Drug repurposing screens reveal FDA approved drugs active against SARS-Cov-2

  1. Mark Dittmar
  2. Jae Seung Lee
  3. Kanupriya Whig
  4. Elisha Segrist
  5. Minghua Li
  6. Kellie Jurado
  7. Kirandeep Samby
  8. Holly Ramage
  9. David Schultz
  10. Sara Cherry

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Abstract

There are an urgent need for antivirals to treat the newly emerged SARS-CoV-2. To identify new candidates we screened a repurposing library of ~3,000 drugs. Screening in Vero cells found few antivirals, while screening in human Huh7.5 cells validated 23 diverse antiviral drugs. Extending our studies to lung epithelial cells, we found that there are major differences in drug sensitivity and entry pathways used by SARS-CoV-2 in these cells. Entry in lung epithelial Calu-3 cells is pH-independent and requires TMPRSS2, while entry in Vero and Huh7.5 cells requires low pH and triggering by acid-dependent endosomal proteases. Moreover, we found 9 drugs are antiviral in lung cells, 7 of which have been tested in humans, and 3 are FDA approved including Cyclosporine which we found is targeting Cyclophilin rather than Calcineurin for its antiviral activity. These antivirals reveal essential host targets and have the potential for rapid clinical implementation.

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  1. ScreenIT

    SciScore for 10.1101/2020.06.19.161042: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIACUC: All work with infectious virus was performed in a Biosafety Level 3 laboratory and approved by the Institutional Biosafety Committee and Environmental Health and Safety. Infections: Cells were plated in 384 well plates (20μL/well) 3,000 cells per well for Vero, 3,000 cells per well Huh7.5, 7,500 cells per well Calu-3.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Cells were blocked (2% BSA/PBST) for 60 minutes and incubated in primary antibody (anti-dsRNA J2) overnight at 4C.
    anti-dsRNA J2
    suggested: (SCICONS Cat# 10010200, RRID:AB_2651015)
    Experimental Models: Cell Lines
    SentencesResources
    Stocks were prepared by infection of Vero E6 cells in 2% serum plus 10mM HEPES for five days, freeze-thawed, and clarified by centrifugation (PO).
    Vero E6
    suggested: None
    This seed stock was amplified in Vero CCL81 (P1) at 1.5×106 TCID50/mL.
    Vero CCL81
    suggested: None
    One hour later cells were infected with SARS-CoV-2 (Vero, MOI=1; Huh7.5 MOI=1; Calu-3 MOI=0.5) Cells were fixed (30hpi Vero and Huh7,5, 48hpi Calu-3) in 4% formaldehyde/PBS for 15min at room temperature and then washed three times with PBST.
    Vero
    suggested: None
    Huh7,5
    suggested: None
    Selectivity index (SI) was calculated as a ratio of drug’s CC50 and IC50 values (SI = CC50/IC50). RT-qPCR: Huh7.5 (750,000 cells/well) or Calu-3 cells (750,000 cells/well) were plated in 6 well plates.
    Calu-3
    suggested: None
    Total RNA was purified using Trizol (Invitrogen) followed by RNA Clean and Concentrate kit (Zymo Researc) 24 hpi for Huh7.5 or 48 hpi for Calu-3.
    Huh7.5
    suggested: RRID:CVCL_YU20)
    Software and Algorithms
    SentencesResources
    The total number of cells and the number of infected cells were measured using MetaXpress 5.3.3 cell scoring module, and the percentage of infected cells was calculated.
    MetaXpress
    suggested: (MetaXpress, RRID:SCR_016654)
    A non-linear regression curve fit analysis (GraphPad Prism 8) was performed on POC Infection and cell viability using log10 transformed concentration values to calculate IC50 values for Infection and CC50 values for cell viability for each drug/cell line combination.
    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: We found the following clinical trial numbers in your paper:

    IdentifierStatusTitle
    NCT04333550RecruitingApplication of Desferal to Treat COVID-19

    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.
    • 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. Version published to 10.1101/2020.06.19.161042v1 on bioRxiv