An Updated Review of the Diagnostic Methods in Delayed Drug Hypersensitivity

Abstract

Delayed drug hypersensitivity reactions are clinically diverse reactions that vary from isolated benign skin conditions that remit quickly with no or symptomatic treatment, drug discontinuation or even continued drug treatment, to the other extreme of severe cutaneous adverse reactions (SCARs) that are associated with presumed life-long memory T-cell responses, significant acute and long-term morbidity and mortality. Diagnostic "in clinic" approaches to delayed hypersensitivity reactions have included patch testing (PT), delayed intradermal testing (IDT) and drug challenges for milder reactions. Patch and IDT are, in general, performed no sooner than 4-6 weeks after resolution of the acute reaction at the maximum non-irritating concentrations. Functional in vitro and ex vivo assays have largely remained the province of research laboratories and include lymphocyte transformation test (LTT) and cytokine release enzyme linked ImmunoSpot (ELISpot) assay, an emerging diagnostic tool which uses cytokine release, typically IFN-γ, after the patient's peripheral blood mononuclear cells are stimulated with the suspected drug(s). Genetic markers such as human leukocyte antigen have shown recent promise for both pre-prescription screening as well as pre-emptive and diagnostic testing strategies.

Keywords: HLA; T cells; delayed hypersensitivity reaction; drug allergy; enzyme linked ImmunoSpot (ELISpot); lymphocyte transformation test (LTT); severe cutaneous adverse reactions; skin testing.

FIGURE 1

T-cell mediated delayed hypersensitivity mechanistic hypotheses. Three non-mutually exclusive hypotheses have been described to clarify drug triggered T-cell activation: (A) The hapten/prohapten model describes how an antigen (drug) that covalently binds to a self-peptide is intracellularly processed and then presented by MHC to T cells. (B) The p-i concept (pharmacological interaction with immune receptor) is based upon non-covalent binding of antigens to HLA or TCR without immune processing. (C) The altered repertoire model indicates that drugs can occupy positions in the peptide binding groove of the MHC, altering the binding cleft and the specificity of MHC binding. HLA, human leukocyte antigens; MHC, major histocompatibility complex; TCR, T-cell receptor.

FIGURE 2

Diagnostic approach to delayed hypersensitivity reactions. 1) Clinical history and identification of possible causal drugs; 2) Patch testing; 3) Intradermal testing reading after 24–48 h; 4) Ex-vivo testing (if available): ELISpot, LTT, HLA allele testing; 5) Single or prolonged drug challenge if conclusive results. ELISpot, enzyme linked ImmunoSpot; HLA, human leukocyte antigens; LTT, lymphocyte transformation test.

FIGURE 3

Detection of cytokine secretion using ELISpot. Cytokine-specific coating antibody is added and incubated overnight at 4°C. The plate is washed and PBMCs and drug(s) are added and incubated for 18–20 h at 37°C. The following day, the plate is washed again and biotin-conjugated detection antibody is added and incubated for 2 h at room temperature. After another wash, streptavidin-bound enzyme is added and incubated for 1 h at room temperature. After the last wash, substrate is added (BCIP-NBT or TMB). Spot development is monitored for ∼15 min (in dark). The plate is washed and left to dry overnight for a final reading on the fourth day. BCIP-NBT, 5-bromo-4-chloro-3-indolyl phosphate (BCIP)/nitro blue tetrazolium (NBT); ELISpot, enzyme linked ImmunoSpot; PBMC, peripheral blood mononuclear cells; TMB, tetramethyl benzidine.

FIGURE 4

Global epidemiology of severe cutaneous adverse drug reactions.