Nodosome inhibition as a novel broad-spectrum antiviral strategy against arboviruses and SARS-CoV-2

  1. Daniel Limonta
  2. Lovely Dyna-Dagman
  3. William Branton
  4. Tadashi Makio
  5. Richard W. Wozniak
  6. Christopher Power
  7. Tom C. Hobman

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Abstract

In the present report, we describe two small molecules with broad-spectrum antiviral activity. These drugs block formation of the nodosome. The studies were prompted by the observation that infection of human fetal brain cells with Zika virus (ZIKV) induces expression of nucleotide-binding oligomerization domain-containing protein 2 (NOD2), a host factor that was found to promote ZIKV replication and spread. A drug that targets NOD2 was shown to have potent broad-spectrum antiviral activity against other flaviviruses, alphaviruses and SARS-CoV-2, the causative agent of COVID-19. Another drug that inhibits the receptor-interacting serine/threonine-protein kinase 2 (RIPK2) which functions downstream of NOD2, also decreased replication of these pathogenic RNA viruses. The broad-spectrum action of nodosome targeting drugs is mediated, at least in part, by enhancement of the interferon response. Together, these results suggest that further preclinical investigation of nodosome inhibitors as potential broad-spectrum antivirals is warranted.

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  1. ScreenIT

    SciScore for 10.1101/2020.11.05.370767: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementConsent: Ethical Approval: Human fetal brain tissues were obtained from 15-19-week aborted fetuses with written consent from the donor parents and prior approval under protocol 1420 (University of Alberta Human Research Ethics Board).
    IRB: Ethical Approval: Human fetal brain tissues were obtained from 15-19-week aborted fetuses with written consent from the donor parents and prior approval under protocol 1420 (University of Alberta Human Research Ethics Board).
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Immunofluorescence staining and cell imaging: Infected cells grown on coverslips were fixed with 4% paraformaldehyde and permeabilized/blocked with a Triton-X100 (0.2%)/BSA (3%) solution and then incubated with mouse anti-Flavivirus Group Antigen 4G2 (Millipore, Burlington, MA), mouse anti-alphavirus capsid (kindly provided by Dr. Andres Merits at University of Tartu), or mouse anti-spike SARS-CoV/SARS-CoV-2 (GenTex, Irvine, CA) at room temperature for 1.5 hour, washed and then incubated with Alexa Fluor secondary antibodies against mouse and DAPI for 1 hour at room temperature.
    anti-Flavivirus Group Antigen 4G2
    suggested: None
    anti-alphavirus capsid
    suggested: None
    anti-spike SARS-CoV/SARS-CoV-2
    suggested: (Sino Biological Cat# 40150-D001-H, RRID:AB_2857930)
    Experimental Models: Cell Lines
    SentencesResources
    SARS-CoV-2 (SARS-CoV-2/CANADA/VIDO 01/2020) was propagated in Vero-E6 cells grown in Dulbecco’s Modified Eagle Medium (DMEM, Thermo Fisher Scientific).
    Vero-E6
    suggested: None
    A549, Huh7, U251, Vero (ATCC, Manassas, VA) and ACE2-hyperexpressing SK-N-SH cells were maintained in DMEM while HEL-18 human primary embryonic pulmonary fibroblasts and Calu-3 cells (ATCC) were maintained in Roswell Park Memorial Institute 1640 medium (RPMI, Thermo Fisher Scientific) and MEM respectively.
    Huh7
    suggested: None
    U251
    suggested: None
    SK-N-SH
    suggested: None
    HEL-18
    suggested: None
    Calu-3
    suggested: None
    Viral titer assay: Titers were determined in Vero CCL-81 and Vero-E6 for arboviruses (flaviviruses and alphaviruses) and coronaviruses respectively.
    Vero
    suggested: None
    A549 cells infected with arboviruses or ACE2-SK-N-SH cells infected with SARS-CoV-2 (MOI=0.05-5) were treated with the RIPK2 inhibitor GSK583 (28) (Sigma-Aldrich) for 24 hours.
    A549
    suggested: None
    ACE2-SK-N-SH
    suggested: None
    Software and Algorithms
    SentencesResources
    Poly(I:C) transfection: HFAs grown in 96-well plates (Greiner, Kremsmünster, Austria) were transfected with polyinosinic:polycytidylic acid (Poly(I:C) (Sigma-Aldrich, St. Louis, MO) at a concentration of 0.02 or 0.1 μg/well using TransIT (0.3 μL/well, Mirus Bio LLC, Madison, WI).
    TransIT
    suggested: None
    Images were analyzed using Volocity or Gen5 software.
    Volocity
    suggested: (Volocity 3D Image Analysis Software, RRID:SCR_002668)
    Gen5
    suggested: (Gen5, RRID:SCR_017317)
    GraphPad Prism software 5.0 (GraphPad Software Inc., La Jolla, CA) was used in all statistical analyses.
    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

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  2. Version published to 10.1101/2020.11.05.370767 on bioRxiv