Combined suicide gene and immunostimulatory gene therapy using AAV-mediated gene transfer to HPV-16 transformed mouse cell: decrease of oncogenicity and induction of protection.
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.
Janousková
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Department of Experimental Virology, Institute of Hematology and Blood Transfusion, 128 00 Prague 2, Czech Republic. olga@uhkt.cz
Síma
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Kunke
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D000818:Animals; D000998:Antiviral Agents / Q000627:therapeutic use*; D018122:B7-1 Antigen / Q000235:genetics*; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor / Q000637:transplantation; D002472:Cell Transformation, Viral; D018932:Chemokine CCL2 / Q000235:genetics*; D003673:Defective Viruses / Q000235:genetics*; D000229:Dependovirus / Q000235:genetics*; D015774:Ganciclovir / Q000627:therapeutic use*; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics / Q000627:therapeutic use*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D007167:Immunotherapy / Q000379:methods*; D007668:Kidney; D007763:Lac Operon; D051379:Mice; D008810:Mice, Inbred C57BL; D009368:Neoplasm Transplantation; D009856:Oncogene Proteins, Viral / Q000502:physiology*; D027383:Papillomaviridae / Q000502:physiology*; D050725:Papillomavirus E7 Proteins; D012097:Repressor Proteins; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics; D013937:Thymidine Kinase / Q000037:antagonists & inhibitors / Q000235:genetics*; D014161:Transduction, Genetic; D023041:Xenograft Model Antitumor Assays
D000998:Antiviral Agents; D018122:B7-1 Antigen; D018932:Chemokine CCL2; C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D012097:Repressor Proteins; C059731:oncogene protein E7, Human papillomavirus type 16; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D013937:Thymidine Kinase; D015774:Ganciclovir