Alpha 1 Antitrypsin is an Inhibitor of the SARS-CoV-2–Priming Protease TMPRSS2

  1. Nurit P Azouz
  2. Andrea Klingler
  3. Victoria Callahan
  4. Ivan Akhrymuk
  5. Katarina Elez
  6. Lluís Raich
  7. Brandon Henry
  8. Justin Benoit
  9. Stefanie Benoit
  10. Frank Noé
  11. Kylene Kehn-Hall
  12. Marc Rothenberg

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Abstract

Background: Host proteases have been suggested to be crucial for dissemination of MERS, SARS-CoV, and SARS-CoV-2 coronaviruses, but the relative contribution of membrane versus intracellular proteases remains controversial. Transmembrane serine protease 2 (TMPRSS2) is regarded as one of the main proteases implicated in the coronavirus S protein priming, an important step for binding of the S protein to the angiotensin-converting enzyme 2 (ACE2) receptor before cell entry.  Methods: We developed a cell-based assay to identify TMPRSS2 inhibitors. Inhibitory activity was established in SARS-CoV-2 viral load systems. Results: We identified the human extracellular serine protease inhibitor (serpin) alpha 1 antitrypsin (A1AT) as a novel TMPRSS2 inhibitor. Structural modeling revealed that A1AT docked to an extracellular domain of TMPRSS2 in a conformation that is suitable for catalysis, resembling similar serine protease inhibitor complexes. Inhibitory activity of A1AT was established in a SARS-CoV-2 viral load system. Notably, plasma A1AT levels were associated with COVID-19 disease severity.  Conclusions: Our data support the key role of extracellular serine proteases in SARS CoV-2 infections and indicate that treatment with serpins, particularly the FDA-approved drug A1AT, may be effective in limiting SARS-CoV-2 dissemination by affecting the surface of the host cells.

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  1. Version published to 10.20411/pai.v6i1.408
  2. Version published to 10.1101/2020.05.04.077826v2 on bioRxiv
  3. ScreenIT

    SciScore for 10.1101/2020.05.04.077826: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: COVID-19 plasma analysis: Adults who presented to the University of Cincinnati Medical Center (UCMC) Emergency Department (ED) with suspected COVID-19 and had a clinically indicated blood draw were prospectively enrolled via an institutional review board–approved waiver of informed consent.
    Consent: COVID-19 plasma analysis: Adults who presented to the University of Cincinnati Medical Center (UCMC) Emergency Department (ED) with suspected COVID-19 and had a clinically indicated blood draw were prospectively enrolled via an institutional review board–approved waiver of informed consent.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Gels were transferred to nitrocellulose membranes (Life Technologies) and probed with the primary antibodies rabbit anti-V5 (Bethyl Laboratories), mouse anti-TMPRSS2 (Santa Cruz), and rabbit anti–human GAPDH (ABCAM) and subsequently with the secondary antibody IRDye 800RD goat anti-rabbit and IRDye 680RD goat anti-mouse (LI-COR Biosciences).
    anti-V5
    suggested: None
    anti-TMPRSS2
    suggested: None
    anti–human GAPDH
    suggested: None
    anti-rabbit
    suggested: (LI-COR Biosciences Cat# 925-68073, RRID:AB_2716687)
    anti-mouse
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    HEK-293T cell culture and transfection: HEK-293T cells were grown in Dulbecco’s modified eagle media (DMEM) supplemented with 10% fetal bovine serum (FBS) and seeded in a black, 96-well plate (75,000 cells/well).
    HEK-293T
    suggested: None
    Caco-2 cells were plated in 96-well plates.
    Caco-2
    suggested: None
    Software and Algorithms
    SentencesResources
    Statistical analyses were performed using GraphPad Prism (GraphPad Software Incorporated).
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    GraphPad
    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 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:
    • 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.

  4. Version published to 10.1101/2020.05.04.077826v1 on bioRxiv