High throughput ELISA chip for drug toxicity screening

药物不良代谢、药物-药物相互作用和直接毒性作用导致了许多药物失败。高通量生物标志物分析提供了预测药物毒性的潜力，有助于减少管道损耗。反应生物公司开发了极低成本的纳米升反应微阵列，以服务于超高通量筛选（uHTS）、大规模IC50测定和药物发现中的选择性分析市场。这些反应比目前广泛用于药物发现的孔板形式小1000到10000倍。基于I期数据显示的细胞抗原的高敏感性分析，我们建议创建能够通过筛选从化合物或其组合处理的细胞中获得的大量细胞裂解物来评估药物诱导肝毒性的微阵列产品和技术。II期研究的以下具体目标包括：目标1：开发和验证涉及药物性肝毒性测试的30多种毒性生物标志物的检测方案。这些标志物将作为heptox芯片（肝细胞毒性生物标志物抗体微阵列）和Mi-HTS（高通量筛选微阵列免疫测定）服务的基础。目的2建立包括常用非处方药和处方药的化合物文库，并利用HepTox-chip(r)和Mi-HTS(r)芯片检测其肝毒性谱。这一目标对于确定新化学实体的意想不到的药物-药物相互作用将特别有用。目标3：通过整合目标1中基因组、蛋白质组学和临床试验中与常见药物相关的公开信息，建立毒性信息学数据库（ToxID(r)），作为肝毒性的预测工具。第二阶段工作的中心目标是开发两种基于抗体微阵列的产品，一种是可销售且用户友好的微阵列芯片，可检测30多种蛋白质毒性生物标志物，另一种是RBC提供的服务包，用于分析大量化学物质对肝细胞毒性生物标志物表达的影响（例如，测试化合物与OTC/处方药库交叉）。药物引起的肝毒性是药物引起死亡的主要原因，也是药物从市场上撤出的主要原因（Waring, 2003），食品和药物管理局（FDA）已经看到了毒物基因组学的潜力，通过对药物治疗动物毒性的机制理解，可以证明支持人类安全使用药物的假设（Lord & Papoian, 2004）。毒性蛋白质组学是一个相对较新的领域，可以具有类似的功能，但更有针对性和更精确的检测，因为监测蛋白质表达变化比监测基因变化更相关（Wetmore & Merrick, 2004； Kikkawa等，2006；Yamamoto等，2005）。

DESCRIPTION (provided by applicant): High throughput ELISA chip for drug toxicity screening Summary Adverse drug metabolism, drug-drug interactions, and direct toxicity effects have caused numerous drug failures. High throughput biomarker profiling offers the potential to predict drug toxicity to help reduce pipeline attrition. Reaction Biology Corporation has developed extremely low cost nanoliter reaction microarrays to serve markets for ultra high throughput screening (uHTS), large scale IC50 determinations, and selectivity profiling in drug discovery. These reactions are 1000 to 10,000-fold smaller than well plate formats currently used widely in drug discovery. Based on Phase I data demonstrating high sensitivity profiling of cellular antigens, we propose to create microarray products and technologies capable of evaluating drug-induced hepatotoxicity by screening large numbers of cell lysates obtained from cells treated with chemical compounds or their combinations. The following specific aims for Phase II research include: Aim 1 Develop and validate detection protocols for over 30 toxic-biomarkers involved in drug-induced liver toxicity testing. These markers will serve as the basis for the HepTox-chip(r) (hepatocyte toxic biomarkers antibody microarray) and for the Mi-HTS(r) (microarray immunoassay for high throughput screening) service offering. Aim 2 Establish a compound library including common over-the-counter and prescription drugs and test their hepatotoxicity profiles with HepTox-chip(r) and Mi-HTS(r) chip. This aim will be particularly useful for identifying unexpected drug- drug interactions of new chemical entities. Aim 3 Develop the Toxic-Informatics Database (ToxID(r)) by integrating the publicly available information related to biomarkers from Aim 1 in genomic, proteomic and clinic tests with common drugs tested in Aim 2 as a predictive tool for hepatotoxicity. The central goal of the proposed Phase II efforts is to develop two antibody microarray based products, one a marketable and user-friendly microarray chip detecting over 30 protein toxic-biomarkers and the other a service package that RBC provides for analyzing a large number of chemicals effects (e.g. test compound crossed against the OTC/prescription drug library) on hepatocyte toxic biomarker expression. Drug-induced liver toxicity is the major cause of drug-induced death and the principle reason for withdrawal of drugs from the market (Waring, 2003) and Food and Drug Adminstration (FDA) already sees toxicogenomic's potential in proving hypotheses that support safe drug use in humans through a mechanistic understanding of toxicities found in drug-treated animals (Lord & Papoian, 2004). The toxicoproteomic is a relatively new area that could have similar function but with more directed and precisely detection, because monitoring protein expression changes is more relevant than monitoring gene changes (Wetmore & Merrick, 2004; Kikkawa et al, 2006; Yamamoto et al, 2005).