Acute lymphoblastic leukemia (ALL) is the leading cause of cancer-related death in children and the relapse rate in adult ALL patients is about 50%, highlighting the need for new therapeutic strategies. Data from our laboratory and others showed that ALL cells are sensitive to drugs that induce endoplasmic reticulum (ER) stress/unfolded protein response (UPR). In search for novel strategies to target the ER stress/UPR in ALL, we tested the efficacy of the NEDD8-activating enzyme (NAE) inhibitor pevonedistat (MLN4924, pevo) in ALL. We found ALL cells exhibited significant in vitro and in vivo sensitivity to pevo-induced ER stress/UPR. Specifically, proteotoxic/ER stress was observed in pevo-treated ALL cells secondary to their inability to halt protein translation following pevo-induced activation of the mTOR pathway and concomitant de-phosphorylation of p-eIF2α (S51). In addition, aberrant activation of MEK/ERK has been correlated with resistance/relapse in pediatric ALL (Blood 2014; 124: 3420-3430). In our Bp- and T-ALL cell line models, we found consistent induction of p-ERK1/2 (T202/Y204) following pevo treatment, suggesting phosphorylation of ERK1/2 as a compensatory survival mechanism in response to pevo's cytotoxicity. Supporting this hypothesis, we observed significant in vitro synergy between the MEK inhibitor selumetinib (SEL) and pevo (CI = 0.017). On this basis, we tested the in vivo efficacy of pevo + SEL in NSG mice injected with NALM6 cells expressing the luciferase gene (NALM6/LUC). Engrafted NSG mice were treated with pevo (s.c., 66 mg/kg) and SEL (p.o., 50 mg/kg) twice daily on weekdays and once per day on weekends. Bioluminescence analysis of animals 21 days post ALL injection revealed significant reduction of tumor burden in mice treated with pevo alone or pevo + SEL (p<0.05 for both vs. control). Kaplan-Meier curves showed a significant survival advantage for mice treated with pevo + SEL compared with the control group (p<0.05). Mechanistic studies showed that pevo led to induction of NOXA and BIM whereas Mcl-1 levels were stabilized, suggesting sequestration of the pro-survival activity of Mcl-1 by NOXA/BIM. Indeed, co-IP analysis demonstrated that binding between NOXA and/or BIM with Mcl-1 was enhanced in pevo-treated ALL cells. Further, significant downregulation of Mcl-1 was observed in ALL cells co-treated with pevo + SEL. We found that the synergy of this combination was prevented by co-treatment with the pan-caspase inhibitor Z-VAD, but observed persistence of Mcl-1downregulation whereas BIM expression remained unchanged. We conclude that MEK/ERK pathway inhibition rebalances the Bcl-2 family proteins in favor of synergistic apoptotic death in ALL cells treated in vitro and in vivo with the NAE inhibitor pevonedistat. Our data supports further investigations of agents targeting NAE and the MEK/ERK pathway in relapsed/refractory ALL.

Citation Format: Shuhua Zheng, Gilles M. Leclerc, Guy J. Leclerc, Joanna DeSalvo, Ronan T. Swords, Julio C. Barredo. Rebalancing of Bcl-2 family proteins mediate the vulnerability of pevonedistat-treated acute lymphoblastic leukemia cells towards MEK/ERK pathway inhibition. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4547.