5.11 Congenital adrenal hyperplasia

Congenital adrenal hyperplasia (CAH) represents a group of autosomal recessive disorders of steroidogenesis caused by defects in steroidogenic enzymes involved in glucocorticoid synthesis or in enzymes providing cofactors to steroidogenic enzymes (1, 2). Congenital lipoid adrenal hyperplasia (CLAH) caused by steroidogenic acute regulatory protein (StAR) deficiency is distinct in origin and presentation from the conventional variants of CAH, with the unique feature of lipid accumulation subsequently leading to destruction of adrenal function. This chapter will also mention aldosterone synthase deficiency, which is the only defect in adrenal steroidogenesis causing deficient mineralocorticoid biosynthesis without affecting glucocorticoid biosynthesis. The disorder cannot strictly be considered a CAH variant as it does not result in increased ACTH drive and thus not in adrenal hyperplasia.

Novel forms of CAH have emerged during recent years. These include P450 oxidoreductase deficiency (ORD), P450 side-chain cleavage (CYP11A1) deficiency, the nonclassic form of CLAH (StAR deficiency), and apparent cortisone reductase deficiency. All forms of congenital adrenal hyperplasia resemble a disease continuum spanning from mild nonclassic presentations to classic onset with severe signs and symptoms.

The adrenal cortex consists of three zones: the outer zona glomerulosa is responsible for mineralocorticoid synthesis, the middle zona fasciculata for glucocorticoid synthesis, and the inner zona reticularis for synthesis of the adrenal androgen precursors dehydroepiandrosterone (DHEA) and androstenedione. All major enzymes involved in adrenal steroidogenesis are located either in the mitochondria or the endoplasmic reticulum. The function of mitochondrial (type I) cytochrome P450 (CYP) enzymes, such as P450 side-chain cleavage (CYP11A1), 11β-hydroxylase (CYP11B1), and aldosterone synthase (CYP11B2), depends on electron transfer facilitated by the proteins adrenodoxin and adrenodoxin reductase. Micrososomal (type II) CYP enzymes localized to the endoplasmic reticulum include 17α-hydroxylase (CYP17A1), 21-hydroxylase (CYP21A2), and P450 aromatase (CYP19A1). The function of CYP type II enzymes crucially depends on P450 oxidoreductase (POR) providing electrons required for monooxygenase reaction catalysed by the CYP enzyme.

Glucocorticoid synthesis is under negative feedback control of the hypothalamic–pituitary–adrenal (HPA) axis (see Chapter 5.9). The pituitary releases ACTH, which binds to its adrenal receptor (melanocortin receptor 2) and stimulates import of cholesterol into the mitochondrion by StAR. In parallel, transcription of genes encoding steroidogenic enzymes and their cofactor enzymes is increased. The rate-limiting step is the conversion of cholesterol into pregnenolone by CYP11A1, which is expressed in all three adrenocortical zones. The biosynthetic directionality of different steroid hormone pathways in the adrenal zones is facilitated by differential expression of steroidogenic enzymes and cofactors.

Glucocorticoids are mainly synthesized in the zona fasciculata, following the route from pregnenolone via progesterone, 17-hydroxyprogesterone (17OHP), or pregnenolone via 17-hydroxypregnenolone and 17OHP. 17-Hydroxyprogesterone is then 21-hydroxylated to 11-deoxycortisol and finally converted to cortisol.

Sex steroids are produced from pregnenolone by 17α-hydroxylation to 17OH-pregnenolone, which is converted by the 17,20-lyase activity of CYP17A1 into DHEA, the universal sex steroid precursor. Sufficient 17,20-lyase activity depends not only on POR but also on the availability of cytochrome b5, which facilitates close interaction between CYP17A1 and its electron donor POR. The conversion from 17OHP to androstenedione is negligible under normal physiological circumstances (Fig. 5.11.1). Androstenedione undergoes conversion to testosterone, which is facilitated by 17β-dehydrogenase type 3 (HSD17B3) in the gonad and also by 17β-dehydrogenase type 5 (AKR1C3) (3) in the adrenal cortex, albeit to a much lesser extent. High-volume production of androgens that bind and activate the androgen receptor, i.e. testosterone and 5α-dihydrotestoserone, and the conversion of androstenedione and testosterone to oestrogens, occurs in the gonad and in part in peripheral target tissues of sex steroid action but not in the adrenal.

Mineralocorticoid synthesis is mainly under the control of the renin–angiotensin–aldosterone system and a potassium feedback loop (see Chapter 5.9). The adrenal zona glomerulosa lacks 17α-hydroxylase activity and pregnenolone is subsequently converted into aldosterone in five enzymatic steps involving the endoplasmic HSD3B2 and CYP21A2 enzymes and mitochondrial CYP11B2 (Fig. 5.11.1). The latter facilitates the three final steps of mineralocorticoid biosynthesis providing 11β-hydroxylase, 18-hydroxylase, and 18-oxidase activities.

The pathophysiology of the different forms of CAH is explained by the specific enzyme deficiency and their consequences on clinical phenotype expression. Table 5.11.1 provides a summary of the various forms.

Steroid 21-hydroxylase deficiency (21OHD) is caused by mutations in the CYP21A2 gene encoding adrenal 21-hydroxylase. 21OHD ranks among the most common inborn errors and accounts for approximately 95% of all cases of CAH. The frequency of the classic form is about 1 in 10 000 to 15 000 livebirths. Nonclassic CAH, caused by milder mutations that do not completely disrupt enzymatic efficiency, is more frequent, with an incidence of about 1 in 500 to 1 in 1000. Glucocorticoid substitution therapy is available since the mid-20th century and the oldest surviving patients with classic CAH are now well within their fifties. Therefore, increasing awareness is necessary not only to address paediatric problems, but also to prevent and treat potential comorbidities during later life (1, 4, 5).

The most severe form due to completely absent 21-hydroxylase enzyme activity comprises mineralocorticoid deficiency, glucocorticoid deficiency, androgen excess (Fig. 5.11.1), and adrenomedullary dysfunction.

Aldosterone action is essential for sodium reabsorption and potassium excretion in the distal renal tubulus. In 21OHD, the deficient conversion of progesterone to 11-deoxycorticosterone results in a lack of aldosterone and its precursors (Fig. 5.11.1). This causes renal salt loss with subsequent severe hyponatraemia, hyperkalaemia, and metabolic acidosis. The clinical course in untreated patients includes dehydration, arterial hypotension, hypovolaemic shock, and finally death due to cardiovascular collapse. This so-called salt-wasting crisis usually develops in the second or third week of life.

Cortisol biosynthesis is impaired due to insufficient enzymatic conversion of 17OHP to 11-deoxycortisol (Fig. 5.11.1). The insufficient cortisol feedback to the hypothalamus and pituitary gland results in increased corticotropin-releasing hormone (CRH) and ACTH secretion leading to the pathological correlate of hyperplastic adrenal glands. Glucocorticoid deficiency results in hypoglycaemia due to impairment of gluconeogenesis, glycogenolysis, proteolysis, and lipolysis. Furthermore, glucocorticoid deficiency lowers myocardial contractility, cardiac output, and decreases the full pressor effects of catecholamines on the cardiovascular system, which can result in cardiovascular shock.

Impaired adrenomedullary development and function has been demonstrated in patients with CAH. Decreased adrenaline levels most likely contribute to the development of hypoglycaemia during intercurrent illness and the development of metabolic consequences such as hyperleptinaemia and hyperinsulinaemia. Adrenaline deficiency is associated with impaired blood glucose response to high-intensity exercise, and explains the failure of hydrocortisone to improve glucose levels during high-intensity exercise.

Accumulated steroid precursors are shunted in the sex hormone biosynthesis pathway resulting in androgen excess (Fig. 5.11.1). Prenatal androgen excess leads to virilization of the external genitalia in 46,XX individuals, i.e. 46,XX disordered sex development (DSD). However, affected patients have normal müllerian structures and thus normal internal female genital organs. The degree of external virilization is classified into five stages according to Prader, spanning a range from isolated mild clitoromegaly (Prader I) to complete labioscrotal fusion with the urethra traversing the penis-like enlarged clitoris (Prader V). External genitalia in affected male 46,XY individuals are normal, but sometimes may be hyperpigmented and slightly enlarged.

A wide range of clinical manifestation of 21OHD exists and can be described as a disease continuum. Commonly, 21OHD is classified into classic (salt-wasting and simple-virilizing forms) and the milder nonclassic form. Disease severity correlates well with the underlying severity of the enzymatic defect. Disease classifications based only on the age of diagnosis is not helpful in clinical practice.

The classic form comprises salt-wasting and simple-virilizing CAH variants with patients usually presenting during childhood. Cortisol deficiency is a characteristic feature. This results in adrenal stimulation and overproduction of steroid precursors that due to the enzymatic block in 21-hydroxylase are redirected towards adrenal androgen synthesis leading to androgen excess. About two-thirds of patients have additional clinically significant aldosterone deficiency and salt loss. Patients with salt-wasting CAH have complete or almost complete absence of 21-hydroxylase function. Patients with glucocorticoid deficiency without apparent mineralocorticoid deficiency are categorized as simple-virilizing CAH.

Almost all patients with classic CAH can be diagnosed by newborn screening within the first 2 weeks of life, before life-threatening salt loss manifests. In countries without implemented CAH newborn screening, girls with ambiguous genitalia are most likely diagnosed soon after birth before manifestation of salt loss. However, the diagnosis in boys with salt-wasting CAH is only established once the patient presents with salt loss. Salt-wasting often manifests with poor feeding, vomiting, failure to thrive, lethargy, and sepsis-like symptoms. The crisis can result in rapid deterioration and if diagnosis is not made in time will lead to a life-threatening situation and consequently death.

Male patients with simple-virilizing CAH, who escaped diagnosis during the neonatal period, commonly present at ages 2 to 7 years with signs of precocious pseudopuberty, including premature pubarche, acne, genitoscrotal hyperpigmentation, increased penile growth, growth acceleration, and advanced bone age. Most patients have a testicular volume in the prepubertal range. However, CAH should be ruled out during the baseline assessment of patients with larger testicular volumes as secondary central precocious puberty, triggered by high circulating androgens of adrenal origin, might have already developed.

The milder, nonclassic form is caused by partial impairment of 21-hydroxylase function and has an estimated incidence of 1:500 to 1 000 in the general population (1, 4, 6). Females are born with normal external genitalia. Most patients can produce sufficient amounts of mineralocorticoids and glucocorticoids, but to the expense of steroid precursor accumulation, leading to increased androgen production. Basal cortisol concentrations are generally normal, but response to 1–24ACTH is insufficient in a significant number of patients (7). Therefore, the need of glucocorticoid substitution has to be established in all patients with nonclassic CAH.

Signs and symptoms at presentation and the age at first presentation of this nonclassic form are highly variable. During childhood premature pubarche, i.e. early onset of pubic hair, and acceleration of growth and bone age are commonly observed signs. Mild clitoromegaly is infrequently found. In later life acne, hirsutism, oligomenorrhoea, sometimes even primary amenorrhoea, and infertility are frequent features. Nonclassic 21OHD is the most common specific cause in women presenting with androgen excess (8). The percentage of undiagnosed patients, in particular males, remains unknown and individuals are regularly diagnosed during family screening after the identification of an affected index patient.

The diagnosis of 21-hydroxylase deficiency has to be considered in all patients with genital ambiguity and/or salt-losing crisis. In case of any genital ambiguity, the karyotype analysis will provide essential information on DSD and guide towards diagnosis of the specific underlying CAH form (Table 5.11.1). The differential diagnosis between 21-hydroxylase deficiency and 11β-hydroxylase deficiency can already be established in the newborn screening using steroid hormone profiling by liquid chromatography–tandem mass spectrometry from filter paper (9). If such a method is unavailable, confirmation tests are similar to the diagnostic procedure for patients diagnosed within a clinical setting without CAH newborn screening (Table 5.11.2).

Randomly timed plasma 17OHP concentrations are significantly increased in classic 21OHD, but should be taken in the morning before 09,00 h to achieve maximal diagnostic value. Commonly, 17OHP concentrations in patients with salt-wasting CAH are higher than in nonsalt-losing patients. A short synacthen test is reserved to investigate borderline cases and is very useful to differentiate between nonclassic CAH and heterozygous carriers (Fig. 5.11.2). A highly specific marker for 21-hydroxylase deficiency is the metabolite 21-deoxycortisol, which is generated by 11-hydroxylation of 17OHP, which only occurs in the absence of 21-hydroxylase activity (Fig. 5.11.1). The analysis of a urine steroid profile is diagnostic with increased metabolites of 17OHP and 21-deoxycortisol. Plasma renin activity should be documented but in the first instance all affected children will be treated with glucocorticoids, mineralocorticoids, and sodium supplementation during the neonatal period and infancy.

In patients with simple-virilizing CAH with delayed diagnosis, plasma 17OHP and urine steroid analysis establish the diagnosis of 21OHD. The degree of 17OHP increase and of sex hormone excess can help to differentiate whether patients are suffering from classic or nonclassic CAH. Measurement of plasma renin activity is needed to assess if patients require additional mineralocorticoid replacement.

Early morning 17OHP concentrations below 2.5 nmol/l in children and below 6.0 nmol/l in women during the follicular phase make the diagnosis of nonclassic CAH unlikely. However, patients with nonclassic CAH may have normal random 17OHP concentrations. A short synacthen test with 17OHP measurements at baseline and after 60 min is the gold standard, and stimulated 17OHP concentrations above 45 nmol/l are diagnostic (Fig. 5.11.2). Cortisol levels should be included in the short synacthen test assessment to identify cases with impaired stress response that would require glucocorticoid cover, at least in increased stress situations such as intercurrent illness. Heterozygous carriers usually have circulating 17OHP levels below 30 nmol/l, but a diagnostic grey area exists for 17OHP concentrations between 30 and 45 nmol/l. Diagnostic sensitivity and specificity can be enhanced by including 21-deoxycortisol and 11-deoxycorticosterone in the measurements before and after ACTH stimulation, but is limited by availability of the tests.

The biochemical diagnosis of 21-hydroxylase deficiency should be confirmed by molecular genetic analysis of the 21-hydroxylase gene, CYP21A2. This provides information on severity of clinical disease expression, facilitates family screening, and aids possible subsequent discussions on future antenatal diagnosis, treatment, and family planning.

The 21-hydroxylase gene (CYP21A2, alias: CYP21, CYP21B) encodes a cytochrome P450 type II enzyme of 495 amino acids. CYP21A2 and its nonfunctional pseudogene (CYP21A1P, alias: CYP21P, CYP21A) are located in the HLA region III on chromosome 6p21.3. Both genes consist of 10 exons sharing a high homology with a nucleotide identity of 98% at exon and of 96% at intron level. They are arranged in tandem repeat with the C4A and C4B genes encoding the fourth factor of the complement system (10).

Complete gene deletions, large gene conversions, chimeric genes, single point mutations, and an 8-bp deletion account for the majority of CYP21A2 mutations (1, 2). Microconversions or apparent gene conversions transferring genetic material from the inactive CYP21A1P pseudogene into the active CYP21A2 gene are the underlying cause for the eight most common point mutations and an 8-bp deletion in exon 3 (Fig. 5.11.3a). In most populations, pseudogene-derived mutations can be detected in similar frequencies. Novel or rare mutations account for about 3–5% of detected mutations in large cohorts. To date, over 90 additional rare pseudogene-independent mutations have been identified (http://www.hgmd.cf.ac.ukhttp://www.cypalleles.ki.se/cyp21.htm). The vast majority of these rare mutations have been identified in single families or small populations. Approximately 1% of CYP21A2-inactivating mutations arise de novo.

About 65–75% of CAH patients are compound heterozygous, i.e. they are affected, but carry different mutations on each chromosome. The clinical phenotype of CAH correlates well with the less severely mutated allele, and consequently with the allele encoding for the higher residual activity of 21-hydroxylase. This has major implications for genetic counselling in patients with nonclassic CAH. If a patient with nonclassic CAH is compound heterozygous for a mild and a severe mutation, the risk of having a child with classic CAH increases significantly to about 1 in 400 (1/50 × 1/2 × 1/4) assuming a heterozygous rate of 1/50 for classic mutations in the general population.

The correlation of genotype with the extent of glucocorticoid and mineralocorticoid deficiency is strong (Fig. 5.11.3b). However, divergence between genotype and phenotype occurs in some cases. Although a trend exists, the correlation between the genotype and the virilization phenotype assessed by Prader genital stages is less pronounced (Fig. 5.11.3b). This implies the importance of other factors modifying clinical androgen effects. This observed variability might be influenced by CAG repeat length of the androgen receptor modulating androgen action (11). Potential variations in the degree of recovery from glucocorticoid and mineralocorticoid deficiency during later life might be explained by significant 21-hydroxylase activity of the extra-adrenal enzymes CYP2C19 and CYP3A4 (12).

Therapeutic management of CAH is challenging and treatment objectives differ with age. Generally, treatment includes glucocorticoid and mineralocorticoid replacement and also aims at the control of adrenal androgen excess. Therapeutic goals in affected children are the prevention of adrenal crisis and precocious pseudopuberty, and protection from long-term complications. Importantly, management in affected girls has to address the issue of genital corrective surgery and psychosexual development. In adults, long-term morbidity and infertility are in the focus.

Hydrocortisone is recommended for replacement therapy from the newborn period to adolescence (13). The average physiological cortisol secretion rate is about 8 mg/m² per day. Typical hydrocortisone doses are 10–15 mg/m² divided in three daily doses, with doses up to 25 mg/m² in infancy only seldom required. These doses are higher than those employed for replacement of adrenal insufficiency, because treatment also aims at normalization of ACTH-driven adrenal androgen excess. Cortisone acetate is not recommended as it requires activation to cortisol by 11β-hydroxysteroid dehydrogenase type 1, which leads to considerable interindividual variability in pharmacokinetics of cortisone acetate. The optimal timing for providing the highest dose of hydrocortisone remains unsolved with no endpoint data supporting either a circadian or reverse-circadian replacement strategy. Providing the highest dose in the evening has no or a minor effect on the ACTH surge occurring during the early morning hours. However, providing the highest dose in the morning, as done in the majority of paediatric endocrine centres in Europe, may also not suppress the ACTH surge sufficiently as hydrocortisone is usually given around 7 am, thus 4–5 h after the surge (14). Ideally, the early glucocorticoid dose is given between 03.00 and 04.00 h, but this approach is rarely tolerated by patients and parents. Long-acting synthetic glucocorticoids, such as prednisone, prednisolone, and dexamethasone, are more likely to be associated with growth suppression and weight gain and should be avoided before final height is achieved.

At final height patients may be changed to treatment with longer-acting glucocorticoids, e.g. with prednisolone (2–4 mg/m² per day) and dexamethasone (0.25–0.375 mg/m² per day, seldom more than 0.5 mg total daily dose). Reassessment of mineralocorticoid deficiency is of paramount importance as prednisolone delivers reduced and dexamethasone exhibits virtually no mineralocorticoid activity. By contrast, hydrocortisone can mask mineralocorticoid deficiency by binding the mineralocorticoid receptor with similar affinity to aldosterone (0.1 mg fludrocortisone is equivalent to 40 mg hydrocortisone).

Several monitoring strategies for corticosteroid therapy in CAH have been described, including clinical and biochemical markers (Table 5.11.3). Data on the superiority of either approach are not available. Similar to the situation in adrenal insufficiency (see Chapter 5.9) cortisol measurements are not useful to monitor quality of glucocorticoid substitution in adrenal insufficiency (15). Treatment of CAH should aim to normalize sex hormones. Commonly, the optimal glucocorticoid dose avoids suppression of 17OHP while maintaining sex hormone concentrations in the mid age- and sex-specific normal range.

Mineralocorticoid replacement is required in all patients with classic CAH, at least during infancy. Fludrocortisone doses during the first year of life are commonly 150 μg/m² per day and should be adjusted according to individual requirements. Total dose of 300 μg per day might be required (13). Sodium needs to be supplemented as milk feeds only provide maintenance sodium requirements. Sodium supplements up to 10 mmol/kg per day may be required at least in the first 6 months of life (16). Sodium supplementation may be discontinued when salt intake is sufficient via food. Sodium supplementation can be beneficial also in later life during episodes of increased sodium loss such as hot weather and intense exercise. Adequate mineralocorticoid replacement generally leads to hydrocortisone dose reduction. The need to continue this therapy should be reassessed after infancy or early childhood using plasma renin activity and blood pressure as reliable markers (13). The relative dose in relation to body surface decreases throughout life. After the first 2 years of life, fludrocortisone doses of 100 μg/m² per day are commonly sufficient. This requirement drops further with adolescents and adults are usually sufficiently supplemented with a total daily dose of 100 to 200 μg (50 to 100 μg/m² per day). Mineralocorticoid substitution is monitored by plasma renin, aiming at the upper normal reference range, and also by blood pressure measurements using age, sex, and height-adjusted references. A significant drop (>15 mmHg) in systolic blood pressure between seated and erect blood pressure recordings indicates postural hypotension suggestive of insufficient mineralocorticoid replacement.

Adrenal crisis due to impaired cortisol response to stress is a serious threat in CAH. During febrile illness (>38.5 °C), trauma, and surgery the daily hydrocortisone dose should be doubled or tripled, with intravenous or intramuscular administration oral doses where appropriate. Fludrocortisone adjustment is commonly not required. However, extra sodium supplement may be necessary. Special attention should be paid towards glucose supply during severe illness as patients tend to hypoglycaemia due to impaired adrenomedullary function. Patients with diarrhoea and vomiting, who are unable to take their medication require IM hydrocortisone (100 mg/m² per dose, maximum 100 mg) and immediate review by a medical professional. Patients with severe illness or major surgery require IV hydrocortisone (Table 5.11.4). All CAH patients must carry a steroid emergency card or MedicAlert bracelet emphasizing the diagnosis ‘adrenal insufficiency’ in addition to CAH. Patients should have an emergency glucocorticoid injection kit and patients (and parents and partners) should undergo self-injection training.

Genital surgery should achieve a genital appearance compatible with gender, unobstructed urinary emptying without incontinence or infections, and good adult sexual and reproductive function (13). A one-stage surgical approach following the latest techniques of vaginoplasty, clitoral, and labial surgery has been recommended (13). Surgery is recommended to be timed at age 2 to 6 months. Surgical procedures between age 12 months and adolescence are usually not recommended in the absence of medical problems. Clitorectomy is absolutely contraindicated at any stage. Vaginal dilations are contraindicated until adolescence. It is important that female CAH patients remain under the follow-up of a specialized gynaecologist throughout adolescence and adulthood.

Prenatal treatment is carried out with dexamethasone, which crosses the placenta and therefore can suppress the fetal HPA axis that drives androgen excess and virilization. However, prenatal treatment is controversial as the safety, including metabolic and psychointellectual long-term consequences of dexamethasone treatment, remains to be fully defined. Prenatal treatment has been shown to be effective to prevent severe genital virilization if started early enough, as major developments in fetal sexual differentiation take place between gestational weeks 4–10. Therefore, dexamethasone treatment needs to be established as soon as pregnancy is confirmed and ideally before the sixth and no later than the eighth week of gestation to achieve significant benefit in preventing 46,XX DSD. Counselling regarding prenatal dexamethasone therapy should be carried out ideally well before conception, involving an endocrinologist, fetal medicine specialist, and clinical geneticist. Patients should be included in ongoing multicentre studies with available protocols (17). The suggested dose is 20–25 μg/kg in three divided doses (total maximum dose 1.5 mg/day) (1). Maternal side effects resemble those of high-dose glucocorticoid treatment including oedema, striae, weight gain, mood fluctuations, and sleep disturbances. Arterial hypertension and impaired glucose tolerance seem not to be increased, but close monitoring for these complications should be carried out. Only one in eight children will benefit from prenatal dexamethasone treatment if the parents are heterozygous carriers, as the chance of carrying an affected girl is 12.5%. All other children are exposed to dexamethasone without benefit. The number of unnecessarily treated cases can be reduced to three out of eight using modern molecular genetic techniques. The fetal sex can be determined as early as week six of gestation by analysing free fetal DNA from maternal blood by using real time PCR approaches. Dexamethasone treatment can be stopped if the fetus is determined to be male. Otherwise, the fetal sex is established from a chorionic villous biopsy. If the fetal karyotype is 46,XX, CYP21A2 mutation analysis is performed and subsequently dexamethasone treatment stopped in pregnancies with an unaffected female. In case of an affected female fetus, current recommendations suggest that dexamethasone treatment should be continued up until delivery.

Two main additional experimental pharmacological therapies in the paediatric setting have focussed on the improvement of final height. A promising approach within a study setting uses a combination of the antiandrogen flutamide to lower glucocorticoid doses (8 mg/m² per day) and the aromatase inhibitor testolactone to reduce oestrogen-mediated fusion of the growth plate. Final outcome data are unavailable, but 2-year follow-up data showed normal linear growth in children treated with this regimen. Another study, including 14 patients, achieved improvement of growth and final height with gonadotropin-releasing-hormone agonists used alone and in combination with growth hormone. Long-term safety and efficiency data are unavailable.

Bilateral adrenalectomy has been proposed as an alternative in severe CAH. However, the experience with bilateral adrenalectomy in CAH is limited. Long-term follow-up data indicated improved signs and symptoms of hyperandrogenism and less obesity after surgery. It has also been reported as a therapeutic approach to achieve successful pregnancies. However, this procedure bears also a number of risk, including surgical and anaesthetic complications and leaving the patients completely adrenal insufficient and thus exposing them to a higher risk of adrenal crisis. Currently, bilateral adrenalectomy for CAH has to be considered an experimental therapeutic strategy and its appropriateness needs to be carefully considered.

Gender-related behaviour demonstrates shifts from female to male even in patients with the milder nonclassic CAH, becoming more obvious in the more severe CAH forms (18). Females with CAH have a more male-typical childhood play behaviour and more male-typical cognitive functioning during childhood than unaffected girls. The impact on adult life is unclear as adult women with CAH do not show a more male-typical cognitive pattern. Psychological health is not compromised and psychological adjustment is not significantly associated with genital virilization or age at genital surgery. Gender assignment is commonly not a problem even in heavily virilized 46,XX individuals. Importantly, most female CAH patients do not experience serious gender identity problems, such as gender dysphoria, reported in only 5% of 250 patients. Similar numbers have been observed in a series of 63 female CAH patients (18). Conversely, 90% of female CAH patients raised as males (n = 33) had a normal male gender identity (19).

Consequences of genital corrective surgery are of importance during adolescence and early adulthood. Inadequate vaginal reconstruction, which may be present in more than 50% of adult patients (20), and reduced clitoral sensitivity, impacts on sexual activity and sexual experience. This often has downstream effects on partner choice, steady relationships, marital status, and fertility (18). Although the surgical approach is continuously improving, physicians involved in the care of adult CAH patients will be confronted with the consequences of previous, now abandoned surgical techniques for several decades to follow. Another important mechanism resulting in sexual dysfunction in CAH patients is near complete suppression of sex hormones due to glucocorticoid overtreatment, with subsequently low libido.

The final height outcome in many patients is not optimal. A meta-analysis, including 18 studies between 1977 and 2001, showed that the mean adult height in classic CAH was 10 cm (−1.4 standard deviation score (SDS)) below the population mean and −1.2 SDS calculated for target height. The pubertal growth spurt occurs earlier and is less pronounced than in the normal population. Hyperandrogenism and overexposure to glucocorticoids both contribute to the problem. Sex hormone excess leads to accelerated growth, early fusion of the epiphysis, and can also trigger secondary central precocious puberty, which further exacerbates the situation.

Glucocorticoid overexposure inhibits growth. Special attention should be drawn towards the infancy growth spurt during the first 2 years of life. This growth phase is characterized by the highest postnatal growth velocity and an impaired growth velocity during this time significantly impacts on final height outcome. Therefore, the lowest optimal dose for glucocorticoid substitution and sex hormone normalization has to be achieved as early in life as possible.

Obesity and increased fat mass is common amongst children and adolescents with CAH (21–23). Birth weight and length, serum leptin concentrations, or type of glucocorticoid and mineralocorticoid dose were not associated with obesity in 89 CAH patients (0.2–17.9 years). However, glucocorticoid dose, chronologic age, advanced bone age maturation, and parental obesity contributed to elevated body mass index (BMI)-SDS (22). CAH females older than 30 years had increased fat mass and higher insulin levels. However, clear evidence of cardiovascular risk factors could not be shown. Women with CAH have a significantly higher rate of gestational diabetes as a risk factor for the development of type 2 diabetes (24). Females with nonclassic CAH (25) and young adult CAH patients (26) have reduced insulin sensitivity. The risk of atherosclerosis might be increased; increased intima media thickness as a marker of atherosclerosis has been detected (26). The intima media thickness was independent of cardiovascular risk factors such as BMI, elevated blood pressure, or lipid profile changes (26).

Daytime systolic blood pressure in children and adolescents with CAH is elevated and the physiological nocturnal dip in blood pressure is absent (27). Elevated systolic blood pressure correlates with the degree of overweight and obesity. CAH patients with normal weight tend to suffer more frequently from diastolic hypotension (28). Older CAH patients had a higher standing diastolic blood pressure than younger patients, which was not observed in the control group (24). Data on mortality are unavailable.

Bone mineral density (BMD) is usually not grossly impaired if patients receive an appropriate glucocorticoid dose. Low BMD appears to be associated with glucocorticoid overtreatment. Androgen excess and subsequent aromatization can lead to enhanced bone density. A reduction of bone turnover markers in conjunction with normal BMD has been noticed by several groups (29, 30).

Fertility prognosis has been improving over the recent years and is now less pessimistic than described in the first reports. Most adult males with CAH are fertile, but the impact on male fertility has possibly been underestimated.

The fertility rate is the lowest in salt-wasting CAH patients, with markedly increasing fertility in patients with simple-virilizing and nonclassic CAH. The aetiology of decreased female fertility is multifactorial (Box 5.11.1). Optimization of fertility can often be achieved by close monitoring and adjustment of glucocorticoid replacement with the longer acting prednisolone. The situation may prove difficult because both over-replacement and under-replacement with glucocorticoids can result in anovulation. Even after achieving good control of 17-hydroxyprogesterone production, serum progesterone levels may remain elevated (31) thereby impairing follicle maturation and implantation of the fertilized egg. The successful use of adrenalectomy to achieve pregnancy has been reported in two patients (32).

Prior to pregnancy, the carrier status of the male partner should be established. If the partner is not a carrier for CAH, the developing child will be an obligatory, clinically healthy carrier. If the partner of a CAH patient desiring pregnancy happens to be a heterozygous CYP21A2 mutation carrier, the patient should be counselled with regard to the possibility of prenatal dexamethasone treatment. Recommendations for the management of women with adrenal insufficiency suggest that the glucocorticoid dose should be increased by 30–50% during the last trimester of pregnancy (33). Although it has been suggested that glucocorticoids have to be rarely adjusted during pregnancy (29), spontaneous miscarriage risk in untreated women with nonclassic CAH is significantly higher than in treated patients (34). Mineralocorticoid requirements may sometimes increase as well, due to the antimineralocorticoid properties of progesterone. Adjustment of the mineralocorticoid dose has to be performed according to postural blood pressure response, and serum sodium and potassium concentrations. Plasma renin activity is physiologically increased during pregnancy and therefore cannot serve as a monitoring tool. Delivery requires glucocorticoid coverage at doses recommended for major surgical stress, i.e. 100–200 mg/24 h, either per continuous intravenous infusion in a 5% glucose solution or per intramuscular injection (e.g. 50 mg four times per day), with rapid tapering after delivery if the clinical situation permits. Patients who underwent corrective surgery for ambiguous genitalia will more likely require a caesarean section.

Two major issues are recognized to impact on male fertility (Box 5.11.1). Hypogonadotrophic hypogonadism is a consequence of increased aromatization of adrenal androgens, in particular androstenedione to oestrone. This results in suppression of pituitary luteinizing hormone and follicle-stimulating hormone secretion impacting on testicular androgen synthesis and spermatogenesis. The condition is reversible after optimization of glucocorticoid therapy.

Benign testicular adrenal rest tumours (TARTs) have been correlated with male infertility (35). Embryologically, testes and adrenal cortex both develop from the urogenital ridge. TARTs arise from adrenal cell nests within the testicular tissue that are subject to continuous ACTH stimulation. TART can result in Leydig cell failure and/or oligospermia. High-dose glucocorticoid treatment may reverse infertility. However, even high doses of steroids may not be sufficient to restore testicular function. Testes-sparing surgery may not reliably restore testicular function. TARTs have been detected in male patients as early as 7 years of age. Early treatment optimization to reduce these hyperplasic areas within the testes appears to be paramount to improve long-term fertility outcome. Of note, TARTs represent a benign entity that responds to glucocorticoid treatment in the early stages and should not be confused with testicular tumours. Treating physicians and urological surgeons need to be aware of this entity to avoid unnecessary gonadectomies based on the suspicion of seminoma.

About 5–8% of CAH cases are due to 11β-hydroxylase deficiency (11OHD), which is equivalent to an incidence of 1 in 100  000 to 200 000 livebirths in nonconsanguineous populations (36). Steroid 11β-hydroxylase (CYP11B1) catalyses the final step in cortisol biosynthesis, the conversion of 11-deoxycortisol to cortisol (Fig. 5.11.1). It also catalyses the conversion of 11-deoxycorticosterone (DOC) to corticosterone, but is lacking noteworthy 18-hydroxylase and 18-oxidase activity (Fig. 5.11.1). Thus 11OHD results in decreased cortisol secretion and accumulation of the glucocorticoid precursor 11-deoxycortisol and the mineralocorticoid precursor DOC (Fig. 5.11.1). DOC activates the mineralocorticoid receptor and may lead to significant arterial hypertension. Accumulated precursors are shunted into the androgen synthesis pathway, leading to hyperandrogenism. Basal concentrations of 17OHP are commonly increased, but may be normal even during the first weeks of life (37).

Classic 11OHD results in virilization of the external genitalia in newborn females, and later on leads to precocious pseudopuberty combined with rapid somatic growth and bone age acceleration in both sexes. Nonclassic 11OHD is a rare condition (36, 38). Affected female patients are born with normal genitalia and present with signs of androgen excess during childhood. Alternatively, they may present as adults with hirsutism and oligomenorrhoea, though certainly only a small minority of women presenting with signs and symptoms suggestive of polycystic ovary syndrome suffer from nonclassic 11β-hydroxylase deficiency.

Steroid 11β-hydroxylase deficiency is caused by mutations in the 11β-hydroxylase gene (CYP11B1), which is localized on chromosome 8q21 approximately 40 kb from the highly homologous aldosterone synthase gene (CYP11B2) (39). CYP11B1-inactivating mutations have been shown to be distributed over the entire coding region consisting of nine exons. Although a cluster is reported in exons 2, 6, 7, and 8 (40, 41), real hot spots such as in 21OHD do not exist. A broad variety of mutations have been reported to cause either classic or nonclassic 11OHD (40, 63).

Glucocorticoid replacement follows the same rules as in 21-hydroxylase deficiency. The blood pressure is often well controlled under glucocorticoid substitution. However, if no blood pressure control can be achieved, antihypertensive treatment should be commenced at an early stage and excessive glucocorticoid exposure should be avoided.

Steroid 17α-hydroxylase deficiency (17OHD) is a rare form of CAH. It accounts for about 1% of all CAH cases and affects adrenal and gonadal steroid biosynthesis. The 17α-hydroxylase enzyme (CYP17A1) catalyses two different enzymatic reactions: firstly, the 17α-hydroxylation of pregnenolone and progesterone and, secondly, via its 17,20 lyase activity, the conversion of 17-hydroxypregenolone to DHEA and with lesser efficiency also that of 17OHP to androstenedione (Fig. 5.11.1). As a consequence, 17OHD results in both glucocorticoid deficiency and sex steroid deficiency. In addition, the mineralocorticoid precursors corticosterone and DOC accumulate (Fig. 5.11.1) Corticosterone has weaker glucocorticoid activity than cortisol, but corticosterone excess production generally prevents adrenal crisis in patients with 17OHD. Accumulation of corticosterone and DOC result in excess mineralocorticoid activity, causing severe hypokalaemic hypertension. Sex steroid deficiency caused by loss of 17,20 lyase activity results in 46,XY DSD presenting as undervirilization in male newborns and in primary amenorrhoea in 46,XX individuals. There is lack of pubertal development due to hypergonadotrophic hypogonadism in both sexes (42).

Due to the low incidence of adrenal crisis in untreated 17OHD, the diagnosis is often only established during adolescence or early adulthood following investigations for hypokalaemic hypertension or delayed pubertal development (42). This fact emphasizes the importance of blood pressure measurement as a clinical screening tool in all patients with delayed puberty. Typical biochemical findings include raised ACTH levels and suppressed plasma renin activity whilst serum aldosterone is decreased, with sex steroid deficiency further confirming the diagnosis (Table 5.11.1). Glucocorticoid replacement commonly normalizes plasma renin activity, aldosterone, blood pressure, and electrolyte disturbances. Doses are lower than required for treatment of 21OHD and 11OHD. Substitution of sex hormones is generally required.

A rare variant of 17OHD has been described, isolated 17,20 lyase deficiency with largely preserved 17α-hydroxylase activity. This manifests with impaired sex steroid biosynthesis only, without concurrent evidence of mineralocorticoid excess or glucocorticoid deficiency.

The CYP17A1 gene consists of eight exons and is located on chromosome 10q24.3. A variety of different mutations have been described, without evidence of a hot spot. Mutations underlying the isolated 17,20 lyase deficiency variant are located within the area of the CYP17A1 molecule that is thought to interact with the cofactor cytochrome b5_, thereby disrupting the electron transfer from POR to CYP17A1, specifically disrupting the conversion of 17OH-pregnenolone to DHEA (43, 44) (Fig. 5.11.1).

Steroid 3β-hydroxysteroid-dehydrogenase type 2 (HSD3B2) deficiency represents a rare CAH variant and data on population-based incidence are lacking. HSD3B2, also termed Δ4/Δ5-isomerase, catalyses three key reactions in adrenal steroidogenesis: the conversion of the Δ5-steroids pregnenolone, 17OH-pregnenolone and DHEA to the Δ4-steroids progesterone, 17OHP and androstenedione, respectively (Fig. 5.11.1). Thereby HSD3B2 deficiency affects all three biosynthetic pathways (mineralocorticoids, glucocorticoids, sex steroids). The clinical spectrum shows a wide variety of disease expression, ranging from a severe salt-wasting form, with or without ambiguous genitalia in affected male neonates, to isolated premature pubarche in infants and children of both sexes and late-onset variant manifesting with hirsutism and menstrual irregularities. Patients with mild biochemical late-onset deficiency are commonly HSD3B2 mutation negative. There is no strong correlation between salt-wasting and male undervirilization, primarily presenting with mostly perineoscrotal hypospadias and bifid scrotum. Female patients diagnosed during neonatal or infant life usually present with normal genitalia, though some cases of minor clitoromegaly have been reported. The diagnosis of HSD3B2 deficiency is often delayed in affected individuals without salt-wasting and with normal genitalia. Furthermore, HSD3B2-deficient patients are at risk to be misdiagnosed as suffering from of 21OHD (45).

The biochemical diagnosis of HSD3B2 deficiency is usually established by the elevated concentrations of Δ5-steroids, such as DHEA, 17OH-pregnenolone, and their metabolites, and a high ratio of Δ5 to Δ4 steroids or their respective urinary metabolites (45) (Fig. 5.11.1). Hormonal criteria have recently been refined for the diagnosis of HSD3B2 deficiency based on genotyping of the HSD3B2 gene. 17OH-pregnenolone concentrations and 17OH-pregnenolone to cortisol ratios at baseline and after ACTH stimulations are of the highest discriminatory value in differentiating between patients affected by HSD3B2 deficiency and patients with milder biochemical abnormalities, who are negative for HSD3B2 mutations (46, 47).

Two isoforms of 3β-hydroxysteroid dehydrogenase, 3β-HSD type 1 and 3β-HSD type 2, exist, which are encoded by the HSD3B1 and HSD3B2 genes, respectively. The HSD3B2 gene is located on chromosome 1p13·1 and consists of four exons. 3β-HSD2 is mainly present in the adrenal and the gonad, while 3β-HSD1 is present in the placenta and almost ubiquitously in peripheral target tissues (45, 48). Both enzymes are NAD-dependant short chain dehydrogenases. HSD3B2 deficiency is caused by mutations in the HSD3B2 gene. A reasonable degree of genotype–phenotype correlation with regard to mineralocorticoid deficiency exists, with major loss of function mutations resulting in the salt-wasting form and partial inactivating mutations allowing for some residual aldosterone synthesis capacity. However, the genotype cannot be used to predict the degree of male undervirilization (45).

ORD is the underlying cause of CAH presenting with apparent combined CYP17A1–CYP21A2 deficiency, which was first described in 1985 (49). However, the molecular pathology has only recently been elucidated as inactivating mutations in the electron donor enzyme POR which provides electrons to all microsomal CYP enzymes including CYP17A1 and CYP21A2 (50, 51). The incidence of ORD is unknown, but a considerable number of patients have been described since the molecular characterization of ORD.

The majority of ORD patients described have skeletal malformations (Box 5.11.2) resembling the Antley–Bixler syndrome phenotype with predominantly craniofacial malformations. Endocrine dysfunction is characterized by adrenal and gonadal insufficiency and disordered sexual development which may occur in affected individuals of both sexes (46,XX DSD and 46,XY DSD). An Antley–Bixler syndrome phenotype can also be caused by autosomal dominant mutations in the fibroblast growth factor receptor 2 gene (FGFR2), which does not manifest with abnormalities of steroid metabolism or ambiguous genitalia (52). Impairment of sterol biosynthesis, specifically of POR-dependent 14α-lanosterol demethylase (CYP51A1), may be causative for the development of skeletal malformation. This is supported by the finding that children born to mothers treated during pregnancy with the CYP51A1 inhibitor fluconazole show evidence of Antley–Bixler syndrome-like skeletal malformations.

Severe sexual ambiguity in ORD can be found in both sexes. Affected girls may present with significant virilization of the external genitalia. Affected boys can be undervirilized, with degrees varying from borderline micropenis to perineoscrotal hypospadias. Progressive postnatal virilization in affected girls does not occur and circulating sex hormone concentrations are invariably low or low normal in both sexes. Mothers pregnant with an affected child may present with virilization manifesting during midgestation and have often low oestriol concentrations. Generally, the androgen excess reverses after delivery (53).

Undervirilization in affected boys is easily conceivable based on the impairment of CYP17A1 function. The potential existence of an alternative ‘backdoor’ pathway towards prenatal androgen synthesis has been described, potentially explaining virilization in affected girls. Postnatally, the alternative pathway ceases and the conventional androgen pathway remains inefficient due to the POR mutations.

Pubertal development in ORD is not well studied yet. It appears to be dominated by the consequences of sex steroid deficiency (54). A common finding in females diagnosed in early adolescence are polycystic ovaries. Females may have large ovarian cysts that have a tendency to rupture and bilateral polycystic ovaries have even been reported in a 2-month old baby with ORD.

Typical biochemical findings include raised 17OHP, albeit not to the extent observed in 21-hydroxylase deficiency. In contrast to 21OHD, sex steroids are low and there is commonly no mineralocorticoid deficiency. The gold standard for diagnosis of ORD is GC/MS analysis of urinary steroid excretion. The metabolome is characterized by accumulation of pregnenolone and progesterone metabolites alongside low androgen metabolites and increased 17OHP metabolites, indicating pathognomonic combined CYP17A1–CYP21A2 deficiency (Fig. 5.11.1). Analysis of serum steroids may lead to misdiagnosis of patients because features of 17OHD and 21OHD are present in variable combinations (55). Prenatal biochemical diagnosis is possible as mothers pregnant with an affected child often present with low serum oestriol and a characteristic urinary steroid profile (2, 56). Recent data suggest that at least 50% of patients can be detected in newborn 17OHP screening (54).

Baseline glucocorticoid secretion is often sufficient, but the cortisol response to stress is usually impaired (54, 56). Affected patients without hydrocortisone replacement are at a high risk for developing a life-threatening adrenal crisis. Glucocorticoids are required in replacement doses only (commonly hydrocortisone 8–10 mg/m² per day) because of absent postnatal androgen excess. Mineralocorticoid production is generally uncompromised, plasma renin activity and serum aldosterone are generally normal. However, some patients show increased excretion of mineralocorticoid metabolites (51) and mild hypertension (50). The POR gene is located on chromosome 7q11.2. It consists of 15 translated exons spanning a region of approximately 32.9 kb and encodes for a protein of 680 amino acids. A variety of POR-inactivating mutations have been reported, including missense, frameshift, and splice site mutations (http://www.cypalleles.ki.se/por.htm). A287P is the most common mutation in Caucasians, while R457H is the most frequent founder mutation in the Japanese population. Although genotype–phenotype correlations are not fully established yet, certain patterns are evolving suggestive of genotype–phenotype correlations predicting the presence and severity of skeletal malformations as well as the correlation of karyotype and presence of genital ambiguity (54).

StAR mobilizes cholesterol from the outer mitochondrial membrane to the inner mitochondrial membrane (Fig. 5.11.1). StAR-independent cholesterol transport only occurs at a low rate. Therefore, a defect in StAR leads to almost no substrate provision for P450 side-chain cleavage and the production of all steroid hormones from adrenal and gonad is severely reduced. In contrast to the conventional CAH forms, the adrenals of individuals affected by CLAH show a characteristic accumulation of lipids, predominantly cholesterol esters (57). The most severe form presents with 46,XY DSD and combined adrenal insufficiency. Salt-wasting typically develops in the neonatal period or after a few weeks of life, but later onset also occurs. Females can show spontaneous pubertal development. Recently a milder form of StAR deficiency has been described with normally virilized 46,XY individuals, who presented with adrenal failure during early childhood (58). Treatment consists of glucocorticoid and mineralocorticoid replacement, and substitution of sex hormones in later life.

The deficiency of P450 side-chain cleavage (CYP11A1) enzyme is a rare inborn error of steroidogenesis. It presents clinically and biochemically with similar signs and symptoms as CLAH caused by StAR mutations. However, all patients with CYP11A1 deficiency had small or normal-sized adrenals (59). Depending on the impairment of CYP11A1 function a spectrum of clinical presentation ranging from 46,XY DSD with severe adrenal insufficiency in the newborn period over midshaft hypospadias and cryptorchidism and later manifestation of adrenal insufficiency during childhood (60). Concentrations of all steroid hormones are characteristically decreased as the first step in steroidogenesis, the conversion of cholesterol to pregnenolone is impaired. Treatment is similar to CLAH.

Aldosterone synthase (CYP11B2, corticosterone methyloxidase, CMO) deficiency (ASD) is a rare condition causing isolated mineralocorticoid deficiency (61). Patients present during the first days to weeks of life. Since patients are not glucocorticoid deficient and can synthesize DOC (and variable levels of corticosterone) the salt-wasting crisis is commonly less pronounced than in 21OHD. Two biochemical forms exist: ASD 1 (CMO I) has an increased ratio of corticosterone to 18OH-corticosterone and decreased 18OH-corticosterone to aldosterone ratio, whereas corticosterone to 18OH-corticosterone is decreased and 18OH-corticosterone to aldosterone is increased in ADS 2 (CMO II). Both forms are associated with mutations in the CYP11B2 gene. The underlying molecular pathology defining these different forms is not fully understood. Patients with CYP11B2 deficiency generally respond well to fludrocortisone (start dose 150 μg/m² per day in neonates and infancy) and will also benefit from salt supplementation. Patients, who manifested with failure to thrive, generally show a good catch-up growth after initiation of treatment. Electrolytes often tend to normalize from age 3 to 4 years. Untreated patients are at significant risk of being growth retarded. Adults are generally asymptomatic, but are more susceptible to salt loss. The need for mineralocorticoid treatment in later life has to be established individually.

Apparent cortisone reductase deficiency is characterized by hyperandrogenism resulting in hirsutism, oligoamenorrhoea, and infertility in females and premature pseudopuberty in males. The condition is caused by mutations in the gene encoding hexose-6-dehydrogenase (62), which provides NADPH to 11β-hydro-xysteroid dehydrogenase type 1 (HSD11B1). HSD11B1 activates inactive cortisone to cortisol within target tissues of glucocorticoid action, namely in the liver and adipose (Fig. 5.11.1). Defects in this system result in increased cortisol clearance leading to activation of the HPA axis and ACTH-mediated adrenal androgen excess.