Patent Application
20080027014
(1) Penta-O-acetyl-5-thio-D-glucopyranose (813 mg) was suspended in ethyl alcohol (20 ml), and thereto was added sodium methoxide (28% methanol solution, 2 drops). The mixture was stirred at room temperature for one hour under argon atmosphere to give a solution of 5-thio-D-glucopyranose. To the solution was added indoline (238 mg), and the resultant mixture was refluxed overnight. Thereto was added acetic acid (2 drops), and the mixture was again refluxed for 7 hours. After being cooled to room temperature, the solvent was evaporated under reduced pressure to give crude 1-(5-thio-β-D-glucopyranosyl)indoline, which was used in the subsequent step without further purification.
(2) The above compound was dissolved in chloroform (20 ml), and thereto were added successively acetic anhydride (1.51 ml), pyridine (1.29 ml) and 4-(dimethylamino)pyridine (24 mg). After being stirred at room temperature for 2.5 days, the organic solvent was evaporated under reduced pressure. The residue was dissolved in ethyl acetate, and the mixture was successively washed with a 1 N aqueous hydrochloric acid solution, water and a saturated aqueous sodium hydrogen carbonate solution. After being dried over magnesium sulfate and treated with activated carbon, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=4:1) and recrystallized from methanol to give 1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indoline (321 mg) as colorless needles. mp 163-165° C. APCI-Mass m/Z 466 (M+H).
(3) The above compound (310 mg) was dissolved in 1,4-dioxane (10 ml), and thereto were added 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (159 mg) and H2O (3 drops). After being stirred at room temperature for 2 hours, thereto was added a saturated aqueous sodium hydrogen carbonate solution (10 ml), and the organic solvent was evaporated under reduced pressure. The residue was extracted with ethyl acetate, and the organic layer was washed with brine. After being dried over magnesium sulfate, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=2:1) to give 1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indole (308 mg) as a colorless solid. APCI-Mass m/Z 481 (M+NH4).
(4) To a stirred solution of the above compound (305 mg) and 4-ethylbenzoyl chloride (166 mg) in dichloromethane (15 ml) was added aluminum chloride (439 mg) at 0° C. After being stirred at 0° C. for 30 minutes and then at room temperature for 5 hours, the mixture was poured into ice-water. Thereto was added a 1 N aqueous hydrochloric acid solution (10 ml), and the resultant mixture was extracted with chloroform twice. The combined organic layer was washed with a saturated aqueous sodium hydrogen carbonate solution and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=3:1-2:1) to give 4-ethylphenyl 1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indol-3-yl ketone (354 mg) as a colorless powder. APCI-Mass m/z 596 (M+H).
(5) To a stirred solution of the above compound (347 mg) in ethyl alcohol (5 ml)-tetrahydrofuran (10 ml) were added cerium(III) chloride heptahydrate (648 mg) and sodium borohydride (66 mg) at 0° C. After being stirred at same temperature for 2 hours, thereto was added a 0.5 N aqueous hydrochloric acid solution (8.5 ml), and the mixture was extracted with ethyl acetate (25 ml) twice. The combined organic layer was washed with a saturated aqueous sodium hydrogen carbonate solution and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure to give crude 4-ethylphenyl 1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indol-3-yl methanol (387 mg), which was used in the subsequent step without further purification.
(6) The above compound was dissolved in acetonitrile (10 ml)-dichloromethane (5 ml), and thereto were added triethylsilane (0.46 ml) and boron trifluoride-diethyl ether complex (0.37 ml) at −10° C. under argon atmosphere. After being stirred at same temperature for one hour, thereto was added a saturated aqueous sodium hydrogen carbonate solution (25 ml), and the organic solvent was evaporated under reduced pressure. The residue was extracted with ethyl acetate (25 ml) twice, and the combined organic layer was dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane ethyl acetate=4:1) to give 3-(4-ethylphenylmethyl)-1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indole (290 mg) as a colorless solid. APCI-Mass m/z 599 (M+NH4).
(7) The above compound (281 mg) was dissolved in methanol (5 ml)-tetrahydrofuran (5 ml), and thereto was added sodium methoxide (28% methanol solution, 2 drops). After being stirred at room temperature for 2 hours under argon atmosphere, the reaction solvent was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:methanol=19:1) to give the titled compound, 3-(4-ethylphenylmethyl)-1-(5-thio-β-D-glucopyranosyl)indole (190 mg) as a colorless powder. APCI-Mass m/z 414 (M+H). 1H-NMR (DMSO-d6) δ 1.14 (t, J=7.5 Hz, 3H), 2.54 (q, J=7.5 Hz, 2H), 3.07 (m, 1H), 3.20-3.45 (m, 2H), 3.57 (m, 1H), 3.79 (m, 1H), 3.91 (m, 1H), 3.96 (s, 2H), 4.69 (t, J=5.5 Hz, 1H), 5.06 (d, J=4.8 Hz, 1H), 5.09 (d, J=4.5 Hz, 1H), 5.15 (d, J=5.5 Hz, 1H), 5.51 (d, J=10.1 Hz, 1H), 6.98 (t, J=7.4 Hz, 1H), 7.10 (d, J=7.9 Hz, 2H), 7.11 (t, J=7.2 Hz, 1H), 7.17 (s, 1H), 7.20 (d, J=8.0 Hz, 2H), 7.42 (d, J=7.9 Hz, 1H), 7.57 (d, J=8.3 Hz, 1H).
(1) Penta-O-acetyl-5-thio-D-glucopyranose (1323 mg) was suspended in ethyl alcohol (30 ml), and thereto was added sodium methoxide (28% methanol solution, 2 drops). The mixture was stirred at room temperature for one hour under argon atmosphere to give a solution of 5-thio-D-glucopyranose. To the solution were added 4-chloroindoline (500 mg) and ammonium chloride (174 mg), and the resultant mixture was refluxed for 22 hours. After being cooled to room temperature, the solvent was evaporated under reduced pressure to give crude 4-chloro-1-(5-thio-β-D-glucopyranosyl)indoline, which was used in the subsequent step without further purification.
(2) The above compound was dissolved in chloroform (20 ml), and thereto were added successively acetic anhydride (2.45 ml), pyridine (2.10 ml) and 4-(dimethylamino)pyridine (40 mg). After being stirred at room temperature overnight, the organic solvent was evaporated under reduced pressure. The residue was dissolved in ethyl acetate, and the mixture was successively washed with a 10% aqueous citric acid solution, water and a saturated aqueous sodium hydrogen carbonate solution. After being dried over magnesium sulfate and treated with activated carbon, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=3:1) to give 4-chloro-1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indoline (1157 mg) as a pale yellow solid. APCI-Mass m/z 500/502 (M+H).
(3) The above compound was treated in a manner similar to Example 1-(3) to give 4-chloro-1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indole as a colorless solid. APCI-Mass m/z 515/517 (M+NH4).
(4) The above compound and 4-ethylbenzoyl chloride were treated in a manner similar to Example 1-(4), (5), (6) and (7) to give the titled compound, 4-chloro-3-(4-ethylphenylmethyl)-1-(5-thio-β-D-glucopyranosyl)indole as a colorless powder. APCI-Mass m/z 448/450 (M+H). 1H-NMR (DMSO-d6) δ 1.15 (t, J=7.5 Hz, 3H), 2.56 (q, J=7.7 Hz, 2H), 3.09 (m, 1H), 3.23 (t, J=8.0 Hz, 1H), 3.41 (m, 1H), 3.57 (m, 1H), 3.79 (d, J=10.0 Hz, 1H), 3.85 (m, 1H), 4.20 (d, J=15.9 Hz, 1H), 4.23 (d, J=15.9 Hz, 1H), 4.71 (m, 1H), 5.05-5.15 (m, 2H), 5.21 (d, J=5.3 Hz, 1H), 5.55 (d, J=10.1 Hz, 1H), 7.01 (d, L=7.5 Hz, 1H), 7.08-7.15 (m, 5H), 7.17 (s, 1H), 7.61 (d, J=8.3 Hz, 1H).
(1) A suspension of 4-chloroindoline (1.00 g) and D-galactose (1.94 g) in H2O (3.0 ml)-ethyl alcohol (20 ml) was refluxed for 29 hours under argon atmosphere. The solvent was evaporated under reduced pressure to give crude 4-chloro-1-(β-D-galactopyranosyl)indoline, which was used in the subsequent step without further purification.
(2) The above compound was suspended in chloroform (20 ml), and thereto were added successively acetic anhydride (4.92 ml), pyridine (4.21 ml) and 4-(dimethylamino)pyridine (80 mg). After being stirred at room temperature for 1.5 hours, the organic solvent was evaporated under reduced pressure. The residue was dissolved in ethyl acetate (70 ml), and the mixture was successively washed with a 10% aqueous copper(II) sulfate solution and a saturated aqueous sodium hydrogen carbonate solution (20 ml). After being dried over magnesium sulfate, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=90:10-60:40) to give 4-chloro-1-(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)indoline (2.34 g) as a pale yellow solid. APCI-Mass m/z 484/486 (M+H).
(3) The above compound was treated in a manner similar to Example 1-(3) to give 4-chloro-1-(2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)indole as a colorless solid. APCI-Mass m/z 499/501 (M+NH4).
(4) The above compound and 4-ethylbenzoyl chloride were treated in a manner similar to Example 1-(4), (5), (6) and (7) to give 4-chloro-3-(4-ethylphenylmethyl)-1-(β-D-galactopyranosyl)indole as a colorless powder. APCI-Mass m/z 432/434 (M+H). 1H-NMR (DMSO-d6) δ 1.15 (t, J=7.5 Hz, 3H), 2.56 (q, J=7.5 Hz, 2H), 3.43-3.55 (m, 3H), 3.67 (t, J=6.0 Hz, 1H), 3.77 (d, J=2.4 Hz, 1H), 3.98 (t, J=9.0 Hz, 1H), 4.23 (s, 2H), 4.64 (t, J=5.5 Hz, 1H), 4.68 (d, J=4.7 Hz, 1H), 4.93 (d, J=5.6 Hz, 1H), 5.04 (d, J=5.8 Hz, 1H), 5.29 (d, J=9.0 Hz, 1H), 7.02 (d, J=7.5 Hz, 1H), 7.06-7.15 (m, 5H), 7.17 (s, 1H), 7.58 (d, J=8.2 Hz, 1H).
(5) The above compound (6.80 g) was suspended in chloroform (300 ml), and thereto were added benzaldehyde dimethyl acetal (3.54 ml) and p-toluenesulfonic acid monohydrate (0.15 g). After being stirred at room temperature for 40 minutes, the solvent was evaporated under reduced pressure, and the residue was dissolved in ethyl acetate (200 ml). The mixture was washed with a saturated aqueous sodium hydrogen carbonate solution (10 ml) and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:methanol=100:0-90:10) to give 4-chloro-3-(4-ethylphenylmethyl)-1-(4,6-O-benzylidene-β-D-galactopyranosyl)indole (7.12 g) as a pale yellow powder. APCI-Mass m/z 520/522 (M+H).
(6) The above compound (7.12 g) was suspended in chloroform (300 ml), and thereto were added pyridine (6.64 ml) and benzoyl chloride (4.77 ml) at 0° C. The mixture was allowed to warm to room temperature, and stirred for 3 days. The reaction mixture was diluted with ethyl acetate (200 ml), and washed successively with a 10% aqueous hydrochloric acid solution (360 ml), a saturated aqueous sodium hydrogen carbonate solution (20 ml) twice and brine (50 ml). After being dried over magnesium sulfate, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure to give crude 4-chloro-3-(4-ethylphenylmethyl)-1-(2,3-di-O-benzoyl-4,6-O-benzylidene-β-D-galactopyranosyl)indole, which was used in the subsequent step without further purification.
(7) A solution of the above compound in acetic acid (240 ml)-H2O (30 ml) was stirred at 70° C. overnight. The solvent was evaporated under reduced pressure, and the residue was dissolved in ethyl acetate (300 ml). The mixture was washed with a saturated aqueous sodium hydrogen carbonate solution (20 ml) twice and brine (30 ml) and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:ethyl acetate=100:0-95:5) to give 4-chloro-3-(4-ethylphenylmethyl)-1-(2,3-di-O-benzoyl-β-D-galactopyranosyl)indole (7.06 g) as a pale yellow solid. APCI-Mass m/z 657/659 (M+NH4).
(8) The above compound (7.06 g) was suspended in 2,4,6-trimethylpyridine (70 ml), and thereto was added benzoyl chloride (1.54 ml) at −40° C. The mixture was allowed to warm to 0° C., and stirred overnight. The reaction mixture was adjusted to pH 5 with a 2 N aqueous hydrochloric acid solution and extracted with ethyl acetate (100 ml)-tetrahydrofuran (100 ml). The mixture was successively washed with water (50 ml) twice, a saturated aqueous sodium hydrogen carbonate solution (20 ml) twice and brine (20 ml) twice and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=90:10-50:50) and recrystallized from ethyl alcohol (100 ml) to give 4-chloro-3-(4-ethylphenylmethyl)-1-(2,3,6-tri-O-benzoyl-β-D-galactopyranosyl)indole (5.17 g) as colorless crystals. mp 192-193° C. APCI-Mass m/z 761/763 (M+NH4).
(9) The above compound (700 mg) was dissolved in dichloromethane (70 ml), and thereto was added (diethylamino)sulfur trifluoride (1.24 ml) at 0° C. under argon atmosphere. After being stirred at room temperature for 24 hours, thereto was added cold water, and the mixture was extracted with chloroform (100 ml) twice. The combined organic layer was washed with brine (30 ml) and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:ethyl acetate=100:0-95:5) and recrystallized from diisopropyl ether (20 ml) to give 4-chloro-3-(4-ethylphenylmethyl)-1-(2,3,6-tri-O-benzoyl-4-fluoro-4-deoxy-β-D-glucopyranosyl)indole (452 mg) as pale yellow crystals. mp 137-138° C. APCI-Mass m/z 746/748 (M+H).
(10) The above compound was treated in a manner similar to Example 1-(7) to give the titled compound, 4-chloro-3-(4-ethylphenylmethyl)-1-(4-fluoro-4-deoxy-β-D-glucopyranosyl)indole as a colorless powder. APCI-Mass m/z 434/436 (M+H). 1H-NMR (DMSO-d6) δ 1.16 (t, J=7.5 Hz, 3H), 2.55 (q, J=7.7 Hz, 2H), 3.50 (ddd, J=12.3, 6.5 and 6.3 Hz, 1H), 3.61 (dd, J=12.1, 5.5 Hz, 1H), 3.70-3.76 (m, 2H), 3.78-3.80 (m, 1H), 4.23 (d, J=3.1 Hz, 2H), 4.32 (dt, J=50.7, 9.2 Hz, 1H), 4.84 (t, J=5.7 Hz, 1H), 5.53 (d, J=5.5 Hz, 1H), 5.56 (d, J=8.5 Hz, 1H), 5.69 (d, J=5.0 Hz, 1H), 7.04 (d, J=7.4 Hz, 1H), 7.09-7.15 (m, 5H), 7.24 (s, 1H), 7.55 (d, J=8.2 Hz, 1H).
4-Chloro-1-(2,3,4,6-tetra-O-acetyl-5-thio-β-D-glucopyranosyl)indole obtained in Example 2-(3) and 4-ethoxybenzoyl chloride were treated in a manner similar to Example 1-(4), (5), (6) and (7) to give the titled compound as a colorless powder. APCI-Mass m/z 464/466 (M+H). 1H-NMR (DMSO-d6) δ 1.30 (t, J=7.0 Hz, 3H), 3.08 (m, 1H), 3.23 (m, 1H), 3.40 (td, J=9.6, 4.8 Hz, 1H), 3.54-3.59 (m, 1H), 3.77-3.88 (m, 2H), 3.97 (q, J=6.9 Hz, 2H), 4.16 and 4.21 (ABq, J=15.8 Hz, 2H), 4.71 (t, J=5.4 Hz, 1H), 5.09 (d, J=4.8 Hz, 1H), 5.11 (d, J=4.5 Hz, 1H), 5.21 (d, J=5.3 Hz, 1H), 5.55 (d, J=9.5 Hz, 1H), 6.82 (d, J=8.5 Hz, 2H), 7.01 (d, J=7.5 Hz, 1H), 7.09-7.15 (m, 2H), 7.11 (d, J=8.7 Hz, 2H), 7.60 (d, J=8.2 Hz, 1H).
(1) 1-(Benzo[b]thiophen-2-ylmethyl)-5-bromo-2-chlorobenzene (1.00 g) was dissolved in tetrahydrofuran (8 ml)-toluene (16 ml), and the mixture was cooled to −78° C. under argon atmosphere. Thereto was added dropwise n-butyl lithium (1.59 M hexane solution, 1.86 ml) over 15 minutes, and the mixture was stirred at the same temperature for 30 minutes. A solution of 2,3,4,6-tetrakis-O-trimethylsilyl-D-glucono-1,5-lactone (see U.S. Pat. No. 6,515,117) (1.26 g) in toluene (10 ml) was added dropwise to the reaction solution over 30 minutes, and the resultant mixture was further stirred at the same temperature for 1.5 hours. Thereto was added a solution of methanesulfonic acid (0.58 ml) in methanol (20 ml), and the mixture was stirred at room temperature overnight. Under ice-cooling, to the mixture was added a saturated aqueous sodium hydrogen carbonate solution (200 ml), and the mixture was extracted with ethyl acetate. The organic layer was washed with brine and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:methanol=10:0-9:1) to give 3-(benzo[b]thiophen-2-ylmethyl)-4-chloro-1-(1-α-methoxy-β-D-glucopyranosyl)benzene (875 mg) as a colorless powder.
(2) A solution of the above compound (1.91 g) and triethylsilane (2.03 ml) in dichloromethane (80 ml) was cooled to −78° C. under argon atmosphere, and thereto was added boron trifluoride-diethyl ether complex (1.61 ml). The mixture was allowed to warm to 0° C. and stirred at the same temperature for 3 hours. To the resultant mixture was added a saturated aqueous sodium hydrogen carbonate solution, and the mixture was extracted with chloroform. The organic layer was washed with brine and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:methanol=10:0-9:1) and crystallization from ethyl acetate-diethyl ether to give 3-(benzo[b]thiophen-2-ylmethyl)-4-chloro-1-(β-D-glucopyranosyl)benzene (1.29 g) as colorless crystals. mp 153-154° C. APCI-Mass m/z 438/440 (M+NH4).
(3) The above compound was treated in a manner similar to Example 3-(5), (6), (7) and (8) to give 3-(benzo[b]thiophen-2-ylmethyl)-4-chloro-1-(2,3,6-tri-O-benzoyl-β-D-glucopyranosyl)benzene as colorless crystals. mp 215-217° C. APCI-Mass m/z 750/752 (M+NH4).
(4) A solution of the above compound (796 mg) and 4-dimethylaminopyridine (780 mg) in dichloromethane (50 ml) was cooled to 0° C. under argon atmosphere, and thereto was added (diethylamino)sulfur trifluoride (0.63 ml). The mixture was allowed to warm to room temperature and stirred at the same temperature for 19 hours. Thereto was added cold water under ice-cooling, and the mixture was extracted with chloroform. The organic layer was washed with brine, dried over magnesium sulfate and treated with activated carbon. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=4:1-2:1) and crystallization from ethyl acetate-hexane to give 3-(benzo[b]thiophen-2-ylmethyl)-4-chloro-1-(2,3,6-tri-O-benzoyl-4-fluoro-4-deoxy-β-D-galactopyranosyl)benzene (305 mg) as colorless crystals. mp 204-207° C. APCI-Mass m/z 752/754 (M+NH4).
(5) The above compound was treated in a manner similar to Example 1-(7) to give the titled compound, 3-(benzo[b]thiophen-2-ylmethyl)-4-chloro-1-(4-fluoro-4-deoxy-β-D-galactopyranosyl)benzene as a colorless powder. APCI-Mass m/z 440/442 (M+NH4). 1H-NMR (DMSO-d6) δ 3.40-3.70 (m, 5H), 4.09 (d, J=9.2 Hz, 1H), 4.34 (d, J=15.9 Hz, 1H), 4.37 (d, J=15.6 Hz, 1H), 4.73 (dd, J=50.5, 2.2 Hz, 1H), 4.86 (t, J=5.6 Hz, 1H), 4.97 (d, J=5.9 Hz, 1H), 5.28 (d, J=5.3 Hz, 1H), 7.14 (s, 1H), 7.24-7.34 (m, 3H), 7.42 (d, J=1.8 Hz, 1H), 7.45 (d, J=8.2 Hz, 1H), 7.73 (d, J=7.7 Hz, 1H), 7.85 (d, J=7.7 Hz, 1H).
(1) 5-bromo-2-chloro-1-(5-phenyl-2-thienylmethyl)benzene was treated in a manner similar to Example 5-(1) and (2) to give 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(β-D-glucopyranosyl)benzene as a colorless powder. APCI-Mass m/z 464/466 (M+NH4).
(2) The above compound (1.00 g) was dissolved in N,N-dimethylformamide (10 ml), and thereto were added trityl chloride (686 mg), 4-dimethylaminopyridine (14 mg) and triethylamine (0.47 ml). After being stirred at room temperature for 6 days, the mixture was diluted with ethyl acetate. The solution was successively washed with a 1 N aqueous hydrochloric acid solution, water twice, a saturated aqueous sodium hydrogen carbonate solution and brine, and the organic layer was dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:methanol=100:1-19:1) to give 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(6-O-trityl-β-D-glucopyranosyl)benzene (870 mg) as a pale yellow powder. ESI-Mass m/z 711/713 (M+Na).
(3) The above compound was treated in a manner similar to Example 1-(2) to give 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(2,3,4-tri-O-acetyl-6-O-trityl-β-D-glucopyranosyl)benzene as a colorless powder. ESI-Mass m/z 837/839 (M+Na).
(4) A solution of the above compound (1.45 g) in formic acid (10 ml) and diethyl ether (5 ml) was stirred at room temperature for 3.5 hours, and then was diluted with water. The mixture was extracted with ethyl acetate, and the organic layer was successively washed with H2O, a saturated aqueous sodium hydrogen carbonate solution and brine. After being dried over magnesium sulfate, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=4:1-1:1) to give 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(2,3,4-tri-O-acetyl-β-D-glucopyranosyl)benzene (689 mg) as colorless crystals. mp 166-168° C. APCI-Mass m/z 590/592 (M+NH4).
(5) The above compound was treated in a manner similar to Example 5-(4) and 1-(7) to give the titled compound, 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(6-fluoro-6-deoxy-β-D-glucopyranosyl)benzene as a colorless powder. APCI-Mass m/z 466/468 (M+NH4). 1H-NMR (DMSO-d6) δ 3.14 (m, 1H), 3.23 (m, 1H), 3.32 (m, 1H), 3.50 (m, 1H), 4.10 (d, J=9.5 Hz, 1H), 4.24 (d, J=15.4 Hz, 1H), 4.28 (d, J=15.7 Hz, 1H), 4.53 (ddd, J=47.8, 10.1 and 5.3 Hz, 1H), 4.58 (dd, J=47.8, 10.3 Hz, 1H), 4.95 (d, J=5.9 Hz, 1H), 5.10 (d, J=5.0 Hz, 1H), 5.27 (d, J=5.5 Hz, 1H), 6.88 (d, J=3.5 Hz, 1H), 7.25 (m, 2H), 7.32 (d, J=3.5 Hz, 1H), 7.35-7.45 (m, 4H), 7.56 (d, J=7.4 Hz, 2H).
(1) 5-Bromo-2-chloro-1-(5-phenyl-2-thienylmethyl)benzene (397 mg) was dissolved in tetrahydrofuran (10 ml), and the mixture was cooled to −78° C. under argon atmosphere. Thereto was added dropwise n-butyl lithium (1.56 M hexane solution, 0.70 ml), and the mixture was stirred at the same temperature for 20 minutes. A solution of 2,3,6-tri-O-benzyl-4-fluoro-4-deoxy-D-glucono-1,5-lactone (496 mg) in tetrahydrofuran (5 ml) was added dropwise to the reaction solution over 5 minutes, and the resultant mixture was further stirred at the same temperature for 1.5 hours. Thereto was added a saturated aqueous ammonium chloride solution (5 ml), and the mixture was allowed to warm to room temperature. The mixture was diluted with H2O and ethyl acetate, and the organic layer was separated, washed with brine and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure to give crude 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(1-hydroxy-2,3,6-tri-O-benzyl-4-fluoro-4-deoxy-D-glucopyranosyl)benzene, which was used in the subsequent step without further purification.
(2) A solution of the above compound in dichloromethane (10 ml) was cooled to −78° C. under argon atmosphere, and thereto were added successively triisopropylsilane (0.68 ml) and boron trifluoride-diethyl ether complex (0.42 ml). After being stirred at −78° C. for 1 hour, the mixture was allowed to warm to 0° C. and stirred at the same temperature for 1 hour. To the resultant mixture was added a saturated aqueous sodium hydrogen carbonate solution, and the mixture was extracted with ethyl acetate. The organic layer was washed with brine and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=19:1-9:1) to give 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(2,3,6-tri-O-benzyl-4-fluoro-4-deoxy-β-D-glucopyranosyl)benzene (226 mg) as a pale yellow caramel. APCI-Mass m/z 736/738 (M+NH4).
(3) To a solution of sodium iodide (550 mg) in acetonitrile (5 ml) was added dropwise chlorotrimethylsilane (0.47 ml) at 0° C. under argon atmosphere, and the mixture was stirred at same temperature for 20 minutes. Thereto was added dropwise a solution of the above compound (220 mg) in acetonitrile (5 ml), and the resultant mixture was stirred at 0° C. for 1 hour and at room temperature for 2.5 hours. The reaction mixture was quenched with a 10% aqueous sodium thiosulfate solution at 0° C., and the mixture was extracted with ethyl acetate. The organic layer was washed successively with H2O, a saturated aqueous sodium hydrogen carbonate solution and brine. After being dried over magnesium sulfate, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (chloroform:methanol=30:1) to give the titled compound, 4-chloro-3-(5-phenyl-2-thienylmethyl)-1-(4-fluoro-4-deoxy-β-D-glucopyranosyl)benzene (123 mg) as a colorless powder. APCI-Mass m/z 466/468 (M+NH4). 1H-NMR (DMSO-d6) δ 3.18 (m, 1H), 3.48-3.67 (m, 4H), 4.14 (d, J=9.5 Hz, 1H), 4.23 (d, J=16.4 Hz, 1H), 4.26 (dt, J=49.3, 9.2 Hz, 1H), 4.27 (d, J=15.9 Hz, 1H), 4.77 (t, J=5.5 Hz, 1H), 5.17 (d, J=5.9 Hz, 1H), 5.49 (d, J=5.5 Hz, 1H), 6.88 (d, J=3.7 Hz, 1H), 7.24-7.29 (m, 2H), 7.32 (d, J=3.7 Hz, 1H), 7.37 (t, J=7.7 Hz, 2H), 7.41-7.45 (m, 2H), 7.57 (d, J=7.4 Hz, 2H).
The chemical structures of the above Examples are shown in Tables 1 and 2 below:
The following compounds shown in Table 3 were prepared in accordance with one of the above Examples.
In the above Tables 1-3, Me is methyl, MeO is methoxy, Et is ethyl, EtO is ethoxy, and “5-thio-β-D-glucopyranosyl”, “4-fluoro-4-deoxy-β-D-glucopyranosyl”, “β-D-galactopyranosyl”, “4-fluoro-4-deoxy-β-D-galactopyranosyl”, and “6-fluoro-6-deoxy-β-D-glucopyranosyl” represent the following chemical formula, respectively:
A solution of 4-chloroindole (3.15 g) and triethylsilane (8.30 ml) in trifluoroacetic acid (32 ml) was stirred at 50° C. for 30 minutes. The solvent was evaporated under reduced pressure, and the residue was basified with a saturated aqueous sodium hydrogen carbonate solution. The mixture was extracted with ethyl acetate twice, and the combined organic layer was dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=100:0-80:20) to give the titled compound (2.89 g) as a colorless oil. APCI-Mass m/z 154/156 (M+H). 1H-NMR (DMSO-d6) δ 2.94 (t, J=8.7 Hz, 2H), 3.46 (t, J=8.7 Hz, 2H), 5.83 (s, 1H), 6.40 (d, J=7.7 Hz, 1H), 6.50 (d, J=8.0 Hz, 1H), 6.90 (t, J=7.9 Hz, 1H).
(1) 5-Bromo-2-chlorobenzoic acid (10.0 g) was suspended in dichloromethane (80 ml), and thereto were added successively N,O-dimethylhydroxylamine hydrochloride (4.56 g), 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (8.96 g), 1-hydroxybenzotriazole (6.31 g) and triethylamine (8.88 ml). After being stirred at room temperature for 64 hours, thereto was added a 10% aqueous hydrochloric acid solution at 0° C., and the mixture was extracted with chloroform. The organic layer was washed with a saturated aqueous sodium hydrogen carbonate solution and brine, and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure to give crude 5-bromo-2-chloro-N-methoxy-N-methylbenzamide, which was used in the subsequent step without further purification.
(2) A solution of the above compound in tetrahydrofuran (200 ml) was cooled to −78° C. under argon atmosphere, and thereto was added dropwise diisobutylaluminum hydride (1.0 M toluene solution, 64 ml). The mixture was stirred at −78° C. for 30 minutes and at 0° C. for 40 minutes. To the resultant mixture was added a 10% aqueous hydrochloric acid solution at 0°, and the mixture was extracted with ethyl acetate. The organic layer was washed with a saturated aqueous sodium hydrogen carbonate solution and brine, and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=19:1) to give 5-bromo-2-chlorobenzaldehyde (8.90 g) as a colorless solid.
(3) To a solution of thianaphthene (5.18 g) in tetrahydrofuran (60 ml) was added dropwise n-butyl lithium (2.44 M hexane solution, 15.8 ml) at −78° C. under argon atmosphere, and the mixture was stirred at −78° C. for 30 minutes and at 0° C. for 30 minutes. The mixture was cooled to −78° C., and thereto was added a solution of the above compound (8.90 g) in tetrahydrofuran (60 ml) in one portion. After being stirred at the same temperature for 1.5 hours, thereto was added a saturated aqueous ammonium chloride solution, and the mixture was extracted with ethyl acetate. The organic layer was washed with brine and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=15:1) to give benzo[b]thiophen-2-yl-(5-bromo-2-chlorophenyl)methanol (12.25 g) as a orange oil.
(4) A solution of the above compound (12.25 g) in dichloromethane (300 ml) was cooled to 0° C., and thereto were added successively triethylsilane (12.17 ml) and boron trifluoride-diethyl ether complex (6.58 ml). After being stirred at the same temperature for 2 hours, thereto was added a saturated aqueous sodium hydrogen carbonate solution, and the mixture was extracted with chloroform. The organic layer was washed with brine and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane only) to give the titled compound, 1-(benzo[b]thiophen-2-ylmethyl)-5-bromo-2-chlorobenzene (9.30 g) as a colorless solid. 1H-NMR (CDCl3) δ 4.30 (s, 2H), 7.02 (d, J=0.9 Hz, 1H), 7.23-7.36 (m, 4H), 7.43 (m, 1H), 7.65-7.69 (m, 1H), 7.73-7.77 (m, 1H).
(1) 5-Bromo-2-chlorobenzoic acid (55.95 g) was suspended in dichloromethane (500 ml), and thereto were added oxalyl chloride (25 ml) and N,N-dimethylformamide (5 drops). After being stirred at room temperature overnight, thereto was added tetrahydrofuran (50 ml), and the mixture was further stirred at the same temperature for 1 hour. The solvent and volatile substance were evaporated under reduced pressure to give 5-bromo-2-chlorobenzoyl chloride. This compound and 2-phenylthiophene (34.63 g) were dissolved in dichloromethane (1000 ml), and thereto was added portionwise aluminum chloride (31.70 g) at 0° C. The mixture was stirred at 0° C. for 30 minutes and at room temperature for 18 hours. The reaction mixture was poured into an ice-water (1500 ml), and the mixture was extracted with chloroform (1000 ml). The organic layer was washed with H2O (1000 ml) twice and a saturated aqueous sodium hydrogen carbonate solution (1000 ml) twice. After being dried over magnesium sulfate and treated with activated carbon, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residual solid was triturated with hexane (1000 ml) and collected by filtration to give 4-bromo-2-chlorophenyl 5-phenyl-2-thienyl ketone (50.22 g) as pale green crystals. mp 132-134° C. APCI-Mass m/z 377/379 (M+H).
(2) To a suspension of the above compound (50.00 g) and triethylsilane (63.44 ml) in dichloromethane (300 ml)-acetonitrile (300 ml) was added dropwise boron trifluoride-diethyl ether complex (50.18 ml) at 0° C. After being stirred at room temperature for 3 hours, thereto was added a saturated aqueous sodium hydrogen carbonate solution (800 ml), and the mixture was extracted with chloroform (300 ml) twice. The combined organic layer was washed with brine (500 ml), dried over magnesium sulfate and treated with activated carbon. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was dissolved in ethyl acetate (300 ml), and the solution was treated with activated carbon. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was crystallized from methanol (150 ml) to give the titled compound,
5-bromo-2-chloro-1-(5-phenyl-2-thienylmethyl)benzene (42.92 g) as pale yellow crystals. mp 80-81° C. 1H-NMR (CDCl3) δ 4.22 (s, 2H), 6.79 (dt, J=3.5, 1.0 Hz, 1H), 7.14 (d, J=3.7 Hz, 1H), 7.21-7.38 (m, 5H), 7.41 (dd, J=2.4, 0.4 Hz, 1H), 7.52-7.57 (m, 2H).
(1) Methyl 2,3,6-tri-O-benzyl-β-D-galactopyranoside (see Sakagami, M.; Hamana, H. Tetrahedron Lett. 2000, 41, 5547-5551.), (16.99 g) was dissolved in dichloromethane (172 ml), and thereto was added (diethylamino)sulfur trifluoride (9.71 ml) at 0° C. The mixture was stirred at 0° C. for 2 hours and at room temperature for 5 hours. The resultant mixture was cooled to 0° C., and thereto was added a saturated aqueous sodium hydrogen carbonate solution. The organic layer was separated and dried over sodium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=100:0-90:10) to give methyl 2,3,6-tri-O-benzyl-4-fluoro-4-deoxy-β-D-glucopyranoside (5.86 g) as a pale yellow oil. APCI-Mass m/z 484 (M+NH4).
(2) A mixture of the above compound (5.39 g) in a 3 M aqueous sulfuric acid solution (10 ml)-acetic acid (50 ml) was stirred at 90° C. for 5 hours. After being cooled to room temperature, thereto was added H2O, and the mixture was extracted with ethyl acetate. The organic layer was washed with successively H2O twice, a saturated aqueous sodium hydrogen carbonate solution and brine. After being dried over magnesium sulfate, the insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=4:1) to give 2,3,6-tri-O-benzyl-4-fluoro-4-deoxy-D-glucose (2.39 g) as a pale yellow oil. APCI-Mass m/z 470 (M+NH4).
(3) The above compound (2.76 g) was dissolved in dimethyl sulfoxide (16 ml), and thereto was added acetic anhydride (11 ml) at 0° C. under argon atmosphere. The mixture was stirred at room temperature overnight, and thereto was added an ice-water at 0° C. The resultant mixture was extracted with ethyl acetate, and the organic layer was washed with a cold water twice and dried over magnesium sulfate. The insoluble materials were filtered off, and the filtrate was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl acetate=19:1-9:1) to give the titled compound, 2,3,6-tri-O-benzyl-4-fluoro-4-deoxy-D-glucono-1,5-lactone (2.08 g) as a colorless solid. APCI-Mass m/z 468 (M+NH4). 1H-NMR (CDCl3) δ 3.74 (ddd, J=11.0, 3.3 and 1.7 Hz, 1H), 3.79 (ddd, J=11.2, 2.6 and 1.7 Hz, 1H), 3.99 (ddd, J=19.8, 6.8 and 6.0 Hz, 1H), 4.15 (d, J=6.8 Hz, 1H), 4.52-4.65 (m, 4H), 4.70 (s, 2H), 4.92 (ddd, J=49.6, 7.6 and 6.0 Hz, 1H), 4.93 (d, J=11.4 Hz, 1H), 7.23-7.38 (m, 15H).
Compounds described in the above examples were used for the SGLT2 inhibition assay.
CHOK1 cells expressing human SGLT2 were seeded in 24-well plates at a density of 400,000 cells/well in F-12 nutrient mixture (Ham's F-12) containing 10% fetal bovine serum, 400 μg/ml Geneticin, 50 units/ml sodium penicillin G (Gibco-BRL) and 50 μg/ml streptomycin sulfate. After 2 days of culture at 37° C. in a humidified atmosphere containing 5% CO2, cells were washed once with the assay buffer (137 mM NaCl, 5 mM KCl, 1 mM CaCl2, 1 mM MgCl2, 50 mM Hepes, and 20 mM Tris, pH 7.4) and incubated with 250 μl of the buffer containing test compounds for 10 min at 37° C. Test compounds were dissolved in DMSO. The final concentration of DMSO was 0.5%. The transport reaction was initiated by addition of 50 μl [14C]-methyl-α-D-glucopyranoside (14C-AMG) solution (final concentration, 0.5 mM). After incubation for 2 hours at 37° C., the uptake was stopped by aspiration of the incubation mixture, the cells were washed three times with ice-cold PBS. Then, cells were solubilized with 0.3 N NaOH and aliquots were taken for determination of radioactivity by a liquid scintillation counter. Nonspecific AMG uptake was defined as that which occurred in the presence of 100 μM of phlorizin, a specific inhibitor of sodium-dependent glucose cotransporter. Specific uptake was normalized for the protein concentrations measured by the method of Bradford. The 50% inhibitory concentration (IC50) values were calculated from dose-response curves by least square method.
Results are shown in the following table:
Compounds described in the above examples were used for the Urinary glucose excretion test in rats.
6-week-old male Sprague-Dawley (SD) rats were housed in individual metabolic cages with free access to food and water from 2 days prior to the experiment. On the morning of the experiment, rats were administered vehicle (0.2% carboxymethyl cellulose solution containing 0.2% Tween80) or test compounds (30 mg/kg) by oral gavage at a volume of 10 ml/kg. Then, urine of the rat was collected for 24 hours, and the urine volume was measured. Subsequently, the glucose concentration in urine was quantified using the enzymatic assay kit and the daily amount of glucose excreted in urine per individual was calculated.
Urinary glucose amounts ranges are depicted by A and B. These ranges are as follows: A≧2400 mg; 2400 mg>B≧2000 mg.
| Number | Date | Country | |
|---|---|---|---|
| 60820759 | Jul 2006 | US |
