title: Non-viral, integrin-mediated gene transfer into fibroblasts from patients with lysosomal storage diseases.
abstract: BACKGROUND
  Non-viral vectors consisting of Lipofectin/integrin-targeting peptide/DNA (LID) complexes have great potential for gene therapy, as they are safe, simple, and able to package large DNA molecules. In this study, these vectors were evaluated in vitro for the therapy of lysosomal storage disorders.
  
  
  METHODS
  Non-viral vectors were designed to deliver therapeutic genes by integrin-mediated uptake into fibroblasts from patients with the lysosomal storage disorders fucosidosis and Fabry disease, which result from deficiencies of alpha-L-fucosidase and alpha-galactosidase A, respectively. The vectors consisted of a complex (LID) of Lipofectin and a peptide containing an integrin-targeting domain and a poly-lysine domain to which was bound plasmid DNA, containing alpha-L-fucosidase (LID-alpha-Fuc) or alpha-galactosidase A (LID-alpha-Gal).
  
  
  RESULTS
  Patients' fibroblasts transfected with LID-alpha-Fuc and LID-alpha-Gal produced the corresponding enzyme at levels which were 10-40% of the total activity in cultures of normal fibroblasts. However, 95-98% of this activity was secreted. Transfection of endothelial cells, the main target cells in Fabry disease, with an LID-alpha-Gal produced a total alpha-galactosidase activity 65% higher than that in untransfected cultures after 6 days, 67% of the activity being secreted. Although transfection of fibroblasts with LID complexes also caused small changes in the distribution of endogenous lysosomal enzymes, it did not appear to affect the viability of the cells.
  
  
  CONCLUSIONS
  The integrin-mediated transfer of genes encoding lysosomal enzymes into cells results in the secretion of large amounts of normal enzyme that could be taken up by other cells. This could be a useful strategy for enzyme-replacement therapy.
authors.affiliation: Biochemistry, Endocrinology and Metabolism Unit, Institute of Child Health at Great Ormond Street Hospital, University College London, England, UK. ewhite@ich.ucl.ac.uk
authors.forename: E J
authors.initials: EJ
authors.lastname: Estruch
authors.forename: S L
authors.initials: SL
authors.lastname: Hart
authors.forename: C
authors.initials: C
authors.lastname: Kinnon
authors.forename: B G
authors.initials: BG
authors.lastname: Winchester
mesh_terms: D002460:Cell Line; D000795:Fabry Disease / Q000201:enzymology*; D005260:Female; D005347:Fibroblasts / Q000201:enzymology; D005645:Fucosidosis / Q000201:enzymology*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D016023:Integrins / Q000235:genetics / Q000378:metabolism*; D008081:Liposomes; D008156:Luciferases / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010957:Plasmids; D000519:alpha-Galactosidase / Q000235:genetics* / Q000378:metabolism; D005644:alpha-L-Fucosidase / Q000235:genetics* / Q000378:metabolism
chemical_list: D016023:Integrins; D008081:Liposomes; D010446:Peptide Fragments; D010714:Phosphatidylethanolamines; C020888:1,2-dielaidoylphosphatidylethanolamine; D008156:Luciferases; D000519:alpha-Galactosidase; D005644:alpha-L-Fucosidase