Massive apoptotic cell death in chemically induced rat urinary bladder carcinomas following in situ HSVtk electrogene transfer.
BACKGROUND
Gene delivery in current gene therapy studies relies largely on recombinant viral vectors. However, the safety of this method is still under investigation. The effectiveness of in vivo electrogene transfer as a means of gene therapy for rat bladder cancers using the herpes simplex virus 1 thymidine kinase (HSVtk) gene in combination with ganciclovir (GCV) was therefore investigated.


METHODS
The killing effects of HSVtk/GCV therapy were evaluated in transitional cell carcinoma (TCC) cells in vitro and in vivo. In animal experiments, electrogene transfer of HSVtk into N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN)-induced rat bladder tumors was conducted followed by GCV administration.


RESULTS
In vitro studies demonstrated that approximately 50-70% of the TCC cells died as a result of transfection with pHSVtk and GCV administration and that this treatment was associated with decreased DNA synthesis and elevated activities of caspase-3, -8 and -9. A significantly decreased mitochondrial membrane potential was also noted in TCC cells given pHSV tk + GCV. A direct single injection of HSVtk into bladder tumors using in vivo electrogene transfer followed by GCV i.p. administration resulted in significant increases in the levels of apoptosis and histopathological necrosis accompanied by marked inflammation. Active caspase-3 was strongly expressed in the cell death areas of the TCC in rats given pHSVtk/GCV therapy.


CONCLUSIONS
In vivo electrogene transfer results in efficient gene transfer in BBN-induced rat bladder tumors and the HSVtk/GCV prodrug system induces significant cell death which appears to be, at least, mediated via the mitochondrial apoptotic pathway.
Shibata
Masa-Aki
MA
Department of Anatomy and Biology, Osaka Medical College, 2-7, Daigaku-machi, Takatsuki, Osaka 569-8686, Japan.
Horiguchi
Taisuke
T
Morimoto
Junji
J
Otsuki
Yoshinori
Y
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D002277:Carcinoma / Q000503:physiopathology* / Q000628:therapy; D018274:Electroporation; D005260:Female; D015774:Ganciclovir / Q000494:pharmacology*; D018014:Gene Transfer Techniques; D018259:Herpesvirus 1, Human / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2; D051381:Rats; D011916:Rats, Inbred F344; D013937:Thymidine Kinase / Q000235:genetics*; D001749:Urinary Bladder Neoplasms / Q000503:physiopathology* / Q000628:therapy; D051028:bcl-2-Associated X Protein
D000998:Antiviral Agents; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D013937:Thymidine Kinase; D015774:Ganciclovir