In vivo electroporetic transfer ofbcl-2 antisense oligonucleotide inhibits the development of hepatocellular carcinoma in rats

Abstract

To investigate the potential use of a bcl-2 antisense oligonucleotide for therapy against hepatocellular carcinoma, we examined the effects of the electroporetic transfer of a bcl-2 antisense oligonucleotide on rat hepatocarcinogenesis induced by N-nitrosomorpholine (NNM). Sprague-Dawley rats were given water containing 175 mg/l NNM for 8 weeks and received intraperitoneal injections of a bcl-2 antisense phosphorothioate oligonucleotide, a sense oligonucleotide or a scrambled sequence oligonucleotide encapsulated in empty liposomes, at a dose of 150 microg oligonucleotide/kg body weight, every 4 weeks. One hour after injection, in vivo electroporation was performed on the liver to achieve selective transfer of the oligonucleotides. By week 16, the rats that had received the bcl-2 antisense oligonucleotide had significantly fewer and smaller precancerous liver lesions positive for glutathione-S-transferase (placental type), and a significantly lower incidence of hepatocellular carcinoma in the electroporation zone than rats that had received the sense or the scrambled oligonucleotides. Moreover, the bcl-2 antisense oligonucleotide significantly increased the apoptotic indices in foci, neoplastic nodules and in hepatocellular carcinomas. The expression of bcl-2 mRNA also decreased, and 3'-fragments of bcl-2mRNA produced by cleavage at the antisense target site were detected in rat liver. Mean cellular fluorescence in the liver increased with higher doses of fluorescein-isothiocyanate-labeled antisense or sense oligonucleotides. Our results show that the electroporetic transfer of bcl-2 antisense oligonucleotide can inhibit rat hepatocarcinogenesis.