Synthesis of verubecestat, a BACE1 inhibitor for the treatment of Alzheimer s disease

Transcription

1 Supporting Information Synthesis of verubecestat, a BACE1 inhibitor for the treatment of Alzheimer s disease David A. Thaisrivongs*, Steven P. Miller*, Carmela Molinaro, Qinghao Chen, Zhiguo J. Song, Lushi Tan, Lu Chen, Wenyong Chen, Azzeddine Lekhal, Sarah K. Pulicare, and Yanke Xu Process Chemistry and Chemical Engineering Research and Development, Process Research and Development, Merck Research Laboratories, P.O. Box 2000, Rahway, New Jersey 07065, WuXi AppTec (Shanghai) Pharmaceutical Co. Ltd., 288 Fute Zhong Road, Waigaoqiao Free Trade Zone, Shanghai , China Experimental procedures and compound characterization data Materials and Methods: All reactions were performed in glassware with overhead stirring under a nitrogen atmosphere using commercial-grade solvents. Air and moisture-sensitive liquids were transferred via stainless steel syringe or cannula and introduced into the reaction vessel through rubber septa. Reaction temperatures were controlled via the reactor jacket, which was supplied with heat exchanging fluid (1:1 water:ethylene glycol or DW-THERM) at the appropriate temperature. Reactions were monitored by reverse-phase HPLC on a Hewlett Packard 1100 Series instrument with a C18 Ascentis Express column by Supelco and an acetonitrile and 0.1% phosphoric acid mobile phase system. All isolated and characterized compounds were >95% pure as judged by 1 H spectroscopic analysis. 1 H and 13 C NMR spectroscopy were performed on Bruker DPX-500 NMR spectrometers. Chemical shifts are reported in ppm relative to residual protio solvent signals; all 13 C NMR spectra are proton decoupled. Data for 1 H are reported as chemical shift (δ ppm), multiplicity (s = singlet, d = doublet, t = triplet, q = quartet, quint = quintet, sex = sextet, sept = septet, oct = octet, m = multiplet, app. = apparent), coupling constant, integration); data for 13 C are reported in terms of chemical shift. High resolution mass spectra were recorded on a Xevo G2 QToF (Waters MS Technologies) mass spectrometer by electrospray ionization. Page 1 of 15

2 O H OMe MeNH2 MeOH N Me OMe NaBH4 MeOH, THF HN Me OMe MsCl, Et3N toluene O O Me S N Me OMe Stage 1 Stage 2 Stage 3 3 N-(4-methoxybenzyl)-N-methylmethanesulfonamide (3) Stage 1: To reactor R1 was charged MeOH (250 ml), followed by methylamine (30% w/w in MeOH, 415 g, 1.2 equivalents). Agitation was begun and the solution cooled to 0 C. To this was charged p-anisaldehyde (500 g, 1.0 equivalents) over 40 minutes, maintaining the internal temperature below 3 C. The reaction was heated to 20 C at a rate of 0.5 C per minute and then aged for 1 hour, at which point it was sampled to confirm that less than 1.0% of p-anisaldehyde remained. Stage 2: To reactor R2 was charged THF (250 ml), followed by sodium borohydride (111 g, 0.8 equivalents); additional THF (250 ml) was used to wash down the sodium borohydride. Agitation was begun and the solution cooled to 0 C. The solution in R1 from Stage 1 was charged to R2 over 2 hours and then the reaction mixture aged for 2 hours, at which point it was sampled to confirm that less than 1.0% of the imine from Stage 1 remained. To reactor R3 was charged water (250 ml) and HCl (35% w/w in water, 750 ml). Agitation was begun and the solution in R3 cooled to 0 C. The solution in R2 was charged to R3 over 8 hours, maintaining the internal temperature in R3 below 3 C, and then the mixture was aged for 1 hour. The solution in R3 was basified to a ph of 13.5 by charged sodium hydroxide (50% w/w in water, 800 ml) over 2 hours, maintaining the internal temperature in R3 between 15 and 25 C, at which point water (750 ml) was added to dissolve residual solids. Toluene (1 L) was charged to R3 and the biphasic mixture agitated for 30 minutes. The layers were allowed to separate, and then the organic layer was transferred to reactor R4. Toluene (500 ml) was charged to R3 and the biphasic mixture agitated for 30 minutes. The layers were allowed to separate, and then the organic layer was transferred to reactor R4. The combined organic layers in R4 were agitated and concentrated by vacuum to approximately 600 ml, maintaining the internal temperature in R4 below 45 C, at which point it was sampled to confirm that the amount of residual water was less than 0.5%. Stage 3: Toluene (2.9 L) and triethylamine (713 ml, 1.4 equivalents) were charged to R4, maintaining the internal temperature between 25 and 30 C. Methanesulfonyl chloride (341 ml, 1.2 equivalents) was charged to R4 over 1 hour, maintaining the internal temperature between 25 and 30 C. The reaction was aged for 2 hours, at which point it was sampled to confirm that less than 1.0% of the amine from Stage 2 remained. Water (1.3 L) was charged to R4 and the biphasic mixture agitated for 30 minutes. The layers were allowed to separate, and then the organic layer was transferred to reactor R5. Toluene (1.2 L) was charged to R4 and the biphasic mixture agitated for 30 minutes. The layers were allowed to separate, and then the organic layer was transferred to reactor R5. To the combined organic layers in R5 was charged sodium chloride (2% w/w in water, 2.5 L) and the biphasic mixture agitated for 30 minutes. The layers Page 2 of 15

3 were allowed to separate, and then the aqueous layer was discarded. The organic solution in R5 was agitated and concentrated by vacuum to approximately 2.5 L, maintaining the internal temperature between 35 and 40 C. Heptane (4 L) was charged to R5 over 12 hours, maintaining the internal temperature between 35 and 40 C, at which point the solution was cooled over 3 hours to an internal temperature between 0 and 5 C. The solids were filtered, washed with heptane (2.5 L) that was cooled to between 0 and 10 C, and then dried for 12 hours between 45 and 50 C under vacuum. The typical purity of the isolated solids by HPLC area percent is greater than 98%, and the typical isolated yield is between 90-95%. 1 H NMR (500 MHz, DMSO-d 6 ) δ 7.25 (d, J = 10.0 Hz, 2 H), 6.94 (d, J = 10.0 Hz, 2 H), 4.15 (s, 2 H), 3.75 (s, 3H), 2.91 (s, 3H), 2.62 (s, 3H); 13 C NMR (500 MHz, DMSO-d 6 ) δ 158.8, 129.6, 128.1, 113.9, 55.0, 52.5, 35.1, 33.9; HRMS [M+NH 4 ] (C 10 H 19 N 2 O 3 S) calc , obs (R,E)-N-(1-(5-bromo-2-fluorophenyl)ethylidene)-2-methylpropane-2-sulfinamide (13) To reactor R1 was charged 1-(5-bromo-2-fluorophenyl)ethan-1-one (400 g, 1 equivalent) and (R)-2-methylpropane-2-sulfinamide (246 g, 1.1 equivalents), followed by EtOAc (1 L). Agitation was begun and the solution heated to 60 C, at which point titanium(iv) ethoxide (85% in isopropyl alcohol, 544 g, 1.1 equivalent) was charged. The reaction was aged for 24 hours, at which point it was sampled to confirm that less than 5% of 1-(5-bromo-2-fluorophenyl)ethan-1- one remained. The solution was then cooled to between 20 and 30 C, at which point EtOAc (3 L) was charged. To reactor R2 was charged potassium glycolate (5 M in water, 4 L) and agitation begun. The solution from R1 was charged to R2 and the biphasic mixture agitated for 1 hour, at which point the layers were allowed to separate and the aqueous layer discarded. To reactor R2 was charged potassium glycolate (5 M in water, 2 L) and agitation begun. The biphasic mixture was agitated for 1 hour, at which point the layers were allowed to separate and the aqueous layer discarded. The final organic layer was agitated and concentrated by vacuum to approximately 2.6 L, maintaining the internal temperature in R2 below 30 C. The solvent in R2 was switched via constant volume distillation using isopropyl alcohol (8 L), maintaining the internal temperature between 20 and 30 C, at which point the solution was sampled to confirm that less than 0.5% w/w EtOAc remained. The batch was treated with 5% w/w CUNO 3 (28.0 g) and then filtered into reactor R3, flushing the CUNO 3 solids with isopropyl alcohol (1.2 L). The solution in R3 was agitated and concentrated by vacuum to approximately 2.0 L, maintaining the Page 3 of 15

4 internal temperatuer between 20 and 30 C. EtOAc (100 ml) and water (500 ml) were charged to R3, and then the solution cooled to 15 C. Product seed (12.0 g) was charged to R3 and the resulting seed bed aged for 2 hours. Water (2.5 L) was charged over 12 hours and then the batch cooled to 0 C over 2 hours. The solids were filtered, washed with a solution of isopropyl alcohol and water (1:4 v/v, 1.2 L) that was cooled to between 0 and 10 C, and then dried for 12 hours at 40 C under vacuum. The typical purity of the isolated solids by HPLC area percent is greater than 96%, and the typical isolated yield is between 80-85%. 1 H NMR (500 MHz, DMSO-d 6 ) δ 7.79 (d, J = 4.6 Hz, 1H), (m, 1H), 7.31 (t, J = 10.2 Hz, 1H), 2.67 (s, 3H), 1.20 (s, 9H); 13 C NMR (500 MHz, DMSO-d 6 ) δ 173.7, 159.7, 157.7, 135.2, 131.7, (d, J = 50 Hz), (d, J = 95 Hz), 116.3, 57.0, 21.9; HRMS [M+H] (C 12 H 15 BrFNOS) calc , obs (R)-2-(5-bromo-2-fluorophenyl)-2-(((R)-tert-butylsulfinyl)amino)-N-(4-methoxybenzyl)-Nmethylpropane-1-sulfonamide (12) To reactor R1 was charged N-(4-methoxybenzyl)-N-methylmethanesulfonamide (3) (116 g, 1.75 equivalents) and THF (630 ml). Agitation was begun and the solution cooled to between 25 and 15 C. n-hexyllithium (2.3 M in hexanes, 206 ml, 1.72 equivalents) was charged, maintaining the internal temperature between 25 and 15 C, and then the reaction aged for 1 hour. The solution in R1 was cooled to an internal temperature below 60 C. To reactor R2 was charged (R,E)-N-(1-(5-bromo-2-fluorophenyl)ethylidene)-2-methylpropane-2-sulfinamide (13) (90 g, 1.0 equivalent) and THF (270 ml). Agitation was begun, and resulting solution was transferred to reactor R1, maintaining the internal temperature of R1 below 60 C. The reaction was aged for 1 hour, at which point it was sampled to confirm that less than 25% of (R,E)-N-(1- (5-bromo-2-fluorophenyl)ethylidene)-2-methylpropane-2-sulfinamide (13) remained. Acetic acid (33 ml, 2.0 equivalents) was charged to R1, maintaining the internal temperature below 35 C, and then the solution was heated to between 20 and 25 C. Water (270 ml) was charged, the biphasic mixture agitated for 15 minutes, at which point the layers were allowed to separate and the aqueous layer discarded. Sodium chloride (10% w/w in water, 270 ml) was charged, the biphasic mixture agitated for 15 minutes, at which point the layers were allowed to separate and the aqueous layer discarded. The organic layer was agitated and concentrated by vacuum to approximately 270 ml, maintaining the internal temperature in R1 below 35 C. The solvent was switched via constant volume distillation using toluene (450 ml), maintaining the internal Page 4 of 15

5 temperature below 35 C, at which point the solution was sampled to confirm that less than 1.0% w/w THF remained. The typical purity of the reaction stream by HPLC area percent is 55 to 65%, and the typical assay yield (including both product diastereomers) is between 70-75%. 1 H NMR (500 MHz, DMSO-d 6 ) δ 7.64 (dd, J = 2.6, 7.1 Hz, 1H), 7.60 (ddd, J = 2.6, 4.2, 8.7 Hz, 1H), (m, 3H), 6.91 (dt, J = 2.9, 8.8 Hz, 2H), 5.58 (s, 1H), 4.08 (dd, J = 14.3, 19.0 Hz, 2H), 3.80 (dd, J = 14.3, 18.8 Hz, 2H), 3.74 (s, 3H), 3.30 (s, 3H), 2.57 (s, 3H), 1.91 (s, 3H), 1.16 (s, 9H); 13 C NMR (500 MHz, DMSO-d 6 ) δ 160.8, 158.8, 158.3, (d, J = 50 Hz), (d, J = 60 Hz), (d, J = 20 Hz), 129.5, 127.7, (d, J = 125 Hz), (d, J = 15 Hz), 114.0, 57.8 (d, J = 15 Hz), 57.2 (d, J = 30 Hz), 55.4 (d, J = 310 Hz), 52.0, 33.5, 27.0, 22.4; HRMS [M+H] (C 22 H 30 BrFN 2 O 4 S 2 ) calc , obs N-(3-((R)-2-(((R)-tert-butylsulfinyl)amino)-1-(N-(4-methoxybenzyl)-N-methylsulfamoyl)propan-2-yl)-4-fluorophenyl)-5-fluoropicolinamide (10) To reactor R1 was charged (R)-2-(5-bromo-2-fluorophenyl)-2-(((R)-tert-butylsulfinyl)amino)-N- (4-methoxybenzyl)-N-methylpropane-1-sulfonamide (12) (100 g diluted in approximately 300 ml of toluene, 1.0 equivalent), 5-fluoropicolinamide (11) (36.0 g, 1.4 equivalents), potassium carbonate (176 g, 7.0 equivalents), and water (164 ml). Agitation was begun and the mixture sparged with argon for 2 hours. To reactor R2 was charged trans-n,n'-dimethyl-cyclohexane- 1,2-diamine (15.5 g, 0.6 equivalent) and toluene (170 ml). Agitation was begun and the solution sparged with argon for 1 hour. Copper(I) iodide (10.5 g, 0.3 equivalent) was charged to R2 and the solution sparged with argon for 1 hour. The solution from R2 was transferred to R1 and the final mixture sparged with argon for 1 hour. The reaction mixture in R1 was heated to 95 C and aged for 20 hours, at which point it was sampled to confirm that less than 1.0% of 12 remained. The mixture was cooled to between 20 and 25 C, at which point ethane-1,2-diamine (32.8 g, 3.0 equivalents) and MeCN (300 ml) were charged. After agitating the biphasic mixture for 15 minutes, the layers were allowed to separate and the aqueous layer discarded. Acetic acid (65.5 g, 6.0 equivalents) and sodium chloride (25% w/w in water, 400 ml) were charged and the biphasic mixture agitated for 15 minutes. The layers were allowed to separate and the aqueous layer discarded. Sodium bicarbonate (7% w/w in water) was charged and the biphasic mixture agitated for 15 minutes. The layers were allowed to separate and the aqueous layer discarded. Sodium chloride (25% w/w in water, 400 ml) were charged and the biphasic mixture agitated Page 5 of 15

6 for 15 minutes. The layers were allowed to separate and the aqueous layer discarded. The solvent was switched via constant volume distillation using isopropyl alcohol (1.1 L), maintaining the internal temperature below 45 C, at which point the solution was sampled to confirm that less than 1.0% w/w toluene remained. The batch was heated to 80 C to bring all of the solids into solution, at which point the batch was cooled to 65 C and product seed (2 g) was charged. The resulting seed bed was aged for 4 hours, and then the slurry was cooled to 50 C over 10 hours. Water (800 ml) was charged over 6 hours and then the slurry was cooled to 20 C over 6 hours. The solids were filtered, washed with isopropyl alcohol (500 ml), and then dried under vacuum at 35 C. The typical purity of the isolated solids by HPLC area percent is 95%, and the typical isolated yield is 70%. 1 H NMR (500 MHz, DMSO-d 6 ) δ 10.8 (s, 1H), 8.73 (d, J = 2.8 Hz, 1H), (m, 2H), 7.97 (dt, J = 2.8, 8.7 Hz, 1H), 7.92 (ddd, J = 2.8, 3.9, 8.8 Hz, 1H), 7.19 (d, J = 8.8 Hz, 2H), 7.19 (t, J = 11.2 Hz, 1H), 6.90 (d, J = 8.7 Hz, 2H), 5.58 (s, 1H), 4.05 (q, J = 14.5 Hz, 2H), 3.86 (q, J = 14.3 Hz, 2H), 3.73 (s, 3H), 3.32 (s, 1H), 2.55 (s, 3H), 1.93 (s, 3H), 1.20 (s, 9H); 13 C NMR (500 MHz, DMSO-d 6 ) δ 161.8, 161.7, 159.8, 158.8, 157.4, 155.5, (d, J = 15 Hz), (d, J = 100 Hz), 134.6, 129.5, (d, J = 45 Hz), 127.8, (d, J = 15 Hz), (d, J = 35 Hz), 120.6, (d, J = 95 Hz), 113.9, 57.9, 57.4, 55.3 (d, J = 235 Hz), 51.9, 33.4, 26.9, 22.4; HRMS [M+H] (C 28 H 34 F 2 N 4 O 5 S 2 ) calc , obs (R)-N-(3-(2-amino-1-(N-methylsulfamoyl)propan-2-yl)-4-fluorophenyl)-5-fluoropicolinamide (18) To reactor R1 was charged N-(3-((R)-2-(((R)-tert-butylsulfinyl)amino)-1-(N-(4-methoxybenzyl)- N-methylsulfamoyl)propan-2-yl)-4-fluorophenyl)-5-fluoropicolinamide (10) (30 g, 1.0 equivalent) and acetic acid (60 ml). Agitation was begun and methanesulfonic acid (30 ml, 9.4 equivalents) was charged and the reaction heated to 70 C and aged for 2 hours, at which point it was sampled to confirm that less than 1% of 10 remained. The solution was then cooled to an internal temperature of between 20 and 25 C. Glycolic acid (20% w/w in water) was charged, maintaining an internal temperature of between 20 and 25 C, and the solution agitated for 20 minutes. Toluene (150 ml) was charged to R1 and the biphasic mixture agitated for 20 minutes. The layers were allowed to separate and the organic layer discarded. Toluene (150 ml) was charged to R1 and the biphasic mixture agitated for 20 minutes. The layers were allowed to Page 6 of 15

7 separate and the organic layer discarded. To reactor R2 was charged ammonium hydroxide (28% w/w in water, 270 ml) and product seed (900 mg). Agitation in R2 was begun, and then the remaining aqueous layer in R1 was charged to R2 over 12 hours, maintaining an internal temperature of between 20 and 25 C. The slurry was agitated for an additional 2 hours and then the solids were filtered, washed with water (60 ml), and the dried under vacuum at 40 to 45 C. The typical purity of the isolated solids by HPLC area percent is greater than 98%, and the typical isolated yield is between 94 and 98%. 1 H NMR (500 MHz, DMSO-d 6 ) δ 10.6 (s, 1H), 8.72 (d, J = 2.9 Hz, 1H), 8.23 (dd, J = 4.7, 8.8 Hz, 1H), 8.14 (dd, J = 2.6, 7.5 Hz, 1H), 7.99 (dt, J = 2.9, 8.7 Hz, 1H), 7.86 (ddd, J = 2.9, 4.2, 8.7 Hz, 1H), 7.10 (dd, J = 8.8, 12.0 Hz, 1H), 6.80 (s, 1H), 3.61 (d, J = 14.3, 1H), 3.44 (d, J = 14.2 Hz, 1H), 3.30 (s, 3H), 1.52 (s, 3H), 1.27 (s, 1H); 13 C NMR (500 MHz, DMSO-d 6 ) δ 162.1, 161.3, 159.5, 157.6, 155.2, (d, J = 20 Hz), (d, J = 125 Hz), (d, J = 10 Hz), (d, J = 65 Hz), (dd, J = 95, 150 Hz), (dd, J = 20, 50 Hz), (d, J = 125 Hz), 59.9 (d, J = 25 Hz), 53.1 (d, J = 15 Hz), 29.3 (d, J = 15 Hz), 28.4; HRMS [M+H] (C 16 H 18 F 2 N 4 O 3 S) calc , obs MK-8931 HBr (19) To reactor R1 was charged (R)-N-(3-(2-amino-1-(N-methylsulfamoyl)propan-2-yl)-4- fluorophenyl)-5-fluoropicolin-amide (18) (10 g, 1.0 equivalent), isopropyl acetate (35 ml), and acetonitrile (15 ml). Agitation was begun and to the mixture was added cyanogen bromide (5 M in acetonitrile, 7.7 ml, 1.6 equivalents). (NOTE: Cyanogen bromide is highly toxic and should be handled in a well-ventilated hood. Residual cyanogen bromide should be quenched with an aqueous caustic solution containing bleach.) The reaction mixture was heated to an internal temperature of 75 C and aged for 4 h, at which point product seed (10 mg) was charged. The resulting slurry was aged for an additional 46 hours before being slowly cooled to between 20 and 25 C over 3 hours. The slurry was aged for an additional 1 hour and then the solids were filtered, washed twice with a solution of isopropyl acetate and acetonitrile (7:3 v/v, 20 ml), and then dried under vacuum at 50 C. The typical purity of the isolated solids by HPLC area percent is greater than 98%, and the typical isolated yield is 85 to 90%. 1 H NMR (500 MHz, DMSO-d 6 ) δ 10.8 (s, 1H), 8.75 (d, J = 2.8 Hz, 1H), 8.24 (dd, J = 4.6, 8.8 Hz, 1H), (m, 3H), 7.29 (dd, J = 8.7, 12.0 Hz, 1H), 4.46 (s, 2H), 3.31 (s, 2H), 3.21 (s, 3H), 1.83 (s, 3H); 13 C NMR (500 MHz, DMSO-d 6 ) δ (d, J = 95 Hz), 159.9, 156.1, 154.2, 154.0, (d, J = 15 Hz), (d, J = 100 Hz), (d, J = 20 Hz), (d, J = 45 Hz), Page 7 of 15

8 124.9 (d, J = 100 Hz), (d, J = 35 Hz), 119.3, (d, J = 100 Hz), 56.1 (d, J = 10 Hz), 54.1 (d, J = 20 Hz), 29.3, 27.3; HRMS [M+H] (C 17 H 17 F 2 N 5 O 3 S) calc , obs MK-8931 (1) To reactor R1 was charged MK-8931 HBr (19) (20 g, 1.0 equivalent), ethyl acetate (200 ml), and potassium carbonate (1 M in water, 4.14 ml). The biphasic mixture was agitated for 15 minutes and then the layers were allowed to separate and the aqueous layer discarded. Water (60 ml) was charged to R1, the biphasic mixture was agitated for 15 minutes, and then the layers were allowed to separate and the aqueous layer discarded. The organic layer was concentrated to 80 ml, maintaining the internal temperature below 35 C. Ethyl acetate (180 ml) was charged to R1 and the solution was concentrated to 120 ml, maintaining the internal temperature below 35 C. The batch was heated to 50 C to bring all of the solids into solution, at which point heptane (20 ml) was charged followed by product seed (120 mg). The resulting seed bed was aged for 4 hours, and then heptane (160 ml) was charged slowly over 6 hours. The slurry was cooled to 20 C over 2 hours, at which point the solids were filtered, washed with heptane (60 ml), and then dried under vacuum at 35 C. The typical purity of the isolated solids by HPLC area percent is greater than 99%, and the typical isolated yield is 91 to 94%. 1 H NMR (500 MHz, DMSO-d 6 ) δ (s, 1H), 8.73 (d, J = 2.8 Hz, 1H), 8.23 (dd, J = 13.5 Hz, 17.6 Hz, 1H), (m, 2H), 7.79 (ddd, J = 3.2, 4.0, 7.1 Hz, 1H), 5.97 (s, 2H), 3.76 (dd, J = 14, 18.5 Hz, 2H), 3.31(s, 1H), 3.10 (s, 3H), 1.61 (s, 3H); 13 C NMR (500 MHz, DMSO-d 6 ) δ 161.8, 161.4, 159.8, 156.9, 155.0, (d, J = 15 Hz), (d, J = 100 Hz), 134.1, (d, J = 50 Hz), (d, J = 75 Hz), (d, J = 25 Hz), 120.9, 120.9, (d, J = 100 Hz), 56.6, 54.2, 29.4 (d, J = 120 Hz); HRMS [M+H] (C 17 H 17 F 2 N 5 O 3 S) calc , obs NMR spectra Page 8 of 15

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