Selective inhibition of the ML2 cell line by EPZ004777. (A) Effect of EPZ004777 on the proliferation of leukemia cell lines bearing an MLL-AF6 (ML2) or MLL-AF9 (MOLM-13) fusion or a cell line lacking an MLL rearrangement (HL60). Cell lines were maintained in the presence of increasing concentrations of EPZ004777: 0.1, 1, and 10 μM. (B) Immunoblots showing the levels of H3K79me2 upon exposure of the different cell lines to indicated concentrations of EPZ004777 or dimethylsulfoxide (DMSO) carrier controls. (C) Cell cycle changes (Hoechst staining) in ML2 cell line upon treatment with 10 μM EPZ004777 for 0, 4, 8, or 10 days. Similar results were obtained in 2 independent experiments, each having 2 technical replicates. (D) Percentage of Annexin V-positive cells plotted 10 d after treatment with 10 μM of EPZ004777 or DMSO control. (E) Time course of HOXA9 and HOXA10 mRNA expression in the ML2 cell line for 7 days of incubation with 10 µM EPZ004777 as measured by quantitative real-time PCR. Expression levels were normalized to GAPDH and expressed relative to those at d 0 (set to 100%). Error bars represent the standard error of the mean (n = 3 independent experiments).