Novel Transcription Profiling in Toxicologic Assays

描述（由申报人提供）：药物开发失败或退出市场的常见原因是肝毒性。这是由于药物代谢、酶诱导或抑制药物代谢酶导致的许多药物不良反应和药物-药物相互作用。不幸的是，缺乏有效的体外和体内方法来准确预测新化学实体对人体的肝毒性。最近有许多报告表明，基因活性的变化可以提供有关毒理学终点的非常有价值的信息。在该领域，RNA谱分析通常是鉴定差异基因表达的标准。然而，这些方法也有局限性。一种新的平台技术，称为转录路径，基于dna转录机制复合物的染色质免疫沉淀，识别细胞内进行活性转录的基因，并非常准确和可重复地量化其转录水平。此外，转录路径在测量细胞和组织内部非常早期的转录变化方面是强大的，这应该允许在代谢途径开始时识别变化。该提案将应用新的转录路径方法，在已知导致化学诱导肝毒性的药物治疗大鼠后的早期时间点鉴定肝脏基因转录变化。此外，将使用相同的药物治疗分离的大鼠肝细胞，也将对其进行转录谱分析，以确定是否在体外观察到相同的基因表达变化。基因转录变化也将与功能性生化变化相关。在第二阶段，来自大鼠研究的诊断谱将用于评估肝毒性终点的跨物种尺度，并确定预测人类肝毒性潜力的新基因表达特征。本文提出的技术发展将大大有助于改善临床前安全性评估，并最终帮助公司、政府机构和学术实体以更短的时间和更低的成本开发更安全的药物。

DESCRIPTION (provided by applicant): A common reason for failure of drugs in development or their removal from the market is hepatotoxicity. This is due to the many adverse drug reactions and drug-drug interactions that result from drug metabolism, enzyme induction or inhibition of drug metabolizing enzymes. Unfortunately, effective in vitro and in vivo approaches to accurately predict hepatotoxicity of new chemical entities in humans are lacking. Numerous reports have shown recently that changes in gene activity can provide highly valuable information about toxicological endpoints. Within that area, RNA profiling has typically been the standard for identifying differential gene expression. However, these methods also have limitations. A new and novel platform technology, called TranscriptionPath and based on chromatin immunoprecipitation of DNA-transcription machinery complexes, identifies genes undergoing active transcription inside cells and quantifies very accurately and reproducibly their levels of transcription. Further, TranscriptionPath is powerful in being able to measure very early transcriptional changes inside cells and tissues, which should permit the identification of changes at the beginnings of metabolic pathways. This proposal will apply the new TranscriptionPath method to the identification of gene transcription changes in liver at early time points after treatment of rats with drugs known to result in chemically-induced hepatotoxicity. In addition, the same drugs will be used to treat isolated rat hepatocytes, which will also be transcriptionally profiled to determine if the same gene expression changes are observed in vitro. Gene transcription changes will also be correlated with functional biochemical changes. In Phase II, the diagnostic profiles derived from the rat studies will be used to assess the cross-species scaling of the hepatotoxic endpoints and to identify new gene expression signatures predictive of hepatotoxic potential in humans. Development of the technology proposed herein should significantly help improve preclinical safety assessment and eventually help companies, governmental agencies and academic entities develop safer drugs in less time and with lower costs.