Abstract 2482A: ARHI (DIRAS3) downregulates Ras-MAPK signaling and inhibits transformation of NIH3T3 cells through direct interaction with GTP-bound Ras

Abstract

Abstract Somatic mutation of Ras was the first genetic alteration documented to drive human oncogenesis. Activating mutations of H-Ras, N-Ras, or K-Ras are associated with 30% of all human cancers. Characterization of wild type Ras proteins has identified their essential role in intra-cellular signaling networks controlling cell proliferation, differentiation and survival. Despite intensive efforts, effective targeted therapy directed against mutated Ras has remained elusive. Regulation of Ras function has been studied in depth, but relatively little attention has been paid to physiological inhibitors of Ras. Our group has previously reported that the maternally imprinted Ras-related tumor suppressor gene ARHI (DIRAS3) is downregulated in more than 60% of ovarian cancers and several other tumor types. Re-expression of ARHI at physiologic levels can inhibit ovarian cancer cell proliferation, block migration, induce autophagy, mediate tumor dormancy and block both PI3K and Ras-MAPK signaling. ARHI shares 50-60% homology with the H- and K-Ras genes, differing primarily in a 34-amino acid N-terminal extension that is required for most of ARHI's functions. The mechanism by which ARHI regulates Ras-MAPK signaling is not fully understood. Here, we show that re-expression of ARHI in NIH3T3 murine fibroblasts inhibits cell proliferation, suppresses clonogenic growth and blocks cell transformation by constitutively active mutant H-Ras. Addition of the N-terminal extension of ARHI to mutant H-Ras also prevents malignant transformation. ARHI interacts directly with mutated Ras on co-immunoprecipitation and can inhibit GTP-bound Ras activity following overexpression of the constitutively active construct. We hypothesize that inhibition of the Ras-MAPK signaling pathways following re-expression of ARHI (DIRAS3) may be, in part, the effect of a direct interaction between GTP-bound Ras and ARHI, resulting in the restoration of intrinsic Ras GTPase activity. Our results suggest that ARHI (DIRAS3) may serve as a novel inhibitor of activated Ras, and re-expression of ARHI in tumors exhibiting an oncogenic mutation of Ras might permit more specifically targeted therapy. Citation Format: Margie N. Sutton. ARHI (DIRAS3) downregulates Ras-MAPK signaling and inhibits transformation of NIH3T3 cells through direct interaction with GTP-bound Ras. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2482A. doi:10.1158/1538-7445.AM2014-2482A