RELATIVE FREQUENCY AND KINETIC-PROPERTIES OF TRANSPORT-DEFECTIVE PHENOTYPES AMONG METHOTREXATE-RESISTANT L1210-CLONAL CELL-LINES DERIVED INVIVO

  • 1 January 1981
    • journal article
    • research article
    • Vol. 41  (11) , 4447-4452
Abstract
Information was sought on the relative extent to which transport-defective, methotrexate-resistant phenotypes emerge among the total subpopulation of resistant phenotypes during therapeutic challenge of leukemic cells in vivo. A number of monoclonal methotrexate-resistant sublines of the L1210 leukemia were derived during methotrexate therapy of leukemic mice and biochemically characterized. Of the total number of 14 sublines derived, 5 exhibited altered [3H]methotrexate tranport alone, 5 exhibited increased dihydrofolate reductase content alone (2- to 18-fold) and 4 showed alterations in both of these properties. Methotrexate binding and substrate turnover rate for dihydrofolate reductase appeared to be unchanged in any of the resistant sublines. The relative resistance of each subline was accounted for by the biochemical alterations observed. Among the transport-defective sublines, 1 subcategory showed a 3- to 4-fold reduction in apparent influx Vmax for [3H]methotrexate, a second category showed both a 5-fold reduction in influx Vmax and a 3-fold increase in the apparent influx Km, and 1 subline showed only a 2-fold increase in Km. Otherwise, Michaelis-Menten saturation kinetics for influx was observed in each case and in the case of the parental line and the other resistant sublines. None of the resistant sublines exhibited altered efflux of [3H]methotrexate. Steady-state levels measured for intracellular exchangeable (osmotically active) fractions of drug accurately reflected the values for specific kinetic parameters determined for each sensitive and resistant cell line. Transport-defective phenotypes represent a major category of methotrexate-resistant cell types which emerge initially from leukemic cell populations under therapy in mice. A similar relative occurrence of this phenotype may result during methotrexate therapy of leukemia patients.