Remdesivir and GS-441524 Retain Antiviral Activity against Delta, Omicron, and Other Emergent SARS-CoV-2 Variants

Figures

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  FIG 1. Representative dose-response curves of remdesivir and GS-441524 against SARS-CoV-2 VOC. Shown are dose-response curves of RDV (A and B) and GS-441524 (C) activity against the WA1 reference isolate and SARS-CoV-2 VOCs in A549-ACE2-TMPRSS2 cells by plaque reduction assay (PRA) (A) or ELISA (B and C). In the PRA, infected cell supernatants were harvested at 48 hpi (solid lines) or 72 hpi (dashed lines) and analyzed by plaque assay on Vero-TMPRSS2 cells. For ELISA, infected cells were fixed at ~48 hpi (solid lines) or ~72 hpi (dashed lines) and processed. The data shown are means and standard deviations from representative experiments that were performed in biological quadruplicates (PRA) or triplicates (ELISA) at each compound concentration. Average calculated EC50 values and fold change from WA1 reference can be found in Table 1.

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  FIG 2. Structural model of Nsp12 highlighting key amino acid substitutions in relation to the active site. Preincorporated remdesivir triphosphate (RDV-TP) was modeled into the cryo-EM structure of the polymerase complex (PDB no. 6XEZ) (37). Nsp12 is represented in green, with Nsp8 in yellow, and Nsp7 in white. The prevalent amino acid substitution P323L, seen in all variants, was measured to be 28.6 Å from RDV-TP (P323 Cα-RDV-TP C1′), whereas G671S, seen in the Delta variant, is at 24.9 Å. Of all the amino acid substitutions reported here, F694Y, seen at low frequency in Delta and Omicron, comes closest to the active site, at 12.2 Å. A computational analysis suggests that the substitutions have no meaningful impact on RDV-TP binding affinity.

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  FIG 3. Recombinant Omicron viruses retain susceptibility to RDV and GS-441524. Shown are ELISA dose-response curves of RDV (solid lines and solid symbols) and GS-441524 (dashed lines and open symbols) activity against recombinant Omicron viruses with (dark purple) or without (light purple) the F694Y substitution compared with WA1 (black), and Omicron (gray) clinical isolates run in parallel. The data shown are means and standard deviations from a representative 72-hpi nucleoprotein ELISA that was performed with biological triplicates at each compound concentration. Average calculated EC50 values are in Table 3.

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  FIG 4. Prevalent Nsp12 substitutions in Delta and Omicron retain susceptibility to RDV and GS-441524. Shown are dose-response curves of RDV (solid lines and solid symbols) and GS-441524 (dashed lines and open symbols) activity against recombinant viruses with/without prevalent Nsp12 substitutions containing a Nano luciferase (Nluc) (A) or firefly luciferase (Fluc) (B) transgene. The data shown are means and standard deviations from a representative experiment that was performed in biological duplicates at each compound concentration. Average calculated EC50 values and fold change from recombinant WA1 references are in Table 4.