Uncovering the Molecular Basis of Phenotypic Diversification in BRAF-mutant Melanoma

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Tumor heterogeneity has been known to contribute to diverse patient outcomes in response to targeted therapies. It has primarily been studied in the context of genetic mutations that are either pre-existing or acquired in response to drug treatment. However, we have discovered signs of phenotypic diversification in normal conditions over time, which we think is driven by non- genetic processes. In order to study this diversification, we have used complimentary concepts from ecological theory known as bet-hedging and phenotypic plasticity. Briefly, we think cancer cell populations diversify their “bets” in normal conditions in order to increase chances of survival to any number of environmental changes. Underlying this theory, phenotypic plasticity suggests cells move around in a landscape of cell states, defined by molecular characteristics. These processes have been implicated in a variety of biological processes, including bacterial survival, stem cell differentiation, and epithelial-to-mesenchymal transitions. Preliminary data in the laboratory suggests complex drug-response dynamics in BRAF-mutant melanoma cell lines, likely resulting from non-genetic heterogeneity. Single-cell derived “clones” of the BRAF-mutant melanoma cell line SKMel5 have been generated and developed into cell sublines, which have shown differential phenotypes to drug treatment. In studying one of these sublines' response to BRAF inhibition, we have observed phenotypic diversification over time. Using a mathematical model that assumes phenotypic plasticity between non-genetically defined states, we have been able to form a theoretical basis for this experimental observation. In this proposal, we attempt to discern the existence of these states, identify molecular determinants, and monitor dynamic phenotypic diversification in real time.