Lifestyle factors are responsible for a considerable portion of cancer incidence worldwide, but credible estimates from the World Health Organization and the International Agency for Research on Cancer (IARC) suggest that the fraction of cancers attributable to toxic environmental exposures is between 7% and 19%. To explore the hypothesis that low-dose exposures to mixtures of chemicals in the environment may be combining to contribute to environmental carcinogenesis, we reviewed 11 hallmark phenotypes of cancer, multiple priority target sites for disruption in each area and prototypical chemical

The high concentrations of individual phytochemicals in vitro studies cannot be physiologically achieved in humans. Our solution for this concentration gap between in vitro and human studies is to combine two or more phytochemicals. We screened 12 phytochemicals by pairwise combining two compounds at a low level to select combinations exerting the synergistic inhibitory effect of breast cancer cell proliferation. A novel combination of luteolin at 30 μM (LUT30) and indole-3-carbinol 40 μM (I3C40) identified that this combination (L30I40) synergistically constrains ERα+ breast cancer cell (MCF7 and T47D) proliferation only, but not triple-negative breast cancer cells. At the same time, the individual LUT30 and I3C40 do not have this anti-proliferative effect in ERα+ breast cancer cells. Moreover, this combination L30I40 does not have toxicity on endothelial cells compared to the current commercial drugs. Similarly, the combination of LUT and I3C (LUT10 mg + I3C10 mg/kg/day) (IP injection) synergistically suppresses tumor growth in MCF7 cells-derived xenograft mice, but the individual LUT (10 mg/kg/day) and I3C (20 mg/kg/day) do not show an inhibitory effect. This combination synergistically downregulates two major therapeutic targets ERα and cyclin dependent kinase (CDK) 4/6/retinoblastoma (Rb) pathway, both in cultured cells and xenograft tumors. These results provide a solid foundation that a combination of LUT and I3C may be a practical approach to treat ERα+ breast cancer cells after clinical trials.

Breast cancer is the most common malignancy among Arab women in Eastern Mediterranean Region (EMR). The incidence of breast cancer has substantially increased in recent years among this women population, especially those younger than 50, and the incidence is expected to double by 2030. Considerable experimental evidence supports the potential role of dietary habits and lifestyle in cancer etiology and cancer prevention. In this review we examined the literature for evidence to link dietary choices and the rise in incidence and mortality of breast cancer among women in EMR. A literature search was conducted in PubMed and Ovid MEDLINE databases up to December 2017. The search terms used are breast cancer prevalence, breast cancer incidence worldwide, breast cancer and: nutrition, protein intake, vitamin D intake, fat intake, phytoestrogens, EMR, Arab, Middle East, Gulf countries, the UAE Arab women, breast cancer risk, diet, and chemoprevention. We found evidence to suggest that there is an alarming epidemic of obesity among women in most of the EMR countries, especially Gulf Cooperation Council (GCC) countries. The rise in the new breast cancer cases among women could be attributed to excess body weight. Their dietary pattern, which correlates with obesity, can be an important factor in the etiology of cancer. Although very few studies were found to support a direct causal relationship between obesity and breast cancer in the EMR, circumstantial evidence clearly points to the possible role of the epidemic, obesity, in this population and the startling rise in cases of breast cancer. Well-designed and systematic studies are urgently needed to confirm these associations and to elucidate potential mechanisms. More urgently, calls to action are needed in many sectors and at all levels of society, to establish intensive strategies for reducing obesity and promoting an overall healthy diet. Continued and expanded research on diet, lifestyle, and breast cancer risk is urgently needed to build the foundation for future progress in evidence-based public health efforts.

Lifestyle factors are responsible for a considerable portion of cancer incidence worldwide, but credible estimates from the World Health Organization and the International Agency for Research on Cancer (IARC) suggest that the fraction of cancers attributable to toxic environmental exposures is between 7% and 19%. To explore the hypothesis that low-dose exposures to mixtures of chemicals in the environment may be combining to contribute to environmental carcinogenesis, we reviewed 11 hallmark phenotypes of cancer, multiple priority target sites for disruption in each area and prototypical chemical disruptors for all targets, this included dose-response characterizations, evidence of low-dose effects and cross-hallmark effects for all targets and chemicals. In total, 85 examples of chemicals were reviewed for actions on key pathways/mechanisms related to carcinogenesis. Only 15% (13/85) were found to have evidence of a dose-response threshold, whereas 59% (50/85) exerted low-dose effects. No dose-response information was found for the remaining 26% (22/85). Our analysis suggests that the cumulative effects of individual (non-carcinogenic) chemicals acting on different pathways, and a variety of related systems, organs, tissues and cells could plausibly conspire to produce carcinogenic synergies. Additional basic research on carcinogenesis and research focused on low-dose effects of chemical mixtures needs to be rigorously pursued before the merits of this hypothesis can be further advanced. However, the structure of the World Health Organization International Programme on Chemical Safety ‘Mode of Action’ framework should be revisited as it has inherent weaknesses that are not fully aligned with our current understanding of cancer biology.

We examined the expression kinetics of some of the aryl hydrocarbon receptor (AhR)-regulated genes in LA1 variant cells compared to wild type (WT) Hepa-1 mouse hepatoma cell lines, and we investigated the stability of AhR protein as a key step in the function of this receptor. Treatment of both cell types with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) resulted in increased CYP1A1 and CYP1B1 mRNA with a subsequent down regulation of AhR. We show here that co-treatment with transcription inhibitor actinomycin D (ActD) has reversed the TCDD-induced depletion of AhR protein in WT. However, the proteolytic degradation of AhR in absence of TCDD was significantly higher in LA1 cells than in WT, and ActD treatment reduced this loss. Induction of CYP1A1 and CYP1B1 mRNA by TCDD in WT cells each exhibited bursts of activity in the initial hour which were about 3-fold greater than in LAI cells. The induced mRNA levels in LA1 exhibited a slow and sustained increase approximating the WT levels by 20h. The induction of two other AhR-regulated genes also showed comparable turnover differences between the two types of cell. Thus, altered regulation of the AhR responsive genes in LA1 may result from a difference in AhR stability.

A correction has been published: Carcinogenesis, Volume 37, Issue 3, March 2016, Page 344, https://doi.org/10.1093/carcin/bgv155.

The purpose of this review is to stimulate new ideas regarding low-dose environmental mixtures and carcinogens and their potential to promote invasion and metastasis. Whereas a number of chapters in this review are devoted to the role of low-dose environmental mixtures and carcinogens in the promotion of invasion and metastasis in specific tumors such as breast and prostate, the overarching theme is the role of low-dose carcinogens in the progression of cancer stem cells. It is becoming clearer that cancer stem cells in a tumor are the ones that assume invasive properties and colonize distant organs. Therefore, low-dose contaminants that trigger epithelial-mesenchymal transition, for example, in these cells are of particular interest in this review. This we hope will lead to the collaboration between scientists who have dedicated their professional life to the study of carcinogens and those whose interests are exclusively in the arena of tissue invasion and metastasis.

The purpose of this review is to stimulate new ideas regarding low-dose environmental mixtures and carcinogens and their potential to promote invasion and metastasis. Whereas a number of chapters in this review are devoted to the role of low-dose environmental mixtures and carcinogens in the promotion of invasion and metastasis in specific tumors such as breast and prostate, the overarching theme is the role of low-dose carcinogens in the progression of cancer stem cells. It is becoming clearer that cancer stem cells in a tumor are the ones that assume invasive properties and colonize distant organs. Therefore, low-dose contaminants that trigger epithelial-mesenchymal transition, for example, in these cells are of particular interest in this review. This we hope will lead to the collaboration between scientists who have dedicated their professional life to the study of carcinogens and those whose interests are exclusively in the arena of tissue invasion and metastasis.

The aryl hydrocarbon receptor (AhR), a transcription factor that is best known for its role in mediating the toxic responses elicited by poly aromatic hydrocarbons as well as many other environmental factors; is also involved in breast cancer progression. We previously reported that stable knockdown of AhR decreased the tumorigenic properties of the highly metastatic MDA-MB-231 breast cancer cell line; whereas ectopic overexpression of AhR was sufficient to transform immortalized human mammary epithelial cells to exhibit malignant phenotypes. In the present study we investigated the genes that are differentially regulated by AhR and are controlling cellular processes linked to breast cancer. We used Affymetrix Human GeneChip 1.0-ST whole transcriptome arrays to analyze alterations of gene expression resulting from stable AhR knockdown in the MDA-MB-231 breast cancer cell line. The expression of 144 genes was significantly altered with a ≥2.0-fold change and a multiple test corrected p-value ≤0.05, as a result of AhR knockdown. We demonstrate that AhR knockdown alters the expression of several genes known to be linked to cancer. These genes include those involved in tryptophan metabolism (KYNU), cell growth (MUC1 and IL8), cell survival (BIRC3 and BCL3), cell migration and invasion (S100A4 and ABI3), multi-drug resistance (ABCC3) and angiogenesis (VEGFA and CCL2). The identification of the genes and pathways affected by AhR depletion provides new insight into possible molecular events that could explain the reported phenotypic changes. In conclusion AhR knockdown alters the expression of genes known to enhance or inhibit cancer progression; tipping the balance towards a state that counteracts tumor progression.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that regulates a battery of genes in response to exposure to a broad class of environmental poly aromatic hydrocarbons (PAH). AhR is historically characterized for its role in mediating the toxicity and adaptive responses to these chemicals, however mounting evidence has established a role for it in ligand-independent physiological processes and pathological conditions, including cancer. The AhR is overexpressed and constitutively activated in advanced breast cancer cases and was shown to drive the progression of breast cancer. In this article we will review the current state of knowledge on the possible role of AhR in breast cancer and how it will be exploited in targeting AhR for breast cancer therapy.

The aryl hydrocarbon receptor (AhR), a ligand‐activated transcription factor that belongs to the basic‐helix‐loop‐helix (bHLH)–Per‐ARNT‐Sim (PAS) superfamily of transcription factors, mediates toxic response induced by environmental chemicals such as polycyclic aromatic hydrocarbons (PAH). AhR is expressed at high levels in several human breast carcinoma cell lines in direct correlation with the degree of their malignancy. Recent studies suggest a possible role for AhR in cancer independent of PAH. Therefore, we established stable AhR knockdown cells of the human breast cancer cell line MDA‐MB‐231 and analyzed their tumorigenic properties in in vitro and in vivo model systems. In addition we analyzed their response to radiation and chemotherapeutic treatment. AhR knockdown attenuated these cells tumorigenic properties in vitro including proliferation, anchorage independent growth, migration and apoptosis and reduced orthotopic xenograft tumor growth and lung metastasis in vivo . Notably, we observed that AhR knockdown enhanced radiation‐induced apoptosis as well as significantly decreased cell clonogenic survival. Furthermore, AhR knockdown in MDA‐MB‐231 cells sensitized them to paclitaxel treatment, evident by a decrease in the required cytotoxic dose. Subsequent analysis revealed AhR knockdown significantly reduced phosphorylation of AKT, which impacts cell proliferation and survival. Apoptosis‐focused gene expression analyses revealed an altered expression of genes regulating apoptosis in MDA‐MB‐231 cells. Collectively, our data identify AhR as a potential novel therapeutic target in the treatment of metastatic breast cancer.

Abstract The aryl hydrocarbon receptor (AhR) is a ligand-activated basic helix-loop-helix transcription factor which binds environmental poly aromatic hydrocarbons (PAH) and mediates their carcinogenic effects. Recent reports, including data from our laboratory implicate AhR in breast cancer development and progression independent of the receptor occupancy by PAH. In this study we investigated the potential of AhR as a stage specific marker of breast cancer. We examined the expression of AhR by immunohistochemistry in tissue microarrays (TMA) containing 192 specimens of clinically defined three stages of invasive breast cancer: node-negative (NN), node-positive (NP) and metastatic (Met) carcinoma. The TMA were obtained from the National Cancer Institute Cooperative Breast Cancer Tissue Resource [NCI-CBCTR] and were stained for AhR using high affinity antibody. The AhR staining was then scored by three evaluators, including two pathologists who were blinded to the study. Statistical analysis showed a significant correlation between the AhR expression and the carcinoma case type (NN, NP or Met) (p-value of ANOVA is < 0.0003), and although there is a significant difference of AhR expression among carcinoma case type of white women (p< 0.0017) there is no significant difference for black women (p<0.0968). There is also a strong positive correlation between the receptor expression and the tumor grade (p <0.0001). While there is no correlation with the status of estrogen or progesterone receptors (p= 0.1643 and 0.1884, respectively), there is strong correlation of high AhR expression and invasive breast carcinoma in women over 50 years old (p<0.0043). In conclusion, our findings identify the AhR as a new predictive clinical marker for metastatic breast cancer and a unique target for the design of novel selective inhibitors for therapeutic intervention of metastatic breast cancer. More importantly, the AhR overexpression identifies a subset of patients who could benefit from therapy targeting this receptor. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 698. doi:1538-7445.AM2012-698

Abstract Breast cancer is the most common cancer in North American women and is the second leading cause of cancer deaths in women today. Despite recent advances in treatment of breast cancer the development of chemo- and radio-resistance remains a major obstacle in treating advanced breast cancer. The aryl hydrocarbon receptor (AhR) is overexpressed and constitutively activated in several human breast carcinoma cell lines and the expression levels showed strong correlation with the degree of the tumor malignancy. In the current study, we evaluated the effect of reducing AhR expression on the chemo- and radio-sensitization of the MDA-MB231 human breast cancer cell line. We stably knocked down (KD) AhR expression in MDA-MB231 by using retroviral vectors expressing AhR-specific shRNA. Results show that reducing AhR expression enhanced radiation-induced apoptosis as shown by FITC-annexin V assay as well as significantly reducing cell clonogenic survival of MDA-MB231 cell line. In addition, knockdown of AhR alone has significantly impacted expression of genes regulating apoptotic pathways in these cells; increasing the expression of a number of pro-apoptotic genes while anti-apoptotic genes were down-regulated. Furthermore, the AhR-KD MDA-MB231 cells were sensitized to paclitaxol treatment; seen as an increase in the dose-dependent cytotoxicity compared to control cells. Our results suggest that reduction of AhR expression increases the chemo- and radio-sensitivity of MDA-MB231 breast cancer cell line. Therefore, AhR could be a novel target for enhancing efficacy of chemo and radiotherapy of breast cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1455. doi:1538-7445.AM2012-1455

The aryl hydrocarbon receptor (AhR) is a ligand activated basic helix-loop-helix transcription factor that binds to environmental poly aromatic hydrocarbons (PAH) and mediates their toxic and carcinogenic responses. There is ample documentation for the role of AhR in PAH-induced carcinogenicity. However, in this report we addressed whether overexpression of AhR alone is sufficient to induce carcinogenic transformation in human mammary epithelial cells (HMEC). Retroviral expression vectors were used to develop a series of stable cell lines expressing varying levels of AhR protein in an immortalized normal HMEC with relatively low endogenous AhR expression. The resulting increase in AhR expression and activity correlated with the development of cellular malignant phenotypes, most significantly epithelial-tomesenchymal transition. Clones overexpressing AhR by more than 3-fold, exhibited a 50% decrease in population doubling time. Cell cycle analysis revealed that this increase in proliferation rates was due to an enhanced cell cycle progression by increasing the percentage of cells transiting into S- and G2/M phases. Cells overexpressing AhR exhibited enhanced motility and migration. Importantly, these cells acquired the ability to invade matrigel matrix, where more than 80% of plated cells invaded the matrigel matrix within 24 h, whereas none of parental or the vector control HMEC were able to invade matrigel. Collectively, these data provide evidence for a direct role of AhR in the progression of breast carcinoma. The results suggest a novel therapeutic target that could be considered for treatment and prevention of breast cancer progression.

Bringing New Minds and New Methods to Bridging Health Disparity and Equity Sakina E. Eltom, DVM, PhD, Paul B. Tchounwou, ScD, FABI, IOM, and Valerie Montgomery Rice, MD This special issue of the Journal of Health Care for the Poor and Underserved highlights selected papers presented at the Twelfth RCMI International Symposium on Health Disparities, held in Nashville, Tennessee, U.S.A., from December 6-9, 2010. The thematic focus of the meeting was on bringing "new minds and new methods" to the efforts of bridging the gap between health disparity and equity. The symposium provided an excellent platform to bring together scientists conducting pre-clinical, clinical, translational, and health policy research, community members, health care providers, government officials, and private industry, to discuss and exchange innovative ideas and strategic approaches to promote and achieve health equity through research, training and education. The symposium featured oral and poster presentations on important topics such as such as cancer, diabetes and obesity, cardiovascular disease, infectious diseases, stroke, mental health, women health, behavioral health, and social health. In addition, professional development workshops were organized on grantsmanship, mentoring and career development, and research collaborations. In their quest for effective solutions to eliminate health disparities among minorities, poor and underserved communities, researchers from the 18 Research Centers in Minority Institutions (RCMI) and other institutions presented and exchanged their research findings at this meeting. Thus, the research presented covered a wide array of basic sciences, clinical, translational, epidemiological, and health policy research. The reports that have been peer-reviewed and published in this issue of JHCPU are snapshots of some of the science presented at the meeting. They represent the diversity of topics as well as institutions that are part of this scientific engagement. The basic science research is represented in this issue by four articles, each of which takes a molecular approach to investigating a disease problem of high relevance to the poor and underserved population. One report focuses on the decrease in intracellular pH of chemosensitive neurons in brainstem as a possible underlying patho-physiology of Sudden Infant Death Syndrome (SIDS). Another report investigates how the CCR5 chemochine receptor haplotypes may affect HIV/AIDS susceptibility and disease progression, by identifying the transcription factors differentially recruited to the polymorphic sites in the CCR5 promoter. Using an in vitro model of human epidermal keratinocytes, the third report explores molecular mechanisms of how exposure to environmental arsenic in combination with ultraviolet B radiation would cause skin cancer. The last report in this group discusses the scientific evidence linking vitamin D [End Page 3] deficiency as a potential contributor to the risk of cardiovascular diseases among African American populations. Five epidemiological studies are included in this issue and they approach different diseases and health problems. The first report examines the epidemiological data of the 2009 outbreak of H1N1 influenza virus in the U.S., while a second one assesses the relationship between obesity and cardiovascular diseases in four U.S. Southern states in comparison with the population of Colorado. Another report evaluates the effect of exposure to silica on respiratory function in Egyptian miner population. One report investigates the patterns of condom use among women who live in public housing development in Puerto Rico while the last report in this group examines the racial/ ethnic and geographic disparities in diagnosis of late stage prostate cancer in Florida. Issues pertaining to clinical practice and translational research are addressed in the remaining seven reports. A report by Chow assesses the rates and predictors of anal dysplasia in HIV-infected Asian and Pacific Islanders in Hawaii, while a report by De Jesus shares the successful field experience of providing affordable and sustainable HIV molecular tests in resource-constrained countries of the Caribbean. The crucial role of subject recruitment for clinical trials and community-participatory research in reducing health disparities is addressed in two reports. In the first one, Cooper et al. describe the successful model of engaging faith-based community of metropolitan Atlanta as providing sustainable participation in clinical trials at the Morehouse School of Medicine. The institutional review boards (IRBs) regulate the ethical standards governing participation of human subjects in clinical trials and community-based research. In the second report...

Abstract Previous studies have found that AhR is overexpressed and constitutively activated in several breast carcinoma cell lines and the expression levels showed strong correlation with the degree of the tumor malignancy. The objective of this study is to examine the effect of reducing the expression levels of AhR in metastatic human breast carcinoma (HBC) cell lines on their tumorigenic properties; specifically cellular growth, invasion into matrigel, and migration. We designed a series of short hairpin RNA (shRNA) targeting AhR and cloned them in pSuper retroviral vectors, and viruses expressing shRNA were used to infect a battery of HBC cell lines including MDA-MB231, MDA-MB468, BT549 and MT2 to generate cell lines that permanently express siRNA targeting AhR. Clonal cell lines with 50%-90% knockdown of AhR were isolated and characterized in vitro for their tumorigenic properties in comparison to the scramble RNA-expressing control cell lines. Cell cycle analysis by fluorescence-activated cell sorting showed that knock down of AhR expression decreases transition of cells from G1 to S and their progression to G2/M phases of cell cycle. This was seen as an increase of cells in G0/G1 (49% to 61%) and a decreased number of cells in S (26% to 21%) and G2/M phases (25% to 17%). This effect was coupled with a prolonged doubling time and a decreased cellular proliferation. Cells with AhR knock-down exhibited a decreased anchorage independent growth; forming less numbers of colonies in soft agar than control cells. Modified Boyden Chamber invasion assay coupled with fluorescence detection demonstrated that clones of HBC with AhR knock down levels exhibited substantially reduced invasiveness and diminished potential to invade matrigel. These clonal cell lines further exhibited reduced ability to migrate, as measured by scratch wound test. These observations collectively demonstrate a major role for the AhR in regulating tumorigenic phenotypes of metastatic breast cancer. Currently breast cancer prevention research focuses on the identification of novel approaches and/or novel targets. Our findings may establish the potential of targeting AhR as one of the prevention strategies for breast cancer. Supported by RR03032 15, CA91408-01 and DAMD17-02-01-0483 grants. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4077.

Although the overall breast cancer incidence rate for African American (AA) is lower than Caucasian (CA) women, among AA women under the age of 50 the rates are twofold higher. Due to their roles in activation of environmental poly aromatic hydrocarbons (PAH) to ultimate carcinogens, the products of CYP1A1 and CYP1B1 genes are implicated as risk factors for breast cancer. The objective of this study is to examine the hypothesis that higher incidence of breast cancer in pre-menopausal AA women is related to a dysregulated expression of CYP1A1 & CYP1B1 genes. To address this hypothesis our laboratory have isolated and characterized primary cultures of mammary epithelial cells (HMEC) from age-matched pre-menopausal AA and CA women, using normal breast tissues from mammoplastic reduction surgery. These isolated HMEC were used as in vitro model system to examine the response of breast tissues to several PAH including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and benzo[a]pyrene (B[a]P). In this study, the expression of CYP1A1 & CYP1B1 was measured in HMEC of ten agematched women donors from each women group ranging in age from 16-47 years. Results from real time RT-PCR analyses showed that CYP1A1 mRNA is expressed at low levels in untreated HMEC samples from both women groups and treatment with either B(a)P or TCDD induced its expression by many folds ranging from 10 to over 400-fold. Western blot analysis of microsomal proteins from these HMEC showed substantial levels of constitutive CYP1B1 protein in HMEC from both AA and CA women, which was further induced by both TCDD and B[a]P. However, HMEC from AA women exhibited higher inducibility of CYP1B1 protein in response to dioxin than HMEC from CA women at each age-match. In contrast, the majority of HMEC from AA women fail to induce CYP1A1 protein in response to dioxin treatment. These findings, although preliminary and require confirmation in larger number of donors, point to the higher risk of these AA women. This is in light of the fact that CYP1B1 more than CYP1A1 is responsible for bioactivation of chemicals, including estrogens into DNA-damaging carcinogens. The study will contribute to a better understanding of differential role of environmental chemicals on the biology of breast cancer in this ethnic minority group.