The G Protein-Coupled Receptor GPR30 Mediates the Proliferative Effects Induced by 17β-Estradiol and Hydroxytamoxifen in Endometrial Cancer Cells

Abstract

The growth of both normal and transformed epithelial cells of the female reproductive system is stimulated by estrogens, mainly through the activation of estrogen receptor alpha (ERalpha), which is a ligand-regulated transcription factor. The selective ER modulator tamoxifen (TAM) has been widely used as an ER antagonist in breast tumor; however, long-term treatment is associated with an increased risk of endometrial cancer. To provide new insights into the potential mechanisms involved in the agonistic activity exerted by TAM in the uterus, we evaluated the potential of 4-hydroxytamoxifen (OHT), the active metabolite of TAM, to transactivate wild-type ERalpha and its splice variant expressed in Ishikawa and HEC1A endometrial tumor cells, respectively. OHT was able to antagonize only the activation of ERalpha by 17beta-estradiol (E2) in Ishikawa cells, whereas it up-regulated c-fos expression in a rapid manner similar to E2 and independently of ERalpha in both cell lines. This stimulation occurred through the G protein-coupled receptor named GPR30 and required Src-related and epidermal growth factor receptor tyrosine kinase activities, along with the activation of both ERK1/2 and phosphatidylinositol 3-kinase/AKT pathways. Most importantly, OHT, like E2, stimulated the proliferation of Ishikawa as well as HEC1A cells. Transfecting a GPR30 antisense expression vector in both endometrial cancer cell lines, OHT was no longer able to induce growth effects, whereas the proliferative response to E2 was completely abrogated only in HEC1A cells. Furthermore, in the presence of the inhibitors of MAPK and phosphatidylinositol 3-kinase pathways, PD 98059 and wortmannin, respectively, E2 and OHT did not elicit growth stimulation. Our data demonstrate a new mode of action of E2 and OHT in endometrial cancer cells, contributing to a better understanding of the molecular mechanisms involved in their uterine agonistic activity.

c-Jun activation is required for 4-hydroxytamoxifen-induced cell death in breast cancer cells

“…Our previous studies have shown that one micromolar OHT induces cell proliferation in thyroid and endometrial cancer cells by activating the EGFR/GPR30/ERK pathway and in turn c-Fos expression (Vivacqua et al, 2006a(Vivacqua et al, , 2006b, whereas it has no effects on the JNK/ c-Jun pathway (Madeo A, unpublished observations). These observations together with this study lead to the hypothesis that the functional output of c-Fos induction by OHT may depend on whether or not there is a concomitant c-Jun activation by JNK signaling.…”

GPR30: A G protein-coupled receptor for estrogen

“…The upregulation of c-fos by estrogen and phytoestrogens has also been shown in breast cancer cells (Maggiolini et al, 2004). Recently, GPR30 has been demonstrated to mediate the proliferative effects of both estrogen and tamoxifen in endometrial cancer cells (Vivacqua et al, 2005) and thyroid cancer cells (Vivacqua et al, 2006). Taken together, these observations suggest that GPR30 may play a role in the regulation of cellular growth, including proliferation and apoptosis.…”

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“…Nevertheless, not all those actions can be attributed to the classical ERs. The ability of E2 to activate G proteins highlighted the role of the orphan G-protein coupled receptor (GPCR) 30 [90, 92, 93], recently renamed GPER (G-Protein coupled Estrogen Receptor).…”

“…Several years later the possible function for GPER was identified by demonstrating protein kinases Erk1 and Erk2 activation induced by E2, as well as by the ER antagonists ICI 182,780 and tamoxifen, in breast cancer cell lines expressing GPER but not in cell lines lacking GPER [89]. Furthermore the up-regulation of c-fos by estrogen and phytoestrogens has also been shown in breast cancer cells [92, 93]. GPER, as the other 906 members of the GPCRs superfamily, is a 7 transmembrane protein (7TM) and its cellular localization is still a matter of debate.…”

“…subsequently, Src promotes the shedding of heparin-binding EGF-like growth factor and therefore EGFR activation [89], which in turn activates several signaling cascades, such as the ERK, PI3 kinase and phospholipase C pathways [200, 202]. Several studies have demonstrated that E2 triggers various biological effects through GPER [89, 90, 92, 93, 198], whereas it has been shown that estriol (E3) exerts an antagonist effect on this receptor [196]. …”