Targeting Lineage-specific MITF Pathway in Human Melanoma Cell Lines by A-485, the Selective Small-molecule Inhibitor of p300/CBP.

Rui Wang,Lloyd T Lam,Anahita Bhathena,Tamar Uziel,Valerie Robinson,Yupeng He,Paul Hessler,Loren M Lasko,Xin Lu,Ziping Yang,Albert Lai

doi:10.1158/1535-7163.mct-18-0511

Abstract

Metastatic melanoma is responsible for approximately 80% of deaths from skin cancer. Microphthalmia-associated transcription factor (MITF) is a melanocyte-specific transcription factor that plays an important role in the differentiation, proliferation, and survival of melanocytes as well as in melanoma oncogenesis. MITF is amplified in approximately 15% of patients with metastatic melanoma. However, no small-molecule inhibitors of MITF currently exist. MITF was shown to associate with p300/CBP, members of the KAT3 family of histone acetyltransferase. p300 and CREB-binding protein (p300/CBP) regulate a wide range of cellular events such as senescence, apoptosis, cell cycle, DNA damage response, and cellular differentiation. p300/CBP act as transcriptional coactivators for multiple proteins in cancers, including oncogenic transcription factors such as MITF. In this study, we showed that our novel p300/CBP catalytic inhibitor, A-485, induces senescence in multiple melanoma cell lines, similar to silencing expression of EP300 (encodes p300) or MITF We did not observe apoptosis and increase invasiveness upon A-485 treatment. A-485 regulates the expression of MITF and its downstream signature genes in melanoma cell lines undergoing senescence. In addition, expression and copy number of MITF is significantly higher in melanoma cell lines that undergo A-485-induced senescence than resistant cell lines. Finally, we showed that A-485 inhibits histone-H3 acetylation but did not displace p300 at promoters of MITF and its putative downstream genes. Taken together, we provide evidence that p300/CBP inhibition suppressed the melanoma-driven transcription factor, MITF, and could be further exploited as a potential therapy for treating melanoma.

Highlights

Melanoma is one of the most frequent cancers with increased incidence in the Western societies
P300/CREB-binding protein (CBP) Inhibition Targets Microphthalmia-associated transcription factor (MITF) in Melanoma Cell Lines comparing with DMSO enrichment relative to negative control (IgG) for chromatin immunoprecipitation (ChIP) is indicated; GAPDH promoter sequence is used as endogenous control for quantitative PCR (qPCR)
Upon performing Ingenuity upstream regulator analysis (IPA) across the different cell lines, we showed that there was an enrichment of transcriptional network regulated by MITF with p300/CBP inhibitor treatment in the two sensitive lines (Fig. 2B)

Summary

Introduction

Melanoma is one of the most frequent cancers with increased incidence in the Western societies. The identification of a constitutively active MAPK pathway due to BRAF V600E mutation in about 40% melanoma has led to the development of selective BRAF inhibitors such as vemurafenib [5]. Observed in patients with BRAF V600E mutation, the long-term benefit of vemurafenib has been compromised because of the development of resistance to the therapy. It was shown that microphthalmiaassociated transcription factor (MITF)-low melanoma is associated with intrinsic resistance to multiple targeted agents including BRAF inhibitor [8, 9]. We evaluated the activity of this specific p300/CBP inhibitor A-485 in a panel of melanoma cell lines. Our results support a rationale for testing p300/CBP inhibitors in patients with melanoma with MITF copy number gain and upregulated expression

Materials and Methods

Results

Discussion

Findings

Disclosure of Potential Conflicts of Interest