The Use of Protein Microarrays to study Autoimmunity and Diabetes The Use of Protein Microarrays to study Autoimmunity and Diabetes Background - Association between islet cell autoantibodies (AA's) and the progression to Type 1 diabetes (T1D) has been well recognized. Although detection rates in individuals possessing all three commonly tested AA's (anti-GAD, -IA-2, -insulin) are thought to be >80%, there exist clinically presenting individuals which fail to present any of these biomarkers at the time of diagnosis. In light of this, it is likely that additional AA's specific for T1D exist. High-content, multiplexed, array-based proteomics solutions are a valuable, unbiased discovery tool that can facilitate the identification of novel AA's. Further, an expanded panel of biomarkers is likely to increase detection rates by combining sensitivity and specificity of a number of AA's and may ultimately prove to be of clinical value as a diagnostic tool. Methods - Thus, we undertook a serological screening effort against a clone library encoding >6000 human proteins using the Nucleic Acid Programmable Protein Array (NAPPA). Employing such a broad set of potential molecular targets our aims were 1) to identify, through multiple confirmatory rounds of screening, an enhanced panel of serum reactive antigens to which AA's associated with T1D can be detected and 2) to test the ability of this panel of antigens to accurately discriminate between those with and without diabetes. To this end, we have completed 1) a pre-screen aimed at eliminating uninformative antigens (displaying no differential AA reactivity between T1D+ (n=50) and T1D- (n=20) serum) and 2) a training screen in which the reproducibility of statistically significant, differentially recognized autoantigens between patient and controls observed in the pre-screen was verified/reproduced in an independent serum set (n=75 T1D+/-). Results - From these efforts we have identified >100 putative autoantigens (including the recently reported ZnT8) that exhibit enhanced AA reactivity with serum from T1D+ patients versus controls (p<0.01). Of these putative targets, a subset of approximately 10 antigens exhibit diagnostic sensitivities in the range of 50%, in addition to specificities between 70 and 80% based upon microarray data. With this panel we are developing an optimized diagnostic algorithm or classifier (using support vector machine analysis) and plan to test performance under blinded conditions. Ultimately, it is our aim to fully validate these biomarkers by orthogonal means and to make use of the combined sensitivities and specificities of this collective panel of potential autoantigens in array format in order to improve diagnostic performance for clinical application. Conclusion - This study represents a significant advance toward comprehensive, unbiased identification of novel AA's associated with T1D and the demonstration of array-based immunodiagnostics based upon these newly identified autoantigens.
|Effective start/end date||8/1/09 → 8/31/10|
- Juvenile Diabetes Research Foundation International (JDRF): $544,659.00
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