Role of Rho/MRTF in Aggressive Vemurafenib-Resistant Murine Melanomas and Immune Checkpoint Upregulation

Cutaneous melanoma is the deadliest skin cancer. Most have Ras-MAPK pathway (BRAF(V600E) or NRAS) mutations and highly effective targeted therapies exist; however, they and immune therapies are limited by resistance, in part driven by small GTPase (Rho and Rac) activation. To facilitate preclinical studies of combination therapies to provide durable responses, we describe the first mouse melanoma lines resistant to BRAF inhibitors. Treatment of mouse lines, YUMM1.7 and YUMMER, with vemurafenib (Vem), the BRAF(V600E)-selective inhibitor, resulted in high-level resistance (IC(50) shifts 20–30-fold). Resistant cells showed enhanced activation of Rho and the downstream transcriptional coactivator, myocardin-related transcription factor (MRTF). Resistant cells exhibited increased stress fibers, nuclear translocation of MRTF-A, and an increased MRTF-A gene signature. Pharmacological inhibition of the Rho/MRTF pathway using CCG-257081 reduced viability of resistant lines and enhanced sensitivity to Vem. Remarkably, co-treatment of parental lines with Vem and CCG-257081 eliminated resistant colony development. Resistant cells grew more slowly in vitro, but they developed highly aggressive tumors with a shortened survival of tumor-bearing mice. Increased expression of immune checkpoint inhibitor proteins (ICIs) in resistant lines may contribute to aggressive in vivo behavior. Here, we introduce the first drug-resistant mouse melanoma models for assessing combinations of targeted and immune therapies.