Reaction Steps
A mixture of 4-cyano-2-fluorobenzoic acid (50.4 mg, 0.305 mmol, Alfa Aesar), N,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)uronium hexafluorophosphate (107 mg, 0.282 mmol) and triethylamine (196 uL, 1.41 mmol) in THF (2.8 mL) was stirred for 15 minutes, followed by the addition of the diastereomeric 4-[(2R)-2-methylpiperazin-1-yl]-3-[4-(7-{[2-(trimethylsilyl)ethoxy]methyl}-7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]butanenitrile hydrochloride salt (130 mg, 0.23 mmol). After stirring for 2 hours, the reaction was partitioned between ethyl acetate and water. The organic layer was washed successively with water, 0.1 N NaOH and brine, dried over sodium sulfate and concentrated. To separate diastereomers, preparative HPLC-MS was used: Waters mass directed fractionation system, Waters SunFire C18 column, 5 μm particle size, 30×100 mm, mobile phase system: Aq(0.1% TFA)/AcN; flow rate 60 mL/min; separating gradient: 30.4-48.4% B in 12 minutes. Peak 1 (first to elute), and Peak 2 (second to elute) were subjected to the deprotection step separately: stirred for 1 hour in a 1:1 mixture of DCM/TFA, concentrated, then stirred for 30 minutes in a solution of ethylenediamine (0.2 mL) in methanol (1.5 mL). Purification via preparative HPLC-MS, eluting with a gradient of MeCN/H2O containing 0.15% NH4OH afforded product (peak 1: 19 mg, peak 2: 23 mg, total 36% of theoretical). Peak 1: 1H NMR (400 MHz, d6-dmso) (rotamers): δ 12.11 (br s, 1H), 8.81 (d, 1H), 8.67 (s, 1H), 8.36 (s, 1H), 7.99-7.93 (m, 1H), 7.79-7.68 (m, 1H), 7.62-7.54 (m, 2H), 6.97 (dd, 1H), 4.93-4.83 (m, 1H), 3.85-3.66 (m, 1H), 3.36-2.08 (m, 10H), 0.90 and 0.71 ((each as d) together=3H); LCMS (M+H)+: 498.1. Peak 2: 1H NMR (400 MHz, d6-dmso) (rotamers): δ □12.08 (br s, 1H), 8.82 (d, 1H), 8.68 (d, 1H), 8.37 (d, 1H), 8.01-7.93 (m, 1H), 7.80-7.71 (m, 1H), 7.63-7.54 (m, 2H), 6.98 (t, 1H), 4.94-4.81 (m, 1H), 3.87-2.26 (m, 11H), 1.05 and 0.87 ((each as d) together=3H); LCMS (M+H)+: 498.1.
