Effect of Herpes Simplex Virus Thymidine Kinase Expression Levels on Ganciclovir-Mediated Cytotoxicity and the “Bystander Effect”

Abstract

Transfer of the herpes simplex virus type-1 thymidine kinase (HSV-tk) gene into tumor cells followed by ganciclovir (GCV) administration, will provide selective tumor cell killing. We studied the effect of herpes simplex virus thymidine kinase (HSV-tk) expression level on the HSV-tk/GCV-mediated "bystander effect." Clones of HSV-tk-transduced rat glioma cells (9L) were isolated that stably expressed with different levels of HSV-tk. All clones studied had similar sensitivity to ganciclovir with IC50 values ranging from 0.45 to 1.3 microM. Within certain enzyme level thresholds, in vitro evaluation of the bystander effect has shown that clones with higher level of HSV-tk expression exhibited a better bystander effect. Interestingly, the bystander effect was observed between different cell types. Both the transduction efficiency and bystander effect are essential factors for the success of the antitumor effect by the HSV-tk/prodrug GCV suicide gene system.

Breast cancer-specific expression of the Candida albicans cytosine deaminase gene using a transcriptional targeting approach

“…Others have reported a minimal threshold level requirement in which they observed no significant difference in cell killing when comparing HSV-tk expression levels from the CMV and Rous sarcoma virus promoters. 15 No significant cell killing was observed when either U2OS or MCF10A cells were transduced with our tissue-specific adenovectors at prodrug concentrations as high as 1000 g/mL, suggesting that these promoters exhibit a remarkable breast tumor specificity when compared with the nontumorigenic MCF10A cells. The ability of these promoters to selectively render breast tumor cells, but not normal breast cells, sensitive to prodrug-mediated cell killing provides another level of specificity when considering targeting tumors within the normal breast tissue.…”

Virally targeted therapies for EBV-associated malignancies

“…Phosphorylated GCV inhibits not only the virally encoded DNA polymerase, but also inhibits the host cell DNA polymerase and is thus cytotoxic (Tiberghien, 1994;Conners, 1995). Furthermore, phosphorylated GCV can be transferred into nearby cells, thus inducing 'bystander' killing (Freeman et al, 1993;Chen et al, 1995;Conners, 1995). In addition, there is evidence that phosphorylated GCV can synergistically kill cells when combined with either certain types of chemotherapy, or irradiation (Kim et al, 1997;Aghi et al, 1998;Wildner et al, 1999).…”

Connexin 43–mediated bystander effect in two rat glioma cell models

“…We used a modified version of the protocol described by Chen et al 12 Cells were treated with trypsin, washed with 10% FCS in RPMI, and then with PBS. The cells were centrifuged and the pellet incubated in lysis buffer (10 mM Tris -HCl pH 7.5, 1 mM dithiothreitol, 1 mM EDTA and 20% glycerol ) in the presence of protease inhibitors: pefabloc SC ( 200 g/ mL ), aprotinin (40 g /mL ), and leupeptin (5 g/ mL ) ( Boehringer Mannheim, Meylan, France).…”