Polyglutamylation, an Important Element in Methotrexate Cytotoxicity and Selectivity in Tumor versus Murine Granulocytic Progenitor Cells in Vitro

Isabelle Fabre, Gerard Fabre, I. David Goldman

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Abstract

Methotrexate (MTX) cytotoxicity was assessed by clonogenic assay in agar with granulocytic progenitor cells from mouse bone marrow and in the Ehrlich ascites tumor, the K562 human chronic myelogenous leukemia, and the P388 murine leukemia. After a 2-hr exposure to MTX, the concentrations necessary to produce 50% inhibition of colony formation were 100, 25,1.2, and 0.25 μM, respectively. This was inversely related to the ability of the tumor cells to accumulate MTX polyglutamyl derivatives and consistent with the observation that no polyglutamyl derivatives were observed in granulocytic progenitor cells after a 2-hr exposure to 5 μM MTX. Continuous exposure to glycine (200 μM)-adenosine (100 μM)thymidine (10 μM) (GAT), along with MTX, protected cells from MTX cytotoxicity by circumventing the requirement for tetrahydrofolate cofactors. However, while the presence of GAT during a 2-hr exposure to 5 μM MTX is sufficient to protect granulocyte progenitor cells from MTX cytotoxicity, the presence of GAT, even after MTX is removed, is required to protect tumor cells. Indeed, if, after a 2-hr exposure of tumor cells to MTX and GAT, both MTX and GAT are removed before plating in agar, cytotoxicity to tumor cells was expressed. This sustained antitumor effect of MTX correlates with the rapid build-up of polyglutamyl derivatives that are retained in the cell even after extracellular and intracellular monoglutamate is eliminated. This is in contrast to granulocytic progenitor cells which appear to be susceptible to the drug only during the period of exposure to the monoglutamate under these conditions. The data strongly suggest that the marked differences in the accumulation of MTX polyglutamyl derivatives between the tumor cells studied and the murine bone marrow granulocytic progenitor cells are an important element in MTX selectivity.

Original languageEnglish (US)
Pages (from-to)3190-3195
Number of pages6
JournalCancer Research
Volume44
Issue number8
Publication statusPublished - Aug 1 1984
Externally publishedYes

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ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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