LAUSR

Oncogenic mutations of both NRAS and KRAS are common in multiple myeloma (MM), an incurable malignancy of plasma cells. However, the mechanisms of pathogenic RAS signaling in MM have remained enigmatic, and there is little evidence to support a dominant role for classical MAPK signaling downstream of RAS in this disease. We recently used an unbiased proteogenomic approach to dissect RAS signaling in MM and discover that mutant isoforms of RAS organized a signaling complex with the amino acid transporter, SLC3A2, and MTOR on lysosomes, which directly activated mTORC1 by co-opting amino acid sensing pathways (PMID: 36115844). To gain further insights into this novel mode of RAS signaling located away from the plasma membrane, we have employed mass spectrometry to characterize proteins associated with lysosomes isolated from MM cell lines. We observed that RAS isoforms, SLC3A2 and several components of the mTORC1 signalosome were purified with lysosomes, consistent with RAS activation of mTORC1 on lysosomes. To identify novel regulators of oncogenic RAS signaling at the lysosome, we used phospho-proteomics to identify proteins with significantly reduced levels of phosphorylation following acute inhibition of KRAS activity with the KRAS G12D inhibitor, MRTX1133, in a MM cell line bearing a KRAS G12D mutation. Many of these phospho-proteins were also associated with lysosomes, including MTOR, which had substantially reduced phosphorylation at the amino-acid dependent phosphorylation site S1261 following KRAS inhibition. Pathway analysis of proteins present in lysosomes and phosphorylated downstream of oncogenic KRAS showed enrichment for proteins that regulate intracellular trafficking. Among these proteins, we found that disruption of a RAB GAP essential for survival in RAS-dependent MM cells substantially reduced associations between RAS and MTOR and nearly abolished RAS-dependent mTORC1 activity. These data suggest that mutant RAS utilizes lysosomal resident proteins to coordinate the colocalization of itself with mTORC1 on lysosomes to drive pathogenic signaling in MM. More generally, these studies suggest that targeting the intracellular localization of RAS may be an alternative route to inhibit oncogenic RAS signaling in MM. Citation Format: Arnold Bolomsky, Ryan M. Young. Oncogenic RAS signals from lysosomes to activate mTORC1 in multiple myeloma [abstract]. In: Proceedings of the AACR Special Conference: Targeting RAS; 2023 Mar 5-8; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Res 2023;21(5_Suppl):Abstract nr A037.