Augmenting transgene expression from carcinoembryonic antigen (CEA) promoter via a GAL4 gene regulatory system.
Though extensively studied, the use of tissue- or cell-type-specific promoters to target transgene expression is hampered by their weak activity. We hypothesized that this problem could be addressed by using a GAL4 gene regulatory system, wherein a weak, tissue-specific promoter would drive expression of the GAL4/VP16 fusion protein (GV16), which in turn would transactivate a minimal synthetic promoter, GAL4/TATA (GT), upstream of a transgene. To test this hypothesis, we constructed adenoviral vectors expressing a lacZ or GV16 gene driven by a carcinoembryonic antigen (CEA) promoter (Ad/CEA-LacZ or Ad/CEA-GV16) and evaluated levels of transgene expression they produced in cultured cells and in subcutaneous tumors after intratumoral administration. In CEA-positive cells, treatment with Ad/CEA-GV16 + Ad/GT-LacZ versus Ad/CEA-LacZ increased transgene expression 20- to 100-fold. In CEA-negative cells, treatment with Ad/CEA-GV16 + Ad/GT-LacZ increased transgene expression to a much lower degree (6- to 8-fold). In addition, analysis of Bax gene-mediated cell death revealed that this system can be used to avoid Bax's toxic effects on CEA-negative cells without compromising its ability to kill CEA-positive cells in vitro and in vivo. Thus, the combination of a tissue-specific promoter with the GAL4 gene regulatory system could be useful for targeting transgene expression.
Koch
P E
PE
Section of Thoracic Molecular Oncology, Department of Thoracic, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
Guo
Z S
ZS
Kagawa
S
S
Gu
J
J
Roth
J A
JA
Fang
B
B
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D002272:Carcinoembryonic Antigen / Q000235:genetics*; D002460:Cell Line; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000235:genetics* / Q000494:pharmacology; D005786:Gene Expression Regulation; D005809:Genes, Regulator; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics*; D006367:HeLa Cells; D017405:Herpes Simplex Virus Protein Vmw65 / Q000235:genetics; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009369:Neoplasms / Q000235:genetics; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011993:Recombinant Fusion Proteins / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D016385:TATA Box; D014157:Transcription Factors / Q000235:genetics* / Q000494:pharmacology; D019076:Transgenes; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; C494827:Bax protein, mouse; D002272:Carcinoembryonic Antigen; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D017405:Herpes Simplex Virus Protein Vmw65; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D051028:bcl-2-Associated X Protein