Effect of ACY738 treatment in human CLL cell lines. (A) HDAC6 protein expression quantified by flow cytometry in normal donor B cells (n = 5) and CLL cell lines; data were compiled from 3 independent experiments. (B) Growth kinetics of OSU-CLL in culture with ACY738 at 1 μM or vehicle for indicated period. (C) Cell-cycle analysis in OSU-CLL determined by Ki67/7AAD staining after 24 hours of indicated treatments. (D-E) Apoptosis was measured by annexin V/viable staining after treatment of OSU-CLL with ACY738 for 24 hours. (F) Phosphorylation of indicated molecules was measured after stimulation with 10 μg/mL of anti-human immunoglobulin M in CLL cell lines. (G) Expression of intracellular MCL1 and BCL-2 protein analyzed by flow cytometry, after incubating OSU-CLL with indicated concentrations of ACY738. All error bars correspond to standard deviations. *P < .05, **P < .005, ***P < .0005. DMSO, dimethyl sulfoxide.