AZD1222/ChAdOx1 nCoV-19 vaccination induces a polyfunctional spike protein-specific TH1 response with a diverse TCR repertoire

Abstract

AZD1222 (ChAdOx1 nCoV-19), a replication-deficient simian adenovirus–vectored vaccine, has demonstrated safety, efficacy, and immunogenicity against coronavirus disease 2019 in clinical trials and real-world studies. We characterized CD4⁺ and CD8⁺ T cell responses induced by AZD1222 vaccination in peripheral blood mononuclear cells from 296 unique vaccine recipients aged 18 to 85 years who enrolled in the phase 2/3 COV002 trial. Total spike protein–specific CD4⁺ T cell helper type 1 (T_H1) and CD8⁺ T cell responses were increased in AZD1222-vaccinated adults of all ages after two doses of AZD1222. CD4⁺ T_H2 responses after AZD1222 vaccination were not detected. Furthermore, AZD1222-specific T_H1 and CD8⁺ T cells both displayed a high degree of polyfunctionality in all adult age groups. T cell receptor β (TCRβ) sequences from vaccinated participants mapped against TCR sequences known to react to SARS-CoV-2 revealed substantial breadth and depth across the SARS-CoV-2 spike protein for both AZD1222-induced CD4⁺ and CD8⁺ T cell responses. Overall, AZD1222 vaccination induced a polyfunctional T_H1-dominated T cell response, with broad CD4⁺ and CD8⁺ T cell coverage across the SARS-CoV-2 spike protein.

Fig. 1.. T_H1 and CD8⁺ T cell responses are elicited after vaccination with AZD1222 or MenACWY.

PBMCs from human participants vaccinated with AZD1222 or MenACWY at the indicated time point after vaccination were stimulated with SARS-CoV-2 spike peptide pools, and the intracellular cytokine response was measured. (A) Frequencies of total spike protein–specific CD4⁺ T cells producing any combination of T_H1 (IFN-γ, IL-2, or TNF) cytokines are shown. (B) Frequencies of total spike protein–specific CD4⁺ T cells producing individual T_H1 cytokines at the indicated time points are shown. (C) Frequencies of total spike protein–specific CD4⁺ T cells producing any combination of T_H2 (IL-4 or IL-13) cytokines are shown. (D) Frequencies of total spike protein–specific CD4⁺ T cells producing individual T_H2 cytokines at the indicated time points are shown. (E) Frequencies of total spike protein–specific CD8⁺ T cells producing any combination of IFN-γ, IL-2, or TNF after vaccination with AZD1222 or MenACWY. (F) Frequencies of individual CD8⁺ T cell cytokines at the indicated time points in participants vaccinated with AZD1222. For all data, responses to each peptide pool were combined to determine the total spike protein–specific response. In the box and whisker plots, the horizontal line represents median, boxes represent IQR, whiskers extend to the minimum and maximum, and symbols represent each participant. Significant differences between AZD1222 and MenACWY at each time point were determined by two-tailed Mann-Whitney tests. Significant differences between time points within each vaccine group were determined by Kruskal-Wallis test with Dunn’s test to correct for multiple comparisons. All comparisons are not significant unless stated as significant; *P < 0.05 and ****P < 0.0001.

Fig. 2.. Age-specific CD4⁺ T cell responses are observed after AZD1222 vaccination.

(A) Median frequencies with IQRs are shown for CD4⁺ T cells from participants within each age cohort producing IFN-γ, IL-2, TNF, or any combination of these cytokines at the indicated time points after stimulation with SARS-CoV-2 spike peptide pools. Significant differences between time points within each vaccine group were determined by Kruskal-Wallis tests with Dunn’s test to correct for multiple comparisons. All comparisons are not significant unless stated as significant; **P < 0.01, ***P < 0.001, and ****P < 0.0001. (B) Frequencies of antigen-stimulated CD4⁺ T cells producing each combination of IFN-γ, IL-2, and TNF cytokines at day 28 (left) or day 56 (right) after vaccination are shown. Individual participant responses are shown with median represented by the horizontal line. (C) Pie graphs indicate the total proportion of spike protein–specific T_H1 cytokine production averaged for all participants within the indicated age groups at day 28 and day 56 after vaccination. A proportion of multicytokine responses are represented by the black (three cytokines) and gray (two cytokines) arcs.

Fig. 3.. Age-specific CD8⁺ T cell responses are elicited after AZD1222 vaccination.

(A) Median frequencies with IQRs of CD8⁺ T cells producing IFN-γ, IL-2, TNF, or any combination of these cytokines are shown for the indicated time points after stimulation with SARS-CoV-2 spike peptide pools. Significant differences between time points within each vaccine group were determined by Kruskal-Wallis tests with Dunn’s test to correct for multiple comparisons. All comparisons are not significant unless stated as significant; *P < 0.05 and **P < 0.01. (B) Frequencies of antigen-stimulated CD8⁺ T cells producing each combination of IFN-γ, IL-2, and TNF cytokines at day 28 (left) or day 56 (right) after vaccination at the individual level are shown. Individual participant responses are shown with the median represented by the horizontal line. (C) Pie graphs indicate the total proportion of spike protein–specific CD8⁺ T cell cytokine production within the indicated age groups at day 28 and day 56 after vaccination. A proportion of multicytokine responses are represented by the black (three cytokines) and gray (two cytokines) arcs.

Fig. 4.. Spike protein–specific TCR breadth and depth increased after vaccination with two doses of AZD1222.

(A) Spike protein–specific TCR breadth after dose two (D2) of vaccination with MenACWY (blue) or AZD1222 (orange) is shown. Breadth indicates SARS-CoV-2–associated unique TCRs of total unique TCRs. (B) Spike protein–specific TCR breadth is shown disaggregated by age. (C) Spike protein–specific TCR depth after vaccination with MenACWY (blue) or AZD1222 (orange) is shown. Depth indicates SARS-CoV-2–associated T cells of total T cells. (D) Spike protein–specific TCR depth is shown disaggregated by age. Data are log₁₀-transformed, where all 0 values have been converted to half of the lowest nonzero value. In the box and whisker plots, the horizontal line represents median, boxes represent IQR, whiskers extend to the minimum and maximum, and symbols represent each participant. The dotted lines represent the lower detected value. Significant differences were determined by one-way ANOVA followed by Sidak’s multiple comparisons tests. All comparisons are not significant unless stated as significant; ***P < 0.001 and ****P < 0.0001.

Fig. 5.. Spike protein–specific CD4⁺ and CD8⁺ T cell responses demonstrate substantial breadth and depth across the entire spike protein.

TCR sequences from participants vaccinated with AZD1222 at day 28 after the second dose were mapped against TCR sequences known to react to SARS-CoV-2. (A and B) CD4⁺ T cell breadth (A) and depth (B) were analyzed for each participant. (C and D) CD8⁺ T cell breadth (C) and depth (D) were analyzed for each participant. The number of participants displaying responses for each epitope region is indicated at the top of each plot (n). Sequences of known variants were aligned to known Multiplex Identification of T cell Receptor Antigen Specificity (MIRA) antigen locations, with any mutations observed in B.1.351 (yellow), B.1.1.7 (blue), or both (red) highlighted. In the box and whisker plots, the horizontal line represents median, boxes represent IQR, whiskers extend to the 5th and 95th percentiles, and individual symbols represent outlier samples.