New Hepatocyte Model Systems to study Hepatitis B Virus Infection

研究乙型肝炎病毒感染的新肝细胞模型系统

• 批准号: 9016547
• 负责人: Robert E Schwartz
• 金额: $ 15.35万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-15 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9016547
• 关键词: Adult; Aging; American; B-Lymphocytes; Biological Models; Biology; Cell Line; Cells; Cellular Tropism; Chemicals; Chronic Hepatitis B; Cirrhosis; Clinical; Genetic; Health; Hepatitis B; Hepatitis B Virus; Hepatocyte; Human; Immunosuppression; Infection; Life Cycle Stages; Metabolic; Modeling; Pathway interactions; Patients; Phenotype; Play; Pluripotent Stem Cells; Primary carcinoma of the liver cells; Process; Production; Regulation; Risk; Role; Stem cells; System; Techniques; United States; Vaccines; Variant; Viral; Viral Pathogenesis; Viral Proteins; Viral hepatitis; Virion; Virus; Virus Diseases; Virus Replication; Work; genetic variant; hepatoma cell; human stem cells; in vitro Model; induced pluripotent stem cell; innovation; insight; new therapeutic target; overexpression; permissiveness; prevent; response; screening; therapeutic development; tool; viral DNA; virus pathogenesis

DESCRIPTION (provided by applicant): Worldwide, hepatitis B virus (HBV) infection is the most common viral hepatitis having infected over two billion people and chronically infecting more than 400 million, putting them at increased risk to develop cirrhosis and hepatocellular carcinoma. In the United States over one million people have chronic hepatitis B infection. Clinical therapy is targeted to the suppression of viral replication but the virus is able to persit in a nonreplicative covalently closed circular form called cccDNA, with the potential to reactivate upon immune suppression or with aging. As a consequence, the hepatitis B virus in chronic HBV infections is challenging to eradicate and cure is rare. The difficulty in developing new HBV therapies has been due to the lack of good model systems. The current model system to study hepatitis B has been hepatoma cell lines in which HBV is over expressed transiently or stably to produce all viral gene products and maintain replication. However such models over express HBV in a non-physiological manner and do not faithfully recapitulate adult hepatocyte phenotype or function as they have undergone a variety of genetic and metabolic changes. As a consequence, the viral entry factors as well as the regulatory processes of viral replication have been poorly understood. This has prevented development of therapeutic strategies that achieve viral control and eradication. This proposal builds on new tools and techniques that enable the long-term culture of metabolically functional primary human hepatocytes and the establishment of pluripotent stem cell derived hepatocyte-like cells. Both innovations enables the exploration of the determinants of HBV entry, viral replication, and the cccDNA state which will open new opportunities to better understand HBV biology and develop new targeted therapies. The establishment of a culture model system that is permissive for HBV infection and also enables the long-term culture of metabolically functional primary human hepatocytes represents a new opportunity to explore the determinants of HBV entry, viral replication, and cccDNA formation and persistence (Aims 1-2). In contrast to current hepatoma cell lines, a functional hepatocyte screening platform will enable the study of the role that biologic pathways play in the HBV viral life cycle's choice towards cccDNA production versus active HBV virion production (Aim 2). These pathways can then be explored and dissected in detail in induced pluripotent derived stem cell derived variants to determine their impact on hepatocyte permissiveness, hepatitis B virus pathogenesis and cccDNA formation (Aim 3). Better insights in HBV pathogenesis, cccDNA formation, virus evasion and the determinants of hepatocyte autonomous responses may also be translational as treatment options for chronically infected HBV patients are available but are unable to eliminate cccDNA leading to lifelong infection and requirements for lifelong treatment.

描述（由申请人提供）：在世界范围内，B型肝炎病毒（HBV）感染是最常见的病毒性肝炎，已感染超过20亿人，慢性感染超过4亿人，使他们患肝硬化和肝细胞癌的风险增加。在美国，超过一百万人患有慢性B型肝炎感染。临床治疗的目标是抑制病毒复制，但病毒能够以称为cccDNA的非复制性共价闭合环状形式存在，并有可能重新激活 免疫抑制或衰老时。因此，慢性HBV感染中的B型肝炎病毒很难根除，治愈率很低。开发新的HBV疗法的困难是由于缺乏良好的模型系统。目前研究B型肝炎的模型系统是肝癌细胞系，其中HBV瞬时或稳定过表达以产生所有病毒基因产物并维持复制。然而，这些模型以非生理方式过度表达HBV，并且不能忠实地再现成体肝细胞表型或功能，因为它们已经经历了各种遗传和代谢变化。因此，病毒进入因子以及病毒复制的调控过程一直知之甚少。这阻碍了实现病毒控制和根除的治疗策略的发展。该提案建立在新的工具和技术的基础上，这些工具和技术能够长期培养具有代谢功能的原代人肝细胞，并建立多能干细胞衍生的肝细胞样细胞。这两项创新使人们能够探索 HBV进入、病毒复制和cccDNA状态的决定因素，这将为更好地理解HBV生物学和开发新的靶向治疗提供新的机会。允许HBV感染并且还能够长期培养代谢功能性原代人肝细胞的培养模型系统的建立代表了探索HBV进入、病毒复制以及cccDNA形成和持久性的决定因素的新机会（目的1-2）。与目前的肝癌细胞系相比，功能性肝细胞筛选平台将能够研究生物途径在HBV病毒生命周期选择cccDNA生产与活性HBV病毒体生产中的作用（目的2）。然后可以在诱导多能干细胞衍生变体中详细探索和剖析这些途径，以确定其对肝细胞容许性、B型肝炎病毒发病机制和cccDNA形成的影响（目的3）。对HBV发病机制、cccDNA形成、病毒逃逸和肝细胞自主反应决定因素的更好理解也可能是转化的，因为慢性感染HBV患者的治疗选择是可用的，但无法消除导致终身感染和终身治疗要求的cccDNA。