nolatrexed and Leukemia-L5178

To determine whether the administration of Thymitaq (AG337), a selective inhibitor of thymidylate synthase (TS), enhances radiation-induced cytotoxicity in vitro and increases tumor control rate in vivo.. In vitro studies were carried out with HT-29 human colon carcinoma cells. In vivo studies were carried out using L5178Y(TK-) murine lymphoma implanted in DBA/2 mice.. Pretreatment of HT-29 cells to nontoxic concentration of AG337 (<10 microM) for a short period of time (< 24 h) significantly enhanced the radiation induced cell lethality. The radiosensitizing enhancement ratio was 1.7. In contrast, there was no increased cell killing when the drug was exposed immediately after irradiation. In studies using L5178Y(TK-) tumors, the drug alone (50 mg/kg, i.p. x 5) had a minimal tumor growth delay, while a single dose of radiation (17 Gy) resulted in < 10% tumor control at day 30. When radiation and drug (17 Gy + AG337, 50 mg/kg, i.p. x 5) were combined, the tumor control rate reached 90% at Day 30. Using the local tumor control assay (TCD50), the radiation dose modification factor after a single dose of radiation was 2.6.. The concentration of drug shown to be of radiosensitizing value in the in vivo studies is achievable in humans. The results of the present study further supports the potential utility of AG337 in the treatment of human tumors by radiotherapy.

Topics: Animals; Antimetabolites, Antineoplastic; Cell Survival; Drug Therapy, Combination; Enzyme Inhibitors; HT29 Cells; Humans; Leukemia L5178; Male; Mice; Mice, Inbred DBA; Quinazolines; Thymidylate Synthase

3,4-Dihydro-2-amino-6 methyl-4-oxo-5-(4-pyridylthio)-quinazoline dihydrochloride (AG337) is a water-soluble, lipophilic inhibitor of thymidylate synthase (TS) designed using X-ray structure - based methodologies to interact at the folate cofactor binding site of the enzyme. The aim of the design program was to identify TS inhibitors with different pharmacological characteristics from classical folate analogs and, most notably, to develop non-glutamate-containing molecules which would not require facilitated transport for uptake and would not undergo intracellular polyglutamylation. One molecule which resulted from this program, AG337, inhibits purified recombinant human TS with a Ki of 11 nM, and displays non-competitive inhibition kinetics. It was further shown to inhibit cell growth in a panel of cell lines of murine and human origin, displaying an IC50 of between 0.39 microM 6.6 microM. TS was suggested as the locus of action of AG337 by the ability of thymidine to antagonize cell growth inhibition and the direct demonstration of TS inhibition in whole cells using a tritium release assay. The demonstration, by flow cytometry, that AG337-treated L1210 cells were arrested in the S phase of the cell cycle was also consistent with a blockage of TS, as was the pattern of ribonucleotide and deoxyribonucleotide pool modulation in AG337-treated cells, which showed significant reduction in TTP levels. The effects of AG337 were quickly reversed on removal of the drug, suggesting, as would be expected for a lipophilic agent, that there is rapid influx and efflux from cells and no intracellular metabolism to derivatives with enhanced retention. In vivo, AG337 was highly active against the thymidine kinase-deficient murine L5178Y/TK-lymphoma implanted either i.p. or i.m. following i.p. or oral delivery. Prolonged dosing periods of 5 or 10 days were required for activity, and efficacy was improved with twice-daily dose administration. Dose levels of 25 mg/kg delivered i.p. twice daily for 10 days, 50 mg/kg once daily for 10 days, or 100 mg/kg once daily for 5 days elicited 100% cures against the i.p. tumor. Doses required for activity against the i.m. tumor were higher (100 mg/kg i.p. twice daily for 5 or 10 days) but demonstrated the ability of AG337 to penetrate solid tissue barriers. Oral delivery required doses of > or = 150 mg/kg twice daily for periods of 5-10 days to produce 100% cure rates against both i.m. and i.p. implanted tumors. These results were consi

Topics: Administration, Oral; Animals; Antimetabolites, Antineoplastic; Cell Cycle; Enzyme Inhibitors; Folic Acid Antagonists; Growth Inhibitors; Humans; Leukemia L1210; Leukemia L5178; Mice; Quinazolines; Rats; Solubility; Thymidylate Synthase