Co‐targeting BET and MEK as salvage therapy for MAPK and checkpoint inhibitor‐resistant melanoma NRAS‐mutant melanoma cells were treated with JQ‐1 alone.

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Co‐targeting BET and MEK as salvage therapy for MAPK and checkpoint inhibitor‐resistant melanoma NRAS‐mutant melanoma cells were treated with JQ‐1 alone or in combination with PD901. Cell viability was determined by Alamar Blue assay after 5 days of treatment and calculated relative to DMSO‐treated controls. Interaction index and 95% confidence interval (CI) were assessed for each cell line. The upper limit of its 95% CI < 1 was considered significant synergy. Cells were cultured in the presence of DMSO, 0.5 μM JQ‐1, 0.1 μM PD901, or combo for 14 days followed by crystal violet staining. Colonies were imaged using a digital camera and quantitated by ImageJ. Representative photographs of crystal violet stained colonies are shown. Cells were treated with DMSO or a single dose of 0.5 μM JQ‐1, 0.1 μM PD901, or combo. At day 6, cells were washed to remove the drugs and refed fresh (drug‐free) medium. Cells were fixed after 7 or 14 days, stained with crystal violet, and relative number of cells quantified. NRAS‐mutant melanoma and non‐transformed cells were treated as in (C) for 7 days. Cells were stained with propidium iodide and Annexin V‐FITC and analyzed by FACS; % Annexin V+/PI+ cells are shown. Data information: Data represent the mean of three independent experiments ± SEM. Statistically significant differences were determined by Student's t‐test; P‐values are shown. IF THIS IMAGE HAS BEEN PROVIDED BY OR IS OWNED BY A THIRD PARTY, AS INDICATED IN THE CAPTION LINE, THEN FURTHER PERMISSION MAY BE NEEDED BEFORE ANY FURTHER USE. PLEASE CONTACT WILEY'S PERMISSIONS DEPARTMENT ON PERMISSIONS@WILEY.COM OR USE THE RIGHTSLINK SERVICE BY CLICKING ON THE 'REQUEST PERMISSIONS' LINK ACCOMPANYING THIS ARTICLE. WILEY OR AUTHOR OWNED IMAGES MAY BE USED FOR NON-COMMERCIAL PURPOSES, SUBJECT TO PROPER CITATION OF THE ARTICLE, AUTHOR, AND PUBLISHER. EMBO Mol Med, Volume: 10, Issue: 5, First published: 11 April 2018, DOI: (10.15252/emmm.201708446)