Modification of in vivo methotrexate antitumor effect in L1210 leukemia by induced impairment of purine salvage

Studies with murine cells have shown that the antitumor action of methotrexate (MTX) may be through a purineless mechanism. If the MTX effect depends, in part, on inhibition of de novo purine synthesis, then the ability of tumor cells to salvage available purine precursors could reduce the cell kill. In the present study, we produced L1210 murine leukemia mutants with impaired purine salvage to determine whether this would affect responsiveness to MTX. Mutant lines L1210/MP, L1210/FAMP, and L1210/555 were produced by developing resistance to the purine analogs 6-mercaptopurine (6-MP), 6-MP + 2-fluoroadenine (2-FA), and 6-MP + 2-FA + 6-methylmercaptopurine riboside respectively. The purine salvage capability of the cell lines was confirmed in vitro by testing the ability of various purines to reverse the growth inhibitory and biochemical effects of MTX in the presence of thymidine. Dose-response curves demonstrated identical in vitro MTX sensitivity for L1210/MP, L1210/FAMP, and the parent line, L1210/S. Despite identical in vitro MTX sensitivity, the cell lines L1210/MP and L1210/FAMP displayed increased sensitivity to the biochemical effects of MTX in an in vivo model, and this was translated into enhanced sensitivity as measured by survival experiments in tumor-bearing mice. The results indicate that impairment of purine salvage sensitizes cells to the antitumor effect of MTX in vivo. This has implications for the clinical use of MTX in view of the variety of rescue techniques that is available.