Discovery and Evaluation of Entry Inhibitors for SARS-CoV-2 and Its Emerging Variants

  1. Arpan Acharya
  2. Kabita Pandey
  3. Michellie Thurman
  4. Elizabeth Klug
  5. Jay Trivedi
  6. Kalicharan Sharma
  7. Christian L. Lorson
  8. Kamal Singh
  9. Siddappa N. Byrareddy

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Abstract

The ongoing coronavirus infectious disease 2019 (COVID-19) pandemic is caused by a novel coronavirus named Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). More than 207 million people have been infected globally, and 4.3 million have died due to this viral outbreak.

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  1. Version published to 10.1128/jvi.01437-21
  2. ScreenIT

    SciScore for 10.1101/2021.04.02.438204: (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

    Experimental Models: Cell Lines
    SentencesResources
    UNCN1T cells (a human bronchial epithelial cell line; Kerafast; cat# ENC011) were cultured in BEGM media (Bronchial Epithelial Cell Growth Medium; Lonza: cat# CC-3170) in FNC (Athena Enzyme Systems; cat# 0407) coated 96-well plates.
    UNCN1T
    suggested: RRID:CVCL_ZC91)
    MTT cell viability assay: Vero and HepG2 cells were seeded at the density of 20,000 cells/well in a 96 well plate containing 100 µL complete DMEM (Gibco, USA) supplemented with 10% FBS (Gibco, USA) and 1% Penstrep (Gibco, USA).
    Vero
    suggested: None
    HepG2
    suggested: None
    In brief, 3X106 HEK293T cells were co-transfected with a plasmid containing lentiviral backbone expressing luciferase and ZsGreen (BEI catalog number NR-52516), a lentiviral helper plasmid expressing HIV Gag-Pol (BEI catalog number NR-52517), a lentiviral helper plasmid expressing HIV Tat (BEI catalog number NR-52518) and a lentiviral helper plasmid expressing HIV Rev (BEI catalog number NR-52519) along with a plasmid expressing spike protein of SARS-CoV-2 7 using jetPRIME transfection reagent (Polyplus-transfection; NY, USA) as per manufacturer’s instruction.
    HEK293T
    suggested: None
    SARS-CoV-2 entry inhibitor screening assay: For screening SARS-CoV-2 entry inhibitors, 24 hrs before starting the assay, 20,000 HEK293T-ACE2 cells were seeded per well in a poly-l-lysine-coated 96-well plate.
    HEK293T-ACE2
    suggested: None
    In brief, Vero E6 cells were seeded in 6-well plates.
    Vero E6
    suggested: None
    The percentage inhibition of SARS-CoV-2 replication in MU-UNMC-1 and MU-UNMC-2 treated cells was calculated with respect to viral loads in untreated control wells that received DMSO (considered 0% inhibition) and negative control wells (uninfected cells).
    MU-UNMC-2
    suggested: None
    Measuring the combinational antiviral potential of MU-UNMC-2 and Remdesivir: To determine the possible synergistic antiviral effect of MU-UNMC-2 on RDV and vice versa against SARS-CoV-2 replication, we tested combined doses of the two in SARS-CoV-2 infected UNCN1T and Vero-STAT1 knockout cells.
    Vero-STAT1
    suggested: ATCC Cat# CCL-81-VHG, RRID:CVCL_YZ45)
    Software and Algorithms
    SentencesResources
    Hitfinder, Zinc database ZincDatabase, Zinc15Database
    Zinc
    suggested: (Zinc, RRID:SCR_008596)
    ChEMBL, Bingo, JChemforExcel, ChemDiff, and BindingMOAD (https://www.click2drug.org/index.php#Databases).
    ChEMBL
    suggested: (ChEMBL, RRID:SCR_014042)
    JChemforExcel
    suggested: None
    The top 500 compounds, the best Glide, was then re-docked using the ‘Induced Fit’ program of the Schrödinger Suite.
    Fit’
    suggested: None
    Statistical analysis: The CC50 and IC50 values were computed using four-parameter variable slope sigmoidal dose-response models using GraphPad Prism (version 8.0).
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    The 3-D interaction landscape between remdesivir and MU-UNMC-2 was calculated based on Loewe additive model using SynergyFinder v.2.
    SynergyFinder
    suggested: (SynergyFinder, RRID:SCR_019318)

    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
    NCT04303299RecruitingFight COVID-19 Trial
    NCT04392427Not yet recruitingNew Antiviral Drugs for Treatment of COVID-19
    NCT04260594Not yet recruitingClinical Study of Arbidol Hydrochloride Tablets in the Treat…
    NCT04355026RecruitingUse of Bromhexine and Hydroxychloroquine for Treatment of CO…
    NCT04276688CompletedLopinavir/ Ritonavir, Ribavirin and IFN-beta Combination for…
    NCT04445272CompletedClinical Trial to Evaluate the Effectiveness and Safety of T…
    NCT04433078RecruitingRepurpoSing Old Drugs TO SuppRess a Modern Threat: COVID-19 …
    NCT04358614CompletedBaricitinib Therapy in COVID-19
    NCT04338958RecruitingRuxolitinib in Covid-19 Patients With Defined Hyperinflammat…
    NCT04280705CompletedAdaptive COVID-19 Treatment Trial (ACTT)
    NCT04315948Active, not recruitingTrial of Treatments for COVID-19 in Hospitalized Adults

    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.
    • 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.

  3. Version published to 10.1101/2021.04.02.438204v1 on bioRxiv