indicate that transfer of the HSVTK - gene renders these cells extremely sensitive to GCV mediated cell 
killing, with concentration needed to kill 50% of cells (IC50) ranging from approximately 0.05 uM 
to 2.0 uM (in one study of normal cell sensitivity to GCV, most cell types were sensitive only in the 
100-3000 uM range (Matthews and Boehme, 1988). 
A Strong "bystander" Effect Occurs in HS.OIORSVTK infected cells in vitro (see Appendix-Smvthe et 
aL 1994bV . To determine if mesothelioma cells would show a "bystander" effect, near-confluent 
culture flasks of malignant mesothelioma cell lines were infected with H5.010RSVTK' at a 
concentration of 100 particles per cell. Twenty-four hours after transfection, the cells were 
trypsinized, mixed with uninfected cells at varying ratios and plated onto multi-well cell culture 
plates. Following an additional 24 hours the cells were treated with a 20 pM solution of ganciclovir 
diluted in normal cell media for 4 days. The cultures were then assessed for viable cell growth by a 
colorimetric assay measuring dehydrogenase activity. There was no diminution in the efficacy of 
GCV treatment until the ratio of infected:uninfected cells exceeded 1:10 (see Figure 3 in Smythe et 
al. 1993b). These data indicate that a strong bystander effect is present, similar to that reported 
for retrovirally-transduced HS MTK experiments. GCV was almost completely effective at dilutions 
of cells of up to 1:10. Significant effects were still noted at dilutions of up to 1:100. 
In summary, adenovirus effectively infects human mesothelioma cells in culture. We have developed 
an adenoviral vector containing the HS \/TK gene that efficiently transduces mesothelioma cells and 
renders them sensitive to the toxic effects of ganciclovir. We have also demonstrated that a strong 
bystander effect is present using the H5.010RSV77VGCV system. 
Recombinant DNA Research, Volume 20 
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