Main protease mutants of SARS-CoV-2 variants remain susceptible to nirmatrelvir (PF-07321332)

Ullrich, Sven; Ekanayake, Kasuni B; Otting, Gottfried; Nitsche, Christoph

doi:10.1101/2021.11.28.470226

Cited by 20 publications

(20 citation statements)

References 47 publications

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“…A comparison of the P132H mutated enzyme potency to that of wildtype M pro by t-Test using the log of individual Ki values with one tail and un-equal variance determined a p-value of 0.07 indicating that the Ki values are not statistically different. These observations are consistent with recently reported biochemical characterization with similar kcat/Km and nirmatrelvir potency values (9). Nirmatrelvir also appears to retain its in vitro antiviral efficacy against Omicron relative to wildtype (10).…”

Section: Main Textsupporting

confidence: 92%

Structural basis for Nirmatrelvir in vitro efficacy against SARS-CoV-2 variants

Greasley

Noell

Plotnikova

et al. 2022

Preprint

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The COVID-19 pandemic continues to be a public health threat with emerging variants of SARS-CoV-2. Nirmatrelvir (PF-07321332) is a reversible, covalent inhibitor targeting the main protease (Mpro) of SARS-CoV-2 and the active protease inhibitor in PAXLOVID (nirmatrelvir tablets and ritonavir tablets). One of the predominant SARS-CoV-2 variants emerging is the B.1.1.529 Omicron harboring a mutation at amino acid position 132 in the Mpro changing a proline to a histidine (P132H). In vitro biochemical enzymatic assay characterization of the enzyme kinetics of the Omicron Mpro (P132H) demonstrate that it is catalytically comparable to wildtype and that nirmatrelvir has similar potency against both wildtype and Omicron (P132H) Mpro with Ki of 0.933nM (wildtype) and 0.635nM (P132H) each, respectively. This observation is reinforced by our structural determination of nirmatrelvir bound to the omicron Mpro at 1.63Å resolution. These in vitro data suggest that PAXLOVID has the potential to maintain plasma concentrations of nirmatrelvir many-fold times higher than the amount required to stop the SARS-CoV-2 variant Omicron from replicating in cells.

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Section: Main Textsupporting

confidence: 92%

Structural basis for Nirmatrelvir in vitro efficacy against SARS-CoV-2 variants

Greasley

Noell

Plotnikova

et al. 2022

Preprint

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“…In this study, we evaluated the currently designated VOCs and two VOIs for susceptibility against Nirmatrelvir to determine in vitro efficacy against emerging SARS-CoV-2 variants. In agreement with initial reports [13, 14], Nirmatrelvir exhibits potent activity against Omicron and other evaluated variants and provides critical data supporting the application of Nirmatrelvir in reducing the disease burden from COVID-19.…”

Section: Introductionsupporting

confidence: 90%

Nirmatrelvir, an orally active Mpro inhibitor, is a potent inhibitor of SARS-CoV-2 Variants of Concern

Devendra

Yurgelonis

McMonagle

et al. 2022

Preprint

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New variants of SARS-CoV-2 with potential for enhanced transmission, replication, and immune evasion capabilities continue to emerge causing reduced vaccine efficacy and/or treatment failure. As of January 2021, the WHO has defined five variants of concern (VOC): B.1.1.7 (Alpha, α), B.1.351 (Beta, β), P.1 (Gamma, γ), B.1.617.2 (Delta, δ), and B.1.1.529 (Omicron, o). To provide a therapeutic option for the treatment of COVID-19 and variants, Nirmatrelvir, the antiviral component of PAXLOVIDTM, an oral outpatient treatment recently authorized for conditional or emergency use treatment of COVID-19, was developed to inhibit SARS-CoV-2 replication. Nirmatrelvir (PF-07321332) is a specific inhibitor of coronavirus main protease (Mpro, also referred to as 3CLpro), with potent antiviral activity against several human coronaviruses, including SARS-CoV-2, SARS-CoV, and MERS (Owen et al, Science 2021. doi: 10.1126/science.abl4784). Here, we evaluated PF-07321332 against the five SARS-CoV-2 VOC (α, β, γ, δ, o) and two Variants of Interest or VOI, C.37 (λ) and B.1.621 (μ), using qRT-PCR in VeroE6 cells lacking the P-glycoprotein (Pgp) multidrug transporter gene (VeroE6 P-gp knockout cells). Nirmatrelvir potently inhibited USA-WA1/2020 strain, and α, β, γ, λ, δ, μ, and o variants in VeroE6 P-gp knockout cells with mean EC50 values 38.0 nM, 41.0 nM, 127.2 nM, 24.9 nM, 21.2 nM, 15.9 nM, 25.7 nM and 16.2 nM, respectively. Sequence analysis of the Mpro encoded by the variants showed ~100% identity of active site amino acid sequences, reflecting the essential role of Mpro during viral replication leading to ability of Nirmatrelvir to exhibit potent activity across all the variants.

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“…also characterized six M pro mutants identified from the circulating SARS-CoV-2 variants. 8 The k cat /K m values for WT and Omicron M pro were 0.016 and 0.023 S -1 µM -1 , which were similar to the values obtained in our study ( Fig. 1B ).…”

Section: Resultssupporting

confidence: 89%

The P132H mutation in the main protease of Omicron SARS-CoV-2 decreases thermal stability without compromising catalysis or small-molecule drug inhibition

Sacco

Gongora

et al. 2022

Preprint

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The main protease (Mpro) of SARS-CoV-2 is essential for viral replication and is considered to be one of the most promising SARS-CoV-2 drug targets. While Mpro mutations have occurred in many variants, including Omicron (B.1.1.529), their structural and biochemical importance have, until this point, remained uncharacterized. Using X-ray crystallography, we show the Omicron Mpro mutant (P132H) induces a small rearrangement of residues distal from the active site. While enzymatic activity and small-molecule inhibition appears unchanged, the melting temperature (Tm) of Omicron Mpro is 2.6 ° C lower than Alpha and Delta Mpro. Yet, when incubated with inhibitors, these enzymes have nearly identical Tm values. The physiological importance of the P132H mutant is unclear; however, we show residue 132 is located at the interface of the dimerization and catalytic domains and is frequently mutated in other variants. Lower thermal stability may indicate increased flexibility that can potentially broaden substrate profile or alter inhibitor binding. Therefore, structural insights are key to anticipating future mutations that may promote drug-resistance and are especially important at present, given the recent approval of nirmatrelvir (PAXLOVID), an oral SARS-CoV-2 Mpro inhibitor.

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Main protease mutants of SARS-CoV-2 variants remain susceptible to nirmatrelvir (PF-07321332)

Cited by 20 publications

References 47 publications

Structural basis for Nirmatrelvir in vitro efficacy against SARS-CoV-2 variants

Structural basis for Nirmatrelvir in vitro efficacy against SARS-CoV-2 variants

Nirmatrelvir, an orally active Mpro inhibitor, is a potent inhibitor of SARS-CoV-2 Variants of Concern

The P132H mutation in the main protease of Omicron SARS-CoV-2 decreases thermal stability without compromising catalysis or small-molecule drug inhibition

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