Combinatorial mRNA vaccination enhances protection against SARS-CoV-2 delta variant

  1. Renee L. Hajnik
  2. Jessica A. Plante
  3. Yuejin Liang
  4. Mohamad-Gabriel Alameh
  5. Jinyi Tang
  6. Chaojie Zhong
  7. Awadalkareem Adam
  8. Dionna Scharton
  9. Grace H. Rafael
  10. Yang Liu
  11. Nicholas C. Hazell
  12. Jiaren Sun
  13. Lynn Soong
  14. Pei-Yong Shi
  15. Tian Wang
  16. Jie Sun
  17. Drew Weissman
  18. Scott C. Weaver
  19. Kenneth S. Plante
  20. Haitao Hu

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Abstract

Emergence of SARS-CoV-2 variants of concern (VOC), including the highly transmissible delta strain, has posed challenges to current COVID-19 vaccines that principally target the viral spike protein (S). Here, we report a nucleoside-modified mRNA vaccine that expresses the more conserved viral nucleoprotein (mRNA-N). We show that mRNA-N alone was able to induce a modest but significant control of SARS-CoV-2 in mice and hamsters. Critically, by combining mRNA-N with the clinically approved S-expressing mRNA vaccine (mRNA-S-2P), we found that combinatorial mRNA vaccination (mRNA-S+N) led to markedly enhanced protection against the SARS-CoV-2 delta variant compared to mRNA-S. In a hamster model, we demonstrated that while mRNA-S alone elicited significant control of the delta strain in the lungs (∼45-fold reduction in viral loads compared to un-vaccinated control), its effectiveness in the upper respiratory tract was weak, whereas combinatorial mRNA-S+N vaccination induced markedly more robust control of the delta variant infection in the lungs (∼450-fold reduction) as well as in the upper respiratory tract (∼20-fold reduction). Immune analyses indicated that induction of N-specific immunity as well as augmented S-specific T-cell response and neutralizing antibody activity were collectively associated the enhanced protection against SARS-CoV-2 delta strain by combinatorial mRNA vaccination. These findings suggest that the combined effects of protection in the lungs and upper respiratory tract could both reduce the risk of severe disease as well as of infection and transmission.

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    SciScore for 10.1101/2021.12.08.471664: (What is this?)

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

    Table 1: Rigor

    EthicsIACUC: Animal ethics statement: The animal study protocols were approved by the Institutional Animal Care and Use Committee (IACUC) at the University of Texas Medical Branch.
    Sex as a biological variableFour groups of 4- to 5-week-old male golden Syrian hamsters (12/group), strain HsdHan: AURA (Envigo, Indianapolis, IN), were vaccinated intramuscularly (i.m.) with either PBS (mock control), mRNA-S (2 µg), mRNA-N (2 µg), or combined mRNA-S+N (2 µg for each) using a prime-boost approach at week 0 (prime) and week 3 (boost), respectively.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Plates were washed again and then incubated with horse radish peroxidase (HRP) conjugated anti-mouse IgG secondary antibody (Biolegend; 1:3000) for 1 hour at 37°C.
    anti-mouse IgG
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Western blot analysis of protein expression by mRNA-N: 293T cells in 6-well plate were directly transfected with 2 µg of mRNA-N-LNP or not transfected (as a cell-only control).
    293T
    suggested: None
    The assays were performed with Vero cells using the SARS-CoV-2 WT or delta strains at BSL-3.
    Vero
    suggested: CLS Cat# 605372/p622_VERO, RRID:CVCL_0059)
    The serum-virus mixtures were placed onto Vero E6 cell monolayer in 6-well plates for incubation for 1 hour at 37°C, followed by addition of 2 ml overlay consisting of MEM with 1.6% agarose, 2% FBS and 1% penicillin–streptomycin to the cell monolayer.
    Vero E6
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    Mouse immunization and SARS-CoV-2 challenge: Vaccine immunogenicity and effectiveness of protection were evaluated in WT Balb/c mice. 6-week-old female BALB/c mice were obtained from the Jackson Laboratories (Wilmington, MA, USA) and were housed in the animal facility of the University of Texas Medical Branch.
    BALB/c
    suggested: RRID:IMSR_ORNL:BALB/cRl)
    Software and Algorithms
    SentencesResources
    Data were analyzed using FlowJo (TreeStar).
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    The plates were washed and scanned using an ImmunoSpot 4.0 analyzer and the spots were counted with ImmunoSpot software (Cellular Technology Ltd, Cleveland, OH) to determine the spot-forming cells (SFC) per 106 splenocytes.
    ImmunoSpot
    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: We detected the following sentences addressing limitations in the study:
    A caveat of the present study is lack of WT SARS-CoV-2 challenge controls in the hamster experiments, which could be used for direct comparison with the delta challenge. However, based on previous preclinical testing of BNT162b2 and mRNA-1273 using WT virus, both mRNA-S vaccines were found to be highly effective in controlling WT SARS-CoV-2 in both upper and lower respiratory tracts [5, 8]. In addition, our analyses of serum neutralizing activity against WT and delta strains in parallel (Fig. 4h) also supported these observations as sera of mRNA-S vaccinated hamsters manifested reduced neutralizing activities by ∼5-fold against the delta strain compared to WT virus. A major finding of our study is that combinatorial mRNA-S+N vaccination led to significant increase in control of the SARS-CoV-2 delta strain infection in the lungs (Fig. 3b) and in the upper respiratory tract (Fig. 3c). Notably, combinatorial vaccination induced an additional 10-fold reduction in viral RNA copies compared to the mRNA-S alone (∼450-fold reduction in the lungs compared to the mock-vaccinated control at 2 DPI) and led to a faster resolution of the infection, indicating that this vaccine approach likely provides even better protection from the disease caused by delta. Furthermore, combinatorial vaccination also induced stronger viral control in the upper respiratory tract (by additional 4-fold compared to mRNA-S alone) (Fig. 3c), albeit to lesser extent than that in the lung. This finding indicates t...

    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.

    Results from scite Reference Check: We found no unreliable references.

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  2. Version published to 10.1101/2021.12.08.471664v1 on bioRxiv