Cristiano R J, Smith L C, Kay M A, Brinkley B R, Woo S L
Department of Cell Biology, Howard Hughes Medical Institute, Baylor College of Medicine, Houston, TX 77030.
Proc Natl Acad Sci U S A. 1993 Dec 15;90(24):11548-52. doi: 10.1073/pnas.90.24.11548.
Receptor-mediated endocytosis is an effective method for gene delivery into target cells. We have previously shown that DNA molecules complexed with asialoglycoprotein can be efficiently endocytosed by primary hepatocytes and the internalized DNA can be released from endosomes by the use of a replication-defective adenovirus. Because the DNA and virus enter target cells independently, activity enhancement requires high concentrations of adenoviral particles. In this study, adenoviral particles were chemically conjugated to poly(L-lysine) and bound ionically to DNA molecules. Quantitative delivery to primary hepatocytes was achieved with significantly reduced viral titer when the asialoorosomucoid-poly(L-lysine) conjugate was included in the complex. The conjugated adenovirus was used to deliver a DNA vector containing canine factor IX to mouse hepatocytes, resulting in the expression of significant concentrations of canine factor IX in the culture medium. The results suggest that receptor-mediated endocytosis coupled with an efficient endosomal lysis vector should permit the application of targeted and efficient gene delivery into the liver for gene therapy of hepatic deficiencies.
受体介导的内吞作用是一种将基因导入靶细胞的有效方法。我们之前已经表明，与去唾液酸糖蛋白复合的DNA分子能够被原代肝细胞有效内吞，并且通过使用复制缺陷型腺病毒，内化的DNA可以从内体中释放出来。由于DNA和病毒独立进入靶细胞，因此活性增强需要高浓度的腺病毒颗粒。在本研究中，腺病毒颗粒通过化学方法与聚-L-赖氨酸偶联，并与DNA分子离子结合。当复合物中包含去唾液酸血清类黏蛋白-聚-L-赖氨酸偶联物时，以显著降低的病毒滴度实现了对原代肝细胞的定量递送。将偶联的腺病毒用于向小鼠肝细胞递送含有犬因子IX的DNA载体，结果在培养基中表达出显著浓度的犬因子IX。这些结果表明，受体介导的内吞作用与有效的内体裂解载体相结合，应该能够实现靶向且高效的基因递送，用于肝脏缺陷的基因治疗。
