Fig. 2: BRD4 regulates CD44/CD44v8-10 expression and modulates oxidative stress and oxidative phosphorylation in T-ALL.

KOPT-K1 and SUP-T1 cell lines were treated with ARV-825 (50 nM for KOPT-K1 and 20 nM for SUP-T1) for 0, 12, and 24 h. A RNA was extracted from cells, and qPCR analysis of CD44 and CD44v8-10 was performed. Gene expression was normalized to the corresponding 18 S rRNA expression level (n = 3). B KOPT-K1 (C) SUP-T1 cell treated with ARV-825 (50 nM) or 20 nM) as above for 24 h and cells were subjected to flow cytometry analysis of surface expression of CD44(left top panel) and CD98(Left bottom panel), as well, (Right top panel)-WCL of duplicate samples cells treated with ARV-825 for 24 h subjected to immunoblots of whole-cell lysates of KOPT-K1 and SUP-T1 cells with the indicated antibodies. (Right bottom panel) results of an assay performed to determine the level of total ROS generation by KOPT-K1 (B) and SUP-T1 (C) cells using an ENZ-51011 kit. D KOPT-K1, SUP-T1, and T-ALL PDX (6506870) treated with ARV-825 in 50 nM, 20 nM, and 100 nM for 24 h respectively subjected to a migration assay 4 h after incubation in media containing HA (150 ng). E CD44- and CD44v8-10–overexpressing KOPT-K1 and SUP-T1 cells (left top panel and bottom panel respectively) treated with ARV-825 (50 or 20 nM) for 24 and subjected to immunoblots with the indicated antibodies and duplicate samples were assayed total ROS (middle panel). Similar samples with 72 h treatment subjected to measure levels of apoptosis (right panel) using flow cytometry. Error bars represent SD from three different biological replicates (*p < 0.05, **p < 0.01, ***p < 0.001 compared to DMSO.