Reaction Steps
N-(4-((1R,3S)-3-aminocyclohexyl)pyridin-3-yl)-2-(2,6-difluorophenyl)imidazo[1,5-b]pyridazin-7-amine bis(2,2,2-trifluoroacetate) and N-(4-((1S,3R)-3-aminocyclohexyl)pyridin-3-3-yl)-2-(2,6-difluorophenyl)imidazo[1,5-b]pyridazin-7-amine bis(2,2,2-trifluoroacetate (1/1). A dry, 5 mL microwave pressure vessel was charged with 2-((1R,3S)-3-(3-((2-(2,6-difluorophenyl)imidazo[1,5-b]pyridazin-7-yl)amino)-4-pyridinyl)cyclohexyl)-1H-isoindole-1,3(2 H)-dione bis(2,2,2-trifluoroacetate) and 2-((1S,3R)-3-(3-((2-(2,6-difluorophenyl)imidazo[1,5-b]pyridazin-7-yl)amino)-4-pyridinyl)cyclohexyl)-1H-isoindole-1,3(2 H)-dione bis(2,2,2-trifluoroacetate) (1/1, 86 mg, 0.06 mmol), a stirbar, EtOH (3 mL) and methylamine (33% wt. solution in absolute EtOH) (0.07 mL, 0.55 mmol). The vessel was crimped with a PTFE-lined seal and placed in a 45° C. heat transfer block for 3 h. The solution was concentrated under a stream of N2 to a volume of 1 mL. The solution was treated with hydrazine, anh. (3.47 μl, 0.11 mmol), and the vessel was crimped with a PTFE line seal. The vessel was placed into a 45° C. heat transfer block for 2 h and was cooled to RT. The solution was concentrated in vacuo. The crude material was purified using prep HPLC method 1 to afford N-(4-((1R,3S)-3-aminocyclohexyl)pyridin-3-yl)-2-(2,6-difluorophenyl)-imidazo[1,5-b]pyridazin-7-amine bis(2,2,2-trifluoroacetate) and N-(4-((1S,3R)-3-aminocyclohexyl)pyridin-3-yl)-2-(2,6-difluorophenyl)imidazo[1,5-b]pyridazin-7-amine bis(2,2,2-trifluoroacetate (1/1) (33 mg, 0.03 mmol, 46% yield). MS (ESI, pos. ion) m/z: 421.0 (M+1). 1H NMR (400 MHz, DMF-d6) δ ppm 1.51 (m, J=10.76 Hz, 2 H) 1.62 (q, J=11.70 Hz, 1 H) 1.85 (q, J=12.00 Hz, 1 H) 1.95 (m, J=6.00, 2.90 Hz, 1 H) 2.02 (d, J=9.78 Hz, 1 H) 2.20 (d, J=11.35 Hz, 1 H) 2.43 (d, J=11.93 Hz, 1 H) 3.44 (q, J=10.40 Hz, 2 H) 7.01 (br. s, 2 H) 6.85 (d, J=9.39 Hz, 1 H) 7.36 (t, J=8.31 Hz, 2 H) 7.47 (s, 1 H) 7.65 7.77 (m, 1 H) 7.72 (tt, J=8.40, 6.70 Hz, 1 H) 8.25 (d, J=9.39 Hz, 1 H) 8.42-8.65 (m, 4 H) 9.02 (br. s., 1 H) 9.07 (br. s., 1 H). 19F NMR (376 MHz, DMF-d) δ ppm −113.30 (t, J=7.15 Hz, 2 F) −74.60 (br. s., 9 F). The DMF from the NMR sample was concentrated in vacuo, and the sample was dried in a vacuum oven (60° C., <1 mm Hg) overnight. Approximately 6 mg was analyzed in D2O to check whether the TFA—aromatic fluorine ratio changed. Sample was a bis-TFA salt by 19F NMR. 1H NMR (400 MHz, D2O) δ ppm 1.38-1.59 (m, 3 H) 1.69 (q, J=11.93 Hz, 1 H) 2.00 (d, J=10.56 Hz, 2 H) 2.10 (d, J=11.15 Hz, 1 H) 2.29 (d, J=11.74 Hz, 1 H) 3.20 (t, J=11.44 Hz, 1 H) 3.35 (t, J=11.15 Hz, 1 H) 6.95 (d, J=9.59 Hz, 1 H) 7.12 (t, J=8.41 Hz, 2 H) 7.54 (tt, J=8.60, 6.50 Hz, 1 H) 7.53 (s, 1 H) 7.90 (d, J=6.06 Hz, 1 H) 8.12 (d, J=9.59 Hz, 1 H) 8.29 (s, 1 H) 8.32 (d, J=6.06 Hz, 1 H). 19F NMR (376 MHz, D2O) δ ppm −117.58 (t, J=7.15 Hz, 2 F) −78.50 (s, 6 F).
