title: Enhanced transgene expression in the mouse skeletal muscle infected by the adeno-associated viral vector with the human elongation factor 1alpha promoter and a human chromatin insulator.
abstract: BACKGROUND
  Efficient and continuous expression of a therapeutic transgene is a key factor for improving the efficacy of gene therapy. Some insulators are known to contribute to continuous high-level expression of a therapeutic transgene.
  
  
  METHODS
  Using the human AAVS1 insulator (DHS) found in the AAVS1 DNase I hypersensitive site, chicken beta-globin insulator (cHS4) and sea urchin arylsufatase insulator (Ars), we newly constructed three recombinant adeno-associated virus vectors (rAAV) and examined their capability of transducing the mouse quadriceps muscle.
  
  
  RESULTS
  DHS increased transgene expression from the human elongation factor 1alpha promoter (EF) by 1000-fold, up to the high level achieved by the human cytomegalovirus immediate early promoter/enhancer (CMV), which comprises an extremely strong promoter for driving a transgene. cHS4 enhanced the expression by 100-fold, whereas Ars did not. The enhanced expression was maintained for at least 24 weeks. Vector copy numbers were similar with and without DHS or cHS4; thus, the enhancement is most likely due to up-regulated transcription. Neither DHS, nor cHS4 affected transgene expression from CMV. DHS enhanced expression from the human muscle creatine kinase promoter/enhancer by 100-fold in mice, as did DHS from EF.
  
  
  CONCLUSIONS
  Although DHS was unable to further enhance high expression from the strong viral enhancer/promoter, it enhanced low expression from the human promoters by 100- to 1000-fold. Thus, DHS may be useful for constructing rAAVs that express a therapeutic transgene from less efficient, tissue specific promoters.
authors.affiliation: Center for Pathogen Genomics, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Mayuyo
authors.initials: M
authors.lastname: Mori-Uchino
authors.forename: Takamasa
authors.initials: T
authors.lastname: Takeuchi
authors.forename: Isao
authors.initials: I
authors.lastname: Murakami
authors.forename: Tetsu
authors.initials: T
authors.lastname: Yano
authors.forename: Toshiharu
authors.initials: T
authors.lastname: Yasugi
authors.forename: Yuji
authors.initials: Y
authors.lastname: Taketani
authors.forename: Keiichi
authors.initials: K
authors.lastname: Nakagawa
authors.forename: Tadahito
authors.initials: T
authors.lastname: Kanda
mesh_terms: D000818:Animals; D002460:Cell Line; D000229:Dependovirus / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D038101:Insulator Elements; D051379:Mice; D018482:Muscle, Skeletal / Q000166:cytology / Q000502:physiology*; D020648:Peptide Elongation Factor 1 / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D019076:Transgenes
chemical_list: D020648:Peptide Elongation Factor 1