Herpes simplex virus thymidine kinase gene therapy in experimental rat BT4C glioma model: Effect of the percentage of thymidine kinase-positive glioma cells on treatment effect, survival time, and tissue reactions

Abstract

Herpes simplex virus thymidine kinase (HSV-tk) gene transfer and ganciclovir (GCV) administration have been suggested for the treatment of malignant gliomas. To understand tissue responses and possible ways to improve the treatment effect, we studied tumor growth, tissue reactions, and survival time after HSV-tk/GCV treatment in a syngeneic BT4C rat glioma model by mixing various ratios of stably transfected HSV-tk-expressing BT4C-tk glioma cells with wild-type BT4C glioma cells (percentage of BT4C-tk cells: 0%, 1%, 10%, 30%, 50%, and 100%), followed by injection into BDIX rat brains (n ϭ 79). With the exception of some animals with end-stage tumors, very little astroglia or microglia reactivity was detected in the wild-type tumors as analyzed by immunocytochemistry using glial fibrillary acid protein (GFAP)-, vimentin-, human histocompatibility leukocyte antigen-DR-, OX-42-, and CD68-specific monoclonal antibodies. After 14 days of GCV treatment, tumors induced with Ն10% BT4C-tk cells showed a significant reduction in tumor size (P Ͻ .05) and prolonged survival time (P Ͻ .01). Astrogliosis, as indicated by a strong GFAP and vimentin immunoreactivity, was seen in the tumor scar area. GFAP and vimentin reactivity was already present after the GCV treatment in tumors induced with 1% BT4C-tk cells. Much less human histocompatibility leukocyte antigen-DR-positive microglia was seen in the treated animals, indicating low microglia reactivity and immunoactivation against the tumor. However, GCV-treated tumors were positive for apoptosis, indicating that apoptosis is an important mechanism for cell death in the BT4C-tk glioma model. Our results suggest that Ն10% transfection efficiency is required for a successful reduction in BT4C glioma tumor size with HSV-tk/GCV treatment in vivo. Tissue reactions after 14 days of GCV treatment are characterized by astrogliosis and apoptosis, whereas microglia response and immunoactivation of the brain cells appear to play a minor role. Stimulation of the microglia response by gene transfer or other means might improve the efficacy of the HSV-tk/GCV treatment in vivo. Cancer Gene Therapy (2000) 7, 413-421

Truncated vesicular stomatitis virus G protein improves baculovirus transduction efficiency in vitro and in vivo

“…Rats were killed on day 4 and perfused with PBS intracardially and fixed with X-gal fixative (4% PFA in phosphate buffer, pH 7.2) for 30 min. 19 After the fixative, the brains were rinsed for 2 h in PBS and imbedded in OCT.…”

Targeting of biotinylated compounds to its target tissue using a low-density lipoprotein receptor–avidin fusion protein

“…BT4C rat glioma cells (10 000 cells/5 ml) were implanted intracranially in the right corpus callosum at a depth of 2.5 mm at the following coordinates: 1 mm caudal to bregma and 2 mm right to sutura sagittalis. 28,38 After 2 weeks of cell inoculations, Av-fusion protein/SFV virus in 0.9% NaCl, or empty SFV (control), was injected by Hamilton syringe with a 27-gauge needle into the tumor (see coordinates above) in 10 ml volumes, first into a depth of 2.5 mm during 2 min and then into a depth of 1.5 mm. To prevent the escape of the virus from the needle track, the syringe was held in place for 8 min.…”

HSV-tk gene therapy for human renal cell carcinoma in nude mice

“…The cell line was tested to be free of mycoplasma and other microbiological contaminants. 7 Wild -type Caki -2 cells (Caki-2 /WT ) were stably transduced with replication -deficient HSV-tk 5 and LacZ ( titer 2Â10 6 cfu /mL ) 8 retroviruses using standard technol-ogy. 9,10 Stably transduced cell lines were produced by geneticin selection: Caki -2 +HSV-tk ( Caki -2/ TK ) and Caki-2 +LacZ (Caki-2 /LacZ ).…”