title: Characterization of the antigen specificity of T-cell clones from piperacillin-hypersensitive patients with cystic fibrosis.
abstract: β-Lactam antibiotics provide the cornerstone of treatment and reduce the rate of decline in lung function in patients with cystic fibrosis, but their use is limited by a high frequency of delayed-type allergic reactions. The objective of this study was to use cloned T-cells expressing a single T-cell receptor from five piperacillin-hypersensitive patients to characterize both the cellular pathophysiology of the reaction and antigen specificity to define the mechanism of activation of T-cells by piperacillin. More than 400 piperacillin-responsive CD4+, CD4+CD8+, or CD8+ T-cell clones were generated from lymphocyte transformation test and ELIspot-positive patients. The T-cell response (proliferation, T helper 2 cytokine secretion, and cytotoxicity) to piperacillin was concentration-dependent and highly specific. Enzyme-linked immunosorbent assay, gel electrophoresis, and mass spectrometry revealed that piperacillin bound exclusively to albumin in T-cell culture. Irreversible piperacillin binding at Lys 190, 195, 199, 432, and 541 on albumin and the stimulation of T-cells depended on incubation time. A synthetic piperacillin albumin conjugate stimulated T-cell receptors via a major histocompatibility complex- and processing-dependent pathway. Flucloxacillin competes for the same Lys residues on albumin as piperacillin, but the resulting conjugate does not stimulate T-cells, indicating that binding of the β-lactam hapten in peptide conjugates confers structural specificity on the activation of the T-cell receptors expressed on drug-specific clones. Collectively, these data describe the cellular processes that underlie the structural specificity of piperacillin antigen binding in hypersensitive patients with cystic fibrosis.
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Department of Pharmacology, Sherrington Building, Ashton Street, University of Liverpool, Liverpool, L69 3GE, United Kingdom.
authors.forename: Sabah
authors.initials: S
authors.lastname: El-Ghaiesh
authors.forename: Manal M
authors.initials: MM
authors.lastname: Monshi
authors.forename: Paul
authors.initials: P
authors.lastname: Whitaker
authors.forename: Rosalind
authors.initials: R
authors.lastname: Jenkins
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Meng
authors.forename: John
authors.initials: J
authors.lastname: Farrell
authors.forename: Ayman
authors.initials: A
authors.lastname: Elsheikh
authors.forename: Daniel
authors.initials: D
authors.lastname: Peckham
authors.forename: Neil
authors.initials: N
authors.lastname: French
authors.forename: Munir
authors.initials: M
authors.lastname: Pirmohamed
authors.forename: B Kevin
authors.initials: BK
authors.lastname: Park
authors.forename: Dean J
authors.initials: DJ
authors.lastname: Naisbitt
mesh_terms: D000293:Adolescent; D000328:Adult; D000900:Anti-Bacterial Agents / Q000276:immunology*; D000938:Antigen-Presenting Cells / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D054407:Chemokine CCL4 / Q000378:metabolism; D002999:Clone Cells; D003550:Cystic Fibrosis / Q000276:immunology*; D004342:Drug Hypersensitivity / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D005260:Female; D006801:Humans; D007378:Interleukins / Q000378:metabolism; D008213:Lymphocyte Activation; D008297:Male; D013058:Mass Spectrometry; D010878:Piperacillin / Q000276:immunology*; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D055815:Young Adult; D047090:beta-Lactams / Q000276:immunology
chemical_list: D000900:Anti-Bacterial Agents; C066507:CCL4 protein, human; D054407:Chemokine CCL4; D000939:Epitopes; D007378:Interleukins; D011948:Receptors, Antigen, T-Cell; D047090:beta-Lactams; D010878:Piperacillin