In-depth analysis of T cell immunity and antibody responses in heterologous prime-boost-boost vaccine regimens against SARS-CoV-2 and Omicron variant.

With the emergence of novel Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) Variants of Concern (VOCs), vaccination studies that elucidate the efficiency and effectiveness of a vaccination campaign are critical to assess the durability and the protective immunity provided by vaccines. SARS-CoV-2 vaccines have been found to induce robust humoral and cell-mediated immunity in individuals vaccinated with homologous vaccination regimens. Recent studies also suggest improved immune response against SARS-CoV-2 when heterologous vaccination strategies are employed. Yet, few data exist on the extent to which heterologous prime-boost-boost vaccinations with two different vaccine platforms have an impact on the T cell-mediated immune responses with a special emphasis on the currently dominantly circulating Omicron strain. In this study, we collected serum and peripheral blood mononuclear cells (PBMCs) from 57 study participants of median 35-year old's working in the health care field, who have received different vaccination regimens. Neutralization assays revealed robust but decreased neutralization of Omicron VOC, including BA.1 and BA.4/5, compared to WT SARS-CoV-2 in all vaccine groups and increased WT SARS-CoV-2 binding and neutralizing antibodies titers in homologous mRNA prime-boost-boost study participants. By investigating cytokine production, we found that homologous and heterologous prime-boost-boost-vaccination induces a robust cytokine response of CD4+ and CD8+ T cells. Collectively, our results indicate robust humoral and T cell mediated immunity against Omicron in homologous and heterologous prime-boost-boost vaccinated study participants, which might serve as a guide for policy decisions.

Superior humoral immunity in vaccinated SARS-CoV-2 convalescence as compared to SARS-COV-2 infection or vaccination.

Emerging variants of concern (VOC) raise obstacles in shaping vaccination strategies and ending the pandemic. Vaccinated SARS-CoV-2 convalescence shapes the current immune dynamics. We analyzed the SARS-CoV-2 VOC-specific cellular and humoral response of 57 adults: 42 convalescent mRNA vaccinated patients (C+V+), 8 uninfected mRNA vaccinated (C-V+) and 7 unvaccinated convalescent individuals (C+V-). While C+V+ demonstrated a superior humoral SARS-CoV-2 response against all analyzed VOC (alpha, delta, omicron) compared to C-V+ and C+V-, SARS-CoV-2 reactive CD4+ and CD8+ T cells, which can cross-recognize the alpha, delta and omicron VOC after infection and/or vaccination were observed in all there groups without significant differences between the groups. We observed a preserved cross-reactive C+V+ and C-V+ T cell memory. An inferior humoral response but preserved cross-reactive T cell memory in C+V- compared to C+V+ was observed, as well as an inferior humoral response but preserved cross-reactive T cell memory in C+V- compared to C-V+. Adaptive immunity generated after SARS-CoV-2 infection and vaccination leads to superior humoral immune response against VOC compared to isolated infection or vaccination. Despite the apparent loss of neutralization potential caused by viral evolution, a preserved SARS-CoV-2 reactive T cell response with a robust potential for cross-recognition of the alpha, delta and omicron VOC was detected in all studied cohorts. Our results may have implications on current vaccination strategies.

Different humoral but similar cellular responses of patients with autoimmune inflammatory rheumatic diseases under disease-modifying antirheumatic drugs after COVID-19 vaccination.

OBJECTIVES: The effect of different modes of immunosuppressive therapy in autoimmune inflammatory rheumatic diseases (AIRDs) remains unclear. We investigated the impact of immunosuppressive therapies on humoral and cellular responses after two-dose vaccination. METHODS: Patients with rheumatoid arthritis, axial spondyloarthritis or psoriatic arthritis treated with TNFi, IL-17i (biological disease-modifying antirheumatic drugs, b-DMARDs), Janus-kinase inhibitors (JAKi) (targeted synthetic, ts-DMARD) or methotrexate (MTX) (conventional synthetic DMARD, csDMARD) alone or in combination were included. Almost all patients received mRNA-based vaccine, four patients had a heterologous scheme. Neutralising capacity and levels of IgG against SARS-CoV-2 spike-protein were evaluated together with quantification of activation markers on T-cells and their production of key cytokines 4 weeks after first and second vaccination. RESULTS: 92 patients were included, median age 50 years, 50% female, 33.7% receiving TNFi, 26.1% IL-17i, 26.1% JAKi (all alone or in combination with MTX), 14.1% received MTX only. Although after first vaccination only 37.8% patients presented neutralising antibodies, the majority (94.5%) developed these after the second vaccination. Patients on IL17i developed the highest titres compared with the other modes of action. Co-administration of MTX led to lower, even if not significant, titres compared with b/tsDMARD monotherapy. Neutralising antibodies correlated well with IgG titres against SARS-CoV-2 spike-protein. T-cell immunity revealed similar frequencies of activated T-cells and cytokine profiles across therapies. CONCLUSIONS: Even after insufficient seroconversion for neutralising antibodies and IgG against SARS-CoV-2 spike-protein in patients with AIRDs on different medications, a second vaccination covered almost all patients regardless of DMARDs therapy, with better outcomes in those on IL-17i. However, no difference of bDMARD/tsDMARD or csDMARD therapy was found on the cellular immune response.

Severe Acute Respiratory Syndrome Coronavirus 2 Cross-Reactive B and T Cell Responses in Kidney Transplant Patients.

BACKGROUND: Immune responses to seasonal endemic coronaviruses might have a pivotal role in protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Those SARS-CoV-2-crossreactive T cells were recently described in immunocompetent individuals. Still, data on cross-reactive humoral and cellular immunity in kidney transplant recipients is currently lacking. METHODS: The pre-existing, cross-reactive antibody B and T cell immune responses against SARS-CoV-2 in unexposed adults with kidney transplantation (Tx, n = 14) and without (non-Tx, n = 12) sampled before the pandemic were compared with 22 convalescent patients with COVID-19 (Cp) applying enzyme-linked immunosorbent assay and flow cytometry. RESULTS: In both unexposed groups, SARS-CoV-2 IgG antibodies were not detectable. Memory B cells binding spike (S) protein SARS-CoV-2 were detected in unexposed individuals (64% among Tx; 50% among non-Tx) and higher frequencies after infection (80% Cp). The numbers of SARS-CoV-2-reactive T cells were comparable between patients who had undergone Tx and those who had not. SARS-CoV-2-reactive follicular T helper cells were present in 61% of the unexposed cohort in both patients who had undergone Tx and those who had not. CONCLUSIONS: Cross-reactive memory B and T cells against SARS-CoV-2 exist also in transplanted adults, suggesting a primed adaptive immunity. The effect on the disease course may depend on the concomitant immunosuppressive drugs.

Neutralizing antibody responses 300 days after SARS-CoV-2 infection and induction of high antibody titers after vaccination.

Neutralizing antibodies against SARS-CoV-2 are important to protect against infection and/or disease. Using an assay to detect antibodies directed against the receptor binding domain (RBD) of SARS-CoV-2 Spike, we identified individuals with SARS-CoV-2 infection after an outbreak at a local health institution. All but one COVID-19 patient developed detectable anti-RBD antibodies and 77% had virus neutralizing antibody titers of >1:25. Antibody levels declined slightly over time. However, we still detected virus neutralizing antibody titers in 64% of the COVID-19 patients at >300 days after infection, demonstrating durability of neutralizing antibody levels after infection. Importantly, full COVID-19 vaccination of these individuals resulted in higher antibody titers compared to fully vaccinated individuals in the absence of prior infection. These data demonstrate long-lived antibody-mediated immunity after SARS-CoV-2 infection, and a clear benefit of two vaccine doses for recovered individuals.

Immune Response in Moderate to Critical Breakthrough COVID-19 Infection After mRNA Vaccination.

SARS-CoV-2 variants of concern (VOCs) can trigger severe endemic waves and vaccine breakthrough infections (VBI). We analyzed the cellular and humoral immune response in 8 patients infected with the alpha variant, resulting in moderate to fatal COVID-19 disease manifestation, after double mRNA-based anti-SARS-CoV-2 vaccination. In contrast to the uninfected vaccinated control cohort, the diseased individuals had no detectable high-avidity spike (S)-reactive CD4+ and CD8+ T cells against the alpha variant and wild type (WT) at disease onset, whereas a robust CD4+ T-cell response against the N- and M-proteins was generated. Furthermore, a delayed alpha S-reactive high-avidity CD4+ T-cell response was mounted during disease progression. Compared to the vaccinated control donors, these patients also had lower neutralizing antibody titers against the alpha variant at disease onset. The delayed development of alpha S-specific cellular and humoral immunity upon VBI indicates reduced immunogenicity against the S-protein of the alpha VOC, while there was a higher and earlier N- and M-reactive T-cell response. Our findings do not undermine the current vaccination strategies but underline a potential need for the inclusion of VBI patients in alternative vaccination strategies and additional antigenic targets in next-generation SARS-CoV-2 vaccines.

Humoral and cellular immunity to SARS-CoV-2 vaccination in renal transplant versus dialysis patients: A prospective, multicenter observational study using mRNA-1273 or BNT162b2 mRNA vaccine.

BACKGROUND: Dialysis and kidney transplant patients are vulnerable populations for COVID-19 related disease and mortality. METHODS: We conducted a prospective study exploring the eight week time course of specific cellular (interferon-γ release assay and flow cytometry) or/and humoral immune responses (ELISA) to SARS-CoV-2 boost vaccination in more than 3100 participants including medical personnel, dialysis patients and kidney transplant recipients using mRNA vaccines BNT162b2 or mRNA-1273. RESULTS: SARS-CoV-2-vaccination induced seroconversion efficacy in dialysis patients was similar to medical personnel (> 95%), but markedly impaired in kidney transplant recipients (42%). T-cellular immunity largely mimicked humoral results. Major risk factors of seroconversion failure were immunosuppressive drug number and type (belatacept, MMF-MPA, calcineurin-inhibitors) as well as vaccine type (BNT162b2 mRNA). Seroconversion rates induced by mRNA-1273 compared to BNT162b2 vaccine were 97% to 88% (p < 0.001) in dialysis and 49% to 26% in transplant patients, respectively. Specific IgG directed against the new binding domain of the spike protein (RDB) were significantly higher in dialysis patients vaccinated by mRNA-1273 (95%) compared to BNT162b2 (85%, p < 0.001). Vaccination appeared safe and highly effective demonstrating an almost complete lack of symptomatic COVID-19 disease after boost vaccination as well as ceased disease incidences during third pandemic wave in dialysis patients. CONCLUSION: Dialysis patients exhibit a remarkably high seroconversion rate of 95% after boost vaccination, while humoral response is impaired in the majority of transplant recipients. Immunosuppressive drug number and type as well as vaccine type (BNT162b2) are major determinants of seroconversion failure in both dialysis and transplant patients suggesting immune monitoring and adaption of vaccination protocols.

Detection of pre-existing SARS-CoV-2-reactive T cells in unexposed renal transplant patients.

BACKGROUND: Recent data demonstrate potentially protective pre-existing T cells reactive against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in samples of healthy blood donors, collected before the SARS-CoV-2 pandemic. Whether pre-existing immunity is also detectable in immunosuppressed patients is currently not known. METHODS: Fifty-seven patients were included in this case-control study. We compared the frequency of SARS-CoV-2-reactive T cells in the samples of 20 renal transplant (RTx) patients to 20 age/gender matched non-immunosuppressed/immune competent healthy individuals collected before the onset of the SARS-CoV-2 pandemic. Seventeen coronavirus disease 2019 (COVID-19) patients were used as positive controls. T cell reactivity against Spike-, Nucleocapsid-, and Membrane- SARS-CoV-2 proteins were analyzed by multi-parameter flow cytometry. Antibodies were analyzed by neutralization assay. RESULTS: Pre-existing SARS-CoV-2-reactive T cells were detected in the majority of unexposed patients and healthy individuals. In RTx patients, 13/20 showed CD4+ T cells reactive against at least one SARS-CoV-2 protein. CD8+ T cells reactive against at least one SARS-CoV-2 protein were demonstrated in 12/20 of RTx patients. The frequency and Th1 cytokine expression pattern of pre-formed SARS-CoV-2 reactive T cells did not differ between RTx and non-immunosuppressed healthy individuals. CONCLUSIONS: This study shows that the magnitude and functionality of pre-existing SARS-CoV-2 reactive T cell in transplant patients is non-inferior compared to the immune competent cohort. Although several pro-inflammatory cytokines were produced by the detected T cells, further studies are required to prove their antiviral protection.

The Magnitude and Functionality of SARS-CoV-2 Reactive Cellular and Humoral Immunity in Transplant Population Is Similar to the General Population Despite Immunosuppression.

BACKGROUND: The ability of transplant (Tx) patients to generate a protective antiviral response under immunosuppression is pivotal in COVID-19 infection. However, analysis of immunity against SARS-CoV-2 is currently lacking. METHODS: Here, we analyzed T cell immunity directed against SARS-CoV-2 spike-, membrane-, and nucleocapsid-protein by flow cytometry and spike-specific neutralizing antibodies in 10 Tx in comparison to 26 nonimmunosuppressed (non-Tx) COVID-19 patients. RESULTS: Tx patients (7 renal, 1 lung, and 2 combined pancreas-kidney Txs) were recruited in this study during the acute phase of COVID-19 with a median time after SARS-CoV-2-positivity of 3 and 4 d for non-Tx and Tx patients, respectively. Despite immunosuppression, we detected antiviral CD4+ T cell-response in 90% of Tx patients. SARS-CoV-2-reactive CD4+ T cells produced multiple proinflammatory cytokines, indicating their potential protective capacity. Neutralizing antibody titers did not differ between groups. SARS-CoV-2-reactive CD8+ T cells targeting membrane- and spike-protein were lower in Tx patients, albeit without statistical significance. However, frequencies of anti-nucleocapsid-protein-reactive, and anti-SARS-CoV-2 polyfunctional CD8+ T cells, were similar between patient cohorts. Tx patients showed features of a prematurely aged adaptive immune system, but equal frequencies of SARS-CoV-2-reactive memory T cells. CONCLUSIONS: In conclusion, a polyfunctional T cell immunity directed against SARS-CoV-2 proteins as well as neutralizing antibodies can be generated in Tx patients despite immunosuppression. In comparison to nonimmunosuppressed patients, no differences in humoral and cellular antiviral-immunity were found. Our data presenting the ability to generate SARS-CoV-2-specific immunity in immunosuppressed patients have implications for the handling of SARS-CoV-2-infected Tx patients and raise hopes for effective vaccination in this cohort.

Robust T Cell Response Toward Spike, Membrane, and Nucleocapsid SARS-CoV-2 Proteins Is Not Associated with Recovery in Critical COVID-19 Patients.

T cell immunity toward SARS-CoV-2 spike (S-), membrane (M-), and nucleocapsid (N-) proteins may define COVID-19 severity. Therefore, we compare the SARS-CoV-2-reactive T cell responses in moderate, severe, and critical COVID-19 patients and unexposed donors. Overlapping peptide pools of all three proteins induce SARS-CoV-2-reactive T cell response with dominance of CD4+ over CD8+ T cells and demonstrate interindividual immunity against the three proteins. M-protein induces the highest frequencies of CD4+ T cells, suggesting its relevance for diagnosis and vaccination. The T cell response of critical COVID-19 patients is robust and comparable or even superior to non-critical patients. Virus clearance and COVID-19 survival are not associated with either SARS-CoV-2 T cell kinetics or magnitude of T cell responses, respectively. Thus, our data do not support the hypothesis of insufficient SARS-CoV-2-reactive immunity in critical COVID-19. Conversely, it indicates that activation of differentiated memory effector T cells could cause hyperreactivity and immunopathogenesis in critical patients.