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Hepatitis C virus polarized cell entry pathways

丙型肝炎病毒极化细胞进入途径

基本信息

批准号：
8523846
负责人：
Matthew J Evans
金额：
$ 35.26万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-15 至 2017-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8523846
关键词：
Address Adverse effects Affect Animal Model Antiviral Agents Biochemical Genetics Biological Assay Biological Models CD81 gene Cell Culture Techniques Cell Line Cell Polarity Cell physiology Cells Clathrin Complement Derivation procedure Development Endocytosis Endosomes Event Foundations Genetic Glycoproteins Glycosaminoglycans Goals Health Hepatitis C Hepatitis C virus Hepatocyte Human Individual Infection Integration Host Factors Kinetics Knowledge Label Life Life Cycle Stages Light Liver Location Low Density Lipoprotein Receptor Malignant neoplasm of liver Mediating Methods MicroRNAs Modeling Mus Mutation Nature Pan Genus Pathway interactions Peptide antibodies Process Proteins Public Health RNA replication Relative (related person)Research Resistance Resistance development Risk Role SR-BI receptor Staging System Testing Tight Junctions Time Viral Viral Proteins Virion Virus Virus Inhibitors Virus Replication Visual base claudin-1 protein genetic analysis human PHEMX protein in vivo inhibitor/antagonist innovation insight liver injury mutant novel occludin polarized cell receptor research study viral RNA

项目摘要

DESCRIPTION (provided by applicant): The hepatitis C virus (HCV) is the leading cause of liver cancer in the Western Hemisphere. Over 170 million people are infected with this virus and thus at increased risk of liver damage and cancer. HCV infection is curable, and thus such health consequences are completely avoidable. However the current HCV therapy is inadequate, as it is associated with severe side effects and only effective in about half of treate individuals. Effective HCV treatment will require antivirals targeting multiple HCV replication steps that can be used in combination to avoid the derivation of resistant viruses. The HCV cell entry process is an attractive target for antiviral development. Inhibiting this stage of the virallife cycle would curtail the persistence of HCV in vivo, which requires constant infection of new cells, as well as the spread of viral mutants that have developed resistance to other HCV inhibitors. The research proposed in this application will advance our understanding of HCV cell entry mechanisms, with the ultimate goal of uncovering novel antiviral targets. Although we now know that numerous host factors are required for HCV cell entry, we do not understand how they mediate HCV infection. The experiments described in this application are based on the hypothesis that the incoming HCV virion utilizes each of these cellular factors in a sequential manner to mediate cell entry. As two of the host proteins that are required for HCV cell entry are tight junction proteins, it is likely that cell polarity, which regulates the localization and funcions of these proteins, impacts the HCV cell entry process. Although the majority of HCV cell entry studies have been conducted in non-polarized cells, we have recently developed the first polarized cell system that permits efficient infection of authentic HCV grown in cell culture. This system will serve as the foundation for the studies proposed in this application. Specifically, we propose experiments to compare HCV cell entry pathways in non-polarized and polarized using a variety of HCV cell entry factor specific inhibitors. Furthermore, we will test how modulating cell polarity influences HCV cell entry, and examine the host factor requirements with entry factor specific inhibitors, silencing, and complementation with a variety of mutants. We will also analyze when and where each entry factor is used by analyzing the kinetics of action of entry factor specific inhibitors and visualizing the HCV cell entry process in non-polarized and polarized cells. Finally, we will identify physical and genetic interactions between host factors and the incoming HCV virion, and explore how such interactions are affected by cell polarity.

描述（由申请人提供）：丙型肝炎病毒（HCV）是西半球肝癌的主要原因。超过 1.7 亿人感染了这种病毒，因此肝损伤和癌症的风险增加。 HCV感染是可以治愈的，因此这种健康后果是完全可以避免的。然而，目前的丙型肝炎病毒治疗还不够，因为它具有严重的副作用，并且仅对大约一半的治疗个体有效。有效的丙型肝炎治疗需要针对多个丙型肝炎病毒复制步骤的抗病毒药物，这些药物可以联合使用以避免产生耐药病毒。 HCV 细胞进入过程是抗病毒开发的一个有吸引力的目标。抑制病毒生命周期的这一阶段将减少丙型肝炎病毒在体内的持续存在，这需要不断感染新细胞，以及对其他丙型肝炎病毒抑制剂产生抗性的病毒突变体的传播。本申请中提出的研究将增进我们对丙型肝炎病毒细胞进入机制的理解，最终目标是发现新的抗病毒靶点。尽管我们现在知道HCV细胞进入需要许多宿主因子，但我们不了解它们如何介导HCV感染。本申请中描述的实验基于以下假设：传入的HCV病毒体以顺序方式利用这些细胞因子中的每一个来介导细胞进入。由于 HCV 细胞进入所需的两种宿主蛋白是紧密连接蛋白，因此调节这些蛋白质的定位和功能的细胞极性很可能会影响 HCV 细胞进入过程。尽管大多数 HCV 细胞进入研究是在非极化细胞中进行的，但我们最近开发了第一个极化细胞系统，该系统允许有效感染在细胞培养物中生长的真实 HCV。这系统将作为本申请中提出的研究的基础。具体来说，我们提出了使用各种 HCV 细胞进入因子特异性抑制剂来比较非极化和极化的 HCV 细胞进入途径的实验。此外，我们将测试调节细胞极性如何影响 HCV 细胞进入，并通过进入因子特异性抑制剂、沉默和与各种突变体的互补来检查宿主因子的要求。我们还将通过分析进入因子特异性抑制剂的作用动力学并可视化非极化和极化细胞中的 HCV 细胞进入过程来分析每个进入因子的使用时间和地点。最后，我们将确定宿主因子与传入的 HCV 病毒粒子之间的物理和遗传相互作用，并探讨细胞极性如何影响这种相互作用。