Inactivation of O6-methylguanine-DNA methyltransferase and sensitization of human tumor cells to killing by chloroethylnitrosourea by O6-methylguanine as a free base

Human fibroblasts and tumor cells with constitutive levels of the DNA repair protein O6-methylguanine-DNA methyltransferase were incubated with mM concentrations of the free base O6-methylguanine for up to 24 h. This treatment depleted the cells of their transferase activity, and sensitized the cell...

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Bibliographic Details
Published in:Cancer research (Chicago, Ill.) Ill.), 1986-04, Vol.46 (4), p.1663-1668
Main Authors: YAROSH, D. B, HURST-CALDERONE, S, BABICH, M. A, DAY, R. S. III
Format: Article
Language:English
Subjects:
Antineoplastic agents
Biological and medical sciences
Cell Survival - drug effects
DNA Repair
Ethylnitrosourea - analogs & derivatives
Ethylnitrosourea - pharmacology
Fibroblasts - enzymology
General aspects
Guanine - analogs & derivatives
Guanine - metabolism
Guanine - pharmacology
Humans
Medical sciences
Methyltransferases - antagonists & inhibitors
Neoplasms - drug therapy
Neoplasms - enzymology
Neoplasms - pathology
O-Methylguanine-DNA Methyltransferase
Pharmacology. Drug treatments
Substrate Specificity
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Summary:Human fibroblasts and tumor cells with constitutive levels of the DNA repair protein O6-methylguanine-DNA methyltransferase were incubated with mM concentrations of the free base O6-methylguanine for up to 24 h. This treatment depleted the cells of their transferase activity, and sensitized the cells to killing by the antineoplastic drug 1-[2-chloroethyl]-1-nitrosourea. Cells constitutively lacking the methyltransferase were not sensitized to cell killing. Cell free extracts incubated with O6-methylguanine also lost methyltransferase activity. Other alkylpurines, such as O6-methylguanosine, S6-methylthioguanine, O6-ethylguanine, and 3-methyladenine, did not have this effect on extracts of human tumor cells, while O6-methylguanosine and O6-methylguanine inactivated purified methyltransferase from Escherichia coli. The data suggest that the free base O6-methylguanine is probably a substrate for the methyltransferase. Calculation of the second order rate constants for free base versus O6-methylguanine in DNA, and experiments in which the free base was mixed with DNA containing O6-methylguanine before reaction with methyltransferase, indicated that the base in DNA is about 4 X 10(7) better as a substrate than is the free base. These results demonstrate that DNA repair capacity of tumor cells can be diminished without DNA damage, and suggest a method for increasing the efficiency of chemotherapy.
ISSN:0008-5472
1538-7445