Abstract LB-265: ARAF activates RAS by antagonizing its interaction with NF1

Abstract

Abstract ARAF, BRAF and CRAF protein kinases are RAS effectors required for activation of ERK signaling. ARAF has relatively modest kinase activity and its biological functions are largely unknown. Here, we show that ectopic expression of ARAF but not B- or CRAF, leads to induction of RAS activation in cells with WT RAS. This phenomenon requires the binding of ARAF to RAS but not its kinase activity. ARAF was found to reduce inactivation of RAS by antagonizing its binding to the major RAS-GAP NF1. By this mechanism, the abundance of ARAF in cells determines cellular RAS-GTP levels, the dynamics of RTK-induced RAS activation and the biological consequences of RAS signaling. We find that a certain expression level of ARAF is required for growth factors induced proliferating or neural differentiation in PC12 cells. In RTK-dependent tumor cells, knocking down of ARAF leads to a reduction of RAS-GTP, thus impairs RAS signaling and the tumor growth. Furthermore, by investigating the tumor samples from MSK patients who are diagnosed with progressing disease after EGFR inhibitor treatment, we find the up-regulation of ARAF (including the ARAF gene amplification) is associated with EGFR inhibitor resistance in the lung cancer patients. We confirmed in cell line and xenograft models that a 5-fold elevation of ARAF expression is sufficient to eliminate RAS inhibition by EGFR inhibition and to cause resistance. Resistance in these cells could be overcome by combining EGFR and SHP2 inhibitors. The latter prevents EGFR activation of RAS-GTP exchange and thus would be expected with weaking the effects of NF1 inhibition by ARAF. Taken together, our work shows that ARAF directly activates RAS by antagonizing its interaction with NF1 and thus affects the duration and cellular response of growth factor activation and sensitivity of tumors to pharmacological inhibition of RTK-RAS signaling. Citation Format: Wenjing Su, Rona Yaeger, Na Na, Jaclyn Hechtman, Viktoriya Paroder, Sandra Misale, Sae-Won Han, Omar I. Abdel-Wahab, Frank McCormick, Neal Rosen, Zhan Yao. ARAF activates RAS by antagonizing its interaction with NF1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-265.