445. CG5757, an Oncolytic Adenovirus for the Treatment of Retinoblstoma (Rb) Pathway-Defective and Telomerase-Positive Cancers

Abstract

Replication-selective oncolytic viruses hold promise for the treatment of cancer. Among this novel group of therapeutics are oncolytic adenoviruses engineered with tumor-specific transcriptional response elements (TRE) controlling essential genes. These vectors replicate selectively in cancer cells, leading to expression of toxic viral products and oncolysis mediated by viral replication. We and others have compared different transcriptional control strategies by placing one or more tumor-specific TREs upstream of different viral genes including E1A, E1B and E4. Relocation of the viral packaging signal was also investigated. One such oncolytic virus, CG5757, was generated by replacing the E1A and E1B endogenous promoters with promoters derived from the human E2F-1 and telomerase reverse transcriptase (hTERT) genes, respectively. The E2F-1 promoter is activated in Rb-defective tumor types, a pathway mutated in approximately 85% of all cancers. Likewise, telomerase is aberrantly expressed in over 90% of tumors. CG5757 also has a deletion in the coding region of the E1B 19k gene, a Bcl2-like viral antiapoptotic protein, to increase vector cytotoxicity. CG5757 shows strong tumor selectivity. In vitro, expression of E1A and E1B genes was highly restricted to Rb-defective and hTERT-positive cancer cells, including Hep3B (hepatocellular carcinoma), LoVo (colorectal carcinoma), A549 (lung cancer), Panc-1 (pancreatic cancer), 253J B-V (bladder cancer), and Hela (cervical cancer). In normal cells, including a lung fibroblast cell line (WI-38) and several other human primary cell lines (HRE, BSMC, PrEC, HMEC and HMVEC-L), no E1 expression could be detected from infection with CG5757. The transcriptional control of E1 gene expression also correlated with selective viral replication in target cells. CG5757 replicates similarly to wild-type virus in tumor cells, but its replication is, on average, 1,000-times less efficient in normal cells. In a viral cytotoxicity assay, CG5757 destroys tumor cells 100- to 10,000-times more efficiently than normal cells. Comparisons of the cytoxicity of CG5757 in tumor cells versus normal cells (normalized for transduction efficiency with wild-type adenovirus 5) yielded high selectivity indices, some of which were greater than 1000. In vivo, strong antitumor activity was seen using CG5757 in NCR nude mice with subcutaneous lung cancer (A549) and bladder transitional cell carcinoma (253J B-V) xenografts. With respect to the 253J B-V model, four weeks after treatment the average tumor volume in animals treated with four consecutive daily intratumoral injections of CG5757 (4×108 particles/mm3 of tumor) decreased to 72% of baseline while the control group had an increase to 944% of baseline. Furthermore, 50% of treated animals had complete regression of the 253J B-V tumor xenografts. The potential therapeutic efficacy of such dual promoter controlled oncolytic adenoviruses in cancers that are Rb-defective and hTERT-positive has been demonstrated.