Afucosylated immunoglobulin G responses are a hallmark of enveloped virus infections and show an exacerbated phenotype in COVID-19

  1. Mads Delbo Larsen
  2. Erik L. de Graaf
  3. Myrthe E. Sonneveld
  4. H. Rosina Plomp
  5. Federica Linty
  6. Remco Visser
  7. Maximilian Brinkhaus
  8. Tonći Šuštić
  9. Steven W. de Taeye
  10. Arthur E.H. Bentlage
  11. Jan Nouta
  12. Suvi Natunen
  13. Carolien A. M. Koeleman
  14. Susanna Sainio
  15. Neeltje A. Kootstra
  16. Philip J.M. Brouwer
  17. Rogier W. Sanders
  18. Marit J. van Gils
  19. Sanne de Bruin
  20. Alexander P.J. Vlaar
  21. Amsterdam UMC COVID-19 biobank study group
  22. Hans L. Zaaijer
  23. Manfred Wuhrer
  24. C. Ellen van der Schoot
  25. Gestur Vidarsson

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Abstract

IgG antibodies are crucial for protection against invading pathogens. A highly conserved N-linked glycan within the IgG-Fc-tail, essential for IgG function, shows variable composition in humans. Afucosylated IgG variants are already used in anti-cancer therapeutic antibodies for their elevated binding and killing activity through Fc receptors (FcγRIIIa). Here, we report that afucosylated IgG which are of minor abundance in humans (∼6% of total IgG) are specifically formed against surface epitopes of enveloped viruses after natural infections or immunization with attenuated viruses, while protein subunit immunization does not elicit this low fucose response. This can give beneficial strong responses, but can also go awry, resulting in a cytokine-storm and immune-mediated pathologies. In the case of COVID-19, the critically ill show aggravated afucosylated-IgG responses against the viral spike protein. In contrast, those clearing the infection unaided show higher fucosylation levels of the anti-spike protein IgG. Our findings indicate antibody glycosylation as a potential factor in inflammation and protection in enveloped virus infections including COVID-19.

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    SciScore for 10.1101/2020.05.18.099507: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementConsent: The ACS have been conducted in accordance with the ethical principles set out in the declaration of Helsinki and all participants provided written informed consent.
    IRB: The study was approved by the Academic Medical Center institutional Medical Ethics Committee of the University of Amsterdam.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    n=24 Live Attenuated vaccine) and HBV antibodies (n=17 natural infection, n=16 HBsAg vaccination)
    HBV
    suggested: None
    Purification of CMV-specific antibodies from sera: CMV-specific antibodies were purified using antigen-coated plates (Serion ELISA classic, Cytomegalovirus IgG, Würzburg, Germany).
    Serion ELISA classic , Cytomegalovirus IgG
    suggested: None
    Purification of Measle- and Mump-virus specific antibodies from sera: Ag-specific antibodies were purified using antigen-coated plates (Serion ELISA classic, Measles IgG and Mumps IgG, Würzburg, Germany).
    Serion ELISA classic , Measles IgG
    suggested: None
    Mumps IgG
    suggested: None
    As positive control, anti-HIV gp120 monoclonal was used (IgG1 b12; 100 µg purified antibody in PBS at 1 mg/ml; NIH Aids Reagent Program, La Jolla, CA, US).
    anti-HIV
    suggested: None
    IgG1
    suggested: None
    Purification of anti-N and anti-S specific antibodies from plasma: SARS-Cov-2-specific antibodies were purified using antigen-coated plates (NUCN, Roskilde, Denmark).
    anti-N
    suggested: None
    anti-S
    suggested: None
    Purification of total IgG from sera: Total IgG1 antibodies were captured from 2 µL of serum using Protein G Sepharose 4 Fast Flow beads (GE Healthcare, Uppsala, Sweden) in a 96-well filter plate (Millipore Multiscreen, Amsterdam, The Netherlands) as described previously (11) or by using Protein G cartridges on the AssayMAP Bravo (Agilent Technologies, Santa Clara, USA) Briefly, 1 µL serum diluted in PBS were applied to the cartridges, followed by washes of PBS, LC-MS pure water and finally eluted with formic acid (1%)
    sera: Total IgG1 antibodies
    suggested: None
    Total IgG1
    suggested: None
    Briefly , 1
    suggested: None
    Mass spectrometric IgG-Fc glycosylation analysis: Eluates containing either antigen-specific antibodies or total IgG were collected in V-bottom plates, dried by vacuum centrifugation for 2.5 hours at 50°C.
    antigen-specific
    suggested: None
    total IgG
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Plates were coated (over-night, 4°C) with recombinant trimerized spike protein produced as described recently (28) or N protein (accession number MN908947, produced in HEK cells with HAVT20 leader peptide, 10xhis tag and a Brit tag as in (23)) in PBS(5 µg/mL and 1 µg/mL, respectively).
    HEK
    suggested: None
    Software and Algorithms
    SentencesResources
    As positive control, anti-HIV gp120 monoclonal was used (IgG1 b12; 100 µg purified antibody in PBS at 1 mg/ml; NIH Aids Reagent Program, La Jolla, CA, US).
    Aids Reagent Program
    suggested: None
    Mass spectrometry results were extracted and evaluated using FlexAnalysis software (Bruker Daltonics) for all samples except for the Measles virus, and Mumps virus cohorts that were analyzed with Skyline software.
    Skyline
    suggested: (Skyline, RRID:SCR_014080)
    Statistical analysis: Statistical analyses were performed using GraphPad Prism version 7.02 for Windows (GraphPad Software Inc.,
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    GraphPad
    suggested: (GraphPad Prism, RRID:SCR_002798)

    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 10.1101/2020.05.18.099507 on bioRxiv