Identification of a First Human Norovirus CD8(+) T Cell Epitope Restricted to HLA-A(*)0201 Allele

Norovirus (NoV) causes a substantial global burden of acute gastroenteritis in all age groups and the development of NoV vaccine is a high priority. There are still gaps in understanding of protective NoV-specific immunity. Antibody mediated immune responses have been widely studied, but in contrast, the research on NoV-specific human T cell-mediated immunity is very limited. We have recently reported NoV capsid VP1-specific 18-mer peptide ((134)SPSQVTMFPHIIVDVRQL(151)) to induce strong CD8(+) T cell immune responses in healthy adult donors. This work extends to identify the precise NoV T cell epitope and the restricting human leucocyte antigen (HLA). Pentamer technology was used to detect HLA-A(*)0201-restricted T cell-mediated responses to 10-mer peptide (139)TMFPHIIVDV(148) of four healthy adult blood donors. Immunogenicity of the 10-mer epitope was confirmed by ELISPOT IFN-γ and intracellular cytokine staining (ICS) on flow cytometry. A population of CD3(+)CD8(+) T lymphocytes binding to HLA-A(*)0201/TMFPHIIVDV pentamers was identified in two HLA-A(*)0201-positive donors. Recognition of the 10-mer epitope by T cells resulted in a strong IFN-γ secretion as shown by ELISPOT assay. In addition, ICS confirmed that high proportion (31 and 59%) of the TMFPHIIVDV epitope-responsive CD3(+)CD8(+) T cells in the two donors had multifunctional phenotype, simultaneously producing IFN-γ, IL-2 and TNF-α cytokines. In the present study novel human NoV HLA-A(*)0201-restricted minimal 10-mer epitope (139)TMFPHIIVDV(148) in the capsid VP1 was identified. The HLA-peptide pentamer staining of T cells from healthy donor PBMCs and cytokine responses in ex-vivo ELISPOT and ICS assays suggest that this epitope is recognized during NoV infection and activates memory phenotype of the epitope-specific multifunctional CD8(+) T cells. The importance of this epitope in protection from NoV infection remains to be determined.