Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency is a disorder which can adopt three clinical expressions: two classical forms -salt-wasting (SW), with residual enzymatic activity (EA) < or = 1% and simple virilizing (SV), with EA 1-2%- and a mild late onset or nonclassical (NC) form, with EA 10-60%. Our objective is to describe clinical characteristics, growth, and bone mass in a group of patients affected by 21-hydroxylase deficiency. Besides, molecular genetics studies were performed in patients, and also when available in their parents and siblings. Nine patients with neonatal diagnosis and 8 with pre or postpubertal diagnosis were studied. Analyses of 10-point mutations in the CYP21A2 gene were performed. We found that all the patients with the classical expression, except one with a de novo mutation R356W in one allele, were fully genotyped with predictive < 2% EA mutations. Signs of hyperandrogenism were present in 5/6 NC patients; one was diagnosed by searching for mutations in asymptomatic siblings. All the NC patients were compound heterozygotes carrying V281L mutation in one allele and a predictive low EA in the other, except for one not yet determined. In patients with neonatal diagnosis, mean height was low at one year of age, though it showed a significant increase before the onset of puberty. We conclude that neonatal diagnosis of classical CAH allows an adequate follow up enhancing growth. Molecular analyses of all members of an affected family may disclose asymptomatic patients. The presence of de novo mutations, as well as, the presence of mutations with low predicted EA in NC patients reinforces the importance of genotyping for appropriate genetic counseling. In fully genotyped NC patients, the lowest value of ACTH-stimulated <em>17OHP</em> was 14 ng/ml. Lower cut-off values might overestimate the diagnosis of the NC form.

We investigated the associations between smoking status and metabolic risk factors and sex hormones in polycystic ovary syndrome (PCOS). The study was designed as a retrospective trans-sectional study including 650 white premenopausal women with the diagnoses hirsutism or PCOS divided according to smoking status: non-smokers (NS-PCOS = 390) and smokers (S-PCOS = 260). One hundred and nineteen healthy women were studied as controls (NS-Control = 105, S-Control = 14). Patients and controls underwent clinical evaluation, hormone analyses, transvaginal ultrasound, oral glucose tolerance tests (OGTT) and adrenocorticotropic hormone (ACTH) tests. S-PCOS has significantly higher fasting lipid profile and 17-hydroxyprogesterone levels (basal and ACTH-stimulated) than NS-PCOS patients, whereas prolactin levels were decreased. No significant differences were found in body composition and measures of insulin resistance between NS-PCOS and S-PCOS. PCO was more prevalent in NS-PCOS patients. During multiple regression analyses, smoking was positively associated with 17-hydroxyprogesterone (<em>17OHP</em>) and cholesterol, triglycerides and low-density lipoprotein and inversely associated with prolactin and high-density lipoprotein. We concluded that smoking was associated with increased adrenal responsiveness, decreased prolactin levels and a more adverse lipid profile in PCOS patients, whereas smoking was unassociated with body composition and insulin resistance. Smoking may be associated with the prevalence of individual Rotterdam criteria.

OBJECTIVE

Several cases of testicular adrenal rest tumours have been reported in men with congenital adrenal hyperplasia (CAH) due to the classical form of 21-hydroxylase deficiency but the prevalence has not been established. The aims of this report were to evaluate the frequency of testicular adrenal rest tissue in this population in a retrospective multicentre study involving eight endocrinology centres, and to determine whether treatment or genetic background had an impact on the occurrence of adrenal rest tissue.

METHODS

Testicular adrenal rest tissue (TART) was sought clinically and with ultrasound examination in forty-five males with CAH due to the classical form of 21-hydroxylase deficiency. When the diagnosis of testicular adrenal rest tumours was sought, good observance of treatment was judged on biological concentrations of 17-hydroxyprogesterone (<em>17OHP</em>), delta4-androstenedione, active renin and testosterone. The results of affected and non-affected subjects were compared.

RESULTS

TART was detected in none of the 18 subjects aged 1 to 15years but was detected in 14 of the 27 subjects aged more than 15years. Five patients with an abnormal echography result had no clinical signs. Therapeutic control evaluated at diagnosis of TART seemed less effective when diagnosis was made in patients with adrenal rest tissue compared to TART-free subjects. Various genotypes were observed in patients with or without TART.

CONCLUSIONS

Due to the high prevalence of TART in classical CAH and the delayed clinical diagnosis, testicular ultrasonography must be performed before puberty and thereafter regularly during adulthood even if the clinical examination is normal.

OBJECTIVE

To investigate whether elevated ACTH-stimulated 17-hydroxyprogesterone (<em>17OHP</em>) levels are caused by CYP21 genetic defects or by a general adrenal hyperresponsiveness in hirsute patients.

METHODS

A total of 337 hirsute patients were evaluated by Ferriman-Gallwey score, serum testosterone, ACTH-stimulated <em>17OHP</em>, and cortisol during the follicular phase. A cutoff value of 16 nmol/liter for maximum ACTH-stimulated <em>17OHP</em> (M<em>17OHP</em>) responses was defined as the upper limit of the 95% confidence interval (CI) for the 97.5 percentile in 42 female controls. All patients were offered total screening of the CYP21 gene, and 252 healthy, premenopausal women with regular menses underwent genetic screening.

RESULTS

Patients were divided into idiopathic hirsutism (IH) (n = 180) and polycystic ovary syndrome (PCOS) (n = 157) groups. M<em>17OHP</em> levels were significantly higher in IH [geometric mean value (nmol/liter +/- 2 sd) 12.2 (4.6-32.3)] and PCOS [11.9 (5.3-27.2)] compared with controls [8.5 (5.1-14.2)] (P < 0.001). A similar percentage of IH and PCOS patients had elevated M<em>17OHP</em> (20.5 vs. 20.8%, not significant), and these also had significantly higher 30-min cortisol levels compared with controls (P < 0.05). The prevalence of CYP21 mutations in patients was 8.6% compared with 6.3% in controls (P = 0.38). Ten of 19 carriers had M<em>17OHP</em> levels below the cutoff limit.

CONCLUSIONS

The significantly higher ACTH-stimulated levels of cortisol and <em>17OHP</em> in hirsute patients indicated adrenal hyperresponsiveness in IH and PCOS. CYP21-carrier status could not explain the observed high prevalence of abnormal ACTH-stimulated <em>17OHP</em> levels in the hirsute population.

OBJECTIVE

Evaluate the Neonatal Screening Program (NSP) for congenital adrenal hyperplasia (CAH) of the Department of Health of the State of Santa Catarina (Secretaria de Estado da Saúde de Santa Catarina, SES/SC), and provide information to improve the program.

METHODS

Descriptive, retrospective study of 748,395 children screened between January 2001 and December 2010. We analyzed the coverage of the NSP-SES/SC prevalence of CAH, child's age when the first sample for 17-hydroxyprogesterone (<em>17OHP</em>) measurement was collected, levels of <em>17OHP</em>, mean age at treatment onset and main clinical manifestations.

RESULTS

The NSP-SES/SC covered 89% of the live newborns in the State. It diagnosed 50 cases of CAH, yielding an incidence of 1:14,967. Mean age at collection of the first sample was 7.3 days and mean level of <em>17OHP</em> was 152.9 ng/mL. The most frequent manifestations were virilized genitalia with nonpalpable gonads, clitoromegaly and genital hyperpigmentation. In three girls, the genre established at birth was incorrect. The salt-wasting form was present in 74% of the cases. There was no occurrence of shock or death. Mean age at treatment onset in the salt-wasting form was 17.4 days compared with 54.9 days in those without the salt-wasting form of the disease. All children were treated with hydrocortisone, and those with salt-wasting CAH were also treated with fludrocortisone.

CONCLUSIONS

The incidence of CAH was 1 case to 14,967 live newborns. Collection of the first sample occurred outside the recommended time, resulting in delays in treatment onset.

The currently used cutoff level for ACTH-stimulated 17- hydroxyprogesterone (<em>17OHP</em>) for the diagnosis of the nonclassical (NC) form of 21-hydroxylase deficiency (21OHD), established before molecular studies, is based on the mean + 2 SD of <em>17OHP</em> levels of obligate heterozygotes. However, carriers of CYP21 mutations present variable ACTH-stimulated <em>17OHP</em> levels, ranging from normal values up to 30 nmol/liter. The aim of this study was to determine whether ACTH-stimulated <em>17OHP</em> levels in obligate carriers for 21OHD would be correlated with the impairment of the enzyme activity caused by these mutations, which would affect the <em>17OHP</em> cutoff level for the diagnosis of the NC form. Fifty-nine parents of patients with the classical and NC forms of 21OHD had their DNA screened for the mutations found in the index case and were divided into three mutation groups according to the impairment of enzyme activity (A = 0%, B = 3%, and C>> 20%). All parents carried mutations in one allele (29 of group A, 9 of group B, and 21 of group C). Blood samples were collected at baseline condition and 60 min after ACTH (250 microg i.v.) to measure <em>17OHP</em> levels. The levels among groups A, B, and C were compared using the Kruskall Wallis test. ACTH-stimulated <em>17OHP</em> levels identified 39% of the carriers (9 in group A, 2 in group B and 12 in group C). The mean +/- SD basal <em>17OHP</em> levels in groups A, B, and C were: 2.94 +/- 1.89, 1.77 +/- 0.81 and 3.90 +/- 2.43 nmol/liter, respectively (P>> 0.05) and for ACTH-stimulated levels were 12.6 +/- 7.2, 13.2 +/- 12.9 and 16.8 +/- 7.8 nmol/liter, respectively (P>> 0.05). Two carriers presented ACTH-stimulated <em>17OHP</em> levels between 30 and 45 nmol/liter and their entire CYP21 sequencing revealed only one mutation in heterozygous state indicating that the current cutoff level might overestimate the diagnosis of the NC form. We conclude that the variable ACTH-stimulated 17-OHP levels in carriers are not related to CYP21 gene mutations with different impairment of enzyme activity.

OBJECTIVE

The objective of the study was to evaluate the efficacy of 17 alpha-hydroxyprogesterone caproate (<em>17OHP</em>-C) in prolonging gestation in patients with a short cervix and other risk factors for preterm delivery, such as previous preterm birth, cervical surgery, uterine anomalies, or prenatal diethylstilbestrol (DES) exposure.

METHODS

This open-label, multicenter, randomized controlled trial included asymptomatic singleton pregnancies from 20(+0) through 31(+6) weeks of gestation with a cervical length less than 25 mm and a history of preterm delivery or cervical surgery or uterine malformation or prenatal DES exposure. Randomization assigned them to receive (or not) 500 mg of intramuscular <em>17OHP</em>-C weekly until 36 weeks. The primary outcome was time from randomization to delivery.

RESULTS

After enrolling 105 patients, an interim analysis demonstrated the lack of efficacy of <em>17OHP</em>-C in prolonging pregnancy. The study was discontinued because of futility. The groups were similar for maternal age, body mass index, parity, gestational age at inclusion, history of uterine anomalies, DES syndrome, previous preterm delivery or midtrimester abortion, and cervical length at randomization. The enrollment-to-delivery interval did not differ between patients allocated to <em>17OHP</em>-C (n = 51) and those allocated to the control group (n = 54) (median [interquartile range] time to delivery: 77 [54-103] and 74 [52-99] days, respectively). The rate of preterm delivery less than 37 (45% vs 44%, P>> .99), less than 34 (24% vs 30%, P = .51), or less than 32 (14% vs 20%, P = .44) weeks was similar in patients allocated to <em>17OHP</em>-C and those in the control group.

CONCLUSIONS

<em>17OHP</em>-C did not prolong pregnancy in women with singleton gestations, a sonographic short cervix, and other risk factors of preterm delivery (prior history, uterine malformations, cervical surgery, or prenatal DES exposure).

Ovarian Sertoli-Leydig cell tumours (SLCT), also termed arrhenoblastomas, are the most frequent virilising tumours in women of reproductive age. Very rare secretory Brenner tumours (BT) have been described, generally after the menopause. A 31-year-old woman sought medical advice for secondary amenorrhoea, progressive hirsutism and a 5-year history of virilisation syndrome with clitoromegaly. Testosterone was markedly high (285 ng/dl, N<85) with moderate elevation of delta 4-androstenedione (D4AD) (311 ng/dl, N <270), dehydroepiandrosterone sulfate (DHEAS) (366 μg/dl, N <340) and 17-hydroxyprogesterone (<em>17OHP</em>) (275 ng/dl). LH was 9 IU/l, FSH 4.3 IU/l, estradiol 60 pg/ml and progesterone 314 ng/100 ml. Cortisol was decreased (1.3 μg/dl) after the dexamethasone suppression test. Pelvic MRI showed a 5-cm right ovarian tumour with a 2.5 cm nodular component and cystic areas, and two nodules measuring 11 mm and 15 mm above the right and left ovaries. After right ovariectomy by laparoscopy, pathological examination concluded on a 3-cm SLCT and a 2-cm BT; the nodules above the ovaries were dysembryoplastic cysts. Postoperatively, testosterone level was normal after 24 h (26 ng/dl), estradiol and progesterone rapidly decreased, cyclic secretion then resumed and the patient menstruated at day 27. To our knowledge, this is the first report of an ovarian tumour associating a Sertoli-Leydig cell tumour and a Brenner tumour in a patient with virilisation syndrome which resolved after ovariectomy.

A sensitive liquid chromatography-electrospray ionization-tandem mass spectrometric (LC-ESI-MS-MS) method for the quantification of 17alpha-hydroxyprogesterone (<em>17OHP</em>) in human saliva has been developed and validated. The saliva was deproteinized with acetonitrile, purified using a Strata-X cartridge, derivatized with a highly proton-affinitive reagent, 2-hydrazinopyridine, and subjected to LC-MS-MS. Quantification was based on the selected reaction monitoring, and deuterated <em>17OHP</em> was used as the internal standard. This method allowed the reproducible and accurate quantification of the salivary <em>17OHP</em> using a 200-mul sample, and the limit of quantitation was 5.0 pg/ml. The developed method was applied to clinical studies. A linear relationship was found to be positive (r(2)=0.975) between the blood <em>17OHP</em> level and the salivary <em>17OHP</em> level measured using the proposed method. The result from the salivary <em>17OHP</em> measurement in patients with congenital adrenal hyperplasia demonstrated that the proposed method is very useful for monitoring of the therapeutic efficacy during hormone replacement therapy.

The binding of mineralocorticoid hormones to the mineralocorticoid receptor is the first step in a cascade of events leading to the stimulation of Na(+) reabsorption by renal cortical collecting duct (CCD) principal cells. The agonist properties of mineralocorticoid hormones are linked to contacts between their 21-hydroxyl group and Asn770, a residue of the ligand-binding domain of the human mineralocorticoid receptor (hMR). Here, we investigate whether the presence of a hydroxyl group at position 11, 17, or 20 could also alter the activity of progesterone (P), a mineralocorticoid antagonist without the 21-hydroxyl group. Both 17 alpha-hydroxyprogesterone (<em>17OHP</em>) and 20 alpha-hydroxyprogesterone (20OHP) antagonized the aldosterone-induced trans-activation activity (IC(50): <em>17OHP</em>, 10(-7) M; 20OHP, 10(-8) M) of the hMR transiently expressed in COS-7 cells lacking steroid receptors. In cultured mouse mpkCCD(cl4) principal cells, <em>17OHP</em> and 20OHP also prevented the aldosterone-stimulated amiloride-sensitive component of the short-circuit current (Ams I(sc)), reflecting Na(+) absorption mediated by the epithelial Na(+) channel (ENaC). In contrast, 11 beta-hydroxyprogesterone (11OHP) activated the transiently expressed hMR in COS-7 cells in a dose-dependent manner (ED(50): 10(-8) M) and, like aldosterone, stimulated Ams I(sc) in mpkCCD(cl4) cells. Docking 11OHP within the hMR-ligand-binding domain homology model revealed that the agonist activity of 11OHP is caused by contacts between its 11 beta-hydroxyl group and Asn770. Furthermore, 11OHP was unable to activate the mutant hMR/N770A, in which Ala is substituted for Asn at position 770. These findings demonstrate that in the absence of the 21-hydroxyl group, the 11 beta-hydroxyl group can produce the contact with the hMR-Asn770 required for the hMR activation leading to stimulated Na(+) absorption.

High levels of 11-ketotestosterone (11KT) were found (49 to 160 ng ml(-1)) in plasma of Siberian sturgeon females during the end of their reproductive cycle. These levels were measured either by specific radioimmunoassay, or both by specific radioimmunoassay and by UV absorption after HPLC (isocratic conditions, 33% methanol, 26% acetonitrile, 41% water). In order to find the origin of 11KT synthesis, ovaries were incubated (30 min and 2h at 20°C) with tritiated 17-hydroxyprogesterone (<em>17OHP</em>) or with tritiated androstenedione (A4). Testosterone (conversion rate from tritiated <em>17OHP</em>: 4%) and 11-ketotestosterone (conversion rate from tritiated A4: 1.6%) were identified as metabolites of respectively <em>17OHP</em> and A4 (TLC, HPLC and crystallization). 11β-hydroxyandrostenedione (11βOHA4) and 11β-hydroxytestosterone (11βOHT) were suggested to be intermediate metabolites. Besides interrenal and blood cells were incubated respectively with tritiated cortisol and tritiated A4. 11βOHA4 was identified in interrenal incubation (yield from tritiated cortisol: 1.2%). 11KT in interrenal (yield from tritiated cortisol: 0.14%), and 11βOHA4 and 11KT in blood cells (yield from tritiated A4: 1.6%), were suspected to be synthesized (TLC, HPLC, acetylation). No significant metabolization of tritiated cortisol could be found in liver. The possible contribution of each of these tissues to high 11KT levels found in plasma is discussed.

Treatment in a group of 19 patients with congenital adrenal hyperplasia (CAH) has been monitored by frequent, serial measurements of saliva 17OH-progesterone (<em>17OHP</em>) concentrations. Detailed <em>17OHP</em> profiles were obtained during consecutive weekend days and every 1-2 h over a separate 24-hour period. Patients showed a marked diurnal rhythm in <em>17OHP</em> levels, particularly when treated with hydrocortisone. In some patients, 10 mg/m2/day of hydrocortisone was sufficient glucocorticoid replacement to produce adequate control, although there was considerable individual variation. Saliva <em>17OHP</em> profiles provided valuable information on the efficacy of hydrocortisone, cortisone acetate, prednisolone and dexamethasone as glucocorticoid suppressive regimes in the treatment of CAH. Preliminary results suggest that hydrocortisone given in two divided doses during the day, supplemented by a small dose of prednisolone at bedtime, is suitable treatment for CAH patients who are still growing. In the patient who has completed statural growth, single daily dose dexamethasone therapy ensures adequate adrenal suppression and is convenient in the longterm.

The pattern of response to 14-day hCG stimulation was studied in 100 apparently normal infants and children and in 2 boys with total adrenal insufficiency. Response was estimated on the levels (nanograms per dl) of testosterone (T), delta4-androstenedione (delta4), and 17-hydroxyprogesterone (<em>17OHP</em>) before and after hCG. Basal levels of T, delta4, and <em>17OHP</em> increased significantly after 9--10 yr of age and before the onset of puberty. It would thus appear that the changes of adrenarche are also expressed by the secretion of delta4 androgens. After hCG, levels of T (629 +/- 218) did not vary with age in infants or prepubertal children but increased significantly at the onset of puberty (1265 +/- 253). Post-hCG levels of delta4 and <em>17OHP</em> increased with age, but their increments (respectively, 31.1 +/- 14.6 and 108.6 +/- 42.2) did not vary between 2--12 yr of age. On the other hand, the rise of delta4 after hCG tended to be higher in infancy and early pubescence. These data suggest that 1) testicular unresponsiveness to gonadotropins does not seem to be responsible for the postnatal decline of testicular activity; 2) during the prepubertal period there are no age-related changes in the absolute rises of T, delta4, or <em>17OHP</em> levels after hCG stimulation; and 3) increasing sensitivity of the testis to gonadotropic stimulation appears to be one of the very first changes occurring at the onset of puberty.

BACKGROUND

Women with polycystic ovary syndrome (PCOS) are at increased risk for cardiovascular (CV) and metabolic disorders. There is a close relationship between elevated androgen plasma levels and the ultrasound findings of stromal hypertrophy. In randomized trials, the administration of metformin has been shown to be followed by an improvement in insulin sensitivity and decrease in androgen levels in most women. In the present study, we investigate the association between reduced ovarian volume in PCOS patients after administration of metformin with improvement in CV risk factors.

METHODS

This was a randomized clinical trial study. A total of 28 women diagnosed with PCOS who referred to the infertility clinic were selected. Anthropometric characteristics of the patients, mean ovarian volume and plasma levels of fasting blood sugar (FBS), lipid profile, luteinizing hormone (LH), follicle stimulating hormone (FSH), estradiol, testosterone, 17-α-OH progesterone (<em>17OHP</em>), dehydroepiandrosterone sulfate (DHEAS), C-reactive protein (CRP) and homocysteine (Hcy) were evaluated before and after treatment with 500 mg metformin, three times daily for three months. Statistics were calculated with the aid of SPSS 16.0 with student's paired t- and Pearson's correlation coefficient tests. Significance was set at p<0.05.

RESULTS

There were significant reductions in mean ovarian volume and body mass index (BMI), in addition to CRP, Hcy, testosterone, FBS, HDL and LDL levels. There was a positive correlation between mean ovarian volume and waist-to-hip ratio (WHR).After treatment, there correlation noted with reduction in mean ovarian volume and decreased BMI, in addition to reductions in CRP, LDL, Hcy and testosterone levels.

CONCLUSIONS

A positive correlation may exist between reduced mean ovarian volume and improvement in cv risk factors after administration of metformin (

BACKGROUND

IRCT138903244176N1).

Congenital adrenal hyperplasia (CAH) [OMIM 201 910] is a group of autosomal recessive disorders most commonly due to 21-hydroxylase deficiency and presenting with a wide range of clinical manifestations. A limited number of inactivating pseudogene-derived mutations account for the majority of 21-hydroxylase gene ( CYP21) mutations, additional rare mutations can be found in single families and small populations. We found three novel CYP21 mutations in CAH patients suffering from the classical form of the disease, of which one is a frameshift mutation (1353-1354insA) leading to a premature termination codon (K277K, Q228A...E294X), one results in a premature stop codon (2551C>T, R444X), and one is a missense mutation (2609T>C; P463L). The frameshift and premature stop mutations can be predicted to result in a CYP21 protein without any residual enzyme activity. To determine the functional consequences of the P463L mutation, the IN VITRO enzyme activity was studied in COS-7 cells and revealed a reduced 21-hydroxylase activity of 2.6+/-0.8 (SD)% for the conversion of 17-hydroxyprogesterone (<em>17OHP</em>) to 11-deoxycortisol and of 3.0+/-0.5 % for the conversion of progesterone to 11-deoxycorticosterone (DOC). We conclude that functional analyses of unknown mutations provide information on the disease severity and should be always performed when novel CYP21 mutations are detected. Knowledge of the residual 21-hydroxylase function improves both genetic counselling and individual clinical management in CAH patients.

To determine the function of Leydig cells in patients with varicocele, hCG-stimulated levels of progesterone (Prog), 17 alpha-Hydroxy-4-pregnene-3,20-dione (<em>17OHP</em>), 4-Androstene-3,17-dione (A-dione), testosterone (T), and estradiol-17 beta (E2) in both spermatic and peripheral veins were measured. Seventy-two patients with idiopathic varicocele were divided into four groups: patients in group 1 were untreated, whereas patients in groups 2, 3, and 4 were given a single i.m. injection of 10,000 IU hCG 24 h, 96 h, and 168 h before surgery, respectively. In the spermatic and peripheral veins, levels of Prog, <em>17OHP</em>, and E2 showed peaks at 24 h, whereas levels of A-dione and T showed peaks at 96 h. Significant increases in the ratios of spermatic veins <em>17OHP</em> to A-dione and <em>17OHP</em> to T, and a significant decrease in the ratio of T to E2, was found 24 h following hCG treatment. These results demonstrate that, following hCG injection, there is a transient inhibition of testicular C17-20-lyase activity, probably mediated by E2, even in subfertile males with varicocele.

Orally administered testosterone undecanoate (TU), an anabolic, androgenic steroid, can potentially be abused by athletes. Indirect evidence for detecting oral TU intake could be deduced from the changes in steroid profile post-administration. Direct evidence could be obtained by detection of unchanged TU in plasma. To this end, both urinary and plasma steroid profiles of six healthy male subjects given a single oral dose of 120 mg of TU were studied by gas chromatography/mass spectrometry (GC/MS) and gas chromatography/tandem mass spectrometry (GC/MS/MS). The increased concentration of glucuronidated testosterone in plasma appears to be the most characteristic sign of oral TU intake. The testosterone glucuronide (TG)/nonconjugated testosterone (T) ratio, TG/17-hydroxyprogesterone (<em>17OHP</em>) ratio, and TG/luteinizing hormone (LH) ratio were observed to be significantly elevated above their basal levels for 10 h, 10 h, and 6 h, respectively. Urinary ratios of TG/epitestosterone glucuronide (EG) were found to be higher than the cut-off value of 6 for the period 4 approximately 8 h post-administration, but only in three subjects. One subject failed to respond with respect to all of the above-mentioned indirect markers, as TG was not significantly increased in either plasma or urine. Unchanged TU was directly detected in plasma of all six subjects from 1 approximately 1.5 h to 4 approximately 6 h after oral TU intake by GC/MS/MS, providing unequivocal proof of exogenous testosterone intake. Distinct and complementary markers for detecting oral TU intake could be obtained from plasma and urine, respectively.

Precocious pubarche (PP) is defined as the onset of pubic hair at 8 years of age in girls and at 9 years of age in boys. PP is idiopathic (IPP) in most children, but it is the earliest manifestation of non-classical congenital adrenal hyperplasia owing to steroid 21-hydroxylase deficiency (NC21OHD) in 5%-20% of cases. 17-Hydroxyprogesterone (<em>17OHP</em>) levels after ACTH stimulation test are used to distinguish the two forms. We studied clinical indicators of NC21OHD in 289 PP children: 14 (4.8%) showed post-ACTH <em>17OHP</em> levels >30 nmol/L and NC21OHD due to CYP21A2 gene mutations was confirmed. NC21OHD children were younger (p: 0.006) and thinner (p: 0.003) than IPP children. Height standard deviation score (SDS) was not different (p: 0.97). NC21OHD girls showed more advanced bone age than IPP girls (p<0.001). Earlier PP onset and bone age advance suggest NC21OHD, which requires confirmation by an ACTH stimulation test. Later, PP appearance in overweight children suggests IPP and could merit only clinical monitoring.

BACKGROUND

Treatment of 21-hydroxylase deficiency (21OHD) is difficult to optimize. Normalization of excessive ACTH and adrenal steroid production commonly requires supraphysiologic doses of glucocorticoids.

OBJECTIVE

We evaluated the safety and tolerability of the selective corticotropin releasing factor type 1 (CRF1) receptor antagonist NBI-77860 in women with classic 21OHD and tested the hypothesis that CRF1 receptor blockade decreases early-morning ACTH and 17α-hydroxyprogesterone (<em>17OHP</em>) in these patients.

METHODS

The study enrolled eight classic 21OHD females, ages 18-58 years, seen at a single tertiary referral university setting.

METHODS

This was a phase Ib, single-blind, placebo-controlled, fixed-sequence, single-dose trial. During three treatment periods separated by 3-week washout intervals, patients sequentially received placebo, NBI-77860 300 mg, and NBI-77860 600 mg at 10 pm; glucocorticoid therapy was withheld for 20 hours. We evaluated ACTH, <em>17OHP</em>, androstenedione, and testosterone as well as NBI-77860 pharmacokinetics over 24 hours.

RESULTS

Dose-dependent reductions of ACTH and/or <em>17OHP</em> were observed in six of eight subjects. Relative to placebo, NBI-77860 led to an ACTH and <em>17OHP</em> reduction by a mean of 43% and 0.7% for the 300 mg dose, respectively, and by 41% and 27% for the 600 mg dose, respectively. Both NBI-77860 doses were well tolerated.

CONCLUSIONS

The meaningful reductions in ACTH and <em>17OHP</em> following NBI-77860 dosing in 21OHD patients demonstrate target engagement and proof of principle in this disorder. These promising data provide a rationale for additional investigations of CRF1 receptor antagonists added to physiologic doses of hydrocortisone and fludrocortisone acetate for the treatment of classic 21OHD.

There is a strong correlation between the severity of genotypes and 17OH-progesterone levels in patients with the nonclassical form of 21-hydroxylase deficiency (NC-CAH); however, there are few studies regarding the correlation with clinical signs. The aim of the study was to evaluate whether genotypes correlate with the severity of the hyperandrogenic phenotype. A cohort of 114 NC-CAH patients were diagnosed by stimulated-<em>17OHP</em> ≥10 ng/ml. CYP21A2 genotypes were divided into 2 groups according to the severity of enzymatic impairment; mild and severe. Clinical data and hormonal profiles were compared between the 2 groups. Age at onset of manifestations did not differ between children or adults carrying both mild and severe genotypes. Frequencies of precocious pubarche and hirsutism, with or without menstrual abnormalities, were similar between the 2 groups. There were no differences in basal testosterone levels of adult symptomatic females carrying both genotypes, but there were differences between adult females with (92.9±49.5 ng/dl) and without hirsutism (43.8±38 ng/dl) (p=0.0002). Similar frequencies of both genotypes were observed in asymptomatic females and in those with clitoromegaly. Nonclassical genotypes do not predict the severity of phenotype. Asymptomatic and virilized females carrying the same genotype suggest that there is a modulatory effect of genes involved in the androgen pathway on the phenotype.

OBJECTIVE

The effectiveness of neonatal screening for reducing morbimortality in children with congenital adrenal hyperplasia (CAH) is the main justification for its implementation. One of the challenges for its implementation is to determine the cutoff value for laboratory measurement of 17-hydroxyprogesterone (<em>17OHP</em>) with appropriate cost-effectiveness. This study identified factors affecting the results of the pilot project of newborn screening for CAH, performed in the state of Minas Gerais, Brazil.

METHODS

Neonatal screening performed between September, 2007 and May, 2008, with <em>17OHP</em> measurements performed in blood samples taken from the heel (filter paper), on the 5(th) day of life, processed by the UMELISA 17-OH Progesterona NEONATAL(®) method. The cutoff value was 80 and 160 nmol/L for healthy children or not, respectively.

RESULTS

The incidence of CAH was 1:19,939 in 159,415 children screened. The 99(th) percentile (p99) of <em>17OHP</em> in the first sample was 108 nmol/L. In 13,298 newborns whose weight had been reported, the p99 of <em>17OHP</em> were, respectively: 344 nmol/L for weight < 1,500 g; 260 nmol/L for weight between 1,500 and 1,999 g; 221 nmol/L for weight between 2,000 and 2,499 g; 109 nmol/L for weight ≥ 2,500g. The rate of recall for medical consultation was 0.31%. The test sensitivity was 100%, specificity was 99.6%, and the positive predictive value was 2.2%. By adjusting the cutoff values of <em>17OHP</em> to 110 nmol/L and 220 nmol/L, a 76% decrease in consultation referrals was projected.

CONCLUSIONS

The use of <em>17OHP</em> cutoff values, considering birth weight, was a cost-effective measure to reduce false positives. The results of this pilot study suggest that screening for CAH might benefit the pediatric population.

BACKGROUND

3β-hydroxysteroid dehydrogenase (3βHSD) type 2 (encoded by HSD3B2) is expressed in the adrenals and gonads. HSD3B2 mutations cause the rare form of congenital adrenal hyperplasia '3βHSD deficiency'. In its classic form, affected individuals have salt wasting early in infancy and may have ambiguous genitalia in both sexes. The presence of peripheral type 1 3βHSD often complicates the hormonal diagnosis of this disorder, in that very high 17α-hydroxypregnenolone levels can be converted extra-adrenally to 17α-hydroxyprogesterone (<em>17OHP</em>).

METHODS

A 46,XX female newborn with no signs of virilization was referred for evaluation of positive <em>17OHP</em> newborn screening, and developed a salt-wasting crisis at 13 days of age. The confirmatory test revealing highly elevated <em>17OHP</em> suggested a 21-hydroxylase deficiency, but sequencing of the CYP21A2 gene was not consistent. Further family history suggested a 3βHSD deficiency. The HSD3B2 gene was then sequenced.

RESULTS

The patient was homozygous for the novel nonsense mutation Q334X in the HSD3B2 gene, inherited from both parents.

CONCLUSIONS

We report a novel mutation of the HSD3B2 gene, Q334X, responsible for a classic 3βHSD deficiency. The clinical and hormonal phenotypes can be complicated in this disorder, and this supports the benefits of <em>17OHP</em> newborn screening to detect various forms of congenital adrenal hyperplasia.

Newborn screening for congenital adrenal hyperplasia (CAH) caused by 21-hydroxylase deficiency is mandated throughout the US. Filter paper blood specimens are assayed for 17-hydroxyprogesterone (<em>17OHP</em>). Prematurity, low birth weight, or critical illness cause falsely elevated results. The purpose of this report is to highlight differences in protocols among US state laboratories. We circulated a survey to state laboratory directors requesting qualitative and quantitative information about individual screening programs. Qualitative and quantitative information provided by 17 state programs were available for analysis. Disease prevalence ranged from 1:9941 to 1:28,661 live births. Four state laboratories mandated a second screen regardless of the initial screening results; most others did so for infants in intensive care units. All but one program utilized birthweight cut-points, but cutoffs varied widely: <em>17OHP</em> values of 25 to 75 ng/mL for birthweights >2250-2500 g. The positive predictive values for normal birthweight infants varied from 0.7% to 50%, with the highest predictive values based in two of the states with a mandatory second screen. Data were unavailable for negative predictive values. These data imply differences in sensitivity and specificity in CAH screening in the US. Standardization of newborn screening protocols could improve the positive predictive value.

Keywords: adrenal hyperplasia; congenital; newborn screening; standardization.