Discovering naturally processed antigenic determinants that confer protective T cell immunity
Pavlo Gilchuk, … , Andrew J. Link, Sebastian Joyce
Pavlo Gilchuk, … , Andrew J. Link, Sebastian Joyce
Published April 1, 2013
Citation Information: J Clin Invest. 2013;123(5):1976-1987. https://doi.org/10.1172/JCI67388.
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Technical Advance Immunology

Discovering naturally processed antigenic determinants that confer protective T cell immunity

Abstract

CD8+ T cells (TCD8) confer protective immunity against many infectious diseases, suggesting that microbial TCD8 determinants are promising vaccine targets. Nevertheless, current T cell antigen identification approaches do not discern which epitopes drive protective immunity during active infection — information that is critical for the rational design of TCD8-targeted vaccines. We employed a proteomics-based approach for large-scale discovery of naturally processed determinants derived from a complex pathogen, vaccinia virus (VACV), that are presented by the most frequent representatives of four major HLA class I supertypes. Immunologic characterization revealed that many previously unidentified VACV determinants were recognized by smallpox-vaccinated human peripheral blood cells in a variegated manner. Many such determinants were recognized by HLA class I–transgenic mouse immune TCD8 too and elicited protective TCD8 immunity against lethal intranasal VACV infection. Notably, efficient processing and stable presentation of immune determinants as well as the availability of naive TCD8 precursors were sufficient to drive a multifunctional, protective TCD8 response. Our approach uses fundamental insights into T cell epitope processing and presentation to define targets of protective TCD8 immunity within human pathogens that have complex proteomes, suggesting that this approach has general applicability in vaccine sciences.

Authors

Pavlo Gilchuk, Charles T. Spencer, Stephanie B. Conant, Timothy Hill, Jennifer J. Gray, Xinnan Niu, Mu Zheng, John J. Erickson, Kelli L. Boyd, K. Jill McAfee, Carla Oseroff, Sine R. Hadrup, Jack R. Bennink, William Hildebrand, Kathryn M. Edwards, James E. Crowe Jr., John V. Williams, Søren Buus, Alessandro Sette, Ton N.M. Schumacher, Andrew J. Link, Sebastian Joyce

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Figure 1

Schema of the approach for characterizing naturally processed, HLA class I–restricted TCD8 epitopes.

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Schema of the approach for characterizing naturally processed, HLA class...
HeLa cells engineered to secrete soluble HLA class I molecules (HeLa-sClass I cells) are inoculated with VACV. Secreted class I molecules are immunoaffinity purified from day 1, 2, and 3 p.i. culture supernatant using W6/32 mAb-bound protein A-Sepharose columns. Associated peptides are isolated and separated by reversed-phase HPLC, and their amino acid sequence was determined by LC-MS/MS. Those peptides derived from VACV that conformed to the canonical A2 and B7 binding motifs are validated using the corresponding synthetic peptide spectra and considered naturally processed class I–restricted determinants. That these are TCD8 epitopes is ascertained by functional assays and dual p/class I tetramer staining. Additionally, the protective potential of the TCD8 epitopes is tested in peptide vaccination and a respiratory VACV challenge model.