Fig. 4: Inhibition of SARS-CoV-2 induced fusion and of spike binding on hACE2 expressing cells by anti-ACE2 VHHs.
a S-fuse assay. Inhibition by B07, B09, B10 VHHs after infection of S-Fuse cells (U2OS-hACE2 GFP1-10 and GFP11) with different SARS-CoV-2 variants (Delta B.1.617.2, BA.1, BQ.1.1, XBB.1.5, XBB1.16.1, EG.5.1.3, BA.2.86.1). Data are mean of two independent experiments. The dashed line indicates the limit of detection. An anti-IgM (or IgE) specific VHH was used as a control (ctl). IC50 values are indicated in the table. b Inhibition of 5 µg/mL spike binding to hACE2-expressing HEK293 cells by increasing concentrations of VHHs B07, B09 or B10. HEK293-hACE2 cells pretreated or not with the VHHs and incubated with soluble spike (S) protein (ancestral spike) were stained with an anti-S antibody. Results were normalized for nonspecific (0%) and specific binding in the absence of inhibitor (100%). Experiments were fitted to a one-site competitive binding model. Data are mean ± SD of three (B07, B09) independent experiments or mean of two independent experiments (B10). c Displacement of S (ancestral spike) binding by B07 (left) or B09 (right) for various concentrations of S (5 µg/mL, 8 µg/mL, 10 µg/mL). Experiments were carried out as in (b) (one representative experiment out of two - 10 µg/mL, or three - 5 µg/mL, 8 µg/mL). d Relationship between the observed IC50 values for B07 or B09 displacement of S binding and initial concentration of S (ancestral spike). Results represent means ± SD of three independent experiments (5 µg/mL, 8 µg/mL) or means of two independent experiments (10 µg/mL). Linear regression analysis: R2 B07 = 0.9005; R2 B09 = 0.7896. Source Data are provided as a Source Data file.
