SARS-CoV-2 mRNA vaccine elicits transient humoral immunity.

(A) Vaccination and sampling timeline of blood donors in this study. (B) Anti-S IgG titer of serum samples was determined by ELISA. Mean ± SEM (left) and individual data (right) are shown. *, P < 0.05 vs. Pre, 3 wks, 24 wks, respectively. (C) Neutralization activity (ID50) of serum samples was determined by pseudo-virus assay. Mean ± SEM (left) and individual data (right) are shown. *, P < 0.05 vs. 3 wks, 24 wks, respectively. Wks, weeks.

Demographic data of the participants.

Demographic data of the reported clinical adverse effects (at injection site)

Demographic data of the reported clinical adverse effects (systemic symptoms)

Antibody sustainers had highly expanded S-reactive Tfh clonotypes.

(A) Anti-S IgG titer of serum samples from sustainers and decliners is shown individually. (B, C, E, and F) UMAP projection of T cells in single-cell analysis of post-vaccinated samples collected from all donors. Each dot corresponds to a single cell and is colored according to the samples from different time points of donors. All samples together with annotated cell types (B), samples grouped by donor type (decliners and sustainers) (C), top 16 expanded clonotypes (16 clonotypes that had the most cell numbers from each donor) grouped by donor type (E), and top 16 expanded clonotypes grouped by time point and donor type (F) are shown. Tcm, central memory T cells; Tem, effector memory T cells; Treg, regulatory T cells; γδT, γδ T cells. (D) Tfh signature score and expression levels of the canonical Tfh cell markers, IL21, ICOS, PDCD1 and CD200, are shown as heat maps in the UMAP plot.

The location of S epitopes recognized by top expanded T clonotypes from post-vaccination samples.

T cell S epitopes recognized by top expanded TCR clonotypes in post-vaccinated samples from sustainers and decliners are mapped by their locations in S protein. Each short bar indicates a 15-mer peptide that activated the TCRs. Epitopes are shown in different colors according to the subsets of the T cells they activated. Relative frequencies of the T cell subsets are shown in pie charts. Numbers of identified epitopes recognized by a dominant T subset in sustainers (Tfh) are shown in blue bars. NTD, N-terminal domain; RBD, receptor-binding domain; FP, fusion peptide; HR1, heptad repeat 1; CH, central helix; CD, connector domain; HR2, heptad repeat 2; TM, transmembrane domain.

TCR clonotypes expanded in post-vaccinated samples and their TCR usages, epitopes and restricting HLAs.

Reactivity of each clonotype to mutated epitopes in SARS-CoV-2 VOCs.

Characteristics and dynamics of S-cross-reactive clonotypes.

(A) UMAP projection of T cells in single-cell analysis of pre-vaccinated T cells from donors #4, #13, #15, #17, and #8. Each dot corresponds to a single cell and is colored according to the samples from different donors. Annotated cell types are shown. (B) Donor, name of reconstituted clonotypes, cell type, clonotype fraction in T cells from each time points, and expansion ratio of clonotypes that were found in pre-vaccinated samples and had more than 50 cells in the combined pre- and post-vaccinated sample set. For clonotypes that showed more than one type, the major type is listed in the front. The expansion ratio was calculated using the maximum cell fraction at post-vaccination points divided by the cell fraction at the pre-vaccination point of each clonotype. Clonotypes that have an expansion ratio larger than 1 are considered as expanded post-vaccination. Cell fractions at individual time points are shown as heat map. Tfr, follicular regulatory T cells; MAIT, mucosal-associated invariant T cells.

The location of S epitopes of pre-existing S-reactive T cells.

S epitopes recognized by top expanded TCR clonotypes in pre-vaccinated samples are mapped by their locations in S protein. Each short bar indicates a 15-mer peptide that activated the TCRs. Epitopes are shown in different colors according to the subtypes of the T cells they activated. Relative frequencies of the T cell subtypes from all five donors are shown in the pie chart. Numbers of identified epitopes recognized by a dominant T subset of pre-existing clonotypes (Treg) from all donors are shown in green bars.

Frequencies of pre-existing S-reactive clonotypes in the public database of uninfected and infected cohorts.

TCRβ sequences of the top expanded clonotypes in pre-vaccinated samples were investigated in the Adaptive database. Frequencies of detected clonotypes are shown in box plot. Healthy, dataset from 786 healthy donors. COVID, dataset from 1487 COVID-19 patients.

S-cross-reactive TCR clonotypes expanded in pre-vaccinated samples and their TCR usages, epitopes, restricting HLAs and cross-reactive epitopes in microbes other than SARS-CoV-2.

Humoral immune response of BNT162b2 vaccinees.

(A and B) Anti-S IgG titer from all donors at 6 weeks after vaccination was compared between male and female vaccinees (A) or among different age groups (20–39, 40–49, 50–59, 60–69) (B).

Humoral and cellular immune responses of sustainers and decliners.

(A) Demographic data and magnitude of anti-S IgG titer reduction of the sustainers and decliners. Anti-S IgG titer reduction is calculated as the titers at (6 wks – 24 wks) /6 wks. (B) Anti-N IgG titer of serum samples from sustainers at 24 weeks after vaccination. (C) S-specific IFNγ release from bulk CD4+ T cells (left) or CD4+ and CD8+ T cells (right) of sustainers (red) and decliners (blue) was measured using QuantiFERON.

Sustainer individuals had more cells in the circled region than decliner individuals.

(A) UMAP projection of single-cell analysis of post-vaccinated samples shown in individuals. (B) The percentage of circled cells in (A) in CD4+ T cells of each individual is shown. P value was calculated using t-test.

Determination of S epitopes, restricting HLAs and mutated epitope antigenicity for post-vaccinated T cell clonotypes expanded in sustainers and decliners.

Reporter cells expressing TCRs were stimulated with 1 µg/ml of indicated S protein peptide in the presence of transformed B cells or HEK 293T cells expressing indicated HLAs for overnight, and analyzed for GFP and CD69 expression. (A and B) Determination of S epitopes of TCR clonotypes expanded in sustainers (A) and decliners (B). (C and D) Determination of restricting HLAs of TCR clonotypes expanded in sustainers (C) and decliners (D). (E and F) Determination of antigenicity of mutated epitopes of TCR clonotypes expanded in sustainers (E) and decliners (F).

Determination of S epitopes, restricting HLAs and cross-reactive epitopes for pre-existing T cell clonotypes expanded by S stimulation.

Reporter cells expressing TCRs were stimulated with 1 µg/ml of indicated S peptides in the presence of transformed B cells or HEK 293T cells expressing indicated HLAs for overnight, and analyzed for GFP and CD69 expression. (A) Determination of S epitopes of T cell clonotypes. (B) Determination of restricting HLAs of T cell clonotypes. (C) Determination of cross-reactive epitopes of T cell clonotypes. Sequences of cross-reactive peptides are in Table 6.

The pre-existing S-reactive T cell clonotypes did not recognize HCoV epitopes.

The only two clonotypes whose epitope sequences were relatively conserved in HCoV strains, donor #8-pre_9 and pre_10, were tested for their reactivity to the similar HCoV epitope counterparts. Reporter cell lines of these clonotypes were co-cultured with indicated peptides as well as APCs, and analyzed for GFP and CD69 expression.