HLA-A*02:01 Presents Penicillin-Modified Cysteinylated Peptides for T Cell Recognition.

Abstract

A subset of patients experience immune-mediated hypersensitivity reactions towards β-lactam antibiotics, with drug-specific T cells implicated as one of the causative factors. The principal mechanism is thought to involve chemical haptenation of self-peptides, resulting in novel peptide drug-adducts that may trigger T cell recognition. Understanding the interactions between the β-lactam drug, the T cell receptor (TCR) and the peptide/human leukocyte antigen (pHLA) complex is critical to gain further mechanistic insights into these hypersensitivity reactions. This study aimed to 1) explore the array of haptenated ligands presented by HLA-A*02:01, 2) evaluate the repertoire of T cells involved in penicillin-induced reactions in a hypersensitive patient and 3) determine if a dominant penicillin-specific TCR clonotype recognises haptenated HLA peptides. An immunopeptidomics approach was applied to identify benzylpenicillin (BP)-modified peptide ligands within the HLA-A*02:01 ligandome. The drug-reactive TCR repertoire was analysed by single-cell sequencing of CD8⁺ T cells expanded in the presence of BP, and the dominant TCR assayed for reactivity in a reporter cell line. We report that BP modifies cysteine in preference to lysine residues within the HLA-A*02:01 immunopeptidome. This modification occurs via cysteine-drug conjugate formation, in conjunction with disulphide-mediated peptide modification, which has not previously been considered in the context of drug hypersensitivities. Furthermore, we demonstrate that a BP-specific TCR expanded from a patient reacts towards a reduction-sensitive epitope, consistent with a BP-cysteine adduct disulphide linked to a cysteine residue within the T cell epitope. Our study provides evidence that cysteine-penicillin adducts can be accommodated by HLA ligands with the potential to induce T cell-mediated allergic reactions.