A Cell Based HTS Approach for the Discovery of New Inhibitors of the H5N1 Virus

描述（由申请人提供）：目前，没有市售疫苗来保护人类免受正在亚洲、欧洲和非洲传播的高致病性H5N1禽流感病毒的侵害。由于人类对任何H5病毒都没有免疫力，美国疾病控制与预防中心（Centers for Disease Control and Prevention）估计，下一次流感大流行的经济影响可能会使美国损失数十亿美元，摧毁世界经济，并可能导致全球10亿人死亡。因此，迫切需要抗病毒药物来补充疫苗开发和现有的化疗药物。高通量筛选（HTS）方法为筛选大型化合物库提供了机会。我们开发并验证了一种384孔的细胞检测方法，该方法使用基于荧光的信号端点检测系统来测量流感病毒感染在Madin Darby犬肾（MDCK）细胞中诱导的CPE。这种分子筛选将为科学界提供一种检测方法，通过在体外评估大型化合物文库来快速鉴定流感病毒的潜在抑制剂。本研究旨在利用分子文库筛选中心网络（MLSCN）的高通量筛选资源，促进发现抑制H5N1禽流感病毒株的新分子探针。为了实现本提案的目标，具体目标是协助MLSCN将经验证的HTS分析转移到适当的筛选中心，并以HTS剂量响应格式为初级分析的单剂量“命中”后续验证性分析提供技术支持。

DESCRIPTION (provided by applicant): Currently, there is no commercially available vaccine to protect humans against the highly pathogenic avian influenza H5N1 virus that is spreading across Asia, Europe, and Africa. Since humans have no immunity against any H5 viruses, the Centers for Disease Control and Prevention estimates that the economic impact of the next influenza pandemic could cost the United States billions of dollars, devastate the world economy and potentially kill one billion people worldwide. Thus, there is a critical need for antiviral drugs to supplement vaccine development and existing chemotherapeutics. A high throughput screening (HTS) approach provides an opportunity to screen large compound libraries. We have developed and validated a 384-well cell-based assay that measures CPE induced in Madin Darby canine kidney (MDCK) cells by influenza virus infection, using a luminescent-based detection system for signal endpoint. This molecular screen will provide the scientific community with an assay that allows for the rapid identification of potential inhibitors of influenza virus by evaluating large compound libraries in vitro. The proposed studies aim to gain access to the Molecular Libraries Screening Center Network (MLSCN) high throughput screening resources to facilitate the discovery of new molecular probes for the inhibition of avian influenza strain H5N1 virus. To achieve the objective of this proposal, the specific aim is to assist the MLSCN in transferring the validated HTS assay to the appropriate screening center and provide technical support for follow up confirmatory assays on single dose 'hits' from the primary assay in a HTS dose response format.