Jan Paul Heribert Depta

T-cell involvement in drug-induced acute generalized exanthematous pustulosis

Acute generalized exanthematous pustulosis (AGEP) is an uncommon eruption most often provoked by drugs, by acute infections with enteroviruses, or by mercury. It is characterized by acute, extensive formation of nonfollicular sterile pustules on erythematous background, fever, and peripheral blood leukocytosis. We present clinical and immunological data on four patients with this disease, which is caused by different drugs. An involvement of T cells could be implied by positive skin patch tests and lymphocyte transformation tests. Immunohistochemistry revealed a massive cell infiltrate consisting of neutrophils in pustules and T cells in the dermis and epidermis. Expression of the potent neutrophil-attracting chemokine IL-8 was elevated in keratinocytes and infiltrating mononuclear cells. Drug-specific T cells were generated from the blood and skin of three patients, and phenotypic characterization showed a heterogeneous distribution of CD4/CD8 phenotype and of T-cell receptor Vbeta-expression. Analysis of cytokine/chemokine profiles revealed that IL-8 is produced significantly more by drug-specific T cells from patients with AGEP compared with drug-specific T cells from patients that had non-AGEP exanthemas. In conclusion, our data demonstrate the involvement of drug-specific T cells in the pathomechanism of this rather rare and peculiar form of drug allergy. In addition, they indicate that even in some neutrophil-rich inflammatory responses specific T cells are engaged and might orchestrate the immune reaction.

Cellular and molecular pathophysiology of cutaneous drug reactions.

Hypersensitivity reactions to drugs can cause a variety of skin diseases like maculopapular, bullous and pustular eruptions. In recent years increasing evidence indicates the important role of T cells in these drug-induced skin diseases. Analysis of such drug-specific T cell clones has revealed that drugs can be recognized by αβ-T cell receptors, not only if bound covalently to peptides, but also if the drug binds in a rather labile way to the presenting major histocompatibility complex (MHC)-peptide. This presentation is sufficient to stimulate T cells.In maculopapular exanthema (MPE), histopathological analysis typically shows a dominant T cell infiltration together with a vacuolar interface dermatitis. Immunohistochemical studies demonstrate the presence of cytotoxic CD4+ and to a lesser degree of CD8+ T cells, which contain perforin and granzyme B. They are close to keratinocytes that show signs of cell destruction. Expression of Fas ligand is barely detectable, suggesting that cytotoxic granule exocytosis may be the dominant pathway leading to keratinocyte cell damage. While in MPE, the killing of cells seems to be predominately mediated by CD4+ T cells, patients with bullous skin disease show a strong CD8+ T cell migration to the epidermis. This is probably due to a preferential presentation of the drug by MHC class I molecules, and a more extensive killing of cells that present drugs on MHC class I molecules. This might lead to bullous skin diseases.In addition to the presence of cytotoxic T cells, drug-specific T cells also orchestrate the inflammatory skin reaction through the release and induction of various cytokines [i.e. interleukin (IL)-5, IL-6, tumor necrosis factor-α and interferon-γ] and chemokines (RANTES, eotaxin or IL-8). The increased expression of these mediators seems to contribute to the generation of tissue and blood eosinophilia, a hallmark of many drug-induced allergic reactions. However, in acute generalized exanthematous pustulosis (a peculiar form of drug allergy), neutrophils represent the predominant cell type within pustules, probably due to their recruitment by IL-8 secreting drug specific T cells and keratinocytes.

Influence of reduced glutathione on the proliferative response of sulfamethoxazole-specific and sulfamethoxazole-metabolite-specific human CD4+ T-cells

Hypersensitivity to the drug sulfamethoxazole (SMX) is thought to be a consequence of bioactivation to the hydroxylamine metabolite (SMX‐NHOH) and further oxidation to the ultimate reactive metabolite, nitroso‐sulfamethoxazole (SMX‐NO). SMX‐NO covalently modifies self proteins which in turn might be recognized as neo‐antigens by T‐cells. The antioxidant glutathione (GSH) is known to protect cells from reactive metabolites by conjugation and subsequent dissociation to SMX‐NHOH and/or SMX. To study the reactivity of T‐cells to SMX metabolites and their respective role in the generation of drug‐specific T‐cells, we analysed the effect of GSH on the response of PBMC to SMX and its metabolites SMX‐NHOH and SMX‐NO. Furthermore, we monitored the proliferative response of drug‐specific T‐cell clones in the presence or absence of GSH. We found that addition of GSH to peripheral blood mononuclear cells had no effect on the SMX‐specific response but enhanced the proliferation to SMX‐metabolites. The response of SMX‐NO‐specific T‐cell clones was abrogated when GSH was present during the covalent haptenation of antigen presenting cells (APC). Conversely, SMX‐specific T‐cell clones gained reactivity through the conversion of SMX‐NO to the parent drug by GSH. While GSH had no effect on the initial activation of T‐cell clones, it prevented covalent binding to APCs, reduced toxicity and thereby led to proliferation of drug‐specific T‐cells to non‐reactive drug metabolites. Our data support the concept that in allergic individuals T‐cells recognize the non‐covalently bound parent drug rather than APC covalently modified by SMX‐NO.

T cell-mediated hypersensitivity to quinolones : mechanisms and cross-reactivity

Background Quinolones are widely used, broad spectrum antibiotics that can induce immediate‐ and delayed‐type hypersensitivity reactions, presumably either IgE or T cell mediated, in about 2–3% of treated patients.

Non-covalent presentation of sulfamethoxazole to human CD4+ T cells is independent of distinct human leucocyte antigen-bound peptides

Background It has been shown that drugs comprise a group of non‐peptide antigens that can be recognized by human T cells in the context of HLA class II and that this recognition is involved in allergic reactions. Recent studies have demonstrated a MHC‐restricted but processing‐ and metabolism‐independent pathway for the presentation of allergenic drugs such as lidocaine and sulfamethoxazole (SMX) to drug‐specific T cells. However, there is little information so far on the precise molecular mechanisms of this non‐covalent drug presentation.

Transfection of drug-specific T-cell receptors into hybridoma cells: tools to monitor drug interaction with T-cell receptors and evaluate cross-reactivity to related compounds.

In the context of drug hypersensitivity, our group has recently proposed a new model based on the structural features of drugs (pharmacological interaction with immune receptors; p-i concept) to explain their recognition by T cells. According to this concept, even chemically inert drugs can stimulate T cells because certain drugs interact in a direct way with T-cell receptors (TCR) and possibly major histocompatibility complex molecules without the need for metabolism and covalent binding to a carrier. In this study, we investigated whether mouse T-cell hybridomas transfected with drug-specific human TCR can be used as an alternative to drug-specific T-cell clones (TCC). Indeed, they behaved like TCC and, in accordance with the p-i concept, the TCR recognize their specific drugs in a direct, processing-independent, and dose-dependent way. The presence of antigen-presenting cells was a prerequisite for interleukin-2 production by the TCR-transfected cells. The analysis of cross-reactivity confirmed the fine specificity of the TCR and also showed that TCR transfectants might provide a tool to evaluate the potential of new drugs to cause hypersensitivity due to cross-reactivity. Recombining the α- and β-chains of sulfanilamide- and quinolone-specific TCR abrogated drug reactivity, suggesting that both original α- and β-chains were involved in drug binding. The TCR-transfected hybridoma system showed that the recognition of two important classes of drugs (sulfanilamides and quinolones) by TCR occured according to the p-i concept and provides an interesting tool to study drug-TCR interactions and their biological consequences and to evaluate the cross-reactivity potential of new drugs of the same class.