A recent study, currently available on bioRxiv* preprint server, shows that a novel protease inhibitor known as nirmatrelvir has potent activity against Omicron variant of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but also other variants evaluated in this paper – which supports its potential role in reducing the disease burden from coronavirus disease 2019 (COVID-19).
Study: Nirmatrelvir, an orally active Mpro inhibitor, is a potent inhibitor of SARS-CoV-2 Variants of Concern. Image Credit: NIAID
*Important notice: bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.
Notwithstanding the increased availability of different vaccine platforms, new variants of SARS-CoV-2 (with potential for enhanced replication, transmission, and immune evasion) continue to emerge, causing treatment failure and/or reduced vaccine efficacy.
Five significant variants of concern are B.1.1.7 (Alpha variant), B.1.351 (Beta variant), P.1 (Gamma variant), B.1.617.2 (Delta variant), and B.1.1.529 (Omicron variant); moreover, B.1.1.1.37 (Lambda variant) and B.1.621 (Mu variant) have been designated as variants of interest and are intensively monitored.
Alongside currently approved antiviral drug remdesivir, some other antivirals are at different stages of development or even approval. More specifically, two orally available oral antiviral drug candidates – PAXLOVID (nirmatrelvir and ritonavir) and molnupiravir – received the Emergency Use Authorization in December of 2021 from the US Food and Drug Administration (FDA).
Nirmatrelvir (PF-07321332), a specific inhibitor of the main protease of coronaviruses (usually abbreviated as Mpro, but sometimes also referred to as 3CLpro), shows an extremely potent antiviral activity against several human coronaviruses – including SARS-CoV-2, original SARS-CoV, and MERS viruses.
But the pertinent question was whether the efficacy would be retained in contact with SARS-CoV-2 variants of concern. A research group led by scientists from the Pfizer Worldwide Research, Development & Medical at Pearl River in the United States aimed to answer that question and described their findings in the recent bioRxiv preprint paper.
Knockout cells infected with SARS-CoV-2
In short, this research group has evaluated nirmatrelavir against the five aforementioned SARS-CoV-2 variants of concern and two variants of interest with the use of real-time reverse transcription PCR (qRT-PCR). A specific type of VeroE6 cell line that lacks the P-glycoprotein (Pgp) multidrug transporter gene (also known as VeroE6 P-gp knockout cells) has been used.
The latter was important, as VeroE6 cells express high levels of P-gp multidrug transporter activities, which means it was necessary to decrease the export of compounds and eliminate the confounding factors that may be introduced by potentially including a co-administered P-gp inhibitor in this type of antiviral activity studies.
Furthermore, for antiviral evaluation, the VeroE6 P-gp knockout cells have been infected at a uniform multiplicity of infection or MOI (which denotes the number of virions added per cell during the infectious process) with each of the tested SARS-CoV-2 variants.
Potent inhibition of all variants
Nirmatrelvir potently inhibited a CDC reference strain known as USA-WA1/2020 strain, but also all tested SARS-CoV-2 variants of concern and variants of interest in VeroE6 P-gp knockout cells. Mean EC50 values were 38.0 nM for the USA-WA1/2020 strain, 41.0 nM for the Alpha variant, 127.2 nM for the Beta variant, 24.9 nM for the Gamma variant, 21.2 nM for the Lambda variant, 15.9 nM for the Delta variant, 25.7 nM for the Mu variant, and 16.2 nM for the Omicron variant.
A detailed sequence analysis of the Mpro encoded by the variants revealed approximately 100% identity of active site amino acid sequences, which reflects the essential role of Mpro during viral replication and results in the ability of nirmatrelvir to exhibit robust activity across all the variants.
Moreover, based on the observed EC50 values (which is the concentration of a compound that gives half-maximal response), the researchers have concluded that nirmatrelvir has potent antiviral activity against all current variants of concern (including Omicron), as well as against two variants of interest (Lambda and Mu).
Against all the mutations
Such a result was actually expected since the Mpro active binding site is highly conserved between different viral agents, even though there is a K90R mutation in the Mpro of Alpha and Beta variants, as well as a P132H mutation in the Omicron variant and V296I mutation in the Delta variant of SARS-CoV-2.
“None of these residues, however, are located in the active site of the enzyme, while K90R and V296I substitutions are conserved and are not expected to induce dramatic structural changes”, explain study authors in this bioRxiv paper. “Therefore, it is not surprising that the observed activity of Nirmatrelvir against these variants was relatively similar”, they add.
Akin to this report, two independent groups reported good pharmacological activity of nirmatrelvir against SARS-CoV-2 variants of concern, further supporting the efficacy of this drug against the circulating variants and adding another option to our therapeutic armamentarium.
*Important notice: bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.
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