Low HLA binding of diabetes-associated CD8+ T-cell epitopes is increased by post translational modifications

Abstract

Background: Type 1 diabetes (T1D) is thought to be an autoimmune disease driven by anti-islet antigen responses and mediated by T-cells. Recent published data suggests that T-cell reactivity to modified peptides, effectively neoantigens, may promote T1D. These findings have given more credence to the concept that T1D may not be solely an error of immune recognition but may be propagated by errors in protein processing or in modifications to endogenous peptides occurring as result of hyperglycemia, endoplasmic reticulum (ER) stress, or general beta cell dysfunction. In the current study, we hypothesized that diabetes-associated epitopes bound human leukocyte antigen (HLA) class I poorly and that post-translational modifications (PTM) to key sequences within the insulin-B chain enhanced peptide binding to HLA class I, conferring the CD8+ T-cell reactivity associated with T1D.

Results: We first identified, through the Immune Epitope Database (IEDB; www.iedb.org ), 138 published HLA class I-restricted diabetes-associated epitopes reported to elicit positive T-cell responses in humans. The peptide binding affinity for their respective restricting allele(s) was evaluated in vitro. Overall, 75% of the epitopes bound with a half maximal inhibitory concentration (IC50) of 8250 nM or better, establishing a reference affinity threshold for HLA class I-restricted diabetes epitopes. These studies demonstrated that epitopes from diabetes-associated antigens bound HLA with a lower affinity than those of microbial origin (binding threshold of 500 nM for 85% of the epitopes). Further predictions suggested that diabetes epitopes also bind HLA class I with lower affinity than epitopes associated with other autoimmune diseases. Therefore, we measured the effect of common PTM (citrullination, chlorination, deamidation, and oxidation) on HLA-A*02:01 binding of insulin-B-derived peptides, compared to native peptides. We found that these modifications increased binding for 44% of the insulin-B epitopes, but only 15% of the control peptides.

Conclusions: These results demonstrate that insulin-derived epitopes, commonly associated with T1D, generally bind HLA class I poorly, but can be subject to PTM that improve their binding capacity and may, in part, be responsible for T-cell activation in T1D and subsequent beta cell death.

Keywords: Class I MHC; Diabetes; HLA-peptide binding affinity; Insulin; T cell epitopes.

Fig. 1

HLA class I binding capacity of diabetes-associated CTL epitopes: A cumulative percentage of diabetes associated epitopes is plotted as a function of the binding capacity of the epitopes assayed for their corresponding HLA class I restricting allele. As shown, about 40% of the epitopes bound with an affinity of 500 nM (red dashed line), or better, and 75% bound at the 8250 nM level (blue dashed line)

Fig. 2

Binding capacity of various types of class I epitopes: The predicted binding affinity of diabetes-associated epitopes (red line) is compared with the predicted binding capacity of virus derived epitopes (blue line), and epitopes associated with other autoimmune diseases (green line). The 500 nM threshold (dashed red line) identifies 60%, 70%, and 78% of the respective epitopes