Adaptive immune responses are larger and functionally preserved in a hypervaccinated individual
2024; Elsevier BV; Volume: 24; Issue: 5 Linguagem: Inglês
10.1016/s1473-3099(24)00134-8
ISSN1474-4457
AutoresKatharina Kocher, Carolin Moosmann, Felix Drost, Christine Schülein, Pascal Irrgang, Philipp Steininger, Jahn Zhong, Johannes Träger, Bernd M. Spriewald, Christoph Bock, Dirk H. Busch, Christian Bogdan, Benjamin Schubert, Thomas Winkler, Matthias Tenbusch, Ev-Marie Schuster, Kilian Schober,
Tópico(s)T-cell and B-cell Immunology
ResumoPrime-boost vaccinations can enhance immune responses,1Pollard AJ Bijker EM A guide to vaccinology: from basic principles to new developments.Nat Rev Immunol. 2021; 21: 83-100Crossref PubMed Scopus (607) Google Scholar whereas chronic antigen exposure can cause immune tolerance.2ElTanbouly MA Noelle RJ Rethinking peripheral T cell tolerance: checkpoints across a T cell's journey.Nat Rev Immunol. 2021; 21: 257-267Crossref PubMed Scopus (0) Google Scholar In humans, the benefits, limitations, and risks of repetitive vaccination remain poorly understood. Here, we report on a 62-year-old male hypervaccinated individual from Magdeburg, Germany (HIM), who deliberately and for private reasons received 217 vaccinations against SARS-CoV-2 within a period of 29 months (figure A; appendix 1 tab 1). HIM's hypervaccination occurred outside of a clinical study context and against national vaccination recommendations. Evidence for 130 vaccinations in a 9 month period was collected by the public prosecutor of Magdeburg, Germany, who opened an investigation of this case with the allegation of fraud, but criminal charges were not filed. 108 vaccinations are individually recorded and partly overlap with the total of 130 prosecutor-confirmed vaccinations (appendix 2 p 12). To investigate the immunological consequences of hypervaccination in this unique situation, we submitted an analysis proposal to HIM via the public prosecutor. HIM then actively and voluntarily consented to provide medical information and donate blood and saliva. This procedure was approved by the local Ethics Committee of the University Hospital of Erlangen, Germany. Throughout the entire hypervaccination schedule HIM did not report any vaccination-related side effects. From November 2019, to October 2023, 62 routine clinical chemistry parameters showed no abnormalities attributable to hypervaccination (appendix 1 tab 2). Furthermore, HIM had no signs of a past SARS-CoV-2 infection, as indicated by repeatedly negative SARS-CoV-2 antigen tests, PCRs and nucleocapsid serology (figure A; appendix 1 tab 1).FigureHypervaccination increases the quantity, but not the quality, of adaptive immunityShow full caption(A) HIM's SARS-CoV-2 vaccination, infection testing and sample collection history. See appendix 2 (p 12) for the numbers and overlap of confirmed and reported, total and individually recorded vaccinations. (B) Timeline of spike-specific IgG in serum. Timepoints are displayed relative to the 3rd vaccination for controls or 215th vaccination for HIM, respectively. For controls with a SARS-CoV-2 breakthrough infection, post-infection timepoints are excluded. n=29 controls. (C) Neutralisation capacity against wildtype and Omicron spike-protein of HIM's and controls' sera in a pseudotype neutralisation assay. (D–E) Frequency of spike epitope (HLA-A*01/LTD)-specific CD8+ T cells. n=5 controls. (F–G) Differentiation phenotype of LTD-specific CD8+ T cells on day 189 after 3rd vaccination (for controls) or 215th vaccination (for HIM). Absolute cell numbers are reported per 1 million PBMCs (G). n=4 controls. (H) T-cell clonality of LTD-specific CD8+ T cells determined by scRNAseq (10× Genomics, Pleasanton CA, USA). Segments of bars indicate individual clones (grey segments=1 cell; blue segments >1 cell). Numbers on top of the bars represent total number of clones and Gini coefficient to indicate clonality. (I) Proliferation index of LTD-specific CD8+ T cells upon stimulation with LTD-peptide for 14 days. n=7 control samples were collected from day 76 to 568 after last spike-exposure (vaccination or infection). (J–L) Cytokine expression of CD8+ T cells after LTD-peptide stimulation. (J) Representative plots of IFNγ/TNFα expression. (K) Quantification of IFNγ+ within CD8+ T cells after stimulation. (L) Normalisation of IFNγ+ to the LTD-specific fraction of CD8+ T cells. Connected dots represent individual donors (B,E). Data points represent individual donors, bars or solid lines and error bars indicate the mean +/- s.d. (C,G,I,K,L). Fold-differences to mean of controls are indicated (C,G,I,K,L). HIM=hypervaccinated individual from Magdeburg, Germany. J&J=Johnson & Johnson Ad26.COV2.S vaccine. AZ=AstraZeneca Vaxzevria vaccine. Moderna=Spikevax vaccine. BNT Original=Pfizer–BioNTech Comirnaty original vaccine. BNT-BA 4–5=Comirnaty original/Omicron BA 4–5. BNT-BA.1=Comirnaty original/Omicron BA.1. GSK/Sanofi=Vidprevtyn Beta vaccine. BNT-XBB.1.5=Comirnaty Omicron XBB.1.5. nd=not detected. IC50=half-maximal inhibitory concentration. LTD=LTDEMIAQY epitope. PE=phycoerythrin. PBMC=peripheral blood mononuclear cells. A*01/LTD-multimer-PE=peptide human leukocyte antigen (HLA) multimer for HLA-A*01:01 presenting the LTD epitope peptide and conjugated to the PE dye. TN-like/SCM=naive-like and stem cell memory T cells. TCM=central memory T cells. TEM=effector memory T cells. TEMRA=effector memory T cells upregulating CD45RA.View Large Image Figure ViewerDownload Hi-res image Download (PPT) (A) HIM's SARS-CoV-2 vaccination, infection testing and sample collection history. See appendix 2 (p 12) for the numbers and overlap of confirmed and reported, total and individually recorded vaccinations. (B) Timeline of spike-specific IgG in serum. Timepoints are displayed relative to the 3rd vaccination for controls or 215th vaccination for HIM, respectively. For controls with a SARS-CoV-2 breakthrough infection, post-infection timepoints are excluded. n=29 controls. (C) Neutralisation capacity against wildtype and Omicron spike-protein of HIM's and controls' sera in a pseudotype neutralisation assay. (D–E) Frequency of spike epitope (HLA-A*01/LTD)-specific CD8+ T cells. n=5 controls. (F–G) Differentiation phenotype of LTD-specific CD8+ T cells on day 189 after 3rd vaccination (for controls) or 215th vaccination (for HIM). Absolute cell numbers are reported per 1 million PBMCs (G). n=4 controls. (H) T-cell clonality of LTD-specific CD8+ T cells determined by scRNAseq (10× Genomics, Pleasanton CA, USA). Segments of bars indicate individual clones (grey segments=1 cell; blue segments >1 cell). Numbers on top of the bars represent total number of clones and Gini coefficient to indicate clonality. (I) Proliferation index of LTD-specific CD8+ T cells upon stimulation with LTD-peptide for 14 days. n=7 control samples were collected from day 76 to 568 after last spike-exposure (vaccination or infection). (J–L) Cytokine expression of CD8+ T cells after LTD-peptide stimulation. (J) Representative plots of IFNγ/TNFα expression. (K) Quantification of IFNγ+ within CD8+ T cells after stimulation. (L) Normalisation of IFNγ+ to the LTD-specific fraction of CD8+ T cells. Connected dots represent individual donors (B,E). Data points represent individual donors, bars or solid lines and error bars indicate the mean +/- s.d. (C,G,I,K,L). Fold-differences to mean of controls are indicated (C,G,I,K,L). HIM=hypervaccinated individual from Magdeburg, Germany. J&J=Johnson & Johnson Ad26.COV2.S vaccine. AZ=AstraZeneca Vaxzevria vaccine. Moderna=Spikevax vaccine. BNT Original=Pfizer–BioNTech Comirnaty original vaccine. BNT-BA 4–5=Comirnaty original/Omicron BA 4–5. BNT-BA.1=Comirnaty original/Omicron BA.1. GSK/Sanofi=Vidprevtyn Beta vaccine. BNT-XBB.1.5=Comirnaty Omicron XBB.1.5. nd=not detected. IC50=half-maximal inhibitory concentration. LTD=LTDEMIAQY epitope. PE=phycoerythrin. PBMC=peripheral blood mononuclear cells. A*01/LTD-multimer-PE=peptide human leukocyte antigen (HLA) multimer for HLA-A*01:01 presenting the LTD epitope peptide and conjugated to the PE dye. TN-like/SCM=naive-like and stem cell memory T cells. TCM=central memory T cells. TEM=effector memory T cells. TEMRA=effector memory T cells upregulating CD45RA. HIM's anti-spike SARS-CoV-2 IgG levels were quantified in Bremen and Erlangen, Germany, starting with measurements before the 214th vaccination. A reference cohort of 29 vaccinees (55% female, 45% male) with a three-dose mRNA regimen served as a control group.3Irrgang P Gerling J Kocher K et al.Class switch toward noninflammatory, spike-specific IgG4 antibodies after repeated SARSCoV-2 mRNA vaccination.Sci Immunol. 2023; 8eade2798Crossref Scopus (50) Google Scholar HIM's anti-spike IgG levels were highest on the day of the 214th vaccination and on day 3 after the 215th vaccination, but contraction kinetics subsequently mirrored those of the control group (figure B). While putative boosting through the 216th vaccination could not be assessed, the 217th vaccination yielded a modest increase in anti-spike IgG levels. Repeated SARS-CoV-2 mRNA vaccination can induce unusual IgG subclass switching to IgG4.3Irrgang P Gerling J Kocher K et al.Class switch toward noninflammatory, spike-specific IgG4 antibodies after repeated SARSCoV-2 mRNA vaccination.Sci Immunol. 2023; 8eade2798Crossref Scopus (50) Google Scholar, 4Buhre JS Pongracz T Künsting I et al.mRNA vaccines against SARS-CoV-2 induce comparably low long-term IgG Fc galactosylation and sialylation levels but increasing long-term IgG4 responses compared to an adenovirus-based vaccine.Front Immunol. 2023; 13: 1-18Crossref Scopus (17) Google Scholar, 5Routhu NK Stampfer SD Lai L et al.Efficacy of mRNA-1273 and Novavax ancestral or BA.1 spike booster vaccines against SARS-CoV-2 BA.5 infection in nonhuman primates.Sci Immunol. 2023; 8: 1-16Crossref Scopus (5) Google Scholar HIM's anti-spike IgG4 antibodies at day 189 after the 215th vaccination were elevated in absolute numbers, but not in relative frequencies, compared with control individuals at day 189 after their 3rd vaccination (appendix 2 p 12 B–C). Notably, IgG4 subclass switching has been described to be scarce in heterologous vaccination schemes with adenoviral-based vaccines (eg, Vaxzevria [Sodertalje, AstraZeneca, Sweden]) as a first dose.3Irrgang P Gerling J Kocher K et al.Class switch toward noninflammatory, spike-specific IgG4 antibodies after repeated SARSCoV-2 mRNA vaccination.Sci Immunol. 2023; 8eade2798Crossref Scopus (50) Google Scholar, 4Buhre JS Pongracz T Künsting I et al.mRNA vaccines against SARS-CoV-2 induce comparably low long-term IgG Fc galactosylation and sialylation levels but increasing long-term IgG4 responses compared to an adenovirus-based vaccine.Front Immunol. 2023; 13: 1-18Crossref Scopus (17) Google Scholar Serum anti-spike IgM and IgA were only mildly elevated compared with controls (appendix 2 p 12 D–E). In saliva, HIM had detectable anti-spike IgG, unlike the control participants (appendix 2 p 12 F). HIM's serum neutralisation capacity was 5·4-fold and 11·5-fold higher compared with the control group vaccinees, for wildtype and Omicron B1.1.529 spike proteins, respectively (figure C). This reflects higher quantities of spike-specific IgG, since antibody avidity was comparable (appendix 2 p 12 G). Compared with control participants, HIM showed mildly elevated frequencies of spike-specific B cells, but a similar phenotype in single-cell RNA sequencing, with most spike-binding cells detectable in CD27+ memory B-cell clusters (appendix 2 p 12 H–I). We did not observe a higher degree of clonal expansion or somatic hypermutation rates (appendix 2 p 12 J). CD8+ T cells specific for the immunodominant spike epitope HLA-A*01/LTD were about 6-fold more frequent than in controls at day 189 after 215th or 3rd vaccination, and were also boosted by the 217th vaccination (figure D–E; appendix 1 tab 3). The T-cell phenotype was biased towards effector memory T cells in absolute and relative numbers, although in absolute numbers HIM had as many cells with an early differentiated stem-like CD62L+/CCR7+ phenotype as control individuals did (figure F–G; appendix 2 p 13 A). T cells specific for an Epstein–Barr virus epitope showed no such differences in size or phenotype (appendix 2 p 13 B–C). Single-cell RNA sequencing of LTD-specific T cells including CITEseq (10x Genomics, Pleasanton CA, USA) for 130 surface proteins confirmed a more differentiated T-cell phenotype (appendix 2 pp 13 D–G; appendix 1 tab 4), and revealed a higher degree of clonal expansion compared with vaccinees in the control group (figure H). Further flow-cytometric analysis of 14 protein markers and metabolic profiling did not show major abnormalities (appendix 2 p 14). The proliferative capacity of HIM's LTD-specific CD8+ T cells was similar to the capacity found in control individuals (figure I), in line with conserved absolute numbers of T cells with an early differentiated stem-like phenotype. HIM had more cytokine-positive cells after epitope-specific stimulation than control individuals (figure J-K; appendix 2 p 15) due to higher absolute numbers of epitope-specific CD8+ effector memory and effector T cells (figure G; appendix 2 p 15 B). However, cytokine release was approximately equal on a per-cell-basis (figure L; appendix 2 p 15 C). Further assessment of ten cytokines in the supernatant confirmed a pattern typical of virus-specific CD8+ T cells (appendix 2 p 15 D). Notably, HIM's CD8+ T cells displayed higher peptide sensitivity than those in the control group (appendix 2 p 15 E–F). Finally, analysis of spike-reactive CD4+ T cells confirmed a lack of nucleocapsid-specific immunity and showed similar amounts of cytokine-producing CD4+ T cells in HIM compared with the control group, with preserved peptide sensitivity (appendix 2 p 15 G). In summary, our case report shows that SARS-CoV-2 hypervaccination did not lead to adverse events and increased the quantity of spike-specific antibodies and T cells without having a strong positive or negative effect on the intrinsic quality of adaptive immune responses. While we found no signs of SARS-CoV-2 breakthrough infections in HIM to date, it cannot be clarified whether this is causally related to the hypervaccination regimen. Importantly, we do not endorse hypervaccination as a strategy to enhance adaptive immunity. We declare no conflicts of interest. Download .xlsx (.04 MB) Help with xlsx files Supplementary appendix 1 Download .pdf (5.5 MB) Help with pdf files Supplementary appendix 2
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