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In vitro drug causality assessment in Stevens-Johnson syndrome - alternatives for lymphocyte transformation test.
Porebski, G. 1,2,3; Pecaric-Petkovic, T. 1,2; Groux-Keller, M. 1,2; Bosak, M. 4; Kawabata, T. T. 5; Pichler, W. J. 1,2
[Article] Clinical & Experimental Allergy. 43(9):1027-1037, September 2013.

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Background: Patients with Stevens-Johnson syndrome (SJS) or toxic epidermal necrolysis (TEN) are often exposed simultaneously to a few potentially culprit drugs. However, both the standard lymphocyte transformation tests (LTT) with proliferation as the assay end-point as well as skin tests, if done, are often negative.

Objective: As provocation tests are considered too dangerous, there is an urgent need to identify the relevant drug in SJS/TEN and to improve sensitivity of tests able to identify the causative drug.

Methods: Fifteen patients with SJS/TEN with the ALDEN score >= 6 and 18 drug-exposed controls were included. Peripheral blood mononuclear cells (PBMC) were isolated and cultured under defined conditions with drugs. LTT was compared to the following end-points: cytokine levels in cell culture supernatant, number of granzyme B secreting cells by ELISpot and intracellular staining for granulysin and IFN[gamma] in CD3 CD4 , CD3 CD8 and NKp46 cells. To further enhance sensitivity, the effect of IL-7/IL-15 pre-incubation of PBMC was evaluated.

Results: Lymphocyte transformation tests was positive in only 4/15 patients (sensitivity 27%, CI: 8-55%). Similarly, with granzyme B-ELISpot culprit drugs were positive in 5/15 patients (sensitivity 33%, CI: 12-62%). The expression of granulysin was significantly induced in NKp46 and CD3 CD4 cells (sensitivity 40%, CI: 16-68% and 53%, CI: 27-79% respectively). Cytokine production could be demonstrated in 38%, CI: 14-68% and 43%, CI: 18-71% of patients for IL-2 and IL-5, respectively, and in 55%, CI: 23-83% for IFN[gamma]. Pre-incubation with IL-7/IL-15 enhanced drug-specific response only in a few patients. Specificities of tested assays were in the range of 95 (CI: 80-99%)-100% (CI: 90-100%).

Conclusions and Clinical Relevance: Granulysin expression in CD3 CD4 , Granzyme B-ELISpot and IFN[gamma] production considered together provided a sensitivity of 80% (CI: 52-96%) and specificity of 95% (80-99%). Thus, this study demonstrated that combining different assays may be a feasible approach to identify the causative drug of SJS/TEN reactions; however, confirmation on another group of patients is necessary.