Comprehensive characterization of the antibody responses to SARS-CoV-2 Spike protein finds additional vaccine-induced epitopes beyond those for mild infection

  1. Meghan E Garrett
  2. Jared G Galloway
  3. Caitlin Wolf
  4. Jennifer K Logue
  5. Nicholas Franko
  6. Helen Y Chu
  7. Frederick A Matsen
  8. Julie M Overbaugh

  • Curated by eLife

    eLife logo

    Evaluation Summary:

    The authors used phage display deep mutational scanning of SARS-CoV-2 Spike protein to profile antibody epitopes outside of the RBD that were recognized in two cohorts of subjects, which together included mRNA vaccinated, SARS-CoV-2 infected with mild or severe COVID-19 and vaccinated with prior infection. Key findings of the study are that severe COVID-19 and vaccinated individuals had higher binding to Spike protein regions NTD, CTD, and SH-H compared to individuals with mild COVID-19, while mild COVID-19 infections had higher binding to FP than vaccinated or severe COVID-19 individuals. They also reported that vaccinated individuals with or without prior infection were not different and that covariates did not appear to impact the antibody recognition profiles. The authors identified potential escape pathways in these epitope regions, some of which differed between vaccination and infection or drifted over time. The authors acknowledge that this approach is limited to linear epitopes and does not include RBD epitopes. However, the study provides novel insight into the major epitope regions targeted by polyclonal antibodies elicited by vaccination vs. infection, as well as potential pathways for the virus to escape recognition.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #2 agreed to share their name with the authors.)

Reviewed by

Read the full article See related articles

Listed in

Log in to save this article

Abstract

Control of the COVID-19 pandemic will rely on SARS-CoV-2 vaccine-elicited antibodies to protect against emerging and future variants; an understanding of the unique features of the humoral responses to infection and vaccination, including different vaccine platforms, is needed to achieve this goal.

Methods:

The epitopes and pathways of escape for Spike-specific antibodies in individuals with diverse infection and vaccination history were profiled using Phage-DMS. Principal component analysis was performed to identify regions of antibody binding along the Spike protein that differentiate the samples from one another. Within these epitope regions, we determined potential sites of escape by comparing antibody binding of peptides containing wild-type residues versus peptides containing a mutant residue.

Results:

Individuals with mild infection had antibodies that bound to epitopes in the S2 subunit within the fusion peptide and heptad-repeat regions, whereas vaccinated individuals had antibodies that additionally bound to epitopes in the N- and C-terminal domains of the S1 subunit, a pattern that was also observed in individuals with severe disease due to infection. Epitope binding appeared to change over time after vaccination, but other covariates such as mRNA vaccine dose, mRNA vaccine type, and age did not affect antibody binding to these epitopes. Vaccination induced a relatively uniform escape profile across individuals for some epitopes, whereas there was much more variation in escape pathways in mildly infected individuals. In the case of antibodies targeting the fusion peptide region, which was a common response to both infection and vaccination, the escape profile after infection was not altered by subsequent vaccination.

Conclusions:

The finding that SARS-CoV-2 mRNA vaccination resulted in binding to additional epitopes beyond what was seen after infection suggests that protection could vary depending on the route of exposure to Spike antigen. The relatively conserved escape pathways to vaccine-induced antibodies relative to infection-induced antibodies suggests that if escape variants emerge they may be readily selected for across vaccinated individuals. Given that the majority of people will be first exposed to Spike via vaccination and not infection, this work has implications for predicting the selection of immune escape variants at a population level.

Funding:

This work was supported by NIH grants AI138709 (PI JMO) and AI146028 (PI FAM). JMO received support as the Endowed Chair for Graduate Education (FHCRC). The research of FAM was supported in part by a Faculty Scholar grant from the Howard Hughes Medical Institute and the Simons Foundation. Scientific Computing Infrastructure at Fred Hutch was funded by ORIP grant S10OD028685.

Article activity feed

  1. Version published to 10.7554/elife.73490 on eLife
  2. eLife

    Evaluation Summary:

    The authors used phage display deep mutational scanning of SARS-CoV-2 Spike protein to profile antibody epitopes outside of the RBD that were recognized in two cohorts of subjects, which together included mRNA vaccinated, SARS-CoV-2 infected with mild or severe COVID-19 and vaccinated with prior infection. Key findings of the study are that severe COVID-19 and vaccinated individuals had higher binding to Spike protein regions NTD, CTD, and SH-H compared to individuals with mild COVID-19, while mild COVID-19 infections had higher binding to FP than vaccinated or severe COVID-19 individuals. They also reported that vaccinated individuals with or without prior infection were not different and that covariates did not appear to impact the antibody recognition profiles. The authors identified potential escape pathways in these epitope regions, some of which differed between vaccination and infection or drifted over time. The authors acknowledge that this approach is limited to linear epitopes and does not include RBD epitopes. However, the study provides novel insight into the major epitope regions targeted by polyclonal antibodies elicited by vaccination vs. infection, as well as potential pathways for the virus to escape recognition.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #2 agreed to share their name with the authors.)

  3. eLife

    Reviewer #1 (Public Review):

    In this study, the authors used a high throughput mutational scanning approach to profile antibody epitopes in the SARS-CoV-2 Spike protein using serum from two cohorts of vaccinated and/or infected subjects. Key findings of the study are that antibody binding to the major epitope regions differed between severe COVID-19 and vaccinated individuals compared to individuals with mild COVID-19. The authors also identified potential viral escape pathways in these epitope regions, some of which differed between vaccination and infection or drifted over time. The authors acknowledged that this approach is limited to the detection of linear epitopes and did not include neutralization epitopes like those found in the receptor binding domain. However, the study provides new insight into the major epitope regions targeted by polyclonal antibodies elicited by vaccination vs. infection, as well as potential pathways that could be used by the virus to escape recognition.

  4. eLife

    Reviewer #2 (Public Review):

    Garrett and Galloway et al. present a study that analyzed the SARS-CoV-2 Spike linear epitopes that are targeted by antibodies following infection or vaccination with the SARS-CoV-2 mRNA vaccine mRNA-1273. The authors use a phage display profiling method termed Phage deep mutational scanning (PhageDMS) where T7 phage display libraries are generated to display overlapping 31 aa peptides that span the entire SARS-CoV-2 spike protein. Serum samples from naïve, SARS-CoV-2 infected hospitalized and non-hospitalized individuals, or individuals vaccinated with two different doses of the Moderna mRNA vaccine mRNA-1273 with and without prior infection are incubated with the PhageDMS library. Antibody-bound phage are then isolated and sequenced to identify the spike peptide targeted by antibodies.

    The authors find both common and unique immunodominant spike protein epitopes between infected hospitalized and non-hospitalized individuals and vaccinated individuals. Infection-induced antibodies targeted the fusion protein (FP) and the stem helix region upstream of heptad repeat 2 (SH-H), whereas vaccination-elicited antibodies also targeted epitopes within the NTD and CTD in addition to the FP and SH-H regions. The authors include a subset of vaccinated individuals who were infected prior to vaccination, yet do not find any significant differences across the epitope binding profiles compared to vaccinated individuals without prior infection.

    The authors performed additional experiments to determine whether mutations within the immunodominant spike epitopes could escape antibody binding. Within the vaccine targeted NTP and CTD regions, a uniform escape profile was observed for the NTD for vaccinated individuals whereas the CTD escape profile was variable. Within the immunodominant epitopes elicited by infection, FP escape at specific sites could be observed, whereas SH-H was more variable. Interestingly, for infected individuals who were subsequently vaccinated, the SH-H mutations became more uniform, and converge on a site known to be targeted by cross-coronavirus monoclonal antibodies.

    Overall, the study is well-designed and presented. The authors data provides insight into the potential mechanisms of escape that could impact long-term vaccine and infection induced humoral immunity. A major limitation of the study that the authors acknowledge is that the PhageDMS method only identifies linear epitopes, and thus antibodies that bind to conformational epitopes including the receptor binding domain, which is a neutralizing antibody target are not captured. Similarly, the mutational scan used to identify potential pathways to escape is also limited to linear epitopes, potentially missing loss of antibody binding that may occur from conformational epitopes that are disrupted upon mutation. That being said, the fact that specific mutations do lead to loss of antibody binding provide strong evidence that mutation at that site would clearly be detrimental to antibody efficacy. The authors note that these mutations have not yet emerged in viral variants, but it is unclear from the current study whether the possible escape mutations would negatively impact viral fitness and thus may not ever emerge on a population level. Regardless, the more information we have regarding potential loss of antibody binding and efficacy, the better prepared we will be to understand the importance of a given mutation should it arise in the next variant.

  5. ScreenIT

    SciScore for 10.1101/2021.10.05.463210: (What is this?)

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

    Table 1: Rigor

    EthicsIRB: All samples were de-identified and thus all work was approved by the Fred Hutchinson Cancer Research Center Institutional Review Board as nonhuman subjects research.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    The index creation and short-read alignment step were done using Bowtie2.
    Bowtie2
    suggested: (Bowtie 2, RRID:SCR_016368)
    Samtools was subsequently used to gather sequencing statistics as well as produce the final peptide counts using the stats and idxstats modules.
    Samtools
    suggested: (SAMTOOLS, RRID:SCR_002105)
    Epitope binding region identification: Principal Component Analysis (PCA) via Singular Value Decomposition (SVD) was performed on each set of batch replicates using the scikit-learn package44.
    scikit-learn
    suggested: (scikit-learn, RRID:SCR_002577)

    Results from OddPub: Thank you for sharing your code and data.

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    Our study has important limitations worth noting. Because the Spike Phage-DMS library displays 31 amino acid peptides, we are unable to detect antibodies that bind to conformational epitopes and/or glycosylated epitopes. This is demonstrated by the lack of observable binding to the RBD region, a domain with complex folding and known target of antibodies from infected and vaccinated individuals. However, prior studies of RBD epitopes have already been reported using an overlapping set of samples from the HAARVI Cohort, however, and together these results paint a more complete picture of epitopes across the Spike protein15,23. Finally, we only have 5 individuals within the hospitalized group and this small sample size limits our ability to make conclusions about epitope binding in those with severe infection. Our finding that vaccinated individuals have a broader response across the Spike protein than infected individuals may have important implications for immune durability against future SARS-CoV-2 variants. Evidence suggests that a polyclonal antibody response that is resistant in the face of multiple mutations is necessary for long-lasting immunity against a mutating viral pathogen39. Thus, the polyclonal response to vaccination may provide greater protection from infection than the more focused response after infection. However, the number of epitopes targeted provides just one benchmark and the ability to escape at the population level could also be influenced by the dive...

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

    IdentifierStatusTitle
    NCT04283461Active, not recruitingSafety and Immunogenicity Study of 2019-nCoV Vaccine (mRNA-1…

    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.

  6. Version published to 10.1101/2021.10.05.463210v1 on bioRxiv