PO-214 Conditional control of MITF reveals cellular subpopulations essential for melanoma survival and recurrence in new zebrafish models

Abstract

Introduction Melanoma is the most lethal type of skin cancer and its incidence continues to rise rapidly. While BRAFV600E inhibitors have led to significant improvements in outcomes for some patients, there are no targeted therapies available for melanoma patients who lack BRAF mutations. An emerging area of new melanoma targets is the melanocyte lineage, which is critical for melanoma survival and contributes to therapeutic resistance. MITF is a conserved ‘master melanocyte regulator’ with a complex role in melanoma. Our lab previously developed a BRAFV600E mitf zebrafish melanoma model carrying a human oncogene and mitfvc7 splice site mutation that enables the conditional control of its endogenous activity. Since there are not many adequate animal models to study the non-BRAF and non-RAS melanomas, we have now also developed a double mutant model p53 mitf. Here we use the new models to investigate MITF-dependent melanoma regression and recurrence. Material and methods We use histopathology and imaging of melanocyte lineage fluorescent reporters (mitf and crestin) in new temperature-sensitive zebrafish models, as well as primary zebrafish melanoma cell line, to study melanoma regression and recurrence. Results and discussions We demonstrate that the MITF activity is crucial for melanocyte survival and that both mutated BRAF and p53 deficiency are oncogenic with low levels of MITF, and result in melanoma resembling the pathology of human disease. This is particularly important considering the new MITF-low subclass of human melanomas that has emerged from the recent TCGA classification. We also show that complete inhibition of MITF activity leads to rapid tumour regression (in both new models), but once its activity is restored the melanomas recur at the same site as the original tumour. This suggests that a subpopulation of cancer initiating cells remains following melanoma regression and is capable of repopulating the tumour. We have been able to show that some cells remaining at the sites of regression express MITF and/or ALDH2 and we are currently searching for other markers. Conclusion We developed new MITF-dependent zebrafish models of melanoma that are relevant to distinct clinical subtypes. We showed that inhibiting MITF in those animals results in melanoma regression, but is not sufficient to eradicate the tumours (once its activity is restored, melanomas recur). We identified subpopulations of cells remaining at the regression sites and are investigating the nature of these proposed cells-of-origin for relapse.