Background - With a few exceptions, the number of new breast cancer cases among women is increasing in almost all Western countries. Although late age at first child birth and genetics are shown to contribute to the increase in breast cancer, the sheer number of newly diagnosed cases cannot solely be explained by these factors. Do environmental influences, including exposure to mixtures of chemicals, also play a role?
Hypothesis - Considering that natural estrogens and man-made estrogens used as pharmaceuticals contribute to breast cancer, concerns arise about the potential role of industrial chemicals and pesticides with hormonal activity. Such chemicals include several that have been banned already, but can still be found in human tissues, such as polychlorinated biphenyls (PCBs) and DDT. A large number of chemicals currently used in consumer products also fall in this category (bisphenol A, UV-filter substances and many more). The studies carried out to date to examine whether certain environmental chemicals are implicated in breast cancer leave much uncertainty about a possible link. But to avoid wrongly dismissing a role for chemicals in breast cancer, the issue of simultaneous exposure to scores of chemicals must be addressed. However, the majority of published epidemiological studies have focused on single chemicals. Methods of dealing with mixture exposure in epidemiology are poorly developed.
Methods/Work performed - The involvement of weakly estrogenic chemicals in breast cancer is often dismissed on two counts: First, it is widely held that their potency is insufficient to contribute significantly to the effects of endogenous steroidal estrogens. Second, it is seen as unlikely that chemicals present at low levels can act together to produce joint effects. Both these ideas were tested experimentally by conducting multi-component mixture studies with combinations of estrogenic chemicals in cell-based assays.
Results and conclusions - Experiments with in vitro estrogenicity assays (yeast estrogen screen, E-Screen) have shown that the combined effects of multi-component mixtures of xenostrogens could be predicted well from information about their individual potency. When the individual chemicals were combined at concentrations below the sensitivity limits of these assays, significant mixture effects were observed. The effects of steroidal estrogens could be exacerbated by co-exposure to low levels of weakly active estrogenic chemicals. These results show that a potential role of such chemicals in breast cancer cannot be dismissed solely on the basis of their low potency. If exposure is to sufficient numbers of weak xenoestrogens, at low levels, significant modulations of the effects of steroidal estrogens may occur. The implications of these findings for epidemiology and risk assessments will be discussed.
The mammary epithelium develops in an ongoing, hormonally driven process that occurs throughout reproductive life. At the onset of puberty, epithelial invasion into the mammary fat pad accelerates dramatically and over the next seven weeks the epithelial network expands dramatically in size and complexity. The mammary epithelium continues to remodel during normal hormonal cycles and during each pregnancy, lactation, and involution cycle. In addition, the mammary gland is capable of extensive regeneration, as isolated fragments of epithelium, or even a single mammary stem cell can regenerate a complete epithelial network with all functional lineages are capable of regenerating.. During puberty, terminal end buds (TEBs) form transiently at the end of each primary duct. TEBs are characteristically different from quiescent ducts, with multiple luminal cell layers enriched in mammary stem cells that increase during puberty and high levels of proliferation. The multilayered TEB gives rise to bilayered ducts, with myoepithelial cells surrounding luminal epithelial cells. Using confocal time-lapse imaging, we showed that mammary epithelium first reorganizes into a multilayered, highly proliferative, incompletely polarized state. New ducts initiate from these preinvasive structures in a Rac- and myosin light chain kinase-dependent fashion. Cells in elongating ducts dynamically rearrange and lack leading cellular extensions. We also found that mammary branching morphogenesis results from the coordinate motility of two different cell types: luminal and myoepithelial cells. They move differently, and the myoepithelial cells appeared to restrain advancing luminal ducts. Importantly, organotypic cultures from normal human breast show the same behavior, indicating that the basic principles underlying breast development are used in both mouse and humans. Because the organization of normal terminal end buds, mammary hyperplasias, and ducts in organotypic culture are similar, suggesting that common cellular mechanisms may underlie normal pubertal development and neoplastic epithelial morphogenesis.
Supported by funds from the National Institute of Environmental Heath Sciences and the National Cancer Institute (ES012801 and CA057621)
Breast cancer is a manifestation of abnormal genetic as well as epigenetic changes. Global hypomethylation, accompanied by promoter hypermethylation, is a common feature of breast tumor cells. Global hypomethylation is thought to induce chromosomal instability, reactivate transposons, promote loss of imprinting, and activate proto-oncogenes. Promoter hypermethylation, on the other hand, appears to be associated with inactivation of genes in virtually all pathways protective of carcinogenesis (e.g. DNA repair, cell cycle control, inflammatory/stress response, detoxification, apoptosis, etc.) including those of breast cancer.
One-carbon metabolism facilitates the cross-talk between genetic and epigenetic processes by playing critical roles in both DNA methylation and DNA synthesis. It provides essential cofactors in the production of primary methyl donors for methylation of DNA, RNA and protein, as well as of dUMP to dTMP in DNA synthesis. A low methyl supply induces DNA global hypomethylation as well as deficient methylation of dUMP to dTMP leading to uracil misincorporation. These processes may result in aberrant DNA repair leading to DNA strand breaks, enhanced mutagenesis and apoptosis. It is important to note that methylation patterns are subject to clonal transmission, and as a result disturbances in one-carbon metabolism can have long-term consequences even after one-carbon metabolism has returned to normal.
This talk will focus on our systematic investigation of the role of one-carbon metabolism in breast cancer etiology and survival. Using a population-based case-control study, Long Island Breast Cancer Study Project, we have examined the inter-relationships among dietary methyl intake, functional polymorphisms related to one-carbon metabolism, and methylation status in tumors. We hope to better elucidate the role of epigenetics in breast cancer development.
Acknowledgement: This work was supported by grants NIH CA109753, DOD BC031746, DOD W81XWH-06-1-0298.
Epidemiological studies suggest that both genetic and environmental factors influence the incidence of sporadic breast cancer in women in different geographic areas and of different ethnicities. However, positive identification of these factors and determination of their functional contributions to cancer etiology have proven difficult. Carcinogenesis involves the sequential acquisition of a complex set of phenotypes over extended time periods. The ability to assay the effects of suspect environmental and genetic factors on presumptive target cells using surrogate endpoints corresponding to the acquired phenotypes aids in the positive identification of such factors. Determining causality is best performed in cultured cells, where complicating variables can be monitored and controlled, and cells can be monitored in real time. Although such culture systems are primitive in comparison to intact tissue, they can be used to model certain aspects of tumor progression. To define molecular events and cellular characteristics that allow tumorigenic progression to proceed in the human breast, others and we have used human mammary epithelial cell (HMEC) cultures derived from surgically discarded phenotypically normal tissue. A notable difference between normal and tumor breast tissue is the ability of the latter to give rise to cells that can proliferate indefinitely in culture. Such “immortality” is thought to be a key predisposing factor for carcinogenesis because it allows the progeny of a single cell to sequentially accumulate the multiple errors needed to gain invasive and metastatic properties. Studies of HMEC and other cell types have led to the hypothesis that human cells have developed extremely stringent mechanisms to prevent immortality, and that malignant precursors must evade this protective gauntlet to give rise to frank malignancies. The p16 tumor suppressor protein is one “sentinel” that responds to potentially oncogenic microenvironmental stresses by causing growth arrest. In rare cases, however, the gene that encodes p16 can be shut down permanently or mutated, allowing HMEC to grow for extended periods and acquire potentially malignant changes. We have assessed the ability of radiation - a known breast carcinogen, to affect evasion of p16-mediated growth arrest by cultured HMEC, and have found that moderate doses of radiation can increase this pre-malignant aberration. This is one example of the use of a simple experimental system employing cultured HMEC to test a potentially carcinogenic function of genetic and environmental factors.
Supported by the UCSF Breast Cancer and the Environment Research Center and a NASA Specialized Center of Research (NSCOR).
Progesterone promotes cell proliferation and development of the normal mammary gland and is implicated in the etiology of breast cancer. The specific functions of progesterone receptor isoforms PRA and PRB in the normal mammary gland are critical to understanding the molecular mechanisms of progesterone action in these processes. PRA is most highly expressed during puberty, an important widow of exposure sensitivity that can increase or decrease breast cancer risk later in life. While PRB is essential for glandular development in pregnancy, the functions of PRA have not been elucidated. To identify PRA-regulated genes, mammary epithelial cells were obtained from pubertal or adult mouse mammary glands and cultured as “organoids” in a 3-D culture system. Progestin (P) treatment causes organoids to develop a cellular organization similar to ducts, mimicking in vivo ductal development. P-induced gene expression was analyzed with whole mouse genome microarrays. We identified 69 P-regulated genes in both pubertal and adult epithelium, 38 genes regulated uniquely in adult epithelium, and 96 genes regulated uniquely in pubertal epithelium. In both pubertal and adult epithelium, inflammation-related genes were prominently upregulated. Many of these genes are not previously known to be P-regulated. Additionally, genes involved in programmed cell death show pubertal upregulation, while those involved in cell adhesion show pubertal and adult upregulation, perhaps reflecting progesterone’s role in organ development. Among the inflammatory genes most dramatically upregulated were serum amyloid A1, A2, and A3. We confirmed P-induced serum amyloid A1 protein expression in organoids. Serum amyloid A proteins are implicated in the induction of inflammation, and are reported to increase expression of proinflammatory factors in white blood cells, and to play a role in recruitment and adhesion of these cells to sites of infection and injury. Supporting the notion that progestins can induce an inflammatory state, we found that mammary glands of ovariectomized mice show increased infiltration of white blood cells upon 5-day progesterone treatment. Inflammatory cells are implicated in normal mammary gland development and the inflammatory state is implicated in tumor progression in many studies. Progestin induction of inflammatory genes suggests a novel role for progestins in mammary gland development and function, and presents a possible link between progestins and inflammation in the etiology of breast cancer. An inflammatory mechanism may underlie the increased breast cancer risk associated with hormone replacement therapy that includes progestins.
This work is supported by the Breast Cancer and the Environment Research Center: NIH/NIEHS/NCI U01 ES12800.
Our work has been designed for testing the hypothesis that estrogenically active environmental chemicals influence the lifetime risk of developing cancer when exposures occur during critical early periods of breast development. For accomplishing this objective Sprague-Dawley rats were used for testing the effects of prenatal and prepubertal exposures to a low (LD) and a high dose (HD) of the hormonally-active compounds bisphenol A (BPA), butyl benzyl phthalate (BBP), and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the genomic and proteomic signatures of mammary glands of 21, 35, 50 or 100 day-old rats. Changes in endocrinological parameters and mammary gland morphology, cell proliferation, gene and protein expression at these ages, which represent critical stages of development and differentiation, are correlated with the response of the mammary gland to chemical carcinogens. In addition, proteins identified in the serum collected from rats at these various stages serve as biomarkers of exposure and risk; correlation of their presence with those found in the serum of a cohort of girls traversing similar windows of development constitutes the basis for prospectively investigating the relationship between hormonally active exposures and pubertal milestones. In rats prenatally or prepubertally-exposed to LD or HD BPA or BBP none of the following parameters were affected: body weight, age at vaginal opening, uterine weight, and estrous cyclicity. Prepubertal HD TCDD exposure, on the other hand, significantly decreased body weight at 21, 35, and 50, but not at 100 days of age, and both, LD and HD TCDD decreased uterine weight at day 35, delayed vaginal opening, and induced irregular estrous cycles. At genomic level, on the other hand, prepubertal exposure to HD BPA resulted in a greater number of dysregulated genes and a greater number of chemically-induced mammary carcinomas than prenatal exposure, an indication that prepuberty is a period of greater sensitivity to the effect of BPA for the mammary gland to undergo specific genomic changes that determine its susceptibility to carcinogenesis. Although these effects were not elicited by the other compounds, the genomic changes induced in the rat mammary gland by BPA, BBP, and TCDD exposure identified a specific set of genes relevant to humans and led us to conclude that each compound at each specific window of exposure selectively affects defined biological processes, cellular components, and molecular functions. In addition, in vitro treatment of human breast epithelial cells with BPA and BBP induces cell transformation and changes in genes dysregulated in animals treated with these compounds in vivo and that exhibit single nucleotide polymorphism (SNP) in the buccal mucosa of the cohort of girls included in these studies. These results have provided new sets of genomic markers and new paradigms for the understanding of the role of environmental exposures in the human population cancer risk.
(This work was supported by NCI and NIEHS Grant UO1 ES012771).
Bisphenol A (BPA) is found as a polymer in food containers such as baby bottles, lining of food and soft drink cans, designer drinking bottles, office water cooler bottles, dental sealents, etc. It is even found in children’s toys. BPA has been measured in greater than 90% of urine of young girls. Our goal is to determine if early exposure to BPA can alter puberty and the ontogeny of mammary protein expression as the offspring matures and if it predisposes for mammary cancer development. Rats were exposed during pregnancy only or during lactation only to 0, 25 and 250 µg BPA/Kg BW orally.
Prenatal only and prepubertal only exposures to BPA did not alter vaginal opening (a marker of puberty). Mammary glands were investigated for protein expression using 2-D gel separation and mass spectrometry (MALDI-TOF-TOF) identification. In offspring of rats exposed prenatally to BPA, 23, 21 and 16 proteins were identified as differentially regulated at 21, 50 and100 days, respectively. Included in these are 14-3-3 proteins and SPARC (secreted protein acidic and rich in cysteine), proteins that are associated with cell signaling, and mammary and cancer development.
In mammary glands of 50 day old rats, but not in 21 day old rats, exposed prepubertally to BPA, we found increased rate of cell proliferation and decreased apoptosis. Using western blot analysis, steroid receptor co-activators 1-3, Akt, phospho-Akt, progesterone receptor A, and erbB3 proteins were determined to be significantly up-regulated at 50 days. In adult rats exposed prepubertally to BPA and then to the carcinogen, dimethylbenzanthracene, we found a dose dependent increase in tumor multiplicity and decreased latency compared to controls.
We conclude that at very low doses, prenatal and postnatal BPA exposures do not significantly alter puberty in rats, but do alter key protein expressions in the mammary of developing rats. Furthermore, we demonstrate that, at a minimum, prepubertal BPA exposure predisposes for increased susceptibility to chemically-induced mammary cancer in rats. If these effects found in rodents carry over to humans, even small, seemingly harmless exposures of BPA early in life could play a role in increased breast cancer susceptibility later in life.
(Supported by NCI/NIEHS Grant UO1 ES012771).
Environmental exposures may alter the timing of breast development, a risk factor for reproductive function, breast cancer, and other chronic diseases. Mount Sinai is one of three Breast Cancer and Environment Research Centers (BCERCs) sites investigating the role of environmental exposures in puberty. We are studying inner city minority girls who are at risk for obesity and early development. We have established a cohort of 412 girls (60% Hispanic, 40% black; 6-8 years old at baseline [BL]). At first annual followup (FU1) we have seen 296 girls as of September 10, 2008. Breast stage was assessed by health providers (BL, FU1); dietary isoflavone and energy intake, urinary biomarkers of phthalates, phenols, and phytoestrogens were measured at BL. Breast development (stage B2+ vs B1) was present in 22% of girls at BL, and in 35% at FU1. Many girls were at-risk-for- or were overweight (40% at BL). Baseline isoflavone and energy intakes were lower among girls who were B2+ vs B1 at BL as well as at FU1 (not significant). Baseline low-molecular-weight phthalate urinary metabolites (LMWPH) were higher among girls who were B2+ vs B1 at FU1; however they did not differ by BL breast stage (unadjusted). We calculated adjusted prevalence ratios (PR) and hazards ratios (HR) for cross-sectional and longitudinal observations. Age- and ethnicity- adjusted prevalence ratios (PR) for B2+ at FU1 were 1.14 per log-LMWPH (95%-CI 0.996-1.30) measured at BL. Other urinary biomarkers had near-null PRs for breast stage at BL or FU1 (benzophenone, bis-phenol A, 2,5-dichlorophenol, triclosan, genistein, daidzein, enterolactone, high-molecular-weight phthalate monoesters). Interaction models indicate that prevalance of breast stage may vary by adiposity for several biomarkers including phthalate metabolites, benzophenone, bisphenol A, and enterolactone. Similarly, HR models predicting age at B2 showed few significant main effects, but there was increased risk for one biomarker among girls with above-median-BMI and above-median-exposure biomarkers at BL. in summary, preliminary results from this longitudinal study suggest few main associations of hormonal environmental exposures with breast development, although effects may be observed among underweight or overweight girls. Further analyses with complete followup may provide additional insights. Disclaimer: The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
This research was supported by grants from NIEHS (ES009584 and ES012645), EPA (R827039 and RD831711), ATSDR (ATU 300014), NCI (CA93447), and NCRR MO1-RR-00071. We thank Charles Dodson for his valuable support of this project.
Onset of pubertal maturation in girls consists of a series of inter-related events, including reactivation of the hypothalamic-pituitary-ovarian axis, adrenarche (activation of the adrenal cortex), gonadarche (marked by the appearance of breast tissue), and pubarche (marked by the appearance of pubic hair). Researchers and clinicians have suggested several different approaches to mark and measure onset and progression of puberty, including breast development alone, onset of breast or pubic hair development (whichever is earlier), an increase in height velocity, bone age or dental age (with exposure to ionizing radiation), and menarche (a relatively late finding in puberty). There are very few studies in the scientific literature examining serial measurements of sex hormones in girls.
The Cincinnati site of the NIH-funded Breast Cancer and the Environment Research Centers (BCERC) recruited 379 girls, age 6-7 years at entry, and followed these girls every six months with assessment of multiple measures by trained examiners. These assessments included (among others) height, weight, maturation, biannual blood specimens for hormone analysis, and annual urine and blood specimens for biomarker analysis.
This presentation will discuss the results of 175 girls who entered puberty during the first three years of the project. Longitudinal comparisons of growth velocity, breast as well as pubic hair stage, and sex hormones will be presented. The sex hormones analyzed include the female hormones estradiol and estrone; the male hormone testosterone; the adrenal hormone DHEA-S; and sex hormone binding globulin (SHBG). Derived variables will include ratio of estradiol to testosterone, and free androgen index (FAI, [testosterone concentration] / SHBG).
Perfluorooctanoic acid (PFOA) is a member of a family of synthetic perfluorinated compounds with extremely high stabilities. PFOA has been widely used in the production of numerous industrial and consumer products (flame retardants, water and oil repellent coatings for fabrics and food packaging, paint additives, electrical insulation and many others). PFOA is one of the most common persistent organic pollutants in the environment. Biomonitoring studies have shown that PFOA and other perfluorinated chemicals are found as global pollutants in air, water and soil, and also are detected in blood samples from almost all organisms sampled including human populations. The Breast Cancer and Environment Research Centers Epidemiology Project identified a subgroup of girls in the University of Cincinnati cohort who had above average concentrations of PFOA in their serum. The peripubertal period in females is thought to be an important window of breast susceptibility to environmental exposures that may affect breast cancer risk later in life. The effects of PFOA in pubertal girls is not known.
We have examined the effect of PFOA exposure in female mice during the peripubertal period (21 through 50 days of age). The effects of PFOA (doses 0.1 – 10 mg/kg BW) were examined in two strains of mice (Balb/c and C57BL/6) which are known to have genetically determined differences in mammary gland development and responses to hormones. PFOA treatment caused: 1) pathological changes in the liver, 2) delayed onset of puberty, 3) changes in uterine development, and 4) alterations in mammary gland development. Changes in the liver and delayed onset of puberty were similar in the 2 mouse strains. However, while Balb/c mice exhibited only inhibition of mammary gland and uterine development (5, 10 mg/kg doses), C57BL/6 mice exhibited stimulatory effects in both organs at low dose (5 mg/kg) and inhibition at higher dose (10 mg/kg).
Summary: These results demonstrate in mice that peripubertal exposure to PFOA has detrimental effects in the liver, uterus, onset of puberty and mammary gland development. However, there were significant differences in the effects based on genetic background. This underscores the need for caution when drawing conclusions about the effects of PFOA and possibly other environmental pollutants on the basis of studies in a single mouse strain. This is particularly relevant for identifying the effects of PFOA in the genetically heterogeneous human population.
This study was supported by the Breast Cancer and the Environment Research Center Grant 1-UO1 ESO12800 01 from National Institute of Environmental Health Sciences
Background: Polyfluoroalkyl compounds (PFCs) and their salts, perfluorooctanoate (PFOA) and perfluorooctane sulfonate, have been reported to change mammary gland structure and function in laboratory animals.
Objective: To determine the relationship between serum PFOA concentration and timing of pubertal maturation in young girls.
Methods: Within the NIH Breast Cancer and the Environment Research Centers (BCERC), we conducted a study of multiple environmental biomarkers in young girls (age 6-7 years at entry), including PFCs. Participants for the study were recruited from area schools in greater Cincinnati, and through members of the Breast Cancer Registry of Greater Cincinnati. Pubertal staging was by conducted by clinicians or trained research staff. Girls have been staged for pubertal maturation every six months, for as long as four years. Blood was collected using a standard protocol and materials provided by the Centers for Disease Control and Prevention (CDC), and analyzed for the PFCs using online solid phase extraction-high-performance liquid chromatography-tandem mass spectrometry. Of the 379 girls in the study, at the time of these statistical analyses we had PFC measures on 266, and of that group, 81 had reached Stage 2 or greater of breast development and 38 had reached Stage 2 of pubic hair appearance. We examined the relationship between PFOA serum concentration, measured at the beginning of the study, with the age at which the girl reached Stage 2 of pubertal maturation. Age, race and BMI percentile were included as covariates in all models. PFOA serum concentration was modeled as both categorical and log-transformed continuous. We also examined the relationship between PFOA serum concentration and BMI percentile, and with serum triglyceride and cholesterol levels.
Results: Detectable serum levels of PFOA were found in all but one of the 266 serum samples. The median was 10.24 ng/ml (range <LOD 0.01 to 55.9 ng/ml), somewhat higher than the median for children 12-19 years age in the NHANES 2003-2004 population (3.9 ng/ml). Only 58.3% of our study participants had serum concentration values below the National Health and Nutrition Examination Survey (NHANES) population 95th percentile value (8.6 ng/ml), and 55 girls (20,7%) had values >11.3 ng/ml. We will present our findings of studies of the relationship between PFOA serum concentration and pubertal maturation, BMI percentile and serum triglyceride and cholesterol levels.
Support for this project provided by the National Institute of Environmental Health Sciences and the National Cancer Institute, to the University of Cincinnati/Cincinnati Children’s Hospital Medical Center, Breast Cancer & the Environment Research Center (U01 ES12770), and Center for Environmental Genetics (P30-ES06096).
The findings and conclusions in this presentation have not been formally disseminated by the Centers for Disease Control and Prevention and should not be construed to represent any agency determination or policy.
The arrival of the 21st century has seen a transition in the conduct of science. The 60 years of NIH-funded investigator-driven experimentation, necessary to break down the complex nature of disease into bite-sized pieces, now creates a platform to perform integrative science. A major step in this new direction was the sequencing of the human and other model organism genomes. This has spawned many new –Omics, transcriptomics, proteomics and metabolomics, each re-invention of older disciplines. However, this technological improvement has not been accompanied by changes in experimental design or in the development of stable statistical measures of the quality of the data acquired in -Omics experiments. In order to extract meaningful information from –Omics experiments, it is first essential to remove all aspects of bias. This requires involvement of statisticians with experience in laboratory science to determine the sources of variation (the order of obtaining animal tissues, distribution of stored samples, order of carrying out analyses, including labeling with Cy-dye or iTRAQ reagents) and to design experimental approaches that balance variation out across experimental groups. Omics approaches should also ensure that the number of experimental replicates is adequate for providing statistical significance. All experiments contain false discovery. In a two-group experiment with one variable, we accept that a 5% chance or lower that the two groups are the same is sufficient to reject the null hypothesis. In –Omics experiments, often with thousands of variables, this assumption is not true. Under the null hypothesis, the expected p-values are uniformally distributed between 0 and 1. Therefore, the false discovery rate (FDR) is predictable. If the number of true changes can be estimated and a particular FDR accepted, the statistician can determine the p-value for an experiment. With this insight, the number of replicates in an experiment can be proposed. In two independent studies carried out by CNGI investigators on changes in gene expression in the mammary gland between days 21 and 50 involving group sizes of 8 animals, microarray analysis determined there were 4982 genes with an FDR of <1% in experiment 1; in experiment 2, 3032 of these genes have a FDR <1%. A high correlation in fold changes was observed, indicating that meaningful interpretations could be made of these data. Pathway analysis revealed that the changes were concentrated in energetics and metabolism, suggesting that the onset of puberty in the mammary gland represents alterations in master switches under transcriptional control.
A principal component of the Breast Cancer and the Environment Research Centers (BCERCs) are the conduct of prospective epidemiologic studies investigating predictors of early puberty, including breast development, in young girls. While there is substantial uncertainty regarding the causes of early puberty, it is known that there are racial differences, and that overweight and obesity predispose to earlier pubertal onset. Dietary factors are of interest as they may be a source of exogenous estrogens (e.g., phytoestrogens), and they may influence sex hormone metabolism and adiposity. In the BCERC epidemiologic studies, over 1,200 girls aged 6-8 years at baseline were enrolled in centers at three sites: the San Francisco Bay Area, Cincinnati, and New York City. Enrollment into these studies occurred from 2004 through 2007, and girls continue to be followed with annual or semi-annual contacts. Pubertal status is determined through clinical exams assessing Tanner stage for breast or pubic hair development. Among other factors, dietary data was collected during the baseline year using four 24-hour telephone recalls, each recall spaced approximately 3 months apart. Data have recently become available for baseline data collection and subsequent clinical exam visits that include anthropometry and Tanner stage assessment. Preliminary analyses of data from the San Francisco Bay Area suggest phytoestrogen intake may be associated with delayed onset of puberty, but this observation needs to be confirmed with more rigorous data analysis and inclusion of data from Cincinnati and New York. Results from these analyses will be presented, along with a broader overview of dietary factors in breast cancer and the role of the BCERCs in examining the role of diet and other environmental factors in the development of breast cancer.
Supported by the UCSF Breast Cancer and the Environment Research Center.
Supported by the Breast Cancer and the Environment Research Centers grant number U01 ES012800 from the National Institute of Environmental Health Sciences and the National Cancer Institute.
The prevalence of obesity among pre-puberty females in the United States has grown to alarming levels in recent years, especially among lower income and minority populations. Because obesity among girls is a key risk factor for later breast cancer, it’s important to address the determinants of obesity and identify approaches for preventing this condition.
The food industry plays an enormous role in influencing the eating habits of young people, including pre-adolescent girls. For several decades, increasing concerns have been raised about the potentially harmful impact of food advertising and marketing practices directed to children and youth. These younger age groups are considered vulnerable because they are unsophisticated consumers who are tempted by enticing TV commercials and tasty products marketed in grocery stores and child-friendly restaurants. Research has shown that these sales practices lead to greater consumption of high-fat high-calorie foods, notably sugary cereals, candy, snack items, soft drinks, and fried fast foods. Traditionally, the food industry has seldom promoted nutritious foods young people; there are few messages that effectively promote foods that may serve as protective factors for breast cancer.
A variety of government agencies, health organizations, and community groups have undertaken extensive prevention campaigns to educate children and their parents about proper eating patterns and effective practices for managing weight. These initiatives have attained only limited success; obesity rates remain stubbornly persistent and solving the problem requires more elaborate forms of intervention than the modest flow of messages that are primarily targeted at the individual level.
To achieve meaningful progress, a comprehensive strategy is needed to leverage and combat the multi-faceted influences in the broader environment. A promising approach focuses on impelling systems-level reforms that will facilitate nutritious choices and restrict glamorization of unhealthy foods. Recently, the first meaningful steps have been undertaken to regulate advertising and marketing, with participation by government agencies, schools, cable and network television, and leading companies in the food industry. This control-oriented approach for reducing unhealthy diets is complemented by positive efforts to promote healthy food options; pro-nutrition programs are aimed at both young people and their parents, who mutually determine eating choices.
The presentation reviews emerging reforms and specifies significant issues that remain to be addressed. An array of advocacy groups is involved in consolidating the tentative progress being made and extending these initiatives in new directions, and the potential role of breast cancer advocates will be discussed.
Women in the United States have among the highest breast cancer rates in the world. This is not due to increased genetic susceptibility, as less than ten percent of breast cancers are attributed to gene mutations such as BRCA1. This observation suggests that environmental factors including nutrition and lifestyle may be major contributors for the tumorigenic process. We have developed a rat model to examine tumor progression and to identify risk factors which develop during puberty and/or during gestation as a result of environmental insults. Specifically we have explored the effects of dietary fatty acids including olive, or safflower oil or butter fat on tumor progression. When rats were exposed to these dietary agents during gestation, and throughout their lifespan (110 days), a significant increase in mammary tumor progression compared to the AIN control diet were observed when a single low dose of DMBA was administered at 21 days. There was an increase in average tumor volume per animal and the progression of CIS to invasive tumors. A significant feature of this model is the range in size of the CIS lesions. Also, the number of invasive tumor was relatively small, which is similar to the human progression of CIS lesions to invasive tumors. We have collected invasive tumors, and large and small CIS lesions for microarray expression analysis. The dietary agents (olive oil and butter fat) were clearly acting as tumor promoters in this model. We examined the effects of these dietary agents on epithelial cells during mammary gland development in the puberty phase. It was clear that these agents not only increased the ongoing cell proliferation during puberty, but also activated the checkpoint pathways suggesting that the exposures to these diets are causing DNA damage. This is one aspect of the tumor promotional properties of these dietary agents. Another study is related to epigenetic changes that may occur during gestation when the dams are exposed to these dietary agents. If epigenetic changes are observed in the pups compared to the AIN diet, then we will determine if these changes predispose these pups to tumorigenesis when a small dose of DMBA is administered during puberty or later in life. This model could be used to examine the effect of various environmental insults on mammary tumorigenesis in young pups or mature animals and mechanisms of tumor progression can also be examined.
Obesity is implicated in breast cancer risk and progression and is associated with poor prognosis in both pre- and post-menopausal incidence of the disease. In particular, the risk of developing breast cancer increases with higher body mass index (BMI) in post-menopausal women. Industrial societies, such as the US, have moved to consumption of diets enriched in fat leading to a positive energy balance, making obesity a national crisis. Significant amounts of saturated fats in the diet have been replaced by polyunsaturated fats (PUFA), particularly the n-6 fatty acids of vegetable oils to decrease cardiovascular risk factors. We sought to study the influence(s) of specific dietary fatty acids on the development of mammary tumors in female rats. Female rats exposed to the diets in utero were dosed with a mammary carcinogen: dimethylbenz[a]anthracene (DMBA) at weaning (DOW) and at puberty (Day of Life DOL 50) and followed for 3 months to monitor tumor development. The diets contained either 10% or 40% fat (% kcal) of one of these fatty acids: olive oil, safflower oil or butter fat. For those rats dosed at DOW, there were statistically (P<0.05) greater number of carcinoma in situ (CIS) and invasive carcinoma (INV) lesions present in rats consuming the 40% olive oil (HFO) and 40% Butter (HFB) diets compared to the control diet. The tumor latency (time to first lesion detected) was shorter (43.4 days) for rats consuming the HFO diet compared to those rats consuming other diets. Additionally 90% of rats on the HFO diet dosed with DMBA at DOW developed more than 2 mammary lesions compared to 75% or less for other diets (45% for AIN). Our findings suggest that HFO and HFB may enhance susceptibility to carcinogenesis through their interaction with obesity. Obesity facilitates the production and availability of numerous mitogenic agents, (including estrogen and IGF-1), immune modulators (including leptin and IL-6), and angiogenic factors (including vascular endothelial growth factor and hepatocyte growth factor) to susceptible target cells throughout the body. Additionally, obesity is associated with increased inflammation and inflammatory markers such as NF-kB and its regulated genes which are involved in invasiveness, proliferation, angiogenesis, and inflammation. Therefore, we will present data suggesting HFO and HFB increase carcinogenesis by influencing obesity and inflammation. These findings begin to suggest mechanisms by which nutrients can influence mammary carcinogenesis.