Premenopausal Serum Research Articles

A sensitive and accurate LC-MS/MS assay with the derivatization of 1-Amino-4-methylpiperazine applied to serum allopregnanolone, pregnenolone and androsterone in pre- and postmenopausal women

The concentrations of allopregnanolone (Allopreg), pregnenolone (Preg) and androsterone (ADT) are very low in the circulation, especially in postmenopausal women, resulting in a considerable challenge for their accurate measurements in serum or plasma. In this report, a sensitive and reliable LC-MS/MS assay method has been developed using a simple sample preparation and the 1-Amino-4-methylpiperazine (AMP) derivatization procedure. A 5pg/ml (0.1pg on column) of low limit of quantitation has been achieved for Allopreg, Preg and ADT, with a sensitivity comparable to data obtained with the commercial reagent. The major benefit of this reagent is to limit the matrix effect since the excess amount of reagent can be removed during the reaction. Multiple reaction monitoring (MRM) from the derivatization of AMP not only increases the detection of these compounds but also provides a good resolution for Allopreg, Preg and ADT from interferences, especially for Allopreg from its isomers. Within the calibration range of 5pg/ml to 2000pg/ml, a good linearity was obtained with R>0.99 where the weighing factor is 1/X. Bias and coefficients of variance are within 15% for all QC levels. The matrix effect has been evaluated, well meeting the acceptance criteria according to the FDA guidelines. With this method, the concentrations of Allopreg, Preg and ADT in postmenopausal serum are in the range of 6.4–53.6pg/ml, 16.2–68.0pg/ml and 23.9–114.0pg/ml, respectively, while the ranges in premenopausal serum are 8.2–701.5pg/ml, 31.2–135.2pg/ml and 47.8–310.0pg/ml, respectively.

Premenopausal serum sex hormone levels in relation to breast cancer risk, overall and by hormone receptor status-Results from the EPIC cohort

Results from prospective studies on premenopausal serum hormone levels in relation to breast cancer risk have been inconclusive, especially with regard to tumor subtypes. Using a case-control study nested within the prospective European Prospective Investigation into Cancer and Nutrition (EPIC) cohort (801 breast cancer cases and 1,132 matched control subjects), we analyzed the relationships of prediagnostic serum estradiol, free estradiol, progesterone, testosterone, free testosterone and sex hormone-binding globulin (SHBG) levels with the risk of breast cancer by estrogen and progesterone receptor-positive and -negative breast tumors and by age at diagnoses. Higher prediagnostic serum levels of testosterone and free testosterone were associated with an increased overall risk of breast cancer [ORQ4-Q1 = 1.56 (95% CI 1.15-2.13), ptrend = 0.02 for testosterone and ORQ4-Q1 = 1.33 (95% CI 0.99-1.79), ptrend = 0.04 for free testosterone], but no significant risk association was observed for estradiol, free estradiol, progesterone and SHBG. Tests for heterogeneity between receptor-positive and -negative tumors were not significant. When analysis were stratified by age at tumor diagnosis, the odds ratios observed for estradiol were stronger and borderline significant for breast cancer diagnosed at age less than 50 [ORQ4-Q1 = 1.32 (95% CI 0.87-2.01), ptrend = 0.05] compared to breast cancer diagnosed at age 50 or above [ORQ4-Q1 = 0.94 (95% CI 0.60-1.47), ptrend = 0.34, phet = 0.04]. In conclusion, our data indicate that higher premenopausal circulating testosterone levels are associated with an increased risk of developing breast cancer, but do not show a significant association of estradiol or progesterone with breast cancer risk, overall, by menstrual cycle phase or by tumor receptor status, although a possible risk increase with higher estradiol levels for tumors diagnosed before age 50 was seen.

Efficacy of intravenously administered ibandronate in postmenopausal Korean women with insufficient response to orally administered bisphosphonates

We investigated rates of insufficient and over-responsiveness to orally administered bisphosphonates in postmenopausal women, and tested the efficacy of intravenous ibandronate in patients with insufficient response to orally administered bisphosphonates. Postmenopausal women were treated with either alendronate (70mg/week; n=88) or risedronate (35mg/week; n=84) for 1year, and their response to orally administered bisphosphonates was assessed using serum C-telopeptide (CTX) levels. Insufficient responders were changed to once-quarterly intravenous ibandronate 3mg injection (n=13) or maintained on orally administered bisphosphonates (n=19), according to patients' preference, for an additional 1year. There was no significant difference in baseline characteristics between two orally administered bisphosphonate groups except the bone mineral density values at the lumbar spine. Insufficient rate was higher in the risedronate group (19.0%) than in the alendronate group (8.0%), using the premenopausal serum CTX median as a cut-off (P=0.043). The over-response rate among the alendronate group (59.1%) was significantly higher than that in the risedronate group (38.1%), based on a serum CTX cut-off value of 0.100ng/ml (P=0.006). Intravenous ibandronate suppressed serum CTX levels to a significantly greater degree at 7days after the second dosing (0.191±0.110ng/mL; P<0.001) and 3months after the fourth dosing (0.274±0.159ng/mL; P=0.004) among insufficient responders, compared with post-oral/pre-intravenous levels (0.450±0.134ng/mL). Rates of insufficient and over-responsiveness to orally administered bisphosphonates were considerable, and a change to intravenous bisphosphonates may be considered in patients showing an insufficient response to orally administered bisphosphonates.

Premenopausal serum androgens and breast cancer risk: a nested case-control study.

IntroductionProspective epidemiologic studies have consistently shown that levels of circulating androgens in postmenopausal women are positively associated with breast cancer risk. However, data in premenopausal women are limited.MethodsA case-control study nested within the New York University Women's Health Study was conducted. A total of 356 cases (276 invasive and 80 in situ) and 683 individually-matched controls were included. Matching variables included age and date, phase, and day of menstrual cycle at blood donation. Testosterone, androstenedione, dehydroandrosterone sulfate (DHEAS) and sex hormone-binding globulin (SHBG) were measured using direct immunoassays. Free testosterone was calculated.ResultsPremenopausal serum testosterone and free testosterone concentrations were positively associated with breast cancer risk. In models adjusted for known risk factors of breast cancer, the odds ratios for increasing quintiles of testosterone were 1.0 (reference), 1.5 (95% confidence interval (CI), 0.9 to 2.3), 1.2 (95% CI, 0.7 to 1.9), 1.4 (95% CI, 0.9 to 2.3) and 1.8 (95% CI, 1.1 to 2.9; Ptrend = 0.04), and for free testosterone were 1.0 (reference), 1.2 (95% CI, 0.7 to 1.8), 1.5 (95% CI, 0.9 to 2.3), 1.5 (95% CI, 0.9 to 2.3), and 1.8 (95% CI, 1.1 to 2.8, Ptrend = 0.01). A marginally significant positive association was observed with androstenedione (P = 0.07), but no association with DHEAS or SHBG. Results were consistent in analyses stratified by tumor type (invasive, in situ), estrogen receptor status, age at blood donation, and menopausal status at diagnosis. Intra-class correlation coefficients for samples collected from 0.8 to 5.3 years apart (median 2 years) in 138 cases and 268 controls were greater than 0.7 for all biomarkers except for androstenedione (0.57 in controls).ConclusionsPremenopausal concentrations of testosterone and free testosterone are associated with breast cancer risk. Testosterone and free testosterone measurements are also highly reliable (that is, a single measurement is reflective of a woman's average level over time). Results from other prospective studies are consistent with our results. The impact of including testosterone or free testosterone in breast cancer risk prediction models for women between the ages of 40 and 50 years should be assessed. Improving risk prediction models for this age group could help decision making regarding both screening and chemoprevention of breast cancer.

Większe przedmenopauzalne stężenia androgenów a pomenopauzalne stężenia estrogenów w surowicy u kobiet z hiperplazją endometrium

Większe przedmenopauzalne stężenia androgenów a pomenopauzalne stężenia estrogenów w surowicy u kobiet z hiperplazją endometrium

Abstract A3: Premenopausal serum androgens and breast cancer risk: A nested case-control study

Abstract Introduction: Prospective epidemiologic studies have consistently shown that levels of circulating androgens in postmenopausal women are positively associated with breast cancer risk. However, data in premenopausal women are limited. Methods: A case-control study nested within a prospective cohort, the NYU Women's Health Study, was conducted. A total of 356 cases (276 invasive and 80 in situ) and 683 individually-matched controls (1:2 matching) were included. Matching variables included age and date of blood donation, as well as phase and day of menstrual cycle. Testosterone, androstenedione, dehydroandrosterone sulfate (DHEAS), and sex hormone-binding globulin (SHBG) were measured using direct radioimmunoassays. Free testosterone was calculated. Samples from a second blood donation at a median of 2 years after the first blood donation were also analyzed for 138 cases and 268 controls. Results: Premenopausal serum testosterone and free testosterone concentrations were positively associated with subsequent breast cancer risk. In models adjusted for known risk factors of breast cancer (BMI, age at menarche, family history of breast cancer, parity/age at first birth, and history of breast biopsy), the odds ratios for increasing quintiles of testosterone were 1.0 (reference), 1.5 (95% confidence interval (CI), 0.9–2.3), 1.2 (95% CI, 0.7–1.9), 1.4 (95% CI, 0.9–2.3) and 1.8 (95% CI, 1.1–2.9) with the P value for trend of 0.04, and for free testosterone were 1.0 (reference), 1.2 (95% CI, 0.7–1.8), 1.5 (95% CI, 0.9–2.3), 1.5 (95% CI, 0.9–2.3), and 1.8 (95% CI, 1.1–2.8) with a P value for trend of 0.01). A marginally significant positive association was observed with androstenedione (p = 0.07), but no association with DHEAS or SHBG. Results were consistent in analyses stratified by tumor type (invasive, in situ), estrogen receptor status, age at blood donation, and menopausal status at diagnosis. Intra-class correlation coefficients (ICCs) for measurements of serial annual blood donations from the same individual were greater than 0.7 for all biomarkers except for androstenedione (0.57 in controls). Conclusions: Premenopausal concentrations of testosterone and free testosterone are positively associated with breast cancer risk. Results from other prospective studies are consistent with our results. Testosterone and free testosterone are also highly reliable (i.e. a single measurement is reflective of a woman's average level over time). Future studies should examine the contribution of including testosterone and free testosterone in breast cancer risk prediction models for women between the ages of 40 and 50. Citation Information: Cancer Prev Res 2011;4(10 Suppl):A3.

Non-nuclear Estrogen Receptor Signaling in the Endothelium

In addition to the classical function of estrogen receptors (ER) as transcription factors, evidence continues to accumulate that they mediate non-nuclear processes in numerous cell types, including the endothelium, in which they activate endothelial NO synthase. Non-nuclear ER signaling entails unique post-translational modifications and protein-protein interactions of the receptor with adaptor molecules, kinases, and G proteins. Recent in vitro and in vivo studies in mice using an estrogen-dendrimer conjugate that is excluded from the nucleus indicate that non-nuclear ER activation underlies the migration and growth responses of endothelial cells to estrogen but not the growth responses of endometrial or breast cancer cells to the hormone. In this minireview, the features of ERα and protein-protein interactions that enable it to invoke extranuclear signaling in the endothelium and the consequences of that signaling are discussed.

Prospective case-control study of premenopausal serum estradiol and testosterone levels and breast cancer risk

IntroductionBreast cancer is frequently a hormonally dependent cancer, and associations of circulating estrogens and androgens with subsequent breast cancer risk are well established in postmenopausal women. Associations of serum estrogens and androgens with breast cancer risk in premenopausal women are less well studied. The objective of this study was to determine whether estradiol and testosterone levels in serum collected before menopause are associated with subsequent breast cancer risk.MethodsWe conducted a prospective case-control study of 266 participants who were registered in the Columbia, Missouri, Serum Bank and not using exogenous hormones at the time of blood collection. Each of 98 in situ or invasive breast cancer cases with prediagnostic serum collected before menopause was matched to two controls by age, date, menstrual cycle day, and time of day of blood collection. Estradiol and testosterone concentrations were quantified by using specific radioimmunoassays, and sex hormone-binding globulin (SHBG) was quantified with a chemiluminescent immunoassay to allow calculation of the non-SHBG bound hormone fractions. Data were analyzed by using conditional logistic regression. All tests of statistical significance were two-sided.ResultsSerum testosterone was strongly and significantly associated with breast cancer risk. The relative odds (OR) for increasing quartiles of total testosterone were 1.0, 2.1 (95% confidence interval (CI) 0.9 to 4.8), 1.5 (95% CI, 0.6 to 3.4), and 3.3 (95% CI, 1.5 to 7.5, Ptrend = 0.006). Comparable ORs for the non-SHBG bound fraction of testosterone that is bioavailable were 1.0, 1.7 (95% CI, 0.7 to 4.2), 1.7 (95% CI, 0.7 to 4.0), and 4.2 (95% CI, 1.6 to 10.9, Ptrend = 0.002). Total and non-SHBG-bound estradiol were not associated with breast cancer, but extreme variation in levels across the menstrual cycle coupled with relatively small numbers, particularly for analyses stratified by cycle phase, limited the power to detect associations.ConclusionsResults suggest that premenopausal women with elevated serum testosterone levels are at an increased risk of breast cancer.

Latest news and product developments

Latest news and product developments

Serum from Postmenopausal Women Directs Differentiation of Human Clonal Osteoprogenitor Cells from an Osteoblastic toward an Adipocytic Phenotype

A consistent observation in osteoporosis is bone volume reduction accompanied by increased marrow adipose tissue. No single cause linking the two phenomena has yet been identified. In a human progenitor cell clone (hOP 7) derived from bone marrow, however, we have demonstrated that rabbit serum can direct differentiation away from an osteoblast lineage to one of adipocytes. We now report on whether human serum has a similar effect. Serum was collected from 10 pre- and 10 postmenopausal women and from the 10 postmenopausal women before and following 6-week hormone replacement therapy (HRT). hOP 7 cells were cultured with the various sera, and after 7-14 days adipocytogenesis was determined by oil red O staining and lipoprotein lipase (LPL) and glycerol 3-phosphate dehydrogenase (G3PDH) expression. Incubation with 10% premenopausal serum led to labeling of 10.9% of cells (P < 0.05) with oil red O, whereas application of 10% postmenopausal serum led to a much larger effect, 43.5% labeling (P < 0.001 with respect to premenopausal serum). Oil red O positivity was accompanied by loss of type I collagen expression and increased LPL and G3PDH expression. HRT did not reverse the adipocytogenic effect of postmenopausal serum. In conclusion, serum from postmenopausal women contains factors that steer hOP 7 bone progenitor cells toward an adipocytic phenotype, irrespective of HRT. The study suggests a role for serum factors in the development of fatty marrow in postmenopausal osteoporosis.

Heart Disease Risk Determines Menopausal Age Rather Than the Reverse

The purpose of this study was to investigate whether a harmful cardiovascular risk profile accelerates menopause. Women with an early menopause are at an increased risk of cardiovascular disease. Although increased cardiovascular risk has been proposed as consequence of menopause, the alternative hypothesis, that increased premenopausal cardiovascular risk promotes early menopause, needs to be examined. We used data from the Framingham Heart Study cohort. This study started in 1948 and has followed up participants biennially since then. Women who were premenopausal at study entry and who reached natural menopause after at least two examination rounds were included in the study (n = 695). Premenopausal age-independent levels of serum total cholesterol, relative weight, blood pressure, and Framingham risk score were determined, as well as premenopausal changes in cholesterol, body weight, and blood pressure. A higher premenopausal serum total cholesterol level was statistically significantly associated with an earlier age at menopause, as were increases in total serum cholesterol, relative weight, and blood pressure in the premenopausal period. A decrease in total serum cholesterol during premenopause was statistically significantly associated with later age at menopause. Decreasing blood pressure was associated with a later menopausal age, but this association was not statistically significant. A decrease in relative weight was associated with a significant earlier age at menopause. Each 1% higher premenopausal Framingham risk score was associated with a decrease in menopausal age of 1.8 years (95% confidence interval -2.72 to -0.92). The findings support the view that heart disease risk determines age at menopause. This offers a novel explanation for the inconsistent findings on cardiovascular disease rate and its relationship to menopausal age and effects of hormone replacement therapy.

Serum Sex Steroids in Premenopausal Women and Breast Cancer Risk Within the European Prospective Investigation into Cancer and Nutrition (EPIC)

Contrasting etiologic hypotheses about the role of endogenous sex steroids in breast cancer development among premenopausal women implicate ovarian androgen excess and progesterone deficiency, estrogen excess, estrogen and progesterone excess, and both an excess or lack of adrenal androgens (dehydroepiandrosterone [DHEA] or its sulfate [DHEAS]) as risk factors. We conducted a case-control study nested within the European Prospective Investigation into Cancer and Nutrition cohort to examine associations among premenopausal serum concentrations of sex steroids and subsequent breast cancer risk. Levels of DHEAS, (Delta4-)androstenedione, testosterone, and sex hormone binding globulin (SHBG) were measured in single prediagnostic serum samples from 370 premenopausal women who subsequently developed breast cancer (case patients) and from 726 matched cancer-free control subjects. Levels of progesterone, estrone, and estradiol were also measured for the 285 case patients and 555 matched control subjects who had provided information about the day of menstrual cycle at blood donation. Conditional logistic regression models were used to estimate relative risks of breast cancer by quartiles of hormone concentrations. All statistical tests were two-sided. Increased risks of breast cancer were associated with elevated serum concentrations of testosterone (odds ratio [OR] for highest versus lowest quartile = 1.73, 95% confidence interval [CI] = 1.16 to 2.57; P(trend) = .01), androstenedione (OR for highest versus lowest quartile = 1.56, 95% CI = 1.05 to 2.32; P(trend) = .01), and DHEAS (OR for highest versus lowest quartile = 1.48, 95% CI = 1.02 to 2.14; P(trend) = .10) but not SHBG. Elevated serum progesterone concentrations were associated with a statistically significant reduction in breast cancer risk (OR for highest versus lowest quartile = 0.61, 95% CI = 0.38 to 0.98; P(trend) = .06). The absolute risk of breast cancer for women younger than 40 followed up for 10 years was estimated at 2.6% for those in the highest quartile of serum testosterone versus 1.5% for those in the lowest quartile; for the highest and lowest quartiles of progesterone, these estimates were 1.7% and 2.6%, respectively. Breast cancer risk was not statistically significantly associated with serum levels of the other hormones. Our results support the hypothesis that elevated blood concentrations of androgens are associated with an increased risk of breast cancer in premenopausal women.

Vaginal pH as a marker for bacterial pathogens and menopausal status

OBJECTIVES: Our purpose was to confirm the elevation of vaginal pH expected in patients with bacterial pathogens in premenopausal women and to examine the relationship of serum follicle-stimulating hormone and estradiol levels to vaginal pH in menopausal patients without and with hormone replacement therapy. STUDY DESIGN: Vaginal pH was determined by phenaphthazine (Nitrazine) pH paper in 253 patients seen in a solo private practice for routine speculum examination. None of the patients were pregnant. Measurements were made of serum levels of follicle-stimulating hormone and estradiol for 172 patients and vaginal cultures were taken from 82 patients. Vaginal pH was correlated with vaginal cultures and serum follicle-stimulating hormone and estradiol levels by use of statistical analysis. RESULTS: Vaginal pH was elevated in all premenopausal patients with documented bacterial pathogens. Serum estradiol levels showed an inverse and serum follicle-stimulating hormone levels a direct statistical correlation with vaginal pH in menopausal patients. CONCLUSIONS: Measurement of vaginal pH is useful, effective, and inexpensive for screening purposes. A vaginal pH of 4.5 is consistent with a premenopausal serum estradiol level and the absence of bacterial pathogens. An elevated vaginal pH in the 5.0 to 6.5 range suggests a diagnosis of either bacterial pathogens or decreased serum estradiol. In patients with an elevated pH, vaginal culture should establish the diagnosis. In the absence of bacterial pathogens, a vaginal pH of 6.0 to 7.5 is strongly suggestive of menopause. Titration of estradiol level by vaginal pH during estrogen replacement therapy may help menopausal women avoid side effects or cessation of therapy. (Am J Obstet Gynecol 1997;176:1270-7.)

An audit of oestradiol levels and implant frequency in women undergoing subcutaneous implant therapy

The aim of the study was to review our long-term use of subcutaneous oestradiol (E2) implant therapy for the treatment of climacteric symptoms in post-menopausal women. On the grounds that the aim is to restore premenopausal serum E2 levels, our declared clinical policy is not to repeat implants even in the presence of symptoms if serum E2 levels are > 400 pmol/l. Therapy was with 50 mg E2 implants inserted subcutaneously in the lower abdominal wall. All women who had attended the gynaecological/endocrinological clinic who had received subcutaneous E2 implants for the relief of climacteric symptoms between December 1981 and 1992 were included. Between December 1981 and December 1992, 275 women received a total of 759 50 mg E2 implants. The median length of implant therapy was 34.2 months (range 3.7-109.5 months), and the median number of implants per patient was 4 and ranged from 1 to 13. One hundred and twenty-nine women had more than four implants and their mean recorded serum E2 level was 425 +/- 187 (mean +/- SD) pmol/l; the mean level over the first 24 months of therapy was 408 +/- 157 pmol/l. This was not different from the mean value of the remaining period of therapy (439 +/- 168 pmol/l). Following the second implant there was no significant progressive rise in serum E2 with time and implant number and the mean E2 level per patient was no higher in those patients who received implants more frequently. The mean time between the first two implants was 9.7 +/- 0.4 months and between subsequent ones was 11.7 +/- 0.5 months. After the first two implants there was no progressive change in this interval with time. This study shows that effective, safe and sympathetic management of women with oestrogen deficient symptoms may be achieved by use of two criteria to determine re-treatment; the return of symptoms, and a serum E2 level no higher than 400 pmol/l. Once therapy is established, E2 implants may need to be prescribed only on an annual basis. There appears to be no justification for giving E2 implants more frequently as this policy achieves satisfactory (physiological) premenopausal E2 levels and good symptomatic relief without any evidence for accumulation of E2 or 'tachyphylaxis'.

Premenopausal estradiol levels and the risk of breast cancer: a new method of controlling for day of the menstrual cycle.

Levels of total estradiol in premenopausal women vary widely over the course of the menstrual cycle with a spike at the time of ovulation and dissimilar patterns pre- and post-ovulation. Evaluating the association between breast cancer and premenopausal measurements of total estradiol when the measurements cannot be taken on a uniform day of the cycle is therefore a difficult methodological challenge. In a matched case-control study of breast cancer nested within a prospective study, premenopausal serum samples obtained up to 7 years before breast cancer diagnosis were available for total estradiol assay. By fitting a three-piece spline model that regressed the logarithm of total estradiol (ln estradiol) on day of menstrual cycle, the authors were able to adjust the measurements for day of the cycle on which they were collected by expressing them in terms of the number of standard deviations above or below the fitted ln estradiol value for that day. Applying the adjusted measurements to the nested case-control study, they found evidence of a 1.5 to 2-fold risk for women in the upper two tertiles of ln estradiol relative to women in the lowest tertile. Conditional logistic regression analysis for day-of-cycle-adjusted ln estradiol treated as a continuous variable resulted in a relative risk estimate of 1.19 (95% confidence interval 0.91-1.55) per standard-deviation increase in adjusted ln estradiol.

An algorithm for testing and reporting serum choriogonadotropin at clinically significant decision levels with use of "pregnancy test" reagents.

We present an algorithm for monitoring the concentration of human choriogonadotropin (hCG) in serum at various clinical decision levels with use of fast, simple, and cost-effective qualitative pregnancy test reagents for hCG in serum. Based on correlation between laboratory data and clinical observations described in the literature, the following concentrations of hCG in serum can be considered as clinically significant decision levels: 5 int. units/L to exclude or "rule out" the presence of increased hCG; 25 int. units/L for "confirming pregnancy" or confirming the presence of increased hCG from sources other than the trophoblast; and 6500 and 82 500 int. units/L to enhance the sonographic diagnoses of ectopic pregnancies and molar pregnancies, respectively. We used Tandem Icon II (Hybritech) pregnancy test reagents and evaluated the reagents for possible "false-positive" findings at the 25 int. units/L limit of hCG detectability by analyzing 100 post-menopausal and more than 4000 premenopausal serum specimens. The performance of the reagents was validated also at 5 int. units/L and at limits of hCG detectability greater than 25 int. units/L.