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Dual-therapeutic reporter genes fusion for enhanced cancer gene therapy and imaging

Abstract

Two of the successful gene-directed enzyme prodrug therapies include herpes simplex virus–thymidine kinase (HSV1–TK) enzyme-ganciclovir prodrug and the Escherichia coli nitroreductase (NTR) enzyme-CB1954 prodrug strategies; these enzyme-prodrug combinations produce activated cytotoxic metabolites of the prodrugs capable of tumor cell death by inhibiting DNA synthesis and killing quiescent cells, respectively. Both these strategies also affect significant bystander cell killing of neighboring tumor cells that do not express these enzymes. We have developed a dual-combination gene strategy, where we identified HSV1-TK and NTR fused in a particular orientation can effectively kill tumor cells when the tumor cells are treated with a fusion HSV1-TK–NTR gene– along with a prodrug combination of GCV and CB1954. In order to determine whether the dual-system demonstrate superior therapeutic efficacy than either HSV1-TK or NTR systems alone, we conducted both in vitro and in vivo tumor xenograft studies using triple negative SUM159 breast cancer cells, by evaluating the efficacy of cell death by apoptosis and necrosis upon treatment with the dual HSV1-TK genes-GCV-CB1954 prodrugs system, and compared the efficiency to HSV1-TK–GCV and NTR-CB1954. Our cell-based studies, tumor regression studies in xenograft mice, histological analyses of treated tumors and bystander studies indicate that the dual HSV1-TK–NTR–prodrug system is two times more efficient even with half the doses of both prodrugs than the respective single gene-prodrug system, as evidenced by enhanced apoptosis and necrosis of tumor cells in vitro in culture and xenograft of tumor tissues in animals.

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Acknowledgements

We thank Dr Srabani Bhaumik from GE Global Research for her support, and providing CytoCy5S substrate for imaging NTR enzyme. We also thank Dr Tim Doyle for support in providing training for the use of different Imaging instruments from Stanford Small Animal Imaging Facility. We thank Dr Sanjiv Sam Gambhir, Chairman, Department of Radiology, Stanford University for providing helpful support and facility for conducting this research. We thank Dr Chaitali Banerjee for her input in preparing this paper. We thank the department of Radiology, Stanford University for funding support (Ramasamy Paulmurugan), and the Canary center at Stanford for providing the facilities.

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Correspondence to R Paulmurugan.

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Sekar, T., Foygel, K., Willmann, J. et al. Dual-therapeutic reporter genes fusion for enhanced cancer gene therapy and imaging. Gene Ther 20, 529–537 (2013). https://doi.org/10.1038/gt.2012.66

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