Related Experiment Video
Updated: Aug 11, 2026
Production and Purification of Non Replicative Canine Adenovirus Type 2 Derived Vectors
Published on: December 3, 2013
Vesicular stomatitis virus G pseudotyped retrovector mediates effective in vivo suicide gene delivery in experimental
1Department of Medicine (Division of Hematology-Oncology), Lady Davis Institute for Medical Research, Montreal, Quebec, Canada. jgalipea@lab.jgh.mcgill.ca
Abstract:
Direct in vivo tumor-targeting with "suicide" viral vectors is limited by either inefficient gene transfer (i.e., retroviral vectors) or indiscriminate transfer of a conditionally toxic gene to surrounding nonmalignant tissue (i.e., adenoviral vectors). Retrovectors pseudotyped with the vesicular stomatitis virus G protein (VSVG) may serve as a remedy to this conundrum. These retroviral particles differ from standard murine retroviruses by their very broad tropism and the capacity to be concentrated by ultracentrifugation without loss of activity. We propose that a VSVG-typed retrovector can be used for efficient and tumor-specific herpes simplex virus thymidine kinase (TK) gene delivery in vivo. To test this hypothesis, we developed a bicistronic retroviral vector that expresses TK and green fluorescence protein (pTKiGFP). The 293GPG packaging cell line was used to generate vTKiGFP retroparticles. In cytotoxicity assays, vTKiGFP-transduced human glioma cell lines were sensitized to the cytotoxic effects of gangciclovir (GCV) 10,000-fold. Subsequently, virus was concentrated by ultracentrifugation to a titer of 2.3 x 10(10) cfu/ml. We tested the antitumor activity of vTKiGFP retroparticles in a rat C6 glioma model of brain cancer. Concentrated retrovector stock (9 microl volume) was injected stereotactically in preestablished intracerebral tumor. Subsequently, rats were treated with GCV for 10 days. Control rats (no GCV) had a mean survival of 38 days (range, 20-52 days). Sections performed on postmortem brain tissue revealed large tumors with evidence of high efficiency retrovector transfer and expression (as assessed by GFP fluorescence). Fluorescence was restricted to malignant tissue. In the experimental group (GCV treated), 8 of 12 remain alive and well >120 days after glioma implantation. In conclusion, vTKiGFP is very efficient at transducing human glioma cell lines in vitro and leads to significant GCV sensitization. Recombinant retroviral particles can be concentrated to titers that allow in vivo intratumoral delivery of large viral doses. The therapeutic efficiency of this reagent has been demonstrated in a preclinical model of brain cancer.
Insights
This study developed a novel vesicular stomatitis virus G protein (VSVG)-pseudotyped retrovector for efficient and tumor-specific gene delivery. This enhanced retroviral vector shows significant promise for brain cancer therapy by sensitizing glioma cells to ganciclovir (GCV).
Area of Science:
- Oncology
- Gene Therapy
- Virology
Background:
- Direct in vivo tumor targeting with viral vectors faces challenges like inefficient gene transfer or off-target effects.
- Vesicular stomatitis virus G protein (VSVG)-pseudotyped retrovectors offer broad tropism and high concentration capacity, potentially overcoming these limitations.
Purpose of the Study:
- To develop and evaluate a VSVG-pseudotyped retrovector for efficient and tumor-specific delivery of the herpes simplex virus thymidine kinase (TK) gene in vivo.
- To assess the therapeutic efficacy of this retrovector in a preclinical brain cancer model.
Main Methods:
- A bicistronic retroviral vector (vTKiGFP) expressing TK and green fluorescence protein (GFP) was engineered.
- Packaging cell lines generated concentrated retroparticles, which were tested in human glioma cell lines and a rat C6 glioma brain cancer model.
- Tumor-specific gene transfer and therapeutic efficacy were assessed by ganciclovir (GCV) treatment and survival analysis.
Main Results:
- vTKiGFP efficiently transduced human glioma cells in vitro, sensitizing them 10,000-fold to GCV.
- Concentrated retroparticles (2.3 x 10(10) cfu/ml) demonstrated high efficiency and tumor-specific gene transfer in a rat brain cancer model, evidenced by GFP fluorescence restricted to malignant tissue.
- GCV treatment of rats receiving intratumoral retrovector injection resulted in 8 of 12 animals surviving beyond 120 days, significantly improving survival compared to controls.
Conclusions:
- VSVG-pseudotyped retrovectors enable efficient in vitro gene transfer and significant ganciclovir sensitization of glioma cells.
- Recombinant retroviral particles can be concentrated to high titers for effective in vivo intratumoral delivery.
- This therapeutic strategy demonstrated significant preclinical efficacy in a brain cancer model, highlighting its potential for clinical translation.
