ACTIVITY OF THE THYMIDYLATE SYNTHASE INHIBITOR 2-DESAMINO-N-10-PROPARGYL-5,8-DIDEAZAFOLIC ACID AND RELATED-COMPOUNDS IN MURINE (L1210) AND HUMAN (W1L2) SYSTEMS INVITRO AND IN L1210 INVIVO

  • 1 September 1990
    • journal article
    • research article
    • Vol. 50  (17) , 5212-5218
Abstract
We examined the in vitro activity of 2-desamino-5,8-dideazafolate and 2-desamino-N10-propargyl-5,8-dideazafolate (desamino-CB3717), the more water-soluble 2-desamino analogues of 5,8-dideazafolate and N10-propargyl-5,8-dideazafolic acid (CB3717). We report Ki values for the inhibition of L1210 thymidylate synthase (TS) of 2 and 0.027 .mu.M for 2-desamino-5,8-dideazafolate and desamino-CB3717, respectively, indicating a 30- and 10-fold loss in TS-inhibitory activity compared with the corresponding 2-NH2 compounds. The synthetic tri- and tetrapolyglutamate derivatives of desamino-CB3717 were 66- and 101-fold more potent than the monoglutamate form as inhibitors of TS. Both desamino compounds were more potent as inhibitors of L1210 and W1L2 cell growth than were their 2-amino counterparts. 2-Desamino-5,8-dideazafolate retains quite good activity against both the TS-overproducing W1L2:C1 line and the L1210 cell line grown in the presence of thymidine, suggesting that a secondary locus of action may be involved. This other target is a folate-dependent enzyme as evidenced by the protection of the inhibition of cell growth by the addition of hypoxanthine or folinic acid together with thymidine. The methotrexate-resistant, dihydrofolate reductase-overproducing L1210:R7A cell line is cross-resistant to 2-desamino-5,8-dideazafolate, which suggests that dihydrofolate reductase is the other target. An L1210 subline (1565) unable to transport reduced folates is 10-fold resistant to desamino-CB3717 and 2-desamino-5,8-dideazafolate but is not cross-resistant to CB3717 or 5,8-dideazafolate. The removal of the 2-amino function of CB3717 did not affect folylpolyglutamate synthetase substrate activity (CB3717 Km = 48 .mu.M, desamino-CB3717 Km = 40 .mu.M). However, both 5,8-dideazafolate and its desamino analogue were about 10-fold better substrates for folylpolyglutamate synthase than were the N1o-propargyl compounds, and this may contribut to their good growth-inhibitory properties. In vivo, desamino-CB3717 cured .apprx.75% of mice bearing the L1210:ICR tumor at doses of 50 mg/kg daily for 5 days and above (maximum tolerated dose > 1000 mg/kg daily for 5 days). This activity is comparable to that observed for CB3717 (maximum tolerated dose = 100 mg/kg daily for 5 days) despite the much more rapid plasma clearance of desamino-CB3717. Considerable TS inhibition (>80%) was observed in the refractory L1210:NCI tumor (as measured by the release of 3H from 5-[3H] 2''-deoxyuridine) removed 2 h after the i.v. injection of 200 mg/kg of desamino-CB3717 and, after a dose of 1000 mg/kg, TS remained inhibited in cells removed 12 h after drug administration. These data are consistent with the rapid formation of an intracellular, noneffluxable pool of desamino-CB3717 which allows the rapid and sustained inhibition of the target enzyme, TS, in tumor cells.

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