Deborah P. Merke

Puberty-related influences on brain development

Puberty is a time of striking changes in cognition and behavior. To indirectly assess the effects of puberty-related influences on the underlying neuroanatomy of these behavioral changes we will review and synthesize neuroimaging data from typically developing children and adolescents and from those with anomalous hormone or sex chromosome profiles. The trajectories (size by age) of brain morphometry differ between boys and girls, with girls generally reaching peak gray matter thickness 1-2 years earlier than boys. Both boys and girls with congenital adrenal hyperplasia (characterized by high levels of intrauterine testosterone), have smaller amygdala volume but the brain morphometry of girls with CAH did not otherwise significantly differ from controls. Subjects with XXY have gray matter reductions in the insula, temporal gyri, amygdala, hippocampus, and cingulate-areas consistent with the language-based learning difficulties common in this group.

Adrenomedullary Dysplasia and Hypofunction in Patients with Classic 21-Hydroxylase Deficiency

BACKGROUND Glucocorticoids are essential for the normal development and functioning of the adrenal medulla. Whether adrenomedullary structure and function are normal in patients with congenital adrenal hyperplasia is not known. METHODS We measured plasma and urinary catecholamines and plasma metanephrines in 38 children with congenital adrenal hyperplasia due to 21-hydroxylase deficiency (25 children with the salt-wasting form and 13 with the simple virilizing form), 39 age-matched normal subjects, and 20 patients who had undergone bilateral adrenalectomy. Adrenal specimens obtained from three other patients with 21-hydroxylase deficiency who had undergone bilateral adrenalectomy and specimens obtained at autopsy from eight other patients were examined histologically. RESULTS Plasma epinephrine and metanephrine concentrations and urinary epinephrine excretion were 40 to 80 percent lower in the patients with congenital adrenal hyperplasia than in the normal subjects (P<0.05), and the values were lowest in the patients with the most severe deficits in cortisol production. Urinary epinephrine excretion and plasma epinephrine concentrations were at or below the limit of detection of the assay in 8 (21 percent) of the patients with congenital adrenal hyperplasia and in 19 (95 percent) of the patients who had undergone adrenalectomy. In the group of patients with congenital adrenal hyperplasia, plasma epinephrine and metanephrine concentrations and urinary epinephrine excretion were approximately 50 percent lower in those who had been hospitalized for adrenal crises than in those who had not. In three patients with congenital adrenal hyperplasia who had undergone bilateral adrenalectomy, the formation of the adrenal medulla was incomplete, and electron-microscopical studies revealed a depletion of secretory vesicles in chromaffin cells. CONCLUSIONS Congenital adrenal hyperplasia compromises both the development and the functioning of the adrenomedullary system.

Modified-Release Hydrocortisone to Provide Circadian Cortisol Profiles

CONTEXT Cortisol has a distinct circadian rhythm regulated by the brains central pacemaker. Loss of this rhythm is associated with metabolic abnormalities, fatigue, and poor quality of life. Conventional glucocorticoid replacement cannot replicate this rhythm. OBJECTIVES Our objectives were to define key variables of physiological cortisol rhythm, and by pharmacokinetic modeling test whether modified-release hydrocortisone (MR-HC) can provide circadian cortisol profiles. SETTING The study was performed at a Clinical Research Facility. DESIGN AND METHODS Using data from a cross-sectional study in healthy reference subjects (n = 33), we defined parameters for the cortisol rhythm. We then tested MR-HC against immediate-release hydrocortisone in healthy volunteers (n = 28) in an open-label, randomized, single-dose, cross-over study. We compared profiles with physiological cortisol levels, and modeled an optimal treatment regimen. RESULTS The key variables in the physiological cortisol profile included: peak 15.5 microg/dl (95% reference range 11.7-20.6), acrophase 0832 h (95% confidence interval 0759-0905), nadir less than 2 microg/dl (95% reference range 1.5-2.5), time of nadir 0018 h (95% confidence interval 2339-0058), and quiescent phase (below the mesor) 1943-0531 h. MR-HC 15 mg demonstrated delayed and sustained release with a mean (sem) maximum observed concentration of 16.6 (1.4) microg/dl at 7.41 (0.57) h after drug. Bioavailability of MR-HC 5, 10, and 15 mg was 100, 79, and 86% that of immediate-release hydrocortisone. Modeling suggested that MR-HC 15-20 mg at 2300 h and 10 mg at 0700 h could reproduce physiological cortisol levels. CONCLUSION By defining circadian rhythms and using modern formulation technology, it is possible to allow a more physiological circadian replacement of cortisol.

Future directions in the study and management of congenital adrenal hyperplasia due to 21-hydroxylase deficiency

Dr. Deborah P. Merke (Warren Grant Magnuson Clinical Center, National Institutes of Health [NIH], Bethesda, Maryland): Congenital adrenal hyperplasia due to 21-hydroxylase deficiency is one of the most common known autosomal recessive disorders (1). In this condition, impaired cortisol production leads to a lack of negative glucocorticoid feedback on the pituitary, hypothalamus, and suprahypothalamic centers, resulting in an increase in corticotropin, a buildup of cortisol precursors, and androgen excess (Figure 1). The carrier frequency of the classic or severe form of 21-hydroxylase deficiency is approximately 1 in 60 persons (2). The carrier frequency of the nonclassic or mild form ranges from 1 in 5 to 1 in 50 persons, depending on ethnicity (3); it is most common in Hispanic and Ashkenazi Jewish populations. Because congenital adrenal hyperplasia has a high frequency, a variable presentation in children and adults, and potential complications, a thorough understanding of the disorder is of great importance to clinicians working in internal medicine, reproductive medicine, and pediatrics. Figure 1. Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency. left middle ACTH right The classic form of congenital adrenal hyperplasia presents in infancy and early childhood as signs and symptoms of virilization with or without adrenal insufficiency. It is subcategorized as salt-losing or non-salt-losing, reflecting the degree of mineralocorticoid deficiency (4). In the early 1950s, cortisone therapy was found to be effective in treating adrenal insufficiency and excess androgen production in patients with congenital adrenal hyperplasia (5). Since the discovery of cortisone therapy and the addition of mineralocorticoid supplementation, the morbidity and mortality of patients with classic disease have markedly decreased, and these patients now have a long life span. Thus, long-term consequences of current treatments are an important consideration. The 21-hydroxylase-deficient mouse, which was described in a Japanese study by Shiroishi and coworkers (6), has been a useful model for gaining new insights into the pathophysiology of the disease in humans and for developing new therapeutic strategies. The mouse model revealed an abnormal hypothalamic-pituitary-adrenal feedback mechanism (7), alterations in the structure and function of the adrenal medulla (8), and a good response to gene therapy (9). Further exploration of the disease process in this animal model is an essential aspect of the bench-to-bedside research approach to improving the human condition. Abnormalities in both the structure and function of the adrenal medulla have been shown in patients with classic congenital adrenal hyperplasia (10). This finding may explain why some children with the severe form of 21-hydroxylase deficiency are prone to adrenal crises, hypoglycemia, and cardiovascular collapse in response to febrile illnesses or other stressful circumstances, despite adequate glucocorticoid replacement. There are many unresolved clinical problems in the management of classic 21-hydroxylase deficiency in both males and females. Among the most critical are inadequate response to glucocorticoid and mineralocorticoid replacement therapy, iatrogenic Cushing syndrome (11), adult short stature (12, 13), and oligo-amenorrhea and infertility in women (14, 15). The new treatment approaches to classic congenital adrenal hyperplasia represent potential solutions to these unresolved issues. In a long-term randomized clinical trial, the NIH is testing a new treatment regimen consisting of reduced hydrocortisone dose, an antiandrogen, and an aromatase inhibitor (16, 17). Bilateral adrenalectomy is being performed in selected cases (18). Future therapies include a new class of drug called corticotropin-releasing hormone antagonists (19) and possibly gene therapy (9). Nonclassic congenital adrenal hyperplasia, the mild form of the disease, is a common cause of hyperandrogenism in women. Although the same gene is involved in both the severe and mild forms, genetic mutations typically associated with the mild form of the disease only partially impair 21-hydroxylase activity. Thus, the patient with nonclassic congenital adrenal hyperplasia is in a fully compensated state; she does not have cortisol deficiency but rather manifestations of hyperandrogenism, usually later in childhood, around puberty, or in early adulthood (20, 21). Nonclassic congenital adrenal hyperplasia is an important consideration in the differential diagnosis of female patients with symptoms or signs of hyperandrogenism, such as severe cystic acne, hirsutism, male pattern baldness, oligo-amenorrhea, or infertility. Nonclassic 21-hydroxylase deficiency, especially when it exists in conjunction with hyperinsulinemia, often results in the polycystic ovary syndrome, with its characteristic reproductive and metabolic comorbid conditions. Recognition of this disorder is crucial for family planning and management in women with hyperandrogenism. Men with nonclassic congenital adrenal hyperplasia are usually asymptomatic but may also present with early puberty or testicular adrenal rests. Another recognized comorbid condition associated with congenital adrenal hyperplasia is activation of ectopic adrenal tissue resulting in adrenal rest tumors (22). These tumors are most commonly found in the testes of men with classic or nonclassic congenital adrenal hyperplasia and often result in oligo-azoospermia and infertility (23). New advances in the diagnostic evaluation and management of these tumors are presented in this paper. Genetics The gene for 21-hydroxylase lies on chromosome 6 within the HLA locus of the major histocompatibility system; thus, 21-hydroxylase deficiency is an HLA-linked disorder (24). Two homologous genes result from ancestral duplication. CYP21B is the active gene; CYP21A is an inactive pseudogene. The location of the CYP21B gene makes it vulnerable to relatively large genomic DNA exchanges with its homologous gene, CYP21A. The proximity of these genes and their location within the HLA region, which has a high rate of recombination, facilitate such exchanges. Therefore, the 21-hydroxylase locus shows an unusual degree of variability between individuals (25, 26). 21-Hydroxylase deficiency is unique because most mutations result from the transfer of sequences between pseudogenes and active genes (27). When deleterious sequences that normally present in the pseudogene are transferred to the active gene, they render the gene incapable of encoding a normal enzyme. The term gene conversion is used to denote the transfer of sequences between homologous genes; however, the mechanism is poorly understood. Specific mutations typically correspond to the three types of 21-hydroxylase deficiency: salt-losing, non-salt-losing (simple virilizing), and nonclassic congenital adrenal hyperplasia (Figure 2). In vitro studies have shown that mutations resulting in complete inactivation of 21-hydroxylase activity are associated with the salt-losing phenotype, those that reduce 21-hydroxylase activity to approximately 2% are associated with the non-salt-losing phenotype, and those that reduce 21-hydroxylase activity to 10% to 75% are associated with the nonclassic phenotype (28, 29) (Figure 2). Most patients are compound heterozygotes, and the severity of the disease is determined by the activity of the less severely affected allele. Figure 2. The 10 most common genetic mutations found in 21-hydroxylase deficiency. SL NSL NC CYP21B The degree of functional impairment predicted by individual mutations usually corresponds to the clinical severity observed in a given patient. However, genotype does not always accurately predict phenotype. This disparity between genotype and phenotype may be due to androgen sensitivity or to other genes that cause differences in steroid metabolism and homeostasis. Animal Model for 21-Hydroxylase Deficiency Dr. Stefan R. Bornstein (Pediatric and Reproductive Endocrinology Branch, National Institute of Child Health and Human Development [NICHD], NIH): In the mouse, the 21-hydroxylase gene also lies within the major histocompatibility locus (30). Mice with spontaneous 21-hydroxylase deficiency have a deletion of the 21-hydroxylase gene (6, 7, 31). 21-Hydroxylase activity is completely absent in newborn mice homozygous for this deletion, and this absence is lethal in the early postnatal stage. The deficiency of glucocorticoids results in adrenocortical hyperplasia and plasma accumulation of precursor steroids in both mice and humans with congenital adrenal hyperplasia. In mice, which normally lack adrenal 17--hydroxylase, the enzymatic blockade results mainly in accumulation of progesterone. Most of the affected mice, if not treated with glucocorticoids and mineralocorticoids, die within 1 week (6, 7, 31). Although the disease state of the 21-hydroxylase-deficient mouse is not completely comparable to human congenital adrenal hyperplasia, it has provided a useful model with which to examine molecular and cellular mechanisms of the disease and test new therapeutic interventions. The adrenal glands of affected mice demonstrate a significant increase in expression of messenger RNA of steroidogenic acute regulatory protein (Figure 3), which is the rate-limiting step for steroidogenesis (32). Corticotropin regulates the expression of this protein, and the increase of messenger RNA reflects impaired negative feedback with increased corticotropin production. Moreover, one study showed that prenatal dexamethasone treatment (0.5 to 2 g/d) failed to adequately suppress fetal adrenal hormones in mice, suggesting hyperactivity of the hypothalamic-pituitary corticotroph axis and insensitivity to glucocorticoid feedback inhibition (7). Intrauterine glucocorticoid deficiency may affect the sensitivity of feedback inhibition postnatally, thus blunting the central effects of treatment

Hypogonadotropic hypogonadism in a female caused by an X-linked recessive mutation in the DAX1 gene.

Adrenal hypoplasia congenita is a rare X-linked disorder characterized by primary adrenal insufficiency and hypogonadotropic hypogonadism.1 All patients described to date have been male, and female...

Clinical characteristics of a cohort of 244 patients with congenital adrenal hyperplasia.

CONTEXT Patients with congenital adrenal hyperplasia (CAH) often suffer from long-term complications secondary to chronic glucocorticoid therapy and suboptimal treatment regimens. OBJECTIVE The aim of the study was to describe clinical characteristics of a large cohort of pediatric and adult CAH patients. DESIGN AND SETTING We conducted a cross-sectional study of 244 CAH patients [183 classic, 61 nonclassic (NC)] included in a Natural History Study at the National Institutes of Health. MAIN OUTCOME MEASURE(S) Outcome variables of interest were height sd score, obesity, hypertensive blood pressure (BP), insulin resistance, metabolic syndrome, bone mineral density, hirsutism (females), and testicular adrenal rest (TART). RESULTS The majority had elevated or suppressed androgens, with varied treatment regimens. Mean adult height SD score was -1.0 ± 1.1 for classic vs. -0.4 ± 0.9 for NC patients (P = 0.015). Obesity was present in approximately one third of patients, across phenotypes. Elevated BP was more common in classic than NC patients (P ≤ 0.01); pediatric hypertensive BP was associated with suppressed plasma renin activity (P = 0.001). Insulin resistance was common in classic children (27%) and adults (38% classic, 20% NC); 18% of adults had metabolic syndrome. The majority (61%) had low vitamin D; 37% of adults had low bone mineral density. Hirsutism was common (32% classic; 59% NC women). TART was found in classic males (33% boys; 44% men). CONCLUSIONS Poor hormonal control and adverse outcomes are common in CAH, necessitating new treatments. Routine monitoring of classic children should include measuring BP and plasma renin activity. Osteoporosis prophylaxis and TART screening should begin during childhood. A longitudinal study is under way.

Comprehensive Genetic Analysis of 182 Unrelated Families with Congenital Adrenal Hyperplasia due to 21-Hydroxylase Deficiency

BACKGROUND Genetic analysis is commonly performed in patients with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency. STUDY OBJECTIVE The objective of the study was to describe comprehensive CYP21A2 mutation analysis in a large cohort of CAH patients. METHODS Targeted CYP21A2 mutation analysis was performed in 213 patients and 232 parents from 182 unrelated families. Complete exons of CYP21A2 were sequenced in patients in whom positive mutations were not identified by targeted mutation analysis. Copy number variation and deletions were determined using Southern blot analysis and PCR methods. Genotype was correlated with phenotype. RESULTS In our heterogeneous U.S. cohort, targeted CYP21A2 mutation analysis did not identify mutations on one allele in 19 probands (10.4%). Sequencing identified six novel mutations (p.Gln262fs, IVS8+1G>A, IVS9-1G>A, p.R408H, p.Gly424fs, p.R426P) and nine previously reported rare mutations. The majority of patients (79%) were compound heterozygotes and 69% of nonclassic (NC) patients were compound heterozygous for a classic and a NC mutation. Duplicated CYP21A2 haplotypes, de novo mutations and uniparental disomy were present in 2.7% of probands and 1.9 and 0.9% of patients from informative families, respectively. Genotype accurately predicted phenotype in 90.5, 85.1, and 97.8% of patients with salt-wasting, simple virilizing, and NC mutations, respectively. CONCLUSIONS Extensive genetic analysis beyond targeted CYP21A2 mutational detection is often required to accurately determine genotype in patients with CAH due to the high frequency of complex genetic variation.

Approach to the adult with congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

Congenital adrenal hyperplasia (CAH) describes a group of autosomal recessive disorders where there is impairment of cortisol biosynthesis. CAH due to 21-hydroxylase deficiency accounts for 95% of cases and shows a wide range of clinical severity. Treatment of the classic or severe form of CAH is targeted at replacing cortisol and aldosterone and effectively controlling excess androgen symptoms by using the lowest possible glucocorticoid dose. Treatment of the mild or nonclassic form is targeted at controlling excess androgen symptoms and may or may not involve glucocorticoid therapy. Hydrocortisone is the treatment of choice for children, but there is no consensus on how patients should be treated as adults. Current glucocorticoid therapy is suboptimal because it is often difficult to reduce excess androgen without giving excess glucocorticoid, and patients may experience hypercortisolism, androgen excess, or a combination of these states. Treatment of CAH, especially in the adult patient, remains controversial given the lack of prospective randomized controlled trials comparing treatment regimens. Nevertheless, patients benefit from careful individualized therapy with avoidance of Cushingoid side effects and optimization of reproductive, sexual, and bone health.

A pharmacokinetic and pharmacodynamic study of delayed- and extended-release hydrocortisone (Chronocort) vs. conventional hydrocortisone (Cortef) in the treatment of congenital adrenal hyperplasia.

Objective  Existing glucocorticoid treatment for congenital adrenal hyperplasia (CAH) is suboptimal and nonphysiological. We compared hormonal profiles during therapy with a new modified‐release hydrocortisone (MR‐HC), Chronocort™, to conventional hydrocortisone (HC), Cortef™, in patients with CAH.

Early androgen exposure modulates spatial cognition in congenital adrenal hyperplasia (CAH)

Major questions remain about the exact role of hormones in cognition. Furthermore, the extent to which early perturbation in steroid function affects human brain development continues to be a wide open area of research. Congenital adrenal hyperplasia (CAH), a genetic disorder of steroid dysfunction characterized in part by in utero over-production of testosterone, was used as a natural model for addressing this question. Here, CAH (n=54, mean age=17.53, 31 female) patients were compared to healthy age- and sex-matched individuals (n=55, mean age=19.02, 22 female) on a virtual equivalent of the Morris Water Maze task [Morris, R., 1984. Developments of a water-maze procedure for studying spatial learning in the rat. J. Neurosci. Methods 11, 47-60], an established measure of sex differences in spatial cognition in rodents. Findings revealed that females with CAH with the most severe form of the disease and expected highest level of in utero exposure to androgens were found to perform similarly to both healthy males and CAH males, whereas strong sex differences were apparent in milder forms of the disorder and in controls. Moreover, advanced bone age, an indicator of long-term childhood exposure to testosterone was correlated with improved performance. The results indicate that individuals exposed to both excess androgens prenatally and prolonged exposure during childhood may manifest long-lasting changes in cognitive function. Such finding suggests a pivotal role of hormonal function on brain development in humans, mirroring results from the animal literature.