Urinary Estrogen Research Articles

Effect of Diindolylmethane on Estrogen-related Hormones, Metabolites and Tamoxifen Metabolism: Results of a Randomized, Placebo-controlled Trial

Abstract Dietary supplement use is high among breast cancer survivors. One compound natural to cruciferous vegetables, diindolylmethane (DIM), is among the supplements commonly used. This bioactive compound has significant experimental evidence for bioactivity in breast chemoprevention. Sparse evidence in the form of well-designed human clinical trials exist to test its efficacy or safety. Methods: In this double-blind placebo-controlled study women taking tamoxifen for breast cancer primary or tertiary prevention were randomly assigned to receive 150 mg DIM (BioResponse(BR)-DIM) twice daily or a placebo for a minimum period of 12 months. Primary outcome was change in urinary estrogen metabolites 2-hydroxyestrone and 16α-hydroxyestrone (baseline to 6 weeks, 6 and 12 months). Secondary endpoints included breast density by mammogram and fat:water ratio MRI (baseline to 12 months) and serum estrogens (baseline to 6, 12 months). Safety data were also evaluated, including tamoxifen metabolites. Results: Adherence to study pills was >91% by pill count and urinary DIM metabolite assessment. In participants assigned DIM there was a significant and sustained shift in urinary estrogen metabolism favoring a higher 2-OH:16α-OH ratio; sex hormone binding globulin (SHBG) was also increased. No change in breast density was demonstrated. Safety analysis showed no appreciable differences in adverse events by treatment arm; however, tamoxifen metabolism for the parent compound as well as endoxifen and 4-OH endoxifen were appreciably reduced in women assigned to the DIM arm. Conclusions In this first large study of DIM in the setting of breast cancer chemoprevention, a favorable shift in estrogen metabolism and SHBG was demonstrated. However, the reduction in tamoxifen metabolites raises concern regarding the potential interaction between DIM and tamoxifen, an area in need of continued research. Impact Given the widespread and generally unsupported use of dietary supplementation by breast cancer survivors, these data will help to inform the use of DIM as a dietary supplement for breast cancer patients receiving tamoxifen.

In-tube solid-phase microextraction based on NH2-MIL-53(Al)-polymer monolithic column for online coupling with high-performance liquid chromatography for directly sensitive analysis of estrogens in human urine

In this work, a novel NH2-MIL-53(Al) incorporated poly(styrene-divinylbenzene-methacrylic acid) (poly(St-DVB-MAA)) monolith was prepared via chemical fabrication. Moreover, it has been efficiently applied to the in-tube solid-phase microextraction (SPME) for online coupling with high-performance liquid chromatography (HPLC) to the direct determination of five estrogens in human urine samples. The NH2-MIL-53(Al)-polymer monolith was suitable for in-tube SPME owing to its good permeability, high extraction efficiency, chemical stability, good reproducibility and long lifetime. The extraction conditions including extraction solvent, pH of sample solution, flow rate of extraction and desorption, and desorption volume were investigated. Under the optimum conditions, the enrichment factors were 180–304 and saturated amounts of extraction were 2326–21393 pmol for estriol, 17β-estradiol, estrone, ethinyl estradiol and progesterone, respectively. The adsorption mechanism was also explored which contributed to its strong extraction to target compounds. The proposed method had low limit of detection (2.0–40ng/L) and good linearity (with R2 between 0.9908 and 0.9978). Four endogenous estrogens were detected in urine samples and the recoveries of all five analytes were ranged from 75.1–120% with relative standard deviations (RSDs) less than 8.7%. The results showed that the proposed online SPME-HPLC method based on NH2-MIL-53(Al)-polymer monolithic column was highly sensitive for directly monitoring trace amount of estrogens in human urine sample.

Soy Food Intake and Biomarkers of Breast Cancer Risk: Possible Difference in Asian Women?

ABSTRACTSoy foods may protect against breast cancer in Asian but not in Western populations. We examined if the levels of various markers of breast cancer risk and inflammation, as well as the effects of soy food consumption on these markers, differ between Asian and non-Asian premenopausal women in two soy intervention trials. One study randomized 220 women to a 2-yr intervention and the other one randomized 96 women in a crossover design to examine the effects of consumption of 2 daily soy servings on nipple aspirate fluid (NAF) volume; estrogens in serum, NAF, and urine; insulin-like growth factor-1 (IGF-1), IGF-binding protein 3, and inflammatory markers in serum; and mammographic densities. Mixed linear models were applied to assess ethnic differences in biomarkers and response to the soy diet. Serum C-reactive protein, serum leptin, NAF volume, and NAF estrone sulfate were lower, while urinary isoflavones were higher in Asian than in non-Asian women. A significant interaction (pinteraction = 0.05) between ethnicity and soy diet was observed for IGF-1 but not for other biomarkers. The current findings suggest possible ethnic differences in levels of biomarkers for breast cancer risk but little evidence that Asian women respond differently to soy foods than non-Asian women.

Systemic Levels of Estrogens and PGE2 Synthesis in Relation to Postmenopausal Breast Cancer Risk

Background: Prostaglandin E2 (PGE2) induces aromatase expression in adipose tissue, leading to increased estrogen production that may promote the development and progression of breast cancer. However, few studies have simultaneously investigated systemic levels of PGE2 and estrogen in relation to postmenopausal breast cancer risk.Methods: Here, we determined urinary estrogen metabolites (EM) using mass spectrometry in a case-cohort study (295 incident breast cancer cases and 294 subcohort members), and using linear regression estimated the effect of urinary levels of a major PGE2 metabolite (PGE-M) on EMs. HRs for the risk of developing breast cancer in relation to PGE-M and EMs were compared between Cox regression models with and without mutual adjustment.Results: PGE-M was a significant predictor of estrone (E1), but not estradiol (E2) levels in multivariable analysis. Elevated E2 levels were associated with an increased risk of developing breast cancer [HRQ5vs.Q1, 1.54; 95% confidence interval (CI), 1.01-2.35], and this association remained unchanged after adjustment for PGE-M (HRQ5vs.Q1, 1.52; 95% CI, 0.99-2.33). Similarly, elevated levels of PGE-M were associated with increased risk of developing breast cancer (HRQ4vs.Q1, 2.01; 95% CI, 1.01-4.29), and this association was only nominally changed after consideration of E1 or E2 levels.Conclusions: Urinary levels of PGE-M and estrogens were independently associated with future risk of developing breast cancer among these postmenopausal women.Impact: Increased breast cancer risk associated with PGE-M might not be fully explained by the estrogens-breast cancer association alone but also by additional effects related to inflammation. Cancer Epidemiol Biomarkers Prev; 26(3); 383-8. ©2016 AACR.

Sex hormones in women with and without migraine: Evidence of migraine-specific hormone profiles.

To compare daily sex hormone levels and rates of change between women with history of migraine and controls. History of migraine, daily headache diaries, and daily hormone data were collected in ovulatory cycles of pre- and early perimenopausal women in the Study of Women's Health Across the Nation. Peak hormone levels, average daily levels, and within-woman day-to-day rates of decline over the 5 days following each hormone peak were calculated in ovulatory cycles for conjugated urinary estrogens (E1c), pregnanediol-3-glucuronide, luteinizing hormone, and follicle-stimulating hormone. Comparisons were made between migraineurs and controls using 2-sample t tests on the log scale with results reported as geometric means. The sample included 114 women with history of migraine and 223 controls. Analyses of within-woman rates of decline showed that E1c decline over the 2 days following the luteal peak was greater in migraineurs for both absolute rate of decline (33.8 [95% confidence interval 28.0-40.8] pg/mgCr vs 23.1 [95% confidence interval 20.1-26.6] pg/mgCr, p = 0.002) and percent change (40% vs 30%, p < 0.001). There was no significant difference between migraineurs and controls in absolute peak or daily E1c, pregnanediol-3-glucuronide, luteinizing hormone, and follicle-stimulating hormone levels. Secondary analyses demonstrated that, among migraineurs, the rate of E1c decline did not differ according to whether a headache occurred during the cycle studied. Migraineurs are characterized by faster late luteal phase E1c decline compared to controls. The timing and rate of estrogen withdrawal before menses may be a marker of neuroendocrine vulnerability in women with migraine.

Endogenous Estrogens, Estrogen Metabolites, and Breast Cancer Risk in Postmenopausal Chinese Women

The role of estrogen metabolism in determining breast cancer risk and differences in breast cancer rates between high-incidence and low-incidence nations is poorly understood. We measured urinary concentrations of estradiol and estrone (parent estrogens) and 13 estrogen metabolites formed by irreversible hydroxylation at the C-2, C-4, or C-16 positions of the steroid ring in a nested case-control study of 399 postmenopausal invasive breast cancer case participants and 399 matched control participants from the population-based Shanghai Women's Health Study cohort. Odds ratios (ORs) and 95% confidence intervals (CIs) of breast cancer by quartiles of metabolic pathway groups, pathway ratios, and individual estrogens/estrogen metabolites were estimated by multivariable conditional logistic regression. Urinary estrogen/estrogen metabolite measures were compared with those of postmenopausal non-hormone-using Asian Americans, a population with three-fold higher breast cancer incidence rates. All statistical tests were two-sided. Urinary concentrations of parent estrogens were strongly associated with breast cancer risk (ORQ4vsQ1 = 1.94, 95% CI = 1.21 to 3.12, Ptrend = .01). Of the pathway ratios, the 2-pathway:total estrogens/estrogen metabolites and 2-pathway:parent estrogens were inversely associated with risk (ORQ4vsQ1 = 0.57, 95% CI = 0.35 to 0.91, Ptrend = .03, and ORQ4vsQ1 = 0.61, 95% CI = 0.37 to 0.99, Ptrend = .04, respectively). After adjusting for parent estrogens, these associations remained clearly inverse but lost statistical significance (ORQ4vsQ1 = 0.65, 95% CI = 0.39 to 1.06, Ptrend = .12 and ORQ4vsQ1 = 0.76, 95% CI = 0.44 to 1.32, Ptrend = .28). The urinary concentration of all estrogens/estrogen metabolites combined in Asian American women was triple that in Shanghai women. Lower urinary parent estrogen concentrations and more extensive 2-hydroxylation were each associated with reduced postmenopausal breast cancer risk in a low-risk nation. Markedly higher total estrogen/estrogen metabolite concentrations in postmenopausal United States women (Asian Americans) than in Shanghai women may partly explain higher breast cancer rates in the United States.

Exercise during energy restriction mitigates bone loss but not alterations in estrogen status or metabolic hormones.

Energy restriction causes bone loss, increasing stress fracture risk. The impact of exercise during energy restriction on bone and endocrine factors is examined. Exercise with energy restriction did not influence endocrine factors, but did mitigate some bone loss seen with energy restriction in sedentary rats. Chronic dietary energy restriction (ER) leads to bone loss and increased fracture risk. Strictly controlled trials of long-term ER with and without vigorous exercise are required to determine whether exercise loading can counterbalance ER-induced bone loss. The aim of this current project is to elucidate the impact of exercise and ER on bone mass, estrogen status, and metabolic hormones. Twenty-four virgin female Sprague-Dawley rats (n = 8/group) were divided into three groups-ad libitum fed + exercise (Adlib + EX), 40% energy restricted + exercise (ER + EX), and 40% energy restricted + sedentary (ER + SED). Energy availability between ER groups was equal. Treadmill running was performed 4days/week at 70% VO2max for 12weeks. Fat and lean mass and areal bone mineral density (aBMD) were lower after 12weeks (p < 0.05) for ER + EX vs Adlib + EX, but ER + EX aBMD was higher than ER + SED (p < 0.0001). Serum leptin and a urinary estrogen metabolite, estrone-1-glucuronide (E1G), were lower at week 12 (p = 0.0002) with ER, with no impact of exercise. Serum insulin-like growth factor I (IGF-I) declined (p = 0.02) from baseline to week 12 in both ER groups. ER + EX exhibited higher cortical volumetric bone mineral density (vBMD) at the midshaft tibia (p = 0.006) vs ER + SED. Exercise during ER mitigated some, but not all, of the bone loss observed in sedentary ER rats, but had little impact on changes in urinary E1G and serum IGF-I and leptin. These data highlight the importance of both adequate energy intake and the mechanical loading of exercise in maintaining bone mass.

Association of Active and Sedentary Behaviors with Postmenopausal Estrogen Metabolism.

Physical activity may reduce endogenous estrogens, but few studies have assessed effects on estrogen metabolism and none have evaluated sedentary behavior in relation to estrogen metabolism. We assessed relationships between accelerometer-measured physical activity and sedentary behavior and 15 urinary estrogens and estrogen metabolites (EM) among postmenopausal controls from a population-based breast cancer case-control study conducted in Poland (2000-2003). Postmenopausal women (N = 542) were ages 40 to 72 yr and not currently using hormone therapy. Accelerometers, worn for 7 d, were used to derive measures of average activity (counts per day) and sedentary behavior (<100 counts per minute per day). Estrogen metabolites were measured in 12-h urine samples using liquid chromatography-tandem mass spectrometry. Estrogen metabolites were analyzed individually, in metabolic pathways (C-2, -4, or -16), and as ratios relative to parent estrogens. Geometric means of estrogen metabolites by tertiles of accelerometer-measures, adjusted for age and body mass, were computed using linear models. High activity was associated with lower levels of estrone and estradiol (P trend = 0.01), whereas increased sedentary time was positively associated with these parent estrogens (P trend = 0.04). Inverse associations were observed between high activity and 2-methoxyestradiol, 4-methoxyestradiol, 17-epiestriol, and 16-epiestriol (P trend = 0.03). Sedentary time was positively associated with methylated catechols in the 2- and 4-hydroxylation pathways (P trend ≤ 0.04). Women in the highest tertile of activity had increased hydroxylation at the C-2, -4, and -16 sites relative to parent estrogens (P trend ≤ 0.02), whereas increased sedentary time was associated with a lower 16-pathway/parent estrogen ratio (P trend = 0.01). Higher activity was associated with lower urinary estrogens, possibly through increased estrogen hydroxylation and subsequent metabolism, whereas sedentary behavior may reduce metabolism.

Oestrogen activity of the serum in adolescents with Type 1 diabetes.

To assess serum oestrogen levels and oestrogenic activity in adolescents with Type 1 diabetes compared with a healthy control group. We conducted a cross-sectional study that evaluated adolescents with Type 1 diabetes (n = 38) and healthy adolescents (control group; n = 32). Serum oestrogens, urinary oestrogen metabolites and serum oestrogenic activity were assessed. Oestrogenic activity was evaluated in an in vitro cell proliferation assay using a modified E-screen assay with MCF-7/BUS cells. Adolescents with Type 1 diabetes had lower oestrogenic activity levels in both phases of the menstrual cycle compared with the control group (follicular phase: 76 vs 94%; luteal phase: 97 vs 131%; P < 0.01), even after adjusting for BMI, oestradiol and oestrone levels. Postmenarcheal adolescents with Type 1 diabetes had lower oestradiol levels compared with control subjects in the follicular phase (63.3 pmol/l vs 89.4 pmol/l; P < 0.01) and higher oestrone levels compared with controls in the luteal phase (196 vs 151.9 pmol/l; P < 0.05). Adolescents with Type 1 diabetes had lower levels of serum oestrogenic activity, and these were lower than expected based on their serum oestradiol levels. We postulate that changes in the serum milieu of oestrogens in patients with Type 1 diabetes may explain their decreased oestrogenic activity and may play a role in their adverse metabolic profile.

Phenolic environmental estrogens in urine and blood plasma from women with uterine leiomyoma: Epidemiological survey

To explore the effect of phenolic environmental estrogens (EE) on women with uterine leiomyoma (UL). Urine and blood plasma samples were collected from 300 patients diagnosed with UL at the Affiliated Zhongda Hospital of Southeast University between December 2013 and December 2014. Control urine and blood plasma samples were collected from 300 women who are either patients without UL or healthy volunteers presenting to the same hospital for physical examination during the same period. Bisphenol A (BPA), nonylphenol (NP) and octylphenol (OP) concentration in these samples was measured using solid phase extraction (SPE) coupled with liquid chromatography-tandem mass spectrometry. The OP concentration in urine and blood plasma was significantly higher in the UL group compared with the control group (r = 0.224, P = 0.001). Urine BPA concentration was not significantly different between the UL group and the control group (r = 0.009, P = 0.896). There was also no statistically significant difference in urine NP concentration between the two groups (r = 0.057, P = 0.419). On logistic regression, exposure concentration of urine BPA (OR, 1.129; 95%CI: 1.081-1.179) and NP (OR, 1.165; 95%CI: 1.025-1.324) was associated with UL genesis (P < 0.05). Nevertheless, there was no significant difference in blood plasma concentration of BPA, OP and NP between the two groups (P > 0.05). Urine and blood plasma EE exposure levels in women, especially the urine level, was related to the incidence of UL.

Alcohol Consumption and Urinary Estrogens and Estrogen Metabolites in Premenopausal Women.

In a cross-sectional analysis, we evaluated the associations of usual total alcohol and wine intake with a comprehensive profile of mid-luteal phase urinary estrogens and estrogen metabolites (referred to jointly as EM) in a sample of 603 premenopausal women participating in the Nurses' Health Study II (NHSII). A total of 15 individual EM (pmol/mg creatinine) were measured by a liquid chromatography/tandem mass spectrometry (LC-MS/MS) method with high accuracy and reproducibility. We used linear mixed models to calculate the adjusted geometric means of individual EM, EM grouped by metabolic pathways, and pathway ratios by category of alcohol intake with non-drinkers of alcohol as the referent. Total alcohol intake was not associated with total EM but was positively associated with estradiol (26% higher among women consuming >15 g/day vs. non-drinkers; P trend = 0.03). Wine consumption was positively associated with a number of EM measures including estradiol (22% higher among women consuming ≥ 5 drinks/week vs. non-drinkers, P trend < 0.0001). In conclusion, the total alcohol intake was positively and significantly associated with urinary estradiol levels. Some differences in urinary estrogen metabolites were observed with wine drinking, when compared with non-drinkers. This study strengthens the evidence that alcohol consumption might play a role in breast cancer and other estrogen-related conditions. Additional studies of premenopausal women are needed to further explore the association of alcohol, particularly the specific types of alcohol, on patterns of estrogen metabolism in blood, urine, and tissue.

Abstract A34: Impact of a lifestyle intervention on metabolic pathways: Results from the Diet, Exercise, Emotional Processing, and Mindfulness (DEEM) intervention

Abstract Background: Breast cancer is the most common non-dermatologic malignancy in women. One-fifth of breast cancers can be attributed to sedentary lifestyles and overweight/obesity. More than two-thirds of U.S. women are overweight or obese. Excess body weight and body fat may affect breast cancer risk through a number of mechanisms, including metabolic pathways (i.e., adipokines, inflammation, and insulin) and estrogen metabolism. To date, limited research has investigated the impact of lifestyle interventions on metabolic changes among women at high risk for breast cancer. The purpose of this pilot study was to test the feasibility of the 6-month exercise and diet intervention and to explore the impact of lifestyle intervention versus control on metabolic changes. Methods: Participants were recruited through the Seattle Cancer Care Alliance. Sixteen women were randomized with half assigned to the control group. Eligible women were 35-65 years of age, had a body mass index (BMI) between 28-40, and were at high risk of breast cancer. Dual-energy X-ray Absorptiometry (DEXA) scans, questionnaire data, blood and urine samples were collected at baseline, 3, 6, and 9- months. The DEEM lifestyle intervention is an individually-tailored intervention that includes motivational enhancement therapy, cognitive and behavioral skills training, and psychological education. Group counseling sessions were conducted weekly for 3 months then tapered over next 3 months. We measured the changes in weight, percent adiposity, central waist/hip circumferences, plasma biomarkers (adiponectin, leptin, C-reactive protein [CRP], Interleukin-6 [IL-6], Homeostatic Model Assessment-Insulin Resistance [HOMA-IR]), and urinary estrogen metabolites (Estrogen DNA Adduct [EDA]). Results: Results showed an average weight loss of 4.1 kg (- 4.6 %) in intervention group versus controls (weight gain 1.4 kg, +1.6 %) at 6 months (p = 0.02). Intervention participants experienced a decrease in percent mean adiposity (-3 %) and waist circumference (-0.2 inches) but no change in hip circumference; controls had increases in all 3 measures (+1.5 %, +0.7 inches, +2.9 inches). These changes were not significantly different. Self-reported physical activity differed between intervention and control participants (6.8 versus.1.4 hrs. /wk., p = 0.01). On average vegetable, fruit, protein intake increased and carbohydrate intake decreased only in intervention group, but not significantly different from controls. Women in the intervention arm reduced plasma CRP (mg/dL) from 0.39 to 0.26 (- 0.13 [34.5 %]) versus from 0.45 to 0.63 (+ 0.18 [39 %]) in controls (p = 0.06); and reduced plasma leptin (ng/ml) from 18.1 to 14.7 (-3.4 [18 %]) versus from 21.8 to 21.9 (+0.1 [0.1 %]) in controls (p = 0.10). Twenty-five percent of the intervention participants went from insulin resistant to non-insulin resistant (HOMA-IR&amp;lt; 2.0), while none of the controls reduced their HOMA-IR. No significant differences were observed in adiponectin, IL-6 and EDA. Conclusion: In this pilot study, the DEEM intervention facilitated weight loss and reduced adiposity. Our findings also suggest that a lifestyle intervention can improve the metabolic health of overweight/obese women at high risk of breast cancer. Future work is needed to confirm this finding in a large-scale study. Citation Format: Claire Jungyoun Han, Larissa Korde, Scott Reding, Kerryn Reding. Impact of a lifestyle intervention on metabolic pathways: Results from the Diet, Exercise, Emotional Processing, and Mindfulness (DEEM) intervention. [abstract]. In: Proceedings of the AACR Special Conference: Metabolism and Cancer; Jun 7-10, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(1_Suppl):Abstract nr A34.

Detection of urinary estrogen conjugates and creatinine using near infrared spectroscopy in Bornean orangutans (Pongo Pygmaeus).

For promoting in situ conservation, it is important to estimate the density distribution of fertile individuals, and there is a need for developing an easy monitoring method to discriminate between physiological states. To date, physiological state has generally been determined by measuring hormone concentration using radioimmunoassay or enzyme immunoassay (EIA) methods. However, these methods have rarely been applied in situ because of the requirements for a large amount of reagent, instruments, and a radioactive isotope. In addition, the proper storage of the sample (including urine and feces) on site until analysis is difficult. On the other hand, near infrared (NIR) spectroscopy requires no reagent and enables rapid measurement. In the present study, we attempted urinary NIR spectroscopy to determine the estrogen levels of orangutans in Japanese zoos and in the Danum Valley Conservation Area, Sabah, Malaysia. Reflectance NIR spectra were obtained from urine stored using a filter paper. Filter paper is easy to use to store dried urine, even in the wild. Urinary estrogen and creatinine concentrations measured by EIA were used as the reference data of partial least square (PLS) regression of urinary NIR spectra. High accuracies (R(2) > 0.68) were obtained in both estrogen and creatinine regression models. In addition, the PLS regressions in both standards showed higher accuracies (R(2) > 0.70). Therefore, the present study demonstrates that urinary NIR spectra have the potential to estimate the estrogen and creatinine concentrations.

The effect of fluphenazine decanoate on glucocorticoid production, reproductive cyclicity, and the behavioral stress response in the Persian onager (Equus hemionus onager).

Artificial insemination, performed to maximize genetic diversity in populations of zoo-housed animals, requires intensive management and has been associated with low success rates in fractious species. In these species, stressors, such as frequent handling, may impact fertility. Long-acting neuroleptic pharmaceuticals (LANs) can attenuate the stress response to handling, but may also disrupt ovulation in some species, compromising their use for artificial insemination. Therefore, the goal of this study was to determine whether LANs may be used to mitigate stress during reproductive management in wild equids without inhibiting ovulation. Six female Persian onagers (Equus hemionus onager) were treated with fluphenazine decanoate (FD; 0.1 mg/kg IM) or saline control in a random crossover design study. Urinary cortisol, progesterone, estrogen metabolites and behavior were monitored, and follicular dynamics were examined using ultrasonography until ovulation. Onagers demonstrated significantly lower cortisol concentrations (P = 0.03) when treated with FD (6.61 ± 3.26 ng/mg creatinine) compared to saline (9.73 ± 3.19 ng/mg creatinine). Overall, there were no differences in peak estrogen (P = 0.51) or progesterone (P = 0.38) concentrations between the two groups, and all animals ovulated within the expected time frame following FD treatment. However, some onagers exhibited only minor reductions in cortisol secretion and one treated female demonstrated a suppressed luteal progesterone peak, indicating a possible reproductive cost to FD administration. While FD may be useful for highly fractious equids for which the stress of handling delays or inhibits ovulation, these results warrant further investigation of dosing.

Estradiol Priming Improves Gonadotrope Sensitivity and Pro-Inflammatory Cytokines in Obese Women.

Obesity is associated with a pro-inflammatory state and relative hypogonadotropic hypogonadism. Estrogen (E2) is a potential link between these phenomena because it exhibits negative feedback on gonadotropin secretion and also inhibits production of pro-inflammatory cytokines. We sought to examine the effect of estrogen priming on the hypothalamic-pituitary-ovarian axis in obesity. This was an interventional study at an academic center of 11 obese and 10 normal-weight (NW) women. A frequent blood-sampling study and one month of daily urinary collection were performed before and after administration of transdermal estradiol 0.1 mg/d for one entire menstrual cycle. Serum LH and FSH before and after GnRH stimulation, and urinary estrogen and progesterone metabolites were measured. E2 increased LH pulse amplitude and FSH response to GnRH (P = .048, and P < .03, respectively) in obese but not NW women. After E2 priming, ovulatory obese but not NW women had a 25% increase in luteal progesterone (P = .01). Obese women had significantly higher baseline IL-6, IL-10, TGF-β, and IL-12 compared with NW (all P < .05); these levels were reduced after E2 (-6% for IL-1β, -21% for IL-8, -5% for TGF-β, -5% for IL-12; all P < .05) in obese but not in NW women. E2 priming seems to improve hypothalamic-pituitary-ovarian axis function and systemic inflammation in ovulatory, obese women. Reducing chronic inflammation at the pituitary level may decrease the burden of obesity on fertility.

Dietary Fat and Fiber Intakes Are Not Associated with Patterns of Urinary Estrogen Metabolites in Premenopausal Women1–

Interindividual differences in the bioavailability of potentially carcinogenic estrogen and estrogen metabolites (EMs) may play a role in the risk of breast cancer. We examined whether dietary intakes of fiber and fat influence premenopausal EM profiles through effects on estrogen synthesis, metabolism, or excretion. We conducted a cross-sectional analysis of 598 premenopausal women who participated in a reproducibility study (n = 109) or served as controls in a nested case-control study of breast cancer (n = 489) within the Nurses' Health Study II. Dietary intakes of fiber and fat were assessed via semiquantitative food frequency questionnaires in 1995 and 1999. Midluteal urine samples were collected between 1996 and 1999 and EMs were quantified with the use of HPLC-tandem mass spectrometry. Linear mixed models were used to estimate creatinine-adjusted geometric means for individual EMs and their pathway groups across categories of dietary intake while controlling for total energy intake and potential confounders. Higher total dietary fiber intake (>25 g/d vs. ≤15 g/d) was associated with significantly higher concentrations of 4-methoxyestradiol (50% difference, P-difference = 0.01, P-trend = 0.004) and lower concentrations of 17-epiestriol (-27% difference, P-difference = 0.03, P-trend = 0.03), but was not associated with any other EMs. The associations did not vary by fiber intake from different sources. Total fat intake (>35% energy vs. ≤25% energy) was suggestively positively associated with 17-epiestriol (22.6% difference, P-difference = 0.14, P-trend = 0.06); the association was significant for polyunsaturated fatty acid (37% difference, P-difference = 0.01, P-trend = 0.01) and trans fat (36.1% difference, P-difference = 0.01, P-trend = 0.01) intakes. Fiber and fat intakes were not strongly associated with patterns of estrogen metabolism in premenopausal women. Our data suggest estrogen metabolism is not a major mechanism through which dietary fiber and fat may affect breast or other hormone-related cancer risks.

Determination of conjugated estrogens in human urine using carrier-mediated hollow-fiber liquid phase microextraction and LC-MS/MS

The extraction conditions of the conjugated estrogens, including estrone-3-glucuronide (E1-3G), 17β-estradiol-3-glucuronide (E2-3G), and 17α-ethynylestradiol-3-glucuronide (EE2-3G), were optimized using carrier-mediated hollow-fiber liquid phase microextraction (HF-LPME) and then applied in the analysis of human urine samples. The optimal extraction conditions were as follows: organic phase composed of 20% (w/w) tri-octylamine (TOA) in 1-octanol; 2M NaCl as the acceptor phase; 800 rpm of agitation; and 50 min of extraction time. The extraction efficiency of carrier-mediated LPME was 11 times higher than that of the three-phase LPME. Estrogens and their conjugates in urine samples from pregnant and non-pregnant women were extracted and analyzed using liquid chromatography tandem mass spectrometry (LC-MS/MS). The results showed that carrier-mediated HF-LPME had good recoveries and linearity in the calibration curve. Estrogens in pregnant and non-pregnant women were mainly excreted in a conjugated form (94.85 ± 4%), whereas free estrogen was detected at a ratio of 5.15%. Both free and conjugated form of E1 were the most dominant species in all forms, which accounted for 72.7% of all the measured estrogens. Consequently, the HF-LPME in carrier-mediated mode was effectively applied to extract the estrogen conjugates in only 10 ml of human urine samples, which is the most significant advantage.

Abstract CT111: Green tea extract supplementation, estrogen metabolism and breast cancer risk in postmenopausal women at high risk of breast cancer

Abstract Background: Green tea intake may be associated with reduced risk of several cancers including breast cancer. One of the proposed mechanisms by which green tea may modify breast cancer risk is through effects on estrogen metabolism. Genetic variation in catechol-O-methyltransferase (COMT), an enzyme involved in the metabolism of both estrogens and tea catechins, may also have a role. Few studies have examined the effects of green tea intake on circulating and urinary estrogens and estrogen metabolites, and even less is known about the potential modifying influence of polymorphisms in the COMT gene. Objectives: To investigate the effects of daily intake of a highly concentrated green tea extract (GTE) for one year on circulating and urinary estrogens and their metabolites in postmenopausal women with different COMT genotypes. Methods: The Minnesota Green Tea Trial (MGTT), a randomized, double-blind, placebo-controlled, phase II clinical trial, enrolled 1075 healthy postmenopausal women with heterogeneously or extremely dense breast tissue (age = 59.78 ± 5.02 years; body mass index = 25.70 ± 8.21 kg/m2). Participants were 93% non-Hispanic white and non-current hormone users. Half of the participants (n = 538) were randomly assigned to the GTE group and were given 4 capsules a day, each containing 200 mg epigallocatechin gallate (EGCG) and the others (n = 537) to the placebo group. Twenty-four hour urine samples were collected at month 0 and at the end of the study, and fasting blood samples were drawn at months 0, 6 and 12. Circulating and urinary estrogens, as well as urinary estrogen metabolites were quantified by liquid chromatography-tandem mass spectrometry. Results: GTE supplementation was associated with reduced urinary estriol (-10.7%, P = 0.08) and 2-methoxy estrone (-12.2%, P = 0.014) compared with baseline, and these changes were significantly greater than the corresponding changes in the placebo group (P = 0.003, and P = 0.02, respectively). GTE supplementation had no significant effects on serum or urinary estrone and estradiol. Among women with the low activity COMT genotype, GTE supplementation reduced urinary estrone (-5.3%) compared with women with high activity COMT (+2.1%) (between groups comparison P = 0.049). Statistical analyses are ongoing to examine the potential modifying effects of COMT genotype on other urinary estrogen metabolites. Conclusion: Daily intake of high-dose green tea extract for 12 months exerts significant effects on urinary excretion of estrogen metabolites, and these effects may be modified by COMT polymorphisms. The potential beneficial effects of green tea extract on estrogen metabolism support the further development of green tea as a potential chemopreventive agents for estrogen-related disease prevention. Funding source: NIH/NCI (R01 CA127236). Citation Format: Hamed Samavat, Renwei Wang, Anna Wu, Jian-Min Yuan, Mindy Kurzer. Green tea extract supplementation, estrogen metabolism and breast cancer risk in postmenopausal women at high risk of breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr CT111. doi:10.1158/1538-7445.AM2015-CT111

Caffeine, coffee, and tea intake and urinary estrogens and estrogen metabolites in premenopausal women.

Prior studies have found weak inverse associations between breast cancer and caffeine and coffee intake, possibly mediated through their effects on sex hormones. High-performance liquid chromatography/tandem mass spectrometry was used to quantify levels of 15 individual estrogens and estrogen metabolites (EM) among 587 premenopausal women in the Nurses' Health Study II with mid-luteal phase urine samples and caffeine, coffee, and/or tea intakes from self-reported food frequency questionnaires. Multivariate linear mixed models were used to estimate geometric means of individual EM, pathways, and ratios by intake categories, and P values for tests of linear trend. Compared with women in the lowest quartile of caffeine consumption, those in the top quartile had higher urinary concentrations of 16α-hydroxyestrone (28% difference; Ptrend = 0.01) and 16-epiestriol (13% difference; Ptrend = 0.04), and a decreased parent estrogens/2-, 4-, 16-pathway ratio (Ptrend = 0.03). Coffee intake was associated with higher 2-catechols, including 2-hydroxyestradiol (57% difference, ≥4 cups/day vs. ≤6 cups/week; Ptrend = 0.001) and 2-hydroxyestrone (52% difference; Ptrend = 0.001), and several ratio measures. Decaffeinated coffee was not associated with 2-pathway metabolism, but women in the highest (vs. lowest) category of intake (≥2 cups/day vs. ≤1-3 cups/month) had significantly lower levels of two 16-pathway metabolites, estriol (25% difference; Ptrend = 0.01) and 17-epiestriol (48% difference; Ptrend = 0.0004). Tea intake was positively associated with 17-epiestriol (52% difference; Ptrend = 0.01). Caffeine and coffee intake were both associated with profiles of estrogen metabolism in premenopausal women. Consumption of caffeine and coffee may alter patterns of premenopausal estrogen metabolism.

The Minnesota Green Tea Trial (MGTT), a randomized controlled trial of the efficacy of green tea extract on biomarkers of breast cancer risk: study rationale, design, methods, and participant characteristics.

The Minnesota Green Tea Trial (MGTT) was a randomized, placebo-controlled, double-blinded trial investigating the effect of daily green tea extract consumption for 12 months on biomarkers of breast cancer risk. Participants were healthy postmenopausal women at high risk of breast cancer due to dense breast tissue with differing catechol-O-methyltransferase (COMT) genotypes. The intervention was a green tea catechin extract containing 843.0 ± 44.0 mg/day epigallocatechin gallate or placebo capsules for 1 year. Annual digital screening mammograms were obtained at baseline and month 12, and fasting blood and 24-h urine samples were provided at baseline and at months 6 and 12. Primary endpoints included changes in percent mammographic density, circulating endogenous sex hormones, and insulin-like growth factor axis proteins; secondary endpoints were changes in urinary estrogens and estrogen metabolites and circulating F2-isoprostanes, a biomarker of oxidative stress. The MGTT screened more than 100,000 mammograms and randomized 1,075 participants based on treatment (green tea extract vs. placebo), stratified by COMT genotype activity (high COMT vs. low/intermediate COMT genotype activity). A total of 937 women successfully completed the study and 138 dropped out (overall dropout rate = 12.8 %). In this paper we report the rationale, design, recruitment, participant characteristics, and methods for biomarker and statistical analyses.