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Methotrexate polyglutamates in cultured human cells.

D S Rosenblatt, V M Whitehead

    Advances in Experimental Medicine and Biology
    |January 1, 1983
    PubMed
    Summary

    Methotrexate forms polyglutamates in cells, persisting even after drug removal. These polyglutamates contribute to toxicity, and their formation can be influenced by reduced folates and other compounds.

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    Area of Science:

    • Biochemistry
    • Cell Biology
    • Pharmacology

    Background:

    • Methotrexate (MTX) is a chemotherapy drug that inhibits dihydrofolate reductase.
    • Intracellular accumulation of MTX and its metabolites, particularly polyglutamates, influences its efficacy and toxicity.
    • Understanding the dynamics of MTX polyglutamate formation and clearance is crucial for optimizing cancer therapy.

    Purpose of the Study:

    • To investigate the accumulation and persistence of methotrexate polyglutamates in cultured human fibroblasts.
    • To examine the impact of reduced folates and other compounds on MTX polyglutamate formation and MTX-induced toxicity.
    • To compare the effects of MTX polyglutamates with antifolates that do not form polyglutamates.

    Main Methods:

    • Culturing human fibroblasts and exposing them to methotrexate.
    • Measuring intracellular drug levels and the proportion of methotrexate polyglutamates.
    • Assessing the effects of reduced folates and thymidine, glycine, and adenosine (GAT) on MTX accumulation and toxicity.
    • Evaluating the impact of 2,4-diamino-5-(3',4'-dichlorophenyl)-6 methyl pyrimidine (DDMP) on deoxyuridine incorporation.

    Main Results:

    • Fibroblasts accumulated MTX polyglutamates exceeding dihydrofolate reductase binding capacity.
    • MTX polyglutamates persisted intracellularly for days after MTX removal, contributing to ongoing toxicity.
    • Reduced folates prevented polyglutamate accumulation and MTX effects when added early, but were less effective after prolonged MTX exposure.
    • GAT prevented MTX toxicity only when continuously present; preincubation allowed polyglutamate accumulation and subsequent toxicity.
    • DDMP, an antifol that doesn't form polyglutamates, inhibited DNA synthesis only while present, with greater reversal upon removal compared to MTX.

    Conclusions:

    • Methotrexate polyglutamates are long-lived intracellular metabolites that contribute significantly to MTX toxicity.
    • The formation and persistence of MTX polyglutamates are influenced by the presence of reduced folates and other compounds.
    • Antifolates like DDMP, which do not form polyglutamates, offer a different toxicity profile with more readily reversible effects upon removal.

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