JQ1 treatment inhibits the growth of pediatric T-ALL cells in vitro. (A-B) Treatment of pediatric T-ALL cells are sensitive to GSI or JQ1 treatment. (A) Diagnostic, relapsed, or IF pediatric T-ALL cells were cultured with vehicle, GSI (1 μM), or increasing concentrations of JQ1 (100-1000 nM) for 5 days and growth and metabolism assayed by MTS. The absorbance levels were normalized to the vehicle control and the GI₅₀ of each cell line was calculated using Graph Pad Prism 5 software. The results are averages of 3 to 5 independent experiments and error bars represent SEM. (B) Absorbance values determined by MTS assay are plotted for each patient sample treated with DMSO, GSI (DBZ, 1 μM), or JQ1 (1 μM). The results are averages of 3 to 5 independent experiments and error bars represent SEM. P < .0001 for all patient samples treated with JQ1; *P < .05, **P < .01, ***P < .001. (C) JQ1 treatment reduces C-MYC expression in diagnostic, relapsed, or IF patient T-ALL samples. Primary leukemic cells from patients were cultured with DMSO, GSI (DBZ, 1 μM), or JQ1 (1 μM) for 24 hours and C-MYC expression levels were measured using quantitative real-time PCR. C-MYC expression was normalized to β-ACTIN and calculated using the ΔΔCT method. The results are averages of 3 independent experiments and error bars represent SEM. P < .001 for all samples treated with JQ1; *P < .05, ***P < .001. (D-E) JQ1 treatment of some diagnostic and relapsed/IF T-ALL samples results in apoptosis. (D) Patient samples TALL-x-13 (relapse) and TALL-x-9 (diagnostic) were treated with DMSO, GSI (DBZ, 1 μM), or JQ1 (1 μM) for 96 hours and apoptotic cells detected by staining with Annexin V and 7AAD followed by flow cytometry. A representative fluorescence-activated cell sorter plot is shown. (E) Collated data of Annexin V–positive cells from relapsed/IF patients treated with DMSO, GSI (DBZ, 1 μM), or JQ1 (1 μM) for 96 hours. The results are averages of 3 independent experiments and error bars represent SEM. *P < .05, ***P < .001.