High throughput ELISA chip for drug toxicity screening

描述(申请人提供)：用于药物毒性筛选的高通量ELISA芯片概述药物的不良代谢、药物间的相互作用和直接毒性作用已导致大量药物失败。高通量生物标志物分析提供了预测药物毒性的潜力，以帮助减少管道磨损。反应生物学公司开发了极低成本的纳升反应微阵列，以服务于药物发现中的超高通量筛选(UHTS)、大规模IC50测定和选择性图谱分析市场。这些反应比目前在药物发现中广泛使用的孔板形式小1000到10000倍。基于展示细胞抗原高敏感性图谱的第一阶段数据，我们建议创建能够通过筛选从化学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的肝毒性的微阵列产品和技术。第二阶段研究的以下具体目标包括：目的1开发和验证用于药物肝毒性测试的30多个毒性生物标志物的检测方案。这些标记物将作为HepTox-CHIP(R)(肝细胞毒性生物标志物抗体微阵列)和Mi-HTS(R)(高通量筛选微阵列免疫分析)服务的基础。目的2建立包含常用非处方药和处方药的化合物文库，并用HepTox-CHIP(R)和Mi-HTS(R)芯片检测其肝毒性。这一目标将特别有助于确定新化学实体的意外药物-药物相互作用。目的3建立毒性信息学数据库(ToxID(R))，将AIM 1中有关生物标记物的公开信息与AIM 2中测试的常见药物进行基因组、蛋白质组和临床试验，作为预测肝毒性的工具。拟议的第二阶段工作的中心目标是开发两个基于抗体微阵列的产品，一个是市场上销售的、用户友好的微阵列芯片，检测30多个蛋白质毒性生物标志物，另一个是RBC提供的服务包，用于分析大量化学物质(例如，与非处方药/处方药文库交叉的测试化合物)对肝细胞毒性生物标志物表达的影响。药物引起的肝脏毒性是药物诱导死亡的主要原因，也是药物从市场上撤出的主要原因(Wering，2003)，食品和药物管理局(FDA)已经看到了通过对药物治疗动物中发现的毒性的机械理解来证明支持在人类中安全使用药物的假说的潜力(Lord&Papoian，2004)。毒素蛋白质组是一个相对较新的领域，可能具有类似的功能，但具有更直接和更精确的检测，因为监测蛋白质表达的变化比监测基因变化更相关(Wetmore&Merrick，2004；Kikkawa等人，2006；Yamamoto等人，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).