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Analysis of Estrogen Agonism and Antagonism of Tamoxifen, Raloxifene, and ICI182780 in Endometrial Cancer Cells: A Putative Role for the Epidermal Growth Factor Receptor Ligand Amphiregulin

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Abstract

Objectives

In different tissues, estrogens, selective estrogen receptor modulators (SERMs), and anti-estrogens exert different biologic activities. For the endometrium, estradiol and tamoxifen induce proliferation, and because of this, tamoxifen treatment of breast cancer patients results in a two- to sevenfold increased risk for development of endometrial cancer. Use of raloxifene, or the anti-estrogen ICI182780, does not result in such an increased risk. The objective of the current study was to generate and analyze gene expression profiles that reflect the transcriptional response of the human endometrium to estradiol, SERMs like tamoxifen and raloxifene, and anti-estrogens like ICI182780.

Methods

Transient transfections were peformed to analyze the transcriptional response of ECC-1 cells to estradiol, tamoxifen, raloxifene, and ICI182780. Subsequently, to reveal the molecular mechanism of action, gene expression profiles were generated and some of the observed regulated genes were confirmed by Northern blotting. Biostatistical methods were employed to analyze the expression profile results further, and amphiregulin effects on ECC-1 cell signaling were investigated using Northern and Western blotting, and 3 H-thymidine incorporation.

Results

Analysis of the profiles revealed that estradiol, tamoxifen, raloxifene, and ICI182780 influence the same biologic processes, but they do so via regulation of different sets of genes. Upon construction of a genetic network it was observed that the largest possible network centered on epidermal growth factor (EGF) receptor signaling. Furthermore, the EGF receptor ligand amphiregulin was differentially regulated by all four ligands. Next it was shown that amphiregulin indeed could stimulate EGF receptor signaling in ECC-1 cells. Based on these results, it was hypothesized that EGF receptor signaling could differentially be affected by estrogen, tamoxifen, raloxifene, and ICI182780 because these four compounds differentially regulate the EGF receptor ligand amphiregulin.

Conclusions

Regulation of amphiregulin coincides with the described in vivo effect of the four ligands on the endometrium. Therefore, it is possible that modulation of EGF receptor signaling is a significant player in estrogen-agonistic growth of the endometrium and needs to be investigated further.

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Authors and Affiliations

  1. Department of Reproduction and Development, PO Box 1738, 3000, DR Rotterdam, The Netherlands

    Susanne C. J. P. Gielen MD, Curt W. Burger MD, PhD, Liesbeth C. M. Kühne BsC, Payman Hanifi-Moghaddam PhD & Leen J. Blok PhD

Authors
  1. Susanne C. J. P. Gielen MD
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  2. Curt W. Burger MD, PhD
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  3. Liesbeth C. M. Kühne BsC
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  4. Payman Hanifi-Moghaddam PhD
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  5. Leen J. Blok PhD
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Corresponding author

Correspondence to Leen J. Blok PhD.

Additional information

The authors thank Wilfred Van IJcken from The Center for Biomics, Erasmus MC, Rotterdam, The Netherlands, for producing custom-made oligoarrays and for providing labeling protocols. In addition, we thank the Center for Biomics for facilitating BioAnalyzer RNA analysis, automated hybridization, array scanning, Imagene software, microarray normalization, and Rosetta Resolver software. Furthermore, we would like to thank Dr P. Giguere (Montreal, Canada) for the C3-luciferase construct, Dr D.P. McDonnell for the pS2-luciferase, and Dr M. Hibner (Scottsdale, AZ) for providing of Raloxifene hydrochloride.

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Gielen, S.C.J.P., Burger, C.W., Kühne, L.C.M. et al. Analysis of Estrogen Agonism and Antagonism of Tamoxifen, Raloxifene, and ICI182780 in Endometrial Cancer Cells: A Putative Role for the Epidermal Growth Factor Receptor Ligand Amphiregulin. Reprod. Sci. 12, e55–e66 (2005). https://doi.org/10.1016/j.jsgi.2005.08.003

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