Ligand-dependent regulation of vascular endothelial growth factor and erythropoietin expression by a plasmid-based autoinducible GeneSwitch system.
We investigated the ability of an improved mifepristone-dependent GeneSwitch system to regulate the expression of genes for two therapeutic proteins: vascular endothelial growth factor (VEGF) and erythropoietin. The GeneSwitch system consisted of two plasmids, one encoding the chimeric GeneSwitch protein, the other an inducible transgene. When the constitutive CMV promoter of the GeneSwitch plasmid was replaced by an autoinducible promoter consisting of four copies of GAL4 DNA binding sites linked to a minimal thymidine kinase promoter, the tightness of transgene regulation was improved by an order of magnitude. Quantitative RT-PCR analysis of GeneSwitch mRNA confirmed that the autoinducible promoter was responsive to mifepristone. We demonstrated the ability of the improved GeneSwitch system to regulate the expression of VEGF or erythropoietin in a biologically relevant manner after delivery of plasmids to the hind-limb muscle of adult mice. This ability of the autoinducible GeneSwitch system to regulate the expression of therapeutic proteins in mice indicates its potential for use in human gene therapy applications.
Abruzzese
R V
RV
Valentis, Inc, The Woodlands, Texas, 77381, USA.
Godin
D
D
Mehta
V
V
Perrard
J L
JL
French
M
M
Nelson
W
W
Howell
G
G
Coleman
M
M
O'Malley
B W
BW
Nordstrom
J L
JL
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D016228:Endothelial Growth Factors / Q000235:genetics*; D004921:Erythropoietin / Q000235:genetics*; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D008024:Ligands; D008222:Lymphokines / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D015735:Mifepristone / Q000494:pharmacology; D018482:Muscle, Skeletal / Q000378:metabolism; D010957:Plasmids; D011401:Promoter Regions, Genetic; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D017931:DNA Primers; D016228:Endothelial Growth Factors; D008024:Ligands; D008222:Lymphokines; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D004921:Erythropoietin; D015735:Mifepristone