Genes In MCF-7 Cells Research Articles

Phytocompounds-based therapeutic approach: Investigating curcumin and green tea extracts on MCF-7 breast cancer cell line

BackgroundBreast cancer (BC) has transcended lung cancer as the most common cancer in the world. Due to the disease's aggressiveness, rapid growth, and heterogeneity, it is crucial to investigate different therapeutic approaches for treatment. According to the World Health Organization (WHO), Plant-based therapeutics continue to be utilized as safe/non-toxic complementary or alternative treatments for cancer, even in developed countries, regardless of how cutting-edge conventional therapies are. Despite their low bioavailability, curcumin (CUR) and green tea (GT) represent safer therapeutic options. Due to their potent molecular-modulating properties on various cancer-related molecules and signaling pathways, they are considered gold-standard therapeutic agents and have been incorporated into the development of one or more therapeutic strategies of BC treatment. MethodsWe investigated the modulatory role of CUR and GT extracts on significant multi molecular targets in MCF-7 BC cell line to assess their potential as BC multi-targeting agents. We analyzed the phytocompounds in GT leaves using High-performance liquid chromatography (HPLC) and Gas chromatography-mass spectrometry (GC-MS) techniques. The mRNA expression levels of Raf-1, Telomerase, Tumor necrosis factor alpha (TNF-α) and Interleukin-8 (IL-8) genes in MCF-7 cells were quantified using quantitative real-time PCR (qRT-PCR). The cytotoxicity of the extracts was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and the released Lactate dehydrogenase (LDH), a valuable marker for identifying the programmed necrosis (necroptosis). Additionally, the concentrations of the necroptosis-related proinflammatory cytokines (TNF-α and IL-8) were measured using enzyme-linked immunosorbent assay (ELISA). ResultsIn contrast to the GT, the results showed the anticancer and cytotoxic properties of CUR against MCF-7 cells, with a relatively higher level of released LDH. The CUR extract downregulated the oncogenic Raf-1, suppressed the Telomerase and upregulated the TNF-α and IL-8 genes. Results from the ELISA showed a notable increase in IL-8 and TNF-α cytokines levels after CUR treatment, which culminated after 72 h. ConclusionsAmong both extracts, only CUR effectively modulated the understudy molecular targets, achieving multi-targeting anticancer activity against MCF-7 cells. Moreover, the applied dosage significantly increased levels of the proinflammatory cytokines, which represent a component of the cytokines-targeting-based therapeutic strategy. However, further investigations are recommended to validate this therapeutic approach.

Impact of Acute or Chronic Acidosis and Hypoxia on Gene Expression Patterns in Tumour Cells: Potential Functional Implications.

Tumours often exhibit pronounced hypoxia and hereby extracellular acidosis due to intensified glycolysis. Since metabolic parameters can modulate gene expression, the aim of the study was to analyse changes in gene expression patterns induced by acute (24h) acidosis or hypoxia and also in tumour cells adapted to long-term acidosis (5weeks). Three tumour cell lines (AT1 prostate carcinoma, MCF-7, and MDA-MB-231 breast carcinoma) were exposed to acidosis (pH6.6) or hypoxia (pO2 1.5mmHg) for 24h. For long-term acidosis, AT1 tumour cells were continuously cultured at pH6.6 for 5weeks. Gene expression was examined by total RNA-sequencing and the functional significance was assessed by gene set enrichment analysis using the Gene Ontology database. Under short-term acidosis (24h), AT1 and MCF-7 cells showed comparable changes. 714 genes were acidosis-dependently regulated in AT1 cells (275 up, 439 down), and 221 genes in MCF-7 cells (95 up, 126 down). MDA-MB-231 cells almost did not respond to low pH (13 regulated genes). Hypoxia affected MCF-7 cells the most (1498 regulated genes), whereas fewer genes were regulated in AT1 and MDA-MB-231 cells. Concerning the function of the regulated genes by short-term acidosis, RNA processing, cell cycle regulation, DNA synthesis, and mitochondrial function were negatively affected. Chronic acidosis showed a different picture. In AT1 cells, 1160 genes were differentially expressed (638 up, 522 down) when cells exposed to low pH for 5weeks. The putatively acidosis-induced changes in functions included tissue structural development, RNA processing, and mitochondrial activity. This study shows that both acute and chronic acidosis of tumour cells lead to altered gene expression and thus affect cell function. Long-term acidosis leads to fundamentally different changes, indicating an adaptation process of the tumour cells.

Estrogen regulated transcription of the non-estrogen-regulated hamster uteroglobin gene in MCF-7 cells

To study the estrogen regulated transcription of the uteroglobin (UG) gene, the founding member of the secretoglobin family widely expressed in many different mammalian species, we re-created functional estrogen response elements (EREs) in the UG gene promoter from a species where UG expression is not regulated by estrogens: the hamster Mesocricetus auratus (Ma), to ascertain if the lack of functional EREs is the real cause of its estrogen insensitivity. Functional EREs in the hamster promoter, including the consensus ERE (cERE), failed to respond to an appropriate estrogen stimulus compared with its estrogen regulated ortholog from the brown hare Lepus capensis (Lc). As the nucleotide sequence is the only difference between genetic constructs from these two species, we suspected that the UG promoter from the hamster probably contains cis-acting genetic elements that negatively impairs the estrogen-regulated transcription mediated by the functional ERE. Accordingly, we prepared chimeric DNA constructs which eventually allowed to identify a region located 29 base pairs (bp) downstream of the ERE as responsible for the lack of estrogen-responsiveness of the Ma-UG gene in the breast cancer cell line MCF-7. This region contains the sequence ACACCCC which has been identified as the core sequence of the Sp/ Krüppel-like factor (KLF) family of transcription factors. This finding is relevant, not only due to the observation on a novel mechanism that control estrogen-induced transcription, but also because it may encourage further investigation for better defining specific genes with an ERE that do not respond to estrogen signaling in MCF-7 cells, a cell line widely employed as an in vitro model in breast cancer research.

THU557 Effects Of Two Experimental Monocarbonyl Analogs Of Curcumin (MACs) On Breast Cancer Growth, Migration, And Epithelial-To-Mesenchymal Transition (EMT)

Abstract Disclosure: R. Stojchevski: None. N. Hadzi-Petrushev: None. M. Mladenov: None. J. Bogdanov: None. S. Velichkovikj: None. L. Poretsky: None. D.B. Avtanski: None. Curcumin, a polyphenol found in turmeric (Curcuma longa), has been reported to have a range of potential therapeutic effects, including anti-inflammatory and antioxidant properties. However, curcumin’s poor bioavailability has limited its use in medicine. Monocarbonyl analogs of curcumin (MACs) have been developed to overcome this challenge and have shown promise in various medical applications, including cancer. Our previous experiments in animals showed that these two analogs possess potent antioxidant properties. The aim of this study was to investigate the in vitro effects of two experimental MACs (C66 (C22H16F6O) and B2BrBC (C20H16Br2O)) on breast cancer cell growth and epithelial-to-mesenchymal transition (EMT). Both C66 and B2BrBC significantly suppressed MCF-7 and BT-474 viability shown by MTT assay. These two analogs also suppressed breast cancer cell migration, demonstrated by scratch migration wound-healing assay using MCF-7 and MDA-MB-231 cells. Further, using qRT-PCR, we investigated the effects of C66 and B2BrBC on the expression of various epithelial (E-cadherin, cytokeratin-18) and mesenchymal (snail, slug, fibronectin, and vimentin) genes in MCF-7 cells subjected to EMT induction (a media supplement containing Wnt-5a, TGFβ1, anti-E-cadherin, anti-sFRP1, and anti-Dkk-1 antibodies). Since the EMT-induction media significantly downregulated the epithelial markers’ and upregulated the mesenchymal markers’ mRNA expression, these effects were significantly abolished when C66 or B2BrBC were introduced. Taken together, these data demonstrate that C66 and B2BrBC are potent agents for suppressing breast cancer cell growth and EMT in vitro. Further investigation is needed to determine the potential clinical use of these compounds in breast cancer treatment. Presentation: Thursday, June 15, 2023

Selective Concurrence of the Long Non-Coding RNA MALAT1 and the Polycomb Repressive Complex 2 to Promoter Regions of Active Genes in MCF7 Breast Cancer Cells.

In cancer cells, the long non-coding RNA (lncRNA) MALAT1 has arisen as a key partner for the Polycomb Repressive Complex 2 (PRC2), an epigenetic modifier. However, it is unknown whether this partnership occurs genome-wide at the chromatin level, as most of the studies focus on single genes that are usually repressed. Due to the genomic binding properties of both macromolecules, we wondered whether there are binding sites shared by PRC2 and MALAT1. Using public genome-binding datasets for PRC2 and MALAT1 derived from independent ChIP- and CHART-seq experiments performed with the breast cancer cell line MCF7, we searched for regions containing PRC2 and MALAT1 overlapping peaks. Peak calls for each molecule were performed using MACS2 and then overlapping peaks were identified by bedtools intersect. Using this approach, we identified 1293 genomic sites where PRC2 and MALAT1 concur. Interestingly, 54.75% of those sites are within gene promoter regions (<3000 bases from the TSS). These analyses were also linked with the transcription profiles of MCF7 cells, obtained from public RNA-seq data. Hence, it is suggested that MALAT1 and PRC2 can concomitantly bind to promoters of actively-transcribed genes in MCF7 cells. Gene ontology analyses revealed an enrichment of genes related to categories including cancer malignancy and epigenetic regulation. Thus, by re-visiting occupancy and transcriptomic data, we identified a key gene subset controlled by the collaboration of MALAT1 and PRC2.

In Vitro Analysis of Cytotoxic Activities of Monotheca buxifolia Targeting WNT/β-Catenin Genes in Breast Cancer Cells.

Breast cancer (BC) is known to be the most common malignancy among women throughout the world. Plant-derived natural products have been recognized as a great source of anticancer drugs. In this study, the efficacy and anticancer potential of the methanolic extract of Monotheca buxifolia leaves using human breast cancer cells targeting WNT/β-catenin signaling was evaluated. We used methanolic and other (chloroform, ethyl acetate, butanol, and aqueous) extracts to discover their potential cytotoxicity on breast cancer cells (MCF-7). Among these, the methanol showed significant activity in the inhibition of the proliferation of cancer cells because of the presence of bioactive compounds, including phenols and flavonoids, detected by a Fourier transform infrared spectrophotometer and by gas chromatography mass spectrometry. The cytotoxic effect of the plant extract on the MCF-7 cells was examined by MTT and acid phosphatase assays. Real-time PCR analysis was performed to measure the mRNA expression of WNT-3a and β-catenin, along with Caspase-1,-3,-7, and -9 in MCF-7 cells. The IC50 value of the extract was found to be 232 μg/mL and 173 μg/mL in the MTT and acid phosphatase assays, respectively. Dose selection (100 and 300 μg/mL) was performed for real-time PCR, Annexin V/PI analysis, and Western blotting using Doxorubicin as a positive control. The extract at 100 μg/mL significantly upregulated caspases and downregulated the WNT-3a and β-catenin gene in MCF-7 cells. Western blot analysis further confirmed the dysregulations of the WNT signaling component (*** p< 0.0001). The results showed an increase in the number of dead cells in methanolic extract-treated cells in the Annexin V/PI analysis. Our study concludes that M. buxifolia may serve as an effective anticancer mediator through gene modulation that targets WNT/β-catenin signaling, and it can be further characterized using more powerful experimental and computational tools.

Preclinical-to-clinical Anti-cancer Drug Response Prediction and Biomarker Identification Using TINDL

Prediction of the response of cancer patients to different treatments and identification of biomarkers of drug response are two major goals of individualized medicine. Here, we developed a deep learning framework called TINDL, completely trained on preclinical cancer cell lines (CCLs), to predict the response of cancer patients to different treatments. TINDL utilizes a tissue-informed normalization to account for the tissue type and cancer type of the tumors and to reduce the statistical discrepancies between CCLs and patient tumors. Moreover, by making the deep learning black box interpretable, this model identifies a small set of genes whose expression levels are predictive of drug response in the trained model, enabling identification of biomarkers of drug response. Using data from two large databases of CCLs and cancer tumors, we showed that this model can distinguish between sensitive and resistant tumors for 10 (out of 14) drugs, outperforming various other machine learning models. In addition, our small interfering RNA (siRNA) knockdown experiments on 10 genes identified by this model for one of the drugs (tamoxifen) confirmed that tamoxifen sensitivity is substantially influenced by all of these genes in MCF7 cells, and seven of these genes in T47D cells. Furthermore, genes implicated for multiple drugs pointed to shared mechanism of action among drugs and suggested several important signaling pathways. In summary, this study provides a powerful deep learning framework for prediction of drug response and identification of biomarkers of drug response in cancer. The code can be accessed at https://github.com/ddhostallero/tindl.

Cluster of resistance-inducing genes in MCF-7 cells by estrogen, insulin, methotrexate and tamoxifen extracted via NMF

Breast cancer has become a leading cause of death for women as the economy has grown and the number of women in the labor force has increased. Several biomarkers with diagnostic, prognostic, and therapeutic implications for breast cancer have been identified in studies, leading to therapeutic advances. Resistance, on the other hand, is one of clinical practice's limitations. In this paper, we use Nonnegative Matrix Factorization to automatically extract two gene signatures from gene expression profiles of wild-type and resistance MCF-7 cells, which were then investigated further using pathways analysis and proved useful in relating resistance pathways to breast cancer regardless of the stimulus that caused it. A few extracted genes (including MAOA, IL4I1, RRM2, DUT, NME4, and SUMO3) represent new elements in the functional network for resistance in MCF-7 ER+breast cancer. As a result of this research, a better understanding of how resistance occurs or the pathways that contribute to it may allow more effective therapies to be developed.

Dietary flavonoid carvacrol triggers the apoptosis of human breast cancer MCF-7 cells via the p53/Bax/Bcl-2 axis

Recently, numerous studies have shown that natural compounds such as carvacrol can be used as therapeutic agents for medicinal purposes. Although carvacrol was found to suppress breast cancer cell lines, however, whether the therapeutic effect of carvacrol on breast cancer is caused by increasing the expression of pro-apoptotic genes in the MCF-7 cell line has not been studied. In this research, we investigate the effect of carvacrol on the expression of P53, pro-apoptotic Bax and anti-apoptotic BCL-2 genes in MCF-7 cells. After preparation and cultivation of MCF-7 cells, the IC50 value of carvacrol on the cells was evaluated by MTT assay, and then apoptosis induction was observed in the cell line treated with different concentrations of carvacrol by DAPI staining. To assess the expression level of Bax, P53 and Bcl-2 both in genes and protein levels QPCR method and western blot analysis were used. According to the results of the research, it was determined that the IC50 of carvacrol compound in MCF-7 cells is 305µM. DAPI staining exhibit apoptosis and morphological changes in treated MCF-7 cells. Real-time PCR assay and western blot showed increasing the Bax and P53 expression and decreasing the expression of the Bcl-2 anti-apoptotic gene and protein. These results showed that carvacrol has cell growth inhibition effects on the MCF-7 cancer cell line. Carvacrol-induced p53-dependent apoptosis, which might be related to the Bax/Bcl-2 associated pathway. These results indicated that carvacrol could be considered suitable for the prevention and treatment of breast cancer.

Chitinase inhibits growth of human breast and colorectal cancer cells in vitro and Ehrlich ascites carcinoma cells in vivo

Trichosanthes dioica seed extract was loaded on a QA-cellulose column and the unbound fraction with the chitinase activity was run on SDS-PAGE. Multiple bands were observed and were separated by a Sephadex G-50 column. The combination of the 6 and 33 kDa masses supported the degradation of chitinase as purified earlier. Only the 33 kDa fraction contained sugar and showed chitinase activity. The chitinase was also isolated by using a chitin column. At 200 µg/ml protein concentration, the chitinase inhibited 49.1 %, 48.8 % and 38.12 % of Ehrlich ascites carcinoma, HCT-116 and MCF-7 cells growth, respectively, in a dose-dependent manner. Exactly, 46 % and 82 % EAC cell growth inhibition were observed after treating the EAC cells bearing Swiss albino mice with the chitinase at the doses of 1.0 and 2.0 mg/Kg/day respectively. EAC, HCT-116 and MCF-7 cells growth inhibitions were due to the induction of apoptosis. ROS was accumulated in HCT-116 and MCF-7 cells. After treatment of HCT-116 cells, the expression level of p53 and TNFα genes increased and PARP gene decreased. On the other hand, elevated expression was observed for PARP, MAPK, NFκB, FAS, FADD, and Caspase-8 genes in MCF-7 cells. The induction of apoptosis in HCT-116 was further confirmed by caspase protein expression. The chitinase causes ‘S’ cell cycle arrest in MCF-7 and HCT-116 cells. T. dioica seed chitinase inhibited EAC, HCT-116 and MCF-7 cells by inducing apoptosis in vitro and EAC in vivo in mice. These promising results indicated that T. dioica seed chitinase can be an anticancer agent.

Anti-proliferative effects of Rhamnus frangula miller leaf and bark extracts on HEK293 and MCF-7 cell lines and evaluation of Bax and Bcl-2 genes expression level in MCF-7 cells

Rhamnus frangula extracts may have anti-proliferative effects on cancer development. This study was exerted to determine the effects of Rhamnus frangula Miller leaf and bark extracts on the viability of MCF-7 cells compared with HEK293 cells and the evaluation of Bax and Bcl-2 gene expression levels in MCF-7 cells. The cell lines were randomly divided into the control group (not exposed to extract) and groups exposed to 0.001, 0.01, 0.1, and 1 mg/ml of Rhamnus frangula leaf and bark extracts. Cell viability was quantified by MTT assay. The expression of Bax and Bcl-2 genes was evaluated by quantitative Real-time PCR analysis. Statistical analysis was performed using ANOVA. HEK293 cells viability significantly increased in groups exposed to 0.001 and 0.01 mg/ml and decreased in group exposed to 1 mg/ml of Rhamnus frangula leaf extract compared to control group (P &lt; 0.01, P &lt; 0.05 and P &lt; 0.01, respectively). Exposure of MCF-7 cells to 0.01, 0.1 and 1 mg/ml of leaf extract led to significant decrease in cell viability (P &lt; 0.05). Exposure of HEK293 cells to 1 mg/ml and of MCF-7 cells to 0.1 and 1 mg/ml of Rhamnus frangula bark extract resulted in significant decrease in cell viability compared to control group (P &lt; 0.01, P &lt; 0.05 and P &lt; 0.01, respectively). The cytotoxic effect of leaf was higher than bark extract on MCF7 cells. 1 mg/ml of leaf and bark extracts significantly increased the expression level of pro-apoptotic Bax and decreased anti-apoptotic Bcl-2 genes in MCF-7 cells (P &lt; 0.01). However, the expression level of the Bax gene was significantly higher in a group exposed to 1 mg/ ml of leaf compared to bark extract (P &lt; 0.01). Despite bark, lower leaf extract concentrations inhibit breast cancer (MCF-7) cells proliferation while having no cytotoxic effects on non-cancerous HEK293 cells. Therefore, using leaf is more appropriate than bark extract to fight against breast cancer cells. We have also shown that Rhamnus frangula leaf and bark extracts mechanism of action has occurred through Bax-dependent apoptotic pathway on breast cancer cell line.

Effects of β-glucan from S. cerevisiae on the expression of Casp9, Ccna2 and Sod1 genes in MCF-7 cells

β-Glucans (βG) are polysaccharides widely distributed in nature with chemopreventive properties. The aim of this study was to investigate the effects of βG and the combined treatment with doxorubicin (Dox) on cell viability and mRNA levels of genes involved in cell cycle, apoptosis and antioxidant response. βG was not cytotoxic. The mRNA levels of CCNA2 of cells exposed to β-glucan was upregulated and the exposure to Dox decreased the expression, while the combination led to an upregulation. Modulation of mRNA levels of CASP9 suggest that βG could inhibit promotion and progression steps of carcinogenesis, eliminating neoplastic cells. The upregulation of CCNA2 gene in combined treatment could be occurred due to ability of βG in restoring the cell cycle distribution pattern after treatment with Dox. The upregulation of SOD1 suggests that βG can enhance the intracellular antioxidant defense, reducing the levels of superoxide dismutase induced by Dox. This response could reduce oxidative damage and attenuate tissue damage during chemotherapeutic treatment. Our data suggest that the drug combination may be less effective in killing tumor cells than the treatment with Dox alone. Thus, future studies should carefully consider this effect on indication of βG during chemotherapy

Diffractaic acid, a novel TrxR1 inhibitor, induces cytotoxicity, apoptosis, and antimigration in human breast cancer cells

Breast cancer represents one of the most frequently encountered cancer types among women worldwide. Thioredoxin reductase 1 (TrxR1) is a therapeutic target for breast cancer therapy due to its overexpression in tumor cells. The current research aims to determine the anticancer effect of diffractaic acid, a lichen acid, in breast cancer, and research whether the anticancer effect of diffractaic acid occurs through TrxR1 targeting. According to the XTT assay results, diffractaic acid induced cytotoxicity in both MCF-7 and MDA-MB-453 cells with IC50 values of 51.32 μg/ml and 87.03 μg/ml, respectively. Flow cytometry and cell migration analyses revealed the apoptotic, necrotic, and antimigratory effects of diffractaic acid. qPCR analysis indicated the upregulation of the BAX/BCL2 ratio and the P53 gene in MCF-7 cells with only the P53 gene in MDA-MB-453 cells. The gene, protein, and enzyme activity of TrxR1 were suppressed in MCF-7 cells, whereas only enzyme activity was suppressed in MDA-MB-453 cells. These findings illustrate the anticancer effect of diffractaic acid on breast cancer targeting TrxR1. In conclusion, these data reveal that diffractaic acid may be considered an effective therapeutic agent for breast cancer treatment.

The Influence of Faradarmani Consciousness Field on the Survival and Death of MCF-7 Breast Cancer Cells: An Optimization Perspective

Mohammad Ali Taheri, the founder of the Faradarmani Consciousness Field (CF) maintains that beyond matter and energy, there are Taheri Consciousness Fields (TCFs) that can be accessed through the connection of the consciousness of the subject of study to the Cosmic Consciousness Network (CCN). This causes an optimization in a subject’s structure and function. Apart from the fact that most studies in the field of cancer are aimed at the death of cancer cells, the purpose of this study was to investigate the MCF-7 cancer cell survival and death according to their optimal nature under the influence of the Faradarmani CF. For the purpose of this investigation, cellular viability was evaluated by MTT assay for 6-24 h under the influence of the Faradarmani CF. Consequently, cell cycle analysis was performed for the evaluation of apoptotic cell death. Finally, the expression of Bcl2 and Bax genes in MCF-7 cells was evaluated as a validation of the obtained cellular scale results. Our findings demonstrate that the Faradarmani CF treatment significantly induced cell proliferation in the MCF-7 cell culture in specific test conditions mediated by increasing cellular viability by about 18% in comparison to the control in a time-dependent manner. Moreover, the S-phase in the cell cycle, as a sign of cell population in the sample treated with the Faradarmani CF, was increased by 56% (up to 24h) in comparison to the control. The real-time RT-PCR reaction results show that in cells treated with the Faradarmani CF, the Bax/Bcl2 ratio decreased after 24 h (more than the 1-fold negative control (untreated)), suggesting a higher cell survival and resistance to death. Considering the concepts of the TCFs, according to Taheri, proliferation and survival appear to become optimized in MCF-7 cancer cell lines under the influence of the Faradarmani CF.

Molecular Mechanisms Underlying the Anti-Breast Cancer Stem Cell Activity of Pterocladia capillacea and Corallina officinalis Polysaccharides.

This study aimed to appraise the activity of Pterocladia capillacea and Corallina officinalis polysaccharides against Breast Cancer Stem Cells (BCSCs). P. capillacea and C. officinalis polysaccharides were characterized to be sulfated polysaccharide-protein complexes. Cytotoxicity of the polysaccharides against MDA-MB-231 and MCF-7 cell lines along with their impact on CD44+/CD24- and aldehyde dehydrogenase 1(ALDH1) positive BCSC population were determined. Their effect on gene expression of CSC markers, Wnt/β-catenin and Notch signaling pathways was evaluated. P. capillacea and C. officinalis polysaccharides inhibited the growth of breast cancer cells and reduced BCSC subpopulation. P. capillacea polysaccharides significantly down-regulated OCT4, SOX2, ALDH1A3 and vimentin in MDA-MB-231 as well as in MCF-7 cells except for vimentin that was up-regulated in MCF-7 cells. C. officinalis polysaccharides exhibited similar effects except for OCT4 that was up-regulated in MDA-MB-231 cells. Significant suppression of Cyclin D1 gene expression was noted in MDA-MB-231 and MCF-7 cells treated with P. capillacea or C. officinalis polysaccharides. β-catenin and c-Myc genes were significantly down-regulated in MDA-MB-231 cells treated with C. officinalis and P. capillacea polysaccharides, respectively, while being up-regulated in MCF-7 cells treated with either of them. Additionally, P. capillacea and C. officinalis polysaccharides significantly down-regulated Hes1 gene in MCF-7 cells despite increasing Notch1 gene expression level. However, significant down-regulation of Notch1 gene was observed in MDA-MB-231 cells treated with P. capillacea polysaccharides. Collectively, this study provides evidence for the effectiveness of P. capillacea and C. officinalis polysaccharides in targeting BCSCs through interfering with substantial signaling pathways contributing to their functionality.

Widespread association of ERα with RMRP and tRNA genes in MCF-7 cells and breast cancers

tRNA gene transcription by RNA polymerase III (Pol III) is a tightly regulated process, but dysregulated Pol III transcription is widely observed in cancers. Approximately 75% of all breast cancers are positive for expression of Estrogen Receptor alpha (ERα), which acts as a key driver of disease. MCF-7 cells rapidly upregulate tRNA gene transcription in response to estrogen and ChIP-PCR demonstrated ERα enrichment at tRNALeu and 5S rRNA genes in this breast cancer cell line. While these data implicate the ERα as a Pol III transcriptional regulator, how widespread this regulation is across the 631 tRNA genes has yet to be revealed. Through analyses of ERα ChIP-seq datasets, we show that ERα interacts with hundreds of tRNA genes, not only in MCF-7 cells, but also in primary human breast tumours and distant metastases. The extent of ERα association with tRNA genes varies between breast cancer cell lines and does not correlate with levels of ERα binding to its canonical target gene GREB1. Amongst other Pol III-transcribed genes, ERα is consistently enriched at the long non-coding RNA gene RMRP, a positive regulator of cell cycle progression that is subject to focal amplification in tumours. Another Pol III template targeted by ERα is the RN7SL1 gene, which is strongly implicated in breast cancer pathology by inducing inflammatory responses in tumours. Our data indicate that Pol III-transcribed non-coding genes should be added to the list of ERα targets in breast cancer.

Non-classical Notch signaling by MDA-MB-231 breast cancer cell-derived small extracellular vesicles promotes malignancy in poorly invasive MCF-7 cells.

Aberrant Notch signaling is implicated in breast cancer progression, and recent studies have demonstrated links between the Notch pathway components Notch1 and Notch1 intracellular domain (N1ICD) with poor clinical outcomes. Growing evidence suggests that Notch signaling can be regulated by small extracellular vesicles (SEVs). Here, we used breast cancer cell models to examine whether SEVs are involved in functional Notch signaling. We found that Notch components are packaged into MDA-MB-231- and MCF-7-derived SEVs, although higher levels of N1ICD were detected in SEVs from the more aggressive MDA-MB-231 cell line than from poorly invasive MCF-7 cells. SEV-Notch components were functional, as SEVs cargo from MDA-MB-231 cells induced the expression of Notch target genes in MCF-7 cells and triggered a more invasive and proliferative phenotype concomitant with the acquisition of mesenchymal features. Neutralization of the N1ICD cargo in MDA-MB-231-derived SEVs significantly reduced their potential to enhance the aggressiveness of MCF-7 cells in vitro and in a xenograft model. Overall, our results indicate that a SEV-mediated non-classical pathway of Notch signal transduction in breast cancer models bypasses the need for classical ligand-receptor interactions, which may have important implications in cancer.

Effects of Endocrine Disruptors o,p'-Dichlorodiphenyltrichloroethane, p,p'-Dichlorodiphenyltrichloroethane, and Endosulfan on the Expression of Estradiol-, Progesterone-, and Testosterone-Responsive MicroRNAs and Their Target Genes in MCF-7 Cells.

Many studies have shown that dichlorodiphenyltrichloroethane (DDT) exposure raises breast cancer risk. Another insecticide with similar properties is endosulfan, which has been actively used in agriculture after DDT prohibition. Previously, we have identified some estradiol-, progesterone-, and testosterone-sensitive microRNAs (miRNAs, miRs). Because DDT and endosulfan have estrogenic, antiandrogenic, and antiprogesterone properties, we hypothesized that these miRNAs are affected by the insecticides. We quantified relative levels of miRNAs and expression levels of their target genes in breast cancer MCF-7 cells treated with p,p′-DDT, o,p′-DDT, or endosulfan. We also quantified miR-19b expression, which, as previously shown, is regulated by estrogen. Here, we observed that miR-19b expression increased in response not only to estradiol but also to testosterone and progesterone. Treatment of MCF-7 cells with p,p′-DDT or endosulfan decreased the protein levels of apoptosis regulators TP53INP1 and APAF1. In cells treated with o,p′-DDT, the TP53INP1 amount decreased after 24 h of incubation, but increased after 48 h of incubation with insecticide. OXTR expression, which is known to be associated with breast carcinogenesis, significantly diminished under the exposure of all insecticides. In cells treated with p,p′-DDT or o,p′-DDT, the observed changes were accompanied by alterations of the levels of hormone-responsive miRNAs: miR-324, miR-190a, miR-190b, miR-27a, miR-193b, and miR-19b.

OCT1 Is a Poor Prognostic Factor for Breast Cancer Patients and Promotes Cell Proliferation via Inducing NCAPH.

Octamer transcription factor 1 (OCT1) is a transcriptional factor reported to be a poor prognostic factor in various cancers. However, the clinical value of OCT1 in breast cancer is not fully understood. In the present study, an immunohistochemical study of OCT1 protein was performed using estrogen receptor (ER)-positive breast cancer tissues from 108 patients. Positive OCT1 immunoreactivity (IR) was associated with the shorter disease-free survival (DFS) of patients (p = 0.019). Knockdown of OCT1 inhibited cell proliferation in MCF-7 breast cancer cells as well as its derivative long-term estrogen-deprived (LTED) cells. On the other hand, the overexpression of OCT1 promoted cell proliferation in MCF-7 cells. Using microarray analysis, we identified the non-structural maintenance of chromosomes condensin I complex subunit H (NCAPH) as a novel OCT1-taget gene in MCF-7 cells. Immunohistochemical analysis showed that NCAPH IR was significantly positively associated with OCT1 IR (p < 0.001) and that positive NCAPH IR was significantly related to the poor DFS rate of patients (p = 0.041). The knockdown of NCAPH inhibited cell proliferation in MCF-7 and LTED cells. These results demonstrate that OCT1 and its target gene NCAPH are poor prognostic factors and potential therapeutic targets for patients with ER-positive breast cancer.

Copper nanoparticles green synthesis and characterization as anticancer potential in breast cancer cells (MCF7) derived from Prunus nepalensis phytochemicals

The green synthesis of nanoparticles from bioactive compounds have attracted a wide range of application, due to increased drug efficacy and less toxicity in the nanosized mediated drug delivery model. In this study, we have fabricated copper nanoparticles (CuNPs) from the fruits of Prunus nepalensis (P. nepalensis) extract. Therefore, the aim of present study was to investigate the anticancer ability of P. nepalensis fruit phytochemical copper nanoaprticles (PNFPCuNP) on cancerous human breast cell line (MCF-7) and healthy (MCFA10) cell lines. Crystalline CuNPs of P. nepalensis synthesis was confirmed by different physicochemical analytical techniques such as UV–Visible Spectroscopy (UV–Vis); Fourier-Transform Infrared Spectroscopy (FT-IR); Scanning Electron Microscopy (SEM); and Transmission Electron Microscopy (TEM). Nanoparticle Size was found to be ranging from 35 to 50 nm with the average size of 42.5 nm. Further, after synthesized compounds were tested anticancer activity on human breast cancer cell lines. Following 72 h treatment to PNFPCuNP, the expression of apoptotic marker genes (P21, p53, P14/P19, Caspase-3) were studied in MCF-7 cells treated at 100 to 200 μg of PNFP-CuNP. Our results showed that PNFP-CuNP increased the gene expression of apoptotic genes in a dose-dependent manner. The real-time PCR data showed a significant upregulation in p53, Bax, caspase-3, and caspase-9 and down regulation in the mRNA expression of Ras and Myc genes in MCF-7 cells exposed to PNFP-CuNP. Collectively, the data from this study stated that P. nepalennsis fruit extract phytochemical derived nanoparticles induced apoptosis via the up regulation of tumour suppressor genes and down regulation of oncogenes in MCF-7 cells. Finally, our study confirmed the CuNPs synthesis from P. nepalensis fruit phytochemical, which showed environmental friendly anticancer activity.