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Title: The phenotype and function of hapten specific T-Cell isolated from hypersensitive patients and healthy human donors
Author: Wang, Eryi
ISNI:       0000 0004 6422 1400
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2015
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Drug hypersensitivity reactions are an important problem for pharmaceutical industry, especially when reactions are observed in late phase clinical trials. Furthermore, management of patients with reactions leads to personal suffering and financial burden on the NHS. Reactions are almost impossible to predict as there is no simple relationship between the dose of drug administered and the development of hypersensitivity. In recent years, pharmacogenetic studies identified strong associations between the expression of specific HLA alleles and susceptibility to different forms of hypersensitivity, which would explain why only a small number of drug-exposed patients develop hypersensitivity. Studies utilizing peripheral blood mononuclear cells have detected drug-specific T-cells in patients with hypersensitivity, but not drug-exposed tolerant controls, suggesting that the adaptive immune system plays an important role in the disease pathogenesis. Despite this, there remains a need to further understand mechanisms as more detailed knowledge will assist the development of diagnostic and predictive assays. Data described herein utilized hypersensitivity reactions to the β-lactam antibiotic piperacillin as a model to investigate the phenotype and function of drug-specific T-cells in blood and skin, focusing specifically on the profile of cytokines secreted. PBMC from hypersensitive patients were activated to proliferate in vitro with piperacillin. T-cell clones responsive to the drug were generated from blood of all patients studied. CD4+ clones were stimulated to proliferate with piperacillin in a concentration-dependent manner and the proliferative response was associated with secretion of Th1 and Th2 cytokines alongside IL-22. In contrast, IL-17 was not secreted from piperacillin-specific clones. Piperacillin-specific CD4+ clones were also isolated from inflamed skins of 2 piperacillin hypersensitive patients. Activation of these clones was associated with secretion of Th1, Th2 cytokines and IL-22, in the absence of IL-17. Finally, CD4+ nitroso sulfamethoxazole (SMX-NO)-responsive clones were isolated from sulfamethoxazole hypersensitive patients and a similar cytokine secretion profile was observed, which suggests that IL-22 secretion might be a common feature of drug hypersensitivity. Evolution of T-cell culture methods means it is now possible to prime naïve T-cells from healthy donors to antigens, including drugs, which they have not previously been exposed to. Piperacillin and SMX-NO were found to prime naïve CD4+ and CD8+ T-cells from healthy donors, when the drug-derived antigens were presented in the context of autologous dendritic cells. Cloned drug-specific T-cells secreted a similar panel of cytokines to that observed with patient cells. Of particular importance was the detection of IL-22 in the absence of IL-17. The final component of the project utilized cloned T-cells with specificity for SMX-NO, piperacillin and flucloxacillin to explore mechanisms of drug-specific T-cell activation and potential cross-reactivity. Clones responsive against all 3 drugs were activated via a hapten mechanism involving (1) formation of protein adducts, (2) antigen processing and (3) presentation of derived peptides in an MHC-restricted manner. No cross-reactivity was observed with the 3 drugs. Collectively, these data showed that drug-haptens activate T-cells from patients with clinically divergent forms of hypersensitivity. T-cells secrete a similar profile of cytokines including the tissue-specific cytokine IL-22 following stimulation through the T-cell receptor. Furthermore, it is possible to prime naïve T-cells with a similar function against drugs using peripheral blood mononuclear cells from healthy donors.
Supervisor: Naisbitt, Dean Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral