Hepatic gene therapy: adenovirus enhancement of receptor-mediated gene delivery and expression in primary hepatocytes.
We have combined a receptor-mediated DNA delivery system with the endosomal lysis ability of adenovirus and shown that DNA can be delivered into primary hepatocytes, resulting in a high level of gene expression. When asialoorosomucoid conjugated with poly(L-lysine) was used to deliver the Escherichia coli beta-galactosidase gene into primary hepatocytes through binding with the hepatic asialoglycoprotein receptor, only a low level of beta-galactosidase was detectable, with less than 0.1% of the hepatocytes being transfected. This level of activity can be greatly enhanced by the cointernalization of the DNA.protein complex with a replication-defective adenovirus, resulting in 100% of the hepatocytes staining blue with 5-bromo-4-chloro-3-indolyl beta-D-galactoside. Quantitative analysis of beta-galactosidase expression also showed a 1000-fold enhancement of activity. To test the applicability of this DNA delivery system for the correction of phenylketonuria, a metabolic disorder that causes severe mental retardation in children, we have delivered the human phenylalanine hydroxylase (PAH) gene to hepatocytes derived from a PAH-deficient mouse strain and demonstrated complete reconstitution of enzymatic activity. This method shows great promise for efficient gene delivery to the liver for correction of hepatic disorders.
Department of Cell Biology, Baylor College of Medicine, Houston, TX 77030.
R J
RJ
Cristiano
L C
LC
Smith
S L
SL
Woo
D016475:3T3 Cells; D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins / Q000378:metabolism; D001692:Biological Transport; D006528:Carcinoma, Hepatocellular; D002478:Cells, Cultured; D004247:DNA / Q000008:administration & dosage / Q000235:genetics*; D004337:Drug Carriers; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000378:metabolism / Q000502:physiology*; D008113:Liver Neoplasms; D051379:Mice; D008810:Mice, Inbred C57BL; D008817:Mice, Mutant Strains; D009961:Orosomucoid / Q000031:analogs & derivatives / Q000378:metabolism; D010651:Phenylalanine Hydroxylase / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D010661:Phenylketonurias / Q000235:genetics / Q000628:therapy*; D011971:Receptors, Immunologic / Q000378:metabolism*; D014162:Transfection / Q000379:methods; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D004337:Drug Carriers; D009961:Orosomucoid; D011971:Receptors, Immunologic; C017030:asialoorosomucoid; D004247:DNA; D010651:Phenylalanine Hydroxylase; D001616:beta-Galactosidase