Use of a High-Density Protein Microarray to Identify Autoantibodies in Subjects with Type 2 Diabetes Mellitus and an HLA Background Associated with Reduced Insulin Secretion

Abstract

New biomarkers for type 2 diabetes mellitus (T2DM) may aid diagnosis, drug development or clinical treatment. Evidence is increasing for the adaptive immune system’s role in T2DM and suggests the presence of unidentified autoantibodies. While high-density protein microarrays have emerged as a useful technology to identify possible novel autoantigens in autoimmune diseases, its application in T2DM has lagged. In Pima Indians, the HLA haplotype (HLA-DRB1*02) is protective against T2DM and, when studied when they have normal glucose tolerance, subjects with this HLA haplotype have higher insulin secretion compared to those without the protective haplotype. Possible autoantibody biomarkers were identified using microarrays containing 9480 proteins in plasma from Pima Indians with T2DM without the protective haplotype (n = 7) compared with those with normal glucose regulation (NGR) with the protective haplotype (n = 11). A subsequent validation phase involving 45 cases and 45 controls, matched by age, sex and specimen storage time, evaluated 77 proteins. Eleven autoantigens had higher antibody signals among T2DM subjects with the lower insulin-secretion HLA background compared with NGR subjects with the higher insulin-secretion HLA background (p<0.05, adjusted for multiple comparisons). PPARG2 and UBE2M had lowest p-values (adjusted p = 0.023) while PPARG2 and RGS17 had highest case-to-control antibody signal ratios (1.7). A multi-protein classifier involving the 11 autoantigens had sensitivity, specificity, and area under the receiver operating characteristics curve of 0.73, 0.80, and 0.83 (95% CI 0.74–0.91, p = 3.4x10-8), respectively. This study identified 11 novel autoantigens which were associated with T2DM and an HLA background associated with reduced insulin secretion. While further studies are needed to distinguish whether these antibodies are associated with insulin secretion via the HLA background, T2DM more broadly, or a combination of the two, this study may aid the search for autoantibody biomarkers by narrowing the list of protein targets.

Autoantibodies against PFDN2 are associated with an increased risk of type 2 diabetes: A case‐control study

“…The reactivity of plasma samples to 14 target antigens ( were selected since they were significantly associated with participants with T2DM who lacked the protective HLA haplotype based on our prior study, 24 and the remaining 3 target proteins (LXR-alpha, interleukin 1 beta [IL1B], and IAPP) were marginally associated in the pilot study but were known to be biologically relevant to obesity and T2DM based on the literature. [27][28][29] Test plasma was previously stored at −70°C.…”

“…24 On the basis of specified statistical criteria or possible role in underlying biology of type diabetes, we then tested 70 of these proteins in a second confirmatory study in a moderately larger group (n = 90) identifying 14 as promising biomarkers. 24 However, in both initial exploratory studies, to maximize the potential difference between groups, we selected individuals with T2DM and without the protective SNP that tags the HLA-DRB1*16:02 haplotype for comparison with those with normal glucose regulation (NGR) and the presence of the protective haplotype. 24 Thus, prior studies were limited by small sample sizes and by the fact that it could not ascertained whether the identified autoantibodies were associated with T2DM, the protective SNP, or a combination.…”

“…24 However, in both initial exploratory studies, to maximize the potential difference between groups, we selected individuals with T2DM and without the protective SNP that tags the HLA-DRB1*16:02 haplotype for comparison with those with normal glucose regulation (NGR) and the presence of the protective haplotype. 24 Thus, prior studies were limited by small sample sizes and by the fact that it could not ascertained whether the identified autoantibodies were associated with T2DM, the protective SNP, or a combination. To further evaluate these autoantibodies, we tested the hypothesis that autoantibodies identified in our prior studies were associated withT2DM, independent of the protective SNP.…”

Key genes and co‐expression network analysis in the livers of type 2 diabetes patients

“…The ubiquitin‐proteasome system was reported to have a role in type 2 diabetes. UBE2M antibody had a higher signal in type 2 diabetes patients’ plasma with the lower insulin‐secretion human leukocyte antigen background. We observed that UBE2M was increased in two hepatic insulin resistance models, indicating that UBE2M possibly plays a key role in type 2 diabetes hepatic insulin resistance.…”

A genomic study on distribution of human leukocyte antigen ( HLA ) -A and HLA-B alleles in Lak population of Iran

“…Clinically, being acquainted with HLA distribution is a sine qua non for bone marrow donating centers [16], [17], forensic medicine [18], studies of HLA related disease such as type 2 diabetes [19], [20], [21] or multiple sclerosis [22], designing peptide vaccine and monoclonal antibodies against tumors [23], [24], infectious agents [25], [26], [27] and autoimmune disease [20], [28], as well as infertility treatment and assisted reproductive technologies [29], [30], [31].…”