New inhibitors of the enzyme thymidylate synthase (TS) are now reaching clinical application. Alteration of the dUTP: dTTP ratio may be critical to TS inhibition-induced tumor cell death. The DNA polymerase assay with modification was used to rapidly and sensitively measure dUTP, dTTP, and dUTP:dTTP ratios in cell extracts of HT29 human colon carcinoma cells treated with the specific TS inhibitor ZD1694 [N-(5-[N-(3,4-dihydro-2-methyl-4-oxoquinazolin-6-ylmethyl)-N-methylamino]-2-thenoyl)-L-glutamic acid]. These results revealed an increase in the dUTP:dTTP ratio at 2 hr after a 2-hr exposure to ZD1694 at concentrations of 0.05 to 0.2 microM with significant normalization at 16 hr after a 2-hr exposure despite evidence of continued TS inhibition. This assay is highly sensitive and reproducible for levels of dUTP and is less labor intensive than traditional assays.
|Number of pages||4|
|Publication status||Published - 01 Sep 1997|
- Antimetabolites, Antineoplastic
- Colonic Neoplasms
- DNA-Directed DNA Polymerase
- Deoxyuracil Nucleotides
- Dose-Response Relationship, Drug
- Enzyme Inhibitors
- Templates, Genetic
- Thymidylate Synthase
- Thymine Nucleotides
- Tumor Cells, Cultured
- Journal Article
- Research Support, U.S. Gov't, P.H.S.