title: Optimization of hCFTR lung expression in mice using DNA nanoparticles.
abstract: Efficient and prolonged human cystic fibrosis transmembrane conductance regulator (hCFTR) expression is a major goal for cystic fibrosis (CF) lung therapy. A hCFTR expression plasmid was optimized as a payload for compacted DNA nanoparticles formulated with polyethylene glycol (PEG)-substituted 30-mer lysine peptides. A codon-optimized and CpG-reduced hCFTR synthetic gene (CO-CFTR) was placed in a polyubiquitin C expression plasmid. Compared to hCFTR complementary DNA (cDNA), CO-CFTR produced a ninefold increased level of hCFTR protein in transfected HEK293 cells and, when compacted as DNA nanoparticles, produced a similar improvement in lung mRNA expression in Balb/c and fatty acid binding protein promoter (FABP) CF mice, although expression duration was transient. Various vector modifications were tested to extend duration of CO-CFTR expression. A novel prolonged expression (PE) element derived from the bovine growth hormone (BGH) gene 3' flanking sequence produced prolonged expression of CO-CFTR mRNA at biologically relevant levels. A time course study in the mouse lung revealed that CO-CFTR mRNA did not change significantly, with CO-CFTR/mCFTR geometric mean ratios of 94% on day 2, 71% on day 14, 53% on day 30, and 14% on day 59. Prolonged CO-CFTR expression is dependent on the orientation of the PE element and its transcription, is not specific to the UbC promoter, and is less dependent on other vector backbone elements.
authors.affiliation: Copernicus Therapeutics, Inc., Cleveland, Ohio 44106, USA.
authors.forename: Linas
authors.initials: L
authors.lastname: Padegimas
authors.forename: Tomasz H
authors.initials: TH
authors.lastname: Kowalczyk
authors.forename: Sam
authors.initials: S
authors.lastname: Adams
authors.forename: Chris R
authors.initials: CR
authors.lastname: Gedeon
authors.forename: Sharon M
authors.initials: SM
authors.lastname: Oette
authors.forename: Karla
authors.initials: K
authors.lastname: Dines
authors.forename: Susannah L
authors.initials: SL
authors.lastname: Hyatt
authors.forename: Ozge
authors.initials: O
authors.lastname: Sesenoglu-Laird
authors.forename: Olena
authors.initials: O
authors.lastname: Tyr
authors.forename: Robert C
authors.initials: RC
authors.lastname: Moen
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Cooper
mesh_terms: D024509:3' Flanking Region; D000281:Administration, Intranasal; D000818:Animals; D003550:Cystic Fibrosis / Q000235:genetics / Q000628:therapy; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics* / Q000378:metabolism; D004247:DNA / Q000737:chemistry*; D004195:Disease Models, Animal; D050556:Fatty Acid-Binding Proteins / Q000235:genetics; D015870:Gene Expression; D005786:Gene Expression Regulation; D023061:Gene Order; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000008:administration & dosage / Q000737:chemistry / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D008168:Lung / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic; D058726:Nanoconjugates / Q000737:chemistry / Q000627:therapeutic use; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D050556:Fatty Acid-Binding Proteins; D058726:Nanoconjugates; D012333:RNA, Messenger; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004247:DNA