Insulin autoantibody polymorphisms with greater discrimination for diabetes in humans

Abstract

Summary.Insulin autoantibodies, like islet cell antibodies, are found not only in the sera of newly diagnosed Type 1 (insulin-dependent) diabetic patients and their relatives, but also in patients with other autoimmunities who do not develop diabetes. Insulin autoantibodies are oligo/monoclonal and frequently binding-site restricted. As determinant selection is genetically determined, we questioned whether certain polymorphisms of insulin autoantibodies, identified by their binding site on the insulin molecule, could better discriminate for Type 1 diabetes, which is also HLA determined. First, we raised monoclonal antibodies to human insulin by classic fusion methods in order to determine the range of antibody polymorphism, and identified five distinct types by their binding profiles to a panel of insulin variants, using an enzyme-linked immunosorbent assay. Two of these polymorphisms, type A and type B, were subsequently found in insulin autoantibody positive human sera using the same panel of insulin variants, and successfully distinguished diabetes-related from diabetes-unrelated individuals. Thus, the type B polymorphism was responsible for binding in 60% of 41 insulin autoantibody positive individuals with polyautoimmune disease but no personal or family history of diabetes (diabetes unrelated), but in only 2% of a group which comprised 17newly-diagnosed insulin autoantibody positive Type 1 diabetic patients, 19 insulin autoantibody positive discordant twins of Type 1 diabetes and six insulin autoantibody positive healthy siblings of Type I diabetic patients (diabetes related) (p < 0.01). Isolation of the type A polymorphism alone reduced the proportion of false negatives in the insulin autoantibody test for diabetes relatedness from 49% to 20% without diminishing its specificity. Thus, insulin autoantibody polymorphisms are more discriminating than the 'nominal' antibody, due possibly to linkage between immune response genes determining response to the type A epitope on the one hand, and susceptibility to Type 1 diabetes on the other.

Relationship of the 37,000- and 40,000-M(r) tryptic fragments of islet antigens in insulin-dependent diabetes to the protein tyrosine phosphatase-like molecule IA-2 (ICA512).

“…Neither GAD nor insulin have been identified by the library screening approach. Diabetes-associated antibodies to these antigens are known to bind epitopes that are highly dependent on protein conformation (26,27), and these may not be displayed by the library. If the 40,000-M, fragments of islet antigen are indeed equivalent to IA-2 and ICA512, then this protein will represent a rare case of an islet antigen identified by both immunochemical and molecular biological approaches.…”

Mature high-affinity immune responses to (pro)insulin anticipate the autoimmune cascade that leads to type 1 diabetes

“…These were not the same as the polyreactive antibodies previously described, since, unlike the polyreactive antibodies, they could not be inhibited by excess tetanus toxoid or single-stranded DNA (data not shown). IAAs with distinct binding characteristics (16), proinsulin autoantibodies (17), and high-titer, low-affinity IAAs in subjects with the rare insulin autoimmune hypoglycemia (18, 19) have also been described. The findings of this study are relevant to both pathogenesis and prediction of T1DM.…”

Developments in the prediction of type 1 diabetes mellitus, with special reference to insulin autoantibodies

“…However, the dynamics and diversity of the autoimmune repertoire can already be demonstrated in subjects with a lone islet-cell autoantibody [145,148,150]. For example, binding studies of IAA-positive sera with insulin variants have shown epitope restriction, allowing the differentiation of diabetes-related IAA from diabetes-unrelated IAA [83]. Another recent study showed that the autoimmune response to GAD in children is initially directed against the middle region of GAD, while later spreading mostly to the NH 2 -terminal epitope [150].…”