Effective Inhibition of SARS-CoV-2 Entry by Heparin and Enoxaparin Derivatives

  1. Ritesh Tandon
  2. Joshua S. Sharp
  3. Fuming Zhang
  4. Vitor H. Pomin
  5. Nicole M. Ashpole
  6. Dipanwita Mitra
  7. Weihua Jin
  8. Hao Liu
  9. Poonam Sharma
  10. Robert J. Linhardt

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Abstract

Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has caused a pandemic of historic proportions and continues to spread globally, with enormous consequences to human health. Currently there is no vaccine, effective therapeutic or prophylactic. Like other betacoronaviruses, attachment and entry of SARS-CoV-2 is mediated by the spike glycoprotein (SGP). In addition to its well-documented interaction with its receptor, human angiotensin converting enzyme 2 (hACE2), SGP has been found to bind to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we pseudotyped SARS-CoV-2 SGP on a third generation lentiviral (pLV) vector and tested the impact of various sulfated polysaccharides on transduction efficiency in mammalian cells. The pLV vector pseudotyped SGP efficiently and produced high titers on HEK293T cells. Various sulfated polysaccharides potently neutralized pLV-S pseudotyped virus with clear structure-based differences in anti-viral activity and affinity to SGP. Concentration-response curves showed that pLV-S particles were efficiently neutralized by a range of concentrations of unfractionated heparin (UFH), enoxaparin, 6- O -desulfated UFH and 6- O -desulfated enoxaparin with an IC 50 of 5.99 µg/L, 1.08 mg/L, 1.77 µg/L, and 5.86 mg/L respectively. The low serum bioavailability of intranasally administered UFH, along with data suggesting that the nasal epithelium is a portal for initial infection and transmission, suggest that intranasal administration of UFH may be an effective and safe prophylactic treatment.

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  1. ScreenIT

    SciScore for 10.1101/2020.06.08.140236: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    BlindingThe total number of cells per well was counted using the Object Count feature in Nikon Elements AR Analysis version 5.02 by a blinded observer.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    HEK293T cells (2 × 106) were plated in a 100-mm tissue culture dish and transfected the next day when they were about 75% confluent with a combination of the following plasmids: 9 µg of pLV-eGFP (a gift from Pantelis Tsoulfas (Addgene plasmid # 36083; http://n2t.net/addgene:36083; RRID:Addgene_36083), 9 µg of psPAX2 (a gift from Didier Trono (Addgene plasmid # 12260 ; http://n2t.net/addgene:12260; RRID:Addgene_12260), and 3 µg of pCAGGS-S (SARS-CoV-2)(Catalog No. NR-52310: BEI Resources) or VSV-G (a gift from Tannishtha Reya (Addgene plasmid # 14888; http://n2t.net/addgene:14888; RRID:Addgene_14888) as control.
    HEK293T
    suggested: None
    Recombinant DNA
    SentencesResources
    HEK293T cells (2 × 106) were plated in a 100-mm tissue culture dish and transfected the next day when they were about 75% confluent with a combination of the following plasmids: 9 µg of pLV-eGFP (a gift from Pantelis Tsoulfas (Addgene plasmid # 36083; http://n2t.net/addgene:36083; RRID:Addgene_36083), 9 µg of psPAX2 (a gift from Didier Trono (Addgene plasmid # 12260 ; http://n2t.net/addgene:12260; RRID:Addgene_12260), and 3 µg of pCAGGS-S (SARS-CoV-2)(Catalog No. NR-52310: BEI Resources) or VSV-G (a gift from Tannishtha Reya (Addgene plasmid # 14888; http://n2t.net/addgene:14888; RRID:Addgene_14888) as control.
    detected: RRID:Addgene_36083)
    detected: RRID:Addgene_12260)
    detected: RRID:Addgene_14888)
    Software and Algorithms
    SentencesResources
    Data was analyzed in Prism 8 (Graphpad Inc).
    Graphpad
    suggested: (GraphPad, RRID:SCR_000306)

    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 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.

  2. Version published to 10.1101/2020.06.08.140236 on bioRxiv