Dual-function vectors for in vivo gene therapy of AAT Liver disease

描述（由申请人提供）：α-1抗胰蛋白酶缺乏症是一种常见的单基因疾病，其中最常见的突变“Z”等位基因（Glu 342 Lys）在北方欧洲人和北美人中发生频率非常高。纯合Z患者患有由于缺乏野生型AAT的正常抗蛋白酶功能而导致的肺病，并且还可能患有肝病，其似乎是由肝细胞内的聚合和/或聚集的Z-AAT蛋白质的保留引发的。表达人Z等位基因的小鼠模型在检查分子终点方面具有实用性，但不能忠实地再现在严重受影响的患者中观察到的炎症和肝硬化。在本申请中，我们建议使用环境挑战和人干细胞衍生的肝嵌合体开发更好的AAT相关肝病模型，并进一步测试组合基因治疗/RNA抑制策略作为潜在的未来治疗。我们的第一个目标是在具有人类Z等位基因的小鼠中使用乙醇诱导肝毒性，以评估肝细胞癌风险增加的风险，并更好地了解这些肝脏中的促炎和促纤维化途径。然后，这一概念将扩展到使用人源化小鼠肝脏与来自iPS细胞的正常或Z等位基因阳性人肝脏干细胞的研究。最后，这两种模型将用于评估基于rAAV的RNAi介导的Z-AAT mRNA敲低的作用。

DESCRIPTION (provided by applicant): Alpha-1 antitrypsin deficiency is a common single-gene disorder, in which the most common mutant "Z" allele (Glu342Lys) occurs with remarkably high frequency in Northern Europeans and North Americans. Homozygous Z patients suffer from a lung disease that is due to the lack of normal antiprotease function of the wild-type AAT, and may also suffer from a liver disease, that appears to be triggered by retention of polymeric and/or aggregated Z-AAT protein within hepatocytes. A mouse model expressing the human Z-allele has utility in examining molecular endpoints but does not faithfully reproduce the inflammation and cirrhosis observed in severely affected patients. In this application, we propose to develop better models of AAT-related liver disease using environmental challenges and human stem cell derived liver chimeras, and further to test a combined gene therapy/RNA inhibition strategy as a potential future therapy. Our first aim will use an ethanol induce hepatotoxicity in mice with the human Z allele to assess the risk of increased risk hepatocellualr carcinoma and to better understand the pro-inflammatory and pro-fibrotic pathways in these livers. This concept will then be extended to studies using humanized mouse livers with either normal or Z-allele positive human liver stem cells derived from iPS cells. Finally both of these models will be utilized to assess the effect of rAAV-based RNAi mediated knockdown of the Z-AAT mRNA.