J Immunol. 2025 Nov 20:vkaf312. doi: 10.1093/jimmun/vkaf312. Online ahead of print.

ABSTRACT

Peptide-based therapeutic vaccines exploit cross-presentation by dendritic cells for the induction of effective T cell responses. Their clinical success, however, has been limited due to incomplete understanding of antigen processing and presentation (APP). Bioorthogonal chemistry (BOC) uses chemical “click” reactions that can be performed selectively without interference of the cellular environment. A “click handle” can be incorporated into a biomolecule with only minor effects on its characteristics, allowing selective visualization and tracking of the biomolecule. We generated 10 analogues of the HLA-A2-restricted, immunogenic hepatitis B virus-derived epitope HBcAg18-27 by replacing each amino acid with the click handle homopropargylglycine. Each analogue was tested for (1) peptide binding affinity to HLA-A2, (2) preservation of immunogenicity, and (3) accessibility for on-cell click reactions. Lastly, we performed a proof-of-concept experiment in which we demonstrate the feasibility of BOC for APP studies. All amino acids could be modified with the click handle without loss of HLA binding. Furthermore, 7 out of 10 analogues retained cognate T cell recognition. Two of the most promising analogues were tested and demonstrated to be accessible for on-cell click as well as suitable for long peptide processing and presentation studies. To our knowledge, we are the first to show the feasibility of BOC to study APP in the human setting. With BOC techniques, we may now be able to sensitively follow antigen routing by visualizing the intracellular location of (long) peptides. Furthermore, this tool enables direct and quantitative epitope studies in a T cell-independent manner.

PMID:41267163 | DOI:10.1093/jimmun/vkaf312