Development of a novel hybrid alphavirus-SARS-CoV-2 particle for rapid in vitro screening and quantification of neutralization antibodies, viral variants, and antiviral drugs

  1. Brian Hetrick
  2. Sijia He
  3. Linda D. Chilin
  4. Deemah Dabbagh
  5. Farhang Alem
  6. Aarthi Narayanan
  7. Alessandra Luchini
  8. Tuanjie Li
  9. Xuefeng Liu
  10. Joshua Copeland
  11. Angela Pak
  12. Tshaka Cunningham
  13. Lance Liotta
  14. Emanuel F. Petricoin
  15. Ali Andalibi
  16. Yuntao Wu

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Abstract

Timely development of vaccines and antiviral drugs is critical to control the COVID-19 pandemic 1–6 . Current methods for quantifying vaccine-induced neutralizing antibodies involve the use of pseudoviruses, such as the SARS-CoV-2 spike protein (S) pseudotyped lentivirus 7–14 . However, these pseudoviruses contain structural proteins foreign to SARS-CoV-2, and require days to infect and express reporter genes 15 . Here we describe the development of a new hybrid alphavirus-SARS-CoV-2 (Ha-CoV-2) particle for rapid and accurate quantification of neutralization antibodies and viral variants. Ha-CoV-2 is a non-replicating SARS-CoV-2 virus-like particle, composed of SARS-CoV-2 structural proteins (S, M, N, and E) and a RNA genome derived from a fast expressing alphavirus vector 16 . We demonstrated that Ha-CoV-2 can rapidly and robustly express reporter genes in target cells within 3-6 hours. We further validated Ha-CoV-2 for rapid quantification of neutralization antibodies, viral variants, and antiviral drugs. In addition, as a proof-of-concept, we assembled and compared the relative infectivity of a panel of 10 Ha-CoV-2 variant isolates (D614G, P.1, B.1.1.207, B.1.351, B.1.1.7, B.1.429, B.1.258, B.1.494, B.1.2, B.1.1298), and demonstrated that these variants in general are 2-10 fold more infectious. Furthermore, we quantified the anti-serum from an infected and vaccinated individual; the one dose vaccination with Moderna mRNA-1273 has greatly increased the anti-serum titer for approximately 6 fold. The post-vaccination serum has also demonstrated various neutralizing activities against all 9 variants tested. These results demonstrated that Ha-CoV-2 can be used as a robust platform for rapid quantification of neutralizing antibodies against SARS-CoV-2 and its variants.

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  1. ScreenIT

    SciScore for 10.1101/2020.12.22.423965: (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

    Antibodies
    SentencesResources
    Membranes were then incubated with Antimouse IgG, HRP-linked Antibody (Cell signaling) (1: 2000 dilution) for 60 min at room temperature.
    Antimouse IgG
    suggested: (Roche Cat# 760-500, RRID:AB_2753116)
    Experimental Models: Cell Lines
    SentencesResources
    Lenti-pseudovirus was assembled by cotransfection of HEK293T cells with SARS-CoV-2 S expression vector (0.5 μg), pCMVΔR8.2 (7.5 μg), and pLTR-Tat-IRES-Luc (10 μg) as previously described 15.
    HEK293T
    suggested: None
    Viral infectivity assay: Ha-CoV-2 particles were used to infect HEK293T(ACE2/TMPRSS2) cells (a gift from Virongy LLC, Manassas, VA), Calu-3 cells (ATCC), HEK293T cells (ATCC) and primary monkey kidney cells provided by Dr. Xuefeng Liu.
    Calu-3
    suggested: None
    Briefly, Vero cells (ATCC) in 12-well plates (2×105 cells per well) were infected with virus for 1 hour at 37 °C.
    Vero
    suggested: None
    HEK293T(ACE2/TMPRSS2) cells were pretreated for 1 hour with serially diluted Arbidol.
    HEK293T(ACE2/TMPRSS2
    suggested: None
    Software and Algorithms
    SentencesResources
    After infection, a 1:1 overlay, consisting of 0.6% agarose and 2X EMEM without phenol red (Quality Biological), supplemented with 10% fetal bovine serum (FBS) (Gibco), was added to each well.
    Quality Biological
    suggested: None

    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 10.1101/2020.12.22.423965 on bioRxiv