Figure 3. Ro 48-8071 inhibits EBOV entry at a post internalization step and does not inhibit endosome acidification or cathepsin levels.

Effects of Ro 48-8071 (indicated concentration in A, 5 µM in B-E) on: (A) VLP-GPΔ and VLP-G entry; one representative of two experiments (done in triplicate). (B) VLP-GPΔ and VLP-LCMV entry; one representative of three experiments (done in duplicate). (C) VLP-GPΔ internalization and entry; 50 µM EIPA was used as the positive control for an inhibitor of EBOV internalization [8]; 10 µM EIPA was used as the control for the entry assay (50 µM EIPA caused high background fluorescence in the entry assay); one representative of two experiments (done in triplicate). (D) Low endosomal pH was detected by incubating cells with Lysotracker Red; 10 mM NH₄Cl was used as the control for pH neutralization; representative images from multiple coverslips from a single experiment. (E) Cathepsin B, L, and combined B/L activity; 10 µM E64d was used as the positive control for inhibition of cysteine protease activity; results from a single experiment performed in duplicate. In all assays, SNB19 cells were pre-treated with the indicated concentration of inhibitor for 1 hr at 37°C, and inhibitors were maintained throughout the assays. Error bars represent standard deviation from the mean of mock-treated samples: * (P<.01), ** (P<.001), or *** (P<.0001).