Fc-Independent Protection from SARS-CoV-2 Infection by Recombinant Human Monoclonal Antibodies

  1. Tal Noy-Porat
  2. Avishay Edri
  3. Ron Alcalay
  4. Efi Makdasi
  5. David Gur
  6. Moshe Aftalion
  7. Yentl Evgy
  8. Adi Beth-Din
  9. Yinon Levy
  10. Eyal Epstein
  11. Olga Radinsky
  12. Ayelet Zauberman
  13. Shirley Lazar
  14. Shmuel Yitzhaki
  15. Hadar Marcus
  16. Angel Porgador
  17. Ronit Rosenfeld
  18. Ohad Mazor

Reviewed by

Read the full article See related articles

Listed in

Log in to save this article

Abstract

The use of passively-administered neutralizing antibodies is a promising approach for the prevention and treatment of SARS-CoV-2 infection. Antibody-mediated protection may involve immune system recruitment through Fc-dependent activation of effector cells and the complement system. However, the role of Fc-mediated functions in the efficacious in-vivo neutralization of SARS-CoV-2 is not yet clear, and it is of high importance to delineate the role this process plays in antibody-mediated protection. Toward this aim, we have chosen two highly potent SARS-CoV-2 neutralizing human monoclonal antibodies, MD65 and BLN1 that target distinct domains of the spike (RBD and NTD, respectively). The Fc of these antibodies was engineered to include the triple mutation N297G/S298G/T299A that eliminates glycosylation and the binding to FcγR and to the complement system activator C1q. As expected, the virus neutralization activity (in-vitro) of the engineered antibodies was retained. To study the role of Fc-mediated functions, the protective activity of these antibodies was tested against lethal SARS-CoV-2 infection of K18-hACE2 transgenic mice, when treatment was initiated either before or two days post-exposure. Antibody treatment with both Fc-variants similarly rescued the mice from death reduced viral load and prevented signs of morbidity. Taken together, this work provides important insight regarding the contribution of Fc-effector functions in MD65 and BLN1 antibody-mediated protection, which should aid in the future design of effective antibody-based therapies.

Article activity feed

  1. Version published to 10.3390/antib10040045
  2. ScreenIT

    SciScore for 10.1101/2021.05.15.443978: (What is this?)

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

    Table 1: Rigor

    EthicsIACUC: Animal experiments: Treatment of animals was in accordance with regulations outlined in the U.S. Department of Agriculture (USDA) Animal Welfare Act and the conditions specified in the Guide for Care and Use of Laboratory Animals (National Institute of Health, 2011).
    Sex as a biological variableFemale K18-hACE2 transgenic (B6.Cg-Tg(K18-ACE2)2Prlmn/J HEMI) were maintained at 20-22 °C and a relative humidity of 50±10% on a 12 hours light/dark cycle, fed with commercial rodent chow (Koffolk Inc.) and provided with tap water ad libitum.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    The secondary antibodies: AP-conjugated Donkey anti-human IgG (Jackson ImmunoResearch, USA, Cat# 709-055-149, lot 130049) or AP-conjugated Donkey anti-mouse IgG (H+L) minimal cross (Jackson ImmunoResearch, USA, Cat# 715-055-150, lot 142717) were applied, followed by the addition of PNPP substrate (Sigma, Israel, Cat# N1891).
    anti-human IgG
    suggested: (Jackson ImmunoResearch Labs Cat# 709-055-149, RRID:AB_2340501)
    anti-mouse IgG
    suggested: (Jackson ImmunoResearch Labs Cat# 715-055-150, RRID:AB_2340777)
    For CD16, plate was incubated with antibodies MD65-YTE and MD65-AG-YTE starting at 100 µg/ml with 2-fold serial dilutions.
    CD16
    suggested: None
    For CD64, antibodies were used starting at 10 µg/ml with serial dilutions.
    CD64
    suggested: None
    Sensors were loaded with CD32 (10 µg/ml), followed by a wash, and then incubated with antibodies MD65-YTE or MD65-AG-YTE.
    CD32
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    The original viral isolate was amplified by 5 passages and quantified by plaque titration assay in Vero E6 cells, and stored at -80°C until use.
    Vero E6
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    Female K18-hACE2 transgenic (B6.Cg-Tg(K18-ACE2)2Prlmn/J HEMI) were maintained at 20-22 °C and a relative humidity of 50±10% on a 12 hours light/dark cycle, fed with commercial rodent chow (Koffolk Inc.) and provided with tap water ad libitum.
    K18-hACE2
    suggested: RRID:IMSR_GPT:T037657)
    B6.Cg-Tg(K18-ACE2)2Prlmn/J HEMI
    suggested: None
    Software and Algorithms
    SentencesResources
    The sensors were then incubated with C1q native protein (250 nM; US Biologicals, USA, Cat# C0010-10D) for 180 s and then transferred to buffer-containing wells for another 180 s.
    US Biologicals
    suggested: None
    Plates were then incubated with blocking solution containing 10% FBS in PBST (0.05% Tween-20), washed and incubated with the collected supernatant for 2 hours, followed by addition of relevant detection mAbs: biotin anti-human IFN-γ (Clone: 4S.B3, BioLegend); biotin anti-human TNF-α (Clone: MP6-XT22, BioLegend); or biotin anti-mouse IL-2 (Clone: JES6-5H4, BioLegend) SA-HRP (Jackson immunoresearch, PA USA) and TMB (Dako, Denmark) used for detection of mIL-2.
    BioLegend)
    suggested: (BioLegend, RRID:SCR_001134)

    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.
    • No funding statement was detected.
    • No protocol registration statement was detected.

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

    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.

  3. Version published to 10.1101/2021.05.15.443978v1 on bioRxiv