[chloro(methyl)phosphoryl]methane

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

Procedure: Diisopropyl ethyl amine (124.28 g, 0.961 mol) was added dropwise to stirred solution of compound tert-butyl 3-(hydroxymethyl)azetidine-1-carboxylate (150 g, 0.80 mol) in dichloromethane (500 mL) at 0° C. Dimethyl phosphinyl chloride (90 g, 0.80 mol) in dichloromethane (300 mL) was added to above reaction mixture over a period of 90 min at same temperature. Then, the reaction mixture was warmed to room temperature and continued stirring for 2 h. After completion of the reaction, the mixture was diluted with water (300 mL). The layers were separated. The organic layer was washed with brine (300 ml), dried over sodium sulfate and concentrated. The resultant concentrated product was purified via column chromatography on silica gel using chloroform/methanol (95/5) to afford tert-butyl 3-{[(dimethylphosphoryl)oxy]methyl}azetidine-1-carboxylate (146.8 g, 68.3%) as brown liquid. After completion of the reaction The layers were separated The organic layer was washed with brine (300 ml) dried over sodium sulfate concentrated The resultant concentrated product was purified via column chromatography on silica gel

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

Procedure: 2-(3-Aminophenylethynyl)-1-ethyl-6-methoxy-1H-indole-3-carbonitrile (100 mg, 0.32 mmol), prepared as described in Example 1H, is dissolved in pyridine (300 μL) at 0° C. Dimethylphosphinic chloride (60 mg, 0.53 mmol) in THF (300 μL) is added. The reaction is stirred at room temperature for 2 hours. The reaction mixture is diluted in EtOAc, and is washed with aqueous HCl followed by brine. The organic layer is dried and concentrated. Purification by silica chromatography (acetone) yields N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-dimethylphosphinic amide (65 mg, 52%), compound 227, as a pure white solid. The silica column is then flushed with 9/1 CH2Cl2/MeOH to yield 9 mg of N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-bis-(dimethylphosphinic)amide as a by-product. is washed with aqueous HCl The organic layer is dried concentrated Purification by silica chromatography (acetone)

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

Procedure: 2-(3-Aminophenylethynyl)-1-ethyl-6-methoxy-1H-indole-3-carbonitrile (100 mg, 0.32 mmol), prepared as described in Example 1H, is dissolved in pyridine (300 μL) at 0° C. Dimethylphosphinic chloride (60 mg, 0.53 mmol) in THF (300 μL) is added. The reaction is stirred at room temperature for 2 hours. The reaction mixture is diluted in EtOAc, and is washed with aqueous HCl followed by brine. The organic layer is dried and concentrated. Purification by silica chromatography (acetone) yields N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-dimethylphosphinic amide (65 mg, 52%), compound 227, as a pure white solid. The silica column is then flushed with 9/1 CH2Cl2/MeOH to yield 9 mg of N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-bis-(dimethylphosphinic)amide as a by-product. is washed with aqueous HCl The organic layer is dried concentrated Purification by silica chromatography (acetone)

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

Procedure: A solution of ManNAc (5.0 g, 2.6 mmol), Ac2O (10 mL) and pyridine (20 mL) was stirred for 10 hours at room temperature, and the mixture was concentrated, followed by coevaporation with toluene. A solution of the residue, benzyl alcohol (20 mL), BF3.OEt2 (1.6 g, 11.3 mmol) in CH3NO2 (150 mL) was gently refluxed for 2.5 hours. After cooling, the mixture was concentrated. The residue was chromatographed on silica gel, with toluene-EtOAc (1:2). The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-a-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) and the solution was stirred for 30 minutes at room temperature, and neutralized by addition of Dowex 50W-X8 (H+). After the resin was filtered off, the filtrate was concentrated, followed by coevaporation with pyridine. A solution of dimethylphosphinic chloride (1 g, 8.8 mmol) in DMF was added to a cold solution of benzyl 2-acetamido-2-deoxy-a-D-mannopyranoside (0.50 g, 1.6 mmol), 2,6-lutidine (0.34 g, 3.2 mmol) in anhydrous DMF (30 mL) in a dry ice-acetone bath, and the reaction was allowed to slowly warm to room temperature. The reaction was monitered by TLC with (1M NH4OAc/2-proponal/EtOAc, 1/2.4/3.4). After 10 hours, the reaction mixture was directly applied to silica gel chromatography, eluted with CHCl3 /EtOAc/MeOH (5/2/1) to give benzyl 2-acetamido-2-deoxy-6-(dimethyl-phosphinyl)-a-D-mannopyranoside (56 percent yield). the mixture was concentrated After cooling the mixture was concentrated The residue was chromatographed on silica gel, with toluene-EtOAc (1:2) The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-a-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) neutralized by addition of Dowex 50W-X8 (H+) After the resin was filtered off the filtrate was concentrated to slowly warm to room temperature After 10 hours eluted with CHCl3 /EtOAc/MeOH (5/2/1)

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

Procedure: To a cooled (0° C.) flask containing 34.0 mL (2.0 M in Et2O, 68.4 mmol) of 4-fluorophenylmagnesium bromide, under an atmosphere of N2, was added a solution of dimethylphosphinic chloride (3.50 g, 31.1 mmol) in 84 mL of THF, dropwise via cannulation, over 20 min. The green reaction mixture was stirred at 0° C. for 1 h, then quenched at 0° C. with 30 mL of saturated NH4Cl resulting in the formation of a white precipitate. The mixture was concentrated on a rotary evaporator and partitioned between EtOAc (200 mL) and H2O (200 mL), upon which the layers were separated. The aqueous layer was extracted with EtOAc (3×100 mL) and the combined organics washed with brine, then dried over MgSO4 and concentrated. The crude product was purified by silica gel flash chromatography (eluted with 5% MeOHIDCM) to provide 2.08 g of an off-white solid: 1H NMR (300 MHz, DMSO-d6) δ 7.83 (m, 2H), 7.35 (td, J=8.9, 1.7 Hz, 2H), 1.66 (s, 3H), 1.61 (s, 3H). 31P NMR (121 MHz, DMSO-d6) δ 37.186. 19F NMR (282 MHz, DMSO-d6) δ−105.14. quenched at 0° C. with 30 mL of saturated NH4Cl resulting in the formation of a white precipitate The mixture was concentrated on a rotary evaporator partitioned between EtOAc (200 mL) and H2O (200 mL), upon which the layers were separated The aqueous layer was extracted with EtOAc (3×100 mL) the combined organics washed with brine dried over MgSO4 concentrated The crude product was purified by silica gel flash chromatography (eluted with 5% MeOHIDCM)

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

Procedure: 2-(3-Aminophenylethynyl)-1-ethyl-6-methoxy-1H-indole-3-carbonitrile (100 mg, 0.32 mmol), prepared as described in Example 1H, is dissolved in pyridine (300 μL) at 0° C. Dimethylphosphinic chloride (60 mg, 0.53 mmol) in THF (300 μL) is added. The reaction is stirred at room temperature for 2 hours. The reaction mixture is diluted in EtOAc, and is washed with aqueous HCl followed by brine. The organic layer is dried and concentrated. Purification by silica chromatography (acetone) yields N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-dimethylphosphinic amide (65 mg, 52%), compound 227, as a pure white solid. The silica column is then flushed with 9/1 CH2Cl2/MeOH to yield 9 mg of N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-bis-(dimethylphosphinic) amide as a by-product. is washed with aqueous HCl The organic layer is dried concentrated Purification by silica chromatography (acetone)

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

Procedure: A solution of ManNAc (5.0 g, 2.6 mmol), Ac2O (10 mL) and pyridine (20 mL) was stirred for 10 hours at room temperature, and the mixture was concentrated, followed by coevaporation with toluene. A solution of the residue, benzyl alcohol (20 mL), BF3.OEt2 (1.6 g, 11.3 mmol) in CH3NO2 (150 mL) was gently refluxed for 2.5 hours. After cooling, the mixture was concentrated. The residue was chromatographed on silica gel, with toluene-EtoAc (1:2). The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-α-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) and the solution was stirred for 30 minutes at room temperature, and neutralized by addition of Dowex 50W-X8 (H+). After the resin was filtered off, the filtrate was concentrated, followed by coevaporation with pyridine. A solution of dimethylphosphinic chloride (1 g, 8.8 mmol) in DMF was added to a cold solution of benzyl 2-acetamido-2-deoxy-α-D-mannopyranoside (0.50 g, 1.6 mmol), 2,6-lutidine (0.34 g, 3.2 mmol) in anhydrous DMF (30 mL) in a dry ice-acetone bath, and the reaction was allowed to slowly warm to room temperature. The reaction was monitored by TLC with (1M NH4 OAc/2-proponal/EtOAc, 1/2.4/3.4). After 10 hours, the reaction mixture was directly applied to silica gel chromatography, eluted with CHCl3 /EtOAc/MeOH (5/2/1) to give benzyl 2-acetamido-2-deoxy-6-(dimethyl-phosphinyl)-α-D-mannopyranoside (56 percent yield). the mixture was concentrated After cooling the mixture was concentrated The residue was chromatographed on silica gel, with toluene-EtoAc (1:2) The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-α-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) neutralized by addition of Dowex 50W-X8 (H+) After the resin was filtered off the filtrate was concentrated to slowly warm to room temperature After 10 hours eluted with CHCl3 /EtOAc/MeOH (5/2/1)

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

Procedure: 2-(3-Aminophenylethynyl)-1-ethyl-6-methoxy-1H-indole-3-carbonitrile (100 mg, 0.32 mmol), prepared as described in Example 1H, was dissolved in pyridine (300 μL) at 0° C. Dimethylphosphinic chloride (60 mg, 0.53 mmol) in THF (300 μL) was added. The reaction was stirred at room temperature for 2 hours. The reaction mixture was diluted in EtOAc, and was washed with aqueous HCl followed by brine. The organic layer was dried and concentrated. Purification by silica chromatography (acetone) yielded N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-dimethylphosphinic amide (65 mg, 52%), compound 227, as a pure white solid. The silica column was then flushed with 9/1 CH2Cl2/MeOH to yield 9 mg of N-[4-(3-cyano-1-ethyl-6-methoxy-1H-indol-2-ylethynyl)-phenyl]-bis-(dimethylphosphinic) amide as a by-product. was washed with aqueous HCl The organic layer was dried concentrated Purification by silica chromatography (acetone)

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

Procedure: To a cooled (0° C.) flask containing 34.0 mL (2.0 M in Et2O, 68.4 mmol) of 4-fluorophenylmagnesium bromide, under an atmosphere of N2, was added a solution of dimethylphosphinic chloride (3.50 g, 31.1 mmol) in 84 mL of THF, dropwise via cannulation, over 20 min. The green reaction mixture was stirred at 0° C. for 1 h, then quenched at 0° C. with 30 mL of saturated NH4Cl resulting in the formation of a white precipitate. The mixture was concentrated on a rotary evaporator and partitioned between EtOAc (200 mL) and H2O (200 mL), upon which the layers were separated. The aqueous layer was extracted with EtOAc (3×100 mL) and the combined organics washed with brine, then dried over MgSO4 and concentrated. The crude product was purified by silica gel flash chromatography (eluted with 5% MeOH/DCM) to provide 2.08 g of an off-white solid: 1H NMR (300 MHz, DMSO-d6) δ 7.83 (m, 2H), 7.35 (td, J=8.9, 1.7 Hz, 2H), 1.66 (s, 3H), 1.61 (s, 3H). 31P NMR (121 MHz, DMSO-d6) δ 37.186. 19F NMR (282 MHz, DMSO-d6) δ−105.14. quenched at 0° C. with 30 mL of saturated NH4Cl resulting in the formation of a white precipitate The mixture was concentrated on a rotary evaporator partitioned between EtOAc (200 mL) and H2O (200 mL), upon which the layers were separated The aqueous layer was extracted with EtOAc (3×100 mL) the combined organics washed with brine dried over MgSO4 concentrated The crude product was purified by silica gel flash chromatography (eluted with 5% MeOH/DCM)

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

Procedure: To a solution of 1-(((5S,7S)-3-(5-(2-(2-hydroxyethoxy)propan-2-yl)pyrazin-2-yl)-7-methyl-2-oxo-1-oxa-3-azaspiro[4.5]decan-7-yl)methyl)-1H-benzo[d]imidazole-6-carbonitrile (60 mg, 0.119 mmol) in dichloromethane (DCM) (1127 μl) was added DIEA (62.3 μl, 0.357 mmol) and dimethylphosphinic chloride (40.1 mg, 0.357 mmol) at RT. The mixture was stirred for 2 hours, concentrated and reconstituted in DMSO/MeOH and filtered. The solution was purified by reverse phase HPLC (Waters Sunfire 30×150 mm Acetonitrile: Water TFA 30-70%, 50 mL/min, 15 min.). The product was free based with PL-HCO3 resin SPE to yield 2-((2-(5-((5S,7S)-7-((6-cyano-1H-benzo[d]imidazol-1-yl)methyl)-7-methyl-2-oxo-1-oxa-3-azaspiro[4.5]decan-3-yl)pyrazin-2-yl)propan-2-yl)oxy)ethyl dimethylphosphinate as a white solid (5.8 mg, 8%). 1H NMR (400 MHz, DMSO-d6) δ 9.24 (d, J=1.51 Hz, 1H), 8.60 (d, J=1.26 Hz, 1H), 8.48 (s, 1H), 8.38 (s, 1H), 7.83 (d, J=8.53 Hz, 1H), 4.12-4.21 (m, 2H), 3.94-4.03 (m, 2H), 3.88 (dd, J=23.09, 10.04 Hz, 2H), 3.42 (t, J=4.77 Hz, 2H), 2.00 (d, J=13.30 Hz, 1H), 1.88 (d, J=14.05 Hz, 1H), 1.59-1.78 (m, 3H), 1.30-1.56 (m, 15H), 1.07 (s, 3H). MS (m/z) 581.3 (M+H+). concentrated filtered The solution was purified by reverse phase HPLC (Waters Sunfire 30×150 mm Acetonitrile: Water TFA 30-70%, 50 mL/min, 15 min.)

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

Procedure: A solution of ManNAc (5.0 g, 2.6 mmol), Ac2O (10 mL) and pyridine (20 mL) was stirred for 10 hours at room temperature, and the mixture was concentrated, followed by coevaporation with toluene. A solution of the residue, benzyl alcohol (20 mL), BF3.OEt2 (1.6 g, 11.3 mmol) in CH3NO2 (150 mL) was gently refluxed for 2.5 hours. After cooling, the mixture was concentrated. The residue was chromatographed on silica gel, with toluene-EtOAc (1:2). The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-a-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) and the solution was stirred for 30 minutes at room temperature, and neutralized by addition of Dowex 50W-X8 (H+). After the resin was filtered off, the filtrate was concentrated, followed by coevaporation with pyridine. A solution of dimethylphosphinic chloride (1 g, 8.8 mmol) in DMF was added to a cold solution of benzyl 2-acetamido-2-deoxy-a-D-mannopyranoside (0.50 g, 1.6 mmol), 2,6-lutidine (0.34 g, 3.2 mmol) in anhydrous DMF (30 mL) in a dry ice-acetone bath, and the reaction was allowed to slowly warm to room temperature. The reaction was monitered by TLC with (1M NH4OAc/2-proponal/EtOAc, 1/2.4/3.4). After 10 hours, the reaction mixture was directly applied to silica gel chromatography, eluted with CHCl3/EtOAc/MeOH (5/2/1) to give benzyl 2-acetamido-2-deoxy-6-(dimethyl-phosphinyl)-a-D-mannopyranoside (56 percent yield). the mixture was concentrated After cooling the mixture was concentrated The residue was chromatographed on silica gel, with toluene-EtOAc (1:2) The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-a-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) neutralized by addition of Dowex 50W-X8 (H+) After the resin was filtered off the filtrate was concentrated to slowly warm to room temperature After 10 hours eluted with CHCl3/EtOAc/MeOH (5/2/1)

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

Procedure: A solution of ManNAc (5.0 g, 2.6 mmol), Ac2O (10 mL) and pyridine (20 mL) was stirred for 10 hours at room temperature, and the mixture was concentrated, followed by coevaporation with toluene. A solution of the residue, benzyl alcohol (20 mL), BF3.OEt2 (1.6 g, 11.3 mmol) in CH3NO2 (150 mL) was gently refluxed for 2.5 hours. After cooling, the mixture was concentrated. The residue was chromatographed on silica gel, with toluene-EtOAc (1:2). The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-a-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) and the solution was stirred for 30 minutes at room temperature, and neutralized by addition of Dowex 50W-X8 (H+). After the resin was filtered off, the filtrate was concentrated, followed by coevaporation with pyridine. A solution of dimethylphosphinic chloride (1 g, 8.8 mmol) in DMF was added to a cold solution of benzyl 2-acetamido-2-deoxy-a-D-mannopyranoside (0.50 g, 1.6 mmol), 2,6-lutidine (0.34 g, 3.2 mmol) in anhydrous DMF (30 mL) in a dry ice-acetone bath, and the reaction was allowed to slowly warm to room temperature. The reaction was monitered by TLC with (1M NH4OAc/2-proponal/EtOAc, 1/2.4/3.4). After 10 hours, the reaction mixture was directly applied to silica gel chromatography, eluted with CHCl3/EtOAc/MeOH (5/2/1) to give benzyl 2-acetamido-2-deoxy-6-(dimethyl-phosphinyl)-a-D-mannopyranoside (56 percent yield). the mixture was concentrated After cooling the mixture was concentrated The residue was chromatographed on silica gel, with toluene-EtOAc (1:2) The isolated benzyl 2-acetamido-4,5,6-tri-O-acetyl-2-deoxy-a-D-mannopyranoside and 0.15 g of NaOMe was dissolved in MeOH (100 mL) neutralized by addition of Dowex 50W-X8 (H+) After the resin was filtered off the filtrate was concentrated to slowly warm to room temperature After 10 hours eluted with CHCl3/EtOAc/MeOH (5/2/1)