In human mutant BRAF melanoma cells, the stemness transcription factor FOXD3 is rapidly induced by inhibition of ERK1/2 signaling and mediates adaptive resistance to RAF inhibitors.,However, the mechanism underlying ERK signaling control of FOXD3 expression remains unknown.,Here we show that SOX10 is both necessary and sufficient for RAF inhibitor-induced expression of FOXD3 in mutant BRAF melanoma cells.,SOX10 activates the transcription of FOXD3 by binding to a regulatory element in FOXD3 promoter.,Phosphorylation of SOX10 by ERK inhibits its transcription activity toward multiple target genes by interfering with the sumoylation of SOX10 at K55, which is essential for its transcription activity.,Finally, depletion of SOX10 sensitizes mutant BRAF melanoma cells to RAF inhibitors in vitro and in vivo.,Thus, our work discovers a novel phosphorylation-dependent regulatory mechanism of SOX10 transcription activity and completes an ERK1/2/SOX10/FOXD3/ERBB3 axis that mediates adaptive resistance to RAF inhibitors in mutant BRAF melanoma cells.,In BRAF mutant melanoma, inhibition of ERK1/2 induces FOXD3 and mediates RAF inhibitor resistance.,Here, the authors show that ERK1/2 mediated phosphorylation regulates sumoylation of SOX10 which activates FOXD3, and depletion of SOX10 sensitises BRAF mutant melanoma cells to RAF inhibitors.
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.