Human Hepatocyte and Discovery Core

项目摘要/摘要 血友病A的基因治疗有望实现持久治愈。腺相关病毒基因 将凝血因子FVIII转移到男性肝脏可以实现治疗纠正，但这是不可持续的。 临床试验中肝毒性和FVIII表达下降的原因尚不清楚。AAV疗法 目前正在研究的靶标是肝脏中的肝细胞。然而，人们对这一问题的了解有限 载体与肝细胞的相互作用以及FVIII表达和免疫应答的影响。 CORE B将提供主要的人类肝细胞模型，以解决这些基本的和机械的相关问题 AAV和FVIII的生物学特性。这一提议的中心假设是多个相互关联的特征 AAV和FVIII的生物学特性限制了治疗表达的持久性，并造成严重的安全问题。这个 核心B，人类肝细胞和发现核心的目标是提供三个原代人类肝细胞 研究AAV载体和FVIII生物学的模型。第一种模式是保持稳定的原代肝细胞培养。 肝细胞功能的二维格式。这些将用于体外研究。第二种模式是 嵌合小鼠，其肝脏高度重新填充原始人类肝细胞。这些将用于在 活体AAV研究。第三种模型是双重植入造血干细胞和肝脏的小鼠 人体肝脏免疫细胞的活体研究。核心B将向调查人员提供这些服务，以解决 FVIII生物学、血友病的基因治疗、肝脏导向的基因转移以及 AAV载体的分子和免疫生物学。

PROJECT SUMMARY/ABSTRACT Gene therapy for hemophilia A holds promise to accomplish a lasting cure. Adeno-associated viral (AAV) gene transfer of clotting factor FVIII to the livers of males can achieve therapeutic correction but this is not sustained. The reasons for hepatotoxicity and the decline in FVIII expression in clinical trials are unclear. The AAV therapies currently under investigation target hepatocytes in the liver. There is, however, a limited understanding of the interactions between the vector and hepatocytes as well as effect of FVIII expression and immune responses. Core B will provide primary human hepatocyte models to address these basic and mechanistic questions related to the biology of AAV and FVIII. The central hypothesis of this proposal is that multiple interconnected features of AAV and FVIII biology limit durability of therapeutic expression and pose serious safety concerns. The objectives of Core B, the Human Hepatocyte and Discovery Core, is to provide three primary human hepatocytes models to study AAV vector and FVIII biology. The first model is primary hepatocyte cultures that retain stable hepatocyte functions in 2-dimensional format. These will be used for in vitro studies. The second model is chimeric mice whose livers are highly repopulated with primary human hepatocytes. These will be used for in vivo AAV studies. And the third model is mice doubly engrafted with hematopoietic stem cells and livers for in vivo studies of human immune cells in the liver. Core B will provide these services to investigators to address major unanswered questions in FVIII biology, gene therapy for hemophilia, liver-directed gene transfer, and molecular and immunobiology of AAV vectors.