Dual Inhibition of Histone Deacetylases and the Mechanistic Target of Rapamycin Promotes Apoptosis in Cell Line Models of Uveal Melanoma.

ABSTRACT

Purpose: Over 90% of uveal melanomas harbor pathogenic variants of the GNAQ or GNA11 genes that activate survival pathways. As previous studies found that Ras-mutated cell lines were vulnerable to a combination of survival pathway inhibitors and the histone-deacetylase inhibitor romidepsin, we investigated whether this combination would be effective in models of uveal melanoma. Methods: A small-scale screen of inhibitors of bromodomain-containing protein 4 (BRD4; OTX-015), extracellular signal-related kinase (ERK; ulixertinib), mechanistic target of rapamycin (mTOR; AZD-8055), or phosphoinositide 3-kinase (PI3K; GDC-0941) combined with a clinically relevant administration of romidepsin was performed on a panel of uveal melanoma cell lines (92.1, Mel202, MP38, and MP41) and apoptosis was quantified by flow cytometry after 48 hours. RNA sequencing analysis was performed on Mel202 cells treated with romidepsin alone, AZD-8055 alone, or the combination, and protein changes were validated by immunoblot. Results: AZD-8055 with romidepsin was the most effective combination in inducing apoptosis in the cell lines. Increased caspase-3 and PARP cleavage were noted in the cell lines when they were treated with romidepsin and mTOR inhibitors. RNA sequencing analysis of Mel202 cells revealed that apoptosis was the most affected pathway in the romidepsin/AZD-8055-treated cells. Increases in pro-apoptotic BCL2L11 and decreases in anti-apoptotic BIRC5 and BCL2L1 transcripts noted in the sequencing analysis were confirmed at the protein level in Mel202 cells. Conclusions: Our data suggest that romidepsin in combination with mTOR inhibition could be an effective treatment strategy against uveal melanoma due in part to changes in apoptotic proteins.