title: In vitro drug causality assessment in Stevens-Johnson syndrome - alternatives for lymphocyte transformation test.
abstract: 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 IFNγ 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 IFNγ. 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
  Granulysin expression in CD3(+) CD4(+) , Granzyme B-ELISpot and IFNγ 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.
authors: ADR-AC GmbH, Bern, Switzerland.
authors: G
authors: G
authors: Porebski
authors: T
authors: T
authors: Pecaric-Petkovic
authors: M
authors: M
authors: Groux-Keller
authors: M
authors: M
authors: Bosak
authors: T T
authors: TT
authors: Kawabata
authors: W J
authors: WJ
authors: Pichler
mesh_terms: D000328:Adult; D000368:Aged; D000945:Antigens, Differentiation, T-Lymphocyte / Q000096:biosynthesis; D016207:Cytokines / Q000378:metabolism / Q000494:pharmacology; D064420:Drug-Related Side Effects and Adverse Reactions / Q000276:immunology; D005260:Female; D053804:Granzymes / Q000378:metabolism; D006801:Humans; D019409:Interleukin-15 / Q000494:pharmacology; D015851:Interleukin-7 / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology*; D008214:Lymphocytes / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D012372:ROC Curve; D013262:Stevens-Johnson Syndrome / Q000175:diagnosis / Q000209:etiology* / Q000276:immunology; D055815:Young Adult
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D016207:Cytokines; C075005:GNLY protein, human; D019409:Interleukin-15; D015851:Interleukin-7; D053804:Granzymes
keywords: Stevens-Johnson syndrome; cytokines; granulysin; granzyme B; lymophocyte transformation test; toxic epidermal necrolysis