RESUMEN
BACKGROUND: Research into drug hypersensitivity associated with the expression of specific HLA alleles has focussed on the interaction between parent drug and the HLA with no attention given to reactive metabolites. For this reason, we have studied HLA-B*13:01-linked dapsone hypersensitivity to (a) explore whether the parent drug and/or nitroso metabolite activate T cells and (b) determine whether HLA-B*13:01 is involved in the response. METHODS: Peripheral blood mononuclear cells (PBMC) from six patients were cultured with dapsone and nitroso dapsone, and proliferative responses and IFN-γ release were measured. Dapsone- and nitroso dapsone-specific T-cell clones were generated and phenotype, function, HLA allele restriction, and cross-reactivity assessed. Dapsone intermediates were characterized by mass spectrometry. RESULTS: Peripheral blood mononuclear cells from six patients and cloned T cells proliferated and secreted Th1/2/22 cytokines when stimulated with dapsone (clones: n = 395; 80% CD4+ CXCR3hi CCR4hi , 20% CD8+CXCR3hi CCR4hi CCR6hi CCR9hi CCR10hi ) and nitroso dapsone (clones: n = 399; 78% CD4+, 22% CD8+ with same chemokine receptor profile). CD4+ and CD8+ clones were HLA class II and class I restricted, respectively, and displayed three patterns of reactivity: compound specific, weakly cross-reactive, and strongly cross-reactive. Nitroso dapsone formed dimers in culture and was reduced to dapsone, providing a rationale for the cross-reactivity. T-cell responses to nitroso dapsone were dependent on the formation of a cysteine-modified protein adduct, while dapsone interacted in a labile manner with antigen-presenting cells. CD8+ clones displayed an HLA-B*13:01-restricted pattern of activation. CONCLUSION: These studies describe the phenotype and function of dapsone- and nitroso dapsone-responsive CD4+ and CD8+ T cells from hypersensitive patients. Discovery of HLA-B*13:01-restricted CD8+ T-cell responses indicates that drugs and their reactive metabolites participate in HLA allele-linked forms of hypersensitivity.