Abstract
ØC31 integrase can integrate targeted plasmid DNA into preferred locations in mammalian genomes, resulting in robust, long-term expression of the integrated transgene. This system represents an effective tool that opens up promising possibilities for gene therapy. The classical treatment for hypoparathyroidism was calcium and vitamin D replacement. Recently, parathyroid hormone (PTH) replacement was reported to be a more potentially physiologic treatment option. However, PTH synthesis is technically difficult and costly. These issues may be minimized by using PTH gene therapy. We attempted to achieve site-specific genomic integration of the PTH gene into a human cell line and mice using this system. We cotransfected 293 HEK cells with PTH-attB plasmid with or without ØC31 integrase plasmid. Expression and secretion of PTH into culture supernatants and site-specific genomic integration of PTH cDNA were assessed by immunoradiometric assays and pseudo-site analysis, respectively. In in vivo experiments, we injected the PTH-attB plasmid with or without ØC31 integrase plasmid into a mouse tail vein using the hydrodynamic method. Plasma PTH concentrations were serially measured, and site-specific integration of PTH cDNA into the mouse genome was confirmed by examining hepatic genomic DNA. PTH was expressed and secreted from 293 HEK cells and mouse hepatocytes, and pseudo-site analysis confirmed the site-specific integration of PTH cDNA into the host genomes. The site-specificity and efficiency of this system are advantageous in many areas, including, potentially, gene therapy. PTH gene therapy is one candidate; however, for clinical applications, we need to regulate PTH expression and secretion in the future.
Original language | English |
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Pages (from-to) | 1033-1041 |
Number of pages | 9 |
Journal | Endocrine Journal |
Volume | 55 |
Issue number | 6 |
DOIs | |
State | Published - 2008 |
Keywords
- ØC31 integrase
- Gene therapy
- Hypoparathyroidism
- Parathyroid hormone