Resveratrol, but not EGCG, in the diet suppresses DMBA-induced mammary cancer in rats

Despite the advent of new and aggressive therapeutics, breast cancer remains a leading killer among women; hence there is a need for the prevention of this disease. Several naturally occurring polyphenols have received much attention for their health benefits, including anti-carcinogenic properties. Two of these are resveratrol, a component of red grapes, and epigallocatechin-3-gallate (EGCG), the major catechin found in green tea. In this study, we tested the hypothesis that these two polyphenols protect against chemically-induced mammary cancer by modulating mammary gland architecture, cell proliferation, and apoptosis. Female Sprague-Dawley CD rats were exposed to either resveratrol (1 g/kg AIN-76A diet), EGCG (0.065% in the drinking water), or control diet (AIN-76A) for the entirety of their life starting at birth. At 50 days postpartum, rats were treated with 60 mg dimethylbenz[a]anthracene (DMBA)/kg body weight to induce mammary cancer. Resveratrol, but not EGCG, suppressed mammary carcinogenesis (fewer tumors per rat and longer tumor latency). Analysis of mammary whole mounts from 50-day-old rats revealed that resveratrol, but not EGCG, treatment resulted in more differentiated lobular structures. Bromodeoxyuridine (BrdU) incorporation studies showed that resveratrol treatment caused a significant reduction in proliferative cells in mammary terminal ductal structures at 50 days postpartum, making them less susceptible to carcinogen insult. The epithelial cells of terminal end buds in the mammary glands of resveratrol-treated rats also showed an increase in apoptotic cells compared to the control or EGCG-treated rats as measured by a DNA fragmentation assay. At the given doses, resveratrol treatment resulted in a serum resveratrol concentration of 2.00 μM, while treatment with EGCG resulted in a serum EGCG concentration of 31.06 nM. 17β-Estradiol, progesterone, and prolactin concentrations in the serum were not significantly affected by resveratrol or EGCG. Neither polyphenol treatment resulted in toxicity as tested by alterations in body weights, diet and drink consumptions, and day to vaginal opening. We conclude that resveratrol in the diet can reduce susceptibility to mammary cancer, while EGCG in the drinking water at the dose used was not effective

Serum profiling and biomarker discovery of rat mammary tumors using mass-coded abundance tags (MCAT)

Advances in Mass-spectrometry techniques allow for the rapid processing and evaluation of complex biological mixtures such as blood/serum. These samples represent a protein rich environment as well as a sentinel monitoring system of the entire organism. The central tenet of these studies is that changes in the microenvironment of a tissue, brought about by a disease process, will lead to sufficient changes in the protein and peptide pattern of the serum, such that the differences can be accurately detected and correctly associated with a particular disease state. Using mass-spectrometry approaches we have developed techniques that allow us to compare samples from tumor-free and tumor present serum samples simultaneously to find biomarkers that indicate the presence of cancer. To examine potentially important but less abundant proteins, ultrafiltration (UF) was used to eliminate the more abundant proteins and combine this with the non-isotopic peptide tags (S-methylthioacetimidate and S-methyl thiopropionimidate) described by Beardsley and Reilley (J. Proteome Res. 2: 15-21, 2003) to differentiate our samples. Use of these mass-coded abundance tags (MCAT) allows for simultaneous evaluation of serum samples from tumor present, and tumor free animals. Using an oa time-of-flight mass-spectrometer (Q-tof) with electrospray ionization we produce high quality spectrums to screen for peptides that have only one tag. Specificity of tagging increases the likelihood that the peptide resulted from a protein unique to either the control or conditioned state. Using the ms/ms function of the Q-tof we sequence the peptide and identify the parent protein. Specifically, our lab is using UF, MCAT and the Q-tof to evaluate rat models of chemically-induced tumors. By using animal models we overcome much of the variability that may exist in human serum samples due to differences in gender, diet and cancer initiation. We have shown that these systems allow for the identification of both small molecules such as Alpha S1 casein precursor (24 kDa) as well as proteins greater than the MCO such as Fibrinogen alpha/alpha E precursor and Coagulation factor 2 (86 and 70 kDa, respectively). With positive sequence identification we can now evaluate the tumors themselves to determine if the proteins are over-expressed in the tumor vs. normal tissues. Using this method of “bottom-up” analysis provides information on the nature and composition of our samples to more rapidly identify those proteins that are unique to the tumor state of the animals

Genistein chemoprevention of prostate cancer in TRAMP mice

Epidemiological studies suggest an inverse association between soy intake and prostate cancer risk. Genistein, the predominant phytoestrogen in soy food, has been proposed as a potential chemopreventive agent due to its anti-estrogen and tyrosine kinase inhibitory effects. To determine the most effective period for genistein chemoprevention, the Transgenic adenocarcinoma mouse prostate (TRAMP) model was used. The treatments were 250 mg genistein/kg AIN-76A diet 1) prepubertally only, 2) in adulthood only or 3) through out life. Controls received AIN-76A diet. By 28 weeks of age, 100% TRAMP mice fed control diet developed prostatic intraepithelial neoplasia (PIN) or adenocarcinomas with 6%, 16%, 44% and 34% developing high grade PIN, well differentiated, moderately differentiated and poorly differentiated prostatic adenocarcinomas, respectively. Prepubertal only (1–35 days postpartum) and adult only genistein treatments (12 – 28 weeks) resulted in 6% and 29% decreases in poorly-differentiated cancerous lesions compared with controls, respectively. The most significant effect was seen in the TRAMP mice exposed to genistein throughout life (1–28 weeks) with a 50% decrease in poorly-differentiated cancerous lesions. In a separate experiment in castrated TRAMP mice, dietary genistein suppressed the development of advanced prostate cancer by 35% compared with controls. Of the tumors that developed in castrated TRAMP mice, 100% were poorly-differentiated in contrast to the 37% of noncastrated TRAMP mice that developed poorly-differentiated tumors. ICI 182,780 (ICI), genistein and estrogen down-regulated androgen receptor (AR), estrogen receptor alpha (ER-α) and progesterone receptor (PR) in the prostates of C57BL/6 mice, and act independently of ER. Our data obtained in intact and castrated transgenic mice suggest that genistein may be a promising chemopreventive agent against androgen-dependent and independent prostate cancers

The plasticizer butyl benzyl phthalate induces genomic changes in rat mammary gland after neonatal/prepubertal exposure

Background: Phthalate esters like n-butyl benzyl phthalate (BBP) are widely used plasticizers. BBP has shown endocrine-disrupting properties, thus having a potential effect on hormone-sensitive tissues. The aim of this study is to determine the effect of neonatal/prepubertal exposure (post-natal days 2-20) to BBP on maturation parameters and on the morphology, proliferative index and genomic signature of the rat mammary gland at different ages of development (21, 35, 50 and 100 days). Results: Here we show that exposure to BBP increased the uterine weight/body weight ratio at 21 days and decreased the body weight at time of vaginal opening. BBP did not induce significant changes on the morphology of the mammary gland, but increased proliferative index in terminal end buds at 35 days and in lobules 1 at several ages. Moreover, BBP had an effect on the genomic profile of the mammary gland mainly at the end of the exposure (21 days), becoming less prominent thereafter. By this age a significant number of genes related to proliferation and differentiation, communication and signal transduction were up-regulated in the glands of the exposed animals. Conclusion: These results suggest that BBP has an effect in the gene expression profile of the mammary gland

Early Exposure to Soy Isoflavones and Effects on Reproductive Health: A Review of Human and Animal Studies

Soy isoflavones are phytoestrogens with potential hormonal activity due to their similar chemical structure to 17-β-estradiol. The increasing availability of soy isoflavones throughout the food supply and through use of supplements has prompted extensive research on biological benefits to humans in chronic disease prevention and health maintenance. While much of this research has focused on adult populations, infants fed soy protein based infant formulas are exposed to substantial levels of soy isoflavones, even when compared to adult populations that consume a higher quantity of soy-based foods. Infant exposure, through soy formula, primarily occurs from birth to one year of life, a stage of development that is particularly sensitive to dietary and environmental compounds. This has led investigators to study the potential hormonal effects of soy isoflavones on later reproductive health outcomes. Such studies have included minimal human data with the large majority of studies using animal models. This review discusses key aspects of the current human and animal studies and identifies critical areas to be investigated as there is no clear consensus in this research field

Chronic Oral Exposure to Bisphenol A Results in a Nonmonotonic Dose Response in Mammary Carcinogenesis and Metastasis in MMTV-erbB2 Mice

Background: Bisphenol A (BPA) is a synthetic compound used to produce plastics and epoxy resins. BPA can leach from these products in appreciable amounts, resulting in nearly ubiquitous daily exposure to humans. Whether BPA is harmful to humans, especially when administered orally in concentrations relevant to humans, is a topic of debate