Recombinant AAV-mediated HSVtk gene transfer with direct intratumoral injections and Tet-On regulation for implanted human breast cancer

Li Zi-Bo,Hu Wei-Xin,Zeng Zhao-Jun,Chen Qian,Luo Sai-Qun

doi:10.1186/1471-2407-6-66

Abstract

BackgroundHSVtk/ganciclovir (GCV) gene therapy has been extensively studied in tumors and relies largely on the gene expression of HSVtk. Most studies, however, have failed to demonstrate any significant benefit of a controlled gene expression strategy in cancer treatment. The Tet-On system is commonly used to regulate gene expression following Dox induction. We have evaluated the antitumor effect of HSVtk/ganciclovir gene therapy under Tet-On regulation by means of adeno-associated virus-2 (AAV-2)-mediated HSVtk gene transfer with direct intratumoral injections in mice bearing breast cancer tumors.MethodsRecombinant adeno-associated virus-2 (rAAV) was constructed and transduced into MCF-7 cell line. GCV treatment to the rAAV infected MCF-7 cells was performed by MTT assay under the doxycycline (Dox) induction or without Dox induction at a vp (viral particle) number of ≥104 /cell. The virus was administered intratumorally to nude mice that had also received GCV intraperitoneally. The antitumor effects were evaluated by measuring tumor regression and histological analysis.ResultsWe have demonstrated that GCV treatment to the infected MCF-7 cells under the Dox induction was of more inhibited effects than those without Dox induction at ≥104 vp/cell. In ex vivo experiments, tumor growth of BALB/C nude mice breast cancer was retarded after rAAV-2/HSVtk/Tet-On was injected into the tumors under the Dox induction. Infiltrating cells were also observed in tumors after Dox induction followed by GCV treatment and cells were profoundly damaged. The expression of HSVtk gene in MCF-7 cells and BALB/C nude mice tumors was up-regulated by Tet-On under Dox induction with reverse transcription-PCR (RT-PCR) analysis.ConclusionThe antitumor effect of rAAV-mediated HSVtk/GCV gene therapy under the Dox induction with direct intratumoral injections may be a useful treatment for breast cancer and other solid tumors.

Highlights

HSVtk/ganciclovir (GCV) gene therapy has been extensively studied in tumors and relies largely on the gene expression of HSVtk
We explored the treatment of human breast cancer by associated virus-2 (AAV-2) expressing HSVtk gene under the regulation of Tet-On, the Recombinant adeno-associated virus-2 (rAAV) was constructed and transduced into MCF-7 cell line or BALB/c nude mice
Almost no difference was found between MCF-7 cells survival as rAAV infection reached a plateau when rAAV titres ranged from 104 to 106vp/cell

Summary

Introduction

HSVtk/ganciclovir (GCV) gene therapy has been extensively studied in tumors and relies largely on the gene expression of HSVtk. The last approach, which can offer the prospect of selectively introducing genes into cancer cells, rendering them susceptible to specific antitumor drugs, is especially appealing for this bystander effect [9]. This is significant since the high therapeutic index that can be achieved with a high concentration of toxic drug will occur only at the tumor site. GCV, a kind of antiviral drug, can be phosphorylated by HSVtK protein into triphosphates, which are potent inhibitors of DNA polymerase, leading to the disruption of cellular DNA synthesis and cell death These properties make the system attractive in cancer therapy [9,10]

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