Diltiazem inhibits SARS-CoV-2 binding.

(A) Cells were subjected to flow cytometry to detect the surface expression of ACE2. (B) Cells were lysed to determine ACE2 mRNA copy numbers by qPCR. (C) Schematic of the viral binding assay. (D) Diltiazem preincubated cells were incubated with HRB25 (M.O.I. = 10) at 4°C for 1 h. The viral RNA level in the cell lysate was measured by qPCR. (E) Diltiazem preincubated Vero-E6 cells were incubated with HRB25 at an M.O.I. of 10, 1, 0.1, or 0.01. The viral RNA level in the cell lysate was measured by qPCR. (F) Vero-E6 cells were treated and infected as described in (C). The cells were incubated with an anti-SARS-CoV-2 nucleocapsid rabbit monoclonal antibody, and visualized with Alexa Fluor 488-conjugated goat anti-rabbit IgG (green). Cell nuclei were stained with Hoechst 33342. Representative images are shown. The fluorescence intensities of cell-bound HRB25 in at least 110 cells per sample were quantified. (G) HEK293T cells were treated and assessed as described in (E). The data shown represent, or are from, three independent experiments or replicates (means ± SDs). The two-tailed unpaired Student’s t-test was used for the statistical analysis. ns, not significant, *p < 0.05, **p < 0.01, ***p < 0.001.