Figure 4

Validation of the Maybridge hits as BAF inhibitors. (a) The molecular structures and masses of the three Maybridge library hits: KM 04416, KM 04550 and JFD 02731. (b,c) Inhibitory effect of the Maybridge hits in the HTS (b) and DFR (c) assays (concentration 10 µM) under oxidizing (w/o GSH) and reducing (w/ GSH) conditions. The fluorescence was normalized with respect to the positive control (DMSO) under oxidizing and reducing conditions, respectively. (d–f) ESI–MS analysis of BAF adduct formation under reducing (w/ GSH, black, dashed box) and oxidizing (w/o GSH, red, clear box) conditions with the Maybridge hits KM 04416, KM 04550 and JFD 02731, respectively. (g,h) FACS analysis of HeLa cells 24 h after the transfection with a GFP encoding plasmid for 1 h or 20 h using the transfection agent X-tremeGENE 9. The transfection was performed in the presence of KM 04416 (10 µM), KM 04450 (10 µM) or JFD 02731 (50 µM). The subfigures depict the percentage of GFP positive cells and the mean GFP intensities, respectively. B, C, G, H: Mean ± SD (N = 3). *p < 0.05, **p < 0.01, ***p < 0.01, NS = not significant respective to positive control.