Functional assays of HLA A2-restricted epitope variant of latent membrane protein 1 (LMP-1) of Epstein-Barr virus in nasopharyngeal carcinoma of Southern China and Taiwan

Human leukocyte antigen (HLA) A2 was consistently associated with increased risk for nasopharyngeal carcinoma (NPC) in Chinese populations. Previously we have reported that an Epstein-Barr virus (EBV) strain carrying an HLA A2-restricted epitope variant of LMP-1 is prevalent in NPC in southern China and Taiwan (Lin et al., J. Gen. Virol. 85: 2023-2034, 2004). The variant has mutation selectively involved one of the two anchor residues in position 2 (125 L -> F) and an additional mutation in position 5 (129 M -> I). Functional assays of the epitope variant were carried out in the present work. The stabilization assay on T2 cells indicated that the variant peptide YFL (YFLEILWRL) prevalent in NPC binds to HLA A2 molecules less efficiently than the prototype peptide YLL (YLLEMLWRL). A dose-dependent binding of the HLA A2 molecules with added peptides was observed. In ex vivo cytotoxic T lymphocyte (CTL) assays with CD8-enriched effectors from A2-positive donors revealed that the YLL-specific CTL was able to lyse EBV-infected B cells expressing HLA A2, whereas the CTL recognition was abrogated with the peptide YFL. Cytokine (IFN-gamma) responses, measured both by intracytoplasmic staining and ELISPOT assays after peptide stimulation, also indicated that the variant epitope peptide failed to give an IFN-gamma response. The IFN-gamma response was almost entirely restricted to those tetramer-positive cells. These results show that EBV isolates from NPC of southern China and Taiwan is dominated by an HLA A2-restricted 'epitope-loss variants' of LMP-1, which would allow the virus to resist immune recognition and may in part contribute to the prevalence of NPC in these populations.