COVID-19 vaccine BNT162b1 elicits human antibody and TH1 T cell responses

  1. Ugur Sahin
  2. Alexander Muik
  3. Evelyna Derhovanessian
  4. Isabel Vogler
  5. Lena M. Kranz
  6. Mathias Vormehr
  7. Alina Baum
  8. Kristen Pascal
  9. Jasmin Quandt
  10. Daniel Maurus
  11. Sebastian Brachtendorf
  12. Verena Lörks
  13. Julian Sikorski
  14. Rolf Hilker
  15. Dirk Becker
  16. Ann-Kathrin Eller
  17. Jan Grützner
  18. Carsten Boesler
  19. Corinna Rosenbaum
  20. Marie-Cristine Kühnle
  21. Ulrich Luxemburger
  22. Alexandra Kemmer-Brück
  23. David Langer
  24. Martin Bexon
  25. Stefanie Bolte
  26. Katalin Karikó
  27. Tania Palanche
  28. Boris Fischer
  29. Armin Schultz
  30. Pei-Yong Shi
  31. Camila Fontes-Garfias
  32. John L. Perez
  33. Kena A. Swanson
  34. Jakob Loschko
  35. Ingrid L. Scully
  36. Mark Cutler
  37. Warren Kalina
  38. Christos A. Kyratsous
  39. David Cooper
  40. Philip R. Dormitzer
  41. Kathrin U. Jansen
  42. Özlem Türeci

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Abstract

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  1. ScreenIT

    SciScore for 10.1101/2020.07.17.20140533: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementConsent: One subject of the 10 µg, and one subject of the 50 µg dose cohort left the study prior to the boosting immunisation due to withdrawal of consent and private reasons.
    IRB: The trial was carried out in Germany in accordance with the Declaration of Helsinki and Good Clinical Practice Guidelines and with approval by an independent ethics committee (Ethik-Kommission of the Landesärztekammer Baden-Württemberg, Stuttgart, Germany) and the competent regulatory authority (Paul-Ehrlich Institute, Langen, Germany).
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variableClinical trial design: Study BNT162-01 (NCT04380701) is an ongoing, first-in-human, Phase 1/2, open-label dose-ranging clinical trial to assess the safety, tolerability, and immunogenicity of ascending dose levels of various intramuscularly administered BNT162 mRNA vaccine candidates in healthy men and non-pregnant women 18 to 55 years (amended to add 56 -85 of age) of age.
    Cell Line AuthenticationContamination: Cell lines were tested for mycoplasma contamination after receipt and before expansion and cryopreservation.

    Table 2: Resources

    Antibodies
    SentencesResources
    A secondary fluorescently labelled goat anti-human polyclonal antibody (Jackson Labs) was added for 90 minutes at room temperature while shaking, before plates were washed once more in a solution containing 0.05% Tween-20.
    A secondary fluorescently labelled goat anti-human polyclonal antibody ( Jackson Labs )
    suggested: None
    anti-human polyclonal antibody
    suggested: (LSBio (LifeSpan Cat# LS-C6444-30, RRID:AB_860431)
    Tests were performed in duplicate and with a positive control (anti-CD3 monoclonal antibody CD3-2 [1:1,000; Mabtech])
    anti-CD3
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Cell culture and primary cell isolation: Vero cells (ATCC CCL-81) and Vero E6 cells (ATCC CRL-1586) were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with GlutaMAX(tm) (Gibco) supplemented with 10% fetal bovine serum (FBS) (Sigma-Aldrich)
    Vero
    suggested: ATCC Cat# CRL-1586, RRID:CVCL_0574)
    Vero E6
    suggested: None
    Serial dilutions of heat-inactivated sera were incubated with the reporter virus (2 × 104 PFU per well to yield approximately a 10-30% infection rate of the Vero CCL81 monolayer) for 1 hour at 37 °C before inoculating Vero CCL81 cell monolayers (targeted to have 8,000 to 15,000 cells per well) in 96-well plates to allow accurate quantification of infected cells.
    Vero CCL81
    suggested: None
    HEK293T cells (ATCC CRL-3216) were seeded (culture medium: DMEM high glucose [Life Technologies] supplemented with 10% heat-inactivated fetal bovine serum [FBS; Life Technologies] and penicillin/ptreptomycin/L-glutamine [Life Technologies]) and transfected the following day with spike expression plasmid using Lipofectamine LTX (Life Technologies) following the manufacturer’s protocol.
    HEK293T
    suggested: None
    Software and Algorithms
    SentencesResources
    Key exclusion criteria included previous clinical or microbiological diagnosis of COVID-19; receipt of medications to prevent COVID-19; previous vaccination with any coronavirus vaccine; a positive serological test for SARS-CoV-2 IgM and/or IgG; and a SARS-CoV-2 NAAT-positive nasal swab; those with increased risk for severe COVID-19; and immunocompromised individuals.
    IgG
    suggested: (DSHB Cat# LEP100 IgG, RRID:AB_528124)
    Total cell counts per well were enumerated by nuclear stain (Hoechst 33342) and fluorescent virally infected foci were detected 16-24 hours after inoculation with a Cytation 7 Cell Imaging Multi-Mode Reader (BioTek) with Gen5 Image Prime version 3.09.
    Gen5
    suggested: (Gen5, RRID:SCR_017317)
    Titers were calculated in GraphPad Prism version 8.4.2 by generating a 4-parameter (4PL) logistical fit of the percent neutralisation at each serial serum dilution.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    FlowJo LLC, BD Biosciences)
    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: We detected the following sentences addressing limitations in the study:
    Limitations of our clinical study include the small sample size and its restriction to participants below 55 years of age. Another constraint is that we did not perform further T cell analysis e.g. deconvolution of epitope diversity, characterisation of HLA restriction and TCR repertoire analysis before and after vaccination, due to the limited blood volumes that were available for biomarker analyses. Further, as vaccine-induced immunity can wane over time, it is important to study persistence of potentially protective immune responses over time. However, samples to assess persistence are not yet available but are planned per study protocol and will be reported elsewhere.

    Results from TrialIdentifier: We found the following clinical trial numbers in your paper:

    IdentifierStatusTitle
    NCT04368728Active, not recruitingStudy to Describe the Safety, Tolerability, Immunogenicity, …
    NCT04380701RecruitingA Trial Investigating the Safety and Effects of Four BNT162 …

    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 10.1038/s41586-020-2814-7
  3. ScreenIT

    SciScore for 10.1101/2020.07.17.20140533: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementOne subject of the 10 µg, and one subject of the 50 µg dose 281 cohort left the study prior to the boosting immunisation due to withdrawal of consent and private 282 reasons.Randomizationnot detected.Blindingnot detected.Power Analysisnot detected.Sex as a biological variableThe study population consisted of healthy males and non-pregnant 85 females with a mean age of 41 years (range 18 to 55 years) with equal gender distribution.Cell Line AuthenticationCell lines were tested for mycoplasma contamination after receipt and 321 before expansion and cryopreservation.

    Table 2: Resources

    Antibodies
    SentencesResources
    CD4+ and CD8+ T cells may confer long-lasting 186 immunity against corona viruses as indicated in SARS-CoV-1 survivors, where CD8+ T-cell immunity 187 persisted for 6-11 years24,27. 188 Some cases of asymptomatic virus exposure have been associated with cellular immune response 189 without seroconversion indicating that SARS-CoV-2 specific T cells could be relevant in disease control 190 even in the absence of neutralising antibodies28.
    CD4+
    suggested: None
          <div style="margin-bottom:8px">
        <div><b>antibodies28</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">A secondary fluorescently labelled goat anti-human 329 polyclonal antibody (Jackson Labs) was added for 90 minutes at room temperature while shaking, before 330 plates were washed once more in a solution containing 0.05% Tween-20.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>A secondary fluorescently labelled goat anti-human 329 polyclonal antibody</b></div>
        <div>suggested: None</div>
      </div>
    
      <div style="margin-bottom:8px">
        <div><b>anti-human 329</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Tests were performed in duplicate and with a positive 391 control (anti-CD3 monoclonal antibody CD3-2 [1:1,000; Mabtech])</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-CD3</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Viral master stocks (2 x 107 PFU/mL) were 344 grown in Vero E6 cells as previously described29.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Vero E6</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Serial dilutions of heat-inactivated sera were incubated with the reporter virus (2 x 104 PFU per 347 well to yield approximately a 10-30% infection rate of the Vero CCL81 monolayer) for 1 hour at 37 ⁰C 348 before inoculating Vero CCL81 cell monolayers (targeted to have 8,000 to 15,000 cells per well) in 96 349 -well plates to allow accurate quantification of infected cells.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Vero CCL81</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">HEK293T cells (ATCC CRL- 364 3216) were seeded (culture medium: DMEM high glucose [Life Technologies] supplemented with 10% 365 heat-inactivated fetal bovine serum [FBS; Life Technologies] and penicillin/ptreptomycin/L-glutamine 366 [Life Technologies]) and transfected the following day with spike expression plasmid using 367 Lipofectamine LTX (Life Technologies) following the manufacturer’s protocol.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>HEK293T</b></div>
        <div>suggested: ATCC Cat# CRL-3216, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0063">CVCL_0063</a></div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">375 For pseudovirus neutralisation assays, Vero cells (ATCC CCL-81) were seeded in 96-well plates in 376 culture medium and allowed to reach approximately 85% confluence before use in the assay (24 hours 377 later).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Vero</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Key exclusion criteria included previous 268 clinical or microbiological diagnosis of COVID‑19; receipt of medications to prevent COVID‑19; 269 previous vaccination with any coronavirus vaccine; a positive serological test for SARS-CoV-2 IgM 270 and/or IgG; and a SARS-CoV-2 NAAT-positive nasal swab; those with increased risk for severe 271 COVID-19; and immunocompromised individuals.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>IgG</b></div>
        <div>suggested: (DSHB Cat# LEP100 IgG, <a href="https://scicrunch.org/resources/Any/search?q=AB_528124">AB_528124</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Total cell counts per well were enumerated 350 by nuclear stain (Hoechst 33342) and fluorescent virally infected foci were detected 16-24 hours after 351 inoculation with a Cytation 7 Cell Imaging Multi-Mode Reader (BioTek) with Gen5 Image Prime 352 version 3.09.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Gen5</b></div>
        <div>suggested: (Gen5, <a href="https://scicrunch.org/resources/Any/search?q=SCR_017317">SCR_017317</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Titers were calculated in GraphPad Prism version 8.4.2 by generating a 4-parameter (4PL) 353 logistical fit of the percent neutralisation at each serial serum dilution.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>GraphPad Prism</b></div>
        <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">FlowJo LLC, BD Biosciences)</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>FlowJo</b></div>
        <div>suggested: (FlowJo, <a href="https://scicrunch.org/resources/Any/search?q=SCR_008520">SCR_008520</a>)</div>
      </div>
    </td></tr></table>

    Data from additional tools added to each annotation on a weekly basis.

    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 is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.

  4. Version published to 10.1101/2020.07.17.20140533v1 on medRxiv