Kianse Networks Controling Flavivirus Replication

Kianse Networks 控制黄病毒复制

• 批准号: 8234067
• 负责人: Klaus J Fruh
• 金额: $ 64.83万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2014-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8234067
• 关键词: Animal Model; Antiviral Agents; Biochemical; Cell model; Cell physiology; Cells; Chemicals; Computer Simulation; Data; Dengue; Dengue Virus; Drug Delivery Systems; Environment; Enzymes; Flavivirus; Flavivirus Infections; Gene Targeting; Genes; Genome; Genomics; Goals; Growth; Immune response; In Vitro; Infection; Infection Control; Japanese Encephalitis; Libraries; Life Cycle Stages; Mitogen-Activated Protein Kinases; Modeling; Mus; Pacific Northwest; Pathway interactions; Pharmaceutical Chemistry; Phosphorylation; Phosphotransferases; Protein Binding; Protein Kinase; Proteins; Screening procedure; Signal Pathway; Small Interfering RNA; Specificity; Testing; Therapeutic; Thiourea; Tissues; Toxic effect; Tyrosine Kinase Inhibitor; Viral; Viral Physiology; Virus; Virus Diseases; Virus Inhibitors; Virus Replication; West Nile virus; Yellow fever virus; analog; base; cell type; drug discovery; functional genomics; in vivo; indexing; inhibitor/antagonist; insight; kinase inhibitor; mouse model; novel; prevent; programs; research study; small molecule; small molecule libraries; src-Family Kinases

Flaviviruses need and manipulate host cell pathways at every step of their infectious cycle. Since many host cell pathways are controlled by protein kinases, this class of host proteins is intimately involved in modulating flaviviral replication both negatively, ie. by controlling innate immune responses, as well as positively, by inducing a virus-supportive environment. The goal of this sub-project is to identify those kinase-controlled host cell pathways that support the flavivirus life cycle. Our preliminary data show that broad-spectrum protein tyrosine kinase inhibitors prevent flaviviral entry whereas MAP-kinase inhibitors interfere with replication. Moreover, we previously demonstrated that Src-kinase inhibitors block flaviviral egress due to a requirement of the c-yes kinase for this step. To identify both specific inhibitors and specific kinases controlling each step of the viral life cycle we will use a chemical genomics approach. We will employ cellbased, high content screening of kinase-directed small molecule libraries to identify kinase inhibitors interfering with different steps of the Dengue-virus (DENV) life cycle. In secondary screens we will select for compounds with a high selectivity index, broad cell type specificity and activity against other flaviviruses (WNV, JEV, YFV). Active compounds will likely inhibit several related kinases thus minimizing the problem of redundancy of signaling pathways and tissue specific expression. The kinases targeted by a given compound will be identified by combining computational, functional genomics and biochemical approaches. Based on these results we will generate models of kinase networks controlling flavivirus infection and examine thise models experimentally by phosphoproteomics and by targeted gene knockdown. Finally, we will examine the therapeutic potential of kinase-targeted antivirally active compounds by examining selected compounds in vivo using a murine model West-Nile Virus infection. The efficacy, toxicity and bio-availability of kinase-directed compounds will be compared with the in vivo activity of TYT-1 analogs (sultam thioureas) generated in sub-project 3 of this program. At the end of these comprehensive studies, we anticipate to have gained further insights into the interaction of flaviviruses with their host cell, identified novel targets for antviral drugs discovery, and characterized novel small molecule inhibitors that are active against flaviviruses in vitro and in vivo.

黄病毒在传染周期的每个步骤中都需要并操纵宿主细胞通路。由于许多主机 细胞途径由蛋白激酶控制，这类宿主蛋白与调节密切相关 黄素复制既负面复制，即。通过控制先天免疫反应以及积极的控制 诱导病毒支持环境。该子项目的目的是识别那些激酶对照 支持黄酮病毒生命周期的宿主细胞通路。我们的初步数据表明广谱 蛋白酪氨酸激酶抑制剂预防黄素进入，而MAP-激酶抑制剂会干扰 复制。此外，我们先前证明了SRC-激酶抑制剂阻止了由于A 此步骤的C-YES激酶的要求。识别特定抑制剂和特定激酶 控制病毒生命周期的每个步骤，我们将使用化学基因组学方法。我们将采用基于细胞的 激酶定向的小分子库的高含量筛选以鉴定激酶抑制剂 干扰登革热病毒（DENV）生命周期的不同步骤。在辅助屏幕中，我们将选择 具有高选择性指数，宽细胞类型特异性和针对其他黄病毒的活性的化合物 （WNV，JEV，YFV）。活性化合物可能会抑制几种相关激酶，从而最大程度地减少 信号通路和组织特异性表达的冗余。由给定的激酶 通过结合计算，功能基因组学和生化方法来识别化合物。 基于这些结果，我们将生成控制黄病毒感染和 通过磷蛋白质组学和靶向基因敲低实验检查这一模型。最后，我们 通过检查选定 使用鼠模型西奈罗病毒感染的体内化合物。功效，毒性和生物可用性 将激酶定向的化合物与TYT-1类似物的体内活性（Sultam thioureas）进行比较 在此程序的子项目中生成。在这些综合研究的结尾，我们预计 进一步了解了黄素病毒与宿主细胞的相互作用，确定了新的目标 抗病毒药物的发现，并表征了针对黄病毒活性的新型小分子抑制剂 体外和体内。