Ampk Activation Vs. Inhibition Induces Apoptosis In Acute Lymphoblastic Leukemia Cells By Differentially Altering Pi3k/Akt/Mtor Or Ras/Craf/Mek/Erk Signaling

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC Acute Lymphoblastic Leukemia (ALL) is the most common hematological malignancy and the main cause of cancer-related deaths in children. Therefore, search for novel treatment strategies is warranted. We identified AMP activated protein kinase (AMPK), a master regulator of bioenergetics, as a potential target for ALL therapy due to its effects on cell proliferation and cell cycle regulation, as well as its crosstalk with critical metabolic and oncogenic pathways. We demonstrated that treatment of NALM6 (Bp-ALL) and CEM (T-ALL) cells with AICAR, an AMPK activator, induced growth inhibition and apoptosis. Using metformin, another AMPK agonist, we found 40% growth inhibition, and up to five- and three-fold greater induction of apoptosis relative to controls in CEM and NALM6, respectively. Unexpectedly, rescue experiments with AMPK inhibitors Ara-A and compound-C (CC) failed to abrogate the cytotoxic effects induced by AICAR. When used alone, Ara-A induced 60% and 40% cell death in NALM6 and CEM cells, respectively, whereas CC induced 75- and 15-fold more apoptotic death relative to controls. To investigate the mechanism by which AMPK activation vs. inhibition induced apoptosis, we determined levels of P-AMPK (T172) and factors associated with the PI3K/Akt/mTOR and RAS/cRAF/Erk signaling pathways in NALM6 and CEM cells treated with either CC, AICAR, or in combination. Our data show that P-AMPK levels were decreased by CC and increased by AICAR. Additional Western blots demonstrated that these agents exerted opposite effects on Akt and RAS signaling. CC decreased P-Akt (S473) and activated the RAS pathway, while AICAR increased P-Akt. We showed that activation of Akt by AICAR down-regulated the RAS pathway via phosphorylation of cRAF (S259). P-mTOR (S2448) and P-4EBP1 (T70) exhibited a greater decrease in cells treated with CC + AICAR as compared to each agent alone. A significant decrease in P-Akt was also detected in cells treated with both agents vs. each drug alone. Together, our data indicate that AICAR and CC induce cell death in ALL cells by two different mechanisms mediated by AMPK: AICAR-activation of AMPK inhibited the RAS-dependent cell proliferation pathway, and CC-inhibition of AMPK by down-regulating the Akt cell survival pathway. These results suggest that alterations in AMPK signaling may regulate the cross-talk between the PI3K/Akt/mTOR and RAS/cRAF/Erk cascades and may dictate the fate of ALL cells by regulating apoptosis after exposure to agents targeting these pathways. Experiments co-targeting AMPK and Akt using AICAR and Akt-inhibitor X, respectively, induced synergistic growth inhibition in CEM (CI=0.90) and NALM6 (CI=0.85) cells compared to each drug alone. These findings provide a rationale for simultaneously targeting AMPK and key signaling factors associated with either PI3K/Akt/mTOR or RAS/cRAF/Erk pathways in ALL. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5089.