loading page

Ex vivo diagnostics using varied cellular inputs in drug-induced severe cutaneous adverse reactions
  • +11
  • Andrew Awad,
  • Effie Mouhtouris,
  • Catriona Vi Nguyen-Robertson,
  • Natasha Holmes,
  • Kyra Chua,
  • Ana Copaescu,
  • Fiona James,
  • Michelle Goh S,
  • Ar K. Aung,
  • Dale Godfrey,
  • Elizabeth Philips,
  • Andrew Gibson,
  • Catarina Almeida F,
  • Jason Trubiano
Andrew Awad
Austin Health

Corresponding Author:[email protected]

Author Profile
Effie Mouhtouris
Austin Health
Author Profile
Catriona Vi Nguyen-Robertson
The Peter Doherty Institute for Infection and Immunity
Author Profile
Natasha Holmes
Austin Health
Author Profile
Kyra Chua
Austin Health
Author Profile
Ana Copaescu
Austin Health
Author Profile
Fiona James
Austin Health
Author Profile
Michelle Goh S
Alfred Health
Author Profile
Ar K. Aung
Alfred Health
Author Profile
Dale Godfrey
The Peter Doherty Institute for Infection and Immunity
Author Profile
Elizabeth Philips
Murdoch University
Author Profile
Andrew Gibson
Murdoch University
Author Profile
Catarina Almeida F
The Peter Doherty Institute for Infection and Immunity
Author Profile
Jason Trubiano
Austin Health
Author Profile

Abstract

Background Drug-induced severe cutaneous adverse reactions (SCARs) are presumed T-cell-mediated hypersensitivities associated with significant morbidity and mortality. Traditional in-vivo testing methods, such as patch or intradermal testing, are limited by a lack of standardisation and poor sensitivity. Modern approaches to testing include measurement of IFN-γ release from patient peripheral blood mononuclear cells (PBMC) stimulated with the suspected causative drug. Objective We sought to improve ex-vivo diagnostics for drug-induced SCAR by comparing enzyme-linked immunospot (ELISpot) sensitivities and flow cytometry-based intracellular cytokine staining (ICS) and cellular composition of separate samples (PBMC or blister fluid cells (BFC)) from the same donor. Methods IFN-γ release ELISpot and flow cytometry analyses were performed on donor-matched PBMC and BFC samples from four SCAR patients with distinct drug-allergies. Results Immune responses to suspected drugs were detected in both PBMC and BFC samples of two donors (Case 1 in response to ceftriaxone and Case 4 to oxypurinol), with BFC eliciting stronger responses. For two other donors, only BFC samples showed a response to meloxicam(Case 2) or sulfamethoxazole and its 4-Nitro metabolite (Case 3). Consistently, flow cytometry revealed a greater proportion of IFN-γ-secreting cells in the BFC compared to PBMC. BFC cells from Case 3 were also enriched for memory/activation/tissue-recruitment markers over PBMC. Conclusion Analysis of BFC samples for drug-allergy diagnostics offers a higher sensitivity for detecting positive responses compared to PBMC. This is consistent with recruitment (and enrichment) of cytokine-secreting cells with a memory/activated phenotype into blisters.