Research Paper Volume 12, Issue 5 pp 4547—4557

ARV-825-induced BRD4 protein degradation as a therapy for thyroid carcinoma

Figure 3. ARV-825 inhibits BRD4 signaling in human thyroid carcinoma cells. TPC-1 cells (A and B) or the primary human thyroid carcinoma cells (“C1”/“C2”) (F) were treated with applied concentration of ARV-825 for 24h, tested by Western blotting and qPCR assays of listed genes. TPC-1 cells were pre-treated with MG-132 (25 μM) for 2h, followed by ARV-825 (100 nM) treated for 24h, expression of listed proteins was shown (C). The stable TPC-1 cells with CRISPR/Cas9 BRD4-KO construct (“BRD4-KO”) cells were treated with or without ARV-825 (100 nM, for indicated time periods), control TPC-1 cells with empty vector (“Cas9-C”) were left untreated; expression of listed proteins was shown (D); Cell viability and proliferation were tested by MTT and EdU staining assays (E), respectively. TPC-1 cells or “C1” human thyroid carcinoma cells were treated with ARV-825 (100 nM), JQ1 (500 nM), CPI203 (500 nM) or GSK1210151A (500 nM) for indicated time periods, cell viability (MTT assay, 96h) (G and J), proliferation (testing nuclear EdU/DAPI ratio, 48h) (H and K) and apoptosis (nuclear TUNEL ratio, 48h) (I and L) were tested, and results were quantified. Expression of listed proteins was quantified and normalized to the corresponding loading control (A, C, D and F). #p < 0.001 vs. “ARV-825” treatment group (GL). The experiments were repeated three times, and similar results obtained.