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Fig. 4 | Clinical Epigenetics

Fig. 4

From: Bromodomain inhibitor i-BET858 triggers a unique transcriptional response coupled to enhanced DNA damage, cell cycle arrest and apoptosis in high-grade ovarian carcinoma cells

Fig. 4

Ovarian cancer cells undergo apoptosis following i-BET858 treatment. Protein lysates of OVCAR-3 cells treated with (A) i-BET858 and B i-BET151 were subjected to western blot analyses to study changes in CDKN1A/p21, cleaved PARP and γH2A.X protein levels after 4, 24 and 48 h of treatment; GAPDH was used as loading control. C, D Protein lysates of OVCAR-3 cells treated with different concentrations of i-BET858 and i-BET151 (10 nM-2.5 µM; 48 h) were subjected to western blot analyses to study dose-dependent changes in CDKN1A/p21, cleaved PARP and γH2A.X protein levels. E Confocal microscopy images of OVCAR-3 spheroids treated with i-BET151, i-BET858 and DMSO control for 48 h. First and second rows show fluorescent-labelled DNA and γH2A.X staining, respectively; third row displays merged images of both fluorescent signals. Scale represents 100 µm. F, G Flow cytometry apoptosis analysis of CAOV3 cells treated with different concentrations of i-BET151, i-BET858 and DMSO vehicle control for (F) 24 h and G 48 h. Cells were stained with propidium iodide and Annexin V-FITC rendering 4 populations: viable (−, −), early apoptotic (−, +), late apoptotic (+, +) and dead (+, −), two of which are highlighted in the panels. Graphs display cell densities, whereby red, green and blue colours indicate high, medium and low cell densities, respectively. H Table detailing specific percentages of OVCAR-3 cells detected in each population following treatments. Percentages of dead cells after 24 h treatment were not significant and are not included in this table

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