Early human B cell signatures of the primary antibody response to mRNA vaccination

  1. Lela Kardava
  2. Nicholas Rachmaninoff
  3. William W. Lau
  4. Clarisa M. Buckner
  5. Krittin Trihemasava
  6. Jana Blazkova
  7. Felipe Lopes de Assis
  8. Wei Wang
  9. Xiaozhen Zhang
  10. Yimeng Wang
  11. Chi-I Chiang
  12. Sandeep Narpala
  13. Genevieve E. McCormack
  14. Can Liu
  15. Catherine A. Seamon
  16. Michael C. Sneller
  17. Sarah O’Connell
  18. Yuxing Li
  19. Adrian B. McDermott
  20. Tae-Wook Chun
  21. Anthony S. Fauci
  22. John S. Tsang
  23. Susan Moir

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Abstract

Messenger RNA (mRNA) vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are highly effective at inducing protective immunity. However, weak antibody responses are seen in some individuals, and cellular correlates of immunity remain poorly defined, especially for B cells. Here we used unbiased approaches to longitudinally dissect primary antibody, plasmablast, and memory B cell (MBC) responses to the two-dose mRNA-1273 vaccine in SARS-CoV-2–naive adults. Coordinated immunoglobulin A (IgA) and IgG antibody responses were preceded by bursts of spike-specific plasmablasts after both doses but earlier and more intensely after dose 2. While antibody and B cell cellular responses were generally robust, they also varied within the cohort and decreased over time after a dose-2 peak. Both antigen-nonspecific postvaccination plasmablast frequency after dose 1 and their spike-specific counterparts early after dose 2 correlated with subsequent antibody levels. This correlation between early plasmablasts and antibodies remained for titers measured at 6 months after vaccination. Several distinct antigen-specific MBC populations emerged postvaccination with varying kinetics, including two MBC populations that correlated with 2- and 6-month antibody titers. Both were IgG-expressing MBCs: one less mature, appearing as a correlate after the first dose, while the other MBC correlate showed a more mature and resting phenotype, emerging as a correlate later after dose 2. This latter MBC was also a major contributor to the sustained spike-specific MBC response observed at month 6. Thus, these plasmablasts and MBCs that emerged after both the first and second doses with distinct kinetics are potential determinants of the magnitude and durability of antibodies in response to mRNA-based vaccination.

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  1. Version published to 10.1073/pnas.2204607119
  2. ScreenIT

    SciScore for 10.1101/2021.07.06.21259528: (What is this?)

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

    Table 1: Rigor

    EthicsIRB: Research phlebotomy was performed at the NIH Clinical Research Center in Bethesda, MD under protocols approved by the NIH Institutional Review Board,
    Consent: All participants provided written informed consent.
    Field Sample Permit: Blood sample collection and processing: Peripheral blood mononuclear cells (PBMC) were isolated by Ficoll density gradient centrifugation from whole blood collected in EDTA vacutainer tubes.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Plates were washed and incubated with MSD SULFO-TAG™ anti-human IgG, IgA or IgM detection antibodies at RT for 60 minutes with shaking.
    anti-human IgG
    suggested: None
    Intracellular flow cytometry: PBMC were first stained with antibodies against cell surface markers CD3, CD19, CD20 and CD27, fixed (Lysing Solution, BD Biosciences), permeabilized (Permeabilizing Solution 2; BD Biosciences) and stained with antibodies against IgG, IgA, IgD and IgM (antibody details in Extended Data Table 5).
    CD3
    suggested: None
    CD19
    suggested: None
    CD20
    suggested: None
    CD27
    suggested: None
    antibodies against IgG
    suggested: None
    IgA , IgD
    suggested: (MyBioSource Cat# MBS531438, RRID:AB_10577295)
    IgM
    suggested: None
    ELISpot assay to enumerate SARS-CoV-2 spike-specific PB: Spike protein S1 and RBD-specific antibody-secreting PB were enumerated by modifying the antigen-specific portion of a previously described ELISpot assay31,32.
    antibody-secreting PB
    suggested: None
    Briefly, wells of Immobilon-P polyvinylidene difluoride (PVDF) membrane 96-well plates (Millipore) were coated with 5ug/ml anti-Ig light-chain antibodies (Rockland Immunochemicals) overnight at 4°C.
    anti-Ig light-chain
    suggested: None
    Plates were incubated at 37°C for 5 hours, followed by overnight incubation at 4°C with biotinylated antibodies (Jackson Immunoresearch) against IgA (catalogue 109-066-011), IgG (catalogue 709-066-149), IgM (catalogue 709-066-073), or biotinylated proteins S1 (Acrobiosystems, catalogue S1N-C82E8) or RBD (Biolegend, catalogue 790904).
    IgA
    suggested: None
    Modeling of association between endpoint antibody concentrations and cluster frequencies: A linear model accounting for the age and gender of the longitudinal vaccine participants was used to estimate whether cell cluster frequencies in response to vaccination were associated with SARS-COV2 spike protein (S-2P/RBD) antibody concentration at endpoint (v2D28):

    Analyses were carried out on both standardized antigen non-specific and specific frequencies, i.e., cluster cell counts as a fraction of total CD19+ cell counts and RBD+ S1+ cells within the clusters, respectively.

    SARS-COV2 spike protein (S-2P/RBD
    suggested: None
    Recombinant DNA
    SentencesResources
    The DNA encoding sequence was cloned into the mammalian cell expression vector pCAGGS and confirmed by sequencing, prior to transient transfection in FreeStyle 293-F cells with 293fectin transfection reagent (ThermoFisher)
    pCAGGS
    suggested: RRID:Addgene_18926)
    Software and Algorithms
    SentencesResources
    Analyses were performed with Excel (Microsoft) and Prism 9.0 (Graphpad) software and antibody concentrations were assigned arbitrary units (AU/ml) by interpolation from the standard curve.
    Excel
    suggested: None
    Prism
    suggested: (PRISM, RRID:SCR_005375)
    Graphpad
    suggested: (GraphPad, RRID:SCR_000306)
    Densities of antibody concentrations at endpoint (v2D28) were estimated using a gaussian kernel with bandwidth automatically selected through biased cross validation using the stat_density function from ggplot2 (3.3.3) with bw = “bcv”.
    ggplot2
    suggested: (ggplot2, RRID:SCR_014601)
    The stained cells were acquired on a FACS Canto II flow cytometer (BD Biosciences) and analyzed using FlowJo software v9.9.6 (BD Biosciences)
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Spectral flow cytometry data processing for FlowSOM clustering and UMAP embedding: FCS files generated from spectral flow cytometry with the 17-color panel and associated manual gates were read into R using FlowWorkspace (4.2.0).
    FlowWorkspace
    suggested: (flowWorkspace, RRID:SCR_001155)
    The following markers were used to perform clustering CD20, CD138, CD38, CD10, CD11c, CD19, CD27, CD21, IgD, IgM, IgG, IgA, using FlowSOM (1.22.0)33, with the number of desired metaclusters (nClus) set to 30.
    FlowSOM
    suggested: (FlowSOM, RRID:SCR_016899)
    The clusters were then grouped by hierarchical clustering of the mean trends using the Euclidean distance at each timepoint and using Ward’s method, as implemented in the hclust function (method = “ward.D2) in R (4.0.2).
    hclust
    suggested: (HCLUST, RRID:SCR_009154)

    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: We found the following clinical trial numbers in your paper:

    IdentifierStatusTitle
    NCT00001281RecruitingStudies of Blood and Reproductive Fluids in HIV-Infected and…
    NCT04411147RecruitingA Longitudinal Study of COVID-19 Sequelae and Immunity
    NCT04280705CompletedAdaptive COVID-19 Treatment Trial (ACTT)
    NCT04579393CompletedFostamatinib for Hospitalized Adults With COVID-19

    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

    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 10.1101/2021.07.06.21259528v1 on medRxiv