Thiol drugs decrease SARS-CoV-2 lung injury in vivo and disrupt SARS-CoV-2 spike complex binding to ACE2 in vitro

  1. Kritika Khanna
  2. Wilfred Raymond
  3. Jing Jin
  4. Annabelle R. Charbit
  5. Irina Gitlin
  6. Monica Tang
  7. Adam D. Werts
  8. Edward G. Barrett
  9. Jason M. Cox
  10. Sharla M. Birch
  11. Rachel Martinelli
  12. Hannah S. Sperber
  13. Sergej Franz
  14. Satish Pillai
  15. Anne Marie Healy
  16. Thomas Duff
  17. Stefan Oscarson
  18. Markus Hoffmann
  19. Stefan Pöhlmann
  20. Graham Simmons
  21. John V. Fahy

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Abstract

Neutrophil-induced oxidative stress is a mechanism of lung injury in COVID-19, and drugs with a functional thiol group (“thiol drugs”), especially cysteamine, have anti-oxidant and anti-inflammatory properties that could limit this injury. Thiol drugs may also alter the redox status of the cysteine-rich SARS-CoV-2 spike glycoprotein (SARS-2-S) and thereby disrupt ACE2 binding. Using ACE2 binding assay, reporter virus pseudotyped with SARS-CoV-2 spikes (ancestral and variants) and authentic SARS-CoV-2 (Wuhan-1), we find that multiple thiol drugs inhibit SARS-2-S binding to ACE2 and virus entry into cells. Pseudoviruses carrying variant spikes were less efficiently inhibited as compared to pseudotypes bearing an ancestral spike, but the most potent drugs still inhibited the Delta variant in the low millimolar range. IC50 values followed the order of their cystine cleavage rates and lower thiol pKa values. In hamsters infected with SARS-CoV-2, intraperitoneal (IP) cysteamine decreased neutrophilic inflammation and alveolar hemorrhage in the lungs but did not decrease viral infection, most likely because IP delivery could not achieve millimolar concentrations in the airways. These data show that thiol drugs inhibit SARS-CoV-2 infection in vitro and reduce SARS-CoV-2-related lung injury in vivo and provide strong rationale for trials of systemically delivered thiol drugs as COVID-19 treatments. We propose that antiviral effects of thiol drugs in vivo will require delivery directly to the airways to ensure millimolar drug concentrations and that thiol drugs with lower thiol pKa values are most likely to be effective.

One Sentence Summary

The effect of cysteamine to decrease SARS-CoV-2 pneumonia in vivo and of multiple thiol drugs to inhibit SARS-CoV-2 infection in vitro provides rationale for clinical trials of thiol drugs in COVID-19.

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  1. ScreenIT

    SciScore for 10.1101/2020.12.08.415505: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    MEXi 293E cells (IBA Lifesciences) were cultured in MEXi culture medium (IBA Lifesciences) at 37°C, 5% CO2 and 125 RPM as described by the manufacturer.
    293E
    suggested: None
    Establishment of HEK293T cells stably expressing ACE2 and TMPRSS2 (293T-ACE2-TMPRSS2): Pseudovirus transduction experiments: 293T-ACE2-TMPRSS2 cells were plated in black 96-well tissue culture treated plates (Greiner Bio-one) 18 hours before the experiment.
    HEK293T
    suggested: None
    293T-ACE2-TMPRSS2
    suggested: None
    Virus-drug mixtures were diluted 12-fold before addition to Vero E6 cell monolayer in 96-well-plate.
    Vero E6
    suggested: None
    For cell viability measurement corresponding to pseudovirus experiment, 293T-ACE-TMPRSS2 cells were seeded in 96 well black plates 18 hours prior to the experiment.
    293T-ACE-TMPRSS2
    suggested: None
    Software and Algorithms
    SentencesResources
    For analysis of the effects of the different thiol-based drugs, area under the curve (AUC) was plotted 27 using Graphpad Prism, and ordinary one -way ANOVA followed by Dunnett’s posttest was performed.
    Graphpad Prism
    suggested: (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: No clinical trial numbers were referenced.

    Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • 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.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1101/2020.12.08.415505 on bioRxiv