BEZ235 induces apoptosis of Eμ-Myc lymphomas and IG-cMYC-translocated cell lines at on-target concentrations. (A) Six independently derived Eμ-Myc lymphomas were cultured in vitro for 24 hours with increasing concentrations of BEZ235 (or dimethylsulfoxide [DMSO] vehicle alone at maximal concentration) and subjected to flow cytometric analysis for annexin-V/PI positivity. The lymphomas have been stratified from most to least BEZ235 sensitive (at 125 nM) from left to right. A unique identifier for each lymphoma is provided above the bar graph. (B) A representative lymphoma (#4242) was treated with BEZ235 or Everolimus for 24 hours prior to flow cytometric analysis for annexin-V/PI positivity. *P < .05 comparing treatments (BEZ235 vs Everolimus) at each concentration (2-way analysis of variance [ANOVA]). (C) The same lymphoma (#4242) was transduced with murine stem-cell virus expressing Bcl2 or empty vector control (MSCV) and treated for 24 hours with BEZ235 prior to flow cytometric analysis for annexin-V/PI positivity. *P < .05 comparing genotypes (Bcl2 vs empty vector) at each concentration (2-way ANOVA). (D) Lymphoma (#4242) was transduced with murine stem-cell virus expressing CrmA or empty vector control (MSCV) and treated for 24 hours with BEZ235 prior to flow cytometric analysis for annexin-V/PI positivity. There was no significant difference in response between genotypes (CrmA vs empty vector) at any drug concentration (P > .05; 2-way ANOVA). Inset: western blot of CrmA expression in the same lymphoma cells with β-actin provided as a loading control. (E) Human IG-cMYC-translocated cell lines were treated for 48 hours with BEZ235 (or DMSO vehicle) and subjected to flow cytometric analysis for annexin-V/PI positivity. (F) The same human cell lines were treated for 48 hours with Everolimus prior to flow cytometric analysis for annexin-V/PI positivity. All graphs represent the mean (± standard error of the mean [SEM]) for at least 3 independent experiments.