Gene Therapy for the Treatment of Malignant Brain Tumors with In Vivo Tumor Transduction with the Herpes Simplex Thymidine Kinase Gene/Ganciclovir System. Iowa Methodist Medical Center, Des Moines, Iowa

Abstract

Murine retroviral vectors can infect a wide variety of proliferating mammalian cell types (e.g. lymphocytes). Non-proliferating tissues (e.g. neurons) are not transduced by murine retroviral vectors. These findings suggest that this type of vector may be useful for the selective introduction of genes into growing tumors in the brain, since the tumor is essentially the only tissue that will integrate and express the vector genes.

A designed equine herpes thymidine kinase (EHV4 TK) variant improves ganciclovir-induced cell-killing

“…Activated GCV thus only causes cell death at the tumor site by DNA damage, and has very little effect on the surrounding cells/tissue. So far, many strategies have targeted the treatment of aggressive malignant brain tumors such as glioblastoma multiforme, however, overall they have had only moderate success in clinical trials [1][2][3][4][5]. Most recently, the HSV1 TK/GCV-based enzyme/prodrug system has been put forward as an efficient strategy for stem cell-based gene therapy [6,7].…”

Superior cytostatic activity of the ganciclovir elaidic acid ester due to the prolonged intracellular retention of ganciclovir anabolites in herpes simplex virus type 1 thymidine kinase gene-transfected tumor cells

“…To investigate the molecular mechanism of improved cytostatic activity of E-GCV over GCV, [8][9][10][11][12][13][14] C]-labeled GCV and E-GCV were exposed to FM3A/0 and FM3Atk − /HSV-1tk + cell cultures for 2, 6 and 24 h. After the incubation period, the cells were carefully washed and cell extracts were prepared for HPLC analysis. As evident from Table 3a and b, conversion of [8][9][10][11][12][13][14] C]GCV to its 5Ј-mono-, 5Ј-di-and 5Ј-tri-phosphate derivatives was markedly more pronounced in HSV-1tk gene-transfected FM3A cells than wild-type FM3A/0 cells.…”

“…As evident from Table 3a and b, conversion of [8][9][10][11][12][13][14] C]GCV to its 5Ј-mono-, 5Ј-di-and 5Ј-tri-phosphate derivatives was markedly more pronounced in HSV-1tk gene-transfected FM3A cells than wild-type FM3A/0 cells. In fact, triphosphate metabolites of GCV appeared at a 30-to 60-fold higher extent in FM3Atk − /HSV-1tk + cells than in FM3A/0 cells after 2, 6 or 24 h incubation of the cells with 1 M radiolabeled drug.…”

“…Since no major differences in metabolic properties of GCV and E-GCV were observed in FM3A cell cultures upon continuous exposure of the cells to both drugs, the retention of the intracellular [8-14 C]GCV metabolite levels upon removal of [8][9][10][11][12][13][14] C]GCV from the culture medium was investigated. FM3A/0 and FM3Atk − /HSV-1tk + cells were incubated with 1 M [8-14 C]GCV or [8][9][10][11][12][13][14] C]E-GCV for a 24-h period, upon which the drug was carefully removed from the extracellular medium after four subsequent washings of the cells (Figure 3).…”

“…FM3A/0 and FM3Atk − /HSV-1tk + cells were incubated with 1 M [8-14 C]GCV or [8][9][10][11][12][13][14] C]E-GCV for a 24-h period, upon which the drug was carefully removed from the extracellular medium after four subsequent washings of the cells (Figure 3). Whereas 50% of the initial intracellular E-GCV/GCV levels persisted for at least 2 to 26 h in E-GCV-treated FM3Atk − /HSV-1tk + cells, GCV levels decreased to 34% within 2 h after careful wash-out of the extracellular drug and then further decayed during the following 68 h (Figure 3a).…”