Abstract 2736: PIM2 is up-regulated in response to metformin-induced cell death triggered by ER stress/UPR in ALL lymphoblasts
Authors: Leclerc Gilles M. , Leclerc Guy J. , Kuznetsov Jeffim N. , Barredo Julio C. 
Published: August 2012

Abstract
Acute lymphoblastic leukemia (ALL) is the most common malignancy in children, and despite significant overall improvements in cure rates, outcome for patients diagnosed with resistant phenotypes or those who relapse is dismal. We investigated the mechanism of cell death induced by metformin (MET) in ALL cell models. We showed that MET induced significant growth inhibition and apoptosis in CCRF-CEM (T-ALL) and NALM6 (Bp-ALL) cells. Western blots revealed that MET activated p-AMPK, p-ACC, p-Akt, and p-4EBP1. The latter suggested that regulation of protein translation may be an important determinant in MET-induced cell death. Indeed, inhibition of mTOR/protein translation with rapamycin (RAPA) rescued MET-induced cell death in ALL cells. In addition, knockdown of AMPK expression in ALL cells using shRNA (shAMPK) abrogated MET-induced apoptosis as compared to control cells expressing scramble shRNA (shCTRL), indicating that AMPK mediated MET's cytotoxicity. Western blots demonstrated that shAMPK cells expressed lower level of total AMPK, p-p38MAPK, p-mTOR, and p-4EBP1 compared to shCTRL cells, indicating that AMPK and protein translation are critical in MET sensitivity. Indeed, pulse labeled 35S-methionine experiments demonstrated increase incorporation confirming the importance of protein synthesis is MET-induced cytotoxicity. More important, we uncovered that MET-induced apoptosis correlated with induction of ER stress as evidenced by up-regulation of IRE1α and CHOP although GRP78 decreased significantly. We further demonstrated that RAPA rescued MET-treated cells by relieving ER stress/UPR mediated cell death. We previously showed that the unfolded protein response (UPR) in ALL cells is regulated by the contextual crosstalk between AMPK and Akt (Mol Cancer Ther 10:437, 2011). Therefore, we evaluated the effects of co-targeting Akt and AMPK using the Akt inhibitor X/perifosine + MET and found these combinations to be synergistic. To further investigate the relationship between protein translation and ER stress/UPR in MET-induced cell death, we examined the role of PIM1/2 kinases. We found that expression of PIM2 was increased in MET-treated ALL with concomitant decreased in the expression of IRE1α, ATF6, and CHOP, suggesting that PIM2 maybe up-regulated as a compensatory survival mechanism aimed at relieving MET-induced ER stress/UPR mediated cell death. To test this hypothesis, we co-treated ALL cells with a PIM1/2 kinase inhibitor + MET and found that PIM2 inhibition synergistically sensitized ALL cells to MET (CI=0.28). Taken together, our data indicate that PIM2 plays a role in buffering cell death in MET-treated cells, and that regulation of protein translation modulates ER stress/UPR induced apoptosis in ALL cells. Consequently, our data support strategies targeting these synthetic lethal interactions as suitable for clinical translation in patients with ALL.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2736. doi:1538-7445.AM2012-2736