FOXA1: a transcription factor with parallel functions in development and cancer

When aberrant, factors critical for organ morphogenesis are also commonly involved in disease progression. FOXA1 (forkhead box A1), also known as HNF3α (hepatocyte nuclear factor 3α), is required for postnatal survival due to its essential role in controlling pancreatic and renal function. In addition to regulating a variety of tissues during embryogenesis and early life, rescue experiments have revealed a specific role for FOXA1 in the postnatal development of the mammary gland and prostate. Activity of the nuclear hormone receptors ERα (oestrogen receptor α) and AR (androgen receptor) is also required for proper development of the mammary gland and prostate respectively. FOXA1 modulates ER and AR function in breast and prostate cancer cells, supporting the postulate that FOXA1 is involved in ER and AR signalling under normal conditions, and that some carcinogenic processes in these tissues stem from hormonally regulated developmental pathways gone awry. In addition to broadly reviewing the function of FOXA1 in various aspects of development and cancer, this review focuses on the interplay of FOXA1/ER and FOXA1/AR, in normal and cancerous mammary and prostate epithelial cells. Given the hormone dependency of both breast and prostate cancer, a thorough understanding of FOXA1's role in both cancer types is critical for battling hormone receptor-positive disease and acquired anti-hormone resistance.     

Estrogen receptor beta in breast cancer—Diagnostic and therapeutic implications

More than 10 years have passed since the discovery of the second estrogen receptor, estrogen receptor β (ERβ). It is now evident that ERα is not the only ER in breast cancer cells; in fact, ERβ is expressed in the majority of breast cancers although at lower levels than in the normal breast. In addition, ERβ is expressed in breast cancer infiltrating lymphocytes, fibroblasts and endothelial cells, all known to influence tumor growth. By overexpressing or knocking-out ERβ in breast cancer cell lines, several researchers have investigated its function with respect to proliferation and tumor growth. It appears that ERβ is anti-proliferative, in many ways antagonising the function of ERα. Furthermore, phytoestrogens have a binding-preference for ERβ and several epidemiological studies indicate a breast cancer preventing effect of this class of compounds. Tamoxifen is one of the standard, adjuvant treatments for ERα positive breast cancer, classically thought to mediate its effect through ERα. However, in several recent studies, ERβ has been described as a potential marker for tamoxifen response. In summary, experimental, epidemiological as well as diagnostic studies point towards ERβ as an important factor in breast cancer, opening up the possibility for novel ERβ-selective therapies in the treatment of breast cancer.     

KLF6 inhibits estrogen receptor-mediated cell growth in breast cancer via a c-Src-mediated pathway

Estrogen receptors play a key role in breast cancer development and progression. Kruppel-like factor 6 (KLF6) is a tumour-suppressing protein. The aim of this study was to identify the role of KLF6 inhibition in estrogen receptor{alpha} (ERα)-elicited breast cancer development. Protein expression levels were examined by western blot analysis and immunoprecipitation was used to analyse interactions between proteins. An MTT assay was used to study cell proliferation. We found that KLF6 mediates cell growth in ERα-positive breast cancer cells through interaction with the c-Src protein. This interaction causes inactivation of the Erk and Akt proteins. These pathways are critical for the proliferation and survival of breast cancer cells. We also established that KLF6 could not mediate cell growth in ERα-negative cells. We conclude that KLF6 can modulate ERα-mediated cell growth in breast cancer cells. The unique role of KLF6 in mediating cell growth in breast cancer cells opens up the possibility of a new therapeutic strategy for treating breast cancer.     

ERα-negative and triple negative breast cancer: Molecular features and potential therapeutic approaches

Triple negative breast cancer (TNBC) is a type of aggressive breast cancer lacking the expression of estrogen receptors (ER), progesterone receptors (PR) and human epidermal growth factor receptor-2 (HER-2). TNBC patients account for approximately 15% of total breast cancer patients and are more prevalent among young African, African-American and Latino women patients. The currently available ER-targeted and Her-2-based therapies are not effective for treating TNBC. Recent studies have revealed a number of novel features of TNBC. In the present work, we comprehensively addressed these features and discussed potential therapeutic approaches based on these features for TNBC, with particular focus on: 1) the pathological features of TNBC/basal-like breast cancer; 2) E₂/ERβ-mediated signaling pathways; 3) G-protein coupling receptor-30/epithelial growth factor receptor (GPCR-30/EGFR) signaling pathway; 4) interactions of ERβ with breast cancer 1/2 (BRCA1/2); 5) chemokine CXCL8 and related chemokines; 6) altered microRNA signatures and suppression of ERα expression/ERα-signaling by micro-RNAs; 7) altered expression of several pro-oncongenic and tumor suppressor proteins; and 8) genotoxic effects caused by oxidative estrogen metabolites. Gaining better insights into these molecular pathways in TNBC may lead to identification of novel biomarkers and targets for development of diagnostic and therapeutic approaches for prevention and treatment of TNBC.     

Cryptotanshinone inhibition of mammalian target of rapamycin pathway is dependent on oestrogen receptor alpha in breast cancer

Cryptotanshinone (CPT) has been demonstrated to inhibit proliferation and mammalian target of rapamycin (mTOR) pathway in MCF‐7 breast cancer cells. However, the same results are unable to be repeated in MDA‐MB‐231 cells. Given the main difference of oestrogen receptor α (ERα) between two types of breast cancer cells, It is possibly suggested that CPT inhibits mTOR pathway dependent on ERα in breast cancer. CPT could significantly inhibit cell proliferation of ERα‐positive cancer cells, whereas ERα‐negative cancer cells are insensitive to CPT. The molecular docking results indicated that CPT has a high affinity with ERα, and the oestrogen receptor element luciferase reporter verified CPT distinct anti‐oestrogen effect. Furthermore, CPT inhibits mTOR signalling in MCF‐7 cells, but not in MDA‐MB‐231 cells, which is independent on binding to the FKBP12 and disrupting the mTOR complex. Meanwhile, increased expression of phosphorylation AKT and insulin receptor substrate (IRS1) induced by insulin‐like growth factor 1 (IGF‐1) was antagonized by CPT, but other molecules of IGF‐1/AKT/mTOR signalling pathway such as phosphatase and tensin homolog (PTEN) and phosphatidylinositol‐4,5‐bisphosphate 3‐kinase (PI3K) were negatively affected. Finally, the MCF‐7 cells transfected with shERα for silencing ERα show resistant to CPT, and p‐AKT, phosphorylation of p70 S6 kinase 1 (p‐S6K1) and eukaryotic initiation factor 4E binding protein 1 (4E‐BP1) were partially recovered, suggesting ERα is required for CPT inhibition of mTOR signalling. Overall, CPT inhibition of mTOR is dependent on ERα in breast cancer and should be a potential anti‐oestrogen agent and a natural adjuvant for application in endocrine resistance therapy.     

Age-associated biomarker profiles of human breast cancer

To explore the hypothesis that aging not only increases breast cancer incidence but also alters breast cancer biology, we correlated patient age and diagnosis with tumor histology, stage and biomarkers independently determined from two different tumor archives: an American collection of approximately 800 paraffin-embedded and immunohistochemically analyzed primary breast cancers, and an European collection of approximately 3000 cryobanked primary breast cancers analyzed by ligand-binding and enzyme immunoassay (EIA). The prognostic biomarkers chosen for comparison represented surrogate measures of tumor: (i). proliferation, growth and genetic instability (mitotic and apoptotic indices, Ki-67/MIB-1-positivity, nuclear grade, p53-positivity), (ii). endocrine-dependence (estrogen receptor (ER), progesterone receptors (PR), pS2, Bcl2), (iii). growth factor receptor-dependence (ErbB2, EGFR/ErbB1), and (iv). angiogenic, invasive and proteolytic potential (uPA, PAI-1, Cathepsin D, VEGF). No biomarker reflecting tumor angiogenic, invasive or proteolytic potential showed a significant correlation with patient age at diagnosis. In contrast, significant inverse correlations (|r|>0.1; P< or =0.05) were observed for all measures of tumor growth and genetic instability as well as growth factor receptor overexpression (ErbB2 or EGFR positivity). Only one marker of endocrine-dependence, ER expression, showed a significant positive correlation with patient age at diagnosis. In summary, these findings support the hypothesis that breast cancer biology is significantly affected by patient age. In particular, breast tumors arising in older patients have slower growth rates, are more likely to be ER-positive, and are less likely to be p53-positive, EGFR-positive or ErbB2-positive.     

Estrogen receptor-β activation in combination with letrozole blocks the growth of breast cancer tumors resistant to letrozole therapy

Treatment with anti-estrogens or aromatase inhibitors (AI) is the main therapeutic strategy used against estrogen receptor ERα-positive breast cancer. Resistance to these therapies presents a major challenge in the management of breast cancer. Little is known about ERβ in breast carcinogenesis. Our aim in this study is to examine potential novel strategies utilizing ERβ activity to overcome AI resistance. We provide evidence that ERβ agonist can reduce the growth of AI-resistant breast cancer cells. Our data further confirm that therapeutic activation of ERβ by DPN, an ERβ agonist, blocks letrozole-resistant tumor growth in a xenograft model. Interestingly, DPN exerted tumor growth inhibition only in the presence of the AI letrozole, suggesting that combination therapy including ERβ activators and AI may be used in the clinical setting treating AI resistant breast cancer. An increase in ERβ levels, with diminished ERα/ERβ ratio, was observed in the tumors from mice treated with DPN/letrozole combination compared to single agents and control. Decreased Cyclin D1 and increased CyclinD1/CDK inhibitors p21 and p27 levels in DPN/letrozole treated tumors were observed, suggesting that the combination treatment may inhibit tumor growth by blocking G1/S phase cell cycle progression. Our data show a decrease in MAPK phosphorylation levels without affecting total levels. In addition to providing evidence suggesting the potential use of ERβ agonists in combination with letrozole in treating AI resistant breast cancer and prolonging sensitivity to AI, we also provide mechanistic evidence supporting the role of ERβ in altering the expression profile associated with resistance.     

Anti-steroidal and anti-growth factor activities of anti-estrogens

Both steroid hormones, such as estrogens and progestins acting via nuclear receptors, and growth factors, such as EGF, IGF-I and IGF-II acting via transmembrane receptors, are able to modulate the growth of human breast cancer cells. In addition to its anti-estrogenic action requiring estrogen receptor (ER) and leading to growth arrest, we have previously shown that the anti-hormone tamoxifen (Tam) is able to block EGF, insulin and IGF-I mitogenic activities in total absence of estrogens (BBRC, 146,1502,1987). This anti-growth factor activity is observe exclusively in ER + cells and is rescued by estradiol addition, thus suggesting that it is mediated by accessible ER sites. In the same culture conditions, progestins and anti-progestins do not display such an inhibition, whereas retinoic acid does, thus indicating that this anti-growth factor effect is not restricted to ER ligands. To progress in the understanding of this inhibition, we first analyzed how Tam could affect EGF and IGF-I binding in responsive cells. We have shown that Tam neither affects EGF and IGF-I binding to their respective receptors by direct competition nor modulates their affinities. However, our recent data suggest that Tam pretreatment (6 days) of MCF7 cells, which similarly prevents EGF and IGF-I mitogenic activities, results in opposite effects on the concentrations of their binding sites.

In conclusion, we propose that some steroid antagonists can inhibit not only the action of agonist ligands of the receptors they are binding to, but can also modulate the action of growth factors by decreasing their receptor concentrations or altering their functionalities.     

表皮生长因子受体与他莫西芬耐药机制

雌激素受体及其信号通路在乳腺癌的发生发展中发挥着关键作用。到目前为止,抑制或阻断雌激素信号通路的内分泌治疗尤其是他莫西芬,仍是对雌激素受体阳性乳腺癌患者最有效的治疗手段之一。然而,他莫西芬的耐药问题直接影响了乳腺癌患者的治疗及预后。最近多项研究表明雌激素受体与表皮生长因子受体家族尤其是HER2介导的信号传导通路在多个点上相互交叉,彼此影响,与他莫西芬的耐药密切相关     

Estrogen and growth factor receptor interactions in human breast and non-small cell lung cancer cells

Extranuclear estrogen receptors may mediate rapid effects of estradiol that communicate with nuclear receptors and contribute to proliferation of human cancers bearing these signaling proteins. To assess these growth-promoting pathways, we undertook controlled homogenization and fractionation of NIH-H23 non-small cell lung cancer cells. As many breast tumors, NIH-H23 cells express estrogen receptors (ER), with the bulk of specific estradiol binding in nuclear fractions. However, as in breast cells, a significant portion of specific, high-affinity estradiol-17beta binding-sites are also enriched in plasma membranes of lung tumor cells. These estrogen binding-sites co-purify with plasma membrane-marker enzymes and are not significantly contaminated by cytosol or nuclei. On further purification of membrane caveolae from lung tumor cells, proteins recognized by monoclonal antibodies to nuclear ER-alpha and to ER-beta were identified in close association with EGF receptor in caveolae. In parallel studies, ER-alpha and ER-beta are also detected in nuclear and extranuclear sites in archival human breast and lung tumor samples and are noted to occur in clusters at the cell membrane by using confocal microscopy to visualize fluorescent-labeled monoclonal antibodies to ER-alpha. Data on site-directed mutagenesis of cysteine-447 in ER-alpha suggest that association of ER forms with membrane sites may depend on acylation of cysteine by palmitate. Estrogen-induced growth of MCF-7 breast cancer and NIH-H23 lung cancer cells in vitro correlated closely with acute hormonal activation of mitogen-activated protein kinase signaling and was significantly reduced by treatment with Faslodex, a pure anti-estrogen. Further, combination of Faslodex with selected growth factor receptor inhibitors elicited a more pronounced inhibiton of tumor cell growth. Thus, extranuclear forms of ER play a role in promoting downstream signaling for hormone-mediated proliferation and survival of breast, as well as lung, cancers and offer a new target for anti-tumor therapy.