Identification of novel CD8+ immune epitopes relevant for EBV-related cancers.

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Background: EBV infects 95% of the world’s population and is associated with EBV related cancers that include Asian prevalent nasopharyngeal carcinoma (NPC) amongst others. Asian HLA restricted EBV viral immune epitope target identification and specifically CD8 epitopes continue to be an unmet need that requires to be addressed with novel methods to inform downstream vaccine development. Methods: Firstly, a lead protein latent membrane protein 2 (LMP2) was identified from 72 proteins on the basis of high: i) expression; ii) predicted immunogenicity; and iii) conservation across EBV strains. Secondly, HLA-A*11:01 restricted peptides predicted in silico (NetMHCPan4.1) identified 7 peptides from three predicted immunogenic regions, then, focusing on a single most immunogenic region only (highest number and affinity in silico predictions), predicted high affinity peptides were experimentally validated using proprietary biophysical peptide MHC binding assay, EZ-MHC. Thirdly, this region was interrogated for additional novel unpredicted immunogenic peptides. Results: A comprehensive 50 peptide panel ranging from 8-14 mer across this single region was designed, peptides procured then tested for experimental peptide HLA-A*11:01 binding (EZ-MHC). Of the 50 peptides screened, 11 novel, in silico unpredicted, EBV LMP2 stable binding peptides were identified from this single region. Conclusions: Additional novel CD8 epitopes were found with unbiased interrogation of peptides derived from a single region that was predicted to be most immunogenic on EBV LMP2 identifying novel lead targets. Further downstream functional validation of these novel targets will further identify the best putative peptide vaccine candidates. These can potentially be harnessed as therapeutic CD8 T cell directed peptide vaccines for not only EBV NPC patients but also patients with other EBV related cancers.