WO2023226959A1 - 烷化剂前药与细胞周期抑制剂联用治疗癌症的方法 - Google Patents

烷化剂前药与细胞周期抑制剂联用治疗癌症的方法 Download PDF

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WO2023226959A1
WO2023226959A1 PCT/CN2023/095699 CN2023095699W WO2023226959A1 WO 2023226959 A1 WO2023226959 A1 WO 2023226959A1 CN 2023095699 W CN2023095699 W CN 2023095699W WO 2023226959 A1 WO2023226959 A1 WO 2023226959A1
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ast
compounds
alkylating agent
inhibitor
inhibitors
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PCT/CN2023/095699
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French (fr)
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孟繁英
齐天阳
段建新
刘星
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深圳艾欣达伟医药科技有限公司
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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
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • A61K31/4523—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems
    • A61K31/4535—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems containing a heterocyclic ring having sulfur as a ring hetero atom, e.g. pizotifen
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/66—Phosphorus compounds
    • A61K31/664—Amides of phosphorus acids
    • 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
    • A61K31/66—Phosphorus compounds
    • A61K31/675—Phosphorus compounds having nitrogen as a ring hetero atom, e.g. pyridoxal phosphate
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the present invention relates to a method for treating cancer, in particular to the combined use of two drugs to treat cancer, and belongs to the field of cancer treatment.
  • DNA alkylating cancer treatment drug AST-3424 developed by our company targets overexpression of aldehyde-keto reductase 1C3 (AKR1C3)
  • ARR1C3 aldehyde-keto reductase 1C3
  • PCT/US2016/021581, publication number WO2016/145092, corresponding to Chinese application number 2016800150788 and publication number CN107530556A discloses compound TH2870; (R)- and (S)-1-(3-(3-N,N-dimethylaminocarbonyl)phenoxy- 4-Niphenyl)-1-ethyl-N,N'-bis(ethylidene)aminophosphate, compositions and their use and preparation methods, corresponding to PCT application number PCT/US2016/062114, publication number WO2017087428A1 , corresponding to the S configuration compound in Chinese application number 2016800446081, publication number CN108290911A), the Chinese name is (S)
  • AST-3424 enters cancer cells and is activated by the AKR1C3 enzyme overexpressed by cancer cells to release the metabolite AST2660 (also known as AST-2660) (Meng, F., Li, WF, Jung ,D.,Wang,CC,Qi,T.,Shia,CS,Hsu,RY,Hsieh,YC,&Duan,J.(2021).A novel selective AKR1C3-activated prodrug AST-3424/OBI-3424 exhibits broad anti -tumor activity.
  • AST-2660 is an alkylating agent, a chemical component of the prodrug AST-3424 that exerts its medicinal effect, and is destroyed by cross-linking with DNA double strands DNA double-stranded structure, thereby triggering cell apoptosis:
  • the clinical trial registration number of the U.S. OBI-3424 drug is NCT03592264
  • the trial stage is Phase II
  • the indications are castration prostate cancer and liver cancer
  • the sponsor is Taiwan OBI-OBI
  • the clinical trial registration number of the 3424 drug is NCT04315324.
  • the trial phase is Phase II.
  • the indication is T-ALLT lymphoblastic acute leukemia.
  • the sponsor is the Southwest Oncology Group SWOG; the clinical trial registration number of the Chinese AST-3424 drug is CTR20191399.
  • the trial phase is Phase II, and the indications are various solid tumors.
  • AST-3424 are a wide range of solid tumors and various leukemias, including Hematological tumors, which is related to the mechanism of action of AST-2660, an alkylating agent that ultimately acts on it.
  • the present invention provides the following treatment methods, drugs for treating cancer and tumors, and pharmaceutical uses.
  • Treatment method using drugs containing alkylating agent prodrug compounds and their salts, esters, solvates and isotopomers and drugs containing cell cycle inhibitor compounds and their salts, esters, solvates and isotopic isomers Combined use to treat cancer and tumor patients.
  • alkylating agent prodrug compounds and their salts, esters, solvates and isotopic isomers to prepare and combine with drugs containing cell cycle inhibitor compounds and their salts, esters, solvates and isotopic isomers Drugs for the treatment of cancer and tumor patients.
  • Drugs for the treatment of cancer and tumors contain alkylating agent prodrug compounds and their salts, esters, solvates and isotopomers.
  • the indications are compounds containing cell cycle inhibitors and their salts, esters and solvates. Drug combinations of substances and isotopes are used to treat cancer and tumor patients.
  • the invention also provides a pharmaceutical composition, which contains an alkylating agent prodrug compound or its salt, ester, solvate, isotope and a cell cycle inhibitor compound or its salt, ester, solvate or isotope.
  • the pharmaceutical composition of the isomer is used for treating cancer and tumor patients.
  • the invention also provides a pharmaceutical use in which an alkylating agent prodrug compound or its salt, ester, solvate or isotopic isomer is prepared with a cell cycle inhibitor compound or its salt, ester, solvate or isotopic isomer.
  • the cell cycle inhibitor is selected from CDK inhibitors, CHK inhibitors, ATR inhibitors, and WEE inhibitors.
  • WEE inhibitors are Wee1 inhibitors.
  • the alkylating agent prodrug compound is selected from the group consisting of AKR1C3 enzyme-activated alkylating agent prodrug compounds and hypoxic-activated alkylating agent prodrug compounds, preferably AKR1C3 enzyme-activated DNA alkylating agent prodrug compounds and hypoxic-activated DNA alkylating agent prodrug compounds. compound.
  • the AKR1C3 enzyme-activating alkylating agent prodrug compound is used in combination with a CDK inhibitor, a WEE inhibitor, a CHK inhibitor, and an ATR inhibitor.
  • hypoxia-activated alkylating agent prodrug compound is combined with a CDK inhibitor, a WEE inhibitor, or an ATR inhibitor.
  • the cell cycle is a highly regulated process that promotes cell growth, replication of genetic material, and cell division.
  • Core cell cycle machinery operating within the cell nucleus drives cells from one phase of the cell cycle to another.
  • Cell cycle inhibitors are substances that inhibit the normal operation of the cell cycle.
  • the entire cell cycle can be divided into the pre-DNA synthesis phase (G1 phase), the DNA synthesis phase (S phase), the late DNA synthesis phase (G2 phase) and the division phase (M phase).
  • G1 phase cells mainly synthesize materials required for mitosis other than DNA, including some ribosomes, proteins, etc.
  • S phase cells mainly complete the synthesis and replication of DNA.
  • topological heterogeneity is usually required.
  • the cell continues to synthesize the materials required for mitosis, including microtubules; in the M phase, under the action of microtubules, the cell divides from one mother cell into two daughter cells.
  • Cyclin-Dependent Kinases CDKs
  • WEE cell cycle checkpoint kinases
  • Checkpoint Kinase CHKs
  • ATR ata xia telangiectasia and Rad3-related
  • CDKs cyclin-dependent kinases
  • cyclin CDK4/6 acts on the transition of cells from G1 phase to S phase
  • CDK1 acts on the transition of cells from G2 phase to M phase.
  • Cell cycle inhibitors specifically act on a specific cell cycle. Specifically, inhibitors act on a specific substance or process, so that the substance or process that regulates the specific cell cycle cannot function normally.
  • CDK inhibitors are inhibitors that inhibit the corresponding kinases. By inhibiting the corresponding kinases, they regulate the progress of the cell cycle, causing cancer cells to be unable to replicate and proliferate normally and undergo apoptosis.
  • CDK1 CDK2, CDK3, CDK4, CDK5, CDK6, CDK7, CDK8, CDK9, CDK11, CDK12, CDK13, CDK14, CDK16, CDK19, CDC, CLK.
  • WEE or Wee1
  • DDR DNA damage repair pathway
  • WEE1 is a serine/threonine protein kinase that blocks the cell cycle in the G2/M phase by inhibiting the activity of CDK1/2, providing time for DNA repair.
  • Wee1 is one of the research targets of anti-cancer drugs that has been initially verified. It has a certain synthetic lethal effect with the TP53 gene mutation that is widely present in tumor cells.
  • DNA damage checkpoints are cell cycle surveillance systems that prevent genomic instability.
  • DNA damage checkpoint kinases CHK1 and CHK2 as elements in the DNA damage checkpoint, are the core of inducing cell cycle arrest, DNA repair and apoptosis.
  • cell cycle checkpoints When cellular DNA is damaged, cell cycle checkpoints are activated, causing cell cycle arrest, which facilitates cell DNA repair to maintain genome integrity and stability.
  • cell cycle checkpoints In the long-term biological evolution, cell cycle checkpoints have a set of conserved signal regulation systems, which are regulated by different CHK subtypes (Checkpoint Kinases, CHKs). Controls the cycle arrest to perform cellular DNA repair.
  • CHKs Checkpoint Kinases
  • ATR ataxia telangiectasia and Rad3-related
  • PIKK phosphatidylinositol 3-kinase
  • ATRIP ATR interacting protein
  • kinase domain for downstream protein phosphorylation, which can phosphorylate target proteins such as cell cycle checkpoint kinase 1 (CHK1) on serine or threonine.
  • ATR kinase can regulate cellular biological processes through a variety of signals, including cell cycle arrest, inhibition of replication origins, promotion of deoxynucleotide synthesis, initiation of replication forks, and repair of DNA double-strand breaks.
  • ATR can activate cellular responses after DNA damage, thereby blocking cell cycle progression, stabilizing replication forks and repairing DNA, thereby avoiding cell apoptosis and is very important for life activities.
  • Alkylating agent prodrug compounds refer to prodrug compounds that can be converted into alkylating agents in vivo.
  • the alkylating agent prodrug compound is selected from the group consisting of AKR1C3 enzyme-activated alkylating agent prodrug compounds and hypoxic-activated alkylating agent prodrug compounds, preferably AKR1C3 enzyme-activated DNA alkylating agent prodrug compounds and hypoxic-activated DNA alkylating agent prodrug compounds. compound.
  • Alkylating agent prodrug compounds include chemical structural formulas (1)-(9):
  • Formula (1) is a hypoxia-activated alkylating agent prodrug compound, more specifically, it is a hypoxia-activated DNA alkylating agent prodrug compound;
  • R is each independently selected from H, -CH 3 , -CH 2 CH 3 , -CF 3 , and X is each independently selected from leaving functional groups such as Cl, Br, MsO, and TsO.
  • TH-302 With TH-302 or its similar compounds
  • Relevant preparations include oral preparations, freeze-dried preparations and concentrated injections, and the relevant prescriptions, preparation methods, clinical compatibility, and administration methods are described in detail and disclosed by Threshold's relevant patents: WO2010048330A1, WO2012142520A2, and WO2008083101A1.
  • Threshold's relevant patents WO2010048330A1, WO2012142520A2, and WO2008083101A1.
  • the present invention will The full text of the above application text is introduced.
  • TH-302 or similar compounds It is a DNA alkylating agent anti-cancer drug with a wide range of cancer treatment potential.
  • Threshold and other pharmaceutical companies such as WO2016011195A2, WO2004087075A1 , WO2007002931A1, WO2008151253A2, WO2009018163A1, WO2009033165A2, WO2010048330A2, WO2012142520A1, WO2008083101A2, WO2020007106A1, WO2020118 251A1, WO2014169035A1, WO2013116385A1, WO2019173799A2, WO2016081547A1, WO2014062856A1, WO2015069489A1, WO2012006032A2, WO2018026606A2, WO2010 048330A2, WO2015171647A1, WO2013096687A1, WO2013126539A2, WO2013096684A2, WO2010 048330A2, WO2015171647A1,
  • Formula (2) is a hypoxia-activated alkylating agent prodrug compound, more specifically, it is a hypoxia-activated DNA alkylating agent prodrug compound;
  • R 1 , R 2 , R 3 and Cx are as described in the claims in patent application PCT/CN2020/114519 and publication number WO2021120717A1.
  • the synthesis and preparation methods of specific compounds are also described in the above application. Herein they are Its full text is incorporated into this application and is specifically defined as:
  • Cx is a 5-10-membered aromatic ring or aromatic heterocyclic ring, aliphatic heterocyclic ring or cycloalkane, which shares two carbon atoms with the nitrobenzene ring to form a fused ring structure;
  • R 1 is connected to any skeleton atom of the Cx ring, and is selected from hydrogen, halogen atom, cyano or isocyanate, hydroxyl, mercapto, amino, OTs, C 1 -C 6 alkyl or Z substituted alkyl, C 2 -C 6 alkenyl or Z substituted alkenyl, C 2 -C 6 alkynyl or Z substituted alkynyl, C 3 -C 8 cycloalkyl or Z substituted cycloalkyl, C 6 -C 10 aryl or Z substituted aryl base, 4-15 membered heterocycle or Z-substituted heterocycle, 5-15-membered heteroaryl or Z-substituted heteroaryl, 1-6 carbon atoms alkoxy group or Z-substituted 1-6 carbon atoms alkane Oxygen group, -CONR 6 R 7 , -SO 2 NR 6 R 7 , -SO 2 R 6 , -OC
  • R 2 and R 3 are each independently hydrogen, C 1 -C 6 alkyl or Z substituted alkyl, C 2 -C 6 alkenyl or Z substituted alkenyl, C 2 -C 6 alkynyl or Z substituted alkynyl, C 3 -C 8 cycloalkyl or Z substituted cycloalkyl, C 6 -C 10 aryl or Z substituted aryl, 4-15 membered heterocycle or Z substituted heterocycle, 5-15 membered heteroaryl or Z substituted
  • the heteroaryl group or R 2 , R 3 and the benzylic carbon atom to which they are bonded together form a 3-6 membered ring;
  • the group can replace the hydrogen atom at any position on the carbon atom of the fused ring, and the number of substitutions is 1;
  • the Z substituent is a halogen atom, cyano group or isocyanate group, hydroxyl group, mercapto group, amino group, C 1 -C 3 alkyl group or substituted alkyl group, C 1 -C 3 alkoxy group or substituted alkoxy group, C 2 - C 3 alkenyl or substituted alkenyl, C 2 -C 3 alkynyl or substituted alkynyl, C 3 -C 8 cycloalkyl or substituted cycloalkyl;
  • R 6 and R 7 are each independently hydrogen, C 1 -C 6 alkyl or Z-substituted C 1 -C 6 alkyl, C 2 -C 6 alkenyl or Z-substituted C 2 -C 6 alkenyl, C 2 -C 6 alkynyl or Z substituted C 2 -C 6 alkynyl, C 3 -C 8 cycloalkyl or Z substituted C 3 -C 8 cycloalkyl, C 6 -C 10 aryl or Z substituted C 6 -C 10 aryl, 4-15 membered heterocyclyl or Z-substituted 4-15-membered heterocyclyl, 5-15-membered heteroaryl or Z-substituted 5-15-membered heteroaryl, or R 6 , R 7 and the atom to which it is bonded together form a 5-7 membered heterocyclyl group or a Z-substituted 5-7 membered heterocyclyl group.
  • Formula (3) is a hypoxia-activated alkylating agent prodrug compound, more specifically, it is a hypoxia-activated DNA alkylating agent prodrug compound;
  • the definition of R 17 is as described in the claims in the patent application PCT/US2016/039092, Publication No. WO2016210175A1 (corresponding to Chinese Application No. 2016800368985, Publication No. CN108024974A).
  • the synthetic preparation method of the specific compound is also recorded in the above application.
  • Its full text is incorporated into this application and is specifically defined as:
  • R 1 is: hydrogen, -N 3 , CN, halo, NR 21 R 22 , -OR 23 , -SO 2 (C1-C6 alkyl), C1-C6 alkyl, C2-C6 alkenyl, C2-C6 Alkynyl, C3-C8 cycloalkyl, C6-C10 aryl, 4-15 membered heterocycle, 5-15 membered heteroaryl or ether;
  • R 21 and R 22 are each independently hydrogen, hydroxyl, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C6-C10 aryl, 4-15 membered heterocycle , 5-15 membered heteroaryl or -SO2 (C1-C6 alkyl); or R 21 and R 22 together with the nitrogen atom to which they are bonded form a 4-15 membered heterocyclic ring or 5-15 membered heteroaryl;
  • R 23 is hydrogen, C1-C6 alkyl or C6-C10 aryl
  • R 2 and R 3 are independently hydrogen or halo
  • R 4 is hydrogen, halo, C1-C6 alkoxy, C1-C6 alkyl or C6-C10 aryl,
  • R 5 , R 7 , R 9 , R 12 and R 15 are independently hydrogen, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C6-C10 aryl, 4-15 membered heterocycle, 5-15 membered heteroaryl; or R 4 and R 5 together with the intervening carbon atoms form a C5-C6 cycloalkyl ring;
  • R 6 and R 10 are independently hydrogen or halo
  • R 8 is hydrogen, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl or 5-15 membered heteroaryl;
  • R 11 is each independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl or C6-C10 aryl;
  • R 13 , R 14 , R 16 and R 17 are independently hydrogen, halo, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl or C1-C6 alkoxy;
  • alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocycle, heteroaryl, alkoxy and ether groups are optionally substituted.
  • Formula (4) is an AKR1C3 enzyme-activated alkylating agent prodrug compound, more specifically, it is an AKR1C3 enzyme-activated DNA alkylating agent prodrug compound;
  • X, Y, Z, R, T, A and X 10 are defined as in patent application PCT/US2016/021581, publication number WO20 16145092A1 (corresponding to Chinese application number 2016800150788, publication number CN107530556A).
  • the synthetic preparation method of the specific compound is also recorded in the above application. The full text is hereby incorporated into this application.
  • the specific definition is:
  • X 10 is O, S, SO or SO 2 ;
  • R 1 and R 2 are each independently hydrogen, C 1 -C 6 alkyl, C 3 -C 8 cycloalkyl, C 6 -C 10 aryl, 4-15 membered heterocycle, ether, -CONR 13 R 14 or -NR 13 COR 14 ;
  • X, Y and Z are each independently hydrogen, CN, halo, C 1 -C 6 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C 3 -C 8 cycloalkyl, C 6 -C 10 aryl, 4-15 membered heterocyclic ring, ether, -CONR 13 R 14 or -NR 13 COR 14 ;
  • R is hydrogen, C 1 -C 6 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C 3 -C 8 cycloalkyl, C 6 -C 10 aryl, 4-15 membered hetero Ring, ether, -CONR 13 R 14 or -NR 13 COR 14 ;
  • R 13 and R 14 are each independently hydrogen, C 1 -C 6 alkyl, C 3 -C 8 cycloalkyl, C 6 -C 10 aryl, 4-15 membered heterocycle or ether
  • T comprises an aminophosphate alkylating agent comprising one or more alkylating agents of the Z 5 -X 5 -Y 5 moiety bonded to the -OP(Z 1 ) moiety, wherein Z 5 is a heteroatom containing nitrogen, sulfur or oxygen, X 5 is a substituted or unsubstituted ethylene group, Y 5 is a halo group or another leaving group, or Z 5 -X 5 -Y 5 are formed together the aziridinyl (NCH 2 CH 2 ) moiety and Z 1 is O or S; and
  • alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocycle, heteroaryl, ether groups are substituted or unsubstituted.
  • Formulas (5) and (6) are AKR1C3 enzyme-activated alkylating agent prodrug compounds, more specifically, they are AKR1C3 enzyme-activated DNA alkylating agent prodrug compounds;
  • R 1 , R 2 , R 3 , R 4 , R 5 , R 8 , R 9 and R 10 are as recorded in the claims in patent application PCT/CN2020/089692 and publication number WO2020228685A1.
  • the synthetic preparation method is also described in the above-mentioned application, and its full text is hereby incorporated into this application. It is specifically defined as:
  • R 1 is C 6 -C 10 aryl or Z-substituted aryl, 4-15-membered heterocycle or Z-substituted heterocycle, 5-15-membered heteroaryl or Z-substituted heteroaryl, 7-15-membered fused ring Or Z substituted fused ring;
  • R 2 is hydrogen, halogen atom, cyano or isocyanate, hydroxyl, mercapto, amine, OTs, OMS, C 1 -C 6 alkyl or Z substituted alkyl, C 2 -C 6 alkenyl or Z substituted alkenyl base, C 2 -C 6 alkynyl or Z substituted alkynyl, C 3 -C 8 cycloalkyl or Z substituted cycloalkyl, C 6 -C 10 aryl or Z substituted aryl, 4-15 membered heterocycle or Z-substituted heterocycle, 5-15 membered heteroaryl or Z-substituted heteroaryl, ether of 1-6 carbon atoms or Z-substituted alkoxy group of 1-6 carbon atoms, -CONR 6 R 7 , -SO 2 NR 6 R 7 , -SO 2 R 6 , -OCOO-R 6 , -COOR 6 , -NR 6 COR
  • R 3 is hydrogen, halogen, cyano or isocyanate, hydroxyl, mercapto, amine, OTs, OMS, C 1 -C 6 alkyl or Z substituted alkyl, C 2 -C 6 alkenyl or Z substituted alkenyl , C 2 -C 6 alkynyl or Z substituted alkynyl, C 3 -C 8 cycloalkyl or Z substituted cycloalkyl, C 6 -C 10 aryl or Z substituted aryl, 4-15 membered heterocycle or Z Substituted heterocycle, 5-15 membered heteroaryl or Z-substituted heteroaryl, C 1 -C 6 alkoxy or Z-substituted C 1 -C 6 alkoxy, -CONR 6 R 7 , -SO 2 NR 6 R 7 , -SO 2 R 6 , -OCO-R 6 , -OCOO-R 6 , -COOR 6 , -
  • R 4 and R 5 are each independently hydrogen, halogen atom, cyano group or isocyanate group, hydroxyl group, mercapto group, amino group, OTs, OMS, C 1 -C 6 alkyl group or Z-substituted alkyl group, C 2 -C 6 Alkenyl or Z-substituted alkenyl, C 2 -C 6 alkynyl or Z substituted alkynyl, C 3 -C 8 cycloalkyl or Z substituted cycloalkyl, C 6 -C 10 aryl or Z substituted aryl, 4 -15-membered heterocycle or Z-substituted heterocycle, 5-15-membered heteroaryl Or Z-substituted heteroaryl, C 1 -C 6 alkoxy or Z-substituted C 1 -C 6 alkoxy, -CONR 6 R 7 , -SO 2 NR 6 R 7 , -SO 2 R 6 , -OC
  • R 6 and R 7 are each independently hydrogen, cyano or isocyanate, C 1 -C 6 alkyl or Z substituted alkyl, C 2 -C 6 alkenyl or Z substituted alkenyl, C 2 -C 6 alkyne base or Z-substituted alkynyl, C 3 -C 8 cycloalkyl or Z substituted cycloalkyl, C 6 -C 10 aryl or Z substituted aryl, 4-15 membered heterocycle or Z substituted heterocycle, 5-15 Metaheteroaryl or Z-substituted heteroaryl, C 1 -C 6 alkoxy or Z-substituted C 1 -C 6 alkoxy, or R 6 and R 7 groups together with the atoms to which they are bonded form 5- 7-membered heterocyclyl or Z-substituted 5-7-membered heterocyclyl;
  • R 8 and R 10 are each independently hydrogen, deuterium, aryl or Z-substituted aryl, C 1 -C 6 alkyl or Z substituted alkyl, C 2 -C 6 alkenyl or Z substituted alkenyl, C 2 - C 6 alkynyl or Z substituted alkynyl, C 3 -C 8 cycloalkyl or Z substituted cycloalkyl and one of them must be hydrogen or deuterium;
  • R 9 is a substituted C 6 -C 10 aryl group with at least one fluorine atom or nitro substitution, a substituted 4-15 membered heterocyclic ring with at least one fluorine atom or nitro substitution, at least one fluorine atom or nitro substitution Substituted 5-15 membered heteroaryl.
  • the Z substituent is a halogen atom, cyano or isocyanate, hydroxyl, mercapto, amine, OTs, OMS, C 1 -C 3 alkyl or substituted alkyl, C 1 -C 3 alkoxy or substituted alkoxy , C 2 -C 3 alkenyl or substituted alkenyl, C 2 -C 3 alkynyl or substituted alkynyl, C 3 -C 8 cycloalkyl or substituted cycloalkyl, aromatic ring, heterocyclic ring, heteroaromatic ring and condensed Ring or substituted aromatic ring, heterocycle, heteroaromatic ring and fused ring, the substitution mode is mono-substitution or geminal disubstitution;
  • the substituents of the substituted C 6 -C 10 aryl group, the substituted 4-15 membered heterocyclic group, and the substituted 5-15 membered heteroaryl group in R 9 are halogen atoms, nitro groups, cyano groups or isocyanate groups, hydroxyl groups, and amino groups , C 1 -C 3 alkyl or alkoxy group, alkenyl, alkynyl, cycloalkyl or benzene ring, substituted benzene ring, C 1 -C 3 alkoxy group or halogen atom substituted alkoxy group.
  • Formula (7) is an AKR1C3 enzyme-activated alkylating agent prodrug compound, more specifically, it is an AKR1C3 enzyme-activated DNA alkylating agent prodrug compound;
  • R 100 , R 101 and R 102 are each independently hydrogen, C1-C8 alkyl, C6-C12 aryl; or R 101 and R 102 together with the nitrogen atom to which they are attached form a 5-7 membered heterocyclic ring;
  • alkyl group and the aryl group are each substituted by 1-3 halo groups or 1-3 C1-C6 alkyl groups;
  • R 1 and R 2 are each independently phenyl or methyl
  • X, Y and Z are each independently hydrogen or halo
  • R is hydrogen or C1-C6 alkyl or halogen substituted alkyl
  • Formula (8) is an AKR1C3 enzyme-activated alkylating agent prodrug compound, more specifically, it is an AKR1C3 enzyme-activated DNA alkylating agent prodrug compound;
  • Rw is as recorded in the claims in the patent application PCT/CN2020/120281 and the publication number WO2021068952A1.
  • the synthesis and preparation method of the specific compound is also recorded in the above application, and its full text is hereby incorporated into this application. Specifically defined as:
  • R 1 is H, C 1-6 alkyl, C 3-6 cycloalkyl, 4-6 membered heterocycloalkyl, 5-6 membered heteroaryl or phenyl, wherein the C 1-6 alkyl, C 3-6 cycloalkyl, 4-6 membered heterocycloalkyl, 5-6 membered heteroaryl and phenyl are optionally substituted by 1, 2 or 3 R a ;
  • Each R a is independently H, F, Cl, Br, I, -CN, -OH, C 1-3 alkoxy or C 1-3 alkyl;
  • R 2 is H or C 1-6 alkyl
  • R 1 and R 2 are connected together, and the N atoms to which they are connected together form a 4-6 membered heterocycloalkyl group, wherein the 4-6 membered heterocycloalkyl group is optionally substituted by 1, 2 or 3 R b ;
  • Each R b is independently H, F, Cl, Br, I, -CN, -OH, -NH 2 , -OCH 3 , -OCH 2 CH 3 , -CH 3 or -CH 2 CH 3 ;
  • R 3 is H, F, Cl, Br, I, -OH, -NH 2 , C 1-3 alkoxy or C 1-3 alkyl;
  • R2 and R3 are connected together to make the structural unit for
  • T 1 is -(CR c R d ) m - or -(CR c R d ) n -O-;
  • n 1, 2 or 3;
  • n 1 or 2;
  • T 2 is N or CH
  • R c and R d are each independently H, F, C 1-3 alkyl or C 1-3 alkoxy;
  • R 4 , R 5 and R 6 are each independently H, F, Cl, Br, I, C 1-3 alkyl or C 1-3 alkoxy;
  • T is N or CH
  • R 7 and R 8 are each independently H, F, Cl, Br or I;
  • R 9 and R 10 are each independently H, F, Cl, Br, I, -CN or
  • the 4-6 membered heterocycloalkyl and 5-6 membered heteroaryl groups each contain 1, 2, 3 or 4 heteroatoms independently selected from N, -O- and -S-.
  • Formula (9) is an AKR1C3 enzyme-activated alkylating agent prodrug compound, more specifically, it is an AKR1C3 enzyme-activated DNA alkylating agent prodrug compound;
  • A is H, C1-C6 alkyl, C1-C6 alkenyl, C1-C6 alkynyl, CFH 2 , CF 2 H, CF 3 , F, Cl, Br, I, OCF 3 , COR or CON(R) 2 ;
  • E is SO or SO 2 ;
  • X is Cl, Br, I or OSO 2 R;
  • Y is Cl, Br, I or OSO 2 R;
  • Each R is independently H or C1-C6 alkyl
  • G is a radical selected from the group consisting of formulas (B)-(AA):
  • R 1 is H, C1-C6 alkyl, CH 2 (CH 2 )nOH, CH 2 CH(OH)CH 2 OH, phenyl, pyridyl, benzyl or pyridylmethyl, provided that when R 1 is benzene group, pyridyl, benzyl or pyridylmethyl, R 1 is optionally replaced by C1-C6 alkyl, C1-C6 alkenyl, C1-C6 alkynyl, OR 6 , N(R 6 )(R 7 ), CFH 2 , CF 2 H, CF 3 , F, Cl, Br, I, OC F 3 , COR 6 , CON(R 6 )(R 7 ), SOR 6 , SON(R 6 )(R 7 ), SO 2 R 6 , SO 2 N(R 6 )(R 7 ), CN or NO 2 replace;
  • R 2 and R 3 are each independently H, C1-C6 alkyl, C1-C6 alkenyl, C1-C6 alkynyl, OR 6 , N(R 6 )(R 7 ), CFH 2 , CF 2 H, CF 3 , F, Cl, Br, I, OCF 3 , COR 6 , CON(R 6 )(R 7 ), SOR 6 , SON(R 6 )(R 7 ), SO2R 6 , SO2N(R 6 )(R 7 ), CN or NO 2 ;
  • R 4 is N(R 6 )(R 7 ), OH, OCH 2 (CH 2 )nN(R 6 )(R 7 ) or CH 2 (CH 2 )nN(R 6 )(R 7 );
  • R 5 is H or C1-C6 alkyl group
  • R 6 and R 7 are each independently H or C1-6 alkyl, or R 6 and R 7 together form a substituted or unsubstituted 5- or 6-membered heterocycle;
  • Z is CH or N
  • W is CH 2 , O, S, SO or SO 2 ;
  • n ranges from 0 to 6;
  • the medicines described herein refer to medicines or preparations, and the medicines prepared contain the hypoxia-activated compound of formula (1) or a salt or solvate thereof as an active ingredient in a specific dosage range, and/or the medicines prepared are specific Dosage form and specific administration method.
  • the above-mentioned drugs should also add pharmaceutically acceptable auxiliary materials or excipients according to the specific drugs, drugs, and preparations.
  • the drug can be in any dosage form for clinical use, such as tablets, suppositories, dispersible tablets, enteric-coated tablets, chewable tablets, orally disintegrating tablets, capsules, sugar-coated agents, granules, dry powders, oral solutions, and small injection needles. , freeze-dried powder for injection or large infusion.
  • pharmaceutically acceptable excipients or excipients in the drug may include one or more of the following: diluents, solubilizers, disintegrants, suspending agents, lubricants, viscosifiers, Mixtures, fillers, flavoring agents, sweeteners, antioxidants, surfactants, preservatives, coating agents, and pigments, etc.
  • Cancer refers to leukemias, lymphomas, carcinomas, and other malignancies (including solid tumors) with potentially unrestrained growth that can expand locally by invasion and systemically by metastasis.
  • cancers that can be treated by AST-3424 or such AKR1C3 enzyme-activated DNA alkylating agent prodrugs include (but are not limited to) adrenal, bone, brain, breast, bronchi, colon and/or rectum, gallbladder, head and cancers of the neck, kidney, larynx, liver, lung, nervous tissue, pancreas, prostate, parathyroid gland, skin, stomach and thyroid gland.
  • cancers include acute and chronic lymphocytic and granulocytic tumors, adenocarcinoma, adenoma, basal cell carcinoma, cervical dysepithelial carcinoma and carcinoma in situ, Ewing's sarcoma, epidermoid carcinoma, giant cell tumor, multiple glioblastoma, pilocytic tumors, intestinal ganglioneuroma, proliferative corneal neurotumor, islet cell carcinoma, Kaposi's sarcoma, leiomyoma, leukemia, lymphoma, malignant carcinoid tumor, malignant melanoma , malignant hypercalcemia, Marfanoid tumor, medullary epithelial carcinoma, metastatic skin cancer, mucosal neuroma, myeloma, mycosis fungoides, neuroblastoma, osteosarcoma, osteogenic and other sarcomas, Ovarian tumors, pheochromocytoma, polycythemia, poly
  • Monotherapy refers to the use of only one anticancer drug in a course of treatment.
  • Combination therapy refers to the simultaneous or sequential use of two or more anti-cancer drugs in one course of treatment.
  • combination therapy requires exploring different dosages and administration cycles based on the characteristics of the disease and the types of combined drugs. Only based on the above conditions, the explored combination drug treatment plan can achieve better therapeutic effects than single drug treatment.
  • the drug dosage and dosage regimen of the combined treatment plan need to be explored through clinical trials with reference to the dosage and dosage regimen of the above-mentioned alkylating agent prodrug compounds and combined drugs.
  • the cancer, tumor patients or their biological samples are found to have specific gene deletion or damage.
  • the specific gene is selected from genes involved in cell cycle checkpoint regulation, preferably p53, p21, CCNB1, WIP1, 14-3-3sigma protein, cdc2/cycB.
  • the cell cycle inhibitors include CDK inhibitors, WEE inhibitors, CHK inhibitors and ATR inhibitors.
  • the CDK inhibitor is selected from the group consisting of Palbociclib, Ribociclib, Abemaciclib, Trilaciclib, dalpiciclib, Adavosertib, Ro-3306, Dinaciclib, Cirtuvivint , Rintodestrant, DS96432529, THZ1, THZ531, Seliciclib, Flavopiridol, AZD4573, SR-4835, Simurosertib, Fadraciclib, NVP-2, SNS-032, (E/Z)-Zotiraciclib, AZD-5438, AT7519, Mevociclib, Kenpaullone, YKL -5-124TFA ⁇ NG 52 ⁇ GSK 3Inhibitor IX ⁇ OTS964 ⁇ Samuraciclib ⁇ Flavopiridol ⁇ KB-0742dihydrochloride ⁇ (+)-Enitociclib ⁇ AUZ 454 ⁇ SY-5609 ⁇ SEL120-34A mono
  • the WEE inhibitor is selected from WEE1-IN-5, WEE1-IN-3, WEE1-IN-4, LEB-03-146, LEB-03-144, Adavosertib, LEB-03-153, LEB-03- 145, PD407824, PD0166285, PD0166285 dihydrochloride, Pomalidomide-C3-adavosertib, DB0614, FMF-06-098-1, etc.
  • MCE MedChemExpress
  • the CHK inhibitor is selected from AZD7762, Prexasertib, SCH900776, GDC-0425, Chk1-IN-6, CCT245737, BML-277, CCT241533, PD407824, CHIR-124, CCT244747, PF477736, GDC-0575, SB-218078, MRT00033659, ANI-7, SAR-020106, CCT241533, CHK1-IN-4, VER-00158411, CHK1-IN-3, Chk1-IN-5, CHK1-IN-2, CHK-IN-1, etc., these compounds
  • the list, specific structure, and supply information can be obtained by visiting commercial reagent websites, such as the https://www.medchemexpress.cn/Targets/Checkpoint%20Kinase%20(Chk).html page of the MedChemExpress (MCE) website.
  • the ATR inhibitor is selected from Ceralasertib, Berzosertib, Gartisertib, BAY1895344, BAY-937, AZ20, ETP-46464, Dactolisib, VE-821, M1774, ATRN-199, RP-3500, ART-0380.
  • Information about these compounds See the literature: Yuan Yinghui, Duan Jilong, Huizi, et al. Research progress of targeting ATR kinase inhibitors in the treatment of cancer [J]. Acta Pharmaceutical Sinica, 2022(057-003).
  • Figure 1 shows the proliferation inhibition curve of AST-3424 alone or combined with Adavosertib on the HT-29 cell line;
  • Figure 2 shows the inhibitory rates of AST-3424 and Adavosertib on HT-29 cell proliferation as single drugs and combinations at fixed concentrations (in each group of histograms, the left side is AST-3424 single drug, the middle is Adavosertib single drug, and the right side It is the combination of AST-3424 and Adavosertib;
  • Figure 3 shows the proliferation inhibition curve of AST-3424 alone or in combination with AZD7762 on the HT-29 cell line;
  • Figure 4 shows the inhibitory rates of single drugs and combinations of AST-3424 and AZD7762 on HT-29 cell proliferation at fixed concentrations (in each group of histograms, the left side is AST-3424 single drug, the middle is AZD7762 single drug, and the right side It is the combination of AST-3424 and AZD7762);
  • Figure 5 shows the proliferation inhibition curve of AST-3424 alone or in combination with Palbociclib on the HT-29 cell line
  • Figure 6 shows the inhibitory rate of single drug and combination drug on HT-29 cell proliferation at fixed concentrations of AST-3424 and Palbociclib (in each set of histograms, the left side is AST-3424 single drug, the middle is Palbociclib single drug, and the right side It is AST-3424 combined with Palbociclib);
  • Figure 7 shows the proliferation inhibition curve of AST-3424 alone or in combination with Adavosertib on H460 cell line;
  • Figure 8 shows the inhibitory rate of single drug and combined drug on H460 cell proliferation at fixed concentrations of AST-3424 and Adavosertib (in each set of histograms, the left side is AST-3424 single drug, the middle is Adavosertib single drug, and the right side is AST -3424 in combination with Adavosertib);
  • Figure 9 shows the proliferation inhibition curve of AST-3424 alone or in combination with AZD7762 on the H460 cell line
  • Figure 10 shows the inhibitory rates of single and combined drugs on H460 cell proliferation at fixed concentrations of AST-3424 and AZD7762. (In each set of histograms, the left side is AST-3424 as a single drug, the middle is AZD7762 as a single drug, and the right side is AST-3424 in combination with AZD7762);
  • Figure 11 shows the proliferation inhibition curve of AST-3424 alone or in combination with Palbociclib on the H460 cell line
  • Figure 12 shows the inhibitory rate of single drug and combination drug on H460 cell proliferation at fixed concentrations of AST-3424 and Palbociclib (in each set of histograms, the left side is AST-3424 single drug, the middle is Palbociclib single drug, and the right side is AST -3424 in combination with Palbociclib);
  • Figure 13 shows the proliferation inhibition curve of AST alone or combined with Advosertib on the HT-29 cell line
  • Figure 14 shows the inhibitory rate of single drug and combined drug on HT-29 cell proliferation at fixed concentrations of AST and Adavosertib (in each group of histograms, the left side is AST single drug, the middle is Adavosertib single drug, and the right side is AST and Adavosertib combined use);
  • Figure 15 shows the proliferation inhibition curve of AST alone or combined with AZD7762 on the HT-29 cell line
  • Figure 16 shows the inhibitory rate of single drug and combined drug on HT-29 cell proliferation at fixed concentrations of AST and AZD7762 (in each group of histograms, the left side is AST single drug, the middle is AZD7762 single drug, and the right side is AST and AZD7762 combined use);
  • Figure 17 is the proliferation inhibition curve of AST alone or combined with Palbociclib on the HT-29 cell line
  • Figure 18 shows the inhibitory rate of single drug and combination drug on HT-29 cell proliferation at fixed concentrations of AST and Palbociclib (in each group of histograms, the left side is AST single drug, the middle is Palbociclib single drug, and the right side is AST and Palbociclib combined use);
  • Figure 19 shows the proliferation inhibition curve of AST alone or combined with Advosertib on H460 cell line
  • Figure 20 shows the inhibitory rate of single drug and combined drug on H460 cell proliferation at fixed concentrations of AST and Adavosertib (in each set of bar graphs, the left side is AST single drug, the middle is Adavosertib single drug, and the right side is AST and Adavosertib combined );
  • Figure 21 is the proliferation inhibition curve of AST alone or combined with AZD7762 on H460 cell line;
  • Figure 22 shows the inhibitory rate of single drug and combined drug on H460 cell proliferation at fixed concentrations of AST and AZD7762 (in each set of histograms, the left side is AST single drug, the middle is AZD7762 single drug, and the right side is the combination of AST and AZD7762 );
  • Figure 23 is the proliferation inhibition curve of AST alone or combined with Palbociclib on H460 cell line;
  • Figure 24 shows the inhibitory rates of single drugs and combinations of AST and Palbociclib on H460 cell proliferation at fixed concentrations of AST and Palbociclib (in each set of histograms, the left side is AST single drug, the middle is Palbociclib single drug, and the right side is AST and Palbociclib combined );
  • Figure 25 shows the proliferation inhibition curve of AST-3424 alone or in combination with Ceralasertib on the HT29/H460 cell line, in which the bottom curve is the AST-3424 single drug group;
  • Figure 26 shows the proliferation inhibition curve of AST-3424 alone or in combination with Ceralasertib on the HT29/H460 cell line in another experiment, in which the bottom curve is the AST-3424 single drug group;
  • Figure 27 shows the proliferation inhibition curve of AST-3424 alone or in combination with Ceralasertib on the HT29 cell line in experiments with different dosing orders of Ceralasertib and AST-3424.
  • the bottom curve is the AST-3424 single drug group;
  • Figure 28 is the proliferation inhibition curve of TH-302 alone or in combination with Adavosertib on the HT29 cell line, in which ⁇ 0.01% O 2 represents hypoxic conditions, and the upper left curve is the combination of TH-302 and 10 ⁇ M Adavosertib under hypoxic conditions. Group;
  • Figure 29 shows the proliferation inhibition curve of TH-302 alone or in combination with Adavosertib on the H460 cell line, in which ⁇ 0.01% O 2 indicates hypoxic conditions, and the combined drug group of TH-302 and 30 ⁇ M Adavosertib under hypoxic conditions is on the left;
  • Figure 30 shows the proliferation inhibition curve of TH-302 alone or in combination with Ceralaserti on the HT29 cell line, in which ⁇ 0.01% O 2 represents hypoxic conditions, and the upper left curve is the combination of TH-302 and 2000 ⁇ M Ceralaserti under hypoxic conditions. Group;
  • Figure 31 is the proliferation inhibition curve of TH-302 alone or combined with Ceralasertib on H460 cell line, where ⁇ 0.01% O 2 represents hypoxic conditions, and the TH-302 normoxic single-drug group is on the right.
  • HT-29 indicates experiments in the HT-29 cell line
  • Adavosertib-2h-pretreatment AST-3424-72h-cotreatment indicates that in combination treatment, Adavosertib is first treated for 2h, and then AST-3424 is added for co-treatment for 72h; in single use, only AST-3424 is added, and then treated for 72h;
  • Adavosertib (alias AZD1775, MK-1775) is a selective inhibitor of Wee1 kinase, a key kinase involved in the cell cycle G2/M checkpoint and DNA damage repair processes.
  • cyclin dependent kinase Cyclin-dependent kinases, CDKs
  • Their activity depends on binding to the cyclin Cyclin to form a complex, and their activity is also regulated by phosphorylation.
  • the phosphorylation status of CDK1 further controls G2/ M checkpoint.
  • Wee1 is a "gatekeeper" at the cell cycle G2/M checkpoint, allowing cells to repair DNA damage before entering mitosis.
  • Wee1 maintains CDK1 in an inactive state by phosphorylating CDK1Tyr15, thereby inhibiting the activity of the CDK1-cyclin B complex, stalling cell division at the G2/M checkpoint, and negatively regulating the cell cycle. Its biological significance is to prevent repair in time. The DNA damage is repaired, preventing cells from entering mitosis carrying DNA damage. Adavosertib selectively inhibits Wee1 kinase and blocks DNA damage repair at the G2/M phase checkpoint in p53 and other gene-deficient tumors, leading to tumor cell death and ultimately achieving the purpose of treating tumors. Its structural formula is:
  • AZD7762 is an effective ATP-competitive cell cycle checkpoint kinase (CHk) inhibitor that eliminates the S and G2 checkpoints induced by DNA damage. It is a CHK inhibitor and its structural formula is
  • Palbociclib is an orally active CDK4 and CDK6 selective inhibitor with the structural formula:
  • Ceralasertib (AZD6738) is a potent ATR kinase inhibitor that controls the G2/M checkpoint. Its structural formula is
  • Combined medication Add 1 ⁇ L of Adavosertib (four concentrations), AZD7762 (four concentrations), Palbociclib (four concentrations) and Ceralasertib (three concentrations) to each well, shake gently to ensure even mixing, and place in culture After incubating for 2 hours, add 1 ⁇ L of test compound AST-3424 or AST at different concentrations, shake gently to ensure uniform mixing, and then place in a 37°C, 5% CO2 incubator.
  • test compound AST-3424 or AST Single use: 24 hours after cell plating, add 99 ⁇ L of growth medium to each well. Add 1 ⁇ L of test compound AST-3424 or AST at different concentrations, shake gently to ensure uniform mixing, and then place in a 37°C, 5% CO2 incubator.
  • Table 1 IC 50 value of AST-3424 alone or in combination with Adavosertib on proliferation inhibition of HT-29 cell line
  • ratio represents the ratio of IC 50 when AST-3424 is used alone and when AST-3424 is combined with Adavosertib.
  • Table 2 IC 50 value of AST-3424 alone or in combination with AZD7762 on the proliferation inhibition of HT-29 cell line
  • ratio represents the ratio of IC 50 when AST-3424 is used alone and when AST-3424 is combined with AZD7762.
  • Table 3 IC 50 value of AST-3424 alone or in combination with Palbociclib on proliferation inhibition of HT-29 cell line
  • ratio represents the ratio of IC 50 when AST-3424 is used alone and when AST-3424 is combined with Palbociclib.
  • the IC 50 value of the proliferation inhibition of H460 cell line by AST-3424 alone or combined with Adavosertib was obtained by fitting.
  • the results are shown in Table 4 below.
  • AST-3424 was combined with different concentrations of Adavosertib, high concentrations
  • the IC 50 value of Adavosertib combination is about 2 times different from the IC 50 value of AST-3424 alone.
  • the IC 50 value of Adavosertib combination at other concentrations is not significantly different from the IC 50 value of AST-3424 alone.
  • Table 4 IC 50 value of AST-3424 alone or in combination with Adavosertib on proliferation inhibition of H460 cell line
  • ratio represents the ratio of IC 50 when AST-3424 is used alone and when AST-3424 is combined with Adavosertib.
  • the IC 50 value of AST-3424 alone or in combination with AZD7762 on the proliferation inhibition of the H460 cell line was obtained.
  • the results are shown in Table 5 below.
  • AST-3424 was combined with different concentrations of AZD7762, 160nM AZD7762
  • the IC 50 value of the combination drug is about 2 times different from the IC 50 value of AST-3424 as a single drug.
  • the IC 50 values of AZD7762 combination drugs at other concentrations are not significantly different from the IC 50 value of AST-3424 as a single drug.
  • Table 5 IC 50 value of AST-3424 alone or in combination with AZD7762 on proliferation inhibition of H460 cell line
  • ratio represents the ratio of IC 50 when AST-3424 is used alone and when AST-3424 is combined with AZD7762.
  • ratio represents the ratio of IC 50 when AST-3424 is used alone and when AST-3424 is combined with Palbociclib.
  • the IC 50 value of the proliferation inhibition of HT-29 cell line by AST alone or combined with Adavosertib was obtained by fitting.
  • the results are shown in Table 7 below. After AST was combined with Adavosertib at different concentrations, the IC of the combined drug was The 50 value is significantly smaller than the single-agent IC 50 value of AST-3424, and shows a certain degree of Adavosertib dose dependence.
  • Table 7 IC 50 value of AST alone or combined with Adavosertib on proliferation inhibition of HT-29 cell line
  • ratio represents the ratio of IC 50 when AST is used alone and when AST is combined with Adavosertib.
  • the IC 50 value of the proliferation inhibition of HT-29 cell line by AST alone or in combination with AZD7762 was obtained by fitting.
  • the results are shown in Table 8 below. After AST was combined with different concentrations of AZD7762, the IC of the combined drug was The 50 value is significantly smaller than the single drug IC 50 value of AST-3424.
  • Table 8 IC 50 value of AST alone or combined with AZD7762 on proliferation inhibition of HT-29 cell line
  • ratio represents the ratio of IC 50 when AST is used alone and when AST is combined with AZD7762.
  • Table 9 IC 50 value of AST alone or combined with Palbociclib on proliferation inhibition of HT-29 cell line
  • ratio, single/combo, single/combo represents the ratio of IC 50 when AST is used alone and when AST is combined with Palbociclib.
  • Table 10 IC 50 value of AST alone or combined with Adavosertib on proliferation inhibition of H460 cell line
  • ratio, single/combo, single/combo represents the ratio of IC 50 when AST is used alone and when AST is combined with Adavosertib.
  • the IC 50 value of AST alone or in combination with AZD7762 on the proliferation inhibition of the H460 cell line was obtained.
  • the results are shown in Table 11 below.
  • the IC 50 of the 160 nM AZD7762 combination was The value is about 3 times different from the single drug IC 50 value of AST.
  • the IC 50 values of other concentrations of AZD7762 combination drugs are not significantly different from the single drug IC 50 value of AST.
  • Table 11 IC 50 value of AST alone or combined with AZD7762 on proliferation inhibition of H460 cell line
  • ratio, single/combo represents the ratio of IC 50 when AST is used alone and when AST is combined with AZD7762.
  • the IC50 value of the proliferation inhibition of the H460 cell line by AST alone or in combination with Palbociclib was obtained by fitting.
  • the results are shown in Table 12 below. After AST was combined with different concentrations of Palbociclib, the IC50 value of the combined drug was the same as that of the combined drug. There is no significant difference in the single-drug IC 50 value of AST, and it is greater than the single-drug IC 50 value.
  • Table 12 IC 50 value of AST alone or combined with Palbociclib on proliferation inhibition of H460 cell line
  • ratio, single/combo represents the ratio of IC 50 when AST is used alone and when AST is combined with Palbociclib.
  • the first experiment was performed to obtain the proliferation inhibition curve of AST-3424 alone or in combination with Ceralasertib on the HT29/H460 cell line in Figure 25. According to the experimental results in Figure 25, the effect of AST-3424 alone or in combination with Ceralasertib on HT29/H460 was obtained by fitting.
  • the IC 50 value of proliferation inhibition of H460 cell line is shown in Table 13 below.
  • Table 13 IC 50 value of AST-3424 alone or in combination with Ceralasertib on proliferation inhibition of HT29/H460 cell line (first experiment)
  • Ratio means ratio
  • Table 14 IC 50 value of AST-3424 alone or in combination with Ceralasertib on proliferation inhibition of HT29/H460 cell line (second experiment)
  • Ratio means ratio
  • Table 15 IC 50 value of AST-3424 alone or in combination with Ceralasertib on the proliferation inhibition of HT29 cell line under different dosing orders
  • ratio, single/combo represents the ratio of IC 50 when AST is used alone and when AST is combined with Palbociclib.
  • AST-3424 and Ceralasertib (ATR inhibitor) has a significant additive effect on the proliferation of H460 cells (p53 wild type).
  • AST-3424 and Ceralasertib (ATR inhibitor) at various concentrations has a very significant additive effect on the proliferation of HT29 cells (p53 deletion), and the additive effect is stronger than that of the H460 cell line.
  • the 24-well plate with glass insert was transferred to the hypoxia workstation.
  • Adavosertib final concentration: 1 ⁇ M, 10 ⁇ M, respectively
  • Ceralasertib final concentration: 200 ⁇ M, 2000 ⁇ M, respectively
  • the final concentrations of Adavosertib added were 3 ⁇ M and 30 ⁇ M, and the final concentrations of Ceralasertib added were 2 ⁇ M, 20 ⁇ M, and 1000 ⁇ M.
  • the IC 50 value of TH-302 alone or combined with Adavosertib on the HT29/H460 cell line under normoxic and hypoxic conditions was obtained, as shown in Table 16 and Table 17, and the corresponding proliferation inhibition curve is shown in Figure 28 , shown in Figure 29.
  • Table 16 Experimental results of inhibition of proliferation of HT29 cell line by TH-302 alone or in combination with Adavosertib under normoxic and hypoxic conditions
  • Compounds represents a compound or a combination of compounds; Ratio represents a ratio.
  • Compounds represents a compound or a combination of compounds; Ratio represents a ratio.
  • the IC 50 value of TH-302 alone or combined with Ceralasertib on the HT29/H460 cell line under normoxic and hypoxic conditions was obtained, as shown in Table 18 and Table 19, and the corresponding proliferation inhibition curve is shown in Figure 30 , shown in Figure 31.
  • Table 18 Experimental results of inhibition of proliferation of HT29 cell line by TH-302 alone or combined with Ceralasertib
  • Compounds represents a compound or a combination of compounds; Ratio represents a ratio.
  • Compounds represents a compound or a combination of compounds; Ratio represents a ratio.
  • TH-302 In p53-deficient (HT29) cell lines, TH-302 has no significant cytotoxicity-enhancing effect when combined with the G2/M cell cycle regulator Adavosertib under normoxic conditions.
  • TH-302 showed a significant cytotoxicity-enhancing effect when combined with the G2/M cell cycle regulator Adavosertib under hypoxic conditions and high concentrations of Adavosertib.
  • TH-302 had no significant cytotoxicity-enhancing effect when combined with the G2/M cell cycle regulator Adavosertib, whether under normoxic or hypoxic conditions.
  • TH-302 had no significant cytotoxicity-enhancing effect when combined with the ATR kinase inhibitor Ceralasertib under normoxic conditions.
  • TH-302 was combined with the ATR kinase inhibitor Ceralasertib. When combined under hypoxic conditions, high concentrations of Ceralasertib showed a significant cytotoxicity-enhancing effect.
  • compounds AST-3424, AST and TH-302 are all DNA alkylating agent prodrugs, which can specifically release the DNA alkylating agent small molecule AST-2660 (or 2660) or Br under specific circumstances. -IPM.
  • the DNA alkylating agents ultimately released by these compounds are cell cycle non-specific, that is, they act on the DNA double-stranded structure at all stages of the cell cycle. Alkylating agents can form very tight bonds between DNA double strands, causing DNA to be unable to complete processes such as transcription, translation, and replication. In other words, they can cause DNA damage and inhibit proliferation of cells in all different cell cycles. .
  • the compound Adavosertib is a selective inhibitor of Wee1 kinase, a key kinase involved in the cell cycle G2/M checkpoint and DNA damage repair processes.
  • Cyclin-dependent kinases (CDKs) regulate the cell cycle by phosphorylating specific substrates. Their activity depends on binding to Cyclin to form a complex, and their activity is also regulated by phosphorylation. The phosphorylation status of CDK1 further controls G2 /M checkpoint.
  • Wee1 maintains CDK1 in an inactive state by phosphorylating CDK1Tyr15, thereby inhibiting the activity of the CDK1-cyclin B complex, stalling cell division at the G2/M checkpoint, and negatively regulating the cell cycle.
  • Adavosertib regulates the phosphorylation status of CDK1 by inhibiting Wee1 kinase, thereby regulating cell cycle progression.
  • Adavosertib is an inhibitor that indirectly acts on CDK and can be considered a broad CDK inhibitor.
  • Palbociclib is an orally active CDK4 and CDK6 selective inhibitor. Therefore, Adavosertib and Palbociclib, as CDK inhibitors, are cell cycle specific and act on specific stages of the cell cycle.
  • AST, AST-3424, and TH-302 are all DNA alkylating agents covered by the general structural formulas (1)-(9) as prodrugs that ultimately act on the DNA double-stranded cross-linking process.
  • Compounds, as well as Adavosertib and Palbociclib, are CDK inhibitors that act on specific cell cycles. It can be inferred that: alkylating agent prodrug compounds and CDK inhibitors that act on specific cell cycles have a cytotoxic additive effect on tumor cell lines, that is, both Drug combination treatment for cancer and tumor patients will have better therapeutic effects, and p53 deletion will enhance the above-mentioned additive effect/combination effect.
  • AZD7762 is a potent ATP-competitive cell cycle checkpoint kinase (Chk) inhibitor that eliminates the S and G2 checkpoints induced by DNA damage and is a ChK inhibitor.
  • Chk cell cycle checkpoint kinase
  • AST and AST-3424 are DNA alkylating agents covered by the general structural formulas (1)-(9) as prodrug compounds that ultimately act on the DNA double-stranded cross-linking process, and AZD7762 As a ChK inhibitor that acts on a specific cell cycle, it can be inferred that the alkylating agent prodrug compound and the CHK inhibitor that acts on a specific cell cycle have a cytotoxic additive effect on tumor cell lines, that is, the two drugs are combined to treat cancer and tumors. Patients will have better treatment outcomes, and p53 deletion will enhance the above-mentioned additive/combination effects.
  • Ceralasertib (AZD6738) is a potent ATR kinase inhibitor that is cell cycle specific and acts on specific stages of the cell cycle.
  • AST, AST-3424, and TH-302 are all DNA alkylating agents covered by the general structural formulas (1)-(9) as prodrugs that ultimately act on the DNA double-stranded cross-linking process.
  • Compounds, and Ceralasertib are ATR inhibitors that act on specific cell cycles. It can be inferred that: alkylating agent prodrug compounds and ATR inhibitors that act on specific cell cycles have additive cytotoxic effects on tumor cell lines, that is, the combination of two drugs Treating cancer and tumor patients will have better therapeutic effects, and p53 deletion will enhance the above-mentioned additive/combination effects.
  • the combination of cell cycle non-specific alkylating prodrug compounds and cell cycle inhibitors that act on specific cell cycles has a cytotoxic additive effect on tumor cells. That is, the combination of two drugs can treat cancer and tumors. Cancer patients will have better therapeutic effects; and further, p53 deletion will enhance the above-mentioned additive effect/combination effect, that is, patients with p53 gene deletion may get more therapeutic benefits after combination.

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Abstract

烷化剂前药与细胞周期抑制剂联用治疗癌症的方法，烷化剂前药可以为AKR1C3酶活化的烷化剂前药或乏氧活化的烷化剂前药，细胞周期抑制剂可以为CDK抑制剂、WEE抑制剂、CHK抑制剂或ATR抑制剂。

Description

烷化剂前药与细胞周期抑制剂联用治疗癌症的方法 技术领域
本发明涉及癌症的治疗方法，特别是两种药物联用治疗癌症，属于癌症治疗领域。
背景技术
我公司开发的以过表达醛酮还原酶1C3(AKR1C3)为标靶的DNA烷化癌症治疗药物AST-3424(参见专利申请：DNA烷化剂，对应PCT申请号PCT/US2016/021581,公开号WO2016/145092，对应中国申请号2016800150788，公开号CN107530556A中公开化合物TH2870；(R)-及(S)-1-(3-(3-N,N-二甲基胺基羰基)苯氧基-4-硝苯基)-1-乙基-N,N’-双(伸乙基)胺基磷酸酯、组合物及其使用及制备方法，对应PCT申请号PCT/US2016/062114,公开号WO2017087428A1，对应中国申请号2016800446081，公开号CN108290911A中的S构型化合物)，中文名为(S)-1-(3-(3-N,N-二甲氨基羰基)苯氧基-4-硝基苯基)-1-乙基-N,N'-双(亚乙基)氨基磷酸酯，也称为OBI-3424、TH-2870的S构型化合物)，CAS号为2097713-69-2，其结构如下：
AST-3424的化学结构式
AST-3424(又名OBI-3424、TH-3424)进入癌细胞内被癌细胞过表达的AKR1C3酶活化而释放代谢产物AST2660(又名AST-2660)(Meng,F.,Li,W.F.,Jung,D.,Wang,C.C.,Qi,T.,Shia,C.S.,Hsu,R.Y.,Hsieh,Y.C.,&Duan,J.(2021).A novel selective AKR1C3-activated prodrug AST-3424/OBI-3424 exhibits broad anti-tumor activity.Americanjournal ofcancer research,11(7),3645–3659.)，AST-2660是烷化剂，是前药AST-3424发挥药效的化学成分，其通过与DNA双链交联而破坏DNA双链结构，进而引发细胞凋亡：
AST-3424代谢为AST-2660(图中2660)的化学反应式
在临床试验中(美国OBI-3424药物的临床试验登记号为NCT03592264，试验阶段为II期，适应症为去势前列腺癌和肝癌，申办方为台湾浩鼎生技股份有限公司OBI；美国OBI-3424药物的临床试验登记号为NCT04315324，试验阶段为II期，适应症为T-ALLT淋巴细胞急性白血病，申办方为美国西南肿瘤协作组SWOG；中国AST-3424药物的临床试验登记号为CTR20191399，试验阶段为II期，适应症为各种实体瘤，申办方为深圳艾欣达伟医药科技有限公司；中国AST-3424药物的临床试验登记号为CTR20201915，试验阶段为II期，适应症为T-ALL和B-ALL T淋巴细胞急性白血病和B淋巴细胞急性白血病，申办方为深圳艾欣达伟医药科技有限公司)，AST-3424的适应症是广泛的实体肿瘤和各种白血病在内的血液肿瘤，这与其最终其作用的AST-2660这一烷化剂的作用机制相关。
发明内容
随着申请人对AST-3424及其类似的AKR1C3激活的烷化剂前药和TH-302及其类似的乏氧激活的烷化剂前药的进一步临床前研究和临床试验研究，申请人发现该类前药化合物与细胞周期抑制剂联用将能达到更好的肿瘤抑制效果。
本发明提供以下的治疗方法，治疗癌症、肿瘤的药物，制药用途。
治疗方法，使用含有烷化剂前药化合物及其盐、酯、溶剂合物、同位素异构体的药物与含有细胞周期抑制剂化合物及其盐、酯、溶剂合物、同位素异构体的药物联用治疗癌症、肿瘤患者。
制药用途，使用烷化剂前药化合物及其盐、酯、溶剂合物、同位素异构体制备与含有细胞周期抑制剂化合物及其盐、酯、溶剂合物、同位素异构体的药物联用治疗癌症、肿瘤患者的药物。
治疗癌症、肿瘤的药物，该药物含有烷化剂前药化合物及其盐、酯、溶剂合物、同位素异构体，其适应症为与含有细胞周期抑制剂化合物及其盐、酯、溶剂合物、同位素异构体的药物联用治疗癌症、肿瘤患者。
本发明还提供一种药物组合物，一种含有烷化剂前药化合物或其盐、酯、溶剂合物、同位素异构体与细胞周期抑制剂化合物或其盐、酯、溶剂合物、同位素异构体的药物组合物，该药物组合物用于治疗癌症、肿瘤患者。
本发明还提供一种制药用途，烷化剂前药化合物或其盐、酯、溶剂合物、同位素异构体在制备与细胞周期抑制剂化合物或其盐、酯、溶剂合物、同位素异构体联用治疗癌症、肿瘤的药物中的用途。
细胞周期抑制剂选自CDK抑制剂、CHK抑制剂、ATR抑制剂、WEE抑制剂。
WEE抑制剂即Wee1抑制剂。
烷化剂前药化合物选自AKR1C3酶活化烷化剂前药化合物、乏氧活化烷化剂前药化合物，优选为AKR1C3酶活化DNA烷化剂前药化合物、乏氧活化DNA烷化剂前药化合物。
优选地，AKR1C3酶活化烷化剂前药化合物与CDK抑制剂、WEE抑制剂、CHK抑制剂、ATR抑制剂联用。
优选地，乏氧活化烷化剂前药化合物与CDK抑制剂、WEE抑制剂联用、ATR抑制剂。
细胞周期是一个高度调控的过程，它能促进细胞生长、遗传物质复制和细胞分裂。在细胞核内运行的核心细胞周期机制会驱动细胞从细胞周期的一个阶段发展到另一个阶段。细胞周期抑制剂就是抑制细胞周期正常运转的物质。
整个细胞周期可分为DNA合成前期(G1期)，DNA合成期(S期)，DNA合成后期(G2期)和分裂期(M期)。G1期，细胞主要合成除DNA以外的有丝分裂所需的物质，包括一些核糖体、蛋白质等；S期，细胞主要完成DNA的合成和复制，同时，为了形成方便DNA复制的结构，通常需要拓扑异构酶的协助；G2期，细胞继续合成有丝分裂所需物质，包括微管等；M期，在微管等的作用下，细胞由一个母细胞分裂成两个子细胞。
上述细胞周期与细胞周期蛋白及其催化搭档细胞周期蛋白依赖激酶(Cyclin-Dependent Kinases，CDKs)、WEE、细胞周期检查点激酶(Checkpoint Kinase，CHKs)以及ATR(ata xia telangiectasia and Rad3-related)即“共济失调毛细血管扩张突变基因Rad3相关激酶”密切相关。
不同的细胞周期蛋白依赖激酶(CDKs)作用于细胞周期的四个不同阶段，比如细胞周期蛋白CDK4/6作用于细胞的G1期转换为S期、CDK1作用于细胞的G2期转换为M期。
细胞周期抑制剂即特异性的作用于特定的细胞周期，具体的，抑制剂作用于某个特定的物质或过程，使得调控该特定细胞周期的物质或过程不能正常发挥作用。
CDK抑制剂(CDKi)就是抑制对应的激酶的抑制剂，通过抑制相应的激酶而调控细胞周期的进度，导致癌细胞无法正常复制增殖而凋亡。
已知的针对不同CDK亚型：CDK1、CDK2、CDK3、CDK4、CDK5、CDK6、CDK7、CDK8、CDK9、CDK11、CDK12、CDK13、CDK14、CDK16、CDK19、CDC、CLK具有不同的抑制剂。
WEE，即Wee1，是DNA损伤修复(DDR)通路相关的一个重要激酶。WEE1是一个丝氨酸/苏氨酸蛋白激酶，它可以通过抑制CDK1/2的活性从而将细胞周期阻滞在G2/M期，为DNA修复提供时间。Wee1是已被初步验证的抗癌药物研究靶点之一，与肿瘤细胞广泛存在的TP53基因突变存在一定的合成致死效应。
DNA损伤检查点是细胞周期监测系统，可防止基因组不稳定。DNA损伤检查点激酶CHK1和CHK2作为DNA损伤检查点中的元素，是诱导细胞周期停滞、DNA修复和细胞凋亡的核心。
细胞DNA受到损伤时会激活细胞周期检查点(checkpoint)，造成细胞周期停滞，从而利于细胞进行DNA修复以保持基因组的完整和稳定。在长期的生物进化中，细胞周期检查点有一套保守的信号调控体系，通过不同的CHK亚型(Checkpoint Kinases，CHKs)来调 控使得周期停滞进行细胞的DNA修复。
ATR(ataxia telangiectasia and Rad3-related)即“共济失调毛细血管扩张突变基因Rad3相关激酶”，属于磷脂酰肌醇3-激酶(PIKK)家族中的丝氨酸-苏氨酸激酶，由2644个氨基酸组成，在其N端存在着ATR相互作用蛋白(ATRIP)结构域，这也是ATR激酶激活的重要区域。在它的C端有下游蛋白磷酸化的激酶结构域，能够将靶蛋白如细胞周期检测点激酶1(CHK1)等丝氨酸或苏氨酸磷酸化。ATR激酶激活后可通过多种信号调控细胞生物过程，包括细胞周期阻滞、抑制复制起点、促进脱氧核苷酸合成、启动复制叉以及修复DNA双链断裂。ATR在DNA损伤后能够激活细胞应答，进而阻滞细胞周期进程并稳定复制叉及修复DNA，从而回避细胞凋亡，对生命活动十分重要。
烷化剂前药化合物是指能在生物体内转化为烷化剂的前药化合物。
烷化剂前药化合物选自AKR1C3酶活化烷化剂前药化合物、乏氧活化烷化剂前药化合物，优选为AKR1C3酶活化DNA烷化剂前药化合物、乏氧活化DNA烷化剂前药化合物。
烷化剂前药化合物包括化学结构式(1)-(9)：
式(1)为乏氧活化烷化剂前药化合物，更具体的，其为乏氧活化DNA烷化剂前药化合物；
其中，R各自独立地选自H、-CH3、-CH2CH3、-CF3，X各自独立地选自Cl、Br、MsO、TsO等离去官能团。
与TH-302或其类似化合物相关的制剂包括口服制剂、冻干制剂和浓缩注射液，并且相关处方、制备方法和临床配伍、施用方法被Threshold公司的相关专利：WO2010048330A1、WO2012142520A2、WO2008083101A1所详细说明并公开，在此本发明将上述申请文本的全文引入。
TH-302或其类似化合物是一个的DNA烷化剂类抗癌药，具有广泛的癌症治疗潜力，这些相关的癌症适应症实验、临床试验被公开在相关的Threshold公司及其他制药公司的专利申请文本中(比如WO2016011195A2、WO2004087075A1、WO2007002931A1、WO2008151253A2、WO2009018163A1、WO2009033165A2、WO2010048330A2、WO2012142520A1、WO2008083101A2、WO2020007106A1、WO2020118251A1、WO2014169035A1、WO2013116385A1、WO2019173799A2、WO2016081547A1、WO2014062856A1、WO2015069489A1、WO2012006032A2、WO2018026606A2、WO2010048330A2、WO2015171647A1、WO2013096687A1、WO2013126539A2、WO2013096684A2、WO2012009288A2、WO2012145684A2、WO2016014390A2、WO2019055786A2、WO2012135757A2、WO2015013448A2、WO2016011328A2、WO2013177633A2、WO2016011195A2、WO2015051921A2)以及FDA登记的临床试验中(NCT02402062、NCT02020226、NCT02076230、 NCT01381822、NCT02093962、NCT01440088、NCT02255110、NCT02342379、NCT01864538、NCT01149915、NCT02433639、NCT00743379、NCT01485042、NCT01721941、NCT02047500、NCT00742963、NCT01497444、NCT00495144、NCT01746979、NCT01144455、NCT01403610、NCT01522872、NCT01833546、NCT02598687、NCT03098160、NCT02496832、NCT02712567)，在此本发明将上述相关申请文本以及临床试验信息全部引入。
式(2)为乏氧活化烷化剂前药化合物，更具体的，其为乏氧活化DNA烷化剂前药化合物；
其中，R1、R2、R3、Cx的定义如专利申请PCT/CN2020/114519，公开号WO2021120717A1中的权利要求书所记载，具体化合物的合成制备方法也记载在上述申请中，在此将其全文引入本申请之中，具体定义为：
Cx为5-10元的芳环或芳杂环、脂杂环或环烷烃，其与硝基苯环共用两个碳原子形成稠环结构；
R1连接在Cx环的任意骨架原子上，选自氢、卤素原子、氰基或异氰基、羟基、巯基、胺基、OTs、C1-C6烷基或Z取代烷基、C2-C6烯基或Z取代烯基、C2-C6炔基或Z取代炔基、C3-C8环烷基或Z取代环烷基、C6-C10芳基或Z取代芳基、4-15元杂环或Z取代杂环、5-15元杂芳基或Z取代杂芳基、1-6个碳原子的烷氧基或Z取代的1-6个碳原子的烷氧基、-CONR6R7、-SO2NR6R7、-SO2R6、-OCOO-R6、-COOR6、-NR6COR7、-OCOR6、-NR6SO2R7、-NR6SO2NR6R7，
R2、R3各自独立地是氢、C1-C6烷基或Z取代烷基、C2-C6烯基或Z取代烯基、C2-C6炔基或Z取代炔基、C3-C8环烷基或Z取代环烷基、C6-C10芳基或Z取代芳基、4-15元杂环或Z取代杂环、5-15元杂芳基或Z取代杂芳基或者R2、R3和与其所键结的苄位碳原子一起形成3-6元环；
基团可取代稠环碳原子上任意位置的氢原子，取代的个数为1；
Z取代基为卤素原子、氰基或异氰基、羟基、巯基、胺基、C1-C3烷基或取代烷基、C1-C3烷氧基或取代烷氧基、C2-C3烯基或取代烯基、C2-C3炔基或取代炔基、C3-C8环烷基或取代环烷基；
R6、R7各自独立地是氢、C1-C6烷基或Z取代的C1-C6烷基、C2-C6烯基或Z取代的C2-C6烯基、C2-C6炔基或Z取代的C2-C6炔基、C3-C8环烷基或Z取代的C3-C8环烷基、C6-C10芳基或Z取代的C6-C10芳基、4-15元杂环基或Z取代的4-15元杂环基、5-15元杂芳基或Z取代的5-15元杂芳基，或者R6、R7和与其所键结的原子一起形成5-7元杂环基或Z取代的5-7元杂环基。

式(3)为乏氧活化烷化剂前药化合物，更具体的，其为乏氧活化DNA烷化剂前药化合物；
其中，R1、R2、R3、R4、R5、R6、R7、R8、R9、R10、R11、R12、R13、R14、R15、R16、R17的定义如专利申请PCT/US2016/039092，公开号WO2016210175A1(对应中国申请号2016800368985，公开号CN108024974A)中的权利要求书所记载，具体化合物的合成制备方法也记载在上述申请中，在此将其全文引入本申请之中，具体定义为：
R1为：氢、-N3、CN、卤基、NR21R22、-OR23、-SO2(C1-C6烷基)、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C8环烷基、C6-C10芳基、4-15元杂环、5-15元杂芳基或醚；
R21和R22各自独立地为氢、羟基、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C8环烷基、C6-C10芳基、4-15元杂环、5-15元杂芳基或-SO2(C1-C6烷基)；或R21和R22与其所键接的氮原子一起形成4-15元杂环或5-15元杂芳基；
R23为氢、C1-C6烷基或C6-C10芳基；
R2和R3独立地为氢或卤基；
R4为氢、卤基、C1-C6烷氧基、C1-C6烷基或C6-C10芳基，
R5、R7、R9、R12和R15独立地为氢、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C8环烷基、C6-C10芳基、4-15元杂环、5-15元杂芳基；或R4和R5与其之间的介入碳原子一起形成C5-C6环烷基环；
R6和R10独立地为氢或卤基；
R8为氢、C1-C6烷基、C2-C6烯基、C2-C6炔基或5-15元杂芳基；
R11各自独立地为C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C8环烷基或C6-C10芳基；
R13、R14、R16和R17独立地为氢、卤基、C1-C6烷基、C2-C6烯基、C2-C6炔基或C1-C6烷氧基；
其中所述烷基、烯基、炔基、环烷基、芳基、杂环、杂芳基、烷氧基和醚基团任选地经取代。
式(4)为AKR1C3酶活化烷化剂前药化合物，更具体的，其为AKR1C3酶活化DNA烷化剂前药化合物；
其中，X、Y、Z、R、T、A以及X10的定义如专利申请PCT/US2016/021581，公开号WO20 16145092A1(对应中国申请号2016800150788，公开号CN107530556A)中的权利要求书所记载，具体化合物的合成制备方法也记载在上述申请中，在此将其全文引入本申请之中，具体定义为：
X10是O、S、SO或SO2；
A是C6-C10芳基、5-15元杂芳基或-N＝CR1R2；
R1及R2各自独立是氢、C1-C6烷基、C3-C8环烷基、C6-C10芳基、4-15元杂环、醚、-CONR13R14或-NR13COR14；
X、Y及Z各自独立是氢、CN、卤基、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C8环烷基、C6-C10芳基、4-15元杂环、醚、-CONR13R14或-NR13COR14；
R是氢、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C8环烷基、C6-C10芳基、4-15元杂环、醚、-CONR13R14或-NR13COR14；
R13及R14各自独立是氢、C1-C6烷基、C3-C8环烷基、C6-C10芳基、4-15元杂环或醚
T包含胺基磷酸酯烷化剂，所述胺基磷酸酯烷化剂包含一或多个键结至-O-P(Z1)部分的Z5-X5-Y5部分的烷化剂，其中Z5是包含氮、硫或氧的杂原子，X5是经取代或未经取代的伸乙基，Y5是卤基或另一离去基，或Z5-X5-Y5一起形成氮丙啶基(NCH2CH2)部分且Z1是O或S；且
其中这些烷基、烯基、炔基、环烷基、芳基、杂环、杂芳基、醚基经取代或未经取代。
式(5)、(6)为AKR1C3酶活化烷化剂前药化合物，更具体的，其为AKR1C3酶活化DNA烷化剂前药化合物；
其中，R1、R2、R3、R4、R5、R8、R9、R10的定义如专利申请PCT/CN2020/089692，公开号WO2020228685A1中的权利要求书所记载，具体化合物的合成制备方法也记载在上述申请中，在此将其全文引入本申请之中，具体定义为：
R1是C6-C10芳基或Z取代的芳基、4-15元杂环或Z取代杂环、5-15元杂芳基或Z取代杂芳基、7-15元的稠环或Z取代稠环；
R2是氢、卤素原子、氰基或异氰基、羟基、巯基、胺基、OTs、OMS、C1-C6烷基或Z取代烷基、C2-C6烯基或Z取代烯基、C2-C6炔基或Z取代炔基、C3-C8环烷基或Z取代环烷基、C6-C10芳基或Z取代芳基、4-15元杂环或Z取代杂环、5-15元杂芳基或Z取代杂芳基、1-6个碳原子的醚或Z取代的1-6个碳原子的烷氧基、-CONR6R7、-SO2NR6R7、-SO2R6、-OCOO-R6、-COOR6、-NR6COR7、-OCOR6、-NR6SO2R7、-NR6SO2NR6R7或者R2和与其所键结的R1基团上的原子一起形成7-15元的稠环或Z取代稠环；
R3是氢、卤素、氰基或异氰基、羟基、巯基、胺基、OTs、OMS、C1-C6烷基或Z取代烷基、C2-C6烯基或Z取代烯基、C2-C6炔基或Z取代炔基、C3-C8环烷基或Z取代环烷基、C6-C10芳基或Z取代芳基、4-15元杂环或Z取代杂环、5-15元杂芳基或Z取代杂芳基、C1-C6烷氧基或Z取代的C1-C6烷氧基、-CONR6R7、-SO2NR6R7、-SO2R6、-OCO-R6、-OCOO-R6、-COOR6、-NR6COR7，-OCOR6、-NR6SO2R7；
R4、R5各自独立地是氢、卤素原子、氰基或异氰基、羟基、巯基、胺基、OTs、OMS、C1-C6烷基或Z取代烷基、C2-C6烯基或Z取代烯基、C2-C6炔基或Z取代炔基、C3-C8环烷基或Z取代环烷基、C6-C10芳基或Z取代芳基、4-15元杂环或Z取代杂环、5-15元杂芳基 或Z取代杂芳基、C1-C6烷氧基或Z取代的C1-C6烷氧基、-CONR6R7、-SO2NR6R7、-SO2R6、-OCOO-R6、-COOR6、-NR6COR6、-OCOR6、-NR6SO2R7或者R4、R5和与其所键结的苯环上的原子一起形成7-15元的稠环或Z取代稠环；
R6和R7各自独立地是氢、氰基或异氰基、C1-C6烷基或Z取代烷基、C2-C6烯基或Z取代烯基、C2-C6炔基或Z取代炔基、C3-C8环烷基或Z取代环烷基、C6-C10芳基或Z取代芳基、4-15元杂环或Z取代杂环、5-15元杂芳基或Z取代杂芳基、C1-C6烷氧基或Z取代的C1-C6烷氧基，或者R6、R7基团与其所键结的原子一起形成5-7元杂环基或Z取代5-7元杂环基；
R8、R10各自独立地为氢、氘、芳基或Z取代芳基、C1-C6烷基或Z取代烷基、C2-C6烯基或Z取代烯基、C2-C6炔基或Z取代炔基、C3-C8环烷基或Z取代环烷基且必有一个为氢、氘；
R9为至少具有一个氟原子或硝基取代的取代C6-C10芳基、至少具有一个氟原子或硝基取代的取代4-15元杂环、至少具有一个氟原子或硝基取代的取代5-15元杂芳基。
Z取代基为卤素原子、氰基或异氰基、羟基、巯基、胺基、OTs、OMS、C1-C3烷基或取代烷基、C1-C3烷氧基或取代烷氧基、C2-C3烯基或取代烯基、C2-C3炔基或取代炔基、C3-C8环烷基或取代环烷基、芳环、杂环、杂芳环和稠环或取代芳环、杂环、杂芳环和稠环，取代的方式为单取代或偕二取代；
R9中的取代C6-C10芳基、取代4-15元杂环、取代5-15元杂芳基的取代基为卤素原子、硝基、氰基或异氰基、羟基、胺基、C1-C3烷基或烷氧基、烯基、炔基、环烷基或苯环、取代苯环、C1-C3烷氧基或卤原子取代烷氧基。
式(7)为AKR1C3酶活化烷化剂前药化合物，更具体的，其为AKR1C3酶活化DNA烷化剂前药化合物；
其中：
A是取代或未经取代的C6-C10的芳基、联芳基或取代的联芳基、5-15元的杂芳基或-N＝C R1R2，其中取代时的取代基选自由以下组成的群：卤基、-CN、-NO2、–O-(CH2)-O-、-CO2H及其盐、-OR100、-CO2R100、-CONR101R102、-NR101R102、-NR100SO2R100、-SO2R100、-SO2NR101R10 2、C1-C6烷基、C3-C10杂环基；
其中，R100、R101及R102各自独立是氢、C1-C8烷基、C6-C12芳基；或R101及R102与其附接至的氮原子一起形成5-7元杂环；
其中烷基及芳基各自是经1-3个卤基或1-3个C1-C6烷基取代；
R1及R2各自独立是苯基或甲基；
X、Y及Z各自独立是氢或卤基；
R是氢或C1-C6烷基或卤素取代烷基；
式(8)为AKR1C3酶活化烷化剂前药化合物，更具体的，其为AKR1C3酶活化DNA烷化剂前药化合物；
其中，Rw的定义如专利申请PCT/CN2020/120281，公开号WO2021068952A1中的权利要求书所记载，具体化合物的合成制备方法也记载在上述申请中，在此将其全文引入本申请之中，具体定义为：
Rw为
R1为H、C1-6烷基、C3-6环烷基、4-6元杂环烷基、5-6元杂芳基或苯基，其中所述C1-6烷基、C3-6环烷基、4-6元杂环烷基、5-6元杂芳基和苯基任选被1、2或3个Ra所取代；
各Ra独立地为H、F、Cl、Br、I、-CN、-OH、C1-3烷氧基或C1-3烷基；
R2为H或C1-6烷基；
或者R1和R2连接在一起，与其相连的N原子一起形成4-6元杂环烷基，其中所述4-6元杂环烷基任选被1、2或3个Rb所取代；
各Rb独立地为H、F、Cl、Br、I、-CN、-OH、-NH2、-OCH3、-OCH2CH3、-CH3或-CH2CH3；
R3为H、F、Cl、Br、I、-OH、-NH2、C1-3烷氧基或C1-3烷基；
或者R2和R3连接在一起使结构单元为
T1为-(CRcRd)m-或-(CRcRd)n-O-；
m为1、2或3；
n为1或2；
T2为N或CH；
Rc和Rd各自独立地为H、F、C1-3烷基或C1-3烷氧基；
R4、R5和R6各自独立地为H、F、Cl、Br、I、C1-3烷基或C1-3烷氧基；
T为N或CH；
R7和R8各自独立地为H、F、Cl、Br或I；
R9和R10各自独立地为H、F、Cl、Br、I、-CN或
所述4-6元杂环烷基和5-6元杂芳基各自包含1、2、3或4个独立选自N、-O-和-S-的杂原子。
式(9)为AKR1C3酶活化烷化剂前药化合物，更具体的，其为AKR1C3酶活化DNA烷化剂前药化合物；
其中，A、E、G、X、Y的定义如专利申请PCT/NZ2019/050030，公开号WO2019190331A1(对应中国申请号2019800234236，公开号CN111918864A)中的权利要求书所记载，具体化合物的合成制备方法也记载在上述申请中，在此将其全文引入本申请之中，具体定义为：
A为H、C1-C6烷基、C1-C6烯基、C1-C6炔基、CFH2、CF2H、CF3、F、Cl、Br、I、OCF3、COR或CON(R)2；
E为SO或SO2；
X为Cl、Br、I或OSO2R；
Y为Cl、Br、I或OSO2R；
每个R独立地为H或C1-C6烷基；
G是选自包括式(B)-(AA)的组中的自由基：
其中：
R1为H、C1-C6烷基、CH2(CH2)nOH、CH2CH(OH)CH2OH、苯基、吡啶基、苄基或吡啶基甲基，条件是当R1为苯基、吡啶基、苄基或吡啶基甲基时，R1任选地在任何可用位置处被C1-C6烷基、C1-C6烯基、C1-C6炔基、OR6、N(R6)(R7)、CFH2、CF2H、CF3、F、Cl、Br、I、OC F3、COR6、CON(R6)(R7)、SOR6、SON(R6)(R7)、SO2R6、SO2N(R6)(R7)、CN或NO2取代；
R2和R3各自独立地为H、C1-C6烷基、C1-C6烯基、C1-C6炔基、OR6、N(R6)(R7)、CFH2、CF2H、CF3、F、Cl、Br、I、OCF3、COR6、CON(R6)(R7)、SOR6、SON(R6)(R7)、SO2R6、SO2N(R6)(R7)、CN或NO2；
R4为N(R6)(R7)、OH、OCH2(CH2)nN(R6)(R7)或CH2(CH2)nN(R6)(R7)；
R5为H或C1-C6烷基基团；
R6和R7各自独立地为H或C1-6烷基，或者R6和R7一起形成取代或未被取代的5元或6元杂环；
Z为CH或N；
W为CH2、O、S、SO或SO2；
n为0至6；
*表示与式(I)的连接点。
关于本文所述药物是指药品或制剂，所制得的药品包含特定剂量范围的有效成分式(1)的乏氧激活化合物或其盐或溶剂合物，和/或所制得的药物为特定剂型、特定给药方式施用。
上述药物除含有式(1)-(9)的烷化剂前药化合物外，还应根据药品、药物、制剂的特定，添加药学上可接受的辅料或赋形剂。所述药物可以为临床施用的任何剂型，例如片剂、栓剂、分散片、肠溶片、咀嚼片、口崩片、胶囊、糖衣剂、颗粒剂、干粉剂、口服溶液剂、注射用小针、注射用冻干粉针或大输液。根据具体剂型和施用方式，所述药物中的药学上可接受的辅料或赋形剂可以包括下述的一种或多种：稀释剂、增溶剂、崩解剂、悬浮剂、润滑剂、粘合剂、填充剂、矫味剂、甜味剂、抗氧化剂、表面活性剂、防腐剂、包裹剂、和色素等。
“癌症”是指可通过侵袭而局部扩展且通过转移而全身扩展的潜在无限制生长的白血病、淋巴瘤、癌及其他恶性肿瘤(包括实体肿瘤)。
在此列举AST-3424或这类AKR1C3酶活化的DNA烷化剂前药能治疗的癌症的实例包括(但不限于)肾上腺、骨、脑、乳房、支气管、结肠及/或直肠、胆囊、头及颈、肾、喉、肝、肺、神经组织、胰脏、前列腺、副甲状腺、皮肤、胃及甲状腺的癌症。癌症的某些其他实例包括急性及慢性淋巴细胞及粒细胞肿瘤、腺癌、腺瘤、基底细胞癌、子宫颈上皮分化不良及原位癌、尤文氏肉瘤、表皮样癌、巨细胞瘤、多型性神经胶母细胞瘤、毛细胞肿瘤、肠神经节细胞瘤、增生性角膜神经肿瘤、胰岛细胞癌、卡波西肉瘤、平滑肌瘤、白血病、淋巴瘤、恶性类癌瘤、恶性黑色素瘤、恶性高钙血症、马方样体型肿瘤、髓样上皮癌、转移性皮肤癌、黏膜神经瘤、骨髓瘤、蕈状肉芽肿、神经胚细胞瘤、骨肉瘤、骨原性及其他肉瘤、卵巢瘤、嗜铬细胞瘤、真性红血球增多症、原发性脑瘤、小细胞肺癌、溃疡型及乳头型二者的鳞状细胞癌、增生、精原细胞瘤、软组织肉瘤、视网膜母细胞瘤、横纹肌肉瘤、肾细胞肿瘤、局部皮肤病灶、网状细胞肉瘤及威尔姆氏肿瘤。
联用，即联合用药治疗。单药治疗是指在一个疗程中仅使用一种抗癌药物。联合治疗是指在一个疗程中同时或先后使用两种或两种以上的抗癌药物。
一般而言，联合治疗需要根据病情特点、联用药物种类探索不同的给药剂量、给药周期，只有根据上述情况，探索得到的联合用药治疗方案才可能取得较单一用药治疗好的治疗效果。
联用治疗方案的药物给药剂量、给药周期均需要在参考上述烷化剂前药化合物和联用药物的剂量、给药方案通过临床试验探索得到。
所述癌症、肿瘤患者或其生物样本被检查出具有特定基因缺失或损伤，所述特定基因选自参与细胞周期Checkpoint调控的基因，优选p53、p21、CCNB1、WIP1、14-3-3sigma protein、cdc2/cycB。
这些基因根据相关研究文献(Molinari,M.(2000),Cell cycle checkpoints and their inact ivation in human cancer.Cell Proliferation,33:261-274.https://doi.org/10.1046/j.1365-2184.2000.00191.x；Donehower L.A.(2014).Phosphatases reverse p53-mediated cell cycle checkpoin ts.Proceedings of the National Academy of Sciences of the United States of America,111(20),7172–7173.https://doi.org/10.1073/pnas.1405663111)可以确定为参与细胞周期Checkpoint调控的基因。
所述细胞周期抑制剂包括CDK抑制剂、WEE抑制剂、CHK抑制剂以及ATR抑制剂。
优选的，CDK抑制剂选自哌柏西利(Palbociclib)、瑞波西利(Ribociclib)、阿贝西利(Abemaciclib)、曲拉西利(Trilaciclib)、达尔西利(dalpiciclib)、Adavosertib、Ro-3306、Dinaciclib、Cirtuvivint、Rintodestrant、DS96432529、THZ1、THZ531、Seliciclib、Flavopiridol、AZD4573、SR-4835、Simurosertib、Fadraciclib、NVP-2、SNS-032、(E/Z)-Zotiraciclib、AZD-5438、AT7519、Mevociclib、Kenpaullone、YKL-5-124TFA、NG 52、GSK 3Inhibitor IX、OTS964、Samuraciclib、Flavopiridol、KB-0742dihydrochloride、(+)-Enitociclib、AUZ 454、SY-5609、SEL120-34A monohydrochloride、CCT-251921、MBQ-167、XL413hydrochloride、BI-1347、THAL-SNS-032、JNJ-7706621、TG003、LDC4297、BMS-265246、Roniciclib、CGP60474、(R)-CR8trihydrochloride、R547、Milciclib、T025、AS2863619、Senexin A、BSJ-4-116、CLK-IN-T3、CDK12-IN-3、CVT-313、Atuveciclib、PHA-793887、Indirubin-3'-monoxime、YKL-5-124、PHA-767491hydrochloride、KH-CB19、Cucurbitacin E、Purvalanol A、BSJ-03-204、ON123300、CDK5inhibitor 20-223、Riviciclib、FN-1501、CP-10、THZ2、Abemaciclib metabolite M2、BS-181、CDK12-IN-E9、Samuraciclib、RGB-286638、CDK2-IN-4、LDC000067、ML167、Purvalanol B、NU6300、CLK1-IN-1、FMF-04-159-2、CKI-7、CDKI-73、MSC2530818、BSJ-04-132、NU6102、Voruciclib、Olomoucine等，这些化合物的名单以及具体结构、供应信息可以通过访问商业试剂网站得到，比如MedChemExpress(MCE)网站的https://www.medchemexpress.cn/Targets/CDK.html？effectName＝Inhibitor网页。
优选的，WEE抑制剂选自WEE1-IN-5、WEE1-IN-3、WEE1-IN-4、LEB-03-146、LEB-03-144、Adavosertib、LEB-03-153、LEB-03-145、PD407824、PD0166285、PD0166285 dihydrochloride、Pomalidomide-C3-adavosertib、DB0614、FMF-06-098-1等，这些化合物的名单以及具体结构、供应信息可以通过访问商业试剂网站得到，比如MedChemExpress(MCE)网站的https://www.medchemexpress.cn/search.html？q＝Wee&ft＝&fa＝&fp＝&fsp＝&ftag＝&fsc＝网页。
优选的，CHK抑制剂选自AZD7762、Prexasertib、SCH900776、GDC-0425、Chk1-IN-6、CCT245737、BML-277、CCT241533、PD407824、CHIR-124、CCT244747、PF477736、GDC-0575、SB-218078、MRT00033659、ANI-7、SAR-020106、CCT241533、CHK1-IN-4、VER-00158411、CHK1-IN-3、Chk1-IN-5、CHK1-IN-2、CHK-IN-1等，这些化合物的名单以及具体结构、供应信息可以通过访问商业试剂网站得到，比如MedChemExpress(MCE)网站的https://www.medchemexpress.cn/Targets/Checkpoint％20Kinase％20(Chk).html网页。
优选的，ATR抑制剂选自Ceralasertib、Berzosertib、Gartisertib、BAY1895344、BAY-937、AZ20、ETP-46464、Dactolisib、VE-821、M1774、ATRN-199、RP-3500、ART-0380，这些化合物的信息参见文献：袁滢惠,段吉隆,惠子,等.靶向ATR激酶抑制剂治疗癌症的研究进展[J].药学学报,2022(057-003)。
式(1)的具体化合物选自以下结构：
式(2)-(9)的具体化合物的化学结构如权利要求7所述，在此不再赘述。
其中，式(2)具体化合物参考专利申请文件PCT/CN2020/114519，公开号WO2021120717A1的权利要求9，在此将该专利申请文件全文引入本申请。
式(3)具体化合物参考专利申请文件CN2016800368985，公开号CN108024974A的【0073-0093】段的表格中化合物，在此将该专利申请文件全文引入本申请。
式(4)具体化合物参考专利申请文件PCT/US2016/021581，公开号WO2016145092A1(对应中国申请号2016800150788，公开号CN107530556A)中的化合物，在此将该专利申请文件全文引入本申请。
式(5)(6)具体化合物参考专利申请文件PCT/CN2020/089692，公开号WO2020228685A1中的化合物，在此将该专利申请文件全文引入本申请。
式(7)具体化合物参考专利申请文件PCT/US2016/021581、公开号WO2016145092A1(对应中国申请号2016800150788、公开号CN107530556A)和PCT/CN2020/089692、公开号WO2020228685A1中的化合物，在此将该两件专利申请文件全文引入本申请。
式(8)具体化合物参考专利申请文件PCT/CN2020/120281，公开号WO2021068952A1中的化合物，在此将该专利申请文件全文引入本申请。
式(9)具体化合物参考专利申请文件PCT/NZ2019/050030，公开号WO2019190331A1(对应中国申请号2019800234236，公开号CN111918864A)中的化合物，在此将该专利申请文件全文引入本申请。
附图说明
图1为AST-3424单独或联用Adavosertib对HT-29细胞系的增殖抑制曲线；
图2为AST-3424和Adavosertib固定浓度下，单药和联用药对HT-29细胞增殖的抑制率(其中每一组柱状图中，左边是AST-3424单药，中间是Adavosertib单药，右边是AST-3424与Adavosertib联用；
图3为AST-3424单独或联用AZD7762对HT-29细胞系的增殖抑制曲线；
图4为AST-3424和AZD7762固定浓度下，单药和联用药对HT-29细胞增殖的抑制率(其中每一组柱状图中，左边是AST-3424单药，中间是AZD7762单药，右边是AST-3424与AZD7762联用)；
图5为AST-3424单独或联用Palbociclib对HT-29细胞系的增殖抑制曲线；
图6为AST-3424和Palbociclib固定浓度下，单药和联用药对HT-29细胞增殖的抑制率(其中每一组柱状图中，左边是AST-3424单药，中间是Palbociclib单药，右边是AST-3424与Palbociclib联用)；
图7为AST-3424单独或联用Adavosertib对H460细胞系的增殖抑制曲线；
图8为AST-3424和Adavosertib固定浓度下，单药和联用药对H460细胞增殖的抑制率(其中每一组柱状图中，左边是AST-3424单药，中间是Adavosertib单药，右边是AST-3424与Adavosertib联用)；
图9为AST-3424单独或联用AZD7762对H460细胞系的增殖抑制曲线；
图10为AST-3424和AZD7762固定浓度下，单药和联用药对H460细胞增殖的抑制率 (其中每一组柱状图中，左边是AST-3424单药，中间是AZD7762单药，右边是AST-3424与AZD7762联用)；
图11为AST-3424单独或联用Palbociclib对H460细胞系的增殖抑制曲线；
图12为AST-3424和Palbociclib固定浓度下，单药和联用药对H460细胞增殖的抑制率(其中每一组柱状图中，左边是AST-3424单药，中间是Palbociclib单药，右边是AST-3424与Palbociclib联用)；
图13为AST单独或联用Adavosertib对HT-29细胞系的增殖抑制曲线；
图14为AST和Adavosertib固定浓度下，单药和联用药对HT-29细胞增殖的抑制率(其中每一组柱状图中，左边是AST单药，中间是Adavosertib单药，右边是AST与Adavosertib联用)；
图15为AST单独或联用AZD7762对HT-29细胞系的增殖抑制曲线；
图16为AST和AZD7762固定浓度下，单药和联用药对HT-29细胞增殖的抑制率(其中每一组柱状图中，左边是AST单药，中间是AZD7762单药，右边是AST与AZD7762联用)；
图17为AST单独或联用Palbociclib对HT-29细胞系的增殖抑制曲线；
图18为AST和Palbociclib固定浓度下，单药和联用药对HT-29细胞增殖的抑制率(其中每一组柱状图中，左边是AST单药，中间是Palbociclib单药，右边是AST与Palbociclib联用)；
图19为AST单独或联用Adavosertib对H460细胞系的增殖抑制曲线；
图20为AST和Adavosertib固定浓度下，单药和联用药对H460细胞增殖的抑制率(其中每一组柱状图中，左边是AST单药，中间是Adavosertib单药，右边是AST与Adavosertib联用)；
图21为AST单独或联用AZD7762对H460细胞系的增殖抑制曲线；
图22为AST和AZD7762固定浓度下，单药和联用药对H460细胞增殖的抑制率(其中每一组柱状图中，左边是AST单药，中间是AZD7762单药，右边是AST与AZD7762联用)；
图23为AST单独或联用Palbociclib对H460细胞系的增殖抑制曲线；
图24为AST和Palbociclib固定浓度下，单药和联用药对H460细胞增殖的抑制率(其中每一组柱状图中，左边是AST单药，中间是Palbociclib单药，右边是AST与Palbociclib联用)；
图25为AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制曲线，其中最下方曲线为AST-3424单药组；
图26为另一次实验中AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制曲线，其中最下方曲线为AST-3424单药组；
图27为Ceralasertib与AST-3424加药顺序不同实验中AST-3424单独或联用Ceralasertib对HT29细胞系的增殖抑制曲线，其中最下方曲线为AST-3424单药组；
图28为TH-302单独或联用Adavosertib对HT29细胞系的增殖抑制曲线，其中＜0.01％O2表示乏氧条件，最左上方曲线为TH-302与10μM Adavosertib在乏氧条件下的联用药组；
图29为TH-302单独或联用Adavosertib对H460细胞系的增殖抑制曲线，其中＜0.01％O2表示乏氧条件，TH-302与30μM Adavosertib在乏氧条件下的联用药组在左边；
图30为TH-302单独或联用Ceralasertib对HT29细胞系的增殖抑制曲线，其中＜0.01％O2表示乏氧条件，最左上方曲线为TH-302与2000μM Ceralaserti在乏氧条件下的联用药组；
图31为TH-302单独或联用Ceralasertib对H460细胞系的增殖抑制曲线，其中＜0.01％ O2表示乏氧条件，TH-302常氧单药组在右边。
附图中的外文与对应的中文释义对照表
在部分图的头部有相关实验的条件说明，以图1为例，其图的头部部分的“HT-29_Adavosertib-2h-pretreatment AST-3424-72h-cotreatment 122421”，各部分含义如下：
HT-29表示在HT-29细胞系中的实验；
Adavosertib-2h-pretreatment AST-3424-72h-cotreatment表示，联合用药中，先用Adavosertib处理2h，然后加入AST-3424共同处理72h；单独用药中，只加入AST-3424，然后处理72h；
122421表示实验编号。
其他附图的类似的头部部分，使用了不同的细胞系、测试化合物或实验编号，其含义参照上述解释。
实施例
AST-3424化学结构式为
其合成方法参见专利申请文本PCT/US2016/062114，公开号WO2017087428A1(对应中国申请号2016800200132，公开号CN108136214A)。
AST化学结构式为
其合成方法参见专利申请文本PCT/CN2020/089692，公开号WO2020228685A1。
TH-302化学结构式为
其合成方法参见专利申请文本CN2012102515573，公开号CN102746336A。
Adavosertib(别名AZD1775、MK-1775)是一种选择性Wee1激酶抑制剂，Wee1激酶是参与细胞周期G2/M检查点和DNA损伤修复过程的关键激酶。细胞周期蛋白依赖性激酶 (Cyclin-dependent kinases,CDKs)通过使特异底物磷酸化调节细胞周期，其活性依赖于与周期素Cyclin结合形成复合物，并且其活性还受到磷酸化调控，CDK1的磷酸化状态进一步控制G2/M检查点。Wee1是细胞周期G2/M检查点的“守门员”，允许细胞在进入有丝分裂之前进行DNA损伤修复。Wee1通过磷酸化CDK1Tyr15，使CDK1维持在非活性状态，从而抑制CDK1-cyclin B复合体的活性，使细胞分裂停滞在G2/M检查点，负调控细胞周期，其生物学意义是使没有来得及修复的DNA损伤被修复，防止细胞携带DNA损伤就进入有丝分裂期。Adavosertib通过选择性抑制Wee1激酶，阻滞p53等基因缺陷型肿瘤在G2/M期检查点的DNA损伤修复，导致肿瘤细胞死亡，最终达到治疗肿瘤的目的，其结构式为
可直接通过商业化的试剂公司购买得到。
AZD7762是一种有效的ATP竞争性的细胞周期检测点激酶(checkpoint kinase，CHk)抑制剂，消除DNA损伤诱导的S和G2检查点，属于CHK抑制剂，其结构式为
可直接通过商业化的试剂公司购买得到。
Palbociclib是一种具有口服活性的CDK4和CDK6选择性抑制剂，结构式为
可直接通过商业化的试剂公司购买得到。
Ceralasertib(AZD6738)是一种有效的ATR激酶抑制剂，控制G2/M检查点，其结构式为
可直接通过商业化的试剂公司购买得到。
一、体外细胞实验
在p53野生型(H460)和p53缺失(HT-29)细胞系中，通过CTG细胞增殖实验检测细胞周期抑制剂是否能够通过抑制肿瘤细胞周期中DNA修复的关键节点来增强DNA烷化剂前药AST、AST-3424、TH-302的体外细胞毒性。
CTG细胞增殖实验实验步骤如下：
1)将H460或者HT-29细胞悬液加入96孔板，每孔100μL，细胞密度分别为2000/孔或10000/孔。
2)细胞在37℃，5％CO2培养箱中培养过夜。
3)化合物处理
联合用药：分别加入1μL Adavosertib(四个浓度)，AZD7762(四个浓度)，Palbociclib(四个浓度)和Ceralasertib(三个浓度)，到每一个孔中，轻轻震荡确保混合均匀，放入培养箱2小时后，加入1μL不同浓度的测试化合物AST-3424或者AST，轻轻震荡确保混合均匀，然后放入37℃，5％CO2培养箱中。
单独用药：细胞铺板24小时以后，每孔补99μL的生长培养基。加入1μL不同浓度的测试化合物AST-3424或者AST，轻轻震荡确保混合均匀，然后放入37℃，5％CO2培养箱中。
4)将细胞板放置培养箱72小时。
5)将细胞待测板放置室温平衡30分钟，每孔弃掉100μL培养基。
6)每孔加入25μL CTG试剂，放置快速振荡器振荡2分钟，室温避光放置30分钟。
7)用多功能酶标仪读取化学发光信号值，读值时间1000ms。
8)用GraphPad Prism 5software计算IC50，得到化合物的IC50(半数抑制浓度)。
1、AST-3424单独或联用Adavosertib对HT-29细胞系的增殖抑制实验结果
根据图1的不同浓度下的增殖抑制率数值实验结果，拟合得到AST-3424单独或联用Adavosertib对HT-29细胞系的增殖抑制IC50值，结果如下表1所示，AST-3424与不同浓度的Adavosertib联用后，联用药的IC50值显著小于AST-3424的单药IC50值，且呈现一定程度的Adavosertib剂量依赖性。
表1：AST-3424单独或联用Adavosertib对HT-29细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST-3424与AST-3424联用Adavosertib时IC50的比值。
我们进一步对图1的数据进行处理，统计几个固定浓度下，即AST-3424和Adavosertib两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST-3424和Adavosertib联用后的抑制率。结果如图2所示。
实验小结：上述结果显示，在p53缺失(HT-29)细胞系中，不同浓度的AST-3424分别与不同浓度的细胞周期抑制剂Adavosertib的联用均有显著的细胞毒性叠加效应；并且这种叠加效应在低浓度AST-3424低浓度的细胞周期抑制剂Adavosertib的作用下依然显著。
2、AST-3424单独或联用AZD7762对HT-29细胞系的增殖抑制实验结果
根据图3的不同浓度下的增殖抑制率数值实验结果，拟合得到AST-3424单独或联用AZD7762对HT-29细胞系的增殖抑制IC50值，结果如下表2所示，AST-3424与不同浓度的AZD7762联用后，联用药的IC50值显著小于AST-3424的单药IC50值，且呈现一定程度的AZD7762剂量依赖性。
表2：AST-3424单独或联用AZD7762对HT-29细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST-3424与AST-3424联用AZD7762时IC50的比值。
进一步对图3的数据进行处理，统计几个固定浓度下，即AST-3424和AZD7762两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST-3424和AZD7762联用后的抑制率。结果如图4所示。
实验小结：上述结果显示，在p53缺失(HT-29)细胞系中，不同浓度的AST-3424分别与不同浓度的细胞周期抑制剂AZD7762的联用均有显著的细胞毒性叠加效应；并且这种叠加效应在低浓度AST-3424和低浓度的细胞周期抑制剂AZD7762的作用下依然显著。
3、AST-3424单独或联用Palbociclib对HT-29细胞系的增殖抑制实验结果
根据图5的实验结果，拟合得到AST-3424单独或联用Palbociclib对HT-29细胞系的增殖抑制IC50值，结果如下表3所示，AST-3424与不同浓度的Palbociclib联用后，联用药的IC50值小于AST-3424的单药IC50值，且呈现一定程度的Palbociclib剂量依赖性。
表3：AST-3424单独或联用Palbociclib对HT-29细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST-3424与AST-3424联用Palbociclib时IC50的比值。
进一步对图5的数据进行处理，统计几个固定浓度下，即AST-3424和AZD7762两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST-3424和AZD7762联用后的抑制率。结果如图6所示。
实验小结：上述结果显示，在p53缺失(HT-29)细胞系中，不同浓度的AST-3424分别与不同浓度的细胞周期抑制剂Palbociclib的联用均有显著的细胞毒性叠加效应；并且这种叠加效应在低浓度AST-3424和低浓度的细胞周期抑制剂Palbociclib的作用下依然显著。
4、AST-3424单独或联用Adavosertib对H460细胞系的增殖抑制实验结果
根据图7的实验结果，拟合得到AST-3424单独或联用Adavosertib对H460细胞系的增殖抑制IC50值，结果如下表4所示，AST-3424与不同浓度的Adavosertib联用后，高浓度的Adavosertib联用药的IC50值与AST-3424的单药IC50值有2倍左右的差异，其他浓度Adavosertib联用药的IC50值与AST-3424的单药IC50值无显著差异。
表4：AST-3424单独或联用Adavosertib对H460细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST-3424与AST-3424联用Adavosertib时IC50的比值。
进一步对图7的数据进行处理，统计几个固定浓度下，即AST-3424和Adavosertib两个单药对H460细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST-3424和Adavosertib联用后的抑制率。结果如图8所示。
实验小结：上述结果显示，在p53野生型(H460)细胞系中，不同浓度的AST-3424分别与不同浓度的细胞周期抑制剂Adavosertib的联用，高浓度的化合物联用有一定程度的细胞毒性叠加效应；这种叠加效应在低浓度AST-3424和低浓度的细胞周期抑制剂Adavosertib的作用下不显著。
5、AST-3424单独或联用AZD7762对H460细胞系的增殖抑制实验结果
根据图9的实验结果，拟合得到AST-3424单独或联用AZD7762对H460细胞系的增殖抑制IC50值，结果如下表5所示，AST-3424与不同浓度的AZD7762联用后，160nM AZD7762联用药的IC50值与AST-3424的单药IC50值有2倍左右的差异，其他浓度AZD7762联用药的IC50值与AST-3424的单药IC50值无显著差异。
表5：AST-3424单独或联用AZD7762对H460细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST-3424与AST-3424联用AZD7762时IC50的比值。
进一步对图9的数据进行处理，统计几个固定浓度下，即AST-3424和AZD7762两个单药对H460细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC2 5)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST-3424和AZD7762联用后的抑制率。结果如图10所示。
实验小结：上述结果显示，在p53野生型(H460)细胞系中，不同浓度的AST-3424分别与不同浓度的细胞周期抑制剂AZD7762的联用，中高浓度的化合物联用有一定程度的细胞毒性叠加效应；这种叠加效应在低浓度AST-3424和低浓度的细胞周期抑制剂AZD7762的作用下不显著。
6、AST-3424单独或联用Palbociclib对H460细胞系的增殖抑制实验结果
根据图11的实验结果，拟合得到AST-3424单独或联用Palbociclib对H460细胞系的增殖抑制IC50值，结果如下表6所示，AST-3424与不同浓度的Palbociclib联用后，联用药的IC50值与AST-3424的单药IC50值无显著差异。
表6：AST-3424单独或联用Palbociclib对H460细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST-3424与AST-3424联用Palbociclib时IC50的比值。
进一步对图11的数据进行处理，统计几个固定浓度下，即AST-3424和Palbociclib两个单药对H460细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST-3424和Palbociclib联用后的抑制率。结果如图12所示。
实验小结：上述结果显示，在p53野生型(H460)细胞系中，不同浓度的AST-3424分别与不同浓度的细胞周期抑制剂Palbociclib的联用，各个药物浓度下均未表现出细胞毒性叠加效应。
7、AST单独或联用Adavosertib对HT-29细胞系的增殖抑制实验结果
根据图13的实验结果，拟合得到AST单独或联用Adavosertib对HT-29细胞系的增殖抑制IC50值，结果如下表7所示，AST与不同浓度的Adavosertib联用后，联用药的IC50值显著小于AST-3424的单药IC50值，且呈现一定程度的Adavosertib剂量依赖性。
表7：AST单独或联用Adavosertib对HT-29细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST与AST联用Adavosertib时IC50的比值。
我们进一步对图13的数据进行处理，统计几个固定浓度下，即AST和Adavosertib两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST和Adavosertib联用后的抑制率。结果如图14所示。
实验小结：上述结果显示，在p53缺失(HT-29)细胞系中，不同浓度的AST分别与不同浓度的细胞周期抑制剂Adavosertib的联用均有显著的细胞毒性叠加效应；并且这种叠加效应在低浓度AST-3424低浓度的细胞周期抑制剂Adavosertib的作用下依然显著。
8、AST单独或联用AZD7762对HT-29细胞系的增殖抑制实验结果
根据图15的实验结果，拟合得到AST单独或联用AZD7762对HT-29细胞系的增殖抑制IC50值，结果如下表8所示，AST与不同浓度的AZD7762联用后，联用药的IC50值显著小于AST-3424的单药IC50值。
表8：AST单独或联用AZD7762对HT-29细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，表示单用AST与AST联用AZD7762时IC50的比值。
进一步对图15的数据进行处理，统计几个固定浓度下，即AST和AZD7762两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST和AZD7762联用后的抑制率。结果如图16所示。
实验小结：上述结果显示，在p53缺失(HT-29)细胞系中，不同浓度的AST分别与不同浓度的细胞周期抑制剂AZD7762的联用均有显著的细胞毒性叠加效应；并且这种叠加效应在低浓度AST-3424低浓度的细胞周期抑制剂AZD7762的作用下依然显著。
9、AST单独或联用Palbociclib对HT-29细胞系的增殖抑制实验结果
根据图17的实验结果，拟合得到AST单独或联用Palbociclib对HT-29细胞系的增殖抑制IC50值，结果如下表9所示，AST与不同浓度的Palbociclib联用后，联用药的IC50值显著与AST-3424的单药IC50值无差异。
表9：AST单独或联用Palbociclib对HT-29细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，single/combo表示单用AST与AST联用Palbociclib时IC50的比值。
进一步对图17的数据进行处理，统计几个固定浓度下，即AST和Palbociclib两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST和Palbociclib联用后的 抑制率。结果如图18所示。
实验小结：上述结果显示，在p53缺失(HT-29)细胞系中，不同浓度的AST分别与不同浓度的细胞周期抑制剂Palbociclib的联用，在低浓度的药物联用条件下显现出一定程度的细胞毒性叠加效应。
10、AST单独或联用Adavosertib对H460细胞系的增殖抑制实验结果
根据图19的实验结果，拟合得到AST单独或联用Adavosertib对H460细胞系的增殖抑制IC50值，结果如下表10所示，AST与不同浓度的Adavosertib联用后，联用药的IC50值与AST-3424的单药IC50值无显著差异。
表10：AST单独或联用Adavosertib对H460细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo，single/combo表示单用AST与AST联用Adavosertib时IC50的比值。
进一步对图19的数据进行处理，统计几个固定浓度下，即AST和Adavosertib两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST和Adavosertib联用后的抑制率。结果如图20所示。
实验小结：上述结果显示，在p53野生型(H460)细胞系中，不同浓度的AST分别与不同浓度的细胞周期抑制剂Adavosertib的联用未表现出毒性叠加效应。
11、AST单独或联用AZD7762对H460细胞系的增殖抑制实验结果
根据21的实验结果，拟合得到AST单独或联用AZD7762对H460细胞系的增殖抑制IC50值，结果如下表11所示，AST与不同浓度的AZD7762联用后，160nM AZD7762联用药的IC50值与AST的单药IC50值有3倍左右的差异，其他浓度AZD7762联用药的IC50值与AST的单药IC50值无显著差异。
表11：AST单独或联用AZD7762对H460细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo表示单用AST与AST联用AZD7762时IC50的比值。
进一步对图21的数据进行处理，统计几个固定浓度下，即AST和AZD7762两个单药对HT-29细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST和AZD7762联用后的 抑制率。结果如图22所示。
实验小结：上述结果显示，在p53野生型(H460)细胞系中，不同浓度的AST分别与不同浓度的细胞周期抑制剂AZD7762的联用，160nM AZD7762的化合物联用有一定程度的细胞毒性叠加效应，其它浓度的药物联用均未显示出细胞毒性的叠加效应。
12、AST单独或联用Palbociclib对H460细胞系的增殖抑制实验结果
根据图23的实验结果，拟合得到AST单独或联用Palbociclib对H460细胞系的增殖抑制IC50值，结果如下表12所示，AST与不同浓度的Palbociclib联用后，联用药的IC50值与AST的单药IC50值无显著差，且大于单药的IC50值。
表12：AST单独或联用Palbociclib对H460细胞系的增殖抑制IC50值
注释说明：Compounds，化合物或化合物联用组合；ratio，single/combo表示单用AST与AST联用Palbociclib时IC50的比值。
进一步对图23的数据进行处理，统计几个固定浓度下，即AST和Palbociclib两个单药对H460细胞的抑制率分别达到50％左右的药物浓度(IC50)、25％左右的药物浓度(IC25)、10％左右的药物浓度(IC10)和0％左右的药物浓度(IC0)时，AST和Palbociclib联用后的抑制率。结果如图24所示。
实验小结：上述结果显示，在p53野生型(H460)细胞系中，不同浓度的AST分别与不同浓度的细胞周期抑制剂Palbociclib的联用，各个药物浓度下均未表现出细胞毒性叠加效应。
13、AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制实验结果
进行第一次实验，得到图25的AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制曲线，根据图25的实验结果，拟合得到AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制IC50值，结果如下表13所示。
表13：AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制IC50值(第一次实验)
注：Ratio表示比率。
进行第二次重复实验，得到图26的AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制曲线，根据图26的实验结果，拟合得到AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制IC50值，结果如下表14所示。
表14：AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制IC50值(第二次实验)
注：Ratio表示比率。
为了进一步探索加药顺序对联用效果的影响，改变加药顺序为先加Ceralasertib，后加AST-3424，实验结果如图27所示增殖抑制曲线，根据图27的实验结果，拟合得到AST-3424单独或联用Ceralasertib对HT29/H460细胞系的增殖抑制IC50值，结果如下表15所示。
表15：不同加药顺序下，AST-3424单独或联用Ceralasertib对HT29细胞系的增殖抑制IC50值
注：Compounds，化合物或化合物联用组合；ratio，single/combo表示单用AST与AST联用Palbociclib时IC50的比值。
实验小结：
AST-3424和Ceralasertib(ATR抑制剂)联用，对H460细胞(p53野生型)的增殖有较显著的叠加效应。
AST-3424分别和各浓度Ceralasertib(ATR抑制剂)联用，对HT29细胞(p53缺失)的增殖有非常显著的叠加效应，且叠加效应强于H460细胞系。
不同加药顺序对AST-3424和Ceralasertib联用的细胞毒性叠加效应无差异。
化合物AST与AST-3424的体外细胞实验总结：
(1)在p53缺失(HT-29)细胞系中，AST或AST-3424分别与细胞周期抑制剂Adavosertib、AZD7762、Palbociclib和Ceralasertib的联用均有显著的细胞毒性叠加效应；并且这种叠加效应在低浓度AST或AST-3424分别与低浓度的细胞周期抑制剂的作用下依然显著。
(2)在p53野生型(H460)细胞中，中高浓度的AST或AST-3424分别与Adavosertib、AZD7762、Ceralasertib联用，有一定程度的细胞毒性叠加效应；这种叠加效应在低浓度 AST或AST-3424分别与低浓度的细胞周期抑制剂的作用下不显著。
(3)在p53野生型(H460)细胞中，AST-3424或AST与Palbociclib联用，均没有表现出显著的细胞毒性叠加效应。
以下进一步考察TH-302单独或联用Adavosertib/Ceralasertib对HT-29/H460细胞系的增殖抑制实验结果。在乏氧、常氧的不同试验条件下，实验方法如下：
1)将HT-29细胞悬液加入两类24孔板中，每孔495μL，细胞密度为5×104/孔。插有glass insert的24孔板用于乏氧实验，普通塑料24孔板用于常氧实验。
2)细胞在37℃，5％CO2培养箱中培养过夜。
3)化合物处理
乏氧条件(O2<0.01％)：
调节低氧工作站至乏氧环境，并利用氧气指示剂确认工作站中的乏氧情况。
细胞铺板24小时以后，将带有玻璃插件的24孔板送入低氧工作站。
将24孔板放至在振荡器上，打开孔板盖振荡，进行气体交换5分钟。
联用板每孔先去掉5μL或者10μL培养液，再加入5μLAdavosertib(终浓度分别为1μM、10μM)或Ceralasertib(终浓度分别为200μM、2000μM)孵育2h后。
每孔加入5μL或者10μL 100X相应浓度化合物，每个实验组3个复孔。
轻轻震荡确保化合物混合均匀，将24孔板盖半打开，并在乏氧工作站内孵育3小时。
常氧条件(O221％)：
细胞铺板24小时以后，联用板每孔先去掉5μL或者10μL培养液，再加入5μL Adavosertib(终浓度分别为1μM、10μM)或Ceralasertib(终浓度分别为200μM、2000μM)孵育2h后。
每孔加入5μL或者10μL 100倍相应浓度化合物，每个实验组3个复孔。
轻轻震荡确保化合物混合均匀，将24孔板在37℃，5％CO2标准培养箱内孵育3小时。
4)化合物共同处理3小时后，所有24孔板用培养基洗2次，每孔、每次500μL。
5)每孔加入500μL培养基。
6)放置37℃，5％CO2培养箱72小时。
7)每孔弃去300μL培养基，加入50μL CTG，震荡混匀2min，室温避光放置15分钟。
8)将培养基从24孔板中，每孔转移100μL至96孔白板中。
9)用多功能酶标仪读取化学发光信号值，读值时间1000ms。
10)用GraphPad Prism 5software计算IC50，得到化合物的IC50(半数抑制浓度)。
对H460细胞的增殖抑制实验中，加入Adavosertib的终浓度分别为3μM、30μM两个浓度，加入Ceralasertib的终浓度分别为2μM、20μM、1000μM三个浓度。
14、TH-302单独或联用Adavosertib/Ceralasertib对HT29/H460细胞系的增殖抑制实验结果
根据上述实验方法，得到TH-302单独或联用Adavosertib在常氧和乏氧条件下对HT29/H460细胞系的IC50值，如表16、表17所示，相应的增殖抑制曲线如图28、图29所示。
表16：TH-302单独或联用Adavosertib对HT29细胞系在常氧、乏氧条件下增殖抑制实验结果

注：Compounds表示化合物或化合物联用组合；Ratio表示比率。
表17：TH-302单独或联用Adavosertib对H460细胞系的增殖抑制实验结果
注：Compounds表示化合物或化合物联用组合；Ratio表示比率。
根据上述实验方法，得到TH-302单独或联用Ceralasertib在常氧和乏氧条件下对HT29/H460细胞系的IC50值，如表18、表19所示，相应的增殖抑制曲线如图30、图31所示。
表18：TH-302单独或联用Ceralasertib对HT29细胞系的增殖抑制实验结果
注：Compounds表示化合物或化合物联用组合；Ratio表示比率。
表19：TH-302单独或联用Ceralasertib对H460细胞系的增殖抑制实验结果
注：Compounds表示化合物或化合物联用组合；Ratio表示比率。
化合物TH-302的体外细胞实验总结
(1)在p53缺失(HT29)细胞系中，TH-302与G2/M细胞周期调节剂Adavosertib，在常氧条件下联用时，无显著的细胞毒性增强效应。
(2)在p53缺失(HT29)细胞系中，TH-302与G2/M细胞周期调节剂Adavosertib，在乏氧条件下联用时，与高浓度的Adavosertib联用，显示出显著的细胞毒性增强效应。
(3)在p53野生型(H460)细胞中，TH-302与G2/M细胞周期调节剂Adavosertib，无论在常氧或乏氧条件下联用时，均无显著的细胞毒性增强效应。
(4)在p53缺失(HT29)细胞系中，TH-302与ATR激酶抑制剂Ceralasertib联用，在常氧条件下联用时，无显著的细胞毒性增强效应。
(5)在p53缺失(HT29)细胞系中，TH-302与ATR激酶抑制剂Ceralasertib联用，在乏氧条件下联用时，高浓度的Ceralasertib联用，显示出显著的细胞毒性增强效应。
(6)在p53野生型(H460)细胞中，TH-302与ATR激酶抑制剂Ceralasertib联用，无论在常氧或乏氧条件下联用时，均无显著的细胞毒性增强效应。
上述实施例中，化合物AST-3424、AST与TH-302均是DNA烷化剂前药，其在特定环境下能够特异性释放出DNA烷化剂小分子AST-2660(或称2660)或Br-IPM。
以AST-3424、TH-302为例：
这些化合物最终释放出的DNA烷化剂是细胞周期非特异性的，即他们作用于细胞周期各阶段的DNA双链结构。烷化剂可以使DNA双链之间形成非常紧密的结合，导致DNA无法完成转录、翻译和复制等过程，也就是说他们是对处于所有不同细胞周期的细胞均能产生DNA损伤和增殖抑制作用。
而化合物Adavosertib是一种选择性Wee1激酶抑制剂，Wee1激酶是参与细胞周期G2/M检查点和DNA损伤修复过程的关键激酶。细胞周期蛋白依赖性激酶(CDKs)通过使特异底物磷酸化调节细胞周期，其活性依赖于与周期素Cyclin结合形成复合物，并且其活性还受到磷酸化调控，CDK1的磷酸化状态进一步控制G2/M检查点。Wee1通过磷酸化CDK1Tyr15，使CDK1维持在非活性状态，从而抑制CDK1-cyclin B复合体的活性，使细胞分裂停滞在G2/M检查点，负调控细胞周期，其生物学意义是使没有来得及修复的DNA损伤被 修复，防止细胞携带DNA损伤就进入有丝分裂期。也就是说，Adavosertib通过抑制Wee1激酶来调控CDK1的磷酸化状态，从而调控细胞周期进程，也就是说Adavosertib是间接作用于CDK的抑制剂，可以认为是广义的CDK抑制剂。Palbociclib是一种具有口服活性的CDK4和CDK6选择性抑制剂。因此，Adavosertib、Palbociclib作为CDK抑制剂均是细胞周期特异性的，作用于细胞周期的特定阶段。
大量文献报道，p53蛋白对细胞周期G1/S期的DNA修复具有重要的调控作用，p53蛋白缺失的细胞在G1/S期的DNA修复受到抑制，因此在这类细胞中使用G2/M细胞周期停滞抑制剂时，可以显著抑制DNA在G2/M周期中的DNA修复能力。
本申请的实验揭示了中高浓度的Adavosertib与AST、AST-3424联用对肿瘤细胞系具有显著的细胞毒性叠加作用，而对于p53缺失的肿瘤细胞系在低浓度的情形下联用即表现显著的细胞毒性叠加作用；Palbociclib与AST、AST-3424共同作用只对p53缺失的肿瘤细胞系具有显著的细胞毒性叠加作用；高浓度的Adavosertib与TH-302联用只对p53缺失的肿瘤细胞系具有显著的细胞毒性叠加作用。
基于以上实验事实，本领域技术人员基于AST、AST-3424、TH-302均为结构通式(1)-(9)涵盖的DNA烷化剂作为最终作用于DNA双链交联过程的前药化合物，以及Adavosertib、Palbociclib均为作用于特定细胞周期的CDK抑制剂，可以推定：烷化剂前药化合物与作用于特定细胞周期的CDK抑制剂对于肿瘤细胞系具有细胞毒性叠加作用，即两种药物联用治疗癌症、肿瘤患者将具有更好的治疗效果，而且p53缺失将增强上述叠加作用/联用效果。
AZD7762是一种有效的ATP竞争性的细胞周期检测点激酶(checkpoint kinase，Chk)抑制剂，消除DNA损伤诱导的S和G2检查点，属于ChK抑制剂。AZD7762作为ChK抑制剂是细胞周期特异性的，作用于细胞周期的特定阶段。
本申请的实验揭示了中高浓度的AZD7762与AST、AST-3424联用对肿瘤细胞系具有显著的细胞毒性叠加作用，而对于p53缺失的肿瘤细胞系在低浓度的情形下联用即表现显著的细胞毒性叠加作用。
基于以上实验事实，本领域技术人员基于AST、AST-3424均为结构通式(1)-(9)涵盖的DNA烷化剂作为最终作用于DNA双链交联过程的前药化合物，以及AZD7762为作用于特定细胞周期的ChK抑制剂，可以推定：烷化剂前药化合物与作用于特定细胞周期的CHK抑制剂对于肿瘤细胞系具有细胞毒性叠加作用，即两种药物联用治疗癌症、肿瘤患者将具有更好的治疗效果，而且p53缺失将增强上述叠加作用/联用效果。
Ceralasertib(AZD6738)是一种有效的ATR激酶抑制剂，是细胞周期特异性的，作用于细胞周期的特定阶段。
本申请的实验揭示了中高浓度的Ceralasertib与AST、AST-3424联用对肿瘤细胞系具有显著的细胞毒性叠加作用，而对于P53缺失的肿瘤细胞系在低浓度的情形下联用即表现显著的细胞毒性叠加作用；高浓度的Ceralasertib与TH-302联用只对p53缺失的肿瘤细胞系具有显著的细胞毒性叠加作用。
基于以上实验事实，本领域技术人员基于AST、AST-3424、TH-302均为结构通式(1)-(9)涵盖的DNA烷化剂作为最终作用于DNA双链交联过程的前药化合物，以及Ceralasertib为作用于特定细胞周期的ATR抑制剂，可以推定：烷化剂前药化合物与作用于特定细胞周期的ATR抑制剂对于肿瘤细胞系具有细胞毒性叠加作用，即两种药物联用治疗癌症、肿瘤患者将具有更好的治疗效果，而且p53缺失将增强上述叠加作用/联用效果。
更进一步，可以推定细胞周期非特异性的烷化剂前药化合物与作用于特定细胞周期的细胞周期抑制剂联用对于肿瘤细胞具有细胞毒性叠加作用，即两种药物联用治疗癌症、肿 瘤患者将具有更好的治疗效果；而且更进一步的，p53缺失将增强上述叠加作用/联用效果，即具有p53基因缺失的患者可能在联用后得到更多治疗受益。

Claims (11)

  1. 治疗方法，使用含有烷化剂前药化合物及其盐、酯、溶剂合物、同位素异构体的药物与含有细胞周期抑制剂化合物及其盐、酯、溶剂合物、同位素异构体的药物联用治疗癌症、肿瘤患者。
  2. 根据权利要求1所述的治疗方法，所述细胞周期抑制剂选自CDK抑制剂、WEE抑制剂、CHK抑制剂、ATR抑制剂。
  3. 根据权利要求2所述的治疗方法，所述烷化剂前药化合物选自AKR1C3酶活化烷化剂前药化合物、乏氧活化烷化剂前药化合物，
    优选的，AKR1C3酶活化烷化剂前药化合物与CDK抑制剂、WEE抑制剂、CHK抑制剂、ATR抑制剂联用；
    乏氧活化烷化剂前药化合物与CDK抑制剂、WEE抑制剂、ATR抑制剂联用。
  4. 根据权利要求1所述的治疗方法，其中，烷化剂前药化合物包括化学结构式(1)-(9)：
    其中，R各自独立地选自H、-CH3、-CH2CH3、-CF3，X各自独立地选自Cl、Br、MsO、TsO等离去官能团；
    其中，R1、R2、R3、Cx的定义如专利申请PCT/CN2020/114519，公开号WO2021120717A1中的权利要求书所记载；

    其中，R1、R2、R3、R4、R5、R6、R7、R8、R9、R10、R11、R12、R13、R14、R15、R16、R17的定义如专利申请PCT/US2016/039092，公开号WO2016210175A1(对应中国申请号2016800368985，公开号CN108024974A)中的权利要求书所记载；
    其中，X、Y、Z、R、T、A以及X10的定义如专利申请PCT/US2016/021581，公开号WO2016145092A1(对应中国申请号2016800150788，公开号CN107530556A)中的权利要求书所记载；
    其中，R1、R2、R3、R4、R5、R8、R9、R10的定义如专利申请PCT/CN2020/089692，公开号WO2020228685A1中的权利要求书所记载；
    其中：
    A是取代或未经取代的C6-C10的芳基、联芳基或取代的联芳基、5-15元的杂芳基或-N＝CR1R2，其中取代时的取代基选自由以下组成的群：卤基、-CN、-NO2、–O-(CH2)-O-、-CO2H及其盐、-OR100、-CO2R100、-CONR101R102、-NR101R102、-NR100SO2R100、-SO2R100、-SO2NR101R10 2、C1-C6烷基、C3-C10杂环基；
    其中，R100、R101及R102各自独立是氢、C1-C8烷基、C6-C12芳基；或R101及R102与其附接至的氮原子一起形成5-7元杂环；
    其中烷基及芳基各自是经1-3个卤基或1-3个C1-C6烷基取代；
    R1及R2各自独立是苯基或甲基；
    X、Y及Z各自独立是氢或卤基；
    R是氢或C1-C6烷基或卤素取代烷基；
    其中，Rw的定义如专利申请PCT/CN2020/120281，公开号WO2021068952A1中的权利要求书所记载；
    其中，A、E、G、X、Y的定义如专利申请PCT/NZ2019/050030，公开号WO2019190331A1(对应中国申请号2019800234236，公开号CN111918864A)中的权利要求书所记载。
  5. 根据权利要求1或2所述的治疗方法，其中，
    所述癌症、肿瘤患者或其生物样本被检查出具有特定基因缺失或损伤，
    所述特定基因选自参与细胞周期Checkpoint调控的基因，优选p53、p21、CCNB1、WIP1、14-3-3 sigma protein、cdc2/cycB。
  6. 根据权利要求3所述的治疗方法，其中，
    CDK抑制剂选自哌柏西利(Palbociclib)、瑞波西利(Ribociclib)、阿贝西利(Abemaciclib)、曲拉西利(Trilaciclib)、达尔西利(dalpiciclib)、Adavosertib、Ro-3306、Dinaciclib、Cirtuvivint、Rintodestrant、DS96432529、THZ1、THZ531、Seliciclib、Flavopiridol、AZD4573、SR-4835、Simurosertib、Fadraciclib、NVP-2、SNS-032、(E/Z)-Zotiraciclib、AZD-5438、AT7519、Mevociclib、Kenpaullone、YKL-5-124 TFA、NG 52、GSK 3 Inhibitor IX、OTS964、Samuraciclib、Flavopiridol、KB-0742 dihydrochloride、(+)-Enitociclib、AUZ 454、SY-5609、SEL120-34A monohydrochloride、CCT-251921、MBQ-167、XL413 hydrochloride、BI-1347、THAL-SNS-032、JNJ-7706621、TG003、LDC4297、BMS-265246、Roniciclib、CGP60474、(R)-CR8 trihydrochloride、R547、Milciclib、T025、AS2863619、Senexin A、BSJ-4-116、CLK-IN-T3、CDK12-IN-3、CVT-313、Atuveciclib、PHA-793887、Indirubin-3'-monoxime、YKL-5-124、PHA-767491 hydrochloride、KH-CB19、Cucurbitacin E、 Purvalanol A、BSJ-03-204、ON123300、CDK5 inhibitor 20-223、Riviciclib、FN-1501、CP-10、THZ2、Abemaciclib metabolite M2、BS-181、CDK12-IN-E9、Samuraciclib、RGB-286638、CDK2-IN-4、LDC000067、ML167、Purvalanol B、NU6300、CLK1-IN-1、FMF-04-159-2、CKI-7、CDKI-73、MSC2530818、BSJ-04-132、NU6102、Voruciclib、Olomoucine；
    WEE抑制剂选自WEE1-IN-5、WEE1-IN-3、WEE1-IN-4、LEB-03-146、LEB-03-144、Adavosertib、LEB-03-153、LEB-03-145、PD407824、PD0166285、PD0166285 dihydrochloride、Pomalidomide-C3-adavosertib、DB0614、FMF-06-098-1；
    CHK抑制剂选自AZD7762、Prexasertib、SCH900776、GDC-0425、Chk1-IN-6、CCT245737、BML-277、CCT241533、PD407824、CHIR-124、CCT244747、PF477736、GDC-0575、SB-218078、MRT00033659、ANI-7、SAR-020106、CCT241533、CHK1-IN-4、VER-00158411、CHK1-IN-3、Chk1-IN-5、CHK1-IN-2、CHK-IN-1；
    ATR抑制剂选自Ceralasertib、Berzosertib、Gartisertib、BAY1895344、BAY-937、AZ20、ETP-46464、Dactolisib、VE-821、M1774、ATRN-199、RP-3500、ART-0380。
  7. 根据权利要求4所述的治疗方法，其中，
    式(1)的乏氧活化烷化剂前药化合物选自以下结构的化合物：
    式(2)的乏氧活化烷化剂前药化合物选自以下结构的化合物：

    式(3)的乏氧活化烷化剂前药化合物选自以下结构的化合物：







    式(4)的AKR1C3酶活化烷化剂前药化合物选自以下结构的化合物：




    式(6)的AKR1C3酶活化前药化合物选自以下结构的化合物：
    式(5)的AKR1C3酶活化前药化合物选自以下结构的化合物：

    式(7)的AKR1C3酶活化前药化合物选自以下结构的化合物：

    以及






    及
    式(8)的AKR1C3酶活化前药化合物选自以下结构的化合物：
    式(9)的AKR1C3酶活化前药化合物选自以下结构的化合物：











  8. 含有烷化剂前药化合物或其盐、酯、溶剂合物、同位素异构体与细胞周期抑制剂化合物或其盐、酯、溶剂合物、同位素异构体的药物组合物，该药物组合物用于治疗癌症、肿瘤患者。
  9. 根据权利要求8所述的药物组合物，其中，
    所述烷化剂前药化合物选自AKR1C3酶活化烷化剂前药化合物、乏氧活化烷化剂前药化合物，优选为AKR1C3酶活化DNA烷化剂前药化合物、乏氧活化DNA烷化剂前药化合物；
    或
    所述细胞周期抑制剂选自CDK抑制剂、WEE抑制剂、CHK抑制剂、ATR抑制剂；
    或
    烷化剂前药化合物包括权利要求4所述化学结构式(1)-(9)；
    或
    所述癌症、肿瘤患者或其生物样本被检查出具有特定基因缺失或损伤，所述特定基因选自参与细胞周期Checkpoint调控的基因，优选p53、p21、CCNB1、WIP1、14-3-3 sigma protein、cdc2/cycB；
    或
    CDK抑制剂选自哌柏西利(Palbociclib)、瑞波西利(Ribociclib)、阿贝西利(Abemaciclib)、曲拉西利(Trilaciclib)、达尔西利(dalpiciclib)、Adavosertib、Ro-3306、Dinaciclib、Cirtuvivint、Rintodestrant、DS96432529、THZ1、THZ531、Seliciclib、Flavopiridol、AZD4573、SR-4835、Simurosertib、Fadraciclib、NVP-2、SNS-032、(E/Z)-Zotiraciclib、AZD-5438、AT7519、Mevociclib、Kenpaullone、YKL-5-124 TFA、NG 52、GSK 3 Inhibitor IX、OTS964、Samuraciclib、Flavopiridol、KB-0742 dihydrochloride、(+)-Enitociclib、AUZ 454、SY-5609、SEL120-34A monohydrochloride、CCT-251921、MBQ-167、XL413 hydrochloride、BI-1347、THAL-SNS-032、JNJ-7706621、TG003、LDC4297、BMS-265246、Roniciclib、CGP60474、(R)-CR8 trihydrochloride、R547、Milciclib、T025、AS2863619、Senexin A、BSJ-4-116、CLK-IN-T3、CDK12-IN-3、CVT-313、Atuveciclib、PHA-793887、Indirubin-3'-monoxime、YKL-5-124、PHA-767491 hydrochloride、KH-CB19、Cucurbitacin E、Purvalanol A、BSJ-03-204、ON123300、CDK5 inhibitor 20-223、Riviciclib、FN-1501、CP-10、THZ2、Abemaciclib metabolite M2、BS-181、CDK12-IN-E9、Samuraciclib、RGB-286638、CDK2-IN-4、LDC000067、ML167、Purvalanol B、NU6300、CLK1-IN-1、FMF-04-159-2、CKI-7、CDKI-73、MSC2530818、BSJ-04-132、NU6102、Voruciclib、Olomoucine；
    WEE抑制剂选自WEE1-IN-5、WEE1-IN-3、WEE1-IN-4、LEB-03-146、LEB-03-144、Adavosertib、LEB-03-153、LEB-03-145、PD407824、PD0166285、PD0166285 dihydrochloride、Pomalidomide-C3-adavosertib、DB0614、FMF-06-098-1；
    CHK抑制剂选自AZD7762、Prexasertib、SCH900776、GDC-0425、Chk1-IN-6、CCT245737、BML-277、CCT241533、PD407824、CHIR-124、CCT244747、PF477736、GDC-0575、SB-218078、MRT00033659、ANI-7、SAR-020106、CCT241533、CHK1-IN-4、VER-00158411、CHK1-IN-3、Chk1-IN-5、CHK1-IN-2、CHK-IN-1；
    ATR抑制剂选自Ceralasertib、Berzosertib、Gartisertib、BAY1895344、BAY-937、AZ20、ETP-46464、Dactolisib、VE-821、M1774、ATRN-199、RP-3500、ART-0380；
    或
    烷化剂前药化合物选自权利要求7所述具体化合物。
  10. 烷化剂前药化合物或其盐、酯、溶剂合物、同位素异构体在制备与细胞周期抑制剂化合物或其盐、酯、溶剂合物、同位素异构体联用治疗癌症、肿瘤的药物中的用途。
  11. 根据权利要求10所述的用途，其中，
    所述烷化剂前药化合物选自AKR1C3酶活化烷化剂前药化合物、乏氧活化烷化剂前药化合物，优选为AKR1C3酶活化DNA烷化剂前药化合物、乏氧活化DNA烷化剂前药化合物；
    或
    所述细胞周期抑制剂选自CDK抑制剂、WEE抑制剂、CHK抑制剂、ATR抑制剂；
    或
    烷化剂前药化合物包括权利要求4所述化学结构式(1)-(9)；
    或
    所述癌症、肿瘤患者或其生物样本被检查出具有特定基因缺失或损伤，所述特定基因选自参与细胞周期Checkpoint调控的基因，优选p53、p21、CCNB1、WIP1、14-3-3 sigma protein、cdc2/cycB；
    或
    CDK抑制剂选自哌柏西利(Palbociclib)、瑞波西利(Ribociclib)、阿贝西利(Abemaciclib)、曲拉西利(Trilaciclib)、达尔西利(dalpiciclib)、Adavosertib、Ro-3306、Dinaciclib、Cirtuvivint、Rintodestrant、DS96432529、THZ1、THZ531、Seliciclib、Flavopiridol、AZD4573、SR-4835、Simurosertib、Fadraciclib、NVP-2、SNS-032、(E/Z)-Zotiraciclib、AZD-5438、AT7519、Mevociclib、Kenpaullone、YKL-5-124 TFA、NG 52、GSK 3 Inhibitor IX、OTS964、Samuraciclib、Flavopiridol、KB-0742 dihydrochloride、(+)-Enitociclib、AUZ 454、SY-5609、SEL120-34A monohydrochloride、CCT-251921、MBQ-167、XL413 hydrochloride、BI-1347、THAL-SNS-032、JNJ-7706621、TG003、LDC4297、BMS-265246、Roniciclib、CGP60474、(R)-CR8 trihydrochloride、R547、Milciclib、T025、AS2863619、Senexin A、BSJ-4-116、CLK-IN-T3、CDK12-IN-3、CVT-313、Atuveciclib、PHA-793887、Indirubin-3'-monoxime、YKL-5-124、PHA-767491 hydrochloride、KH-CB19、Cucurbitacin E、Purvalanol A、BSJ-03-204、ON123300、CDK5 inhibitor 20-223、Riviciclib、FN-1501、CP-10、THZ2、Abemaciclib metabolite M2、BS-181、CDK12-IN-E9、Samuraciclib、RGB-286638、CDK2-IN-4、LDC000067、ML167、Purvalanol B、NU6300、CLK1-IN-1、FMF-04-159-2、CKI-7、CDKI-73、MSC2530818、BSJ-04-132、NU6102、Voruciclib、Olomoucine；
    WEE抑制剂选自WEE1-IN-5、WEE1-IN-3、WEE1-IN-4、LEB-03-146、LEB-03-144、Adavosertib、LEB-03-153、LEB-03-145、PD407824、PD0166285、PD0166285 dihydrochloride、Pomalidomide-C3-adavosertib、DB0614、FMF-06-098-1；
    CHK抑制剂选自AZD7762、Prexasertib、SCH900776、GDC-0425、Chk1-IN-6、CCT245737、BML-277、CCT241533、PD407824、CHIR-124、CCT244747、PF477736、GDC-0575、SB-218078、MRT00033659、ANI-7、SAR-020106、CCT241533、CHK1-IN-4、VER-00158411、CHK1-IN-3、Chk1-IN-5、CHK1-IN-2、CHK-IN-1；
    ATR抑制剂选自Ceralasertib、Berzosertib、Gartisertib、BAY1895344、BAY-937、AZ20、ETP-46464、Dactolisib、VE-821、M1774、ATRN-199、RP-3500、ART-0380；
    或
    烷化剂前药化合物选自权利要求7所述具体化合物。
PCT/CN2023/095699 2022-05-23 2023-05-23 烷化剂前药与细胞周期抑制剂联用治疗癌症的方法 WO2023226959A1 (zh)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20150005263A1 (en) * 2011-12-22 2015-01-01 Threshold Pharmaceuticals, Inc. Administration of hypoxia activated prodrugs in combination with chk1 inhibitors for treating cancer
CN107530556A (zh) * 2015-03-10 2018-01-02 深圳艾衡昊医药科技有限公司 Dna烷化剂
CN108290911A (zh) * 2015-11-16 2018-07-17 深圳艾衡昊医药科技有限公司 (r)-及(s)-1-(3-(3-n,n-二甲基胺基羰基)苯氧基-4-硝苯基)-1-乙基-n,n’-双(伸乙基)胺基磷酸酯、组合物及其使用及制备方法
CN113853379A (zh) * 2019-05-13 2021-12-28 深圳艾欣达伟医药科技有限公司 含氟化合物及抗癌医药用途

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20150005263A1 (en) * 2011-12-22 2015-01-01 Threshold Pharmaceuticals, Inc. Administration of hypoxia activated prodrugs in combination with chk1 inhibitors for treating cancer
CN107530556A (zh) * 2015-03-10 2018-01-02 深圳艾衡昊医药科技有限公司 Dna烷化剂
CN108290911A (zh) * 2015-11-16 2018-07-17 深圳艾衡昊医药科技有限公司 (r)-及(s)-1-(3-(3-n,n-二甲基胺基羰基)苯氧基-4-硝苯基)-1-乙基-n,n’-双(伸乙基)胺基磷酸酯、组合物及其使用及制备方法
CN113853379A (zh) * 2019-05-13 2021-12-28 深圳艾欣达伟医药科技有限公司 含氟化合物及抗癌医药用途

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
SUN YING, HU HAO, JING XIAODONG, MENG QINGYE, YU BING, CONG HAILIN, SHEN YOUQING: "Co-delivery of Chemotherapeutic Drugs and Cell Cycle Regulatory Agents Using Nanocarriers for Cancer Therapy", SCIENCE CHINA MATERIALS, vol. 64, no. 8, 1 August 2021 (2021-08-01), pages 1827 - 1848, XP093111712, ISSN: 2095-8226, DOI: 10.1007/s40843-020-1627-4 *

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