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Targeted therapeutic options for the 15-25% of melanomas driven by oncogenic N-RAS mutations are currently unavailable. Therefore, immune-based therapies are the front-line treatment for these tumors. Unfortunately, only 40% of patients respond to immune-based therapies and numerous responders exhibit severe adverse events. Thus, it is important to find new therapeutic options for N-RAS-mutant melanomas. To understand the mechanism by which mutant N-RAS promotes melanoma formation, our laboratory developed a suite of genetically engineered mouse models (GEMMs) that enable conditional, melanocyte-specific knock-in of N-Ras mutations into endogenous alleles. We developed a total of five GEMMS containing N-Ras codon 61 mutations found commonly (Q61R, Q61K, Q61L) or uncommonly (Q61H, Q61P) in melanoma. Recent data from the laboratory show that GEMMs expressing melanoma-common N-Ras mutants exhibit reduced tumor latency and increased tumor burden as compared to those expressing N-Ras mutations privy to other tumor types.

I hypothesized that the melanoma-common N-RAS mutations differentially utilize the canonical N-RAS signaling pathway (PI3K-AKT, RAF-MEK-ERK) to alter cellular proliferation and more effectively drive melanoma formation. Analyses of cell proliferation in mouse embryonic fibroblasts (MEFs) demonstrated that all of the aforementioned mutations increased proliferation levels similarly as compared to wild-type (WT) MEFs. However, immunoblots examining downstream pathways revealed no differences in the levels of phosphorylated ERK and AKT. Thus, analyzing these pathways in melanocytes within the skin of our GEMMs could illustrate the differences between the potent and non-potent melanoma-driving N-RAS mutations as tumor microenvironment could affect the signaling pathways. Additionally, testing of WT RAS activity revealed that there is a difference in levels of GTP-bound WT H-/K-Ras within our MEFs expressing various N-RasQ61 mutations. Further elucidation of the signaling pathways and functions that enable a subset of N-RAS mutations to more effectively drive melanoma formation will allow us to identify novel therapeutic targets for this disease.