SARS-CoV-2 mRNA vaccines induce robust anti-spike (S) antibody and CD4 + T cell responses. It is not yet clear whether vaccine-induced follicular helper CD4 + T (T FH ) cell responses contribute to this outstanding immunogenicity. Using fine needle aspiration of draining axillary lymph nodes from individuals who received the BNT162b2 mRNA vaccine, we evaluated the T cell receptor sequences and phenotype of lymph node T FH . Mining of the responding T FH T cell receptor repertoire revealed a strikingly immunodominant HLA-DPB1 ∗ 04-restricted response to S 167-180 in individuals with this allele, which is among the most common HLA alleles in humans. Paired blood and lymph node specimens show that while circulating S-specific T FH cells peak one week after the second immunization, S-specific T FH persist at nearly constant frequencies for at least six months. Collectively, our results underscore the key role that robust T FH cell responses play in establishing long-term immunity by this efficacious human vaccine.

Although mRNA vaccine efficacy against severe COVID-19 remains high, variant emergence has prompted booster immunizations. However, repeated antigen exposure effects on SARS-CoV-2 memory T cells are poorly understood. Here, we utilize MHC-multimers with scRNAseq to profile SARS-CoV-2-responsive T cells ex vivo from humans with one, two, or three antigen exposures, including vaccination, primary, and breakthrough infection. Exposure order determined the distribution between spike- and non-spike-specific responses, with vaccination after infection leading to expansion of spike-specific T cells and differentiation to CCR7-CD45RA+ effectors. In contrast, individuals after breakthrough infection mount vigorous non-spike-specific responses. Analysis of over 4,000 epitope-specific T cell receptor sequences demonstrates that all exposures elicit diverse repertoires characterized by shared TCR motifs, confirmed by monoclonal TCR characterization, with no evidence for repertoire narrowing from repeated exposure. Our findings suggest that breakthrough infections diversify the T cell memory repertoire and current vaccination protocols continue to expand and differentiate spike-specific memory.

SARS-CoV-2 mRNA vaccines generate high and persistent levels of circulating anti-spike (S) antibodies and S-specific CD4+ T cells following prime-boost vaccination. It is not yet clear whether vaccine-induced follicular helper CD4+ T (TFH) cell responses in the draining lymph nodes contribute to this outstanding immunogenicity. Using fine needle aspiration of draining axillary lymph nodes from individuals who received the BNT162b2 mRNA vaccine, we show that frequency of TFH correlates with that of S-binding germinal center B cells. Mining of of the responding TFH T cell receptor repertoire revealed a strikingly immunodominant HLA-DPB1*04-restricted response to S167-180 in individuals with this allele, which is itself among the most common HLA alleles in humans. Analysis of paired blood and lymph node specimens show that circulating S-specific TFH cells peak one week after the second immunization while S-specific lymph node TFH persist at nearly constant frequencies for at least six months following mRNA vaccination. Collectively, our results underscore the key role that robust TFH cell responses play in establishing long-term immunity by this very efficacious human vaccine.

SARS-CoV-2 mRNA vaccines, including Pfizer/Biontech BNT162b2, were shown to be effective for COVID-19 prevention, eliciting both robust antibody responses in naive individuals and boosting pre-existing antibody levels in SARS-CoV-2-recovered individuals. However, the magnitude, repertoire, and phenotype of epitope-specific T cell responses to this vaccine, and the effect of vaccination on pre-existing T cell memory in SARS-CoV-2 convalescent patients, are still poorly understood. Thus, in this study we compared epitope-specific T cells elicited after natural SARS-CoV-2 infection, and vaccination of both naive and recovered individuals. We collected peripheral blood mononuclear cells before and after BNT162b2 vaccination and used pools of 18 DNA-barcoded MHC-class I multimers, combined with scRNAseq and scTCRseq, to characterize T cell responses to several immunodominant epitopes, including a spike-derived epitope cross-reactive to common cold coronaviruses. Comparing responses after infection or vaccination, we found that T cells responding to spike-derived epitopes show similar magnitudes of response, memory phenotypes, TCR repertoire diversity, and αβTCR sequence motifs, demonstrating the potency of this vaccination platform. Importantly, in COVID-19-recovered individuals receiving the vaccine, pre-existing spike-specific memory cells showed both clonal expansion and a phenotypic shift towards more differentiated CCR7-CD45RA+ effector cells. In-depth analysis of T cell receptor repertoires demonstrates that both vaccination and infection elicit largely identical repertoires as measured by dominant TCR motifs and receptor breadth, indicating that BNT162b2 vaccination largely recapitulates T cell generation by infection for all critical parameters. Thus, BNT162b2 vaccination elicits potent spike-specific T cell responses in naive individuals and also triggers the recall T cell response in previously infected individuals, further boosting spike-specific responses but altering their differentiation state. Overall, our study demonstrates the potential of mRNA vaccines to induce, maintain, and shape T cell memory through vaccination and revaccination.

SARS-CoV-2 mRNA vaccines are effective for COVID-19 prevention, eliciting both robust antibody responses in naive individuals and boosting antibody levels in convalescent donors. However, the effect of repeated antigen exposures, such as vaccination following infection or breakthrough infections, on the magnitude, repertoire, and phenotype of pre-existing memory T cells, is still poorly understood. Thus, we compared epitope-specific CD8 T cells elicited after SARS-CoV-2 infection, vaccination of both naive and recovered individuals, and breakthrough infection cases. We used pools of 18 DNA-barcoded MHC-class I multimers, combined with scRNAseq and scTCRseq, to characterize T cell responses as defined by magnitude, specificity, T cell receptor (TCR) repertoire, and gene expression profile to both spike-derived and non-spike derived epitopes. In-depth analysis of over 4000 unique epitope-specific TCR sequences demonstrates that both vaccination and infection, including breakthrough cases, elicit identical repertoires as measured by dominant TCR motifs and repertoire diversity, indicating that BNT162b2 vaccination largely recapitulates spike-specific T cell repertoire generation by infection. Importantly, in COVID-19-recovered individuals receiving the vaccine, pre-existing spike-specific memory cells showed both clonal expansion and a phenotypic shift towards more differentiated CCR7-CD45RA+ effector cells, demonstrating the potency of this vaccine to recall spike-specific CD8 memory T cells. Importantly, in breakthrough infections, we observed a high proportion of T cells targeting non-spike epitopes, showing that new immune memory could be formed during SARS-CoV-2 infection after vaccination. The work was funded by 75N93019C00052, HHSN272201400006C, 3U01AI144616-02S1, R01AI136514