HUMAN BIOMARKER DISCOVERY AND VALIDATION STUDIES

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The ultimate goal of the Exposure Biology Program is to understand the development and progression of complex disease by accurately, and quantitatively assessing the individual's exposure to environmental stressors and the individual's responses to these stressors. Two of the most important risk factors for human morbidity and mortality are exposure to cigarette smoke and obesity. Smoking and obesity are each associated with systemic chronic inflammatory and oxidative stress. There is increasing evidence that the oxidative stress resulting from chronic inflammatory stimuli is the unifying mechanism underlying the development of co-morbidities in cigarette smoke and obesity and for the interaction of these risk factors with other environmental toxicants and the genome. We hypothesize that reactive nitrogen and reactive oxygen species (RNS/ROS) modified proteins in plasma will provide important biomarkers for exposure to cigarette smoke, presence of obesity, and the combined response. The overarching objective for Project 1 is to identify, verify, and validate plasma protein biomarkers of oxidative stress in a human population through four aims: Aim 1. Identify RNS/ROS-modified peptides as candidate biomarkers in human plasma using MS/MS. Aim 2 will Verify RNS/ROS-modified peptides as specific biosignatures in 120 human plasma samples by data-directed MS and FTICR-MS (Core A). Aim 3. Validate RNS/ROS-modified proteins for use as specific biomarkers using custom-designed sandwich ELISA microarrays (Core B). The validation will include development and evaluation of antibody reagents and assay reproducibility and sensitivity (Core B). The selection of proteins for validation will be informed by Project 2. Aim 4. Test in the laboratory and in the clinic, a detector system for exposure and for both specific and general markers of RNS/ROS response (with Project 3). The sensor system is nanoparticle-based multiplexed Immunochromatographic / Electrochemical Biosensor (IEB) that will support the measurement of markers for exposure (cotinine) and specific RNS/RNS modified proteins and generic response markers of oxidative stress and chronic inflammation on a single platform. Project 1 will apply state-of-the-art proteomic analysis of human plasma samples from robust, well characterized cohort to provide NIEHS a database of candidate biomarkers, and reagents for selected markers for exposure to cigarette smoke, presence of obesity, and the combined effect. The biomarkers will be tested and validated a cohort containing 500 humans and informed by parallel studies in mice, and deployed on robust, in-clinic deployable nanoparticle-based sensors.