Targeted killing of carcinoembryonic antigen (CEA)-producing cholangiocarcinoma cells by polyamidoamine dendrimer-mediated transfer of an Epstein-Barr virus (EBV)-based plasmid vector carrying the CEA promoter.
The present study reports a novel nonviral method to efficiently and specifically target carcinoembryonic antigen (CEA)-producing cholangiocarcinoma (CC) cells in vitro. Epstein-Barr virus (EBV)-based and conventional plasmid vectors were constructed that possess the beta-galactosidase (beta-gal) or herpes simplex virus-1 (HSV-1) thymidine kinase (Tk) genes as well as tandem repeats of the human genomic sequence -82 to -42 bp from the transcriptional start site of the CEA gene. The plasmids were transfected by means of polyamidoamine dendrimer into CEA-positive (HuCC-T1) or -negative cell lines. Transfection of the conventional plasmid vector with the CEA promoter and beta-gal gene resulted in a very low or undetectable level of marker gene expression even in the CEA-positive cell line. Transferring the HSV-1 Tk gene by conventional plasmid did not affect the susceptibility of HuCC-T1 cells to ganciclovir. In marked contrast, strong beta-gal expression was specifically obtained in HuCC-T1 cells by transfecting the EBV-based plasmid in which the CEA promoter and a ubiquitous promoter (SRalpha) are employed to drive the EBV-encoded nuclear antigen 1 (EBNA1) and beta-gal genes, respectively (pTES.beta). Furthermore, CEA-positive but not -negative tumor cells were rendered highly susceptible to ganciclovir when transfected with the EBV-based vector that carries the CEA promoter-EBNA1 and SRalpha-HSV-1 Tk genes (pTES.Tk). These results strongly suggest that the EBV-based plasmid vector/cationic polymer system (EBV/polyplex) equipped with the CEA promoter provides an efficient nonviral method for the targeted gene therapy of CEA-producing malignancies.
Tanaka
S
S
Third Department of Internal Medicine, Kyoto Prefectural University of Medicine, Kamikyo, Japan.
Iwai
M
M
Harada
Y
Y
Morikawa
T
T
Muramatsu
A
A
Mori
T
T
Okanoue
T
T
Kashima
K
K
Maruyama-Tabata
H
H
Hirai
H
H
Satoh
E
E
Imanishi
J
J
Mazda
O
O
D002272:Carcinoembryonic Antigen / Q000235:genetics / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D018281:Cholangiocarcinoma / Q000378:metabolism / Q000473:pathology / Q000628:therapy*; D003110:Colonic Neoplasms / Q000378:metabolism / Q000473:pathology / Q000628:therapy*; D004396:Coloring Agents; D017931:DNA Primers / Q000737:chemistry; D019309:Epstein-Barr Virus Nuclear Antigens / Q000235:genetics*; D015774:Ganciclovir / Q000494:pharmacology; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors; D006801:Humans; D010078:Oxazines; D011073:Polyamines / Q000627:therapeutic use*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics*; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects; D014966:Xanthenes; D001616:beta-Galactosidase / Q000378:metabolism
D002272:Carcinoembryonic Antigen; D004396:Coloring Agents; D017931:DNA Primers; D019309:Epstein-Barr Virus Nuclear Antigens; D010078:Oxazines; D011073:Polyamines; D014966:Xanthenes; C005843:resazurin; D001616:beta-Galactosidase; D015774:Ganciclovir