title: Episomal maintenance of S/MAR-containing non-viral vectors for RPE-based diseases.
abstract: The efficacy of non-viral genetic therapies has historically been limited by transient gene expression and vector loss. Scaffold matrix attachment regions (S/MARs) have been shown to augment transcription, promote episomal maintenance, and provide insulator-like function to DNA in in vitro and in vivo systems. Here we explore the ability of S/MAR elements to mediate these effects in retinal pigment epithelial (RPE) cells with the eventual goal of improving the persistence of expression of our non-viral gene delivery tools. We engineered an RPE-specific reporter vector with or without an S/MAR immediately downstream of the eGFP expression cassette. We show that the S/MAR vector is maintained as an episome for up to 1 year. Experiments in which rhodamine-labeled DNA was delivered to the subretinal space of mice show better persistence of the S/MAR-containing vector in the RPE than the non-S/MAR vector. These results suggest that inclusion of the S/MAR region promotes episomal maintenance of plasmid DNA in the RPE after subretinal delivery and that inclusion of this DNA element may be beneficial for non-viral ocular gene transfer.
authors.affiliation: Department of Cell Biology, University of Oklahoma Health Sciences Center, 940 Stanton L. Young Blvd., BMSB 781, 73104, Oklahoma City, OK, USA, adarsha_koirala@meei.harvard.edu.
authors.forename: Adarsha
authors.initials: A
authors.lastname: Koirala
authors.forename: Shannon M
authors.initials: SM
authors.lastname: Conley
authors.forename: Muna I
authors.initials: MI
authors.lastname: Naash
mesh_terms: D000818:Animals; D004247:DNA / Q000493:pharmacokinetics; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics; D045170:Matrix Attachment Regions / Q000235:genetics*; D051379:Mice; D008807:Mice, Inbred BALB C; D053758:Nanoparticles / Q000627:therapeutic use*; D010957:Plasmids / Q000235:genetics*; D012162:Retinal Degeneration / Q000235:genetics / Q000628:therapy*; D055213:Retinal Pigment Epithelium / Q000502:physiology; D019745:cis-trans-Isomerases / Q000235:genetics*; D000099178:Retinoid Isomerohydrolase
chemical_list: D004247:DNA; D049452:Green Fluorescent Proteins; D019745:cis-trans-Isomerases; D000099178:Retinoid Isomerohydrolase; C485184:enhanced green fluorescent protein