Quantitative annotations of T-Cell repertoire specificity.

The specificity of a T-cell receptor (TCR) repertoire determines personalized immune capacity. Existing methods have modeled the qualitative aspects of TCR specificity, while the quantitative aspects remained unaddressed. We developed a package, TCRanno, to quantify the specificity of TCR repertoires. We created deep-learning-based, epitope-aware vector embeddings to infer individual TCR specificity. Then we aggregated clonotype frequencies of TCRs to obtain a quantitative profile of repertoire specificity at epitope, antigen and organism levels. Applying TCRanno to 4195 TCR repertoires revealed quantitative changes in repertoire specificity upon infections, autoimmunity and cancers. Specifically, TCRanno found cytomegalovirus-specific TCRs in seronegative healthy individuals, supporting the possibility of abortive infections. TCRanno discovered age-accumulated fraction of severe acute respiratory syndrome coronavirus 2 specific TCRs in pre-pandemic samples, which may explain the aggressive symptoms and age-related severity of coronavirus disease 2019. TCRanno also identified the encounter of Hepatitis B antigens as a potential trigger of systemic lupus erythematosus. TCRanno annotations showed capability in distinguishing TCR repertoires of healthy and cancers including melanoma, lung and breast cancers. TCRanno also demonstrated usefulness to single-cell TCRseq+gene expression data analyses by isolating T-cells with the specificity of interest.

SARS-CoV-2 epitope-specific T cells: Immunity response feature, TCR repertoire characteristics and cross-reactivity.

The devastating COVID-19 pandemic caused by SARS-CoV-2 and multiple variants or subvariants remains an ongoing global challenge. SARS-CoV-2-specific T cell responses play a critical role in early virus clearance, disease severity control, limiting the viral transmission and underpinning COVID-19 vaccine efficacy. Studies estimated broad and robust T cell responses in each individual recognized at least 30 to 40 SARS-CoV-2 antigen epitopes and associated with COVID-19 clinical outcome. Several key immunodominant viral proteome epitopes, including S protein- and non-S protein-derived epitopes, may primarily induce potent and long-lasting antiviral protective effects. In this review, we summarized the immune response features of immunodominant epitope-specific T cells targeting different SRAS-CoV-2 proteome structures after infection and vaccination, including abundance, magnitude, frequency, phenotypic features and response kinetics. Further, we analyzed the epitopes immunodominance hierarchy in combination with multiple epitope-specific T cell attributes and TCR repertoires characteristics, and discussed the significant implications of cross-reactive T cells toward HCoVs, SRAS-CoV-2 and variants of concern, especially Omicron. This review may be essential for mapping the landscape of T cell responses toward SARS-CoV-2 and optimizing the current vaccine strategy.

Resolving SARS-CoV-2 CD4+ T cell specificity via reverse epitope discovery.

The current strategy to detect immunodominant T cell responses focuses on the antigen, employing large peptide pools to screen for functional cell activation. However, these approaches are labor and sample intensive and scale poorly with increasing size of the pathogen peptidome. T cell receptors (TCRs) recognizing the same epitope frequently have highly similar sequences, and thus, the presence of large sequence similarity clusters in the TCR repertoire likely identify the most public and immunodominant responses. Here, we perform a meta-analysis of large, publicly available single-cell and bulk TCR datasets from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected individuals to identify public CD4+ responses. We report more than 1,200 αßTCRs forming six prominent similarity clusters and validate histocompatibility leukocyte antigen (HLA) restriction and epitope specificity predictions for five clusters using transgenic T cell lines. Collectively, these data provide information on immunodominant CD4+ T cell responses to SARS-CoV-2 and demonstrate the utility of the reverse epitope discovery approach.

Functional changes in cytotoxic CD8+ T-cell cross-reactivity against the SARS-CoV-2 Omicron variant after mRNA vaccination.

Understanding the T-cell responses involved in inhibiting COVID-19 severity is crucial for developing new therapeutic and vaccine strategies. Here, we characterized SARS-CoV-2 spike-specific CD8+ T cells in vaccinees longitudinally. The BNT162b2 mRNA vaccine can induce spike-specific CD8+ T cells cross-reacting to BA.1, whereas the T-cell receptor (TCR) repertoire usages decreased with time. Furthermore the mRNA vaccine induced spike-specific CD8+ T cells subpopulation expressing Granzyme A (GZMA), Granzyme B (GZMB) and Perforin simultaneously in healthy donors at 4 weeks after the second vaccination. The induced subpopulation was not maintained at 12 weeks after the second vaccination. Incorporating factors that efficiently induce CD8+ T cells with highly cytotoxic activity could improve future vaccine efficacy against such variants.

Interpretable artificial intelligence model for accurate identification of medical conditions using immune repertoire.

Underlying medical conditions, such as cancer, kidney disease and heart failure, are associated with a higher risk for severe COVID-19. Accurate classification of COVID-19 patients with underlying medical conditions is critical for personalized treatment decision and prognosis estimation. In this study, we propose an interpretable artificial intelligence model termed VDJMiner to mine the underlying medical conditions and predict the prognosis of COVID-19 patients according to their immune repertoires. In a cohort of more than 1400 COVID-19 patients, VDJMiner accurately identifies multiple underlying medical conditions, including cancers, chronic kidney disease, autoimmune disease, diabetes, congestive heart failure, coronary artery disease, asthma and chronic obstructive pulmonary disease, with an average area under the receiver operating characteristic curve (AUC) of 0.961. Meanwhile, in this same cohort, VDJMiner achieves an AUC of 0.922 in predicting severe COVID-19. Moreover, VDJMiner achieves an accuracy of 0.857 in predicting the response of COVID-19 patients to tocilizumab treatment on the leave-one-out test. Additionally, VDJMiner interpretively mines and scores V(D)J gene segments of the T-cell receptors that are associated with the disease. The identified associations between single-cell V(D)J gene segments and COVID-19 are highly consistent with previous studies. The source code of VDJMiner is publicly accessible at https://github.com/TencentAILabHealthcare/VDJMiner. The web server of VDJMiner is available at https://gene.ai.tencent.com/VDJMiner/.

SARS-CoV-2-specific T cell responses in patients with multisystem inflammatory syndrome in children.

Multisystem inflammatory syndrome in children (MIS-C) is a severe complication of SARS-CoV-2 infections that occurs in the pediatric population. We sought to characterize T cell responses in MIS-C compared to COVID-19 and pediatric hyperinflammatory syndromes. MIS-C was distinct from COVID-19 and hyperinflammatory syndromes due to an expansion of T cells expressing TRBV11-2 that was not associated with HLA genotype. Children diagnosed with MIS-C, but who were negative for SARS-CoV-2 by PCR and serology, did not display Vß skewing. There was no difference in the proportion of T cells that became activated after stimulation with SARS-CoV-2 peptides in children with MIS-C compared to convalescent COVID-19. The frequency of SARS-CoV-2-specific TCRs and the antigens recognized by these TCRs were comparable in MIS-C and COVID-19. Expansion of Vß11-2⁺ T cells was a specific biomarker of MIS-C patients with laboratory confirmed SARS-CoV-2 infections. Children with MIS-C had robust antigen-specific T cell responses to SARS-CoV-2.

SARS-CoV-2 mRNA vaccination elicits a robust and persistent T follicular helper cell response in humans.

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 (TFH) 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 TFH. Mining 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 among the most common HLA alleles in humans. Paired blood and lymph node specimens show that while circulating S-specific TFH cells peak one week after the second immunization, S-specific TFH persist at nearly constant frequencies for at least six months. Collectively, our results underscore the key role that robust TFH cell responses play in establishing long-term immunity by this efficacious human vaccine.

Protective heterologous T cell immunity in COVID-19 induced by the trivalent MMR and Tdap vaccine antigens.

BACKGROUND: T cells control viral infection, promote vaccine durability, and in coronavirus disease 2019 (COVID-19) associate with mild disease. We investigated whether prior measles-mumps-rubella (MMR) or tetanus-diphtheria-pertussis (Tdap) vaccination elicits cross-reactive T cells that mitigate COVID-19. METHODS: Antigen-presenting cells (APC) loaded ex vivo with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), MMR, or Tdap antigens and autologous T cells from COVID-19-convalescent participants, uninfected individuals, and COVID-19 mRNA-vaccinated donors were co-cultured. T cell activation and phenotype were detected by interferon-γ (IFN-γ) enzyme-linked immunospot (ELISpot) assays and flow cytometry. ELISAs (enzyme-linked immunosorbant assays) and validation studies identified the APC-derived cytokine(s) driving T cell activation. TCR clonotyping and single-cell RNA sequencing (scRNA-seq) identified cross-reactive T cells and their transcriptional profile. A propensity-weighted analysis of COVID-19 patients estimated the effects of MMR and Tdap vaccination on COVID-19 outcomes. FINDINGS: High correlation was observed between T cell responses to SARS-CoV-2 (spike-S1 and nucleocapsid) and MMR and Tdap proteins in COVID-19-convalescent and -vaccinated individuals. The overlapping T cell population contained an effector memory T cell subset (effector memory re-expressing CD45RA on T cells [TEMRA]) implicated in protective, anti-viral immunity, and their detection required APC-derived IL-15, known to sensitize T cells to activation. Cross-reactive TCR repertoires detected in antigen-experienced T cells recognizing SARS-CoV-2, MMR, and Tdap epitopes had TEMRA features. Indices of disease severity were reduced in MMR- or Tdap-vaccinated individuals by 32%-38% and 20%-23%, respectively, among COVID-19 patients. CONCLUSIONS: Tdap and MMR memory T cells reactivated by SARS-CoV-2 may provide protection against severe COVID-19. FUNDING: This study was supported by a National Institutes of Health (R01HL065095, R01AI152522, R01NS097719) donation from Barbara and Amos Hostetter and the Chleck Foundation.

Specific CD8+ TCR Repertoire Recognizing Conserved Antigens of SARS-CoV-2 in Unexposed Population: A Prerequisite for Broad-Spectrum CD8+ T Cell Immunity.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has developed variants escaping neutralization antibody immunity established against the original virus. An understanding of broad-spectrum adaptive immunity, including CD8+ T cell immunity to wide range of epitopes, could help translational efforts to improve coronavirus disease 2019 (COVID-19) prevention and therapy. However, there have been few direct studies in which such immunity exists in a population. METHODS: We selected SARS-CoV-2-conserved structural peptides that are not prone to mutation as antigens for broad-spectrum CD8+ T cell immunity. Peripheral blood mononuclear cells (PBMCs) from unexposed healthy donors were stimulated with these peptides in vitro and CD8+ T cell-specific response was monitored. The conserved peptide-specific CD8+ T cells were sorted for T cell receptor (TCR) repertoire sequencing. The presence of specific complementary determining region 3 (CDR3) clones was analyzed in a healthy cohort. RESULTS: For each structural protein, including S, E, M, N, the conserved peptides could potentially provide the largest number of major histocompatibility complex-I (MHC-I) epitopes in the Oriental and Caucasian populations. For conserved peptides from spike (S), envelope (E), membrane (M), nucleocapsid (N) proteins, we found that there were no cross-reactive memory T cells in the unexposed individuals. Instead, their T cells contain naïve TCR repertoire recognizing these conserved peptides. Using TCR sequencing and CDR3 clustering for the conserved peptides specific T cells, we found that the recovered patients had a higher proportion of TCR repertoire similar with that of specific CD8+ T cells in unexposed individuals. Meanwhile, CDR3 clones of the above T cells were widely present in the healthy population. CONCLUSIONS: This study provides evidence of broad-spectrum SARS-CoV-2 specific CD8+ TCR repertoire in unexposed healthy population, which is implicated in the development and implementation of broad-spectrum vaccines against COVID-19.

Profiling of T Cell Repertoire in SARS-CoV-2-Infected COVID-19 Patients Between Mild Disease and Pneumonia.

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a public health emergency. The most common symptoms of COVID-19 are fever, cough, and fatigue. While most patients with COVID-19 present with mild illness, some patients develop pneumonia, an important risk factor for mortality, at early stage of viral infection, putting these patients at increased risk of death. So far, little has been known about differences in the T cell repertoires between COVID-19 patients with and without pneumonia during SARS-CoV-2 infection. Herein, we aimed to investigate T cell receptor (TCR) repertoire profiles and patient-specific SARS-CoV-2-associated TCR clusters between COVID-19 patients with mild disease (no sign of pneumonia) and pneumonia. The TCR sequencing was conducted to characterize the peripheral TCR repertoire profile and diversity. The TCR clustering and CDR3 annotation were exploited to further discover groups of patient-specific TCR clonotypes with potential SARS-CoV-2 antigen specificities. Our study indicated a slight decrease in the TCR repertoire diversity and a skewed CDR3 length usage in patients with pneumonia compared to those with mild disease. The SARS-CoV-2-associated TCR clusters enriched in patients with mild disease exhibited significantly higher TCR generation probabilities and most of which were highly shared among patients, compared with those from pneumonia patients. Importantly, using similarity network-based clustering followed by the sequence conservation analysis, we found different patterns of CDR3 sequence motifs between mild disease- and pneumonia-specific SARS-CoV-2-associated public TCR clusters. Our results showed that characteristics of overall TCR repertoire and SARS-CoV-2-associated TCR clusters/clonotypes were divergent between COVID-19 patients with mild disease and patients with pneumonia. These findings provide important insights into the correlation between the TCR repertoire and disease severity in COVID-19 patients.

Comprehensive analysis of TCR repertoire in COVID-19 using single cell sequencing.

T-cell receptor (TCR) is crucial in T cell-mediated virus clearance. To date, TCR bias has been observed in various diseases. However, studies on the TCR repertoire of COVID-19 patients are lacking. Here, we used single-cell V(D)J sequencing to conduct comparative analyses of TCR repertoire between 12 COVID-19 patients and 6 healthy controls, as well as other virus-infected samples. We observed distinct T cell clonal expansion in COVID-19. Further analysis of VJ gene combination revealed 6 VJ pairs significantly increased, while 139 pairs significantly decreased in COVID-19 patients. When considering the VJ combination of α and ß chains at the same time, the combination with the highest frequency on COVID-19 was TRAV12-2-J27-TRBV7-9-J2-3. Besides, preferential usage of V and J gene segments was also observed in samples infected by different viruses. Our study provides novel insights on TCR in COVID-19, which contribute to our understanding of the immune response induced by SARS-CoV-2.

Deciphering the TCR Repertoire to Solve the COVID-19 Mystery.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected several millions and killed more than quarter of a million worldwide to date. Important questions have remained unanswered: why some patients develop severe disease, while others do not; and what roles do genetic variabilities play in the individual immune response to this viral infection. Here, we discuss the critical role T cells play in the orchestration of the antiviral response underlying the pathogenesis of the disease, COVID-19. We highlight the scientific rationale for comprehensive and longitudinal TCR analyses in COVID-19 and reason that analyzing TCR repertoire in COVID-19 patients would reveal important findings that may explain the outcome disparity observed in these patients. Finally, we provide a framework describing the different strategies, the advantages, and the challenges involved in obtaining useful TCR repertoire data to advance our fight against COVID-19.