The M5076 murine "ovarian"
tumor which is naturally refractive to
methotrexate was found to be highly responsive to the lipophilic
antifolate,
metoprine. M5076 cells were markedly deficient in mediated entry of
methotrexate. This was in contrast to the
L1210 leukemia, a
tumor highly responsive to
methotrexate but poorly responsive to
metoprine. Two
L1210 leukemia sublines, with acquired resistance to
methotrexate by virtue of a deficiency in mediated entry of
drug similar to that seen for M5076 cells, were found to be collaterally sensitive to
metoprine. The insensitivity to
methotrexate of the M5076
tumor and the two L1210 sublines is associated with low saturability (high Km) and reduced capacity (low Vmax) for mediated influx of
drug.
5-Methyltetrahydrofolate, the major circulating
folate in blood but not
metoprine, shares this mediated route for entry. Therefore, a relatively low level of accumulation of this natural
folate in these
methotrexate-resistant
tumors, in the face of a
metoprine-induced blockade at the level of
dihydrofolate reductase, probably accounts for the high sensitivity of these
tumors to this lipophilic agent. Evidence for this notion was derived during transport and growth experiments in vitro using
5-formyltetrahydrofolate as a model
folate coenzyme. The value for influx Vmax of this
folate compound in a transport-deficient
methotrexate-resistant subline compared to the parental L1210 was reduced to the same extent as that shown for
methotrexate. Growth of this resistant L1210 subline showed a greater requirement for this model compound than did the parental line. Also, the concentration necessary for 50% inhibition by
metoprine in the presence of this reduced
folate was lower in the resistant subline. Inhibition of each cell line by
metoprine, on the other hand, was the same when
folic acid was used as the
folate source. The implications of these findings for the use of lipophilic
antifolates as alternative
therapy for some
methotrexate-resistant
tumors are discussed.