An ACAT inhibitor regulates SARS-CoV-2 replication and antiviral T cell activity
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Abstract
The severity of disease following infection with SARS-CoV-2 is determined by viral replication kinetics and host immunity, with early T cell responses and/or suppression of viraemia driving a favourable outcome. Recent studies have uncovered a role for cholesterol metabolism in the SARS-CoV-2 life cycle and in T cell function. Here we show that blockade of the enzyme Acyl-CoA:cholesterol acyltransferase (ACAT) with Avasimibe inhibits SARS-CoV-2 entry and fusion independent of transmembrane protease serine 2 expression in multiple cell types. We also demonstrate a role for ACAT in regulating SARS-CoV-2 RNA replication in primary bronchial epithelial cells. Furthermore, Avasimibe boosts the expansion of functional SARS-CoV-2-specific T cells from the blood of patients sampled in the acute phase of infection. Thus, re-purposing of available ACAT inhibitors provides a compelling therapeutic strategy for the treatment of COVID-19 to achieve both antiviral and immunomodulatory effects.
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SciScore for 10.1101/2022.04.12.487988: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IRB: Ethics: The COVIDsortium cohort was approved by the ethical committee of UK National Research Ethics Service (20/SC/0149) and registered at https://ClinicalTrials.gov (NCT04318314).
Consent: All study participants gave written informed consent prior to inclusion in the study and all storage of samples obtained complied with the Human Tissue Act 2004.Sex as a biological variable The Acute Cohort was recruited from hospitalized patients at the Royal Free Hospital, London, and SARS-CoV-2 infection was confirmed by PCR (n=22; median age 82 years; 45% female, 55% male; 73% white, 4% black, 14% Asian, 9% other). Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line … SciScore for 10.1101/2022.04.12.487988: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IRB: Ethics: The COVIDsortium cohort was approved by the ethical committee of UK National Research Ethics Service (20/SC/0149) and registered at https://ClinicalTrials.gov (NCT04318314).
Consent: All study participants gave written informed consent prior to inclusion in the study and all storage of samples obtained complied with the Human Tissue Act 2004.Sex as a biological variable The Acute Cohort was recruited from hospitalized patients at the Royal Free Hospital, London, and SARS-CoV-2 infection was confirmed by PCR (n=22; median age 82 years; 45% female, 55% male; 73% white, 4% black, 14% Asian, 9% other). Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources On d7, PBMC were restimulated with 1μg/mL peptide + anti-CD28 in the presence of 1μg/ml Brefeldin A (Sigma-Aldrich) for 16h at 37°C, followed by antibody staining and flow cytometric analysis. anti-CD28suggested: NoneExperimental Models: Cell Lines Sentences Resources SARS-CoV-2 pseudoparticle genesis and infection: Lentiviral pseudoparticles were generated by transfecting 293T cells with p8.91 (Gag-pol), pCSFW 293Tsuggested: KCB Cat# KCB 200744YJ, RRID:CVCL_0063)Viral titres were determined by infecting Calu-3 cells with a serial dilution of virus and 48h later measuring cellular luciferase. Calu-3suggested: NoneBriefly, VeroE6 cells were seeded into 96-well flat culture plates with transparent-bottom to reach confluency (~ 5 × 104 per well). VeroE6suggested: NoneRecombinant DNA Sentences Resources SARS-CoV-2 pseudoparticle genesis and infection: Lentiviral pseudoparticles were generated by transfecting 293T cells with p8.91 (Gag-pol), pCSFW p8.91suggested: None(luciferase reporter) and a codon optimised expression construct pcDNA3.1-SARS-CoV-2-Spike, as previously described (Thompson et al., 2020). pcDNA3.1-SARS-CoV-2-Spikesuggested: NoneSoftware and Algorithms Sentences Resources The full peptide sequences can be found in (Reynolds et al., 2020)) in cRPMI (RPMI 1640 (Thermo Fisher Scientific)+2 Thermo Fisher Scientific)+2suggested: Nonerecombinant human IL-2 (PeproTech)+ 5μg/ml anti-CD28 (Invitrogen). PeproTech)+suggested: NoneAll samples were acquired on a BD Biosciences Fortessa-X20 or Fortessa and analysed using FlowJo v. FlowJosuggested: (FlowJo, RRID:SCR_008520)The Zen software also has many rendering options including removing localization errors and outliers based on brightness and size of fluorescent signals. Zensuggested: NoneStatistical analysis: Statistical analyses were performed with Prism 7.0 (GraphPad) as indicated in figure legends (Wilcoxon matched-pairs signed-rank test, Mann–Whitney test, Spearman correlation, Kruskall Wallis, unpaired t test) with significant differences marked on all figures. Prismsuggested: (PRISM, RRID:SCR_005375)GraphPadsuggested: (GraphPad Prism, RRID:SCR_002798)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: We found the following clinical trial numbers in your paper:
Identifier Status Title NCT04318314 Recruiting COVID-19: Healthcare Worker Bioresource: Immune Protection a… Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- No conflict of interest statement was detected. If there are no conflicts, we encourage authors to explicit state so.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
Results from scite Reference Check: We found no unreliable references.
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