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Inherent Resistance of Human Squamous Carcinoma Cell Lines to Methotrexate as a Result of Decreased Polyglutamylation of This Drug¹

Department of Pharmacology and Medicine, Yale University School of Medicine, New Haven, Connecticut 06510 [G. P., Y-M. C., J. R. B.], and Grace Cancer Drug Center, Roswell Park Memorial Institute, Buffalo, New York 14263 [J. J. M.]

Three human squamous carcinoma cell lines (FaDu, A253, and SQCC/Y₁) were tested for sensitivity to methotrexate (MTX) and trimetrexate, a second generation folate antagonist in clinical trials. Two of the three cell lines (A253 and SQCC/Y₁) showed inherent resistance to methotrexate, when cytotoxicity was evaluated after short term exposure (4 and 24 h). In contrast, all three cell lines were markedly sensitive to trimetrexate, an antifolate which is taken up by cells by a different transport system than is methotrexate and which is not polyglutamylated. The basis for the natural resistance to methotrexate shown by two of the three cell lines was examined. Levels of dihydrofolate reductase activity, inhibition of this enzyme by methotrexate, and influx of MTX did not differ significantly between the three cell lines; however, resistance was correlated to the amounts of polyglutamates of methotrexate synthesized by the three cell lines. After a 24-h incubation with 10 µM MTX, the A253 cell line was able to form only 35.0 pmol/10⁷ cells of polyglutamates, compared to 250 pmol/10⁷ cells synthesized by the FaDu cell line, while the SQCC/Y₁ cell line, intermediate in sensitivity to methotrexate, was able to form 145 pmol/10⁷ cells of MTX polyglutamates. The A253 cell line contained less folylpolyglutamate synthetase activity compared to the FaDu and SQCC/Y₁ cell lines. However, it is not clear if this difference is sufficient to explain the marked differences in polyglutamates of methotrexate found between the cell lines. We conclude that decreased polyglutamylation of methotrexate in some human squamous cell carcinomas may be the major contributing factor in inherent resistance to high dose pulse administration of this drug.

¹ Supported by USPHS Grants CA 08010, CA 43500, and CA 24538.

² Present address: Memorial Sloan Kettering Institute, 1275 York Ave., New York, NY 10021.

³ Scholar of the Leukemia Society of America.

⁴ American Cancer Society Professor of Pharmacology and Medicine. To whom requests for reprints should be addressed.

Received 2/ 3/89. Revised 6/22/89. Accepted 6/28/89.

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