To a solution of 4-(3-amino-6-((1S,3S,4S)-3-fluoro-4-hydroxycyclohexyl)pyrazin-2-yl)-2-fluorobenzoic acid (20 mg, 0.057 mmol) in DMF (573 μL) was added (S)-2-amino-2-(3-bromo-5-fluorophenyl)ethanol HCl salt (18.59 mg, 0.069 mmol), aza-HOBt (11.69 mg, 0.086 mmol), EDC (21.95 mg, 0.115 mmol), and DIEA (30.0 μl, 0.172 mmol). The reaction mixture was stirred for 15 h. Water was added, and the reaction mixture was extracted with EtOAc three times. The organic layers were dried over Na2SO4, filtered and concentrated in vacuo. The crude product was purified by HPLC. Pure fractions were lyophilized to provide 4-(3-amino-6-((1S,3S,4S)-3-fluoro-4-hydroxycyclohexyl)pyrazin-2-yl)-N—((S)-1-(3-bromo-5-fluorophenyl)-2-hydroxyethyl)-2-fluorobenzamide (12.9 mg, 0.019 mmol, 32.8%) as a TFA salt. LCMS (m/z): 565.1/567.1 (MH+), 0.72 min; 1H NMR (400 MHz, CD3OD) δ ppm 7.78 (s, 2H), 7.63-7.47 (m, 2H), 7.37 (s, 1H), 7.22-7.02 (m, 2H), 5.15-5.03 (m, 1H), 4.43-4.15 (m, 1H), 3.77 (t, J=5.7 Hz, 2H), 3.62-3.47 (m, 1H), 2.76 (br. s., 1H), 2.19 (dd, J=3.1, 6.3 Hz, 1H), 2.02-1.92 (m, 1H), 1.89-1.66 (m, 2H), 1.56 (dd, J=3.3, 12.3 Hz, 1H), 1.42 (br. s., 1H). The absolute stereochemistry was determined based on the information of X-ray co-structure of 4-(3-amino-6-((1S,3S,4S)-3-fluoro-4-hydroxycyclohexyl)pyrazin-2-yl)-N—((S)-1-(3-bromo-5-fluorophenyl)-2-hydroxyethyl)-2-fluorobenzamide.
