Antibodies that potently inhibit or enhance SARS-CoV-2 spike protein-ACE2 interaction isolated from synthetic single-chain antibody libraries

  1. Matthew D. Beasley
  2. Sanja Aracic
  3. Fiona M. Gracey
  4. Ruban Kannan
  5. Avisa Masarati
  6. S. R. Premaratne
  7. Madhara Udawela
  8. Rebecca E. Wood
  9. Shereen Jabar
  10. Nicole L. Church
  11. Thien-Kim Le
  12. Dahna Makris
  13. Bradley K. McColl
  14. Ben R. Kiefel

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Abstract

Antibodies with high affinity against the receptor binding domain (RBD) of the SARS-CoV-2 S1 ectodomain were identified from screens using the Retained Display™ (ReD) platform employing a 1 × 10 11 clone single-chain antibody (scFv) library. Numerous unique scFv clones capable of inhibiting binding of the viral S1 ectodomain to the ACE2 receptor in vitro were characterized. To maximize avidity, selected clones were reformatted as bivalent diabodies and monoclonal antibodies (mAb). The highest affinity mAb completely neutralized live SARS-CoV-2 virus in cell culture for four days at a concentration of 6.7 nM, suggesting potential therapeutic and/or prophylactic use. Furthermore, scFvs were identified that greatly increased the interaction of the viral S1 trimer with the ACE2 receptor, with potential implications for vaccine development.

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  1. ScreenIT

    SciScore for 10.1101/2020.07.27.224089: (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

    Antibodies
    SentencesResources
    The scFvs were re-formatted to IgG1 antibodies and transiently expressed in HEK SUS cells and purified with Protein A resin.
    IgG1
    suggested: None
    ACE2-S1 inhibition assay: The ability of RBD-binding antibodies to block the high-affinity interaction between SARS-CoV-2 RBD and human ACE2 protein was tested in a bead-binding assay.
    human ACE2
    suggested: None
    SARS-CoV-2 virus neutralization assay: The potency of the antibody clones in different formats (scFv, diabody, IgG1) to protect cells in culture from infection by SARS-CoV-2 virus was assayed using quadruplicate wells of Vero cells in culture for four days with antibody and virus25.
    virus25
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    The scFvs were re-formatted to IgG1 antibodies and transiently expressed in HEK SUS cells and purified with Protein A resin.
    HEK SUS
    suggested: ATCC Cat# CRL-1573.3, RRID:CVCL_4W07)
    The neutralization titer is the lowest antibody concentration that neutralized the infectivity of 100 TCID50 of virus, read as the absence of cytopathic effect (CPE) in Vero cells on day 5 after infection.
    Vero
    suggested: CLS Cat# 605372/p622_VERO, RRID:CVCL_0059)

    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.
    • No funding statement was detected.
    • 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. ScreenIT

    SciScore for 10.1101/2020.07.27.224089: (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

    Antibodies
    SentencesResources
    Results Prior to the COVID-19 outbreak, previous studies of SARS-CoV and MERS-CoV had identified the RBD within the S1 subunit of the viral spike protein as the binding site of multiple neutralizing antibodies19,20.
    antibodies19,20
    suggested: None
    Discussion Rapid development of anti-SARS CoV-2 RBD antibodies from the ReD platform is demonstrated.
    anti-SARS CoV-2 RBD
    suggested: None
    However, B-cell derived antibodies may have intrinsic issues with production at the scale required for response to COVID-19.
    COVID-19
    suggested: None
    The remarkably high affinity of S1 RBD for ACE2 (KD between 1.2 nM22 and 44 nM5) indicates that competition with this binding event requires antibodies with very high affinity for the S1 protein.
    ACE2
    suggested: None
    At the time of writing, this appears to be the first report of antibodies with these properties for the SARS-CoV-2 S1 glycoprotein.
    SARS-CoV-2 S1 glycoprotein.
    suggested: (Abcam Cat# ab273074, AB_2847846)
    ACE2-S1 inhibition assay The ability of RBD-binding antibodies to block the high-affinity interaction between SARSCoV-2 RBD and human ACE2 protein was tested in a bead-binding assay.
    human ACE2
    suggested: None
    SARS-CoV-2 virus neutralization assay The potency of the antibody clones in different formats (scFv, diabody, IgG1) to protect cells in culture from infection by SARS-CoV-2 virus was assayed using quadruplicate wells of Vero cells in culture for four days with antibody and virus25.
    IgG1
    suggested: None
          <div style="margin-bottom:8px">
        <div><b>virus25</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The scFvs were re-formatted to IgG1 antibodies and transiently expressed in HEK SUS cells and purified with Protein A resin.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>HEK SUS</b></div>
        <div>suggested: ATCC Cat# CRL-1573.3, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_4W07">CVCL_4W07</a></div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The neutralization titer is the lowest antibody concentration that neutralized the infectivity of 100 TCID50 of virus, read as the absence of cytopathic effect (CPE) in Vero cells on day 5 after infection.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Vero</b></div>
        <div>suggested: CLS Cat# 605372/p622_VERO, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0059">CVCL_0059</a></div>
      </div>
    </td></tr></table>

    Data from additional tools added to each annotation on a weekly basis.

    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 is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.

  3. Version published to 10.1101/2020.07.27.224089 on bioRxiv