Abstract 4692: IAP inhibitors prime childhood leukemia cells to chemotherapy-induced apoptosis in a strictly RIP1-dependent manner and exert anti-leukemic activity in a NOD/SCID mouse model in vivo
Authors: Löder Sandra , Fakler Melanie , Vanlangenakker Nele , Bertrand Mathieu , Vandenabeele Peter , Jeremias Irmela , Debatin Klaus-Michael , Fulda Simone 
Published: June 2012

Abstract
Defects in apoptosis contribute to poor outcome in high risk pediatric acute lymphoblastic leukemia (ALL), calling for novel strategies that counter apoptosis resistance. “Inhibitor of Apoptosis” (IAP) proteins present promising targets, since they are expressed at high levels in acute leukemia.
In the present study, we investigated the effect of small molecule IAP inhibitors alone and in combination with chemotherapeutics in ALL cell lines, primary leukemic blasts, normal peripheral blood lymphocytes and in a mouse model of pediatric ALL.
Here, we report that IAP inhibitors at subtoxic concentrations cooperate with various anticancer drugs (i.e. AraC, Gemcitabine, Cyclophosphamide, Doxorubicin, Etoposide, Vincristine, Taxol) to induce apoptosis in ALL cells in a highly synergistic manner as calculated by combination index (CI index 0.3). Also, IAP inhibitors act in concert with AraC to reduce clonogenic survival, thus demonstrating also a long-term anti-leukemic effect. Importantly, we identify RIP1 as a critical regulator of this synergism of IAP inhibitors and AraC that mediates the formation of a RIP1/FADD/caspase-8 complex via an autocrine/paracrine loop of TNFα. Knockdown of RIP1 abolishes formation of this complex and subsequent activation of caspase-8 and -3, mitochondrial perturbations and apoptosis. Similarly, pharmacological inhibition of RIP1 kinase activity by Necrostatin-1 inhibits IAP inhibitor- and AraC-triggered interaction of RIP1, FADD and caspase-8 and apoptosis. In addition, blockage of TNFα by a TNFα-neutralizing antibody profoundly attenuates the cooperativity of IAP inhibitor and AraC to induce formation of the RIP1/FADD/caspase-8 complex and apoposis. Of note, IAP inhibitors potently trigger cell death in leukemic blasts from children with ALL ex vivo and also sensitize primary leukemia cells for chemotherapy-induced cell death. In contrast to malignant cells, IAP inhibitors and AraC at equimolar concentrations are non-toxic to both resting and activated normal peripheral blood lymphocytes, pointing to a therapeutic window. Most importantly, IAP inhibitors exert potent anti-leukemic activity in vivo in a mouse model of pediatric ALL engrafted in NOD/SCID mice.
In conclusion, our findings provide first evidence that IAP inhibitors present a promising strategy to prime childhood ALL cells for chemotherapy-induced apoptosis in a RIP1-dependent manner. These data have important implications for the development of apoptosis-targeted therapies in childhood leukemia.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4692. doi:10.1158/1538-7445.AM2011-4692