WO2022233316A1 - Composé macrocyclique à douze éléments - Google Patents

Composé macrocyclique à douze éléments Download PDF

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Publication number
WO2022233316A1
WO2022233316A1 PCT/CN2022/091154 CN2022091154W WO2022233316A1 WO 2022233316 A1 WO2022233316 A1 WO 2022233316A1 CN 2022091154 W CN2022091154 W CN 2022091154W WO 2022233316 A1 WO2022233316 A1 WO 2022233316A1
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compound
added
alkyl
pharmaceutically acceptable
acceptable salt
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PCT/CN2022/091154
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English (en)
Chinese (zh)
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张杨
付志飞
孙继奎
陈健
黎健
陈曙辉
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南京明德新药研发有限公司
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Priority to CN202280027673.9A priority Critical patent/CN117177980A/zh
Publication of WO2022233316A1 publication Critical patent/WO2022233316A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/529—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim forming part of bridged ring systems
    • 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
    • C07D498/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D498/12—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and oxygen atoms as the only ring hetero atoms in which the condensed system contains three hetero rings
    • C07D498/18—Bridged systems

Definitions

  • the present invention relates to a series of twelve-membered macrocyclic compounds, in particular to the compounds represented by formula (III) and their pharmaceutically acceptable salts.
  • RAS protein is the product expressed by the RAS gene.
  • RAS protein can bind to guanine trinucleotide phosphate (GTP) or guanine dinucleotide phosphate (GDP), and the active state of RAS protein has effects on cell growth, differentiation, cytoskeleton, protein transport and secretion, etc. , whose activity is regulated by binding to GTP or GDP: when the RAS protein binds to GDP, it is in a dormant state, that is, in an "inactive" state; when stimulated by upstream specific cell growth factors, the RAS protein is induced The exchange of GDP, combined with GTP, is called the "activated" state at this time.
  • the RAS protein bound to GTP can activate downstream proteins for signal transmission.
  • the RAS protein itself has weak hydrolysis GTP hydrolysis activity and can hydrolyze GTP to GDP. In this way, the transition from the activated state to the deactivated state can be achieved.
  • GAP GTPase activating proteins, GTP hydrolase activating proteins
  • RAS protein can interact with RAS protein, greatly promoting its ability to hydrolyze GTP to GDP. Mutation of the RAS protein will affect its interaction with GAP, which also affects its ability to hydrolyze GTP to GDP, making it always active. Activated RAS proteins continue to give downstream proteins growth signals, which eventually lead to continuous cell growth and differentiation, and ultimately produce tumors.
  • KRAS Kirsten rat sarcoma virus oncogene homolog
  • HRAS Harvey rat sarcoma virus oncogene homolog
  • NRAS neuronal Blastoma rat sarcoma virus oncogene homolog
  • KRAS G12C mutation is a relatively common subtype of KRAS gene mutation, which refers to the mutation of glycine No. 12 to cysteine.
  • KRAS G12C mutation is the most common in lung cancer. According to the data reported in the literature (Nat Rev Drug Discov 2014; 13: 828-851), KRAS G12C mutation accounts for about 10% of all lung cancer patients.
  • the present invention provides a compound represented by formula (III) or a pharmaceutically acceptable salt thereof,
  • T 1 , T 2 and T 3 are each independently selected from CH and N;
  • T 4 is selected from CR 6 and N;
  • Ring A is selected from piperazinyl,
  • R 1 is selected from said optionally substituted with 1, 2 or 3 Ra ;
  • each R 2 is independently selected from H and C 1-3 alkyl optionally substituted with 1 , 2 or 3 R ;
  • R 3 is selected from C 1-6 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R4 is selected from H, F, Cl, Br and I;
  • R 5 is selected from H and F
  • R is selected from H, F, Cl and CN ;
  • n 0, 1, 2 and 3;
  • Each R a is independently selected from H, F, Cl, Br, I, CN, C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl,
  • the C 1-3 alkyl and cyclobutenyl groups are optionally substituted with 1, 2 or 3 R;
  • each R b is independently selected from H, F, Cl, Br, I and CN;
  • each R is independently selected from F, OCH3 , N( CH3 ) 2 , NH2 , and morpholinyl;
  • R 1 is selected from when, the Substituted with 1, 2 or 3 R a selected from C 1-3 alkyl, -C ( O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl , the C 1-3 alkyl and cyclobutenyl groups are optionally substituted with 1, 2 or 3 Rs, each R is independently selected from F, OCH 3 , N(CH 3 ) 2 , NH 2 and morpholine base.
  • each R a is independently selected from H, F, Cl, Br, I, CN, CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O) OCH 3 , -C(O)NHCH 3 and cyclobutenyl, the CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O)OCH 3 , -C(O)NHCH 3 and cyclic Butenyl is optionally substituted with 1, 2 or 3 R, other variables are as defined herein.
  • each R a is independently selected from H, F, CN, CHF 2 , CH 2 F, CH 2 OCH 3 , CH 2 N(CH 3 ) 2 , CH 2 NH 2 , -C(O)OCH 3 , -C(O)NHCH 3 , Other variables are as defined in the present invention.
  • the R 1 is selected from Other variables are as defined in the present invention.
  • each R 2 is independently selected from H and CH 3 , and other variables are as defined herein.
  • the R3 is selected from CH( CH3 ) 2 , and other variables are as defined herein.
  • said R4 is selected from F and Cl, and other variables are as defined herein.
  • the present invention provides a compound represented by formula (III) or a pharmaceutically acceptable salt thereof,
  • T 1 , T 2 and T 3 are each independently selected from CH and N;
  • T 4 is selected from CR 6 and N;
  • Ring A is selected from piperazinyl,
  • R 1 is selected from said optionally substituted with 1, 2 or 3 Ra ;
  • each R 2 is independently selected from H and C 1-3 alkyl optionally substituted with 1 , 2 or 3 R ;
  • R 3 is selected from C 1-6 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R4 is selected from H, F, Cl, Br and I;
  • R 5 is selected from H and F
  • R is selected from H, F, Cl and CN ;
  • n 0, 1, 2 and 3;
  • Each R a is independently selected from H, F, Cl, Br, I, CN, C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl,
  • the C 1-3 alkyl and cyclobutenyl groups are optionally substituted with 1, 2 or 3 R;
  • each R b is independently selected from H, F, Cl, Br, I and CN;
  • each R is independently selected from F, OCH3 , N( CH3 ) 2 , NH2 , and morpholinyl;
  • R 1 is selected from when, the Optionally substituted with 1, 2 or 3 R a selected from C 1-3 alkyl, -C ( O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutane alkenyl, the C 1-3 alkyl and cyclobutenyl are optionally substituted with 1, 2 or 3 R, each R independently selected from F, OCH 3 , N(CH 3 ) 2 , NH 2 and Morpholine.
  • each R a is independently selected from H, F, Cl, Br, I, CN, CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O) OCH 3 , -C(O)NHCH 3 and cyclobutenyl, the CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O)OCH 3 , -C(O)NHCH 3 and cyclic Butenyl is optionally substituted with 1, 2 or 3 R, other variables are as defined herein.
  • each R a is independently selected from H, F, CN, CHF 2 , CH 2 F, CH 2 OCH 3 , CH 2 N(CH 3 ) 2 , CH 2 NH 2 , -C(O)OCH 3 , -C(O)NHCH 3 , Other variables are as defined in the present invention.
  • the R 1 is selected from Other variables are as defined in the present invention.
  • each R 2 is independently selected from H and CH 3 , and other variables are as defined herein.
  • the R3 is selected from CH( CH3 ) 2 , and other variables are as defined herein.
  • said R4 is selected from F and Cl, and other variables are as defined herein.
  • the present invention provides a compound represented by formula (III) or a pharmaceutically acceptable salt thereof,
  • T 1 , T 2 and T 3 are each independently selected from CH and N;
  • T 4 is selected from CR 6 and N;
  • Ring A is selected from piperazinyl,
  • R 1 is selected from said optionally substituted with 1, 2 or 3 Ra ;
  • each R 2 is independently selected from H and C 1-3 alkyl optionally substituted with 1 , 2 or 3 R ;
  • R 3 is selected from C 1-6 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R4 is selected from H, F, Cl, Br and I;
  • R 5 is selected from H and F
  • R is selected from H, F, Cl and CN ;
  • n 0, 1, 2 and 3;
  • Each R a is independently selected from H, F, Cl, Br, I, CN, C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl, the C 1-3 alkyl and cyclobutenyl are optionally substituted with 1, 2 or 3 R;
  • each R b is independently selected from H, F, Cl, Br, I and CN;
  • each R is independently selected from F, OCH3 , N( CH3 ) 2 , NH2 , and morpholinyl;
  • R 1 is selected from said optionally substituted with 1, 2 or 3 Ra ;
  • R 1 is selected from said by 1, 2 or 3 C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl, the C 1-3 alkane substituted with 1, 2 or 3 F, OCH 3 , NH 2 and morpholinyl; or
  • each R a is independently selected from H, F, Cl, Br, I, CN, CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O) OCH 3 , -C(O)NHCH 3 and cyclobutenyl, the CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O)OCH 3 , -C(O)NHCH 3 and cyclic Butenyl is optionally substituted with 1, 2 or 3 R, other variables are as defined herein.
  • each R a is independently selected from H, F, CN, CHF 2 , CH 2 F, CH 2 OCH 3 , CH 2 N(CH 3 ) 2 , CH 2 NH 2 , -C(O)OCH 3 , -C(O)NHCH 3 , Other variables are as defined in the present invention.
  • the R 1 is selected from Other variables are as defined in the present invention.
  • each R 2 is independently selected from H and CH 3 , and other variables are as defined herein.
  • the R3 is selected from CH( CH3 ) 2 , and other variables are as defined herein.
  • said R4 is selected from F and Cl, and other variables are as defined herein.
  • the present invention provides a compound represented by formula (III) or a pharmaceutically acceptable salt thereof,
  • T 1 , T 2 and T 3 are each independently selected from CH and N;
  • T 4 is selected from CR 6 and N;
  • Ring A is selected from piperazinyl,
  • R 1 is selected from said optionally substituted with 1, 2 or 3 Ra ;
  • each R 2 is independently selected from H and C 1-3 alkyl optionally substituted with 1 , 2 or 3 R ;
  • R 3 is selected from C 1-6 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R4 is selected from H, F, Cl, Br and I;
  • R 5 is selected from H and F
  • R is selected from H, F, Cl and CN ;
  • n 0, 1, 2 and 3;
  • Each R a is independently selected from H, F, Cl, Br, I, CN, C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl, the C 1-3 alkyl and cyclobutenyl are optionally substituted with 1, 2 or 3 R;
  • each R b is independently selected from H, F, Cl, Br, I and CN;
  • each R is independently selected from F, OCH3 , N( CH3 ) 2 , NH2 , and morpholinyl;
  • T 4 is selected from N
  • ring A is selected from piperazinyl
  • R 1 is selected from when, the optionally substituted with 1, 2 or 3 Ra ;
  • T 4 is selected from N
  • ring A is selected from piperazinyl
  • R 1 is selected from when, the by 1, 2 or 3 C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl, the C 1-3 alkane group is substituted with 1, 2 or 3 F, OCH3 , NH2 and morpholinyl.
  • each R a is independently selected from H, F, Cl, Br, I, CN, CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O) OCH 3 , -C(O)NHCH 3 and cyclobutenyl, the CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O)OCH 3 , -C(O)NHCH 3 and cyclic Butenyl is optionally substituted with 1, 2 or 3 R, other variables are as defined herein.
  • each R a is independently selected from H, F, CN, CHF 2 , CH 2 F, CH 2 OCH 3 , CH 2 N(CH 3 ) 2 , CH 2 NH 2 , -C(O)OCH 3 , -C(O)NHCH 3 , Other variables are as defined in the present invention.
  • the R 1 is selected from Other variables are as defined in the present invention.
  • each R 2 is independently selected from H and CH 3 , and other variables are as defined herein.
  • the R3 is selected from CH( CH3 ) 2 , and other variables are as defined herein.
  • said R4 is selected from F, and other variables are as defined herein.
  • the present invention provides a compound represented by formula (II) or a pharmaceutically acceptable salt thereof,
  • T 1 , T 2 and T 3 are each independently selected from CH and N;
  • Ring A is selected from piperazinyl,
  • R 1 is selected from said optionally substituted with 1, 2 or 3 Ra ;
  • R 2 is selected from H and C 1-3 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R 3 is selected from C 1-6 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R4 is selected from H, F, Cl, Br and I;
  • R 5 is selected from H and F
  • n 0, 1, 2 and 3;
  • Each R a is independently selected from H, F, Cl, Br, I, CN, C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl, the C 1-3 alkyl and cyclobutenyl are optionally substituted with 1, 2 or 3 R;
  • each R b is independently selected from H, F, Cl, Br, I and CN;
  • each R is independently selected from F, OCH3 , N( CH3 ) 2 , NH2 , and morpholinyl;
  • R 1 is selected from when, the optionally substituted with 1, 2 or 3 Ra ;
  • R 1 is selected from when, the By 1 , 2 or 3 C 1-3 alkyl, -C(O)OC 1-3 alkyl, -C(O)NHC 1-3 alkyl and cyclobutenyl, the C 1-3 alkane group is substituted with 1, 2 or 3 F, OCH3 , NH2 and morpholinyl.
  • each R a is independently selected from H, F, Cl, Br, I, CN, CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O) OCH 3 , -C(O)NHCH 3 and cyclobutenyl, the CH 3 , CH 2 CH 3 , CH(CH 3 ) 2 , -C(O)OCH 3 , -C(O)NHCH 3 and cyclic Butenyl is optionally substituted with 1, 2 or 3 R, other variables are as defined herein.
  • each R a is independently selected from H, F, CN, CHF 2 , CH 2 F, CH 2 OCH 3 , CH 2 N(CH 3 ) 2 , CH 2 NH 2 , -C(O)OCH 3 , -C(O)NHCH 3 , Other variables are as defined in the present invention.
  • the R 1 is selected from Other variables are as defined in the present invention.
  • the R2 is selected from H and CH3 , and other variables are as defined herein.
  • the R3 is selected from CH( CH3 ) 2 , and other variables are as defined herein.
  • said R4 is selected from F, and other variables are as defined herein.
  • the present invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,
  • Ring A is selected from piperazinyl and
  • R 1 is selected from said optionally substituted with 1, 2 or 3 Ra ;
  • R 2 is selected from H and C 1-3 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R 3 is selected from C 1-6 alkyl optionally substituted with 1 , 2 or 3 R b ;
  • R4 is selected from H, F, Cl, Br and I;
  • n 0, 1, 2 and 3;
  • Each R a is independently selected from H, F, Cl, Br, I, CN, C 1-3 alkyl, -C(O)OC 1-3 alkyl and -C(O)NHC 1-3 alkyl , the C 1-3 alkyl is optionally substituted by 1, 2 or 3 F;
  • each R b is independently selected from H, F, Cl, Br, I and CN;
  • R 1 is selected from said Optionally substituted with 1, 2 or 3 Ra .
  • each R a is independently selected from H, F, Cl, Br, I, CN, CH 3 , CH 2 CH 3 and CH(CH 3 ) 2 , the CH 3 , CH2CH3 and CH( CH3 ) 2 are optionally substituted with 1, 2 or 3 F, other variables are as defined in the present invention.
  • each R a is independently selected from H, F, CN, CH 2 F, -C(O)OCH 3 and -C(O)NHCH 3 , and other variables are as defined in the present invention definition.
  • the R 1 is selected from Other variables are as defined in the present invention.
  • the R2 is selected from H and CH3 , and other variables are as defined herein.
  • the R3 is selected from CH( CH3 ) 2 , and other variables are as defined herein.
  • said R4 is selected from F, and other variables are as defined herein.
  • the compound, or a pharmaceutically acceptable salt thereof is selected from
  • R 1 , R 2 , R 3 , R 4 , R 5 and m are as defined in the present invention.
  • the present invention also provides the following compounds or pharmaceutically acceptable salts thereof,
  • the compound, or a pharmaceutically acceptable salt thereof is selected from
  • the compound, or a pharmaceutically acceptable salt thereof is selected from
  • the present invention also provides the application of the above-mentioned compounds or their pharmaceutically acceptable salts in the preparation of medicaments for treating tumors.
  • the tumor refers to a tumor associated with a KRAS G12C mutation.
  • the present invention also provides following synthetic method:
  • the present invention also provides the following test methods:
  • DMEM medium fetal bovine serum was purchased from Biosera, and horse serum was purchased from Gibco.
  • CellTiter-Glo Cell Viability Chemiluminescence Detection Reagent
  • MIA-PA-CA-2 cell line was purchased from Nanjing Kebai Biotechnology Co., Ltd. EnVision Multilabel Analyzer (PerkinElmer).
  • MIA-PA-CA-2 cells were seeded in a white 96-well plate, 80 ⁇ L of cell suspension per well, which contained 1000 MIA-PA-CA-2 cells. Cell plates were incubated overnight in a carbon dioxide incubator.
  • the compounds to be tested were diluted 5-fold to the 8th concentration, that is, from 2 mM to 26 nM, and a double-well experiment was set up.
  • the concentration of compounds transferred to the cell plate ranged from 10 [mu]M to 0.13 nM.
  • the cell plates were placed in a carbon dioxide incubator for 3 days. Another cell plate was prepared, and the signal value was read on the day of drug addition as the maximum value (Max value in the following equation) to participate in data analysis.
  • the IC 50 value can be obtained by curve fitting with four parameters (“log(” in GraphPad Prism software). inhibitor)/response--variable scope” mode derived).
  • test compound was mixed with 10% dimethyl sulfoxide/60% polyethylene glycol 400/30% aqueous solution, vortexed and sonicated to prepare a 1 mg/mL clear solution, which was filtered through a microporous membrane for use.
  • Male SD mice aged 7 to 10 weeks were selected, and the candidate compound solution was administered intravenously at a dose of 2-3 mg/kg.
  • Candidate compound solutions were administered orally at a dose of 10 mg/kg.
  • Whole blood was collected for a certain period of time, plasma was prepared, drug concentration was analyzed by LC-MS/MS method, and pharmacokinetic parameters were calculated by Phoenix WinNonlin software (Pharsight, USA).
  • Human pancreatic cancer Mia PaCa-2 cells (ATCC-CRL-1420) were cultured in vitro in monolayer, and the culture conditions were DMEM/F12 medium with 20% fetal bovine serum, 1% double antibody, and incubated at 37°C with 5% carbon dioxide. box cultivation. Conventional digestion with trypsin-EDTA was performed twice a week for passage. When the cell saturation is 80%-90% and the number reaches the requirement, collect the cells, count them, resuspend in an appropriate amount of PBS, add Matrigel 1:1 to obtain a cell suspension with a cell density of 25 x 10 6 cells/mL .
  • Tumor diameters were measured with vernier calipers twice a week.
  • Relative tumor proliferation rate T/C (%) TRTV/CRTV ⁇ 100% (TRTV: RTV of treatment group; CRTV: RTV of negative control group).
  • the mean tumor volume, TRTV and CRTV were taken on the same day.
  • TGI (%) reflecting tumor growth inhibition rate.
  • TGI(%) [(1-(average tumor volume at the end of administration of a certain treatment group-average tumor volume at the beginning of administration of this treatment group))/(average tumor volume at the end of treatment in the solvent control group-the start of treatment in the solvent control group time average tumor volume)] ⁇ 100%.
  • Cell culture Human non-small cell lung cancer NCI-H358 was cultured in monolayer in vitro, and the culture conditions were DMEM/F12 medium with 20% fetal bovine serum, 1% double antibody, and 37°C 5% carbon dioxide incubator. Conventional digestion with trypsin-EDTA was performed twice a week for passage. When the cell saturation is 80%-90% and the number reaches the requirement, the cells are collected, counted, resuspended in an appropriate amount of PBS, and added with Matrigel 1:1 to obtain a cell suspension with a cell density of 25 ⁇ 10 6 cells/mL .
  • Tumor diameters were measured with vernier calipers twice a week.
  • Relative tumor proliferation rate T/C (%) TRTV/CRTV ⁇ 100% (TRTV: RTV of treatment group; CRTV: RTV of negative control group).
  • the mean tumor volume, TRTV and CRTV were taken on the same day.
  • TGI (%) reflecting tumor growth inhibition rate.
  • TGI(%) [(1-(average tumor volume at the end of administration of a certain treatment group-average tumor volume at the beginning of administration of this treatment group))/(average tumor volume at the end of treatment in the solvent control group-the start of treatment in the solvent control group time average tumor volume)] ⁇ 100%.
  • the compound of the present invention has inhibitory effect on KRAS G12C target, and exhibits good inhibitory activity on the proliferation of MIA-PA-CA-2 and NCI-H358 cells.
  • the tumor growth of MiaPaCa-2 xenograft and NCI-H358 xenograft has a good inhibitory effect, and the compounds of the present invention have excellent pharmacokinetic properties.
  • the compounds of the present invention have good plasma stability, whole blood stability and GSH phosphate buffer solution stability.
  • the compounds of the present invention have no PXR positive activation and no CYP-induced activation.
  • the compounds of the present invention have significant antitumor effect without obvious intolerance.
  • the term "pharmaceutically acceptable” refers to those compounds, materials, compositions and/or dosage forms that, within the scope of sound medical judgment, are suitable for use in contact with human and animal tissue , without excessive toxicity, irritation, allergic reactions or other problems or complications, commensurate with a reasonable benefit/risk ratio.
  • salts refers to salts of the compounds of the present invention, prepared from compounds with specific substituents discovered by the present invention and relatively non-toxic acids or bases.
  • base addition salts can be obtained by contacting such compounds with a sufficient amount of base in neat solution or in a suitable inert solvent.
  • acid addition salts can be obtained by contacting such compounds with a sufficient amount of acid in neat solution or in a suitable inert solvent.
  • Certain specific compounds of the present invention contain both basic and acidic functional groups and thus can be converted into either base or acid addition salts.
  • the pharmaceutically acceptable salts of the present invention can be synthesized from the acid or base containing parent compound by conventional chemical methods. Generally, such salts are prepared by reacting the free acid or base form of these compounds with a stoichiometric amount of the appropriate base or acid in water or an organic solvent or a mixture of the two.
  • the compounds of the present invention may exist in specific geometric or stereoisomeric forms.
  • the present invention contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers isomers, (D)-isomers, (L)-isomers, and racemic mixtures thereof and other mixtures, such as enantiomerically or diastereomerically enriched mixtures, all of which belong to this within the scope of the invention.
  • Additional asymmetric carbon atoms may be present in substituents such as alkyl. All such isomers, as well as mixtures thereof, are included within the scope of the present invention.
  • the compounds of the present invention may contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute the compound.
  • compounds can be labeled with radioisotopes, such as tritium ( 3 H), iodine-125 ( 125 I) or C-14 ( 14 C).
  • deuterated drugs can be formed by replacing hydrogen with deuterium, and the bonds formed by deuterium and carbon are stronger than those formed by ordinary hydrogen and carbon. Compared with non-deuterated drugs, deuterated drugs can reduce toxic side effects and increase drug stability. , enhance the efficacy, prolong the biological half-life of drugs and other advantages. All transformations of the isotopic composition of the compounds of the present invention, whether radioactive or not, are included within the scope of the present invention.
  • substituted means that any one or more hydrogen atoms on a specified atom are replaced by a substituent, which may include deuterium and hydrogen variants, as long as the valence of the specified atom is normal and the substituted compound is stable.
  • oxygen it means that two hydrogen atoms are substituted. Oxygen substitution does not occur on aromatic groups.
  • any variable eg, R
  • its definition in each case is independent.
  • the group may optionally be substituted with up to two Rs, with independent options for R in each case.
  • combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds.
  • linking group When the number of a linking group is 0, such as -(CRR) n -, it means that the linking group is a single bond.
  • the direction of attachment is arbitrary, for example,
  • the linking group L in the middle is -MW-, at this time -MW- can connect ring A and ring B in the same direction as the reading order from left to right. It is also possible to connect ring A and ring B in the opposite direction to the reading order from left to right.
  • Combinations of the linking groups, substituents and/or variants thereof are permissible only if such combinations result in stable compounds.
  • any one or more sites in the group can be linked to other groups by chemical bonds.
  • connection method of the chemical bond is not located, and there is an H atom at the linkable site, when the chemical bond is connected, the number of H atoms at the site will be correspondingly reduced with the number of chemical bonds connected to the corresponding valence. the group.
  • the chemical bond connecting the site to other groups can be represented by straight solid line bonds straight dotted key or wavy lines express.
  • a straight solid bond in -OCH3 indicates that it is connected to other groups through the oxygen atom in this group;
  • the straight dashed bond in the group indicates that it is connected to other groups through the two ends of the nitrogen atom in the group;
  • the wavy line in the phenyl group indicates that it is connected to other groups through the 1 and 2 carbon atoms in the phenyl group;
  • C 1-6 alkyl is used to denote a straight or branched chain saturated hydrocarbon group consisting of 1 to 6 carbon atoms.
  • the C 1-6 alkyl includes C 1-5 , C 1-4 , C 1-3 , C 1-2 , C 2-6 , C 2-4 , C 6 and C 5 alkyl and the like; it can be Is monovalent (eg methyl), divalent (eg methylene) or polyvalent (eg methine).
  • C 1-6 alkyl examples include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), butyl (including n-butyl, isobutyl , s-butyl and t-butyl), pentyl (including n-pentyl, isopentyl and neopentyl), hexyl, etc.
  • C 1-3 alkyl is used to denote a straight or branched chain saturated hydrocarbon group consisting of 1 to 3 carbon atoms.
  • the C 1-3 alkyl group includes C 1-2 and C 2-3 alkyl groups, etc.; it can be monovalent (eg methyl), divalent (eg methylene) or multivalent (eg methine) .
  • Examples of C1-3 alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.
  • Cn-n+m or Cn - Cn+m includes any particular instance of n to n+ m carbons, eg C1-12 includes C1 , C2 , C3, C 4 , C 5 , C 6 , C 7 , C 8 , C 9 , C 10 , C 11 , and C 12 , also including any range from n to n+ m , eg C 1-12 includes C 1-3 , C 1-6 , C 1-9 , C 3-6 , C 3-9 , C 3-12 , C 6-9 , C 6-12 , and C 9-12 , etc.; in the same way, n yuan to n +m-membered means that the number of atoms in the ring is from n to n+m, for example, 3-12-membered ring includes 3-membered ring, 4-membered ring, 5-membered ring, 6-membered ring, 7-membered ring, 8-membere
  • the compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments enumerated below, embodiments formed in combination with other chemical synthesis methods, and those well known to those skilled in the art Equivalent to alternatives, preferred embodiments include, but are not limited to, the embodiments of the present invention.
  • the compounds of the present invention exist in stereo configuration, which can be determined by single crystal diffraction, optical rotation, CD and other test methods.
  • the structure of the compound of the present invention can be confirmed by conventional methods well known to those skilled in the art. If the present invention relates to the absolute configuration of the compound, the absolute configuration can be confirmed by conventional technical means in the art. For example, single crystal X-ray diffraction method (SXRD), the cultured single crystal is collected by Bruker D8 venture diffractometer, the light source is CuK ⁇ radiation, and the scanning mode is: After scanning and collecting relevant data, the crystal structure was further analyzed by the direct method (Shelxs97), and the absolute configuration could be confirmed.
  • SXRD single crystal X-ray diffraction method
  • the cultured single crystal is collected by Bruker D8 venture diffractometer
  • the light source is CuK ⁇ radiation
  • the scanning mode is: After scanning and collecting relevant data, the crystal structure was further analyzed by the direct method (Shelxs97), and the absolute configuration could be confirmed.
  • FIG. 1 Tumor volume diagram of the Balb/c Nude mouse model of human pancreatic cancer Mia PaCa-2 cells in nude mice subcutaneously transplanted;
  • FIG. 1 Body weight changes in the Balb/c Nude mouse model study of human pancreatic cancer Mia PaCa-2 cells in nude mice subcutaneously transplanted with tumors.
  • the low-energy conformation of AMG510 was calculated by the Macromodel module of Schrodinger's Maestro software.
  • the binding mode of AMG510 and KRAS G12C protein is shown in Figure 1
  • the 2D map of the binding mode of AMG510 and KARS G12C is shown in Figure 2
  • the low-energy conformation of compounds A to E and the active conformation of AMG510 are superimposed as shown in Figures 3 to 8 Show.
  • the reaction system was cooled to room temperature, diluted with 1.5 liters of water and 2.0 liters of ethyl acetate, the organic phase was collected after separation, and the aqueous phase was extracted with ethyl acetate (1.0 liters, twice). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a residue.
  • the reactions were quenched by adding saturated ammonium chloride (1.0 L) to the respective reaction systems.
  • the two systems were combined for extraction.
  • 1.0 L of water and 1.0 L of ethyl acetate were added to the system for dilution, and the organic phase was collected after standing for liquid separation.
  • the aqueous phase was extracted with ethyl acetate (600 mL, 1 time).
  • the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a residue.
  • Step 8 Synthesis of Compounds 001-12
  • compound 001-11 (91.85g, 181.14mmol, 1.00eq) and diisopropylethylamine (37.46g, 289.82mmol, 50.48mL, 1.6eq) were added to a pre-dried 3.0-liter three-necked flask. and tetrahydrofuran (920 mL). After the addition, the temperature of the system was lowered to 0-5° C., and phosphorus oxychloride (41.66 g, 271.71 mmol, 25.25 mL, 1.5 eq) was slowly added to the system. After the addition, the system was slowly heated to 40° C. (system temperature), and stirred at this temperature for 2.0 hours to obtain compound 001-12. The reaction solution was used directly in the next step
  • the reaction system containing compound 001-12 (95.19g, 181.14mmol, 1.00eq) was cooled to 0-10°C and maintained, and diisopropylethylamine (58.52g, 452.84mmol, 78.87mL, 2.5eq) and pre-dissolved solution of compound 001-13 (72.55g, 362.27mmol, 2.0eq) in tetrahydrofuran (210mL). After the addition was complete, the system was stirred at 25°C for 12.0 hours. Combine 2 parallel reactions. The temperature of the system was controlled at 20 to 25°C, and 1.0 liter of saturated sodium chloride was added to the system to quench the reaction. The system is left to stand for liquid separation.
  • the aqueous phase was extracted with ethyl acetate (0.5 L, 2 times).
  • the organic phases were combined, washed with brine (0.5L, once), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to give the crude intermediate compound, which was used in the next step without purification.
  • reaction solution was cooled to room temperature, filtered, and the filter cake was washed with 50 mL of 2-methyltetrahydrofuran.
  • the white filter cake was collected and concentrated under reduced pressure to remove the solvent.
  • the white solid was dried in a vacuum oven at 45°C for 20 hours to obtain compound 001-16.
  • Step 12 Synthesis of Compounds 001-18
  • compound 001-17 (9.1 g, 14.38 mmol, 1 eq) was dissolved in anhydrous dichloromethane (45 mL), trifluoroacetic acid (15 mL) was added, and the mixture was stirred for 5 hours.
  • the reaction solution was concentrated under reduced pressure, and the crude product was slurried with methyl tert-butyl ether ((100 mL*4), filtered, and the solid was collected.
  • the solid was dissolved in 100 mL of dichloromethane, 100 mL of saturated aqueous sodium bicarbonate solution was added, and the liquid was separated. Extraction with chloromethane ((40 mL*2). The organic phases were combined, washed with saturated brine ((40 mL)), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain compound 001-18.
  • compound 002-1 (0.8g, 1.78mmol, 1eq) was dissolved in tetrahydrofuran (12mL), N,N-diisopropylethylamine (688.61mg, 5.33mmol, 928.05 ⁇ L, 3eq) was added, then added Phosphorus oxychloride (680.81 mg, 4.44 mmol, 412.61 ⁇ L, 2.5 eq) was reacted at 40° C. for 3 hours. Compound 002-2 was obtained, and the reaction system was directly used in the next step.
  • compound 002-2 (0.8 g, 1.71 mmol, 1 eq) was dissolved in tetrahydrofuran (13 mL), and N,N-diisopropylethylamine (1.10 g, 8.53 mmol, 1.49 mL, 5 eq) was added. ), then compound 003-1 (596.07 mg, 2.22 mmol, 1.3 eq, HCl) was added, and the temperature was raised to 20° C. to react for 15 hours.
  • N,N-diisopropylethylamine (1.10 g, 8.53 mmol, 1.49 mL, 5 eq) was added, and the reaction was carried out at 20° C. for 15 hours. Since the reaction was not complete, the reaction was carried out at 50°C for 15 hours. N,N-diisopropylethylamine (1.10 g, 5 eq) was added, and the reaction was carried out at 70° C. for 30 hours. The reaction system was added to ice water (50 mL), the aqueous phase was extracted with ethyl acetate (50 mL*3), and the layers were separated.
  • compound 003-2 (0.6g, 902.66 ⁇ mol, 1eq) was dissolved in tetrahydrofuran (30mL), palladium/carbon (0.5g, 902.66 ⁇ mol, 10% content, 1.00eq) was added, hydrogen was replaced three times, and then hydrogen atmosphere (15Psi) at 25°C for 8 hours.
  • the reaction system was directly filtered, and the filtrate was concentrated under reduced pressure to obtain compound 003-3, which could be directly used in the next reaction without purification.
  • Step 4 Synthesis of Compounds 003 and 004
  • compound 003-3 (0.1g, 188.48 ⁇ mol, 1eq) was dissolved in dichloromethane (5mL), N,N-diisopropylethylamine (121.79mg, 942.39 ⁇ mol, 164.14 ⁇ L, 5eq) was added, Then, acryloyl chloride (34.12 mg, 376.96 ⁇ mol, 30.74 ⁇ L, 2 eq) was added, and the reaction was carried out at -60° C. for 10 minutes. Saturated sodium bicarbonate solution (20 mL) was added to the reaction system, and the layers were separated. The aqueous phase was extracted with dichloromethane (50 mL*3), and the layers were separated.
  • reaction solution was concentrated to dryness under reduced pressure, acetonitrile (2 mL) was added to dissolve, and the crude product was subjected to high performance liquid preparative chromatography (chromatographic column: Waters Xbridge BEH C18 100*25mm*5 ⁇ m; mobile phase: [water (ammonium bicarbonate)-acetonitrile] ; acetonitrile%: 25%-55%, 10min) separation and purification to obtain compound 005.
  • chromatographic column Waters Xbridge BEH C18 100*25mm*5 ⁇ m; mobile phase: [water (ammonium bicarbonate)-acetonitrile] ; acetonitrile%: 25%-55%, 10min
  • reaction solution was concentrated to dryness under reduced pressure, dissolved in acetonitrile (2 mL), filtered, and the crude product was subjected to high performance liquid preparative chromatography (chromatographic column: Waters Xbridge BEH C18 100*25mm*5 ⁇ m; mobile phase: [water (ammonium bicarbonate)- Acetonitrile]; acetonitrile %: 30%-60%, 10 min) was separated and purified to obtain compound 006.
  • chromatographic column Waters Xbridge BEH C18 100*25mm*5 ⁇ m; mobile phase: [water (ammonium bicarbonate)- Acetonitrile]; acetonitrile %: 30%-60%, 10 min
  • Step 1 Synthesis of Compound 007-1
  • cyanoacetic acid (3.64g, 42.78mmol, 6eq) was dissolved in dichloromethane (35mL), oxalyl chloride (8.15g, 64.17mmol, 5.62mL, 9eq) was added, followed by N,N-dimethyl Formamide (104.23 mg, 1.43 mmol, 109.72 ⁇ L, 0.2 eq) was reacted at 25° C. for 3 hours. After the completion of the reaction, the reaction system was directly concentrated under reduced pressure to obtain an intermediate state of the acid chloride.
  • compound 001-8 (2.2 g, 7.13 mmol, 1 eq) was dissolved in dichloromethane (40 mL), triethylamine (2.89 g, 28.52 mmol, 3.97 mL, 4 eq) was added, and acid chloride was added slowly at 0 °C as an intermediate state , and then the temperature was slowly raised to 25 °C for 2 hours. Water (50 mL) was added to the reaction system, and the layers were separated. The aqueous phase was extracted with dichloromethane (30 mL*2), and the layers were separated.
  • compound 007-1 (2.1 g, 5.59 mmol, 1 eq) was dissolved in tetrahydrofuran (30 mL), tetramethylammonium fluoride (10.42 g, 111.83 mmol, 20 eq) was added, and the reaction was carried out at 20 °C for 23 hours.
  • Water (50 mL) was added to the reaction system, the aqueous phase was extracted with ethyl acetate (100 mL*3), and the layers were separated. The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product compound 007-2, which was directly used in the next reaction without purification.
  • compound 007-2 (1.5g, 5.74mmol, 1eq) was dissolved in dichloromethane (45mL), imidazole (1.17g, 17.22mmol, 3eq) and tert-butyldiphenylchlorosilane (2.37g) were added at 0°C g, 8.61 mmol, 2.21 mL, 1.5 eq), react at 20°C for 6 hours. Water (100 mL) was added to the reaction system, the aqueous phase was extracted with dichloromethane (100 mL*3), and the layers were separated.
  • compound 007-3 (1.76g, 3.52mmol, 1eq) was dissolved in tetrahydrofuran (30mL), sodium tert-butoxide (676.93mg, 7.04mmol, 2eq) was added at 0°C, stirred at 0°C for 0.5 hours, then 0°C The acid chloride intermediate tetrahydrofuran (24 mL) solution was slowly added, and then the temperature was slowly raised to 25° C. to react for 3 hours. Water (50 mL) was added to the reaction system, the aqueous phase was extracted with ethyl acetate (100 mL*2), and the layers were separated.
  • compound 007-5 (0.91g, 1.32mmol, 1eq) was dissolved in tetrahydrofuran (35mL), sodium hydride (263.10mg, 6.58mmol, 60% content, 5eq) was slowly added at 0°C, and then reacted at 50°C for 4 hours .
  • Saturated ammonium chloride solution 50 mL was added to the reaction system to quench the reaction, the aqueous phase was extracted with ethyl acetate (3*100 mL), and the layers were separated. The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product.
  • compound 007-6 (1.17g, 1.79mmol, 1eq) was dissolved in tetrahydrofuran (18mL), N,N-diisopropylethylamine (692.32mg, 5.36mmol, 933.05 ⁇ L, 3eq) was added, then added Phosphorus oxychloride (684.48 mg, 4.46 mmol, 414.83 ⁇ L, 2.5 eq) was reacted at 40° C. for 3 hours to obtain the crude product compound 007-7, and the reaction system could be directly used in the next reaction.
  • N,N-diisopropylethylamine (692.32mg, 5.36mmol, 933.05 ⁇ L, 3eq) was added, then added Phosphorus oxychloride (684.48 mg, 4.46 mmol, 414.83 ⁇ L, 2.5 eq) was reacted at 40° C. for 3 hours to obtain the crude product compound 007-7, and the reaction system
  • compound 007-7 (1.1 g, 1.63 mmol, 1 eq) was dissolved in tetrahydrofuran (17.2 mL), N,N-diisopropylethylamine (844.11 mg, 6.53 mmol, 1.14 mL) was added, 4eq), then compound 001-13 (425.12 mg, 2.12 mmol, 1.3 eq) was added, and the temperature was raised to 20° C. to react for 16 hours. The reaction system was added to water (50 mL), the aqueous phase was extracted with ethyl acetate (50 mL*3), and the layers were separated.
  • Step 8 Synthesis of Compound 007-9
  • compound 007-8 (0.69 g, 823.89 ⁇ mol, 1 eq) and compound 001-15 (256.92 mg, 1.65 mmol, 2 eq) were dissolved in 1,4-dioxane (14 mL) and water (1.4 mL) , added sodium carbonate (261.97mg, 2.47mmol, 3eq) and tetrakis(triphenylphosphine)palladium (95.21mg, 82.39 ⁇ mol, 0.1eq), and reacted at 80°C for 1 hour.
  • compound 007-10 (0.34g, 503.90 ⁇ mol, 1eq) was dissolved in tetrahydrofuran (21mL), triphenylphosphine (462.59mg, 1.76mmol, 3.5eq) was added, and diethyl azodicarboxylate was added at 20°C (307.16 mg, 1.76 mmol, 320.62 ⁇ L, 3.5 eq), and reacted at 20° C. for 2 hours. Water (20 mL) was added to the reaction system, the aqueous phase was extracted with ethyl acetate (30 mL*3), and the layers were separated.
  • Step 11 Synthesis of the trifluoroacetate salt of compound 007-12
  • Step 12 Synthesis of Compounds 007 and 008
  • the trifluoroacetate salt of compound 007-12 (0.5 g, 351.33 ⁇ mol, 1 eq) was dissolved in dichloromethane (12 mL), and N,N-diisopropylethylamine (681.08 mg, 5.27 mmol) was added. 917.90 ⁇ L, 15eq), then acryloyl chloride (63.60mg, 702.65 ⁇ mol, 57.29 ⁇ L, 2eq) was added, and the reaction was performed at -60°C for 10 minutes. Saturated sodium bicarbonate solution (20 mL) was added to the reaction system, and the layers were separated.
  • the aqueous phase was extracted with dichloromethane (50 mL*3), and the layers were separated. The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product.
  • the reaction solution was concentrated under reduced pressure, methyl tert-butyl ether (200 mL) was added, and the mixture was filtered, and the filter cake was thoroughly rinsed with methyl tert-butyl ether until no product remained.
  • the filtrate was extracted with methyl tert-butyl ether (200 mL*3), the organic phases were collected and combined into saturated aqueous sodium chloride solution (40 mL), the organic phase was separated and dried by adding anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure.
  • Step 8 Synthesis of Compound 009-9
  • Step 12 Synthesis of Compound 009-13
  • Step 14 Synthesis of Compounds 009 and 010
  • compound 007-6 (1.8g, 2.75mmol, 1eq) was dissolved in tetrahydrofuran (27mL), N,N-diisopropylethylamine (1.07g, 8.24mmol, 1.44mL, 3eq) was added, then added Phosphorus oxychloride (1.05 g, 6.87 mmol, 638.21 ⁇ L, 2.5 eq) was reacted at 40° C. for 3 hours. Compound 011-1 was obtained, and the reaction system was directly used in the next step.
  • compound 011-1 (1.8g, 2.67mmol, 1eq) was dissolved in tetrahydrofuran (26mL) at 0°C, and N,N-diisopropylethylamine (1.38g, 10.68mmol, 1.86mL, 4eq) was added. ), then compound 011-2 (646.94 mg, 3.47 mmol, 1.3 eq) was added, and the temperature was raised to 20° C. to react for 1 hour. Compound 011-2 (248.64 mg, 1.34 mmol, 0.5 eq) was added, and the reaction was carried out at 20° C. for 1 hour.
  • compound 011-3 (1.92g, 2.33mmol, 1eq) and compound 001-15 (727.08mg, 4.66mmol, 2eq) were dissolved in 1,4-dioxane (40mL) and water (4mL), Sodium carbonate (741.38 mg, 6.99 mmol, 3 eq) and tetrakis(triphenylphosphine) palladium (269.43 mg, 233.16 ⁇ mol, 0.1 eq) were added, and the mixture was reacted at 80° C. for 1 hour. Water (50 mL) was added to the reaction system, the aqueous phase was extracted with ethyl acetate (60 mL*3), and the layers were separated.
  • compound 011-4 (1.6g, 1.78mmol, 1eq) was dissolved in tetrahydrofuran (48mL), tetramethylammonium fluoride (5.80g, 62.28mmol, 35eq) was added, and the reaction was carried out at 20°C for 23 hours.
  • Water (40 mL) was added to the reaction system, the aqueous phase was extracted with ethyl acetate (50 mL*3), and the layers were separated. The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain compound 011-5, which was directly used in the next reaction without purification.
  • compound 011-5 (1.6g, 2.42mmol, 1eq) was dissolved in tetrahydrofuran (96mL), triphenylphosphine (2.22g, 8.48mmol, 3.5eq) was added, and diethyl azodicarboxylate was added at 20°C (1.48g, 8.48mmol, 1.54mL, 3.5eq), react at 20°C for 1 hour. Water (20 mL) was added to the reaction system, the aqueous phase was extracted with ethyl acetate (30 mL*3), and the layers were separated.
  • compound 011-6 (0.5 g, 777.98 ⁇ mol, 1 eq) was dissolved in dichloromethane (15 mL), trifluoroacetic acid (3.55 g, 31.12 mmol, 2.30 mL, 40 eq) was added, and the reaction was carried out at 20° C. for 1 hour.
  • the reaction system is directly concentrated under reduced pressure to obtain compound 011-7, which can be directly used in the next reaction without purification.
  • compound 011-7 (0.96g, 692.47 ⁇ mol, 1eq) was dissolved in dichloromethane (24mL), N,N-diisopropylethylamine (1.34g, 10.39mmol, 1.81mL, 15eq) was added, Then, acryloyl chloride (125.35 mg, 1.38 mmol, 112.93 ⁇ L, 2 eq) was added, and the reaction was carried out at -60° C. for 10 minutes. Saturated sodium bicarbonate solution (20 mL) was added to the reaction system, and the layers were separated. The aqueous phase was extracted with dichloromethane (50 mL*3), and the layers were separated.
  • DMEM medium fetal bovine serum was purchased from Biosera, and horse serum was purchased from Gibco.
  • CellTiter-Glo Cell Viability Chemiluminescence Detection Reagent
  • MIA-PA-CA-2 cell line was purchased from Nanjing Kebai Biotechnology Co., Ltd. EnVision Multilabel Analyzer (PerkinElmer).
  • MIA-PA-CA-2 cells were seeded in a white 96-well plate, 80 ⁇ L of cell suspension per well, which contained 1000 MIA-PA-CA-2 cells. Cell plates were incubated overnight in a carbon dioxide incubator.
  • the compounds to be tested were diluted 5-fold to the 8th concentration, that is, from 2 mM to 26 nM, and a double-well experiment was set up.
  • the concentration of compounds transferred to the cell plate ranged from 10 [mu]M to 0.13 nM.
  • the cell plates were placed in a carbon dioxide incubator for 3 days. Another cell plate was prepared, and the signal value was read on the day of drug addition as the maximum value (Max value in the following equation) to participate in data analysis.
  • the IC 50 value can be obtained by curve fitting with four parameters (“log(” in GraphPad Prism software). inhibitor)/response--variable scope” mode derived).
  • Table 1 provides the inhibitory activity of the compounds of the present invention on the proliferation of MIA-PA-CA-2 cells.
  • the compounds of the present invention show good inhibitory activity on the proliferation of MIA-PA-CA-2 cells.
  • Test Example 2 In vivo pharmacokinetic study
  • test compound was mixed with 10% dimethyl sulfoxide/60% polyethylene glycol 400/30% aqueous solution, vortexed and sonicated to prepare a 1 mg/mL clear solution, which was filtered through a microporous membrane for use.
  • Male SD mice aged 7 to 10 weeks were selected, and the candidate compound solution was administered intravenously at a dose of about 2 mg/kg.
  • Candidate compound solutions are administered orally at a dose of about 10 mg/kg.
  • Whole blood was collected for a certain period of time, plasma was prepared, drug concentration was analyzed by LC-MS/MS method, and pharmacokinetic parameters were calculated by Phoenix WinNonlin software (Pharsight, USA).
  • the compound of the present invention has excellent total systemic exposure, peak concentration and bioavailability after oral administration, and exhibits excellent pharmacokinetic properties.
  • the compounds of the present invention have good stability in CD-1 mice, SD rats, beagle dogs, cynomolgus monkeys and human plasma.
  • Experimental operation Prepare 96-well incubation plates, named T0, T60, T120, T240, and NGSH, respectively. Add 1500 ⁇ L of GSH-potassium phosphate buffer solution (final test GSH concentration is 5 ⁇ M) to the corresponding incubation plate, and then add the working solution of the test compound or control compound to the corresponding incubation plate, and prepare two parallels for each sample. hole. All samples were incubated in a 37°C water bath. The final incubation concentrations of the test compound and the control compounds afatinib and ibrutinib were 1 ⁇ M and 10 ⁇ M, respectively.
  • Test Example 6 Bidirectional permeability and efflux rate studies
  • OBJECTIVE The bidirectional permeability and efflux rate of the test compounds were determined using the MDR1-MDCKII monolayer cell model.
  • MDR1-MDCK II cells (passage 11) were seeded into 96-well cell culture plates, and were used for transport experiments after 4-7 days of continuous culture.
  • Test compounds were diluted from stock solutions to 2 ⁇ M concentrations (DMSO ⁇ 1%) in transport buffer (HBSS with 10 mM Hepes, pH 7.4) and applied to the apical or basolateral side of the cell monolayer. Penetration of test compounds from A to B direction or B to A direction was repeated with and without P-gp inhibitor (GF120918, 10 ⁇ M), respectively.
  • Digoxin was tested bidirectionally at 10 ⁇ M in the presence or absence of 10 ⁇ M GF120918, while nadolol and metoprolol were tested bidirectionally at 2 ⁇ M in the absence of GF120918. Plates were incubated for 2.5 hours without shaking in a carbon dioxide incubator at 37 ⁇ 1°C with a saturated humidity of 5% carbon dioxide. In addition, the jet ratio of each compound was also measured. According to the peak area ratio of analyte/IS, LC-MS/MS analysis method was used to quantify the test articles and reference compounds. The experimental results are shown in Table 6.
  • PXR receptor and live cell multiplex assay (activity): Prepare live cell multiplex assay reagent, add 6.7 ⁇ L 300x live cell multiplex assay substrate to 2mL live cell multiplex assay buffer, and store at room temperature. Remove the cells from the incubator, aspirate the dosing solution, and rinse once with 200 ⁇ L of viable cell multiplex assay buffer at room temperature. Add 50 ⁇ L of viable cell multiplex detection reagent to each well, tilt left and right 2-3 times, and incubate the plate for 15 minutes at room temperature in the dark.
  • Luciferase Assay Reagent Add 2 mL of Assay Substrate to 2 mL of Assay Buffer. After the 15-minute incubation, aspirate the live cell multiplex detection reagent, add 100 ⁇ L of luciferase detection reagent to each well, and incubate for at least 5 minutes. Fluorescence values were read in a SpectraMax M4 plate reader (fluorescence relative units at 485nm/535nm, activity). After adding the luciferase detection reagent and incubating for at least 5 minutes, the luminescence (relative units of fluorescence, PXR activation) was read with a SpectraMax M4 plate reader reading time of 500ms/well.
  • the culture system After the culture system is established, discard the upper medium of the sandwich medium, and add 200 ⁇ L of pre-warmed to 37°C and freshly prepared dosing working solution (including test sample, positive control and matrix) into each cell culture well. control), the cell culture plate was placed in an incubator for 24 hours. After 24 hours of incubation, the freshly prepared dosing solution was replaced and the incubation was continued for 24 hours. The entire incubation time was 48 hours. Three parallels were made for each drug concentration and control concentration.
  • the cell wells were washed twice with 0.5 mL of HBSS solution preheated to 37 °C, and 100 ⁇ L preheated to 37 °C was added to each well. Incubate for 30 minutes with the enzymatically labeled substrate working solution. After 30 minutes of incubation, 75.0 ⁇ L of the supernatant sample per well was added to a 96-well deep-well plate containing 150 ⁇ L of stop solution.
  • the plate was shaken for 10 minutes, centrifuged at 4°C and 3220g for 20 minutes, and then the supernatant solution was taken and diluted with an aqueous solution containing 0.1% formic acid at a ratio of 1:4. After the diluted samples were shaken for 10 minutes, the generation of metabolites was detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS).
  • LC-MS/MS liquid chromatography-tandem mass spectrometry
  • the potential toxicity of the test article was assessed by the amount of lactate dehydrogenase (LDH) released from hepatocytes. 100 ⁇ L of the administration working solution after incubation with hepatocytes for 24 hours and 48 hours was taken out, and the concentration of lactate dehydrogenase was detected with a commercial LDH kit. The cell lysis solution was used as a positive control for the experiment, and the incubation medium was used as a blank control.
  • LDH lactate dehydrogenase
  • RNA samples were randomly selected from different positions of the sample plate, and the OD values at 260 nm and 280 nm were measured by ND2000 microspectrophotometer, and the purity of total RNA was judged by calculating the ratio of the two.
  • Reverse transcription to obtain cDNA The selected genes were quantitatively analyzed in real time with a real-time PCR instrument. The reaction conditions were set as follows: 50°C for two minutes; 95°C for ten minutes; 40 cycles of the following two steps: 95°C for fifteen seconds, and 60°C for one minute.
  • the endogenous control 18S rRNA was used as an internal standard.
  • the experimental data show the production of enzymatic metabolites of CYP1A2, CYP2B6 and CYP3A4. Changes in enzymatic activity are shown by comparing the fold induction of the corresponding cytochrome enzymes in the presence or absence of the compound.
  • the calculation method of the induction fold and the calculation method of the induction ratio with the control compound are as follows:
  • Induction fold enzymatic activity in samples treated with test article (or control compound)/enzyme activity in samples treated with matrix control
  • Induction ratio to control compound (%) (induction fold of samples treated with test substance-1)/(induction fold of samples treated with control compound-1) ⁇ 100
  • This project uses the ⁇ Ct relative quantification method to compare the differences in gene expression between different treatment groups, and uses 18S rRNA as the internal reference gene to correct the gene expression level of each sample.
  • the 2- ⁇ Ct method was used for statistical analysis to compare the fold changes between the treatment group and the matrix control group.
  • the calculation method of the induction ratio of the test article and the control compound is as follows:
  • Induction ratio to control compound (%) (induction fold of samples treated with test substance-1)/(induction fold of samples treated with control compound-1) ⁇ 100
  • Test Example 9 In vivo pharmacodynamic study of human pancreatic cancer Mia PaCa-2 cells in nude mice subcutaneously transplanted tumor Balb/c Nude mouse model Cell culture and tumor tissue preparation
  • Cell culture Human pancreatic cancer Mia PaCa-2 cells (ATCC-CRL-1420) were cultured in vitro in monolayer, and the culture conditions were DMEM/F12 medium with 20% fetal bovine serum, 1% double antibody, and incubated at 37°C with 5% carbon dioxide. box cultivation. Conventional digestion with trypsin-EDTA was performed twice a week for passage. When the cell saturation is 80%-90% and the number reaches the requirement, the cells are collected, counted, resuspended in an appropriate amount of PBS, and added with Matrigel 1:1 to obtain a cell suspension with a cell density of 25 ⁇ 10 6 cells/mL .
  • MiaPaCa-2 cells were subcutaneously inoculated into the right back of each mouse, when the average tumor volume reached 190mm 3 , randomization was performed according to tumor volume, and dosing was started according to the schedule in Table 9.
  • PO oral administration
  • QD once a day
  • Tumor diameters were measured with vernier calipers twice a week.
  • Relative tumor proliferation rate T/C (%) TRTV/CRTV ⁇ 100% (TRTV: RTV of treatment group; CRTV: RTV of negative control group).
  • the mean tumor volume, TRTV and CRTV were taken on the same day.
  • TGI (%) reflecting tumor growth inhibition rate.
  • TGI(%) [(1-(average tumor volume at the end of administration of a certain treatment group-average tumor volume at the beginning of administration of this treatment group))/(average tumor volume at the end of treatment in the solvent control group-the start of treatment in the solvent control group time average tumor volume)] ⁇ 100%.
  • the mean tumor volumes of the analytes compound 009 (1.5 mg/kg) and compound 009 (5 mg/kg) were 1325 mm 3 and 506 mm 3 , respectively; T/C were 56.99% and 21.89%, respectively; TGI was 44.37% and 82.59%, respectively , significantly inhibited tumor growth at both concentrations (both p values were less than 0.001).
  • the mean tumor volume of the test substance AMG510 (5mg/kg) was 963 mm 3 , the T/C was 41.78%, the TGI was 61.28%, and the p values were all ⁇ 0.001. It also had a significant tumor inhibitory effect, and the mice in each dose group had a significant tumor-inhibiting effect.
  • the body weight was stable, and there was no obvious intolerance.

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Abstract

L'invention concerne une série de composés macrocycliques à douze éléments, et spécifiquement un composé tel que représenté par la formule (III) et un sel pharmaceutiquement acceptable de celui-ci.
PCT/CN2022/091154 2021-05-06 2022-05-06 Composé macrocyclique à douze éléments WO2022233316A1 (fr)

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WO2020259513A1 (fr) * 2019-06-24 2020-12-30 Guangdong Newopp Biopharmaceuticals Co., Ltd. Composés hétérocycliques utilisés en tant qu'inhibiteurs de kras g12c
US20210122764A1 (en) * 2019-10-28 2021-04-29 Merck Sharp & Dohme Corp. Small Molecule Inhibitors of KRAS G12C Mutant
WO2021147965A1 (fr) * 2020-01-21 2021-07-29 南京明德新药研发有限公司 Composé macrocyclique servant d'inhibiteur de kras
CN113248521A (zh) * 2020-02-11 2021-08-13 上海和誉生物医药科技有限公司 一种k-ras g12c抑制剂及其制备方法和应用

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WO2020259513A1 (fr) * 2019-06-24 2020-12-30 Guangdong Newopp Biopharmaceuticals Co., Ltd. Composés hétérocycliques utilisés en tant qu'inhibiteurs de kras g12c
US20210122764A1 (en) * 2019-10-28 2021-04-29 Merck Sharp & Dohme Corp. Small Molecule Inhibitors of KRAS G12C Mutant
WO2021147965A1 (fr) * 2020-01-21 2021-07-29 南京明德新药研发有限公司 Composé macrocyclique servant d'inhibiteur de kras
CN113248521A (zh) * 2020-02-11 2021-08-13 上海和誉生物医药科技有限公司 一种k-ras g12c抑制剂及其制备方法和应用

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