Center for Novel Biomarkers of Response

暴露生物学计划的最终目标是通过准确、准确和定量地评估个人对环境应激源的暴露以及个人对这些应激源的反应来了解复杂疾病的发展和进展。吸烟和肥胖是导致人类发病率和死亡率的两个最重要的风险因素，两者都与全身慢性炎症和氧化应激有关。该中心的组织主题是，对血浆中持久修饰蛋白的识别和验证将提供关于应激源、其作用方式和靶器官的特定信息。人类和动物项目的环境应激集中在主流和侧流卷烟烟雾上，肥胖是一个令人困惑的生理因素。我们提出了一个综合的、多学科的中心，包括三个研究项目(人、鼠和传感器)和两个技术核心(蛋白质组学和ELISA微阵列)，它们总共将有四个目标。目的1.以MS/MS为候选生物标志物，寻找活性氮和活性氧基团(RNS/ROS)修饰多肽。目的2.利用数据定向MS验证RNS/ROS修饰多肽作为特异性生物标志物。目的3.使用定制设计的夹心ELISA微阵列验证RNS/ROS修饰蛋白作为环境应激源的特异性生物标志物的有效性。目的4.开发、测试和部署两个探测器系统，用于暴露以及RNS/ROS反应的特异性和一般性标志物。基于实验室的ELISA微阵列平台将用于提供高通量， 对数十种分析物进行多路分析。经过验证的生物标记物还将被部署在一个原型、临床可部署的探测器系统上，用于现场分析。该传感器系统是基于纳米颗粒的多路免疫层析/电化学生物传感器(IEB)，将支持在单一平台上测量暴露(可替宁)和反应的标志物，包括氧化应激和慢性炎症的特定(修饰蛋白)和通用标志物，如肿瘤坏死因子-α。关键成果和可交付成果。PNNL U54中心将应用最先进的蛋白质组和传感器技术来提供 NIEHS具有告知反应的生物标志物的作用模式数据库，选定的标志物的试剂在人类中进行测试和验证，并通过小鼠的平行研究获得信息，并部署在(A)基于实验室的ELISA微阵列平台和(B)适合用于大规模人体生物监测研究的坚固、临床可部署的纳米颗粒传感器上，以评估基因与环境的相互作用。这个U54中心的结果还将提高科学界比较、对比和推断人类和老鼠之间的生物标志物的能力。

The ultimate goal of the Exposure Biology Program is to understand the development and progression of complex disease by precisely, 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, both assocaiated with systemic chronic inflammation and oxidative stress. The organizing theme the Center is that identification and validation of persistent modified proteins in plasma will provide specific information about the stressor, its mode of action, and the target organ. The environmental stress of the human and animal projects is focused on main-stream and side-stream cigarette smoke with obesity as a confounding physiological factor. We propose an integrated, multidisciplinary center with three research projects (human, mouse, and sensor) and two technology cores (proteomics and ELISA Microarray) which collectively will have four goals. Aim 1. Discover reactive nitrogen and reactive oxygen species (RNS/ROS) modified peptides using MS/MS as candidate biomarkers. Aim 2. Verification of RNS/ROS modified peptides as specific biosignatures using data-directed MS. Aim 3. Validation of RNS/ROS modified proteins for use as specific biomarkers for environmental stressors using custom-designed sandwich ELISA microarrays. Aim 4. Develop, test, and deploy two detector systems for exposure and for both specific and general markers of RNS/ROS response. A laboratory-based ELISA Microarray platform will be used to provide high-throughput, multiplexed analysis of dozens of analytes. The validated biomarkers will be also be deployed on a prototype, clinic-deployable detector system for on-site analysis. The sensor system is nanoparticle-based multiplexed Immunochromatographic / Electrochemical Biosensor (IEB) that will support the measurement of markers for exposure (cotinine) and response, both specific (modified proteins) and generic markers of oxidative stress and chronic inflammation, such as TNF-alpha, in a single platform. Critical Outcomes and Deliverables. The PNNL U54 Center will apply state-of-the-art proteomic and sensor technologies to provide NIEHS with a database of mode-of-action informing biomarkers of response, reagents for selected markers tested and validated in humans and informed by parallel studies in mice, and deployed on (a) a laboratory based ELISA Microarray platform, and (b) on a robust, clinic-deployable nanoparticle-based sensor suitable for use in large-scale human biomonitoring studies to evaluate the interaction of genes and the environment. The results of this U54 Center also will improve the scientific community's ability to compare, contrast and extrapolate biomarkers between humans and mice.