Heterogeneity of oestrogen receptor expression in normal and malignant breast tissue

We have investigated, by image analysis, the cell cycle expression of estrogen receptors (ER) on MCF-7 cell line and on MCF-7 xenografts. The results demonstrate, in vitro as well as in vivo, an increase of ER concentration during the G0/G1-phase, followed by a decrease during the S-phase until the late S-phase where a rapid increase was noted. These results confirm that estrogens are involved in the DNA synthesis since ER is expressed in vivo at a maximal level in the late G1. In presence of saturating concentrations of 17 beta-estradiol, the mean ER concentration in G0/G1 phase is significantly decreased compared with the control cells cultured in estrogen-deprived medium. This indicates that 17 beta-estradiol down-regulates ER preferentially in the G0/G1 phase. These data suggest that ER in S and G2/M phases is unable to interact with its ligand. Consequently, estrogens may have no effects on the entry of cells in mitosis. Finally, after long-term tamoxifen treatment of MCF-7 xenografts, a tamoxifen-resistant tumor was developed which was characterized by a change in the profile of ER concentration during the G0/G1 phase. In conclusion, it is possible that the differences in cell cycle distribution of ER could be correlated with different phenotypes of breast cancer and also with different clinical phases of tumoral evolution. However, it remains to be known what is the clinical significance of the ER cell cycle expression in relation to tumor aggressiveness and survival.

Section: Introductionsupporting

confidence: 84%

Cell cycle expression of estrogen receptors determined by image analysis on human breast cancer cellsin vitro andin vivo

Rostagno

Moll

Birtwisle-Peyrottes

et al. 1996

“…Most of the published studies, based on comparative studies performed on receptor-negative and positive cells, have demonstrated that the effects of antiestrogen are mediated via ER [11]. However, in vivo the presence of cellular heterogeneity in the expression of ER and PR content [12] did not permit any direct demonstration of ERmediated growth effects of tamoxifen. Indeed, the biochemical assay performed on cell homogenates measured mean receptor levels and did not permit single-cell hormonal receptor level measurements [13].…”

Section: Introductionmentioning

confidence: 95%

Cell cycle expression of steroid receptors determined by image analysis on human breast cancer cell line: A hypothesis on the effects of antiestrogens

Rostagno

Caldani

Lahlou

1996

Tamoxifen is the widespread anti-hormonal compound used for the treatment of human breast cancer. It is admitted that its effects are mediated via estrogen receptors (ER) but the molecular basis of its activity has yet to be clearly defined. In this work, we have developed a new image cytometry procedure in order to clarify the interactions between steroid receptors and tamoxifen at the cell cycle kinetic level. On untreated cells, an increase of ER level and a decrease of progesterone receptor (PR) level during the G0/G1 phase were demonstrated. Then, the ER and PR levels fell during the S-phase until the beginning of G2/M phase, where an increase was observed, especially for PR. These results suggest that ER is synthesized preferentially during the G0/G1 transition and PR during the S/G2 transition. After short-term tamoxifen treatment an augmentation of ER level was observed which was not dose-dependent, suggesting an increase in receptor translation rather than an augmentation of ER synthesis. PR level declined in the majority of the population leading to a selection of a subset of proliferating PR negative cells after treatment. These data demonstrate that the synthesis of steroid receptors is linked with the progression of cells through the cell cycle and indicate that tamoxifen blocks MCF-7 cells in G1 via its interactions with ER. Our multifluorescence imaging procedure appears to provide a rapid and quantitative approach which is useful for investigating alterations in steroid receptors after endocrine treatment.

“…Upon excision, representative portions of tumor specimens were snap frozen on dry ice or in liquid nitrogen and stored at -70 ° C. Portions of the frozen tissues were mounted for cryostat sectioning in a suitable embedding medium and sections prepared and fixed for ER immunocytochemical assay, using an ER-ICA kit (Abbott Diagnostics Inc., Chicago, USA) and previously described methodologies for staining and slide assessment [26,27].…”

Section: Tumor Sample Preparation Immunohistochemistry and Rna Analmentioning

confidence: 99%

Effects of short-term antiestrogen treatment of primary breast cancer on estrogen receptor mRNA and protein expression and on estrogen-regulated genes

McClelland

Manning

Gee

et al. 1996

Effects of the pure antiestrogen ICI182780 and tamoxifen on ER-protein, ER-mRNA, and estrogen-regulated mRNA expression were analysed using matched pretreatment core-cut biopsies and post-treatment mastectomy samples from 43 ER positive human breast cancers. Sixteen controls received either no preoperative treatment (n = 9) (7 days) or placebo (n = 7) (median 21 days) prior to primary surgery. Nineteen patients received ICI182780 6 mg/day (n = 10) or 18 mg/day (n = 9) for 7 days. Eight patients were given preoperative tamoxifen (4 x 40 mg-day 1, 20 mg/day thereafter, median 21 days). ER-protein expression was assessed on pre and post treatment samples by immunocytochemistry. ER, pS2, pLIV1, and actin-mRNA expression was determined by northern analysis on post-treatment samples only. ER-mRNA levels were similar to controls following ICI182780 or tamoxifen treatment. However ER-protein levels were significantly suppressed by ICI182780, particularly at the higher dosage (p = 0.0013). Tamoxifen had no significant effect on ER-protein levels. The ER-mRNA and ER-protein contents of control tumors were linearly related (Spearman r = 0.719, p = 0.006). A similar relationship between pretreatment protein and post ICI182780 treatment mRNA levels was observed (r = 0.652, p = 0.005). However, comparison of post ICI182780 treatment protein and mRNA results shows a loss of linearity through a reduction in protein without concurrent loss of mRNA (r = 0.28, p = 0.257). pS2 mRNA hybridization was lower in ICI182780 treated samples than controls (Mann-Whitney p = 0.035) but was unaffected by tamoxifen. pLIV1 mRNA hybridization was uninfluenced by either treatment. Short term exposure of breast tumors to ICI182780 appears to produce a greater inhibition of estrogen-induced transcriptional events than tamoxifen. These effects appear to occur without a concurrent reduction in ER mRNA levels.