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Combined suicide gene and immunostimulatory gene therapy using AAV-mediated gene transfer to HPV-16 transformed mouse
O Janousková1, P Síma, D Kunke
1Department of Experimental Virology, Institute of Hematology and Blood Transfusion, 128 00 Prague 2, Czech Republic. olga@uhkt.cz
Abstract:
To test the effects of combined transduction of a suicide gene and genes coding for various immunostimulatory factors on the oncogenicity and immunogenicity of TC-1 cells (HPV-16 transformed C57BL/6 mouse cells), several bicistronic recombinant adeno-associated viruses (rAAV) were constructed. Each of these constructs carried, and in infected cells expressed, the herpes simplex type 1 thymidine-kinase gene (HSV-TK) and the gene of one of the following immunostimulatory factors: human monocyte chemoattractant protein 1 (MCP-1), mouse B7.1 costimulatory molecule (B7.1), or mouse granulocyte-macrophage colony-stimulating factor (GM-CSF). For control purposes, an rAAV carrying the HSV-TK gene and neomycin resistance gene (neo) and an rAAV containing the lacZ gene were used. All of these constructs proved functional both in mouse TC-1 and human 293T cells. For experiments in mice, TC-1 cells were infected in vitro with the AAV recombinants at an input multiplicity of 50 particles/cell; these cells were then administered to 5-week-old mice. As from day 5, half of the animals were given ganciclovir (GCV) (2.5 mg/day) for 10 days. With a single exception, none of the mice inoculated with cells treated with rAAV expressing HSV-TK + B7.1 or HSV-TK + MCP-1 developed tumour irrespective of GCV treatment. The tumour suppressive effect was less marked in animals inoculated with TC-1 cells infected with rAAV expressing HSV-TK + GM-CSF, and among these it was somewhat more pronounced in GCV-untreated animals. A clear antitumour effect of GCV treatment was only observed in mice inoculated with TC-1 cells transduced with rAAV expressing HSV-TK but no immunostimulatory factor. Mice that remained tumour-free on day 54 were challenged with untreated TC-1 cells. The tumour resistance rates found were related not only to the immunostimulatory gene used for the transduction, but also to GCV treatment. The best protection was recorded in mice pre-inoculated with TC-1 cells transduced with either B7.1 or MCP-1-expressing rAAV and not given GCV.
Insights
Combining suicide gene therapy with immunostimulatory factors like B7.1 or MCP-1 in TC-1 cells effectively suppressed tumor growth in mice. The best tumor resistance was observed without ganciclovir treatment, highlighting the potential of these combined immunotherapies.
Area of Science:
- Oncology
- Immunotherapy
- Gene Therapy
Background:
- TC-1 cells, derived from HPV-16-transformed C57BL/6 mouse cells, are used to study tumor development and potential treatments.
- Recombinant adeno-associated viruses (rAAV) are effective vectors for delivering therapeutic genes.
- Suicide gene therapy (using HSV-TK) combined with immunostimulatory factors can enhance anti-tumor immune responses.
Purpose of the Study:
- To evaluate the combined effects of suicide gene (HSV-TK) and immunostimulatory factor (MCP-1, B7.1, GM-CSF) transduction via rAAV on TC-1 cell oncogenicity and immunogenicity.
- To assess the impact of ganciclovir (GCV) treatment on the anti-tumor efficacy of these combined gene therapies.
Main Methods:
- Construction of bicistronic rAAV vectors encoding HSV-TK and one of three immunostimulatory factors (MCP-1, B7.1, GM-CSF) or control genes (neo, lacZ).
- In vitro transduction of TC-1 cells with rAAV vectors.
- Inoculation of mice with transduced TC-1 cells, followed by administration of ganciclovir (GCV) to a subset of animals.
- Tumor monitoring and challenge with untreated TC-1 cells in tumor-free survivors.
Main Results:
- TC-1 cells transduced with rAAV expressing HSV-TK + B7.1 or HSV-TK + MCP-1 prevented tumor formation in most mice, regardless of GCV treatment.
- Tumor suppression was less pronounced with HSV-TK + GM-CSF, with better results in GCV-untreated animals.
- GCV treatment showed a clear anti-tumor effect only when TC-1 cells were transduced with HSV-TK alone.
- Mice that rejected tumors showed enhanced resistance to subsequent challenge with untreated TC-1 cells, with the best protection seen in mice treated with B7.1 or MCP-1 expressing rAAV and not given GCV.
Conclusions:
- Combined transduction of suicide genes and immunostimulatory factors, particularly B7.1 and MCP-1, demonstrates significant potential in suppressing tumor growth and enhancing anti-tumor immunity.
- The efficacy of combined gene therapy is influenced by the specific immunostimulatory factor used and the administration of ganciclovir.
- This strategy offers a promising approach for developing novel cancer immunotherapies.