title: Treatment of phenylketonuria using minicircle-based naked-DNA gene transfer to murine liver.
abstract: Host immune response to viral vectors, persistence of nonintegrating vectors, and sustained transgene expression are among the major challenges in gene therapy. To overcome these hurdles, we successfully used minicircle (MC) naked-DNA vectors devoid of any viral or bacterial sequences for the long-term treatment of murine phenylketonuria, a model for a genetic liver defect. MC-DNA vectors expressed the murine phenylalanine hydroxylase (Pah) complementary DNA (cDNA) from a liver-specific promoter coupled to a de novo designed hepatocyte-specific regulatory element, designated P3, which is a cluster of evolutionary conserved transcription factor binding sites. MC-DNA vectors were subsequently delivered to the liver by a single hydrodynamic tail vein (HTV) injection. The MC-DNA vector normalized blood phenylalanine concomitant with reversion of hypopigmentation in a dose-dependent manner for more than 1 year, whereas the corresponding parental plasmid did not result in any phenylalanine clearance. MC vectors persisted in an episomal state in the liver consistent with sustained transgene expression and hepatic PAH enzyme activity without any apparent adverse effects. Moreover, 14-20% of all hepatocytes expressed transgenic PAH, and the expression was observed exclusively in the liver and predominately around pericentral areas of the hepatic lobule, while there was no transgene expression in periportal areas.
  
  
  CONCLUSIONS
  This study demonstrates that MC technology offers an improved safety profile and has the potential for the genetic treatment of liver diseases.
authors.affiliation: Division of Metabolism, Department of Pediatrics, University of Zurich, Zurich, Switzerland; and affiliated with the Children's Research Center Zurich.
authors.forename: Hiu Man
authors.initials: HM
authors.lastname: Viecelli
authors.forename: Richard P
authors.initials: RP
authors.lastname: Harbottle
authors.forename: Suet Ping
authors.initials: SP
authors.lastname: Wong
authors.forename: Andrea
authors.initials: A
authors.lastname: Schlegel
authors.forename: Marinee K
authors.initials: MK
authors.lastname: Chuah
authors.forename: Thierry
authors.initials: T
authors.lastname: VandenDriessche
authors.forename: Cary O
authors.initials: CO
authors.lastname: Harding
authors.forename: Beat
authors.initials: B
authors.lastname: Thöny
mesh_terms: D000818:Animals; D004278:DNA, Superhelical; D004195:Disease Models, Animal; D005260:Female; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D008099:Liver / Q000201:enzymology*; D008297:Male; D008810:Mice, Inbred C57BL; D010649:Phenylalanine / Q000097:blood; D010651:Phenylalanine Hydroxylase / Q000378:metabolism; D010661:Phenylketonurias / Q000628:therapy*; D011401:Promoter Regions, Genetic
chemical_list: D004278:DNA, Superhelical; D010649:Phenylalanine; D010651:Phenylalanine Hydroxylase