Ion Channel Assay for Identifying Influenza Therapeutics

描述（由申请方提供）：流感病毒是正粘病毒科的一种包膜RNA病毒，是NIAID关注的C类生物防御病原体。尽管在疫苗和药物的开发方面取得了重大进展，但流感病毒每年感染5-15%的人口，每年导致300 - 500万例严重疾病和250，000 - 500，000例死亡。每年都会出现对疫苗有抗药性的新流感毒株，而现有的抗流感药物往往很容易被规避。我们需要新的抗流感药物来抑制不同的病毒株，并抵抗病毒逃逸。这一提议将导致一种新的检测方法，用于发现抗流感的治疗方法。FDA批准的四种流感治疗药物中有两种靶向M2离子通道，使M2成为治疗流感感染的有效分子靶标。然而，开发靶向M2的新疗法一直很困难。离子通道一直是传统高通量筛选（HTS）的棘手靶标，这在很大程度上是因为基于细胞的离子通道测定限制了优选HTS策略的应用。需要操纵离子通道并检测其活性的新方法来帮助开发针对困难的离子通道靶点如M2的新药。该提案的目标是开发一种高通量筛选测定法，该测定法可以鉴定针对流感M2蛋白的新型抑制剂。同样对NIAID具有重要意义的是，由该提议产生的方法和技术也将有助于发现新的病毒离子通道、其他病毒内的离子通道的新抑制剂，以及研究理解病毒离子通道对病毒复制和病理学的贡献。该提案的具体目标是： I.建立阻断M2活性的化合物的主要HTS试验。 二.在概念验证HTS中测试M2活性测定。

DESCRIPTION (provided by applicant): Influenza is an enveloped RNA virus of the orthomyxovirus family and a Category C biodefense pathogen of interest to the NIAID. Despite significant advances in the development of both vaccines and drugs, influenza viruses infect 5-15% of the human population every year, resulting in 3-5 million cases of severe illness and 250,000-500,000 deaths annually. New strains of influenza that are resistant to vaccines emerge every year, and existing drugs against influenza are often easily evaded. New drugs against influenza are needed that inhibit diverse strains of the virus and that resist viral evasion. This proposal will result in a novel assay for the discovery of therapeutics against influenza. Two of the four FDA-approved therapeutics against influenza target the M2 ion channel, making M2 a validated molecular target for the treatment of influenza infection. However, developing new therapeutics that target M2 has been difficult. Ion channels have been troublesome targets for traditional high-throughput screening (HTS), in large part because cell-based ion channel assays restrict the application of preferred HTS strategies. New methods of manipulating ion channels and detecting their activity are needed to aid the development of new drugs against difficult ion channel targets such as M2. The goal of this proposal is to develop a high-throughput screening assay that can identify novel inhibitors against influenza M2 proteins. Also of significance to the NIAID, the methods and techniques that result from this proposal will also be useful for the discovery of new viral ion channels, new inhibitors of ion channels within other viruses, and research into understanding the contribution of viral ion channels to viral replication and pathology. The Specific Aims of this proposal are to: I. Establish a Primary HTS Assay for Compounds that Block M2 Activity. II. Test M2 Activity Assay in Proof-of-concept HTS.