Estrogen-dependent Cancer Research Articles

17β-Estradiol promotes endometrial cancer proliferation and invasion through IL-6 pathway.

Accumulating evidence revealed that the leading risk factor of endometrial cancer is exposure to endogenous and exogenous estrogens, while the exact mechanism underlying estrogen contribution to endometrial cancer progression has not been elucidated clearly. Interleukin (IL)-6 has been verified to be critical for tumor progression in several human cancers. In this study, we provided evidence that 17β-estradiol (E2) could significantly promote endometrial cancer cells viability, migration and invasion through activation of IL-6 pathway, which involved in its downstream pathway and target genes (p-Stat3, Bcl-2, Mcl-1, cyclin D1 and MMP2). Meanwhile, utilization of IL-6-neutralizing antibody could partially attenuate the increased cancer growth and invasion abilities in Ishikawa and RL95-2 endometrial cancer cell lines and an orthotopic endometrial cancer model. We established a causative link between estrogen and IL-6 signaling activation in the development of endometrial cancer. The molecular mechanism defined in this study provided the evidence that E2 promotes endometrial carcinoma progression via activating the IL-6 pathway, indicating that interruption of IL-6 might be an essential therapeutic strategy in estrogen-dependent endometrial cancer.

Dying for a good night of sleep?: Researchers are investigating how poor sleep can increase the risk of cancer or worsen the course of the disease.

Dying for a good night of sleep?: Researchers are investigating how poor sleep can increase the risk of cancer or worsen the course of the disease.

MON-034 Novel Estrogen Activity and Gene Regulation Induced by Personal Care Products in Breast Cancer Cells

The contribution of personal care product (PCP) use to hormone regulated cancer and cancer disparities is unknown. Considering that environmental estrogens and endocrine disrupting chemicals (EDCs) have the potential to contribute to cancer promotion and progression, evaluating chronic use PCPs and hair products for estrogen activity may be of use for risk assessment. In this study, we tested the hypothesis that the ethanol extracts of 11 PCPs (including hair products) would induce estrogen agonist or antagonist activity in breast cancer cells in culture. We have previously shown that three of these PCP extracts induce estrogen agonist activity in the MCF-7 E3 estrogen proliferation assay while six extracts induce estrogen antagonist effects in the T47dkbluc estrogen reporter gene assay. In the current study, the LanthaScreen TR-FRET ER Alpha Competitive Binding Assay was employed to confirm that all nine estrogenic PCP extracts bind to the ER with up to 85% of the activity of E2. Then, to compare endogenous gene regulation induced by the estrogenic PCP extracts with treatment with E2, the Qiagen RT2 Profiler PCR Array for Human Estrogen Receptor Signalling was utilized. MCF-7 cells were treated with solvent control, 1 nM E2 or one of the nine PCP extracts shown to have estrogen activity in bioassays. The three extracts with agonist activity in bioassays produced a gene activation pattern similar to E2 with fewer total genes induced. At the same time, these extracts had no effect on some genes E2 inhibited (compared to control). While the extracts with estrogen antagonist activity in the bioassays inhibited the transcription of most genes induced by E2, these extracts activated the transcription of a number of genes E2 either down regulated or did not regulate. The antagonist PCP extracts repressed transcription of as many as 20 genes such as GPER1, PGR, NR5A2, CAV1 relative to E2. At the same time, in contrast to E2, these extracts induced the transcription of genes such as CYP1A1, CKB, CTGF, WNT4, CTGF, S100A6, FST, NCOA1 CTGF, S100A6, FST, NCOA1, APBB1, PTCH1, MAFF, WNT5A, and/or NOV. We conclude that chronic exposure to these estrogenic or antiestrogenic PCPs has the potential to impact estrogen mediated signaling in exposed individuals that exposure may have an influence on the proliferation and survival of estrogen dependent cancer.

Estrogen Sulfotransferase Induction Inhibits Breast Cancer Cell Line MCF-7 Proliferation

Estrogens are known for their proliferative effects, resulting in tumorigenesis and causing cancers. The majority of breast cancers are estrogen-dependent. Estrogenb-locking drugs developed for treating estrogen-dependent breast cancers include antagonists of estrogen receptors (ERs) and inhibitors of estrogen synthesis enzymes such as aromatase and steroid sulfatase (STS). However, drugs targeting estrogen inactivation enzyme, estrogen sulfotransferase (SULT1E1), have not been developed for estrogen-dependent cancer treatment or prevention. Estrogens must be inactivated after their actions in vivo by SULT1E1, uncontrolled estrogen activity in certain tissues will cause cancers. The majority of human breast cancer cell lines are known to express very low levels of SULT1E1 compared to normal mammary cells. Therefore, gene up-regulation of SULT1E1 could provide novel possibilities for the treatment of estrogen-dependent breast and endometrial cancers. Our data suggest that certain nutritional flavonoids induce SULT1E1 and inhibit cell proliferation in estrogen-dependent breast cancer MCF-7 cells. Our results also suggest that SULT1E1 inducer has an additive or synergistic effect on the inhibition of MCF-7 cell proliferation when combined with other known breast cancer drugs. Naturally occurring flavonoids are safe and inexpensive; they could be used for the prevention of certain breast cancers and/or used as co-drug for the treatment of estrogen-dependent cancers. These results may lead to creating innovative approaches for the treatment or prevention of estrogen-dependent cancers and may lead to the discovery of novel drugs or co-drugs.

Trends in incidence of uterine cancer in Songkhla, Southern Thailand.

ObjectiveTo define the present incidence of uterine cancer and the trends of incidence in the future. Additional, subgroup analysis in religion and socio-demographic factors will be analyzed.MethodsThe data of uterine cancer from The Songkhla Cancer Registry between; 1989 and 2016, was extracted. The population denominators of Songkhla Province were referred from the information of the population censuses surveyed by the National Statistical Office. The incidence was estimate by; the age specific rate, and analyzed in trend incidence using Jointpoint and Age-period-cohort; APC regression models and trend projection in 2017–2030 using by Jointpoint, the NordPred and APC model. The comparison of incidence according to habitat and religion were unpaired t-test.ResultsThe incidence of uterine cancer in Songkhla Province increased from 1.5, to 5.3 per 100,000 women-years in 2016, and 8 per 100,000 women-years in 2030. The incidence was more in urban areas and in Buddhists.ConclusionThe incidence of uterine cancer is increasing. The policy, which promotes protective factors and control risk factors, would decrease incidence of endometrial cancer, and other estrogen dependent cancers additional to chronic diseases from the cardiovascular system.

Clinicopathological and prognostic significance of long noncoding RNA MALAT1 in human cancers: a review and meta-analysis

BackgroundThe aberrant regulation of MALAT1 has been indicated to be involved in various carcinogenic pathways contributing to the tumourigenesis and progression of cancers. The current meta-analysis summarized the research advances of MALAT1 functions and analyzed its prognostic value among multiple types of cancers.MethodsEligible studies were identified through retrieving the PubMed, Web of Science, and CNKI databases, up to Mar 1, 2018. 28 studies of 5436 patients and 36 studies of 3325 patients were enrolled in the meta-analysis to evaluate the association of MALAT1 expression with survival outcomes and clinical parameters.ResultsThe results demonstrated that over-expression of MALAT1 may predict lymph node metastasis (pooled OR = 2.335, 95% CI 1.606–3.395, P = 0.000) and distant metastasis (pooled OR = 2.456, 95% CI 1.407–4.286, P = 0.002). Moreover, MALAT1 was also related with tumour size (pooled OR = 1.875, 95% CI 1.257–2.795, P = 0.002) and TNM stage (pooled OR = 2.034, 95% CI 1.111–3.724, P = 0.021). Additionally, elevated MALAT1 expression could predict poor OS (pooled HR = 2.298, 95% CI 1.953–2.704, P = 0.000), DFS (pooled HR = 2.036, 95% CI 1.240–3.342, P = 0.005), RFS (pooled HR = 2.491, 95% CI 1.505–4.123, P = 0.000), DSS (pooled HR = 2.098, 95% CI 1.372–3.211, P = 0.001) and PFS (pooled HR = 1.842, 95% CI 1.138–2.983, P = 0.013) in multivariate model. Importantly, subgroup analyses disclosed that increased MALAT1 expression had a poor OS among different cancer types (Estrogen-dependent cancer: pooled HR = 2.656, 95% CI 1.560–4.523; urological cancer: pooled HR = 1.952, 95% CI 1.189–3.204; glioma: pooled HR = 2.315, 95% CI 1.643–3.263; digestive cancer: pooled HR = 2.451, 95% CI 1.862–3.227).ConclusionsThe present findings demonstrated that MALAT1 may be a novel biomarker for predicting survival outcome, lymph node metastasis and distant metastasis.

Significance of prohibitin domain family in tumorigenesis and its implication in cancer diagnosis and treatment

Prohibitin (PHB) was originally isolated and characterized as an anti-proliferative gene in rat liver. The evolutionarily conserved PHB gene encodes two human protein isoforms with molecular weights of ~33 kDa, PHB1 and PHB2. PHB1 and PHB2 belong to the prohibitin domain family, and both are widely distributed in different cellular compartments such as the mitochondria, nucleus, and cell membrane. Most studies have confirmed differential expression of PHB1 and PHB2 in cancers compared to corresponding normal tissues. Furthermore, studies verified that PHB1 and PHB2 are involved in the biological processes of tumorigenesis, including cancer cell proliferation, apoptosis, and metastasis. Two small molecule inhibitors, Rocaglamide (RocA) and fluorizoline, derived from medicinal plants, were demonstrated to interact directly with PHB1 and thus inhibit the interaction of PHB with Raf-1, impeding Raf-1/ERK signaling cascades and significantly suppressing cancer cell metastasis. In addition, a short peptide ERAP and a natural product xanthohumol were shown to target PHB2 directly and prohibit cancer progression in estrogen-dependent cancers. As more efficient biomarkers and targets are urgently needed for cancer diagnosis and treatment, here we summarize the functional role of prohibitin domain family proteins, focusing on PHB1 and PHB2 in tumorigenesis and cancer development, with the expectation that targeting the prohibitin domain family will offer more clues for cancer therapy.

Abstract B28: Development of estrogen-dependent and estrogen receptor-negative cervical cancer in HPV transgenic mice

Abstract Estradiol (E2) and human papillomavirus (HPV) cooperate to cause cervical cancer. The role of estrogen receptor α (ERα) in cervical cancer has been underappreciated. Chronic treatment with E2 promotes the development of cervical cancer in K14E7 transgenic mice expressing HPV16 E7 oncoprotein. In the present study, we bred K14E7 to mice with conditional ablation of Esr1 (ERα-coding gene) in the cervical epithelial cells (Wnt7aCre/Esr1f/f; Esr1ed/ed). We found that E2 promoted the development of ERα cervical cancer in K14E7/Esr1ed/ed mice. In addition, proliferation and apoptotic indices of ERα cancer in K14E7/Esr1ed/ed mice were similar to those of ERα+ cancer in K14E7/Esr1f/f mice. We also found that E2 increased the epithelium thickness and epithelial cell proliferation in the cervix of Esr1ed/ed mice. Our published results have demonstrated that deletion of Esr1 in the cervical stroma promotes the regression of cervical precancer lesions in K14E7 mice. In patients, ERα is expressed in the cancer stroma but not cancer epithelial cells. These observations support that stromal ERα, rather than epithelial ERα, mainly mediates oncogenic E2 signaling for cervical cancer. They also suggest that stromal ERα might be an effective therapeutic target for cervical cancer. Note: This abstract was not presented at the conference. Citation Format: Jieun Son, Sang-Hyuk Chung. Development of estrogen-dependent and estrogen receptor-negative cervical cancer in HPV transgenic mice [abstract]. In: Proceedings of the AACR Special Conference: Advances in Modeling Cancer in Mice: Technology, Biology, and Beyond; 2017 Sep 24-27; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(10 Suppl):Abstract nr B28.

Fingolimod interrupts the cross talk between estrogen metabolism and sphingolipid metabolism within prostate cancer cells

Sphingolipids are critical regulators of tumor microenvironments and play an important role in estrogen-dependent cancers. Estrogen and estrogen metabolites were found to be involved in prostate cancer. Fingolimod (FTY720) is a sphingokinase-1 (SphK1) inhibitor with anticancer properties against various tumor cell types. Herein, we investigated the interference of FTY720 with the cross talk between sphingolipid metabolism and estrogen metabolism within prostate cancer cells. FTY720 showed cytotoxic antiproliferative effects against androgen-dependent and -independent prostate cancer cells with IC50 ranging from 3.0 ± 0.3 to 6.8 ± 1.7 μM. Exposure of prostate cancer cells to FTY720 resulted in a dramatic decrease in the concentration of estradiol, estrone, 4-hydroxyestradiol and 16α-hydroxyestrone compared to control cells. However, FTY720 significantly increased the concentration of 2-methoxyestrone and 2-methoxyestradiol within prostate cancer cells. This was mirrored by significant downregulating of the expression of estrogen and catechol estrogen-synthesizing enzymes (CYP19, CYP1A1 and CYP1B1) within prostate cancer cells. On the other hand, FTY720 significantly upregulated the expression of catechol estrogen-detoxifying enzyme (COMT). Additionally, FTY720 abolished estrogen-stimulated expression of ERα and basal expression of ERβ within prostate cancer cells. Furthermore, FTY720 suppressed the expression of the ER-downstream regulated genes, CXCR4 and cyclin D1. Reciprocally, it was found that estradiol and catechol estrogens significantly induced the expression of SphK1 while methoxylated catechol estrogen suppressed its expression within prostate cancer cells in a dose-dependent manner. Current research has highlighted the hazardous influence of the estrogenic component to prostate cancer. We found that fingolimod (FTY720) could modulate the estrogenic micromilieu and interrupt its cross talk with sphingolipid metabolism.

Выбор врача: есть ли альтернатива гормональной терапии?

The breast pathology is associated with a female sex hormones failure. It can cause estrogen-dependent cancers. The problem of the mastopathy and mastodynia (fibrocystic breast disease, benign mammary dysplasia) therapy is not completely solved. The hormone therapy is associated with complications and has a large number of contraindications, which makes finding new strategies for the prevention and treatment of these diseases. The aim of the study was to compare the effects of hormonal and non-hormonal drugs. Combined application of capsules and cream Mammodol® in women with mastopathy and mastodynia leads to pronounced clinical, ultrasound and laboratory effects. This allows to recommend it as an alternative to hormone therapy. Key words: mastopathy, benign mammary dysplasia, estrogens, psychoemotional state, indole-3-carbinol, Vitex Agnus castus, hormone therapy, joint application, prolactin.

SMURF1 facilitates estrogen receptor \u0251 signaling in breast cancer cells

BackgroundEstrogen receptor alpha (ER alpha) is expressed in the majority of breast cancers and promotes estrogen-dependent cancer progression. ER alpha positive breast cancer can be well controlled by ER alpha modulators, such as tamoxifen. However, tamoxifen resistance is commonly observed by altered ER alpha signaling. Thus, further understanding of the molecular mechanisms, which regulates ER alpha signaling, is important to improve breast cancer therapy.MethodsSMURF1 and ER alpha protein expression levels were measured by western blot, while the ER alpha target genes were measured by real-time PCR. WST-1 assay was used to measure cell viability; the xeno-graft tumor model were used for in vivo study. RNA sequencing was analyzed by Ingenuity Pathway Analysis. Identification of ER alpha signaling was accomplished with luciferase assays, real-time RT-PCR and Western blotting. Protein stability assay and ubiquitin assay was used to detect ER alpha protein degradation. Immuno-precipitation based assays were used to detect the interaction domain between ER alpha and SMURF1. The ubiquitin-based Immuno-precipitation based assays were used to detect the specific ubiquitination manner happened on ER alpha.ResultsHere, we identify the E3 ligase SMURF1 facilitates ER alpha signaling. We show that depletion SMURF1 decreases ER alpha positive cell proliferation in vitro and in vivo. SMURF1 depletion based RNA-sequence data shows SMURF1 is necessary for ER alpha target gene expression in the transcriptomic scale. Immunoprecipitation indicates that SMURF1 associates with the N-terminal of ER alpha in the cytoplasm via its HECT domain. SMURF1 increases ER alpha stability, possibly by inhibiting K48-specific poly-ubiquitination process on ER alpha protein. Interestingly, SMURF1 expression could be induced via estradiol treatment.ConclusionsOur study reveals a novel positive feedback between SMURF1 and ER alpha signaling in supporting breast cancer growth. Targeting SMURF1 could be one promising strategy for ER alpha positive breast cancer treatment.

In vitro and in vivo antiproliferative activity of organo-nickel SCS-pincer complexes on estrogen responsive MCF7 and MC4L2 breast cancer cells. Effects of amine fragment substitutions on BSA binding and cytotoxicity.

A family of organonickel complexes has been prepared, fully characterized, and tested for their antiproliferative activity against estrogen-responsive human breast cancer cells (MCF7). The three SCS-type pincer ligands HL1, HL2, and HL3 and their corresponding Ni(ii) complexes NiL1, NiL2, and NiL3 have been synthesized and fully characterized, including by single crystal diffraction studies for the complexes. The complexes possess square planar geometry with two symmetrical 5-membered nickellacycles. Fluorescence spectroscopy, circular dichroism measurements, molecular modeling, colorimetric based assay and tumor transplantation studies were used to evaluate the protein binding and antiproliferative activities of these organometallic complexes both in vitro and in vivo. Fluorescence quenching was used to investigate bovine serum albumin (BSA) interaction at different temperatures (293, 303 and 313 K), and the results were analyzed using the classical Stern-Volmer equation, allowing us to propose a dynamic quenching mechanism. Studies in vitro on the antiproliferative activity of the three organonickel complexes against estrogen-responsive human breast cancer cells (MCF7) showed promising antitumor activity for NiL1 containing pyrrolidine fragments. In vivo administration of this compound significantly inhibits tumor growth in estrogen-dependent MC4L2 cancer cells in female BALB/c mice.

Safety Aspects of the Use of Quercetin as a Dietary Supplement.

The flavonoid quercetin is frequently found in low amounts as a secondary plant metabolite in fruits and vegetables. Isolated quercetin is also marketed as a dietary supplement, mostly as the free quercetin aglycone, and frequently in daily doses of up to 1000mgd-1 exceeding usual dietary intake levels. The present review is dedicated to safety aspects of isolated quercetin used as single compound in dietary supplements. Among the numerous published human intervention studies, adverse effects following supplemental quercetin intake have been rarely reported and any such effects were mild in nature. Published adequate scientific data for safety assessment in regard to the long-term use (>12 weeks) of high supplemental quercetin doses (≥1000mg) are currently not available. Based on animal studies involving oral quercetin application some possible critical safety aspects could be identified such as the potential of quercetin to enhance nephrotoxic effects in the predamaged kidney or to promote tumor development especially in estrogen-dependent cancer. Furthermore, animal and human studies with single time or short-term supplemental quercetin application revealed interactions between quercetin and certain drugs leading to altered drug bioavailability. Based on these results, some potential risk groups are discussed in the present review.

Sex differences in behavioral and neurochemical effects of gonadectomy and aromatase inhibition in rats

Aromatase inhibitors, which are widely used for the treatment of estrogen-dependent cancers, have been associated with psychiatric side effects ranging from mania to depression. In the present study, we investigated sex differences in the behavioral and neurochemical effects of aromatase inhibition on male and female, sham-operated or gonadectomized adult rats. Three weeks after surgery, rats received chronic treatment with the aromatase inhibitor letrozole or vehicle and were then subjected to the open field test, which assesses general activity. Half of the subjects were subsequently exposed to the stressful procedure of the forced swim test (FST), which is also a test of antidepressant activity. Aromatase activity was analyzed in the hypothalamus and testosterone and corticosterone were assayed in the blood serum of all rats. The hippocampus and prefrontal cortex (PFC) were analyzed for monoamine (noradrenaline, dopamine and serotonin), as well as amino acid (GABA, glutamate, glycine, taurine, alanine and histidine) levels. The observed decrease in hypothalamic aromatase activity confirmed the efficacy of letrozole treatment in both sexes. Moreover, letrozole enhanced testosterone levels in sham-operated females. In the open field test, females were overall more active and explorative than males and gonadectomy eliminated this sex difference. In the FST, females exhibited overall higher immobility than males and gonadectomy further enhanced this passive behavior in both sexes. However, sustained aromatase inhibition had no effect on open field and FST behaviors. Head shakes during FST, which were fewer in females than in males, were reduced by castration in males and by letrozole treatment in ovariectomized females, suggesting a role of testosterone and extra-gonadal estrogens in the expression of this behavior. Sustained aromatase inhibition also decreased noradrenaline and the dopaminergic turnover rates [DOPAC/DA, HVA/DA] in the hippocampus and PFC of male and female rats, irrespectively of gonadectomy. Moreover, letrozole treatment enhanced the serotonergic turnover [5HIAA/5HT] rate in the hippocampus of males and females, irrespectively of gonadectomy. Amino acid levels were not influenced by letrozole, but sex differences were demonstrated with higher levels in the PFC of females vs. males. Present findings suggest that the neuropsychiatric effects of aromatase inhibition can be attributed to the inhibition of extragonadal estrogen synthesis, presumably in the brain, and could be further associated with serotonergic and catecholaminergic changes in brain regions involved in mood and cognition. Importantly, present data could be linked with the neurobiology of affective side-effects in post-menopausal women receiving aromatase inhibitors.

SHARPIN stabilizes estrogen receptor α and promotes breast cancer cell proliferation.

Estrogen receptor α is expressed in the majority of breast cancers and promotes estrogen-dependent cancer progression. In our study, we identified the novel E3 ubiquitin ligase SHARPIN function to facilitate ERα signaling. SHARPIN is highly expressed in human breast cancer and correlates with ERα protein level by immunohistochemistry. SHARPIN expression level correlates with poor prognosis in ERα positive breast cancer patients. SHARPIN depletion based RNA-sequence data shows that ERα signaling is a potential SHARPIN target. SHARPIN depletion significantly decreases ERα protein level, ERα target genes expression and estrogen response element activity in breast cancer cells, while SHARPIN overexpression could reverse these effects. SHARPIN depletion significantly decreases estrogen stimulated cell proliferation in breast cancer cells, which effect could be further rescued by ERα overexpression. Further mechanistic study reveals that SHARPIN mainly localizes in the cytosol and interacts with ERα both in the cytosol and the nuclear. SHARPIN regulates ERα signaling through protein stability, not through gene expression. SHARPIN stabilizes ERα protein via prohibiting ERα protein poly-ubiquitination. Further study shows that SHARPIN could facilitate the mono-ubiquitinaiton of ERα at K302/303 sites and facilitate ERE luciferase activity. Together, our findings propose a novel ERα modulation mechanism in supporting breast cancer cell growth, in which SHARPIN could be one suitable target for development of novel therapy for ERα positive breast cancer.

Estrogen-like and tissue-selective effects of 7-methoxycoumarin from Ficus umbellata (Moraceae): an in vitro and in vivo study

BackgroundFicus umbellata is a medicinal plant previously shown to endow estrogenic properties. Its major component was isolated and characterized as 7-methoxycoumarin (MC). Noteworthy, coumarins and the respective active metabolite 7-hydroxycoumarin analogs have shown aromatase inhibitory activity, which is of particular interest in the treatment of estrogen-dependent cancers. The present work aimed at evaluating the estrogenic/antiestrogenic effects of MC in vitro and in vivo.MethodsTo do so, in vitro assays using E-screen and reporter gene were done. In vivo, a 3-day uterotrophic assay followed by a postmenopausal-like rat model to characterize MC as well as F. umbellata aqueous extract in ovariectomized Wistar rats was performed. The investigations focused on histological (vaginal and uterine epithelial height) and morphological (uterine wet weight, vagina stratification and cornification) endpoints, bone mass, biochemical parameters and lipid profile.ResultsMC induced a significant (p < 0.05) MCF-7 cell proliferation at a concentration of 0.1 μM, but did not inhibit the effect induced by estradiol in both E-screen and reporter gene assays. In vivo, MC treatment did not show an uterotrophic effect in both rat models used. However, MC (1 mg/kg) induced a significant increase (p < 0.01) of vaginal epithelial height. No significant change was observed with MC in abdominal fat weight, serum lipid levels and bone weight.ConclusionThese results suggest that MC has a weak estrogenic activity in vitro and in vivo that accounts only in part to the estrogenicity of the whole plant extract. MC could be beneficial with regard to vagina dryness as it showed a tissue specific effect without exposing the uterus to a potential tumorigenic growth.

Abstract 3121: Validation of protein kinase D1 as a prognostic factor and pharmacological target for the treatment of breast cancer

Abstract Tamoxifen-resistant ERα positive (ERα+) breast cancers and triple-negative breast cancers represent two aggressive subgroups of mammary tumors with limited treatment options and a poor prognosis. Thus, there is still an urgent need to develop and improve targeted pharmacological treatments against these estrogen-independent cancers. Recent data from our group suggest that the protein kinase D1 (PKD1) participates in the development and/or maintenance of the estrogen-independent phenotype in breast cancer. Indeed, up-regulating PKD1 conferred anchorage-independent growth to estrogen-dependent MCF-7 cells in the absence of β-estradiol. In addition, we found that high PKD1 mRNA levels were associated with a worse prognosis in tamoxifen-treated tumors. PKD1 is a serine-threonine kinase encoded by the prkd1 gene and belongs to the protein kinase D (PKD) family, a novel subgroup of the calcium and calmodulin-regulated kinases (CAMK). The PKD family also includes PKD2 and PKD3 which share a high homology with PKD1. The objective of the present study was to determine whether PKD1 could be a prognostic factor and/or a pharmacological target for the treatment of breast cancer. Expression of prkd1, prkd2 and prkd3 was analyzed by RT-qPCR in 527 breast cancer specimens (181 ERα- tumors including 102 triple negative tumors and 346 ERα+ tumors). We showed that higher prkd1 mRNA levels were associated with a lower metastasis-free survival in the overall breast cancer population (p=0.015). prkd1 prognostic value was even stronger in ERα- tumors (p=0.00042) and in triple negative tumors (p=0.0043). Conversely, prkd1 expression was not associated with prognosis in ERα+ tumors. Interestingly, high prkd2 and prkd3 mRNA levels were associated with a better prognosis in the entire cohort (p=0.0074 and p=0.028 respectively). We have developed PKD1 inhibitors in collaboration with the AB Science pharmaceutical company and screened the ability of thirty-five compounds to inhibit anchorage-independent growth of MCF-7 cells in the absence of estrogens. The antitumor activity of two PKD1 inhibitors is currently being evaluated in vivo against patient-derived breast cancer xenografts. In conclusion, our results show that high prkd1 expression is a bad-prognosis factor in breast cancer and that PKD1 could be a relevant therapeutic target for the treatment of breast cancer, in particular those harboring an estrogen-independent phenotype. Citation Format: Caroline Spasojevic, Marie Regairaz, Sophie Vacher, Franck Assayag, Jean Marc Ricort, Elisabetta Marangoni, Ivan Bièche, Christian Auclair. Validation of protein kinase D1 as a prognostic factor and pharmacological target for the treatment of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3121. doi:10.1158/1538-7445.AM2017-3121

Clinical relevance of Cyr61 expression in patients with hormone-dependent breast cancer.

Tumor resistance to endocrine therapy triggers estrogen-independent cancer progression, which is a major obstacle to the successful treatment of hormone receptor positive breast cancer (BC). The underlying molecular mechanisms of endocrine resistance are not fully understood yet. The matricellular protein cysteine-rich angiogenic inducer 61 (Cyr61) is associated with tumor invasiveness and the induction of tumorigenesis in various malignancies in vivo and the induction of estrogen-independence and endocrine therapy resistance in BC. The present study evaluated the potential effects and clinical relevance of Cyr61 expression levels in 67 patients with primary non-metastatic BC. Immunohistochemical analysis of formalin-fixed paraffin-embedded tissue sections was performed, and the association between Cyr61 protein expression and clinicopathological factors and survival was analyzed. Cyr61 overexpression was revealed to be significantly associated with a positive estrogen receptor (ER)/progesterone receptor (PR) status (P=0.016) and to the molecular subtype of BC (P=0.039). Compared with patients without Cyr61 overexpression, patients with Cyr61 overexpression exhibited an increased recurrence rate (30.6 vs. 22.6%) and decreased long-term survival (10-year overall survival, 62.9 vs. 69.7%); however, these associations did not reach statistically significant levels in Cox regression model analysis. Similar results were identified in the subgroup analysis of patients with ER/PR positive BC. These results indicate that Cyr61 serves a role in the development of endocrine therapy resistance in BC and is thus a potential therapeutic target to overcome endocrine therapy resistance. However, additional long-term survival analyses with large patient populations are required.

Development of a novel recombinant LHRH fusion protein for therapy of androgen and estrogen dependent cancers

LHRH based vaccines are promising candidates for therapy of androgen and estrogen dependent cancers. We report in this communication development of a novel recombinant protein vaccine candidate against LHRH. A synthetic gene was designed in which the codon sequence in the LHRH decapeptide was modified by substituting the codon for 6-glycine with that of l-leucine. Further the LHRH(6leu) gene was linked to heat-labile enterotoxin of E. coli (LTB) as carrier. This LHRH(6leu)-LTB gene was cloned into a prokaryotic expression vector under the control of inducible and strong bacteriophage T7 promoter to over-express LHRH(leu) fused to LTB as recombinant protein in E. coli. Recombinant LHRH(leu)-LTB protein of ∼14 kDa size, was purified from inclusion bodies using in-situ refolding on the column and Ni-NTA based immobilized affinity chromatography. Western blot confirmed the immunoreactivity of purified LHRH(leu)-LTB fusion protein with anti-LHRH monoclonal antibody. The vaccine protein was further characterized by mass spectroscopy, circular dichroism and fluorescence spectroscopy. This communication reports a recombinant LHRH fusion protein with potential for blocking of sex hormones production for eventual therapy of sex hormones dependent neoplasms.

Effects of bisphenol compounds on the growth and epithelial mesenchymal transition of MCF-7 CV human breast cancer cells

Bisphenol-A (BPA) has been considered as an endocrine disrupting chemical (EDC) because it can exert estrogenic properties. For bisphenol-S (BPS) and bisphenol-F (BPF) that are BPA analogs and substitutes, their risk to estrogen-dependent cancer has been reported rarely compared with the numerous cases of BPA. In this study, we examined whether BPA, BPS, and BPF can lead to the proliferation, migration, and epithelial mesenchymal transition (EMT) of MCF-7 clonal variant (MCF-7 CV) breast cancer cells expressing estrogen receptors (ERs). In a cell viability assay, BPA, BPS, and BPF significantly increased proliferation of MCF-7 CV cells compared to control (DMSO) as did 17β-estradiol (E2). In Western blotting assay, BPA, BPS, and BPF enhanced the protein expression of cell cycle progression genes such as cyclin D1 and E1. In addition, MCF-7 CV cells lost cell to cell contacts and acquired fibroblast-like morphology by the treatment of BPA, BPS, or BPF for 24 hours. In cell migration assay, BPA, BPS, and BPF accelerated the migration capability of MCF-7 CV cells as did E2. In relation with the EMT process, BPA, BPS, and BPF increased the protein expression ofN-cadherin, while they decreased the protein expression of E-cadherin. When BPA, BPS, and BPF were co-treated with ICI 182,780, an ER antagonist, proliferation effects were reversed, the expression of cyclin D1 and cyclin E1 was downregulated, and the altered cell migration and expression ofN-cadherin and E-cadherin by BPA, BPS, and BPF were restored to the control level. Thus, these results imply that BPS and BPF also have the risk of breast cancer progression as much as BPA in the induction of proliferation and migration of MCF-7 CV cells by regulating the protein expression of cell cycle-related genes and EMT markersvia the ER-dependent pathway.