CN116102534A - 共价PARP PROTACs衍生物及其应用 - Google Patents
共价PARP PROTACs衍生物及其应用 Download PDFInfo
- Publication number
- CN116102534A CN116102534A CN202111319693.7A CN202111319693A CN116102534A CN 116102534 A CN116102534 A CN 116102534A CN 202111319693 A CN202111319693 A CN 202111319693A CN 116102534 A CN116102534 A CN 116102534A
- Authority
- CN
- China
- Prior art keywords
- synthesis
- covalent
- parp
- compound
- reaction
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 229920000776 Poly(Adenosine diphosphate-ribose) polymerase Polymers 0.000 title claims abstract description 31
- 238000011865 proteolysis targeting chimera technique Methods 0.000 title claims abstract description 7
- 108010026668 snake venom protein C activator Proteins 0.000 title claims abstract description 7
- 102100023712 Poly [ADP-ribose] polymerase 1 Human genes 0.000 claims abstract description 32
- 239000002246 antineoplastic agent Substances 0.000 claims abstract description 19
- 229940041181 antineoplastic drug Drugs 0.000 claims abstract description 19
- 101001113440 Homo sapiens Poly [ADP-ribose] polymerase 2 Proteins 0.000 claims abstract description 18
- 102100023652 Poly [ADP-ribose] polymerase 2 Human genes 0.000 claims abstract description 18
- 102000038566 DCAFs Human genes 0.000 claims abstract description 8
- 108091007824 DCAFs Proteins 0.000 claims abstract description 8
- 102000003960 Ligases Human genes 0.000 claims abstract description 8
- 108090000364 Ligases Proteins 0.000 claims abstract description 8
- 238000006243 chemical reaction Methods 0.000 claims description 143
- 239000003814 drug Substances 0.000 claims description 21
- 229940079593 drug Drugs 0.000 claims description 18
- 108010064218 Poly (ADP-Ribose) Polymerase-1 Proteins 0.000 claims description 12
- 108090000623 proteins and genes Proteins 0.000 claims description 12
- 102000004169 proteins and genes Human genes 0.000 claims description 12
- 125000000539 amino acid group Chemical group 0.000 claims description 9
- 238000002360 preparation method Methods 0.000 claims description 9
- 230000015556 catabolic process Effects 0.000 claims description 8
- 238000006731 degradation reaction Methods 0.000 claims description 8
- 150000001335 aliphatic alkanes Chemical group 0.000 claims description 6
- -1 cyanoacetamido group Chemical group 0.000 claims description 6
- 125000005843 halogen group Chemical group 0.000 claims description 6
- 125000001425 triazolyl group Chemical group 0.000 claims description 6
- HRPVXLWXLXDGHG-UHFFFAOYSA-N Acrylamide Chemical group NC(=O)C=C HRPVXLWXLXDGHG-UHFFFAOYSA-N 0.000 claims description 4
- 229910052801 chlorine Inorganic materials 0.000 claims description 4
- 125000001309 chloro group Chemical group Cl* 0.000 claims description 4
- AFOSIXZFDONLBT-UHFFFAOYSA-N divinyl sulfone Chemical group C=CS(=O)(=O)C=C AFOSIXZFDONLBT-UHFFFAOYSA-N 0.000 claims description 4
- 125000003277 amino group Chemical group 0.000 claims description 3
- 125000001033 ether group Chemical group 0.000 claims description 3
- 125000002534 ethynyl group Chemical group [H]C#C* 0.000 claims description 3
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 3
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 3
- 125000000449 nitro group Chemical group [O-][N+](*)=O 0.000 claims description 3
- 125000003396 thiol group Chemical group [H]S* 0.000 claims description 2
- 125000003700 epoxy group Chemical group 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 26
- 101710179684 Poly [ADP-ribose] polymerase Proteins 0.000 abstract description 21
- FAQDUNYVKQKNLD-UHFFFAOYSA-N olaparib Chemical compound FC1=CC=C(CC2=C3[CH]C=CC=C3C(=O)N=N2)C=C1C(=O)N(CC1)CCN1C(=O)C1CC1 FAQDUNYVKQKNLD-UHFFFAOYSA-N 0.000 abstract description 19
- 229960000572 olaparib Drugs 0.000 abstract description 19
- 210000004881 tumor cell Anatomy 0.000 abstract description 12
- 230000002401 inhibitory effect Effects 0.000 abstract description 8
- 239000012661 PARP inhibitor Substances 0.000 abstract description 7
- 229940121906 Poly ADP ribose polymerase inhibitor Drugs 0.000 abstract description 7
- 102100029586 DDB1- and CUL4-associated factor 16 Human genes 0.000 abstract description 5
- 101000917435 Homo sapiens DDB1- and CUL4-associated factor 16 Proteins 0.000 abstract description 5
- 102000006275 Ubiquitin-Protein Ligases Human genes 0.000 abstract description 5
- 108010083111 Ubiquitin-Protein Ligases Proteins 0.000 abstract description 5
- 230000000593 degrading effect Effects 0.000 abstract description 5
- 239000003446 ligand Substances 0.000 abstract description 5
- 239000000203 mixture Substances 0.000 abstract description 5
- 230000035755 proliferation Effects 0.000 abstract description 5
- 238000013461 design Methods 0.000 abstract description 2
- 238000012986 modification Methods 0.000 abstract description 2
- 230000004048 modification Effects 0.000 abstract description 2
- 230000015572 biosynthetic process Effects 0.000 description 210
- 238000003786 synthesis reaction Methods 0.000 description 210
- 150000001875 compounds Chemical class 0.000 description 162
- 238000005481 NMR spectroscopy Methods 0.000 description 96
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 93
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 92
- 239000000543 intermediate Substances 0.000 description 75
- 239000011734 sodium Substances 0.000 description 65
- 210000004027 cell Anatomy 0.000 description 52
- 239000002994 raw material Substances 0.000 description 44
- 239000000047 product Substances 0.000 description 41
- 239000007787 solid Substances 0.000 description 36
- 239000000243 solution Substances 0.000 description 36
- VGCXGMAHQTYDJK-UHFFFAOYSA-N Chloroacetyl chloride Chemical compound ClCC(Cl)=O VGCXGMAHQTYDJK-UHFFFAOYSA-N 0.000 description 31
- 206010028980 Neoplasm Diseases 0.000 description 30
- 238000000034 method Methods 0.000 description 26
- 239000012044 organic layer Substances 0.000 description 22
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 14
- 238000010898 silica gel chromatography Methods 0.000 description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 13
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 description 12
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-dimethylformamide Substances CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 12
- 230000004663 cell proliferation Effects 0.000 description 12
- 102000015087 Poly (ADP-Ribose) Polymerase-1 Human genes 0.000 description 11
- 239000007928 intraperitoneal injection Substances 0.000 description 11
- 239000000463 material Substances 0.000 description 11
- 239000012043 crude product Substances 0.000 description 10
- 230000005764 inhibitory process Effects 0.000 description 10
- 239000002904 solvent Substances 0.000 description 9
- 241000699670 Mus sp. Species 0.000 description 8
- 230000000259 anti-tumor effect Effects 0.000 description 8
- 238000011282 treatment Methods 0.000 description 8
- 239000007821 HATU Substances 0.000 description 7
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical class [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 7
- 238000000605 extraction Methods 0.000 description 7
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- 201000011510 cancer Diseases 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 230000005918 in vitro anti-tumor Effects 0.000 description 6
- 125000005647 linker group Chemical group 0.000 description 6
- 238000012544 monitoring process Methods 0.000 description 6
- 230000037396 body weight Effects 0.000 description 5
- 230000008859 change Effects 0.000 description 5
- 238000011161 development Methods 0.000 description 5
- 230000018109 developmental process Effects 0.000 description 5
- 210000003494 hepatocyte Anatomy 0.000 description 5
- 238000004896 high resolution mass spectrometry Methods 0.000 description 5
- 239000000843 powder Substances 0.000 description 5
- 239000007858 starting material Substances 0.000 description 5
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- 206010006187 Breast cancer Diseases 0.000 description 4
- 208000026310 Breast neoplasm Diseases 0.000 description 4
- 239000003112 inhibitor Substances 0.000 description 4
- 239000003921 oil Substances 0.000 description 4
- DYHSDKLCOJIUFX-UHFFFAOYSA-N tert-butoxycarbonyl anhydride Chemical compound CC(C)(C)OC(=O)OC(=O)OC(C)(C)C DYHSDKLCOJIUFX-UHFFFAOYSA-N 0.000 description 4
- 231100000419 toxicity Toxicity 0.000 description 4
- 230000001988 toxicity Effects 0.000 description 4
- 230000004614 tumor growth Effects 0.000 description 4
- 238000005160 1H NMR spectroscopy Methods 0.000 description 3
- UOXJNGFFPMOZDM-UHFFFAOYSA-N 2-[di(propan-2-yl)amino]ethylsulfanyl-methylphosphinic acid Chemical compound CC(C)N(C(C)C)CCSP(C)(O)=O UOXJNGFFPMOZDM-UHFFFAOYSA-N 0.000 description 3
- SFHYNDMGZXWXBU-LIMNOBDPSA-N 6-amino-2-[[(e)-(3-formylphenyl)methylideneamino]carbamoylamino]-1,3-dioxobenzo[de]isoquinoline-5,8-disulfonic acid Chemical compound O=C1C(C2=3)=CC(S(O)(=O)=O)=CC=3C(N)=C(S(O)(=O)=O)C=C2C(=O)N1NC(=O)N\N=C\C1=CC=CC(C=O)=C1 SFHYNDMGZXWXBU-LIMNOBDPSA-N 0.000 description 3
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- 206010008342 Cervix carcinoma Diseases 0.000 description 3
- 206010009944 Colon cancer Diseases 0.000 description 3
- 208000001333 Colorectal Neoplasms Diseases 0.000 description 3
- 206010033128 Ovarian cancer Diseases 0.000 description 3
- 206010061535 Ovarian neoplasm Diseases 0.000 description 3
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 description 3
- 230000001028 anti-proliverative effect Effects 0.000 description 3
- 201000010881 cervical cancer Diseases 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 238000011534 incubation Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 201000007270 liver cancer Diseases 0.000 description 3
- 208000014018 liver neoplasm Diseases 0.000 description 3
- VMGAPWLDMVPYIA-HIDZBRGKSA-N n'-amino-n-iminomethanimidamide Chemical compound N\N=C\N=N VMGAPWLDMVPYIA-HIDZBRGKSA-N 0.000 description 3
- 208000002154 non-small cell lung carcinoma Diseases 0.000 description 3
- 230000005311 nuclear magnetism Effects 0.000 description 3
- 238000010791 quenching Methods 0.000 description 3
- 230000000171 quenching effect Effects 0.000 description 3
- BOLDJAUMGUJJKM-LSDHHAIUSA-N renifolin D Natural products CC(=C)[C@@H]1Cc2c(O)c(O)ccc2[C@H]1CC(=O)c3ccc(O)cc3O BOLDJAUMGUJJKM-LSDHHAIUSA-N 0.000 description 3
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 3
- 229960005322 streptomycin Drugs 0.000 description 3
- BNWCETAHAJSBFG-UHFFFAOYSA-N tert-butyl 2-bromoacetate Chemical compound CC(C)(C)OC(=O)CBr BNWCETAHAJSBFG-UHFFFAOYSA-N 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 208000029729 tumor suppressor gene on chromosome 11 Diseases 0.000 description 3
- FPVCVHVTMPCZTH-UHFFFAOYSA-N 2-[2-[2-(2-azidoethoxy)ethoxy]ethoxy]ethanamine Chemical compound NCCOCCOCCOCCN=[N+]=[N-] FPVCVHVTMPCZTH-UHFFFAOYSA-N 0.000 description 2
- 102000052609 BRCA2 Human genes 0.000 description 2
- 108700020462 BRCA2 Proteins 0.000 description 2
- 101150008921 Brca2 gene Proteins 0.000 description 2
- 208000005623 Carcinogenesis Diseases 0.000 description 2
- IAZDPXIOMUYVGZ-WFGJKAKNSA-N Dimethyl sulfoxide Chemical compound [2H]C([2H])([2H])S(=O)C([2H])([2H])[2H] IAZDPXIOMUYVGZ-WFGJKAKNSA-N 0.000 description 2
- 241000699660 Mus musculus Species 0.000 description 2
- NSGDYZCDUPSTQT-UHFFFAOYSA-N N-[5-bromo-1-[(4-fluorophenyl)methyl]-4-methyl-2-oxopyridin-3-yl]cycloheptanecarboxamide Chemical compound Cc1c(Br)cn(Cc2ccc(F)cc2)c(=O)c1NC(=O)C1CCCCCC1 NSGDYZCDUPSTQT-UHFFFAOYSA-N 0.000 description 2
- 206010061902 Pancreatic neoplasm Diseases 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 230000036952 cancer formation Effects 0.000 description 2
- 231100000504 carcinogenesis Toxicity 0.000 description 2
- 238000004113 cell culture Methods 0.000 description 2
- 230000003833 cell viability Effects 0.000 description 2
- 239000007810 chemical reaction solvent Substances 0.000 description 2
- 239000012230 colorless oil Substances 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 238000010828 elution Methods 0.000 description 2
- 150000002118 epoxides Chemical group 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 238000001704 evaporation Methods 0.000 description 2
- 230000008020 evaporation Effects 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 230000014509 gene expression Effects 0.000 description 2
- 229910052736 halogen Inorganic materials 0.000 description 2
- 150000002367 halogens Chemical class 0.000 description 2
- 238000000338 in vitro Methods 0.000 description 2
- 208000032839 leukemia Diseases 0.000 description 2
- 208000015486 malignant pancreatic neoplasm Diseases 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 210000004940 nucleus Anatomy 0.000 description 2
- 238000011580 nude mouse model Methods 0.000 description 2
- 201000002528 pancreatic cancer Diseases 0.000 description 2
- 208000008443 pancreatic carcinoma Diseases 0.000 description 2
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 230000007115 recruitment Effects 0.000 description 2
- 229920006395 saturated elastomer Polymers 0.000 description 2
- 150000003384 small molecules Chemical class 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- WWUQRUIBJJBKLS-UHFFFAOYSA-N (2-methylpropan-2-yl)oxycarbonylcarbamic acid Chemical compound CC(C)(C)OC(=O)NC(O)=O WWUQRUIBJJBKLS-UHFFFAOYSA-N 0.000 description 1
- CTJSPUFGQNVJJP-UHFFFAOYSA-N 1,2,3,4-tetrahydroquinolin-6-ol Chemical compound N1CCCC2=CC(O)=CC=C21 CTJSPUFGQNVJJP-UHFFFAOYSA-N 0.000 description 1
- DUNPXMOBBKBKHK-UHFFFAOYSA-N 10-[(2-methylpropan-2-yl)oxycarbonylamino]decanoic acid Chemical compound CC(C)(C)OC(=O)NCCCCCCCCCC(O)=O DUNPXMOBBKBKHK-UHFFFAOYSA-N 0.000 description 1
- HPTPZJBSQUULAV-UHFFFAOYSA-N 11-[(2-methylpropan-2-yl)oxycarbonylamino]undecanoic acid Chemical compound CC(C)(C)OC(=O)NCCCCCCCCCCC(O)=O HPTPZJBSQUULAV-UHFFFAOYSA-N 0.000 description 1
- FALRKNHUBBKYCC-UHFFFAOYSA-N 2-(chloromethyl)pyridine-3-carbonitrile Chemical compound ClCC1=NC=CC=C1C#N FALRKNHUBBKYCC-UHFFFAOYSA-N 0.000 description 1
- VRPJIFMKZZEXLR-UHFFFAOYSA-N 2-[(2-methylpropan-2-yl)oxycarbonylamino]acetic acid Chemical compound CC(C)(C)OC(=O)NCC(O)=O VRPJIFMKZZEXLR-UHFFFAOYSA-N 0.000 description 1
- OUPMUGIXWNVDSN-UHFFFAOYSA-N 2-[hydroxy-(1-oxido-4-oxoquinoxalin-4-ium-2-yl)methyl]-4-methoxyphenol Chemical compound COC1=CC=C(O)C(C(O)C=2N(C3=CC=CC=C3[N+](=O)C=2)[O-])=C1 OUPMUGIXWNVDSN-UHFFFAOYSA-N 0.000 description 1
- WCFJUSRQHZPVKY-UHFFFAOYSA-N 3-[(2-methylpropan-2-yl)oxycarbonylamino]propanoic acid Chemical compound CC(C)(C)OC(=O)NCCC(O)=O WCFJUSRQHZPVKY-UHFFFAOYSA-N 0.000 description 1
- AEKCPQVMPJWNHL-UHFFFAOYSA-N 3-bromoprop-1-ene 3-bromoprop-1-yne Chemical compound BrCC=C.BrCC#C AEKCPQVMPJWNHL-UHFFFAOYSA-N 0.000 description 1
- HIDJWBGOQFTDLU-UHFFFAOYSA-N 4-[(2-methylpropan-2-yl)oxycarbonylamino]butanoic acid Chemical compound CC(C)(C)OC(=O)NCCCC(O)=O HIDJWBGOQFTDLU-UHFFFAOYSA-N 0.000 description 1
- UZOVYGYOLBIAJR-UHFFFAOYSA-N 4-isocyanato-4'-methyldiphenylmethane Chemical compound C1=CC(C)=CC=C1CC1=CC=C(N=C=O)C=C1 UZOVYGYOLBIAJR-UHFFFAOYSA-N 0.000 description 1
- GFMRZAMDGJIWRB-UHFFFAOYSA-N 5-[(2-methylpropan-2-yl)oxycarbonylamino]pentanoic acid Chemical compound CC(C)(C)OC(=O)NCCCCC(O)=O GFMRZAMDGJIWRB-UHFFFAOYSA-N 0.000 description 1
- VNTPPTFQTSGONS-UHFFFAOYSA-N 6-amino-2-[(2-methylpropan-2-yl)oxycarbonyl]hexanoic acid Chemical compound CC(C)(C)OC(=O)C(C(O)=O)CCCCN VNTPPTFQTSGONS-UHFFFAOYSA-N 0.000 description 1
- FPRZYWCRQHFPSX-UHFFFAOYSA-N 8-[(2-methylpropan-2-yl)oxycarbonylamino]octanoic acid Chemical compound CC(C)(C)OC(=O)NCCCCCCCC(O)=O FPRZYWCRQHFPSX-UHFFFAOYSA-N 0.000 description 1
- QCCWTNAQYOOPQP-UHFFFAOYSA-N 9-[(2-methylpropan-2-yl)oxycarbonylamino]nonanoic acid Chemical compound CC(C)(C)OC(=O)NCCCCCCCCC(O)=O QCCWTNAQYOOPQP-UHFFFAOYSA-N 0.000 description 1
- 208000010507 Adenocarcinoma of Lung Diseases 0.000 description 1
- NOWKCMXCCJGMRR-UHFFFAOYSA-N Aziridine Chemical compound C1CN1 NOWKCMXCCJGMRR-UHFFFAOYSA-N 0.000 description 1
- 102000036365 BRCA1 Human genes 0.000 description 1
- 108700020463 BRCA1 Proteins 0.000 description 1
- 101150072950 BRCA1 gene Proteins 0.000 description 1
- 102000015367 CRBN Human genes 0.000 description 1
- SAVPIJHSAJQLGK-UHFFFAOYSA-N ClCC(=O)Cl.ClCC(=O)Cl Chemical compound ClCC(=O)Cl.ClCC(=O)Cl SAVPIJHSAJQLGK-UHFFFAOYSA-N 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 206010064571 Gene mutation Diseases 0.000 description 1
- 101000941994 Homo sapiens Protein cereblon Proteins 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- 206010024305 Leukaemia monocytic Diseases 0.000 description 1
- XCHARIIIZLLEBL-UHFFFAOYSA-N Medicagenic acid 3-O-beta-D-glucoside Chemical compound C12CC(C)(C)CCC2(C(O)=O)CCC(C2(CCC3C4(C)C(O)=O)C)(C)C1=CCC2C3(C)CC(O)C4OC1OC(CO)C(O)C(O)C1O XCHARIIIZLLEBL-UHFFFAOYSA-N 0.000 description 1
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 1
- 229940125941 PARP1/2 inhibitor Drugs 0.000 description 1
- 102000014160 PTEN Phosphohydrolase Human genes 0.000 description 1
- 108010011536 PTEN Phosphohydrolase Proteins 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 235000005811 Viola adunca Nutrition 0.000 description 1
- 240000009038 Viola odorata Species 0.000 description 1
- 235000013487 Viola odorata Nutrition 0.000 description 1
- 235000002254 Viola papilionacea Nutrition 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- HSFWRNGVRCDJHI-UHFFFAOYSA-N alpha-acetylene Natural products C#C HSFWRNGVRCDJHI-UHFFFAOYSA-N 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 150000001540 azides Chemical class 0.000 description 1
- 238000011001 backwashing Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 239000007805 chemical reaction reactant Substances 0.000 description 1
- 210000001072 colon Anatomy 0.000 description 1
- 201000010897 colon adenocarcinoma Diseases 0.000 description 1
- 208000029742 colonic neoplasm Diseases 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- DEZRYPDIMOWBDS-UHFFFAOYSA-N dcm dichloromethane Chemical compound ClCCl.ClCCl DEZRYPDIMOWBDS-UHFFFAOYSA-N 0.000 description 1
- 230000006837 decompression Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- UXGNZZKBCMGWAZ-UHFFFAOYSA-N dimethylformamide dmf Chemical compound CN(C)C=O.CN(C)C=O UXGNZZKBCMGWAZ-UHFFFAOYSA-N 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 238000009509 drug development Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 125000001301 ethoxy group Chemical group [H]C([H])([H])C([H])([H])O* 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 238000003304 gavage Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- ACGUYXCXAPNIKK-UHFFFAOYSA-N hexachlorophene Chemical compound OC1=C(Cl)C=C(Cl)C(Cl)=C1CC1=C(O)C(Cl)=CC(Cl)=C1Cl ACGUYXCXAPNIKK-UHFFFAOYSA-N 0.000 description 1
- 231100000171 higher toxicity Toxicity 0.000 description 1
- 239000005457 ice water Substances 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000005917 in vivo anti-tumor Effects 0.000 description 1
- 230000002197 limbic effect Effects 0.000 description 1
- 201000005249 lung adenocarcinoma Diseases 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000006996 mental state Effects 0.000 description 1
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Natural products C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 239000012046 mixed solvent Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 201000006894 monocytic leukemia Diseases 0.000 description 1
- WOOWBQQQJXZGIE-UHFFFAOYSA-N n-ethyl-n-propan-2-ylpropan-2-amine Chemical compound CCN(C(C)C)C(C)C.CCN(C(C)C)C(C)C WOOWBQQQJXZGIE-UHFFFAOYSA-N 0.000 description 1
- XBXCNNQPRYLIDE-UHFFFAOYSA-M n-tert-butylcarbamate Chemical compound CC(C)(C)NC([O-])=O XBXCNNQPRYLIDE-UHFFFAOYSA-M 0.000 description 1
- 230000009701 normal cell proliferation Effects 0.000 description 1
- 238000010899 nucleation Methods 0.000 description 1
- 230000002018 overexpression Effects 0.000 description 1
- RNGQTPOUTNKYLA-UHFFFAOYSA-N oxolane-2,5-dione Chemical compound O=C1CCC(=O)O1.O=C1CCC(=O)O1 RNGQTPOUTNKYLA-UHFFFAOYSA-N 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 229910000027 potassium carbonate Inorganic materials 0.000 description 1
- 230000003389 potentiating effect Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- RZWZRACFZGVKFM-UHFFFAOYSA-N propanoyl chloride Chemical compound CCC(Cl)=O RZWZRACFZGVKFM-UHFFFAOYSA-N 0.000 description 1
- YORCIIVHUBAYBQ-UHFFFAOYSA-N propargyl bromide Chemical compound BrCC#C YORCIIVHUBAYBQ-UHFFFAOYSA-N 0.000 description 1
- 230000018883 protein targeting Effects 0.000 description 1
- 230000017854 proteolysis Effects 0.000 description 1
- 229940124823 proteolysis targeting chimeric molecule Drugs 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 229940014800 succinic anhydride Drugs 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000002626 targeted therapy Methods 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 125000000999 tert-butyl group Chemical group [H]C([H])([H])C(*)(C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- AOCSUUGBCMTKJH-UHFFFAOYSA-N tert-butyl n-(2-aminoethyl)carbamate Chemical compound CC(C)(C)OC(=O)NCCN AOCSUUGBCMTKJH-UHFFFAOYSA-N 0.000 description 1
- POHWAQLZBIMPRN-UHFFFAOYSA-N tert-butyl n-(3-aminopropyl)carbamate Chemical compound CC(C)(C)OC(=O)NCCCN POHWAQLZBIMPRN-UHFFFAOYSA-N 0.000 description 1
- ZFQWJXFJJZUVPI-UHFFFAOYSA-N tert-butyl n-(4-aminobutyl)carbamate Chemical compound CC(C)(C)OC(=O)NCCCCN ZFQWJXFJJZUVPI-UHFFFAOYSA-N 0.000 description 1
- DPLOGSUBQDREOU-UHFFFAOYSA-N tert-butyl n-(5-aminopentyl)carbamate Chemical compound CC(C)(C)OC(=O)NCCCCCN DPLOGSUBQDREOU-UHFFFAOYSA-N 0.000 description 1
- RVZPDKXEHIRFPM-UHFFFAOYSA-N tert-butyl n-(6-aminohexyl)carbamate Chemical compound CC(C)(C)OC(=O)NCCCCCCN RVZPDKXEHIRFPM-UHFFFAOYSA-N 0.000 description 1
- BEHVGNKIRNVBPF-UHFFFAOYSA-N tert-butyl n-(8-aminooctyl)carbamate Chemical compound CC(C)(C)OC(=O)NCCCCCCCCN BEHVGNKIRNVBPF-UHFFFAOYSA-N 0.000 description 1
- CUPBLDPRUBNAIE-UHFFFAOYSA-N tert-butyl n-[2-[2-[2-(2-aminoethoxy)ethoxy]ethoxy]ethyl]carbamate Chemical compound CC(C)(C)OC(=O)NCCOCCOCCOCCN CUPBLDPRUBNAIE-UHFFFAOYSA-N 0.000 description 1
- WCNWLERBLMBSOT-UHFFFAOYSA-N tert-butyl n-[2-[2-[2-[2-(2-aminoethoxy)ethoxy]ethoxy]ethoxy]ethyl]carbamate Chemical compound CC(C)(C)OC(=O)NCCOCCOCCOCCOCCN WCNWLERBLMBSOT-UHFFFAOYSA-N 0.000 description 1
- ISTGQSQWSKCNFJ-UHFFFAOYSA-N tert-butyl n-ethylcarbamate Chemical compound CCNC(=O)OC(C)(C)C ISTGQSQWSKCNFJ-UHFFFAOYSA-N 0.000 description 1
- 238000011269 treatment regimen Methods 0.000 description 1
- 230000005760 tumorsuppression Effects 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
- C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
- C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
-
- 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
- A61K31/50—Pyridazines; Hydrogenated pyridazines
- A61K31/502—Pyridazines; Hydrogenated pyridazines ortho- or peri-condensed with carbocyclic ring systems, e.g. cinnoline, phthalazine
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
- A61K47/545—Heterocyclic compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
- A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
- C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Engineering & Computer Science (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Molecular Biology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
本发明涉及共价PARP PROTACs衍生物及其应用，属于抗肿瘤药物技术领域。本发明解决的技术问题是提供一种具有选择性降解PARP2的新型共价PROTACs衍生物，该共价PARPPROTACs衍生物的结构通式为式Ⅰ所示。本发明基于DCAF16 E3连接酶配体KB02和PARP抑制剂Olaparib，通过讨论Linker链接方式、长度和组成成分等，设计并合成了一系列新型共价PARP PROTACs分子，这些分子可以通过共价修饰DCAF16发挥招募E3连接酶的作用，并依赖泛素‑蛋白酶体降解细胞核内的PARP2蛋白，达到良好的抑制肿瘤细胞增殖、治疗多种肿瘤细胞的效果。
Description
技术领域
本发明涉及共价PARP PROTACs衍生物及其应用，属于抗肿瘤药物技术领域。
背景技术
抗肿瘤药物(anti-tumor drug)为治疗肿瘤疾病的一类药物。近年来，分子肿瘤学、分子药理学的发展使肿瘤本质正在逐步阐明；大规模快速筛选、组合化学、基因工程等先进技术的发明和应用加速了药物开发进程；抗肿瘤药物的研究与开发已进入一个崭新的时代。
当前，在治疗肿瘤过程中发现小分子抗肿瘤药物出现包括耐药性、副作用和对多种蛋白的抑制活性减弱等问题，小分子抑制剂药物的开发已处于瓶颈期。靶向蛋白降解成为药物开发中的一项新技术，也拓展了小分子药物的开发领域。利用蛋白靶向降解嵌合体(PROTACs)技术来降解对肿瘤发生有重要作用的蛋白质，已成为一种潜在的癌症治疗策略。到目前为止，已开发的大多数PROTACs都利用配体招募在肿瘤和正常组织中普遍表达的E3连接酶(如CRBN和VHL等)。如果目标蛋白不是肿瘤特异性的，这些PROTACs可能会引起靶标毒性。因此，识别和招募在肿瘤中表达丰富的，而在正常组织中表达最低的E3连接酶具有开发肿瘤特异性/选择性PROTACs的潜力。另外，研究发现PARP2与PARP1蛋白在许多促进肿瘤发生的生理功能上表现出明显的差异，而目前已有的靶向降解PARP蛋白的PROTACs分子中，几乎都选择性降解PARP1，而对PARP2降解的研究极少。因此开发对PARP1和PARP2具有更高选择性的抑制剂或PROTACs分子将有助于开发更安全的针对癌症的PARP靶向疗法。
发明内容
针对以上缺陷，本发明解决的技术问题是提供一种具有选择性降解PARP2的新型共价PROTACs衍生物，用于治疗非BRCA基因突变的肿瘤细胞，促进基于靶向PARP的肿瘤个体化治疗。
共价PARP PROTACs衍生物，其特征在于，其结构通式为式Ⅰ所示：
其中，X选自卤素原子、甲基、羟基、氨基、硝基或可与目标蛋白的氨基酸残基发生共价反应的亲电基团；linker为直链烷烃类链、PEG类链、醚类链或三氮唑类链中的至少一种。
在本发明的一个实施方式中，所述X为卤素原子或可与目标蛋白的氨基酸残基发生共价反应的亲电基团；其中，所述可与目标蛋白的氨基酸残基发生共价反应的亲电基团包括丙烯酰胺基、α-卤酮基、氰基乙酰胺基、巯基、环氧化物基、乙烯基砜基或活化乙炔基。
在本发明的一个具体实施例中，所述X为氯原子。
在本发明的一个实施方式中，所述直链烷烃的结构式为式⑴、式⑵或式⑶所示：
式⑴为
其中n1为1～12中的任意整数；
式⑵为
其中n2为1～5中的任意整数；
式⑶为
其中n3为1～7中的任意整数；
所述PEG单元链的结构式为式⑷或式⑸所示：
式⑷为
其中n4为1～4中的任意整数；
式⑸为
其中n5为1～4中的任意整数；
所述三氮唑类链的结构式为
其中n6、n7为1～5中的任意整数。
本发明还提供本发明所述共价PARP PROTACs衍生物在制备抗肿瘤药物中的应用。
本发明共价PARP PROTACs衍生物，可以招募DCAF16 E3连接酶，从而达到抗肿瘤的效果。作为一种实施方式，所述抗肿瘤药物为招募DCAF16 E3连接酶的药物。
在一种具体实施方式中，所述抗肿瘤药物为靶向降解PARP2蛋白的药物。
在一个实施例中，所述抗肿瘤药物为靶向降解PARP1和PARP2蛋白的药物。
与现有技术相比，本发明具有如下有益效果：
本发明基于DCAF16 E3连接酶配体KB02和PARP抑制剂Olaparib，通过讨论Linker链接方式、长度和组成成分等，设计并合成了一系列新型共价PARP PROTACs分子，这些分子可以通过共价修饰DCAF16发挥招募E3连接酶的作用，并依赖泛素-蛋白酶体降解细胞核内的PARP2蛋白，达到良好的抑制肿瘤细胞增殖、治疗多种肿瘤细胞的效果。
附图说明
图1为目标化合物对MDA-MB-231细胞内的PARP1和PARP2的降解活性。
图2为化合物C8体外抗增殖活性及适用性评估。
图3为化合物C8对MDA-MB-231肿瘤体积随时间的变化。
图4为实验裸鼠体重随时间的变化。
具体实施方式
本发明共价PARP PROTACs衍生物，其结构通式为式Ⅰ所示：
其中，X选自卤素原子、甲基、羟基、氨基、硝基或可与目标蛋白的氨基酸残基发生共价反应的亲电基团；linker为直链烷烃类链、PEG类链、醚类链或三氮唑类链中的至少一种。
本发明化合物，基于DCAF16 E3连接酶配体KB02和PARP抑制剂Olaparib(奥拉帕尼)，通过linker连接，得到一系列新型共价PARP PROTACs衍生物。这些分子可以通过共价修饰DCAF16发挥招募E3连接酶的作用，可以成功实现选择性靶向降解PARP2蛋白，从而发挥优于PARP抑制剂的抗肿瘤作用。
在本发明一个具体实施方式中，所述X为卤素原子或可与目标蛋白的氨基酸残基发生共价反应的亲电基团；其中，所述可与目标蛋白的氨基酸残基发生共价反应的亲电基团包括丙烯酰胺、α-卤酮、环氧化物、氮丙环、乙烯基砜或活化乙炔。
其中，所述丙烯酰胺基为
α-卤酮基
X1为卤素；氰基乙酰胺基为
巯基为-SH，环氧化物基为
乙烯基砜基
活化乙炔基为
X2为卤素。
在本发明一个具体实施例中，所述X为氯原子。
本发明共价PARP PROTACs衍生物中，所述直链烷烃的结构式为式⑴、式⑵或式⑶所示：
式⑴为
其中n1为1～11中的任意整数；
式⑵为
其中n2为1～5中的任意整数；
式⑶为
其中n3为1～7中的任意整数；
所述PEG单元链的结构式为式⑷或式⑸所示：
式⑷为
其中n4为1～4中的任意整数；
式⑸为
其中n5为1～4中的任意整数；
所述三氮唑类链的结构式为
在一些具体的实施例中，所述共价PARP PROTACs衍生物分为A～D类化合物，其具体结构式如下：
A系列化合物A1～A8结构如下：
B系列化合物B1～B10结构如下：
C系列化合物C1～C9，C8-M结构如下：
D系列化合物D1～D3结构如下：
本发明共价PARP PROTACs衍生物，可以招募DCAF16 E3连接酶，从而达到抗肿瘤的效果。作为一种实施方式，所述抗肿瘤药物为招募DCAF16 E3连接酶的药物。
在一种具体实施方式中，所述抗肿瘤药物为靶向降解PARP2蛋白的药物。
在一个实施例中，所述抗肿瘤药物为靶向降解PARP1和PARP2蛋白的药物。
下面结合实施例对本发明的具体实施方式做进一步的描述，并不因此将本发明限制在所述的实施例范围之中。
实施例1 PARP PROTACs分子的制备
1、PARP PROTACs分子中间体的合成
试剂与条件:(a)二碳酸二叔丁酯Di-tert-butyl decarbonate,二氯甲烷DCM,50℃,大气压pressure,4h；(b)溴乙酸叔丁酯Tert-butyl Bromoacetate,K2CO3,N,N-二甲基甲酰胺DMF,室温rt,6h；(c)DCM/TFA,rt,1～2h.
中间体化合物K1的合成
称取6-羟基-1,2,3,4-四氢喹啉K0(500mg，3.36mmol，1eq)置于50mL加压封管中，用20mL二氯甲烷(DCM)溶解。取二碳酸二叔丁酯(735mg，3.36mmol，1eq)溶于溶液中，在常温下搅拌溶解后移入50℃油浴锅中进行加热加压反应3-4h。TLC监测反应结束，减压蒸馏除去DCM，加水后用EA萃取三次，合并有机层，用饱和食盐水反洗有机层，经无水硫酸钠干燥，过滤的溶液经减压浓缩。得到粗产物经300～400目硅胶柱层析(PE/EA洗脱体系＝3:1)分离，得到目标产物K1，为无色油状物758mg，产率为93.8％。1H NMR(400MHz,DMSO-d6)δ9.09(s,1H),7.29(d,J＝8.8Hz,1H),6.52(dd,J＝8.8,2.9Hz,1H),6.48(d,J＝2.8Hz,1H),3.56(t,J＝6.6Hz,2H),2.63(t,J＝6.6Hz,2H),1.77(p,J＝6.5Hz,2H),1.43(s,9H).HRMS(DART-TOF)calculated for C14H19NO3 +[M+Na]+272.1257，found 272.1263.
中间体化合物K2的合成
称取K1(800mg，3.2mmol，1eq)和碳酸钾(1.3g，9.6mmol，3eq)置于50mL圆底烧瓶中，用15mL DMF溶解，在室温搅拌下缓慢滴加溴乙酸叔丁酯(749mg，3.84mmol，1.2eq)，然后在室温下搅拌6h，TLC监测反应结束。加入水淬灭反应，用EA萃取三次，合并有机层，用饱和食盐水反洗有机层，经无水硫酸钠干燥，过滤的溶液经减压浓缩。得到粗产物经300-400目硅胶柱层析(PE/EA洗脱体系＝5:1)分离，得到目标产物K2，为无色油状物1.12g，产率为95.9％。1H NMR(400MHz,Chloroform-d)δ7.53(d,J＝9.0Hz,1H),6.75–6.58(m,2H),4.45(s,2H),3.70–3.62(m,2H),2.72(t,J＝6.6Hz,2H),1.89(p,J＝6.5Hz,2H),1.49(d,J＝7.4Hz,18H).HRMS(DART-TOF)calculated for C19H29NNaO4 +[M+Na]+m/z 386.1953,found386.1946.
中间体化合物K3的合成
选用50mL的圆底烧瓶，将K2(1g，4.83mmol，1eq)溶解于20mL DCM/TFA＝2：1的溶剂中。将反应混合物在室温下搅拌1～2h，TLC监测反应结束。减压浓缩除去溶剂后，得到581mg乳白色固体粉末K3，产率为98％。K3可不经纯化，直接用于下一步反应。1H NMR(400MHz,DMSO-d6)δ7.02(d,J＝8.6Hz,1H),6.84–6.76(m,2H),4.65(s,2H),3.36–3.29(m,2H),2.77(t,J＝6.5Hz,2H),1.98–1.89(m,2H).13C NMR(101MHz,DMSO-d6)δ170.55,122.53,115.80,114.16,65.19,42.13,25.66,20.14.HRMS(DART-TOF)calculated for C11H14NO4 +[M+H]+m/z208.0895,found 208.0893.
通用合成步骤A：合成中间体化合物G1～G9
称取L1(120mg，0.328mmol，1eq)，HATU(150mg，0.39mmol，1.2eq)和不同链长的含N-BOC-氨基某羧酸的linker(1.2eq)溶于无水DMF中，加入DIPEA(129mg，1mmol，3eq)在室温下搅拌反应，TLC监测反应结束。后处理：加大量水淬灭反应后，用EA萃取3次，合并有机层，用后饱和食盐水反洗2次。有机层用无水硫酸钠干燥，过滤，减压浓缩后用硅胶柱层(DCM/CH3OH体系)分离纯化，得到目标产物，经1H-NMR和高分辨质谱确认化合物结构。
中间体化合物G1的合成
G1的合成参考通用合成步骤C，以L1和BOC-甘氨酸为反应原料即可得到中间体G1，淡黄色固体，产率为89％。1H NMR(400MHz,Chloroform-d)δ10.62(s,1H),8.50–8.43(m,1H),7.80–7.68(m,3H),7.38–7.30(m,2H),7.04(t,J＝9.0Hz,1H),5.49(d,J＝15.6Hz,1H),4.29(s,2H),4.02–3.27(m,10H),1.44(s,9H).HRMS(DART-TOF)calculated forC27H30FN5NaO5 +[M+Na]+m/z 546.2118,found 546.2125.
中间体化合物G2的合成
G2的合成参考通用合成步骤C，以L1和Boc-beta-丙氨酸为反应原料即可得到中间体G2，淡黄色固体，产率为90％。1H NMR(400MHz,Chloroform-d)δ11.68(s,1H),8.47–8.40(m,1H),7.78–7.70(m,3H),7.36–7.28(m,2H),7.02(t,J＝9.7,5.1Hz,1H),4.74(s,1H),4.28(s,2H),3.79–3.32(m,10H),2.60–2.44(m,2H),1.39(s,9H).HRMS(DART-TOF)calculated for C28H32FN5NaO5 +[M+Na]+m/z 560.2286,found 560.2286.
中间体化合物G3的合成
G3的合成参考通用合成步骤C，以L1和N-BOC-GAMMA-氨基丁酸为反应原料即可得到中间体G3，淡黄色固体，产率为85％。1H NMR(400MHz,Chloroform-d)δ11.46(br,1H),8.48–8.41(m,1H),7.79–7.71(m,3H),7.38–7.28(m,2H),7.06–6.98(m,1H),4.87(s,1H),4.28(s,2H),3.74–3.13(m,10H),2.44–2.30(m,2H),2.12–2.05(m,2H),1.40(s,9H).HRMS(DART-TOF)calculated for C29H34FN5NaO5 +[M+Na]+m/z 574.2441,found 574.2442.
中间体化合物G4的合成
G4的合成参考通用合成步骤C，以L1和Boc-5-氨基戊酸为反应原料即可得到中间体G4，淡黄色固体，产率为81％。1H NMR(400MHz,Chloroform-d)δ10.33(br,1H),8.47-8.41(m,1H),7.80–7.70(m,3H),7.35–7.30(m,2H),7.05(t,J＝9.0Hz,1H),4.64(s,1H),4.29(s,2H),3.84–3.09(m,10H),2.36(dt,J＝26.4,7.5Hz,2H),1.70–1.64(m,2H),1.58–1.51(m,2H),1.43(s,9H).HRMS(DART-TOF)calculated for C30H36FN5NaO5 +[M+Na]+m/z 588.2583,found 588.2596.
中间体化合物G5的合成
G5的合成参考通用合成步骤C，以L1和叔丁氧羰酰基6-氨基己酸为反应原料即可得到中间体G5，淡黄色固体，产率为82％。1H NMR(400MHz,Chloroform-d)δ10.55(br,1H),8.50–8.43(m,1H),7.83–7.69(m,3H),7.38–7.30(m,2H),7.05(t,J＝9.1Hz,1H),4.59(s,1H),4.29(s,2H),3.86–3.10(m,10H),2.33(dt,J＝28.0,7.6Hz,2H),1.65(q,J＝8.0Hz,2H),1.43(s,9H),1.41–1.35(m,4H).HRMS(DART-TOF)calculated for C30H36FN5NaO5 +[M+Na]+m/z 602.2753,found 602.2757.
中间体化合物G6的合成
G6的合成参考通用合成步骤C，以L1和BOC-8-氨基辛酸为反应原料即可得到中间体G6，淡黄色固体，产率为87％。1H NMR(400MHz,Chloroform-d)δ10.64(br,1H),8.51–8.44(m,1H),7.82–7.68(m,3H),7.39–7.29(m,2H),7.05(t,J＝9.1Hz,1H),4.54(s,1H),4.29(s,2H),3.80–3.07(m,10H),2.40–2.24(m,2H),1.73–1.61(m,4H),1.44(s,9H),1.35–1.29(m,6H).HRMS(DART-TOF)calculated for C33H42FN5NaO5 +[M+Na]+m/z 630.3067,found630.3071.
中间体化合物G7的合成
G7的合成参考通用合成步骤C，以L1和9-叔丁氧羰基氨基-壬酸为反应原料即可得到中间体G7，淡黄色固体，产率为86％。1H NMR(400MHz,Chloroform-d)δ10.65(br,1H),8.50–8.45(m,1H),7.81–7.68(m,3H),7.38–7.29(m,2H),7.05(t,J＝9.1Hz,1H),4.53(s,1H),4.29(s,2H),3.82–3.08(m,10H),2.42–2.27(m,2H),1.70–1.63(m,2H),1.44(s,9H),1.35–1.29(m,10H).HRMS(DART-TOF)calculated for C34H44FN5NaO5 +[M+Na]+m/z 644.3224,found 644.3231.
中间体化合物G8的合成
G8的合成参考通用合成步骤C，以L1和10-((叔丁氧基羰基)氨基)癸酸为反应原料即可得到中间体G8，淡黄色固体，产率为92％。1H NMR(400MHz,Chloroform-d)δ10.68(br,1H),8.51–8.44(m,1H),7.82–7.68(m,3H),7.39–7.30(m,2H),7.05(t,J＝9.1Hz,1H),4.53(s,1H),4.29(s,2H),3.84–3.06(m,10H),2.34–2.26(m,2H),1.63–1.59(m,2H),1.44(s,9H),1.31–1.26(m,12H).HRMS(DART-TOF)calculated for C35H46FN5NaO5 +[M+Na]+m/z658.3381,found 658.3325.
中间体化合物G9的合成
G9的合成参考通用合成步骤C，以L1和11-(Boc-氨基)十一烷酸为反应原料即可得到中间体G9，淡黄色固体，产率为89％。1H NMR(400MHz,Chloroform-d)δ10.66(br,1H),8.50–8.43(m,1H),7.80–7.67(m,3H),7.39–7.30(m,2H),7.05(t,J＝9.1Hz,1H),4.53(s,1H),4.29(s,2H),3.85–3.02(m,10H),2.34–2.26(m,2H),1.60–1.56(m,2H),1.44(s,9H),1.33–1.26(m,14H).HRMS(DART-TOF)calculated for C36H48FN5NaO5 +[M+Na]+m/z 672.3537,found 672.3535.
中间体化合物G10的合成
G10的合成参考通用合成步骤C，以L1和12-(Boc-氨基)十一烷酸为反应原料即可得到中间体G10，淡黄色固体，产率为75％。1H NMR(400MHz,Chloroform-d)δ10.67(br,1H),8.50–8.43(m,1H),7.81–7.67(m,3H),7.38–7.29(m,2H),7.05(t,J＝9.1Hz,1H),4.54(s,1H),4.28(s,2H),3.86–3.01(m,10H),2.35–2.27(m,2H),1.61–1.57(m,2H),1.45(s,9H),1.35–1.25(m,16H).HRMS(DART-TOF)calculated for C37H50FN5NaO5 +[M+Na]+m/z 686.3796,found 686.3795.
中间体化合物G11的合成
G11的合成参考通用合成步骤C，以L1和13-(BOC-氨基)十一烷酸为反应原料即可得到中间体G11，淡黄色固体，产率为72％。1H NMR(400MHz,Chloroform-d)δ10.66(br,1H),8.51–8.44(m,1H),7.81–7.66(m,3H),7.38–7.29(m,2H),7.05(t,J＝9.1Hz,1H),4.53(s,1H),4.29(s,2H),3.85–3.01(m,10H),2.34–2.26(m,2H),1.61–1.58(m,2H),1.44(s,9H),1.37–1.26(m,18H).HRMS(DART-TOF)calculated for C38H52FN5NaO5 +[M+Na]+m/z 700.3952,found 700.3953.
通用合成步骤B：合成中间体C1-1～C9-1
在25mL圆底烧瓶中加入中间体G1～G9(1.2eq)，加入TFA/DCM溶液溶解，在室温下搅拌反应1～2h，TLC监测反应结束后，减压浓缩除去溶剂，得到的粗产物即可用于下一步反应。将前一步得到的粗产物溶于无水DMF溶解，再向反应液中加入K3(1eq)、HATU(1.2eq)和DIPEA(3eq)继续在室温下反应3～4h，TLC监测反应结束。后处理：加大量水淬灭反应后，用EA萃取3次，合并有机层，用后饱和食盐水反洗1～2次。有机层用无水硫酸钠干燥，过滤，减压浓缩后用硅胶柱层析(DCM/CH3OH体系＝20:1)分离纯化，得到目标产物，经1H-NMR和高分辨质谱确认化合物结构。
中间体化合物C1-1的合成
化合物C1-1的合成参考通用合成步骤H，以G1和K3为反应原料即可得到中间体C1-1，淡黄色粉末，产率为59.2％。1H NMR(400MHz,Chloroform-d)δ11.19(s,1H),8.49–8.42(m,1H),7.79–7.62(m,4H),7.36–7.29(m,3H),7.04(t,J＝8.9Hz,1H),6.63–6.58(m,2H),6.40(d,J＝8.2Hz,1H),4.42(d,J＝5.6Hz,2H),4.28(s,2H),4.21(S,2H),3.87–3.21(m,10H),2.74–2.67(m,2H),1.94–1.86(m,2H).13C NMR(101MHz,DMSO-d6)δ170.55,155.83,130.87,127.75,122.53,115.80,114.16,65.19,42.13,25.66,20.14.HRMS(DART-TOF)calculated for C33H33FN6NaO5 +[M+Na]+m/z 635.2394,found 635.2388.
中间体化合物C2-1的合成
化合物C2-1的合成参考通用合成步骤H，以G2和K3为反应原料即可得到中间体C2-1，淡黄色粉末，产率为61.4％。1H NMR(400MHz,Chloroform-d)δ10.49(d,J＝14.2Hz,1H),8.46(t,J＝4.4Hz,1H),7.80–7.70(m,4H),7.32(q,J＝8.1Hz,3H),7.05(t,J＝8.7Hz,1H),6.58(d,J＝3.8Hz,2H),6.40(d,J＝8.2Hz,1H),4.38(s,2H),4.28(s,2H),3.88–3.19(m,10H),2.78–2.70(m,4H),2.67–2.50(m,2H),2.00–1.92(m,2H).HRMS(DART-TOF)calculatedfor C34H35FN6NaO5 +[M+Na]+m/z 649.2551,found 649.2558.
中间体化合物C3-1的合成
化合物C3-1的合成参考通用合成步骤H，以G3和K3为反应原料即可得到中间体C3-1，乳白色固体，产率为52.5％。1H NMR(400MHz,Chloroform-d)δ10.57(br,1H),8.46(dd,J＝6.9,2.4Hz,1H),7.80–7.69(m,3H),7.36–7.28(m,2H),7.04(t,J＝8.9Hz,1H),6.93(s,1H),6.57(d,J＝10.5Hz,2H),6.41(t,J＝8.0Hz,1H),4.37(s,2H),4.28(s,2H),3.83–3.24(m,12H),2.72(t,J＝6.5Hz,2H),2.43–2.29(m,2H),1.94–1.88(m,4H).HRMS(DART-TOF)calculated for C35H37FN6NaO5 +[M+Na]+m/z 663.2707,found 663.2689.
中间体化合物C4-1的合成
化合物C4-1的合成参考通用合成步骤H，以G4和K3为反应原料即可得到中间体C4-1，乳白色固体，产率为55.3％。1H NMR(400MHz,Chloroform-d)δ10.69(br,1H),8.46(dd,J＝6.9,2.4Hz,1H),7.80–7.69(m,3H),7.34–7.28(m,2H),7.04(t,J＝8.9Hz,1H),6.77(s,1H),6.60–6.53(m,2H),6.42(d,J＝8.3Hz,1H),4.37(s,2H),4.28(s,2H),3.83–3.23(m,12H),2.72(t,J＝6.5Hz,2H),2.43–2.31(m,2H),1.96–1.86(m,2H),1.76–1.54(m,4H).HRMS(DART-TOF)calculated for C36H39FN6NaO5 +[M+Na]+m/z 677.2863,found 677.2863.
中间体化合物C5-1的合成
化合物C5-1的合成参考通用合成步骤H，以G5和K3为反应原料即可得到中间体C5-1，乳白色固体，产率为55.3％。1H NMR(400MHz,Chloroform-d)δ10.37(br,1H),8.49–8.43(m,1H),7.82–7.68(m,3H),7.32(t,J＝6.6Hz,3H),7.04(t,J＝8.8Hz,1H),6.67(s,1H),6.57(d,J＝10.4Hz,2H),6.43(d,J＝8.3Hz,1H),4.38(s,2H),4.28(s,2H),3.86–3.18(m,12H),2.73(t,J＝6.5Hz,2H),2.39–2.27(m,2H),1.97–1.86(m,2H),1.66–1.53(m,4H),1.40–1.31(m,2H).HRMS(DART-TOF)calculated for C37H41FN6NaO5 +[M+Na]+m/z 691.3020,found 691.3021.
中间体化合物C6-1的合成
化合物C6-1的合成参考通用合成步骤H，以G6和K3为反应原料即可得到中间体C6-1，乳白色固体，产率为51.9％。1H NMR(400MHz,Chloroform-d)δ10.46(br,1H),8.48-8.42(m,1H),7.82-7.53(m,3H),7.34–7.29(m,3H),7.05(t,J＝8.8Hz,1H),6.80–6.50(m,4H),4.43(s,2H),4.28(s,2H),3.87–3.23(m,14H),2.76(t,J＝6.6Hz,2H),2.39–2.26(m,2H),1.68–1.46(m,4H),1.34–1.25(m,6H).HRMS(DART-TOF)calculated for C39H45FN6NaO5 +[M+Na]+m/z 719.3333,found719.3324.
中间体化合物C7-1的合成
化合物C7-1的合成参考通用合成步骤H，以G7和K3为反应原料即可得到中间体C7-1，乳白色固体，产率为52.1％。1H NMR(400MHz,Chloroform-d)δ10.48(br,1H),8.48-8.42(m,1H),7.82-7.57(m,3H),7.34–7.29(m,3H),7.05(t,J＝8.8Hz,1H),6.82–6.75(m,2H),6.68–6.50(m,2H),4.43(s,2H),4.28(s,2H),3.87–3.23(m,14H),2.76(t,J＝6.6Hz,2H),2.38–2.25(m,2H),1.65–1.49(m,4H),1.34–1.23(m,8H).HRMS(DART-TOF)calculated forC40H47FN6NaO5 +[M+Na]+m/z 733.3480,found 733.3488.
中间体化合物C8-1的合成
化合物C8-1的合成参考通用合成步骤H，以G8和K3为反应原料即可得到中间体C8-1，乳白色固体，产率为60.7％。1H NMR(400 MHz,Chloroform-d)δ10.53(br,1H),8.49–8.43(m,1H),7.82–7.69(m,3H),7.32(t,J＝7.1 Hz,2H),7.04(t,J＝8.6 Hz,1H),6.74(d,J＝5.7 Hz,2H),6.59–6.37(m,3H),4.47(s,2H),4.28(s,2H),3.87–3.24(m,12H),2.69(t,J＝6.6 Hz,2H),2.38–2.25(m,2H),2.00–1.92(m,2H),1.66–1.49(m,4H),1.31–1.27(m,10H).HRMS(DART-TOF)calculated for C41H49FN6NaO5 +[M+Na]+m/z 747.3646,found 747.3645.
中间体化合物C9-1的合成
化合物C9-1的合成参考通用合成步骤H，以G9和K3为反应原料即可得到中间体C9-1，乳白色固体，产率为61.4％。1H NMR(400 MHz,Chloroform-d)δ10.60(br,1H),8.50–8.44(m,1H),7.84-7.52(m,3H),7.33(t,J＝7.1 Hz,3H),7.09–7.02(m,1H),6.64–6.54(m,3H),6.43(d,J＝8.4 Hz,1H),4.38(s,2H),4.29(s,2H),3.85–3.19(m,12H),2.73(t,J＝6.5 Hz,2H),2.40–2.25(m,2H),1.97–1.87(m,2H),1.65–1.49(m,4H),1.36–1.23(m,12H).HRMS(DART-TOF)calculated for C42H52FN6O5 +[M+H]+m/z 739.3975,found 739.3978.
试剂与条件:(a)氯乙酰氯Chloroacetyl chloride,TEA，THF,0℃～rt,1～2h.(b)3-溴丙炔3-Bromopropyne,K2CO3,CH3CN,60℃,4h.(c)Tert-butyl Bromoacetate,K2CO3,DMF,rt,4h；DCM/TFA,rt,1～2h.(d)1-氨基-11-叠氮-3,6,9-三氧杂十一烷11-Azido-3,6,9-trioxaundecan-1-amine,HATU,DIPEA,rt,3h.
中间体化合物KB02的合成
在25mL圆底烧瓶中，称取K0(100mg，0.67mmol，1eq)溶于THF中。向溶液中滴加TEA(203mg，2mmol，3eq)，在冰水浴条件下搅拌反应15分钟，再向反应液中缓慢滴加氯乙酰氯(91mg，0.8mmol，1.2eq),观察溶液由澄清变浑浊，继续反应1h左右，用TLC监测反应结束。减压蒸发除去反应溶剂，加水，用EA萃取3次，有机层经无水硫酸钠干燥，过滤，减压浓缩得到粗品经硅胶柱层析(PE/EA体系＝3:1)分离纯化。得到乳白色固体，产率91％。1H NMR(400MHz,Chloroform-d)δ7.03(s,1H),6.72(d,J＝7.1Hz,2H),4.20(s,2H),3.81(t,J＝6.9Hz,2H),2.68–2.64(m,2H),2.01–1.90(m,2H).HRMS(DART-TOF)calculated forC11H13ClNO2 +[M+H]+m/z 226.0635,found 226.0636.
中间体化合物K4的合成
在25mL反应瓶中加入KB02(100mg，0.44mmol，1eq)和K2CO3(182mg，1.32mmol，3eq)，用乙腈溶解。向搅拌中的反应液中滴加3-溴丙炔(63mg，0.53mmol，1.2eq)，再将反应液移入60℃油浴锅中继续反应4h。TLC监测反应结束。处理：反应液经减压浓缩后，加入水，用EA萃取3次，合并有机层，经无水硫酸钠干燥，过滤，有机层减压浓缩后，用硅胶柱层析(PE/EA体系＝5:1)分析纯化，得到白色固体，产率78％。1H NMR(400MHz,Chloroform-d)δ7.15(s,1H),6.88–6.79(m,2H),4.69(s,2H),4.20(s,2H),3.81(t,J＝6.5Hz,2H),2.79–2.66(m,2H),2.54(s,1H),2.01–1.96(m,2H).HRMS(DART-TOF)calculated for C14H15ClNO2 +[M+H]+m/z 264.0791,found 264.0793.
中间体化合物K5的合成
在25mL反应瓶中加入KB02(100mg，0.44mmol，1eq)和K2CO3(182mg，1.32mmol，3eq)，用DMF溶解。向搅拌的反应液中滴加溴乙酸叔丁酯(104mg，0.53mmol，1.2eq)，再室温下搅拌反应4h，TLC监测反应结束。处理：反应液经减压浓缩后，加入水，用EA萃取3次，合并有机层，经无水硫酸钠干燥，过滤，有机层减压浓缩后，用硅胶柱层析(PE/EA体系)分析纯化，得到棕色油状物。再向上述所得产物中加入TFA/DCM混合溶剂溶解，在室温下搅拌反应1～2h，TLC监测反应结束后，减压浓缩充分除去溶剂，最终得到灰棕色固体，总产率为65％。1H NMR(400MHz,Chloroform-d)δ7.23(s,1H),6.81–6.75(m,2H),4.68(s,2H),4.20(s,2H),3.83–3.79(m,2H),2.78–2.66(m,2H),2.07–1.94(m,2H).HRMS(DART-TOF)calculated forC13H15ClNO4 +[M+H]+m/z 284.0690,found 284.0689.
中间体化合物K5-N的合成
在25mL圆底烧瓶中加入K5(100mg，0.35mmol，1eq)和HATU(160mg，0.42mmol，1.2eq)，溶于6mL DMF中，将N,N-二异丙基乙胺DIPEA(135mg，1.05mmol，3eq)滴加入反应液中。最后将1-氨基-11-叠氮-3,6,9-三氧杂十一烷(92mg，0.42mmol，1.2eq)的DMF溶液滴加入反应液中，室温下搅拌反应3～4h，TLC监测反应结束。减压浓缩除去溶剂，加水淬灭反应，用EA萃取3次，饱和食盐水反洗2次，合并有机层加无水硫酸钠干燥后，过滤并减压浓缩有机层，然后经硅胶柱层析(DCM/CH3OH体系＝15:1)分离纯化，得到乳白色固体，产率为42％。1HNMR(400MHz,Chloroform-d)δ7.09–7.00(m,2H),6.84(dd,J＝8.9,2.9Hz,1H),6.76(d,J＝2.9Hz,1H),4.50(s,2H),4.05(s,2H),3.71–3.52(m,16H),3.43–3.34(m,2H),2.90–2.83(m,2H),2.21–2.07(m,2H).HRMS(DART-TOF)calculated for C21H30ClN5NaO6 +[M+Na]+m/z506.1782,found506.1785.
试剂与条件：(a)丁二酸酐Succinic anhydride,DCM,50℃,6h；(b)TFA/DCM,rt,1～2h；HATU,DIPEA,DMF,rt,3～4h.
中间体化合物M1的合成
称取L1(1g，2.73mmol，1eq)和丁二酸酐(819mg，8.19mmol，3eq)置于封管中，用DCM作溶剂，将反应液移入50℃油浴锅中加热加压反应约6h。TLC监测反应结束后减压蒸发除去DCM，加入饱和食盐水，并用稀盐酸将反应液PH调节为酸性，再用EA萃取3次，有机层经无水硫酸钠干燥后，减压浓缩经硅胶柱层析(DCM/CH3OH体系＝15:1)纯化，得到产物1.2g，产率为94％。1H NMR(400MHz,Chloroform-d)δ11.38(br,1H),8.45(dd,J＝7.8,1.5Hz,1H),7.80–7.70(m,3H),7.33(q,J＝10.6,8.7Hz,2H),7.05(t,J＝8.7Hz,1H),4.30(s,2H),3.72(m,3H),3.54(d,J＝8.1Hz,2H),3.42–3.22(m,3H),2.42–2.30(m,2H),2.15–2.11(m,2H).HRMS(DART-TOF)calculated for C24H23FN4NaO5 +[M+Na]+m/z 489.2283,found 489.2287.
通用合成步骤I：合成中间体化合物M2～M10
在25mL圆底烧瓶中加入M1(120mg，0.258mmol，1eq)，用无水DMF溶解，再向反应液中加入HATU(118mg，0.31mmol，1.2eq)和DIPEA(100mg，0.77mmol，3eq)，反应液在室温下搅拌反应15～30分钟，称取不同链长和类型的Linker(1.2eq)溶于DMF，缓慢滴加入反应液中，继续在室温下反应3～4h，TLC监测反应结束。后处理：加大量水淬灭反应后，用EA萃取3次，合并有机层，用后饱和食盐水反洗1～2次。有机层用无水硫酸钠干燥，过滤，减压浓缩后用硅胶柱层析(DCM/CH3OH体系＝20:1)分离纯化，得到目标产物，经1H-NMR和高分辨质谱确认化合物结构。
中间体化合物M2的合成
M2的合成参考通用合成步骤I，以M1和N-叔丁氧羰基-1,2-乙二胺为反应原料即可得到中间体M2，淡黄色固体，产率为82％。1H NMR(400MHz,Chloroform-d)δ11.51(br,1H),8.47–8.41(m,1H),7.80–7.68(m,3H),7.34–7.28(m,2H),7.01(t,J＝8.8Hz,1H),6.79(s,1H),5.30(s,1H),4.27(s,2H),3.86–3.09(m,12H),2.75–2.59(m,2H),2.54–2.48(m,2H),2.39–2.20(m,2H),1.39(s,9H).HRMS(DART-TOF)calculated for C31H37FN6NaO6 +[M+Na]+m/z631.2656,found 631.2661.
中间体化合物M3的合成
M3的合成参考通用合成步骤I，以M1和N-叔丁氧羰基-1,3-丙二胺为反应原料即可得到中间体M3，淡黄色固体，产率为75％。1H NMR(400MHz,Chloroform-d)δ11.51(br,1H),8.47–8.41(m,1H),7.80–7.68(m,3H),7.34–7.28(m,2H),7.04(t,J＝8.8Hz,1H),6.79(s,1H),5.30(s,1H),4.27(s,2H),3.86–3.08(m,12H),2.75–2.59(m,2H),2.54–2.48(m,2H),2.39–2.20(m,2H),1.64-1.57(m,2H),1.43(s,9H).HRMS(DART-TOF)calculated forC32H39FN6NaO6 +[M+Na]+m/z 645.2812,found 645.2811.
中间体化合物M4的合成
M4的合成参考通用合成步骤I，以M1和N-叔丁氧羰基-1,4-丁二胺为反应原料即可得到中间体M4，淡黄色固体，产率为63％。1H NMR(400MHz,Chloroform-d)δ10.90(s,1H),8.46(d,J＝7.1Hz,1H),7.80–7.71(m,3H),7.33(q,J＝4.6Hz,2H),7.04(t,J＝9.0Hz,1H),6.34(s,1H),4.74(s,1H),4.29(s,2H),3.90–3.00(m,12H),2.77–2.63(m,2H),2.59–2.47(m,2H),2.08–1.90(m,2H),1.58–1.49(m,4H),1.43(s,9H).HRMS(DART-TOF)calculatedfor C33H41FN6NaO6 +[M+Na]+m/z 659.2969,found 659.2961.
中间体化合物M5的合成
M5的合成参考通用合成步骤I，以M1和N-(5-氨基戊基)氨基甲酸叔丁酯为反应原料即可得到中间体M5，淡黄色固体，产率为69％。1H NMR(400MHz,Chloroform-d)δ11.01(br,1H),8.46–8.40(m,1H),7.78–7.67(m,3H),7.33-7.30(m,2H),7.04(t,J＝8.7Hz,1H),6.35(s,1H),4.76(s,1H),4.29(s,2H),3.84–3.00(m,12H),2.75–2.59(m,2H),2.55–2.49(m,2H),2.09–2.02(m,2H),1.49–1.45(m,2H),1.44(s,9H),1.30–1.23(m,2H).HRMS(DART-TOF)calculated for C34H43FN6NaO6 +[M+Na]+m/z 673.3126,found 673.3119.
中间体化合物M6的合成
M6的合成参考通用合成步骤I，以M1和N-(6-氨基己基)氨基甲酸叔丁酯为反应原料即可得到中间体M6，淡黄色固体，产率为65％。1H NMR(400MHz,Chloroform-d)δ11.21(br,1H),8.48–8.41(m,1H),7.78–7.68(m,3H),7.33-7.30(m,2H),7.02(t,J＝8.8Hz,1H),6.35(s,1H),4.66(s,1H),4.28(s,2H),3.84–3.00(m,12H),2.75–2.59(m,2H),2.55–2.49(m,2H),2.09–2.02(m,2H),1.49–1.45(m,2H),1.44(s,9H),1.32–1.19(m,4H).HRMS(DART-TOF)calculated for C35H45FN6NaO6 +[M+Na]+m/z 687.3282,found 687.3281.
中间体化合物M7的合成
M7的合成参考通用合成步骤I，以M1和N-(8-氨基辛基)氨基甲酸叔丁酯为反应原料即可得到中间体M7，淡黄色固体，产率为79％。1H NMR(400MHz,Chloroform-d)δ10.73(br,1H),8.50–8.43(m,1H),7.80–7.70(m,3H),7.37–7.30(m,2H),7.05(t,J＝9.1Hz,1H),6.44–6.33(m,1H),4.58(s,1H),4.29(s,2H),3.85–3.26(m,12H),3.12–3.06(m,2H),2.62–2.49(m,2H),2.13(t,J＝7.6Hz,2H),1.64–1.56(m,2H),1.44(s,9H),1.31–1.27(m,6H).HRMS(DART-TOF)calculated for C36H47FN6NaO6 +[M+Na]+m/z 701.3448,found 701.3425.
中间体化合物M8的合成
M8的合成参考通用合成步骤I，以M1和[2-(2-氨基乙氧基)乙基]氨基甲酸叔丁酯为反应原料即可得到中间体M8，淡黄色固体粉末，产率为72％。1H NMR(400MHz,Chloroform-d)δ11.08(s,1H),8.49–8.43(m,1H),7.79–7.71(m,3H),7.36–7.31(m,2H),7.04(t,J＝9.1Hz,1H),6.62(s,1H),5.27(s,1H),4.29(s,2H),3.85–3.21(m,16H),2.76–2.62(m,2H),2.58–2.53(m,2H),1.44(s,9H).HRMS(DART-TOF)calculated forC33H41FN6NaO7 +[M+Na]+m/z 675.2918,found 675.2898.
中间体化合物M9的合成
M9的合成参考通用合成步骤I，以M1和2-(2-(2-氨基乙氧基)乙氧基)乙基氨基甲酸叔丁酯为反应原料即可得到中间体M9，淡黄色固体粉末，产率为79％。1H NMR(400MHz,Chloroform-d)δ11.53(s,1H),8.45–8.38(m,1H),7.75–7.67(m,3H),7.36-7.29(m,2H),7.02(t,J＝8.7Hz,1H),6.74(s,1H),5.28(s,1H),4.26(s,2H),3.81–3.17(m,20H),2.73–2.49(m,4H),1.39(s,9H).HRMS(DART-TOF)calculated for C35H45FN6NaO8 +[M+Na]+m/z719.3181,found 719.3162.
中间体化合物M10的合成
M10的合成参考通用合成步骤I，以M1和13-氨基-5,8,11-三氧杂-2-氮杂十三烷酸1,1-二甲基乙酯为反应原料即可得到中间体M10，黄色油状物，产率为67％。1H NMR(400MHz,Chloroform-d)δ11.13(br,1H),8.40–8.35(m,1H),7.78–7.65(m,3H),7.36-7.29(m,2H),6.99(t,J＝8.7Hz,1H),6.69(s,1H),5.28(s,1H),4.28(s,2H),3.80–3.18(m,24H),2.74–2.49(m,4H),1.39(s,9H).HRMS(DART-TOF)calculated for C37H49FN6NaO9 +[M+Na]+m/z763.3443,found 763.3429.
中间体化合物M11的合成
M11的合成参考通用合成步骤I，以M1和16-氨基-5,8,11,14-四氧杂-2-氮杂十六烷酸1,1-二甲基乙酯为反应原料即可得到中间体M11，橙黄色油状物，产率为64％。1H NMR(400MHz,Chloroform-d)δ11.28(s,1H),8.49–8.43(m,1H),7.79–7.71(m,3H),7.36–7.31(m,2H),7.02(t,J＝9.1Hz,1H),6.65(s,1H),5.27(s,1H),4.29(s,2H),3.87–3.15(m,28H),2.73–2.64(m,2H),2.56–2.50(m,2H),1.44(s,9H).HRMS(DART-TOF)calculated forC39H53FN6NaO10+[M+Na]+m/z 807.3705,found 807.3713.
2、PARP PROTACs分子的合成
合成路线1:目标化合物A1～A8和B1～B10的合成
通用合成步骤J：化合物的A1～A8和B1～B10合成
该合成步骤分三步进行：
(1)在25mL圆底烧瓶中加入中间体L3～L10或M2～M10(1.2eq)，用TFA/DCM溶解，室温下搅拌反应1～2h，TLC监测反应结束后，将反应液减压浓缩，除去反应溶剂，得到粗品可直接用于下一步反应。
(2)向上述装有粗产物溶于无水DMF溶解，再向反应液中加入K3(1eq)、HATU(1.2eq)和DIPEA(3eq)继续在室温下反应3～4h，TLC监测反应结束。后处理：加大量水淬灭反应后，用EA萃取3次，合并有机层，用后饱和食盐水反洗两次。有机层用无水硫酸钠干燥，过滤，减压浓缩后用硅胶柱层析(DCM/CH3OH体系＝18:1)分离纯化，得到目标产物用于下一步反应。
(3)将上一步反应得到的目标产物(1eq)置于25mL反应瓶中，用约15mL DCM溶解，再向反应液中加入TEA(3eq)，在冰浴和氮气保护下缓慢滴加氯乙酰氯的DCM溶液(将1.2eq的氯乙酰氯溶解于2mL DCM中)，滴加完于室温下反应30分钟，减压浓缩除去溶剂，加入水，用DCM萃取三次，合并有机层，再用饱和NaCl溶液进行反洗，最后有机层经无水硫酸钠干燥，过滤后减压浓缩，粗品经硅胶柱层析(DCM/CH3OH体系＝20:1)得到目标化合物，经核磁和高分辨质谱确认正确结构。
目标化合物A1的合成
化合物A1的合成参考通用合成步骤J，以L3为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A1，黄色固体，总体产率为31％。1H NMR(400MHz,Chloroform-d)δ10.53(s,1H),8.48–8.41(m,1H),7.83–7.70(m,4H),7.35–7.29(m,3H),7.03(t,J＝9.4Hz,2H),6.88–6.80(m,1H),6.77(d,J＝2.8Hz,1H),4.50(s,2H),4.28(s,2H),4.04(t,J＝6.4Hz,2H),3.95–3.34(m,10H),3.22–3.08(m,2H),2.85(t,J＝6.6Hz,2H),2.76–2.49(m,4H),2.16–2.02(m,2H).HRMS(DART-TOF)calculated for C37H39ClFN7NaO6 +[M+Na]+m/z 754.2532,found 754.2514.
目标化合物A2的合成
化合物A2的合成参考通用合成步骤J，以L4为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A2，总体产率为36％。1H NMR(400MHz,Chloroform-d)δ10.70(s,1H),8.48–8.41(m,1H),7.84–7.56(m,4H),7.34–7.27(m,3H),7.08–6.98(m,2H),6.90–6.83(m,1H),6.81–6.76(m,1H),4.51(s,2H),4.27(s,2H),4.03(t,J＝6.4Hz,2H),3.94–3.15(m,12H),2.89–2.81(m,2H),2.78–2.53(m,4H),2.14–2.05(m,2H),1.74–1.67(m,2H).HRMS(DART-TOF)calculated for C38H41ClFN7NaO6 +[M+Na]+m/z768.2688,found 768.2682.
目标化合物A3的合成
化合物A3的合成参考通用合成步骤J，以L5为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A3，总体产率为32％。1H NMR(400MHz,Chloroform-d)δ10.77(s,1H),8.48–8.41(m,1H),7.82–7.68(m,4H),7.34–7.28(m,3H),7.08–6.97(m,2H),6.87–6.59(m,2H),4.45(s,2H),4.28(s,2H),4.03(t,J＝6.4Hz,2H),3.92–3.09(m,12H),2.91–2.81(m,2H),2.72–2.53(m,4H),2.16–2.05(m,2H),1.59–1.56(m,6H).HRMS(DART-TOF)calculated for C39H43ClFN7NaO6 +[M+Na]+m/z 782.2845,found782.2846.
目标化合物A4的合成
化合物A4的合成参考通用合成步骤J，以L6为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A4，总体产率为29％。1H NMR(400MHz,Chloroform-d)δ10.60(s,1H),8.49–8.42(m,1H),7.82–7.68(m,3H),7.31(dd,J＝7.2,5.2Hz,2H),7.21(s,1H),7.09–6.98(m,2H),6.86–6.68(m,2H),6.68–6.55(m,1H),4.49(s,2H),4.28(s,2H),4.04(t,J＝6.4Hz,2H),3.92–3.03(m,12H),2.85(t,J＝6.7Hz,2H),2.73–2.50(m,4H),2.17–2.06(m,2H),1.56–1.49(m,4H),1.36–1.32(m,4H).HRMS(DART-TOF)calculated for C41H47ClFN7NaO6 +[M+Na]+m/z 810.3158,found 810.3156.
目标化合物A5的合成
化合物A5的合成参考通用合成步骤J，以L7为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A5，总体产率为34％。1H NMR(400MHz,Chloroform-d)δ10.55(s,1H),8.48–8.41(m,1H),7.83–7.70(m,4H),7.33–7.27(m,3H),7.08–6.98(m,2H),6.88–6.80(m,1H),6.80–6.74(m,1H),4.52(s,2H),4.27(s,2H),4.04(s,2H),3.94–3.15(m,16H),2.85(t,J＝6.7Hz,2H),2.76–2.53(m,4H),2.16–2.05(m,2H).HRMS(DART-TOF)calculated for C39H43ClFN7NaO7 +[M+Na]+m/z 798.2794,found 798.2770.
目标化合物A6的合成
化合物A6的合成参考通用合成步骤J，以L8为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A6，总体产率为42％。1H NMR(400MHz,Chloroform-d)δ10.68(s,1H),8.50–8.41(m,1H),7.82–7.69(m,3H),7.55–7.41(m,1H),7.34–7.28(m,2H),7.07–6.98(m,3H),6.86–6.78(m,1H),6.78–6.73(m,1H),4.51(s,2H),4.27(s,2H),4.03(s,2H),3.87–3.29(m,18H),3.19–3.05(m,2H),2.84(t,J＝6.7Hz,2H),2.75–2.50(m,4H),2.16–2.05(m,2H).13C NMR(101MHz,Chloroform-d)δ180.89,167.99,165.45,165.00,155.80,145.55,134.27,133.66(d,J＝9.9Hz),131.64,129.55,129.23,128.76,128.30,127.12,126.14,125.03,123.98,123.35,115.91,114.38,113.09,102.41,84.31,70.33–69.73(m),67.57,61.40,53.72,47.35,46.65,41.40,38.79,37.57,26.08,23.20.HRMS(DART-TOF)calculated for C41H47ClFN7NaO8 +[M+Na]+m/z 842.3056,found842.3062.
目标化合物A7的合成
化合物A7的合成参考通用合成步骤J，以L9为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A7，总体产率为45％。1H NMR(400MHz,Chloroform-d)δ10.57(s,1H),8.48–8.41(m,1H),7.82–7.69(m,3H),7.52(s,1H),7.33–7.27(m,2H),7.13(s,1H),7.07–6.98(m,2H),6.86–6.78(m,1H),6.77–6.72(m,1H),4.51(s,2H),4.27(s,2H),4.03(s,2H),3.91–3.31(m,22H),3.19–3.08(m,2H),2.84(t,J＝6.6Hz,2H),2.75–2.50(m,4H),2.16–2.05(m,2H).13C NMR(101MHz,Chloroform-d)δ180.89,165.44,164.97,158.26,155.87,145.52,133.72,133.56,131.65,129.55,129.25,127.13,126.11,125.01,116.04,114.36,113.18,102.42,84.30,71.10–69.33(m),67.57,63.45,52.57,48.17,46.77,38.84,37.29,26.08,23.20.HRMS(DART-TOF)calculated forC43H51ClFN7NaO9 +[M+Na]+m/z 886.3318,found 886.3317.
目标化合物A8的合成
化合物A8的合成参考通用合成步骤J，以L10为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物A8，总体产率为37％。1H NMR(400MHz,Chloroform-d)δ10.60(s,1H),8.45(dd,J＝7.5,1.8Hz,1H),7.82–7.69(m,3H),7.62–7.51(m,1H),7.33–7.27(m,2H),7.22(s,1H),7.07–6.98(m,2H),6.85–6.80(m,1H),6.78–6.72(m,1H),4.50(s,2H),4.27(s,2H),4.03(s,2H),3.89–3.27(m,26H),3.16–3.12(m,2H),2.84(t,J＝6.7Hz,2H),2.76–2.53(m,4H),2.16–2.06(m,2H).13C NMR(101MHz,Chloroform-d)δ180.89,170.2,167.27,165.72,164.92,158.26,155.87,145.52,133.72,133.56,131.65,129.55,129.25,127.13,126.11,125.01,116.04,114.36,113.18,102.42,84.30,71.10–69.33(m),67.57,63.45,52.57,48.17,46.77,38.84,37.29,26.08,23.20.HRMS(DART-TOF)calculated for C45H55ClFN7NaO10 +[M+Na]+m/z 930.3581,found 930.3588.
目标化合物B1的合成
化合物B1的合成参考通用合成步骤J，以M2为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B1，淡黄色固体，总体产率为33％。1H NMR(400MHz,Chloroform-d)δ10.80(br,1H),8.48–8.42(m,1H),7.75-7.68(m,3H),7.31(d,J＝6.2Hz,3H),7.08–6.99(m,1H),6.77–6.70(m,1H),6.60–6.49(m,2H),6.43–6.35(m,1H),4.46(s,2H),4.38(s,2H),4.27(s,2H),3.78–3.18(m,14H),2.77–2.59(m,4H),2.51–2.43(m,2H),1.90–1.85(m,2H).13C NMR(101MHz,Chloroform-d)δ172.73,170.75,168.81,165.23,165.06,160.49,145.44,134.43,133.69,131.64,129.54,128.34,127.17,124.99,112.93,67.56,49.92,48.82,45.02,41.95,37.61,36.05,35.58,31.28,29.62,28.52,26.89,23.63.HRMS(DART-TOF)calculated for C39H41ClFN7NaO7 +[M+Na]+m/z 796.2637,found 796.2635.
目标化合物B2的合成
化合物B2的合成参考通用合成步骤J，以M3为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B2，总体产率为32％。
1H NMR(400MHz,Chloroform-d)δ11.01(d,J＝10.3Hz,1H),8.48–8.41(m,1H),7.78–7.69(m,3H),7.36–7.27(m,2H),7.24–7.13(m,2H),7.02(t,J＝8.8Hz,1H),6.86–6.70(m,2H),6.66–6.55(m,1H),4.47(s,2H),4.27(s,2H),4.18(s,2H),3.84–3.18(m,14H),2.74–2.53(m,6H),1.94–1.91(m,2H),1.70–1.63(m,2H).13C NMR(101MHz,Chloroform-d)δ172.83,170.79,165.23,165.06,160.49,145.44,134.43,133.69,131.64,129.54,128.34,127.17,124.99,112.93,67.56,45.02,41.95,37.61,36.05,35.58,31.28,29.62,28.52,26.89,23.63.HRMS(DART-TOF)calculated for C40H43ClFN7NaO7 +[M+Na]+m/z 810.2794,found 810.2796.
目标化合物B3的合成
化合物B3的合成参考通用合成步骤J，以M4为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B3，总体产率为41％。1H NMR(400MHz,Chloroform-d)δ10.84(d,J＝11.4Hz,1H),8.48–8.42(m,1H),7.79–7.70(m,3H),7.36–7.28(m,2H),7.03(t,J＝9.0Hz,2H),6.81–6.66(m,3H),6.34–6.22(m,1H),4.47(s,2H),4.28(s,2H),4.19(s,2H),3.87–3.22(m,14H),2.76–2.60(m,4H),2.54–2.49(m,2H),2.03–1.91(m,2H),1.63–1.46(m,4H).13C NMR(101MHz,Chloroform-d)δ172.31,171.04,166,22,160.53,146.21,136.40,133.74,131.68,129.54,128.27,127.19,125.00,120.08,116.32,106.07,67.56,58.94,48.63,41.93,39.06,38.67,36.43,28.83,26.92,26.77,23.63.HRMS(DART-TOF)calculated for C41H45ClFN7NaO7 +[M+Na]+m/z 824.2951,found 824.2943.
目标化合物B4的合成
化合物B4的合成参考通用合成步骤J，以M5为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B4，白色固体，总体产率为50％。1H NMR(400MHz,Chloroform-d)δ10.87(d,J＝9.9Hz,1H),8.48–8.41(m,1H),7.78–7.70(m,3H),7.36–7.28(m,2H),7.03(t,J＝9.1Hz,2H),6.83–6.64(m,3H),6.21(d,J＝18.7Hz,1H),4.47(s,2H),4.28(s,2H),4.19(s,2H),3.81–3.18(m,14H),2.83–2.57(m,4H),2.50(t,J＝6.4Hz,2H),2.00–1.95(m,2H),1.58–1.44(m,4H),1.36–1.29(m,2H).13C NMR(101MHz,Chloroform-d)δ172.22,170.83,168.13,160.43,145.44,134.39,133.69,131.81,131.74,131.65,129.54,128.35,127.18,125.00,116.10,112.78,67.58,41.91,39.27,38.75,37.62,29.14,28.96,28.53,26.89,23.87,23.62.HRMS(DART-TOF)calculated for C42H47ClFN7NaO7 +[M+Na]+m/z838.3107,found 838.3104.
目标化合物B5的合成
化合物B5的合成参考通用合成步骤J，以M6为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B5，白色固体，总体产率为53％。1H NMR(400MHz,Chloroform-d)δ11.02(d,J＝13.8Hz,1H),8.47–8.41(m,1H),7.78–7.69(m,3H),7.35–7.28(m,2H),7.02(t,J＝9.0Hz,1H),6.80–6.72(m,2H),6.65(t,J＝6.0Hz,1H),6.28–6.13(m,1H),4.46(s,2H),4.27(s,2H),4.18(s,2H),3.80–3.18(m,14H),2.74–2.60(m,4H),2.55–2.47(m,2H),2.01–1.93(m,2H),1.54–1.40(m,4H),1.34–1.27(m,4H).13C NMR(101MHz,Chloroform-d)δ170.85,167.26,155.77,145.49,134.42,133.70,131.65,129.54,128.31,127.15,125.01,116.10,112.73,67.59,63.13,45.61,44.99,41.92,40.58,39.27,38.76,37.62,29.70,29.35,28.53,26.89,26.17,23.62,23.44.HRMS(DART-TOF)calculated forC43H49ClFN7NaO7 +[M+Na]+m/z 852.3264,found 852.3261.
目标化合物B6的合成
化合物B6的合成参考通用合成步骤J，以M7为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B6，乳白色固体，总体产率为47％。1H NMR(400MHz,Chloroform-d)δ11.00(br,1H),8.48–8.40(m,1H),7.78–7.69(m,3H),7.36–7.29(m,2H),7.03(t,J＝8.8Hz,2H),6.80–6.74(m,2H),6.58(t,J＝6.0Hz,1H),6.51–6.37(m,1H),4.46(s,2H),4.28(s,2H),4.19(s,2H),3.81–3.24(m,14H),2.77–2.47(m,4H),2.12(t,J＝7.6Hz,2H),1.99–1.95(m,2H),1.63–1.47(m,4H),1.31–1.26(m,6H).13C NMR(101MHz,Chloroform-d)δ173.41,173.25,170.57,170.37,167.31,165,41,165.14,160.54,145.42,158.02,155.47,145.42,134.46,133.67,131.63,128.35,127.16,125.00,123,67,123.45,117.88,112.82,67.62,46.69,45.49,44.91,41.70,41.19,38.99,37.65,36.60,34.92,33.11,29.69,29.45,29.06,28.85,26.90,26.58,25.49,23.62.HRMS(DART-TOF)calculated for C44H51ClFN7NaO7 +[M+Na]+m/z 866.3422,found 866.3420.
目标化合物B7的合成
化合物B7的合成参考通用合成步骤J，以M8为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B7，总体产率为25％。1H NMR(400MHz,Chloroform-d)δ10.98(br,1H),8.47–8.41(m,1H),7.82–7.66(m,3H),7.35–7.28(m,2H),7.12(d,J＝5.5Hz,2H),7.03(t,J＝9.0Hz,1H),6.80–6.72(m,2H),6.58–6.50(m,1H),4.48(s,2H),4.27(s,2H),4.18(s,2H),3.82–3.25(m,18H),2.75–2.59(m,4H),2.54–2.49(m,2H),2.17–2.05(m,2H).13C NMR(101MHz,Chloroform-d)δ170.89,δ164.98,160.30,151.49,145.50,139.56,135.10,134.26,133.69,131.61,129.55,129.23,128.33,127.14,125.00,120.80,116.13(d,J＝21.9Hz),112.91,70.52–69.03(m),67.65,52.86,38.80,37.55,26.89,23.46.HRMS(DART-TOF)calculated for C41H45ClFN7NaO8 +[M+Na]+m/z 840.2791,found 840.2789.
目标化合物B8的合成
化合物B8的合成参考通用合成步骤J，以M9为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B8，总体产率为28％。1H NMR(400MHz,Chloroform-d)δ11.03(br,1H),8.48–8.41(m,1H),7.78–7.69(m,3H),7.34–7.29(m,2H),7.11(d,J＝6.1Hz,2H),7.02(t,J＝8.9Hz,1H),6.80–6.71(m,2H),6.56–6.48(m,1H),4.47(s,2H),4.27(s,2H),4.18(s,2H),3.83–3.18(m,24H),2.77–2.52(m,6H),1.99–1.94(m,2H).13C NMR(101MHz,Chloroform-d)δ172.20,170.74,168.25,165.22,160.56,155.77,145.49,134.40,133.69,131.80,131.64,129.53,128.31,127.16,125.00,116.31,116.08,112.91,70.52–69.03(m),67.66,44.98,41.91,39.29,38.86,37.64,31.12,28.34,26.89,23.62.HRMS(DART-TOF)calculated for C43H49ClFN7NaO9 +[M+Na]+m/z 884.3162,found884.3156.
目标化合物B9的合成
化合物B9的合成参考通用合成步骤J，以M10为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B9，总体产率为31％。1H NMR(400MHz,Chloroform-d)δ10.53(s,1H),8.45(d,J＝7.1Hz,1H),7.82–7.68(m,3H),7.32(dd,J＝8.7,5.1Hz,2H),7.20–7.12(m,1H),7.03(t,J＝9.0Hz,1H),6.82–6.57(m,4H),4.48(s,2H),4.27(s,2H),4.18(s,2H),3.85–3.23(m,28H),2.72–2.50(m,6H),2.03–1.94(m,2H).13C NMR(101MHz,Chloroform-d)δ172.18,170.84,168.15,165.40,160.52,156.17,145.49,134.40,133.69,131.80,131.64,129.53,128.31,127.16,125.00,116.31,116.08,112.91,70.94–69.14(m),67.91,45.02,41.83,39.26,38.34,37.24,32.01,28.54,26.79,23.35.HRMS(DART-TOF)calculated for C45H53ClFN7NaO10 +[M+Na]+m/z 928.3424,found928.3433.
目标化合物B10的合成
化合物B10的合成参考通用合成步骤J，以M11为初始反应原料，经过与K3反应后，再与氯乙酰氯进行反应，最终得到目标产物B10，总体产率为21％。
1H NMR(400MHz,Chloroform-d)δ10.58(s,1H),8.49–8.42(m,1H),7.75(d,J＝18.2Hz,3H),7.34–7.30(m,3H),7.14–6.99(m,2H),6.81–6.61(m,3H),4.48(s,2H),4.27(s,2H),4.18(s,2H),3.90–3.24(m,32H),2.81–2.52(m,6H),2.01–1.94(m,2H).13C NMR(101MHz,Chloroform-d)δ172.23,170.97,169.01,165.89,160.71,157.17,146.12,134.89,133.23,131.54,131.13,129.53,128.31,127.16,125.00,116.31,116.08,112.91,70.28–69.09(m),67.89,44.87,41.44,39.43,38.26,37.24,32.01,28.54,26.73,23.61.HRMS(DART-TOF)calculated for C47H57ClFN7NaO11+[M+Na]+m/z 972.3686,found972.3683.
合成路线二：目标化合物C1～C9的合成
通用合成步骤K：化合物C1～C9的合成
在25mL圆底烧瓶中加入中间体C1-1～C9-1(1eq)，用约15mL DCM溶解，再向反应液中加入TEA(1eq)，在冰浴中和氮气保护下缓慢滴加氯乙酰氯的DCM溶液(将1.2eq的氯乙酰氯溶解于2mL DCM中)，滴加完于室温下反应30分钟，减压浓缩除去溶剂，加入水，用DCM萃取三次，合并有机层，再用饱和NaCl溶液进行反洗，最后有机层经无水硫酸钠干燥，过滤后减压浓缩，粗品经硅胶柱层析(DCM/CH3OH体系＝20:1)得到目标化合物，经核磁和高分辨质谱确认正确结构。
目标化合物C1的合成
化合物C1的合成参考通用合成步骤K，将C1-1和氯乙酰氯作为反应原料即可得到目标产物C1，乳白色固体，产率87.2％。1H NMR(400MHz,Chloroform-d)δ11.38(d,J＝13.8Hz,1H),8.48–8.41(m,1H),7.78–7.59(m,4H),7.37–7.29(m,2H),7.03(t,J＝9.1Hz,2H),6.80–6.74(m,2H),4.51(s,2H),4.28(s,2H),4.23–4.11(m,4H),3.82–3.31(m,10H),2.75–2.60(m,2H),1.97–1.93(m,2H).13C NMR(101MHz,Chloroform-d)δ168.26,166.45,165.26,165.12,160.76,158.22,155.75,145.46,134.56(d,J＝3.4Hz),133.68,131.91,131.64,129.53,129.30,128.29,127.15,124.99,123.37(d,J＝16.1Hz),116.22(d,J＝21.9Hz),114.89,112.63,67.47,46.68(d,J＝16.7Hz),44.69,44.20,42.21,41.75,40.77,37.64,29.69,26.85,23.60.HRMS(DART-TOF)calculated for C35H34ClFN6NaO6 +[M+Na]+m/z711.2110,found 711.2113.
目标化合物C2的合成
化合物C2的合成参考通用合成步骤K，将C2-1和氯乙酰氯作为反应原料即可得到目标产物C2，乳白色固体，产率86.7％。1H NMR(400MHz,Chloroform-d)δ10.96(br,1H),8.48–8.42(m,1H),7.79–7.69(m,3H),7.44–7.28(m,3H),7.04(t,J＝8.8Hz,1H),6.79–6.73(m,2H),4.45(s,2H),4.28(s,2H),4.18(s,2H),3.85–3.30(m,12H),2.75–2.55(m,4H),1.98–1.89(m,2H).13C NMR(101MHz,Chloroform-d)δ170.02,168.57,166.46,165.26,164.92,160.71,158.23,155.35,145.76,134.49,133.71,132.04,131.52,129.62,129.01,128.29,127.15,125.23,123.65,116.23,114.91,112.73,67.49,46.75,44.56,44.32,42.37,41.57,40.89,37.62,29.65,26.75,23.58.HRMS(DART-TOF)calculated forC36H36ClFN6NaO6 +[M+Na]+m/z 725.2426,found 725.2416.
目标化合物C3的合成
化合物C3的合成参考通用合成步骤K，将C3-1和氯乙酰氯作为反应原料即可得到目标产物C3，乳白色固体，产率90.3％。1H NMR(400MHz,Chloroform-d)δ11.13(br,1H),8.48–8.41(m,1H),7.79–7.69(m,3H),7.36–7.28(m,2H),7.11–6.98(m,3H),6.81–6.74(m,2H),4.45(s,2H),4.28(s,2H),4.17(s,2H),3.83–3.21(m,12H),2.72–2.67(m,2H),2.48–2.32(m,2H),1.98–1.89(m,4H).13C NMR(101MHz,Chloroform-d)δ172.23,168.91,167.23,165.21,164.47,159.98,158.23,155.35,145.76,134.49,132.89,132.21,131.12,129.62,129.01,128.29,127.15,125.23,123.65,118.03,115.01,113.11,67.49,49.92,48.23,46.75,44.56,44.32,42.37,39.32,37.62,31.45,29.65,27.15,26.53,22.38.HRMS(DART-TOF)calculated for C37H38ClFN6NaO6 +[M+Na]+m/z 739.2423,found 739.2426.
目标化合物C4的合成
化合物C4的合成参考通用合成步骤K，将C4-1和氯乙酰氯作为反应原料即可得到目标产物C4，乳白色固体，产率91.6％。1H NMR(400MHz,Chloroform-d)δ11.00(s,1H),8.49–8.42(m,1H),7.79–7.69(m,3H),7.36–7.28(m,2H),7.03(t,J＝8.8Hz,2H),6.85–6.59(m,3H),4.47(s,2H),4.28(s,2H),4.18(s,2H),3.84–3.22(m,12H),2.75–2.61(m,2H),2.41–2.28(m,2H),2.01–1.93(m,2H),1.69–1.59(m,4H).13C NMR(101MHz,Chloroform-d)δ171.34,168.02,165.05,163.47,160.55,158.25,155.79,145.48,134.38,133.69,131.65,129.54,128.33,127.19,125.00,116.11,114.31,112.98,67.57,46.78,41.90(d,J＝54.0Hz),38.60,37.64,32.01,29.05,26.89,23.62,21.94.HRMS(DART-TOF)calculatedfor C38H40ClFN6NaO6 +[M+Na]+m/z 753.2357,found 753.2352.
目标化合物C5的合成
化合物C5的合成参考通用合成步骤K，将C5-1和氯乙酰氯作为反应原料即可得到目标产物C5，淡黄色固体，产率89.5％。1H NMR(400MHz,Chloroform-d)δ11.21(br,1H),8.48–8.41(m,1H),7.78–7.68(m,3H),7.34–7.28(m,2H),7.13(s,1H),7.02(t,J＝8.8Hz,1H),6.79–6.63(m,3H),4.45(s,2H),4.28(s,2H),4.17(s,2H),3.83–3.27(m,12H),2.76–2.60(m,2H),2.42–2.26(m,2H),2.00-1.87(m,2H),1.69–1.55(m,4H),1.39–1.35(m,2H).13CNMR(101MHz,Chloroform-d)δ172.03,168.92,167.43,165.61,164.23,159.78,157.91,154.35,145.86,134.21,132.89,132.24,131.62,129.03,128.87,128.29,127.15,125.93,123.71,117.93,115.21,113.30,65.49,49.93,48.93,46.79,43.86,43.32,42.37,39.42,36.96,31.51,28.95,27.25,26.53,25.21,22.38,21.32.HRMS(DART-TOF)calculated forC39H42ClFN6NaO6 +[M+Na]+m/z 767.2536,found 767.2530.
目标化合物C6的合成
化合物C6的合成参考通用合成步骤K，将C6-1和氯乙酰氯作为反应原料即可得到目标产物C6，淡黄色固体，产率84.2％。1H NMR(400MHz,Chloroform-d)δ10.91(br,1H),8.48–8.43(m,1H),7.79–7.69(m,3H),7.34–7.28(m,2H),7.15(s,1H),7.03(t,J＝8.8Hz,1H),6.80–6.74(m,2H),6.58(t,J＝6.0Hz,1H),4.46(s,2H),4.28(s,2H),4.19(s,2H),3.83–3.22(m,12H),2.78–2.63(m,2H),2.38–2.25(m,4H),1.66–1.50(m,4H),1.36–1.26(m,6H).13C NMR(101MHz,Chloroform-d)δ173.25,170.57,170.37,167.31,165,41,165.14,160.54,145.42,158.02,155.47,145.42,134.46,133.67,131.63,128.35,127.16,125.00,123,67,123.45,117.88,112.82,67.62,49.98,48.73,46.69,45.49,44.91,41.70,41.19,38.99,37.65,36.60,34.92,33.11,29.69,29.45,29.06,28.85,26.90,26.58,25.49,23.62,21.66.HRMS(DART-TOF)calculated for C41H46ClFN6NaO6 +[M+Na]+m/z 795.3049,found 795.3019.
目标化合物C7的合成
化合物C7的合成参考通用合成步骤K，将C7-1和氯乙酰氯作为反应原料即可得到目标产物C7，淡黄色固体，产率75.1％。1H NMR(400MHz,Chloroform-d)δ10.56(br,1H),8.49–8.42(m,1H),7.80–7.69(m,3H),7.34–7.28(m,2H),7.15(s,1H),7.03(t,J＝8.8Hz,1H),6.81–6.70(m,2H),6.61–6.49(m,1H),4.47(s,2H),4.28(s,2H),4.19(s,2H),3.89–3.24(m,12H),2.79–2.64(m,2H),2.37–2.23(m,2H),2.01–1.96(m,2H),1.66–1.49(m,4H),1.33–1.26(m,10H).13C NMR(101MHz,Chloroform-d)δ172.25,170.37,168.99,168.31,165,41,163.14,160.54,158.02,155.47,145.42,134.46,133.67,131.63,128.35,127.16,125.04,123,67,123.45,117.78,112.82,65.22,49.98,48.73,46.69,45.49,44.91,41.70,41.19,38.99,37.65,36.60,34.92,33.11,29.69,29.45,29.06,28.85,26.90,26.58,25.49,23.62,22.31.HRMS(DART-TOF)calculated for C42H48ClFN6NaO6 +[M+Na]+m/z809.3205,found 809.3202.
目标化合物C8的合成
化合物C8的合成参考通用合成步骤K，将C8-1和氯乙酰氯作为反应原料即可得到目标产物C8，淡黄色固体，产率70.8％。1H NMR(400MHz,Chloroform-d)δ11.16(br,1H),8.48–8.41(m,1H),7.78–7.68(m,3H),7.35–7.28(m,2H),7.15(s,1H),7.03(t,J＝8.8Hz,1H),6.80–6.73(m,2H),6.57(t,J＝6.0Hz,1H),4.46(s,2H),4.28(s,2H),4.18(s,2H),3.84–3.20(m,12H),2.72–2.68(m,2H),2.36–2.24(m,2H),1.99–1.94(m,2H),1.66–1.52(m,4H),1.29–1.20(m,12H).13C NMR(101MHz,Chloroform-d)δ171.98,168.92,167.92,165.29,160.69,156.25,145.55,134.41,133.71,131.81,131.73,131.64,129.54,128.27,127.16,125.02,112.84,67.62,46.85,45.19,41.66,39.08,37.66,33.26,29.49,29.33,29.29,29.12,26.90,26.77,25.16,23.62.HRMS(DART-TOF)calculated for C43H50ClFN6NaO6 +[M+Na]+m/z 823.3362,found 823.3358.
目标化合物C9的合成
化合物C9的合成参考通用合成步骤K，将C9-1和氯乙酰氯作为反应原料即可得到目标产物C9，淡黄色固体，产率67.2％。1H NMR(400MHz,Chloroform-d)δ10.65(br,1H),8.49–8.42(m,1H),7.79–7.69(m,3H),7.34–7.28(m,2H),7.15(s,1H),7.03(t,J＝8.8Hz,1H),6.81–6.74(m,2H),6.55(t,J＝6.0Hz,1H),4.47(s,2H),4.28(s,2H),4.19(s,2H),3.83–3.23(m,12H),2.76–2.62(m,2H),2.39–2.25(m,2H),2.00–1.96(m,2H),1.64–1.49(m,4H),1.29–1.24(m,14H).13C NMR(101MHz,Chloroform-d)δ172.98,170.27,168.65,165.29,160.69,145.55,134.41,133.71,131.81,131.73,131.64,129.54,128.27,127.16,125.02,112.84,67.62,46.85,45.19,41.66,39.08,37.66,33.26,29.49,29.33,29.29,29.12,28.63,26.90,26.77,25.16,23.62.HRMS(DART-TOF)calculated for C44H52ClFN6NaO6 +[M+Na]+m/z 837.3518,found 837.3516.
目标化合物C8-M的合成
化合物C8-M的合成参考通用合成步骤K，将C8-1和丙酰氯作为反应原料即可得到目标产物C8-M，乳白色固体，产率89.3％。1H NMR(400MHz,Chloroform-d)δ10.88(br,1H),8.50–8.41(m,1H),7.80–7.69(m,3H),7.34–7.28(m,2H),7.09–6.98(m,1H),6.77–6.68(m,2H),6.56(t,J＝5.8Hz,1H),4.46(s,2H),4.28(s,2H),3.85–3.26(m,12H),2.71–2.66(m,2H),2.55–2.31(m,4H),1.99–1.90(m,2H),1.64–1.51(m,4H),1.31–1.26(m,12H),1.18–1.10(m,3H).13C NMR(101MHz,Chloroform-d)δ172.01,168.96,168.60,163.41,159.89,145.55,134.41,133.71,132.02,131.62,131.14,129.52,128.27,127.16,125.02,112.84,67.62,46.85,45.19,41.66,39.08,37.66,33.26,29.49,29.33,29.29,29.12,26.90,26.77,25.16,23.62,11.23.HRMS(DART-TOF)calculated for C44H53FN6NaO6 +[M+Na]+m/z803.3908,found 803.3907.
合成路线三：目标化合物D1～D3的合成
通用步骤L：化合物D1～D3的合成
在25mL圆底烧瓶中加入叠氮化合物(1eq)，用THF：H2O＝2：1混合液溶解，加入CuSO4.5H2O(1.5eq)后溶液呈橙色，在室温下搅拌反应15min溶液变浑浊，再向搅拌的溶液中加入炔烃类化合物(1.2eq)，最后加入维生素钠(2eq)，观察反应液由浑浊变澄清，TLC监测反应结束。处理反应液：向反应液中加水，用DCM萃取3次，收集有机层，用无水硫酸钠干燥，减压浓缩得到粗品。经硅胶柱层析(DCM:CH3OH体系＝20:1)分离得到目标化合物，经高分辨质谱和核磁确认正确结构。
目标化合物D1的合成
化合物D1的合成参考通用合成步骤L，以N1和K5-N为反应原料即可得到目标产物D1，淡黄色固体，产率52.2％。1H NMR(400MHz,Chloroform-d)δ10.61(s,1H),8.49–8.40(m,1H),7.82–7.68(m,4H),7.32–7.28(m,2H),7.10–6.97(m,3H),6.82(dd,J＝9.0,2.9Hz,1H),6.74(d,J＝2.9Hz,1H),4.54(t,J＝5.1Hz,2H),4.50(s,2H),4.27(s,2H),4.04(s,2H),3.90–3.76(m,6H),3.63–3.53(m,14H),3.35–3.30(m,2H),2.85(t,J＝6.7Hz,2H),2.66–2.47(m,4H),2.20–2.06(m,2H).HRMS(DART-TOF)calculated for C44H51ClFN9NaO8 +[M+Na]+m/z 910.3431,found 910.3428.
目标化合物D2的合成
化合物D2的合成参考通用合成步骤L，以N2和K5-N为反应原料即可得到目标产物D2，乳白色固体，产率57.5％。1H NMR(400MHz,Chloroform-d)δ10.65(br,1H),8.49–8.43(m,1H),7.84–7.69(m,3H),7.53–7.46(m,1H),7.37–7.29(m,2H),7.09–7.00(m,3H),6.86–6.79(m,1H),6.74(d,J＝2.9Hz,1H),4.50(d,J＝4.7Hz,4H),4.29(s,2H),4.04(t,J＝6.3Hz,2H),3.88–3.24(m,24H),2.85(t,J＝6.7Hz,2H),2.78–2.73(m,2H),2.45–2.33(m,2H),2.17–2.06(m,2H),1.80–1.64(m,4H).13C NMR(101MHz,Chloroform-d)δ180.88,167.87,165.45,155.85,145.49,134.31,133.59,131.65,129.55,128.74,128.32,127.15,126.14,125.03,114.34,113.20,102.41,84.30,71.14–68.96(m),67.61,59.06,50.19,47.00,38.86,36.42,32.85,29.69,26.10,24.55,23.22.HRMS(DART-TOF)calculated forC48H57ClFN9NaO9 +[M+Na]+m/z 980.3850,found 980.3853.
目标化合物D3的合成
化合物D3的合成参考通用合成步骤L，以K4和L2-N为反应原料即可得到目标产物D3，乳白色固体，产率52.1％。1H NMR(400MHz,Chloroform-d)δ10.93(s,1H),8.48–8.41(m,1H),7.82(s,1H),7.78–7.69(m,3H),7.44(s,1H),7.34–7.27(m,2H),7.01(t,J＝9.1Hz,2H),6.83(d,J＝8.9Hz,2H),5.17(s,2H),4.55(t,J＝5.1Hz,2H),4.27(s,2H),4.18(s,2H),3.92–3.77(m,6H),3.63–3.29(m,16H),3.10–3.06(m,2H),2.75–2.47(m,4H),1.99–1.90(m,2H).13C NMR(101MHz,Chloroform-d)δ180.88,δ164.94,160.47,158.24,155.78,145.49,143.70,134.34,133.69,131.59(d,J＝8.2Hz),129.55,129.22,128.34,127.15,125.03,123.92(d,J＝31.4Hz),116.24,116.03,112.78,71.16–70.08(m),69.82,69.40,62.19,61.16,53.34,52.85,50.34,46.81,41.77,38.73,37.63,29.69,26.89,23.67.HRMS(DART-TOF)calculated for C44H51ClFN9NaO8 +[M+Na]+m/z 910.3431,found 910.3428.
实施例2化合物活性测试
(1)实验方法：
1)细胞来源与培养
本发明中使用的细胞主要包括：人乳腺癌细胞(MDA-MB-436，MDA-MB-468，MDA-MB-231)，人胰腺癌细胞(Capan-1)，人结直肠癌细胞(SW620)，人宫颈癌细胞(Hela),人肝癌细胞(HepG2)，人非小细胞肺癌细胞(H3122和H2228)，人卵巢癌细胞(SKOV3)，人髓性单核细胞白血病细(MV-4-11)，人正常肝细胞(L-O2)均来源于中国科学院典型培养物保藏委员会细胞库(https://www.cellbank.org.cn/)。细胞培养方法如下：
①人乳腺癌细胞(MDA-MB-468，MDA-MB-231)，人结直肠癌细胞(SW620)，人宫颈癌细胞(Hela)人非小细胞肺癌细胞(H3122，H2228)，人卵巢癌细胞(SKOV3)，人髓性单核细胞白血病细(MV-4-11)，人正常肝细胞(LO2)的培养条件：10％FBS，89％DMEM高糖培养基，1％双抗(Penicillin-Streptomycin Liquid，100×)。
②人乳腺癌细胞(MDA-MB-436)，人肝癌细胞(HepG2)的培养条件：10％FBS，89％RPMI-1640培养基，1％双抗(Penicillin-Streptomycin Liquid，100×)。
③人胰腺癌细胞(Capan-1)培养条件：20％FBS，79％IMDM培养基，1％双抗(Penicillin-Streptomycin Liquid，100×)。
上述所有细胞均在细胞孵箱中培养，培养条件：5％CO2，37℃饱和湿度。根据细胞生生长密度，一般细胞每周传代2～3次，细胞复苏传代达到最优状态时可以进行相关细胞实验。
2)MTT法测定细胞增殖抑制实验
①称取待检测的化合物及阳性药对照品(2～3mg)，使用摩尔浓度计算器，用DMSO做溶剂，将化合物配制成浓度为10mM和20mM，所有化合物贮存在-20℃冰箱中保存。
②将处于对数生长期的待检测细胞作传代处理后，用细胞计数仪计算细胞浓度。随后调整细胞悬液浓度，将细胞均匀接种在96孔板中，使得接种细胞数约为800～2000个/每孔。其中H2228，SW620，Hela，MV-4-11，HepG2为800个/孔；MDA-MB-468，MDA-MB-231，SKOV3为1000个/孔；MDA-MB-436，LO2为1500个/孔；Capan-1，H3122为2000个/每孔，边缘孔用无菌PBS填充，置于细胞培养孵箱中培养12～24h至细胞贴壁(悬浮细胞无需考虑)。
③将待测化合物加入含血清的培养基中稀释，使得最初浓度为20μM或10μM，依次按3倍稀释9个浓度，每个浓度设置2～3个复孔，将对应浓度药物加入含有细胞的96孔板中。
④加药完毕后，将96孔板置于5％CO2,37℃饱和湿度培养孵箱中孵育5-7天，其中Capan-1需孵育约10天。
⑤终止培养，在避光条件下，向每孔中加入20μL MTT溶液(用PBS配制成5mg/mL)，放入孵箱中继续孵育4h。
⑥在显微镜下观察细胞中沉积有明显的蓝紫色结晶甲瓒(Formazan)后，用排枪去除96孔板中的混合培养液，加入DMSO溶液(100～150μL/孔)溶解细胞中的甲瓒，置摇床上低速振荡10min，使结晶物充分溶解。
⑦用酶联免疫检测仪在562nm波长处测定其光吸收值(OD值)，每个实验重复3次，取平均值为最终结果。采用Excel计算细胞增殖抑制率，计算公式：细胞增殖抑制率＝[1-(A样品孔-A空白孔)/(A对照孔-A空白孔)]，再利用GraphPad Prism 8计算待测化合物的细胞增殖抑制活性IC50。
(2)实验结果
实验结果见表1、表2、表3和表4。
表1化合物A1～A8的体外抗肿瘤细胞增殖活性
表2化合物B1～B10的体外抗肿瘤细胞增殖活性
表3化合物D1～D3的体外抗肿瘤细胞增殖活性
表4化合物C1～C9的体外抗肿瘤细胞增殖活性
图1为目标化合物对MDA-MB-231细胞内的PARP1/2降解活性。化合物的浓度为1μM在细胞中作用24h。
由以上IC50数据可知，大多目标化合物对不同肿瘤细胞株有一定的增殖抑制活性。其中C系列的抗肿瘤的活性较好，化合物C8对PARP2的降解活性最优，且对多种肿瘤细胞表现出优于阳性药奥拉帕尼的抗肿瘤活性。因此我们选择C8作为候选PROTAC分子进行后续抗肿瘤机制研究。其结果见图2。
图2为化合物C8体外抗增殖活性及适用性评估。其中，A为PARP1的活性测试；B，C为四种类型癌细胞中C8和Olaparib的IC50值；D为C8、Olaparib和C8-M在不同人源肿瘤细胞系中的IC50值；E为化合物C8、C8-M或Olaparib处理的正常肝细胞L-O2的细胞存活率
首先，我们通过PARP1活性测定来评估C8与PARP的结合亲和力。研究证实，与抑制剂Olaparib的IC50值为3.99nM相比，C8与PARP1的结合IC50值为2.74nM，这表明C8没有脱靶，与PARP1仍然具有很强结合亲和力(图2的A图)。
体外抗肿瘤细胞增殖活性主要采用MTT方法进行测试，目前研究表明，BRCA1和BRCA2突变的癌细胞对PARP抑制剂更为敏感，因此我们评估了化合物C8对MDA-MB-436(BRCA1-/-)、Capan-1(BRCA2-/-)、MDA-MB-468(PTEN-/-)和MDA-MB-231(wild type)四种不同的肿瘤细胞的增殖抑制活性。从图2的B图结果分析可知，C8对MDA-MB-436和Capan-1细胞的增值抑制活性IC50分别为26nM和61nM，表现出与PARP1/2抑制剂Olaparib相似的活性(20nM和53nM)。但有趣的是，C8能有效抑制MDA-MB-468和MDA-MB-231乳腺癌细胞的增殖，其细胞增值抑制活性IC50分别为0.69μM和0.53μM，抑制活性远高于Olaparib的抑制活性(4.19μM和4.47μM)，活性提高约6～8.5倍(如图2的C图)。
随后，我们根据DCAF16在肿瘤细胞中的表达量以及PARP过量表达癌细胞的分析[229]，我们筛选了多种人源肿瘤细胞，包括Hela(人宫颈癌细胞)、HepG2(人肝癌细胞)、SKOV3(人卵巢癌细胞)、H2228和H3122(人非小细胞肺癌)、SW620(人结直肠癌)和MV-4-11(人白血病细胞)等7种细胞，检测C8、C8-M和抑制剂Olaparib的增殖抑制活性。结果如图2的D图所示，C8在所有测试细胞中比Olaparib更有效，特别是在DCAF16过量表达的癌细胞如结肠癌和肺腺癌中，其活性IC50值提升了2～5倍，而其非活性化合物C8-M比C8的活性降低了2～3倍。另外，我们在正常人源L-O2肝细胞中对化合物C8进行初步安全性的评估，结果如图2的E图所示。C8在高浓度(>5μM)时表现比C8-M和Olaparib更高的抑制正常细胞增殖活性，这说明C8分子结构中KB02配体结构上共价键氯原子的引入会导致毒性增加。但是，在低浓度时，C8与PARP抑制剂Olaparib的细胞存活率相当。因此，综合考虑C8良好的降解PARP2活性，以及对MDA-MB-231肿瘤细胞优秀的抗增殖活性，我们认为C8对正常肝细胞的毒性是可以接受的。
实施例3 C8体内抗肿瘤活性研究
(1)实验方法
基于化合物C8在MDA-MB-231细胞中表现出优秀的体外抗肿瘤细胞增殖活性，以及对肿瘤细胞核内PARP2的良好降解作用，我们选用MDA-MB-231异种移植瘤模型研究C8在体内的抗肿瘤活性。首先，我们将Balb/c nude雌性裸鼠皮下接种MDA-MB-231细胞建立异种移植肿瘤模型，待肿瘤生长至约100mm3时，将其随机分为5组，每组5只。为了研究C8在不同剂量与不同给药方式的抗肿瘤活性，我们选择已上市的PARP抑制剂Olaparib作为阳性对照药。因此，在本实验中，将实验组分别设置为：空白组(Vehicle control，100μL，腹腔注射ip)，阳性药组(Olaparib，100mg/kg，灌胃ig)，C8口服组(100mg/kg，灌胃ig)，C8低剂量组(12.5mg/kg，腹腔注射ip)和C8高剂量组(25mg/kg，腹腔注射ip)。所有实验组每天给药一次(灌胃组的给药体积为200μL，腹腔注射组的给药体积为100μL)，连续给药21天，给药期间关注小鼠的进食、体重以及肿瘤生长情况，每隔两天对小鼠体重(Body weight)和肿瘤大小进行测量，并记录数据。用游标卡尺测量并记录肿瘤长径(a，mm)和垂直于长径的方向的肿瘤短径(b，mm)。肿瘤体积(Tumor volume)计算公式为：V(mm3)＝a×b×b/2肿瘤抑瘤率的计算公式为：100×[1-(最终肿瘤体积实验组-肿瘤初始体积实验组)/(最终肿瘤体积空白组-肿瘤初始体积空白组)]。数据处理用GraphPad Prism 8进行统计和计算。
(2)实验结果
实验结果如图3所示，与空白组相比，当以相同的给药方式和剂量(口服给药，剂量为100mg/kg)时，化合物C8对肿瘤的抑制率为63.5％，优于阳性药Olaparib对肿瘤的抑制率57.8％。而C8以腹腔注射方式给药时，C8能够以剂量依赖性的抑制肿瘤的生长，当腹腔注射C8剂量为25mg/kg和12.5mg/kg时，对肿瘤生长的抑制率分别为98.3％和82.9％，远高于阳性药Olaparib抑制率57.8％。另外，C8口服给药剂量比腹腔注射增加8倍后，其肿瘤生长抑制率仍低于腹腔注射组。由此，我们可以从本次实验中得出结论：C8在腹腔注射低剂量下即可有效抑制MDA-MB-231异种移植肿瘤的生长，随剂量增加肿瘤抑制效果增加，有剂量依赖性；C8的腹腔注射药物生物利用度远高于口服，C8口服生物利用度较差。
根据小鼠给药期间的体重变化，我们可以初步判断化合物C8的毒性。因此，在给药期间我们测量并记录了小鼠的体重变化。在给药期间实验小鼠均没有发生死亡，根据实验结果分析如图4所示，我们发现阳性药组，C8口服组和C8低剂量腹腔注射组小鼠的体重均无明显变化，精神状态良好且能正常进食，这说明化合物在低剂量或低吸收的情况下，对小鼠没有明显的毒性。
Claims (9)
1.共价PARP PROTACs衍生物，其特征在于，其结构通式为式Ⅰ所示：
其中，X选自卤素原子、甲基、羟基、氨基、硝基或可与目标蛋白的氨基酸残基发生共价反应的亲电基团；linker为直链烷烃类链、PEG类链、醚类链或三氮唑类链中的至少一种。
2.根据权利要求1所述的共价PARP PROTACs衍生物，其特征在于：所述X为卤素原子或可与目标蛋白的氨基酸残基发生共价反应的亲电基团；其中，所述可与目标蛋白的氨基酸残基发生共价反应的亲电基团，包括丙烯酰胺基、α-卤酮基、氰基乙酰胺基、巯基、环氧化物基、乙烯基砜基或活化乙炔基。
3.根据权利要求2所述的共价PARP PROTACs衍生物，其特征在于：所述X为氯原子。
4.根据权利要求1～3任一项所述的共价PARP PROTACs衍生物，其特征在于：所述直链烷烃的结构式为式⑴、式⑵或式⑶所示：
式⑴为
其中n1为1～12中的任意整数；
式⑵为
其中n2为1～5中的任意整数；
式⑶为
其中n3为1～7中的任意整数；
所述PEG单元链的结构式为式⑷或式⑸所示：
式⑷为
其中n4为1～4中的任意整数；
式⑸为
其中n5为1～4中的任意整数；
所述三氮唑类链的结构式为
其中n6、n7为1～5中的任意整数。
5.根据权利要求1所述的共价PARP PROTACs衍生物，其特征在于，所述共价PARPPROTACs衍生物的结构式为以下结构式中的任意一种：
6.权利要求1～5任一项所述的共价PARP PROTACs衍生物在制备抗肿瘤药物中的应用。
7.根据权利要求6所述的共价PARP PROTACs衍生物在制备抗肿瘤药物中的应用，其特征在于：所述抗肿瘤药物为招募DCAF16 E3连接酶的药物。
8.根据权利要求6所述的共价PARP PROTACs衍生物在制备抗肿瘤药物中的应用，其特征在于：所述抗肿瘤药物为靶向降解PARP2蛋白的药物。
9.根据权利要求6所述的共价PARP PROTACs衍生物在制备抗肿瘤药物中的应用，其特征在于：所述抗肿瘤药物为靶向降解PARP1和PARP2蛋白的药物。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111319693.7A CN116102534A (zh) | 2021-11-09 | 2021-11-09 | 共价PARP PROTACs衍生物及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111319693.7A CN116102534A (zh) | 2021-11-09 | 2021-11-09 | 共价PARP PROTACs衍生物及其应用 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116102534A true CN116102534A (zh) | 2023-05-12 |
Family
ID=86258422
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111319693.7A Pending CN116102534A (zh) | 2021-11-09 | 2021-11-09 | 共价PARP PROTACs衍生物及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116102534A (zh) |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20150301058A1 (en) * | 2012-11-26 | 2015-10-22 | Caris Science, Inc. | Biomarker compositions and methods |
| CN110551102A (zh) * | 2018-06-01 | 2019-12-10 | 四川大学 | Alk共价抑制剂及其用途 |
| CN110684015A (zh) * | 2018-07-06 | 2020-01-14 | 四川大学 | 靶向alk的protac及其应用 |
| CN110845474A (zh) * | 2019-11-07 | 2020-02-28 | 四川大学 | 一种靶向i型prmt的化合物及其制备方法和应用 |
| CN111606969A (zh) * | 2020-05-13 | 2020-09-01 | 四川大学 | 一种parp1蛋白降解剂及其在抗肿瘤中的应用 |
| WO2021074414A1 (en) * | 2019-10-16 | 2021-04-22 | Cemm - Forschungszentrum Für Molekulare Medizin Gmbh | Oxazole and thioazole-type cullin ring ubiquitin ligase compounds and uses thereof |
| WO2021074418A1 (en) * | 2019-10-16 | 2021-04-22 | Cemm - Forschungszentrum Für Molekulare Medizin Gmbh | Carbazole-type cullin ring ubiquitin ligase compounds and uses thereof |
| WO2021119159A1 (en) * | 2019-12-10 | 2021-06-17 | Kymera Therapeutics, Inc. | Irak degraders and uses thereof |
-
2021
- 2021-11-09 CN CN202111319693.7A patent/CN116102534A/zh active Pending
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20150301058A1 (en) * | 2012-11-26 | 2015-10-22 | Caris Science, Inc. | Biomarker compositions and methods |
| CN110551102A (zh) * | 2018-06-01 | 2019-12-10 | 四川大学 | Alk共价抑制剂及其用途 |
| CN110684015A (zh) * | 2018-07-06 | 2020-01-14 | 四川大学 | 靶向alk的protac及其应用 |
| WO2021074414A1 (en) * | 2019-10-16 | 2021-04-22 | Cemm - Forschungszentrum Für Molekulare Medizin Gmbh | Oxazole and thioazole-type cullin ring ubiquitin ligase compounds and uses thereof |
| WO2021074418A1 (en) * | 2019-10-16 | 2021-04-22 | Cemm - Forschungszentrum Für Molekulare Medizin Gmbh | Carbazole-type cullin ring ubiquitin ligase compounds and uses thereof |
| CN110845474A (zh) * | 2019-11-07 | 2020-02-28 | 四川大学 | 一种靶向i型prmt的化合物及其制备方法和应用 |
| WO2021119159A1 (en) * | 2019-12-10 | 2021-06-17 | Kymera Therapeutics, Inc. | Irak degraders and uses thereof |
| CN111606969A (zh) * | 2020-05-13 | 2020-09-01 | 四川大学 | 一种parp1蛋白降解剂及其在抗肿瘤中的应用 |
Non-Patent Citations (6)
| Title |
|---|
| CHUNLAN PU，等: "Selective degradation of PARP2 by PROTACs via recruiting DCAF16 for triple-negative breast cancer", EUROPEAN JOURNAL OF MEDICINAL CHEMISTRY, vol. 236, 3 April 2022 (2022-04-03), pages 1 - 19, XP087049662, DOI: 10.1016/j.ejmech.2022.114321 * |
| HUIFANG SHAN，等: "Discovery of a novel covalent CDK4/6 inhibitor based on palbociclib scaffold", EUROPEAN JOURNAL OF MEDICINAL CHEMISTRY, vol. 219, 3 April 2021 (2021-04-03), pages 1 - 20 * |
| ROBERT I. TROUP, 等: "Current strategies for the design of PROTAC linkers: a critical review", EXPLOR TARGET ANTITUMOR THER., vol. 5, no. 1, 31 December 2020 (2020-12-31), pages 273 * |
| 季鸣，等: "新型PARP1/2抑制剂YHP-743的抗肿瘤作用", 药学学报, no. 06, 2 May 2018 (2018-05-02), pages 112 - 117 * |
| 张文娟，等: "新型 PI3K 抑制剂邻苯二甲酰亚胺及其衍生物的设计合成与抗肿瘤活性", 中国药物化学杂志, vol. 23, no. 3, 30 June 2013 (2013-06-30), pages 181 - 186 * |
| 罗 超，等: "奥拉帕尼通过PARP-1通路调控脂多糖诱导的A549细胞炎症反应", 中国药理学与毒理学杂志, vol. 33, no. 3, 31 March 2019 (2019-03-31), pages 193 - 199 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103282365B (zh) | 结晶的(8S,9R)‑5‑氟‑8‑(4‑氟苯基)‑9‑(1‑甲基‑1H‑1,2,4‑三唑‑5‑基)‑8,9‑二氢‑2H‑吡啶并[4,3,2‑de]酞嗪‑3(7H)‑酮甲苯磺酸盐 | |
| NO335100B1 (no) | Isetionatsalt, fremgangsmåte for fremstilling av slike, farmasøytisk doseform omfattende slike samt slike forbindelser for behandling av sykdom | |
| JP6704422B2 (ja) | キナゾリン誘導体の塩およびその製造方法 | |
| WO2017045583A1 (zh) | 一种富集了nuc‐1031单一异构体的组合物及其制备方法和用途 | |
| CN114736214B (zh) | 一种倍半萜衍生物、其药物组合物及其制备方法和用途 | |
| JP2023538090A (ja) | Ｂｔｋ阻害剤としての架橋二環式化合物 | |
| JP6827942B2 (ja) | トリプトリドのｃ１４ヒドロキシルエステル化アミノ酸誘導体、ならびにその製造方法および使用 | |
| CN116390728A (zh) | 喹唑啉衍生物及其制备方法和用途 | |
| CN111518020A (zh) | Magl抑制剂及其制备方法、用途 | |
| CN106995449B (zh) | 鬼臼毒素-维甲酸杂合物合成方法和应用于预防、治疗肿瘤的药物 | |
| CN106317033B (zh) | 一种水飞蓟宾23-取代衍生物及其注射剂的制备方法和用途 | |
| EP2896623A1 (en) | CRYSTAL OF N-[2-({2-[(2S)-2-CYANOPYRROLIDIN-1-YL]-2- OXOETHYL}AMINO)-2-METHYLPROPYL]-2-METHYLPYRAZOLO[1,5-a]PYRIMIDINE-6-CARBOXAMIDE | |
| CN116102534A (zh) | 共价PARP PROTACs衍生物及其应用 | |
| CN101495485B (zh) | 具有抗肿瘤活性的喜树碱衍生物 | |
| CN107892691B (zh) | 2,8,9-三取代-9h-嘌呤类化合物及其盐和应用 | |
| TW201922690A (zh) | 環－amp反應元素結合蛋白的抑制劑 | |
| US9499552B2 (en) | Pyrazolo[1,5-A]pyrimidine derivative and use of anti-tumor thereof | |
| CN115433207A (zh) | 作为egfr抑制剂的大环杂环类化合物及其应用 | |
| CN107011312A (zh) | 片叶苔素d含氮衍生物及其制备方法和在治疗肿瘤疾病中的用途 | |
| CN109942665A (zh) | 雷公藤内酯醇衍生物及其制备方法和应用 | |
| CN110590778B (zh) | 3，10二对甲氧基苯基6，12二氮杂四高立方烷类化合物及合成方法和药物组合物 | |
| DK161833B (da) | Analogifremgangsmaade til fremstilling af n-(vinblastinoyl-23)-derivater af aminosyrer eller farmaceutisk acceptable syreadditionssalte deraf | |
| WO2023142754A1 (zh) | Ezh1/2抑制剂及其制备和抗肿瘤治疗中的应用 | |
| EP3825315B1 (en) | Crystal form as ask1 inhibitor, preparation method therefor, and application thereof | |
| JP7348214B2 (ja) | Ｈｄａｃ６選択的阻害剤の結晶形及びその使用 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |