Abstract 3964: NVP-BEZ235 enhances dexamethasone-induced BIM expression and apoptosis in models of T-ALL with PTEN dysfunction and increased PI3K/AKT activity
Authors: Hall Connor , Kang Min 
Published: May 2015

Introduction: Glucocorticoid (GC) resistance presents a major challenge in pediatric acute lymphoblastic leukemia (ALL) treatment. Increased activation of the PI3K/AKT pathway, which may occur following loss of PTEN function, may be a possible mechanism of GC resistance. We therefore investigated the effects of inhibiting the PI3K/AKT pathway with the dual PI3K/mTOR inhibitor NVP-BEZ235 on the enhancement of dexamethasone activity in in vitro and in vivo models of ALL and the role the PI3K/AKT pathway plays in resistance to GCs.

Methods: Cytotoxicity of NVP-BEZ235, dexamethasone, and the combination were evaluated using DIMSCAN in in vitro models of T- and B-ALL and combination indices (CI) were calculated using CalcuSyn. Apoptotic activity for each single agent and the combination was determined by flow cytometry. Protein and mRNA expression were measured by immunoblotting and RT-PCR, respectively. Knockdown of BIM was accomplished using siRNA. In vivo effects were assessed following 28 days of treatment using NOD/SCID mice systemically engrafted with T-ALL model COG-LL-317h.

Results: The combination of dexamethasone plus NVP-BEZ235 showed strong synergistic activity (CI &lt; 0.3) in 6/6 T-ALL models tested, while such activity was seen in only 1/6 B-ALL models. Synergistic cytotoxicity corresponded with increased AKT phosphorylation. A systemic xenograft model of COG-LL-317h also showed significant increase in event-free survival in combination treated mice compared to vehicle control mice (P &lt; 0.01), while there was no significant increase in survival with mice treated with either agent alone. In COG-LL-317h, the percentage of apoptotic cells were significantly greater in cells treated with the combination (24.8 ± 1.0%) compared with vehicle control (2.1 ± 0.2%, P &lt; 0.0001), dexamethasone (4.4 ± 0.3%, P &lt; 0.0001), or NVP-BEZ235 (2.0 ± 0.2%, P &lt; 0.0001). In COG-LL-317h, the pro-apoptotic Bcl-2 family protein BIM was substantially increased at both the mRNA and protein levels when treated with the combination compared with either agent alone, while the B-cell ALL RS4;11 did not show any enhancement with the combination. Compared with non-targeting siRNA, transfection with siRNA against BIM resulted in significant reduction of cells undergoing apoptosis (39.3 ± 3.3% vs. 20.3 ± 0.7%, P=0.0007) and reduced caspase activation in response to the combination.

Conclusion: These data suggest activation of the PI3K/AKT pathway following PTEN loss plays a significant role in reducing GC responses and use of the combination may offset that effect by increasing BIM expression and apoptosis. Our results show that NVP-BEZ235 may provide a new adjunct for ALL patients with hyperactivated PI3K/AKT pathway by enhancing dexamethasone activity.

Citation Format: Connor Hall, Min Kang. NVP-BEZ235 enhances dexamethasone-induced BIM expression and apoptosis in models of T-ALL with PTEN dysfunction and increased PI3K/AKT activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3964. doi:10.1158/1538-7445.AM2014-3964