Targeting BET proteins in melanoma: A novel treatment approach.

Abstract

9091 Background: Manipulation of key epigenetic regulators in melanoma proliferation is emerging as a new therapeutic strategy. Bromodomain-containing proteins such as the extraterminal domain (BET) family are components of transcription factor complexes and determinants of epigenetic memory. We investigated the expression of BRD4, a BET family member in melanoma cell lines and tissues, and the effects of its inhibition with the small molecule compounds MS436 and MS417 in in vitro and in vivo models of melanoma. Methods: BRD2 and BRD4 expression were analyzed by immunohistochemistry. We tested the effects of pharmacological or RNAi-mediated inhibition of BRD4 in melanoma cells using crystal violet-based assays for proliferation/colony formation and flow-cytometry for cell cycle analysis. The molecular effects of BRD4 suppression were examined using RNA sequencing, Real-Time quantitative PCR and western blots for p27, p21, MYC, ERK1 and SKP2. In the in vivo xenograft experiments NOD/SCID/IL2γR-/-mice were injected with melanoma cells and treated with MS417. Statistical significance was determined by unpaired t-test (GraphPad). Results: BRD4 was found significantly upregulated in primary and metastatic melanoma tissues compared to melanocytes and nevi (p<0.001). Treatment with BET inhibitors impaired melanoma cell proliferation in vitro and tumor growth and metastatic behavior in vivo, effects that were mostly recapitulated by individual silencing of BRD4. Rapidly after BET displacement, key cell cycle genes (SKP2, ERK1 and c-MYC) were downregulated concomitantly with the accumulation of CDK inhibitors (p21, p27), followed by melanoma cell cycle arrest. BET inhibitor efficacy was not influenced by BRAF or NRAS mutational status. Conclusions: Our results demonstrate for the first time a role for BRD4 in melanoma maintenance and support the role of BET proteins as novel targets in melanoma. Further investigation in the clinical setting is warranted.

Inhibition of bromodomain and extra-terminal proteins (BET) as a potential therapeutic approach in haematological malignancies: emerging preclinical and clinical evidence

“…In the HIV-1 Tg26 transgenic mice, MS417 blocks BRD-4 binding to acetylated NF-κB and effectively attenuates the inflammatory response in HIV-induced nephropathy [Zhang et al 2012a]. MS417 induces cell cycle arrest in preclinical melanoma models, but it has not been tested in haematological malignancies [Paoluzzi et al 2013]. OTX015, originally developed for the treatment of inflammatory bowel disease, is among the thienodiazepine inhibitors with a promising antitumour profile in haematological malignancies [Miyoshi et al 2009;Gautschi and Minikis, 2014].…”