methyl (2S,4R)-4-hydroxypyrrolidine-2-carboxylate

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

Procedure: A solution of 2(S)-tert-butoxycarbonylamino-8-nonenoic acid (purchased from RSP Amino Acids)(3.5 g, 12.9 mmol) in 200 mL of DCM was treated sequentially with 4(R)-hydroxypyrrolidine-2(S)-carboxylic acid methyl ester hydrocholoride (2.15 g, 11.8 mmol), N-methyl morpholine (4.25 mL, 38.6 mmol), and HATU (5.37 g, 14.1 mmol). The reaction mixture was stirred at rt under N2 for 3 days, and then concentrated in vacuo. The residue was partitioned between ethyl acetate and pH 4 buffer (biphthalate). The organic phase was washed with sat. aq. NaHCO3, dried (MgSO4), and concentrated in vacuo to give the crude product. Flash chromatography (50% ethyl acetate/hexane to 100% ethyl acetate) gave 4.7 g (˜100%) of 1-(2(S)-tert-Butoxycarbonylamino-non-8-enoyl)-4(R)-hydroxy-pyrrolidine-2(S)-carboxylic acid methyl ester as a colorless oil: 1H NMR (500 MHz, CD3OD) δ 1.33-1.50 (m, 8H), 1.46 (s, 9H), 1.57 (m, 1H), 1.72 (m, 1H) 2.08 (m, 2H), 2.28 (m, 1H), 3.72 (s, 3H,) 3.75-3.87 (m, 2H), 4.36 (m, 1H), 4.51 (bs, 1H), 4.57 (t, J=8.2 Hz, 1H), 4.95 (d, J=10.4 Hz, 1H), 5.01 (m, 1H), 5.83 (m, 1H). LC-MS (Method J, retention time: 3.01 min), MS m/z 399 (M++1). Step 26ii: Preparation of 1-{[1-(2(S)-tert-Butoxycarbonylamino-non-8-enoyl)-4(R)-hydroxy-pyrrolidine-2(S)carbonyl]-(1R)-amino}-2(S)-vinylcyclopropanecarboxylic acid ethyl ester concentrated in vacuo The residue was partitioned between ethyl acetate and pH 4 buffer (biphthalate) The organic phase was washed with sat. aq. NaHCO3 dried (MgSO4) concentrated in vacuo to give the crude product

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

Procedure: To a solution of (3R)-2-(tert-butoxycarbonylamino)-3-ethyl-5-methylnon-8-enoic acid (2.0 g, 6.4 mmole) in dichloromethane (20 mL) was added DIPEA (1.93 mL, 19.2 mmole) and HATU (2.42 g, 6.38 mmole) followed by (2S,4R)-methyl 4-hydroxypyrrolidine-2-carboxylate (1.15 g, 6.38 mmole) at room temperature. The reaction mixture was stirred at room temperature for 2 h. The reaction mass was diluted with DCM and washed with water. The organic layer was dried over anhydrous Na2SO4; filtered; then concentrated under reduced pressure to get crude compound as mixture of diastereomers. The material was subjected to SFC purification to afford (2S,4R)-methyl 1-((2S,3R)-2-(tert-butoxycarbonylamino)-3-ethyl-5-methylnon-8-enoyl)-4-hydroxypyrrolidine-2-carboxylate (1.5 g, 53%). MS: MS m/z 441.6 (M++1). washed with water The organic layer was dried over anhydrous Na2SO4 filtered then concentrated under reduced pressure to get crude compound as mixture of diastereomers The material was subjected to SFC purification

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

Procedure: To a solution of (3R)-2-(tert-butoxycarbonylamino)-3-ethyl-5-methylnon-8-enoic acid (2.0 g, 6.4 mmole) in dichloromethane (20 mL) was added DIPEA (1.93 mL, 19.2 mmole) and HATU (2.42 g, 6.38 mmole) followed by (2S,4R)-methyl 4-hydroxypyrrolidine-2-carboxylate (1.15 g, 6.38 mmole) at room temperature. The reaction mixture was stirred at room temperature for 2 h. The reaction mass was diluted with DCM and washed with water. The organic layer was dried over anhydrous Na2SO4; filtered; then concentrated under reduced pressure to get crude compound as mixture of diastereomers. The material was subjected to SFC purification to afford (2S,4R)-methyl 1-((2S,3R)-2-(tert-butoxycarbonylamino)-3-ethyl-5-methylnon-8-enoyl)-4-hydroxypyrrolidine-2-carboxylate (1.5 g, 53%). MS: MS m/z 441.6 (M++1). washed with water The organic layer was dried over anhydrous Na2SO4 filtered then concentrated under reduced pressure to get crude compound as mixture of diastereomers The material was subjected to SFC purification

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

Procedure: Diisopropylethylamine (0.782 g, 5.5 mmol) was dropped into a suspension of compound (2S,4R)-4-Hydroxy-pyrrolidine-2-carboxylic acid methyl ester (1.0 g, 5.5 mmol) in anhydrous dichloromethane (50 mL), the mixture was stirred at room temperature for 10 minutes. 1-isocyanato-4-(trifluoromethoxy)benzene (1.116 g, 5.5 mmol) was added to the suspension and the mixture was continued to be stirred for 2 hours at room temperature. The mixture was poured into water (50 mL), the resulting mixture was filtrated. The liquid was extracted with dichloromethane (3×50 mL). The organic layers was combined, washed with brine (50 mL), dried over anhydrous sodium sulphate, filtered and concentrated. The residue combined with the filter cake yielded 1.86 g (97%) of the target compound. 1H NMR (300 MHz, CDCl3): δ 7.41 (d, 2H, J=9.0 Hz), 7.14 (d, 2H, J=8.7 Hz), 6.59 (s, 1H), 4.64 (t, 2H, J=7.5 Hz), 3.78-3.73, (m, 4H), 3.59 (d, 1H, J=9.9 Hz), 2.36-2.21 (m, 2H); LC-MS: 349.1 [M+1]+. to be stirred for 2 hours at room temperature the resulting mixture was filtrated The liquid was extracted with dichloromethane (3×50 mL) washed with brine (50 mL) dried over anhydrous sodium sulphate filtered concentrated

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

Procedure: Thionyl chloride (47 g, 393 mmol) was added dropwise at 0° C. to a stirred solution of (2S,4R)-4-hydroxy-pyrrolidine-2-carboxylic acid (40 g, 0.31 mol) in anhydrous methanol (200 mL). The reaction was allowed to proceed overnight, and concentrated to give the title intermediate (50 g). 1H NMR (400 MHz, CD3OD) δ (ppm) 4.56 (m, 2H), 3.85 (s, 3H), 3.41 (m, 1H), 3.28 (m, 1H), 2.40 (m, 1H), 2.22 (m, 1H). concentrated

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

Procedure: Thionyl chloride (47 g, 393 mmol) was added dropwise at 0° C. to a stirred solution of (2S,4R)-4-hydroxy-pyrrolidine-2-carboxylic acid (40 g, 0.31 mol) in anhydrous methanol (200 mL). The reaction was allowed to proceed overnight, and concentrated to give the title intermediate (50 g). 1H NMR (400 MHz, CD3OD) δ (ppm) 4.56 (m, 2H), 3.85 (s, 3H), 3.41 (m, 1H), 3.28 (m, 1H), 2.40 (m, 1H), 2.22 (m, 1H). concentrated

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

Procedure: Methyl (2S,4R)-4-hydroxypyrrolidine-2-carboxylate 3a (5.53 g, 38 mmol) was dissolved in 80 mL of methanol in an ice-water bath, followed by addition of 40% formaldehyde solution (31 mL, 380 mmol) and sodium cyanoborohydride (12 g, 190 mmol) in batches. The reaction mixture was stirred for 0.5 hours, then warmed up to room temperature and stirred for 3 hours. The mixture was quenched with 40 mL of water, concentrated under reduced pressure and extracted with dichloromethane (80 mL×3). The combined organic extracts were washed with saturated brine (30 mL×2), dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain the title compound methyl (2S,4R)-4-hydroxy-1-methyl-pyrrolidine-2-carboxylate 3b crude product as a colourless oil, which was directly used in the next step. stirred for 3 hours The mixture was quenched with 40 mL of water concentrated under reduced pressure extracted with dichloromethane (80 mL×3) The combined organic extracts were washed with saturated brine (30 mL×2) dried over anhydrous sodium sulfate filtered The filtrate was concentrated under reduced pressure

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

Procedure: Thionyl chloride (47 g, 393 mmol) was added dropwise at 0° C. to a stirred solution of (2S,4R)-4-hydroxy-pyrrolidine-2-carboxylic acid (40 g, 0.31 mol) in anhydrous methanol (200 mL). The reaction was allowed to proceed overnight, and concentrated to give the title intermediate (50 g). 1H NMR (400 MHz, CD3OD) δ (ppm) 4.56 (m, 2H), 3.85 (s, 3H), 3.41 (m, 1H), 3.28 (m, 1H), 2.40 (m, 1H), 2.22 (m, 1H). concentrated

Conditions: temp:120.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: 500 mg 4-chloro-2-[4-(4-chloro-phenyl)-piperazin-1-yl]-6,7-dihydro-thieno[3,2-d]pyrimidine 5-oxide (cf Example 124), 284.19 mg methyl (2S,4R)-4-hydroxy-pyrrolidine-2-carboxylate and 691.65 μl diisopropylethylamine are placed in 8 ml dioxane, heated to 120° C. in the microwave for 0.3 hours. Then the reaction mixture is combined with water and dichloromethane and extracted. The organic phase is separated off using a phase separation cartridge and evaporated to dryness. The diastereomers are separated by preparative HPLC (method B). 200 mg of Diastereomer 1 (Example 125) and 160 mg of Diastereomer 2 (Example 126) are obtained as powders. Analytical HPLC-MS (method A): Diastereomer 1, RT=2.51 min, Diastereomer 2, RT=2.57 min. extracted The organic phase is separated off a phase separation cartridge evaporated to dryness The diastereomers are separated by preparative HPLC (method B) 200 mg of Diastereomer 1 (Example 125) and 160 mg of Diastereomer 2 (Example 126) are obtained as powders

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

Procedure: A suspension of 4(R)-hydroxyproline (35.12 g, 267.8 mmol) in methanol (500 ml) was saturated with gaseous hydrochloric acid. The resulting solution was allowed to stand for 16 hrs and the solvent evaporated in vacuo to afford the title compound as a white solid. the solvent evaporated in vacuo

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

Procedure: Thionyl chloride (270 mL) was added dropwise to methanol (1500 mL) over 1 hour, followed by the addition of (4R)-4-hydroxy-L-proline (150 g). The mixture was stirred at reflux for 36 hours, and then concentrated under reduced pressure. Methanol (1500 mL) was added, followed by a slow addition of triethylamine (310 mL). The mixture was cooled to 0° C., filtered, and the filtrate was concentrated to afford methyl (4R)-4-hydroxy-L-prolinate. The residue was used directly without further purification. at reflux for 36 hours concentrated under reduced pressure Methanol (1500 mL) was added followed by a slow addition of triethylamine (310 mL) filtered the filtrate was concentrated

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

Procedure: To a solution of (3R)-2-(tert-butoxycarbonylamino)-3-ethyl-5-methylnon-8-enoic acid (2 g, 6.38 mmole) in dichloromethane (20 mL) was added DIPEA (1.93 mL, 19.19 mmole), HATU (2.42 g, 6.38 mmole) followed by (2S,4R)-methyl 4-hydroxypyrrolidine-2-carboxylate (1.15 g, 6.38 mmole) at room temperature. The reaction mixture was stirred at room temperature for 2 h. The reaction mass was diluted with DCM and washed with water. The organic layer was dried over anhydrous Na2SO4 and concentrated under reduced pressure to get crude compound. The crude compound was taken to the next step without further purification (1.5 g, 53.4%). MS: MS m/z 441.6 (M++1) washed with water The organic layer was dried over anhydrous Na2SO4 concentrated under reduced pressure to get crude compound The crude compound was taken to the next step without further purification (1.5 g, 53.4%)