Reaction Steps
To a solution of (6R,6aS,14aR)-1,6,8,14a-tetrahydroxy-11-((3R,4R,5R,6S)-4-hydroxy-3,5-dimethoxy-6-methyltetrahydro-2H-pyran-2-ylamino)-6a-methoxy-3-methyl-7,9,12,14-tetraoxo-5,6,6a,7,9,12,14,14a-octahydrobenzo[a]tetracene-2-carboxylic acid (60 mg, 0.09 mmol) in THF (2 mL) was added polystyrene-carbodiimide (156 mg, 0.18 mmol), 1-hydroxybenzotriazole hydrate (24 mg, 0.18 mmol) and 3-(aminomethyl)pyridine (11 mg, 0.10 mmol). The reaction mixture was stirred at room temperature under nitrogen for 7 h. The reaction was filtered through a micro filter and concentrated. The crude material was purified by preparative HPLC (10:90 acetonitrile/water to 60:40 acetonitrile/water with 0.05% TFA over 10 min, then isocratic) to afford a dark red solid. The solid was dissolved in methanol (2 mL) and treated with HCl (0.20 mL, 1.25 M solution in methanol) at 0° C. The solution was concentrated and triturated with diethyl ether to afford 6R,6aS,14aR)-1,6,8,14a-tetrahydroxy-11-((3R,4R,5R,6S)-4-hydroxy-3,5-dimethoxy-6-methyltetrahydro-2H-pyran-2-ylamino)-6a-methoxy-3-methyl-7,9,12,14-tetraoxo-N-(pyridin-3-ylmethyl)-5,6,6a,7,9,12,14,14a-octahydrobenzo[a]tetracene-2-carboxamide hydrochloride as a mixture of diastereomers (12 mg, 21%) and as a red solid: 1H NMR (500 MHz, CD3OD) δ 8.84-8.80 (m, 1H), 8.80-8.65 (m, 1H), 8.64-8.58 (m, 1H), 8.10 (s, 1H), 8.09-8.06 (m, 1H), 6.63 (s, 1H), 6.16 (s, 0.3H), 6.01 (s, 0.7H), 5.20 (s, 0.3H), 4.95 (s, 0.7H), 4.67 (br s, 1H), 3.92-3.89 (m, 1H), 3.75 (s, 3H), 3.70-3.67 (m, 0.3H), 3.64-3.62 (m, 0.7H), 3.57 (s, 3H), 3.54 (s, 1H), 3.50 (s, 1H), 3.49-3.37 (m, 5H), 3.24-3.17 (m, 1H), 3.10-3.02 (m, 1H), 2.23 (s, 3H), 1.28 (d, J=6.5 Hz, 2H), 1.23 (d, J=6.5 Hz, 1H); MS (ESI−) m/z 772 (M−H); HPLC 96.7% (AUC), tR 10.53 min.
