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Context: Chronic supraphysiological glucocorticoid therapy controls the androgen excess of 21-hydroxylase deficiency (21OHD) but contributes to the high prevalence of obesity, glucose intolerance, and reduced bone mass in these patients. Abiraterone acetate (AA) is a prodrug for abiraterone, a potent CYP17A1 inhibitor used to suppress androgens in the treatment of prostate cancer., Objective: The objective of the study was to test the hypothesis that AA added to physiological hydrocortisone and 9α-fludrocortisone acetate corrects androgen excess in women with 21OHD without causing hypertension or hypokalemia., Design: This was a phase 1 dose-escalation study., Setting: The study was conducted at university clinical research centers., Participants: We screened 14 women with classic 21OHD taking hydrocortisone 12.5-20 mg/d to enroll six participants with serum androstenedione greater than 345 ng/dL (>12 nmol/L)., Intervention: AA was administered for 6 days at 100 or 250 mg every morning with 20 mg/d hydrocortisone and 9α-fludrocortisone acetate., Main Outcome Measure: The primary endpoint was normalization of mean predose androstenedione on days 6 and 7 (< 230 ng/dL [<8 nmol/L)] in greater than 80% of participants. Secondary end points included serum 17-hydroxyprogesterone and testosterone (T), electrolytes, plasma renin activity, and urine androsterone and etiocholanolone glucuronides., Results: With 100 mg/d AA, mean predose androstenedione fell from 764 to 254 ng/dL (26.7-8.9 nmol/L). At 250 mg/d AA, mean androstenedione normalized in five participants (83%) and decreased from 664 to 126 ng/dL (23.2-4.4 nmol/L), meeting the primary end point. Mean androstenedione declined further during day 6 to 66 and 38 ng/dL (2.3 and 1.3 nmol/L) at 100 and 250 mg/d, respectively. Serum T and urinary metabolites declined similarly. Abiraterone exposure was strongly negatively correlated with mean androstenedione. Hypertension and hypokalemia were not observed., Conclusion: AA 100-250 mg/d added to replacement hydrocortisone normalized several measures of androgen excess in women with classic 21OHD and elevated serum androstenedione.

Context: Polycystic ovary syndrome (PCOS) is a triad of anovulation, insulin resistance, and hyperandrogenism. Androgen excess may correlate with metabolic risk and PCOS consensus criteria define androgen excess on the basis of serum T. Here we studied the utility of the androgen precursor serum androstenedione (A) in conjunction with serum T for predicting metabolic dysfunction in PCOS., Patients and Methods: Eighty-six PCOS patients fulfilling Rotterdam diagnostic consensus criteria and 43 age- and body mass index-matched controls underwent measurement of serum androgens by tandem mass spectrometry and an oral glucose tolerance test with homeostatic model assessment of insulin resistance and insulin sensitivity index calculation. We analyzed 24-hour urine androgen excretion by gas chromatography/mass spectrometry., Results: PCOS patients had higher levels of serum androgens and urinary androgen metabolites than controls (all P < .001). Within the PCOS cohort, both serum A and T were positively correlated with the free androgen index (T × 100/SHBG) and total androgen metabolite excretion (all P < .001). All subjects with T above the normal reference range [high T (HT)] also had high A (HA/HT group, n = 56). However, the remaining 30 patients had normal T levels, either in the presence of HA (HA/NT; n = 20) or normal A (NA/NT; n = 10). The groups did not differ in age or BMI. The HA/HT and HA/NT groups had higher total androgen excretion than NA/NT (P < .01 and P < .05, respectively). Multiple linear regression showed a strong negative association between serum androstenedione and insulin sensitivity. The incidence of dysglycemia according to an oral glucose tolerance test increased with the severity of androgen phenotype (NA/NT, 0%; HA/NT, 14%; HA/HT, 25%, P = .03)., Conclusion: Simultaneous measurement of serum T and A represents a useful tool for predicting metabolic risk in PCOS women. HA levels are a sensitive indicator of PCOS-related androgen excess.

The steroidogenic enzyme 21-hydroxylase is necessary for the synthesis of both glucocorticoids and mineralocorticoids. 21-hydroxylase is a cytochrome P-450 enzyme and is encoded by the gene CYP21A2. Here we report a 68-year-old phenotypically 'male' but genetically female patient with 21-hydroxylase deficiency (21OHD) and the concomitant virilizing adrenocortical carcinoma. This patient grew up as a male and has not encountered any episodes of adrenal insufficiency without glucocorticoid replacement in his lifetime. A chromosome test at admission, however, identified the 46, XX karyotype, and serum 17-hydroxyprogesterone and urine pregnanetriolone and 11β-hydroxyandrostendione were all elevated, consistent with 21OHD. Moreover, serum testosterone was 1.90 ng/ml, much higher than the female standard levels, and serum cortisol was 5.7 µg/ml, slightly lower than standard levels. Genetic analysis identified the patient as a heterozygote of the two pathogenic mutations in the CYP21A2 gene: IVS2-13C(A)>G and R356W. Magnetic resonance imaging (MRI) revealed the presence of left adrenal tumor measuring 6 cm, which was subsequently diagnosed as adrenocortical carcinoma based on the criteria of Weiss. Immunohistochemical analysis of the tumor specimens revealed the expression of various enzymes involved in testosterone production, including 3β-hydroxysteroid dehydrogenase, 17α-hydroxylase/17,20-lyase, and 17β-hydroxysteroid dehydrogenase. Importantly, the expression of immunoreactive 21-hydroxylase was detected in these tumor cells. The levels of adrenal tumor-derived steroid metabolites were all markedly decreased following the surgery. This is the first report on a virilized 21OHD patient associated with the adrenocortical tumor that produces testosterone. Moreover, the concomitant adrenocortical tumor may ameliorate adrenocortical insufficiency by producing cortisol.

This study describes an on-line stacking CE approach by sweeping with whole capillary sample filling for analyzing five anabolic androgenic steroids in urine samples. The five anabolic steroids for detection were androstenedione, testosterone, epitestosterone, boldenone, and clostebol. Anabolic androgenic steroids are abused in sport doping because they can promote muscle growth. Therefore, a sensitive detection method is imperatively required for monitoring the urine samples of athletes. In this research, an interesting and reliable stacking capillary electrophoresis method was established for analysis of anabolic steroids in urine. After liquid-liquid extraction by n-hexane, the supernatant was dried and reconstituted with 30 mM phosphate buffer (pH 5.00) and loaded into the capillary by hydrodynamic injection (10 psi, 99.9 s). The stacking and separation were simultaneously accomplished at -20 kV in phosphate buffer (30 mM, pH 5.0) containing 100 mM sodium dodecyl sulfate and 40 % methanol. During the method validation, calibration curves were linear (r≥0.990) over a range of 50-1,000 ng/mL for the five analytes. In the evaluation of precision and accuracy for this method, the absolute values of the RSD and the RE in the intra-day (n=3) and inter-day (n=5) analyses were all less than 6.6 %. The limit of detection for the five analytes was 30 ng/mL (S/N=5, sampling 99.9 s at 10 psi). Compared with simple MECK, this stacking method possessed a 108- to 175-fold increase in sensitivity. This simple and sensitive stacking method could be used as a powerful tool for monitoring the illegal use of doping.

Because of their perceived and reported effects on self-image, muscle development, performance, and similar factors, anabolic-androgenic steroids (AAS) and their precursors are among the most abused substances by professional, amateur, and recreational athletes. However, AAS abuse is not limited to athletes, but is also prevalent in the workplace, especially those professions in which image, strength, and endurance are coveted attributes. The detection of many steroids in biological specimens is analogous to the detection of an abused drug such as cocaine. Identification of the parent drug or its characteristic metabolite(s) in a donor's sample with a drug screening technique and confirmation of the drug/metabolite with a suitable alternative technology provides evidence of use. These analyses and subsequent interpretive scenarios become far more complex when the ingested AAS is an endogenous compound such as dehydroepiandrosterone (DHEA), androstenedione (Adione), or dihydrotestosterone (DHT). These compounds and their metabolites are present in specimens such as urine as a course of our natural endocrine function. Therefore, it becomes much more challenging for the laboratory to establish testing and interpretative paradigms that can distinguish "normal" urinary profiles of these steroids and their metabolites from profiles indicative of exogenous use. Distinguishing "normal" from "abnormal" urine profiles is particularly challenging during screening when literally tens of steroids and their metabolites may be tested simultaneously in a single chromatographic analysis. The purpose of this paper is to review the relevant literature about DHEA, Adione, and DHT administration, detection, and interpretation specifically as it relates to changes in the urinary AAS profile that may be identified during the routine laboratory screening of donor urine specimens.

Background: To evaluate the metabolic changes in urinary steroids in pre- and post-menopausal women and men with papillary thyroid carcinoma (PTC)., Methods: Quantitative steroid profiling combined with gas chromatography-mass spectrometry was used to measure the urinary concentrations of 84 steroids in both pre- (n = 21, age: 36.95 ± 7.19 yr) and post-menopausal female (n = 19, age: 52.79 ± 7.66 yr), and male (n = 16, age: 41.88 ± 8.48 yr) patients with PTC. After comparing the quantitative data of the patients with their corresponding controls (pre-menopause women: n = 24, age: 33.21 ± 10.48 yr, post-menopause women: n = 16, age: 49.67 ± 8.94 yr, male: n = 20, age: 42.75 ± 4.22 yr), the levels of steroids in the patients were normalized to the mean concentration of the controls to exclude gender and menopausal variations., Results: Many urinary steroids were up-regulated in all PTC patients compared to the controls. Among them, the levels of three active androgens, androstenedione, androstenediol and 16α-hydroxy DHEA, were significantly higher in the pre-menopausal women and men with PTC. The corticoid levels were increased slightly in the PTC men, while progestins were not altered in the post-menopausal PTC women. Estrogens were up-regulated in all PTC patients but 2-hydroxyestrone and 2-hydroxy-17β-estradiol were remarkably changed in both pre-menopausal women and men with PTC. For both menopausal and gender differences, the 2-hydroxylation, 4-hydroxylation, 2-methoxylation, and 4-methoxylation of estrogens and 16α-hydroxylation of DHEA were differentiated between pre- and post-menopausal PTC women (P < 0.001). In particular, the metabolic ratio of 2-hydroxyestrone to 2-hydroxy-17β-estradiol, which could reveal the enzyme activity of 17β-hydroxysteroid dehydrogenase, showed gender differences in PTC patients (P < 1 × 10-7)., Conclusions: These results are expected be helpful for better understanding the pathogenic differences in PTC according to gender and menopausal conditions.

Background: Congenital adrenal hyperplasia is a group of disorders caused by defects in the adrenal steroidogenic pathways. In its most common form, 21-hydroxylase deficiency, patients develop varying degrees of glucocorticoid and mineralocorticoid deficiency as well as androgen excess. Therapy is guided by monitoring clinical parameters as well as adrenal hormone and metabolite concentrations., Content: We review the evidence for clinical and biochemical parameters used in monitoring therapy for congenital adrenal hyperplasia. We discuss the utility of 24-h urine collections for pregnanetriol and 17-ketosteroids as well as serum measurements of 17-hydroxyprogesterone, androstenedione, and testosterone. In addition, we examine the added value of daily hormonal profiles obtained from salivary or blood-spot samples and discuss the limitations of the various assays., Summary: Clinical parameters such as growth velocity and bone age remain the gold standard for monitoring the adequacy of therapy in congenital adrenal hyperplasia. The use of 24-h urine collections for pregnanetriol and 17-ketosteroid may offer an integrated view of adrenal hormone production but target concentrations must be better defined. Random serum hormone measurements are of little value and fluctuate with time of day and timing relative to glucocorticoid administration. Assays of daily hormonal profiles from saliva or blood spots offer a more detailed assessment of therapeutic control, although salivary assays have variable quality.

Introduction: Quantities of various anabolic/androgenic steroids have been found in dietary supplements without their presence being disclosed on the label. The aim of this study was to quantify the excretion patterns of the diagnostic metabolites, 19-norandrosterone (19-NA), and 19-noretiocholanolone (19-NE) after ingestion of small doses of 19-nor-4-androstene-3,17-dione (19-norandrostenedione)., Methods: Eleven males and nine females entered the laboratory in the morning after an overnight fast. An initial urine sample was collected, and volunteers then ingested 500 mL of water containing 5 g of creatine monohydrate and 1.0, 2.5, or 5.0 microg of 19-norandrostenedione. The volume of each urine void was measured, and an aliquot was taken. Samples were analyzed for the metabolites 19-NA and 19-NE by GCMS., Results: Baseline urinary 19-NA concentrations were 0.19 +/- 0.14 ng x mL. Ingestion of the supplement resulted in peak mean urinary 19-NA concentrations of 0.68 +/- 0.36, 1.56 +/- 0.86, and 3.89 +/- 3.11 ng.mL in the 1.0-, 2.5-, or 5.0-microg trials, respectively. Under current WADA regulations, ingestion of the 1.0-microg dose produced 0 positive doping tests, 5 subjects (20%) tested positive in the 2.5-microg trial, and 15 subjects (75%) had urinary 19-NA concentrations exceeding 2 ng x mL after ingesting creatine containing 5.0 microg of the steroid. The recovery of the ingested dose was highly variable between individuals, with values ranging from 11% to 84% (mean +/- SD = 47% +/- 18%)., Conclusions: Ingestion of trace amounts of 19-norandrostenedione can result in transient elevations of urinary 19-NA and 19-NE concentrations. The addition of as little as 2.5 microg of 19-norandrostenedione to a supplement (0.00005% contamination) appears sufficient to result in a doping violation in some individuals.

Progesterone (P) is a strong mineralocorticoid receptor (MR) antagonist in vitro. The high P concentrations seen in normal pregnancy only moderately increase renin and aldosterone concentrations. In previous in vitro studies we hypothesized that this may be explained by intrarenal conversion of P to less potent metabolites. To investigate the in vivo anti-MR potency of P, we performed an infusion study in patients with adrenal insufficiency (n = 8). They omitted 9alpha-fluorocortisol for 4 d and hydrocortisone for 0.5 d before a continuous iv infusion of aldosterone for 8.5 h, with an additional iv P infusion commenced at 4 h. During aldosterone infusions the initially elevated urinary sodium to potassium ratio decreased significantly. Despite the 1000-fold excess of P over aldosterone, the urinary sodium to potassium ratio and urinary sodium excretion increased only slightly after 3 h of P infusion. We detected inhibition of renal 11beta-hydroxysteroid dehydrogenase type 2 by P, thus giving cortisol/prednisolone access to the MR. Urinary and plasma concentrations of 17alpha-hydroxyprogesterone, a major metabolite of renal P metabolism, and those of serum androstenedione and deoxycorticosterone, a mineralocorticoid itself, increased significantly during P infusion. This supports the hypothesis of an effective protection of the MR from P by efficient extraadrenal downstream conversion of P.

Dehydroepiandrosterone (DHEA) and androstenedione are weak androgens, which need conversion to more potent testosterone in order to enhance anabolic action. Consequences of oral dosing at 1 mg/kg on the urinary and plasma androgen profile of mare and gelding have been evaluated with an analytical method involving conjugate fractionation and selective hydrolysis, group separation, and quantitation by gas chromatography-mass spectrometry with selected ion monitoring of trimethylsilyl ethers. Peak levels of testosterone total conjugates in urine (range 300-6000 microg/L) were attained a few hours after dosing. Renal clearance was fast, so the testosterone detection period lasted only 20 to 33 h, the longest time being generated by androstenedione. The urinary testosterone/epitestosterone ratio for detection of exogenous testosterone in the mare was inoperative after DHEA administration because there was a concomitant increase of epitestosterone, which thereby acted as a masking agent. Androstanediols and androstenediols, as well as some 17-ketosteroids, were additional markers. A transient increase of circulating free testosterone has been evidenced, and this would support possible anabolic/androgenic action by supplementation with DHEA and androstenedione along the oral route.

Androstenedione is a steroid hormone and the major precursor to testosterone. It is available without prescription and taken with the expectation that it will be converted to testosterone endogenously and increase strength and athletic performance. The metabolism of orally administered testosterone has not been well studied. We randomly assigned 37 healthy men to receive 0, 100, or 300 mg oral androstenedione in a single daily dose for 7 d. Single 8-h urine collections were performed on the day before the start of the androstenedione administration and on d 1 and 7 to assess excretion rates of free and glucuronide- conjugated testosterone, androsterone, etiocholanolone, and dihydrotestosterone. Serum testosterone glucuronide concentrations were measured by frequent blood sampling over 8 h on d 1 in 16 subjects (5 each in the 0 and 100 mg group and 6 in the 300 mg group). In the control group, mean (+/-SE) d 1 and 7 excretion rates for testosterone, androsterone, etiocholanolone, and dihydrotestosterone were 3 +/- 1, 215 +/- 26, 175 +/- 26, and 0.4 +/- 0.1 microg/h, respectively. In the 100 mg group, mean d 1 and 7 excretion rates for testosterone, androsterone, etiocholanolone, and dihydrotestosterone were 47 +/- 11, 3,836 +/- 458, 4,306 +/- 458, and 1.6 +/- 0.2 microg/h, respectively. In the 300 mg group, mean d 1 and 7 excretion rates for testosterone, androsterone, etiocholanolone, and dihydrotestosterone were 115 +/- 39, 8,142 +/- 1,362, 10,070 +/- 1,999, and 7.7 +/- 1.5 microg/h, respectively. Urinary excretion rates of all metabolites were greater in both the 100 and 300 mg groups than in controls (P < 0.0001). Urinary excretion rates of testosterone (P = 0.007), androsterone (P = 0.009), etiocholanolone (P = 0.0005), and dihydrotestosterone (P < 0.0001) were greater in the subjects who received 300 mg androstenedione than in those who received 100 mg. In the treated groups, excretion of free testosterone accounted for less than 0.1% of the total excreted testosterone measured. Serum testosterone glucuronide levels increased significantly during frequent blood sampling in both the 100 and 300 mg groups compared with controls (P = 0.0005 for the 100 mg group; P < 0.0001 for the 300 mg group). The net mean changes in area under the curve for serum testosterone glucuronide were -18 +/- 25%, 579 +/- 572%, and 1267 +/- 1675% in the groups receiving 0, 100, and 300 mg/d androstenedione, respectively. We conclude that the administration of both 100 and 300 mg androstenedione increases the excretion rates of conjugated testosterone, androsterone, etiocholanolone, and dihydrotestosterone and the serum levels of testosterone glucuronide in men. The magnitude of these increases is much greater than the changes observed in serum total testosterone concentrations. These findings demonstrate that orally administered androstenedione is largely metabolized to testosterone glucuronide and other androgen metabolites before release into the general circulation.

Since the appearance of 4-androsten-3,17-dione (I) as a nutritional supplement in early 1997, we have frequently observed a characteristic deterioration of endogenous steroid profiles in athletes' urine in routine anabolic steroid testing in which concentrations of major endogenous urinary steroids and testosterone exceed normal. Human excretion studies are performed with I and newer, over-the-counter "supplements" 4-androsten-3beta,17beta-diol (II) and 19-nor-4-androsten-3,17-dione (III). Endogenous urinary steroids affected by I and II are androsterone, etiocholanolone, their hydroxylated derivatives 5alpha- and 5beta-androstan-3alpha,17beta-diols, testosterone, and epitestosterone. Their concentrations briefly increase by one to two orders of magnitude and return to normal 24 h after oral administration of I and II. The average male may test positive for testosterone because testosterone concentration rises faster than that of epitestosterone, causing the testosterone/epitestosterone (T/E) ratio to rise above the positive cutoff of 6:1. A remarkable distinction in excretion patterns was observed in eastern Asian men, for whom I and II did not affect urinary concentrations of testosterone and did not increase the T/E ratio. First-pass metabolism deactivates most of the orally administered drugs I and II, rapidly converting them into inactive androsterone and etiocholanolone. Drug II is a more effective testosterone booster because of its different metabolic pathway. After the use of III, a precursor of the potent anabolic nandrolone, high concentrations of norandrosterone and noretiocholanolone appear in urine, similar to nandrolone. These are detectable in urine for 7-10 days after a single oral dose of III (50 mg).