Discordance Between the Predicted Versus the Actually Recognized CD8+ T Cell Epitopes of HCMV pp65 Antigen and Aleatory Epitope Dominance

CD8+ T cell immune monitoring aims at measuring the size and functions of antigen-specific CD8+ T cell populations, thereby providing insights into cell-mediated immunity operational in a test subject. The selection of peptides for ex vivo CD8+ T cell detection is critical because within a complex antigen exists a multitude of potential epitopes that can be presented by HLA class I molecules. Further complicating this task, there is HLA class I polygenism and polymorphism which predisposes CD8+ T cell responses towards individualized epitope recognition profiles. In this study, we compare the actual CD8+ T cell recognition of a well-characterized model antigen, human cytomegalovirus (HCMV) pp65 protein, with its anticipated epitope coverage. Due to the abundance of experimentally defined HLA-A(*)02:01-restricted pp65 epitopes, and because in silico epitope predictions are most advanced for HLA-A(*)02:01, we elected to focus on subjects expressing this allele. In each test subject, every possible CD8+ T cell epitope was systematically covered testing 553 individual peptides that walk the sequence of pp65 in steps of single amino acids. Highly individualized CD8+ T cell response profiles with aleatory epitope recognition patterns were observed. No correlation was found between epitopes’ ranking on the prediction scale and their actual immune dominance. Collectively, these data suggest that accurate CD8+ T cell immune monitoring may necessitate reliance on agnostic mega peptide pools, or brute force mapping, rather than electing individual peptides as representative epitopes for tetramer and other multimer labeling of surface antigen receptors.