Abstract C72: Counteracting autophagy allows to overcome resistance to the mTOR everolimus in mantle cell lymphoma.

Abstract

Abstract Background: Mantle cell lymphoma (MCL) is an aggressive B-cell lymphoid neoplasm genetically characterized by the t(11;14)(q13;q32) leading to the overexpression of cyclin D1, which causes cell cycle deregulation at the G1-S phase transition. As a consequence of its poor response to conventional chemotherapy and relatively short patient survival, new therapeutic strategies are required. The phosphatidylinositol 3-kinase/Akt/mTOR survival pathway is constitutively activated in MCL, thereby making the mTOR inhibition a very attractive strategy for its treatment. The first clinical studies conducted in relapsed MCL patients have reported a significant, but relative short duration of response to the oral rapamycin derivative everolimus (RAD001). In this context, our aim was to analyze the mechanism of action related to resistance/sentitivity of this drug in MCL cells, and to propose further combination strategies. Methods: The sensitivity to everolimus was analyzed in MCL primary cultures, MCL cell lines and peripheral blood lymphocytes from healthy donors. Cells were treated for up to 72h with increasing doses of everolimus, followed by cytotoxicity quantification, cell-cycle determination and western blot analysis of the PI3K/Akt/mTOR pathway. Autophagy was studied by LC3I/II expression and Cell-ID™ Green staining of autophagolysosomes by flow cytometry and fluorescence microsopy. Results: Everolimus exerted heterogeneous antitumoral effect on MCL cells, while sparing normal cells. This effect was associated with G1 cell-cycle arrest and reduced phosphorylation of the mTOR downstream targets, 4E-BP1 and S6. However, Akt re-phosphorylation was observed after 48h of everolimus treatment, suggesting feedbacks loops within the mTOR signaling pathway. Accordingly, everolimus combination studies with the Akt inhibitor VIII, isozyme-selective (Akti-1/2) showed synergistic cytotoxic effect in MCL cells and overcame Akt mechanism of resistance in a sub-set of MCL samples. When compared to these samples, increased levels of autophagic hallmarks such as autophagic vacuoles and LC3 processing were found in everolimus/Akti-1/2 low-responsive cells. In these cells, the combination with the autophagy inhibitor hydroxychloroquine overcame the resistance to mTOR/Akt inhibitors, and efficiently induced apoptosis, thus suggesting a prosurvival role of autophagy. Conclusions: This study suggests that targeting autophagy as anticancer therapy may represent an attractive sensitizing strategy for everolimus-based therapy in MCL. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C72.