特异性导向肝细胞表达和复制的EB病毒载体系统的组建

发展了一种高效的、能将携带外源基因的质粒型ＥＢ病毒载体特异性地导入肝细胞表达的方法。将ＥＢ病毒复制子载体ｐＤＲ２经改造得到携带外源基因的ｐＥＢｌｕｃ质粒。将ｐＥＢｌｕｃ与人工制备的、既保留了ＤＮＡ结合活性又能为肝细胞受体识别并内在化的半乳糖基化组蛋白的ｆｌ组分结合，形成ｐＥＢｌｕｃ－半乳糖基化组蛋白复合物。该复合物通过脱唾液酸糖蛋白受体介导的内吞进入肝细胞并表达。ｐＥＢｌｕｃ质粒在细胞内能稳定存在并自主复制。该系统具有应用于以肝细胞为靶组织的基因治疗的前景。

THE CONSTRUCTION OF A NOVEL HEPATOCYTE TARGETED AND AUTONOMOUSLY REPLICATED GENEDELIVERYSYSTEM WITH AN EPSTEIN-BARR VIRUS BASED EXPRESSION VECTOR

An artificial virus like DNA-protein complex was constructed with an autonomously replicated vector based Epstein-Barr virus replicon and galactosylated histone.This complex could be specially uptaken by hepatocytes through the asialoglycoprotein receptor mediated endocytosis pathway.The plasmid pEBluc,an EBV based expression vector harboring the luciferase report gene,was used to transfect different human cell lines,and only in the hepatic cell lines,the luciferase activity could be detected.Compared with the transfection mediated vialiposome,the report gene expressed not only in hepatoma cells,but also in non-hepatic cell lines.The result of the Southern-blotting of the DNA extracted from the cells transfected by this kind of DNA/protein complex illustrated that the pEBluc existed and replicated as an episome.