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Therapeutic Relevance of Differences in the Structural Specificity of the Transport Systems for Folate Analogs in L1210 Tumor Cells and in Isolated Murine Intestinal Epithelial Cells¹

Memorial Sloan-Kettering Cancer Center, New York, New York 10021

The kinetics of aminopterin, methotrexate (10-methylaminopterin), and 10-ethylaminopterin transport by isolated murine intestinal epithelial cells was determined and compared with that of L1210 tumor cells. The Michaelis constant (K_m) for the influx of aminopterin into L1210 tumor cells was 1.2 µM, as contrasted to 7.2 µM for influx into isolated intestinal epithelial cells. Substitution of a methyl or an ethyl moiety at position 10 of the aminopterin molecule reduced the affinity (increased the K_m) of the L1210 carrier system for either derivative approximately 3-fold. In contrast, the affinity of the carrier system of isolated intestinal epithelia for methotrexate or for 10-ethylaminopterin was 12- and 17-fold less than that for aminopterin. The rate of efflux of each of the folate analogs within a particular cell type was approximately the same; efflux was approximately 3 times slower from isolated intestinal epithelial cells than from L1210 cells.

Following the s.c. administration of aminopterin (3.0 mg/kg) to mice bearing L1210 ascites, levels of drug in excess of that bound to dihydrofolate reductase were maintained in both small intestine and L1210 cells for at least 24 hr. After 3.0 mg/kg s.c., levels of methotrexate or of 10-ethylaminopterin in excess of that bound to dihydrofolate reductase were maintained for only 4 to 6 hr in small intestine but for 16 to 24 hr in L1210 tumor cells. Therapy trials in which a s.c. dose was given every other day until death confirmed that the optimum dose range for aminopterin (0.15 to 0.3 mg/kg) was lower than that for either methotrexate (6 to 9 mg/kg) or 10-ethylaminopterin (6 to 9 mg/kg) and that the increase in life span at the maximum tolerated range was higher for the derivatives substituted at position 10 (150 to 180%) than it was for aminopterin (75%).

The higher affinity of the carrier system in tumor cells as compared to that of the carrier system in normal cells for each folate analog results in a greater persistence of drug in tumor in vivo and explains the selective antitumor action of each of the three analogs. Substitution at position 10 of aminopterin reduces the affinity of the transport system in both tumor and normal cells for these drugs, but to a different extent. The greater reduction in the affinity of the carrier in normal cells than in tumor cells accounts for the improved therapeutic index of both the methyl and the ethyl-substituted derivatives.

¹ Supported in part by Grants CA-16153 and CA-08748 from the National Cancer Institute and Grant CH-26 from the American Cancer Society.

² To whom requests for reprints should be addressed.

Received 5/12/77. Accepted 8/24/77.

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