Molecular mechanism of the inhibition of estradiol-induced endometrial epithelial cell proliferation by clomiphene citrate

Endocrinology. 2010 Jan;151(1):394-405. doi: 10.1210/en.2009-0721. Epub 2009 Nov 24.


We examined the molecular mechanisms of the antiestrogenic effects of clomiphene citrate (CC) in the endometrium using two types of cell lines, Ishikawa and EM-E6/E7/hTERT cells. CC or ICI182780 inhibited 17beta-estradiol (E2)-induced endometrial cell proliferation and transcriptional activation of the estrogen response element (ERE) gene. We directly visualized the ligand-estrogen receptor (ER)alpha interaction using green fluorescent protein (GFP)-tagged ER alpha in a single living cell. Whereas E2 changed the nuclear localization of GFP-ER alpha to a punctate distribution within 5 min, CC or ICI182780 changed the slower and less mobilization of GFP-ER alpha compared with E2. Pretreatment with CC or ICI182780 partly prevented the E2-induced nuclear redistribution of GFP-ER alpha. Fluorescence recovery after photobleaching revealed that GFP-ER alpha mobility treated with E2 was more rapid than that treated by CC or ICI182780. As coactivator recruitment to the ER is essential for ER-dependent transcription, we examined the interaction between ER alpha and steroid receptor coactivator-1 (SRC-1). The complex formation between ER alpha and SRC-1 was significantly increased by E2 but was prevented in the presence of CC or ICI182780 by coimmunoprecipitation. Moreover, the E2-induced colocalization of GFP-ER alpha and SRC-1 was prevented in the presence of CC or ICI182780 according to an immunofluorescence assay. We also observed that the reduction of SRC-1 using small interfering RNA for SRC-1 resulted in the inhibition of E2-induced cell proliferation and transcriptional activation of the ERE gene. Collectively, these results suggest that CC may inhibit E2-induced endometrial epithelial cell proliferation and ERE transactivation by inhibiting the recruitment of SRC-1 to ER alpha.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Proliferation / drug effects*
  • Cells, Cultured
  • Clomiphene / pharmacology*
  • Down-Regulation / drug effects
  • Endometrium / drug effects*
  • Endometrium / metabolism
  • Endometrium / physiology
  • Estradiol / pharmacology*
  • Estrogen Receptor alpha / antagonists & inhibitors
  • Estrogen Receptor alpha / metabolism
  • Female
  • Green Fluorescent Proteins / metabolism
  • Humans
  • Nuclear Receptor Coactivator 1 / metabolism
  • Protein Binding / drug effects
  • RNA, Small Interfering / pharmacology
  • Response Elements / drug effects
  • Response Elements / physiology
  • Signal Transduction / drug effects*
  • Signal Transduction / genetics
  • Tissue Distribution
  • Transcriptional Activation / drug effects


  • ESR1 protein, human
  • Estrogen Receptor alpha
  • RNA, Small Interfering
  • Green Fluorescent Proteins
  • Clomiphene
  • Estradiol
  • Nuclear Receptor Coactivator 1