5-Fluoro-1-(2'-deoxy-2'-fluoro-beta-D-ribofuranosyl) uracil trapping in Morris hepatoma cells expressing the herpes simplex virus thymidine kinase gene.

Abstract

UNLABELLED:

The planning and individualization of gene therapy with suicide genes such as herpes simplex virus thymidine kinase (HSV-tk) necessitates the assessment of the enzyme activity expressed in the tumor. This can be done by uptake measurements of specific substrates for HSV-tk. Due to the molecular structure of 5-fluoro-1-(2'-deoxy-fluoro-beta-D-ribofuranosyl)uracil (FFUdR), it may be a substrate for both the mammalian thymidine kinase and HSV-tk.

METHODS:

Using a HSV-tk-expressing rat hepatoma cell line and a control cell line (bearing the empty vector) the uptake of 3H-FFUdR was determined with increasing incubation periods. Furthermore, measurements with graded mixtures of HSV-tk-expressing cells and control cells were made. To elucidate the mechanism of FFUdR transport into cells, a series of inhibition/competition experiments was performed with challenge inhibitors of the nucleoside and the nucleobase transport systems.

RESULTS:

The uptake studies with tritiated FFUdR revealed a 14- to 19-fold higher accumulation in the HSV-tk-expressing cell line compared to the control cell line. While the 3H-FFUdR uptake was 3- to 4-fold higher than the 3H-ganciclovir uptake in the HSV-tk-expressing cells, it was also higher in control cells (5-fold). Furthermore, FFUdR accumulation was linearly correlated with the amount of HSV-tk-expressing cells. FFUdR uptake and growth inhibition by therapeutic doses of ganciclovir were highly correlated, with r = 0.96. Inhibition/competition experiments showed that FFUdR is transported mainly by the equilibrative and the concentrative nucleoside transporter but not by the nucleobase transport systems.

CONCLUSION:

The FFUdR uptake is an indicator of the HSV-tk activity in tumor cells and can be used as a prognostic marker during gene therapy with HSV-tk. The relative merits of ganciclovir and FFUdR as specific substrates for HSV-tk will need to be further explored in vivo.

• Authors: C Germann, A F Shields, J R Grierson, I Morr, U Haberkorn
• Journal: Journal of nuclear medicine : official publication, Society of Nuclear Medicine (J Nucl Med) Vol. 39 Issue 8 Pg. 1418-23 (Aug 1998) ISSN: 0161-5505 [Print] United States
• PMID: 9708520 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• 5-fluoro-1-(2'-deoxy-2'-fluoro-ribofuranosyl)uracil
• Antiviral Agents
• Floxuridine
• Thymidine Kinase
• Ganciclovir

Topics

• Animals
• Antiviral Agents (pharmacokinetics, pharmacology)
• Floxuridine (analogs & derivatives, pharmacokinetics, pharmacology)
• Ganciclovir (pharmacokinetics, pharmacology)
• Genetic Therapy
• Liver Neoplasms, Experimental (genetics, pathology, therapy)
• Rats
• Simplexvirus (genetics)
• Thymidine Kinase (genetics)
• Tomography, Emission-Computed