The impact of viral mutations on recognition by SARS-CoV-2 specific T cells

Abstract

We identify amino acid variants within dominant SARS-CoV-2 T cell epitopes by interrogating global sequence data. Several variants within nucleocapsid and ORF3a epitopes have arisen independently in multiple lineages and result in loss of recognition by epitope-specific T cells assessed by IFN-γ and cytotoxic killing assays. Complete loss of T cell responsiveness was seen due to Q213K in the A∗01:01-restricted CD8+ ORF3a epitope FTSDYYQLY_207-215; due to P13L, P13S, and P13T in the B∗27:05-restricted CD8+ nucleocapsid epitope QRNAPRITF_9-17; and due to T362I and P365S in the A∗03:01/A∗11:01-restricted CD8+ nucleocapsid epitope KTFPPTEPK_361-369. CD8+ T cell lines unable to recognize variant epitopes have diverse T cell receptor repertoires. These data demonstrate the potential for T cell evasion and highlight the need for ongoing surveillance for variants capable of escaping T cell as well as humoral immunity.

Keywords: Immune response; Immunology; Molecular biology; Phylogenetics; Virology.

Graphical abstract

Figure 1

Functional impact of mutations in key SARS-CoV-2 dominant epitopes (A–F) Recognition of wild-type (black) and mutant (red) peptide titrations by polyclonal epitope-specific T cell lines in IFN-γ ELISpot assays. SFU, spot forming units. Shown are mean values from three or more replicates +/− standard deviation. (G–J) Ability of CD8+ T cell lines to kill autologous B cells loaded with wild-type (black) or mutant (red) peptides in carboxyfluorescein succinimidyl ester (CFSE) assays. The effector:target ratio denotes the proportion of CD8+ T cells:B cells in each assay. (K–N) Recognition of wild-type (black) and mutant (red) peptide titrations by a polyclonal CD8+ T cell line specific for the HLA∗A01:01-restricted ORF1a epitope TTDPSFLGRY_1637-1646, using intra-cellular cytokine staining for interferon-gamma (IFNg, K), tumor necrosis factor (TNFa, L), and the degranulation factor CD107a (M), and a killing assay (N). Similar findings were seen with a T cell line generated from another donor (Figure S4).

Figure 2

T cell receptor (TCR) repertoire of polyclonal CD8+ T cell lines specific for A∗01:01-restricted ORF3a epitope FTSDYYKLY_207-215 (A, C, E, and G) Data are shown from T cell lines generated using peripheral blood mononuclear cells from four donors. The Q213K variant (red) showed complete loss of recognition in each case using peptide titrations in IFN-γ ELISpot assays compared with recognition of the wild-type (black) peptide. Shown are mean values from three or more replicates +/− standard deviation. SFU, spot forming units. (B, D, F, and H) A diverse range of TCRs was observed.

Figure 3

Global presence of variants in key dominant SARS-CoV-2 epitopes (A) Weekly frequency over time since beginning of SARS-CoV-2 pandemic of all variants studied in functional experiments. Mutation counts were obtained from COG-UK global metadata (dated August 4, 2021). Variants named with prefix of SARS-CoV-2 protein (S, spike; N, nucleocapsid), followed by wild-type amino acid, position within protein, and variant amino acid. Epiweek relates to week number since start of global SARS-CoV-2 pandemic was declared on March 11, 2020. (B–E) Phylogenies representing global SARS-CoV-2 genomes depicting the presence of epitopes variants impacting T cell responses. In each case, phylogenies represent all available variant sequences (red tips), along with a selection of non-variant sequences, which were subsampled for visualization purposes. The bar to the right of each phylogeny is annotated by main ancestral lineages only and not each individual PANGO lineage that viruses belong to. The grapevine pipeline (https://github.com/COG-UK/grapevine) was used for generating the phylogeny based on all data available on GISAID and COG-UK up until August 4, 2021.