Kosovac, D. and Wild, J. and Ludwig, C. and Meissner, S. and Bauer, A. P. and Wagner, R. (2011) Minimal doses of a sequence-optimized transgene mediate high-level and long-term EPO expression in vivo: challenging CpG-free gene design. GENE THERAPY, 18 (2). pp. 189-198. ISSN 0969-7128,
Full text not available from this repository. (Request a copy)Abstract
Advanced gene delivery techniques can be combined with rational gene design to further improve the efficiency of plasmid DNA (pDNA)-mediated transgene expression in vivo. Herein, we analyzed the influence of intragenic sequence modifications on transgene expression in vitro and in vivo using murine erythropoietin (mEPO) as a transgene model. A single electro-gene transfer of an RNA-and codon-optimized mEPOopt gene into skeletal muscle resulted in a 3- to 4-fold increase of mEPO production sustained for >1 year and triggered a significant increase in hematocrit and hemoglobin without causing adverse effects. mEPO expression and hematologic levels were significantly lower when using comparable amounts of the wild type (mEPOwt) gene and only marginal effects were induced by mEPO Delta CpG lacking intragenic CpG dinucleotides, even at high pDNA amounts. Corresponding with these observations, in vitro analysis of transfected cells revealed a 2- to 3-fold increased (mEPOopt) and 50% decreased (mEPO Delta CpG) erythropoietin expression compared with mEPOwt, respectively. RNA analyses demonstrated that the specific design of the transgene sequence influenced expression levels by modulating transcriptional activity and nuclear plus cytoplasmic RNA amounts rather than translation. In sum, whereas CpG depletion negatively interferes with efficient expression in postmitotic tissues, mEPOopt doses <0.5 mu g were sufficient to trigger optimal long-term hematologic effects encouraging the use of sequence-optimized transgenes to further reduce effective pDNA amounts. Gene Therapy (2011) 18, 189-198; doi:10.1038/gt.2010.134; published online 14 October 2010
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | PLASMID DNA; SKELETAL-MUSCLE; MAMMALIAN-CELLS; ERYTHROPOIETIN PRODUCTION; AUTOIMMUNE ANEMIA; NAKED DNA; THERAPY; ELECTROPORATION; DELIVERY; ELECTROTRANSFER; codon optimization; CpG dinucleotides; in vivo transfection; electro-gene transfer; erythropoietin; transgene expression |
| Subjects: | 600 Technology > 610 Medical sciences Medicine |
| Divisions: | Medicine > Lehrstuhl für Medizinische Mikrobiologie und Hygiene |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 26 Jun 2020 09:30 |
| Last Modified: | 26 Jun 2020 09:30 |
| URI: | https://pred.uni-regensburg.de/id/eprint/21327 |
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