Sterilizing Immunity against SARS-CoV-2 Infection in Mice by a Single-Shot and Modified Imidazoquinoline TLR7/8 Agonist-Adjuvanted Recombinant Spike Protein Vaccine

  1. Sonia Jangra
  2. Jana De Vrieze
  3. Angela Choi
  4. Raveen Rathnasinghe
  5. Gabriel Laghlali
  6. Annemiek Uvyn
  7. Simon Van Herck
  8. Lutz Nuhn
  9. Kim Deswarte
  10. Zifu Zhong
  11. Niek Sanders
  12. Stefan Lienenklaus
  13. Sunil David
  14. Shirin Strohmeier
  15. Fatima Amanat
  16. Florian Krammer
  17. Hamida Hammad
  18. Bart N. Lambrecht
  19. Lynda Coughlan
  20. Adolfo García-Sastre
  21. Bruno G. De Geest
  22. Michael Schotsaert

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Abstract

The search for vaccines that protect from severe morbidity and mortality as a result of infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes coronavirus disease 2019 (COVID-19) is a race against the clock and the virus. Several vaccine candidates are currently being tested in the clinic. Inactivated virus and recombinant protein vaccines can be safe options but may require adjuvants to induce robust immune responses efficiently. In this work we describe the use of a novel amphiphilic imidazoquinoline (IMDQ-PEG-CHOL) TLR7/8 adjuvant, consisting of an imidazoquinoline conjugated to the chain end of a cholesterol-poly(ethylene glycol) macromolecular amphiphile). This amphiphile is water soluble and exhibits massive translocation to lymph nodes upon local administration, likely through binding to albumin. IMDQ-PEG-CHOL is used to induce a protective immune response against SARS-CoV-2 after single vaccination with trimeric recombinant SARS-CoV-2 spike protein in the BALB/c mouse model. Inclusion of amphiphilic IMDQ-PEG-CHOL in the SARS-CoV-2 spike vaccine formulation resulted in enhanced immune cell recruitment and activation in the draining lymph node. IMDQ-PEG-CHOL has a better safety profile compared to native soluble IMDQ as the former induces a more localized immune response upon local injection, preventing systemic inflammation. Moreover, IMDQ-PEG-CHOL adjuvanted vaccine induced enhanced ELISA and in vitro microneutralization titers, and a more balanced IgG2a/IgG1 response. To correlate vaccine responses with control of virus replication in vivo , vaccinated mice were challenged with SARS-CoV-2 virus after being sensitized by intranasal adenovirus-mediated expression of the human angiotensin converting enzyme 2 (ACE2) gene. Animals vaccinated with trimeric recombinant spike protein vaccine without adjuvant had lung virus titers comparable to non-vaccinated control mice, whereas animals vaccinated with IMDQ-PEG-CHOL-adjuvanted vaccine controlled viral replication and infectious viruses could not be recovered from their lungs at day 4 post infection. In order to test whether IMDQ-PEG-CHOL could also be used to adjuvant vaccines currently licensed for use in humans, proof of concept was also provided by using the same IMDQ-PEG-CHOL to adjuvant human quadrivalent inactivated influenza virus split vaccine, which resulted in enhanced hemagglutination inhibition titers and a more balanced IgG2a/IgG1 antibody response. Enhanced influenza vaccine responses correlated with better virus control when mice were given a lethal influenza virus challenge. Our results underscore the potential use of IMDQ-PEG-CHOL as an adjuvant to achieve protection after single immunization with recombinant protein and inactivated vaccines against respiratory viruses, such as SARS-CoV-2 and influenza viruses.

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  2. Rapid Reviews Infectious Diseases

    Yang Shi

    Review of "Sterilizing Immunity against SARS-CoV-2 Infection in Mice by a Single-Shot and Modified Imidazoquinoline TLR7/8 Agonist-Adjuvanted Recombinant Spike Protein Vaccine"

    Reviewer: Yang Shi (Aachen University) | 📘📘📘📘📘

  3. ScreenIT

    SciScore for 10.1101/2020.10.23.344085: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIACUC: Mice, Cell lines and reagents: 6-8 weeks old female BALB/c mice were obtained from Charles River Laboratories, MA and were housed in a specified pathogen-free facility at Icahn school of medicine at Mount Sinai, with food and water ad libitum, adhering to the guidelines from Institutional Animal Care and Use Committee.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variableMice, Cell lines and reagents: 6-8 weeks old female BALB/c mice were obtained from Charles River Laboratories, MA and were housed in a specified pathogen-free facility at Icahn school of medicine at Mount Sinai, with food and water ad libitum, adhering to the guidelines from Institutional Animal Care and Use Committee.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    After 72 hours of incubation at 37°C, 5% CO2, the plates were fixed in 4% formaldehyde, followed by immune-staining of infected cells with anti-mouse SARS-CoV-2 nucleoprotein and anti-mouse SARS-CoV-2 spike monoclonal antibodies.
    anti-mouse SARS-CoV-2 nucleoprotein
    suggested: None
    anti-mouse SARS-CoV-2
    suggested: None
    After incubation in primary antibodies, HRP-conjugated anti-mouse secondary antibody was added for 1 hour.
    anti-mouse
    suggested: None
    Isolated lymph nodes were collected in ice cold PBS, smashed through 70 μm cell strainers, washed with PBS and stained for 30min at 4°C with following primary labeled antibodies: CD3, CD20, CD11c, MHCII, CD86, CD40.
    CD3
    suggested: None
    CD20
    suggested: None
    CD11c
    suggested: None
    CD86
    suggested: None
    CD40
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Both cells were maintained in Dulbecco’s Modified Eagle’s Medium supplemented with 10% Fetal bovine serum (FBS) and additionally with 1% non-essential amino acids for Vero-E6 cells.
    Vero-E6
    suggested: None
    Murine RAW blue 264.7 macrophages were cultured in DMEM medium supplemented with 10 % heat-inactivated FBS, antibiotics (50 units/mL penicillin and 50 μg/mL streptomycin), 2 mM L-glutamine and 0.01 % Zeocin.
    RAW blue 264.7
    suggested: None
    For IVR-180, MDCK cells were incubated with the lung homogenate dilutions for 1 hour at 37°C, 5% CO2 and then overlaid with a mixture of 2% oxoid agar and 2X minimal essential medium (MEM) supplemented with 1% diethyl-aminoethyl (DEAE)-dextran and 1 μg/ml
    MDCK
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    In order to make SARS-CoV-2 Spike-vaccinated BALB/c mice susceptible to challenge with wild type SARS-CoV-2 virus, airway expression of human ACE-2, the receptor for SARS-CoV-2, was obtained by intranasal transduction of mice with 2.5×108 PFU of adenovirus expressing h-ACE-2 (Ad5-hACE2), 4.5-weeks post-vaccination as described in (28).
    BALB/c
    suggested: None
    Analysis of in vivo lymphatic drainage: 20 μL (1 mg/mL in PBS) of Cyanine5-PEG-CHOL or Cyanine5-PEG were injected into the footpad of female C57BL/6 WT mice.
    C57BL/6
    suggested: None
    Software and Algorithms
    SentencesResources
    Image processing was done using the ImageJ software package.
    ImageJ
    suggested: (ImageJ, RRID:SCR_003070)
    Data were processed using the FlowJo software package.
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)

    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.

  4. Version published to 10.1101/2020.10.23.344085 on bioRxiv