Conditional control of MITF reveals cellular subpopulations essential for melanoma survival and recurrence in new zebrafish models
Melanoma is the most lethal type of skin cancer with over 132,000 cases occurring globally each year and continually rising incidence. BRAFV600E inhibitors have led to clinically significant improvements in outcomes for melanoma patients, yet many patients with metastatic melanoma rapidly succumb to the disease due to eventual chemoresistance or insensitivity to the drug. Thus, it is critical to identify new therapies that can act alone, or be combined with available treatments for enhanced efficacy and/or to overcome drug resistance. Evidence from human melanoma indicates that the melanocyte lineage is critical for melanoma survival and contributes to therapeutic resistance. MITF is a highly conserved “master melanocyte transcription factor” with a complex role in melanoma. Our lab has previously developed a temperature sensitive BRAFV600E mitfavc7 zebrafish melanoma model carrying a human oncogene and mitfavc7 splice site mutation that enables the conditional control of its endogenous activity by changes to water temperature. As part of my PhD project, I characterized and compared two new models developed since then: a very aggressive BRAFV600E mitfavc7p53M214K melanoma model with three driving mutations and a slower developing BRAF-‐independent mitfavc7p53M214K. I showed that the MITF activity is crucial for melanocyte survival in both models and that both mutated BRAF and p53 deficiency are oncogenic with low levels of MITF, and result in fish nevi and melanoma resembling the pathology of human disease. Both models are also relevant to a low-‐MITF subclass of human melanomas that emerged from a recent classification by The Cancer Genome Atlas Network. In addition, I established that, similarly to the BRAFV600Emitfavc7, complete inhibition of MITF activity leads to rapid tumour regression, but once its activity is restored the melanomas recur at the same site as the original tumour. I used histopathology studies and melanocyte lineage transgenes to identify and visualize subpopulations of cells remaining at the site of regression in these new zebrafish melanoma models. I hypothesised that these are the cells of origin for tumour recurrence (melanoma stem or progenitor cells), showed that some of them express a cancer stem cell marker aldehyde dehydrogenase, and attempted to target these subpopulations using 5-‐nitrofurans (a prodrug NFN1, shown previously by our lab to target ALDHhigh subpopulations in context of melanoma) in fish after melanoma regression. Finally, I also developed and described a new primary zebrafish melanoma cell line that I derived from one of these zebrafish tumours. This study is still in progress, but the cell line will be a useful tool for further investigation of these proposed melanoma progenitor cells in vitro, with potential applications for lineage tracing and transplantations.