title: Physiological levels of HBB transgene expression from S/MAR element-based replicating episomal vectors.
abstract: Replicating episomal vectors (REV) are in principle able to provide long-term transgene expression in the absence of integration into the target cell genome. The scaffold/matrix attachment region (S/MAR) located 5' of the human beta-interferon gene (IFNB1) has been shown to confer a stable episomal replication and retention function within plasmid vectors when stably transfected and selected in mammalian cells. The minimal requirement for the IFNB1 S/MAR to function in DNA replication and episomal retention is transcription through this element. We used the erythroid beta-globin locus control region-beta-globin gene (betaLCR-HBB) microlocus cassette as a model to assess tissue-specific expression from within an IFNB1 S/MAR-based plasmid REV. The betaLCR-HBB plus S/MAR combination constructs provided either high or low levels of transcription through the S/MAR element. Our results show that the betaLCR-HBB microlocus is able to reproducibly and stably express at full physiological levels on an episome copy number basis. In addition, our data show that even low levels of transcription from betaLCR-HBB through the S/MAR element are sufficient to allow efficient episomal replication and retention. These data provide the principles upon which generic and flexible expression cassette-S/MAR-based REVs can be designed for a wide range of applications.
authors: King's College London School of Medicine, Nuclear Biology Group, Department of Medical and Molecular Genetics, 8th Floor Tower Wing, Guy's Hospital, London SE1 9RT, UK.
authors: Argyro
authors: A
authors: Sgourou
authors: Samantha
authors: S
authors: Routledge
authors: Dionysios
authors: D
authors: Spathas
authors: Aglaia
authors: A
authors: Athanassiadou
authors: Michael N
authors: MN
authors: Antoniou
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D003001:Cloning, Molecular / Q000379:methods*; D018628:Gene Dosage; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D016899:Interferon-beta / Q000235:genetics / Q000378:metabolism; D020014:K562 Cells; D045170:Matrix Attachment Regions; D051379:Mice; D010957:Plasmids / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D019076:Transgenes; D055544:beta-Globins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism
chemical_list: D055544:beta-Globins; D016899:Interferon-beta