INTRINSIC RESISTANCE TO METHOTREXATE OF CULTURED MAMMALIAN-CELLS IN RELATION TO INHIBITION KINETICS OF THEIR DIHYDROFOLATE REDUCTASES

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  • 1 January 1976
    • journal article
    • research article
    • Vol. 36  (6) , 1991-1997
Abstract
Four cultured mammalian cell lines [mouse L1210 and L5178Y leukemias, human WI-L2 lymphoblasts, rat Yoshida sarcoma] differing in intrinsic resistance to methotrexate over a 70-fold range, were compared with respect to several biochemical factors that might influence response to the drug. Cellular activity of the enzymes dihydrofolate reductase [EC-1.5.1.3] and thymidylate synthetase [EC-2.1.1.6] and the total levels of folate cofactors did not vary by more than a factor of 2 among the cell lines. All cell types were able to transport extracellular methotrexate efficiently across the cell membrane, and at comparable rates. A kinetic study of highly purified dihydrofolate reductases from the 4 sources revealed small differences in the Km values for dihydrofolate and NADPH. A study was made of the inhibition of the 4 dihydrofolate reductases by methotrexate, and Ki values were obtained by fitting the zone B equation of Goldstein to the resulting data. Values of Ki determined by this method correlated with intrinsic resistance of the cell lines and showed a 25-fold range from the most sensitive to the most resistant line. The response of a cell to methotrexate is significantly influenced by the dissociation constant of its dihydrofolate reductase-methotrexate complex.