Antiviral potential of helicase inhibitors

解旋酶抑制剂的抗病毒潜力

• 批准号: 8018993
• 负责人: David N Frick
• 金额: $ 36.56万
• 依托单位: UNIVERSITY OF WISCONSIN MILWAUKEE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-01 至 2013-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8018993
• 关键词: ATP Hydrolysis; ATP phosphohydrolase; Acquired Immunodeficiency Syndrome; Antiviral Agents; Basic Science; Biological Assay; Boxing; Categories; Cells; Chemicals; Chemistry; Clinic; Clinical; Clinical Trials; Collaborations; Collection; DDX1 gene; DDX1 protein; DNA; DNA Binding; DNA Viruses; Data; Dengue; Dengue Virus; Development; Direct Costs; Drug Delivery Systems; Enzymes; Flavivirus; Funding; Funding Opportunities; Future; Goals; HIV; Hepatitis C; Hepatitis C virus; Human; Japanese Encephalitis; Japanese Encephalitis Viruses; Japanese encephalitis virus; Kansas; Learning; Ligand Binding; Link; Molecular; Molecular Bank; Monitor; New York; Patients; Peptide Hydrolases; Peptides; Pharmaceutical Preparations; Polymerase; Preclinical Drug Evaluation; Production; Proteins; RNA; RNA Viruses; RNA helicase A; RNA replication; Research Institute; Research Project Grants; Role; Screening procedure; Specificity; Testing; Translating; United States National Institutes of Health; Virus; Virus Replication; West Nile virus; Yellow Fever; Yellow fever virus; anti-hepatitis C; base; biodefense; burden of illness; design; drug development; helicase; high throughput screening; human WRN protein; inhibitor/antagonist; innovation; medical schools; pathogen; public health relevance; response; scale up; small molecule; tool; viral RNA

DESCRIPTION (provided by applicant): In response to the Funding Opportunity Announcement "Development of Assays for High-Throughput Drug Screening," we propose here to develop assays to identify and analyze inhibitors of helicases needed for RNA virus replication. All viruses require helicases to separate duplex DNA or RNA, yet helicases remain undeveloped antiviral drug targets. Although some helicase inhibitors that target DNA viruses are progressing in clinical trials, no antiviral agents targeting RNA viruses have entered the clinical arena. We therefore will target RNA viruses, focusing on pathogens with abnormally high global burdens of disease: hepatitis C virus (HCV), Dengue virus (DENV), West Nile virus (WNV), Japanese encephalitis virus (JEV), yellow fever virus (YFV), and human immunodeficiency virus (HIV). The assays to be developed are needed as part of an ongoing collaborative hypothesis-driven research project to develop HCV NS3 helicase inhibitors as chemical probes to facilitate basic science and for drug development. In that endeavor, we have initiated an uHTS at the MLPCN that uses our recently developed molecular beacon based helicase assay (MBHA) as a primary screen. To evaluate HCV helicase inhibitor specificity, MBHAs will be developed using helicases encoded by viruses related to HCV (DNV, WNV, YFV, and JEV) and human cellular helicases. The human proteins to be analyzed have all been linked to HIV replication and include the DEAD-box proteins DDX1, DDX3, DDX24, MDA-5, RNA helicase A (RHA), and Werner syndrome protein (WRN). In aim 2, assays will be developed to analyze the cellular antiviral efficacy, and mechanism of action of HCV helicase inhibitors. Mechanistic assays include ATPase assays, ligand binding assays, protein oligomerization assays, and simultaneous protease/helicase assays. Most of these mechanistic assays could be used with either HCV helicase or one of the ten other helicases targeted here to screen for additional classes of helicase inhibitors. PUBLIC HEALTH RELEVANCE: In collaboration with a NIH high throughput-screening center, my lab is searching a collection of hundreds of thousands of small molecules for compounds that might inhibit the helicase encoded by the hepatitis C virus (HCV). In this project, we will develop tests to rapidly search these HCV helicase inhibitors for compounds that could be used to treat hepatitis C. We will also develop similar assays to identify compounds that might inhibit viruses that cause AIDS, yellow fever, Dengue fever, West Nile and Japanese encephalitis.

描述（由申请人提供）：为了响应资助机会公告“高通量药物筛选测定的开发”，我们在此建议开发测定来鉴定和分析RNA病毒复制所需的解旋酶抑制剂。所有病毒都需要解旋酶来分离双链DNA或RNA，但解旋酶仍然是未开发的抗病毒药物靶标。尽管一些靶向DNA病毒的解旋酶抑制剂正在临床试验中取得进展，但还没有靶向RNA病毒的抗病毒剂进入临床竞技场。因此，我们将针对RNA病毒，重点关注全球疾病负担异常高的病原体：丙型肝炎病毒（HCV），登革热病毒（DENV），西尼罗河病毒（WNV），日本脑炎病毒（JEV），黄热病病毒（YFV）和人类免疫缺陷病毒（HIV）。作为正在进行的合作假设驱动的研究项目的一部分，开发HCV NS 3解旋酶抑制剂作为化学探针，以促进基础科学和药物开发，需要开发的检测方法。在这奋进，我们已经在MLPCN启动了uHTS，该uHTS使用我们最近开发的基于分子信标的解旋酶测定（MBHA）作为初级筛选。为了评价HCV解旋酶抑制剂特异性，将使用HCV相关病毒（DNV、WNV、YFV和JEV）编码的解旋酶和人细胞解旋酶开发MBHA。待分析的人类蛋白质都与HIV复制有关，包括DEAD盒蛋白DDX 1、DDX 3、DDX 24、MDA-5、RNA解旋酶A（RHA）和沃纳综合征蛋白（WRN）。在目标2中，将开发分析HCV解旋酶抑制剂的细胞抗病毒功效和作用机制的测定法。机理测定包括ATP酶测定、配体结合测定、蛋白质寡聚化测定和同时蛋白酶/解旋酶测定。大多数这些机制的测定可以与HCV解旋酶或其他十个解旋酶之一，在这里针对筛选其他类别的解旋酶抑制剂。 公共卫生相关性：在与NIH高通量筛选中心的合作中，我的实验室正在搜索数十万个小分子的集合，以寻找可能抑制丙型肝炎病毒（HCV）编码的解旋酶的化合物。在这个项目中，我们将开发测试，以快速搜索这些HCV解旋酶抑制剂的化合物，可用于治疗丙型肝炎。我们还将开发类似的检测方法，以确定可能抑制导致艾滋病、黄热病、登革热、西尼罗河和日本脑炎的病毒的化合物。