A Novel Cancer-Targeting Adenovirus Expressing Interferon Alpha For Pancreatic Cancer Therapy

Abstract

Introduction: More effective systemic therapy is clearly needed for pancreatic adenocarcinoma. Interferon alpha (IFNα) is promising in multimodality therapy, but has a short half-life and strong side effects. We have generated a novel oncolytic adenovirus using several strategies to enhance efficacy while reducing toxicity. The COX2 promoter is used to drive viral replication due to known overexpression in pancreatic tumor cells, while avoiding replication toxicity to normal organs, e.g., liver. Our design uses the Ad5/Ad3 fiber-knob chimera for optimal infectivity, and adenoviral death protein for enhanced apoptosis and viral spread. Moreover, our oncolytic adenovirus system allows massive transgene expression in target cells. We hypothesize that localized IFNα expression from oncolytic adenovirus will enhance anti-tumor effect with minimal toxicity. Methods: Qualitative assessment of effect was done with crystal violet staining at serial time points. Quantitative cell viability was assessed colorimetrically using a commercially-available kit. COX2 expression in cell lines was assessed by RT-PCR, and promoter function by a luciferase gene reporter assay with known COX2-positive and negative controls. IFNα expression levels were quantitiated by ELISA from supernatant. In vivo, nude mice were injected on each flank with 1×10^6 S2O13 tumor cells, then intratumorally with virus at 1×10^10 vp/50μl or PBS when tumors were 5-8mm in size. Results: in vitro a robust effect on highly aggressive metastatic pancreatic cancer S2 cell lines is seen. When compared to Ad5 wt, the standard control virus without cancer specificity, the new virus has a similarly potent effect on S2VP10, and is more powerful than Ad5 wt when used on S2O13. Our novel virus is also more powerful than the conventional non-replicating IFNα virus. Analysis of COX2 expression in pancreatic cancer cell lines shows that both S2 cell lines are COX2-positive. Our novel virus shows no toxicity to COX2-negative BT474 cells to day 8 post infection, indicating cancer specificity. IFNα expression is dependent on viral replication and reaches a much higher concentration using the novel virus than from the non-replicating IFNα virus. In vivo experiments show a strong suppression of tumor growth from the new virus (Fig 1). Conclusions: A new cancer-targeting oncolytic adenovirus was successfully designed to selectively replicate in cancer cells with a powerful antitumor effect. The effect of the targeted vector was as strong as the viruses without replication selectivity. By expressing IFNα locally in the tumor environment, toxicity from systemic delivery seen in current therapy should be avoided. We are actively working in an in vivo syngeneic model using immunocompetent hamsters.