IMPROVED METHODS FOR IN VIVO HEPATIC GENE THERAPY

逆转录病毒载体介导的肝脏基因治疗可以永久 纠正代谢紊乱，如鸟氨酸转氨甲基酶 血友病或血栓症等缺陷。我们最近做了 证明了稳定的和治疗水平的两种表达 在大鼠体内可以获得血浆蛋白。然而，两者的风险都 体外和活体给药方法目前限制了其应用 对人类的基因治疗。体内给药系统涉及门静脉 肝再生期间注射逆转录病毒载体。多数 研究人员使用70%的肝部分切除来诱导肝细胞 基于MoMLV的转导所必需的复制 逆转录病毒载体。这些研究的目标是减少或消除 体内逆转录病毒载体传递方法对啮齿动物的毒性研究 还有猪。存在诱导肝细胞复制的生长因子 体外和体内。这些生长因子将用于啮齿动物 通过腺病毒载体或S纯化的蛋白。腺病毒载体 会导致肝脏的短期表达，因为免疫 对腺病毒转导细胞的反应。在某些情况下， 生长因子的管理将与门户网站相结合 分支闭塞，这应该会使肝脏对 较低剂量的生长因子。门静脉分支闭塞诱导 阻断肝组织中肝细胞的凋亡及其代偿性 肝细胞在非闭塞肝组织中的复制。的影响 这些对肝细胞复制和肝功能的治疗将是 下定决心。它们促进逆转录病毒载体转导的能力 将通过向门静脉注射逆转录病毒载体进行评估 在肝细胞复制高峰期，并确定 报告基因的表达水平。如果实验成功了 在啮齿动物身上，我们将尝试使用这些方法来诱导 肝细胞复制和促进逆转录病毒载体转导 猪。这里提出的实验可能会确定一个可以接受的 将逆转录病毒载体输送到慢性肝炎患者肝脏的程序 遗传性疾病。

Retroviral vector-mediated hepatic gene therapy could permantly correct metabolic disorders such as ornithine transcarbamylase deficiencies such as hemophilia or thrombophilia. We have recently demonstrated that stable and therapeutic levels of expression of two plasma proteins can be achieved in rats. However, the risks of both ex vivo and in vivio delivery methods currently limit the application of gene therapy to humans. In vivo delivery systems involve portal vein injection of retroviral vector during liver regeneration. Most investigators use a 70% partial hepatectomy to induce the hepatocyte replication that is necessary for transduction with a MoMLV-based retroviral vector. The goal of these studies is to decrease or eliminate the toxicity of the in vivo retroviral vector delivery methods in rodents and pigs. Growth factors exist that induce hepatocyte replication in vitro and invivo. These growth factors will be administered to rodents via a adenoviral vector or a s purified protein. The adenoviral vector will result in short-term expression in the liver because of the immune response to adenoviral-transduced cells. In some cases, the administration of the growth factor will be combined with portal branch occlusion, which should make the liver more responsive to lower doses of the growth factors. Portal branch occlusion induces apoptosis of hepatocytes from the occluded liver, and compensatory replication of hepatocytes in the non-occluded liver. the effect of these treatments upon hepatocyte replication and liver function will be determined. Their ability to facilitate retroviral vector transduction will be assessed by injecting a retroviral vector into the portal vein during the peak period of hepatocyte replication, and determining the level of expression of the reporter gene. If experiments are successful in rodents, we will attempt to use these approaches to induce hepatocyte replication and facilitate retroviral vector transduction in pigs. The experiments proposed here might identify an acceptable procedure for delivering retroviral vectors to livers of patients with genetic disorders.