Purpose: Multiple BRAF inhibitor resistance mechanisms have been described, however, their relative frequency, clinical correlates, and effect on subsequent therapy have not been assessed in patients with metastatic melanoma.Experimental Design: Fifty-nine BRAF V600 -mutant melanoma metastases from patients treated with dabrafenib or vemurafenib were analyzed. The genetic profile of resistance mechanisms and tumor signaling pathway activity was correlated with clinicopathologic features and therapeutic outcomes.Results: Resistance mechanisms were identified in 58% progressing tumors and BRAF alterations were common. Gene expression analysis revealed that mitogen-activated protein kinase (MAPK) activity remained inhibited in 21% of resistant tumors, and the outcomes of patients with these tumors were poor. Resistance mechanisms also occurred in pretreatment biopsies and heterogeneity of resistance mechanisms occurred within patients and within tumors. There were no responses to subsequent targeted therapy, even when a progressing tumor had a resistance mechanism predicted to be responsive.Conclusions: Selecting sequential drugs based on the molecular characteristics of a single progressing biopsy is unlikely to provide improved responses, and first-line therapies targeting multiple pathways will be required.

One-third of BRAF-mutant metastatic melanoma patients treated with combined BRAF and MEK inhibition progress within 6 months. Treatment options for these patients remain limited. Here we analyse 20 BRAF V600 -mutant melanoma metastases derived from 10 patients treated with the combination of dabrafenib and trametinib for resistance mechanisms and genetic correlates of response. Resistance mechanisms are identified in 9/11 progressing tumours and MAPK reactivation occurred in 9/10 tumours, commonly via BRAF amplification and mutations activating NRAS and MEK2. Our data confirming that MEK2 C125S , but not the synonymous MEK1 C121S protein, confers resistance to combination therapy highlight the functional differences between these kinases and the preponderance of MEK2 mutations in combination therapy-resistant melanomas. Exome sequencing did not identify additional progression-specific resistance candidates. Nevertheless, most melanomas carried additional oncogenic mutations at baseline (for example, RAC1 and AKT3) that activate the MAPK and PI3K pathways and are thus predicted to diminish response to MAPK inhibitors.