The microphthalmia-associated transcription factor (MITF) is the “master melanocyte transcription factor” with a complex role in melanoma.,MITF protein levels vary between and within clinical specimens, and amplifications and gain- and loss-of-function mutations have been identified in melanoma.,How MITF functions in melanoma development and the effects of targeting MITF in vivo are unknown because MITF levels have not been directly tested in a genetic animal model.,Here, we use a temperature-sensitive mitf zebrafish mutant to conditionally control endogenous MITF activity.,We show that low levels of endogenous MITF activity are oncogenic with BRAFV600E to promote melanoma that reflects the pathology of the human disease.,Remarkably, abrogating MITF activity in BRAFV600Emitf melanoma leads to dramatic tumor regression marked by melanophage infiltration and increased apoptosis.,These studies are significant because they show that targeting MITF activity is a potent antitumor mechanism, but also show that caution is required because low levels of wild-type MITF activity are oncogenic.
Knowledge of tumor mutation status is becoming increasingly important for the treatment of cancer, as mutation-specific inhibitors are being developed for clinical use that target only sub-populations of patients with particular tumor genotypes.,Melanoma provides a recent example of this paradigm.,We report here development, validation, and implementation of an assay designed to simultaneously detect 43 common somatic point mutations in 6 genes (BRAF, NRAS, KIT, GNAQ, GNA11, and CTNNB1) potentially relevant to existing and emerging targeted therapies specifically in melanoma.,The test utilizes the SNaPshot method (multiplex PCR, multiplex primer extension, and capillary electrophoresis) and can be performed rapidly with high sensitivity (requiring 5-10% mutant allele frequency) and minimal amounts of DNA (10-20 nanograms).,The assay was validated using cell lines, fresh-frozen tissue, and formalin-fixed paraffin embedded tissue.,Clinical characteristics and the impact on clinical trial enrollment were then assessed for the first 150 melanoma patients whose tumors were genotyped in the Vanderbilt molecular diagnostics lab.,Directing this test to a single disease, 90 of 150 (60%) melanomas from sites throughout the body harbored a mutation tested, including 57, 23, 6, 3, and 2 mutations in BRAF, NRAS, GNAQ, KIT, and CTNNB1, respectively.,Among BRAF V600 mutations, 79%, 12%, 5%, and 4% were V600E, V600K, V600R, and V600M, respectively. 23 of 54 (43%) patients with mutation harboring metastatic disease were subsequently enrolled in genotype-driven trials.,We present development of a simple mutational profiling screen for clinically relevant mutations in melanoma.,Adoption of this genetically-informed approach to the treatment of melanoma has already had an impact on clinical trial enrollment and prioritization of therapy for patients with the disease.