Modulation of Antifolate Cytotoxicity by Metabolites from Dying Cells in a Lymphocyte Clonal Assay

Overview

Journal Br J Cancer

Specialty Oncology

Date 1988 May 1

PMID 3395552

Citations 2

Authors

J M Hughes

A deFazio

M H Tattersall

Affiliations

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Abstract

A lymphocyte clonal assay developed to quantitate in vivo somatic cell mutations at the hypoxanthine-guanine phosphoribosyltransferase locus was modified in order to study resistance to methotrexate. Even though nucleoside-free culture conditions were used methotrexate was not lethal to lymphocytes plated into micro-wells at greater than 10(2) cells/well. HPLC analysis of supernatants from wells plated initially with 10(4) cells/well in 100 microM methotrexate revealed the presence of micro-molar levels of hypoxanthine and thymidine by the 5th and 8th day of culture respectively. When lymphocytes were plated at less than or equal to 10(2) cells/well in nucleoside free medium, methotrexate was cytotoxic and micro-molar levels of thymidine together with hypoxanthine protected lymphocytes cultured under these conditions from toxicity. Modulation of nucleic acid antimetabolite cytotoxicity by nucleosides and bases has been recognised for some years. Nucleoside free culture conditions have been advocated for studying cellular sensitivity to antifolates to avoid such interfering factors. However our results indicate that metabolites from dying or damaged cells can prevent methotrexate cytotoxicity, further complicating the development of a suitable clonogenic assay for investigating antifolate sensitivity.

Citing Articles

Methotrexate cytotoxicity determination using the MTT assay following enzymatic depletion of thymidine and hypoxanthine.

Haber M, Madafiglio J, Norris M J Cancer Res Clin Oncol. 1993; 119(6):315-7.

PMID: 8449966 DOI: 10.1007/BF01208836.

Potentiation of methotrexate lymphocytotoxicity in vitro by inhibitors of nucleoside transport.

Hughes J, Tattersall M Br J Cancer. 1989; 59(3):381-4.

PMID: 2930702 PMC: 2247054. DOI: 10.1038/bjc.1989.76.

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