Extended Data Fig. 8. Antibody resistant mutants selected with VSV-SARS-CoV2 or authentic SARS-CoV-2 virus.

a. The method for assessing monoclonal antibody resistant spike protein variants is shown. SARS-CoV-2 was passaged serially in the presence of monoclonal antibodies at the increasing concentrations indicated in the figure or without antibody (no monoclonal antibody). Following passage at IC₉₀ concentrations, samples were treated with 10× IC₉₀ concentrations of monoclonal antibodies and any resultant resistant virus collected, and the genome was sequenced. Red viruses in the schematic represent selection of escape variants.

b. The escape phenotype of 6 independent plaques selected with AZD1061 was validated by demonstration of escape by testing in a PRNT. Antibody neutralization as measured by PRNT against the 6 plaque-purified, AZD1061-resistant SARS-CoV-2 viruses (blue) was compared to the parent virus WA-1 (orange) during treatment with AZD1061 or AZD7442. All plaque-purified viruses resulted from the same monoclonal antibody passage as detailed in (a). Data shown are from a single technical replicate for each of the six selected escape mutants.

c,d. The escape phenotype of independent plaques selected with AZD1061 also was validated by demonstration of loss of binding to proteins incorporating variant residues in the selected plaques using biolayer interferometry (BLI). Data shown are from a single experiment.

c. Binding traces of AZD8895 and AZD1061 to various spike trimers with kinetics curve fits. An inability to fit AZD1061 binding to the K444E S variant is due to a lack of detectable binding even at 5 µM for AZD1061.

d. Summary of AZD8895 and AZD1061 kinetic binding values to the S trimer variants from binding traces with R² indicating goodness of the fit. Relative fold-change in K_D is shown in comparison to wild-type. No detectable binding is indicated as NB.