The Cyto-LTT: A multiplex cytokine assay to detect and assess the strength of T cell reactivity in drug hypersensitivity.

Background The conventional in vitro lymphocyte transformation test (LTT) for diagnosing drug hypersensitivity reactions (DHRs) is limited by low sensitivity. To improve detection and assess T cell activation strength, we adapted this test to a cytokine-based assay (Cyto-LTT) by measuring IL-5, IL-13, IFN-γ, granzyme B and granulysin secretion. Methods We retrospectively analyzed 851 positive Cyto-LTT results (from a total 6058 Cyto-LTT) from a Swiss drug hypersensitivity diagnostic laboratory's database, including 97 patients with Drug Rash with Eosinophilia and Systemic Symptoms (DReSS) and 754 patients with exanthems, including urticarial, maculo- and maculopapular-rash. Cytokine responses measured across three drug concentrations of amoxicillin (n = 734), aromatic sulfonamides (n = 81), or vancomycin (n = 36) were evaluated for dose-dependency and the correlation to the clinical manifestations. A grading system defined strong responses as cytokine stimulation indices (SI) above the 75th percentile (per cytokine) in the exanthem group which served as a comparator for the DReSS cases. Results Cytokine responses increased dose-dependently. DReSS cases exhibited significantly stronger cytokine secretions compared to exanthem cases (p < 0.001), with strong responses observed in 62.9 % of DReSS patients versus 33.6 % of exanthem cases. Conclusions Cyto-LTT reveals dose-dependent T cell activation in delayed drug hypersensitivity reactions, challenging the notion that drug allergic reactions are dose-independent. The assay not only identifies the culprit drug but also quantifies immune activation strength. Stronger responses were more frequent in DReSS, suggesting the test may also inform risk assessment-particularly in guiding caution with structurally related compounds or in patients at risk for severe or recurrent reactions.