Abstract Purpose: The aim of our study is to evaluate the preclinical therapeutic activity and mechanism of action of BEZ235, a dual PI3K/mTOR inhibitor, in combination with dexamethasone in acute lymphoblastic leukemia (ALL). Experimental Design: The cytotoxic effects of BEZ235 and dexamethasone as single agents and in combination were assessed in a panel of ALL cell lines and xenograft models. The underlying mechanism of BEZ235 and dexamethasone was evaluated using immunoblotting, TaqMan RT-PCR, siRNA, immunohistochemistry, and immunoprecipitation. Results: Inhibition of the PI3K/AKT/mTOR pathway with the dual PI3K/mTOR inhibitor BEZ235 enhanced dexamethasone-induced anti-leukemic activity in in vitro (continuous cell lines and primary ALL cultures) and systemic in vivo models of T-ALL (including a patient-derived xenograft). Through inhibition of AKT1, BEZ235 was able to alleviate AKT1-mediated suppression of dexamethasone-induced apoptotic pathways leading to increased expression of the proapoptotic BCL-2 protein BIM. Downregulation of MCL-1 by BEZ235 further contributed to the modulation of dexamethasone resistance by increasing the amount of BIM available to induce apoptosis, especially in PTEN-null T-ALL where inhibition of AKT only partially overcame AKT-induced BIM suppression. Conclusions: Our data support the further investigation of agents targeting the PI3K/mTOR pathway to modulate glucocorticoid resistance in T-ALL. Clin Cancer Res; 22(3); 621–32. ©2015 AACR.