A Hypoxia- and α-Fetoprotein-Dependent Oncolytic Adenovirus Exhibits Specific Killing of Hepatocellular Carcinomas

Purpose: Oncolytic adenoviruses (Ad) constitute a new promising modality of cancer gene therapy that displays improved efficacy over nonreplicating Ads. We have previously shown that an E1B 19-kDa-deleted oncolytic Ad exhibits a strong cell-killing effect but lacks tumor selectivity. To achieve hepatoma-restricted cytotoxicity and enhance replication of Ad within the context of tumor microenvironment, we used a modified human alpha-fetoprotein (hAFP) promoter to control the replication of Ad with a hypoxia response element (HRE). Experimental Design: We constructed Ad-HRE6/hAFP Delta 19 and Ad-HRE12/hAFP Delta 19 that incorporated either 6 or 12 copies of HRE upstream of promoter. The promoter activity and specificity to hepatoma were examined by luciferase assay and fluorescence-activated cell sorting analysis. In addition, the AFP expression-and hypoxia-dependent in vitro cytotoxicity of Ad-HRE6/hAFP Delta 19 and Ad-HRE12/hAFP Delta 19 was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and cytopathic effect assay. In vivo tumoricidal activity on subcutaneous and liver orthotopic model was monitored by noninvasive molecular imaging. Results: Ad-HRE12/hAFP Delta 19 exhibited enhanced tumor selectivity and cell-killing activity when compared with Ad-hAFP Delta 19. The tumoricidal activity of Ad-HRE12/hAFP Delta 19 resulted in significant inhibition of tumor growth in both subcutaneous and orthotopic models. Histologic examination of the primary tumor after treatment confirmed accumulation of viral particles near hypoxic areas. Furthermore, Ad-HRE12/hAFP Delta 19 did not cause severe inflammatory immune response and toxicity after systemic injection. Conclusions: The results presented here show the advantages of incorporating HREs into a hAFP promoter-driven oncolytic virus. This system is unique in that it acts in both a tissue-specific and tumor environment-selective manner. The greatly enhanced selectivity and tumoricidal activity of Ad-HRE12/hAFP Delta 19 make it a promising therapeutic agent in the treatment of liver cancers. Clin Cancer Res; 16(24); 6071-82. (C) 2010 AACR.