title: Minicircle DNA Provides Enhanced and Prolonged Transgene Expression Following Airway Gene Transfer.
abstract: Gene therapy for cystic fibrosis using non-viral, plasmid-based formulations has been the subject of intensive research for over two decades but a clinically viable product has yet to materialise in large part due to inefficient transgene expression. Minicircle DNA give enhanced and more persistent transgene expression compared to plasmid DNA in a number of organ systems but has not been assessed in the lung. In this study we compared minicircle DNA with plasmid DNA in transfections of airway epithelial cells. In vitro, luciferase gene expression from minicircles was 5-10-fold higher than with plasmid DNA. In eGFP transfections in vitro both the mean fluorescence intensity and percentage of cells transfected was 2-4-fold higher with minicircle DNA. Administration of equimolar amounts of DNA to mouse lungs resulted in a reduced inflammatory response and more persistent transgene expression, with luciferase activity persisting for 2 weeks from minicircle DNA compared to plasmid formulations. Transfection of equal mass amounts of DNA in mouse lungs resulted in a 6-fold increase in transgene expression in addition to more persistent transgene expression. Our findings have clear implications for gene therapy of airway disorders where plasmid DNA transfections have so far proven inefficient in clinical trials.
authors.affiliation: UCL Institute of Child Health, 30 Guilford Street, London, WC1N 1EH, United Kingdom.
authors.forename: Mustafa M
authors.initials: MM
authors.lastname: Munye
authors.affiliation: UCL Institute of Child Health, 30 Guilford Street, London, WC1N 1EH, United Kingdom.
authors.forename: Aristides D
authors.initials: AD
authors.lastname: Tagalakis
authors.affiliation: UCL Respiratory Centre for Inflammation and Tissue Repair, 5 University Street, London, WC1E 6JF, United Kingdom.
authors.forename: Josephine L
authors.initials: JL
authors.lastname: Barnes
authors.affiliation: UCL MRC Laboratory for Molecular Cell Biology, Gower Street, London WC1E 6BT, United Kingdom.
authors.forename: Rachel E
authors.initials: RE
authors.lastname: Brown
authors.affiliation: UCL Respiratory Centre for Inflammation and Tissue Repair, 5 University Street, London, WC1E 6JF, United Kingdom.
authors.forename: Robin J
authors.initials: RJ
authors.lastname: McAnulty
authors.affiliation: UCL Institute of Child Health, 30 Guilford Street, London, WC1N 1EH, United Kingdom.
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Howe
authors.affiliation: UCL Institute of Child Health, 30 Guilford Street, London, WC1N 1EH, United Kingdom.
authors.forename: Stephen L
authors.initials: SL
authors.lastname: Hart
mesh_terms: D000818:Animals; D056444:Axonemal Dyneins / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism; D004270:DNA, Circular / Q000235:genetics*; D004847:Epithelial Cells / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D049409:Luciferases, Firefly / Q000235:genetics* / Q000378:metabolism; D008168:Lung / Q000378:metabolism*; D051379:Mice; D010957:Plasmids; D014162:Transfection; D019076:Transgenes
chemical_list: D016207:Cytokines; D004270:DNA, Circular; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D049409:Luciferases, Firefly; D056444:Axonemal Dyneins; C497507:DNAH5 protein, human