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Breast cancer comprises about 30% of all new female cancers each year and is the most common malignant cancer in women in the United States. Breast cancer cell lines have been harnessed for many years as a foundation for in vitro analytic studies to understand the use of cancer prevention and therapy. There has yet to be a compilation of works to analyze the pitfalls, novel discoveries, and essential techniques for breast cancer cell line studies in a scientific context. In this article, we review the history of breast cancer cell lines and their origins, as well as analyze the molecular pathways that pharmaceutical drugs apply to breast cancer cell lines in vitro and in vivo. Controversies regarding the origins of certain breast cancer cell lines, the benefits of utilizing Patient-Derived Xenograft (PDX) versus Cell-Derived Xenograft (CDX), and 2D versus 3D cell culturing techniques will be analyzed. Novel outcomes from epigenetic discovery with dietary compound usage are also discussed. This review is intended to create a foundational tool that will aid investigators when choosing a breast cancer cell line to use in multiple expanding areas such as epigenetic discovery, xenograft experimentation, and cancer prevention, among other areas. [ABSTRACT FROM AUTHOR]

Breast cancer (BC) is among the most frequently diagnosed malignant cancers in women in the United States. Diet and nutrition supplementation are closely related to BC onset and progression, and inulin is commercially available as a health supplement to improve gut health. However, little is known with respect to inulin intake for BC prevention. We investigated the effect of an inulin-supplemented diet on the prevention of estrogen receptor-negative mammary carcinoma in a transgenic mouse model. Plasma short-chain fatty acids were measured, the gut microbial composition was analyzed, and the expression of proteins related to cell cycle and epigenetics-related genes was measured. Inulin supplementation greatly inhibited tumor growth and significantly delayed tumor latency. The mice that consumed inulin had a distinct microbiome and higher diversity of gut microbial composition compared to the control. The concentration of propionic acid in plasma was significantly higher in the inulin-supplemented group. The protein expression of epigenetic-modulating histone deacetylase 2 (Hdac2), Hdac8, and DNA methyltransferase 3b decreased. The protein expression of factors related to tumor cell proliferation and survival, such as Akt, phospho-PI3K, and NF-kB, also decreased with inulin administration. Furthermore, sodium propionate showed BC prevention effect in vivo through epigenetic regulations. These studies suggest that modulating microbial composition through inulin consumption may be a promising strategy for BC prevention. [ABSTRACT FROM AUTHOR]

Simple Summary: Certain life stages, known as critical periods, during growth and development are thought to be important for later-life breast cancer initiation and progression. Nutritional factors, especially those found in plant-based diets, are believed to be key to the impact of these critical periods on cancer. However, there is currently little known with respect to how nutrition during critical periods can affect breast cancer. In this study we evaluated nutritional intervention during the critical period of puberty and whether it could have a significant effect on tumor phenotype, as well as underlying gene expression, protein expression and DNA methylation patterns. We found that sulforaphane-containing broccoli sprout extracts administered during the peripubertal period in mice were able to reduce tumor size and incidence while delaying latency. We also found gross changes to gene expression patterns, including many cancer-associated genes, as well as potentially important changes to methylation profiles in our treatment groups. Breast cancer (BC) is a nearly ubiquitous malignancy that effects the lives of millions worldwide. Recently, nutritional prevention of BC has received increased attention due to its efficacy and ease of application. Chief among chemopreventive compounds are plant-based substances known as dietary phytochemicals. Sulforaphane (SFN), an epigenetically active phytochemical found in cruciferous vegetables, has shown promise in BC prevention. In addition, observational studies suggest that the life stage of phytochemical consumption may influence its anticancer properties. These life stages, called critical periods (CPs), are associated with rapid development and increased susceptibility to cellular damage. Puberty, a CP in which female breast tissue undergoes proliferation and differentiation, is of particular interest for later-life BC development. However, little is known about the importance of nutritional chemoprevention to CPs. We sought to address this by utilizing two estrogen receptor-negative [ER(-)] transgenic mouse models fed SFN-containing broccoli sprout extract during the critical period of puberty. We found that this treatment resulted in a significant decrease in tumor incidence and weight, as well as an increase in tumor latency. Further, we found significant alterations in the long-term expression of cancer-associated genes, including p21, p53, and BRCA2. Additionally, our transcriptomic analyses identified expressional changes in many cancer-associated genes, and bisulfite sequencing revealed that the antiproliferation-associated gene Erich4 was both hypomethylated and overexpressed in our experimental group. Our study indicates that dietary interventions during the CP of puberty may be important for later-life ER(-) BC prevention and highlights potential important genetic and epigenetic targets for treatment and study of the more deadly variants of BC. [ABSTRACT FROM AUTHOR]

Breast cancer (BC) is the most common malignancy and the second leading cause of cancer death among women in the United States. The consumption of natural dietary components such as broccoli sprouts (BSp) and green tea polyphenols (GTPs) has demonstrated exciting potential in reducing the risk of BC through the regulation of epigenetic mechanisms. However, little is known about their impacts on reversing epigenomic aberrations that are centrally involved in the initiation and progression of BC. Previously, we have determined the efficacy of combined BSp and GTPs treatment on the inhibition of the growth of a mammary tumor in a transgenic Her2/neu mouse model. We sought to extend our previous study to identify universal biomarkers that represent common mechanistic changes among different mouse models in response to this dietary regime by including a new transgenic mouse model, C3(1)-SV40 TAg (SV40). As a result, we identified novel target genes that were differentially expressed and methylated in response to dietary botanicals when administered singly (BSp and GTPs) and in combination (BSp + GTPs) in both mouse models. We discovered more differentially expressed and methylated genes in the combination treatment group compared to the singly administered groups. Subsequently, several biological pathways related to epigenetic regulations were identified in response to the combination treatment. Furthermore, when compared to the BSp and GTPs treatment alone, the combinatorial treatment showed a more significant impact on the regulation of the epigenetic modifier activities involved in DNA methylation and histone modifications. Our study provides key insights about the impact of the combined administration of BSp and GTPs on BC using a multi-omics analysis, suggesting a combinatorial approach is more efficacious in preventing and inhibiting BC by impacting key tumor-related genes at transcriptomic and methylomic levels. Our findings could be further extrapolated as a comprehensive source for understanding the epigenetic modifications that are associated with the effects of these dietary botanicals on BC prevention. [ABSTRACT FROM AUTHOR]

Breast cancer (BC) is the most common cancer in women in the United States. There has been an increasing incidence and decreasing mortality rate of BC cases over the past several decades. Many risk factors are associated with BC, such as diet, aging, personal and family history, obesity, and some environmental factors. Recent studies have shown that healthy individuals and BC patients have different microbiota composition, indicating that microbiome is a new risk factor for BC. Gut and breast microbiota alterations are associated with BC prognosis. This review will evaluate altered microbiota populations in gut, breast tissue, and milk of BC patients, as well as mechanisms of interactions between microbiota modulation and BC. Probiotics and prebiotics are commercially available dietary supplements to alleviate side-effects of cancer therapies. They also shape the population of human gut microbiome. This review evaluates novel means of modulating microbiota by nutritional treatment with probiotics and prebiotics as emerging and promising strategies for prevention and treatment of BC. The mechanistic role of probiotic and prebiotics partially depend on alterations in estrogen metabolism, systematic immune regulation, and epigenetics regulation. [ABSTRACT FROM AUTHOR]

There is conflicting evidence on the role of lipid biomarkers in breast cancer (BC), and no study to our knowledge has examined this association among African women. We estimated odds ratios (ORs) and 95% confidence intervals (95% CI) for the association of lipid biomarkers—total cholesterol, high-density lipoprotein (HDL), low-density lipoprotein (LDL), and triglycerides—with odds of BC overall and by subtype (Luminal A, Luminal B, HER2-enriched and triple-negative or TNBC) for 296 newly diagnosed BC cases and 116 healthy controls in Nigeria. Each unit standard deviation (SD) increase in triglycerides was associated with 39% increased odds of BC in fully adjusted models (aOR: 1.39; 95% CI: 1.03, 1.86). Among post-menopausal women, higher total cholesterol (aOR: 1.65; 95% CI: 1.06, 2.57), LDL cholesterol (aOR: 1.59; 95% CI: 1.04, 2.41), and triglycerides (aOR: 1.91; 95% CI: 1.21, 3.01) were associated with increased odds of BC. Additionally, each unit SD increase in LDL was associated with 64% increased odds of Luminal B BC (aOR 1.64; 95% CI: 1.06, 2.55). Clinically low HDL was associated with 2.7 times increased odds of TNBC (aOR 2.67; 95% CI: 1.10, 6.49). Among post-menopausal women, higher LDL cholesterol and triglycerides were significantly associated with increased odds of Luminal B BC and HER2 BC, respectively. In conclusion, low HDL and high LDL are associated with increased odds of TN and Luminal B BC, respectively, among African women. Future prospective studies can definitively characterize this association and inform clinical approaches targeting HDL as a BC prevention strategy. [ABSTRACT FROM AUTHOR]

Breast cancer has strong developmental origins and maternal nutrition composition may influence later-life breast cancer risk in the offspring. Our study focused on a bioactive dietary component, genistein (GE) enriched in soybean products, to investigate specific timing of maternal GE exposure that may influence preventive efficacy of GE on offspring breast cancer later in life, and to explore the potential epigenetic mechanisms. Our results indicate a time-dependent effect of maternal GE exposure on early-life breast cancer development in offspring mice. Through integrated transcriptome and methylome analyses, we identified several candidate genes showing significantly differential gene expression and DNA methylation changes. We further found maternal long-term GE treatment can induce inherited epigenetic landmark changes in a candidate tumor suppressor gene, Trp63 , resulting in transcriptional activation of Trp63 and induction of the downstream target genes. Our results suggest that maternal long-term exposure to soybean GE may influence early-life epigenetic reprogramming processes, which may contribute to its temporal preventive effects on breast cancer in the offspring. This study provides important mechanistic insights into an appropriate maternal administration of soybean products on prevention of breast cancer later in offspring life. [ABSTRACT FROM AUTHOR]

Background: The association between obesity and breast cancer (BC) has been extensively studied among US, European and Asian study populations, with often conflicting evidence. However, despite the increasing prevalence of obesity and associated conditions in Africa, the continent with the highest age-standardized BC mortality rate globally, few studies have evaluated this association, and none has examined in relation to molecular subtypes among African women. The current analysis examines the association between body composition, defined by body mass index (BMI), height, and weight, and BC by molecular subtype among African women.Methods: We estimated odds ratios (ORs) and 95% confidence intervals (95% CI) for the association between measures of body composition and BC and molecular subtypes among 419 histologically confirmed cases of BC and 286 healthy controls from the Mechanisms for Established and Novel Risk Factors for Breast Cancer in Women of Nigerian Descent (MEND) case-control study.Results: Higher BMI (aOR: 0.79; 95% CI: 0.67, 0.95) and weight (aOR: 0.83; 95% CI: 0.69, 0.98) were associated with reduced odds of BC in adjusted models, while height was associated with non-statistically significant increased odds of BC (aOR: 1.07, 95% CI: 0.90, 1.28). In pre/peri-menopausal, but not post-menopausal women, both higher BMI and weight were significantly associated with reduced odds of BC. Further, higher BMI was associated with reduced odds of Luminal A, Luminal B, and HER2-enriched BC among pre/peri-menopausal women, and reduced odds of triple-negative BC among post-menopausal women.Conclusions: Higher BMI and weight were associated with reduced odds of BC overall and by molecular subtype among West African women. Larger studies of women of African descent are needed to definitively characterize these associations and inform cancer prevention strategies. [ABSTRACT FROM AUTHOR]

Abstract Background Estrogen receptor-α (ERα)-negative breast cancer is clinically aggressive and normally does not respond to conventional estrogen target-directed therapies. The soybean isoflavone, genistein (GE), has been shown to prevent and inhibit breast cancer and recent studies have suggested that GE can enhance the anticancer capacity of an estrogen antagonist, tamoxifen (TAM), especially in ERα-positive breast cancer cells. However, the role of GE in ERα-negative breast cancer remains unknown. Methods We have evaluated the in vitro and in vivo epigenetic effects of GE on ERα reactivation by using MTT assay, real-time reverse transcription-polymerase chain reaction (RT-PCR) assay, western-blot assay, immunoprecipitation (ChIP) assay, immunohistochemistry and epigenetic enzymatic activity analysis. Preclinical mouse models including xenograft and spontaneous breast cancer mouse models were used to test the efficacy of GE in vivo. Results We found that GE can reactivate ERα expression and this effect was synergistically enhanced when combined with a histone deacetylase (HDAC) inhibitor, trichostatin A (TSA), in ERα-negative MDA-MB-231 breast cancer cells. GE treatment also re-sensitized ERα-dependent cellular responses to activator 17β-estradiol (E2) and antagonist TAM. Further studies revealed that GE can lead to remodeling of the chromatin structure in the ERα promoter thereby contributing to ERα reactivation. Consistently, dietary GE significantly prevented cancer development and reduced the growth of ERα-negative mouse breast tumors. Dietary GE further enhanced TAM-induced anti-cancer efficacy due at least in part to epigenetic ERα reactivation. Conclusions Our studies suggest that soybean genistein can epigenetically restore ERα expression, which in turn increases TAM-dependent anti-estrogen therapeutic sensitivity in vitro and in vivo. The results from our studies reveal a novel therapeutic combination approach using bioactive soybean product and anti-hormone therapy in refractory ERα-negative breast cancer which will provide more effective options in breast cancer therapy.

Background: Aberrations in the regulation of cell proliferation perturb cellular homeostasis and lead to malignancies in which dysregulation of the cell cycle and suppressed apoptosis are 2 common phenomena. Combinatorial nutritional approaches could be efficacious in ameliorating these aberrations.Objectives: We sought to investigate the effect of dietary broccoli sprouts (BSp) and green tea polyphenol (GTP) administration on cell cycle progression and apoptosis in mammary tumors.Methods: Forty female HER2/neu transgenic mice were randomly divided into 4 groups and treated with control, 26% BSp (wt:wt) in food, 0.5% GTPs (wt:vol) in drinking water, or combined BSp and GTPs from dams' conception until their pups were killed at 29 wk of age. Pups' tumor growth was monitored weekly for 27 wk. Tumor cell cycle- and apoptosis-related protein expression was measured. Data were analyzed with 2-factor or 3-factor (repeated-measures) ANOVA.Results: Compared with the control group, BSp and/or GTPs decreased tumor incidence (P < 0.05) and combined BSp and GTPs synergistically [combination index (CIn) < 1] reduced tumor volume over time (P-time < 0.01). BSp and/or GTPs upregulated the expression of phosphatase and tension homolog, P16, and P53 (P < 0.05) and downregulated myelocytomatosis oncogene, Bmi1 polycomb ring finger oncogene, and telomerase reverse transcriptase (P < 0.05) compared with the control group. Combined BSp and GTPs synergistically (CIn < 1) downregulated the expression of cyclin B1, D1, and E1 and cyclin-dependent kinase 1, 2, and 4 (P < 0.05) compared with the control group. Moreover, combined BSp and GTPs induced apoptosis by regulating Bcl-2-associated X protein and B-cell lymphoma 2 (P < 0.05). BSp and/or GTPs also reduced the expression of DNA methyltransferase 1, 3A, and 3B and histone deacetylase 1 compared with the control group (P < 0.05).Conclusions: Collectively, lifelong BSp and GTP administration can prevent estrogen receptor-negative mammary tumorigenesis through cell cycle arrest and inducing apoptosis in HER2/neu mice. [ABSTRACT FROM AUTHOR]

Background: Nigeria reports the highest age-standardized mortality rate for breast cancer (BC) among African countries and disproportionately high rates of high-grade cancer. Histological grade is a strong predictor of mortality, and evidence suggests that educational attainment influences cancer outcomes. Objective: We characterize the association between educational trends across the lifecourse and BC grade at diagnosis. Methods: Data on 224 BC patients enrolled in the Mechanisms for Established and Novel Risk Factors for Breast Cancer in Nigerian Women (MEND) study was analyzed. Participant and parental (mother and father) education was categorized as low (primary school or less) or high (secondary school or greater). Accordingly, the educational trend across the life-course was determined for each participant relative to each parent: stable high, increasing, decreasing, or stable low. BC grade was classified as high (grade 3) or low (grades 1–2). Findings: About 34% of participants, 71% of fathers, and 85% of mothers had low education. Approximately one-third of participants were diagnosed with high-grade BC. Participants with low-grade BC were more likely to have highly educated fathers (p = 0.04). After adjusting for age, comorbidities, marital status and mammogram screening, participants with highly educated fathers were 60% less likely to have high-grade BC (aOR 0.41; 95% CI 0.20 to 0.84) compared to those with less-educated fathers. Stable high life-course education relative to father was also associated with a significantly lower likelihood of having high-grade BC (aOR 0.36; 95% CI 0.15 to 0.87) compared to stable low life-course education. No significant associations were observed for the participant’s education, mother’s education, or life-course education relative to mother. Conclusions: Early-life socioeconomic status (SES) may influence BC grade. This deserves further study to inform policies that may be useful in reducing high-grade BC in Nigeria. [ABSTRACT FROM AUTHOR]

Since dietary polyphenols can have beneficial effects in prevention and treatment of cancer, we tested the hypothesis that breast cancer patients’ intestinal microbiota is modulated by genistein (GE), an isoflavone found in soy, and that microbial alterations may offset the side effects brought about by chemotherapy. We demonstrated successful humanization of germ-free mice by transplanting fecal samples from breast cancer patients before and after chemotherapy. Mice were then grouped based on chemotherapy status and GE or control diet. We did not find any significant differences between pre-chemotherapy and post-chemotherapy bacterial composition and abundances. Germ-free mice on a GE diet showed differences in microbial composition as compared to mice on control diet. Four weeks after introduction of the customized GE diet, there was distinct clustering of GE-fed mice as compared to the control-fed group. In the gut microbiome of GE-treated humanized mice, there was an increase in abundance of genera Lactococcus and Eubacterium. Phylum Verrucomicrobia showed statistically significant (p = 0.02) differences in abundances between the GE-fed and control-fed groups. There was an increase in bacteria belonging to family Lachnospiraceae and Ruminococcaceae in GE-fed mice. Marked changes were observed in GE catabolism in mice humanized with fecal material from two of three patients’ post-chemotherapy with complete disappearance of 4-ethylphenol and 2-(4-hydroxyphenol) propionic acid conjugates. The post-tumor samples did not show any distinct clustering of the gut microbiota between the two diet groups. There was an increase in latency of about 25% for tumor growth of the humanized mice that were on a GE diet as compared to humanized mice on a control diet. The average tumor size for the GE group was significantly decreased compared to the non-GE group. Collectively, our results suggest that the intestinal microbiota becomes altered with a GE diet before induction of tumor. Our findings indicate that GE modulates the microbiome in humanized mice that may contribute to its effects on increasing the latency of breast tumor and reducing tumor growth. [ABSTRACT FROM AUTHOR]

With cancer often classified as a disease that has an important epigenetic component, natural compounds that have the ability to regulate the epigenome become ideal candidates for study. Humans have a complex diet, which illustrates the need to elucidate the mechanisms of interaction between these bioactive compounds in combination. The natural compounds withaferin A (WA), from the Indian winter cherry, and sulforaphane (SFN), from cruciferous vegetables, have numerous anti-cancer effects and some report their ability to regulate epigenetic processes. Our study is the first to investigate the combinatorial effects of low physiologically achievable concentrations of WA and SFN on breast cancer cell proliferation, histone deacetylase1 (HDAC1) and DNA methyltransferases (DNMTs). No adverse effects were observed on control cells at optimal concentrations. There was synergistic inhibition of cellular viability in MCF-7 cells and a greater induction of apoptosis with the combinatorial approach than with either compound administered alone in both MDA-MB-231 and MCF-7 cells. HDAC expression was down-regulated at multiple levels. Lastly, we determined the combined effects of these bioactive compounds on the pro-apoptotic BAX and anti-apoptotic BCL-2 and found decreases in BCL-2 and increases in BAX. Taken together, our findings demonstrate the ability of low concentrations of combinatorial WA and SFN to promote cancer cell death and regulate key epigenetic modifiers in human breast cancer cells. [ABSTRACT FROM AUTHOR]

Breast cancer is the second most common cancer and a leading cause of cancer death in women. Specifically, estrogen receptor-α (ERα)-negative breast cancers are clinically more aggressive and normally do not respond to conventional hormone-directed therapies such as tamoxifen. Although epigenetic-based therapies such as 5-aza-2’-deoxycytidine and/or trichostatin A as DNA methyltransferase (DNMT) and histone deacetylase (HDAC) inhibitors, respectively, can regulate the expression of ERα, this can often lead to a number of side effects. Plant-based dietary compounds such as resveratrol and pterostilbene in novel combinatorial therapy provides new avenues to target these side effects and provide similar results with a higher level of safety. Here, we report that combinatorial resveratrol and pterostilbene leads to the reactivation of ERα expression in ERα-negative breast cancer cells in a time-dependent manner. Chromatin immunoprecipitation analysis of the ERα promoter in each cell type revealed an increase in enrichment of acetyl-H3, acetyl-H3lysine9 (H3K9) and acetyl-H4 active chromatin markers in the ERα promoter region after combinatorial treatment. This treatment also resulted in a significant change in HDAC and histone acetyl transferase (HAT) enzyme activity in these cells after 3 days of treatments. The combination resulted in a significant decrease in DNMT enzyme activity and 5-methylcytosine levels in MDA-MB-157 breast cancer cells. Moreover, reactivation of ERα expression by resveratrol combined with pterostilbene was found to sensitize ERα-dependent response to 17β-estradiol (E2)-mediated cellular proliferation and antagonist 4-hydroxytamoxifen (4-OHT)-mediated inhibition of cellular proliferation in ERα-negative breast cancer cells. E2 and 4-OHT further affected the ERα-responsive downstream progesterone receptor (PGR) gene in ERα reactivated MDA-MB-157 cells. Collectively, our findings provide a new and safer way of restoring ERα expression by regulating epigenetic mechanisms with the use of phytochemicals in combinatorial therapy. This combination can further provide effective treatment options for hormonal refractory breast cancer with available anti-hormonal therapy. [ABSTRACT FROM AUTHOR]

Dietary phytochemicals are currently being studied with great interest due to their ability to regulate the epigenome resulting in prevention of cancer. Some natural botanicals have been reported to have enhanced and synergistic impact on cancer suppression when administered at optimum concentrations and in-conjunction. Sulforaphane (SFN) is an isothiocyanate found in cruciferous vegetables and sodium butyrate (NaB) is a short-chain fatty acid produced by gut microbiota. They have been intensively explored due to numerous anti-cancerous properties and ability to modulate epigenetic machinery by inhibition of histone deacetylase (HDAC). Genistein (GE), present in soy, is a known DNA methyltransferase (DNMT) inhibitor. While combined chemoprotective epigenetic effects induced by SFN and GE have been investigated, the key impact of combinatorial SFN-NaB, GE-NaB, and SFN-GE-NaB bioactive components in regulation of various mechanisms are poorly defined. In the present study, we found that combinations of dietary compounds had synergistic effects in decreasing cellular viability at lower dosages than their single dosages in breast cancer cell lines. The respective combinations limited growth and increased apoptosis and necrosis in cancerous cells among which the tri-combination displayed the most significant impact. Additionally, the respective combinations of compounds arrested MDA-MB-231 and MCF-7 breast cancer cells at G2/M phase. Our further mechanistic evaluation revealed that respective di-combinations and tri-combination had higher impact in down-regulation of DNMTs (DNMT3A and DNMT3B), HDACs (HDAC1, HDAC6 and HDAC11), histone methyltransferases (EZH2 and SUV39H1) and histone acetyltransferases (GCN5, PCAF, P300 and CBP) levels as compared to singly administered compounds. We also found that these combinations exhibited global epigenetic changes by inhibition of DNMT and HDAC activity, histone H3 at lysine 27 methylation (H3K27me) and histone H3 at lysine 9 methylation (H3K9me) levels, and by induction of histone acetyltransferases activity. Collectively, our investigation indicates that combined SFN, GE and NaB is highly effective in inhibiting breast cancer genesis by, at least in part, regulating epigenetic modifications, which may have implications in breast cancer therapy. • The combined SFN-GE, SFN-NaB, GE-NaB and all dietary compounds synergistically inhibited breast cancer cells proliferation. • The respective combination treatments induced apoptosis and cell cycle arrest in MDA-MB-231 and MCF-7 breast cancer cells. • The di-combinations and tri-combination had the most promising impact on the expression of key DNMTs, HDACs, HMTs and HATs. • The combinations displayed global epigenetic changes by regulating key enzymatic activities and histone methylation levels. [ABSTRACT FROM AUTHOR]

Background: Estrogen receptor-α (ERα)-negative breast cancer is clinically aggressive and normally does not respond to conventional estrogen target-directed therapies. The soybean isoflavone, genistein (GE), has been shown to prevent and inhibit breast cancer and recent studies have suggested that GE can enhance the anticancer capacity of an estrogen antagonist, tamoxifen (TAM), especially in ERα-positive breast cancer cells. However, the role of GE in ERα-negative breast cancer remains unknown. Methods: We have evaluated the in vitro and in vivo epigenetic effects of GE on ERα reactivation by using MTT assay, real-time reverse transcription-polymerase chain reaction (RT-PCR) assay, western-blot assay, immunoprecipitation (ChIP) assay, immunohistochemistry and epigenetic enzymatic activity analysis. Preclinical mouse models including xenograft and spontaneous breast cancer mouse models were used to test the efficacy of GE in vivo. Results: We found that GE can reactivate ERα expression and this effect was synergistically enhanced when combined with a histone deacetylase (HDAC) inhibitor, trichostatin A (TSA), in ERα-negative MDA-MB-231 breast cancer cells. GE treatment also re-sensitized ERα-dependent cellular responses to activator 17β-estradiol (E2) and antagonist TAM. Further studies revealed that GE can lead to remodeling of the chromatin structure in the ERα promoter thereby contributing to ERα reactivation. Consistently, dietary GE significantly prevented cancer development and reduced the growth of ERα-negative mouse breast tumors. Dietary GE further enhanced TAM-induced anti-cancer efficacy due at least in part to epigenetic ERα reactivation. Conclusions: Our studies suggest that soybean genistein can epigenetically restore ERα expression, which in turn increases TAM-dependent anti-estrogen therapeutic sensitivity in vitro and in vivo. The results from our studies reveal a novel therapeutic combination approach using bioactive soybean product and anti-hormone therapy in refractory ERα-negative breast cancer which will provide more effective options in breast cancer therapy. [ABSTRACT FROM AUTHOR]

Breast cancer is the most common cancer and the leading cause of cancer death in women. Although tamoxifen therapy is successful for some patients, it does not provide adequate benefit for those who have estrogen receptor (ER)-negative cancers. Therefore, we approached novel treatment strategies by combining two potential bioactive dietary supplements for the reactivation of ERa expression for effective treatment of ERα-negative breast cancer with tamoxifen. Bioactive dietary supplements such as green tea polyphenols (GTPs) and sulforaphane (SFN) inhibit DNA methyltransferases (DNMTs) and histone deacetylases (HDACs), respectively, which are of central importance to cancer prevention. In the present study, we have observed that treatment of ERa-negative breast cancer cells with GTPs and SFN alone or in combination leads to the reactivation of ERa expression. The combination of 20 mg/mL GTPs and 5 μM SFN was found to be the optimal dose of ERareactivation at 3 days in MDA-MB-231 cells. The reactivation of ERa expression was consistently correlated with ERa promoter hypomethylation and hyperacetylation. Chromatin immunoprecipitation (ChIP) analysis of the ERa promoter revealed that GTPs and SFN altered the binding of ERα-transcriptional co-repressor complex thereby contributing to ERareactivation. In addition, treatment with tamoxifen in combination with GTPs and SFN significantly increased both cell death and inhibition of cellular proliferation in MDA-MB-231 cells in comparison to treatment with tamoxifen alone. Collectively, our findings suggest that a novel combination of bioactive-HDAC inhibitors with bioactive-demethylating agents is a promising strategy for the effective treatment of hormonal refractory breast cancer with available anti-estrogens. [ABSTRACT FROM AUTHOR]

Background: The status of estrogen receptor-a (ERα) is critical to the clinical prognosis and thERαpeutic approach in breast cancer. ERα-negative breast cancer is clinically aggressive and has a poor prognosis because of the lack of hormone target-directed thERαpies. Previous studies have shown that epigenetic regulation plays a major role in ERα silencing in human breast cancer cells. Dietary green tea polyphenol, (-)-epigallocatechin-3-gallate (EGCG), is believed to be an anticancer agent in part through its regulation of epigenetic processes. Results: In our current studies, we found that EGCG can reactivate ERα expression in ERα-negative MDA-MB-231 breast cancer cells. Combination studies using EGCG with the histone deacetylase (HDAC) inhibitor, trichostatin A (TSA), revealed a synergistic effect of reactivation of ERα expression in ERα-negative breast cancer cells. Reactivation of ERα expression by EGCG and TSA treatment was found to sensitize ERα-dependent cellular responses to activator 17b-estradiol (E2) and antagonist tamoxifen in ERα-negative breast cancer cells. We also found that EGCG can lead to remodeling of the chromatin structure of the ERα promoter by altering histone acetylation and methylation status thereby resulting in ERα reactivation. A decreased binding of the transcription repressor complex, Rb/p130-E2F4/5-HDAC1-SUV39H1-DNMT1, in the regulatory region of the ERα promoter also contributes to ERα transcriptional activation through treatment with EGCG and/or TSA. Conclusions: Collectively, these studies show that green tea EGCG can restore ERα expression by regulating epigenetic mechanisms, and this effect is enhanced when combined with an HDAC inhibitor. This study will facilitate more effective uses of combination approaches in breast cancer thERαpy and will help to explore more effective chemothERαpeutic strategies toward hormone-resistant breast cancer. [ABSTRACT FROM AUTHOR]

With the recent advancement of genetic screening for testing susceptibility to mammary oncogenesis in women, the relevance of the gene−environment interaction has become progressively apparent in the context of aberrant gene expressions. Fetal exposure to external stressors, hormones, and nutrients, along with the inherited genome, impact its traits, including cancer susceptibility. Currently, there is increasing interest in the role of epigenetic biomarkers such as genomic methylation signatures, plasma microRNAs, and alterations in cell-signaling pathways in the diagnosis and primary prevention of breast cancer, as well as its prognosis. Polyphenols like natural stilbenes have been shown to be effective in chemoprevention by exerting cytotoxic effects that can stall cell proliferation. Besides possessing antioxidant properties against the DNA-damaging effects of reactive oxygen species, stilbenes have also been observed to modulate cell-signaling pathways. With the increasing trend of early-life screening for hereditary breast cancer risks, the potency of different phytochemicals in harnessing the epigenetic biomarkers of breast cancer risk demand more investigation. This review will explore means of exploiting the abilities of stilbenes in altering the underlying factors that influence breast cancer risk, as well as the appearance of associated biomarkers. [ABSTRACT FROM AUTHOR]

Breast cancer is a sporadic disease with genetic and epigenetic components. Genomic instability in breast cancer leads to mutations, copy number variations, and genetic rearrangements, while epigenetic remodeling involves alteration by DNA methylation, histone modification and microRNAs (miRNAs) of gene expression profiles. The accrued scientific findings strongly suggest epigenetic dysregulation in breast cancer pathogenesis though genomic instability is central to breast cancer hallmarks. Being reversible and plastic, epigenetic processes appear more amenable toward therapeutic intervention than the more unidirectional genetic alterations. In this review, we discuss the epigenetic reprogramming associated with breast cancer such as shuffling of DNA methylation, histone acetylation, histone methylation, and miRNAs expression profiles. As part of this, we illustrate how epigenetic instability orchestrates the attainment of cancer hallmarks which stimulate the neoplastic transformation-tumorigenesis-malignancy cascades. As reversibility of epigenetic controls is a promising feature to optimize for devising novel therapeutic approaches, we also focus on the strategies for restoring the epistate that favor improved disease outcome and therapeutic intervention. [ABSTRACT FROM AUTHOR]

Triple-negative breast cancer comprises approximately 15–20% of all breast cancers diagnosed and is nearly twice as common in black women than white women in the United States. We evaluated the effects of two epigenetic-modifying compounds on markers of growth potential in several triple-negative breast cancer cell lines. Suberoylanilide hydroxamic acid (SAHA), a histone deacetylase (HDAC) inhibitor currently used in the treatment of cutaneous T cell lymphoma, was administered to triple-negative breast cancer cells alone or in combination with epigallocatechin-3-gallate (EGCG), a DNA methyltransferase (DNMT) inhibitor isolated from green tea. The compounds affected the expression of oncogenic miR-221/222 and tumor suppressors, p27 and PTEN, in addition to estrogen receptor alpha (ERα). E-cadherin expression was increased while N-cadherin was decreased, indicating a more epithelial phenotype. In addition, the activity of DNMTs was diminished with the treatments, and there was a significant enrichment of AcH3 within the promoter of p27 and PTEN, suggesting a role of epigenetic mechanisms for the aforementioned changes. These results translated to reduced migration of the triple-negative breast cancer cells with the treatments. Together, these findings support the role of SAHA and EGCG in limiting growth and proliferation of breast cancer cells. [ABSTRACT FROM AUTHOR]

Breast cancer is the second most common cancer and the second leading cause of death from cancer among women in the United States (US). Cancer prevention and therapy through the use of phytochemicals that have epigenetic properties has gained considerable interest during the past few decades. Such dietary components include, but are not limited to, grape seed proanthocyanidins (GSPs) and resveratrol (Res), both of which are present in red wine. In this study, we report for the first time the synergistic effects of GSPs and Res on inhibiting MDA-MB-231 and MCF-7 human breast cancer cells. Our results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays and clonogenic assays indicate that treatments with the combinations of GSPs and Res synergistically decreased cell viability and posttreatment cell proliferation in both cell lines. Additional analyses show that treatments with GSPs and Res in combination synergistically induced apoptosis in MDA-MB-231 cells by upregulating Bax expression and down-regulating Bcl-2 expression. DNA methyltransferase (DNMT) activity and histone deacetylase (HDAC) activity were greatly reduced in MDA-MB-231 and MCF-7 cells after treatments with GSPs and Res in combination. Collectively, our findings suggest that GSPs and Res synergistically inhibit human breast cancer cells through inducing apoptosis, as well as modulating DNA methylation and histone modifications. [ABSTRACT FROM AUTHOR]

Dietary compounds that possess the properties of altering epigenetic processes are gaining popularity as targets for cancer prevention studies. These compounds when administered at optimal concentrations and especially in combination can have enhanced effects in cancer prevention or therapy. It is important to study the interaction of two or more compounds in order to assess their role in enhancing prevention. Genistein (GEN), found in soy, has been extensively studied for its role as an epigenetic modifier especially as a DNA methyltransferase (DNMT) inhibitor and sulforaphane (SFN), found in cruciferous vegetables, is known as a histone deacetylase (HDAC) inhibitor. However, very little is known about the effects of these two compounds in conjunction in breast cancer prevention or therapy. In our current study, we determined that, at certain doses, the compounds have synergistic effects in decreasing cellular viability of breast cancer cell lines. Our results indicate that the combination of GEN and SFN is much more effective than their single doses in increasing the rate of apoptosis and lowering the colony forming potential of these cells. We determined that these compounds inhibit cell cycle progression to G2 phase in MDA-MB-231 and G1 phase in MCF-7 breast cancer cell lines. Additionally, we determined that the combination is effective as an HDAC and histone methyltransferase (HMT) inhibitor. Furthermore, we demonstrated that this combination downregulates the levels of HDAC2 and HDAC3 both at the mRNA and protein levels. We also found that these compounds have the potential to downregulate KLF4 levels, which plays an important role in stem cell formation. The combination of GEN and SFN is also effective in downregulating hTERT levels, which is known to be activated when KLF4 binds to its promoter region. Our hypothesis is further strengthened by in vivo studies, where the combination is administered to transgenic mice in the form of genistein and SFN-enriched broccoli sprouts. We have demonstrated that the combination is more effective in preventing or treating mammary cancer via extending tumor latency and reducing tumor volumes/sizes than either of these dietary components administered alone. These results are consistent with our in vitro study suggesting potential preventive and therapeutic effects of this novel dietary combinatorial approach against breast cancer. [ABSTRACT FROM AUTHOR]

Little is known about the effects of combinatorial dietary compounds on the regulation of epigenetic mechanisms involved in breast cancer prevention. The human diet consists of a multitude of components, and there is a need to elucidate how certain compounds interact in collaboration. Withaferin A (WA), found in the Indian winter cherry and documented as a DNA methyltransferase (DNMT) inhibitor, and sulforaphane (SFN), a well-known histone deacetylase (HDAC) inhibitor found in cruciferous vegetables, are two epigenetic modifying compounds that have only recently been studied in conjunction. The use of DNMT and HDAC inhibitors to reverse the malignant expression of certain genes in breast cancer has shown considerable promise. Previously, we found that SFN + WA synergistically promote breast cancer cell death. Herein, we determined that these compounds inhibit cell cycle progression from S to G2 phase in MDA-MB-231 and MCF-7 breast cancer. Furthermore, we demonstrate that this unique combination of epigenetic modifying compounds down-regulates the levels of Cyclin D1 and CDK4 , and pRB; conversely, the levels of E2F mRNA and tumor suppressor p21 are increased independently of p53. We find these events coincide with an increase in unrestricted histone methylation. We propose SFN + WA-induced breast cancer cell death is attributed, in part, to epigenetic modifications that result in the modulated expression of key genes responsible for the regulation of cancer cell senescence. [ABSTRACT FROM AUTHOR]

Overnutrition-induced obesity and metabolic dysregulation are considered major risk factors contributing to breast cancer. The origin of both obesity and breast cancer can retrospect to early development in human lifespan. Genistein (GE), a natural isoflavone enriched in soybean products, has been proposed to associate with a lower risk of breast cancer and various metabolic disorders. Our study aimed to determine the effects of maternal exposure to soybean dietary GE on prevention of overnutrition-induced breast cancer later in life and explore potential mechanisms in different mouse models. Our results showed that maternal dietary GE treatment improved offspring metabolic functions by significantly attenuating high-fat diet-induced body fat accumulation, lipid panel abnormalities and glucose intolerance in mice offspring. Importantly, maternal dietary GE exposure effectively delayed high-fat diet-simulated mammary tumor development in female offspring. Mechanistically, we found that maternal dietary GE may exert its chemopreventive effects through affecting essential regulatory gene expression in control of metabolism, inflammation and tumor development via, at least in part, regulation of offspring gut microbiome, bacterial metabolites and epigenetic profiles. Altogether, our findings indicate that maternal GE consumption is an effective intervention approach leading to early-life prevention of obesity-related metabolic disorders and breast cancer later in life through dynamically influencing the interplay between early-life gut microbiota, key microbial metabolite profiles and offspring epigenome. [ABSTRACT FROM AUTHOR]

Available treatment strategies against estrogen receptor (ER)-negative breast cancer patients are limited due to their poor response to hormonal therapy. We have shown previously that the combinations of green tea polyphenols (GTPs), a dietary DNA methyltransferase inhibitor, and sulforaphane (SFN), a dietary histone deacetylase inhibitor, reactivate ERα expression in ERα-negative MDA-MB-231 cells. Here, we investigated the functional significance of ERα reactivation in the reactivation of silenced tumor suppressor genes (TSGs) in ERα-negative human breast cancer cells. We found that the treatment of MDA-MB-231 cells with the combinations of GTPs and SFN leads to the reactivation of silenced TSGs such as p21 CIP1/WAF1 and KLOTHO through active chromatin modifications. Further, GTPs- and SFN-mediated reactivation of TSGs was, at least in part, dependent on ERα reactivation in ERα-negative MDA-MB-231 cells. Collectively, our findings suggest that a novel combination of bioactive dietary supplements could further be explored as an effective therapeutic option against hormonal refractory breast cancer. [ABSTRACT FROM AUTHOR]