Fig. 3
ECDD-S18 decreases vacuolar ATPase activity in HNSCC cells. (a) Cal-27 and (b) FaDu cells were seeded onto 96-black well plates. Cells were treated with 0.1, 1, and 2 µM ECDD-S18 or 0.1 µM bafilomycin A1 (positive control) for 24 h. Cells were stained with LysoTracker for 2 h to indicate the lysosomal acidification and then fixed with 4% paraformaldehyde. Fluorescence intensity and images were captured using the Operetta High Content Imaging System. The bar graphs showed the relative fluorescence intensity of the mean LysoTracker (red) intensity normalized with DAPI intensity. Scale bar = 50 µm. (c) Molecular docking result of ECDD-S18 bound at the interface between two adjacent c subunits and subunit a of V-ATPase VO region, along with a close-up view of the binding residues surrounding ECDD-S18. (d) 2D interaction diagrams of the V-ATPase-ECDD-S18 complex. Note that the asterisk in (c) indicates the residues from the second (adjacent) subunit c. Data were illustrated as mean ± S.E.M. (n = 3). *p < 0.05 relative to DMSO control using one-way ANOVA with a Tukey’s multiple comparison test.
