JP3965455B2 - Vinylated deoxyguanosine derivatives - Google Patents
Vinylated deoxyguanosine derivatives Download PDFInfo
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- JP3965455B2 JP3965455B2 JP22764895A JP22764895A JP3965455B2 JP 3965455 B2 JP3965455 B2 JP 3965455B2 JP 22764895 A JP22764895 A JP 22764895A JP 22764895 A JP22764895 A JP 22764895A JP 3965455 B2 JP3965455 B2 JP 3965455B2
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- YKBGVTZYEHREMT-KVQBGUIXSA-N 2'-deoxyguanosine Chemical class C1=NC=2C(=O)NC(N)=NC=2N1[C@H]1C[C@H](O)[C@@H](CO)O1 YKBGVTZYEHREMT-KVQBGUIXSA-N 0.000 title claims description 11
- 150000003839 salts Chemical class 0.000 claims description 13
- 125000000037 tert-butyldiphenylsilyl group Chemical group [H]C1=C([H])C([H])=C([H])C([H])=C1[Si]([H])([*]C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H])C1=C([H])C([H])=C([H])C([H])=C1[H] 0.000 claims description 6
- 125000002777 acetyl group Chemical group [H]C([H])([H])C(*)=O 0.000 claims description 5
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 3
- 150000001875 compounds Chemical class 0.000 description 88
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 36
- -1 di (p-methoxyphenyl) phenylmethyl Chemical group 0.000 description 30
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 29
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 description 27
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 25
- 239000002904 solvent Substances 0.000 description 25
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 24
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 22
- 239000002253 acid Substances 0.000 description 22
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 21
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 20
- XKRFYHLGVUSROY-UHFFFAOYSA-N Argon Chemical compound [Ar] XKRFYHLGVUSROY-UHFFFAOYSA-N 0.000 description 18
- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 18
- 230000000692 anti-sense effect Effects 0.000 description 18
- 239000000126 substance Substances 0.000 description 16
- 239000000243 solution Substances 0.000 description 15
- 238000005160 1H NMR spectroscopy Methods 0.000 description 13
- 238000004992 fast atom bombardment mass spectroscopy Methods 0.000 description 13
- 238000004519 manufacturing process Methods 0.000 description 13
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 12
- WTDHULULXKLSOZ-UHFFFAOYSA-N Hydroxylamine hydrochloride Chemical compound Cl.ON WTDHULULXKLSOZ-UHFFFAOYSA-N 0.000 description 12
- 239000010410 layer Substances 0.000 description 12
- 150000002148 esters Chemical class 0.000 description 11
- 239000000203 mixture Substances 0.000 description 10
- 239000002798 polar solvent Substances 0.000 description 10
- 238000010898 silica gel chromatography Methods 0.000 description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 10
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 9
- TWSDQEODKUAQFN-JIMJEQGWSA-N CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO[Si](C2=CC=CC=C2)(C3=CC=CC=C3)C(C)(C)C)N4C=NC5=C(N=C(N=C54)N)C=C Chemical compound CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO[Si](C2=CC=CC=C2)(C3=CC=CC=C3)C(C)(C)C)N4C=NC5=C(N=C(N=C54)N)C=C TWSDQEODKUAQFN-JIMJEQGWSA-N 0.000 description 9
- KSCYUGTVWSTJHL-ZGIBFIJWSA-N CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO[Si](C2=CC=CC=C2)(C3=CC=CC=C3)C(C)(C)C)N4C=NC5=C(N=C(N=C54)N)C=C[Si](C)(C)C Chemical compound CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO[Si](C2=CC=CC=C2)(C3=CC=CC=C3)C(C)(C)C)N4C=NC5=C(N=C(N=C54)N)C=C[Si](C)(C)C KSCYUGTVWSTJHL-ZGIBFIJWSA-N 0.000 description 9
- XAHRUDZQJUBLHV-YTFSRNRJSA-N CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO[Si](C2=CC=CC=C2)(C3=CC=CC=C3)C(C)(C)C)N4C=NC5=C4N=C(NC5=O)N Chemical compound CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO[Si](C2=CC=CC=C2)(C3=CC=CC=C3)C(C)(C)C)N4C=NC5=C4N=C(NC5=O)N XAHRUDZQJUBLHV-YTFSRNRJSA-N 0.000 description 9
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 9
- 229910052786 argon Inorganic materials 0.000 description 9
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 9
- INPFZARXMRVOCI-PWRODBHTSA-N 2-amino-9-[(2r,4s,5r)-5-[[tert-butyl(diphenyl)silyl]oxymethyl]-4-hydroxyoxolan-2-yl]-3h-purin-6-one Chemical compound C([C@@H]1[C@H](C[C@@H](O1)N1C2=C(C(N=C(N)N2)=O)N=C1)O)O[Si](C(C)(C)C)(C=1C=CC=CC=1)C1=CC=CC=C1 INPFZARXMRVOCI-PWRODBHTSA-N 0.000 description 8
- VBCDWCFFKJVYKA-OYUWMTPXSA-N CC=CC1=C2C(=NC(=N1)N)N(C=N2)[C@H]3C[C@@H]([C@H](O3)CO[Si](C4=CC=CC=C4)(C5=CC=CC=C5)C(C)(C)C)OC(=O)C Chemical compound CC=CC1=C2C(=NC(=N1)N)N(C=N2)[C@H]3C[C@@H]([C@H](O3)CO[Si](C4=CC=CC=C4)(C5=CC=CC=C5)C(C)(C)C)OC(=O)C VBCDWCFFKJVYKA-OYUWMTPXSA-N 0.000 description 8
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 description 8
- 238000004132 cross linking Methods 0.000 description 8
- NFHFRUOZVGFOOS-UHFFFAOYSA-N palladium;triphenylphosphane Chemical compound [Pd].C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1.C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1.C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1.C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1 NFHFRUOZVGFOOS-UHFFFAOYSA-N 0.000 description 8
- NMHKBHRSEZEPKA-HBNTYKKESA-N CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO)N2C=NC3=C(N=C(N=C32)N)C=C Chemical compound CC(=O)O[C@H]1C[C@@H](O[C@@H]1CO)N2C=NC3=C(N=C(N=C32)N)C=C NMHKBHRSEZEPKA-HBNTYKKESA-N 0.000 description 7
- HYXUTZYGZXVHFS-OYUWMTPXSA-N [Si](C1=CC=CC=C1)(C1=CC=CC=C1)(C(C)(C)C)OC[C@@H]1[C@H](C[C@@H](O1)N1C2=NC(=NC(=C2N=C1)C=C[Si](C)(C)C)N)O Chemical compound [Si](C1=CC=CC=C1)(C1=CC=CC=C1)(C(C)(C)C)OC[C@@H]1[C@H](C[C@@H](O1)N1C2=NC(=NC(=C2N=C1)C=C[Si](C)(C)C)N)O HYXUTZYGZXVHFS-OYUWMTPXSA-N 0.000 description 7
- 150000008282 halocarbons Chemical class 0.000 description 7
- 108020004707 nucleic acids Proteins 0.000 description 7
- 150000007523 nucleic acids Chemical class 0.000 description 7
- 102000039446 nucleic acids Human genes 0.000 description 7
- WFDIJRYMOXRFFG-UHFFFAOYSA-N Acetic anhydride Chemical compound CC(=O)OC(C)=O WFDIJRYMOXRFFG-UHFFFAOYSA-N 0.000 description 6
- SJRJJKPEHAURKC-UHFFFAOYSA-N N-Methylmorpholine Chemical compound CN1CCOCC1 SJRJJKPEHAURKC-UHFFFAOYSA-N 0.000 description 6
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 6
- 150000004945 aromatic hydrocarbons Chemical class 0.000 description 6
- RAXXELZNTBOGNW-UHFFFAOYSA-N imidazole Natural products C1=CNC=N1 RAXXELZNTBOGNW-UHFFFAOYSA-N 0.000 description 6
- KWGKDLIKAYFUFQ-UHFFFAOYSA-M lithium chloride Chemical compound [Li+].[Cl-] KWGKDLIKAYFUFQ-UHFFFAOYSA-M 0.000 description 6
- 239000012046 mixed solvent Substances 0.000 description 6
- NYHBQMYGNKIUIF-UUOKFMHZSA-N Guanosine Chemical class C1=NC=2C(=O)NC(N)=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O NYHBQMYGNKIUIF-UUOKFMHZSA-N 0.000 description 5
- WORJEOGGNQDSOE-UHFFFAOYSA-N chloroform;methanol Chemical compound OC.ClC(Cl)Cl WORJEOGGNQDSOE-UHFFFAOYSA-N 0.000 description 5
- 239000012044 organic layer Substances 0.000 description 5
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 5
- RYHBNJHYFVUHQT-UHFFFAOYSA-N 1,4-Dioxane Chemical compound C1COCCO1 RYHBNJHYFVUHQT-UHFFFAOYSA-N 0.000 description 4
- YYROPELSRYBVMQ-UHFFFAOYSA-N 4-toluenesulfonyl chloride Chemical compound CC1=CC=C(S(Cl)(=O)=O)C=C1 YYROPELSRYBVMQ-UHFFFAOYSA-N 0.000 description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 4
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 4
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 4
- 239000003054 catalyst Substances 0.000 description 4
- 239000003431 cross linking reagent Substances 0.000 description 4
- 239000013078 crystal Substances 0.000 description 4
- 239000004210 ether based solvent Substances 0.000 description 4
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 4
- 108020004999 messenger RNA Proteins 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 description 4
- FPGGTKZVZWFYPV-UHFFFAOYSA-M tetrabutylammonium fluoride Chemical compound [F-].CCCC[N+](CCCC)(CCCC)CCCC FPGGTKZVZWFYPV-UHFFFAOYSA-M 0.000 description 4
- KBDWGFZSICOZSJ-UHFFFAOYSA-N 5-methyl-2,3-dihydro-1H-pyrimidin-4-one Chemical compound N1CNC=C(C1=O)C KBDWGFZSICOZSJ-UHFFFAOYSA-N 0.000 description 3
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 3
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 description 3
- MUBZPKHOEPUJKR-UHFFFAOYSA-N Oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 description 3
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 150000007513 acids Chemical class 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- 238000001914 filtration Methods 0.000 description 3
- 229910052751 metal Inorganic materials 0.000 description 3
- 239000002184 metal Substances 0.000 description 3
- 238000010992 reflux Methods 0.000 description 3
- 238000003756 stirring Methods 0.000 description 3
- UHDGCWIWMRVCDJ-UHFFFAOYSA-N 1-beta-D-Xylofuranosyl-NH-Cytosine Natural products O=C1N=C(N)C=CN1C1C(O)C(O)C(CO)O1 UHDGCWIWMRVCDJ-UHFFFAOYSA-N 0.000 description 2
- YKBGVTZYEHREMT-UHFFFAOYSA-N 2'-deoxyguanosine Natural products C1=2NC(N)=NC(=O)C=2N=CN1C1CC(O)C(CO)O1 YKBGVTZYEHREMT-UHFFFAOYSA-N 0.000 description 2
- 125000001731 2-cyanoethyl group Chemical group [H]C([H])(*)C([H])([H])C#N 0.000 description 2
- NSPMIYGKQJPBQR-UHFFFAOYSA-N 4H-1,2,4-triazole Chemical compound C=1N=CNN=1 NSPMIYGKQJPBQR-UHFFFAOYSA-N 0.000 description 2
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 2
- MIKUYHXYGGJMLM-GIMIYPNGSA-N Crotonoside Natural products C1=NC2=C(N)NC(=O)N=C2N1[C@H]1O[C@@H](CO)[C@H](O)[C@@H]1O MIKUYHXYGGJMLM-GIMIYPNGSA-N 0.000 description 2
- UHDGCWIWMRVCDJ-PSQAKQOGSA-N Cytidine Natural products O=C1N=C(N)C=CN1[C@@H]1[C@@H](O)[C@@H](O)[C@H](CO)O1 UHDGCWIWMRVCDJ-PSQAKQOGSA-N 0.000 description 2
- NYHBQMYGNKIUIF-UHFFFAOYSA-N D-guanosine Natural products C1=2NC(N)=NC(=O)C=2N=CN1C1OC(CO)C(O)C1O NYHBQMYGNKIUIF-UHFFFAOYSA-N 0.000 description 2
- 108020004414 DNA Proteins 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
- ROSDSFDQCJNGOL-UHFFFAOYSA-N Dimethylamine Chemical compound CNC ROSDSFDQCJNGOL-UHFFFAOYSA-N 0.000 description 2
- XPDWGBQVDMORPB-UHFFFAOYSA-N Fluoroform Chemical compound FC(F)F XPDWGBQVDMORPB-UHFFFAOYSA-N 0.000 description 2
- VZCYOOQTPOCHFL-OWOJBTEDSA-N Fumaric acid Chemical compound OC(=O)\C=C\C(O)=O VZCYOOQTPOCHFL-OWOJBTEDSA-N 0.000 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 description 2
- AFVFQIVMOAPDHO-UHFFFAOYSA-N Methanesulfonic acid Chemical compound CS(O)(=O)=O AFVFQIVMOAPDHO-UHFFFAOYSA-N 0.000 description 2
- KDLHZDBZIXYQEI-UHFFFAOYSA-N Palladium Chemical compound [Pd] KDLHZDBZIXYQEI-UHFFFAOYSA-N 0.000 description 2
- 239000012345 acetylating agent Substances 0.000 description 2
- SKCNIGRBPJIUBQ-UHFFFAOYSA-N chloroform;ethyl acetate Chemical compound ClC(Cl)Cl.CCOC(C)=O SKCNIGRBPJIUBQ-UHFFFAOYSA-N 0.000 description 2
- 230000000295 complement effect Effects 0.000 description 2
- UHDGCWIWMRVCDJ-ZAKLUEHWSA-N cytidine Chemical compound O=C1N=C(N)C=CN1[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O1 UHDGCWIWMRVCDJ-ZAKLUEHWSA-N 0.000 description 2
- VGONTNSXDCQUGY-UHFFFAOYSA-N desoxyinosine Natural products C1C(O)C(CO)OC1N1C(NC=NC2=O)=C2N=C1 VGONTNSXDCQUGY-UHFFFAOYSA-N 0.000 description 2
- JXTHNDFMNIQAHM-UHFFFAOYSA-N dichloroacetic acid Chemical compound OC(=O)C(Cl)Cl JXTHNDFMNIQAHM-UHFFFAOYSA-N 0.000 description 2
- XBDQKXXYIPTUBI-UHFFFAOYSA-N dimethylselenoniopropionate Natural products CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- OAYLNYINCPYISS-UHFFFAOYSA-N ethyl acetate;hexane Chemical compound CCCCCC.CCOC(C)=O OAYLNYINCPYISS-UHFFFAOYSA-N 0.000 description 2
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 2
- 229940029575 guanosine Drugs 0.000 description 2
- 229910052500 inorganic mineral Inorganic materials 0.000 description 2
- 239000011777 magnesium Substances 0.000 description 2
- 229910052749 magnesium Inorganic materials 0.000 description 2
- 239000011707 mineral Substances 0.000 description 2
- 235000010755 mineral Nutrition 0.000 description 2
- PSHKMPUSSFXUIA-UHFFFAOYSA-N n,n-dimethylpyridin-2-amine Chemical compound CN(C)C1=CC=CC=N1 PSHKMPUSSFXUIA-UHFFFAOYSA-N 0.000 description 2
- 230000000269 nucleophilic effect Effects 0.000 description 2
- 150000007524 organic acids Chemical class 0.000 description 2
- VLTRZXGMWDSKGL-UHFFFAOYSA-N perchloric acid Chemical compound OCl(=O)(=O)=O VLTRZXGMWDSKGL-UHFFFAOYSA-N 0.000 description 2
- 239000011574 phosphorus Substances 0.000 description 2
- 229910052698 phosphorus Inorganic materials 0.000 description 2
- FAIAAWCVCHQXDN-UHFFFAOYSA-N phosphorus trichloride Chemical compound ClP(Cl)Cl FAIAAWCVCHQXDN-UHFFFAOYSA-N 0.000 description 2
- XHXFXVLFKHQFAL-UHFFFAOYSA-N phosphoryl trichloride Chemical compound ClP(Cl)(Cl)=O XHXFXVLFKHQFAL-UHFFFAOYSA-N 0.000 description 2
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 description 2
- NROKBHXJSPEDAR-UHFFFAOYSA-M potassium fluoride Chemical compound [F-].[K+] NROKBHXJSPEDAR-UHFFFAOYSA-M 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- 229910052708 sodium Inorganic materials 0.000 description 2
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- JOXIMZWYDAKGHI-UHFFFAOYSA-N toluene-4-sulfonic acid Chemical compound CC1=CC=C(S(O)(=O)=O)C=C1 JOXIMZWYDAKGHI-UHFFFAOYSA-N 0.000 description 2
- VZCYOOQTPOCHFL-UHFFFAOYSA-N trans-butenedioic acid Natural products OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 description 2
- QIWRFOJWQSSRJZ-UHFFFAOYSA-N tributyl(ethenyl)stannane Chemical compound CCCC[Sn](CCCC)(CCCC)C=C QIWRFOJWQSSRJZ-UHFFFAOYSA-N 0.000 description 2
- 125000000026 trimethylsilyl group Chemical group [H]C([H])([H])[Si]([*])(C([H])([H])[H])C([H])([H])[H] 0.000 description 2
- PDVFSPNIEOYOQL-UHFFFAOYSA-N (4-methylphenyl)sulfonyl 4-methylbenzenesulfonate Chemical compound C1=CC(C)=CC=C1S(=O)(=O)OS(=O)(=O)C1=CC=C(C)C=C1 PDVFSPNIEOYOQL-UHFFFAOYSA-N 0.000 description 1
- XMNIXWIUMCBBBL-UHFFFAOYSA-N 2-(2-phenylpropan-2-ylperoxy)propan-2-ylbenzene Chemical compound C=1C=CC=CC=1C(C)(C)OOC(C)(C)C1=CC=CC=C1 XMNIXWIUMCBBBL-UHFFFAOYSA-N 0.000 description 1
- KMEMIMRPZGDOMG-UHFFFAOYSA-N 2-cyanoethoxyphosphonamidous acid Chemical compound NP(O)OCCC#N KMEMIMRPZGDOMG-UHFFFAOYSA-N 0.000 description 1
- 125000002103 4,4'-dimethoxytriphenylmethyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C(*)(C1=C([H])C([H])=C(OC([H])([H])[H])C([H])=C1[H])C1=C([H])C([H])=C(OC([H])([H])[H])C([H])=C1[H] 0.000 description 1
- 125000004172 4-methoxyphenyl group Chemical group [H]C1=C([H])C(OC([H])([H])[H])=C([H])C([H])=C1* 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- LSNNMFCWUKXFEE-UHFFFAOYSA-M Bisulfite Chemical compound OS([O-])=O LSNNMFCWUKXFEE-UHFFFAOYSA-M 0.000 description 1
- WKBOTKDWSSQWDR-UHFFFAOYSA-N Bromine atom Chemical group [Br] WKBOTKDWSSQWDR-UHFFFAOYSA-N 0.000 description 1
- FTWFDZDKTPWDJA-UHFFFAOYSA-N CC(C)(C)[SiH](C1=CC=CC=C1)C1=CC=CC=C1.Cl Chemical compound CC(C)(C)[SiH](C1=CC=CC=C1)C1=CC=CC=C1.Cl FTWFDZDKTPWDJA-UHFFFAOYSA-N 0.000 description 1
- 0 CCCC(N)=NC(CCCC1OC(C*)C(*)C1)=C(C(CC(*)N)=C)N Chemical compound CCCC(N)=NC(CCCC1OC(C*)C(*)C1)=C(C(CC(*)N)=C)N 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- 108020004635 Complementary DNA Proteins 0.000 description 1
- YCKRFDGAMUMZLT-UHFFFAOYSA-N Fluorine atom Chemical compound [F] YCKRFDGAMUMZLT-UHFFFAOYSA-N 0.000 description 1
- AVXURJPOCDRRFD-UHFFFAOYSA-N Hydroxylamine Chemical compound ON AVXURJPOCDRRFD-UHFFFAOYSA-N 0.000 description 1
- 125000000393 L-methionino group Chemical group [H]OC(=O)[C@@]([H])(N([H])[*])C([H])([H])C(SC([H])([H])[H])([H])[H] 0.000 description 1
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 description 1
- 239000012359 Methanesulfonyl chloride Substances 0.000 description 1
- PXHVJJICTQNCMI-UHFFFAOYSA-N Nickel Chemical compound [Ni] PXHVJJICTQNCMI-UHFFFAOYSA-N 0.000 description 1
- 229910021586 Nickel(II) chloride Inorganic materials 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- OFOBLEOULBTSOW-UHFFFAOYSA-N Propanedioic acid Natural products OC(=O)CC(O)=O OFOBLEOULBTSOW-UHFFFAOYSA-N 0.000 description 1
- WETWJCDKMRHUPV-UHFFFAOYSA-N acetyl chloride Chemical compound CC(Cl)=O WETWJCDKMRHUPV-UHFFFAOYSA-N 0.000 description 1
- 239000012346 acetyl chloride Substances 0.000 description 1
- 230000002152 alkylating effect Effects 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 235000019270 ammonium chloride Nutrition 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229910052801 chlorine Inorganic materials 0.000 description 1
- 125000001309 chloro group Chemical group Cl* 0.000 description 1
- MMFVFNVXXDFELX-UHFFFAOYSA-N chloroform;n,n-diethylethanamine Chemical compound ClC(Cl)Cl.CCN(CC)CC MMFVFNVXXDFELX-UHFFFAOYSA-N 0.000 description 1
- WDJQAYUPVJEQJW-UHFFFAOYSA-N chloroform;n,n-diethylethanamine;methanol Chemical compound OC.ClC(Cl)Cl.CCN(CC)CC WDJQAYUPVJEQJW-UHFFFAOYSA-N 0.000 description 1
- SBUXRMKDJWEXRL-ROUUACIJSA-N cis-body Chemical compound O=C([C@H]1N(C2=O)[C@H](C3=C(C4=CC=CC=C4N3)C1)CC)N2C1=CC=C(F)C=C1 SBUXRMKDJWEXRL-ROUUACIJSA-N 0.000 description 1
- 239000012230 colorless oil Substances 0.000 description 1
- GYOZYWVXFNDGLU-XLPZGREQSA-N dTMP Chemical class O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](COP(O)(O)=O)[C@@H](O)C1 GYOZYWVXFNDGLU-XLPZGREQSA-N 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 229960005215 dichloroacetic acid Drugs 0.000 description 1
- HPNMFZURTQLUMO-UHFFFAOYSA-N diethylamine Chemical compound CCNCC HPNMFZURTQLUMO-UHFFFAOYSA-N 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 229910052731 fluorine Inorganic materials 0.000 description 1
- 239000011737 fluorine Substances 0.000 description 1
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- 125000000524 functional group Chemical group 0.000 description 1
- 230000014509 gene expression Effects 0.000 description 1
- 238000001415 gene therapy Methods 0.000 description 1
- 230000002140 halogenating effect Effects 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- RMGJCSHZTFKPNO-UHFFFAOYSA-M magnesium;ethene;bromide Chemical compound [Mg+2].[Br-].[CH-]=C RMGJCSHZTFKPNO-UHFFFAOYSA-M 0.000 description 1
- VZCYOOQTPOCHFL-UPHRSURJSA-N maleic acid Chemical compound OC(=O)\C=C/C(O)=O VZCYOOQTPOCHFL-UPHRSURJSA-N 0.000 description 1
- 239000011976 maleic acid Substances 0.000 description 1
- 150000002739 metals Chemical class 0.000 description 1
- 229940098779 methanesulfonic acid Drugs 0.000 description 1
- IZDROVVXIHRYMH-UHFFFAOYSA-N methanesulfonic anhydride Chemical compound CS(=O)(=O)OS(C)(=O)=O IZDROVVXIHRYMH-UHFFFAOYSA-N 0.000 description 1
- QARBMVPHQWIHKH-UHFFFAOYSA-N methanesulfonyl chloride Chemical compound CS(Cl)(=O)=O QARBMVPHQWIHKH-UHFFFAOYSA-N 0.000 description 1
- BMGNSKKZFQMGDH-FDGPNNRMSA-L nickel(2+);(z)-4-oxopent-2-en-2-olate Chemical compound [Ni+2].C\C([O-])=C\C(C)=O.C\C([O-])=C\C(C)=O BMGNSKKZFQMGDH-FDGPNNRMSA-L 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 235000006408 oxalic acid Nutrition 0.000 description 1
- 229910052763 palladium Inorganic materials 0.000 description 1
- QJPQVXSHYBGQGM-UHFFFAOYSA-N palladium;triphenylphosphane Chemical compound [Pd].C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1.C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1 QJPQVXSHYBGQGM-UHFFFAOYSA-N 0.000 description 1
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- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 229910000027 potassium carbonate Inorganic materials 0.000 description 1
- 239000011698 potassium fluoride Substances 0.000 description 1
- 235000003270 potassium fluoride Nutrition 0.000 description 1
- 235000019260 propionic acid Nutrition 0.000 description 1
- IUVKMZGDUIUOCP-BTNSXGMBSA-N quinbolone Chemical compound O([C@H]1CC[C@H]2[C@H]3[C@@H]([C@]4(C=CC(=O)C=C4CC3)C)CC[C@@]21C)C1=CCCC1 IUVKMZGDUIUOCP-BTNSXGMBSA-N 0.000 description 1
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- MHYGQXWCZAYSLJ-UHFFFAOYSA-N tert-butyl-chloro-diphenylsilane Chemical compound C=1C=CC=CC=1[Si](Cl)(C(C)(C)C)C1=CC=CC=C1 MHYGQXWCZAYSLJ-UHFFFAOYSA-N 0.000 description 1
- HWCKGOZZJDHMNC-UHFFFAOYSA-M tetraethylammonium bromide Chemical compound [Br-].CC[N+](CC)(CC)CC HWCKGOZZJDHMNC-UHFFFAOYSA-M 0.000 description 1
- CZDYPVPMEAXLPK-UHFFFAOYSA-N tetramethylsilane Chemical compound C[Si](C)(C)C CZDYPVPMEAXLPK-UHFFFAOYSA-N 0.000 description 1
- SBUXRMKDJWEXRL-ZWKOTPCHSA-N trans-body Chemical compound O=C([C@@H]1N(C2=O)[C@H](C3=C(C4=CC=CC=C4N3)C1)CC)N2C1=CC=C(F)C=C1 SBUXRMKDJWEXRL-ZWKOTPCHSA-N 0.000 description 1
- FJXYMVBFBYAWDR-UHFFFAOYSA-N tributyl(prop-1-enyl)stannane Chemical compound CCCC[Sn](CCCC)(CCCC)\C=C\C FJXYMVBFBYAWDR-UHFFFAOYSA-N 0.000 description 1
- PIILXFBHQILWPS-UHFFFAOYSA-N tributyltin Chemical compound CCCC[Sn](CCCC)CCCC PIILXFBHQILWPS-UHFFFAOYSA-N 0.000 description 1
- GRGCWBWNLSTIEN-UHFFFAOYSA-N trifluoromethanesulfonyl chloride Chemical compound FC(F)(F)S(Cl)(=O)=O GRGCWBWNLSTIEN-UHFFFAOYSA-N 0.000 description 1
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- JDQLLFOVXMXKNW-UHFFFAOYSA-N trimethyl(2-tributylstannylethenyl)silane Chemical compound CCCC[Sn](CCCC)(CCCC)\C=C\[Si](C)(C)C JDQLLFOVXMXKNW-UHFFFAOYSA-N 0.000 description 1
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Description
【０００１】
【発明の属する技術分野】
本発明はビニル化デオキシグアノシン誘導体又はその塩に関する。より詳細にいうと、本発明は、核酸のクロスリンク剤あるいはクロスリンク能を有するアンチセンスオリゴ核酸の製造に有用なビニル化デオキシグアノシン誘導体又はその塩に関する。
【０００２】
【従来の技術】
特定の遺伝子やメッセンジャーRNA (m-RNA) などの核酸に対して、その核酸配列と相補的な塩基配列を有するアンチセンスオリゴ核酸を特異的に結合させ、遺伝子発現を抑制する方法が知られている。この方法は、生化学的な実験ツールとして利用されるばかりではなく、本格的な遺伝子治療に応用できる可能性があることから、生化学及び医学・薬学の分野において大きな関心がもたれている。
【０００３】
この方法を効率的に行うためには、相補的結合により生成する二重鎖又は三重鎖複合体の安定化が必要とされており、安定化のための種々の手段が提案されている。その代表的なものとして、例えば、アルキル化能を有する官能基をオリゴ核酸中に導入し、遺伝子DNA やm-RNA などの標的核酸と共有結合しうる能力（クロスリンク能）を付与したアンチセンスオリゴ核酸が知られている（例えば、Volssov, V.V. et al., Nucleic Acid Res., 14, 4065, 1986; Webb, T.R. et al., J. Am. Chem. Soc., 108, 2764, 1986; Baker, B.F. et al., J. Am. Chem. Soc., 111, 2700, 1989などを参照）。
【０００４】
しかしながら、このように修飾されたアンチセンスオリゴ核酸の多くは、クロスリンク速度が遅いという問題があり、また反応性が高いものは化学的安定性が低いという欠点を有している。このようなアンチセンスオリゴ核酸を生体に適用することは望ましくない(Rokita, S.E. et al., J. Am. Chem. Soc., 116, 1690, 1994）。
【０００５】
【発明が解決しようとする課題】
本発明の課題は、標的核酸に効率的にクロスリンクする性質を有するアンチセンスオリゴ核酸中の製造に有用な化合物を提供することにある。また、本発明の課題は、上記のような化合物であって、化学的に安定な化合物を提供することにある。さらに本発明の別な課題は、上記の特徴を有する化合物であって、生体に適用可能なアンチセンスオリゴ核酸の製造に有用な化合物を提供することである。
【０００６】
【課題を解決するための手段】
本発明者らは上記の課題を解決すべく鋭意検討を重ねた結果、下記構造のビニル化グアノシン誘導体が化学的に安定であり、かつ求核性ヌクレオチドと容易に反応して強固なクロスリンクを形成できることを見い出した。また、上記の化合物が、標的遺伝子核酸に対して効率的にクロスリンクする性質を有するアンチセンスオリゴ核酸の製造に有用であることを見い出した。本発明はこれらの知見を基にして完成されたものである。
【０００７】
すなわち本発明は、下記式(I):
【化３】
【０００８】
【発明の実施の形態】
上記式(I) 中のR1、R2、及びR3は、下記の(1) 〜(3) のいずれかにより定義される。
(1)R1 は水素原子を示し、R2は水素原子、tert- ブチルジフェニルシリル基、若しくは下記の式：
【化４】
(2) R1はメチル基を示し、R2はtert- ブチルジフェニルシリル基を示し、かつR3はアセチル基を示す；又は
(3) R1はトリメチルシリル基を示し、R2はtert- ブチルジフェニルシリル基を表し、かつR3は水素原子、アセチル基、若しくは-P(O)(OH)Hを示す。
【０００９】
上記化合物は、酸付加塩又は塩基付加塩を形成する場合があるが、このような塩も本発明の範囲に包含される。本発明の化合物自体をクロスリンク剤として生体に適用する場合や、生体に適用可能なアンチセンスオリゴ核酸の製造のための製造中間体として用いる場合には、塩としては生理的に許容されるものが好ましい。塩基付加塩としては、例えば、トリエチルアミン、ジメチルアミン、アンモニア、トリエチルアミン、若しくはジエチルアミン等のアミン類の塩；又はナトリウム、カリウム、カルシウム、若しくはマグネシウム等の金属類の塩を挙げることができる。酸付加塩としては、例えば、塩酸、硫酸、若しくは過塩素酸等の鉱酸類の塩；又はシュウ酸、フマル酸、マレイン酸、酢酸、プロピオン酸、メタンスルホン酸、若しくはp-トルエンスルホン酸等の有機酸等との塩を挙げることができる。
【００１０】
本発明の化合物を以下の表１に具体的に示す。表中、R2として「化４」と示されている場合には、R2がチミジル酸誘導体の基（上記〔化４〕で示される基）であり、該基におけるR4がそれぞれ表中に示されたものであることを示す。また、表中、Ac、Me、Et、TBDPS 、TMS 、及びDMTrは、それぞれ、アセチル基、メチル基、エチル基、tert- ブチルジフェニルシリル基、トリメチルシリル基、及びジ (p-メトキシフェニル）フェニルメチル基を表す（以下、本明細書中において同様である）。
【００１１】
【表１】
本発明化合物の製造方法の例を下記スキームに従って説明するが、本発明の化合物の製造方法はこれらの方法に限定されることはない。スキーム中、X は塩素原子、臭素原子、p-トルエンスルホニルオキシ基、又はトリフルオロメタンスルホニルオキシ基などの脱離基を表し、Met はトリブチルスズ、マグネシウムハライド等の金属を表す（以下のスキームにおいて同様である）。
【００１３】
【化５】
化合物(2):
ジクロロメタン若しくはクロロホルム等のハロゲン化炭化水素系溶媒、又はジメチルホルムアミド若しくはジメチルスルホキシド等の非プロトン性極性溶媒等の適当な溶媒中、2'- デオキシグアノシン(1) をトリエチルアミン、イミダゾール等の塩基の存在下で0.2 〜10当量の塩化 tert-ブチルジフェニルシランと反応させることにより化合物(2) を製造することができる。通常の場合、反応は-20 ℃〜+150℃の温度で５分間〜24時間行えばよい。
【００１５】
化合物(3):
ベンゼン若しくはトルエン等の芳香族炭化水素系溶媒、ジクロロメタン若しくはクロロホルム等のハロゲン化炭化水素系溶媒、テトラヒドロフラン若しくはジエチルエーテル等のエーテル系溶媒、又はそれらの混合溶媒中で、化合物(2) を0.2 〜 100当量の無水酢酸若しくは塩化アセチルなどのアセチル化剤とピリジン若しくはトリエチルアミン等の塩基存在下に反応させるか、あるいは上記アセチル化剤を溶媒兼用で用いて、必要により上記塩基存在下に反応させることにより、化合物(3) を製造することができる。通常の場合、反応は０〜 150℃の温度で５分間〜24時間行えばよい。
【００１６】
化合物(4):
ベンゼン若しくはトルエン等の芳香族炭化水素系溶媒、ジクロロメタン若しくはクロロホルム等のハロゲン化炭化水素系溶媒、テトラヒドロフラン若しくはジエチルエーテル等のエーテル系溶媒、ジメチルホルムアミド、水、若しくはメタノール等の極性溶媒、又はそれらの混合溶媒中で、化合物(3) を例えば、0.2 〜 100当量のp-トルエンスルホニルクロリド、メタンスルホニルクロリド、若しくはトリフルオロメタンスルホニルクロリド、又はp-トルエンスルホン酸無水物、メタンスルホン酸無水物、若しくはトリフルオロメタンスルホン酸無水物などのスルホニル化剤とピリジン若しくはトリエチルアミン等の塩基存在下に反応させるか、あるいは上記スルホニル化剤を溶媒兼用で用いて、必要により上記塩基存在下に反応させることにより、化合物(4) を製造することができる。通常の場合、反応は０〜 150℃の温度で５分間〜24時間行えばよい。上記反応において、ジメチルアミノピリジン等の反応促進剤を0.01〜10当量共存させてもよい。
【００１７】
また、ベンゼン若しくはトルエン等の芳香族炭化水素系溶媒、ジクロロメタン若しくはクロロホルム等のハロゲン化炭化水素系溶媒、テトラヒドロフラン若しくはジエチルエーテル等のエーテル系溶媒、ジメチルホルムアミド、水、若しくはメタノール等の極性溶媒、又はそれらの混合溶媒中、あるいは溶媒の非存在下に、化合物(3) を0.2 〜 100当量のオキシ塩化リン、オキシ臭化リン、五塩化リン、若しくは塩化チオニル等のハロゲン化剤で処理することによっても化合物(4) を製造することができる。通常の場合、反応は０〜 150℃の温度で５分間〜24時間行えばよい。
【００１８】
化合物(Ia):
ベンゼン若しくはトルエン等の芳香族炭化水素系溶媒、ジクロロメタン若しくはクロロホルム等のハロゲン化炭化水素系溶媒、テトラヒドロフラン若しくはジオキサン等のエーテル系溶媒、ジメチルホルムアミド、水、若しくはメタノール等の極性溶媒、又はそれらの混合溶媒中で、ビニルトリブチル錫若しくはビニルマグネシウムブロミド等のビニル金属化合物(5) 0.2 〜 100当量と化合物(4) とを0.0001〜１当量の触媒の存在下に反応させることにより化合物(Ia)を製造することができる。触媒としては、テトラキス（トリフェニルホスフィン）パラジウム若しくは塩化ビス（トリフェニルホスフィン）パラジウム等のパラジウム触媒、又は塩化ビス（アセチルアセトナート）ニッケル等のニッケル触媒を用いることができる。上記反応をピリジン若しくはトリエチルアミン等の塩基存在下に行ってもよい。通常の場合、反応は０〜 150℃の温度で５分間〜24時間行えばよい。
【００１９】
化合物(Ib):
テトラヒドロフラン若しくはエーテル等のエーテル系溶媒、ジメチルホルムアミド、水、若しくはメタノール等の極性溶媒、又はそれらの混合溶媒中で、化合物(Ia)をフッ化テトラブチルアンモニウム若しくはフッ化カリウム等の含フッ素化合物 0.2〜 100当量と反応させることにより化合物(Ib)を製造することができる。通常の場合、反応は０〜 100℃の温度で５分間〜24時間行えばよい。
【００２０】
化合物(Ic):
アセトニトリル、ジメチルホルムアミド、若しくはジメチルスルホキシド等の適当な溶媒中、化合物(Ib)を0.2 〜 100当量の1-テトラゾール存在下に0.2 〜10当量のβ−シアノエチルホスホラミダイトデオキシチミン(6) と０〜 100℃で５分間〜24時間反応させた後、得られた生成物とt-ブチルハイドロペルオキシド若しくはクメンペルオキシド等の過酸化物とを反応させることにより、化合物(Ic)を製造することができる。通常の場合、反応は -20〜50℃で５分間〜24時間程度行えばよい。
【００２１】
化合物(Id):
ベンゼン若しくはトルエン等の芳香族炭化水素系溶媒、ジクロロメタン若しくはクロロホルム等のハロゲン化炭化水素系溶媒、テトラヒドロフラン若しくはジエチルエーテル等のエーテル系溶媒、ジメチルホルムアミド、水、若しくはメタノール等の極性溶媒、又はそれらの混合溶媒中、あるいは溶媒の非存在下に、化合物(Ic)を0.2 〜 100当量のトリフルオロ酢酸若しくはジクロロ酢酸等の有機酸、又は塩酸若しくは硫酸等の鉱酸で処理することにより化合物(Id)を製造することができる。反応は一般的に -20〜50℃の温度で５分間〜24時間行えばよい。
【００２２】
【化６】
化合物(IIa):
スキーム１中の化合物(4) と化合物(5) から化合物(Ia)を製造する反応の条件に準じて、化合物(4) と化合物(7) とを反応させることにより、化合物(IIa) を製造することができる（上記スキーム２を参照）。
【００２４】
【化７】
化合物(IIIa)及び化合物(IIIb):
スキーム１中の化合物(4) と化合物(5) から化合物(Ia)を製造する反応の条件に準じて、化合物(4) と化合物(8) とを反応させることにより、化合物(IIIa)を製造することができる（上記スキーム３を参照）。テトラヒドロフラン若しくはエーテル等のエーテル系溶媒、ジメチルホルムアミド、水、若しくはメタノール等の極性溶媒、又はそれらの混合溶媒中で、化合物(IIIa)を 0.2〜 100当量の水酸化ナトリウム若しくは水酸化カリウム等の金属水酸化物と反応させることにより、化合物(IIIb)を製造することができる。反応は、一般的に０〜100 ℃の温度で５分間〜24時間行えばよい。
【００２６】
化合物(IIIc):
ベンゼン若しくはトルエン等の芳香族炭化水素系溶媒、ジクロロメタン若しくはクロロホルム等のハロゲン化炭化水素系溶媒、テトラヒドロフラン若しくはジエチルエーテル等のエーテル系溶媒、ジメチルホルムアミド等の極性溶媒、又はそれらの混合溶媒中、0.5 〜50当量のN-メチルモルホリン若しくはトリエチルアミン等の塩基存在下に、化合物(IIIb)をリン化合物(9) と反応させた後、生成物にトリエチルアミンを作用させることにより化合物(IIIc)を製造することができる。リン化合物(9) は、例えば0.5 〜10当量の三塩化リンと0.5 〜10当量の1,2,4-トリアゾールを -20〜50℃で５分間〜24時間反応させることにより調製することができる。通常の場合、上記反応は-100〜50℃の温度で５分間〜48時間行えばよい。
【００２７】
本発明の化合物は、シチジンやグアノシンと速やかに付加体を形成することができるので、本発明の化合物を製造中間体として用いることにより、遺伝子DNA やm-RNA などの標的核酸に対して相補的に結合し、かつ共有結合によるクロスリンクを形成するアンチセンスオリゴ核酸を製造することができる。従って、本発明の別の態様により、本発明の化合物からなるアンチセンスオリゴ核酸製造用中間体、好ましくはクロスリンク能を有するアンチセンスオリゴ核酸製造用中間体が提供される。また、上記化合物を製造用中間体として用いて製造されたアンチセンスオリゴ核酸、好ましくはクロスリンク能を有するアンチセンスオリゴ核酸も、本発明の範囲に包含される。もっとも、本発明の上記化合物の用途は、これらの用途に限定されることはない。
【００２８】
以下、本発明を実施例によりさらに具体的に説明するが、本発明の範囲は下記の実施例に限定されることはない。また、上記の一般的な製造方法の説明及び下記の実施例に記載した試薬や反応条件には適宜の修飾や改変が可能であることはいうまでもない。なお、実施例中の化合物番号は上記スキーム中の化合物番号に対応している。
【００２９】
【実施例】
例１：5'-O-tert-ブチルジフェニルシリル-2'-デオキシグアノシン(2)
アルゴン気流下、2'- デオキシグアノシン (2.0 g 、7.2 mmol) の無水 DMF (30 ml)懸濁液に t- ブチルクロロジフェニルシラン (3.0 ml、12 mmol)とイミダゾール (1.5 g 、21 mmol)を加え、室温にて５時間攪拌した。５時間後、反応層に水 (30 ml)を加えて得られた結晶を濾取し、酢酸エチルで洗浄した。メタノールから再結晶し、化合物(2) を無色粉末状結晶として得た (1.5 g 、3.0 mmol、収率 41%) 。m.p.: 163-166 ℃。
【００３０】
1H-NMR (DMSO-d6) δ 10.60 (1H,brs), 7.80 (1H,s), 7.7-7.3 (10H,m), 6.48 (2H,s), 6.14 (1H,t,J=6.6Hz), 5.38 (1H,d,J=4.0Hz), 4.50-4.40 (1H,m), 3.95-3.65 (2H,s), 2.65-2.45 (1H,m), 2.35-2.20 (1H,m), 0.99 (9H,s)
IR (cm-1, neat) : 3600-2700, 1680, 1630
FABMS (m/z) : 506(M+1)+ , 528(M+23) +
例２：3'-O- アセチル-5'-O-tert- ブチルジフェニルシリル-2'-デオキシグアノシン(3)
アルゴン気流下、化合物(2) (870 mg 、1.7 mmol) の無水ピリジン (35 ml)溶液に無水酢酸 (0.8 ml、8.5 mmol) を加え、85℃で攪拌を行った。５時間後、反応層を酢酸エチル (50 ml)で希釈し、水 (20 ml ×2)及び飽和食塩水 (20 ml)で洗浄、無水硫酸ナトリウムで乾燥後、減圧下留去した。残渣をアセトニトリルから再結晶し 化合物(3) を無色粉末状結晶として得た (651 mg、1.2 mmol、収率 70%) 。m.p.: 226-229 ℃。
【００３２】
1H-NMR (CDCl3) δ 12.01 (1H,brs), 7.80-7.30 (10H,m), 6.23 (3H,dd,J=8.7, 5.8Hz), 5.53 (1H,d,J=5.9Hz), 4.20 (1H,m), 3.89 (2H,d,J=3.6Hz), 2.85-2.70 (1H,m), 2.60-2.45 (1H,m), 2.10 (3H,s), 0.99 (9H,s)
IR (cm-1, neat) : 3500-2600, 1740, 1680, 1600
FABMS (m/z) : 548(M+1)+ , 570(M+23) +
例３：3'-O- アセチル-5'-O-tert- ブチルジフェニルシリル -6-O-p-トルエンスルホニル-2'-デオキシグアノシン(4)
アルゴン気流下、化合物(3) (625 mg 、1.14 mmol)の無水ジクロロメタン (15 ml)溶液を０℃に冷却し、トリエチルアミン (0.5 ml、3.59 mmol)、p-トルエンスルホニルクロリド (1.08 g、5.65 mmol)及びジメチルアミノピリジン (43.3 mg 、0.355 mmol) を加えて攪拌を行った。20時間後、反応層をクロロホルム (15 ml)で希釈し、有機層を水 (10 ml)で洗浄、水層をクロロホルム (10 ml ×2)で逆抽出した。有機層を合わせて、無水硫酸ナトリウムで乾燥後、減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (FL-60D、クロロホルム−酢酸エチル 9:1→4:1)にて精製し 化合物(4) を無色カラメル状物質として得た (800 mg、1.14 mmol 、収率 100%)。
【００３４】
1H-NMR (CDCl3) δ 8.02 (2H,d,J=8.3Hz), 7.87 (1H,s), 7.70-7.60 (4H,m), 7.50-7.30 (8H,m), 6.26 (1H,dd,J=8.7, 5.4Hz), 5.55 (1H,m), 4.91 (2H,s), 4.25-4.15 (1H,m), 3.90 (2H,m), 2.90-2.75 (1H,m), 2.60-2.40 (1H,m), 2.45 (3H,s), 2.11 (3H,s), 1.06 (9H,s)
IR (cm-1, neat) : 3600-3000, 1740, 1620
FABMS (m/z) : 702(M+1)+ , 644(M-57) +
例４：9-(3-O- アセチル-5-O-tert-ブチルジフェニルシリル -2-デオキシ -β-D- リボフラノシル)-2-アミノ -6-ビニルプリン(Ia)
アルゴン気流下、化合物(4) (320 mg 、0.46 mmol)の無水ジオキサン (5 ml) 溶液に塩化リチウム (40 mg 、0.94 mmol)及びテトラキス（トリフェニルホスフィン）パラジウム(0) (110 mg 、0.095 mmol) を加えて室温にて攪拌した。30分後、ビニルトリ (n-ブチル）錫 (760 μl 、2.3 mmol) を加えて、加熱還流した。90分後、反応層を酢酸エチル-10%アンモニア水 (30 ml: 3 ml)にあけ、有機層を 10%アンモニウム水 (3 ml) 、飽和食塩水 (3 ml×2) で洗浄、無水硫酸ナトリウムで乾燥後、減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (クロロホルム−酢酸エチル 4:1) にて精製し化合物(Ia)を淡黄色カラメル状物質として得た (150 mg、0.27 mmol 、収率 59%) 。
【００３６】
1H-NMR (CDCl3) δ 7.93 (1H,s), 7.75-7.30 (10H,m), 7.13 (1H,dd,J=17.6, 10.7Hz), 6.86 (1H,dd,J=17.6, 1.9Hz), 6.34 (1H,dd,J=8.9, 5.6Hz), 5.85 (1H,dd,J=10.7, 1.9Hz), 5.56 (1H,m), 4.87 (2H,s), 4.20 (1H,m), 3.90 (2H,m), 2.95-2.75 (1H,m), 2.60-2.50 (1H,m), 2.12 (3H,s), 1.07 (9H,s)
IR (cm-1, neat) : 3600-3100, 1740, 1600
FABMS (m/z) : 558(M+1)+ , 500(M-57) +
例５：9-(3-O- アセチル -2-デオキシ -β-D- リボフラノシル)-2-アミノ -6-ビニルプリン(Ib)
アルゴン気流下、化合物(Ia) (25 mg 、46μmol)の無水テトラヒドロフラン (0.1 ml) 溶液を０℃に冷却し、テトラブチルアンモニウムフルオリド(100μl 、100 μmol 、THF 中 1M 溶液）を滴下した。１時間後、反応層をシリカゲルカラムクロマトグラフィー (FL-60D、ヘキサン−酢酸エチル 9:1) にて精製し化合物(Ib)を無色油状物質として得た (14 mg 、44μmol 、収率 98%) 。
【００３８】
1H-NMR (CDCl3) δ 7.77 (1H,s), 7.12 (1H,dd,J=17.5, 10.9Hz), 6.89 (1H,dd,J=17.6, 1.8Hz), 6.21 (1H,dd,J=9.9, 5.6Hz), 5.89 (1H,dd,J=10.6, 2.0Hz), 5.53 (1H,d,J=5.6Hz), 5.07 (2H,brs), 4.25 (1H,m), 3.20 (1H,dd,J=9.2, 5.0Hz), 2.38 (1H,dd,J=14.0, 5.4Hz), 2.13 (3H,s)
IR (cm-1,neat) : 3350, 3200, 1730
FABMS (m/z) : 320(M+1)+ , 550(M-31) +
【００３９】
例６：5'-O- ジ（パラメトキシフェニル）フェニルメチルチミジリル-(3'-5')-9-(3-O-アセチル -2-デオキシ -β-D- リボフラノシル)-2-アミノ -6-ビニルプリン-3'-[P- β-(シアノエチル）エステル(Ic)
アルゴン気流下、化合物(Ib) (24 mg 、75μmol)とT-ベータアミダイト(84 mg、110 μmol)の無水アセトニトリル (0.5 ml) 溶液に、1-テトラゾール(6) (73 mg、90μmol)を加えて室温にて攪拌を行った。２時間攪拌後、反応層を０℃に冷却し、t-ブチルヒドロパーオキサイド (16μl 、86μmol 、トルエン中 5.4M 溶液）を加えて攪拌を続けた。２時間後、不溶物を濾去し減圧下溶媒留去した。残渣をシリカゲルカラムクロマトグラフィー (FL-60D、クロロホルム−メタノール 99:1 →95:5) にて精製し化合物(Ic)を無色油状物質として得た (29 mg 、30μmol 、収率 40%) 。
【００４０】
1H-NMR (CDCl3) δ 7.86 (1H,s), 7.53 (0.5H,d,J=0.99Hz), 7.46 (0.5H,d,J=13Hz), 7.42-7.17 (10H,m), 7.10 (1H,ddd,J=17.5, 10.9, 2.6Hz), 6.90-6.77 (4H,m), 6.44-6.33 (1H,m), 6.30-6.23 (1H,m), 5.84 (1H,ddd,J=10.9, 3.6, 1.98Hz), 5.47 (1H,dd,J=15.8, 5.3Hz), 5.35-5.25 (3H,m), 5.22-5.10 (1H,m), 4.60-4.05 (6H,m), 3.79 (6H,s), 3.55-3.27 (3H,m), 2.72-2.24 (5H,m), 2.12 (1.5H,s), 2.11 (1.5H,s), 1.41 (1.5H,d,J=0.99Hz), 1.38 (1.5H,d,J=0.99Hz)
IR (cm-1,neat) : 3480, 3340, 3200, 1740, 1690, 1600
FABMS (m/z) : 979(M+1)+
【００４１】
例７：チミジリル-(3'-5')-9-(3-O-アセチル -2-デオキシ -β-D- リボフラノシル)-2-アミノ -6-ビニルプリン-3'-[P- β-(シアノエチル）エステル(Id)
化合物(Ic) (2.0 mg、2.0 μmol)に、 CHCl3 0.1 ml 中のトリフルオロ酢酸 (0.2 ml、3 μmol)を滴下した。５分後、反応層にエーテル (ca. 1 ml) を加えて、析出する結晶を濾取し、化合物(Id)を無色粉状物質として得た (0.6 mg、0.9 μmol 、収率 45%) 。
【００４２】
1H-NMR (CDCl3) δ 8.36 (0.5H,s), 8.32 (0.5H,s), 7.44 (0.5H,s), 7.43 (0.5H,s), 7.12-7.03 (1H,m), 6.35-6.26 (1H,m), 6.22-6.13 (1H,m), 6.06-5.96 (1H,m), 5.58-5.48 (1H,m), 5.22-5.03 (1H,m), 4.63-4.17 (7H,m), 3.91-3.73 (3H,m), 3.54-3.19 (3H,m), 2.82-2.72 (2H,m), 2.60-2.36 (2H,m), 2.16 (1.5H,s), 2.15 (1.5H,s), 1.91 (1.5H,d,J=0.99Hz), 1.89 (1.5H,d,J=0.99Hz)
FABMS (m/z) : 677(M+1)+
【００４３】
例８：9-(3-O- アセチル-5-O-tert-ブチルジフェニルシリル -2-デオキシ -β-D- リボフラノシル)-2-アミノ -6-メチルビニルプリン(IIa)
アルゴン気流下、化合物(4) (350 mg 、0.5 mmol) の無水ジオキサン (5 ml) 溶液に塩化リチウム (42 mg 、1.0 mmol) 及びテトラキス（トリフェニルホスフィン）パラジウム(0) (116 mg 、0.1 mmol) を加え室温にて攪拌した。30分後、メチルビニルトリ (n-ブチル）錫 (800 μl 、2.5 mmol、cis/trans=2.5/1)を加えて、加熱還流した。６時間後、反応層を酢酸エチル (100 ml) にあけ、有機層を 10%アンモニウム水 (30 ml)、飽和食塩水(30 ml×2)で洗浄、無水硫酸ナトリウムで乾燥後、減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (クロロホルム−メタノール = 99:1)にて精製し化合物(IIa) を淡黄色カラメル状物質として得た (238 mg、0.42 mmol 、収率 84%) 。
【００４４】
1H-NMR (CDCl3) δ 7.90 (1H,s), 7.68-7.63 (4H,m), 7.51-7.32 (6.6H,m), 6.87 (0.4H,dq,J=11.2, 2.0Hz), 6.83 (0.6H,dq,J=15.5, 1.6Hz), 6.28 (0.4H,dq,J=11.5, 7.2Hz), 6.38 (1H,t,J=6.3Hz), 5.55 (1H,d,J=4.6Hz), 4.83 (1H,bs), 4.19 (1H,dd,J=3.3, 2.0Hz), 3.91-3.89 (2H,m), 2.94-2.81 (1H,m), 2.56-2.51 (1H,m), 2.25 (3H,dd,J=1.7, 7.6Hz), 2.02 (3H,dd,J=1.7, 6.9Hz), 2.02 (3H,s), 1.06 (9H,s)
IR (cm-1,neat) : 3500, 1740, 1600
FABMS (m/z) : 572(M+1)+ , 514(M-57) +
【００４５】
例９：9-(3-O- アセチル-5-O-tert-ブチルジフェニルシリル -2-デオキシ -β-D- リボフラノシル)-2-アミノ-6-(2-トリメチルシリルビニル）プリン(IIIa)
アルゴン気流下、化合物(4) (104 mg 、0.15 mmol)の無水ジオキサン (2 ml) 溶液に塩化リチウム (13 mg 、0.3 mmol) 及びテトラキス（トリフェニルホスフィン）パラジウム(0) (35 mg、0.03 mmol)を加え室温にて攪拌した。30分後、トリメチルシリルビニルトリ (n-ブチル）錫 (1.7 ml、1.5 mmol) を加えて、加熱還流した。１時間後、反応層を酢酸エチル (50 ml)にあけ、有機層を 10%アンモニウム水 (10 ml)、飽和食塩水 (10 ml ×2)で洗浄、無水硫酸ナトリウムで乾燥後、減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (ヘキサン−酢酸エチル 20:1 →10:1) にて精製し化合物(IIIa)を淡黄色カラメル状物質として得た (80 mg 、0.13 mmol 、収率 87%) 。
【００４６】
1H-NMR (CDCl3) δ 7.95 (1H,s), 7.73 (1H,d,J=19.1Hz), 7.68-7.64 (4H,m), 7.45-7.33 (6H,m), 7.36 (1H,d,J=19.1Hz), 6.34 (1H,dd,J=8.9, 5.3Hz), 5.57 (1H,bd,J=6.3Hz), 4.97 (2H,b), 4.91 (1H,d,J=2.0Hz), 3.92-3.90 (2H,m), 2.88-2.79 (1H,m), 2.56-2.49 (1H,m), 2.12 (3H,s), 1.08 (9H,s), 0.21 (9H,s)
IR (cm-1,neat) : 3300, 1740, 1595, 1570
FABMS (m/z) : 630(M+1)+
【００４７】
例10：9-(5-O-tert-ブチルジフェニルシリル -2-デオキシ -β-D- リボフラノシル)-2-アミノ-6-(2-トリメチルシリルビニル）プリン(IIIb)
化合物(IIIa) (72 mg 、0.11 mmol)のメタノール (1.0 ml) 溶液に炭酸カリウム (110 mg、0.8 mmol) を加え室温にて30分間攪拌した。反応層を塩化メチレン(30 ml) で希釈し飽和塩化アンモニウム水溶液 (10 ml)、飽和食塩水 (10 ml)で洗浄、無水硫酸ナトリウムで乾燥後、減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (クロロホルム−メタノール = 98:2 → 95:5)にて精製し化合物(IIIb)を淡黄色油状物質として得た (67 mg 、0.11 mmol 、収率 100%)。
【００４８】
1H-NMR (CDCl3) δ 7.90 (1H,s), 7.71 (1H,d,J=19.1Hz), 7.68-7.64 (4H,m), 7.45-7.33 (6H,m), 7.34 (1H,d,J=19.1Hz), 4.81 (2H,bs), 4.70 (1H,dt,J=3.0, 2.6Hz), 4.10 (1H,dd,J=7.3, 4.3Hz), 3.87 (1H,dd,J=10.9, 5.3Hz), 3.81 (1H,dd,J=11.2, 4.3Hz), 2.70 (1H,dt,J=13.5, 7.3Hz), 2.44 (1H,ddd,J=13.2, 5.9, 3.3Hz), 1.07 (9H,m), 0.20 (9H,s)
IR (cm-1,neat) : 3500, 1605, 1590
FABMS (m/z) : 588(M+1)+ , 530(M-57)
【００４９】
例11：9-(5-O-tert-ブチルジフェニルシリル -2-デオキシ -3-O-H-ホスホネート -β-D- リボフラノシル)-2-アミノ-6-(2-トリメチルシリルビニル）プリン・トリエチルアンモニウム塩(IIIc)
アルゴン気流下、三塩化リン (4.4 μl 、0.05 mmol)を塩化メチレン (1.5 ml) に溶かし、室温中で N- メチルモルホリン (4.4 μl 、0.05 mmol) 1,2,4- トリアゾールを加え室温で30分間攪拌した。反応液を -78℃に冷却し、化合物(IIIb) (11 mg 、0.03 mmol)の塩化メチレン (0.5 ml) 溶液を滴下し -60℃で10時間攪拌後、TEABバッファー(10 ml) 中に注ぎ分離、水層を塩化メチレン (30 ml)で４回抽出、無水硫酸ナトリウムで乾燥後、減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (クロロホルム−トリエチルアミン = 98:2 →クロロホルム−メタノール−トリエチルアミン = 88:10:2) にて精製し化合物(IIIc)を淡黄色油状物質として得た (14 mg 、0.02 mmol 、収率 73%) 。
【００５０】
1H-NMR (CDCl3) δ 7.91 (1H,s), 7.68 (1H,d,J=19.1Hz), 7.68-7.64 (4H,m), 7.45-7.33 (6H,m), 7.33 (1H,d,J=19.1Hz), 6.37 (1H,dd,J=8.2, 6.3Hz), 5.12-5.10 (1H,m), 4.93 (2H,bs), 4.30 (1H,dd,J=6.3, 3.6Hz), 3.91 (1H,dd,J=11.2, 4.3Hz), 3.82 (1H,dd,J=11.2, 4.0Hz), 3.05-2.73 (2H,m), 3.07 (6H,q,J=7.3Hz), 1.34 (9H,t,J=7.3Hz), 1.03 (9H,s), 0.18 (9H,s)
IR (cm-1,neat) : 3500, 1605, 1590
FABMS (m/z) : 652(M+1)+ , 594(M-57)
【００５１】
例12：本発明の化合物のクロスリンク剤としての機能
本発明のビニル化デオキシグアノシン誘導体のクロスリンク剤としての機能評価を行った。下記のスキームに従い、本発明の化合物(IIa) 及び(IIIa)を用いて、ビニル化デオキシグアノシン誘導体とヒドロキシルアミンとを反応させ、ビニル化デオキシグアノシン誘導体の半減期(t1/2)を求めた。反応条件の詳細は以下のとおりである。その結果、化合物IIa のt1/2は１時間であり、化合物(IIIa)のt1/2は18時間以上であった。
【００５２】
【化８】
例13：化合物(IIa) とヒドロキシルアミン塩酸塩との付加体(10)
化合物(IIa) (30 mg、70μmol)のエタノール (7.0 ml) 溶液にヒドロキシルアミン塩酸塩 (24 mg 、350 μmol)を加え、室温にて攪拌した。24時間後、溶媒を減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (FL-60D、クロロホルム→クロロホルム−メタノール 95:5)にて精製して付加体(10)を黄色油状物質として得た (16 mg 、26μmol 、収率 38%) 。
【００５４】
1H-NMR (CDCl3) δ 7.93 (1H,s), 7.68-7.63 (4H,m), 7.51-7.32 (6H,m), 6.33-6.27 (1H,m), 5.75 (2H,bs), 5.53 (1H,d,J=5.9Hz), 4.18 (1H,t,J=1.7Hz), 3.92-3.85 (3H,m), 3.52 (1H,d,J=16.5Hz), 3.36-3.28 (1H,m), 2.78-2.71 (1H,m), 2.57-2.49 (1H,m), 2.11 (3H,s), 1.52 (3H,d,J=6.6Hz), 1.02 (9H,s),
IR (cm-1,neat) : 3500, 1720, 1600
FABMS (m/z) : 605(M+ +1) 、 572(M+ -33)
例14：化合物(IIIa)とヒドロキシルアミン塩酸塩との付加体(11)
化合物(IIIa) (10 mg 、1.6 μmol)のエタノール (1 ml) 溶液にヒドロキシルアミン塩酸塩 (5.5 mg、8 μmol 、エタノール 0.5 ml 中) を加え、室温にて攪拌した。24時間後、溶媒を減圧下留去した。残渣をシリカゲルカラムクロマトグラフィー (FL-60D、クロロホルム→クロロホルム−メタノール 95:5)にて精製し付加体(11)を無色油状物質(cis体＋trans 体混合物) として得た (4.0 mg、0.68μmol 、収率 44%) 。
【００５６】
1H-NMR (CDCl3) δ 7.92 (1H,s), 7.18 (1H,t,J=4.9Hz), 7.75 (1H,t,J=5.9Hz,minor), 7.68-7.63 (4H,m), 7.51-7.32 (6H,m), 6.31 (1H,dd,J=5.6, 8.9Hz), 5.56 (1H,d,J=5.9Hz), 5.01 (2H,bs), 4.19 (1H,dd,J=3.3, 2.0Hz), 4.15 (2H,d,J=4.9Hz), 3.94 (2H,d,J=6.3Hz,minor), 3.91-3.89 (2H,m), 2.94-2.81 (1H,m), 2.56-2.51 (1H,m), 2.12 (3H,s), 1.07 (9H,s),
IR (cm-1,neat) : 3600-3300, 1605, 1590
FABMS (m/z) : 589(M+ +1) 、 573(M+ -15)
【発明の効果】
本発明の化合物は化学的に安定であり、かつシチジンやグアノシンなどの求核性分子と容易に反応して共有結合を形成する性質を有している。本発明の化合物を製造用中間体として用いると、クロスリンク能を有するアンチセンスオリゴ核酸を製造することができる。[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a vinylated deoxyguanosine derivative or a salt thereof. More specifically, the present invention relates to a vinylated deoxyguanosine derivative or a salt thereof useful for the production of a nucleic acid cross-linking agent or an antisense oligonucleic acid having cross-linking ability.
[0002]
[Prior art]
There is a known method of specifically suppressing antisense gene expression by binding an antisense oligonucleic acid having a complementary nucleotide sequence to a specific gene or nucleic acid such as messenger RNA (m-RNA). Yes. This method is not only used as a biochemical experimental tool but also has a great interest in the fields of biochemistry and medicine / pharmaceuticals because it can be applied to full-scale gene therapy.
[0003]
In order to efficiently perform this method, it is necessary to stabilize a double-stranded or triple-stranded complex formed by complementary bonding, and various means for stabilization have been proposed. A typical example is an antisense in which a functional group having an alkylating ability is introduced into an oligonucleic acid to give it the ability to covalently bind to a target nucleic acid such as gene DNA or mRNA (cross-linking ability). Oligonucleic acids are known (eg, Volssov, VV et al., Nucleic Acid Res., 14, 4065, 1986; Webb, TR et al., J. Am. Chem. Soc., 108, 2764, 1986; Baker, BF et al., J. Am. Chem. Soc., 111, 2700, 1989, etc.).
[0004]
However, many of the antisense oligonucleic acids modified in this way have a problem that the cross-linking speed is low, and those having high reactivity have a defect that the chemical stability is low. It is not desirable to apply such an antisense oligonucleic acid to living bodies (Rokita, S.E. et al., J. Am. Chem. Soc., 116, 1690, 1994).
[0005]
[Problems to be solved by the invention]
An object of the present invention is to provide a compound useful for production in an antisense oligonucleic acid having a property of efficiently cross-linking to a target nucleic acid. Another object of the present invention is to provide a compound as described above, which is chemically stable. Still another object of the present invention is to provide a compound having the above-described characteristics, which is useful for producing an antisense oligonucleic acid applicable to a living body.
[0006]
[Means for Solving the Problems]
As a result of intensive studies to solve the above problems, the present inventors have found that a vinylated guanosine derivative having the following structure is chemically stable and easily reacts with a nucleophilic nucleotide to form a strong crosslink. I found out that it can be formed. Moreover, it discovered that said compound was useful for manufacture of the antisense oligonucleic acid which has the property which crosslinks with a target gene nucleic acid efficiently. The present invention has been completed based on these findings.
[0007]
That is, the present invention provides the following formula (I):
[Chemical 3]
[0008]
DETAILED DESCRIPTION OF THE INVENTION
R in the above formula (I)1, R2And RThreeIs defined by one of the following (1) to (3).
(1) R1 Indicates a hydrogen atom, R2Is a hydrogen atom, tert-butyldiphenylsilyl group, or the following formula:
[Formula 4]
(2) R1Represents a methyl group and R2Represents a tert-butyldiphenylsilyl group and RThreeRepresents an acetyl group; or
(3) R1Represents a trimethylsilyl group, R2Represents a tert-butyldiphenylsilyl group and RThreeRepresents a hydrogen atom, an acetyl group, or -P (O) (OH) H.
[0009]
The above compounds may form acid addition salts or base addition salts, and such salts are also included in the scope of the present invention. When the compound of the present invention itself is applied to a living body as a cross-linking agent, or when used as a production intermediate for the production of an antisense oligonucleic acid applicable to a living body, the salt is physiologically acceptable. Is preferred. Examples of the base addition salt include salts of amines such as triethylamine, dimethylamine, ammonia, triethylamine, or diethylamine; or salts of metals such as sodium, potassium, calcium, or magnesium. Examples of the acid addition salt include salts of mineral acids such as hydrochloric acid, sulfuric acid, or perchloric acid; or oxalic acid, fumaric acid, maleic acid, acetic acid, propionic acid, methanesulfonic acid, or p-toluenesulfonic acid. Mention may be made of salts with organic acids and the like.
[0010]
The compounds of the present invention are specifically shown in Table 1 below. In the table, R2As "Chemical 4", R2Is a group of a thymidylate derivative (the group represented by the above [Chemical Formula 4]), and R in the groupFourIndicates that each is shown in the table. In the table, Ac, Me, Et, TBDPS, TMS, and DMTr are acetyl group, methyl group, ethyl group, tert-butyldiphenylsilyl group, trimethylsilyl group, and di (p-methoxyphenyl) phenylmethyl, respectively. Represents a group (hereinafter the same in this specification).
[0011]
[Table 1]
Although the example of the manufacturing method of this invention compound is demonstrated according to the following scheme, the manufacturing method of the compound of this invention is not limited to these methods. In the scheme, X represents a leaving group such as a chlorine atom, bromine atom, p-toluenesulfonyloxy group, or trifluoromethanesulfonyloxy group, and Met represents a metal such as tributyltin or magnesium halide (the same applies in the following schemes). is there).
[0013]
[Chemical formula 5]
Compound (2):
In a suitable solvent such as a halogenated hydrocarbon solvent such as dichloromethane or chloroform, or an aprotic polar solvent such as dimethylformamide or dimethyl sulfoxide, 2'-deoxyguanosine (1) is present in the presence of a base such as triethylamine or imidazole. The compound (2) can be produced by reacting with 0.2 to 10 equivalents of tert-butyldiphenylsilane chloride. Usually, the reaction may be performed at a temperature of -20 ° C to + 150 ° C for 5 minutes to 24 hours.
[0015]
Compound (3):
Compound (2) is added in an amount of 0.2 to 100 in an aromatic hydrocarbon solvent such as benzene or toluene, a halogenated hydrocarbon solvent such as dichloromethane or chloroform, an ether solvent such as tetrahydrofuran or diethyl ether, or a mixed solvent thereof. By reacting with an acetylating agent such as acetic anhydride or acetyl chloride in the presence of a base such as pyridine or triethylamine, or by using the acetylating agent in combination with a solvent, and optionally in the presence of the base, Compound (3) can be produced. Usually, the reaction may be performed at a temperature of 0 to 150 ° C. for 5 minutes to 24 hours.
[0016]
Compound (4):
Aromatic hydrocarbon solvents such as benzene or toluene, halogenated hydrocarbon solvents such as dichloromethane or chloroform, ether solvents such as tetrahydrofuran or diethyl ether, polar solvents such as dimethylformamide, water, or methanol, or a mixture thereof In a solvent, compound (3) is, for example, 0.2 to 100 equivalents of p-toluenesulfonyl chloride, methanesulfonyl chloride, or trifluoromethanesulfonyl chloride, or p-toluenesulfonic acid anhydride, methanesulfonic acid anhydride, or trifluoromethane. By reacting with a sulfonylating agent such as sulfonic acid anhydride in the presence of a base such as pyridine or triethylamine, or by using the sulfonylating agent in combination with a solvent and reacting in the presence of the base as necessary. Compound (4) can be produced. Usually, the reaction may be performed at a temperature of 0 to 150 ° C. for 5 minutes to 24 hours. In the above reaction, 0.01 to 10 equivalents of a reaction accelerator such as dimethylaminopyridine may be present together.
[0017]
In addition, aromatic hydrocarbon solvents such as benzene or toluene, halogenated hydrocarbon solvents such as dichloromethane or chloroform, ether solvents such as tetrahydrofuran or diethyl ether, polar solvents such as dimethylformamide, water, or methanol, or these Or by treating compound (3) with a halogenating agent such as 0.2 to 100 equivalents of phosphorus oxychloride, phosphorus oxybromide, phosphorus pentachloride, or thionyl chloride in a mixed solvent of Compound (4) can be produced. Usually, the reaction may be performed at a temperature of 0 to 150 ° C. for 5 minutes to 24 hours.
[0018]
Compound (Ia):
Aromatic hydrocarbon solvents such as benzene or toluene, halogenated hydrocarbon solvents such as dichloromethane or chloroform, ether solvents such as tetrahydrofuran or dioxane, polar solvents such as dimethylformamide, water, or methanol, or a mixed solvent thereof A compound (Ia) is produced by reacting 0.2 to 100 equivalents of a vinyl metal compound (5) such as vinyltributyltin or vinylmagnesium bromide with 0.004 to 1 equivalent of a catalyst in the presence of a catalyst. be able to. As the catalyst, a palladium catalyst such as tetrakis (triphenylphosphine) palladium or bis (triphenylphosphine) palladium, or a nickel catalyst such as bis (acetylacetonato) nickel chloride can be used. The above reaction may be performed in the presence of a base such as pyridine or triethylamine. Usually, the reaction may be performed at a temperature of 0 to 150 ° C. for 5 minutes to 24 hours.
[0019]
Compound (Ib):
In a polar solvent such as tetrahydrofuran or ether, a polar solvent such as dimethylformamide, water, or methanol, or a mixed solvent thereof, compound (Ia) is converted to a fluorine-containing compound such as tetrabutylammonium fluoride or potassium fluoride 0.2 to Compound (Ib) can be produced by reacting with 100 equivalents. Usually, the reaction may be performed at a temperature of 0 to 100 ° C. for 5 minutes to 24 hours.
[0020]
Compound (Ic):
In a suitable solvent such as acetonitrile, dimethylformamide, or dimethyl sulfoxide, compound (Ib) is mixed with 0.2 to 10 equivalents of β-cyanoethyl phosphoramidite deoxythymine (6) and 0 to 0 in the presence of 0.2 to 100 equivalents of 1-tetrazole. After reacting at 100 ° C. for 5 minutes to 24 hours, compound (Ic) can be produced by reacting the resulting product with a peroxide such as t-butyl hydroperoxide or cumene peroxide. Usually, the reaction may be performed at -20 to 50 ° C for about 5 minutes to 24 hours.
[0021]
Compound (Id):
Aromatic hydrocarbon solvents such as benzene or toluene, halogenated hydrocarbon solvents such as dichloromethane or chloroform, ether solvents such as tetrahydrofuran or diethyl ether, polar solvents such as dimethylformamide, water, or methanol, or a mixture thereof Compound (Id) is treated with 0.2 to 100 equivalents of an organic acid such as trifluoroacetic acid or dichloroacetic acid, or a mineral acid such as hydrochloric acid or sulfuric acid in a solvent or in the absence of a solvent. Can be manufactured. The reaction is generally carried out at a temperature of -20 to 50 ° C for 5 minutes to 24 hours.
[0022]
[Chemical 6]
Compound (IIa):
Compound (IIa) is produced by reacting Compound (4) with Compound (7) according to the reaction conditions for producing Compound (Ia) from Compound (4) and Compound (5) in Scheme 1. (See Scheme 2 above).
[0024]
[Chemical 7]
Compound (IIIa) and Compound (IIIb):
Compound (IIIa) is produced by reacting Compound (4) with Compound (8) according to the reaction conditions for producing Compound (Ia) from Compound (4) and Compound (5) in Scheme 1. (See Scheme 3 above). In a polar solvent such as tetrahydrofuran or ether, a polar solvent such as dimethylformamide, water, or methanol, or a mixed solvent thereof, 0.2 to 100 equivalents of metal water such as sodium hydroxide or potassium hydroxide. Compound (IIIb) can be produced by reacting with an oxide. The reaction may be generally performed at a temperature of 0 to 100 ° C. for 5 minutes to 24 hours.
[0026]
Compound (IIIc):
In an aromatic hydrocarbon solvent such as benzene or toluene, a halogenated hydrocarbon solvent such as dichloromethane or chloroform, an ether solvent such as tetrahydrofuran or diethyl ether, a polar solvent such as dimethylformamide, or a mixed solvent thereof, 0.5 to Compound (IIIc) can be produced by reacting compound (IIIb) with phosphorus compound (9) in the presence of 50 equivalents of a base such as N-methylmorpholine or triethylamine, and then reacting the product with triethylamine. it can. The phosphorus compound (9) can be prepared, for example, by reacting 0.5 to 10 equivalents of phosphorus trichloride with 0.5 to 10 equivalents of 1,2,4-triazole at −20 to 50 ° C. for 5 minutes to 24 hours. . Usually, the above reaction may be performed at a temperature of -100 to 50 ° C for 5 minutes to 48 hours.
[0027]
Since the compound of the present invention can rapidly form an adduct with cytidine or guanosine, the compound of the present invention is complementary to a target nucleic acid such as gene DNA or m-RNA by using it as a production intermediate. An antisense oligonucleic acid that binds to and forms a covalent crosslink can be produced. Accordingly, another aspect of the present invention provides an intermediate for producing an antisense oligonucleic acid, preferably an intermediate for producing an antisense oligonucleic acid having a cross-linking ability, comprising the compound of the present invention. In addition, an antisense oligonucleic acid produced using the above compound as an intermediate for production, preferably an antisense oligonucleic acid having cross-linking ability, is also encompassed in the scope of the present invention. But the use of the said compound of this invention is not limited to these uses.
[0028]
Hereinafter, the present invention will be described more specifically with reference to examples. However, the scope of the present invention is not limited to the following examples. Needless to say, the reagents and reaction conditions described in the above description of the general production method and the following examples can be appropriately modified or altered. In addition, the compound number in an Example respond | corresponds to the compound number in the said scheme.
[0029]
【Example】
Example 1: 5'-O-tert-butyldiphenylsilyl-2'-deoxyguanosine (2)
Under a stream of argon, a suspension of 2'-deoxyguanosine (2.0 g, 7.2 mmol) in anhydrous DMF (30 ml) was mixed with t-butylchlorodiphenylsilane (3.0 ml, 12 mmol) and imidazole (1.5 g, 21 mmol). The mixture was further stirred at room temperature for 5 hours. After 5 hours, water (30 ml) was added to the reaction layer, and the resulting crystals were collected by filtration and washed with ethyl acetate. Recrystallization from methanol gave Compound (2) as colorless powdery crystals (1.5 g, 3.0 mmol, yield 41%). m.p .: 163-166 ° C.
[0030]
1H-NMR (DMSO-d6) δ 10.60 (1H, brs), 7.80 (1H, s), 7.7-7.3 (10H, m), 6.48 (2H, s), 6.14 (1H, t, J = 6.6Hz) , 5.38 (1H, d, J = 4.0Hz), 4.50-4.40 (1H, m), 3.95-3.65 (2H, s), 2.65-2.45 (1H, m), 2.35-2.20 (1H, m), 0.99 (9H, s)
IR (cm-1, neat): 3600-2700, 1680, 1630
FABMS (m / z): 506 (M + 1)+, 528 (M + 23)+
Example 2: 3'-O-acetyl-5'-O-tert-butyldiphenylsilyl-2'-deoxyguanosine (3)
Acetic anhydride (0.8 ml, 8.5 mmol) was added to a solution of compound (2) (870 mg, 1.7 mmol) in anhydrous pyridine (35 ml) under an argon stream, and the mixture was stirred at 85 ° C. After 5 hours, the reaction layer was diluted with ethyl acetate (50 ml), washed with water (20 ml × 2) and saturated brine (20 ml), dried over anhydrous sodium sulfate, and evaporated under reduced pressure. The residue was recrystallized from acetonitrile to obtain compound (3) as colorless powder crystals (651 mg, 1.2 mmol, yield 70%). m.p .: 226-229 ° C.
[0032]
1H-NMR (CDClThree) δ 12.01 (1H, brs), 7.80-7.30 (10H, m), 6.23 (3H, dd, J = 8.7, 5.8Hz), 5.53 (1H, d, J = 5.9Hz), 4.20 (1H, m) , 3.89 (2H, d, J = 3.6Hz), 2.85-2.70 (1H, m), 2.60-2.45 (1H, m), 2.10 (3H, s), 0.99 (9H, s)
IR (cm-1, neat): 3500-2600, 1740, 1680, 1600
FABMS (m / z): 548 (M + 1)+, 570 (M + 23)+
Example 3: 3'-O-acetyl-5'-O-tert-butyldiphenylsilyl-6-O-p-toluenesulfonyl-2'-deoxyguanosine (4)
Under a stream of argon, a solution of compound (3) (625 mg, 1.14 mmol) in anhydrous dichloromethane (15 ml) was cooled to 0 ° C., triethylamine (0.5 ml, 3.59 mmol), p-toluenesulfonyl chloride (1.08 g, 5.65 mmol). ) And dimethylaminopyridine (43.3 mg, 0.355 mmol) were added and stirred. After 20 hours, the reaction layer was diluted with chloroform (15 ml), the organic layer was washed with water (10 ml), and the aqueous layer was back extracted with chloroform (10 ml × 2). The organic layers were combined, dried over anhydrous sodium sulfate, and evaporated under reduced pressure. The residue was purified by silica gel column chromatography (FL-60D, chloroform-ethyl acetate 9: 1 → 4: 1) to obtain compound (4) as a colorless caramel-like substance (800 mg, 1.14 mmol, yield 100%) ).
[0034]
1H-NMR (CDClThree) δ 8.02 (2H, d, J = 8.3Hz), 7.87 (1H, s), 7.70-7.60 (4H, m), 7.50-7.30 (8H, m), 6.26 (1H, dd, J = 8.7, 5.4 Hz), 5.55 (1H, m), 4.91 (2H, s), 4.25-4.15 (1H, m), 3.90 (2H, m), 2.90-2.75 (1H, m), 2.60-2.40 (1H, m) , 2.45 (3H, s), 2.11 (3H, s), 1.06 (9H, s)
IR (cm-1, neat): 3600-3000, 1740, 1620
FABMS (m / z): 702 (M + 1)+, 644 (M-57)+
Example 4: 9- (3-O-acetyl-5-O-tert-butyldiphenylsilyl-2-deoxy-β-D-ribofuranosyl) -2-amino-6-vinylpurine (Ia)
Under a stream of argon, a solution of compound (4) (320 mg, 0.46 mmol) in anhydrous dioxane (5 ml) was added to lithium chloride (40 mg, 0.94 mmol) and tetrakis (triphenylphosphine) palladium (0) (110 mg, 0.095 mmol). ) Was added and stirred at room temperature. After 30 minutes, vinyltri (n-butyl) tin (760 μl, 2.3 mmol) was added and heated to reflux. After 90 minutes, the reaction layer was poured into ethyl acetate-10% aqueous ammonia (30 ml: 3 ml), and the organic layer was washed with 10% aqueous ammonium (3 ml) and saturated brine (3 ml × 2). After drying with sodium, it was distilled off under reduced pressure. The residue was purified by silica gel column chromatography (chloroform-ethyl acetate 4: 1) to obtain compound (Ia) as a pale yellow caramel-like substance (150 mg, 0.27 mmol, yield 59%).
[0036]
1H-NMR (CDClThree) δ 7.93 (1H, s), 7.75-7.30 (10H, m), 7.13 (1H, dd, J = 17.6, 10.7Hz), 6.86 (1H, dd, J = 17.6, 1.9Hz), 6.34 (1H, dd, J = 8.9, 5.6Hz), 5.85 (1H, dd, J = 10.7, 1.9Hz), 5.56 (1H, m), 4.87 (2H, s), 4.20 (1H, m), 3.90 (2H, m ), 2.95-2.75 (1H, m), 2.60-2.50 (1H, m), 2.12 (3H, s), 1.07 (9H, s)
IR (cm-1, neat): 3600-3100, 1740, 1600
FABMS (m / z): 558 (M + 1)+, 500 (M-57)+
Example 5: 9- (3-O-acetyl-2-deoxy-β-D-ribofuranosyl) -2-amino-6-vinylpurine (Ib)
Under a stream of argon, a solution of compound (Ia) (25 mg, 46 μmol) in anhydrous tetrahydrofuran (0.1 ml) was cooled to 0 ° C., and tetrabutylammonium fluoride (100 μl, 100 μmol, 1M solution in THF) was added dropwise. After 1 hour, the reaction layer was purified by silica gel column chromatography (FL-60D, hexane-ethyl acetate 9: 1) to obtain Compound (Ib) as a colorless oil (14 mg, 44 μmol, yield 98%). .
[0038]
1H-NMR (CDClThree) δ 7.77 (1H, s), 7.12 (1H, dd, J = 17.5, 10.9Hz), 6.89 (1H, dd, J = 17.6, 1.8Hz), 6.21 (1H, dd, J = 9.9, 5.6Hz) , 5.89 (1H, dd, J = 10.6, 2.0Hz), 5.53 (1H, d, J = 5.6Hz), 5.07 (2H, brs), 4.25 (1H, m), 3.20 (1H, dd, J = 9.2 , 5.0Hz), 2.38 (1H, dd, J = 14.0, 5.4Hz), 2.13 (3H, s)
IR (cm-1, neat): 3350, 3200, 1730
FABMS (m / z): 320 (M + 1)+, 550 (M-31)+
[0039]
Example 6: 5′-O-di (paramethoxyphenyl) phenylmethylthymidylyl- (3′-5 ′)-9- (3-O-acetyl-2-deoxy-β-D-ribofuranosyl) -2- Amino-6-vinylpurine-3 '-[P- β- (cyanoethyl) ester (Ic)
Under a stream of argon, add 1-tetrazole (6) (73 mg, 90 μmol) to a solution of compound (Ib) (24 mg, 75 μmol) and T-beta amidite (84 mg, 110 μmol) in anhydrous acetonitrile (0.5 ml). The mixture was stirred at room temperature. After stirring for 2 hours, the reaction layer was cooled to 0 ° C., t-butyl hydroperoxide (16 μl, 86 μmol, 5.4 M solution in toluene) was added and stirring was continued. After 2 hours, insolubles were removed by filtration, and the solvent was distilled off under reduced pressure. The residue was purified by silica gel column chromatography (FL-60D, chloroform-methanol 99: 1 → 95: 5) to obtain compound (Ic) as a colorless oily substance (29 mg, 30 μmol, yield 40%).
[0040]
1H-NMR (CDClThree) δ 7.86 (1H, s), 7.53 (0.5H, d, J = 0.99Hz), 7.46 (0.5H, d, J = 13Hz), 7.42-7.17 (10H, m), 7.10 (1H, ddd, J = 17.5, 10.9, 2.6Hz), 6.90-6.77 (4H, m), 6.44-6.33 (1H, m), 6.30-6.23 (1H, m), 5.84 (1H, ddd, J = 10.9, 3.6, 1.98Hz ), 5.47 (1H, dd, J = 15.8, 5.3Hz), 5.35-5.25 (3H, m), 5.22-5.10 (1H, m), 4.60-4.05 (6H, m), 3.79 (6H, s), 3.55-3.27 (3H, m), 2.72-2.24 (5H, m), 2.12 (1.5H, s), 2.11 (1.5H, s), 1.41 (1.5H, d, J = 0.99Hz), 1.38 (1.5 (H, d, J = 0.99Hz)
IR (cm-1, neat): 3480, 3340, 3200, 1740, 1690, 1600
FABMS (m / z): 979 (M + 1)+
[0041]
Example 7: Thymidylyl- (3'-5 ')-9- (3-O-acetyl-2-deoxy-β-D-ribofuranosyl) -2-amino-6-vinylpurine-3'-[P-β- (Cyanoethyl) ester (Id)
Compound (Ic) (2.0 mg, 2.0 μmol) was added to CHCl.Three Trifluoroacetic acid (0.2 ml, 3 μmol) in 0.1 ml was added dropwise. After 5 minutes, ether (ca. 1 ml) was added to the reaction layer, and the precipitated crystals were collected by filtration to obtain Compound (Id) as a colorless powder (0.6 mg, 0.9 μmol, yield 45%). .
[0042]
1H-NMR (CDClThree) δ 8.36 (0.5H, s), 8.32 (0.5H, s), 7.44 (0.5H, s), 7.43 (0.5H, s), 7.12-7.03 (1H, m), 6.35-6.26 (1H, m ), 6.22-6.13 (1H, m), 6.06-5.96 (1H, m), 5.58-5.48 (1H, m), 5.22-5.03 (1H, m), 4.63-4.17 (7H, m), 3.91-3.73 (3H, m), 3.54-3.19 (3H, m), 2.82-2.72 (2H, m), 2.60-2.36 (2H, m), 2.16 (1.5H, s), 2.15 (1.5H, s), 1.91 (1.5H, d, J = 0.99Hz), 1.89 (1.5H, d, J = 0.99Hz)
FABMS (m / z): 677 (M + 1)+
[0043]
Example 8: 9- (3-O-acetyl-5-O-tert-butyldiphenylsilyl-2-deoxy-β-D-ribofuranosyl) -2-amino-6-methylvinylpurine (IIa)
Under a stream of argon, a solution of compound (4) (350 mg, 0.5 mmol) in anhydrous dioxane (5 ml) was added to lithium chloride (42 mg, 1.0 mmol) and tetrakis (triphenylphosphine) palladium (0) (116 mg, 0.1 mmol). ) Was added and stirred at room temperature. After 30 minutes, methylvinyltri (n-butyl) tin (800 μl, 2.5 mmol, cis / trans = 2.5 / 1) was added and heated to reflux. After 6 hours, the reaction layer was poured into ethyl acetate (100 ml), and the organic layer was washed with 10% aqueous ammonium (30 ml) and saturated brine (30 ml × 2), dried over anhydrous sodium sulfate, and then distilled under reduced pressure. Left. The residue was purified by silica gel column chromatography (chloroform-methanol = 99: 1) to obtain compound (IIa) as a pale yellow caramel-like substance (238 mg, 0.42 mmol, yield 84%).
[0044]
1H-NMR (CDClThree) δ 7.90 (1H, s), 7.68-7.63 (4H, m), 7.51-7.32 (6.6H, m), 6.87 (0.4H, dq, J = 11.2, 2.0Hz), 6.83 (0.6H, dq, J = 15.5, 1.6Hz), 6.28 (0.4H, dq, J = 11.5, 7.2Hz), 6.38 (1H, t, J = 6.3Hz), 5.55 (1H, d, J = 4.6Hz), 4.83 (1H , bs), 4.19 (1H, dd, J = 3.3, 2.0Hz), 3.91-3.89 (2H, m), 2.94-2.81 (1H, m), 2.56-2.51 (1H, m), 2.25 (3H, dd , J = 1.7, 7.6Hz), 2.02 (3H, dd, J = 1.7, 6.9Hz), 2.02 (3H, s), 1.06 (9H, s)
IR (cm-1, neat): 3500, 1740, 1600
FABMS (m / z): 572 (M + 1)+, 514 (M-57)+
[0045]
Example 9: 9- (3-O-acetyl-5-O-tert-butyldiphenylsilyl-2-deoxy-β-D-ribofuranosyl) -2-amino-6- (2-trimethylsilylvinyl) purine (IIIa)
Under a stream of argon, lithium chloride (13 mg, 0.3 mmol) and tetrakis (triphenylphosphine) palladium (0) (35 mg, 0.03 mmol) were added to a solution of compound (4) (104 mg, 0.15 mmol) in anhydrous dioxane (2 ml). ) And stirred at room temperature. After 30 minutes, trimethylsilylvinyltri (n-butyl) tin (1.7 ml, 1.5 mmol) was added and heated to reflux. After 1 hour, the reaction layer was poured into ethyl acetate (50 ml), and the organic layer was washed with 10% aqueous ammonium (10 ml) and saturated brine (10 ml × 2), dried over anhydrous sodium sulfate, and distilled under reduced pressure. Left. The residue was purified by silica gel column chromatography (hexane-ethyl acetate 20: 1 → 10: 1) to obtain compound (IIIa) as a pale yellow caramel-like substance (80 mg, 0.13 mmol, yield 87%).
[0046]
1H-NMR (CDClThree) δ 7.95 (1H, s), 7.73 (1H, d, J = 19.1Hz), 7.68-7.64 (4H, m), 7.45-7.33 (6H, m), 7.36 (1H, d, J = 19.1Hz) , 6.34 (1H, dd, J = 8.9, 5.3Hz), 5.57 (1H, bd, J = 6.3Hz), 4.97 (2H, b), 4.91 (1H, d, J = 2.0Hz), 3.92-3.90 ( 2H, m), 2.88-2.79 (1H, m), 2.56-2.49 (1H, m), 2.12 (3H, s), 1.08 (9H, s), 0.21 (9H, s)
IR (cm-1, neat): 3300, 1740, 1595, 1570
FABMS (m / z): 630 (M + 1)+
[0047]
Example 10: 9- (5-O-tert-butyldiphenylsilyl-2-deoxy-β-D-ribofuranosyl) -2-amino-6- (2-trimethylsilylvinyl) purine (IIIb)
To a solution of compound (IIIa) (72 mg, 0.11 mmol) in methanol (1.0 ml) was added potassium carbonate (110 mg, 0.8 mmol), and the mixture was stirred at room temperature for 30 minutes. The reaction layer was diluted with methylene chloride (30 ml), washed with saturated aqueous ammonium chloride solution (10 ml) and saturated brine (10 ml), dried over anhydrous sodium sulfate, and evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform-methanol = 98: 2 → 95: 5) to obtain compound (IIIb) as a pale yellow oily substance (67 mg, 0.11 mmol, yield 100%).
[0048]
1H-NMR (CDClThree) δ 7.90 (1H, s), 7.71 (1H, d, J = 19.1Hz), 7.68-7.64 (4H, m), 7.45-7.33 (6H, m), 7.34 (1H, d, J = 19.1Hz) , 4.81 (2H, bs), 4.70 (1H, dt, J = 3.0, 2.6Hz), 4.10 (1H, dd, J = 7.3, 4.3Hz), 3.87 (1H, dd, J = 10.9, 5.3Hz), 3.81 (1H, dd, J = 11.2, 4.3Hz), 2.70 (1H, dt, J = 13.5, 7.3Hz), 2.44 (1H, ddd, J = 13.2, 5.9, 3.3Hz), 1.07 (9H, m) , 0.20 (9H, s)
IR (cm-1, neat): 3500, 1605, 1590
FABMS (m / z): 588 (M + 1)+, 530 (M-57)
[0049]
Example 11: 9- (5-O-tert-butyldiphenylsilyl-2-deoxy-3-OH-phosphonate-β-D-ribofuranosyl) -2-amino-6- (2-trimethylsilylvinyl) purine triethylammonium salt (IIIc)
Under argon flow, phosphorus trichloride (4.4 μl, 0.05 mmol) was dissolved in methylene chloride (1.5 ml), and N-methylmorpholine (4.4 μl, 0.05 mmol) 1,2,4-triazole was added at room temperature to 30 Stir for minutes. The reaction solution was cooled to -78 ° C, a solution of compound (IIIb) (11 mg, 0.03 mmol) in methylene chloride (0.5 ml) was added dropwise, and the mixture was stirred at -60 ° C for 10 hours, and then poured into TEAB buffer (10 ml). The aqueous layer was separated and extracted four times with methylene chloride (30 ml), dried over anhydrous sodium sulfate, and evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform-triethylamine = 98: 2 → chloroform-methanol-triethylamine = 88: 10: 2) to obtain compound (IIIc) as a pale yellow oil (14 mg, 0.02 mmol, Yield 73%).
[0050]
1H-NMR (CDClThree) δ 7.91 (1H, s), 7.68 (1H, d, J = 19.1Hz), 7.68-7.64 (4H, m), 7.45-7.33 (6H, m), 7.33 (1H, d, J = 19.1Hz) , 6.37 (1H, dd, J = 8.2, 6.3Hz), 5.12-5.10 (1H, m), 4.93 (2H, bs), 4.30 (1H, dd, J = 6.3, 3.6Hz), 3.91 (1H, dd , J = 11.2, 4.3Hz), 3.82 (1H, dd, J = 11.2, 4.0Hz), 3.05-2.73 (2H, m), 3.07 (6H, q, J = 7.3Hz), 1.34 (9H, t, J = 7.3Hz), 1.03 (9H, s), 0.18 (9H, s)
IR (cm-1, neat): 3500, 1605, 1590
FABMS (m / z): 652 (M + 1)+, 594 (M-57)
[0051]
Example 12: Function of a compound of the present invention as a cross-linking agent
The functional evaluation of the vinylated deoxyguanosine derivative of the present invention as a crosslinking agent was performed. According to the following scheme, using the compounds (IIa) and (IIIa) of the present invention, a vinylated deoxyguanosine derivative and hydroxylamine are reacted to give a half-life of the vinylated deoxyguanosine derivative (t1/2) Details of the reaction conditions are as follows. As a result, t of Compound IIa1/2Is 1 hour, t of compound (IIIa)1/2Was over 18 hours.
[0052]
[Chemical 8]
Example 13: Adduct of Compound (IIa) and Hydroxylamine Hydrochloride (10)
Hydroxylamine hydrochloride (24 mg, 350 μmol) was added to a solution of compound (IIa) (30 mg, 70 μmol) in ethanol (7.0 ml), and the mixture was stirred at room temperature. After 24 hours, the solvent was distilled off under reduced pressure. The residue was purified by silica gel column chromatography (FL-60D, chloroform → chloroform-methanol 95: 5) to obtain the adduct (10) as a yellow oily substance (16 mg, 26 μmol, yield 38%).
[0054]
1H-NMR (CDClThree) δ 7.93 (1H, s), 7.68-7.63 (4H, m), 7.51-7.32 (6H, m), 6.33-6.27 (1H, m), 5.75 (2H, bs), 5.53 (1H, d, J = 5.9Hz), 4.18 (1H, t, J = 1.7Hz), 3.92-3.85 (3H, m), 3.52 (1H, d, J = 16.5Hz), 3.36-3.28 (1H, m), 2.78-2.71 (1H, m), 2.57-2.49 (1H, m), 2.11 (3H, s), 1.52 (3H, d, J = 6.6Hz), 1.02 (9H, s),
IR (cm-1, neat): 3500, 1720, 1600
FABMS (m / z): 605 (M++1), 572 (M+-33)
Example 14: Adduct of Compound (IIIa) and Hydroxylamine Hydrochloride (11)
Hydroxylamine hydrochloride (5.5 mg, 8 μmol in 0.5 ml of ethanol) was added to a solution of compound (IIIa) (10 mg, 1.6 μmol) in ethanol (1 ml), and the mixture was stirred at room temperature. After 24 hours, the solvent was distilled off under reduced pressure. The residue was purified by silica gel column chromatography (FL-60D, chloroform → chloroform-methanol 95: 5) to obtain an adduct (11) as a colorless oily substance (cis body + trans body mixture) (4.0 mg, 0.68 μmol, Yield 44%).
[0056]
1H-NMR (CDClThree) δ 7.92 (1H, s), 7.18 (1H, t, J = 4.9Hz), 7.75 (1H, t, J = 5.9Hz, minor), 7.68-7.63 (4H, m), 7.51-7.32 (6H, m), 6.31 (1H, dd, J = 5.6, 8.9Hz), 5.56 (1H, d, J = 5.9Hz), 5.01 (2H, bs), 4.19 (1H, dd, J = 3.3, 2.0Hz), 4.15 (2H, d, J = 4.9Hz), 3.94 (2H, d, J = 6.3Hz, minor), 3.91-3.89 (2H, m), 2.94-2.81 (1H, m), 2.56-2.51 (1H, m), 2.12 (3H, s), 1.07 (9H, s),
IR (cm-1, neat): 3600-3300, 1605, 1590
FABMS (m / z): 589 (M++1), 573 (M+-15)
【The invention's effect】
The compounds of the present invention are chemically stable and have the property of readily reacting with nucleophilic molecules such as cytidine and guanosine to form covalent bonds. When the compound of the present invention is used as an intermediate for production, an antisense oligonucleic acid having cross-linking ability can be produced.
Claims (1)
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