HLA-DR BINDING ANALYSIS OF PEPTIDES FROM ISLET ANTIGENS IN IDDM

HLA molecules are essential for thymic education and HLA restriction o f T-cell responses. We therefore analyzed the HLA-DR binding affinitie s of synthetic peptides covering the entire sequences of GAD65, islet cell antigen 69 (ICA69), and (pro)insulin, which are candidate antigen s in the autoimmune process of T-cell-mediated destruction of the panc reatic beta-cells. Subsequently, peptide HLA-DR binding was correlated to peptide antigenicity by comparing known T-cell epitopes with their HLA-binding affinities defined in this study. The results demonstrate the following. 1) (Pro)insulin peptides display a strong binding affi nity for HLA-DR2, which is associated with negative genetic predisposi tion to IDDM, whereas poor binding was observed for HLA-DR molecules n eutrally or positively associated with IDDM. This suggests that the ab sence of insulin-reactive T-cells in DR2+ individuals may be explained by negative selection on high-affinity DR2 binding insulin peptides. 2) Most autoantigenic peptides display promiscuous H-LA-DR binding pat terns. This promiscuity in itself is not sufficient to explain the gen etic association of HLA-DR with development of IDDM. 3) HLA-DR3 bindin g of autoantigenic GAD65 peptides is relatively weak compared with tha t of other known T-cell epitopes. 4) All peptide epitopes recognized b y HLA-DR-restricted T-cells from either IDDM patients or GAD65-immuniz ed HLA-DR transgenic mice bind with high affinity to their HLA-DR rest riction molecule (P < 0.0006). In contrast, T-cell epitopes recognized by nondiabetic controls bind DR molecules with weak or undetectable a ffinity. These results thus indicate a strong correlation between anti genicity and HLA-DR binding affinity of GAD65 peptides in IDDM. Furthe rmore, negative thymic selection of insulin peptides in low-risk (HLA- DR2 expressing) subjects may explain the lack of autoreactivity to ins ulin in such individuals.