PT  - JOURNAL ARTICLE
AU  - Sabah El-Ghaiesh
AU  - Manal M. Monshi
AU  - Paul Whitaker
AU  - Rosalind Jenkins
AU  - Xiaoli Meng
AU  - John Farrell
AU  - Ayman Elsheikh
AU  - Daniel Peckham
AU  - Neil French
AU  - Munir Pirmohamed
AU  - B. Kevin Park
AU  - Dean J. Naisbitt
TI  - Characterization of the Antigen Specificity of T-Cell Clones from Piperacillin-Hypersensitive Patients with Cystic Fibrosis
AID  - 10.1124/jpet.111.190900
DP  - 2012 Jun 01
TA  - Journal of Pharmacology and Experimental Therapeutics
PG  - 597--610
VI  - 341
IP  - 3
4099  - http://jpet.aspetjournals.org/content/341/3/597.short
4100  - http://jpet.aspetjournals.org/content/341/3/597.full
SO  - J Pharmacol Exp Ther2012 Jun 01; 341
AB  - 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.