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¹ Department of Obstetrics and Gynecology, Catholic University of the Sacred Heart, Rome, Italy
² Department of Oncology, Catholic University of the Sacred Heart, 86100 Campobasso, Italy

(Requests for offprints should be addressed to G Scambia who is now at Department of Obstetrics and Gynecology, Catholic University of the Sacred Heart, Largo A Gemelli, 8-00168 Rome, Italy; Email: giovanni.scambia{at}rm.unicatt.it)

	Abstract

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The study reported here was designed to determine whether a phytoestrogen-containing soy extract (SSE) could negate/overwhelm the inhibitory effects of ICI 182 780 on the growth of estrogen-sustained human breast cancer xenografts (MCF-7), in ovariectomized athymic mice. As expected, estradiol-supplemented tumors did not grow over the study period in ICI 182 780-treated females; concomitant administration of 50 mg/kg per day SSE slightly potentiated the inhibitory activity of the drug, while at 100 mg/kg per day, SSE partially negated ICI 182 780 activity. In keeping with these in vivo outcomes, we observed that the level of cyclin D1 (and progesterone receptor) in MCF-7 xenografts was considerably reduced by ICI 182 780, an effect enhanced by concomitant treatment with 50 SSE, but reduced by the higher dosage (i.e. 100 mg/kg per day). Thrombospondin-1 (TSP-1) and kallikrein 6 (KLK6) levels were also reduced following ICI 182 780, although to a lesser degree; again, combined anti-estrogen and SSE produced a dose-dependent regulation in TSP-1 and KLK6 tumor level, with a further reduction in the mRNA gene expression at 50 SSE (compared with ICI 182 780) and a partial reversion of the drug-induced down-regulation at 100 mg/kg per day. No modulation was detected in the serum concentration of IGF-1 (a potent mitogen for estrogen receptor-positive breast cancer cell lines) either upon treatment with ICI 182 780 or concomitant administration of the anti-estrogen with SSE. In conclusion, results from this study raise concerns about the consumption of isoflavone supplements in conjunction with ICI 182 780 therapy, in postmenopausal women with estrogen-dependent breast cancer.

	Introduction

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Breast cancer is a major Public Health issue worldwide and among women aged 20–59 years it represents the leading cause of cancer death in the United States (Jemal et al. 2006). In postmenopausal women, endocrine therapy is a widely accepted treatment modality for hormone receptor-positive breast cancer, with endocrine agents designed to inhibit the synthesis of estrogen or to block the activity of estrogen at the estrogen receptor (ER; Howell 2005). Tamoxifen, a selective ER modulator was the first endocrine agent to become widely available: in women with ER-positive breast cancer, 5 years of adjuvant tamoxifen have been shown to reduce disease recurrence rates by almost half and decrease mortality by a third (Early Breast Cancer Trialists’ Collaborative Group 2005). However, this drug has been associated with increased risks of potentially life-threatening events, such as endometrial cancer (Bergman et al. 2000), uterine sarcoma (Wickerham et al. 2002), and thromboembolic events (Decensi et al. 2005). These adverse effects have been largely attributed to the partial estrogen agonist activity of tamoxifen.

Over recent years, as the mechanisms underlying the responses to hormonal interventions were elucidated, a number of new endocrine agents have been developed, including aromatase inhibitors (AIs) and anti-estrogens without partial agonist activity. AIs, which block the enzyme aromatase that catalyses the conversion of androgens into estrogens, are replacing tamoxifen as standard treatment for early and advanced breast cancer. The ER antagonist ICI 182 780 (fulvestrant), which unlike tamoxifen has no estrogen agonist activity (Wakeling et al. 1991), is also starting to challenge standard treatments. Fulvestrant (Faslodex) is currently approved in both the United States and Europe for the treatment of hormone receptor-positive metastatic breast cancer in postmenopausal women with disease progression following anti-estrogen therapy. Fulvestrant has a unique mode of action: it binds, blocks, and degrades the ER and consequently lacks cross-resistance with other endocrine agents (Wakeling et al. 1991, Dauvois et al. 1993, McClelland et al. 1996). Hot flushes represent the most common adverse events observed during treatment with the drug.

Soy isoflavones are a class of estrogen-like compounds that have become widely used among postmenopausal women as a ‘natural’ alternative to hormone replacement therapy (Duncan et al. 2003). Since the use of isoflavone supplements to relieve menopausal symptoms (and to enhance general well-being) is more frequent among breast cancer patients (Newton et al. 2002), their use in this specific setting of patients is emerging as an area of increasing interest, relevant to the safety of isoflavones (Messina & Loprinzi 2001, Messina et al. 2006). Much of the concern arises from studies of ovariectomized, athymic mice that have demonstrated the ability of genistein and soy protein to stimulate the growth of breast cancer cells in a dose-dependent manner (Allred et al. 2001, Ju et al. 2001), as well as the ability of genistein to antagonize the inhibitory effects of tamoxifen on tumor growth (Ju et al. 2002). However, Ju et al.(2006) recently reported that dietary daidzein only slightly stimulated the in vivo growth of estrogen-dependent human breast tumor (MCF-7), while (±)-equol did not induce any stimulatory activity. Overall findings from these and our previous study (Gallo et al. 2006a) support the concept that the type, combination, and doses of isoflavones are critical factors and are likely modifiers of the different biological effects observed.

The present study was designed to investigate the potential interaction between a phytoestrogen-containing soy extract (SSE) and the anti-estrogen ICI 182 780 in a well established preclinical model of estrogen-dependent breast cancer. The primary endpoint was tumor proliferation as determined by tumor growth curves; secondary endpoints included markers of ER expression and activation (i.e. Cyclin D1, progesterone receptor, PR) as well as regulation of molecules related to angiogenesis and cell cycle control (i.e. thrombospondin-1, TSP-1 and kallikrein 6, KLK6). Finally, since there is considerable evidence for a crosstalk between the insulin-like growth factor-1 (IGF-I) and estrogen signaling pathways in the regulation of breast cancer cell proliferation, IGF-I level were measured in serum samples from all animals.

	Materials and methods

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Cell line

MCF-7 cells were purchased from the European Collection of Cell Cultures (ECACC, Salisbury, UK). According to ECACC suggestions, cells were grown in Dulbecco’s modified eagle medium supplemented with 10% fetal bovine serum, 1% non-essential amino acids mixture, and 1% kanamycin. Cells, propagated as a monolayer culture, were trypsinized twice weekly and plated at a density of 1 x 10⁵ cells/ml. All cultures were incubated at 37 °C, under 5% CO₂, in a high humidity atmosphere. On the day of dosing, cells were trypsinized and a suspension of 8 x 10⁶ cells was injected subcutaneously in the right flank of each animal (0.2 ml/mice).

Animals

Female athymic mice (homozygous Athymic Nude-nu), 5-weeks old and within a weight range of ~18–22 g, were obtained from Charles River (Calco, Lecco, Italy). Animals were housed in a purpose-built facility with a controlled environment and maintained in an isolator in which control was set to keep temperature and relative humidity at 26±2 °C and 50% respectively. Artificial lighting provided a 24-h cycle of 12 h light:12 h dark. Sterile water and food were supplied ad libitum during the study. A phytoestrogen-free semi-purified diet was used (Harlan, Carrezzana, Milano, Italy). The diet contained the following ingredients: wheat starch (42%), casein (20%), dextrin (13%), powdered sucrose (10%), cellulose (6%), corn oil (4%), mineral and vitamin premix (3%), DL-methionine (0.45%), sodium chloride, calcium carbonate, magnesium oxide, and potassium chloride. Proximate analysis: protein 19%, fat 4%, fiber 6%, carbohydrate 53.5%, ash 5.5%, and moisture 12%. Procedures and facilities followed the requirements of Commission Directive 86/609/EEC concerning the protection of animals used for experimental and other scientific purposes. Italian legislation is defined in the Decreto Legislativo No. 116 of 27 January 1992. The project was approved by the local ethics committee.

Human tumor xenograft growth

SOYSELECT (SSE, Indena spa, Milan, Italy) is a standardized extract from soy with a double standardization (13–17% isoflavone glycosides genistin and daidzin, and not <18% B-group saponins, by HPLC). The product is prepared by extracting ripe whole soy beans or oil-free soy flour with aliphatic alcohols, through an industrial manufacturing proprietary process (patents US 6 280 777 and US 6 607 757). One gram of extract also contains 0.058 g protein, 0.035 g fat, and 0.023 g ash, with the remaining matter undefined. The batch used in the study contained 14.7% isoflavone glycosides and 21.2% B-group saponins. SSE was dissolved in sterile water at the concentrations required for dosing. ICI 182 780, purchased from Tocris (Bristol, UK), was dissolved in peanut oil.

One week after the arrival, a total of 40 mice were ovariectomized and allowed a week recovery. There were five experimental groups in the study, each consisting of eight mice. Groups 2 (referred to as 17ß-estradiol (17ß-E2) group), 3 (ICI 182 780 group), 4 (SSE 50 group), and 5 (SSE 100 group) were implanted subcutaneously with 60-day release, 0.18 mg 17ß-E2 pellets (Innovative Research of America, Sarasota, Fl, USA) one week after ovariectomy (i.e. the day before tumor inoculation); these pellets are designed to produce 80–100 pg/ml of serum estradiol (as indicated by the supplier). Mice in groups 3, 4, and 5 also received ICI 182 780 via the s.c. route, at 5 mg/mouse, once a week for 4 weeks. Groups 4 and 5 also received SSE at 50 and 100 mg/kg per day respectively, by oral gavage, 5 consecutive days per week, until the end of the study (i.e. day 51). Group 1 was used as negative control (NC, i.e. no estrogen supplementation). Groups 1, 2, and 3 received the vehicle (sterile water) by oral gavage. Treatments started one day after tumor inoculation.

Dosages of SSE were selected on the basis of previous studies carried out in our laboratory (Gallo et al. 2005, 2006a,Gallo et al. b); these dosages were seen to produce physiological plasma levels of isoflavones. In fact, a previous pilot study carried out in our laboratory showed that following administration of 50 and 100 mg/ kg per day SSE to healthy athymic mice, plasma concentrations of total daidzein and genistein were as follows: total daidzein 1.1±0.3 and 1.6±0.7 µM, total genistein 0.8±0.1 and 0.9±0.2 µM (mean±S.D.), for the low- and high-dose groups respectively (unpublished data). Actually, these isoflavone blood levels are in the range of those found in Japanese women on a traditional soy diet: daidzein mean concentrations, 246.8 nmol/l (range 0–2407); genistein mean concentrations, 501.9 nmol/l (range 0–4192; Morton et al. 2002). In addition, similar blood concentrations of isoflavones were detected in a previous clinical trial in menopausal women receiving the tested extract (Scambia et al. 2000). During the study, mice were checked daily for any adverse clinical reactions. Body weight was also recorded twice per week; food intake was determined over several 24 h periods randomly throughout the study. At the end of the study, animals were killed by carbon dioxide; blood was collected and serum frozen at –20 °C for analysis; uteri were rapidly removed, free of fat and weighed. All tumors were also removed, immediately placed in liquid nitrogen and stored at –80 °C for analysis.

Evaluation of anti-tumor activity

Tumor dimensions were measured twice per week using a caliper. The tumor weight was calculated from two-dimensional measurements (mm; Corbett et al. 1997): Tumor weight = length x width²/2. Differences in efficacy between treatment groups were expressed as the percentage of maximum tumor weight inhibition (TWI%). The ratio between the median tumor weights of treated tumors and that of control tumors x 100 (T/C%) was assessed on each day of measurement and used to calculate the TWI%: TWI% = 100–T/C%.

Real-time quantitative PCR

Four mRNA targets were selected to evaluate the ability of SSE to modulate these targets. Analysis was done on cyclin D1, PR, TSP-1, and KLK6. Analysis was carried out on MCF-7 tumors excised at the end of the study. A real-time quantitative RT-PCR was performed using the iCycler iQ system (Bio-Rad). cDNA was prepared starting from 1 µg total RNA using the iScrypt cDNA Synthesis Kit, according to the manufacture’s instructions. For those groups receiving ICI 182 780 treatment, the yield of RNA extraction from xenografts was low and, for this reason, RNA was pooled for each group of animals; specifically, after extraction, equal amounts of RNA were taken from each tumor and pooled per experimental group. RNA polymerase II was used to normalize gene expression data. Each analysis was repeated in triplicate, at least thrice for each condition, and results were then averaged. Amplifications were carried out using the primers reported in Table 1. To each primer, iQ SYBR Green Supermix (Bio-Rad) was used in a final volume of 25 µl, starting with a 3-min template denaturation step at 95 °C, followed by 40 cycles of 15 s at 95 °C, and 1 min at 60 °C. Standard curves were generated using a serial dilution of the initial amount of control cDNA to determine the range of template concentrations, which showed a good linearity and efficiency for the different reactions. Melt curves of the reaction products were also generated to assess the specificity of the measured fluorescence. The mean of threshold cycles (Ct) for each specimen was used to obtain the fold change expression level applying the following equation: Fold change = 2^–(Ct), where Ct = Ct specific gene-Ct housekeeping gene and (Ct) = Ct specimen–Ct control. A fold change equal to 1 represents a sample with an expression level equal to the selected control condition (i.e. 17ß-E2). This operation was done using the Excel spreadsheet RelQuant (Bio-Rad).

Circulating levels of IGF-I were determined by RIA following the instructions provided by the manufacturer (product code IGF-R22, Mediagnost, Reutlingen, Germany).

Statistical analysis

The Steel–Dwass test was used to detect the statistical significance of the differences between treatments on tumor weight growth curves. Data from plasma IGF-1 analysis and uterus weight were analyzed by one-way ANOVA followed by Dunnett’s or Tukey-Kramer multiple comparison test. P-values were considered to be significant when <0.05. Statistical analysis was performed using the Kyplot freeware package (Kyens Lab. Inc., Tokyo, Japan).

	Results

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Effect of treatments on the growth of MCF-7 in ovariectomized athymic female mice

Ovariectomized athymic mice implanted with MCF-7 cells were divided into five treatment groups. As expected, MCF-7 xenografts did not form proliferating tumors in the ovariectomized hosts without estrogenic supplementation (negative controls; Fig. 1). Tumors in the positive control group (17ß-E2) grew rapidly, statistical analysis showing significant differences when compared with negative controls (P < 0.01). Treatment with ICI 182 780 (5 mg/mouse, once weekly, for 4 weeks) significantly inhibited (P < 0.01) the estradiol-sustained growth of MCF-7 xenografts in athymic mice. Concomitant administration of 50 mg/kg per day SSE slightly potentiated the growth inhibitory activity of ICI 182 780, being the group mean tumor weight consistently lower than ICI 182 780 group throughout the study; this difference, however, did not achieve statistical significance. Conversely, treatment with 100 mg/kg per day SSE partially negated ICI activity, growth curves being significantly different from both ICI 182 780- and SSE 50-treated group (P < 0.01); the tumor growth remained, however, significantly lower when compared with the positive control (P < 0.01). At the end of the study, a TWI% of 93, 93, and 76% was calculated for groups 3 (ICI 182 780), 4 (SSE 50), and 5 (SSE 100) respectively. Body weight and food consumption did not significantly differ among treatment groups (data not shown).

View larger version (19K): [in this window] [in a new window]   Figure 1 Effect of treatments on MCF7-tumor growth in ovariectomized athymic mice (mean±S.D., n = 8 mice/group). One day before tumor inoculation, mice were implanted with 17ß-estradiol pellet (17ß-E2, 0.18 mg/pellet). Starting from the day after tumor cell implantation, mice were treated with 0 (positive control) or ICI 182 780 (5 mg/mouse, once weekly, 4 consecutive weeks); the SSE 50 and SSE 100 groups also received either 50 or 100 mg/kg per day SSE, by oral gavage, 5 consecutive days per week, until the end of the study. A negative control group (NC), without 17ß-estradiol supplementation, was also included in the study. Groups with different letters are significantly different (P < 0.01, by Steel-Dwass test).

To evaluate the ability of SSE to modulate mRNA expression of selected genes, we conducted quantitative RT-PCR analysis using mRNA isolated from MCF-7 tumors in positive controls (17ß-E2), ICI 182 780-treated mice, 50 and 100 mg/kg per day SSE-treated mice. Specifically, the following genes were evaluated: cyclin D1, PR, TSP-1, and KLK6. The analysis was performed through direct quantification of the expression of the selected genes versus the positive control (i.e. 17ß-E2 group). Results obtained are shown in Table 2. In ICI 182 780-treated tumors, the expression of cyclin D1 (0.52) and PR (0.05) was considerably reduced; likewise, mRNA expressions of TSP-1 (0.6) and KLK6 (0.34) were down-regulated in this group, although to a lesser degree. These findings are in keeping with the ability of pure anti-estrogens to affect the expression of direct targets of 17ß-E2. In SSE-treated mice, the modulation of the selected genes was seen to be dose-dependent. Specifically, at a low dosage (SSE 50) there was a tendency towards an increase of the anti-estrogen effect, with a further reduction in the mRNA expression of cyclin D1, PR, TSP-1, and KLK6 (compared with ICI 182 780); on the other hand, at the higher dosages (SSE 100), the enhancement of the anti-estrogen effect disappeared and, instead, there was a tendency towards increased expression of the 17ß-E2 targets, particularly evident for TSP-1 and KLK6.

To explore the possible effects of treatments on IGF-I, we quantitated IGF-I concentrations in the serums derived from mice at the end of the study. Results obtained (Table 3) did not show any significant difference among treatment groups.

Treatment with 17ß-E2, 0.18 mg/pellet, significantly increased the relative uterus weight (mg/g body weight) when compared with ovariectomized controls (P < 0.001; Fig. 2). Administration of ICI 182 780 (5 mg/mouse, once weekly, for 4 weeks) significantly decreased uterus weight with respect to mice receiving 17ß-E2 (P < 0.001). At both doses tested, SSE did not induce any significant change in the relative uterine weight compared with ICI 182 780-treated mice: in each group, weight means were significantly lower than 17ß-E2-treated mice and not significantly different from negative controls.

View larger version (19K): [in this window] [in a new window]   Figure 2 Effect of treatments on relative uterus weight (mg/g body weight) of ovariectomized athymic mice (mean±S.D. n = 8 mice/group). One day before tumor inoculation, mice were implanted with 17ß-estradiol pellet (17ß-E2, 0.18 mg/pellet). Starting from the day after tumor cell implantation, mice were treated with 0 (positive control) or ICI 182 780 (5 mg/mouse, once weekly, 4 consecutive weeks); the SSE 50 and SSE 100 groups also received either 50 or 100 mg/kg per day SSE, by oral gavage, five consecutive days per week, until the end of the study. A negative control group (NC), without 17ß-estradiol supplementation, was also included in the study. Groups with different letters are significantly different (P < 0.001, by Tukey-Kramer test).

	Discussion

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Data from a recent study showed that hKLK6 was specifically up-regulated upon estradiol treatment in MCF-7, as well as in other breast cancer cell lines (Paliouras & Diamandis 2007). Notably, findings from the present study confirm that the regulation of KLK6 expression is under the control of estradiol, since the ER antagonist ICI 182 780 down-regulated its expression in 17ß-E2-supplemented tumors. The discovery that kallikreins are able to hydrolyze a number of different substrates, taken together with the dysregulated expression of these proteins in breast, prostate, and ovarian cancer, raises the possibility that kallikreins could contribute to the invasiveness and/or progression of these cancers (Borgono & Diamandis 2004). Also TSP-1 production is under the control of estradiol: Ghosh et al.(2000) found an approximately fourfold up-regulation of expression in MCF-7 cells in response to 17ß-E2 treatment and, more recently, Buterin et al.(2006) reported that TSP-1 is up-regulated in an estrogen-dependent manner in T47D cells, an effect also elicited by genistein, although to a lesser degree. TSP-1 is a multi-functional protein that has received great attention due to observations which suggest that it reduces angiogenesis, leading to the inhibition of tumor growth (Ren et al. 2006). However, the role of TSP-1 in breast cancer is controversial, with studies showing that paradoxically, TSP-1 production by breast cancer cells is associated with tumor cell proliferation rather than regression (Wang et al. 1996). It has also been demonstrated that patients with breast, lung, colon, and gynecological malignancies have significantly elevated serum levels of TSP-1 compared with non-tumor bearing human controls (Esemuede et al. 2004).

IGF-1 and estrogens have been shown to be intimately linked to the progression of a number of human cancers, notably breast cancer (Yee & Lee 2000). Evidence for a cross-talk between the IGF-I and estrogen signaling pathways in the regulation of breast cancer cell proliferation have been recently reported, with studies showing an up- and a down-regulation of IGF-1 receptor expression following exposure of MCF-7 cells to 17ß-E2 (Stewart et al. 1990) and ICI 182 780 (Huynh et al. 1996) respectively. Recent studies also suggest a role for soybean phytochemicals in the modulation of IGF-1 signaling pathway: specifically, a reduction in circulating levels of this growth factor was reported in mice bearing human prostate or bladder tumors following consumption of dietary soy products (Zhou et al. 1999, Singh et al. 2006). Results obtained in the current study did not show any modulation of IGF-1 serum levels either upon treatment with ICI 182 780 or upon concomitant administration of the anti-estrogen and SSE. The reason why effects of soy isoflavones on IGF-I signaling are seen in some experimental systems but not others are not understood, but may involve differences in both cell lines and/or experimental conditions.

In conclusion, results from this study raise concerns about the consumption of isoflavone supplements in conjunction with ICI 182 780 (fulvestrant) therapy in postmenopausal women with estrogen-dependent breast cancer. Additional studies are warranted to develop a more complete understanding of the mechanisms involved in the interaction between the complex soy products that people actually consume and anti-cancer therapies. Indeed, the consideration of whether a particular exposure could have effects and whether these would be positive or negative will likely depend on the product, level of exposure and dosing regimen, and the particular end point being examined. Such fundamental studies are long overdue to allow individuals, including physicians, to make informed decisions on the value or otherwise of the current unregulated, and potentially harmful, consumption of phytoestrogen products by breast cancer patients.

	Funding

 
The authors declare that there is no conflict of interest that would prejudice the impartiality of this scientific work.

	References

Top Abstract Introduction Materials and methods Results Discussion References

 
Allred CD, Allred KF, Ju YH, Virant SM & Helferich WG 2001 Soy diets containing varying amounts of genistein stimulate growth of estrogen-dependent (MCF-7) tumors in a dose-dependent manner. Cancer Research 61 5045–5050.[Abstract/Free Full Text]

Bergman L, Beelen ML, Gallee M, Hollema H, Benraadt J & van Leeuwen FE 2000 Risk and prognosis of endometrial cancer after tamoxifen for breast cancer. Comprehensive Cancer Centres’ ALERT Group. Assessment of liver and endometrial cancer risk following tamoxifen. Lancet 356 881–887.[CrossRef][ISI][Medline]

Borgono CA & Diamandis EP 2004 The emerging roles of human tissue kallikreins in cancer. Nature Reviews. Cancer 4 876–890. Review.[CrossRef][ISI][Medline]

Buterin T, Koch C & Naegeli H 2006 Convergent transcriptional profiles induced by endogenous estrogen and distinct xenoestrogens in breast cancer cells. Carcinogenesis 27 1567–1578.[Abstract/Free Full Text]

Corbett T, Valeriote F, LoRusso P, Polin L, Panchapor C, Pugh S, White K, Knight J, Demchik L, Jones J et al. 1997 In vivo methods for screening and preclinical testing. In Anticancer Drug Development Guide, pp 75–99. Ed BA Teicher. Totowa, New Jersey: Humana Press.

Dauvois S, White R & Parker MG 1993 The antiestrogen ICI 182780 disrupts estrogen receptor nucleocytoplasmic shuttling. Journal of Cell Science 106 1377–1388.[Abstract]

Decensi A, Maisonneuve P, Rotmensz N, Bettega D, Costa A, Sacchini V, Salvioni A, Travaglini R, Oliviero P, D’Aiuto G et al. 2005 Effect of tamoxifen on venous thromboembolic events in a breast cancer prevention trial. Circulation 111 650–656.[Abstract/Free Full Text]

Duncan AM, Phipps WR & Kurzer MS 2003 Phytooestrogens. Best Practice and Research in Clinical Endocrinology and Metabolism 17 253–271. Review.[CrossRef]

Early Breast Cancer Trialists’ Collaborative Group 2005 Effects of chemotherapy and hormonal therapy for early breast cancer on recurrence and 15-year survival: an overview of the randomised trials. Lancet 365 1687–1717.[CrossRef][ISI][Medline]

Esemuede N, Lee T, Pierre-Paul D, Sumpio BE & Gahtan V 2004 The role of Thrombospondin-1 in human disease. Journal of Surgical Research 122 135–142. Review.[CrossRef][ISI][Medline]

Gallo D, Zannoni GF, Apollonio P, Martinelli E, Ferlini C, Passetti G, Riva A, Morazzoni P, Bombardelli E & Scambia G 2005 Characterization of the pharmacologic profile of a standardized soy extract in the ovariectomized rat model of menopause: effects on bone, uterus, and lipid profile. Menopause 12 589–600.[CrossRef][ISI][Medline]

Gallo D, Ferlini C, Fabrizi M, Prislei S & Scambia G 2006a Lack of stimulatory activity of a phytoestrogen-containing soy extract on the growth of breast cancer tumors in mice. Carcinogenesis 27 1404–1409.[Abstract/Free Full Text]

Gallo D, Zannoni GF, Martinelli E, Ferlini C, Fabrizi M, Riva A, Morazzoni P, Bombardelli E & Scambia G 2006b Estradiol and phytoestrogens differently influence the rodent postmenopausal mammary gland. Menopause 13 72–79.[CrossRef][ISI][Medline]

Ghosh MG, Thompson DA & Weigel RJ 2000 PDZK1 and GREB1 are estrogen-regulated genes expressed in hormone-responsive breast cancer. Cancer Research 60 6367–6375.[Abstract/Free Full Text]

Howell A 2005 New developments in the treatment of postmenopausal breast cancer. Trends in Endocrinology and Metabolism 16 420–428.[ISI][Medline]

Huynh H, Nickerson T, Pollak M & Yang X 1996 Regulation of insulin-like growth factor I receptor expression by the pure antiestrogen ICI 182780. Clinical Cancer Research 2 2037–2042.[Abstract]

Jemal A, Siegel R, Ward E, Murray T, Xu J, Smigal C & Thun MJ 2006 Cancer statistics, 2006. CA: A Cancer Journal for Clinicians 56 106–130.[Abstract/Free Full Text]

Ju YH, Allred CD, Allred KF, Karko KL, Doerge DR & Helferich WG 2001 Physiological concentrations of dietary genistein dose-dependently stimulate growth of estrogen-dependent human breast cancer (MCF-7) tumors implanted in athymic nude mice. Journal of Nutrition 131 2957–2962.[Abstract/Free Full Text]

Ju YH, Doerge DR, Allred KF, Allred CD & Helferich WG 2002 Dietary genistein negates the inhibitory effect of tamoxifen on growth of oestrogen-dependent human breast cancer (MCF-7) cells implanted in athymic mice. Cancer Research 62 2474–2477.[Abstract/Free Full Text]

Ju YH, Fultz J, Allred KF, Doerge DR & Helferich WG 2006 Effects of dietary daidzein and its metabolite, equol, at physiological concentrations on the growth of estrogen-dependent human breast cancer (MCF-7) tumors implanted in ovariectomized athymic mice. Carcinogenesis 27 856–863.[Abstract/Free Full Text]

McClelland RA, Gee JM, Francis AB, Robertson JF, Blamey RW, Wakeling AE & Nicholson RI 1996 Short-term effects of pure antioestrogen ICI 182780 treatment on oestrogen receptor, epidermal growth factor receptor and transforming growth factor-alpha protein expression in human breast cancer. European Journal of Cancer 32A 413–416.[CrossRef]

Messina MJ & Loprinzi CL 2001 Soy for breast cancer survivors: a critical review of the literature. Journal of Nutrition 131 3095S–3108S. Review.[Abstract/Free Full Text]

Messina M, McCaskill-Stevens W & Lampe JW 2006 Addressing the soy and breast cancer relationship: review, commentary, and workshop proceedings. Journal of the National Cancer Institute 98 1275–1284. Review.[Abstract/Free Full Text]

Morton MS, Arisaka O, Miyake N, Morgan LD & Evans BA 2002 Phytoestrogen concentrations in serum from Japanese men and women over forty years of age. Journal of Nutrition 132 3168–3171.[Abstract/Free Full Text]

Newton KM, Buist DS, Keenan NL, Anderson LA & LaCroix AZ 2002 Use of alternative therapies for menopause symptoms: results of a population-based survey. Obstetrics and Gynecology 100 18–25.[Abstract/Free Full Text]

Osborne CK, Coronado-Heinsohn EB, Hilsenbeck SG, McCue BL, Wakeling AE, McClelland RA, Manning DL & Nicholson RI 1995 Comparison of the effects of a pure steroidal antiestrogen with those of tamoxifen in a model of human breast cancer. Journal of the National Cancer Institute 87 746–750.[Abstract/Free Full Text]

Paliouras M, Diamandis EP 2007 Coordinated steroid hormone-dependent and independent expression of multiple kallikreins in breast cancer cell lines. Breast Cancer Research and Treatment 102 7–18.[CrossRef][ISI][Medline]

Ren B, Yee KO, Lawler J & Khosravi-Far R 2006 Regulation of tumor angiogenesis by thrombospondin-1. Biochimica et Biophysica Acta 1765 178–188. Review.[Medline]

Scambia G, Mango D, Signorile PG, Anselmi R, Palena C, Gallo D, Bombardelli E, Morazzoni P, Riva A & Mancuso S 2000 Clinical effects of a standardized soy extract in postmenopausal women: a pilot study. Menopause 7 105–111.[ISI][Medline]

Singh AV, Franke AA, Blackburn GL & Zhou JR 2006 Soy phytochemicals prevent orthotopic growth and metastasis of bladder cancer in mice by alterations of cancer cell proliferation and apoptosis and tumor angiogenesis. Cancer Research 66 1851–1858.[Abstract/Free Full Text]

Stewart AJ, Johnson MD, May FE & Westley BR 1990 Role of insulin-like growth factors and the type I insulin-like growth factor receptor in the estrogen-stimulated proliferation of human breast cancer cells. Journal of Biological Chemistry 265 21172–21178.[Abstract/Free Full Text]

Wakeling AE, Dukes M & Bowler J 1991 A potent specific pure antiestrogen with clinical potential. Cancer Research 51 3867–3873.[Abstract/Free Full Text]

Wang TN, Qian X, Granick MS, Solomon MP, Rothman VL, Berger DH & Tuszynski GP 1996 Thrombospondin-1 (TSP-1) promotes the invasive properties of human breast cancer. Journal of Surgical Research 63 39–43.[CrossRef][ISI][Medline]

Wickerham DL, Fisher B, Wolmark N, Bryant J, Costantino J, Bernstein L & Runowicz CD 2002 Association of tamoxifen and uterine sarcoma. Journal of Clinical Oncology 20 2758–2760. Review.[Free Full Text]

Yee D & Lee AV 2000 Crosstalk between the insulin-like growth factors and estrogens in breast cancer. Journal of Mammary Gland Biology and Neoplasia 5 107–115. Review.[CrossRef][ISI][Medline]

Zhou JR, Gugger ET, Tanaka T, Guo Y, Blackburn GL & Clinton SK 1999 Soybean phytochemicals inhibit the growth of transplantable human prostate carcinoma and tumor angiogenesis in mice. Journal of Nutrition 129 1628–1635.[Abstract/Free Full Text]

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