Abstract
The ability of an antifolate to generate polyglutamates [via folyl polyglutamate sythetase (FPGS)] is an important determinant of its activity as this results in retention of the antimetabolite within the cells and may increase its affinity for the target enzyme. The estimation of TS activity in intact cells can be used to evaluate the retention of 5, 8-dideazafolic acid (DAF) analogues by monitoring [3H]2′-deoxyuridine metabolism in cells following the attainment of a state of dynamic equilibrium. CB3717 (2-amino 10-propargyl DAF) is an inhibitor of TS which has demonstrable clinical efficacy but also exhibits unpredictable toxicities. An in vitro structure activity study, with variations at positions 2 and 10, gave results which ranged from well retained (e.g. 2-methyl, 10-H) through intermediate (e.g. CB3717) to essentially unretained (e.g. 2-methoxy, 10-propargyl). Retention within cells was found to correlate with FPGS substrate activity but independent of both isolated TS inhibition and in vitro cytotoxicity. In mice it is possible to estimate the effect of analogues upon TS in ascitic tumour cells which are totally resistant to CB3717 due to the high circulating thymidine. CB3717 exhibited modest inhibition of TS in vivo with only 43% inhibition 2hrs after the i.v. administration of the maximum tolerated dose (200 mgs/kg). Modification at the 2 position had a dramatic effect such that the dose required to generate retained metabolites which inhibited TS activity by 50% after 2hrs, was »500, 240, 27, and 2 mgs/kg for 2-methoxy, CB3717, desamino (2-H), and 2-methyl analogues respectively. Measuring the drug/target interaction in intact cells has allowed us to identify analogues which are considerably more potent in vivo TS inhibitors.
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Taylor, G., Jackman, A., Balmanno, K. et al. 157 ESTIMATION OF THE IN VITRO AND IN VIVO INHIBITORY EFFECTS OF ANTIFOLATES UPON THYMIDYLATE SYNTHASE (TS) IN WHOLE CELLS. Pediatr Res 24, 137 (1988). https://doi.org/10.1203/00006450-198807000-00181
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DOI: https://doi.org/10.1203/00006450-198807000-00181