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• W2023705807 abstract "Gene therapy for neurodegenerative diseases depends critically on the vector system to direct sustained and stable expression of the transgene. It is, however, a commonly observed phenomenon that transgene expression from currently available vectors is down-regulated following ex vivo gene transfer to the central nervous system (CNS). In an attempt to circumvent this problem, we have systematically evaluated the potential of different cis-acting elements to increase and stabilize transgene expression in vitro and after grafting of engineered cell lines to the CNS.Plasmid vector constructs incorporating Woodchuck hepatitis post-transcriptional regulatory element (WPRE), cHS4 insulator elements and/or the translational enhancer element SP163 were produced. Stable, polyclonal cultures of HiB5 cells were generated by transfection with reporter constructs, and in vitro transgene mRNA and protein levels were determined. Finally, HiB5 clones engineered to express the enhanced green fluorescent protein (EGFP) were grafted to the rat striatum and expression levels were evaluated.Inserting the WPRE element downstream of the open reading frame (ORF) of a reporter gene and flanking the transcriptional unit with cHS4 insulator elements significantly increased protein and mRNA expression levels. Surprisingly, the SP163 element, previously reported to be a translational enhancer, apparently did not promote any translational enhancing activity. Furthermore, the SP163 element exerted a negative effect on transcription. The ability of cHS4 and WPRE elements to stabilize in vivo transgene expression was demonstrated by transplantation of HiB5 clones containing expression constructs into the rat striatum.The data suggest that incorporating cis-acting elements in gene therapy vectors may result in improvements to currently available therapeutic vectors." @default.
• W2023705807 created "2016-06-24" @default.
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• W2023705807 date "2003-07-21" @default.
• W2023705807 modified "2023-10-15" @default.
• W2023705807 title "Increased<i>in vitro</i>and<i>in vivo</i>transgene expression levels mediated through<i>cis</i>-acting elements" @default.
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• W2023705807 doi "https://doi.org/10.1002/jgm.444" @default.
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