Induction by estrogens of methotrexate resistance in MCF-7 breast cancer cells [published erratum appears in Carcinogenesis 1998 Nov;19(11):2059]

Abstract

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-Delbrück 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.

Irreversible inhibition of estrogen receptor α signaling and the emergence of hormonal resistance in MCF7 breast cancer cells induced by DNA damage agents

“…Significantly, MCF7 cells are characterized as p53-positive tumor cells, suggesting potential interactions between p53 and ERα signaling. Currently, only few studies describe the interrelation between p53 and ERα, highlighting changes in p53 activity under estrogen stimulation (67,68). MCF7 cells are wtERα and wtp53 positive (69) but the interplay between the two transcription factors in 5-FU and UVC-treated cells has not been investigated in the present study.…”

17β‐Oestradiol attenuates nucleotide excision repair

“…Fusenig (Deutsches Krebsforschungszentrum, Heidelberg, Germany [13]) and cell culture medium (RPMI 1640) was from Gibco BRL/Life Technologies (Paisley, UK). Foetal bovine serum was stripped of endogenous oestrogens using charcoal as described by Thibodeau et al [14]. Anti‐T〈〉T antibody was produced and characterised ‘in house’ as described by Ahmad et al [15].…”