2,8-dimethyl-1-thia-3,8-diazaspiro[4.5]decane

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a solution of dicyclohexylcarbodiimide (DCC) (1.3 g, 6.3 mmol) in dichloromethane (50 ml) at room temperature was added 3-(1-methyl-indole-3-yl)propanoic acid (1.18 g, 5.8 mmol) and a solution of 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (1.2 g, 6.4 mmol) in dichloromethane (50 ml). The resulting solution was stirred at room temperature for 48 h. During the reaction a white solid precipitated. After filtration, the solvent was evaporated and the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 100/10/1) to give the title compound (0.7 g) as a colorless oil. 1H NMR (CDCl3, 300 MHz) δ 7.60 (d, J=7.82 Hz, 1H, CHC arom), 7.35-7.2 (m, 2H, 2CHC arom), 7.23 (dt, J=1.06, 7.45 Hz, 1H, CHCH arom), 6.90 (s, 1H, CHNH arom), 5.54, 5.07 (2q, J=6.16 and J=6.25 Hz, 1H, CHCH3), 4.63, 3.68 (2d, J=12.0 and J=11.5 Hz, 1H, CHHNCO), 3.75, 3.74 (2s, 3H, NCH3), 3.32 (d, J=11.5 Hz, 0.6H, CHHNCO), 3.17-3.05 (m, 2.4H), 2.8-2.4 (m), 2.68 [t, J=7.6 Hz, C(O)CH2], 2.28, 2.26 (2s, 3H, NCH3), 2.3-2.2 (m), 2.1-2.0 (m), 2.0-1.8 (m), 1.7-1.4 (m), 1.48, 1.43 (2d, J=6.18 and J=6.26 Hz, 3H, CH3—CH) ppm; 13C NMR (CDCl3, 300 MHz) δ 170.6 (C), 137.1 (C), 126.8 (C), 126.7 (CH), 121.8 (CH), 119.0 (CH), 118.9 (CH), 113.7 (C), 109.4 (CH), 59.0 (C), 57.5, 57.2 (CH), 54.6, 54.2 (CH2), 53.2, 52.9 (CH2), 46.3, 46.2 (CH3), 38.1, 37.4 (CH2), 36.9, 36.6 (CH2), 36.4, 34.7 (CH2), 32.7 (CH3), 25.5, 23.5 (CH3), 21.0, 20.9 (CH2) ppm. During the reaction a white solid precipitated After filtration the solvent was evaporated the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 100/10/1)

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a solution of dicyclohexylcarbodiimide (DCC) (1.3 g, 6.3 mmol) in dichloromethane (50 ml) at room temperature was added 3-(1-methyl-indole-3-yl)propanoic acid (1.18 g, 5.8 mmol) and a solution of 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (1.2 g, 6.4 mmol) in dichloromethane (50 ml). The resulting solution was stirred at room temperature for 48 h. During the reaction a white solid precipitated. After filtration, the solvent was evaporated and the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 100/10/1) to give the title compound (0.7 g) as a colorless oil. 1H NMR (CDCl3, 300 MHz) δ 7.60 (d, J=7.82 Hz, 1H, CHC arom), 7.35-7.2 (m, 2H, 2CHC arom), 7.23 (dt, J=1.06, 7.45 Hz, 1H, CHCH arom), 6.90 (s, 1H, CHNH arom), 5.54, 5.07 (2q, J=6.16 and J=6.25 Hz, 1H, CHCH3), 4.63, 3.68 (2d, J=12.0 and J=11.5 Hz, 1H, CHHNCO), 3.75, 3.74 (2s, 3H, NCH3), 3.32 (d, J=11.5 Hz, 0.6H, CHHNCO), 3.17-3.05 (m, 2.4H), 2.8-2.4 (m), 2.68 [t, J=7.6 Hz, C(O)CH2], 2.28, 2.26 (2s, 3H, NCH3), 2.3-2.2 (m), 2.1-2.0 (m), 2.0-1.8 (m), 1.7-1.4 (m), 1.48, 1.43 (2d, J=6.18 and J=6.26 Hz, 3H, CH3—CH) ppm; 13C NMR (CDCl3, 300 MHz) δ 170.6 (C), 137.1 (C), 126.8 (C), 126.7 (CH), 121.8 (CH), 119.0 (CH), 118.9 (CH), 113.7 (C), 109.4 (CH), 59.0 (C), 57.5, 57.2 (CH), 54.6, 54.2 (CH2), 53.2, 52.9 (CH2), 46.3, 46.2 (CH3), 38.1, 37.4 (CH2), 36.9, 36.6 (CH2), 36.4, 34.7 (CH2), 32.7 (CH3), 25.5, 23.5 (CH3), 21.0, 20.9 (CH2) ppm. During the reaction a white solid precipitated After filtration the solvent was evaporated the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 100/10/1)

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a stirred solution of 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (Example 1) (0.86 g, 4.62 mmol) in dichloromethane (90 ml) at room temperature was added dicyclohexylcarbodiimide (DCC) (1.28 g, 6.2 mmol) followed by addition of propionic acid (0.41 ml, 5.5 mmol). The resulting solution was stirred at room temperature overnight. During the reaction a white solid precipitated. After filtration the solvent was evaporated and the crude product was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 90/10/1) to give the title compound (0.5 g) as a colorless oil. 1H NMR (CDCl3, 300 MHz) δ 5.51, 5.19 (2q, J=6.15 and J=6.22 Hz, 1H, CHCH3), 4.63, 3.74 (2d, J=11.88 and J=11.57 Hz, 1H, CHHNCO), 3.44, 3.08 (2d, J=11.55 and J=12.09 Hz, 1H, CHHNCO), 2.8-2.6 (m, 2H), 2.5-2.2 (m, 7H, CH2, CH2CH3, NCH3), 2.2-2.0 (m, 1H), 2.0-1.8 (m, 3H) 1.52, 1.48 (2d, J=6.30 and J=6.18 Hz, 3H, CH3—CH), 1.19-1.12 (m, 3H, CH3) ppm; 13C NMR (CDCl3, 300 MHz) δ 171.6 (C), 57.5, 57.2 (CH), 55.5 (C), 54.7, 54.3 (CH2), 53.2, 53.0 (CH2), 46.3 (CH3), 38.4, 37.7 (CH2), 37.0, 36.4 (CH2), 28.8, 26.8 (CH2), 25.6, 23.6 (CH3), 9.6, 9.4 (CH3) ppm. During the reaction a white solid precipitated After filtration the solvent was evaporated the crude product was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 90/10/1)

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A solution of dicyclohexylcarbodiimide (985 mg, 4.77 mmol) in dry and distilled dichloromethane (10 ml) was added to a stirred solution of 3,5-di-tert-butyl-4-hydroxybenzoic acid (1.14 g, 4.55 mmol) in dichloromethane (15 ml) at room temperature under an argon atmosphere. Dicyclohexylurea began to precipitate as a white solid. 1-Hydroxybenzotriazole (645 mg, 4.77 mmol) was added and the resulting solution was stirred at room temperature for 5 min. 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (846 mg, 4.55 mmol) in dichloromethane (5 ml) was then added and the resulting mixture was kept at room temperature overnight. The next day mixture was heated at 30° C. (temperature of water bath) for 5 hrs and then kept at room temperature for another 4 days. The resulting suspension was filtered and the solvent evaporated under reduced pressure. The residue was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 140/10/1) to give two fractions: AF723 (224 mg, 99.16% chemical purity) and a mixture of AF723 with a by-product (943 mg). The mixture of AF723 with by-product was purified by flash chromatography in a COMBI-flash system using linear gradient (silica, CH2Cl2/EtOH/NH4OH 220/10/1 to 140/10/1). After evaporation and drying under vacuum, AF723 (516 mg, 100% chemical purity) was obtained as a white solid. 1H NMR (CDCl3, 500 MHz) δ 7.27 (s, 2H, arom CH), 5.55 (m, 1H, CH—S), 5.44 (s, 1H, OH), 4.10 (m, 1H, CHH—N—CO), 3.40 (m, 1H, CHH—N—CO), 2.60 (m, 2H, CH2—NCH3), 2.28 (m, 1H, CHH—NCH3), 2.25 (s, 3H, NCH3), 2.11 (m, 1H, CHH—NCH3), 1.95-1.66 (m, 4H, two CH2—CS), 1.60 (d, J=6.15 Hz, 3H, CH3—CH), 1.44 (s, 18H, t-butyl) ppm; 13C NMR (CDCl3, 500 MHz) δ 170.78 (C═O), 155.52 (C), 135.93 (two C), 127.39 (C), 124.30 (two CH), 59.56 (C), 58.00 (CH), 56.40 (CH2), 54.23 (CH2), 52.82 (CH2), 45.88 (CH3), 37.68 (CH2), 36.50 (CH2), 34.28 (C), 30.21 (CH3), 23.99 (CH3) ppm; FTIR (HATR) 2943.89, 1619.92, 1388.67 cm−1; GC-MS (EI) 7.31 min m/z: 419 (M+1). distilled dichloromethane (10 ml) to precipitate as a white solid the resulting mixture was kept at room temperature overnight kept at room temperature for another 4 days The resulting suspension was filtered the solvent evaporated under reduced pressure The residue was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 140/10/1) to give two fractions The mixture of AF723 with by-product was purified by flash chromatography in a COMBI-flash system After evaporation drying under vacuum AF723 (516 mg, 100% chemical purity) was obtained as a white solid GC-MS (EI) 7.31 min m/z

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A solution of dicyclohexylcarbodiimide (985 mg, 4.77 mmol) in dry and distilled dichloromethane (10 ml) was added to a stirred solution of 3,5-di-tert-butyl-4-hydroxybenzoic acid (1.14 g, 4.55 mmol) in dichloromethane (15 ml) at room temperature under an argon atmosphere. Dicyclohexylurea began to precipitate as a white solid. 1-Hydroxybenzotriazole (645 mg, 4.77 mmol) was added and the resulting solution was stirred at room temperature for 5 min. 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (846 mg, 4.55 mmol) in dichloromethane (5 ml) was then added and the resulting mixture was kept at room temperature overnight. The next day mixture was heated at 30° C. (temperature of water bath) for 5 hrs and then kept at room temperature for another 4 days. The resulting suspension was filtered and the solvent evaporated under reduced pressure. The residue was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 140/10/1) to give two fractions: AF723 (224 mg, 99.16% chemical purity) and a mixture of AF723 with a by-product (943 mg). The mixture of AF723 with by-product was purified by flash chromatography in a COMBI-flash system using linear gradient (silica, CH2Cl2/EtOH/NH4OH 220/10/1 to 140/10/1). After evaporation and drying under vacuum, AF723 (516 mg, 100% chemical purity) was obtained as a white solid. 1H NMR (CDCl3, 500 MHz) δ 7.27 (s, 2H, arom CH), 5.55 (m, 1H, CH—S), 5.44 (s, 1H, OH), 4.10 (m, 1H, CHH—N—CO), 3.40 (m, 1H, CHH—N—CO), 2.60 (m, 2H, CH2—NCH3), 2.28 (m, 1H, CHH—NCH3), 2.25 (s, 3H, NCH3), 2.11 (m, 1H, CHH—NCH3), 1.95-1.66 (m, 4H, two CH2—CS), 1.60 (d, J=6.15 Hz, 3H, CH3—CH), 1.44 (s, 18H, t-butyl) ppm; 13C NMR (CDCl3, 500 MHz) δ 170.78 (C═O), 155.52 (C), 135.93 (two C), 127.39 (C), 124.30 (two CH), 59.56 (C), 58.00 (CH), 56.40 (CH2), 54.23 (CH2), 52.82 (CH2), 45.88 (CH3), 37.68 (CH2), 36.50 (CH2), 34.28 (C), 30.21 (CH3), 23.99 (CH3) ppm; FTIR (HATR) 2943.89, 1619.92, 1388.67 cm−1; GC-MS (EI) 7.31 min m/z: 419 (M+1). distilled dichloromethane (10 ml) to precipitate as a white solid the resulting mixture was kept at room temperature overnight kept at room temperature for another 4 days The resulting suspension was filtered the solvent evaporated under reduced pressure The residue was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 140/10/1) to give two fractions The mixture of AF723 with by-product was purified by flash chromatography in a COMBI-flash system After evaporation drying under vacuum AF723 (516 mg, 100% chemical purity) was obtained as a white solid GC-MS (EI) 7.31 min m/z

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (982 mg, 5.28 mmol) was dissolved in dry dichloromethane (5 ml) under argon atmosphere. Distilled triethylamine (1.10 ml, 7.92 mmol) was added and the resulting solution was cooled to 0° C. Valproyl chloride (910 mg, 5.60 mmol) was added dropwise with a syringe. The reaction flask was allowed to warm to room temperature with stirring, and white solid started to precipitate. After 4 h of stirring, dichloromethane (100 ml) was added and the resulting solution was washed with water (10 ml). The two phases were separated and the aqueous phase was extracted with dichloromethane (2×50 ml). The combined organic phase was dried with sodium sulfate, filtered and evaporated under reduced pressure. The residue was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 150/10/1) to give AF717 (521 mg, 99.2% chemical purity) as an off-white powder. 1H NMR (CDCl3, 300 MHz) δ 5.53, 5.26 (2q, J=6.12 and J=6.3 Hz, 1H, CHCH3), 4.67, 3.90 (2d, J=12.0 and J=11.35 Hz, 1H, CHHNCO), 3.48, 3.09 (2d, J=11.35 and J=12.0 Hz, 1H, CHHNCO), 2.78-2.59 (m, 2H, CH2N), 2.53-2.47 (m, 1H, CHCO), 2.30, 2.19 (2s, 3H, NCH3), 2.15-2.05 (m, 2H, CH2N), 2.02-1.84 (m, 2H, CH2CS), 1.74-1.61 (m, 4H, CH2CHCO), 1.55, 1.50 (2d, J=6.3 and J=6.12 Hz, 3H, CH3CH), 1.44-1.36 (m, 2H, CH2CS), 1.34-1.21 (m, 4H, CH2CH3), 0.93-0.87 (m, 6H, CH3CH2) ppm; 13C NMR (CDCl3, 300 MHz) δ 174.35, 174.04 (C), 59.13 (C), 57.35, 57.19 (CH), 54.52, 54.06 (CH2), 53.13, 52.77 (CH2), 46.09 (CH3), 43.77, 42.55 (CH), 38.37, 37.58 (CH2), 37.04, 36.20 (CH2), 35.76, 35.35 (CH2), 35.08, 34.77 (CH2), 26.01, 23.01 (CH3), 21.08, 20.98 (CH2), 20.73, 20.56 (CH2), 14.27, 14.22 (CH3) ppm. to warm to room temperature to precipitate After 4 h of stirring dichloromethane (100 ml) was added the resulting solution was washed with water (10 ml) The two phases were separated the aqueous phase was extracted with dichloromethane (2×50 ml) The combined organic phase was dried with sodium sulfate filtered evaporated under reduced pressure The residue was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 150/10/1) to give AF717 (521 mg, 99.2% chemical purity) as an off-white powder

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (573 mg, 3.08 mmol) was dissolved in dry dichloromethane (2 ml) under an argon atmosphere. Distilled triethylamine (644 μl, 4.62 mmol) was added and the resulting solution was cooled to 0° C. A solution of p-fluorobenzenesulfonyl chloride (600 mg, 3.08 mmol) in dry dichloromethane (2 ml) was added dropwise with a syringe. The reaction flask was allowed to warm to room temperature with stirring, and a white solid started to precipitate. After 1 h of stirring, dichloromethane (50 ml) was added and the resulted solution was washed with water (2×10 ml). The organic phase was dried with anhydrous magnesium sulfate, filtered and evaporated to give the crude mixture as an oil. The crude product was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 93/7/1) to provide AF716 (496 mg, 98.8% chemical purity) as an off-white powder. 1H NMR (CDCl3, 300 MHz) δ 7.89-7.85 (m, 2H, two CHCSO2), 7.27-7.19 (m, 2H, CHCF), 5.02 (q, J=6.12 Hz, 1H, CH—CH3), 3.66 (d, J=11.37 Hz, 1H, CHHNS), 3.48 (d, J=11.37 Hz, 1H, CHHNS), 2.72-2.50 (m, 2H, CH2NCH3), 2.25 (s, 3H, NCH3), 2.16-2.07 (m, 2H, CH2NCH3), 1.95-1.88 (m, 2H, CH2CS), 1.55 (d, J=6.12 Hz, 3H, CH3CH), 1.48 (m, 2H, CH2CS) ppm; 13C NMR (CDCl3, 300 MHz) δ 166.92 and 163.54 (C), 130.05 (CH), 129.93 (CH), 129.58 (C), 116.60 (CH), 116.30 (CH), 60.76 (C), 60.24 (CH), 54.05 (CH2), 53.22 (CH2), 46.03 (CH3), 37.16 (CH2), 37.05 (CH2), 25.56 (CH3) ppm. to warm to room temperature to precipitate After 1 h of stirring dichloromethane (50 ml) was added the resulted solution was washed with water (2×10 ml) The organic phase was dried with anhydrous magnesium sulfate filtered evaporated

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (573 mg, 3.08 mmol) was dissolved in dry dichloromethane (2 ml) under an argon atmosphere. Distilled triethylamine (644 μl, 4.62 mmol) was added and the resulting solution was cooled to 0° C. A solution of p-fluorobenzenesulfonyl chloride (600 mg, 3.08 mmol) in dry dichloromethane (2 ml) was added dropwise with a syringe. The reaction flask was allowed to warm to room temperature with stirring, and a white solid started to precipitate. After 1 h of stirring, dichloromethane (50 ml) was added and the resulted solution was washed with water (2×10 ml). The organic phase was dried with anhydrous magnesium sulfate, filtered and evaporated to give the crude mixture as an oil. The crude product was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 93/7/1) to provide AF716 (496 mg, 98.8% chemical purity) as an off-white powder. 1H NMR (CDCl3, 300 MHz) δ 7.89-7.85 (m, 2H, two CHCSO2), 7.27-7.19 (m, 2H, CHCF), 5.02 (q, J=6.12 Hz, 1H, CH—CH3), 3.66 (d, J=11.37 Hz, 1H, CHHNS), 3.48 (d, J=11.37 Hz, 1H, CHHNS), 2.72-2.50 (m, 2H, CH2NCH3), 2.25 (s, 3H, NCH3), 2.16-2.07 (m, 2H, CH2NCH3), 1.95-1.88 (m, 2H, CH2CS), 1.55 (d, J=6.12 Hz, 3H, CH3CH), 1.48 (m, 2H, CH2CS) ppm; 13C NMR (CDCl3, 300 MHz) δ 166.92 and 163.54 (C), 130.05 (CH), 129.93 (CH), 129.58 (C), 116.60 (CH), 116.30 (CH), 60.76 (C), 60.24 (CH), 54.05 (CH2), 53.22 (CH2), 46.03 (CH3), 37.16 (CH2), 37.05 (CH2), 25.56 (CH3) ppm. to warm to room temperature to precipitate After 1 h of stirring dichloromethane (50 ml) was added the resulted solution was washed with water (2×10 ml) The organic phase was dried with anhydrous magnesium sulfate filtered evaporated

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a stirred solution of 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (2.18 g, 11.7 mmol) in dichloromethane (230 ml) at room temperature was added dicyclohexylcarbodiimide (DCC) (3.24 g, 15.7 mmol) followed by addition of 3-indolepropionic acid (2.87 g, 15.2 mmol). The resulting solution was stirred at room temperature overnight. During the reaction a white solid precipitated. After filtration the solvent was evaporated and the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 90/10/1) to give AF710 (2.5 g, 100% chemical purity) as a white solid. 1H NMR (CDCl3, 300 MHz) δ 8.17 (br s, 1H, NH-indole), 7.60 (d, J=7.81 Hz, 1H, CHC arom), 7.35 (d, J=8.08 Hz, 1H, CHC arom), 7.19 (app t, J=7.53 Hz, 1H, CHCH arom), 7.12 (app t, J=7.45 Hz, 1H, CHCH arom), 7.02 (d, J=1.86 Hz, 1H, CHNH arom), 5.52, 5.09 (2q, J=6.15 and J=6.22 Hz, 1H, CHCH3), 4.62, 3.66 (2d, J=11.76 and J=11.5 Hz, 1H, CHHNCO), 3.29, 3.08 (2d, J=11.48 and J=12.0 Hz, 1H, CHHNCO), 3.18-3.11 (m, 2H), 2.72-2.66 (m, 2H), 2.64-2.46 (m, 2H), 2.26, 2.25 (2s, 3H, NCH3), 2.32-2.19, 2.12-2.02 (2m, 2H), 1.87-1.80, 1.68-1.51 (2m, 4H), 1.48, 1.43 (2d, J=6.21 and J=6.19 Hz, 3H, CH3—CH) ppm; 13C NMR (CDCl3, 500 MHz) δ 170.58 (C), 136.39 (C), 127.25 (C), 122.19 (CH), 121.80 (CH), 119.52 (CH), 118.73 (CH), 115.17 (C), 111.31 (CH), 57.48, 57.19 (CH), 55.46 (C), 54.55, 54.12 (CH2), 53.11, 52.86 (CH2), 46.21, 46.15 (CH3), 38.05, 37.32 (CH2), 36.82, 36.31 (CH2), 34.41 (CH2), 25.44, 23.47 (CH3), 21.04, 20.96 (CH2) ppm. During the reaction a white solid precipitated After filtration the solvent was evaporated the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 90/10/1) to give AF710 (2.5 g, 100% chemical purity) as a white solid

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a stirred solution of 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]dec-2-ene (10.4 ml, 60 mmol) in methanol (150 ml) at room temperature, bromocresol green (5 mg) was added and the solution became blue. 4N HCl/MeOH was added to the stirred solution until the color changed to yellow. Sodium cyanoborohydride (3.9 gr, 62 mmol) was then added in one portion and the resulting mixture was stirred at room temperature for 2 h. During this period, each time that the reaction color changed to green, more 4N HCl/MeOH was added to keep the solution color yellow. At the end of this time, the solvent was evaporated under reduced pressure to give blue-green oil. Dichloromethane (100 ml) was added to the residue and the mixture was washed with 2N NaOH (50 ml). The two phases were separated and the aqueous phase was extracted with dichloromethane (100 ml). The organic phases were combined, dried with anhydrous magnesium sulfate, filtered and the solvent was evaporated under reduced pressure. The residue was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 90/10/1) to give the title compound (2.18 g) as an almost colorless oil. 1H NMR (CDCl3, 500 MHz) δ 4.61 (q, J=6.18 Hz, 1H, CHCH3), 3.10 (d, J=12.6 Hz, 1H, CHHNH), 2.74 (d, J=12.6 Hz, 1H, CHHNH), 2.27-2.19 (m, 1H), 2.23 (s, 3H, NCH3), 2.10 (m, 2H), 1.85-1.70 (m, 5H), 1.46 (d, J=6.18 Hz, 3H, CH3CH) ppm. During this period was added At the end of this time, the solvent was evaporated under reduced pressure to give blue-green oil the mixture was washed with 2N NaOH (50 ml) The two phases were separated the aqueous phase was extracted with dichloromethane (100 ml) dried with anhydrous magnesium sulfate filtered the solvent was evaporated under reduced pressure The residue was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 90/10/1)

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a solution of dicyclohexylcarbodiimide (DCC) (1.3 g, 6.3 mmol) in dichloromethane (50 ml) at room temperature was added 3-(1-methyl-indole-3-yl)propanoic acid (1.18 g, 5.8 mmol) and a solution of 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (1.2 g, 6.4 mmol) in dichloromethane (50 ml). The resulting solution was stirred at room temperature for 48 h. During the reaction a white solid precipitated. After filtration, the solvent was evaporated and the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 100/10/1) to give the title compound (0.7 g) as a colorless oil. 1H NMR (CDCl3, 300 MHz) δ 7.60 (d, J=7.82 Hz, 1H, CHC arom), 7.35-7.2 (m, 2H, 2CHC arom), 7.23 (dt, J=1.06, 7.45 Hz, 1H, CHCH arom), 6.90 (s, 1H, CHNH arom), 5.54, 5.07 (2q, J=6.16 and J=6.25 Hz, 1H, CHCH3), 4.63, 3.68 (2d, J=12.0 and J=11.5 Hz, 1H, CHHNCO), 3.75, 3.74 (2s, 3H, NCH3), 3.32 (d, J=11.5 Hz, 0.6H, CHHNCO), 3.17-3.05 (m, 2.4H), 2.8-2.4 (m), 2.68 [t, J=7.6 Hz, C(O)CH2], 2.28, 2.26 (2s, 3H, NCH3), 2.3-2.2 (m), 2.1-2.0 (m), 2.0-1.8 (m), 1.7-1.4 (m), 1.48, 1.43 (2d, J=6.18 and J=6.26 Hz, 3H, CH3—CH) ppm; 13C NMR (CDCl3, 300 MHz) δ 170.6 (C), 137.1 (C), 126.8 (C), 126.7 (CH), 121.8 (CH), 119.0 (CH), 118.9 (CH), 113.7 (C), 109.4 (CH), 59.0 (C), 57.5, 57.2 (CH), 54.6, 54.2 (CH2), 53.2, 52.9 (CH2), 46.3, 46.2 (CH3), 38.1, 37.4 (CH2), 36.9, 36.6 (CH2), 36.4, 34.7 (CH2), 32.7 (CH3), 25.5, 23.5 (CH3), 21.0, 20.9 (CH2) ppm. During the reaction a white solid precipitated After filtration the solvent was evaporated the crude product was purified by flash chromatography (silica, CH2Cl2/EtOH/NH4OH 100/10/1)

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a stirred solution of 2,8-dimethyl-1-thia-3,8-diaza-spiro[4.5]decane (Example 1) (0.86 g, 4.62 mmol) in dichloromethane (90 ml) at room temperature was added dicyclohexylcarbodiimide (DCC) (1.28 g, 6.2 mmol) followed by addition of propionic acid (0.41 ml, 5.5 mmol). The resulting solution was stirred at room temperature overnight. During the reaction a white solid precipitated. After filtration the solvent was evaporated and the crude product was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 90/10/1) to give the title compound (0.5 g) as a colorless oil. 1H NMR (CDCl3, 300 MHz) δ 5.51, 5.19 (2q, J=6.15 and J=6.22 Hz, 1H, CHCH3), 4.63, 3.74 (2d, J=11.88 and J=11.57 Hz, 1H, CHHNCO), 3.44, 3.08 (2d, J=11.55 and J=12.09 Hz, 1H, CHHNCO), 2.8-2.6 (m, 2H), 2.5-2.2 (m, 7H, CH2, CH2CH3, NCH3), 2.2-2.0 (m, 1H), 2.0-1.8 (m, 3H) 1.52, 1.48 (2d, J=630 and J=6.18 Hz, 3H, CH3—CH), 1.19-1.12 (m, 3H, CH3) ppm; 13C NMR (CDCl3, 300 MHz) δ 171.6 (C), 57.5, 57.2 (CH), 55.5 (C), 54.7, 54.3 (CH2), 53.2, 53.0 (CH2), 46.3 (CH3), 38.4, 37.7 (CH2), 37.0, 36.4 (CH2), 28.8, 26.8 (CH2), 25.6, 23.6 (CH3), 9.6, 9.4 (CH3) ppm. During the reaction a white solid precipitated After filtration the solvent was evaporated the crude product was purified by flash chromatography (silica, CH2Cl2/MeOH/NH4OH 90/10/1)