Discordance between the predicted vs. the actually recognized CD8+ T cell epitopes of HCMV pp65 antigen and aleatory epitope dominance

Abstract

CD8+ T cell immune monitoring aims at measuring the numbers and functions of antigen-specific CD8+ T cell populations engaged during immune responses, providing insights into the magnitude and quality of cell-mediated immunity operational in a test subject. The selection of peptides for ex vivo CD8+ T cell detection is critical, however, because for each restricting HLA class I molecule present in a human individual there is a multitude of potential epitopes within complex antigens, and HLA diversity between the test subjects predisposes CD8+ T cell responses to individualized epitope recognition profiles. We report here on a brute force CD8+ T cell epitope mapping approach for the human cytomegalovirus (HCMV) pp65 antigen on ten HLA-A*02:01-matched HCMV infected human subjects. In this approach, in each test subject, every possible CD8+ T cell epitope was systematically tested; that is 553 individual peptides that walk the sequence of the HCMV pp65 protein in steps of single amino acids. Highly individualized CD8+ T cell response profiles with aleatory epitope recognition patterns were observed. We compared the actually detected epitope utilization in each individual with epitope prediction ranking for the shared HLA-A*02:01 allele, and for additional HLA class I alleles expressed by each individual. No correlation was found between epitopes ranking on the prediction scale and their actual immune dominance. The data suggest that accurate CD8+ T cell immune monitoring might depend on the agnostic reliance on mega peptide pools, or brute force mapping, rather than individualized epitope predictions.

CERI, CEFX, and CPI: Largely Improved Positive Controls for Testing Antigen-Specific T Cell Function in PBMC Compared to CEF

“…Rather pp65 495–503 is just one of several potential CD8+ T cell epitopes to which CD8+ T cells respond in an apparently aleatory (dice-like) manner [ 9 ]. We confirmed this notion in a follow up study testing 10 additional HLA-A*02:01-positive subjects [ 12 ].…”

“…Similar results were obtained for the CMV pp65 495–503 peptide in HLA-A*02:01-positive, CMV-infected donors in the aforementioned study [ 20 ]. This observation was confirmed in a follow-up study that involved 52 HLA-A*02:01-positive, CMV-infected subjects: even though all these subjects developed strong T cell immunity to CMV, 8% of them either did not respond to the CMV pp65 495–503 peptide at all, and 27% displayed a subdominant/cryptic response to the pp65 495–503 peptide [ 12 ].…”

“…There is also an urgent need for negative controls, in particular when using mega peptide pools that consist of hundreds of peptides. For the latter, we refer to a recent communication from our laboratory [ 12 ].…”

CERI, CEFX, and CPI: largely improved positive controls for testing antigen-specific T cell function in PBMC compared to CEF

“…Similar results were obtained for the CMV pp65 495-503 peptide in HLA-A*02:01-positive, CMV-infected donors in the aforementioned study[10]. This observation was confirmed in a follow-up study that involved 52 HLA-A*02:01-positive, CMV-infected subjects: even though all these subjects have developed strong T cell immunity to CMV, 8% of them did not respond to the CMV pp65 495-503 peptide at all, and 27% displayed subdominant/cryptic response to the pp65 495-503 peptide[23].…”

“…Major progress has been made regarding allele-specific epitope prediction [20], but we are still far from customizing peptides for the reliable assessment of CD8+ T cell immunity [19,21,22]. However, selecting a single predicted peptide for immune monitoring that is not actually targeted by CD8+ T cells in an individual will result in false negative results [23]. Therefore, immune monitoring of CD8+ T cells increasingly moves towards usage of large peptide libraries in an attempt to cover all possible CD8+ T cell epitopes, rather than relying on individualized predictions of peptides [24].…”