Immune response to vaccine candidates based on different types of nanoscaffolded RBD domain of the SARS-CoV-2 spike protein

  1. Duško Lainšček
  2. Tina Fink
  3. Vida Forstnerič
  4. Iva Hafner-Bratkovič
  5. Sara Orehek
  6. Žiga Strmšek
  7. Mateja Manček-Keber
  8. Peter Pečan
  9. Hana Esih
  10. Špela Malenšek
  11. Jana Aupič
  12. Petra Dekleva
  13. Tjaša Plaper
  14. Sara Vidmar
  15. Lucija Kadunc
  16. Mojca Benčina
  17. Neža Omersa
  18. Gregor Anderluh
  19. Florence Pojer
  20. Kelvin Lau
  21. David Hacker
  22. Bruno Correia
  23. David Peterhoff
  24. Ralf Wagner
  25. Roman Jerala

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Abstract

Effective and safe vaccines against SARS-CoV-2 are highly desirable to prevent casualties and societal cost caused by Covid-19 pandemic. The receptor binding domain (RBD) of the surface-exposed spike protein of SARS-CoV-2 represents a suitable target for the induction of neutralizing antibodies upon vaccination. Small protein antigens typically induce weak immune response while particles measuring tens of nanometers are efficiently presented to B cell follicles and subsequently to follicular germinal center B cells in draining lymph nodes, where B cell proliferation and affinity maturation occurs. Here we prepared and analyzed the response to several DNA vaccines based on genetic fusions of RBD to four different scaffolding domains, namely to the foldon peptide, ferritin, lumazine synthase and β-annulus peptide, presenting from 6 to 60 copies of the RBD on each particle. Scaffolding strongly augmented the immune response with production of neutralizing antibodies and T cell response including cytotoxic lymphocytes in mice upon immunization with DNA plasmids. The most potent response was observed for the 24-residue β-annulus peptide scaffold that forms large soluble assemblies, that has the advantage of low immunogenicity in comparison to larger scaffolds. Our results support the advancement of this vaccine platform towards clinical trials.

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  1. ScreenIT

    SciScore for 10.1101/2020.08.28.244269: (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 variableMouse immunization studies: To test the immunogenicity of DNA vaccines female 8-10 weeks BALB/c OlaHsd mice (Envigo, Italy) were used for immunization protocols.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    For total IgG determination, goat anti mouse IgG (H+L)-HRP antibodies (Jackson ImmunoResearch; 115-035-003) were used.
    anti mouse IgG
    suggested: None
    H+L)-HRP
    suggested: None
    To determine specific types of anti-RBD antibodies in mouse sera goat anti-mouse IgG1-HRP (Abcam; ab97240)
    anti-RBD
    suggested: None
    anti-mouse IgG1-HRP
    suggested: (Santa Cruz Biotechnology Cat# sc-2060, RRID:AB_631760)
    The next day, cells were infected with VSVΔG*/G virus in serum-free medium for 1h, after which medium was removed and cells were washed with PBS before complete medium supplemented with anti-VSV-G antibody (8G5F11, Kerafast) was added to the cells.
    anti-VSV-G
    suggested: (Absolute Antibody Cat# Ab01401-2.0, RRID:AB_2883992)
    Experimental Models: Cell Lines
    SentencesResources
    Cell culture: The human embryonic kidney (HEK) 293, HEK293T (ATCC) cell line and mouse NIH-3T3 cell (ATCC) line was cultured in DMEM (Invitrogen) supplemented with 10% FBS (BioWhittaker) at 37 °C in a 5% CO2 environment.
    HEK
    suggested: None
    HEK293T
    suggested: None
    HEK293E cells were cultured in EX-CELL 293 serum-free medium (Sigma) and incubated with agitation at 37°C and 4.5% CO2.
    HEK293E
    suggested: None
    For neutralization assay, HEK293 were seeded (2.5*104/well) a day before transfection with plasmids encoding ACE2 (pCG1-ACE2, a kind gift of Stefan Pölhmann), pCMV3-C-Myc-TMPRSS2 and phRL-TK (encoding Renilla luciferase).
    HEK293
    suggested: None
    To determine RBD specific cytotoxicity, mouse NIH-3T3 cells were seeded into 24-well plates (1*105/well); next day cells were transfected with pCG1-hACE2 (900 ng/well) and pCMV-TMPRSS2 (30 ng/well) plasmids.
    NIH-3T3
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    ACE-2 (hsACE-2, residues S19 to D615) was codon optimized for H. sapiens and cloned into the pTwist_CMV_BetaGlobin_WPRE_Neo (Twist Biosciences).
    ACE-2 (hsACE-2
    suggested: None
    Mouse immunization studies: To test the immunogenicity of DNA vaccines female 8-10 weeks BALB/c OlaHsd mice (Envigo, Italy) were used for immunization protocols.
    BALB/c OlaHsd
    suggested: None
    Surrogate assay of protection of viral infection by immunization: Balb/c mice were immunized with RBD-bann according to scheme, presented above.
    Balb/c
    suggested: RRID:IMSR_ORNL:BALB/cRl)
    Software and Algorithms
    SentencesResources
    Modelling of designed RBD-scaffolded protein cages: Molecular model structures of designed nanovaccines were prepared by performing homology modelling with Modeller (version 9.23)56.
    Modeller
    suggested: (MODELLER, RRID:SCR_008395)
    For total IgG determination, goat anti mouse IgG (H+L)-HRP antibodies (Jackson ImmunoResearch; 115-035-003) were used.
    Jackson ImmunoResearch
    suggested: (Jackson ImmunoResearch, RRID:SCR_010488)

    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: Please consider improving the rainbow (“jet”) colormap(s) used on page 20. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

    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 10.1101/2020.08.28.244269v1 on bioRxiv