Carcinogenesis, Vol 19, 1545-1552, Copyright © 1998 by Oxford University Press
PA Thibodeau, N Bissonnette, SK Bedard, D Hunting and B Paquette
Development of drug resistance is a major factor that limits the
effectiveness of chemotherapy treatments. In this study, we determined
whether estradiol or its metabolites 2-, 4- and 16alpha-hydroxyestrone
could enhance the development of methotrexate resistance in the breast
carcinoma cell line, MCF-7. Cells were incubated with the estrogens at a
concentration of 10(-8) M for 12 cell doublings and enhancement of
methotrexate resistance was measured with the Luria-Delbruck assay. The
most efficient estrogens were the 4-hydroxyestrone and 16alpha-
hydroxyestrone, which both stimulated methotrexate resistance by 88- fold
as compared with the control without estrogen. 2-Hydroxyestrone had an
enhancement factor of 33-fold, whereas estradiol showed a slight effect
with an enhancement factor of 3.2-fold. To determine whether the estrogen
receptor was involved in the development of resistance, expression of the
pS2 gene, which contains an estrogen-responsive element, was measured. Both
estradiol and 16alpha-hydroxyestrone stimulated expression of the pS2 gene.
In contrast, 2- and 4- hydroxyestrone did not increase the level of pS2
mRNA. This suggests that tumors classified as estrogen receptor negative
could also develop methotrexate resistance as the result of exposure to
estrogens. The status of the tumor suppressor gene p53 was analyzed in
methotrexate sensitive and resistant clones. In all the methotrexate
resistant clones analyzed, the western blots indicated that the p53 protein
was still present and transcriptionally competent, as measured by its
capacity to stimulate transcription of the p21waf1/cip1 gene following UVB
irradiation. However, the basal level of p53 was higher in resistant clones
and addition of 2- or 4-hydroxyestrone increased p53 to levels equivalent
to those observed following UVB irradiation. However, this induction of p53
accumulation by estrogens failed to stimulate the transcription of
p21waf1/cip1, which indicates that a transcriptionally inactive form of p53
accumulated in methotrexate resistant cells.
ARTICLES
Induction by estrogens of methotrexate resistance in MCF-7 breast cancer cells [published erratum appears in Carcinogenesis 1998 Nov;19(11):2059]
Department of Radiobiology, Faculty of Medicine, Universite de Sherbrooke, Quebec, Canada.
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