IMPROVED ADENOVIRAL VECTORS FOR HEPATIC GENE THERAPY

这项研究的目标是开发出改良的腺病毒 可用于将基因转移至肝脏以进行治疗的载体 肝脏缺陷 虽然腺病毒介导的基因转移， 在动物模型中，体内肝细胞是非常有效的， 因为基因表达的瞬时性和不能 以进行重复转导。具体目标是：1）制定一个 E4基因功能缺陷的重组腺病毒载体。的E4 缺陷型载体除了为克隆外源基因提供更多的空间外， 应该导致转导细胞中较少内源性腺病毒基因表达 细胞因此，对宿主细胞基因的干扰可能较少 调节和载体产生抗原的可能性较小， 最终将限制体内转导细胞的寿命。（二） 建立抗原特异性免疫应答在a）丧失 和B）不能在肝细胞中表达腺病毒介导的基因， 为了在腺病毒介导的肝细胞转染后实现肝细胞的二次转导， 体内基因转移这些免疫反应的描述将允许 非免疫原性载体的合理设计。3)探讨 腺病毒E3区编码的单个蛋白在 在体内肝细胞基因转导后重组腺病毒载体。 某些E3基因产物的组成性高水平基因表达可能 保护转导细胞免受免疫介导的破坏。我们建议 直接在体内解决这些问题。4)研究腺病毒介导的 将基因转移到胆管上皮细胞中，以开发这种方法， 治疗多种疾病。 5)评估胆道 道作为体内将基因递送到肝细胞中的手段。的该模式 基因递送具有优于血管递送途径的潜在优势 并将被探索作为当前方法的替代方案。 该项目的成功完成将具有普遍的应用， 目前腺病毒作为载体靶向的所有细胞类型 用于基因转移。使用肝脏作为靶器官将有一般 申请基因疗法治疗大量遗传病 由肝脏缺陷引起的。

The goal of the research proposed here is to develop improved adenoviral vectors that can be used for gene transfer to the liver for the treatment of hepatic deficiencies. Although adenoviral-mediated gene transfer to hepatocytes in vivo is very efficient in animal models, improvements are needed because of the transient nature of gene expression and the inability to perform repeat transduction. The specific aims are to: 1) Develop a recombinant adenoviral vector deficient in E4 gene function. The E4 deficient vectors besides allowing more room for cloning exogenous genes, should lead to less endogenous adenoviral gene expression in transduced cells. As a result, there may be less interference with host cell gene regulation and less probability that the vector will produce antigens that ultimately will limit the life-span of transduced cells in vivo. 2) Establish the role of the antigen-specific immune response in a) the loss of adenoviral-mediated gene expression in hepatocytes and b) the inability to achieve secondary transduction of hepatocytes after adenoviral-mediated gene transfer in vivo. Delineation of these immune responses will allow for rational design of vectors which are non-immunogenic. 3) Investigate the significance of individual proteins encoded in the adenoviral E3 region in recombinant adenoviral vectors after in vivo hepatocyte gene transduction. Constitutive high level gene expression of some of the E3 gene products may protect transduced cells from immune-mediated destruction. We propose to directly address these issues in vivo. 4) Investigate adenoviral-mediated gene transfer into biliary epithelial cells to develop this as a method for the treatment of a number of medical disorders. 5) Evaluate the biliary tract as a means of delivering genes into hepatocytes in vivo. This mode of gene delivery has potential advantages over the vascular routes of delivery and will be explored as an alternative to current methods. The successful completion of this project will have general applications to all cell types that are currently being targeted by adenovirus as a vehicle for gene transfer. Using the liver as a target organ will have general application for gene therapy for a large number of genetic disorders resulting from hepatic deficiencies.