Germinal center-derived broadly neutralizing antibodies adapt to SARS-CoV-2 antigenic drift

  1. Kiyomi Shitaoka
  2. Yohei Kawano
  3. Akifumi Higashiura
  4. Yoko Mizoguchi
  5. Takao Hashiguchi
  6. Norihisa Nishimichi
  7. Shiyu Huang
  8. Ayano Ito
  9. Akima Yamamoto
  10. Shun Ohki
  11. Miyuki Kanda
  12. Tomohiro Taniguchi
  13. Yasuyuki Yokosaki
  14. Satoshi Okada
  15. Takemasa Sakaguchi
  16. Tomoharu Yasuda

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Abstract

The outbreak of SARS-CoV-2 variant Omicron which harbors a striking number of mutations in the spike protein has been raising concerns about the effectiveness of vaccines and antibody treatment 1 . Here, we confirmed a substantial reduction in neutralizing potency against Omicron in all convalescent and vaccinated sera. However, we found that some people infected by the early strain show relatively higher neutralization to Omicron. From those B cells, we developed neutralizing antibodies inhibiting broad variants including Delta and Omicron. Unlike reported antibodies, one had an extremely large interface and widely covered receptor binding motif of spike, thereby interfering with diversified variants. Somatic mutations introduced by long-term germinal center reaction contributed to the key structure of antibodies and the universal interaction with spike variants. Recalling such rare B cells may confer sustainable protection against SARS-CoV-2 variants emerging one after another.

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  1. ScreenIT

    SciScore for 10.1101/2022.01.26.477937: (What is this?)

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

    Table 1: Rigor

    EthicsField Sample Permit: Crystallization, X-ray diffraction data collection, and processing: Crystallization was carried out by the sitting-drop vapor diffusion method at 20 °C.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    The serum dilution or antibody concentration causing a 50% reduction of RLU compared to control (ED50 or IC50, respectively) were reported as the neutralizing antibody titers.
    ED50
    suggested: None
    IC50
    suggested: (GeneTex Cat# GTX21096, RRID:AB_384608)
    Experimental Models: Cell Lines
    SentencesResources
    The vector was transfected into Expi293 cells and incubated at 37°C for 4 days.
    Expi293
    suggested: RRID:CVCL_D615)
    Lentiviral vector, pWPI-ffLuc-P2A-EGFP for luciferase reporter assay and pTRC2puro-ACE2-P2A-TMPRSS2 for the generation of 293T cell line susceptible to SARS-CoV-2 infection was created from pWPI-IRES-Puro-Ak-ACE2-TMPRSS2, a gift from Sonja Best (Addgene, no.154987) by In-Fusion® technology (Takara Bio).
    293T
    suggested: None
    Pseudovirus production and neutralization: Pseudoviruses bearing SARS-Cov2 S-glycoprotein and carrying a firefly luciferase (ffLuc) reporter gene were produced in LentiX-293T cells by transfecting with pWPI-ffLuc-P2A-EGFP, psPAX2, and either of S variant from Wuhan, D614G
    LentiX-293T
    suggested: None
    Pseudovirus titers were measured by infecting 293T/TRCAT cells for 72 hours before measuring luciferase activity (ONE-Glo™ Luciferase Assay System, Promega, Madison, WI)
    293T/TRCAT
    suggested: None
    Neutralization assay with authentic SARS-CoV-2 viruses: VeroE6/TMPRSS2 cells (African green monkey kidney-derived cells expressing human TMPRSS2, purchased from the Japanese Collection of Research Bioresources (JCRB) Cell Bank, JCRB1819) were maintained in DMEM containing 10% FBS and 1 mg/mL G418 at 37 °C in 5% CO2.
    VeroE6/TMPRSS2
    suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
    Recombinant DNA
    SentencesResources
    PCR fragments were assembled into a linearized pcDNA vector using NEBuilder HiFi DNA Assembly Master Mix (New England Biolabs) according to the manufacturer’s instructions.
    pcDNA
    suggested: RRID:Addgene_66792)
    The pcDNA3 (Invitrogen) vectors containing an Ig light chain gene and the pcDNA4 (Invitrogen) vectors containing an Ig heavy chain gene were simultaneously transfected into Expi293 cells using Expi293 Expression System Kit (Thermo Fisher Scientific).
    pcDNA3
    suggested: RRID:Addgene_15475)
    pcDNA4
    suggested: RRID:Addgene_45907)
    A soluble version of the S protein (amino acids 1–1213), including the T4 foldon trimerization domain, a histidine tag, and a strep-tag, was cloned into the mammalian expression vector pCMV.
    pCMV
    suggested: RRID:Addgene_16459)
    psPAX2 (Addgene, no.12260) was a gift from Didier Trono. pCDNA3.3_CoV2_B.1.1.7 (Addgene, no.170451) for Alpha-S and pcDNA3.3-SARS2-B.1.617.2 (Addgene, no.172320) for Delta-S proteins, were gifts from David Nemazee25.
    psPAX2
    suggested: RRID:Addgene_12260)
    pCDNA3.3_CoV2_B.1.1.7
    suggested: RRID:Addgene_170451)
    pcDNA3.3-SARS2-B.1.617.2
    suggested: RRID:Addgene_172320)
    Lentiviral vector, pWPI-ffLuc-P2A-EGFP for luciferase reporter assay and pTRC2puro-ACE2-P2A-TMPRSS2 for the generation of 293T cell line susceptible to SARS-CoV-2 infection was created from pWPI-IRES-Puro-Ak-ACE2-TMPRSS2, a gift from Sonja Best (Addgene, no.154987) by In-Fusion® technology (Takara Bio).
    pWPI-IRES-Puro-Ak-ACE2-TMPRSS2
    suggested: RRID:Addgene_154987)
    pcDNA3.4 expression plasmids encoding SARS-CoV-2 S proteins with human codon optimization and 19 a.a deletion of C-terminus (C-del19) from Wuhan, D614G, and Omicron were generated by assembly of PCR products, annealed oligonucleotides, or artificial synthetic gene fragments (Integrated DNA Technologies, IDT) using In-Fusion® technology.
    pcDNA3.4
    suggested: RRID:Addgene_131198)
    lentiviral vector VSV-G-pseudotyped lentivirus carrying ACE2 and TMPRSS2 genes were produced in LentiX-293T cells by transfecting with pTRC2puro-ACE2-P2A-TMPRSS2, psPAX2 (gag-pol), and pMD2G-VSV-G (envelope) using PEI-MAX (Polysciences)
    pTRC2puro-ACE2-P2A-TMPRSS2
    suggested: None
    pMD2G-VSV-G
    suggested: None
    Pseudovirus production and neutralization: Pseudoviruses bearing SARS-Cov2 S-glycoprotein and carrying a firefly luciferase (ffLuc) reporter gene were produced in LentiX-293T cells by transfecting with pWPI-ffLuc-P2A-EGFP, psPAX2, and either of S variant from Wuhan, D614G
    pWPI-ffLuc-P2A-EGFP
    suggested: None
    Software and Algorithms
    SentencesResources
    ED50 or IC50 were calculated using a nonlinear regression curve fit (GraphPad Prism software Inc., La Jolla, CA)
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    Paired-end, 300 bp sequencing was performed using MiSeq (Illumina) with the MiSeq reagent kit v3 (Illumina, MS-102-3003).
    MiSeq
    suggested: (A5-miseq, RRID:SCR_012148)
    Structure determination and analyses: Phase determinations were carried out by the molecular replacement method using the program Phaser in the PHENIX package32 and the program Molrep33 with the combination between RBD structure (PDB ID:7EAM) and Fab structures (PDB ID:7CHB and 7CHP) as search models.
    PHENIX
    suggested: (Phenix, RRID:SCR_014224)
    All figures of structures were generated by the program pymol (The PyMOL Molecular Graphics System, Version 1.2r3pre, Schrödinger, LLC.).
    PyMOL
    suggested: (PyMOL, RRID:SCR_000305)

    Results from OddPub: Thank you for sharing your data.

    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.

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

    About SciScore

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  2. Version published to 10.1101/2022.01.26.477937v1 on bioRxiv