Cellular and humoral responses to SARS-CoV-2 vaccination in immunosuppressed patients.

OBJECTIVE: SARS-CoV-2 vaccinations have demonstrated vaccine-immunogenicity in healthy volunteers, however, efficacy in immunosuppressed patients is less well characterised. There is an urgent need to address the impact of immunosuppression on vaccine immunogenicity. METHODS: Serological, T-cell ELISpot, cytokines and immunophenotyping were used to assess vaccine responses (either BNT162b2 mRNA or ChAdOx1 nCoV-19) in double-vaccinated patients receiving immunosuppression for renal transplants or haematological malignancies (n = 13). Immunological responses in immunosuppressed patients (VACC-IS) were compared to immunocompetent vaccinated (VACC-IC, n = 12), unvaccinated (UNVACC, n = 11) and infection-naïve unvaccinated (HC, n = 3) cohorts. RESULTS: No significant different differences in T-cell responses were observed between VACC-IS and VACC-IC (92%) to spike-peptide (S) stimulation. UNVACC had the highest T-cell non-responders (n = 3), whereas VACC-IC and VACC-IS both had one T-cell non-responder. No significant differences in humoral responses were observed between VACC-IC and VACC-IS, with 92% (12/13) of VACC-IS patients demonstrating seropositivity. One VACC-IS failed to seroconvert, however had detectable T-cell responses. All VACC-IC participants were seropositive for anti-spike antibodies. VACC-IS and VACC-IC participants elicited strong Th1 cytokine response with immunodominance towards S-peptide. Differences in T-cell immunophenotyping were seen between VACC-IS and VACC-IC, with lower CD8+ activation and T-effector memory phenotype observed in VACC-IS. CONCLUSION: SARS-CoV-2 vaccines are immunogenic in patients receiving immunosuppressive therapy, with responses comparable to vaccinated immunocompetent participants. Lower humoral responses were seen in patients treated with B-cell depleting therapeutics, but with preserved T-cell responses. We suggest further work to correlate both protective immunity and longevity of these responses in both healthy and immunosuppressed patients.

Trilogy of COVID-19: Infection, Vaccination, and Immunosuppression.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative respiratory pathogen responsible for coronavirus disease 2019 (COVID-19). In 2020, the power of open science was visible to all, as novel vaccinology led to rapid establishment of vaccine clinical trials, and subsequent authorization of SARS-CoV-2 at an unprecedented pace. This evoked rapid deployment of SARS-CoV-2 vaccines and booster doses to keep with the ever-changing landscape of SARS-CoV-2. In this review, we provide an overview of vaccine efficacy studies, which have been well characterized in healthy individuals. Nevertheless, vaccine efficacy within the immunosuppressed is less well characterized, as these individuals were omitted from initial efficacy studies. Consequently, vaccine-induced responses in this group are relatively unknown. Currently, limited evidence investigating vaccine efficacy within the immunosuppressed is available. Here, we provide an overview of SARS-CoV-2 infection and associated pathogenesis. Furthermore, we undertake a critical analysis of observed vaccine responses from clinical studies, conducted in healthy and immunosuppressed populations. Whilst vaccine deployment has curbed mortality, there are significant challenges that lie ahead. This includes correlating vaccine responses with protective immunity and ensuring that global vaccine equity is met.

Antibody Responses to SARS-CoV-2 Infection-Comparative Determination of Seroprevalence in Two High-Throughput Assays versus a Sensitive Spike Protein ELISA.

Robust assay development for SARS-CoV-2 serological testing requires assessment of asymptomatic and non-hospitalised individuals to determine if assays are sensitive to mild antibody responses. Our study evaluated the performance characteristics of two high-throughput SARS-CoV-2 IgG nucleocapsid assays (Abbott Architect and Roche) and The Binding Site (TBS) Anti-Spike IgG/A/M ELISA kit in samples from healthcare workers (HCWs). The 252 samples were collected from multi-site NHS trusts and analysed for SARS-CoV-2 serology. Assay performance was evaluated between these three platforms and ROC curves were used to redefine the Abbott threshold. Concordance between Abbott and TBS was 66%. Any discrepant results were analysed using Roche, which showed 100% concordance with TBS. Analysis conducted in HCWs within 58 days post-PCR result demonstrated 100% sensitivity for both Abbott and Roche. Longitudinal analysis for >100 days post-PCR led to sensitivity of 77.2% and 100% for Abbott and Roche, respectively. A redefined Abbott threshold (0.64) increased sensitivity to 90%, producing results comparable to TBS and Roche. The manufacturer's threshold set by Abbott contributes to lower sensitivity and elevated false-negative occurrences. Abbott performance improved upon re-optimisation of the cut-off threshold. Our findings provided evidence that TBS can be used as bespoke alternative for SARS-CoV-2 serology analysis where high-throughput platforms are not feasible on site.

HLA class I mono-specific APCs and target cells: a method to standardise in vitro CD8+ T cell expansion and functional assays.

The introduction of in vitro T cell expansion and assay methods that are robust and easy to use would be welcome in cancer vaccine and infectious disease research. By coating HLA class I--ve B cells with recombinant HLA class I peptide complexes, we are able to produce antigen presenting cells and target cells expressing a single defined antigen in the context of costimulatory and adhesion molecules. HLA class I mono-specific cells promoted the in vitro expansion of CMV epitope specific CD8+ T cells from 0.03% to 30.6% in 2 weeks, which was comparable to using peptide-loaded dendritic cells. The HLA class I mono-specific cells were also shown to promote in vitro antigen specific T cell function in assays based on measuring cytokine production and cytolytic activity. HLA class I mono-specific cells are simple to prepare, can be used with any recombinant HLA class I allele/peptide combination and should provide a useful system for in vitro T cell expansion and functional analysis.

Brief Report: Immunization of HIV-Infected Adults in the UK With Haemophilus influenzae b/Meningococcal C Glycoconjugate and Pneumococcal Polysaccharide Vaccines.

U.K. guidelines for vaccinating HIV-infected adults against bacteria are based on limited data. We compared antibody responses between 211 HIV-infected and 73 HIV-uninfected adults vaccinated with pneumococcal polysaccharide vaccine (PPV) and Haemophilus influenzae b/meningococcal C polysaccharide-tetanus toxoid glycoconjugate vaccine (Hib/MenC-TT). IgG responses to Hib/MenC-TT were not significantly different. PPV induced median IgGs >1.3 µg/mL for 10/12 serotypes among HIV-uninfected participants and 5/12 in HIV-infected participants. HIV-uninfected adults had higher post-vaccination IgGs than HIV-infected adults for 4/12 serotypes (P < 0.001). Responses did not associate with CD4 count or viral suppression. In a U.K. HIV-infected population, Hib/MenC-TT induced similar responses to HIV-uninfected adults, whereas PPV induced poor responses.

Serum hepcidin-25 and response to intravenous iron in patients with non-dialysis chronic kidney disease.

BACKGROUND: Hepcidin-25 is an iron regulator which reduces iron absorption and promotes sequestration in the reticulo-endothelial system. We investigated hepcidin and traditional iron storage marker utility in predicting haemoglobin increment following bolus intravenous iron. METHODS: The cohort included 129 consecutive non-dialysis chronic kidney disease patients that attended for intravenous iron over a 6-month period. Serum hepcidin-25 levels (determined by mass spectrometry) pre iron infusion and 6 weeks post were compared with ferritin and transferrin saturation in multivariate models. RESULTS: Log10 ferritin [coefficient 0.559 (0.435-0.684) p < 0.001] and log10 high-sensitive C-reactive protein [coefficient 0.092 (0.000-0.184) p = 0.049] were significantly associated with baseline log10 hepcidin-25 levels. Log10 estimated glomerular filtration rate was the only independent determinant of pre-infusion haemoglobin [coefficient 1.37 (0.16-2.59) p = 0.027]. Log10 hepcidin-25 was an independent predictor of haemoglobin increment 6 weeks following iron infusion [coefficient -0.84 (-1.38 to -0.31) p = 0.002]. Ferritin, transferrin saturation and hepcidin had similar predictive utility for a 1 g/dl haemoglobin increase (c-statistics: 0.68, 0.70, 0.69). CONCLUSIONS: Hepcidin is an iron sensor marker which predicts the magnitude of haemoglobin increment following protocolised intravenous iron infusion. Although displaying similar predictive performance to ferritin and transferrin saturation, hepcidin may also play a mechanistic role.

Detection of vimentin-specific autoreactive CD8+ T cells in cardiac transplant patients.

BACKGROUND: Evidence is emerging that autoimmunity can play a role in allograft rejection. Reports have described the presence of autoantibodies in transplant patients and CD4+ autoreactive T cells in rodent models of allograft rejection. The objective of this study was to seek evidence of CD8+ T-cell-mediated autoimmunity in the transplant setting. The author have previously observed autoimmunity to the non-polymorphic cytoskeletal protein vimentin in cardia transplant patients. In this study, vimentin antibody positive patients were screened for the presence of vimentin-specific self-major histocompatibility complex class I-restricted CD8+ T cells. METHODS: Two peptide sequences from vimentin that bound HLA-A*0201 were identified and fluorochrome-labeled A*0201 tetramers with each peptide were constructed to screen for vimentin-specific T cells. RESULTS: Tetramer-binding CD8+ T cells were detected in peripheral blood lymphocytes from two of six patients after expansion by in vitro stimulation with peptide. Tetramer-binding T cells produced interferon-gamma in an antigen-specific fashion. No autoreactive T cells specific for vimentin were detected after peptide stimulation of T cells from eight healthy A*0201-positive volunteers. CONCLUSIONS: This finding is the first evidence of CD8+ T-cell-mediated autoimmunity in human transplant patients.

The role of hepcidin-25 in kidney transplantation.

BACKGROUND: Hepcidin-25 is a peptide hormone involved in iron absorption and homeostasis and found at increased serum levels in conditions involving systemic inflammation, renal dysfunction, and increased adiposity. Hepcidin may play a role in the pathogenesis of anemia, but its role in kidney transplantation is undefined. METHODS: This study enrolled 100 stable patients beyond 12 months after transplantation, from a large single United Kingdom center. Serum hepcidin-25 level, and relevant demographic and laboratory data pertinent to posttransplantation anemia, were measured and collected. Independent predictors of serum hepcidin were evaluated, and the relationship between hepcidin and hemoglobin, assessed. RESULTS: Independent associations were seen between higher hepcidin levels and allograft dysfunction (estimated glomerular filtration rate), increased inflammation (high-sensitivity C-reactive peptide), higher transferrin saturation (a marker of iron stores), and the use of marrow-suppressive medication (P<0.05 for all). Higher fat tissue index (whole-body multifrequency bioimpedance measurement) was also associated with higher hepcidin levels, but this relationship did not persist after adjustment for inflammation (high-sensitivity C-reactive peptide). In turn, inflammation was associated with increased fat tissue index (P=0.01) and male gender (P=0.04). A nonlinear association between serum hepcidin level and hemoglobin was seen, with a progressive fall in hemoglobin as hepcidin levels rose to 100 ng/mL, but little effect thereafter (P=0.009). This association was independent of renal dysfunction and female gender, both of which were also independently associated with a lower hemoglobin level. CONCLUSIONS: These results highlight possible mechanisms of hemoglobin reduction in kidney transplantation patients, and the therapeutic opportunities from understanding the role of hepcidin in this context.

Measurement of antibodies to pneumococcal, meningococcal and haemophilus polysaccharides, and tetanus and diphtheria toxoids using a 19-plexed assay.

The measurement of antibody responses to vaccination is useful in the assessment of immune status in suspected immune deficiency. Previous reliance on enzyme-linked immunoabsorbent assays (ELISA) has been cumbersome, time-consuming and expensive. The availability of flow cytometry systems has led to the development of multiplexed assays enabling simultaneous measurement of antibodies to several antigens. We optimized a flow cytometric bead-based assay to measure IgG and IgM concentrations in serum to 19 antigens contained in groups of bacterial subunit vaccines: pneumococcal vaccines, meningococcal vaccines, Haemophilus influenzae b (Hib), and tetanus and diphtheria toxoid vaccines. 89-SF was employed as the standard serum. The assay was used to determine specific antibody levels in serum from 193 healthy adult donors. IgG and pneumococcal IgM antibody concentrations were measurable across 3 log10 ranges encompassing the threshold protective IgG antibody levels for each antigen. There was little interference between antibody measurements by the 19-plexed assay compared with monoplexed assays, and a lack of cross-reactive IgG antibody, but evidence for cross-reacting IgM antibody for 3/19 pneumococcal antigens. 90th centile values for 15/19 IgG concentrations and 12/12 IgM concentrations of the 193 adult sera were within these ranges and percentages of sera containing protective IgG antibody levels varied from 4% to 95% depending on antigen. This multiplexed assay can simultaneously measure antibody levels to 19 bacterial vaccine antigens. It is suitable for use in standard clinical practice to assess the in vivo immune response to test vaccinations and measure absolute antibody levels to these antigens.

Investigation of peptide involvement in T cell allorecognition using recombinant HLA class I multimers.

Alloreactive T cells are involved in injurious graft rejection and graft-vs-host disease. However, they can also evoke beneficial responses to tumor Ags restricted by foreign MHC molecules. Manipulation of these alloreactivities requires information on the basis of T cell allorecognition. The vigorous T cell response to foreign MHC molecules may arise from peptide-independent recognition of polymorphic residues of foreign MHC molecules or peptide-specific recognition of novel peptides presented by foreign MHC molecules. We investigated CD8+ T cell allorecognition using recombinant HLA class I/peptide complexes. Peptide-specific allorecognition was examined using tetramers of HLA-A*0201 representing five peptides derived from ubiquitously expressed self-proteins that are known to bind endogenously to HLA-A*0201. Distinct subsets of CD8+ T cells specific for each HLA-A*0201/peptide combination were detected within four in vitro-stimulated T cell populations specific for foreign HLA-A*0201. Peptide-independent allorecognition was investigated using artificial Ag-presenting constructs (aAPCs) coated with CD54, CD80, and functional densities of a single HLA-A*0201/peptide combination for four different peptides. None of the four T cell populations specific for foreign HLA-A*0201 were stimulated by the aAPCs, whereas they did produce IFN-gamma upon stimulation with cells naturally expressing HLA-A*0201. Thus, aAPCs did not stimulate putative peptide-independent allorestricted T cells. The results show that these alloreactive populations comprise subsets of T cells, each specific for a self-peptide presented by foreign class I molecules, with no evidence of peptide-independent components.