Modeling human hepatotropic infections in complex tissue organoids

描述(由申请人提供)：人体肝脏是几种重要的人类病原体的储存库，包括乙肝病毒(乙肝)、丙型肝炎病毒(丙型肝炎病毒)和疟原虫物种，所有这些都代表着严重的全球健康问题。据估计，仅乙肝病毒和丙型肝炎病毒就感染了全球5亿人，每年死亡总数超过150万人。慢性乙肝病毒和丙型肝炎病毒感染可能会造成严重的健康后果，包括肝炎、肝硬变、肝功能衰竭和肝细胞癌。与乙肝病毒和丙型肝炎病毒混合感染是很常见的，并会导致明显更糟糕的预后。目前存在针对乙肝病毒的预防性疫苗，但针对该病毒的根治疗法尚不存在。此外，据报道，一线抗病毒药物增加了对乙肝病毒的耐药性。丙型肝炎病毒的疫苗尚未开发出来，虽然丙型肝炎病毒特异性的蛋白酶和聚合酶抑制剂在早期临床开发中显示出希望，但迅速出现的耐药性表明，将需要更多的靶点和抗病毒药物的组合来有效控制。缺乏忠实地模拟肝脏生物学的体外和体内系统以及对人类嗜肝病原体的敏感性，严重阻碍了药物和疫苗的开发。在这里，我们建议采取跨学科的方法，将组织工程与分子病毒学和人源化小鼠技术相结合，创建平台，促进对基本病毒-宿主和病毒-病毒相互作用的研究，促进对肝病进展机制的了解，并提供预测系统来测试药物和疫苗的有效性和毒性。具体地说，我们的目标是在微图案化的原代人类肝细胞培养(MPCCs)中表征乙肝病毒和丙型肝炎病毒的生物学特征和相关肝脏疾病的模型--这是Sangeeta Bhatia博士的实验室最近开发的一项突破性技术。我们将把这一方法扩展到开发三维肝脏器官，并研究移植了这些结构的小鼠的乙肝和丙型肝炎病毒感染。由于慢性炎症在肝脏疾病的发展中起着重要作用，我们的目标是在静态和动态组织培养系统中整合免疫系统的组件，并最终使用与人类造血系统重组的小鼠来接受允许乙肝病毒/丙型肝炎病毒的肝脏有机物质。通过这些平台的开发和使用，我们希望揭示乙肝病毒和丙型肝炎病毒的病毒学和发病机制，并揭示治疗干预的新途径。 公共卫生相关性：乙型和丙型肝炎病毒是包括癌症在内的人类肝病的主要原因。该建议旨在建立和利用新的原代人肝细胞组织培养系统和复杂的三维肝脏器官来研究丙型肝炎病毒和乙肝病毒的单一和联合感染。由此产生的发现应该有助于开发更有效的治疗方法，旨在根除这些致命的病毒疾病。

DESCRIPTION (provided by applicant): The human liver serves as the reservoir for several important human pathogens, including hepatitis B (HBV) and C viruses (HCV) and Plasmodium species, all of which represent serious global health concerns. HBV and HCV alone chronically infect an estimated 500 million people worldwide, with annual deaths totaling more than 1.5 million. Chronic HBV and HCV infections can have severe health consequences, including hepatitis, cirrhosis, liver failure, and hepatocellular carcinoma. Co-infection with HBV and HCV is common, and leads to a significantly worse prognosis. A preventative vaccine for HBV exists, but curative treatments targeting the virus are not available. Furthermore, increasing HBV resistance has been reported to first-line antiviral drugs. A vaccine for HCV has not yet been developed, and, while HCV-specific protease and polymerase inhibitors are showing promise in early clinical development, rapid emergence of resistance indicates that additional targets and combinations of antivirals will be needed for effective control. The scarcity of in vitro and in vivo systems that faithfully mimic liver biology and susceptibility to human hepatotropic pathogens has severely hampered drug and vaccine development. Here, we propose to take an interdisciplinary approach and combine tissue engineering with molecular virology and humanized mouse technology to create platforms that will facilitate studies of basic virus-host and virus-virus interactions, promote understanding of the mechanisms of liver disease progression, and provide predictive systems to test drug and vaccine efficacy and toxicity. Specifically, we aim to characterize HBV and HCV biology and model associated liver disease in micropatterned primary human hepatocyte cultures (MPCCs) - a breakthrough technology that was recently developed in Dr. Sangeeta Bhatia's laboratory. We will extend this methodology to develop three-dimensional liver organoids, and investigate HBV and HCV infection in mice transplanted with these structures. Since chronic inflammation plays a significant role in liver disease progression, we aim to incorporate components of the immune system in static and dynamic tissue culture systems and ultimately to use mice reconstituted with a human hematopoietic system recipients for HBV/HCV-permissive liver organoids. Through the development and use of these platforms, we hope to shed light on HBV and HCV virology and pathogenesis, and to uncover novel avenues for therapeutic intervention. PUBLIC HEALTH RELEVANCE: Hepatitis B and C viruses are leading causes of human liver disease including cancer. This proposal aims to create and utilize novel primary human hepatocyte tissue culture systems and complex 3D liver organoids to study HCV and HBV mono- and co-infections. The resulting findings should aid development of more effective treatments aimed at eradicating these deadly viral diseases.