Estrogen Receptor Signaling Pathways Involved in Invasion and Colony Formation of Androgen-Independent Prostate Cancer Cells PC-3.

Castration-resistant prostate cancer (CRPC) is an advanced and androgen-independent form of prostate cancer. Recent studies of rapid actions mediated by estrogen in the prostate and its relationship with CRPC are emerging. We have previously shown that estrogen receptor (ER) promotes migration and invasion of the androgen-independent prostate cancer cells PC-3, but the signaling pathways involved in these events remain to be elucidated. Therefore, this study aimed to analyze the role of ERα and ERß in the activation of SRC, and the involvement of SRC and PI3K/AKT on invasion and colony formation of the PC-3 cells. Our results showed that the activation of ERα (using ERα-selective agonist PPT) and ERß (using ERß-selective agonist DPN) increased phosphorylation of SRC in PC-3 cells. In the presence of the selective inhibitor for SRC-family kinases PP2, the effects of DPN and PPT on transmigration and soft agar colony formation assays were decreased. Furthermore, SRC is involved in the expression of the non-phosphorylated ß-catenin. Finally, using PI3K specific inhibitor Wortmannin and AKT inhibitor MK2206, we showed that PI3K/AKT are also required for invasion and colony formation of PC-3 cells simulated by ER. This study provides novel insights into molecular mechanisms of ER in PC-3 cells by demonstrating that ER, located outside the cell nucleus, activates rapid responses molecules, including SRC and PI3K/AKT, which enhance the tumorigenic potential of prostate cancer cells, increasing cell proliferation, migration, invasion, and tumor formation.

Estrogen Receptors Promote Migration, Invasion and Colony Formation of the Androgen-Independent Prostate Cancer Cells PC-3 Through ß-Catenin Pathway.

Prostate cancer is initially dependent on the androgen, gradually evolves into an androgen-independent form of the disease, also known as castration-resistant prostate cancer (CRPC). At this stage, current therapies scantily improve survival of the patient. Androgens and estrogens are involved in normal prostate and prostate cancer development. The mechanisms by which estrogens/estrogen receptors (ERs) induce prostate cancer and promote prostate cancer progression have not yet been fully identified. Our laboratory has shown that androgen-independent prostate cancer cells PC-3 express both ERα and ERß. The activation of ERß increases the expression of ß-catenin and proliferation of PC-3 cells. We now report that the activation of ERß promotes the increase of migration, invasion and anchorage-independent growth of PC-3 cells. Furthermore, the activation of ERα also plays a role in invasion and anchorage-independent growth of PC-3 cells. These effects are blocked by pretreatment with PKF 118-310, compound that disrupts the complex ß-catenin/TCF/LEF, suggesting that ERs/ß-catenin are involved in all cellular characteristics of tumor development in vitro. Furthermore, PKF 118-310 also inhibited the upregulation of vascular endothelial growth factor A (VEGFA) induced by activation of ERs. VEGF also is involved on invasion of PC-3 cells. In conclusion, this study provides novel insights into the signatures and molecular mechanisms of ERß in androgen-independent prostate cancer cells PC-3. ERα also plays a role on invasion and colony formation of PC-3 cells.

Estrogen receptors localization and signaling pathways in DU-145 human prostate cancer cells.

The aim of the present study was to investigate the subcellular localization of estrogen receptors ERα and ERß in androgen-independent prostate cancer cell line DU-145, and the possible role of exportin CRM1 on ERs distribution. In addition, we evaluated the ERs contribution to activation of ERK1/2 and AKT. Immunostaining of ERα and ERß was predominantly found in the extranuclear regions of DU-145 cells. CRM1 inhibitor Leptomycin B reduced drastically the presence of ERα and ERß in the extranuclear regions and increased in the nuclei, indicating the possible involvement of CRM1 on ERs nuclear-cytoplasmic shuttling. 17ß-estradiol (E2), ERα-selective agonist PPT and ERß-selective agonist DPN induced a rapid increase on ERK1/2 phosphorylation. E2-induced ERK1/2 activation was partially inhibited when cells were pretreated with ERα- or ERß-selective antagonists, and blocked by simultaneous pretreatment with both antagonists, suggesting ERα/ß heterodimers formation. Furthermore, E2 treatment did not activate AKT pathway. Therefore, we highlighted a possible crosstalk between extranuclear and nuclear ERs and their upstream and downstream signaling molecules as an important mechanism to control ER function as a potential therapeutic target in prostate cancer cells.

Activation of estrogen receptor beta (ERß) regulates the expression of N-cadherin, E-cadherin and ß-catenin in androgen-independent prostate cancer cells.

The aim of the present study was to investigate the impact of the activation of estrogen receptors on expression and localization of N-cadherin, E-cadherin and non-phosphorylated ß-catenin in androgen-independent prostate cancer cells (PC-3 and DU-145) and in human post pubertal prostate epithelial cells (PNT1A). Expression of N-cadherin was detected in PNT1A and PC-3 cells, but not in DU-145 cells. E-cadherin was detected only in DU-145 cells and ß-catenin was detected in all cells studied. N-cadherin and ß-catenin were located preferentially in the cellular membrane of PNT1A cells and in the cytoplasm of PC-3 cells. E-cadherin and ß-catenin were located preferentially in the cellular membrane of DU-145 cells. 17ß-estradiol (E2) or the ERα-selective agonist PPT did not affect the content and localization of N-cadherin in PC-3 and PNT1A cells or E-cadherin in DU-145 cells. In PC-3 cells, ERß-selective agonist DPN decreased the expression of N-cadherin. DPN-induced downregulation of N-cadherin was blocked by pretreatment with the ERß-selective antagonist (PHTPP), indicating that ERß1 is the upstream receptor regulating the expression of N-cadherin. In DU-145 cells, the activation of ERß1 by DPN increased the expression of E-cadherin. Taken together, these results suggest that activation of ERß1 is required to maintain an epithelial phenotype in PC-3 and DU-145 cells. The activation of ERß1 also increased the expression of ß-catenin in cytoplasm of PC-3 and in the cellular membrane of DU-145 cells. In conclusion, our results indicate differential expression and localization of N-cadherin, E-cadherin and ß-catenin in androgen-independent prostate cancer cells. The reduction of N-cadherin content by activation of ERß, exclusively observed in androgen-independent prostate cancer cells (PC-3), may be related to the activation of signaling pathways, such as the release of ß-catenin into the cytoplasm, translocation of ß-catenin to the nucleus and activation of gene transcription.

Estrogen receptor beta (ERß) mediates expression of ß-catenin and proliferation in prostate cancer cell line PC-3.

The aim of the present study was to characterize the mechanism underlying estrogen effects on the androgen-independent prostate cancer cell line PC-3. 17ß-estradiol and the ERß-selective agonist DPN, but not the ERα-selective agonist PPT, increased the incorporation of [methyl-(3)H]thymidine and the expression of Cyclin D2, suggesting that ERß mediates the proliferative effect of estrogen on PC-3 cells. In addition, upregulation of Cyclin D2 and incorporation of [methyl-(3)H]thymidine induced by 17ß-estradiol and DPN were blocked by the ERß-selective antagonist PHTPP in PC-3 cells. Upregulation of Cyclin D2 and incorporation of [methyl-(3)H]thymidine induced by DPN were also blocked by PKF118-310, a compound that disrupts ß-catenin-TCF (T-cell-specific transcription factor) complex, suggesting the involvement of ß-catenin in the estradiol effects in PC-3 cells. A diffuse immunostaining for non-phosphorylated ß-catenin was detected in the cytoplasm of PC-3 cells. Low levels of non-phosphorylated ß-catenin immunostaining were also detected near the plasma membrane and in nuclei. Treatment of PC-3 cells with 17ß-estradiol or DPN markedly increased non-phosphorylated ß-catenin expression. These effects were blocked by pretreatment with the ERß-selective antagonist PHTPP, PI3K inhibitor Wortmannin or AKT inhibitor MK-2206, indicating that ERß-PI3K/AKT mediates non-phosphorylated ß-catenin expression. Cycloheximide blocked the DPN-induced upregulation of non-phosphorylated ß-catenin, suggesting de novo synthesis of this protein. In conclusion, these results suggest that estrogen may play a role in androgen-independent prostate cancer cell proliferation through a novel pathway, involving ERß-mediated activation of ß-catenin.

Physiopathological aspects of the Wnt/ß-catenin signaling pathway in the male reproductive system.

The Wnt/ß-catenin signaling pathway controls several biological processes throughout development and adult life. Dysregulation of Wnt/ß-catenin signaling underlies a wide range of pathologies in animals and humans, including cancer in different tissues. In this review, we provide an update of the Wnt/ß-catenin signaling pathway and the possible roles of the Wnt/ß-catenin signaling in the biology of testis, epididymis and prostate. Data from our laboratory suggest the involvement of 17ß-estradiol and estrogen receptors (ERs) on the regulation of ß-catenin expression in rat Sertoli cells. We also provide emerging evidences of the involvement of Wnt/ß-catenin pathway in testis and prostate cancer. Our understanding of the role of Wnt/ß-Catenin signaling in male reproductive tissues is still evolving, and several questions are open to be addressed in the future.