CD8+ T-Cell Responses Identify β-Cell Autoimmunity in Human Type 1 Diabetes

CD8 + T-Cell Responses Identify β-Cell Autoimmunity in Human Type 1 Diabetes Roberto Mallone 1 2 , Emanuela Martinuzzi 1 2 , Philippe Blancou 3 4 , Giulia Novelli 5 , Georgia Afonso 1 2 , Manuel Dolz , Graziella Bruno 5 , Lucy Chaillous , Lucienne Chatenoud 1 2 , Jean-Marie Bach 3 4 and Peter van Endert 1 2 1 INSERM, U580, Paris, France 2 Université Paris Descartes, Faculté de Médecine René Descartes, Paris, France 3 INRA, Immuno-Endocrinology Unit, ENVN, Nantes, France; the Université de Nantes, Nantes, France 4 Università di Torino, Department of Internal Medicine, Torino, Italy 5 CHU de Nantes, Hôpital Hôtel-Dieu, Clinique d’Endocrinologie, Nantes, France Address correspondence and reprint requests to Roberto Mallone or Peter van Endert, INSERM U580, Hôpital Necker, 161 rue de Sèvres, 75743 Paris Cedex 15, France. E-mail: mallone{at}necker.fr or vanendert{at}necker.fr Abstract Despite the understanding that type 1 diabetes pathogenesis is mediated by T-cells, detection of these rare lymphocytes remains largely elusive. Suitable T-cell assays are highly needed, since they could offer preclinical diagnoses and immune surrogate end points for clinical trials. Although CD4 + T-cell assays have met with limited success, CD8 + T-cells are increasingly recognized as key actors in the diabetes of the NOD mouse. CD8 + T-cells are likely to play a role also in humans and may provide new markers of β-cell autoimmunity. Taking advantage of a panel of HLA-A2–restricted β-cell epitopes derived from preproinsulin, GAD, and islet glucose-6-phosphatase catalytic subunit-related protein (IGRP), we have implemented an islet-specific CD8 + T-cell interferon-γ enzyme-linked immunospot (ISL8Spot) assay. The ISL8Spot assay is capable of detecting and quantifying β-cell–reactive CD8 + T-cells directly ex vivo, without any preliminary expansion, using either fresh or frozen samples. Positive ISL8Spot responses separate new-onset diabetic and healthy samples with high accuracy (86% sensitivity, 91% specificity), using as few as five immunodominant epitopes. Moreover, sensitivity reaches 100% when the ISL8Spot assay is complemented by antibody determinations. Combination of CD8 + T-cell measurements with immune intervention strategies may open new avenues toward type 1 diabetes prediction and prevention. Ab, antibody ELISpot, enzyme-linked immunospot IFN, interferon IGRP, islet glucose-6-phosphatase catalytic subunit–related protein ISL8Spot, islet-specific CD8+ T-cell interferon