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John W. Crabb, Ph.D.Professor and Staff
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Research
Proteomic biomarker discovery for ocular diseases is a major focus of our laboratory. Our recent progress in assessing susceptibility to age-related macular degeneration (AMD) is outlined below. Other projects in the laboratory using proteomic technology are highlighted in the attached posters and include mechanistic studies of AMD, primary open angle glaucoma, diabetic retinopathy, uveal melanoma and the visual cycle.
Biomarkers for Age-related Macular Degeneration
Age-related macular degeneration (AMD) is the most common cause of legal blindness in the elderly in developed countries. Both genetic and environmental factors contribute to the disease and presently there are no cures. There is growing consensus that AMD is an inflammatory disease involving dysregulation of the complement system. AMD has also been characterized as a systemic disease, based in part on the presence of retinal drusen in patients with membranoproliferative glomerulonphritis type II and on systemic complement activation in AMD. The molecular cause of the disease remains unknown, however oxidative stress appears to be a catalyst of the AMD inflammatory response as smoking significantly increases the risk of AMD, antioxidant vitamins can selectively slow AMD progression, and a host of oxidative protein and DNA modifications have been detected at elevated levels in AMD Bruch’s membrane, drusen, retina, RPE and plasma. While AMD susceptibility genes now account for over 50% of AMD cases, many individuals with AMD risk-genotypes may never develop advanced disease with severe visual loss. Nevertheless, the prevalence of advanced AMD is increasing and early identification of AMD risk could help slow or prevent disease progression. Some years ago we found elevated levels of carboxyethylpyrrole (CEP), an oxidative protein modification generated from docosahexaenoate (DHA)-containing lipids, in Bruch’s membrane and drusen from AMD patients [2002 Proc Natl Acad Sci USA 99:14682]. Subsequently, we found CEP adducts as well as CEP autoantibodies to be elevated in plasma from AMD donors [2003 J Biol Chem 278: 42027] and CEP adducts stimulated neovascularization in vivo, suggesting a role in the induction of choroidal neovascularization (CNV) [2006 Proc Nal Acad Sci USA 103: 13480]. Recently we showed in a relatively large study population that combined CEP proteomic and genomic biomarker measurements are more effective in predicting AMD than either method alone [2009 Mol & Cell Proteomics, PMID: 19202148]. In another recent study, we quantified by LC MS/MS and LC-fluorimetry two advanced glycation endproducts (AGEs), namely carboxymethyllysine (CML) and pentosidine and demonstrated higher mean levels in AMD in plasma proteins [2009 Mol & Cell Proteomics, PMID: 19435712]. AGEs are a heterogeneous group of mostly oxidative modifications resulting from the Maillard nonenzymatic glycation reaction and their association with AMD by several laboratories support the hypothesis that AMD is a systemic disease. Our analyses suggest that plasma protein levels of CML together with pentosidine discriminate between AMD and control patients with 89% accuracy and that pentosidine in combination with CEP adducts can discriminate with 92% accuracy. In this technology, concurrent analyses of plasma protein fructosyl-lysine facilitates detection of possible confounding factors like diabetes. A much larger investigation of plasma protein CML, pentosidine and CEP as AMD biomarkers is currently underway. The long-term goal of this research is the development of routine prognostic technology for use in clinical medicine to monitor and predict AMD susceptibility and progression, much like serum cholesterol measurements are used to monitor susceptibly to cardiovascular disease.