Allergic drug reactions are immune-mediated idiosyncratic reactions. The main problem in the clinical management of drug allergic patients is the unpredictable nature of the reaction and a lack of understanding of the relationship between drug exposure, drug antigen formation and the stimulation of a pathogenic immune response. Drug-responsive T lymphocytes are thought to play a crucial role in the pathogenesis of many forms of allergic drug reactions, especially reactions that develop in the skin. Studies have shown that cytotoxic CD4+ and CD8+ T cells migrate to the skin of allergic patients and directly kill autologous keratinocytes following drug stimulation. The objective of these studies was to further investigate the role of T cells in (1) patients with cystic fibrosis presenting with allergic reactions to the β-lactam antibiotic piperacillin and (2) a patient that developed severe hepatitis following exposure to trimethoprim. Cloned T cells responsive against the culprit drugs were utilized to define mechanisms of drug antigen presentation, cross-reactivity at individual T cell receptors and the cellular pathophysiology of different forms of allergic reaction. Mass spectrometric methods were employed to detect and fully characterize the nature of piperacillin derived-epitopes on protein which can function as an antigen to stimulate T cells. Albumin modification was time- and concentration- dependent, with selective modification of Lys541 observed at low concentrations, whereas at higher concentrations up to 13/59 lysine residues were modified. Piperacillin-specific T lymphocyte responses (proliferation and cytokines) were detected ex vivo with cells from hypersensitive patients, and analysis of incubation medium showed that modification of the same lysine residues in albumin occurred in situ. The antigenicity of piperacillin-modified albumin was confirmed by stimulation of T lymphocytes with characterized synthetic conjugates. Analysis of minimally-modified T cell stimulatory albumin conjugates revealed peptide sequences incorporating Lys 190, 432 and 541 as principal functional epitopes for T cells. Over 400 piperacillin-responsive CD4+, CD4+8+ or CD8+ T cell clones were generated from five allergic patients. The T cell response was highly specific; clones were stimulated with piperacillin, but not structurally-related compounds. The synthetic piperacillin albumin conjugate stimulated clones via a MHC- restricted pathway involving processing. Flucloxacillin, which modifies essentially the same Lys residues on albumin as piperacillin and occupies non- covalent albumin binding sites, reduced piperacillin albumin binding and the piperacillin-albumin conjugate-specific T cell response. Piperacillin also stimulated T cells in the absence of processing and when antigen presenting cells
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:569887 |
| Date | January 2011 |
| Creators | El-Ghaiesh, Sabah Hussien |
| Publisher | University of Liverpool |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
Page generated in 0.0017 seconds