CN109574871A - A kind of acetylamino azobenzene derivative and its preparation and application - Google Patents
A kind of acetylamino azobenzene derivative and its preparation and application Download PDFInfo
- Publication number
- CN109574871A CN109574871A CN201811403987.6A CN201811403987A CN109574871A CN 109574871 A CN109574871 A CN 109574871A CN 201811403987 A CN201811403987 A CN 201811403987A CN 109574871 A CN109574871 A CN 109574871A
- Authority
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- China
- Prior art keywords
- acetylamino
- reaction
- halogen
- azobenzene derivative
- substituent
- Prior art date
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Links
- -1 acetylamino azobenzene derivative Chemical class 0.000 title claims abstract description 24
- 238000002360 preparation method Methods 0.000 title claims abstract description 19
- 150000001875 compounds Chemical class 0.000 claims abstract description 25
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 11
- 239000003814 drug Substances 0.000 claims abstract description 6
- 206010069755 K-ras gene mutation Diseases 0.000 claims abstract description 5
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims abstract description 5
- 229940079593 drug Drugs 0.000 claims abstract description 5
- 201000005202 lung cancer Diseases 0.000 claims abstract description 5
- 208000020816 lung neoplasm Diseases 0.000 claims abstract description 5
- 201000001275 rectum cancer Diseases 0.000 claims abstract description 4
- 206010060862 Prostate cancer Diseases 0.000 claims abstract description 3
- 208000000236 Prostatic Neoplasms Diseases 0.000 claims abstract description 3
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 35
- 238000006243 chemical reaction Methods 0.000 claims description 30
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 24
- 229910052736 halogen Inorganic materials 0.000 claims description 23
- 150000002367 halogens Chemical class 0.000 claims description 23
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 22
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 18
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 18
- 239000000243 solution Substances 0.000 claims description 17
- 125000001424 substituent group Chemical group 0.000 claims description 16
- 238000003756 stirring Methods 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- 229960000583 acetic acid Drugs 0.000 claims description 7
- 239000012362 glacial acetic acid Substances 0.000 claims description 7
- 235000011121 sodium hydroxide Nutrition 0.000 claims description 7
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 claims description 6
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical class [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 claims description 6
- 230000035484 reaction time Effects 0.000 claims description 6
- 238000010992 reflux Methods 0.000 claims description 6
- 229910000342 sodium bisulfate Inorganic materials 0.000 claims description 6
- 125000000217 alkyl group Chemical group 0.000 claims description 5
- 238000001914 filtration Methods 0.000 claims description 5
- 230000001376 precipitating effect Effects 0.000 claims description 5
- 125000003342 alkenyl group Chemical group 0.000 claims description 4
- 125000003545 alkoxy group Chemical group 0.000 claims description 4
- 229910052799 carbon Inorganic materials 0.000 claims description 4
- 125000004432 carbon atom Chemical group C* 0.000 claims description 4
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 4
- 125000000449 nitro group Chemical group [O-][N+](*)=O 0.000 claims description 4
- 239000007864 aqueous solution Substances 0.000 claims description 3
- 125000001841 imino group Chemical group [H]N=* 0.000 claims description 3
- 239000000126 substance Substances 0.000 claims description 3
- 238000001035 drying Methods 0.000 claims description 2
- 230000000155 isotopic effect Effects 0.000 claims description 2
- 239000000203 mixture Substances 0.000 claims description 2
- 238000012986 modification Methods 0.000 claims description 2
- 230000004048 modification Effects 0.000 claims description 2
- 150000003839 salts Chemical class 0.000 claims description 2
- 238000010898 silica gel chromatography Methods 0.000 claims description 2
- 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 claims 1
- 125000002252 acyl group Chemical group 0.000 claims 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-M hydroxide Chemical compound [OH-] XLYOFNOQVPJJNP-UHFFFAOYSA-M 0.000 claims 1
- 230000003287 optical effect Effects 0.000 claims 1
- 229910052708 sodium Inorganic materials 0.000 claims 1
- 239000011734 sodium Substances 0.000 claims 1
- 101710113436 GTPase KRas Proteins 0.000 abstract description 17
- 230000002401 inhibitory effect Effects 0.000 abstract description 13
- 108090000623 proteins and genes Proteins 0.000 abstract description 10
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- 230000000694 effects Effects 0.000 abstract description 8
- 230000008859 change Effects 0.000 abstract description 6
- 238000005286 illumination Methods 0.000 abstract description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 15
- XKMLYUALXHKNFT-UUOKFMHZSA-N Guanosine-5'-triphosphate Chemical compound C1=2NC(N)=NC(=O)C=2N=CN1[C@@H]1O[C@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)[C@@H](O)[C@H]1O XKMLYUALXHKNFT-UUOKFMHZSA-N 0.000 description 13
- 239000007787 solid Substances 0.000 description 13
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 12
- 239000000543 intermediate Substances 0.000 description 11
- 238000005160 1H NMR spectroscopy Methods 0.000 description 10
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical class [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 10
- 102000016914 ras Proteins Human genes 0.000 description 10
- 108010014186 ras Proteins Proteins 0.000 description 9
- QGWNDRXFNXRZMB-UUOKFMHZSA-N GDP Chemical compound C1=2NC(N)=NC(=O)C=2N=CN1[C@@H]1O[C@H](COP(O)(=O)OP(O)(O)=O)[C@@H](O)[C@H]1O QGWNDRXFNXRZMB-UUOKFMHZSA-N 0.000 description 8
- 150000001412 amines Chemical class 0.000 description 8
- 210000004027 cell Anatomy 0.000 description 8
- 239000000460 chlorine Substances 0.000 description 8
- QGWNDRXFNXRZMB-UHFFFAOYSA-N guanidine diphosphate Natural products C1=2NC(N)=NC(=O)C=2N=CN1C1OC(COP(O)(=O)OP(O)(O)=O)C(O)C1O QGWNDRXFNXRZMB-UHFFFAOYSA-N 0.000 description 8
- METKIMKYRPQLGS-UHFFFAOYSA-N atenolol Chemical compound CC(C)NCC(O)COC1=CC=C(CC(N)=O)C=C1 METKIMKYRPQLGS-UHFFFAOYSA-N 0.000 description 7
- 239000012460 protein solution Substances 0.000 description 7
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 6
- 238000011534 incubation Methods 0.000 description 6
- 230000035772 mutation Effects 0.000 description 6
- 239000002773 nucleotide Substances 0.000 description 6
- 125000003729 nucleotide group Chemical group 0.000 description 6
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 description 6
- 235000018102 proteins Nutrition 0.000 description 6
- 238000012360 testing method Methods 0.000 description 6
- 210000004881 tumor cell Anatomy 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 5
- 235000002639 sodium chloride Nutrition 0.000 description 5
- ZPQOPVIELGIULI-UHFFFAOYSA-N 1,3-dichlorobenzene Chemical compound ClC1=CC=CC(Cl)=C1 ZPQOPVIELGIULI-UHFFFAOYSA-N 0.000 description 4
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 4
- PAYRUJLWNCNPSJ-UHFFFAOYSA-N Aniline Chemical compound NC1=CC=CC=C1 PAYRUJLWNCNPSJ-UHFFFAOYSA-N 0.000 description 4
- VGCXGMAHQTYDJK-UHFFFAOYSA-N Chloroacetyl chloride Chemical compound ClCC(Cl)=O VGCXGMAHQTYDJK-UHFFFAOYSA-N 0.000 description 4
- IAZDPXIOMUYVGZ-WFGJKAKNSA-N Dimethyl sulfoxide Chemical compound [2H]C([2H])([2H])S(=O)C([2H])([2H])[2H] IAZDPXIOMUYVGZ-WFGJKAKNSA-N 0.000 description 4
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 4
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 description 4
- VYFYYESMHPKEAD-FMTVUPSXSA-N [(2r,3s,5r)-5-(2-amino-6-oxo-3h-purin-9-yl)-3-hydroxy-3-[2-(methylamino)benzoyl]oxolan-2-yl]methyl phosphono hydrogen phosphate Chemical compound CNC1=CC=CC=C1C(=O)[C@@]1(O)[C@@H](COP(O)(=O)OP(O)(O)=O)O[C@@H](N2C3=C(C(N=C(N)N3)=O)N=C2)C1 VYFYYESMHPKEAD-FMTVUPSXSA-N 0.000 description 4
- 201000011510 cancer Diseases 0.000 description 4
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- 230000005764 inhibitory process Effects 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 239000002994 raw material Substances 0.000 description 4
- 239000011780 sodium chloride Substances 0.000 description 4
- 238000010189 synthetic method Methods 0.000 description 4
- DLFVBJFMPXGRIB-UHFFFAOYSA-N Acetamide Chemical compound CC(N)=O DLFVBJFMPXGRIB-UHFFFAOYSA-N 0.000 description 3
- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 3
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 3
- 101150040459 RAS gene Proteins 0.000 description 3
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 3
- 230000003281 allosteric effect Effects 0.000 description 3
- 229910052801 chlorine Inorganic materials 0.000 description 3
- 239000003112 inhibitor Substances 0.000 description 3
- 239000012074 organic phase Substances 0.000 description 3
- 229910000027 potassium carbonate Inorganic materials 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- HIXDQWDOVZUNNA-UHFFFAOYSA-N 2-(3,4-dimethoxyphenyl)-5-hydroxy-7-methoxychromen-4-one Chemical compound C=1C(OC)=CC(O)=C(C(C=2)=O)C=1OC=2C1=CC=C(OC)C(OC)=C1 HIXDQWDOVZUNNA-UHFFFAOYSA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 101001065501 Escherichia phage MS2 Lysis protein Proteins 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 239000007995 HEPES buffer Substances 0.000 description 2
- 230000027455 binding Effects 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 230000004663 cell proliferation Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 2
- 235000018417 cysteine Nutrition 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 235000019441 ethanol Nutrition 0.000 description 2
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 2
- 230000005284 excitation Effects 0.000 description 2
- 239000006260 foam Substances 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 239000003292 glue Substances 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 229910001629 magnesium chloride Inorganic materials 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 229910000474 mercury oxide Inorganic materials 0.000 description 2
- UKWHYYKOEPRTIC-UHFFFAOYSA-N mercury(ii) oxide Chemical compound [Hg]=O UKWHYYKOEPRTIC-UHFFFAOYSA-N 0.000 description 2
- CHMBIJAOCISYEW-UHFFFAOYSA-N n-(4-aminophenyl)acetamide Chemical compound CC(=O)NC1=CC=C(N)C=C1 CHMBIJAOCISYEW-UHFFFAOYSA-N 0.000 description 2
- 150000002832 nitroso derivatives Chemical class 0.000 description 2
- 239000003208 petroleum Substances 0.000 description 2
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 108700042226 ras Genes Proteins 0.000 description 2
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- 150000003536 tetrazoles Chemical class 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- AYGXDXVGROLVJK-UHFFFAOYSA-N 1-chloro-2-nitrosobenzene Chemical compound ClC1=CC=CC=C1N=O AYGXDXVGROLVJK-UHFFFAOYSA-N 0.000 description 1
- GUMCAKKKNKYFEB-UHFFFAOYSA-N 2,4,5-trichloroaniline Chemical class NC1=CC(Cl)=C(Cl)C=C1Cl GUMCAKKKNKYFEB-UHFFFAOYSA-N 0.000 description 1
- KQCMTOWTPBNWDB-UHFFFAOYSA-N 2,4-dichloroaniline Chemical compound NC1=CC=C(Cl)C=C1Cl KQCMTOWTPBNWDB-UHFFFAOYSA-N 0.000 description 1
- SDYWXFYBZPNOFX-UHFFFAOYSA-N 3,4-dichloroaniline Chemical compound NC1=CC=C(Cl)C(Cl)=C1 SDYWXFYBZPNOFX-UHFFFAOYSA-N 0.000 description 1
- IZUUGCNHLFFAGM-UHFFFAOYSA-N 4-chloro-3-cyclopropylaniline Chemical compound NC1=CC=C(Cl)C(C2CC2)=C1 IZUUGCNHLFFAGM-UHFFFAOYSA-N 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 238000009010 Bradford assay Methods 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- 208000001333 Colorectal Neoplasms Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 206010064571 Gene mutation Diseases 0.000 description 1
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- 101001024703 Homo sapiens Nck-associated protein 5 Proteins 0.000 description 1
- NGESVDVCRSHORY-UHFFFAOYSA-N N-bromo-4-chloroaniline Chemical compound ClC1=CC=C(NBr)C=C1 NGESVDVCRSHORY-UHFFFAOYSA-N 0.000 description 1
- 102100036946 Nck-associated protein 5 Human genes 0.000 description 1
- 206010061902 Pancreatic neoplasm Diseases 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 125000000738 acetamido group Chemical group [H]C([H])([H])C(=O)N([H])[*] 0.000 description 1
- 125000002777 acetyl group Chemical group [H]C([H])([H])C(*)=O 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 125000003647 acryloyl group Chemical group O=C([*])C([H])=C([H])[H] 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 150000001335 aliphatic alkanes Chemical class 0.000 description 1
- 230000008856 allosteric binding Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 150000001555 benzenes Chemical class 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
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- 238000004113 cell culture Methods 0.000 description 1
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- 238000004440 column chromatography Methods 0.000 description 1
- 229940126214 compound 3 Drugs 0.000 description 1
- NKLPQNGYXWVELD-UHFFFAOYSA-M coomassie brilliant blue Chemical compound [Na+].C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=C1 NKLPQNGYXWVELD-UHFFFAOYSA-M 0.000 description 1
- 125000001559 cyclopropyl group Chemical group [H]C1([H])C([H])([H])C1([H])* 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 229960000935 dehydrated alcohol Drugs 0.000 description 1
- 238000010612 desalination reaction Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 238000007876 drug discovery Methods 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 229960004756 ethanol Drugs 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 230000005496 eutectics Effects 0.000 description 1
- 239000012894 fetal calf serum Substances 0.000 description 1
- 239000012065 filter cake Substances 0.000 description 1
- 235000013922 glutamic acid Nutrition 0.000 description 1
- 239000004220 glutamic acid Substances 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
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- 239000011159 matrix material Substances 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- MDUFZRDLZSSHAU-UHFFFAOYSA-N n-(4-nitrosophenyl)acetamide Chemical compound CC(=O)NC1=CC=C(N=O)C=C1 MDUFZRDLZSSHAU-UHFFFAOYSA-N 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 208000002154 non-small cell lung carcinoma Diseases 0.000 description 1
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- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C245/00—Compounds containing chains of at least two nitrogen atoms with at least one nitrogen-to-nitrogen multiple bond
- C07C245/02—Azo compounds, i.e. compounds having the free valencies of —N=N— groups attached to different atoms, e.g. diazohydroxides
- C07C245/06—Azo compounds, i.e. compounds having the free valencies of —N=N— groups attached to different atoms, e.g. diazohydroxides with nitrogen atoms of azo groups bound to carbon atoms of six-membered aromatic rings
- C07C245/08—Azo compounds, i.e. compounds having the free valencies of —N=N— groups attached to different atoms, e.g. diazohydroxides with nitrogen atoms of azo groups bound to carbon atoms of six-membered aromatic rings with the two nitrogen atoms of azo groups bound to carbon atoms of six-membered aromatic rings, e.g. azobenzene
-
- 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
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C2601/00—Systems containing only non-condensed rings
- C07C2601/02—Systems containing only non-condensed rings with a three-membered ring
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmacology & Pharmacy (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The present invention relates to a kind of acetylamino azobenzene derivative and its preparation and application, which has general formulaWithIn structure.Acetylamino azobenzene derivative according to the present invention has the characteristic that structure change and activity change are generated under illumination condition, and the structure in structural formula I is presented in a dark environment, and the structure in formula II is presented after ultraviolet lighting；Structure I I has K-Ras (G12C) protein inhibiting activity, and structure I activity is weaker.Such compound can be used for treating the relevant tumour of K-Ras gene mutation, such as carcinoma of the rectum, the selective drug of lung cancer and prostate cancer etc..
Description
Technical field
The present invention relates to field of pharmaceutical chemistry technology, and in particular to a kind of acetylamino azobenzene derivative and its system
It is standby with application.
Background technique
The mutation of Ras gene is related with many cancers and the relevant Ras gene of cancer is essentially all single base mutation,
99% mutation all occur 12,13 glycine and 61 glutamic acid (Nature reviews cancer, 2011,
11,761-774).Being distributed in for Ras mutation is not consistent in cancer, and K-Ras is the highest gene of the frequency of mutation, is reached
The mutation of 86%, K-Ras are most commonly in colorectal cancer, lung cancer (mainly non-small cell lung cancer) and cancer of pancreas.
K-Ras albumen has always been considered as being " can not patent medicine " for many years.New technology in drug discovery, which promotes, to be directed to
The new treatment (McCormick, 2015) of RAS.Currently, for Ras albumen inhibitor research be concentrated mainly on it is following several
A aspect: the formation of Ras-GTP is prevented, the effect of inhibition Ras- effect protein, the positioning of Ras is reduced and inhibits GTP enzyme
Activity.
Ras protein inhibitor research difficult point be, Ras and GDP (guanosine diphosphate), GTP
Very strong (reaching picomole rank) (Nature reviews of compatibility between (guanosine triphosphate)
Molecular cell biology, 2012,13,39-51.), therefore directly act on the competitiveness of GTP/GDP binding pocket
Inhibitor is difficult to weaken the combination of Ras albumen and GTP.Shokat seminar reports a kind of band electrophilic group (such as ethenesulfonyl
Base, acryloyl group) inhibitor, the allosteric binding pocket that eutectic is not found before one as the result is shown can lead to Ras
The change of middle Switch I and Switch II structure, can weaken K-ras (G12C) and GTP combination (Nature, 2013,
503(7477):548).Janes also reports a kind of compound (Cell, 2018,172,578-589) on the basis of them,
Have the function of preferably inhibiting K-Ras albumen, and also obtains relatively good result in mouse experiment in vivo.These suppressions
Although preparation has the selectivity for saltant type K-Ras (G12C) albumen without being directed to wild type K-Ras albumen, but not
Has the feature for inhibiting situation to be controlled in K-Ras albumen at different conditions.
Summary of the invention
It is an object of the present invention to overcome the above-mentioned drawbacks of the prior art and provide a kind of acetylamino azos
Benzene analog derivative and its preparation and application.
The purpose of the present invention can be achieved through the following technical solutions: a kind of acetylamino azobenzene derivative, should
The structural formula of derivative is as shown in Formulas I or Formula II:
Above structure has hydrophobic benzene ring structure in conjunction with albumen allosteric pocket, has electrophilic group prominent with Ras
12 white cysteine covalent bonds of a kink of preserved egg simultaneously inhibit protein activation, and can further pass through illumination after albumen covalent bond
Inhibitory activity is controlled, realizes the selectivity for specific organization or region.
Preferably, the substituent R in structural formula on A ring1In H, halogen, hydroxyl, nitro, C1~C10 or alkoxy
One kind；Substituent R on A ring2Selected from one of H or halogen；Substituent R on A ring3Selected from one of H or halogen；
Substituent R on A ring4Selected from one of H, halogen or C1~C10；The n is the integer in 1~3, and the X is selected from halogen
Element, alkenyl or dimaleoyl imino；C1~the C10 is alkyl of the carbon atom number 1~10, including straight chained alkyl, branched alkane
Base or cyclic alkyl, such as methyl, ethyl, isopropyl, cyclopropyl.
It is furthermore preferred that R1For halogen, R2For H, R3For halogen, R4For H, n 1, X are halogen.
Most preferably, R1For chlorine, R2For H, R3For chlorine, R4For H, n 1, X are chlorine.
Preferably, the derivative includes that the isotopic compound, racemic modification, optically-active of substance shown in Formulas I or Formula II are living
One of property isomers, polymorphic, pharmaceutically acceptable salt or mixture.
A kind of preparation method of acetylamino azobenzene derivative as described above, which is characterized in that including following step
It is rapid:
(1) by ammonium persulfate-sodium bisulfate respectively withMixing, is stirred
It mixes after completion of the reaction, obtains
(2) step (1) is obtainedIt is dissolved in glacial acetic acid, is addedStirring
Terminate to reaction, is obtained by filtration
Alternatively, step (1) is obtainedIt is dissolved in glacial acetic acid, is addedIt stirs
Mixing to reaction terminates, and is obtained by filtration
(3) willIt is dissolved in methanol, hydrochloric acid is added, is heated to reflux, reaction terminates
PH is adjusted with 3M sodium hydrate aqueous solution afterwards, saturated sodium bicarbonate solution then is added dropwise until generating precipitating, it is separation, dry
It arrives
(4) willIt is dissolved in methylene chloride, is then addedIt stirs to anti-
It should terminate, column chromatographic purifying is up to described
Preferably, the substituent R on A ring occurred in reaction step1Selected from H, halogen, hydroxyl, nitro, C1~C10 or
One of alkoxy；Substituent R on A ring2Selected from one of H or halogen；Substituent R on A ring3Selected from H or halogen
One of；Substituent R on A ring4Selected from one of H, halogen or C1~C10；The n is the integer in 1~3, described
X is selected from halogen, alkenyl or dimaleoyl imino；C1~the C10 is alkyl of the carbon atom number 1~10.
Preferably, ammonium persulfate-sodium bisulfate described in step (1) with Molar ratio be 1:(1.2~2), the rate of stirring is 500~1000rpm, and the temperature of reaction is
20~30 DEG C, the reaction time is 2~12 hours.
Preferably, in step (2),WithMolar ratio be (1.5~3): 1,
It is describedWithMolar ratio be (1.5~3): 1, the rate of the stirring is 500
~1000rpm, the temperature of reaction are 20~30 DEG C, and the reaction time is 12~15 hours.
Preferably, in step (3), the methanol of addition and the volume ratio of hydrochloric acid are 1:(0.5~2),Molar ratio with HCl in hydrochloric acid is 1:(50~100), the temperature of back flow reaction is
80~90 DEG C, the time is 10~12 hours, and reaction system pH is adjusted to 8~9 by 3M sodium hydrate aqueous solution, the drying
Temperature is 55~65 DEG C.
Preferably, in step (4),Molar ratio be 1:(2~3),
The temperature of reaction is 20~30 DEG C, and the reaction time is 2~12 hours, and purifying uses silica gel column chromatography.
A kind of application of acetylamino azobenzene derivative as described above, the derivative are used to prepare treatment K-Ras
The drug of the relevant tumour of gene mutation, the relevant tumour of the K-Ras gene mutation includes the carcinoma of the rectum, lung cancer or prostate
One of cancer.
Compared with prior art, the beneficial effects of the present invention are embodied in following several respects:
(1) derivative has the characteristic that structure change and activity change are generated under illumination condition, is in a dark environment
The structure in formula II is presented in structure in existing structural formula I after ultraviolet lighting；Structure I I has K-Ras (G12C) albumen
Inhibitory activity, structure I activity are weaker；
(2) it can be used for treating the relevant tumour of K-Ras gene mutation, such as carcinoma of the rectum, lung cancer and prostate cancer etc. have
The drug of selectivity.
(3) preparation method is easy, can be easily separated purifying.
Detailed description of the invention
Fig. 1 is that the cis-trans structural derivative that embodiment 2 is prepared exchanges the nucleotide of K-Ras (G12C) albumen
Experimental result；
Fig. 2 is the transconfiguration derivative that is prepared of embodiment 1 to the active experimental result of inhibiting tumour cells；
Fig. 3 is the cis-structure derivative that is prepared of embodiment 1 to the active experimental result of inhibiting tumour cells；
Fig. 4 is the cis-structure derivative that is prepared of embodiment 2 to the active experimental result of inhibiting tumour cells；
Fig. 5 is the transconfiguration derivative that is prepared of embodiment 2 to the active experimental result of inhibiting tumour cells；
Specific embodiment
Elaborate below to the embodiment of the present invention, the present embodiment under the premise of the technical scheme of the present invention into
Row is implemented, and the detailed implementation method and specific operation process are given, but protection scope of the present invention is not limited to following realities
Apply example.
Embodiment 1
(E) preparation flow of the chloro- N- of -2- (4- ((4- chlorphenyl) azo) phenyl) acetamide is as follows:
Reaction reagent and condition: (a) K2CO3,H2O,10min；(b)AcOH,DCM,r.t.overnight(c)1) HCl,
MeOH,reflux,9h；2)NaOH；(d)chloroacetyl chloride,dry DCM,DIPEA,r.t., overnight.
Specifically, comprising the following steps:
(1), intermediate 1 [N- (4- nitroso phenyl) acetamide]
Ammonium persulfate-sodium bisulfate is dissolved in 50ml water, addition potassium carbonate adjusting pH is weakly acidic, then adds rapidly
Enter in the 4- amino acetanilide having been dissolved in water, solution generates rapidly celadon foam and celadon precipitating, to ash
Green foam disappears, and after precipitating sedimentation, reaction continues to stop for 10 minutes again.Filtering obtains celadon precipitating, while hot with hot second
Alcohol, which dissolves and is filtered to remove impurity, obtains the solution of green, is spin-dried for ethyl alcohol, obtains target product intermediate 1 (green solid).Directly
Investment is connect to react in next step.
(2), intermediate 2 [(E)-N- (4- ((4- chlorphenyl) azo) phenyl) acetamide]
In glacial acetic acid, parachloroanilinum is added in the nitroso compound dissolution that back is obtained in system.At room temperature
It is stirred overnight.Reaction terminates isolated cake solids, filters and wash solid, in methylene chloride by solid dissolution, has
Machine mutually uses saturated sodium bicarbonate solution and saturated common salt water washing, dry with anhydrous sodium sulfate, is concentrated to get 2 (yellow of intermediate
Solid, 1.2428g, 50.02%).1H NMR(400MHz,CDCl3) δ 2.229 (s, 3H), 7.473 (d, J=2.2Hz, 2H),
7.677 (d, J=2.1Hz, 2H), 7.845 (d, J=2.2Hz, 2H), 7.91 (d, J=2.2Hz, 2H) ppm.
(3), intermediate 3 [(E) -4- ((4- chlorphenyl) azo) aniline]
Reactant 2 and 3M hydrochloric acid are added in methyl alcohol, is heated to reflux 9 hours.It is molten that 3M sodium hydroxide is added dropwise after reaction
Liquid adjusts pH=8-9, is then slowly added into saturated sodium bicarbonate solution until generating yellow mercury oxide.Isolated yellow is heavy
Shallow lake is dried to obtain intermediate 3 (yellow solid, 134.3mg, 52.9%) at 60 DEG C or less1H NMR(400MHz,CDCl3)δ
6.741 (d, J=2.2Hz, 2H), 7.441 (d, J=2.2Hz, 2H), 7.795 (dd, J1=2.2Hz, J2=2.2Hz, 4H)
ppm。
(4), compound 4 [the chloro- N- of (E) -2- (4- ((4- chlorphenyl) azo) phenyl) acetamide]
The compound 3 that back reacts is dissolved in the methylene chloride steamed again, then chloracetyl chloride is added anti-
It answers in system, is stirred overnight at room temperature.After reaction, with methylene chloride: being eluted under the conditions of petroleum ether=2:1, column chromatography point
From obtaining target compound 4 (yellow solid, 150.06mg, 44.92%).1H NMR(400 MHz,CDCl3) δ 4.238 (s,
2H), 7.483 (d, J=2.2Hz, 2H), 7.739 (d, J=2.2Hz, 1H), 7.860 (d, J=2.2Hz, 2H), 7.948 (d, J
=2.2Hz, 2H) ppm.[M-H]–C14H10Cl2N3O, target compound Exact mass calculated value: 306.0201, observed value:
306.0204。
Embodiment 2
(E) preparation flow of the chloro- N- of -2- (4- ((2,4 dichloro benzene base) azo) phenyl) acetamide is as follows:
Reaction reagent and condition: (a) K2CO3,H2O,30min；(b)AcOH,DCM,r.t.overnight(c)1) HCl,
MeOH,reflux,9h；2)NaOH；(d)chloroacetyl chloride,dry DCM,DIPEA,r.t., overnight.
Specifically, comprising the following steps:
(1), intermediate 5 [the chloro- 1- nitrosobenzene of 2,4- bis-]
2,4- dichloroaniline is dissolved in methylene chloride, ammonium persulfate-sodium bisulfate is dissolved in the water, in nitrogen
It is stirred overnight at room temperature after the lower mixing of protection.After reaction, organic phase is washed with 1M hydrochloric acid (10mL) and saturated brine.Finally,
With the dry organic phase of anhydrous sodium sulfate, enriched product intermediate 5 is directly placed into and reacts in next step.
(2), intermediate 6 [(E)-N- (4- ((2,4 dichloro benzene base) azo) phenyl) acetamide]
In glacial acetic acid, 4- amino acetanilide is added in the nitroso compound dissolution that back is obtained in system.
It is stirred overnight at room temperature.After reaction, the separation of filter cake shape solid filters and washs solid.In methylene chloride by solid dissolution,
With saturated sodium bicarbonate solution and saturated common salt water washing, organic phase is dry in anhydrous sodium sulfate, is concentrated to get among product
Body 6 (yellow solid, 2.6532g, 43.2%).1H NMR(400MHz,CDCl3) δ 2.238 (s, 3H), 7.308 (d, J=
0.6Hz, 1H), 7.569 (d, J=0.6Hz, 1H), 7.677 (d, J=2.1 Hz, 2H), 7.679 (s, 1H), 7.962 (d, J=
2.2Hz, 2H) ppm.
(3), intermediate 7 [(E) -4- ((2,4 dichloro benzene base) azo) aniline]
Reactant 6 and 3M hydrochloric acid are added in methyl alcohol, is heated to reflux 9h.3M sodium hydroxide solution is added dropwise after reaction
Reaction solution pH to 8-9 is adjusted, saturated sodium bicarbonate solution then is added until generating yellow mercury oxide.Isolated yellow is heavy
It forms sediment in 60 DEG C or less dry intermediates 7 (yellow solid, 112.3mg, 64.92%).1H NMR(400MHz,CDCl3)δ
6.738 (d, J=1.9Hz, 2H), 7.269 (s, 1H), 7.531 (d, J=2.2Hz, 1H), 7.641 (d, J=2.2Hz, 1H),
7.845 (d, J=2.1Hz, 2H) ppm.
(4), compound 8 [the chloro- N- of (E) -2- (4- ((2,4 dichloro benzene base) azo) phenyl) acetamide]
The compound that back is reacted is dissolved in the methylene chloride steamed again, is then added dropwise into reaction system
Chloracetyl chloride is stirred overnight at room temperature.Silica gel chromatograph column purification is used after reaction, in methylene chloride: petroleum ether=2:1 item
It is eluted under part, obtains target compound 8 (yellow solid, 152.4mg, 57.5%).1H NMR(400 MHz,DMSO-d6)δ
4.316(s,2H),7.562(dd,J1=2.2Hz, J2=2.2Hz, 1H), 7.683 (d, J=2.1Hz, 1H), 7.841 (d, J=
2.1Hz, 2H), 7.895 (d, J=0.6Hz, 1H), 7.941 (d, J=2.1Hz, 2H), 10.719 (s, 1H) ppm；HRMS
(ESI):[M-H]–C14H10Cl3N3O, target compound Exact mass calculated value: 340.9889, observed value: 340.9886.
Embodiment 3
(E) the chloro- N- of -2- (4- ((3,4- dichloro) azo) phenyl) acetamide
(9) preparation
Using synthetic method similar to Example 2, the difference is that:
(a), raw material amine described in step 1, using 3,4-DCA
Final detection result is as follows: 1H NMR (400MHz, DMSO-d6): δ 10.71 (s, 1H), 8.02 (s, 1H), 7.92
(d, J=7.6Hz, 2H), 7.85 (s, 1H), 7.84 (s, 1H), 7.82 (d, J=9.2Hz, 2H), 4.32 (s, 2H) ppm.HRMS
(ESI):[M+H]–C14H11Cl3N3O, target compound Exact mass calculated value: 341.9959, observed value: 341.9967.
Embodiment 4
(E) the chloro- N- of -2- (4- ((2,4,5- trichlorophenyl) azo) phenyl) acetyl
The preparation of amine (10)
Using synthetic method similar to Example 2, the difference is that:
(a), raw material amine described in step 1, using 2,4,5- trichloroanilines
Final detection result is as follows: 1H NMR (600MHz, DMSO) δ 10.75 (s, 1H), 8.11 (s, 1H), 7.95 (d, J
=8.8Hz, 2H), 7.85 (d, J=8.8Hz, 2H), 7.80 (s, 1H), 4.33 (s, 2H) ppm. HRMS (ESI): [M+H]–
C14H10Cl4N3O, target compound Exact mass calculated value: 375.9578, observed value: 375.9568.
Embodiment 5
(E) the chloro- N- of -2- (4- ((the bromo- 4- chlorphenyl of 3-) azo) phenyl) acetyl
The preparation of amine (11)
Using synthetic method similar to Example 2, the difference is that:
(a), raw material amine described in step 1, using the bromo- 4- chloroaniline of 3-
Final detection result is as follows: 1H NMR (600MHz, DMSO) δ 10.72 (s, 1H), 8.17 (s, 1H), 7.95 (d, J
=7.6Hz, 2H), 7.91 (d, J=8.5Hz, 1H), 7.88-7.86 (m, 1H), 7.85 (d, J=8.4 Hz, 2H), 4.33 (s,
2H)ppm.HRMS(ESI):[M+H]–C14H11BrCl2N3O, target compound Exact mass calculated value: 385.9462, observation
Value: 385.9456.
Embodiment 6
(E) the chloro- N- of -2- (4- ((the chloro- 3- cyclopropyl phenyl of 4-) azo) benzene
Base) acetamide (12) preparation
Using synthetic method similar to Example 2, the difference is that:
(a), raw material amine described in step 1, using 3- cyclopropyl -4- chloroaniline
Final detection result is as follows: 1H NMR (600MHz, CDCl3) δ 8.42 (s, 1H), 7.96 (dd, J=11.7,
5.3Hz, 2H), 7.76 (d, J=8.7Hz, 2H), 7.67 (dt, J=14.2,7.1Hz, 1H), 7.53 (d, J=1.7Hz, 1H),
7.50 (d, J=8.4Hz, 1H), 2.37-2.22 (m, 1H), 1.16-1.04 (m, 2H), 0.90-0.79 (m, 2H) ppm.HRMS
(ESI):[M+H]–C17H16Cl2N3O, target compound Exact mass calculated value: 348.0670, observed value: 348.0663.
Three, biological activity test
(1) compound and K-Ras (G12C) albumen are incubated for
For embodiment 1 and embodiment 2, K-Ras (FL, G12C) albumen is made into incubation buffer to 50 μM of 150 μ L
Protein solution is separately added into the DMSO solution of the compound of cis-structure and transconfiguration to final concentration of 500 μM thereto
(amount of DMSO is 2% (v/v)), 4 DEG C are incubated for for 24 hours.Take 80 μ L protein solutions in 1.5mL EP pipe, be added 20 μ L 5 ×
SDS.90 DEG C of heating 5min run SDS-PAGE glue.With coomassie brilliant blue staining rear decoloring (destainer: 30% dehydrated alcohol,
10% glacial acetic acid, 60% distilled water).Protein band is found according to albumen Maker, cuts glue, Trypsin enzymatic lysis, Zip- desalination
Sample introduction does protein spectrum experiment afterwards.
(2) nucleotide exchange test
(a) compound and protein binding
Albumen is added in buffer solution (20mm HEPEs, pH 7.5,150mM NaCl, 1mM EDTA), until most
Final concentration of 20 μM, compound storing liquid (20mM) is placed on wavelength and is added to irradiate after 4min to be protected from light under the ultraviolet lamp of 400nm
Enter system to final concentration of 400 μM, final volume is 300 μ L, and 4 DEG C are incubated for for 24 hours.
(b) combination of albumen and mant-dGDP
After the completion of compound incubation, edta buffer solution (the 20mM HEPEs, pH of 40 μ L are added into protein solution
7.5,150mM NaCl, 15mM EDTA, 1mM DTT), make final concentration of 2.5 mM of EDTA；It is added simultaneously into protein solution
The mant-dGDP (initial concentration 10mM) of 14.4 μ L, mant-dGDP concentration is 400 μM at this time, is incubated at room temperature 1h.Then it is added
2μL MgCl2, final concentration of 10mM, and in 4 DEG C of incubation 1h, the mant-dGDP being not bound with finally is removed with NAP-5 pillar,
Bradford method surveys protein concentration.
(c) it titrates
With albumen reaction buffer (20mM HEPES, pH7.5,150mM NaCl, 1mM MgCl2, 1mM DTT) and by albumen
It is diluted to 1.5 μM.Into black matrix plate, 10 μ L of protein solution is added in every hole, totally 16 parallel samples, each parallel sample concentration
All set two multiple holes.Being added dropwise to complete rear slight shaking makes the protein solution inflow hole bottom for hanging over hole wall, microplate reader (excitation light wave
It is long: 360nm；Wavelength of transmitted light: 440nm) protein fluorescence value in gaging hole.After having surveyed, added with the volley of rifle fire into protein solution sample
Enter 5 μ L various concentrations GDP or GTP solution (be dissolved in edta buffer liquid: 20mM HEPES, pH 7.5,150mM NaCl,
15mM EDTA, 1mM DTT).GDP/GTP concentration be respectively as follows: 3000 μM, 1200 μM, 480.0 μM, 192.0 μM, 76.80 μM,
30.72μM、12.29μM、4.915μM、1.966μM、0.786μM、0.314μM、0.126 μM、0.050μM、0.020μM、
0.008μM,0.003μM.It shakes and mixes after should sufficiently melting when use.
Experiment divides ultraviolet irradiation group and non-irradiated group, and ultraviolet irradiation group first uses 4 min of 400nm length ultraviolet light irradiation, then
Plus nucleotide (GDP and GTP solution), the distance between 384 orifice plates and light source are 5cm or so, and room temperature is protected from light incubation on shaking table
2h.Non-irradiated group is not required to illumination, and room temperature, which is protected from light, directly after nucleotide is added dropwise is incubated for 2h.After incubation, measured with microplate reader
Fluorescence intensity (excitation wavelength: 360nm；Launch wavelength: 440 nm).Compare the fluorescent value being added before and after GDP or GTP solution, and
IC is calculated with 5 software of GraphPad Prism50Value.The fluorescent value calculation formula before and after GDP or GTP solution is added:
Fn=FluAfter n/FluBefore n, Rn=(Fn-Fmin)/(Fmax-Fmin)
(FluAfter n、FluBefore nIt is the average value of two multiple holes fluorescent values, gained FnAs normalized numerical value.According to Rn
To concentration (logCn) S type curve is done, calculate nucleotide IC50).
Nucleotide exchange test result is as shown in Figure 1, wherein after the compound cis-trans that embodiment 2 is prepared is incubated for
8-trans represents the experimental result of transconfiguration, and 8-cis becomes cis- after representing transconfiguration 400nm ultraviolet lighting completely
Experimental result after structure.From the figure we can see that after light-operated molecular switch compound 8 is incubated for K-Ras (G12C),
Part trans-compound 8 and K-Ras mutain 12 cysteine covalent bonds enter allosteric pocket.In ultraviolet lighting
Afterwards, trans-compound 8 and 8 allosteric of cis-compound, protein structure also change therewith.GTP IC50 is shown as in experiment
Value increases, that is, weakens the affinity of K-Ras (G12C) and GTP.
(3) cell experiment
1, cell inhibitory effect is tested
(1) cell culture.Tumour cell ties up to DMEM complete medium, and (10% tire ox blood is added in DMEM in high glucose culture medium
Clearly, 100units/mL penicillin, 100mg/mL streptomysin) in culture.H358 is in (the DMEM in high glucose culture of DMEM complete medium
Base, be added 10% fetal calf serum) in culture.Cell is in CO237 DEG C of cultures in cell incubator.Three are passed on after cell recovery
More than secondary, it can be used for active testing when length to 80% expires and when in good condition.
(2) concrete operations.With the proliferation inhibition activity of tetrazole (MTT) method test compound on intracellular.In short, thin
Born of the same parents plant in 96 orifice plates, are protected from light the compound incubation 72h that various concentration is added.Then 20 μ L MTT (5mg/ are added in every hole
ML) and it is incubated for 4h.Supernatant is sucked, every hole is added 150 μ L DMSO and shakes 20min.Microplate reader (Thermo Varioskan
Flash the OD value (OD) in each hole under 550nm) is read.Each compound divides into three multiple holes in each concentration.
(3) drug is calculated as follows to the proliferation inhibition rate of cell line: cell proliferation inhibition rate=(ODNegative control examination-ODTest)/
(ODNegative control-ODBlank) × 100%.Dose response can be obtained with the various concentration cell proliferation inhibiting rate mapping of same sample
Curve is analyzed with software GraphPad Prism 5, therefrom finds out the half-inhibitory concentration IC of sample50。
The half-inhibitory concentration IC of above-mentioned 4 kinds of samples50As a result as shown in Figure 2-5.From the figure we can see that chemical combination
Object has good inhibiting effect to the tumour cell that Ras is mutated, and cis-structure inhibiting effect is obvious, and transconfiguration inhibits to make
With weaker, selectivity and controllability are embodied.
Claims (10)
1. a kind of acetylamino azobenzene derivative, which is characterized in that the structural formula of the derivative is as shown in Formulas I or Formula II:
2. a kind of acetylamino azobenzene derivative according to claim 1, which is characterized in that in structural formula on A ring
Substituent R1Selected from one of H, halogen, hydroxyl, nitro, C1~C10 or alkoxy；Substituent R on A ring2Selected from H or
One of halogen；Substituent R on A ring3Selected from one of H or halogen；Substituent R on A ring4Selected from H, halogen or C1
One of~C10；The n is the integer in 1~3, and the X is selected from halogen, alkenyl or dimaleoyl imino；The C1~
C10 is alkyl of the carbon atom number 1~10.
3. a kind of acetylamino azobenzene derivative according to claim 1, which is characterized in that the derivative includes
It is the isotopic compound of substance shown in Formulas I or Formula II, racemic modification, optical active isomers, polymorphic, pharmaceutically acceptable
One of salt or mixture.
4. a kind of preparation method of the acetylamino azobenzene derivative as described in claims 1 to 3 is any, which is characterized in that
The following steps are included:
(1) by ammonium persulfate-sodium bisulfate respectively withMixing, is stirred to react
After, it obtains
(2) step (1) is obtainedIt is dissolved in glacial acetic acid, is addedIt stirs to anti-
It should terminate, be obtained by filtration
Alternatively, step (1) is obtainedIt is dissolved in glacial acetic acid, is addedStirring is extremely
Reaction terminates, and is obtained by filtration
(3) willIt is dissolved in methanol, hydrochloric acid is added, is heated to reflux, is added dropwise after reaction
3M sodium hydrate aqueous solution adjusts pH, and saturated sodium bicarbonate solution then is added dropwise until generating precipitating, separates, be dried to obtain
(4) willIt is dissolved in methylene chloride, is then addedStirring to reaction is tied
Beam, column chromatographic purifying is up to described
5. a kind of preparation method of acetylamino azobenzene derivative according to claim 4, which is characterized in that reaction
The substituent R on A ring occurred in step1Selected from one of H, halogen, hydroxyl, nitro, C1~C10 or alkoxy；On A ring
Substituent R2Selected from one of H or halogen；Substituent R on A ring3Selected from one of H or halogen；Substituent group on A ring
R4Selected from one of H, halogen or C1~C10；The n is the integer in 1~3, and the X is selected from halogen, alkenyl or Malaysia acyl
Imido grpup；C1~the C10 is alkyl of the carbon atom number 1~10.
6. a kind of preparation method of acetylamino azobenzene derivative according to claim 4, which is characterized in that step
(1) ammonium persulfate-sodium bisulfate described inMolar ratio be 1:(1.2
~2), the rate of stirring is 500~1000rpm, and the temperature of reaction is 20~30 DEG C, and the reaction time is 2~12 hours.
7. a kind of preparation method of acetylamino azobenzene derivative according to claim 4, which is characterized in that step
(2) in,WithMolar ratio be (1.5~3): 1, it is describedWithMolar ratio be (1.5~3): 1, the rate of the stirring is 500~
1000rpm, the temperature of reaction are 20~30 DEG C, and the reaction time is 12~15 hours.
8. a kind of preparation method of acetylamino azobenzene derivative according to claim 4, which is characterized in that step
(3) in, the methanol of addition and the volume ratio of hydrochloric acid are 1:(0.5~2),With hydrochloric acid
The molar ratio of middle HCl is 1:(50~100), the temperature of back flow reaction is 80~90 DEG C, and the time is 10~12 hours, 3M hydroxide
Reaction system pH is adjusted to 8~9 by sodium water solution, and the temperature of the drying is 55~65 DEG C.
9. a kind of preparation method of acetylamino azobenzene derivative according to claim 4, which is characterized in that step
(4) in,Molar ratio be 1:(2~3), the temperature of reaction is 20~30
DEG C, the reaction time is 2~12 hours, and purifying uses silica gel column chromatography.
10. a kind of application of the acetylamino azobenzene derivative as described in claims 1 to 3 is any, which is characterized in that described
Derivative is used to prepare the drug of the relevant tumour for the treatment of K-Ras gene mutation, the relevant tumour packet of the K-Ras gene mutation
Include the carcinoma of the rectum, one of lung cancer or prostate cancer.
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| WO2025034702A1 (en) | 2023-08-07 | 2025-02-13 | Revolution Medicines, Inc. | Rmc-6291 for use in the treatment of ras protein-related disease or disorder |
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| WO2022235870A1 (en) | 2021-05-05 | 2022-11-10 | Revolution Medicines, Inc. | Ras inhibitors for the treatment of cancer |
| WO2022235864A1 (en) | 2021-05-05 | 2022-11-10 | Revolution Medicines, Inc. | Ras inhibitors |
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| CN114349657B (en) * | 2021-12-14 | 2023-12-22 | 杭州医学院 | Azo microtubulin inhibitor and preparation method and application thereof |
| WO2024206858A1 (en) | 2023-03-30 | 2024-10-03 | Revolution Medicines, Inc. | Compositions for inducing ras gtp hydrolysis and uses thereof |
| WO2024229406A1 (en) | 2023-05-04 | 2024-11-07 | Revolution Medicines, Inc. | Combination therapy for a ras related disease or disorder |
| WO2025034702A1 (en) | 2023-08-07 | 2025-02-13 | Revolution Medicines, Inc. | Rmc-6291 for use in the treatment of ras protein-related disease or disorder |
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