Abstract 1325: DIRAS3 (ARHI) is required for amino acid-mediated autophagy and nutrient deprivation in dormant ovarian cancers

The persistence of dormant, drug-resistant cancer cells after cytoreductive surgery and combination chemotherapy is a major challenge that contributes to poor outcomes for ovarian cancer patients. Despite normalization of CA125 and negative imaging following primary treatment, “second look” surgery can detect small, quiescent, poorly vascularized nodules of persistent ovarian cancer in ~50% of patients. After positive second look operations, persistent ovarian cancer may take months or years to become clinically evident, consistent with tumor dormancy. Autophagy is one mechanism by which persistent, potentially dormant cancer cells could survive in a nutrient poor environment. Autophagy or “self-eating” is a catabolic process by which organelles and misfolded proteins are degraded and recycled to provide energy that could protect dormant cancer cells from acute starvation in the face of an inadequate blood supply. Autophagy can be induced by multiple mechanisms. Our group has found that, in contrast to primary ovarian cancers, the cancer cells in >80% of positive second look biopsies exhibit autophagy and express DIRAS3 (ARHI). DIRAS3 is an imprinted tumor suppressor gene that is downregulated in >60% of primary ovarian cancers by multiple mechanisms including loss of heterozygosity, CpG promoter methylation, transcriptional repression (E2F1/4) and miRNA regulation (miR-221/222). Re-expression of DIRAS3 prevents ovarian cancer growth, inhibits motility, induces autophagy and establishes tumor dormancy in xenografts. When DIRAS3 is upregulated in human ovarian carcinoma cell lines from a doxycycline-inducible promoter in nu/nu murine xenografts cancer cells remain dormant until DIRAS3 is downregulated permitting cell cycling, vascularization and progressive growth. Autophagy is required for survival of dormant xenografts in that treatment with chloroquine, a functional inhibitor of autophagy, markedly delays outgrowth of ovarian cancer xenografts when DIRAS3 is downregulated. Mechanisms by which DIRAS3 is upregulated and autophagy induced in positive second look specimens remains poorly understood. We have found that amino acid deprivation upregulates DIRAS3 and induces autophagy. Knockdown of DIRAS3 markedly decreases induction of autophagy following nutrient deprivation. We have identified that amino acid deprivation results in decreased E2F1/E2F4 expression and transcriptional upregulation of DIRAS3, but changes in promoter methylation or miRNA regulation are not observed. Knockdown of E2F1 or E2F4, both genetically and pharmacologically increase DIRAS3 expression and autophagy. Taken together these results suggest that nutrient deprivation results in transcriptional upregulation of DIRAS3-mediated autophagy and thus likely presents a mechanism by which DIRAS3 is upregulated in dormant, second look ovarian cancer tumor specimens. Citation Format: Margie Nicole Sutton, Gilbert Y. Huang, Jinhua Zhou, Zhen Lu, Robert C. Bast. DIRAS3 (ARHI) is required for amino acid-mediated autophagy and nutrient deprivation in dormant ovarian cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1325.