mTOR Inhibition by Everolimus in Childhood Acute Lymphoblastic Leukemia Induces Caspase-Independent Cell Death
Figure 5
Cell death is dependent on new protein synthesis but not JNK signalling.
(A) NALM6 cells were treated for the indicated time with 16 µM everolimus and cell lysates prepared. Lysates were probed for phosphorylated JNK (p-JNK) and loading assessed using β-actin. (B) NALM6 cells were treated with the indicated concentrations of everolimus for 16 h, with or without a 1 h pre-incubation with the JNK inhibitor SP600125 (5 µM) and viability assessed using annexin V/PI staining. The percentage of viable cells is shown with the bars representing the mean ± SE of 2 experiments. (C) NALM6 or REH cells were incubated with indicated concentrations of cyclohexamide for 1 h prior to the addition of the indicated concentration of everolimus. The mean ± SD of ≥3 independent experiments is shown. *p<0.05 when comparing cells with and without cyclohexamide. (D) NALM6 cells were treated with the indicated concentrations of everolimus for the specified times and cell lysates prepared. The indicated proteins were assessed by Western blotting and where indicated the bands were quantitated by densitometry using β-actin for normalization.