Improved Neutralization of Omicron BA.4/5, BA.4.6, BA.2.75.2, BQ.1.1, and XBB.1 with Bivalent BA.4/5 Vaccine

The BNT162b2 bivalent BA.4/5 COVID-19 vaccine has been authorized to mitigate COVID-19 due to current Omicron and potentially future variants. New sublineages of SARS-CoV-2 Omicron continue to emerge and have acquired additional mutations, particularly in the spike protein, that may lead to improved viral fitness and immune evasion. The present study characterized neutralization activities against new Omicron sublineages BA.4.6, BA.2.75.2, BQ.1.1, and XBB.1 after a 4th dose (following three doses of BNT162b2) of either the original monovalent BNT162b2 or the bivalent BA.4/5 booster in individuals >55 years of age. For all participants, the 4th dose of monovalent BNT162b2 vaccine induced a 3.0x, 2.9x, 2.3x, 2.1x, 1.8x, and 1.5x geometric mean neutralizing titer fold rise (GMFR) against USA/WA1-2020 (a strain isolated in January 2020), BA.4/5, BA.4.6, BA.2.75.2, BQ.1.1, and XBB.1, respectively; the bivalent vaccine induced 5.8x, 13.0x, 11.1x, 6.7x, 8.7x, and 4.8x GMFRs. For individuals without SARS-CoV-2 infection history, BNT162b2 monovalent induced 4.4x, 3.0x, 2.5x, 2.0x, 1.5x, and 1.3x GMFRs, respectively; the bivalent vaccine induced 9.9x, 26.4x, 22.2x, 8.4x, 12.6x, and 4.7x GMFRs. These data suggest the bivalent BA.4/5 vaccine is more immunogenic than the original BNT162b2 monovalent vaccine against circulating Omicron sublineages, including BQ.1.1 that is becoming prevalent globally.

Genetic surveillance of SARS-CoV-2 Mpro reveals high sequence and structural conservation prior to the introduction of protease inhibitor Paxlovid

SARS-CoV-2 continues to represent a global health emergency as a highly transmissible, airborne virus. An important coronaviral drug target for treatment of COVID-19 is the conserved main protease (Mpro). Nirmatrelvir is a potent Mpro inhibitor and the antiviral component of Paxlovid. The significant viral sequencing effort during the ongoing COVID-19 pandemic represented a unique opportunity to assess potential nirmatrelvir escape mutations from emerging variants of SARS-CoV-2. To establish the baseline mutational landscape of Mpro prior to the introduction of Mpro inhibitors, Mpro sequences and its cleavage junction regions were retrieved from [~]4,892,000 high-quality SARS-CoV-2 genomes in GISAID. Any mutations identified from comparison to the reference sequence (Wuhan-hu-1) were cataloged and analyzed. Mutations at sites key to nirmatrelvir binding and protease functionality (e.g., dimerization sites) were still rare. Structural comparison of Mpro also showed conservation of key nirmatrelvir contact residues across the extended Coronaviridae family (alpha-, beta-, and gamma-coronaviruses). Additionally, we showed that over time the SARS-CoV-2 Mpro enzyme remained under purifying selection and was highly conserved relative to the spike protein. Now, with the EUA approval of Paxlovid and its expected widespread use across the globe, it is essential to continue large-scale genomic surveillance of SARS-CoV-2 Mpro evolution. This study establishes a robust analysis framework for monitoring emergent mutations in millions of virus isolates, with the goal of identifying potential resistance to present and future SARS-CoV-2 antivirals. ImportanceThe recent authorization of oral SARS-CoV-2 antivirals, such as Paxlovid, has ushered in a new era of the COVID-19 pandemic. Emergence of new variants, as well as selective pressure imposed by antiviral drugs themselves, raise concern for potential escape mutations in key drug binding motifs. To determine the potential emergence of antiviral resistance in globally circulating isolates and its implications for the clinical response to the COVID-19 pandemic, sequencing of SARS-CoV-2 viral isolates before, during, and after the introduction of new antiviral treatments is critical. The infrastructure built herein for active genetic surveillance of Mpro evolution and emergent mutations will play an important role in assessing potential antiviral resistance as the pandemic progresses and Mpro inhibitors are introduced. We anticipate our framework to be the starting point in a larger effort for global monitoring of the SARS-CoV-2 Mpro mutational landscape.

Innovative Randomized Phase 1 Study and Dosing Regimen Selection to Accelerate and Inform Pivotal COVID-19 Trial of Nirmatrelvir

BackgroundCOVID-19 is a continued leading cause of hospitalization and death. Safe and efficacious COVID-19 antivirals are needed urgently. Nirmatrelvir (PF-07321332), the first orally bioavailable, SARS-CoV-2 Mpro inhibitor against the coronaviridae family, has demonstrated potent preclinical antiviral activity and benign safety profile. MethodsWe report safety, tolerability, and pharmacokinetic data of nirmatrelvir with and without ritonavir as a pharmacokinetic enhancer, from an accelerated randomized, double-blind, placebo-controlled, phase 1 study. Two interleaving single-ascending dose (SAD) cohorts were evaluated in a 3-period crossover. Multiple-ascending dose (MAD) with nirmatrelvir/ritonavir twice daily (BID) dosing was evaluated over 10 days in 5 parallel cohorts. Safety was assessed, including in a supratherapeutic exposure cohort. Dose and dosing regimen for clinical efficacy evaluation in phase 2/3 clinical trials were supported by integrating modelling and simulations of SAD/MAD data with nonclinical data and a quantitative systems pharmacology model (QSP). ResultsIn SAD, MAD, and supratherapeutic exposure cohorts, nirmatrelvir/ritonavir was safe and well tolerated. Nirmatrelvir exposure and half-life were considerably increased by ritonavir, enabling selection of nirmatrelvir/ritonavir dose and regimen for phase 2/3 trials (300/100 mg BID), to achieve concentrations continuously above those required for 90% inhibition of viral replication in vitro. The QSP model suggested that a 5-day regimen would significantly decrease viral load in SARS-CoV-2-infected patients and prevent development of severe disease, hospitalization, and death. ConclusionsAn innovative and seamless trial design expedited establishment of phase 1 safety and pharmacokinetics of nirmatrelvir/ritonavir, enabling high confidence in phase 2/3 dose selection and accelerated pivotal trials initiation. NCT04756531

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

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, ), B.1.351 (Beta, {beta}), P.1 (Gamma, {gamma}), B.1.617.2 (Delta, {delta}), 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 PAXLOVID, 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 (, {beta}, {gamma}, {delta},, o) and two Variants of Interest or VOI, C.37 ({lambda}) and B.1.621 (), 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 , {beta}, {gamma}, {lambda}, {delta}, , 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.

Nirmatrelvir, Molnupiravir, and Remdesivir maintain potent in vitro activity against the SARS-CoV-2 Omicron variant

Variants of SARS-CoV-2 have become a major public health concern due to increased transmissibility, and escape from natural immunity, vaccine protection, and monoclonal antibody therapeutics. The highly transmissible Omicron variant has up to 32 mutations within the spike protein, many more than previous variants, heightening these concerns of immune escape. There are now multiple antiviral therapeutics that have received approval for emergency use by the FDA and target both the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) and the main protease (Mpro), which have accumulated fewer mutations in known SARS-CoV-2 variants. Here we test nirmatrelvir (PF-07321332), and other clinically relevant SARS-CoV-2 antivirals, against a panel of SARS-CoV-2 variants, including the novel Omicron variant, in live-virus antiviral assays. We confirm that nirmatrelvir and other clinically relevant antivirals all maintain activity against all variants tested, including Omicron.

An Oral SARS-CoV-2 Mpro Inhibitor Clinical Candidate for the Treatment of COVID-19

AbstractThe worldwide outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become an established global pandemic. Alongside vaccines, antiviral therapeutics are an important part of the healthcare response to counter the ongoing threat presented by COVID-19. Here, we report the discovery and characterization of PF-07321332, an orally bioavailable SARS-CoV-2 main protease inhibitor with in vitro pan-human coronavirus antiviral activity, and excellent off-target selectivity and in vivo safety profiles. PF-07321332 has demonstrated oral activity in a mouse- adapted SARS-CoV-2 model and has achieved oral plasma concentrations exceeding the in vitro antiviral cell potency, in a phase I clinical trial in healthy human participants. ClinicalTrials.gov Identifier: NCT04756531 One-Sentence SummaryPF-07321332 is disclosed as a novel, orally active, investigational small-molecule inhibitor of the SARS-CoV-2 main protease, which is being evaluated in clinical trials for the treatment of COVID-19.

Discovery of a Novel Inhibitor of Coronavirus 3CL Protease as a Clinical Candidate for the Potential Treatment of COVID-19

COVID-19 caused by the SARS-CoV-2 virus has become a global pandemic. 3CL protease is a virally encoded protein that is essential across a broad spectrum of coronaviruses with no close human analogs. The designed phosphate prodrug PF-07304814 is metabolized to PF-00835321 which is a potent inhibitor in vitro of the coronavirus family 3CL pro, with selectivity over human host protease targets. Furthermore, PF-00835231 exhibits potent in vitro antiviral activity against SARS-CoV-2 as a single agent and it is additive/synergistic in combination with remdesivir. We present the ADME, safety, in vitro, and in vivo antiviral activity data that supports the clinical evaluation of this compound as a potential COVID-19 treatment.