Estrogen-dependent Carcinoma Research Articles

Active Heme Metabolism Suppresses Macrophage Phagocytosis via the TLR4/Type I IFN Signaling/CD36 in Uterine Endometrial Cancer.

Uterine endometrial cancer (UEC) is a common gynecological estrogen-dependent carcinoma, usually accompanied by intermenstrual bleeding. Active heme metabolism frequently plays an increasingly important role in many diseases, especially in cancers. Tumor-associated macrophages (TAMs) are the major population in the immune microenvironment of UEC. However, the roles of heme metabolisms in the crosstalk between UEC cells (UECCs) and macrophages are unclear. In our study, by using TCGA database analysis, integration analysis of the protein-protein interaction (PPI) network and sample RNA transcriptome sequencing were done. The expression level of both heme-associated molecules and iron metabolism-related molecules were measured by quantitative real-time polymerase chain reaction. Heme level detection was done through dehydrohorseradish peroxidase assay. In addition to immunohistochemistry, phagocytosis assay of macrophages, immunofluorescence staining, intracellular ferrous iron staining, as well as enzyme-linked immune sorbent assay were performed. In the study, we verified that heme accumulation in UECCs is apparently higher than in endometrial epithelium cells. Low expression of succinate dehydrogenase B under the regulation of estrogen contributes to over-production of succinate and heme accumulation in UECC. More importantly, excessive heme in UECCs impaired macrophage phagocytosis by regulation of CD36. Mechanistically, this process is dependent on toll-like receptor (TLR4)/type I interferons alpha (IFN Iα) regulatory axis in macrophage. Collectively, these findings elucidate that active heme metabolism of UECCs directly decreases phagocytosis by controlling the secretion of TLR4-mediated IFN Iα and the expression of CD36, and further contributing to the immune escape of UEC.

Estrogen pathway mutations and cancer

Cancer is caused by an accumulation of mutations in a stem cell. A defective mismatch repair (MMR) system can lead to such an accumulation of mutations. MMR defects are found in a cancer syndrome called Lynch Syndrome, and tumors of this syndrome are indeed characterized by such an accumulation of mutations, particularly in short repetitive DNA sequences, called microsatellites. When such mutated microsatellites are located in the coding sequences of genes with essential roles for tumorigenesis, we speak of ‘target genes’. Many such target genes have been found and in this review we focus the possible involvement of target genes involved in the estrogen-receptor pathway (ER). We recently identified NRIP1, encoding the nuclear receptor-interacting protein 1, as the most frequently mutated gene in microsatellite instable (MSI) endometrial cancer (EC). NRIP1 is a known corepressor of the ER pathway, the pathway essential in regulating the concentrations of estrogens, a hormone for which the endometrium is highly responsive. This in combination with the fact that high exposure to estrogens is currently considered the major risk factor for EC - approximately 80% of all sporadic EC tumors are estrogen dependent carcinomas - make NRIP1 the perfect target gene for EC. Interestingly, mutations in NRIP1 were also detected in MSI colorectal carcinoma (CRC) samples. Finding mutations in an estrogen receptor signaling protein in colorectal tissue might not be that expected, as colon is not typically associated with being responsive to estrogens. However, evidence is accumulating to better understand this finding. For instance, it was shown that NRIP1 , in colon tissue, stimulates APC gene transcription and inhibits β-catenin activation. Moreover, some studies suggested that estrogens can increase the expression of MLH1 in colon cancer cells, highlighting the implications of estrogen protecting against colon cancer, by regulating the MMR system. All in all we conclude that genes involved in the estrogen pathway are the perfect candidates to be studied in MMR-deficient tumors, especially those developing in hormonal responsive tissues.

Trastuzumab-induced HER2 phosphorylation: exploring the mechanisms and implications

Cancer is caused by an accumulation of mutations in a stem cell. A defective mismatch repair (MMR) system can lead to such an accumulation of mutations. MMR defects are found in a cancer syndrome called Lynch Syndrome, and tumors of this syndrome are indeed characterized by such an accumulation of mutations, particularly in short repetitive DNA sequences, called microsatellites. When such mutated microsatellites are located in the coding sequences of genes with essential roles for tumorigenesis, we speak of ‘target genes’. Many such target genes have been found and in this review we focus the possible involvement of target genes involved in the estrogen-receptor pathway (ER). We recently identified NRIP1, encoding the nuclear receptor-interacting protein 1, as the most frequently mutated gene in microsatellite instable (MSI) endometrial cancer (EC). NRIP1 is a known corepressor of the ER pathway, the pathway essential in regulating the concentrations of estrogens, a hormone for which the endometrium is highly responsive. This in combination with the fact that high exposure to estrogens is currently considered the major risk factor for EC - approximately 80% of all sporadic EC tumors are estrogen dependent carcinomas - make NRIP1 the perfect target gene for EC. Interestingly, mutations in NRIP1 were also detected in MSI colorectal carcinoma (CRC) samples. Finding mutations in an estrogen receptor signaling protein in colorectal tissue might not be that expected, as colon is not typically associated with being responsive to estrogens. However, evidence is accumulating to better understand this finding. For instance, it was shown that NRIP1, in colon tissue, stimulates APC gene transcription and inhibits β-catenin activation. Moreover, some studies suggested that estrogens can increase the expression of MLH1 in colon cancer cells, highlighting the implications of estrogen protecting against colon cancer, by regulating the MMR system. All in all we conclude that genes involved in the estrogen pathway are the perfect candidates to be studied in MMR-deficient tumors, especially those developing in hormonal responsive tissues.

Abstract A127: A novel class of compounds that selectively target (ER +ve) breast cancer cells.

Abstract Estrogens play an important role in breast cancer development, with approximately 60% of premenopausal and 75% of postmenopausal breast cancer patients having estrogen-dependent carcinomas. In recent decades endocrine therapy has emerged for the treatment of estrogen receptor (ER) positive breast cancer, including aromatase inhibitors (anastrozole) and estrogen receptor antagonists (tamoxifen). However, drug resistance is often induced by these therapies, tumor selectivity is sometimes poor with non-tumor populations being affected, and target selectivity is also a problem with “off-target-effects” culminating in adverse toxicities and/or limited efficacy. For the past ten years collaboration between the Calvary Mater Newcastle Hospital and the University of Newcastle (Australia) has seen the development of many small molecule drugs for the treatment of malignancy. Our recent findings have discovered a family of 2-phenylacrylonitriles showing growth inhibition against a panel of human cancer derived cell lines. Focused library approaches facilitated the identification of a simple pharmacophore, comprising two terminal aromatic moieties linked via a conjugated cyano moiety. Multiple libraries led to the discovery of two key compounds, MT-CN-07 and MT-CN-18. Both exhibit broad spectrum growth inhibition with GI50 values of 3.2–46μM and 3.1–69μM, respectively, in a panel of nine human cancer cell lines derived from colon, ovarian, lung, skin, prostate and pancreatic carcinomas, neuroblastoma and glioblastoma1. An initial screen in one breast cancer ER+ve cell line (MCF-7) suggested that these compounds may selectively target estrogen dependent breast cancer cells. Indeed recent studies have shown that MT-CN-07 and MT-CN-18 are up to 645 fold more potent at inhibiting the growth of ER+ve breast cancer cells (ZR-75–1, T47D, MCF-7) with GI50 values of 0.08–0.6μM, and 0.107–2.25μM, respectively, when compared with the nine cell lines derived from other tumor types. Moreover, these compounds are up to 450 fold more potent in ER+ve breast cancer cell lines when compared with the estrogen independent cell line MDA-MB-231 (ER-ve, GI50 = 36μM and 26μM, respectively) and the non-tumorigenic breast epithelial cell line MCF10A (GI50 = 36μM and 37μM, respectively). Screening of these compounds in the drug resistant, etoposide-selected breast cancer cell line, MCF7/VP, which has amplification and over-expression of the ABCC1 gene, showed they were not substrates for this drug resistant mechanism. The ability to specifically target estrogen dependent tumors, while having little or no effect on normal breast cells, or other tumor types is a unique finding. Thus we have identified a novel group of molecules that could be exploited for the treatment of estrogen sensitive breast cancer. Reference: 1. Tarleton M, Gilbert J, Robertson MJ, McCluskey A, Sakoff JA. (2011) Library synthesis and cytotoxicity of a family of 2-phenylacrylonitriles and discovery of an estrogen dependent breast cancer lead compound. Med Chem Comm. 2, 31–37 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr A127.

Bone and Men's Health. Effects of aromatase inhibitors on human osteoblasts

Aromatase inhibitors (AIs) have been widely employed as an adjuvant endocrine therapy in postmenopausal patients with estrogen receptor (ER) positive breast carcinoma in place of tamoxifen. AIs work on suppressing both systemic and local estrogen biosynthesis in an almost complete manner resulting in estrogen depletion and subsequently inhibition of estrogen dependent carcinoma cell proliferation. Despite marked therapeutic effects in these patients, potential side effects related to long term and marked estrogen depletion should be considered for the benefits of these patients. Among these possible side effects, bone loss and/or increased rates of pathological fractures may be considered one of the most serious complications in terms of effecting activities of daily life of the patients. AIs can be classified into two types, steroidal and non-steroidal in terms of their structure and modes of actions. The former is also characterized to display androgenic actions. Estrogens are well-known to exert their effects primarily through osteoblasts in human bone tissues. Therefore, in this review, we will summarize in vitro effects of AIs, especially androgenic effects of steroidal AIs upon human osteoblasts in order to further understand their effects on human skeletal systems.

MMP7 expression regulated by endocrine therapy in ERβ-positive colon cancer cells

BackgroundMany studies have shown that colon cancer is an estrogen-dependent carcinoma. This study explored the efficacy of endocrine therapy in colon cancer cells with high metastatic potential (HT29). We investigated the proliferation of HT29 cells after exposure to endocrine therapy (tamoxifen) and 5-FU.MethodsApoptosis was evaluated using flow cytometry. The expression of matrix metalloproteinases 7 (MMP-7) and estrogen receptor beta (ERβ) was measured by reverse transcription-polymerase chain reaction (RT-PCR) and western blot. The migration capability of treated cells was determined with wound scratch assay.ResultsTamoxifen alone, 5-FU alone, and the combination of the two drugs can significantly inhibit HT29 cell proliferation and migration, block the cells in G2/M phase and induce cell apoptosis. These drugs also can down-regulate MMP7 and ERβ expression.ConclusionOur findings suggest that endocrine therapy is an efficient therapy for inhibiting ERβ-positive colon cancer cell proliferation and migration via down-regulation of MMP7.

Cytochrome P450 aromatase gene (CYP19) expression in gastric cancer

The secretion of biologically active estrogen through the conversion of circulating precursor androgens has been suggested to play important roles in the pathophysiology of estrogen-dependent carcinomas. In the present study, we examined aromatase expression in gastric carcinoma. Nineteen specimens of gastric carcinoma were obtained from Japanese patients at the Department of Surgery, Tottori University Hospital, Japan. Nontumoral tissues adjacent to the carcinoma were also available for analysis. The histological features of the gastric carcinomas were as follows: 8 intestinal-type and 11 diffuse-type adenocarcinomas. Tissue specimens removed at surgery were used for the preparation of RNA or for immunohistochemical analysis. Six cell lines derived from human gastric cancers were also used as a model system. Using conventional and real-time reverse transcription-polymerase chain reactions, aromatase mRNA expression and promoter usage were assayed. Immunohistochemical analysis was performed using an anti-aromatase antibody. We demonstrated the molecular basis of aromatase mRNA expression, which depended on three proximal promoters in tumoral and nontumoral tissues, for the first time. The tumoral tissues exhibited positive staining for anti-aromatase antibody. At the same time, positive staining was also observed in nontumoral mucosa, predominantly in the parietal cells. We provide evidence suggesting a mechanism for the secretion of estrogen through the conversion of a precursor androgen in tumoral and nontumoral tissues in the stomach.

Characterization of a novel silencer element in the human aromatase gene PII promoter.

Approximately two thirds of breast cancer patients have estrogen-dependent carcinomas. The biosynthesis of estrogens is catalyzed by the microsomal enzyme aromatase. Mechanisms controlling human aromatase gene expression are complicated by the existence of multiple tissue specific promoters. The most proximally located Pll promoter is mainly active in ovarian granulosa cells. PlI can be switched on in human breast cancer cells. Since there are strong silencer elements located within the 3' portion of the PlI promoter, we propose that the function of these silencer elements could be reversed by breast cancer cell specific signals/factors, resulting in aberrant expression of aromatase. We have identified and characterized a novel silencer element, S2, which is upstream of S1, a silencer element recently identified by another group. S2, a 54-bp fragment 100% conserved between humans and rodents, functions in both orientation- and promoter-independent manners. The core region of S2 contains two consensus binding sites for members of the GATA transcription factors. GATA-4 was found to be expressed in three out of four human breast cancer cell lines examined by RT-PCR, and transfection with GATA-4 partially reversed the repressive function of S2. However, we were unable to demonstrate that DNA-protein complexes formed between nuclear extracts of human breast and ovarian cancer cells and S2 contain GATA-4 using a supershifting approach. We suggest that the expression of GATA-4, and more importantly, other yet to be identified GATA or GATA-related factor(s), are implicated in provoking aberrant expression of aromatase, and therefore, the biosynthesis of estrogens, in human breast cancer cells.

From Endocrinology to Intracrinology.

Recently, in situ formation of active sex steroids at the sites of their actions from biologically inactive precursors in the circulation have been demonstrated to play very important roles in sex steroid-dependent neoplasms. These tissues in which the conversion occurs are designated as intracrine tissues and their mechanisms of actions can be designated as intracrinology in contrast to endocrinology. Aromatase, which converts serum androgens to estrone, and 17B-hydroxysteroid dehydrogenase I, which is involved primarily in the conversion of estrone to estradiol, are two major enzymes which function in the in situ formation of biologically active estrogens from circulatory androgens. In human estrogen-dependent neoplasms, including breast, endometrioid endometrial, and common epithelial ovarian carcinoma, we recently demonstrated overexpression of aromatase, especially in stromal cells at sites of frank invasion possibly under a new promoter usage and that of 17B-hydroxysteroid dehydrogenase I in these carcinoma cells. These estrogen-dependent carcinomas are considered to have a common characteristic in estrogen metabolism (i.e., the expression of aromatase in the stromal cells and of 17B-hydroxysteroid dehydrogenase I in the epithelial cells). With these in situ mechanisms of generating biologically active estrogens from circulating androgens, that is, "intracrine manner," these estrogen-dependent neoplasms can exert estrogenic actions on carcinoma cells despite low circulating serum estrogen levels, as observed in postmenopausal women. Evaluation of intracrine mechanisms can provide new insights into various estrogen-related biological phenomena in humans.

Flavonoid inhibition of aromatase enzyme activity in human preadipocytes

Eleven flavonoid compounds were compared with aminoglutethimide (AG), a pharmaceutical aromatase inhibitor, for their abilities to inhibit aromatase enzyme activity in a human preadipocyte cell culture system. Flavonoids exerting no effect on aromatase activity were catechin, daidzein, equol, genistein, β-naphthoflavone (BNF), quercetin and rutin. The synthetic flavonoid, α-naphthoflavone (ANF), was the most potent aromatase inhibitor, with an I 50 value of 0.5 μM. Three naturally-occurring flavonoids, chrysin, flavone, and genistein 4′-methyl ether (Biochanin A) showed I 50 values of 4.6, 68, and 113 μM, respectively, while AG showed an I 50 value of 7.4 μM. Kinetic analyses showed that both AG and the flavonoids acted as competitive inhibitors of aromatase. The K i values, indicating the effectiveness of inhibition, were 0.2, 2.4, 2.4, 22, and 49 μM for ANF, AG, chrysin, flavone, and Biochanin A, respectively. Chrysin, the most potent of the naturally-occurring flavonoids, was similar in potency and effectiveness to AG, a pharmaceutical aromatase inhibitor used clinically in cases of estrogen-dependent carcinoma. These data suggest that flavonoid inhibition of peripheral aromatase activity may contribute to the observed cancer-preventive hormonal effects of plant-based diets.

Growth inhibition of estrogen-dependent rat mammary cancer by EMD 16-795, a new synthetic anti-estrogen.

To test the effect of a synthetic anti-estrogen EMD 16-795 (7-METHOXY-4 (p-(2-pyrrolidino-ethoxy) pheny1)-isoflavanol-4)) on estrogen-dependent mammary cancer adult female rats bearing dimethyl benzanthracene-induced mammary carcimoma were treated orally every third day with doses of 15 or 10 mg per kg bodyweight EMD 16-795 dissolved in water or 35% ethanol. The 1 mg dose inhibited tumor growth and did not affect bodyweight but the higher doses caused a partial regression of tumor tissue and a loss of bodyweight of nearly 10%. The higher the dose the longer the inhibitory effect lasted after cessation of EMD treatment. It is believed that the EMD 16-795 BLOCKS the binding of the natural estrogen to the specific hormone receptors thus inhibiting estrogen-dependent carcinoma.

PROPHYLACTIC CASTRATION IN CARCINOMA OF THE BREAST

The beneficial effects of oophorectomy as a palliative procedure in advanced mammary carcinoma have been known for more than 60 years (Schinzinger, 1889; Beatson, 1896). With increasing knowledge of the relationship of certain malignancies to hormonal functions, particularly the estrogendependent carcinomas of the breast, castration by surgical means or by irradiation of the ovaries has become more widely practiced in the management of metastatic cancer of the breast. In contrast to the acceptance of castration as a palliative measure in the treatment of advanced cancers, the suppression of ovarian function, either by surgery or by irradiation as a prophylactic method before metastases occur, has been a controversial problem. The majority of publications on this subject have been in disfavor of such procedures, as exemplified by Ackerman and del Regato 1 by the statement: There is no evidence that sterilization results in the prolongation of life or that the systematic sterilization

Prophylactic castration in carcinoma of the breast.

The beneficial effects of oophorectomy as a palliative procedure in advanced mammary carcinoma have been known for more than 60 years (Schinzinger, 1889; Beatson, 1896). With increasing knowledge of the relationship of certain malignancies to hormonal functions, particularly the estrogendependent carcinomas of the breast, castration by surgical means or by irradiation of the ovaries has become more widely practiced in the management of metastatic cancer of the breast. In contrast to the acceptance of castration as a palliative measure in the treatment of advanced cancers, the suppression of ovarian function, either by surgery or by irradiation as a prophylactic method before metastases occur, has been a controversial problem. The majority of publications on this subject have been in disfavor of such procedures, as exemplified by Ackerman and del Regato 1 by the statement: There is no evidence that sterilization results in the prolongation of life or that the systematic sterilization