FIGURE 4 from Targeting the Clear Cell Sarcoma Oncogenic Driver Fusion Gene EWSR1::ATF1 by HDAC Inhibition

Depletion of BRD4 from the EWSR1 promoter region suppresses the transcription of EWSR1::ATF1.A, CCS cells were treated with 0–10 μmol/L vorinostat for 24 hours, and the protein expression of BRD4 was detected using Western blotting. B, Genome browser screenshot of the epigenome maps at EWSR1. Tracks visualize CUT&RUN-seq data for BRD4 (n = 2), and scATAC-seq data are shown for treatment with 3 μmol/L vorinostat or vehicle for 24 hours. Blue highlight shows the EWSR1 promoter region. C, CCS cells were treated with 0–3 μmol/L JQ1 for 48 hours, and the number of viable cells was estimated using a WST-8 assay (n = 3). The calculated IC₅₀ values are shown in Table 1. D, CCS cells were treated with 0, 0.3, 1.0, 3.0, or 10.0 μmol/L JQ1 for 24 hours. EWSR1::ATF1 protein expression levels were estimated via Western blotting. E, CCS cells were treated with 0, 0.3, and 3.0 μmol/L JQ1 for 24 hours. EWSR1::ATF1 mRNA levels in CCS cells were quantified using qRT-PCR (normalized to GAPDH; n = 3). Data in C and E are means ± SDs. *, P < 0.05; **, P < 0.01 (Student t test).