A Single Dose of Self-Transcribing and Replicating RNA Based SARS-CoV-2 Vaccine Produces Protective Adaptive Immunity In Mice

  1. Ruklanthi de Alwis
  2. Esther S Gan
  3. Shiwei Chen
  4. Yan Shan Leong
  5. Hwee Cheng Tan
  6. Summer L Zhang
  7. Clement Yau
  8. Daiki Matsuda
  9. Elizabeth Allen
  10. Paula Hartman
  11. Jenny Park
  12. Maher Alayyoubi
  13. Hari Bhaskaran
  14. Adrian Dukanovic
  15. Belle Bao
  16. Brenda Clemente
  17. Jerel Vega
  18. Scott Roberts
  19. Jose A. Gonzalez
  20. Marciano Sablad
  21. Rodrigo Yelin
  22. Wendy Taylor
  23. Kiyoshi Tachikawa
  24. Suezanne Parker
  25. Priya Karmali
  26. Jared Davis
  27. Sean M. Sullivan
  28. Steve G. Hughes
  29. Pad Chivukula
  30. Eng Eong Ooi

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Abstract

A self-transcribing and replicating RNA (STARR™) based vaccine (LUNAR ® -COV19) has been developed to prevent SARS-CoV-2 infection. The vaccine encodes an alphavirus-based replicon and the SARS-CoV-2 full length spike glycoprotein. Translation of the replicon produces a replicase complex that amplifies and prolong SARS-CoV-2 spike glycoprotein expression. A single prime vaccination in mice led to robust antibody responses, with neutralizing antibody titers increasing up to day 60. Activation of cell mediated immunity produced a strong viral antigen specific CD8 + T lymphocyte response. Assaying for intracellular cytokine staining for IFN-γ and IL-4 positive CD4 + T helper lymphocytes as well as anti-spike glycoprotein IgG2a/IgG1 ratios supported a strong Th1 dominant immune response. Finally, single LUNAR-COV19 vaccination at both 2 μg and 10 μg doses completely protected human ACE2 transgenic mice from both mortality and even measurable infection following wild-type SARS-CoV-2 challenge. Our findings collectively suggest the potential of Lunar-COV19 as a single dose vaccine.

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  1. ScreenIT

    SciScore for 10.1101/2020.09.03.280446: (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.

    Table 2: Resources

    Antibodies
    SentencesResources
    The protein lysate (10 μg) was resolved on a 7.5% NuPAGE Tris-Acetate gel (Thermo Fisher Scientific), and the spike protein expression was analyzed by LI-COR Quantitative Western Blot system using a rabbit antibody detecting S1 (40150-T62-COV2, Sino Biologic) and a mouse antibody for S2 region (GTX632604, GeneTex) along with appropriate secondary antibodies (goat anti-rabbit 800 and goat anti-mouse 680).
    GTX632604
    suggested: (GeneTex Cat# GTX632604, RRID:AB_2864418)
    anti-rabbit
    suggested: (Rockland Cat# 18-4416-32, RRID:AB_2610839)
    anti-mouse 680
    suggested: None
    Flow cytometry: Surface staining was performed on freshly-isolated splenocytes using the following panel of antibodies and reagents: B220 (RA3-6B2), CD3 (17A2), CD4 (RM4-5), CD8α (53-6.7), CD44 (IM7), CD62L (MEL-14) and DAPI.
    B220
    suggested: None
    CD3
    suggested: (Abcam Cat# ab52305, RRID:AB_955118)
    CD4
    suggested: (Miltenyi Biotec Cat# 130-109-536, RRID:AB_2657974)
    CD8α
    suggested: None
    CD44
    suggested: None
    CD62L
    suggested: None
    Software and Algorithms
    SentencesResources
    For LUNAR-CoV19 vaccine, the reaction for RNA was performed as previously described [43] with proprietary modifications to allow highly efficient co-transcriptional incorporation of a proprietary Cap1 analogue and to achieve high quality RNA molecule of over 11,000-nt long the STARR mRNA.
    STARR
    suggested: (Starr, RRID:SCR_001071)
    Log2 Fold Changes generated were used for volcano plots constructed using Prism v8.1.0 software.
    Prism
    suggested: (PRISM, RRID:SCR_005375)
    Data acquisition was performed on a BD LSRFortessa and analyzed using FlowJo.
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Plates were imaged using ImmunoSpot analyzer and quantified using ImmunoSpot software.
    ImmunoSpot
    suggested: (ImmunoSpot Software for Analyzing ELISPOT Assays, RRID:SCR_011082)

    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

    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 10.1101/2020.09.03.280446 on bioRxiv