ERα Gene Research Articles

Estrogen receptor alpha deficiency in cardiomyocytes reprograms the heart-derived extracellular vesicle proteome and induces obesity in female mice.

Dysregulation of estrogen receptor alpha (ERα) has been linked with increased metabolic and cardiovascular disease risk. Here, we generate and characterize cardiomyocyte-specific ERα knockout (ERαHKO) mice to assess the role of ERα in the heart. The most striking phenotype was obesity in female ERαHKO but not male ERαHKO mice. Female ERαHKO mice showed cardiac dysfunction, mild glucose and insulin intolerance and reduced ERα gene expression in skeletal muscle and white adipose tissue. Transcriptomic, proteomic, lipidomic and metabolomic analyses revealed evidence of contractile and/or metabolic dysregulation in heart, skeletal muscle and white adipose tissue. We show that heart-derived extracellular vesicles from female ERαHKO mice contain a distinct proteome associated with lipid and metabolic regulation, and have the capacity to metabolically reprogram the target skeletal myocyte proteome with functional impacts on glycolytic capacity and reserve. This multi-omics study uncovers a cardiac-initiated and sex-specific cardiometabolic phenotype regulated by ERα and provides insights into extracellular vesicle-mediated interorgan communication.

ER-/PR+ breast cancer is controlled more effectively with an inflammatory inhibitor than hormonal inhibitor.

The anti-estrogen tamoxifen is a highly effective hormonal therapy for hormonal-positive (HR+) breast cancer patients; however, the estrogen receptor-negative, progesterone receptor-positive (ER-/PR+) subtype does not give the benefits of tamoxifen. Therefore ER-/PR+ breast cancer has a poor clinical outcome, and novel drug therapy for ER-/PR+ breast cancer could benefit these patients. 53,805 gene expressions were characterized into HR+ BC and triple-negative breast cancer (TNBC) and analyzed through Breast Cancer Gene Expression Miner in 4319 breast cancer patient samples. The clinical outcomes including overall survival, distant metastasis-free survival, and relapse-free survival were obtained from the PrognoScan database containing 1190 human breast cancer patient samples. To determine the function of ERα and inflammation-related genes such as USP1, CDC20, and CASP1, we used the CRISPR-Cas9 system or gene knockdown (KD) system. To check tumor cell proliferation and migration of ERα KO breast cancer cell line, we used tamoxifen and the inflammation inhibitor Ac-YVAD-CHO. For further confirmation, cancer growth was checked with the inflammation inhibitor in ERα KO breast cancer cell line using a three-dimensional (3D) organoid tissue culture system (ex vivo). We found that gene expression in ER-/PR+ hormonal-positive breast cancer is positively related to ER-/PR- very similar to TNBC, not other HR+ breast cancer using a 4319 breast cancer patient database. Especially, inflammation-related genes, USP1, CDC20, and CASP1, which are highly expressed in TNBC, are also upregulated in ER-/PR+ HR+ breast cancer. Suppression of USP1, CDC20, and CASP1 inhibited tumor cell growth and metastasis in ERα KO (ER-/PR +) cell lines. Interestingly, loss of ERα in HR+ cell lines is not responsive to tamoxifen, but highly sensitive to the inflammation inhibitor, Ac-YVAD-CHO. In in vitro and ex vivo (3D organoid) models, inflammation inhibitor-specific blocks ER-/PR+ tumor proliferation and migration. These findings suggest that an inflammation inhibitor might be a potential option for therapy for ER-/PR+ HR breast cancer patients.

Abstract P3-11-05: Efficacy of Gonadal steroids on aromatase inhibitor-resistant ER+ breast cancer: Insights from single-cell trajectory analysis of a patient-derived xenograft model

Abstract Background: While estrogen typically promotes the progression of hormone-dependent breast cancer via the activation of estrogen receptor (ER)-α, estrogen-induced suppression of ER+ breast cancer has been clinically observed. Our previous study demonstrated that estrogen increased the percentage of cells expressing IL24, linking to the estrogen-dependent growth inhibition of an ER+ aromatase inhibitor (AI)-resistant tumor (Mori et al. Cancers, 2021). To continue our evaluation, we investigated whether progesterone (P4) and dihydrotestosterone (DHT) would affect the growth of this estrogen-suppressive tumor. Methods: An estrogen-suppressive patient-derived xenograft (PDX) model (named GS3) was established from an AI resistant ER+/HER2– brain metastatic breast cancer. E2 (1mg), P4 (10mg), DHT (12.5mg), or placebo pellets were implanted in mice carrying GS3 for in vivo drug efficacy examination. Beside tumor growth response, immunohistochemistry (IHC) and RNA sequencing of PDX specimens were conducted to decipher molecular changes after each treatment. The single-cell RNAseq analysis was further performed to examine gene expression profiles in individual cells. Results: ERα, ERβ, Progesterone receptor (PR), and androgen receptor (AR) genes in GS3 are wild-type and are not amplified. Measurements of tumor volume showed that E2, E2+P4, and DHT suppressed the growth of GS3. The Tumor growth was not modulated by P4 treatment. IHC indicated that the number of Ki-67+ cells were decreased after E2, E2+P4, and DHT treatments, but were not changed after P4 treatment. PR+ cells appeared after E2 and E2+P4 treatments, and the AR+ cells increased after DHT treatment. Bulk-RNA sequencing indicated that E2 and E2+P4 treatments resulted in comparable gene expression patterns, while those of placebo and P4-treated tumors were similar. GSEA analysis showed that in E2 treatment, the hallmark estrogen response gene sets were upregulated, and the hallmark G2M checkpoint gene set was downregulated. However, in DHT treatment, the hallmark interferon alpha/gamma response gene sets and androgen response gene set were upregulated, and the hallmark TNFA signaling via NFkB gene set was downregulated. Single-cell RNA sequencing analysis of Placebo/E2/DHT samples revealed 9 clusters; cells from E2-treated and DHT-treated tumors were placed in different clusters based on principle component analysis of Highly Variable Genes, although both E2 and DHT treatments resulted in tumor regression. DHT promoted cell cycle arrest, but it did not increase IL24 expression. The hallmark oxidative phosphorylation and androgen response gene sets were upregulated in all clusters from DHT-treated tumors. Trajectory analysis of single cells revealed that three major branches associated with clusters selective to the treatments of placebo, E2, and DHT were separated from a common branch which are consist of G2M phase cells. Conclusions: E2 and DHT were effective suppressor of GS3, but not P4. Based on the results from IHC, bulk-RNAseq, and Single-cell RNAseq, the mechanism of DHT-induced tumor regression is different from that by E2. Our results suggest that DHT/E2 could be treatment options for patients with relapsed ER+ AI-resistance breast cancer. Citation Format: Hitomi Mori, Kohei Saeki, Gregory Chang, Jinhui Wang, Xiwei Wu, Noriko Kanaya, George Somlo, Shiuan Chen. Efficacy of Gonadal steroids on aromatase inhibitor-resistant ER+ breast cancer: Insights from single-cell trajectory analysis of a patient-derived xenograft model [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-11-05.

Association of the KISS1, LIN28B, VDR and ERα gene polymorphisms with early and fast puberty in Chinese girls

Objectives To explore the association of KISS1, LIN28B, vitamin D receptor (VDR), and estrogen receptor α (ERα) gene polymorphisms and the risk of early with fast puberty (EFP) risk, and with hormone levels in EFP cases, in Chinese girls. Methods The analysis was based on the data of 141 girls with EFP and 152 girls without EFP. Clinical features were documented, and all SNP genotyping was conducted using SNaPshot method. Statistical analysis was performed to assess the association of the SNPs with EFP risk, and with hormone levels in EFP cases. Results There was a significant association between rs7759938-C polymorphism in the LIN28B gene and the risk for EFP in the recessive (TT + CT vs. CC) model (p = 0.040). Remarkably, rs5780218-delA polymorphism in the KISS1 gene and rs2234693-C polymorphism in the ERα gene were significantly associated with peak LH (luteinizing hormone) levels (p = 0.008, 0.045) and peak LH/FSH (follicle-stimulating hormone) ratio (p = 0.007, 0.006). Additionally, on 7 of the 8 variant loci the alleles associated with increased levels of both peak LH levels and peak LH/FSH ratio in EFP cases were also associated with increased CPP risk. Conclusions Our findings indicate that rs7759938-C polymorphism in the LIN28B gene might have a protective effect on EFP susceptibility. The most striking findings of this study is that, rs5780218-delA polymorphism in the KISS1 gene and rs2234693-C polymorphism in the ERα gene influenced levels of GnRH-stimulated peak LH and LH/FSH ratio, and in general CPP risk genes might also contributes to the abnormality of hormonal levels in EFP.

The Aryl Hydrocarbon Receptor Ligand 6-Formylindolo(3,2-b)carbazole Promotes Estrogen Receptor Alpha and c-Fos Protein Degradation and Inhibits MCF-7 Cell Proliferation and Migration

Introduction: Worldwide, breast cancer is the most common cancer in women and is the main cause of death among all neoplasia in this group. Luminal A breast cancer represents approximately 70% of all breast cancers and is treated with hormone therapies targeting estrogen receptor alpha (ERα). Unfortunately, patients develop drug resistance leading to recurrence of neoplasia due to estrogen-independent ERα reactivation. Therefore, it is crucial to identify new molecular targets downstream ERα signaling pathway that allows the implementation of better treatments to improve the outcome of breast cancer patients. Overexpression of c-Fos, an ERα gene target, has been associated with increased cell motility, malignancy, metastasis, and invasion while its neutralization results in decreased breast cancer tumorigenesis. The aryl hydrocarbon receptor (AHR) ligands halogenated and polycyclic aromatic hydrocarbons, highly toxic compounds, down regulate c-Fos and ERα levels. The present study aimed to evaluate whether 6-formylindolo(3,2-b)carbazole (FICZ), a no toxic AHR agonist, modifies c-Fos levels in MCF-7 mammary carcinoma cells as well as to determine its effects on cell proliferation and migration. In addition, the possible mechanism through which FICZ mediates c-Fos levels in MCF-7 cells was investigated. Methods: Initially, the effect of FICZ on c-Fos mRNA and protein levels in MCF-7 cells, untreated or treated with estradiol, was evaluated by qPCR and Western blot. 2,3,7,8-Tetrachloro-dibenzo-p-dioxin, an AHR prototype agonist, was used as a positive control. Next, we examined the effect of FICZ on MCF-7 cell proliferation and migration by cell counting, MTT, 3H-thymidine incorporation, and scratch-wound assays. Finally, the involvement of proteasome 26S on ERα and c-Fos protein degradation was investigated by the use of MG132 and Western blot. Results: The data show that FICZ treatment downregulates c-Fos mRNA and protein levels, most likely by promoting ERα proteasome degradation, blocking MCF-7 cell proliferation and migration. The results also demonstrate that liganded ERα was required for FICZ-mediated ERα degradation. Conclusions: Activation of AHR results in a decreased MCF-7 cell proliferation and migration by ERα and c-Fos down regulation. Targeting AHR might be a promising therapy for breast cancer treatment, particularly when estrogen-independent ERα reactivation presents.

Detection of Calreticulin as a Candidate Prognostic Biomarker in Invasive Breast Carcinoma from a Biological Scaffold-Based 3D Co-culture System

Introduction: Breast cancer is the most common cancer in women and the world’s second leading cause of death in women, after lung cancer. Calreticulin (CRT), an endoplasmic reticulum (ER) multipurpose protein, has been proposed as a potential biomarker for breast cancer. However, reports on the correlation between CRT expression and cell invasiveness in breast cancer micro-tissues are scarce. Thus, in the current study, we analyzed the potential correlation between CRT and invasiveness of breast cancer in a biological scaffold-based 3D co-culture system. Methods: MCF7, MDA-MB-231 and MCF-10A breast cell lines were co-cultured in a 3-dimensional (3D) system with MRC-5 lung fibroblast cell line in the cell density ratio of 3:1. Thereafter, calreticulin gene and protein expression levels were determined based on quantitative reverse transcription-polymerase chain reaction (RT-qPCR) and immunohistochemistry, respectively. Moreover, via RT-qPCR analysis, the gene expression levels of calreticulin-related candidate metastasis genes in breast cancer micro-tissues were carried out. Results: The results showed occasional foci of lumen-like morphology in the non-cancerous breast micro-tissues and the formation of solid clusters for breast cancer micro-tissues. Moreover, immunohistochemistry results revealed protein expression of calreticulin in non-cancerous and cancerous breast micro-tissues with cytoplasmic and nucleic acid localizations. It was found that PCMT1 and ER-α genes were significantly downregulated (p < 0.01) in invasive breast cancer micro-tissues. Conclusion: This study suggests that CRT and CRT-related candidate metastasis genes may potentially serve as prognostic biomarkers in invasive breast carcinoma.

P38 and ERK1/2-Dependent Activation of c-Jun Is Required for the Downregulation of Oxidative Stress-Induced ERα in Hypothalamic Astrocytes

Introduction: Gonadotropin-releasing hormone (GnRH) is a hypothalamic neuropeptide that plays important roles in the female fertility. Accumulating evidence suggests that ERα present in the astrocytes of the hypothalamus region is essential for production of GnRH. The astrocytes display age-related senescence associated to oxidative stress induced by the estrogen metabolites. However, it is still unclear whether and how ERα expression changes during astrocyte aging. Methods: Immunofluorescence was performed to analyze the ERα gene levels in hypothalamic astrocytes of naturally aging C57BL/6J female mice. We employed an oxidative stress cell model receiving 2-hydroxyestradiol (2OH-E2) intervention to confirm the downregulation of ERα expression in primary astrocytes. Western blot analysis was used to explore which oxidative stress signaling pathways induced loss of the ERα gene. Finally, ChIP-qPCR was employed to evaluate whether the c-Jun protein is able to regulate ERα gene expression. Results: Compared to young mice, we found that the ERα expression of mid-aged mice was significantly decreased. In hypothalamic astrocytes, 2OH-E2 treatment significantly reduced the expression of the ERα gene. Moreover, we observed that transcription factor c-Jun could directly inhibit transcriptional ERα gene expression and might also reduce it by decreasing H3K27 acetylation at promoter regions. Administration of the antioxidants Rg1 and astaxanthin significantly attenuated the decrease in ERα gene expression induced by oxidative stress. Conclusions: The current data demonstrate that oxidative stress leads to loss of ERα involving the activation of the p38 and ERK1/2 pathways and the induction of the c-Jun protein in hypothalamic astrocytes. C-Jun protein regulates ERα gene expression via direct transcriptional repression or involving histone acetylation modifications at ERα gene promoter sites.

ИССЛЕДОВАНИЕ ERα/BGLI SNP ПОЛИМОРФИЗМА У КРУПНОГО РОГАТОГО СКОТА, АССОЦИРОВАННОГО С РЕПРОДУКТИВНОЙ ФУНКЦИЕЙ

The article presents the results of genotyping of Holstein cows of Bayserke-Agro LLP, Talgar district, Almaty region, for the estrogen receptor gene locus, ERα/BGLI SNP polymorphism. In the studied animals, the distribution of genetic variants by the ERα gene locus was established; The results of DNA testing indicate that the DNA of the tested cows has a violation of gene balance, in cows the G allele is predominantly found, the frequency of which was 0.83. The reproductive function of 120 cows with different genetic variants of the ERα gene was monitored and it was found that animals with good indicators of reproductive function had a heterozygous AG genotype. Also, to determine the associative effect of ERα gene alleles on the reproductive ability of cows, the index of non-return of cows on the 58th day after artificial insemination was determined, and according to this criterion, cows with a heterozygous AG genotype had advantages compared to animals with homozygous AA and GG genotypes. Conducting genotyping of cows at the ERα gene locus by PCR-RFLP analysis allows determining the genotype of animals and ERα/BGLI SNP polymorphism is recommended as a DNA marker of reproductive function in cows.

BDE-47, -99, -209 and Their Ternary Mixture Disrupt Glucose and Lipid Metabolism of Hepg2 Cells at Dietary Relevant Concentrations: Mechanistic Insight through Integrated Transcriptomics and Proteomics Analysis.

Polybrominated diphenyl ethers (PBDEs) are persistent organic chemicals implied as flame retardants. Humans are mainly exposed to BDE-47, -99, and -209 congeners by diet. PBDEs are metabolic disruptors with the liver as the main target organ. To investigate their mode of action at a human-relevant concentration, we exposed HepG2 cells to these congeners and their mixture at 1 nM, analyzing their transcriptomic and proteomic profiles. KEGG pathways and GSEA Hallmarks enrichment analyses evidenced that BDE-47 disrupted the glucose metabolism and hypoxia pathway; all the congeners and the MIX affected lipid metabolism and signaling Hallmarks regulating metabolism as mTORC1 and PI3K/AKT/MTOR. These results were confirmed by glucose secretion depletion and increased lipid accumulation, especially in BDE-47 and -209 treated cells. These congeners also affected the EGFR/MAPK signaling; further, BDE-47 enriched the estrogen pathway. Interestingly, BDE-209 and the MIX increased ERα gene expression, whereas all the congeners and the MIX induced ERβ and PPARα. We also found that PBDEs modulated several lncRNAs and that HNRNAP1 represented a central hub in all the four interaction networks. Overall, the PBDEs investigated affected glucose and lipid metabolism with different underlying modes of action, as highlighted by the integrated omics analysis, at a dietary relevant concentration. These results may support the mechanism-based risk assessment of these compounds in relation to liver metabolism disruption.

Evaluation of tooth eruption rate of incisor teeth in rats with estrogen deficiency.

To assess the influence of estrogen deficiency on tooth eruption rate (TER) and gene expression of estrogen receptor alpha and beta (ERα and ERβ) in the odontogenic region of teeth with continuous formation in a rat model. Ovariectomies (OVX; n = 25) and sham surgeries (SHAM; n = 25) were performed in female Wistar rats when animals were 25days old. The TER of the lower incisors, both in impeded (hyperfunction condition) and unimpeded (trimmed incisal edge-hypofunction condition) conditions, was evaluated using standardized digital photographs acquired every 48-72h for 3weeks (35th-53rd day of life), using a camera coupled to a stereomicroscope. Quantitative real-time PCR was performed to evaluate the relative gene expression of ERα and ERβ in the odontogenic region. The OVX group showed a significant reduction in TER when compared to the SHAM group, only in the impeded condition (p = 0.03). There was no statistically significant difference between the groups in ERα gene expression (p = 0.33). ERβ showed a significantly higher gene expression in the OVX group (p ≤ 0.05). Estrogen deficiency decreases TER in teeth under impeded condition. Estrogen deficiency also increases ERβ gene expression in the odontogenic region of teeth with continuous formation. Hormonal disturbances affecting estrogen levels can cause alterations in dental formation and teeth eruption.

Correlation between ERα gene polymorphism and multiple sclerosis and neuromyelitis optica.

Objective:To study the polymorphism distribution of estrogen receptor (ER) α gene and the correlation between different types of polymorphism in multiple sclerosis (MS) and neuromyelitis optica (NMO) patients.Methods:Forty-six cases of MS and NMO diagnosed from June 2018 to December 2019 were collected. Peripheral venous blood samples were collected. The patient’s gender, age of onset, course of disease, and other clinical data were recorded. Fifty-eight healthy volunteers of the same age and sex were selected. By means of Pvu II and Xba I restriction fragment length polymorphism enzyme recognition sites of ER α gene, polymerase chain reaction-restriction fragment length polymorphism analysis was conducted.Results:There was no significant difference in the frequency distribution of ER α gene’s PP, Pp, and pp genotype between MS and NMO case group and control group (P = .598). Frequency distribution of ER α gene’s XX, Xx, and xx was statistically significant between MS and NMO case group and control group (P = .021). Among them, distribution of Xx and Xx gene frequency between patient group and the control group was statistically significant (P = .001, OR = 4.622, 95% CI: 1.803–11.852). There was no significant correlation between ER α genotypes and the onset age in patient group (P > .05). The difference was statistically significant in disease duration of XX and Xx genotype (P = .006). The comparison of Xx and xx genotype frequency distribution in gender exists a difference(P = .047, OR = 7.500, 95% CI: 1.023–54.996).Conclusions:Xba I gene polymorphisms in the ER α gene have correlation with MS and NMO. Xba I gene could be a risk factor of MS and NMO pathogenesis, especially the women with Xx genotype are more vulnerable. Xba I gene polymorphisms in the ER α gene may impact the disease duration of MS and NMO, or rather, the disease duration of Xx genotype persists longer than Xx genotype. Pvu II gene polymorphisms in the ER α gene has no correlation with MS and NMO.

Hypomorphism of a Novel Long ERα Isoform Causes Severe Reproductive Dysfunctions in Female Mice.

Estrogen receptor alpha (ERα)-mediated estrogen signaling plays a pivotal role in both reproductive and nonreproductive functions. Transcriptional regulation of the ERα gene is highly complex, with multiple transcript variants being differentially produced across the tissues. However, tissue-specific variation and physiological specificity of the ERα variants are not yet fully understood. In an attempt to generate a Cre-dependently restorable ERα-null mouse for functional genetic studies, we unexpectedly produced ERα hypomorphic mice with biased downregulation of a previously unappreciated long ERα isoform that is enriched in the female reproductive organs (uterus and ovaries) and the pituitary but minimally expressed in the brain. Female homozygous mutant mice were capable of pregnancy but displayed irregular estrus cycle and rarely kept newborn pups alive. No significant morphological and pathological changes in reproductive system or disruption of body weight homeostasis were seen in female homozygous mutant mice. Collectively, our results define a tissue-specific enriched long ERα isoform and its preferential role in female reproductive function rather than body weight homeostasis.

Evaluation of antitumoral effect of Hibiscus sabdariffa extract on human breast cancer cells

BackgroundBreast cancer is the most frequent tumor in women. Natural substances represent an important source of innovative therapeutic solutions, eventually integrating or substituting conventional drugs and chemicals. Hibiscus sabdariffa L. is a plant of the Malvaceae family that has raised interest thanks to its anti-inflammatory, antioxidant and anticancer effects. In this work, we evaluated the antitumoral effects of an enriched fraction of Hibiscus sabdariffa L. extract (HsEF) in two human breast cancer cell lines, MCF-7(ERα +) and MDA-MB-231 (triple negative).Methods and resultsCell viability was assessed by MTT and Trypan blue assays. HsEF reduced both cell lines viability in a dose and time dependent manner and this effect results irreversible. In MCF-7 cells immunofluorescence experiments, demonstrated that HsEF induced ERα trans-location from nucleus to perinuclear area and in cytoplasmic compartment. qRT-PCR and western blotting high-lighted that HsEF reduced ERα, BRCA1 and caveolin1 gene and protein expression in MCF-7 cells, but not in MDA-MB-231 cells. Moreover, we demonstrated that HsEF reduced proteasome activity, an increased autophagy, impair migration and invasion in both cell lines.ConclusionsOur data suggest HsEF has an antitumoral effects on both breast tumor cells examined and that ERα involvement could explain the differences observed between the two cell lines.

Epigenetic restoration and activation of ERβ: an inspiring approach for treatment of triple-negative breast cancer

BackgroundTriple-negative breast cancer (TNBC) is one of the most aggressive subtypes of breast cancer. TNBC lacks targeted therapy receptors, rendering endocrine and HER2-targeted therapies ineffective. TNBC is typically treated with cytotoxic chemotherapy followed by surgery. Targeting epigenetic modifications could potentially be a new effective TNBC target therapy. The aim of this study is to examine the effects of epigenetic drugs, decitabine as DNA methyltransferase inhibitor (DNMTI) and vorinostat as histone deacetylase inhibitor (HDACI), and the ERβ agonist DPN on ERα and ERβ re-expressions in the MDA-MB-231 cells as a model of TNBC.MethodsUsing MTT assay, the IC50 of decitabine, vorinostat, and DPN on MDA-MB-231 cells were determined. The effects of all drugs alone or in combinations on MDA-MB-231 cells were evaluated. qRT-PCR was used to determine ERα & ERβ gene expression. Caspase-3 activity and the protein expression levels of VEGF, Cyclin D1, and IGF-1 were assessed.ResultsBoth ERα and ERβ mRNA were re-expressed in different high levels in all treated groups, especially in the triple therapy group compared with control. Significantly, the triple drugs therapy showed the lowest levels of VEGF, Cyclin D1, and IGF-1 and the highest level of Caspase-3 activity, indicating a possible antitumor effect of ERβ activation through decreasing proliferation and angiogenesis and increasing apoptosis in MDA-MB-231 cells.ConclusionsThe antiproliferative effect of ERβ could be retained when co-expressed with Erα using a powerful epigenetic combination of Decitabine and vorinostat with DPN.

Gold nanoparticles conjugated DNA-tile nanomaterials for simultaneous delivery of morpholino antisense oligonucleotides and doxorubicin

Current therapies have limited treatment effect on cancer. Combinational delivery of multiple therapeutics has been commonly investigated to enhance the therapeutic outcome of the disease. The carrier systems have the great importance for effective co-delivery of the multiple drugs. In this report, we demonstrated a DNA nanomaterial for simultaneous delivery of morpholino antisense oligonucleotides (AONs) and doxorubicin to breast cancer cells. DNA-tile nanostructures were prepared by embedding morpholino oligos against ERα and HER2 genes and the morpholino embedded DNA-tiles were modified with gold nanoparticles by sticky-ended association to obtain increased internalization. Then the DNA-carrier was loaded with doxorubicin. Morpholino and doxorubicin therapeutics were successfully delivered to breast cancer cells. It was observed that the co-delivery system showed 50% enhanced intracellular uptake of the chemotherapy drug, better cytotoxic effect and significantly higher level of cell cycle arrest than free doxorubicin at targeted receptor-positive breast cancer cells. The obtained results indicated that the proposed nanosystem is an effective delivery vehicle for combinational cancer therapies and increased anti-cancer activity of chemotherapy.

Integrated biomarker responses in European seabass Dicentrarchus labrax (Linnaeus, 1758) chronically exposed to PVC microplastics

Few studies evaluated long-term effects of polyvinyl chloride (PVC) microplastics (MPs) ingestion in fish. The present study aimed to investigate the integrated biomarker responses in the liver and blood of 162 European seabass, Dicentrarchus labrax, exposed for 90 days to control, virgin and marine incubated PVC enriched diets (0.1 % w/w) under controlled laboratory condition.Enzymatic and tissue alterations, oxidative stress, gene expression alterations and genotoxicity were examined. Additives and environmental contaminants levels in PVC-MPs, control feed matrices and in seabass muscles were also detected.The results showed that the chronic exposure at environmentally realistic PVC-MPs concentrations in seabass, cause early warning signs of toxicological harm in liver by induction of oxidative stress, the histopathological alterations and also by the modulation of the Peroxisome proliferator-activated receptors (PPARs) and Estrogen receptor alpha (ER-α) genes expression. A trend of increase of DNA alterations and the observation of some neoformations attributable to lipomas suggest also genotoxic and cancerogenic effects of PVC.This investigation provides important data to understand the regulatory biological processes affected by PVC-MPs ingestion in marine organisms and may also support the interpretation of results provided by studies on wild species.

The Isoforms of Estrogen Receptor Alpha and Beta in Thyroid Cancer

The incidence of thyroid cancer was predominant in women, indicating that the sex hormone may have a role in thyroid cancer development. Generally, the sex hormone exerts its function by binding to the correspondent nuclear receptors. Therefore, aberrant of these receptors may be involved in the development of thyroid cancer. Estrogen receptor alpha (ERα) and beta (ERβ), two main estrogen receptors, have been reported to have an important role in the pathogenesis of thyroid cancer. When the ERα and ERβ genes undergo the alternative RNA splicing, some ERα and ERβ isoforms with incomplete functional domains may be formed. To date, several isoforms of ERα and ERβ have been identified. However, their expression and roles in thyroid cancer are far from clear. In this review, we summarized the expressions and roles of ERα and ERβ isoforms in thyroid cancer, aiming to provide the perspective of modulating the alternative RNA splicing of ERα and ERβ against thyroid cancer.

YAP inhibits ERα and ER+ breast cancer growth by disrupting a TEAD-ERα signaling axis

Hippo signaling restricts tissue growth by inhibiting the transcriptional effector YAP. Here we uncover a role of Hippo signaling and a tumor suppressor function of YAP in estrogen receptor positive (ER+) breast cancer. We find that inhibition of Hippo/MST1/2 or activation of YAP blocks the ERα transcriptional program and ER+ breast cancer growth. Mechanistically, the Hippo pathway transcription factor TEAD physically interacts with ERα to increase its promoter/enhancer occupancy whereas YAP inhibits ERα/TEAD interaction, decreases ERα occupancy on its target promoters/enhancers, and promotes ERα degradation by the proteasome. Furthermore, YAP inhibits hormone-independent transcription of ERα gene (ESR1). Consistently, high levels of YAP correlate with good prognosis of ER+ breast cancer patients. Finally, we find that pharmacological inhibition of Hippo/MST1/2 impeded tumor growth driven by hormone therapy resistant ERα mutants, suggesting that targeting the Hippo-YAP-TEAD signaling axis could be a potential therapeutical strategy to overcome endocrine therapy resistance conferred by ERα mutants.

Activity of Estrogen Receptor β Agonists in Therapy-Resistant Estrogen Receptor-Positive Breast Cancer.

BackgroundAmong women, breast cancer is the leading cause of cancer-related death worldwide. Estrogen receptor α-positive (ERα+) breast cancer accounts for 70% of all breast cancer subtypes. Although ERα+ breast cancer initially responds to estrogen deprivation or blockade, the emergence of resistance compels the use of more aggressive therapies. While ERα is a driver in ERα+ breast cancer, ERβ plays an inhibitory role in several different cancer types. To date, the lack of highly selective ERβ agonists without ERα activity has limited the exploration of ERβ activation as a strategy for ERα+ breast cancer.MethodsWe measured the expression levels of ESR1 and ESR2 genes in immortalized mammary epithelial cells and different breast cancer cell lines. The viability of ERα+ breast cancer cell lines upon treatments with specific ERβ agonists, including OSU-ERb-12 and LY500307, was assessed. The specificity of the ERβ agonists, OSU-ERb-12 and LY500307, was confirmed by reporter assays. The effects of ERβ agonists on cell proliferation, cell cycle, apoptosis, colony formation, cell migration, and expression of tumor suppressor proteins were analyzed. The expression of ESR2 and genes containing ERE-AP1 composite response elements was examined in ERα+ human breast cancer samples to determine the correlation between ESR2 expression and overall survival and that of putative ESR2-regulated genes.ResultsIn this study, we demonstrate the efficacy of highly selective ERβ agonists in ERα+ breast cancer cell lines and drug-resistant derivatives. ERβ agonists blocked cell proliferation, migration, and colony formation and induced apoptosis and S and/or G2/M cell-cycle arrest of ERα+ breast cancer cell lines. Also, increases in the expression of the key tumor suppressors FOXO1 and FOXO3a were noted. Importantly, the strong synergy between ERβ agonists and ERα antagonists suggested that the efficacy of ERβ agonists is maximized by combination with ERα blockade. Lastly, ESR2 (ERβ gene) expression was negatively correlated with ESR1 (ERα gene) and CCND1 RNA expression in human metastatic ERα+/HER2- breast cancer samples.ConclusionOur results demonstrate that highly selective ERβ agonists attenuate the viability of ERα+ breast cancer cell lines in vitro and suggest that this therapeutic strategy merits further evaluation for ERα+ breast cancer.

Bone Mass and Osteoblast Activity Are Sex-Dependent in Mice Lacking the Estrogen Receptor α in Chondrocytes and Osteoblast Progenitor Cells.

While estrogen receptor alpha (ERα) is known to be important for bone development and homeostasis, its exact function during osteoblast differentiation remains unclear. Conditional deletion of ERα during specific stages of osteoblast differentiation revealed different bone phenotypes, which were also shown to be sex-dependent. Since hypertrophic chondrocytes can transdifferentiate into osteoblasts and substantially contribute to long-bone development, we aimed to investigate the effects of ERα deletion in both osteoblast and chondrocytes on bone development and structure. Therefore, we generated mice in which the ERα gene was inactivated via a Runx2-driven cyclic recombinase (ERαfl/fl; Runx2Cre). We analyzed the bones of 3-month-old ERαfl/fl; Runx2Cre mice by biomechanical testing, micro-computed tomography, and cellular parameters by histology. Male ERαfl/fl; Runx2Cre mice displayed a significantly increased cortical bone mass and flexural rigidity of the femurs compared to age-matched controls with no active Cre-transgene (ERαfl/fl). By contrast, female ERαfl/fl; Runx2Cre mice exhibited significant trabecular bone loss, whereas in cortical bone periosteal and endosteal diameters were reduced. Our results indicate that the ERα in osteoblast progenitors and hypertrophic chondrocytes differentially contributes to bone mass regulation in male and female mice and improves our understanding of ERα signaling in bone cells in vivo.