Co-operative induction of apoptosis using A-1210477.

A. Cells were co-incubated with 1 μM A-1210477 (477) and with 10 nM ABT-199 (199), 10nM pladienolide B (PLB), 1 μM torin1, 1 μM etoposide or 10 nM AC220 for 4 hours. Alternatively, cells were incubated with JQ1 for 48 hours and A-1210477 was added for the final four hours of the incubation. Cells were then fixed and processed for Cytochrome C release. (Mean+/- SD for n = 3). B. Cells were incubated with 1 μM A-1210477 for four hours and 10 nM ABT-199 was added either before, after or concurrently (final 2 hours). In the two-step conditions, cells were pelleted and rinsed twice in RPMI at 4°C in between agents. R10 = medium without drug.