A novel compound which sensitizes BRAF wild-type melanoma cells to vemurafenib in a TRIM16-dependent manner

Selina K Sutton,Benjamin D Noll,Greg M Arndt,Jonathan Baell,Belamy B Cheung,Shudong Wang,Xu Dong Zhang,Daniel R Carter,Patrick Kim,Naresh Kumar,Owen Tan,Grant A Mcarthur,Glenn M Marshall

doi:10.18632/oncotarget.10700

Abstract

There is an urgent need for better therapeutic options for advanced melanoma patients, particularly those without the BRAFV600E/K mutation. In melanoma cells, loss of TRIM16 expression is a marker of cell migration and metastasis, while the BRAF inhibitor, vemurafenib, induces melanoma cell growth arrest in a TRIM16-dependent manner. Here we identify a novel small molecule compound which sensitized BRAF wild-type melanoma cells to vemurafenib. High throughput, cell-based, chemical library screening identified a compound (C012) which significantly reduced melanoma cell viability, with limited toxicity for normal human fibroblasts. When combined with the BRAFV600E/K inhibitor, vemurafenib, C012 synergistically increased vemurafenib potency in 5 BRAFWT and 4 out of 5 BRAFV600E human melanoma cell lines (Combination Index: CI < 1), and, dramatically reduced colony forming ability. In addition, this drug combination was significantly anti-tumorigenic in vivo in a melanoma xenograft mouse model. The combination of vemurafenib and C012 markedly increased expression of TRIM16 protein, and knockdown of TRIM16 significantly reduced the growth inhibitory effects of the vemurafenib and C012 combination. These findings suggest that the combination of C012 and vemurafenib may have therapeutic potential for the treatment of melanoma, and, that reactivation of TRIM16 may be an effective strategy for patients with this disease.

Highlights

Melanoma is a highly aggressive malignancy, and it accounts for only a small percentage of skin cancers overall, it is responsible for the majority of skin cancer deaths [1]
To determine whether C012 had specific cytotoxicity to melanoma cells over other cancer cells, C012 was screened at 10 μM as a single agent for its effects on cell viability using the Alamar Blue assay against a panel of human cancer cell lines, normal human fibroblasts (MRC-5 and WI-38) and 10 human melanoma cell lines (Figure 1B)
We found no significant change in levels of MEK and ERK phosphorylation with the C012/vemurafenib combination treatment compared to single agent vemurafenib treatment of BRAFWT/NRASQ61R (Mel-JD and Mel-RM) cells (Supplementary Figure 2A) which instead displayed the expected MAPK pathway activation with vemurafenib treatment

Summary

Introduction

Melanoma is a highly aggressive malignancy, and it accounts for only a small percentage of skin cancers overall, it is responsible for the majority of skin cancer deaths [1]. The limited availability of therapeutic options highlight BRAFWT/ NRAS mutant patients as a subgroup most in need of new therapeutic treatment strategy and a better understanding of disease pathology. Identification of patient subgroups that are likely to respond to immune based therapy is still largely unknown [20, 21] This emphasises the important role targeted therapies have to play in melanoma treatment. We showed high levels of TRIM16 in melanoma tissues from patients treated with vemurafenib correlated with clinical response. These data suggest the potential of TRIM16 as a candidate tumor suppressor protein in melanoma and that restoration of TRIM16 expression may be a potential therapeutic strategy for melanoma treatment. Our findings have identified a novel small molecule with clinical potential in the treatment of melanoma, including BRAFWT melanoma, and provide mechanistic insights into the role of TRIM16 in this process

RESULTS

DISCUSSION

ACKNOWLEDGMENTS and funding