Abstract B12: The importance of the RASA1/R-Ras/Ral-A signaling axis in melanoma tumorigenesis

The Ras family of small GTP-binding proteins is frequently activated by mutations, including NRAS (20%), KRAS (2%), and HRAS (1%), in melanoma. In addition to mutations, Ras isoforms can also be activated by the inactivation of Ras GTPase activating proteins (RasGAPs), such as NF1 , RASA1 , and RASA2 . In our recent study, we observed that the inactivation of RASA1 (RAS p21 protein activator 1, also called p120RasGAP) suppressed melanoma via its RasGAP activity toward the R-Ras (related RAS viral (r-ras) oncogene homolog) isoform. We hypothesized that, although not mutated, R-Ras is activated in melanoma through the inactivation of RasGAPs and that RasGAP/R-Ras pathway activation cooperates with BRAF activation in melanoma tumorigenesis. In this study, we addressed the importance of R-Ras, a previously less-appreciated member of the Ras family, in melanoma tumorigenesis and investigated the molecular mechanisms underlying R-Ras signaling in BRAF mutant melanoma. We observed frequent activation of R-Ras in BRAF mutant human melanoma cell lines. In addition, RNAi-mediated reduced expression of R-Ras suppressed anchorage-independent colony growth and tumor growth. Moreover, among the three major RAS effector pathways, reduced R-Ras expression suppressed Ral-A activation, which may explain the mechanism of Ral-A activation in BRAF mutant melanoma. Interestingly, anchorage-independent growth driven by RASA1 inactivation and subsequent R-Ras activation was suppressed by both genetic (siRNA targeting Ral-A) and pharmacologic (Ral inhibitor BQU57) inhibition of Ral-A. To further investigate the impact of RASA1 loss, and thus R-Ras activation, on BRAF mutant melanoma development in vivo, we generated a RASA1 L/L ; BRAF CA/CA ; Tyr-CreERT2 mouse model in which treatment with 4OHT results in the expression of constitutively activated mutant BRAF and the deletion of RASA1 in melanocytic lineage cells. Preliminary analysis shows hyperpigmentation of the ear, tail, and foot pad in RASA1 L/L BRAF CA/CA mice compared to RASA1 +/+ BRAF CA/CA littermates and the development of melanoma in RASA1 mutant mice. This study demonstrates the importance of the RASA1/R-Ras/Ral-A signaling pathway in BRAF mutant melanoma and supports the possible combinatorial treatment strategy targeting both the BRAF/MAPK and Ral signaling pathways. Citation Format: Kristen S. Hill, Xue Wang, Evan R. Roberts, Youngchul Kim, Jane Messina, Minjung Kim. The importance of the RASA1/R-Ras/Ral-A signaling axis in melanoma tumorigenesis [abstract]. In: Proceedings of the AACR Special Conference: Advances in Modeling Cancer in Mice: Technology, Biology, and Beyond; 2017 Sep 24-27; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(10 Suppl):Abstract nr B12.