Pharmacokinetic/pharmacodynamic study of ZD9331, a nonpolyglutamatable inhibitor of thymidylate synthase, in a murine model following two curative administration schedules.

ZD9331 is a nonpolyglutamatable antifolate inhibitor of thymidylate synthase currently in clinical development. This enzyme is crucial for DNA synthesis and catalyzes the reductive methylation of dUMP to form thymidylate, which is subsequently converted to dTTP. The pharmacokinetics of two curative antitumor doses of ZD9331 administered by either a single i.p. bolus injection (50 mg/kg) or by 24-h s.c. infusion (3 mg/kg) have been measured in a thymidine salvage-incompetent murine lymphoma model (L5178Y) using a sensitive and specific ELISA. To gain an understanding of the relationship between the pharmacokinetics of ZD9331 and antitumor activity perturbations in tumor, dTTP and dUMP concentrations were also determined. After bolus administration, ZD9331 was eliminated from plasma and tissues relatively rapidly, with terminal elimination (lambda(z) 0-24 h) of 4-6 h. Liver concentrations were 8-fold higher than those measured in the plasma. Kidney and lymphoma drug concentrations were similar to those of plasma, although there was evidence of a slower overall elimination of drug at later time points. Steady-state concentrations of ZD9331 were obtained 4-5 h after the start of the 24 h s.c. infusion. At the end of infusion, elimination rates were similar for plasma and tissues (approximately 3.5 h) but appeared to be slower in the tumor at later time points. Liver concentrations were approximately 4-fold higher, and kidney and tumor concentrations were similar to those in the circulation. Depletion of dTTP and elevation in dUMP in the tumor were consistent with inhibition of thymidylate synthase after both administration schedules, although the time for which dTTP was decreased was longer (approximately 24 h) for the infusional route than for the bolus injection (<16 h). The results suggest that antitumor activity is dependent on attaining adequate drug concentrations to affect dTTP pools as well as on the duration of effective drug levels.

Comparison of plasma and tissue levels of ZD1694 (Tomudex), a highly polyglutamatable quinazoline thymidylate synthase inhibitor, in preclinical models.

ZD1694 (Tomudex, raltitrexed) is a specific quinazoline antifolate thymidylate synthase inhibitor that relies on polyglutamation for high potency. Antibodies to ZD1694 have been used to establish a sensitive radioimmunoassay as an alternative to high-performance liquid chromatography (HPLC). The radioimmunoassay is reproducible, accurate and provides a means of determining low levels of ZD1694 in plasma (< 1 nM). By virtue of the high cross-reactivity of the antibodies with polyglutamated forms of ZD1694, it is also possible to measure the total concentration of drug in tissues. Results obtained in L1210 mouse leukaemia cells and in mouse tissues were similar to those previously determined using radiolabelled drug. Pharmacokinetic studies in mice have confirmed that the compound is rapidly eliminated from the plasma and that there is a prolonged terminal elimination phase. ZD1694 was measured in plasma (0.56 ng ml(-1); 1.2 pmol ml(-1)) up to 7 days after a single i.p. dose of 100 mg kg(-1) ZD1694. Liver, kidney and gut epithelium had a substantially higher level of ZD1694 immunoreactivity than plasma. For example, 24 h after a single i.p. dose at 1, 10 and 100 mg kg(-1), total drug levels in the liver were 480, 325 and 152 times higher than plasma levels respectively. In kidney and gut epithelium, total drug levels at these doses were approximately 55 and 34 times those of plasma. The high tissue to plasma ratios were maintained for at least 7 days after administration. Similarly, high tissue to plasma ratios (> 100) were found in dogs treated with a clinically relevant dose of ZD1694. These were maintained for 4 weeks in liver and kidney tissue (> 100). Total gastrointestinal concentrations of ZD1694 were approximately 10 times higher than plasma 3 days after administration, but levels were near to the limit of detection at 4 weeks. These results are consistent with extensive polyglutamation of ZD1694 within tissues in both mice and dog and provide further support for the infrequent schedule that has been used clinically. Although it has not been possible to measure individual polyglutamated forms of ZD1694, the radioimmunoassay provides a convenient means of assessing total drug levels in tissues and is currently the only method suitable for measuring the extent of drug retention in normal tissue and tumour biopsies obtained from patients treated with ZD1694.