Identifying SARS-CoV-2 antiviral compounds by screening for small molecule inhibitors of Nsp3 papain-like protease

  1. Chew Theng Lim
  2. Kang Wei Tan
  3. Mary Wu
  4. Rachel Ulferts
  5. Lee A. Armstrong
  6. Eiko Ozono
  7. Lucy S. Drury
  8. Jennifer C. Milligan
  9. Theresa U. Zeisner
  10. Jingkun Zeng
  11. Florian Weissmann
  12. Berta Canal
  13. Ganka Bineva-Todd
  14. Michael Howell
  15. Nicola O'Reilly
  16. Rupert Beale
  17. Yogesh Kulathu
  18. Karim Labib
  19. John F.X. Diffley

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Abstract

The COVID-19 pandemic has emerged as the biggest life-threatening disease of this century. Whilst vaccination should provide a long-term solution, this is pitted against the constant threat of mutations in the virus rendering the current vaccines less effective. Consequently, small molecule antiviral agents would be extremely useful to complement the vaccination program. The causative agent of COVID-19 is a novel coronavirus, SARS-CoV-2, which encodes at least nine enzymatic activities that all have drug targeting potential. The papain-like protease (PLpro) contained in the nsp3 protein generates viral non-structural proteins from a polyprotein precursor, and cleaves ubiquitin and ISG protein conjugates. Here we describe the expression and purification of PLpro. We developed a protease assay that was used to screen a custom compound library from which we identified dihydrotanshinone I and Ro 08-2750 as compounds that inhibit PLpro in protease and isopeptidase assays and also inhibit viral replication in cell culture-based assays.

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  1. Version published to 10.1042/bcj20210244
  2. ScreenIT

    SciScore for 10.1101/2021.04.07.438804: (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.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    The His-TEV bacterial sequence was codon optimized (Table S2) and cloned into plasmid pET11a at NdeI/BamHI sites (synthesised and cloned by GeneWiz) (Addgene ID 169192).
    GeneWiz
    suggested: (GENEWIZ, RRID:SCR_003177)
    Data analysis: MATLAB was used to process data (more details in nsp13 paper in this issue) Gel-based assay: 0.5 μM of enzyme was pre-incubated with the selected inhibitor drugs for 10 minutes.
    MATLAB
    suggested: (MATLAB, RRID:SCR_001622)
    Data Availability Statement: All data in this paper can be found in FigShare (https://figshare.com/).
    FigShare
    suggested: (FigShare, RRID:SCR_004328)

    Results from OddPub: Thank you for sharing your data.

    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

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  3. Version published to 10.1101/2021.04.07.438804v1 on bioRxiv