(57) Abstract:The invention concerns new derivatives of 3'-oximino-2',3'-dideoxynucleosides formula:
< / BR>where B is unsubstituted or substituted thymine-1-yl, uracil-1-yl, cytosine-1-yl, adenin-9-yl and guanine-9-yl, and R - C1-C6alkyl, C6-C9aryl or C1-C6acyl. 3 tab., 2 Il. The invention relates to the field of organic chemistry and Virology and applies to new nucleoside analogues containing the carbohydrate components of the 3-oximino-2-deoxyribofuranosyl, 3 Allexinno-2-deoxyribofuranosyl (acyl= acetyl, propionyl, isobutyryl, pivaloyl, benzoyl and other) or a 3-methoxyimino-2-deoxyribofuranosyl possessing antiviral activity of a broad spectrum of activity against the human immunodeficiency viruses (HIV), herpes simplex (HSV) and hepatitis B (VHB), which can find application in medicine.It is known the use of Retrovir (zidovudine, AZT, 3'-azido-2',3'- dideoxythymidine) for treatment of patients with acquired immunodeficiency syndrome .It is known the use of acyclovir (ACG, zovirax) for the treatment of diseases caused by herpes simplex virus .Also known application EP is Rus hepatitis .An object of the invention is the creation of new nucleoside analogues with antiviral activity broad-spectrum, more selective antiviral effect and lack of resistance to these analogues from mutant strains of viruses or cells deficient in respect fosfauriliruetsa enzymes.Compounds according to the invention by the well-known organic chemistry methods, such as oxidation, oxymorphine, acylation, the introduction of protective groups and their removal in accordance with schemes 1 and 2. Formula and numbering of some of the synthesized compounds are given in Fig. 1. Synthesis of 3'-oximino-2'-3'-dideoxynucleosides lead from natural nucleosides containing 2-deoxyribofuranosyl as carbohydrate components. Position 5' 2-deoxyribofuranosyl protect monomethoxypolyethylene, dimethoxytrityl or tributyltinhydride group, then oxidizes the hydroxyl in position 3' in the keto-group (oxidant - pyridinium dichromate or the reagent dess-Martin), eximious in situ (hydroxylamine hydrochloride in pyridine) and remove the protective group at position 5'. The output 30 - 70%. Virological tests show that 3'-oximino-2'-3'-digesic is it human (HIV), virus hepatitis B and herpes simplex virus (HSV). Connections demonstrate antovic activity in cells defective in timedancing, as well as activity against HSV strains defective in timedancing.Example 1. Synthesis of 3'-oximino-2'3'-dideoxynucleosides, 3'-acetoxidans-2'3'-dideoxynucleosides and 3'-methoxyimino-2'3'-dideoxynucleosides containing 5-substituted derivatives of uracil as a nucleic bases (for example, derivatives of thymine).3'-Keto-2'3'-dideoxythymidine 29 was synthesized , oxidation reagent dess-Martin conducted . NMR spectra were obtained on a spectrometer Bruker AC 250 V solution CDCl3or D2O using TMC or acetonitrile, respectively, as an internal standard. In the description of the spectra used the following abbreviations: s - singlet, DD - doublet of doublets, DDD - doublet of doublets of doublets, m = multiplet, Quartet, Fri - pseudotable. The signals of the protective groups were not mentioned in the description of the spectra. Mass spectra were filmed in positive or negative mode, the spectrometer Jeol DX-300 operating system JMA-DA 5000 and use nitrobenzyl alcohol as a matrix.UV spectra were obtained on Uvicon-931 spectrophotome the el 60 (Merck, Art. 15111) using dichloromethane and methanol as eluents. The melting points were measured on the apparatus Reichter (Austria) and were not corrected. Reversed-phase chromatography was carried out on LiChroprep RP-18 (40-63 µm, Merck, Art. 13900). X-ray exposure was performed on the diffractometer CAD-4 (Nonius, the Netherlands). The structures were solved by direct method and refined by least squares method with anisotropic approximation for non-hydrogen atoms. The coordinates of the hydrogen atoms were determined from difference Fourier syntheses and refined using isotropic temperature factors. The final values of R-factors were 4.2% and 3.0 percent for compounds 1E and 2Z, respectively. The crystals of these compounds were obtained from the water.5'-Monoethoxylate-3'-oximino-2', 3'-dideoxythymidine (30E + 30Z). To a saturated solution of hydroxylamine hydrochloride in pyridine (5 ml) was added compound 29 (1.45 g, and 2.83 mmol). After 15 min the reaction mixture was one stripped off in vacuo and to the residue were added dichloromethane (50 ml) and water (50 ml). After extraction the organic layer was dried with anhydrous sodium sulfate, one stripped off and preapared with toluene. After column chromatography on silica gel (gradient of methanol in dichloromethane, 0_2,5 is (H1', 1H), 4,70 m (H4', 1H), 3,61 DD (H5', J5',4'= 3,0 Hz, J5',5"= situated 10.5 Hz, 1H), 3,48 DD (H5", J5",4'= 1,8 Hz, 1H), 3,55 DD (H2, J2",1'=6,8 Hz, J2",2'=-18,7 Hz, 1H), 2,79 DDD (H2', J2',1'=7,7 Hz, J2',4'=1,9 Hz, 1H), 1,36 d (Me-C5, 3H). 30Z 7,76 to (H6, 1H), 6,39 PT (H1', 1H), 4,94 m (H4', 1H), 3,94 DD (H5', J5',4'= 1,7 Hz, J5',5"=-10,2 Hz, 1H), 3,29 DD (H5", J5",4'=1,9 Hz, 1H), 3,23 DD (H2, J2",1'=6,5 Hz, J2",2'=-16,0 Hz, 1H), 3,06 DDD (H2', J2',1'=9,2 Hz, J2',4'=1,9 Hz, 1H), 1,29 d (Me-C5, 3H). m/e (FAB MS<0) 527 (M-H)-.3'-Oximino-2', 3'-dideoxythymidine (1E). Solution mixture of compounds 30E+30Z (193 mg, and 0.37 mmol) in 80% aqueous acetic acid (5 ml) was stirred for 12 h at room temperature, evaporated to dryness and persuasively with toluene. To the residue was added water (5 ml), dichloromethane (5 ml), the aqueous layer was separated and washed with dichloromethane (5 ml), filtered through a wet filter paper and evaporated to dryness. The residue was purified by reversed-phase chromatography (gradient of methanol in water, 0_5%). After freeze drying was obtained 64 mg of compound 1E as a colorless foam (68%).Subsequent crystallization from water allowed to obtain 43 mg of crystalline 1E. T. pl. 117-119oC.1H NMR (D2O): EUR 7.57 (H6, 1H), of 6.31 PT (H1', 1H), with 4.64 m (H4', 1H), 3,89 DD (H5', J5',4'=2,7 Hz, J5',5"=by 12,8 Hz, 1H), 3,82 DD (H5", J5",4'P>13C NMR (20% CD3OD in H2O): 168,2 (C4), 161,6 (C3'), 153,4 (C2), 139,2 (C6), 113,8 (C5), 85,1 (C1'), 81,0 (C4'), 62,4 (C5'), 34,9 (C2'), 12,6 (Me-C5). m/e (FAB MS<0) 254 (M - H)-, (FAB MS>0) 256 (M+H)+. UV:max= 267 nm ( 964000).Elemental analysis: C10H13N3O5H2O.Found, %: C 43,65; H 5,47; IN SHED 15.37;
Calculated, %: C 43,96; H 5,52; IN lower than the 5.37.X-ray analysis: conformational parameters 1E, are presented in table. 1. Three dimensional structure of the E1 shown in Fig. 2.5'-Monoethoxylate-3'-methoxyimino-2', 3'-dideoxythymidine (31E+31Z). The reaction of the compound 29 (1.45 g, 0.88 mmol) with a saturated solution of O-methylhydroxylamine hydrochloride in pyridine (2 ml) followed by purification on silica gel (gradient of methanol in dichloromethane, 0 _ 2%, as described for 30E+30Z allowed to obtain 431 mg of the mixture 31E+31Z as a colourless foam (91%).1H NMR (CDCl3): 31E 7,58 to (H6, 1H), 6,38 DD (H1', 1H), 4.65 m (H4', 1H), 3,97, with (N-OMe, 3H), 3,59 DD (H5', J5',4'=3,1 Hz, J5',5"=-10,4 Hz, 1H), 3.40 in DD (H5", J5",4'=2,0 Hz, 1H), 3,44 DD (H2, J2",1'=6,8 Hz, J2",2'=is 18.5 Hz, 1H), 2,72 DDD (H2', J2',1'=7,6 Hz, J2',4'=2,0 Hz, 1H), 1,37 d (Me-C5, 3H). 31Z 7,74 to (H6, 1H), 6,39 DD (H1', 1H), 4,82 m (H4', 1H), 3,89 with (N-OMe, 3H), 3,84 DD (H5', J5',4'= 1,8 Hz, J5',5"=-10,2 Hz, 1H), 3,23 DD (H5", J5",4'or = 1.5 Hz, 1H), 3,19 DD (H2, J2",1'=6,3 Hz, J2",2'=-16,3 Hz, 1H'-dideoxythymidine (2E+2Z . Solution mixture of compounds 31E+31Z (222 mg, 0.41 mmol) in 80% aqueous acetic acid (5 ml) was stirred for 12 h at room temperature and processed as described for the connection. 1E. The residue was purified by reversed-phase chromatography (gradient of methanol in water, 0_7%). After freeze drying was obtained 79 mg of a mixture of compounds 2E+2Z as a colourless foam (72%). Subsequent crystallization from water allowed to obtain 45 mg of the crystalline 2Z. T. pl. 121-123oC.1H NMR (D2O): 2E 7,55 to (H6, 1H), 6,28 DD (H1', 1H), 4,68 m (H4', 1H), 3,86 with (N-OMe, 3H), 3,90 DD (H5', J5',4'=2,9 Hz, J5',5"=-13,0 Hz, 1H), 3,82 DD (H5", J5",4'=4,3 Hz, 1H), 3,28 DD (H2, J2",1'=7,3 Hz, J2",2'= -19,2 Hz, 1H), 2,92 DDD (H2', J2',1'=6,1 Hz, J2',4'=1,9 Hz, 1H), 1,82 d (Me-C5, 3H). 2Z 7,71 to (H6, 1H), 6,27 DD (H1', 1H), 4.65 m (H4', 1H), 3,82(N-OMe, 3H), 4,08 DD (H5', J5',4'=3,3 Hz, J5',5"=-12,6 Hz, 1H), 3,80 DD (H5", J5",4'=2,3 Hz, 1H), is 3.08 DD (H2, J2",1'=6,5 Hz, J2",2'=-17,4 Hz, 1H), 2,90 DDD (H2', J2',1'=8,2 Hz, J2',4'=1,8 Hz, 1H), 1,84 d (Me-C5, 3H).13WITH NMR (20% CD3OD in H2O): 2E 166,9 (C4), 159,5 (C3'), to 152.0 (C2), 139,0 (C6), 112,1 (C5), and 84.6 (C1'), 79,8 (C4'), 62,3 (C5'), 61,1 (OMe), 33,1 (C2'), 11,9 (Me-C5). 2Z 166,8 (C4), 159,8 (C3'), 152,1 (C2), 137,7 (C6), 112,7 (C5) of 83.4 (C1'), 79,6 (C4'), 62,4 (C5'), 60,8 (OMe), 35,0 (C2'), 12,0 (Me-C5). m/e (FAB MS<0) 268 (M-H)-, (FAB MS>0) 270 (M+H)+. UV: max= 267 nm ( 96500).Elementary analysis:
C11X-ray analysis: conformational parameters 2Z presented in table. 1. Three-dimensional structure 2Z shown in Fig. 2.5'-Monoethoxylate-3'-acetoxidans-2',3'-dideoxythymidine (32E+32Z). To the solution mixture 30E+30Z (460 mg, 0.87 mmol) in pyridine ( 5 ml) under stirring at 0oC was added acetyl chloride (71 μl, 1.0 mmol). The reaction mixture is allowed to warm to room temperature and after 6 h was added saturated aqueous sodium hydrogen carbonate solution (2 ml). The solution was one stripped off, and the residue was treated and purified on silica gel as described for 30E+30Z (gradient of methanol in dichloromethane, 0_2%. Received 327 mg of the mixture 32E+32Z as a colourless foam (66%).1H NMR (CDDl3): 32E 7,60 to (H6, 1H), 6,28 DD (Hl', 1H), 4,82 m (H4', 1H), 3,74 DD (H5', J5',4'=3,0 Hz, J5',5"=-10,7 Hz, 1H), 3,48 DD (H5", J5",4'=2,1 Hz, 1H), 3,62 DD (H2, J2",1'=6,5 Hz, J2",2'= -18,6 Hz, 1H), 3,13 DDD (H2', J2',1'=7,8 Hz, J2',4'=1,8 Hz, 1H), 2,23 with (N-OCOMe, 3H), 1,37 d (Me-C5, 3H). 32Z 7,73 to (H6, 1H), 6.48 in DD (H1', 1H), 4,92 m (H4', 1H), 3,94 DD (H5', 1H), 3,47 DD (H5", J5",4'=1,8 Hz, J5',5"= -10,4 Hz, 1H), 3,39 DD (H2, J2",1'=6,1 Hz, J2",2'=-17,1 Hz, 1H), 3,18 DDD (H2', J2',1'= 9,2 Hz, J2',4'=1,1 Hz, 1H), 1,94 c (N-OCOMe, 3H), 1.34 in (Me-C5, 3H). m/e (FAB MS < 0) 568 (M-H)-.3'-Acetoxidans-2',3'-dideoxythymidine (3E+3Z). Solution mixture of compounds 32E+32Z is relatively, as described for compound 1E. After purification by chromatography on silica gel (gradient of acetone in dichloromethane, 0_50% and freeze drying there was obtained 110 mg of a mixture of compounds 3E+3Z in the form of a colorless hygroscopic foam (62%).1H NMR (D2O): 3E 7,58 to (H6, 1H), 6.30-in PT (H1', 1H), 4,78 m (H4', 1H), 3,98 DD (H5', J5',4'=2,7 Hz, J5',5"=-16,0 Hz, 1H), 3,91 DD (H5", J5",4'= 3,9 Hz, 1H), 3,50 DD (H2, J2",1'=7,4 Hz, J2",2'=-19,6 Hz, 1H), 3,13 DDD (H2', J2',1'=6,2 Hz, J2',4'=1,3 Hz, 1H), 2,16 with (N-OCOMe, 3H), 1,83 d (Me-C5, 3H). 3Z 7,73 to (H6, 1H), 6,34 DD (H1', 1H), 5.00 m (H4', 1H), 3,98 DD (H5', J5',4'=3,2 Hz, J5',5"=-12,9 Hz, 1H), 3,92 DD (H5", J5",4'=2,8 Hz, 1H), 3.25 to DD (H2, J2",1'=6,3 Hz, J2",2'=17, 7C Hz, 1H), 3,10 DDD (H2', J2',1'=8,2 Hz, J2',4'=1,3 Hz, 1H), 1,82 with (N-OCOMe, 3H), 1,83 d (Me-C5, 3H).13C NMR (20% CD3OD in H2O): 3E 172,0 (COCH3), 168,9 (C3'), 162,9 (C4), to 152.0 (C2), 138,3 (C6), to 112.2 (C5), 84,9 (C1'), 80,4 (C4'), 62,1 (C5'), 34,5 (C2'), 19,0 (COCH3), 11,9 (Me-C5). m/e (FAB MS<0) 296 (M-H)-, (FAB MS>0) 298 (M+H)+. UV:max= 267 nm ( 95900).Elemental analysis: C12H15N3O6H2O.Found, %: C 45,73; H 5,25; N 13,20;
Calculated, %: C 45,72; H 5,43; N 13,32.Example 2. Synthesis of 3'-oximino-2',3'-dideoxynucleosides, acetoxidans-2',3'-dideoxynucleosides containing cytosine, adenine and guanine as nucleic onono-2', 3'-dideoxyadenosine (34E+34Z). To a solution of N6-dimethoxytrityl-5'-tributylammonium-2'- deoxyadenosine 33 (447 mg, 0.67 mmol) in dichloromethane (6 ml) at 0oC was added a reagent dess-Martin (530 mg, 1.25 mmol) in dichloromethane (6 ml) and pyridine (0.1 ml). The solution was allowed to warm to room temperature and after 30 min was added saturated aqueous sodium thiosulfate solution (5 ml). The organic layer was separated, saturated solution of hydroxylamine hydrochloride in pyridine (2 ml) was added, and the solution was one stripped off in vacuum and preapared with toluene. To the residue was added saturated aqueous sodium hydrogen carbonate solution (10 ml) and dichloromethane (10 ml). The organic layer after extraction was washed with water (2x10 ml), dried with sodium sulfate and one stripped off to dryness. After purification by chromatography on silica gel (gradient of methanol in dichloromethane containing a trace of triethylamine, 0_2%) was obtained 227 mg of the mixture 34E+34Z as a colourless foam (49,8%).1H NMR (CDCl3): 34E 8,18 with (H8, 1H), 8,05 (H2, 1H), 6.42 per PT (H1', 1H), 4,98 m (H4', 1H), 4,03 DD (H5', J5',4'= 2,3 Hz, J5',5"=-11,4 Hz, 1H) 3,88 DD (H5", J5",4'=4,1 Hz, 1H), 3,54 DD (H2, J2",1'=7,0 Hz, J2",2'=-18,4 Hz, 1H), 3,09 DDD (H2', J2',1'=6,7 Hz, J2',4'=1,9 Hz, 1H). 34Z 8,29 (H8, 1H), 8,05 (H2, 1H), 6,36 DD (H1', 1H), 4,66 m (H4', 1H), 4,22 DD (H5', J5',4H2', J2',1'=a 8.9 Hz, J2',4'=1,6 Hz, 1H. m/e (FAB MS<0) 680 (M-H)-.3'-Oximino-2',3'-dideoxyadenosine (14E+14Z). To a solution of the mixture of compounds 34E+34Z (226 mg, 0.33 mmol) in tetrahydrofuran (5 ml) at 0oC was added 1,1 M solution of tetrabutylammonium fluoride in tetrahydrofuran (0.33 ml). The solution was allowed to warm to room temperature and evaporated to dryness. After purification by chromatography on silica gel (gradient of methanol in dichloromethane 0_4%, containing traces of triethylamine) and evaporation the residue was dissolved in 20 ml of 80% aqueous acetic acid, stirred 30 min, one stripped off and double-preapared with toluene. To the residue was added water (10 ml) and dichloromethane (10 ml). The aqueous layer was separated and washed with dichloromethane (3x10 ml). After lyophilization of the aqueous layer obtained 64 mg of a mixture 14E+14Z in the form of a colourless foam (73,5%).1H NMR (CDCl3): MAY 14 8,29 (H8, 1H), 8,16 (H2, 1H), 6,50 PT (H1', 1H), 4,70 m (H4', 1H), 3.96 points DD (H5', J5',4'=2,4 Hz, J5',5",=-12,6 Hz, 1H), 3,84 DD (H5", J5",4'=4,0 Hz, 1H), to 3.58 DD (H2, J2",1'=7,1 Hz, J2",2= -18,7 Hz, 1H), 3,48-3.25 m (H2', 1H). 14Z compared to 8.26 (H8, 1H), 8,16 (H2, 1H), 6,44 DD (H1', 1H), 4,70 m (H4', 1H), 4,21 DD (H5', J5',4'=2,8 Hz, J5',5"= -12,6 Hz, 1H), 3,92 DD (H5", J5",4'=1,8 Hz, 1H), 3,48-3.25 m (H2', H2", 2H). m/e (FAB MS<0) 263 (M-H)-, (FAB MS>0) 265 (M+H)+. UV:max=260 nm 15000).3'-AcetoxyoC was added acetyl chloride (18 μl, 0.25 mmol). Then the reaction mixture allowed to warm to room temperature and after 1 h was added saturated aqueous sodium hydrogen carbonate solution (1 ml). The solution was one stripped off in vacuo, and the residue was treated and purified on silica gel as described for 32E+32Z (gradient of methanol in dichloromethane 0_3%) containing traces of triethylamine). To the residue (60 mg, 0,083 mmol) in tetrahydrofuran (2 ml) at 0oC was added 1.1 M solution of tetrabutylammonium fluoride (100 ml) and then was done as described for 14E+14Z. After purification by chromatography on silica gel (gradient of methanol in dichloromethane 0_4%, containing traces of triethylamine), the dry residue was dissolved in 5 ml of 80% aqueous acetic acid and then was done as described for 14E+14Z. After lyophilization received 9 mg of the mixture 15E+15Z in the form of a yellowish foam (32%). 1H NMR (CDCl3): 15E 8,24 (H8, 1H), 8.17 and C (H2, 1H), 6.48 in PT (H1', 1H), 4,66 m (H4', 1H), 3,90 DD (H5', J5',4'=2,4 Hz, J5',5"=-12,2 Hz, 1H), 3,80 DD (H5", J5",4= 4,1 Hz, 1H), 3,67 DD (H2, J2",1'=6,9 Hz, J2",2'= -18,8 Hz, 1H), 3,55 DDD (H2', J2',1'=6,9 Hz, J2',4'=1,7 Hz, 1H), 2,20 (N-OCOMe, 3H). 15Z 8,30 (H8, 1H), 8.17 and C (H2, 1H), 6.42 per DD (H1', 1H), 4,60 m (H4', 1H), 3,98 DD (H5', J5',4'=2,6 Hz, J5',5"=-12,3 Hz, 1H), 4.92 in DD (H5", J5",4'=4,0 Hz, 1H), 3,48 m (H2, 1H), 3.05 m (H2', 1H). m/e (FAB MS<0) 305 (M-H)-, (FAB MS&dicesaro steps in cell cultures.Study of antiviral activity. Antiviral research excluding studies with HIV-1 and HIV-2 was based on inhibition of virus-induced cytopathogenicity in cell cultures E6SM or HEL [6-7]. Appropriate cell culture in Microlitre quantities were inoculated with 100 CCID50of the virus. 1 CCID50virus was sufficient to infect 50% cell culture. After a period of absorption (1 h) residual virus was removed and cell cultures were ingibirovany in the presence of different concentrations (400, 200, 100,... ág/ml) of the compounds under study. Cytopathogenic effect of the virus was assessed by completion cytopathogenic process in the control infected by the virus in cell culture.Inhibition of HIV-induced syncytium formation. Culture of CEM cells were suspended at a concentration of 250,000-300,000 cells/ml of culture medium and infected with 100 CCID50HIV-1 (IIIB) or HIV-2 (ROD). Then 100 μl of the suspension of infected cells were transferred into a tablet of 200 μl containing 100 µl respectively diluted solutions of the investigated compounds. After 4 days incubation at 37oC syncytium formation in cell cultures was suctions activity was expressed, as the concentration of compound that reduces the number of surviving cells by 50% (CC50). Measurement of cytotoxicity was based on microscopically visible alteration of normal cell morphology (E6SM) or the inhibition of normal cell growth (HEL) .Activity against hepatitis B. Human 2.2.15 cells infected UGB, were isolated from cell lines HEP G2 and cultivated . Cells, cultured in a modified environment Dabelko with the addition of 4% bovine serum and 0.5 mm glutamine, and treated with investigational agents within 9 days. The culture fluid was changed every 3 days. Cells HEP G2 and raw 2.2.15 cells served as negative and positive control, respectively. Then the medium was removed and cells lysed. Full of intracellular DNA was isolated and subjected to analysis southern blot, using32P-labeled specific probe (pTHBV plasmid contains the gene UGB full length). Determined the inhibition of viral replicative DNA intermediate in the treated cells compared to control. The study of the cytotoxicity of the compounds was performed in cells HEP G2, located in the tablet, measuring penetration into cells-neutral is adelene antiviral activity.Results.Activity against HIV. Antovic activity of the synthesized compounds was studied using HIV-1 (strain IIIBand HIV-2 (strain ROD) in cultures of human MT-4, CEM/0 and CEM/TK-cells (PL. 1). Compounds 1E, 3E+3Z showed pronounced activity against both HIV-1 and HIV-2 in cultures of cells MT-4 and CEM/0. The activity against HIV-1 and HIV-2 in MT-4 cells was 10-20 times higher than in CEM/0 cells. Cytotoxic activity of the studied compounds was 5-15 times higher for MT-4 cells than for CEM/0 cells.3'-Methoxyimino-2', 3'-dideoxythymidine 2E+2Z were less active. Compounds 1E, 3E+3Z has also been active in cell culture CEM/TK-scarce on timedancing (EC5020 μg/ml and 20 μg/ml), whereas AZT was completely inactive in this cell line.3'-Aryloxypropanolamine-2',3'-dideoxynucleosides (compounds 4a-4e) showed similar values inhibiting concentrations.3'-Oximino-2', 3'-dideoxynucleoside containing natural nucleic acid bases other than thymine (compound 11E+11Z, 14E+14Z, 15E+15Z) also showed antovic activity in CEM cells, although less than 1E and 3E+3Z. Compound 11E+11Z were equally active in the culture of CEM-TK-kletok or not depends on phosphorylation, catalyzed encoded thymidine kinase, and the mechanism of action is different from the conventional mechanism of action inherent in the nucleoside analogue and is associated with subsequent cellular phosphorylation.Activity against HSV. Antiitch activity of the synthesized compounds was studied using four strains of HSV-1 and three strains of HSV-2 in cultures of E6SM and HEL cells (PL. 2). Compound 1E showed pronounced inhibitory properties against several strains of HSV-1 (EC50of 0.4 to 1.3 µg/ml) and slightly lower activity against HSV-2 (EC50from 0.5 to 11.0 µg/ml). Compounds 1E and 3E+3Z were less toxic to cells that have studied Antiherpes virus effect of activity (> 400 μg/ml and 400 μg/ml, respectively) than ACG and 5-(E)-bromovinyl-2'-deoxyuridine (BVDU) (400 μg/ml and 300 μg/ml, respectively) (table. 1).3'-Aryloxypropanolamine-2',3'-dideoxynucleosides (compounds 4a-4e) showed similar values inhibiting concentrations.Compounds 1E and 3E+3Z was inactive against the strain of HSV-1 (TK-) (B2006), deficient timedancing and compounds 2E+2Z for all studied strains of HSV, with the exception of minor activity to strains of HSV-1 (KOS) and HSV-2 (Lyons) in HEL cells with values EC50against UGB was studied in cultured human cell line 2.2.15, infected UGB. Compounds 1E and 4a showed significant activity against hepatitis B EC500.25 microgram/ml and 1.5 µg/ml, respectively, inhibiting viral replicative DNA intermediate in comparison with control and showed no cytotoxicity up to a concentration of 50 μg/ml and 200 μg/ml, respectively (SI>200). 3'-Oximino-2',3'-dideoxynucleoside containing other natural nucleic acid bases (compounds 11E+11Z, 14E+14Z and 17E+17Z) also showed activity against UGB with a few less units EC5010 μg/ml, 10 μg/ml and 7 mg/ml, respectively.Activity against other viruses. The compounds did not show activity and cytotoxicity against several DNA and RNA viruses, namely: coxsackie virus, poliovirus, parainfluenza-3, reovirus-1, sindbis, semiliki forest, cytomegalovirus in different cell cultures (E6SM, HeLa, Vero, HEL). Compounds 1E, 3E+3Z, and some compounds of structure 4 showed low activity against varicella zoster virus with values EC5020-50 mcg/ml.Cytotoxic properties. Data on the study of the cytotoxic activity show that the compounds according to the invention are moderately toxic to cell cultures CEM/0 and MT-4 and low toxicity in relation to E6SM, HEL and cell culture HEP G2.the viruses HIV-1 and HIV-2, hepatitis B, HSV-1 and HSV-2, comparable with the activity of the compounds used in medicine for the treatment of diseases caused by these viruses (so far there are no commercial treatment for VGB);
compound 1E is the first example of a nucleoside analogue, showing activity against three different classes of viruses [9-(2-phosphonylmethoxyethyl)adenine (PMEA) active against HIV and HSV, but inactive against hepatitis B;3TC active against HIV and hepatitis B, but is inactive against HSV];
one of the mechanisms of emergence of resistance to activits nucleoside analogues and, in particular, to AZT is the loss of ability to undergo phosphorylation of human-encoded thymidine kinase in cells, which prevents their further transformation into active nucleoside 5'-triphosphate. Partial preservation of antovic activity in CEM/TK-cells terminowymi oximino-nucleosides 1E and 3E+3Z and the almost complete conservation activity casinogame nucleosides 11E+11Z indirectly testifies to the differences in their mechanisms of action in comparison with the structural analogs, which can help to avoid problems in cellular resistance to chemotherapy 3'-Oh is currently nucleoside analogues, is relevant to study the possibility of chemotherapy other nucleoside analogues nature;
the most active compounds according to the invention - 1E, its acetyl derivative 3E+3Z are synthetically easily accessible connections. So, 1E is synthesized from thymidine in 3 stages with sub-optimal total yield of 63%. 1. 3'-Oximino-2',3'-dideoxynucleoside General formula I
< / BR>where B is unsubstituted or substituted thymine-I-yl, uracil-I-yl, cytosine-L-yl, adenin-9-yl, guanine-9-yl;
R - C1-C6-alkyl or C1-C6-acyl.
< / BR>where R1radical containing nitroxyl group >NO, and R2=R1or H, which possess antiviral activity against RNA-containing viruses (human immunodeficiency virus and vesicular stomatitis virus) and DNA-containing virus (cytomegalovirus)
FIELD: organic chemistry, biochemistry, medicine, virology.
SUBSTANCE: invention relates to derivatives of 2'=amino-2'-deoxynucleosides of the formula:
wherein R means hydrogen atom (H), alkyl, aminoalkyl; R1 means -(R2NR3) wherein R2 and/or R3 means H, -OH, -NH2, alkyl, benzyl under condition that R doesn't represent H or methyl when R2 and R3 mean H. Compounds elicit an antiviral activity with respect to measles and Marburg viruses exceeding that of ribavirin.
EFFECT: valuable properties of compounds.
4 tbl, 2 dwg, 18 ex
FIELD: medicine, pharmacology, bioorganic chemistry, pharmacy.
SUBSTANCE: invention relates to the effective using amount of β-L-2'-deoxynucleoside of the formula (I) or (II) used in manufacturing a medicinal agent used in treatment of hepatitis B, pharmaceutical compositions containing thereof, and methods for treatment of hepatitis B. Proposed agent shows the enhanced effectiveness in treatment of hepatitis B.
EFFECT: enhanced and valuable medicinal properties of agent.
83 cl, 6 tbl, 11 ex
SUBSTANCE: claimed invention relates to method of gemcitabine hydrochloride purification, which includes enriching gemcitabine hydrochloride with its p-anomer, according to which solution of gemcitabine hydrochloride in water is taken with ratio of water to gemcitabine hydrochloride from 3:1 to 12:1 (wt/vol); solution is processed with activated coal, activated coal being taken in amount from 0.1 to 10 wt % of gemcitabine hydrochloride amount in solution; activated coal is removed from solution with formation of filtered solution; concentration of gemcitabine hydrochloride in filtered solution is increased until ratio of filtered solution to gemcitabine hydrochloride equals from 1:1 to 1:5 (wt/vol), efficient for gemcitabine hydrochloride sedimentation; deposited gemcitabine hydrochloride is isolated; and in case admixture content in deposited gemcitabine hydrochloride is not reduced to required level, stages (a)-(e) are repeated. Claimed invention also relates to method of obtaining gemcitabine hydrochloride using claimed purification method.
EFFECT: creation of efficient method of gemcitabine hydrochloride purification.
5 cl, 1 tbl, 5 dwg, 8 ex
SUBSTANCE: present invention relates to (2'R)-2'-dezoxy-2'-fluoro-2'-C-methylnucleoside (β-D or (β-L) , where X represents O; R1 and R7 independently represent H; R3 represents hydrogen and R4 represents NH2; or its pharmaceutically acceptable salt. The invention also pertains to the method of producing the said compounds, which involves glycosylation of N4-benzoylcytosine with a compound of formula 1-4, where R represents methyl, Pg is chosen from C(O)Ph, CH2Ph or both Pg groups can be included in 1,3-(1,1,3,3-tetraisopropyldisiloxanylidene); with further removal of protection of 3'-OPg and 5'-OPg and N-benzoyl of the obtained product.
EFFECT: invented compounds or their pharmaceutically acceptable salts are used as active ingredients against Flaviviridae family viruses in pharmaceutical compositions and liposomal pharmaceutical compositions.
4 cl, 9 tbl, 5 ex, 4 dwg
SUBSTANCE: invention relates to method of obtaining enriched with β-anomer 2'-desoxy-2',2'-difluorocytidine of formula (I)
, which includes stages: (i) interaction of enriched with α-anomer compound of 1-halogenribofuranose of formula (III) with nucleic base of formula (IV) in solvent obtaining enriched with β-anomer nucleoside of formula (II) , with constant removal of formed in reaction process silylhalogenide of formula R3SiX (V) by distillation using carrier or running inert gas through reaction mixture; and (ii) removal of protective group from enriched with β-anomer nucleoside of formula (II). Invention also relates to method of obtaining hydrate of enriched with β-anomer 2'-desoxy-2',2'-difluorocytidine of formula (I), which at stage (ii) after removal of protective group additionally includes stages of dissolving formula (I) nucleoside in water; heating of obtained solution to temperature from 40 to 60°C; cooling of solution to temperature ranging from 10 to 25°C with or without mixing and without changing pH; and filtering of deposited solid substances.
EFFECT: method improvement.
17 cl, 2 tbl, 7 ex
SUBSTANCE: invention relates to the method of producing 2'-desoxy-β-L-thymidine, which involves reacting 5'-O-trityl- or 5'-O-dimethoxytrityl- substituted 2,2' -anhydro-1 -β-L- arabinofuranosylthymine with a reducing agent RedAl and a complexing agent 15-crown-5-ether in a polar solvent 1,2-dimethoxyethane (DME) or tetrahydrofuran, obtaining 5'-O-trityl- or 5'-O-dimethoxytrityl- substituted 2,2'-desoxy-β-L-thymidine, subjected to protection removal if necessary. The invention also relates to the method of producing 2'-desoxy-β-L-thymidine, which involves reacting L-arabinose with cyanamide with subsequent reaction of the intermediate product - L-arabinofuranosylaminooxazoline - with a cycling or condensing agent, forming 2,2' -anhydro-1-β-L-arabinofuranosylthymine; reaction of the latter with a reducing agent RedAl and a complexing agent 15-crown-5-ether in a polar solvent 1,2-dimethoxyethane (DME) or tetrahydrofuran, obtaining 2'-desoxy-β-L-thymidine, where L-arabinofuranosylaminooxazoline can be protected by trityl or dimethoxytrityl in position 5' before or after reaction with the cycling or condensing agent; and protection removal of optionally protected 2'-desoxy-β-L-thymidine, if this is necessary or desired. Use in the given methods of such a reducing agent as Red-Al, and such a complexing agent as 15-crown -5-ether, causes a reaction of intramolecular protection and production of the required nucleoside product with good output.
EFFECT: compound is of great importance as an antiviral or antineoplastic preparation.
13 cl, 29 dwg, 28 ex
SUBSTANCE: invention relates to a pyrimidine nucleoside compound of general formula (1) , in which one of X and Y is a cyano group and the other is hydrogen; R1 is hydrogen, (R3)(R4)(R5)Si- or a carbonyl group which includes an alkyl monosubstituted with an amino group; R2 is hydrogen or (R6)(R7)(R8)Si-, provided that at least one of R1 and R2 is not hydrogen; or R1 and R2 together form a 6-member cyclic group -Si(R9)(R10)-, where each of R9 and R10 is a straight or branched alkyl; R3, R4 and R5 denote a straight or branched alkyl optionally substituted alkoxy, or cycloalkyl; R6, R7 and R8 denote a straight or branched alkyl optionally substituted alkoxy, cycloalkyl or phenyl, or to pharmacologically acceptable salts thereof. The invention also relates to a range of specific compounds of formula (1) or to their pharmacologically acceptable salts: 5'-O-triisopropylsilyl-2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; 5'-O-diethylisopropylsilyl-2'-cyano-2,-desoxy-1-β-D-arabinofuranosylcytosine; 5'-O-dimethylthexylsilyl-2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; 5'-O-(dimethyl-n-octylsilyl)-2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; 3'-O-dimethylthexylsilyl-2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; 3'-O-diethylisopropylsilyl -2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; 3'-O-(tert-butyldimethylsily)-2'-cyano-2'-desoxy-1-β-O-arabinofuranosylcytosine; 3'-O-triisopropylsilyl-2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; 3'-O-dimethylthexylsilyl-5'-O-(L-valyl)-2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; 5'-O-(L-valyl)-3'-O-(tert-butyldimethylsilyl)-2'-cyano-2'-desoxy-1-β-D-arabinofuranosylcytosine; and 3'-O-cyclopropyl-diisopropylsilyl-2'-cyano-2'-desoxy-1-β-D- arabinofuranosylcytosine.
EFFECT: obtaining formula (1) compounds or their pharmacologically acceptable salts for preparing a medicinal agent for treating tumours.
9 cl, 20 tbl, 1 dwg, 73 ex
FIELD: medicine, pharmaceutics.
SUBSTANCE: present invention refers to compounds of formula (I) where R1 is chosen from ethyl, n-propyl, isopropyl or isobutyl, and to its pharmaceutically acceptable salts. Besides, the invention refers to a pharmaceutical composition on the basis of said compounds used for treating a hepatitis C virus (HCV) mediated disease, and also to a method of treating the hepatitis C virus (HCV) mediated disease, and to the method of selective O-acylation nucleoside II for producing O-acyl nucleoside I in an alkaline reaction medium including the stages: (i) dissolution of II and DMAP in a heterogeneous mixture of water and a solvent and addition of a water base for pH control between approximately 7.5 to approximately 12; (ii) optional addition of a sufficient amount of saturated aqueous NaCl for preparing a diphase reaction mixture; (iii) addition of an acidating agent and an accessory base sufficient for pH preservation between approximately 7.5 to approximately 12; (iv) reaction monitoring and interruption of adding said acidating agent and said base after sufficient conversion provided; (v) optional contact of O-acylnucleoside with the pharmaceutically acceptable acid to produce a pharmaceutically acceptable salt.
EFFECT: production of the pharmaceutical composition for treating the hepatitis C virus (HCV) mediated disease.
9 cl, 2 tbl, 8 ex
SUBSTANCE: invention relates to 5'-urethane AZT derivatives of general formula
where X = -NH2, -NHMe, -NHEt, .
EFFECT: compounds have low toxicity, can efficiently inhibit reproduction of the immunodeficiency virus type 1 virus in a CEM SS cell culture.
1 cl, 2 tbl, 7 ex
SUBSTANCE: nucleic base (e.g. uracil, cytosine, adenine, guanine, hypoxanthine, xanthine or similar) reacts with perfluoroalkyl halide in the presence of sulphoxide, peroxide and an iron compound to obtain a perfluoroalkyl-substituted nucleic base.
EFFECT: high cost effectiveness as an intermediate compound for producing medicinal agents.
15 cl, 6 tbl