D2.21 - The Cyto-LTT: A multiplex cytokine assay to detect and assess the strength of T cell reactivity in drug hypersensitivity

The conventional in vitro lymphocyte transformation test (LTT) for diagnosing drug hypersensitivity reactions (DHRs) is limited by low sensitivity and provide little information on T cell activation magnitude. We developed a cytokine-based LTT (Cyto-LTT) measuring IL-5, IL-13, IFN-γ, granzyme B, and granulysin, to improve culprit drug detection and quantify immune activation.

We retrospectively analyzed 851 positive Cyto-LTT results from 6058 tests in a Swiss drug hypersensitivity reaction diagnostic laboratory. Patients included 97 with Drug Rash with Eosinophilia and Systemic Symptoms (DReSS) and 754 with exanthems (urticarial, macular, or maculopapular rashes). Cytokine responses to three concentrations of amoxicillin (n=734), aromatic sulfonamides (n=81), orvancomycin (n=36) were assessed for dose-dependency and correlation with clinical phenotype. Strong responses were defined as stimulation indices (SI) above the 75th percentile for each cytokine in the exanthem cohort.

Cytokine secretion increased dose-dependently for all tested drugs. DReSS patients exhibited significantly stronger cytokine responses than those with exanthems (p<0.001), with strong responses observed in 62.9% of DReSS versus 33.6% of exanthem cases.

Cyto-LTT not only identifies culprit drugs but also quantifies T cell activation, challenging the notion that delayed DHRs are dose-independent. Stronger cytokine responses were more frequent in DReSS, highlighting the assay’s potential to inform risk assessment and guide caution when prescribing structurally related drugs or managing patients at risk of severe or recurrent reactions.