Resistance to methotrexate in thymidylate synthetase-deficient mutants of cultured mouse mammary tumor FM3A cells.
Growth inhibition by methotrexate (MTX) of a cultured mouse mammary carcinoma FM3A line and its mutants deficient in thymidine kinase and thymidylate synthetase was studied under a variety of conditions. In medium containing 10 microM thymidine, the thymidylate synthetase-deficient mutants were slightly more resistant to MTX than the wild-type line and the thymidine kinase-deficient mutant. The addition of both 10 microM thymidine and 50 microM hypoxanthine to the medium completely eliminated the inhibition by MTX of the wild-type and thymidylate synthetase-deficient mutants but had no effect on inhibition of the thymidine kinase-deficient mutant. Thus, addition of either thymidine or purine alone was not sufficient to protect FM3A cells against the growth inhibitory effect of MTX. In contrast, addition of a small amount of 5-methyltetrahydrofolate to medium containing 10 microM thymidine caused an increase of several orders of magnitude in the resistance of thymidylate synthetase-deficient mutants to MTX but did not affect that of the wild-type line. Wild-type cells became almost as resistant to MTX as did the mutant cells by addition of 1 microM 5-fluorodeoxyuridine as well as a small amount of 5-methyltetrahydrofolate with thymidine. These results show directly that thymidylate synthetase is essential in determining the cytotoxicity of MTX by modulating the intracellular tetrahydrofolate pool.
D000818:Animals; D002460:Cell Line; D004351:Drug Resistance; D005467:Floxuridine / Q000494:pharmacology; D008325:Mammary Neoplasms, Experimental / Q000188:drug therapy* / Q000201:enzymology; D008727:Methotrexate / Q000627:therapeutic use*; D008780:Methyltransferases / Q000378:metabolism*; D051379:Mice; D009154:Mutation; D013763:Tetrahydrofolates / Q000378:metabolism* / Q000494:pharmacology; D013936:Thymidine / Q000494:pharmacology; D013937:Thymidine Kinase / Q000378:metabolism; D013940:Thymidylate Synthase / Q000378:metabolism*
D013763:Tetrahydrofolates; D005467:Floxuridine; D008780:Methyltransferases; D013940:Thymidylate Synthase; D013937:Thymidine Kinase; C005984:5-methyltetrahydrofolate; D013936:Thymidine; D008727:Methotrexate