290. Demonstration of Antitumor Efficacy with an Oncolytic Adenovirus CG8840 for the Treatment of Bladder Cancer in an Orthotopic Tumor Model in Mice

Abstract

Cancer of the bladder is a good target for oncolytic virotherapy because the bladder has several unique features that allow for facile instillation of virus with minimal systemic exposure. CG8840 is a conditionally replication competent oncolytic adenovirus (OAV) designed to preferentially replicate and kill uroplakin II-positive bladder tumors. CG8840 replicates in and kills bladder transitional carcinoma cells (TCC) in vitro, with highly attenuated cytotoxicity in other cell types and human primary cells. In vivo, CG8840 elicits reproducible and dose-dependent anti-tumor effects in subcutaneous human xenograft tumor models in nude mice. We have developed an orthotopic bladder tumor model in nude mice to closely mimic the location and behavior of bladder tumors in humans. Employing the human bladder transitional cell carcinoma cell line SW780 we established tumors on the luminal surface of the nude mice bladder. In order to visualize the tumor growth and anti-tumor effect following virus infection, the TCC cell line SW780 was stably transduced to express the luciferase gene product. The bladder lumen was initially seeded with 1 × 106 SW780-luc tumor cells in a volume of 100 μl after which the mouth of the urethra was closed with a purse string for 1hr to allow for tumor cell attachment to the bladder wall. Following IP injection of luciferin, the orthotopic tumors were visualized in vivo by luminescence imaging. Measurement of the photon counts emitted from the tumors following luciferin injection allowed us to monitor the growth of the orthotopic tumors. Within 2 weeks tumor size was large enough to initiate efficacy studies. Immuno-histochemical analyses of the orthotopic tumor-bearing bladder with human anti-cytokeratin staining confirmed the human origin of the tumors. Histologically the SW780-Luc orthotopic tumors closely resembled superficial bladder tumors in humans. The need for pretreatment with dodecyl-β-D-maltoside to enhance infectivity of bladder epithelium with adenovirus was demonstrated earlier employing a beta galactosidase-expressing non-replicative adenovirus (Ramesh et al., 2003). For efficient infection of orthotopic tumors with adenovirus similar pretreatment was found to be essential. Following intravesical instillations of the uroplakin-II positive bladder tumor-specific OAV CG8840 we studied the anti-tumor efficacy of the virus. Three doses of CG8840 (1 × 1010 vp/treatment weekly for 3 consecutive weeks) were efficacious in reducing the tumor burden. Similar antitumor activity was not observed following intravesical instillation of a non-bladder tumor specific OAV. Six weeks after the initiation of OAV treatment, immuno-histochemical examination of the bladder showed complete absence of tumor cells in CG8840-treated mice. This simplified technique consistently produced a high incidence of orthotopic tumors in mice. This will be a useful model to conduct intravesical anti-tumor efficacy studies of OAV or other viruses both as single agents and in combination with other therapeutic agents.