Fig 4. Combination treatment with ACY-241 and pomalidomide reduces S phase frequency.

H929 (A), U266 (B), and MM.1s (C) cells were treated with 1 μM or 0.05 μM pomalidomide or 3 μM ACY-241 alone or in combination for 3 days followed by incubation for 1 hour with EdU and stained for EdU incorporation and FxCycle Far Red to measure S phase frequency. Percent cells in S phase was determined by gating EdU positive cells. Representative flow dot plots are shown for each cell line (left) and the mean ± SD or three independent experiments is plotted (right). * p < 0.05, ** p < 0.01; *** p < 0.001. D) Cells treated as in A-C were harvested after 48 hours. Total protein was isolated and probed with antibodies for Myc, IRF4, IKZF3 and β-Actin. Results are representative of at least 3 independent experiments for each antibody.