Oral Presentation 26th Annual Lorne Proteomics Symposium 2021

The Benzylpenicillin haptenome and its impact on the HLA-A*02:01 immunopeptidome (#44)

Shawn Goh 1 , Katherine Scull 1 , Kirti Pandey 1 , Robert Puy 2 , Robyn O'Hehir 2 , Anthony Purcell 1 , Nicole Mifsud 1 , Patricia Illing 1
  1. Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia
  2. Department of Allergy, Immunology and Respiratory Medicine, Monash University, Central Clinical School, The Alfred Hospital, Melbourne, VIC, Australia

Penicillins are a class of β-lactam antibiotics used as a first-line treatment against a broad array of bacteria. However, following administration around 10% of individuals experience immune-mediated hypersensitivity reactions, which restricts treatment options in these individuals. Clinical investigations suggest that penicillin hypersensitivity is associated with HLA-A*02:01, a common HLA allotype found in 40% of the Caucasian population that is involved in T cell-mediated immune responses. We have previously demonstrated the presence of Benzylpenicillin-specific CD8+ T cells in a HLA-A*02:01+ hypersensitive patient and hypothesise that the presentation of drug-modified peptides by HLA-A*02:01 is driving the immunological drug response.

 

Study Aims:

1) Identify intracellular targets of Benzylpenicillin and predict if peptides encompassing these haptenation sites can furnish ligands for HLA allotypes associated with increased risk of penicillin-hypersensitivity

2) Identify naturally presented HLA ligands with the Benzylpenicillin adduct

 

To map the Benzylpenicillin haptenome of treated antigen presenting cells (APCs), we employed an MS-based bottom-up approach. Benzylpenicillin-modified proteins and sites were identified based on a mass addition at Lys or Arg residues, and further validated by the presence of diagnostic Benzylpenicillin ions in spectra used for peptide assignment. Utilising a predictive tool (NetMHCpan, v4.1b), we have shown that peptides encompassing haptenation sites are predicted ligands of HLA allotypes associated with increased risk of penicillin-hypersensitivity. This suggests that haptenated intracellular proteins undergo antigen processing to be presented by HLA presentation for T cell recognition. Furthermore, we have isolated drug modified peptides directly from immunoaffinity purified HLA-A*02:01 of drug treated cells.

 

This study provides essential knowledge of drug modified antigens generated by Benzylpenicillin and their capacity to enter the antigen presentation pathway for display at the cell surface by HLA molecules. Together, the data will help inform rational intervention strategies based on the immunogenicity of identified drug-modified peptides and their interactions with T cells.