,1H),8.33(d,J ＝8.2Hz,1H),8.23(s,1H),8.00(s,1H),7.51(d,J＝8.2Hz,1H),4.71(s,2H),3.56–3.47(m,1H),2.65 (s,3H),2.15(s,3H),1.27(d,J＝6.8Hz,3H),1.13(d,J＝8.6Hz,1H),0.62–0.14(m,4H).
实施例7化合物S7的制备
Example 7 Preparation of Compound S7
步骤1：将中间体1(300mg，0.77mmol,1eq)、中间体6(673mg，2.30mmol,3eq)、K2CO3(212mg，1.53mmol,2eq)溶于1,4-二氧六环(20mL)中，再加屈氏蒸馏水(4mL)，氩气置换，加入Pd(dppf)Cl2(56mg，0.077mmol,0.1eq)，氩气置换，氩气保护状态下，100℃搅拌反应2小时。送LCMS检测，反应液成功。反应液冷却后，二氯甲烷与水萃取，饱和氯化钠水洗，无水硫酸钠干燥。旋干纯化得化合物S7-1粗品(453mg，纯度:76.59％，收率:74.99％)。Step 1: Dissolve intermediate 1 (300mg, 0.77mmol, 1eq), intermediate 6 (673mg, 2.30mmol, 3eq), K 2 CO 3 (212mg, 1.53mmol, 2eq) in 1,4-dioxane (20 mL), add Troubles distilled water (4 mL), replace with argon, add Pd(dppf)Cl 2 (56 mg, 0.077 mmol, 0.1 eq), replace with argon, stir the reaction at 100°C under argon protection 2 Hour. Sent LCMS for testing and the reaction solution was successful. After the reaction solution was cooled, it was extracted with dichloromethane and water, washed with saturated sodium chloride, and dried over anhydrous sodium sulfate. Spin-dry and purify to obtain crude compound S7-1 (453 mg, purity: 76.59%, yield: 74.99%).
步骤2：将化合物S7-1(453mg，纯度:76.59％，0.58mmol，1eq)溶于二氯甲烷(50mL)中，加入三氟乙酸(3.7g,32.81mmol，57eq),氩气保护，室温反应10小时。反应结束，调pH为中性，旋干，加K2CO3(398mg，2.88mmol，5eq)、甲醇50mL，屈氏蒸馏水10mL，反应2小时。反应结束，二氯甲烷与水萃取，氯化钠水洗，无水硫酸钠干燥，旋干，送制备纯化，得化合物S7(26.90mg，0.057mmol,收率:32.76％)。LCMS：MS m/z(ESI):473.2[M+H]+.1H NMR(400MHz,DMSO-d6))δ12.20(s,1H),11.84(s,1H),8.04(d,J＝8.1Hz,1H),7.93(s,1H),7.57(s,1H),7.42(d,J＝8.1Hz,1H),6.53(s,1H),4.69(s,2H),3.52(s,1H),2.65(s,3H),2.15(s,3H),1.27(d,J＝6.8Hz,3H),1.12(s,1H),0.60–0.19(m,4H).Step 2: Dissolve compound S7-1 (453mg, purity: 76.59%, 0.58mmol, 1eq) in dichloromethane (50mL), add trifluoroacetic acid (3.7g, 32.81mmol, 57eq), protect with argon, room temperature Reaction takes 10 hours. At the end of the reaction, adjust the pH to neutral, spin to dryness, add K 2 CO 3 (398 mg, 2.88 mmol, 5 eq), 50 mL of methanol, and 10 mL of Treconite distilled water, and react for 2 hours. At the end of the reaction, extract with dichloromethane and water, wash with sodium chloride, dry with anhydrous sodium sulfate, spin dry, and send to preparation and purification to obtain compound S7 (26.90 mg, 0.057 mmol, yield: 32.76%). LCMS: MS m/z(ESI):473.2[M+H] + . 1 H NMR(400MHz, DMSO-d6))δ12.20(s,1H),11.84(s,1H),8.04(d,J ＝8.1Hz,1H),7.93(s,1H),7.57(s,1H),7.42(d,J＝8.1Hz,1H),6.53(s,1H),4.69(s,2H),3.52(s ,1H),2.65(s,3H),2.15(s,3H),1.27(d,J＝6.8Hz,3H),1.12(s,1H),0.60–0.19(m,4H).
实施例8化合物S8的制备
Example 8 Preparation of Compound S8
步骤1：将中间体1(300mg，0.77mmol,1eq)、中间体7(432mg，1.15mmol,1.5eq)、K2CO3(212mg，1.53mmol,2eq)溶于1,4-二氧六环(20mL)中，再加屈氏蒸馏水(4mL)，氩气置换，加入Pd(dppf)Cl2(56mg，0.077mmol,0.1eq)，氩气置换，氩气保护状态下，100℃搅拌反应2小时。送LCMS检测，反应液成功。反应液冷却后，二氯甲烷与水萃取，饱和氯化钠水洗，无水硫酸钠干燥。旋干得化合物S8-1粗品(500mg，纯度:74.49％，收率:80.37％)。Step 1: Dissolve intermediate 1 (300mg, 0.77mmol, 1eq), intermediate 7 (432mg, 1.15mmol, 1.5eq), K 2 CO 3 (212mg, 1.53mmol, 2eq) in 1,4-dioxane ring (20 mL), add Tribune distilled water (4 mL), replace with argon, add Pd(dppf)Cl 2 (56 mg, 0.077 mmol, 0.1 eq), replace with argon, and stir the reaction at 100°C under argon protection. 2 hours. Sent LCMS for testing and the reaction solution was successful. After the reaction solution was cooled, it was extracted with dichloromethane and water, washed with saturated sodium chloride, and dried over anhydrous sodium sulfate. Spin to dryness to obtain crude compound S8-1 (500 mg, purity: 74.49%, yield: 80.37%).
步骤2：将化合物S8-1(500mg，纯度:74.49％，0.617mmol,1eq)溶于二氯甲烷(50mL)中，加入三氟乙酸(4.0g,35.16mmol，57eq)，氩气保护，室温反应10小时。反应结束，调pH为中性，旋干，加K2CO3(426mg，3.08mmol，5eq)、甲醇50mL，屈氏蒸馏水10mL，反应2小时。反应结束，二氯甲烷与水萃取，氯化钠水洗，无水硫酸钠干燥，旋干，送制备纯化，得化合物S8(118.15mg，0.249mmol,收率:40.45％)。LCMS：MS m/z(ESI):474.2[M+H]+.1H NMR(400MHz,DMSO-d6))δ13.61(s,1H),12.27(s,1H),9.03(d,J＝1.7Hz,1H),8.64(s,1H),8.38(s,1H),7.96(s,1H),4.72(s,2H),3.61(dd,J＝9.0,6.8Hz,1H),2.67(s,3H),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.16(dd,J＝8.4,4.3Hz,1H),0.59(dt,J＝8.5,5.1Hz,1H),0.46–0.36(m,2H),0.25(dt,J＝8.5,4.2Hz,1H).Step 2: Dissolve compound S8-1 (500mg, purity: 74.49%, 0.617mmol, 1eq) in dichloromethane (50mL), add trifluoroacetic acid (4.0g, 35.16mmol, 57eq), protect with argon, room temperature Reaction takes 10 hours. After the reaction is completed, adjust the pH to neutral, spin to dryness, add K 2 CO 3 (426 mg, 3.08 mmol, 5 eq), 50 mL of methanol, and 10 mL of Trecroft distilled water, and react for 2 hours. At the end of the reaction, extract with dichloromethane and water, wash with sodium chloride, dry with anhydrous sodium sulfate, spin dry, and send to preparation and purification to obtain compound S8 (118.15 mg, 0.249 mmol, yield: 40.45%). LCMS: MS m/z(ESI):474.2[M+H] + . 1 H NMR(400MHz, DMSO-d6))δ13.61(s,1H),12.27(s,1H),9.03(d,J ＝1.7Hz,1H),8.64(s,1H),8.38(s,1H),7.96(s,1H),4.72(s,2H),3.61(dd,J＝9.0,6.8Hz,1H),2.67 (s,3H),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.16(dd,J＝8.4,4.3Hz,1H),0.59(dt,J＝8.5,5.1Hz, 1H),0.46–0.36(m,2H),0.25(dt,J＝8.5,4.2Hz,1H).
实施例9化合物S9的制备
Example 9 Preparation of Compound S9
步骤1：将中间体1(500mg,1.28mmol,1eq)溶于1,4-二氧六环(25mL)，水(5mL)中，加入中间体8(1.5g,5.1mmol,4eq)，Pd(dppf)Cl2(94.2mg,0.13mmol,0.1eq),碳酸钠(1.25g,0.77mmol,3eq)，氩气保护,100℃搅拌反应2小时。LCMS监测有产物生成。反应液冷却后加水(30mL)稀释，二氯甲烷萃取三次(30mL×3)分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，得到化合物S9-1粗品(1g)。LCMS：MS m/z(ESI):605.3[M+H]+。Step 1: Dissolve intermediate 1 (500mg, 1.28mmol, 1eq) in 1,4-dioxane (25mL), water (5mL), add intermediate 8 (1.5g, 5.1mmol, 4eq), Pd (dppf)Cl 2 (94.2mg, 0.13mmol, 0.1eq), sodium carbonate (1.25g, 0.77mmol, 3eq), protected by argon, stirred and reacted at 100°C for 2 hours. LCMS monitored product formation. After cooling, the reaction solution was diluted with water (30 mL), extracted with dichloromethane three times (30 mL × 3), and separated. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude compound S9-1 (1 g). LCMS: MS m/z(ESI):605.3[M+H] + .
步骤2：将化合物S9-1(1g,粗品)溶于DCM(35mL)中,加入三氟乙酸(5mL)，室温反应10小时。反应液浓缩干，产物粗品加25mL甲醇及5mL水溶解，加入碳酸钾(1.37g,10mmol,5eq),室温搅拌2小时，过滤，用少量甲醇冲洗滤饼，溶剂减压浓缩。粗品加少量二甲基甲酰胺溶解，制备分离(0.1％NH3.H2O)纯化，冻干得到化合物S9(127.21mg，收率：16.2％)。LCMS：MS m/z(ESI):475.3[M+H]+.1H NMR(400MHz,DMSO-d6)δ14.20(s,1H),12.22(s,1H),8.92(s,1H),8.52(s,1H),8.02(s,1H),4.71(s,2H),3.58–3.42(m,1H),2.62(s,3H),2.12(s,3H),1.24(d,J＝6.8Hz,3H),1.09(dq,J＝8.3,4.2,3.6Hz,1H),0.53(dq,J＝8.4,4.3Hz,1H),0.45–0.27(m,2H),0.18(dt,J＝9.5,4.7Hz,1H).Step 2: Dissolve compound S9-1 (1g, crude product) in DCM (35mL), add trifluoroacetic acid (5mL), and react at room temperature for 10 hours. The reaction solution was concentrated to dryness, the crude product was dissolved in 25 mL of methanol and 5 mL of water, potassium carbonate (1.37 g, 10 mmol, 5 eq) was added, stirred at room temperature for 2 hours, filtered, the filter cake was washed with a small amount of methanol, and the solvent was concentrated under reduced pressure. The crude product was dissolved by adding a small amount of dimethylformamide, purified by preparative separation (0.1% NH 3 .H 2 O), and lyophilized to obtain compound S9 (127.21 mg, yield: 16.2%). LCMS: MS m/z(ESI):475.3[M+H] + . 1 H NMR(400MHz, DMSO-d 6 )δ14.20(s,1H),12.22(s,1H),8.92(s,1H ),8.52(s,1H),8.02(s,1H),4.71(s,2H),3.58–3.42(m,1H),2.62(s,3H),2.12(s,3H),1.24(d, J＝6.8Hz,3H),1.09(dq,J＝8.3,4.2,3.6Hz,1H),0.53(dq,J＝8.4,4.3Hz,1H),0.45–0.27(m,2H),0.18(dt ,J=9.5,4.7Hz,1H).
实施例10化合物S10的制备
Example 10 Preparation of Compound S10
将中间体1(250mg,0.64mmol，1eq)、中间体9(263mg,1.92mmol，3eq)、K2CO3(177mg,1.28mmol，2eq)、溶于1,4-二氧六环(20mL)，加水(4mL)，氩气置换，加入Pd(dppf)Cl2(56mg,0.077mmol，0.1eq)氩气置换，氩气保护状态下，100℃搅拌2小时。送LCMS检测反应成功。反应液冷却后，将反应液转移至100mL水中，用二氯甲烷(100mL)萃取，萃取3次。将收集的有机相用无水硫酸钠干燥，旋干。送制备纯化得化合物S10(65.49mg，0.146mmol，收率:22.88％)。LCMS：MS m/z(ESI):448.2[M+H]+.1H NMR(400MHz,DMSO-d6))δ12.24(s,1H),8.94(d,J＝2.1Hz,1H),8.21(dd,J＝8.0,2.3Hz,1H),7.90(s,1H),7.38(d,J＝8.1Hz,1H),4.68(s,2H),3.63–3.54(m,1H),2.65(s,3H),2.55(s,3H),2.17(s,3H),1.29(d,J＝6.8Hz,3H),1.17–1.09(m,1H),0.61–0.22(m,4H).Dissolve Intermediate 1 (250mg, 0.64mmol, 1eq), Intermediate 9 (263mg, 1.92mmol, 3eq), K 2 CO 3 (177mg, 1.28mmol, 2eq) in 1,4-dioxane (20mL ), add water (4mL), replace with argon, add Pd(dppf)Cl 2 (56mg, 0.077mmol, 0.1eq) and replace with argon, stir at 100°C for 2 hours under argon protection. Send LCMS to detect the success of the reaction. After the reaction liquid was cooled, the reaction liquid was transferred to 100 mL of water, and extracted with dichloromethane (100 mL) three times. The collected organic phase was dried over anhydrous sodium sulfate and spun to dryness. Compound S10 (65.49 mg, 0.146 mmol, yield: 22.88%) was prepared and purified. LCMS: MS m/z (ESI): 448.2[M+H] + . 1 H NMR (400MHz, DMSO-d6)) δ12.24 (s, 1H), 8.94 (d, J = 2.1Hz, 1H), 8.21(dd,J＝8.0,2.3Hz,1H),7.90(s,1H),7.38(d,J＝8.1Hz,1H),4.68(s,2H),3.63–3.54(m,1H),2.65 (s,3H),2.55(s,3H),2.17(s,3H),1.29(d,J＝6.8Hz,3H),1.17–1.09(m,1H),0.61–0.22(m,4H).
实施例11化合物S11的制备
Example 11 Preparation of Compound S11
将中间体1(350mg,0.89mmol)溶于1,4-二氧六环(15mL)，水(3mL)中，加入中间体10(370mg,2.68mmol,3eq)，Pd(dppf)Cl2(64.5mg,0.089mmol,0.1eq)，碳酸钠(284.1mg,2.68mmol,3eq)，氩气保护,100℃搅拌反应3小时。LCMS监测有产物生成。反应液冷却后加水(30mL)稀释，二氯甲烷萃取三次(30mL×3)，分液，有机相用无水硫酸钠干燥，过滤，减压浓缩，粗品加少量二甲基甲酰胺溶解，制备(制备柱：Waters-SunFire C18，洗脱相：0.04vol％FA水溶液/乙腈，洗脱梯度：开始54vol％乙腈-结束62vol％乙腈，流速：30mL/min)分离纯化得到化合物S11(112.18mg，纯度：99.3％)。LCMS：MS m/z(ESI):448.2[M+H]+.1HNMR(400MHz,DMSO-d6)δ12.24(s,1H),8.57(d,J＝5.1Hz,1H),7.96(s,1H),7.74(s,1H),7.66(d,J＝4.9Hz,1H),4.69(s,2H),3.66–3.51(m,1H),2.65(s,3H),2.56(s,3H),2.16(s,3H),1.28(d,J＝6.8Hz,3H),1.16–1.03(m,1H),0.57(t,J＝4.1Hz,1H),0.49–0.32(m,2H),0.24(d,J＝5.1Hz,1H).Dissolve intermediate 1 (350mg, 0.89mmol) in 1,4-dioxane (15mL), water (3mL), add intermediate 10 (370mg, 2.68mmol, 3eq), Pd(dppf)Cl 2 ( 64.5 mg, 0.089 mmol, 0.1 eq), sodium carbonate (284.1 mg, 2.68 mmol, 3 eq), protected by argon, and stirred at 100°C for 3 hours. LCMS monitored product formation. After cooling, the reaction solution was diluted with water (30 mL), extracted with methylene chloride three times (30 mL (Preparative column: Waters-SunFire C18, elution phase: 0.04vol% FA aqueous solution/acetonitrile, elution gradient: starting at 54vol% acetonitrile - ending at 62vol% acetonitrile, flow rate: 30mL/min) Compound S11 (112.18mg, Purity: 99.3%). LCMS: MS m/z (ESI): 448.2[M+H] + . 1 HNMR (400MHz, DMSO-d 6 ) δ 12.24 (s, 1H), 8.57 (d, J = 5.1Hz, 1H), 7.96 (s,1H),7.74(s,1H),7.66(d,J＝4.9Hz,1H),4.69(s,2H),3.66–3.51(m,1H),2.65(s,3H),2.56( s,3H),2.16(s,3H),1.28(d,J＝6.8Hz,3H),1.16–1.03(m,1H),0.57(t,J＝4.1Hz,1H),0.49–0.32(m ,2H),0.24(d,J＝5.1Hz,1H).
实施例12化合物S12的制备
Example 12 Preparation of Compound S12
将中间体1(250mg,0.640mmol，1eq)、包含中间体11a和11b的混合物(314mg,1.279 mmol，2eq)、K2CO3(177mg,1.28mmol，2eq)、溶于1,4-二氧六环(20mL)，加水(4mL)，氩气置换。再加入Pd(dppf)Cl2(56mg,0.077mmol，0.1eq)，氩气置换，氩气保护状态下，100℃搅拌2小时。送LCMS检测反应成功。反应液冷却后，将反应液转移至100mL水中，用二氯甲烷(100mL)萃取，萃取3次。将收集的有机相用无水硫酸钠干燥，旋干。送制备纯化得化合物S12(33.38mg，0.070mmol，收率:11.02％)。LCMS：MS m/z(ESI):474.2[M+H]+.1H NMR(400MHz,DMSO-d6)δ12.23(s,1H),8.90(d,J＝1.7Hz,1H),8.18(dd,J＝8.1,2.2Hz,1H),7.89(s,1H),7.41(d,J＝8.1Hz,1H),4.67(s,2H),3.63–3.54(m,1H),2.65(s,3H),2.20–2.14(m,4H),1.29(d,J＝6.8Hz,3H),1.14(s,1H),1.01(d,J＝6.4Hz,4H),0.61–0.23(m,4H).Intermediate 1 (250 mg, 0.640 mmol, 1 eq), a mixture containing intermediates 11a and 11b (314 mg, 1.279 mmol, 2eq), K 2 CO 3 (177mg, 1.28mmol, 2eq), dissolved in 1,4-dioxane (20mL), added water (4mL), and replaced with argon. Then add Pd(dppf)Cl 2 (56 mg, 0.077 mmol, 0.1 eq), replace with argon, and stir at 100°C for 2 hours under argon protection. Send LCMS to detect the success of the reaction. After the reaction liquid was cooled, the reaction liquid was transferred to 100 mL of water, and extracted with dichloromethane (100 mL) three times. The collected organic phase was dried over anhydrous sodium sulfate and spun to dryness. Compound S12 (33.38 mg, 0.070 mmol, yield: 11.02%) was prepared and purified. LCMS: MS m/z (ESI): 474.2[M+H] + . 1 H NMR (400MHz, DMSO-d 6 ) δ 12.23 (s, 1H), 8.90 (d, J = 1.7Hz, 1H), 8.18(dd,J＝8.1,2.2Hz,1H),7.89(s,1H),7.41(d,J＝8.1Hz,1H),4.67(s,2H),3.63–3.54(m,1H),2.65 (s,3H),2.20–2.14(m,4H),1.29(d,J＝6.8Hz,3H),1.14(s,1H),1.01(d,J＝6.4Hz,4H),0.61–0.23( m,4H).
实施例13化合物S13的制备
Example 13 Preparation of Compound S13
将中间体1(300mg,0.767mmol，1eq)、中间体12(176mg,0.920mmol，1.2eq)、K2CO3(212mg,1.534mmol，2eq)、溶于1,4-二氧六环(20mL)，加水(4mL)，氩气置换。再加入Pd(dppf)Cl2(56mg,0.077mmol，0.1eq)氩气置换，氩气保护状态下，100℃搅拌2小时。送LCMS检测反应成功。反应液冷却后，将反应液转移至100mL水中，用二氯甲烷(100mL)萃取，萃取3次。将收集的有机相用无水硫酸钠干燥，旋干。送制备纯化得化合物S13(60.10mg，0.120mmol，收率:15.61％)。LCMS：MS m/z(ESI):502.1[M+H]+.1H NMR(400MHz,DMSO-d6))δ12.24(s,1H),9.21(d,J＝1.6Hz,1H),8.57(dd,J＝8.1,1.7Hz,1H),8.07(d,J＝8.1Hz,1H),8.00(s,1H),4.72(s,2H),3.59(dq,J＝13.5,6.8Hz,1H),2.66(s,3H),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.14(dt,J＝13.2,4.8Hz,1H),0.63–0.55(m,1H),0.46–0.35(m,2H),0.28–0.22(m,1H).Intermediate 1 (300mg, 0.767mmol, 1eq), intermediate 12 (176mg, 0.920mmol, 1.2eq), K 2 CO 3 (212mg, 1.534mmol, 2eq) were dissolved in 1,4-dioxane ( 20 mL), add water (4 mL), and replace with argon. Then add Pd(dppf)Cl 2 (56 mg, 0.077 mmol, 0.1 eq) for argon replacement, and stir at 100°C for 2 hours under argon protection. Send LCMS to detect the success of the reaction. After the reaction liquid was cooled, the reaction liquid was transferred to 100 mL of water, and extracted with dichloromethane (100 mL) three times. The collected organic phase was dried over anhydrous sodium sulfate and spun to dryness. Compound S13 (60.10 mg, 0.120 mmol, yield: 15.61%) was prepared and purified. LCMS: MS m/z(ESI):502.1[M+H] + . 1 H NMR(400MHz, DMSO-d6))δ12.24(s,1H),9.21(d,J＝1.6Hz,1H), 8.57(dd,J＝8.1,1.7Hz,1H),8.07(d,J＝8.1Hz,1H),8.00(s,1H),4.72(s,2H),3.59(dq,J＝13.5,6.8Hz ,1H),2.66(s,3H),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.14(dt,J＝13.2,4.8Hz,1H),0.63–0.55(m, 1H),0.46–0.35(m,2H),0.28–0.22(m,1H).
实施例14化合物S14的制备
Example 14 Preparation of Compound S14
将中间体1(100mg，0.25mmol),中间体13(103mg,0.37mmol)，[1,1'-双(二苯基膦)二茂铁]二氯化钯二氯甲烷络合物(18.3mg,0.025mmol)，碳酸铯(244mg,0.75mmol)溶于1,4-二氧六环(8mL)和水(2mL)中，氮气保护，油浴100℃反应2小时。LCMS检测，反应成功。反应液冷却后，加水(20mL)稀释，二氯甲烷(30mL×3)萃取，合并有机相用饱和氯化钠水洗涤，无水硫酸钠干燥。旋干送制备纯化，得化合物S14(2.4mg，纯度:100％)。LCMS：MS m/z(ESI):502.3[M+H]+.1H NMR(400MHz,DMSO-d6)δ12.21(s,1H),8.06(s,1H),7.87(s,1H),7.80(s,1H),7.77(d,J＝7.9Hz,1H),7.31(d,J＝7.9Hz,1H),4.66(s,2H),4.40(s,2H),3.62–3.54(m,1H),3.52(s,2H),2.64(s,3H),2.16(s,3H),1.28(d,J＝6.8Hz,3H),0.85(s, 1H),0.57(s,1H),0.48–0.31(m,2H),0.23(s,1H).Intermediate 1 (100 mg, 0.25 mmol), intermediate 13 (103 mg, 0.37 mmol), [1,1'-bis(diphenylphosphine)ferrocene]dichloride palladium dichloromethane complex (18.3 mg, 0.025 mmol), cesium carbonate (244 mg, 0.75 mmol) was dissolved in 1,4-dioxane (8 mL) and water (2 mL), protected by nitrogen, and reacted in an oil bath at 100°C for 2 hours. LCMS detection showed that the reaction was successful. After the reaction solution was cooled, it was diluted with water (20 mL) and extracted with dichloromethane (30 mL × 3). The combined organic phases were washed with saturated sodium chloride water and dried over anhydrous sodium sulfate. Spin dry and send for preparation and purification to obtain compound S14 (2.4 mg, purity: 100%). LCMS: MS m/z(ESI):502.3[M+H] + . 1 H NMR(400MHz, DMSO-d6)δ12.21(s,1H),8.06(s,1H),7.87(s,1H) ,7.80(s,1H),7.77(d,J＝7.9Hz,1H),7.31(d,J＝7.9Hz,1H),4.66(s,2H),4.40(s,2H),3.62–3.54( m,1H),3.52(s,2H),2.64(s,3H),2.16(s,3H),1.28(d,J＝6.8Hz,3H),0.85(s, 1H),0.57(s,1H),0.48–0.31(m,2H),0.23(s,1H).
实施例15化合物S15的制备
Example 15 Preparation of Compound S15
将中间体1(100mg，0.26mmol)，吡啶-4-基硼酸(62.9g,0.51mmol)，[1,1'-双(二苯基膦)二茂铁]二氯化钯二氯甲烷络合物(18.3mg,0.025mmol)，碳酸铯(250mg,0.77mmol)溶于1,4-二氧六环(6mL)和水(1.5mL)中，氩气保护，油浴100℃反应2小时。反应液冷却到室温，过滤，用乙酸乙酯(10mL×3)萃取，合并有机相，干燥，旋干，送制备纯化得化合物S15(62.7mg,纯度:99％)。LCMS：MS m/z(ESI):434.1[M+H]+.1H NMR(400MHz,DMSO-d6)δ11.89(s,1H),8.72(d,J＝5.9Hz,2H),7.97(s,1H),7.93–7.87(m,2H),4.70(s,2H),3.59(dq,J＝13.8,6.9Hz,1H),2.65(s,3H),2.17(s,3H),1.29(d,J＝6.8Hz,3H),1.18–1.06(m,1H),0.70–0.51(m,1H),0.50–0.31(m,2H),0.25(dd,J＝11.4,6.3Hz,1H).Intermediate 1 (100 mg, 0.26 mmol), pyridin-4-ylboronic acid (62.9 g, 0.51 mmol), [1,1'-bis(diphenylphosphine)ferrocene]dichloropalladium dichloromethane complex The compound (18.3 mg, 0.025 mmol), cesium carbonate (250 mg, 0.77 mmol) was dissolved in 1,4-dioxane (6 mL) and water (1.5 mL), protected by argon, and reacted in an oil bath at 100°C for 2 hours. . The reaction solution was cooled to room temperature, filtered, and extracted with ethyl acetate (10 mL×3). The organic phases were combined, dried, and spin-dried to obtain compound S15 (62.7 mg, purity: 99%). LCMS: MS m/z (ESI): 434.1[M+H] + . 1 H NMR (400MHz, DMSO-d6) δ11.89 (s, 1H), 8.72 (d, J = 5.9Hz, 2H), 7.97 (s,1H),7.93–7.87(m,2H),4.70(s,2H),3.59(dq,J＝13.8,6.9Hz,1H),2.65(s,3H),2.17(s,3H), 1.29(d,J＝6.8Hz,3H),1.18–1.06(m,1H),0.70–0.51(m,1H),0.50–0.31(m,2H),0.25(dd,J＝11.4,6.3Hz, 1H).
实施例16化合物S16的制备
Example 16 Preparation of Compound S16
将中间体1(0.3g，0.77mmol,1eq)置于50mL洁净单口烧瓶中，加1,4-二氧六环(12mL)、水(3mL)溶解，室温下加入中间体14(0.4g,1.46mmol,1.9eq)，碳酸钠(0.246g,2.31mmol,3eq)和Pd(dppf)Cl2(0.056g，0.077mmol,0.1eq)，氩气保护，100℃搅拌反应2小时。反应液冷却至室温，向反应液中加入20mL水，乙酸乙酯(30mL×3)萃取三次，有机相中加入氯化钠饱和溶液洗涤，分液，有机相用无水硫酸钠干燥，过滤浓缩，制备分离，得到化合物S16(86.73mg,收率：22.5％)。LCMS：MS m/z(ESI):502.1[M+H]+.1HNMR(400MHz,DMSO-d6)δ12.26(s,1H),8.95(t,J＝6.1Hz,1H),8.50(s,1H),8.27(d,J＝5.0Hz,1H),8.02(s,1H),4.73(s,2H),3.62(dq,J＝13.7,6.8Hz,1H),2.66(s,3H),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.20–1.07(m,1H),0.68–0.54(m,1H),0.48–0.35(m,2H),0.32–0.18(m,1H).Place Intermediate 1 (0.3g, 0.77mmol, 1eq) in a 50mL clean single-neck flask, add 1,4-dioxane (12mL) and water (3mL) to dissolve, and add Intermediate 14 (0.4g, 1eq) at room temperature. 1.46mmol, 1.9eq), sodium carbonate (0.246g, 2.31mmol, 3eq) and Pd(dppf)Cl 2 (0.056g, 0.077mmol, 0.1eq), under argon protection, stir and react at 100°C for 2 hours. The reaction solution was cooled to room temperature, 20 mL of water was added to the reaction solution, and extracted three times with ethyl acetate (30 mL , prepared and separated to obtain compound S16 (86.73 mg, yield: 22.5%). LCMS: MS m/z (ESI): 502.1[M+H] + . 1 HNMR (400MHz, DMSO-d6) δ12.26 (s, 1H), 8.95 (t, J = 6.1Hz, 1H), 8.50 ( s,1H),8.27(d,J＝5.0Hz,1H),8.02(s,1H),4.73(s,2H),3.62(dq,J＝13.7,6.8Hz,1H),2.66(s,3H ),2.17(s,3H),1.30(d,J＝6.8Hz,3H),1.20–1.07(m,1H),0.68–0.54(m,1H),0.48–0.35(m,2H),0.32– 0.18(m,1H).
实施例17化合物S17的制备
Example 17 Preparation of Compound S17
将中间体1(0.30g，0.77mmol,1eq)置于100mL洁净单口烧瓶中，加1,4-二氧六环(12mL)、水(3mL)溶解，室温下加入中间体15(0.33g,1.46mmol,1.9eq)，碳酸钠(0.25g,2.31mmol,3eq)和Pd(dppf)Cl2(0.056g，0.077mmol,0.1eq)，100℃搅拌反应2小时。反应液冷却至室温，向反应液中加入20mL水，加乙酸乙酯(30mL×3)萃取三次，有机相中加入氯化钠饱和溶液洗涤，分液，有机相用无水硫酸钠干燥，过滤浓缩，制备分离，得到化合物S17(9.2mg,纯度:99.7％)。LCMS：MS m/z(ESI):452.1[M+H]+.1H NMR(400MHz,DMSO-d6)δ11.98(s,1H),8.52(s,1H),8.27(t,J＝7.3Hz,1H),7.68(s,1H),7.09(d,J＝7.6Hz,1H),4.45(s,2H),3.39–3.32(m,1H),2.40(s,3H),1.93(s,3H),1.05(d,J＝6.4Hz,3H),0.90(s,1H),0.34(s,1H),0.16(d,J＝6.9Hz,2H),0.01(d,J＝4.9Hz,1H).Place Intermediate 1 (0.30g, 0.77mmol, 1eq) in a 100mL clean single-necked flask, add 1,4-dioxane (12mL) and water (3mL) to dissolve, and add Intermediate 15 (0.33g, 1eq) at room temperature. 1.46mmol, 1.9eq), sodium carbonate (0.25g, 2.31mmol, 3eq) and Pd(dppf)Cl 2 (0.056g, 0.077mmol, 0.1eq), stir and react at 100°C for 2 hours. The reaction solution was cooled to room temperature, 20 mL of water was added to the reaction solution, and ethyl acetate (30 mL × 3) was added for extraction three times. A saturated solution of sodium chloride was added to the organic phase to wash, and the liquids were separated. The organic phase was dried with anhydrous sodium sulfate and filtered. Concentrate and preparatively isolate to obtain compound S17 (9.2 mg, purity: 99.7%). LCMS: MS m/z(ESI):452.1[M+H] + . 1 H NMR (400MHz, DMSO-d6) δ11.98(s,1H),8.52(s,1H),8.27(t,J＝ 7.3Hz,1H),7.68(s,1H),7.09(d,J＝7.6Hz,1H),4.45(s,2H),3.39–3.32(m,1H),2.40(s,3H),1.93( s,3H),1.05(d,J＝6.4Hz,3H),0.90(s,1H),0.34(s,1H),0.16(d,J＝6.9Hz,2H),0.01(d,J＝4.9 Hz,1H).
实施例18化合物S18的制备
Example 18 Preparation of Compound S18
将化合物1(400mg，1.02mmol，1eq)，中间体16(1.5g，6.12mmol，6eq)，Na2CO3(325mg，3.06mmol，3eq)，置于100mL三口瓶中，加入40mL 1,4-二氧六环，8mL水，氩气置换，加入Pd(dppf)Cl2(73mg，0.1mmol，0.1eq)氩气置换，搅拌升温至100℃，反应2小时，取样检测，原料反应完全，反应液加入30mL水，50mL乙酸乙酯分层，水相20mL乙酸乙酯萃取三次，合并有机相，盐水洗涤分层，有机相无水硫酸钠干燥脱溶，送制备得化合物S18(110mg，0.211mmol，淡黄色固体，收率:22.88％)。LCMS：MS m/z(ESI):474.2[M+H]+.1HNMR(400MHz,DMSO-d6)δ12.24(s,1H),8.51(d,J＝4.8Hz,1H),7.96(s,1H),7.75(s,1H),7.59(dd,J＝5.1,1.6Hz,1H),4.69(s,2H),3.60(dd,J＝9.1,6.8Hz,1H),2.65(s,3H),2.16(s,3H),2.00(dd,J＝14.9,7.1Hz,1H),1.29(d,J＝6.8Hz,4H),1.16–1.05(m,1H),1.00(t,J＝2.1Hz,1H),0.98(d,J＝4.5Hz,2H),0.64–0.52(m,1H),0.46–0.33(m,2H),0.25(dd,J＝9.1,5.0Hz,1H).Place compound 1 (400mg, 1.02mmol, 1eq), intermediate 16 (1.5g, 6.12mmol, 6eq) and Na 2 CO 3 (325mg, 3.06mmol, 3eq) into a 100mL three-necked flask, and add 40mL 1,4 -Dioxane, 8mL water, argon replacement, add Pd(dppf)Cl 2 (73mg, 0.1mmol, 0.1eq) argon replacement, stir and heat to 100°C, react for 2 hours, take a sample and check, the raw material reaction is complete, Add 30 mL of water to the reaction solution, layer 50 mL of ethyl acetate, extract the aqueous phase three times with 20 mL of ethyl acetate, combine the organic phases, wash the layers with brine, dry the organic phase with anhydrous sodium sulfate to remove the solvent, and prepare compound S18 (110 mg, 0.211 mmol, light yellow solid, yield: 22.88%). LCMS: MS m/z (ESI): 474.2[M+H] + . 1 HNMR (400MHz, DMSO-d6) δ12.24 (s, 1H), 8.51 (d, J = 4.8Hz, 1H), 7.96 ( s,1H),7.75(s,1H),7.59(dd,J＝5.1,1.6Hz,1H),4.69(s,2H),3.60(dd,J＝9.1,6.8Hz,1H),2.65(s ,3H),2.16(s,3H),2.00(dd,J＝14.9,7.1Hz,1H),1.29(d,J＝6.8Hz,4H),1.16–1.05(m,1H),1.00(t, J＝2.1Hz,1H),0.98(d,J＝4.5Hz,2H),0.64–0.52(m,1H),0.46–0.33(m,2H),0.25(dd,J＝9.1,5.0Hz,1H ).
对比例1化合物S-34的制备
Comparative Example 1 Preparation of Compound S-34
步骤1：取化合物1(100mg,0.256mmol)溶于N,N-二甲基甲酰胺(3mL)中，分别加入氰化锌(150mg,1.28mmol),1,1'-双(二苯基膦)二茂铁(56.7mg,0.1mmol)，活化锌粉(16.6mg,0.256mmol),向反应液中吹入氩气30秒，加入三(二亚苄基丙酮)二钯(46.6mg,0.05mmol),继续向反应液中吹入氩气1分钟。微波125℃，反应一个小时。将反应液放至室温，过滤，滤饼加二氯甲烷洗涤两次。滤饼继续用二甲基亚砜(5mL)打浆30分钟，过滤用油泵抽干，得到化合物S-34-a(50mg,灰色固体，收率：51％)。1H-NMR(400MHz,DMSO-d6)δ 12.92–11.68(m,1H),8.20(s,1H),4.71(s,2H),3.71–3.49(m,1H),2.63(s,3H),2.18(s,3H),1.31(d,J＝6.8Hz,3H),1.24(s,1H),0.59(t,J＝7.0Hz,1H),0.48–0.35(m,2H),0.27(d,J＝4.9Hz,1H).Step 1: Dissolve compound 1 (100mg, 0.256mmol) in N,N-dimethylformamide (3mL), add zinc cyanide (150mg, 1.28mmol) and 1,1'-bis(diphenyl) respectively. Phosphine) ferrocene (56.7 mg, 0.1 mmol), activated zinc powder (16.6 mg, 0.256 mmol), blow argon gas into the reaction solution for 30 seconds, and add tris(dibenzylideneacetone) dipalladium (46.6 mg, 0.05 mmol), continue to blow argon gas into the reaction solution for 1 minute. Microwave at 125°C for one hour. The reaction solution was brought to room temperature, filtered, and the filter cake was washed twice with dichloromethane. The filter cake was continued to be beaten with dimethyl sulfoxide (5 mL) for 30 minutes, filtered and drained with an oil pump to obtain compound S-34-a (50 mg, gray solid, yield: 51%). 1 H-NMR (400MHz, DMSO-d 6 )δ 12.92–11.68(m,1H),8.20(s,1H),4.71(s,2H),3.71–3.49(m,1H),2.63(s,3H),2.18(s,3H),1.31(d, J＝6.8Hz,3H),1.24(s,1H),0.59(t,J＝7.0Hz,1H),0.48–0.35(m,2H),0.27(d,J＝4.9Hz,1H).
步骤2：将化合物S-34-a(30mg，0.079mmol),叠氮基三丁基锡烷(261mg,0.79mmol)溶于甲苯(2mL)中，置换氩气三次，油浴120℃反应36小时。将反应液放至室温，过滤，滤饼用二氯甲烷洗涤两次。滤饼用二甲基亚砜(5mL)溶解，用碱法[NH4HCO3]条件制备，得化合物S-34(1.21mg,白色固体，纯度99％)。1H-NMR(400MHz,DMSO-d6)δ12.29(s,1H),8.10(s,1H),5.33(t,J＝4.9Hz,1H),4.82(s,2H),3.68–3.53(m,1H),2.69(s,3H),2.19(s,3H),1.33(d,J＝6.8Hz,3H),0.86(t,J＝6.7Hz,1H),0.61(t,J＝6.8Hz,1H),0.44(d,J＝5.4Hz,2H),0.28(d,J＝5.5Hz,1H).Step 2: Dissolve compound S-34-a (30 mg, 0.079 mmol) and azidotributylstannane (261 mg, 0.79 mmol) in toluene (2 mL), replace the argon gas three times, and react in an oil bath at 120°C for 36 hours. The reaction solution was brought to room temperature, filtered, and the filter cake was washed twice with dichloromethane. The filter cake was dissolved in dimethyl sulfoxide (5 mL) and prepared using alkaline method [NH 4 HCO 3 ] conditions to obtain compound S-34 (1.21 mg, white solid, purity 99%). 1 H-NMR (400MHz, DMSO-d 6 ) δ12.29 (s, 1H), 8.10 (s, 1H), 5.33 (t, J = 4.9Hz, 1H), 4.82 (s, 2H), 3.68–3.53 (m,1H),2.69(s,3H),2.19(s,3H),1.33(d,J＝6.8Hz,3H),0.86(t,J＝6.7Hz,1H),0.61(t,J＝ 6.8Hz,1H),0.44(d,J＝5.4Hz,2H),0.28(d,J＝5.5Hz,1H).
测试例1：对Raw264.7细胞Akt S473磷酸化水平的抑制Test Example 1: Inhibition of Akt S473 phosphorylation level in Raw264.7 cells
本实验用ELISA方法检测。This experiment uses ELISA method for detection.
1.材料及仪器来源1. Source of materials and instruments
Raw264.7细胞购自ATCC；mC5a蛋白购自R&D；DMEM无血清培养基购自Gibco；胎牛血清(BSA)购自Gibco；RIPA缓冲液购自CST；蛋白酶磷酸酶抑制剂购自CST；磷酸化Akt S473的ELISA检测试剂盒购自R&D；96孔板购自Falcon；高结合(high binding)96孔板购自Costar；Infinite M1000荧光酶标仪购自Tecan。Raw264.7 cells were purchased from ATCC; mC5a protein was purchased from R&D; DMEM serum-free medium was purchased from Gibco; fetal calf serum (BSA) was purchased from Gibco; RIPA buffer was purchased from CST; protease phosphatase inhibitor was purchased from CST; phosphate The ELISA detection kit for Akt S473 was purchased from R&D; the 96-well plate was purchased from Falcon; the high binding 96-well plate was purchased from Costar; the Infinite M1000 fluorescence microplate reader was purchased from Tecan.
2.待测化合物溶液的配制2. Preparation of solution of compound to be tested
分别用100％DMSO将表1中的待测化合物的粉末配制成10mM的DMSO溶液，然后取4μL加入196μL的100％DMSO中，并用100％DMSO进行3倍梯度稀释，共8个浓度点。分别取10μL各浓度待测化合物溶液加入190μL无血清培养基中。Use 100% DMSO to prepare the powder of the test compound in Table 1 into a 10mM DMSO solution, then add 4 μL to 196 μL of 100% DMSO, and perform a 3-fold gradient dilution with 100% DMSO for a total of 8 concentration points. Take 10 μL of each concentration of the compound solution to be tested and add it to 190 μL of serum-free medium.
3.Raw264.7细胞处理3.Raw264.7 cell processing
取对数生长期Raw264.7细胞(PI3Kγ)(培养条件：DMEM+10％FBS(v/v))，以2.2×106个/mL的浓度90μL铺种于96孔板中(Falcon 353072)，于37℃，5％CO2条件下培养，过夜待细胞贴壁后，分别加入10μL上述制备得到的不同浓度的待测化合物的DMSO溶液(对照孔和空白孔只加DMSO)，于37℃，5％CO2条件下培养，2小时后，加入5μL mC5a蛋白的DMEM无血清培养基溶液(R&D 2150-C5)(空白孔只加培养基)，室温孵育5分钟后，去掉培养基，加入100μL 1×细胞裂解液(由RIPA缓冲液(CST 9806S)和蛋白酶磷酸酶抑制剂配制(CST 5872S)，按照相应试剂说明书配制)，800rpm振荡1分钟，分别形成对照组、空白组和实验组，储存于-80℃待用。Take Raw264.7 cells (PI3Kγ) in the logarithmic growth phase (culture conditions: DMEM+10% FBS (v/v)), and seed them in a 96-well plate (Falcon 353072) at a concentration of 2.2×10 6 cells/mL in 90 μL. , incubate at 37°C and 5% CO 2 overnight. After the cells adhere to the wall, add 10 μL of DMSO solutions of different concentrations of the test compounds prepared above (only DMSO is added to the control wells and blank wells), and incubate at 37°C. , culture under 5% CO 2 conditions. After 2 hours, add 5 μL of mC5a protein DMEM serum-free medium solution (R&D 2150-C5) (only add medium to the blank well). After incubation at room temperature for 5 minutes, remove the medium and add 100μL 1× cell lysis solution (prepared from RIPA buffer (CST 9806S) and protease phosphatase inhibitor (CST 5872S), prepared according to the corresponding reagent instructions), shake at 800rpm for 1 minute, and form a control group, blank group and experimental group respectively. Store at -80°C until use.
4.检测步骤4. Detection steps
按照磷酸化Akt S473的ELISA检测试剂盒(R&D DYC887BE)的说明书配制捕获抗体(Capture)溶液，将Capture溶液以每孔100μL加入高结合(high binding)96孔板中(Costar 42592)，室温孵育过夜。用300μL PBST清洗三次，加入封闭液(含1％(w/v)BSA的PBS溶液)(Genview FA016-100G)，室温孵育2小时，再用300μL PBST清洗三次；分别加入90μL对照组、空白组和实验组的细胞样本，室温孵育2小时，用300μL PBST清洗三次；加入100μL anti-pAkt S473稀释液，室温孵育2小时，用300μL PBST清洗三次；加入100μL辣根过氧化物酶标记的链霉亲和素(Streptavidin-HRP)稀释液，室温孵育40分钟，用300μL PBST清洗三次；加入100μL ELISA反应底物四甲基联苯胺(TMB)，室温孵育20分钟，加入50μL 2N(mol/L)的H2SO4溶液终止反应。于Infinite M1000荧光酶标仪(Tecan)上读取OD值，吸收波长为450nm和570nm。Prepare the capture antibody (Capture) solution according to the instructions of the ELISA detection kit for phosphorylated Akt S473 (R&D DYC887BE). Add 100 μL of the Capture solution per well into a high binding 96-well plate (Costar 42592) and incubate at room temperature overnight. . Wash three times with 300 μL PBST, add blocking solution (PBS solution containing 1% (w/v) BSA) (Genview FA016-100G), incubate at room temperature for 2 hours, then wash three times with 300 μL PBST; add 90 μL control group and blank group respectively. With the cell samples of the experimental group, incubate at room temperature for 2 hours, wash three times with 300 μL PBST; add 100 μL anti-pAkt S473 diluent, incubate at room temperature for 2 hours, wash three times with 300 μL PBST; add 100 μL horseradish peroxidase-labeled streptomyces Avidin (Streptavidin-HRP) dilution, incubate at room temperature for 40 minutes, wash three times with 300 μL PBST; add 100 μL ELISA reaction substrate tetramethylbenzidine (TMB), incubate at room temperature for 20 minutes, add 50 μL 2N (mol/L) The reaction was terminated with H 2 SO 4 solution. The OD value was read on an Infinite M1000 fluorescence microplate reader (Tecan), and the absorption wavelengths were 450nm and 570nm.
5.数据分析5.Data analysis
加DMSO不加mC5a蛋白为空白组，加DMSO及mC5a蛋白为对照组。 Adding DMSO without mC5a protein was the blank group, and adding DMSO and mC5a protein was the control group.
Inhibition％＝[1-(Total mean(实验组)-total mean(空白组))/(Total mean(对照组)-total mean(空白组))]×100％；其中，Inhibition％：化合物对Raw264.7细胞Akt S473磷酸化水平的抑制百分率；total mean：总平均值。Inhibition%=[1-(Total mean(experimental group)-total mean(blank group))/(Total mean(control group)-total mean(blank group))]×100%; among them, Inhibition%: compound’s effect on Raw264 .7% inhibition of Akt S473 phosphorylation level in cells; total mean: total mean.
6.将计算得到的抑制率用XLFIT 5.0软件计算出IC50值，测试结果如表1。6. Use the calculated inhibition rate to calculate the IC 50 value using XLFIT 5.0 software. The test results are shown in Table 1.
表1：化合物对RAW264.7细胞活性抑制IC50值
Table 1: IC 50 value of compounds inhibiting RAW264.7 cell activity
测试例2：对T47D和PC3细胞Akt S473磷酸化水平的抑制Test Example 2: Inhibition of Akt S473 phosphorylation levels in T47D and PC3 cells
本实验用Incell ELISA方法检测。In this experiment, Incell ELISA method was used for detection.
1.材料及仪器来源1. Source of materials and instruments
T47D细胞购自ATCC；PC3细胞购自SIBS；RPMI 1640和F12K培养基购自Gibco；磷酸化Akt S473(Ser473-p-Akt)抗体购自CST；96孔板购自Falcon；Infinite M1000荧光酶标仪购自Tecan；YZJ-0673为专利WO2016095833中的化合物S-37。T47D cells were purchased from ATCC; PC3 cells were purchased from SIBS; RPMI 1640 and F12K culture media were purchased from Gibco; phosphorylated Akt S473 (Ser473-p-Akt) antibody was purchased from CST; 96-well plates were purchased from Falcon; Infinite M1000 fluorescent enzyme label The instrument was purchased from Tecan; YZJ-0673 is compound S-37 in patent WO2016095833.
2.待测化合物溶液的配制2. Preparation of solution of compound to be tested
分别用100％DMSO将表2中的待测化合物的粉末配制成10mM的DMSO溶液，然后取20μL加入80μL的100％DMSO中，并用100％DMSO进行3倍梯度稀释，共8个浓度点。取10μL各浓度待测化合物溶液加入190μL无血清培养基中。Use 100% DMSO to prepare the powder of the test compound in Table 2 into a 10mM DMSO solution, then add 20 μL to 80 μL of 100% DMSO, and perform 3-fold gradient dilution with 100% DMSO for a total of 8 concentration points. Take 10 μL of the compound solution to be tested at each concentration and add it to 190 μL of serum-free medium.
3.T47D和PC3细胞处理3.T47D and PC3 cell processing
取对数生长期T47D(PI3Kα)(培养条件：RPMI 1640+10％FBS(v/v))和PC3(PI3Kβ)(培养条件：F12K+10％FBS(v/v))细胞，以2.2×105个/mL(PC3)或4.4×105个/mL(T47D)的浓度90μL铺种于96孔板中(Falcon 353072)，于37℃，5％CO2条件下培养，过夜待细胞贴壁后，分别加入10mL上述制备得到的不同浓度的待测化合物的DMSO溶液(对照孔只加DMSO，背景孔加10μM YZJ-0673)，于37℃，5％CO2条件下培养，2小时后，加入100μL 4％多聚甲醛(Biosharp BL539A)，室温孵育45分钟后，取出，再加入100μL 0.1％Triton X-100(v/v)溶液，继续孵育30分钟，分别形成对照组、背景组和实验组。Take T47D (PI3Kα) (culture conditions: RPMI 1640+10% FBS (v/v)) and PC3 (PI3Kβ) (culture conditions: F12K + 10% FBS (v/v)) cells in the logarithmic growth phase and incubate them at 2.2× 90 μL of a concentration of 10 5 cells/mL (PC3) or 4.4 × 10 5 cells/mL (T47D) was plated in a 96-well plate (Falcon 353072), cultured at 37°C, 5% CO 2 , and left overnight until the cells were attached. After the wall, add 10 mL of DMSO solutions of different concentrations of the test compound prepared above (only DMSO is added to the control well, and 10 μM YZJ-0673 is added to the background well), and incubate at 37°C and 5% CO2 for 2 hours. , add 100 μL 4% paraformaldehyde (Biosharp BL539A), incubate at room temperature for 45 minutes, take it out, add 100 μL 0.1% Triton X-100 (v/v) solution, and continue to incubate for 30 minutes to form a control group, background group and test group.
4.检测步骤4. Detection steps
对照组、背景组和实验组分别取出Triton X-100溶液，用200μL PBS清洗两次(每次300rpm，1分钟)，加入封闭液(含1％(w/v)BSA的PBS溶液)(Genview FA016-100G)，室温孵育2小时后取出，用200μL PBS清洗一次(300rpm，1分钟)，加入30μL Ser473-p-Akt抗体0.1％(w/v)BSA稀释液(cell signaling 4060L)，4℃孵育过夜。取出Ser473-p-Akt抗体，用200μL PBS清洗两次(每次300rpm，1分钟)，加入100μL辣根过氧化物酶标记的抗兔免疫球蛋白G(HRP anti-rabbit IgG)、0.1％BSA稀释液(w/v)(CST 7074S)，室温孵育1.5小时。用200μL PBS清洗两次(每次300rpm，1分钟)，加入100μL ELISA反应底物四甲基联苯胺(TMB)，室温孵育0.5小时后，加入50μL 2N硫酸终止反应，室温孵育20min后，于Infinite M1000荧 光酶标仪(Tecan)上读取OD值，吸收波长为450nm。The control group, background group and experimental group respectively took out the Triton X-100 solution, washed it twice with 200 μL PBS (300 rpm, 1 minute each time), and added blocking solution (PBS solution containing 1% (w/v) BSA) (Genview FA016-100G), incubate at room temperature for 2 hours, take it out, wash once with 200 μL PBS (300 rpm, 1 minute), add 30 μL Ser473-p-Akt antibody 0.1% (w/v) BSA dilution (cell signaling 4060L), 4°C Incubate overnight. Remove the Ser473-p-Akt antibody, wash it twice with 200 μL PBS (300 rpm each time, 1 minute), and add 100 μL horseradish peroxidase-labeled anti-rabbit immunoglobulin G (HRP anti-rabbit IgG) and 0.1% BSA. Dilution (w/v) (CST 7074S), incubate at room temperature for 1.5 hours. Wash twice with 200 μL PBS (300 rpm, 1 minute each time), add 100 μL ELISA reaction substrate tetramethylbenzidine (TMB), incubate at room temperature for 0.5 hours, add 50 μL 2N sulfuric acid to terminate the reaction, incubate at room temperature for 20 min, and incubate in Infinite M1000 fluorescent The OD value was read on a light microplate reader (Tecan), and the absorption wavelength was 450 nm.
取出TMB溶液，用200μL PBS清洗三次(每次300rpm，1分钟)，加入100μL 0.1％Janus水溶液(w/v)(Abcam ab111622)，室温孵育10分钟，取出Janus溶液，用200μL双重水清洗5次(每次300rpm，1分钟)，加入100μL 1.5M的盐酸，室温孵育10分钟，于Infinite M1000荧光酶标仪(Tecan)上读取OD值，吸收波长为595nm。Take out the TMB solution, wash it three times with 200μL PBS (300rpm each time, 1 minute), add 100μL 0.1% Janus aqueous solution (w/v) (Abcam ab111622), incubate at room temperature for 10 minutes, take out the Janus solution, wash it 5 times with 200μL double water (300rpm, 1 minute each time), add 100μL 1.5M hydrochloric acid, incubate at room temperature for 10 minutes, read the OD value on an Infinite M1000 fluorescence microplate reader (Tecan), and the absorption wavelength is 595nm.
5.数据分析5.Data analysis
加10μΜYZJ-0673为背景组，加DMSO为对照组。Add 10 μM YZJ-0673 as the background group, and add DMSO as the control group.
Inhibition％＝[1-(Ratio(实验组)–Ratio(背景组))/(Ratio(对照组)–Ratio(背景组))]×100％，其中，Inhibition％为化合物对T47D和PC3细胞活性抑制百分率，Ratio＝OD450/OD595。将计算得到的抑制率用XLFIT 5.0软件计算出IC50值，测试结果如表2。Inhibition%=[1-(Ratio(experimental group)-Ratio(background group))/(Ratio(control group)-Ratio(background group))]×100%, where Inhibition% is the activity of the compound on T47D and PC3 cells Inhibition percentage, Ratio=OD450/OD595. Use XLFIT 5.0 software to calculate the IC 50 value from the calculated inhibition rate. The test results are shown in Table 2.
表2化合物对T47D和PC3细胞活性抑制IC50值
Table 2 Compounds inhibit the activity of T47D and PC3 cells with IC 50 values
从表1和表2可以看出，本申请化合物对Raw264.7细胞Akt S473磷酸化水平具有较高的抑制活性，对T47D和PC3细胞抑制活性低，说明了本申请化合物对PI3Kγ具有较高的选择抑制活性。As can be seen from Table 1 and Table 2, the compound of the present application has high inhibitory activity on the phosphorylation level of Akt S473 in Raw264.7 cells and low inhibitory activity on T47D and PC3 cells, indicating that the compound of the present application has high inhibitory activity on PI3Kγ. Select inhibitory activity.
测试例3：对PI3K激酶抑制活性的测定Test Example 3: Determination of PI3K kinase inhibitory activity
1.材料与仪器来源1. Source of materials and instruments
人源PI3Kγ和PI3Kδ激酶购自Millipore，人源PI3Kα激酶购自Invitrogen，人源PI3Kβ激酶购自Eurofins，ADP-Glo激酶检测试剂盒购自Promega，PIP2脂质底物购自Life Technologies，384孔板购自PE。Human PI3Kγ and PI3Kδ kinase were purchased from Millipore, human PI3Kα kinase was purchased from Invitrogen, human PI3Kβ kinase was purchased from Eurofins, ADP-Glo kinase detection kit was purchased from Promega, and PIP2 lipid substrate was purchased from Life Technologies, 384-well plate Purchased from PE.
2.测试方法步骤2. Test method steps
本实验使用ADP-Glo的方法检测化合物对PI3Kγ、PI3Kα、PI3Kβ和PI3Kδ活性的抑制，所用试剂为PI3Kγ(Millipore 14-558)，PI3Kα(Invitrogen PV4788)、PI3Kβ(Eurofins 14-603M)、PI3Kδ(Millipore 14-604M)、PIP2(Life Technologies PR8982B)，ADP-Glo(Promega V9102/3)。This experiment uses the ADP-Glo method to detect the inhibition of the activities of PI3Kγ, PI3Kα, PI3Kβ and PI3Kδ by compounds. The reagents used are PI3Kγ (Millipore 14-558), PI3Kα (Invitrogen PV4788), PI3Kβ (Eurofins 14-603M), PI3Kδ (Millipore 14-604M), PIP2 (Life Technologies PR8982B), ADP-Glo (Promega V9102/3).
通过下述方法测定表3中待测化合物对PI3K激酶活性的抑制作用待测化合物用100％DMSO配制成实验所需最大浓度的100×的DMSO溶液，然后用100％DMSO进行3倍梯度稀释，共10个浓度点。在384孔板(PE 6008289)中加入50nL配制好的待测化合物溶液，其中空白孔和对照孔加入DMSO。配制1×缓冲液，并用1×缓冲液配制2×的激酶酶稀释液(反应浓度：PI3Kγ2.5μg/mL、PI3Kα0.15μg/mL、PI3Kβ0.3μg/mL和PI3Kδ1.2μg/mL)和2×反应底物稀释液ATP(反应浓度：25μM)/PIP2，每孔中分别加入2.5μL的激酶稀释液和2.5μL的反应底物稀释液，其中空白孔加入2.5μL的1×缓冲液和2.5μL的反应底物稀释液。震荡混匀后，于25℃孵育1小时；加入5μL ADP-Glo试剂盒中的ADP-Glo reagent终止反应，震荡混匀，25℃孵育2小时；加入10μL ADP-Glo试剂盒中的Kinase Detection Reagent，震荡混匀，25℃孵育30分钟，分别形成对照组、空白组和实验组，用Envision荧光酶标仪测定各孔化学发光强度。The inhibitory effect of the test compounds on PI3K kinase activity in Table 3 was determined by the following method. The test compounds were prepared with 100% DMSO into a 100× DMSO solution with the maximum concentration required for the experiment, and then 3-fold gradient dilution was performed with 100% DMSO. There are 10 concentration points in total. Add 50nL of the prepared test compound solution to the 384-well plate (PE 6008289), and add DMSO to the blank wells and control wells. Prepare 1× buffer, and use 1× buffer to prepare 2× kinase enzyme dilution (reaction concentration: PI3Kγ2.5 μg/mL, PI3Kα0.15 μg/mL, PI3Kβ0.3 μg/mL and PI3Kδ1.2 μg/mL) and 2× Reaction substrate diluent ATP (reaction concentration: 25 μM)/PIP2, add 2.5 μL of kinase diluent and 2.5 μL of reaction substrate diluent to each well, and add 2.5 μL of 1× buffer and 2.5 μL to the blank hole. reaction substrate dilution. After shaking and mixing, incubate at 25°C for 1 hour; add 5 μL of ADP-Glo reagent in the ADP-Glo kit to terminate the reaction, shake and mix, and incubate at 25°C for 2 hours; add 10 μL of Kinase Detection Reagent in the ADP-Glo kit , shake and mix, and incubate at 25°C for 30 minutes to form a control group, a blank group, and an experimental group. Use an Envision fluorescent microplate reader to measure the chemiluminescence intensity of each well.
3.数据分析3.Data analysis
抑制率％＝[1-(平均值(实验组)-平均值(空白组))/(平均值(对照组)-平均值(空白组))]×100％Inhibition rate % = [1-(average value (experimental group)-average value (blank group))/(average value (control group)-average value (blank group))]×100%
各待测化合物不同浓度各设2个平行孔。测得的本申请化合物对PI3K激酶4个亚型的 抑制活性的IC50值见表3和表4。Two parallel wells were set up at different concentrations for each compound to be tested. The measured activity of the compound of the present application on the four subtypes of PI3K kinase IC50 values for inhibitory activity are shown in Tables 3 and 4.
表3：化合物对PI3Kγ激酶活性抑制IC50值
Table 3: IC 50 value of compounds inhibiting PI3Kγ kinase activity
表4：化合物对PI3Kα、PI3Kβ和PI3Kδ激酶活性抑制IC50值
Table 4: IC 50 values of compounds inhibiting PI3Kα, PI3Kβ and PI3Kδ kinase activity
从表3和表4可以看出，本申请化合物对PI3Kγ激酶具有较高的抑制活性，对PI3Kα、PI3Kβ和PI3Kδ激酶抑制活性低，说明了本申请化合物对PI3Kγ具有较高的选择抑制活性。It can be seen from Table 3 and Table 4 that the compound of the present application has high inhibitory activity against PI3Kγ kinase and low inhibitory activity against PI3Kα, PI3Kβ and PI3Kδ kinase, indicating that the compound of the present application has high selective inhibitory activity against PI3Kγ.
测试例4：小鼠体内药代试验Test Example 4: In vivo pharmacokinetic test in mice
应用LC/MS/MS法测定了小鼠分别静脉注射给药和口服灌胃给药本申请化合物后不同时刻血浆中的药物浓度，以已知化合物D1和本申请的化合物S11、S5和S18为例，研究本申请化合物在小鼠体内的药代动力学行为，评价其药动学特征。The LC/MS/MS method was used to determine the drug concentration in the plasma of mice at different times after intravenous injection and oral gavage administration of the compound of the present application. The known compound D1 and the compounds of the present application S11, S5 and S18 were For example, study the pharmacokinetic behavior of the compound of the present application in mice and evaluate its pharmacokinetic characteristics.
其中，化合物D1(CAS号2504036-13-7)的结构如下所示，可参照现有文献进行制备或通过市购得到。
Among them, the structure of compound D1 (CAS No. 2504036-13-7) is as follows, which can be prepared by referring to existing literature or purchased commercially.
1.材料及仪器来源1. Source of materials and instruments
成年雄性ICR小鼠由北京维通利华实验动物技术有限公司提供；聚乙二醇15-羟基硬脂酸酯(Solutol HS15)购自Sigma，货号为42966；磺丁基-β-环糊精(Captisol)购自Sigma-ALDRICH，货号为332607；抗凝剂乙二胺四乙酸二钾(EDTA-K2)购自Sigma，货号为V900157；二甲基亚砜购自Sigma，货号为D5879；地塞米松(品牌NIFDC，批号:6TUC-T4C2)。Adult male ICR mice were provided by Beijing Weitonglihua Experimental Animal Technology Co., Ltd.; polyethylene glycol 15-hydroxystearate (Solutol HS15) was purchased from Sigma, product number 42966; sulfobutyl-β-cyclodextrin (Captisol) was purchased from Sigma-ALDRICH, the product number is 332607; the anticoagulant dipotassium ethylenediaminetetraacetate (EDTA-K 2 ) was purchased from Sigma, the product number is V900157; dimethyl sulfoxide was purchased from Sigma, the product number is D5879; Dexamethasone (brand name NIFDC, lot number: 6TUC-T4C2).
2.实验方案：2. Experimental plan:
2.1试验动物：健康成年雄性ICR小鼠(体重25-40g，每6只一组，静脉注射组小鼠自由饮水和饮食；灌胃给药组禁食过夜，给药4h后自由饮水和饮食)；2.1 Experimental animals: healthy adult male ICR mice (weight 25-40g, each group of 6, mice in the intravenous injection group can drink water and eat freely; mice in the intragastric administration group are fasted overnight, and can drink water and eat freely 4 hours after administration) ;
2.2给药方式与剂量：给药前挑选符合实验要求(检疫合格，通过实验室适应期，体重符 合要求)的动物，称重标记。ICR小鼠尾静脉给药，给药剂量2mg/kg,溶媒为5％DMSO+30％(10％Solutol HS15)+65％(20％Captisol，pH 7.4)，v/v/v；灌胃给药剂量为10mg/kg,溶媒为5％DMSO+30％(10％Solutol HS15)+65％(20％Captisol，pH 7.4)，v/v/v。化合物称取一定重量，按照0.4mg/mL(静脉注射组)或1mg/mL(灌胃给药组)浓度溶于溶媒中，化合物给予量按5mL/kg(静脉注射组)或10mL/kg(灌胃给药组)以体重换算，即如小鼠称重后体重为25g，静脉给药化合物量为：25/1000kg×0.4mg/mL×5mL/kg＝0.05mg，灌胃给药化合物量以此类推计算。2.2 Dosing method and dosage: Select the drug that meets the experimental requirements before administration (qualifies for quarantine, passes the laboratory adaptation period, and has a weight that meets the requirements of the experiment) Animals that meet the requirements are weighed and marked. ICR mice were administered via tail vein, the dosage was 2 mg/kg, the solvent was 5% DMSO + 30% (10% Solutol HS15) + 65% (20% Captisol, pH 7.4), v/v/v; administered by gavage The dosage is 10 mg/kg, the solvent is 5% DMSO + 30% (10% Solutol HS15) + 65% (20% Captisol, pH 7.4), v/v/v. Weigh a certain weight of the compound and dissolve it in the solvent at a concentration of 0.4 mg/mL (intravenous injection group) or 1 mg/mL (gavage administration group). The dosage of the compound is 5 mL/kg (intravenous injection group) or 10 mL/kg ( Gastrointestinal administration group) is converted by body weight, that is, if the weight of the mouse after weighing is 25g, the amount of compound administered intravenously is: 25/1000kg×0.4mg/mL×5mL/kg=0.05mg, the amount of compound administered intragastrically Calculate by analogy.
2.3血样采集：采集血样前，绑定小鼠，每一只给药的小鼠在预定的采血时间点(静脉给药：分别于给药后的0.083，0.25，0.5，1，2，4，7，24h采血，共8个时间点；灌胃给药：分别于给药后的0.083，0.25，0.5，1，2，4，7，24h采血，共8个时间点)，通过眼眦静脉采血约100μL。全血转移至预先加入抗凝剂EDTA-K2的1.5mL试管中，离心6min(8000rpm,4℃)，分离出血浆，整个过程在采血后15min内完成。所有的样品都需要存放于-20℃冰箱直至样品分析。2.3 Blood sample collection: Before collecting blood samples, bind the mice, and each administered mouse will collect blood at predetermined blood collection time points (intravenous administration: respectively at 0.083, 0.25, 0.5, 1, 2, 4, Collect blood at 7 and 24 hours, a total of 8 time points; intragastric administration: Collect blood at 0.083, 0.25, 0.5, 1, 2, 4, 7, and 24 hours after administration, a total of 8 time points), through the canthus vein Collect approximately 100 μL of blood. Transfer the whole blood to a 1.5 mL test tube pre-added with the anticoagulant EDTA-K 2 , centrifuge for 6 minutes (8000 rpm, 4°C), and separate the plasma. The entire process is completed within 15 minutes after blood collection. All samples need to be stored in a -20°C refrigerator until sample analysis.
2.4分析方法：采用液相色谱-串联质谱法(型号：Triple QuadTM 4000)分析血浆样品，色谱条件为：色谱柱为：Waters XBridge-C18(2.1×50mm,3.5μm)，流动相A:含0.05％甲酸和5mmol/L乙酸铵的水溶液(v/v)，流动相B:含0.05％甲酸和5mmol/L乙酸铵的甲醇溶液(v/v)，流速为0.6mL/min，柱温为40℃，进样量为2μL，液相梯度为：
2.4 Analysis method: Use liquid chromatography-tandem mass spectrometry (model: Triple Quad TM 4000) to analyze plasma samples. The chromatographic conditions are: the chromatographic column is: Waters XBridge-C18 (2.1×50mm, 3.5μm), mobile phase A: containing Aqueous solution of 0.05% formic acid and 5mmol/L ammonium acetate (v/v), mobile phase B: methanol solution (v/v) containing 0.05% formic acid and 5mmol/L ammonium acetate, the flow rate is 0.6mL/min, and the column temperature is 40°C, the injection volume is 2 μL, and the liquid phase gradient is:
质谱条件为：扫描模式：多反应离子监测(正离子模式)；Mass spectrometry conditions are: scan mode: multiple reaction ion monitoring (positive ion mode);
离子源：涡轮喷雾；电离模式：电喷雾离子化。Ion source: turbine spray; ionization mode: electrospray ionization.
2.5标准曲线的配制：将待测化合物称量适当粉末，按2mg/mL的浓度加入二甲基亚砜，作为试验的储备液。将储备液使用纯乙腈分别配制成20ng/mL，40ng/mL，100ng/mL，400ng/mL，2000ng/mL，10000ng/mL，34000ng/mL，40000ng/mL浓度的标准曲线工作溶液和60ng/mL，16000ng/mL，30000ng/mL浓度的质控样品工作溶液。将标准曲线工作液和质控样品工作液使用空白ICR小鼠血浆基质20倍稀释，配制待测化合物在空白ICR小鼠血浆基质中浓度为1.00ng/mL，2.00ng/mL，5.00ng/mL，20.0ng/mL，100ng/mL，500ng/mL，1700ng/mL，2000ng/mL的标准曲线和3ng/mL，800ng/mL，1500ng/mL的低中高浓度的质控样品。2.5 Preparation of standard curve: Weigh appropriate powder of the compound to be tested, and add dimethyl sulfoxide at a concentration of 2 mg/mL as the stock solution for the test. Use pure acetonitrile to prepare the stock solution into standard curve working solutions with concentrations of 20ng/mL, 40ng/mL, 100ng/mL, 400ng/mL, 2000ng/mL, 10000ng/mL, 34000ng/mL, 40000ng/mL and 60ng/mL. , 16000ng/mL, 30000ng/mL quality control sample working solution. Dilute the standard curve working solution and the quality control sample working solution 20 times using the blank ICR mouse plasma matrix, and prepare the concentration of the test compound in the blank ICR mouse plasma matrix to be 1.00ng/mL, 2.00ng/mL, and 5.00ng/mL. , 20.0ng/mL, 100ng/mL, 500ng/mL, 1700ng/mL, 2000ng/mL standard curves and 3ng/mL, 800ng/mL, 1500ng/mL low, medium and high concentration quality control samples.
2.6生物样品的处理：将冻存的血浆样品放置湿冰上解冻，待样品解冻后，放置涡旋仪上涡旋5min。分别取20μL血浆样品、标准曲线及质控指控样品加至96孔板中，再加入200μL含内标地塞米松(配制浓度2000ng/mL)的乙腈溶液，沉淀蛋白，涡旋混合5min，然后在3700rpm，4℃离心15min，取上清在同样条件下二次离心，最后进2μL上清溶液进行LC-MS/MS分析。测试结果参数及口服生物利用度F如表5。2.6 Processing of biological samples: Thaw the frozen plasma samples on wet ice. After the samples are thawed, place them on a vortexer and vortex for 5 minutes. Take 20 μL of plasma sample, standard curve and quality control sample and add them to the 96-well plate, then add 200 μL of acetonitrile solution containing internal standard dexamethasone (prepared concentration 2000ng/mL), precipitate the protein, vortex and mix for 5 minutes, and then Centrifuge at 3700 rpm and 4°C for 15 min. Take the supernatant and centrifuge it twice under the same conditions. Finally, add 2 μL of the supernatant solution for LC-MS/MS analysis. The test result parameters and oral bioavailability F are shown in Table 5.
表5化合物小鼠体内药物动力学(PK)参数

Table 5 In vivo pharmacokinetic (PK) parameters of compounds in mice

注：表5中“Cmax”为达峰浓度，“AUC0-t”为从0时到最终可定量时间点的血药浓度-时间曲线下面积，“F”为口服生物利用度。Note: “C max ” in Table 5 is the peak concentration, “AUC 0-t ” is the area under the plasma concentration-time curve from time 0 to the final quantifiable time point, and “F” is the oral bioavailability.
从表5可以看出，化合物S11、S18和化合物S5的暴露量(AUC0-t)和生物利用度(F)均高于化合物D1。说明本申请化合物具有更优异的体内药代参数。As can be seen from Table 5, the exposure (AUC 0-t ) and bioavailability (F) of compounds S11, S18 and compound S5 are higher than those of compound D1. This shows that the compound of the present application has better in vivo pharmacokinetic parameters.
测试例5：在小鼠结肠癌细胞MC38与体外分化的M2巨噬细胞同种混合皮下移植的肿瘤模型中的药效学测试Test Example 5: Pharmacodynamic test in a tumor model in which mouse colon cancer cells MC38 and in vitro differentiated M2 macrophages were mixed subcutaneously and transplanted
1.实验目的1. Experimental purpose
评价本申请化合物在小鼠结肠癌细胞MC38与体外培养的巨噬细胞同种混合皮下移植的肿瘤模型中的抗肿瘤作用。The anti-tumor effect of the compound of the present application was evaluated in a tumor model in which mouse colon cancer cell MC38 cells and macrophages cultured in vitro were mixed subcutaneously and transplanted.
2.受试物配制2. Preparation of test substance
2.1溶媒配制2.1 Solvent preparation
5％DMSO(二甲基亚砜)(v/v)+40％PEG400(v/v)+55％纯水(v/v)，给药体积10μL/g。5% DMSO (dimethyl sulfoxide) (v/v) + 40% PEG400 (v/v) + 55% pure water (v/v), dosage volume 10 μL/g.
2.2化合物S11给药制剂配制2.2 Preparation of compound S11 dosage preparation
称量适量化合物S11至离心管中，加入溶媒，涡旋至全溶溶液，浓度为1mg/mL，每周配制两次。Weigh an appropriate amount of compound S11 into a centrifuge tube, add solvent, and vortex until the solution is completely dissolved. The concentration is 1 mg/mL. Prepare twice a week.
2.3化合物D1给药制剂配制2.3 Preparation of compound D1 dosage preparation
称量适量化合物D1(CAS号2504036-13-7，结构参见测试例4)至离心管中，加入溶媒，涡旋至全溶溶液，浓度为1mg/mL。Weigh an appropriate amount of compound D1 (CAS No. 2504036-13-7, see Test Example 4 for structure) into a centrifuge tube, add solvent, and vortex until the solution is completely dissolved. The concentration is 1 mg/mL.
3.实验动物3. Experimental animals
C57BL/6小鼠，雌性，8-9周(肿瘤细胞接种时的小鼠周龄)，体重18.0-20.0g，每组8只。购自上海吉辉实验动物饲养有限公司。C57BL/6 mice, female, 8-9 weeks old (the age of mice when tumor cells were inoculated), weight 18.0-20.0g, 8 mice in each group. Purchased from Shanghai Jihui Experimental Animal Breeding Co., Ltd.
4.小鼠结肠癌细胞MC38培养4. Culture of mouse colon cancer cell MC38
小鼠结肠癌细胞MC38(购于上海交通大学)培养在含10％(v/v)胎牛血清(Gibco；10099-141C)、100U/mL青霉素和100pg/mL链霉素(Gibco；15140122)的RPMI-1640培养液(Gibco；61870-036)中。收集指数生长期的MC38细胞，PBS重悬至适合浓度用于小鼠皮下肿瘤接种。Mouse colon cancer cell MC38 (purchased from Shanghai Jiao Tong University) was cultured in a medium containing 10% (v/v) fetal calf serum (Gibco; 10099-141C), 100 U/mL penicillin and 100 pg/mL streptomycin (Gibco; 15140122). in RPMI-1640 culture medium (Gibco; 61870-036). MC38 cells in the exponential growth phase were collected and resuspended in PBS to a suitable concentration for subcutaneous tumor inoculation in mice.
5.小鼠巨噬细胞的体外培养5. In vitro culture of mouse macrophages
从C57BL/6小鼠腿骨中分离骨髓细胞，制备成单细胞悬液，添加50ng/mL M-CSF(Peprotech；315-02)诱导为骨髓来源的M0巨噬细胞；在M0巨噬细胞诱导过程中添加20ng/mL白细胞介素4(IL4)(Peprotech；214-14)培养48h，获得极化的M2巨噬细胞；体外培养的M0巨噬细胞和M2巨噬细胞分别与小鼠结肠癌细胞系MC38按1:1(个数)混合，皮下接种，总计106个细胞/小鼠。Bone marrow cells were isolated from the leg bones of C57BL/6 mice, prepared into single cell suspension, and added with 50ng/mL M-CSF (Peprotech; 315-02) to induce bone marrow-derived M0 macrophages; in M0 macrophage induction During the process, 20ng/mL interleukin 4 (IL4) (Peprotech; 214-14) was added and cultured for 48 hours to obtain polarized M2 macrophages; M0 macrophages and M2 macrophages cultured in vitro were closely related to mouse colon cancer. The cell line MC38 was mixed at a ratio of 1:1 (number) and inoculated subcutaneously, with a total of 10 6 cells/mouse.
6.动物接种分组及给药6. Animal vaccination grouping and administration
6.1动物接种分组6.1 Animal vaccination grouping
随机选取8只雌性C57BL/6小鼠背部皮下分别接种5×105个MC38细胞作为对照组；随机选取8只雌性C57BL/6小鼠背部皮下分别接种5×105个MC38细胞+5×105个M0巨噬细胞作为M0组；随机选取24只雌性C57BL/6小鼠背部皮下分别接种5×105个MC38细胞+5×105个M2巨噬细胞(肿瘤细胞与体外分化的M2巨噬细胞混合接种肿瘤模型的建立方法参考文 献Nature.2016；539(7629):437-442.)作为M2组；接种当天定义为第0天，第1天开始给药。Eight female C57BL/6 mice were randomly selected and inoculated subcutaneously on the back with 5 × 10 5 MC38 cells as the control group; 8 female C57BL/6 mice were randomly selected and inoculated subcutaneously with 5 × 10 5 MC38 cells + 5 × 10 cells on the back. 5 M0 macrophages were used as the M0 group; 24 female C57BL/6 mice were randomly selected and inoculated subcutaneously on the back with 5 × 10 5 MC38 cells + 5 × 10 5 M2 macrophages (tumor cells and in vitro differentiated M2 macrophages). Reference text for establishment of phagocyte mixed inoculation tumor model Nature.2016;539(7629):437-442.) was used as the M2 group; the day of vaccination was defined as day 0, and administration began on day 1.
6.2给药方式和剂量6.2 Administration method and dosage
本实验采用灌胃给药的方式，对照组小鼠分别灌胃溶媒作为溶媒组，给药体积10μL/g；M0组小鼠分别灌胃溶媒作为与M0巨噬细胞混接的溶媒组，给药体积10μL/g；随机选取8只M2组小鼠分别灌胃溶媒作为与M2巨噬细胞混接的溶媒组，给药体积10μL/g；随机选取8只M2组分别灌胃1mg/mL的化合物S11给药制剂作为与M2巨噬细胞混接的化合物S11组，给药剂量为10mg/kg；剩余M2组小鼠分别灌胃1mg/mL的化合物D1给药制剂作为与M2巨噬细胞混接的化合物D1组，给药剂量为10mg/kg。This experiment adopted the method of intragastric administration. The mice in the control group were administered with vehicle as the vehicle group, and the administration volume was 10 μL/g; the mice in the M0 group were administered with vehicle as the vehicle group mixed with M0 macrophages. The drug volume is 10 μL/g; 8 mice in the M2 group are randomly selected to be gavaged with the vehicle as the vehicle group mixed with M2 macrophages, and the dosage volume is 10 μL/g; 8 mice in the M2 group are randomly selected to be gavaged with 1 mg/mL of the drug. The compound S11 administration preparation was used as the compound S11 group mixed with M2 macrophages, and the dosage was 10 mg/kg; the mice in the remaining M2 group were administered 1 mg/mL compound D1 administration preparation as the compound D1 mixed with M2 macrophages. For the compound D1 group, the dosage was 10 mg/kg.
7.动物给药后观察7. Observation of animals after administration
常规监测包括观察肿瘤生长及治疗对动物正常行为的影响，具体内容有实验动物的活动性，摄食和饮水情况，体重增加或降低情况，眼睛、被毛及其它异常情况。每周称重和测量肿瘤体积2次，给药周期27天，于第27天称量体重测量肿瘤体积后处死小鼠取瘤块称重(TW)，计算肿瘤体积(TV)、相对肿瘤重量增长率(T/C)和肿瘤生长抑制率(瘤重)(TGI)，做统计学检测。T/C(％)＝T/C×100％，TGI(％)＝(1-T/C)×100％，T和C分别为治疗组(与M2巨噬细胞混接的化合物S11组或与M2巨噬细胞混接的化合物D1组)和对照组(溶媒组)某一特定时间点的相对肿瘤体积或重量。Routine monitoring includes observing the impact of tumor growth and treatment on the normal behavior of animals, including the activity of experimental animals, food and water intake, weight gain or loss, eyes, coat and other abnormalities. Weigh and measure the tumor volume twice a week. The dosing period is 27 days. On the 27th day, the body weight is measured and the tumor volume is measured. The mice are then killed and the tumor block is taken out and weighed (TW). The tumor volume (TV) and relative tumor weight are calculated. Growth rate (T/C) and tumor growth inhibition rate (tumor weight) (TGI) were analyzed statistically. T/C (%) = T/C × 100%, TGI (%) = (1-T/C) × 100%, T and C are the treatment groups respectively (compound S11 group mixed with M2 macrophages or The relative tumor volume or weight at a specific time point between the compound D1 group mixed with M2 macrophages) and the control group (vehicle group).
8.结果8.Results
实验期间，小鼠肿瘤生长曲线如图1所示。图1中，溶媒：溶媒组；M0+溶媒：与M0巨噬细胞混接的溶媒组；M2+溶媒：与M2巨噬细胞混接的溶媒组；M2+化合物D1：与M2巨噬细胞混接的化合物D1组，10mg/kg(mpk)，灌胃给药，每天一次给药；M2+化合物S11：与M2巨噬细胞混接的化合物S11组，10mg/kg，灌胃给药，每天一次给药。**，P<0.01；***，P<0.001；ns，P>0.05。(n＝8，平均值±标准误)。During the experiment, the mouse tumor growth curve is shown in Figure 1. In Figure 1, vehicle: vehicle group; M0+ vehicle: vehicle group mixed with M0 macrophages; M2+ vehicle: vehicle group mixed with M2 macrophages; M2+ compound D1: compound mixed with M2 macrophages D1 group, 10 mg/kg (mpk), administered by gavage, once a day; M2+ compound S11: compound S11 group mixed with M2 macrophages, 10 mg/kg, administered by gavage, once a day. **, P<0.01; ***, P<0.001; ns, P>0.05. (n=8, mean ± standard error).
给药后第27天模型各组小鼠肿瘤重量分析如图2所示。图2中，溶媒：溶媒组；M0+溶媒：与M0巨噬细胞混接的溶媒组；M2+溶媒：与M2巨噬细胞混接的溶媒组；M2+化合物D1：与M2巨噬细胞混接的化合物D1组，10mg/kg，灌胃给药，每天一次给药；M2+化合物S11：与M2巨噬细胞混接的化合物S11组，10mg/kg，灌胃给药，每天一次给药。**，P<0.01；ns，P>0.05。(n＝8，平均值±标准误)。The tumor weight analysis of mice in each group of the model on the 27th day after administration is shown in Figure 2. In Figure 2, vehicle: vehicle group; M0+ vehicle: vehicle group mixed with M0 macrophages; M2+ vehicle: vehicle group mixed with M2 macrophages; M2+ compound D1: compound mixed with M2 macrophages D1 group, 10 mg/kg, administered by gavage, once a day; M2+ compound S11: compound S11 group mixed with M2 macrophages, 10 mg/kg, administered by gavage, once a day. **, P<0.01; ns, P>0.05. (n=8, mean ± standard error).
给药后第27天模型中各组药效参数如下表6所示：The pharmacodynamic parameters of each group in the model on the 27th day after administration are shown in Table 6 below:
表6给药后第27天在小鼠MC38和巨噬细胞混合接种模型中各组药效分析表
Table 6 Analysis of drug efficacy of each group in the mouse MC38 and macrophage mixed inoculation model on the 27th day after administration
注：表6中“T/C”表示相对肿瘤重量增长率，“TGI”表示肿瘤生长抑制率(瘤重)，“\”表示无对应参数。Note: “T/C” in Table 6 indicates relative tumor weight growth rate, “TGI” indicates tumor growth inhibition rate (tumor weight), and “\” indicates no corresponding parameter.
由图1、图2和表6可知，在小鼠MC38和巨噬细胞混合接种模型中，本发明实施例化合物S11表现出显著的抑制肿瘤生长作用。说明本申请化合物具有更优异的抑制肿瘤生长作用。It can be seen from Figure 1, Figure 2 and Table 6 that in the mouse MC38 and macrophage mixed inoculation model, the compound S11 of the embodiment of the present invention showed a significant inhibitory effect on tumor growth. It shows that the compound of the present application has better inhibitory effect on tumor growth.
在本申请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。此外应理解，在阅读了本申请的上述讲授内容之后，本领域技术人员可以对本申请作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。All documents mentioned in this application are incorporated by reference in this application to the same extent as if each individual document was individually incorporated by reference. As a reference. In addition, it should be understood that after reading the above teaching content of this application, those skilled in the art can make various changes or modifications to this application, and these equivalent forms also fall within the scope defined by the appended claims of this application.
以上所述实施例的各技术特征可以进行任意的组合，为使描述简洁，未对上述实施例中的各个技术特征所有可能的组合都进行描述，然而，只要这些技术特征的组合不存在矛盾，都应当认为是本说明书记载的范围。The technical features of the above-described embodiments can be combined in any way. To simplify the description, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, All should be considered to be within the scope of this manual.
以上所述实施例仅表达了本申请的几种实施方式，其描述较为具体和详细，但并不能因此而理解为对申请专利范围的限制。应当指出的是，对于本领域的普通技术人员来说，在不脱离本申请构思的前提下，还可以做出若干变形和改进，这些都属于本申请的保护范围。因此，本申请专利的保护范围应以所附权利要求为准。 The above-described embodiments only express several implementation modes of the present application, and their descriptions are relatively specific and detailed, but they should not be construed as limiting the scope of the patent application. It should be noted that, for those of ordinary skill in the art, several modifications and improvements can be made without departing from the concept of the present application, and these all fall within the protection scope of the present application. Therefore, the protection scope of this patent application should be determined by the appended claims.

Claims (15)

  1. 一种式(I)所示的化合物、或其药学上可接受的盐、或其立体异构体：
    A compound represented by formula (I), or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof:
    其中，in,
    环A为5或6元杂芳基环；Ring A is a 5- or 6-membered heteroaryl ring;
    (R3)n表示环A上的氢被n个R3取代，n为0、1、2、3或4；每个R3相同或不同，各自独立地为氘、卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、NRa1Rb1、-N(Ra3)-C(O)C1-3烷基、-N(Ra3)-C(O)-氘代C1-3烷基、-N(Ra3)-C(O)OC1-3烷基、-SO2C1-3烷基、-SO2C3-6环烷基、-C(O)NRa1Rb1、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基、3至6元杂环烷基、苯基或5至6元杂芳基；其中所述3至6元杂环烷基、苯基、5至6元杂芳基各自独立地为未取代的或被1、2或3个各自独立地选自取代基组Q的取代基取代；(R 3 ) n means that the hydrogen on ring A is replaced by n R 3s , n is 0, 1, 2, 3 or 4; each R 3 is the same or different, and each R 3 is independently deuterium, halogen, cyano, or hydroxyl , carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halo C 1-3 alkyl, halo C 1-3 alkoxy , NR a1 R b1 , -N(R a3 )-C(O)C 1-3 alkyl, -N(R a3 )-C(O)-deuterated C 1-3 alkyl, -N(R a3 )-C(O)OC 1-3 alkyl, -SO 2 C 1-3 alkyl, -SO 2 C 3-6 cycloalkyl, -C(O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy, 3 to 6 membered heterocycloalkyl, phenyl or 5 to 6-membered heteroaryl; wherein the 3- to 6-membered heterocycloalkyl, phenyl, and 5- to 6-membered heteroaryl groups are each independently unsubstituted or 1, 2, or 3 are independently selected from substituents Substituent substitution of group Q;
    Z为N或CRZ；T为N或CRT；U为N或CRU；W为N或CRW；Y为N或CRY；Z is N or CR Z ; T is N or CR T ; U is N or CR U ; W is N or CR W ; Y is N or CR Y ;
    RZ、RT、RU各自独立地为氢、卤素、NRa1Rb1或C1-8烷基；其中所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基；R Z , RT , and RU are each independently hydrogen, halogen, NR a1 R b1 or C 1-8 alkyl; wherein the C 1-8 alkyl is preferably C 1-6 alkyl, more preferably C 1-3 alkyl;
    RW、RY各自独立地为氢、卤素、C1-8烷基、卤代C1-8烷基、卤代C1-8烷氧基、NRa1Rb1或C3-6环烷基；其中，所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基；R W and R Y are each independently hydrogen, halogen, C 1-8 alkyl, halogenated C 1-8 alkyl, halogenated C 1-8 alkoxy, NR a1 R b1 or C 3-6 cycloalkyl base; wherein, the C 1-8 alkyl group is preferably a C 1-6 alkyl group, and more preferably a C 1-3 alkyl group;
    或者RW、RY相连，与相邻6元环形成9或10元杂芳基环、9或10元苯基并杂环烷基环或9或10元杂芳基并杂环烷基环；所述9或10元杂芳基环、9或10元苯基并杂环烷基环、9或10元杂芳基并杂环烷基环各自独立地为未取代的或被1、2或3个各自独立地选自取代基组Q的取代基取代；Or R W and R Y are connected to form a 9- or 10-membered heteroaryl ring, a 9- or 10-membered phenyl heterocycloalkyl ring, or a 9- or 10-membered heteroaryl heterocycloalkyl ring with the adjacent 6-membered ring. ; The 9- or 10-membered heteroaryl ring, the 9- or 10-membered phenyl heterocycloalkyl ring, and the 9- or 10-membered heteroaryl heterocycloalkyl ring are each independently unsubstituted or substituted by 1 or 2 Or substituted by 3 substituents each independently selected from the substituent group Q;
    R1、R2各自独立地为C1-8烷基、卤代C1-8烷基或C3-6环烷基；其中，所述C3-6环烷基为未取代的或被1或2个各自独立地选自卤素和C1-3烷基的取代基取代；所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基；所述卤代C1-8烷基优选为卤代C1-6烷基，更优选为卤代C1-3烷基；R 1 and R 2 are each independently C 1-8 alkyl, halogenated C 1-8 alkyl or C 3-6 cycloalkyl; wherein, the C 3-6 cycloalkyl is unsubstituted or substituted 1 or 2 substituents each independently selected from halogen and C 1-3 alkyl are substituted; the C 1-8 alkyl is preferably C 1-6 alkyl, more preferably C 1-3 alkyl; so The halogenated C 1-8 alkyl group is preferably a halogenated C 1-6 alkyl group, and more preferably a halogenated C 1-3 alkyl group;
    取代基组Q选自卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、NRa1Rb1、-SO2C1-3烷基、-S(O)C1-3烷基、-C(O)NRa1Rb1、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基、3至6元杂环烷基、苯基或5至6元杂芳基；The substituent group Q is selected from halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halogenated C 1-3 Alkyl group, halogenated C 1-3 alkoxy group, NR a1 R b1 , -SO 2 C 1-3 alkyl group, -S(O)C 1-3 alkyl group, -C(O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy, 3 to 6-membered heterocycloalkyl , phenyl or 5 to 6-membered heteroaryl;
    Ra1、Rb1各自独立地为氢、C1-3烷基或乙酰基，或Ra1、Rb1与相连的氮原子共同形成4至6元饱和单杂环；所述4至6元饱和单杂环为未取代的或被1、2或3个各自独立地选自下述的取代基取代：氘、卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、-SO2C1-3烷基、-S(O)C1-3烷基、-C(O)NH2、-C(O)NH(C1-3烷基)、-C(O)N(C1-3烷基)2、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基或3至6元杂环烷基；R a1 and R b1 are each independently hydrogen, C 1-3 alkyl or acetyl, or R a1 , R b1 and the connected nitrogen atom together form a 4- to 6-membered saturated monoheterocyclic ring; the 4- to 6-membered saturated The monoheterocycle is unsubstituted or substituted by 1, 2 or 3 substituents each independently selected from the following: deuterium, halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkyl Oxygen group, C 2-4 alkenyl group, C 2-4 alkynyl group, halogenated C 1-3 alkyl group, halogenated C 1-3 alkoxy group, -SO 2 C 1-3 alkyl group, -S(O )C 1-3 alkyl, -C(O)NH 2 , -C(O)NH(C 1-3 alkyl), -C(O)N(C 1-3 alkyl) 2 , -C( O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyloxy or 3 to 6-membered heterocycloalkyl;
    Ra3为氢或C1-8烷基；其中所述C1-8烷基优选为C1-6烷基，更优选为C1-3烷基。 R a3 is hydrogen or C 1-8 alkyl; wherein the C 1-8 alkyl is preferably C 1-6 alkyl, more preferably C 1-3 alkyl.
  2. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，T为CRT；U为CRU；W为N或CRW；Y为N或CRY；且W、Y不同时为N。The compound of claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein T is CRT ; U is CRU ; W is N or CRW ; Y is N or CR Y ; and W and Y are not N at the same time.
  3. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，T为N；U为CRU；W为CRW；Y为N。The compound of claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein T is N; U is CRU ; W is CRW ; Y is N.
  4. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，结构为选自式(A)、式(B)和式(C)所示结构：
    The compound according to claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, characterized in that the structure The structure is selected from the group consisting of formula (A), formula (B) and formula (C):
  5. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，结构选自以下结构中的一种：

    The compound according to claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, characterized in that the structure Choose one of the following structures:

  6. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，环A选自下组的5或6元杂芳基环：噻吩环、呋喃环、噻唑环、异噻唑环、咪唑环、噁唑环、吡咯环、吡唑环、1,2,3-三唑环、1,2,4-三唑环、1,2,5-三唑环、1,3,4-三唑环、四唑环、异噁唑环、1,2,3-噁二唑环、1,2,4-噁二唑环、1,2,5-噁二唑环、1,3,4-噁二唑环、噻二唑环、吡啶环、哒嗪环、嘧啶环、吡嗪环、三嗪环或四嗪环。The compound of claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein ring A is selected from the following group of 5- or 6-membered heteroaryl rings: thiophene ring, furan ring , thiazole ring, isothiazole ring, imidazole ring, oxazole ring, pyrrole ring, pyrazole ring, 1,2,3-triazole ring, 1,2,4-triazole ring, 1,2,5-triazole Ring, 1,3,4-triazole ring, tetrazole ring, isoxazole ring, 1,2,3-oxadiazole ring, 1,2,4-oxadiazole ring, 1,2,5-oxadiazole ring Diazole ring, 1,3,4-oxadiazole ring, thiadiazole ring, pyridine ring, pyridazine ring, pyrimidine ring, pyrazine ring, triazine ring or tetrazine ring.
  7. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，式(I)中，为式(a)所示结构：
    The compound according to claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, characterized in that, in formula (I), It is the structure shown in formula (a):
    R01、R02各自独立地为氘、卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、C2-4烯基、C2-4炔基、卤代C1-3烷基、卤代C1-3烷氧基、NRa1Rb1、-N(Ra3)-C(O)C1-3烷基、-N(Ra3)-C(O)-氘代C1-3烷基、-N(Ra3)-C(O)OC1-3烷基、-SO2C1-3烷基、-SO2C3-6环烷基、-C(O)NRa1Rb1、-C(O)OC1-3烷基、-OC(O)C1-3烷基、C3-6环烷基、C3-6环烷基氧基、3至6元杂环烷基、苯基或5至6元杂芳基；其中所述3至6元杂环烷基、苯基、5至6元杂芳基各自独立地为未取代的或被1、2或3个各自独立地选自取代基组Q的取代基取代。R 01 and R 02 are each independently deuterium, halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1-3 alkoxy, C 2-4 alkenyl, C 2-4 alkynyl, halogen Substituted C 1-3 alkyl, halogenated C 1-3 alkoxy, NR a1 R b1 , -N(R a3 )-C(O)C 1-3 alkyl, -N(R a3 )-C( O)-deuterated C 1-3 alkyl, -N(R a3 )-C(O)OC 1-3 alkyl, -SO 2 C 1-3 alkyl, -SO 2 C 3-6 cycloalkyl , -C(O)NR a1 R b1 , -C(O)OC 1-3 alkyl, -OC(O)C 1-3 alkyl, C 3-6 cycloalkyl, C 3-6 cycloalkyl Oxygen, 3 to 6-membered heterocycloalkyl, phenyl or 5 to 6-membered heteroaryl; wherein the 3 to 6-membered heterocycloalkyl, phenyl, 5 to 6-membered heteroaryl are each independently Substituted or substituted by 1, 2 or 3 substituents each independently selected from the substituent group Q.
  8. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，R01为卤素、氰基、羟基、羧基、C1-3烷基、C1-3烷氧基、卤代C1-3烷基或卤代C1-3烷氧基；R02为-NH-C(O)C1-3烷基、-NH-C(O)-氘代C1-3烷基或-NH-C(O)OC1-3烷基。The compound of claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein R 01 is halogen, cyano, hydroxyl, carboxyl, C 1-3 alkyl, C 1 -3 alkoxy, halogenated C 1-3 alkyl or halogenated C 1-3 alkoxy; R 02 is -NH-C(O)C 1-3 alkyl, -NH-C(O)- Deuterated C 1-3 alkyl or -NH-C(O)OC 1-3 alkyl.
  9. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，R1为一氯甲基、二氯甲基、三氯甲基、一氯乙基、1,2-二氯乙基、三氯乙基、一溴乙基、一氟甲基、二氟甲基、三氟甲基、一氟乙基、二氟乙基、三氟乙基、环丙基、环丁基、环戊基、环己基、一氟取代的环丙基或一氟取代的环丁基；R2为甲基、乙基、正丙基、异丙基、环丙基、环丁基、环戊基、环己基、一氟取代的环丙基或一氟取代的环丁基。The compound of claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein R1 is monochloromethyl, dichloromethyl, trichloromethyl, or monochloroethyl. base, 1,2-dichloroethyl, trichloroethyl, monobromoethyl, monofluoromethyl, difluoromethyl, trifluoromethyl, monofluoroethyl, difluoroethyl, trifluoroethyl , cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, monofluoro-substituted cyclopropyl or monofluoro-substituted cyclobutyl; R 2 is methyl, ethyl, n-propyl, isopropyl, cyclobutyl Propyl, cyclobutyl, cyclopentyl, cyclohexyl, monofluoro-substituted cyclopropyl or monofluoro-substituted cyclobutyl.
  10. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，结构选自以下结构中的一种：
    The compound according to claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, characterized in that the structure Choose one of the following structures:
  11. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在 于，结构选自以下结构中的一种：
    The compound according to claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, characterized in that Yu, structure Choose one of the following structures:
  12. 如权利要求1所述的化合物、或其药学上可接受的盐、或其立体异构体，其特征在于，式(I)化合物选自以下结构中的任意一种：

    The compound of claim 1, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, wherein the compound of formula (I) is selected from any one of the following structures:

  13. 一种药物组合物，其特征在于，包括权利要求1-12中任一项所述的化合物、或其药学上可接受的盐、或其立体异构体；以及药学可接受的载体。A pharmaceutical composition, characterized by comprising the compound according to any one of claims 1 to 12, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof; and a pharmaceutically acceptable carrier.
  14. 一种如权利要求1-12任一所述的化合物、或其药学上可接受的盐、或其立体异构体，或权利要求13所述的药物组合物在制备治疗和/或预防与PI3Kγ活性相关的或由PI3Kγ活性介导的疾病的药物中的应用。A compound as claimed in any one of claims 1 to 12, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof, or a pharmaceutical composition as claimed in claim 13 in the preparation of the treatment and/or prevention of PI3Kγ Drug applications in diseases related to or mediated by PI3Kγ activity.
  15. 如权利要求14所述的应用，其特征在于，所述与PI3Kγ活性相关的或由PI3Kγ活性介导的疾病为炎症、代谢性疾病或癌症。 The application according to claim 14, wherein the disease related to or mediated by PI3Kγ activity is inflammation, metabolic disease or cancer.
PCT/CN2023/116425 2022-09-01 2023-09-01 Heteroaryl-substituted pyridopyrrolidone derivative, and pharmaceutical composition and use thereof WO2024046454A1 (en)

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