Glycans are a critical recognition element in cell-cell interactions, signaling, and intrinsic/extrinsic stimulation of the immune system among other process. Improper regulation of glycosylation pathways resulting from genetic disorders, over production of protein, or steric hindrance of proper checkpoint pathways can result in atypical glycan display. Irregularities in the O-linked glycosylation pathway in certain disease states can result in the truncated display of N acetyl galactosamine (GalNAc), referred to as Tn antigen, rather than fully elongated oligosaccharide chains. While Tn antigen is not displayed on normal cells, the existence of anti-GalNAc antibodies can be part of a healthy immune repertoire; however, high levels of anti-GalNAc titers represent an important indicator of several diseases. In one example, GalNAc displayed on the hinge region of IgA1 inherently stimulates increasing levels of anti-GalNAc autoantibodies. This results in IgA nephropathy in which large immune-complex aggregates accumulate ending in kidney failure. GalNAc is also displayed by helminth parasites, the coat glycoprotein of HIV, and 90% of carcinomas representing an important immune response element in several disorders. We propose a clinically relevant assay for quantifying anti-GalNAc concentrations from sera with the long term goal of improving diagnostic algorithms to identify patients with such disorders where enhanced distribution of anti-GalNAc antibodies is apparent. The impact of our proposed multimodal assay will provide not only improved diagnostic capabilities for accurately stratifying patients but also inherently building a more comprehensive understanding of how enhancements in glycan antibody levels correlate with disease states. Here we focus specifically on a liposomal assay displaying the irregular O-linked GalNAc (Tn antigen) for quantifying anti-GalNAc antibody concentrations seen as a feature in several disorders. Our stimuli-responsive liposome formulations display agglutination, color change, as well as fluorescence signals and provide a straightforward method for integration with existing pathology lab workflows. As such we anticipate this straightforward approach will lead to improved accuracy in diagnostic decision making and improved understanding of the GalNAc-specific immune response in different diseases. Our laboratory is perfectly matched to perform this research given our expertise in developing sensors and assays based on this well characterized liposome platform. Having been at the forefront in discovering new tools for the fabrication, characterization, and study of such stimuli-responsive liposomes related to those outlined in this proposal, we are confident that the technical objectives set forth can be met successfully within our proposed time frame.
The appearance of abnormal sugar (glycan) structures can be seen in numerous diseases including IgA nephropathy, Crohn's, and various cancers, giving rise to increased numbers of antibodies against those sugars. Detection of anti-glycan antibodies in serum could provide a means for assessing how the immune response has been stimulated thereby offering a window into more effective diagnosis of distinct health conditions. This study thereby holds significant value in providing a simple, clinically relevant strategy to determine the concentration of antibodies against a sugar antigen (Tn antigen) to offer a more comprehensive serum analysis which will inherently augment our understanding of the correlation between Tn antigen specific antibodies in various diseases.
