Basic and clinical immunology
Toxic epidermal necrolysis: Effector cells are drug-specific cytotoxic T cells

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Background

Toxic epidermal necrolysis (TEN) is a very rare but extremely severe drug reaction characterized by widespread apoptosis of epidermis with extensive blisters. We previously found drug-specific cytotoxic CD8 T lymphocytes in the blisters of a single patient.

Objective

To confirm the role of drug specific cytotoxic lymphocytes in a larger series, to test the cytotoxicity on keratinocytes, and to look for cross-reactivity between chemically related drugs.

Methods

The phenotype of lymphocytes present in the blister fluids of 6 patients with TEN was analyzed by flow cytometry. Cytotoxic functions were tested by chromium release assay on a variety of target cells (autologous or MHC class I–matched EBV-transformed lymphocytes, autologous keratinocytes) after nonspecific (CD3 monoclonal antibody) or specific (suspected and potentially cross-reactive drugs) activation.

Results

Blister lymphocytes were CD8+HLA-DR+CLA+CD56+. In all 6 cases, they were cytotoxic after nonspecific activation. A drug-specific cytotoxicity was observed in 4 cases (3 related to cotrimoxazole and 1 to carbamazepine) toward lymphocytes. Blister cells also killed IFN-γ–activated autologous keratinocytes in the presence of drug in the 2 patients tested. Blister cells showed a strong immunoreactivity for granzyme B, and cytotoxicity was abolished by EGTA, but not by anti-Fas/CD95, suggesting perforin/granzyme–mediated killing.

By using several sulfonamides for testing the specificity of the drug T-cell receptor interaction, we observed cross-reactivity only between 4 structurally closely related medications.

Conclusion

These results strongly suggest that drug-specific, MHC class I–restricted, perforin/granzyme–mediated cytotoxicity probably has a primary role in TEN.

Section snippets

Cells from patients

The study was performed on the blister fluid lymphocytes of patients hospitalized for TEN in a specialized unit. Among 25 consecutive patients with biopsy-proven TEN, we studied all 6 with enough blister fluid cells (n = 15), a single drug suspected as responsible (8/15), and availability of a soluble form of this drug (6/8). An ethical committee (Comité Consultatif pour la Protection des Personnes se prêtant à une Recherche Biomédicale Hôpital Pitié-Salpêtrière, Paris, France) approved the

Immunophenotype of blister fluid cells

Phenotypic analysis of T lymphocytes present in blister fluids from 6 patients with TEN is presented in Table I. The majority of the lymphocytes were CD3+T-cell receptor (TCR) αβ+, with a huge predominance of the CD8+ subset. In most patients, these lymphocytes were activated as they expressed HLA-DR and had been targeted to home in the skin because they were stained by anti–cutaneous lymphocyte-associated antigen (CLA) monoclonal antibody. In 5 of the 6 patients, there was a significant

Discussion

We had previously found CD8+ T lymphocytes exhibiting drug-specific cytotoxicity in the blister fluid of a single patient with TMP-SMX–related TEN.14 These specific CTLs were MHC class I–restricted and used a perforin/granzyme–mediated pathway to exert their killing.

The current data extend these initial findings in a total of 6 patients, demonstrate that the blister fluid cells can also kill autologous keratinocytes in the presence of the drug, and suggest cross-reactivity of

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Supported in part by a grant from the French Ministry of Health (No AOM 98027) and through contracts between Institut National de la Santé Et de la Recherche Médicale and the following drug companies: Bayer, Glaxo-Wellcome, Hoechst-Marion-Roussel, Leo, Lilly, Novartis, Parke-Davis-Jouveinal, Pfizer, Rhone-Poulenc-Rorer, Sanofi-Winthrop, and Servier. Dr Nassif was the recipient of grants from Fondation pour la Recherche Médicale and Société pour l'Investigation Dermatologique Et Allergologique. Dr Moslehi received a grant from the Fondation René Touraine.

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