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.

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.

Determination of Patulin in Health Foods Containing Hawthorn by HPLC / 中国药师

Objective:To establish an HPLC method for the determination of patulin in health foods containing hawthorn to im-prove the quality control of patulin in related health foods. Methods:An Agilent TC-C18(2)(250 mm ×4.6 mm, 5 μm) column was used with the temperature of 30℃. The mobile phase consisted of 0. 8% tetrahydrofuran and the flow rate was 1. 0 ml·min-1 . The detection wavelength was set at 276nm. Results:The calibration curve of patulin was linear within the range of 1-20ng(r=1. 000 0), the average recovery was 92. 1% and RSD was 2. 2%(n=6). Conclusion:The method is simple, reliable and accurate, which can be used in the content determination of patulin in health foods containing hawthorn.

Effect of autologous blood transfusion on cytoimmunity during orthopedic operation / 中国医师进修杂志

Objective To explore the effect of autologous blood transfusion on cytoimmunity during orthopedic operation.Methods Two hundred and twenty-six patients undertaken selective operation from June 2007 to October 2011 were divided into observation group ( 154 cases) and control group (72 cases).The observation group was received autologous blood transfusion and the control group was given homologous transfusion.T lymphocyte subsets and natural killer (NK) cell counts in blood samples were detected by flow cytometry before operation and 1,6 days after operation.Results CD3+,CD4+,CD4+/CD8+,NK cell of the observation group and control group on 1st day of postoperation were 0.7184 ±0.0921,0.3878 ±0.0611,1.64 ± 0.27,0.1627 ± 0.0633 and 0.6548 ± 0.0852,0.3137 ± 0.0726,1.18 ± 0.31,0.1465 ± 0.0514,respectively,which decreased obviously compared with those of preoperation in both groups (0.7436 ± 0.1069,0.4301 ±0.0818,1.68 ±0.31,0.1945 ±0.0572 and 0.7537 ±0.0940,0.4453 ±0.0608,1.62 ± 0.32,0.1821 ± 0.0571 ) (P ＜ 0.05 ).The above parameters of the observation group were significandy higher than those of the control group on 1 st day of postoperation (P ＜ 0.05 ).There were no significant differences of CD3+,CD4+,CD4+/CD8+ and NK cell on the 6th day of postoperation compared with the preoperation in the observation group (P ＞ 0.05).But CD3+,CD4+,CD4+/CDs+ and NK cell on the 6th day of the postoperation were 0.6266 ± 0.0905,0.3048 ± 0.0425,1.07 ± 0.27,0.1408 ± 0.0716,which were significantly lower than the preoperation in the control group (P ＜ 0.05).Conclusions Intraoperative autologous blood transfusion have less inhibitory effect on cytoimmunity than homologous transfusion.The patients having autologous blood transfusion have a rapid recovery of cytoimmunity.