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.
Section of Thoracic Molecular Oncology, Department of Thoracic, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
P E
PE
Koch
Z S
ZS
Guo
S
S
Kagawa
J
J
Gu
J A
JA
Roth
B
B
Fang
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