Materiały źródłowe

• #1 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Congenital adrenal hyperplasia (CAH) encompasses a group of autosomal recessive conditions caused by genetic mutations that disrupt enzymes responsible for producing glucocorticoids, mineralocorticoids, and sex steroids from cholesterol in the adrenal glands. […] These mutations impair cortisol synthesis, prompting a compensatory increase in adrenocorticotropic hormone, which leads to adrenal cortex hyperplasia hence the name „CAH.” The condition manifests in infants, children, or adults with symptoms of corticosteroid deficiency, often accompanied by either a deficiency or an excess of mineralocorticoids and sex steroids, depending on the specific enzyme defect. […] The most common form, 21-hydroxylase (21-OH) deficiency, accounts for approximately 95% of cases. Clinical presentations vary by enzyme defect, ranging from ambiguous genitalia in genotypic females to potentially life-threatening adrenal insufficiency in males.

• #2 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Congenital adrenal hyperplasia (CAH) refers to a group of autosomal recessive conditions caused by mutations in genes encoding enzymes involved in the production of glucocorticoids, and sometimes mineralocorticoids and sex steroids, from cholesterol in the adrenal glands. […] These mutations impair cortisol synthesis, triggering a compensatory increase in adrenocorticotropic hormone (ACTH), which leads to adrenal cortex hyperplasia hence the term „CAH.” […] Gene mutations that cause defects in steroidogenesis are classified as CAH and involve the following enzymes and proteins: 21-Hydroxylase (21-OH), 11-Hydroxylase (11-OH), 3-Hydroxysteroid dehydrogenase type-2 (3-HSD-2), 17-Hydroxylase/17,20-lyase (17-OH), P450 oxidoreductase (POR), Steroidogenic acute regulatory protein (StAR), Cholesterol side-chain cleavage enzyme (SCC).

• #3 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Congenital adrenal hyperplasia (CAH) encompasses a group of autosomal recessive conditions caused by genetic mutations that disrupt enzymes responsible for producing glucocorticoids, mineralocorticoids, and sex steroids from cholesterol in the adrenal glands. […] These mutations impair cortisol synthesis, prompting a compensatory increase in adrenocorticotropic hormone, which leads to adrenal cortex hyperplasia hence the name „CAH.” The condition manifests in infants, children, or adults with symptoms of corticosteroid deficiency, often accompanied by either a deficiency or an excess of mineralocorticoids and sex steroids, depending on the specific enzyme defect. […] The most common form, 21-hydroxylase (21-OH) deficiency, accounts for approximately 95% of cases. Clinical presentations vary by enzyme defect, ranging from ambiguous genitalia in genotypic females to potentially life-threatening adrenal insufficiency in males.

• #4 Congenital adrenal hyperplasia (CAH) | Great Ormond Street Hospital

  https://www.gosh.nhs.uk/conditions-and-treatments/conditions-we-treat/congenital-adrenal-hyperplasia-cah/

  Congenital adrenal hyperplasia is group of inherited conditions that are present at birth (congenital) where the adrenal gland is larger than usual (hyperplasia). In CAH, the body is missing an enzyme (chemical substance) that stimulates the adrenal glands to release the cortisol hormone. Lacking this hormone means that the body is less able to cope with physiological stress, which can be life threatening. It also makes the level of androgen (male hormone) increase, which causes male characteristics to appear early in boys or inappropriately in girls. […] CAH is an inherited disorder, that is, it is passed on from parent to child. Most types of CAH are autosomal recessive disorders; this means that both parents are carriers of the disease. […] In a small proportion of people, CAH is caused by a gene mutation (change) that happens by chance and cannot be predicted. A number of genes have been identified as causing different types of CAH for instance, the most common form of CAH is called 21-hydroxylase deficiency and results from the gene labelled CYP21 being absent or changed. This means that aldosterone and cortisol are not produced but production of androgen is unaffected. The gene affected results in different types of CAH, affecting people to varying degrees from mild to severe.

• #5 Congenital adrenal hyperplasia (CAH) | Great Ormond Street Hospital

  https://www.gosh.nhs.uk/conditions-and-treatments/conditions-we-treat/congenital-adrenal-hyperplasia-cah/

  Congenital adrenal hyperplasia is group of inherited conditions that are present at birth (congenital) where the adrenal gland is larger than usual (hyperplasia). In CAH, the body is missing an enzyme (chemical substance) that stimulates the adrenal glands to release the cortisol hormone. Lacking this hormone means that the body is less able to cope with physiological stress, which can be life threatening. It also makes the level of androgen (male hormone) increase, which causes male characteristics to appear early in boys or inappropriately in girls. […] CAH is an inherited disorder, that is, it is passed on from parent to child. Most types of CAH are autosomal recessive disorders; this means that both parents are carriers of the disease. […] In a small proportion of people, CAH is caused by a gene mutation (change) that happens by chance and cannot be predicted. A number of genes have been identified as causing different types of CAH for instance, the most common form of CAH is called 21-hydroxylase deficiency and results from the gene labelled CYP21 being absent or changed. This means that aldosterone and cortisol are not produced but production of androgen is unaffected. The gene affected results in different types of CAH, affecting people to varying degrees from mild to severe.

• #6 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Gene mutations that cause defects in steroidogenesis are classified as CAH and involve the following enzymes and proteins: 21-Hydroxylase (21-OH), 11-Hydroxylase (11-OH), 3-Hydroxysteroid dehydrogenase type-2 (3-HSD-2), 17-Hydroxylase/17,20-lyase (17-OH), P450 oxidoreductase (POR), Steroidogenic acute regulatory protein (StAR), Cholesterol side-chain cleavage enzyme (SCC). […] Defects in the CYP21A2 gene, which causes 21-OH deficiency, account for approximately 95% of cases. […] Loss or impairment of CYP21A2 results in reduced production of the enzyme cytochrome P450c21 (21-OH). As a result, 21-OH deficiency hinders the conversion of 17-OHP to 11-deoxycortisol, the penultimate step in cortisol synthesis, and the conversion of progesterone to deoxycorticosterone (DOC) in aldosterone synthesis.

• #7 Congenital Adrenal Hyperplasia: Practice Essentials, Background, Pathophysiology

  https://emedicine.medscape.com/article/919218-overview

  The term congenital adrenal hyperplasia (CAH) encompasses a group of autosomal recessive disorders, each of which involves a deficiency of an enzyme involved in the synthesis of cortisol, aldosterone, or both. Deficiency of 21-hydroxylase, resulting from mutations or deletions of CYP21A, is the most common form of congenital adrenal hyperplasia, accounting for more than 90% of cases. The diagnosis of congenital adrenal hyperplasia depends on the demonstration of inadequate production of cortisol and/or aldosterone in the presence of accumulation of excess concentrations of precursor hormones. […] A mutation or deletion of any of the genes that code for enzymes involved in cortisol or aldosterone synthesis results in congenital adrenal hyperplasia. The particular phenotype that results depends on the sex of the individual, the location of the block in synthesis, and the severity of the genetic deletion or mutation.

• #8 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Congenital adrenal hyperplasia (CAH) encompasses a group of autosomal recessive conditions caused by genetic mutations that disrupt enzymes responsible for producing glucocorticoids, mineralocorticoids, and sex steroids from cholesterol in the adrenal glands. […] These mutations impair cortisol synthesis, prompting a compensatory increase in adrenocorticotropic hormone, which leads to adrenal cortex hyperplasia hence the name „CAH.” The condition manifests in infants, children, or adults with symptoms of corticosteroid deficiency, often accompanied by either a deficiency or an excess of mineralocorticoids and sex steroids, depending on the specific enzyme defect. […] The most common form, 21-hydroxylase (21-OH) deficiency, accounts for approximately 95% of cases. Clinical presentations vary by enzyme defect, ranging from ambiguous genitalia in genotypic females to potentially life-threatening adrenal insufficiency in males.

• #9 Genetics and Pathophysiology of Congenital Adrenal Hyperplasia | Oncohema Key

  https://oncohemakey.com/genetics-and-pathophysiology-of-congenital-adrenal-hyperplasia-2/

  Defective 21-OHD also promotes accumulation of other steroid hormone intermediates such as 21-deoxycortisol, 16-hydroxyprogesterone, 11-ketoandrostenedione, and 11-ketotestosterone. […] The enzyme 17-hydroxysteroid dehydrogenase type 5 also known as aldo-keto reductase 1C3 (AKR1C3) can convert DHEA and androstenedione to androstanediol and testosterone, respectively. […] The specific molecular mechanisms responsible for the altered hypothalamic-pituitary-adrenal (HPO) axis function accompanied by apparent ovarian androgen excess are unclear. Increased circulating concentrations of adrenal androgens and progestins likely influence HPO axis function. […] The CYP21A2 gene is located in a complex genetic region at chromosome 6p21.3 where it lies in close proximity to a highly homologous pseudogene, CYP21A1P.

• #10 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Congenital adrenal hyperplasia (CAH) refers to a group of autosomal recessive conditions caused by mutations in genes encoding enzymes involved in the production of glucocorticoids, and sometimes mineralocorticoids and sex steroids, from cholesterol in the adrenal glands. […] These mutations impair cortisol synthesis, triggering a compensatory increase in adrenocorticotropic hormone (ACTH), which leads to adrenal cortex hyperplasia hence the term „CAH.” […] Gene mutations that cause defects in steroidogenesis are classified as CAH and involve the following enzymes and proteins: 21-Hydroxylase (21-OH), 11-Hydroxylase (11-OH), 3-Hydroxysteroid dehydrogenase type-2 (3-HSD-2), 17-Hydroxylase/17,20-lyase (17-OH), P450 oxidoreductase (POR), Steroidogenic acute regulatory protein (StAR), Cholesterol side-chain cleavage enzyme (SCC).

• #11 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Congenital adrenal hyperplasia (CAH) refers to a group of autosomal recessive conditions caused by mutations in genes encoding enzymes involved in the production of glucocorticoids, and sometimes mineralocorticoids and sex steroids, from cholesterol in the adrenal glands. […] These mutations impair cortisol synthesis, triggering a compensatory increase in adrenocorticotropic hormone (ACTH), which leads to adrenal cortex hyperplasia hence the term „CAH.” […] Gene mutations that cause defects in steroidogenesis are classified as CAH and involve the following enzymes and proteins: 21-Hydroxylase (21-OH), 11-Hydroxylase (11-OH), 3-Hydroxysteroid dehydrogenase type-2 (3-HSD-2), 17-Hydroxylase/17,20-lyase (17-OH), P450 oxidoreductase (POR), Steroidogenic acute regulatory protein (StAR), Cholesterol side-chain cleavage enzyme (SCC).

• #12 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Defects in the CYP21A2 gene, which causes 21-OH deficiency, account for approximately 95% of cases. […] Loss or impairment of CYP21A2 results in reduced production of the enzyme cytochrome P450c21 (21-OH). […] As a result, 21-OH deficiency hinders the conversion of 17-OHP to 11-deoxycortisol, the penultimate step in cortisol synthesis, and the conversion of progesterone to deoxycorticosterone (DOC) in aldosterone synthesis. […] Disease severity and phenotypic presentation vary depending on the location and extent of gene mutations or deletions, resulting in complex allelic variations. […] Cortisol deficiency triggers feedback stimulation of ACTH release from the pituitary gland. […] Elevated ACTH levels can cause adrenal cortex hyperplasia, accumulation of cortisol precursors, and diversion of steroidogenic pathways, resulting in excess production of androgens or mineralocorticoid precursors.

• #13 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Defects in the CYP21A2 gene, which causes 21-OH deficiency, account for approximately 95% of cases. […] Loss or impairment of CYP21A2 results in reduced production of the enzyme cytochrome P450c21 (21-OH). […] As a result, 21-OH deficiency hinders the conversion of 17-OHP to 11-deoxycortisol, the penultimate step in cortisol synthesis, and the conversion of progesterone to deoxycorticosterone (DOC) in aldosterone synthesis. […] Disease severity and phenotypic presentation vary depending on the location and extent of gene mutations or deletions, resulting in complex allelic variations. […] Cortisol deficiency triggers feedback stimulation of ACTH release from the pituitary gland. […] Elevated ACTH levels can cause adrenal cortex hyperplasia, accumulation of cortisol precursors, and diversion of steroidogenic pathways, resulting in excess production of androgens or mineralocorticoid precursors.

• #14 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Defects in the CYP21A2 gene, which causes 21-OH deficiency, account for approximately 95% of cases. […] Loss or impairment of CYP21A2 results in reduced production of the enzyme cytochrome P450c21 (21-OH). […] As a result, 21-OH deficiency hinders the conversion of 17-OHP to 11-deoxycortisol, the penultimate step in cortisol synthesis, and the conversion of progesterone to deoxycorticosterone (DOC) in aldosterone synthesis. […] Disease severity and phenotypic presentation vary depending on the location and extent of gene mutations or deletions, resulting in complex allelic variations. […] Cortisol deficiency triggers feedback stimulation of ACTH release from the pituitary gland. […] Elevated ACTH levels can cause adrenal cortex hyperplasia, accumulation of cortisol precursors, and diversion of steroidogenic pathways, resulting in excess production of androgens or mineralocorticoid precursors.

• #15 Congenital Adrenal Hyperplasia – Endotext – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK278953/

  Congenital Adrenal Hyperplasia (CAH) is a term used to describe a group of genetically determined disorders of defective steroidogenesis that result in variable deficiency of the end products cortisol and/or aldosterone and their deleterious, including life-threatening, effects on metabolism and electrolytes with simultaneous diversion to the accumulation of androgens and their virilizing effects. […] The production of cortisol in the zona fasciculata of the adrenal cortex occurs in five major enzyme-mediated steps. CAH results from deficiency in any one of these enzymes. Impaired cortisol synthesis leads to chronic elevations of ACTH via the negative feedback system, causing overstimulation of the adrenal cortex and resulting in hyperplasia and over-secretion of the precursors to the enzymatic defect.

• #16 Congenital Adrenal Hyperplasia – Endotext – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK278953/

  Adrenal steroidogenesis occurs in three major pathways: glucocorticoids, mineralocorticoids, and sex steroids as shown in Figure 1. The adrenal gland architecture suggests that the adrenal acts as three separate glands: zona glomerulosa, zona fasciculate, zona reticularis. The hypothalamic-pituitary-adrenal feedback system is mediated through the circulating level of plasma cortisol by negative feedback of cortisol on CRF and ACTH secretion. Therefore, any CAH condition that results in a decrease in cortisol secretion leads to increased ACTH production, which in turn stimulates (1) excessive synthesis of adrenal products in those pathways unimpaired by the enzyme deficiency and (2) a build-up of precursor molecules in pathways blocked by the enzyme deficiency. […] In the most common form 21OHD-CAH, the function of 21-hydroxylating cytochrome P450 is deficient, creating a block in the P450 cortisol production pathway. This leads to an accumulation of 17-hydroxyprogesterone (17-OHP), a precursor of the 21-hydroxylation step. Excess 17-OHP is then shunted into the intact androgen pathway, where the 17,20-lyase enzyme converts 17-OHP to 4-androstenedione, the major adrenal androgen. […] The clinical symptoms of the five different forms of CAH result from the specific hormones that are deficient and those that are produced in excess.

• #17 Congenital Adrenal Hyperplasia – Endotext – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK278953/

  Adrenal steroidogenesis occurs in three major pathways: glucocorticoids, mineralocorticoids, and sex steroids as shown in Figure 1. The adrenal gland architecture suggests that the adrenal acts as three separate glands: zona glomerulosa, zona fasciculate, zona reticularis. The hypothalamic-pituitary-adrenal feedback system is mediated through the circulating level of plasma cortisol by negative feedback of cortisol on CRF and ACTH secretion. Therefore, any CAH condition that results in a decrease in cortisol secretion leads to increased ACTH production, which in turn stimulates (1) excessive synthesis of adrenal products in those pathways unimpaired by the enzyme deficiency and (2) a build-up of precursor molecules in pathways blocked by the enzyme deficiency. […] In the most common form 21OHD-CAH, the function of 21-hydroxylating cytochrome P450 is deficient, creating a block in the P450 cortisol production pathway. This leads to an accumulation of 17-hydroxyprogesterone (17-OHP), a precursor of the 21-hydroxylation step. Excess 17-OHP is then shunted into the intact androgen pathway, where the 17,20-lyase enzyme converts 17-OHP to 4-androstenedione, the major adrenal androgen. […] The clinical symptoms of the five different forms of CAH result from the specific hormones that are deficient and those that are produced in excess.

• #18 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Cortisol deficiency triggers feedback stimulation of ACTH release from the pituitary gland. Elevated ACTH levels can cause adrenal cortex hyperplasia, accumulation of cortisol precursors, and diversion of steroidogenic pathways, resulting in excess production of androgens or mineralocorticoid precursors. […] Inadequate sex steroid production may occur depending on the extent of the enzyme defect in steroidogenesis (eg, 17-OH, POR, StAR, and SCC). This disruption can lead to clinical manifestations in infants, including hyperpigmentation from elevated ACTH, failure to thrive, electrolyte imbalances, acidosis or alkalosis, and shock due to adrenal insufficiency. […] A defect in StAR (STARD1) impairs the transport of cholesterol to the inner mitochondrial membrane, disrupting the production of all steroid hormones.

• #19 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Defects in the CYP21A2 gene, which causes 21-OH deficiency, account for approximately 95% of cases. […] Loss or impairment of CYP21A2 results in reduced production of the enzyme cytochrome P450c21 (21-OH). […] As a result, 21-OH deficiency hinders the conversion of 17-OHP to 11-deoxycortisol, the penultimate step in cortisol synthesis, and the conversion of progesterone to deoxycorticosterone (DOC) in aldosterone synthesis. […] Disease severity and phenotypic presentation vary depending on the location and extent of gene mutations or deletions, resulting in complex allelic variations. […] Cortisol deficiency triggers feedback stimulation of ACTH release from the pituitary gland. […] Elevated ACTH levels can cause adrenal cortex hyperplasia, accumulation of cortisol precursors, and diversion of steroidogenic pathways, resulting in excess production of androgens or mineralocorticoid precursors.

• #20 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #21 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #22 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #23 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #24 Congenital adrenal hyperplasia due to 21-hydroxylase deficiency – Wikipedia

  https://en.wikipedia.org/wiki/Congenital_adrenal_hyperplasia_due_to_21-hydroxylase_deficiency

  In the insufficiency of 21-hydroxylase to participate in the biosynthesis of cortisol, the 21-hydroxylation in the zona fasciculata of the adrenal cortex is impaired, so 17OHP and progesterone will not be properly converted into 11-deoxycortisol and 11-deoxycorticosterone, respectively the precursors for cortisol and aldosterone. […] As the plasma concentration of cortisol and aldosterone decreases, ACTH levels increase, leading to excessive production and accumulation of cortisol precursors (especially 17OHP), which are eventually transferred to androsterone that is a feedstock for other androgens. […] Other androgens may be additionally produced from 17OHP, due to its elevated levels, that leads, inter alia, to its 5-reduction. […] Some of the androgens produced by the backdoor pathway are those that cannot be converted to estrogens by aromatase, causing prenatal virilization, and making them the dominant androgens in classic 21-hydroxylase deficiency.

• #25 Genetics and Pathophysiology of Congenital Adrenal Hyperplasia | Oncohema Key

  https://oncohemakey.com/genetics-and-pathophysiology-of-congenital-adrenal-hyperplasia-2/

  Defective 21-OHD also promotes accumulation of other steroid hormone intermediates such as 21-deoxycortisol, 16-hydroxyprogesterone, 11-ketoandrostenedione, and 11-ketotestosterone. […] The enzyme 17-hydroxysteroid dehydrogenase type 5 also known as aldo-keto reductase 1C3 (AKR1C3) can convert DHEA and androstenedione to androstanediol and testosterone, respectively. […] The specific molecular mechanisms responsible for the altered hypothalamic-pituitary-adrenal (HPO) axis function accompanied by apparent ovarian androgen excess are unclear. Increased circulating concentrations of adrenal androgens and progestins likely influence HPO axis function. […] The CYP21A2 gene is located in a complex genetic region at chromosome 6p21.3 where it lies in close proximity to a highly homologous pseudogene, CYP21A1P.

• #26 Genetics and Pathophysiology of Congenital Adrenal Hyperplasia | Oncohema Key

  https://oncohemakey.com/genetics-and-pathophysiology-of-congenital-adrenal-hyperplasia-2/

  Most mutations result from gene conversion events in which the functional gene acquires deleterious CYP21A1P sequences or from misalignment during meiosis that can give rise to duplication or deletions of the RCCX unit. […] Despite these potential obstacles, genetic analysis can be a useful adjunct to newborn screening.

• #27 Nationwide carrier screening for congenital adrenal hyperplasia: integrated approach of CYP21A2 pathogenic variant genotyping and comprehensive large gene deletion analysis | BMC Medical Genomics | Full Text

  https://bmcmedgenomics.biomedcentral.com/articles/10.1186/s12920-025-02089-5

  The CYP21A2 gene encodes the 21-Hydroxylase enzyme, which is located on the short arm of chromosome 6 (6p21.3) within the human leukocyte antigen complex. Coexisting with neighboring genes for tenascin XB (TNXB), complement C4B (C4B), and serine/threonine kinase 19 pseudogene (RP2), the CYP21A2 gene forms a genetic module known as RCCX (RP2-C4B-CYP21A2-TNXB). Typically, RCCX haplotypes comprise two modular units: the previously described unit and another containing RP1, CYP21A1P, TNXA, and C4A. CYP21A1P, a non-functional pseudogene, shares 98% coding sequence homology with CYP21A2. Copy Number Variation (CNV) of RCCX module haplotypes has also been reported. Approximately 95% of defective CYP21A2 mutations in CAH fall into three categories: 1) 70% involve deleterious variants originating from the pseudogene CYP21A1P due to small gene conversions; 2) 25-30% encompass large gene rearrangements resulting from unequal meiotic crossing over, leading to a 30-kb deletion and a large gene conversion spanning the 3 end of CYP21A1P and the 5 end of CYP21A2, known as the chimeric CYP21A1P/CYP21A2 gene; and 3) approximately 5% involve other spontaneous missense or small indel variants.

• #28 Nationwide carrier screening for congenital adrenal hyperplasia: integrated approach of CYP21A2 pathogenic variant genotyping and comprehensive large gene deletion analysis | BMC Medical Genomics | Full Text

  https://bmcmedgenomics.biomedcentral.com/articles/10.1186/s12920-025-02089-5

  The CYP21A2 gene encodes the 21-Hydroxylase enzyme, which is located on the short arm of chromosome 6 (6p21.3) within the human leukocyte antigen complex. Coexisting with neighboring genes for tenascin XB (TNXB), complement C4B (C4B), and serine/threonine kinase 19 pseudogene (RP2), the CYP21A2 gene forms a genetic module known as RCCX (RP2-C4B-CYP21A2-TNXB). Typically, RCCX haplotypes comprise two modular units: the previously described unit and another containing RP1, CYP21A1P, TNXA, and C4A. CYP21A1P, a non-functional pseudogene, shares 98% coding sequence homology with CYP21A2. Copy Number Variation (CNV) of RCCX module haplotypes has also been reported. Approximately 95% of defective CYP21A2 mutations in CAH fall into three categories: 1) 70% involve deleterious variants originating from the pseudogene CYP21A1P due to small gene conversions; 2) 25-30% encompass large gene rearrangements resulting from unequal meiotic crossing over, leading to a 30-kb deletion and a large gene conversion spanning the 3 end of CYP21A1P and the 5 end of CYP21A2, known as the chimeric CYP21A1P/CYP21A2 gene; and 3) approximately 5% involve other spontaneous missense or small indel variants.

• #29 Nationwide carrier screening for congenital adrenal hyperplasia: integrated approach of CYP21A2 pathogenic variant genotyping and comprehensive large gene deletion analysis | BMC Medical Genomics | Full Text

  https://bmcmedgenomics.biomedcentral.com/articles/10.1186/s12920-025-02089-5

  The CYP21A2 gene encodes the 21-Hydroxylase enzyme, which is located on the short arm of chromosome 6 (6p21.3) within the human leukocyte antigen complex. Coexisting with neighboring genes for tenascin XB (TNXB), complement C4B (C4B), and serine/threonine kinase 19 pseudogene (RP2), the CYP21A2 gene forms a genetic module known as RCCX (RP2-C4B-CYP21A2-TNXB). Typically, RCCX haplotypes comprise two modular units: the previously described unit and another containing RP1, CYP21A1P, TNXA, and C4A. CYP21A1P, a non-functional pseudogene, shares 98% coding sequence homology with CYP21A2. Copy Number Variation (CNV) of RCCX module haplotypes has also been reported. Approximately 95% of defective CYP21A2 mutations in CAH fall into three categories: 1) 70% involve deleterious variants originating from the pseudogene CYP21A1P due to small gene conversions; 2) 25-30% encompass large gene rearrangements resulting from unequal meiotic crossing over, leading to a 30-kb deletion and a large gene conversion spanning the 3 end of CYP21A1P and the 5 end of CYP21A2, known as the chimeric CYP21A1P/CYP21A2 gene; and 3) approximately 5% involve other spontaneous missense or small indel variants.

• #30 Congenital adrenal hyperplasia due to 21-hydroxylase deficiency – Wikipedia

  https://en.wikipedia.org/wiki/Congenital_adrenal_hyperplasia_due_to_21-hydroxylase_deficiency

  In the insufficiency of 21-hydroxylase to participate in the biosynthesis of cortisol, the 21-hydroxylation in the zona fasciculata of the adrenal cortex is impaired, so 17OHP and progesterone will not be properly converted into 11-deoxycortisol and 11-deoxycorticosterone, respectively the precursors for cortisol and aldosterone. […] As the plasma concentration of cortisol and aldosterone decreases, ACTH levels increase, leading to excessive production and accumulation of cortisol precursors (especially 17OHP), which are eventually transferred to androsterone that is a feedstock for other androgens. […] Other androgens may be additionally produced from 17OHP, due to its elevated levels, that leads, inter alia, to its 5-reduction. […] Some of the androgens produced by the backdoor pathway are those that cannot be converted to estrogens by aromatase, causing prenatal virilization, and making them the dominant androgens in classic 21-hydroxylase deficiency.

• #31 Congenital adrenal hyperplasia due to 21-hydroxylase deficiency – Wikipedia

  https://en.wikipedia.org/wiki/Congenital_adrenal_hyperplasia_due_to_21-hydroxylase_deficiency

  In the insufficiency of 21-hydroxylase to participate in the biosynthesis of cortisol, the 21-hydroxylation in the zona fasciculata of the adrenal cortex is impaired, so 17OHP and progesterone will not be properly converted into 11-deoxycortisol and 11-deoxycorticosterone, respectively the precursors for cortisol and aldosterone. […] As the plasma concentration of cortisol and aldosterone decreases, ACTH levels increase, leading to excessive production and accumulation of cortisol precursors (especially 17OHP), which are eventually transferred to androsterone that is a feedstock for other androgens. […] Other androgens may be additionally produced from 17OHP, due to its elevated levels, that leads, inter alia, to its 5-reduction. […] Some of the androgens produced by the backdoor pathway are those that cannot be converted to estrogens by aromatase, causing prenatal virilization, and making them the dominant androgens in classic 21-hydroxylase deficiency.

• #32 Recommendations for the diagnosis and treatment of classic forms of 21-hydroxylase-deficient congenital adrenal hyperplasia | Anales de Pediatría

  https://www.analesdepediatria.org/en-recommendations-for-diagnosis-treatment-classic-articulo-S2341287917301023

  The gene at the root of 21OHD is known as CYP21A2. It is located in the in the HLA class III region of the short arm of chromosome 6p21.3, adjacent to the CYP21A1P pseudogene. […] Ninety-five percent of the pathological variants of the CYP21A2 gene are recurrent and result from two different mechanisms involving the CYP21A2 and the CYP21A1 pseudogene, namely unequal crossing over and gene conversion. Five percent of the total alleles correspond to mutations of the CYP21A2 gene that cannot be attributed to gene microconversion. […] There is a strong genotype/phenotype correlation, as the severity of symptoms depends on the extent of enzymatic disruption, which in turn is determined by the genetic changes found in each allele.

• #33 Clinical Update on Congenital Adrenal Hyperplasia: Recommendations from a Multidisciplinary Adrenal Program

  https://www.mdpi.com/2077-0383/12/9/3128

  As explained, alterations in the CYP21A2 gene translate into enzymatic deficiency of the 21-hydroxylase (21OHD) activity causing a decrease in cortisol biosynthesis. However, in certain severe cases, CYP21A2 mutations can also affect aldosterone production. Then, the lack of cortisol feedback results in increased ACTH, which promotes the accumulation of 17-hydroxyprogesterone (17OHP) and other steroids that serve as substrates for androgen excess. […] Clinical CAH phenotypes are characterized by decreased cortisol synthesis and increased androgen secretion and depend on both the age at presentation and the severity of the CYP21A2 mutation. Hence, CAH cases can be divided into three categories: (a) Salt-wasting (SW) represents 65–75% of the classic CAH cases. These are manifested in infancy and are characterized by a marked cortisol and aldosterone deficiency along with hyperandrogenism. Residual enzymatic activity of SW-CAH is typically < 1%. (b) Simple virilizing CAH comprises 25–35% of the classic CAH. Unlike SW-CAH, this form manifests later in life and is characterized by a severe cortisol deficit, but unaltered aldosterone. Residual enzymatic activity of this form of CAH is 1–2%; and (c) Non-classic CAH is the most frequently seen in the clinic. It usually manifests during puberty and is characterized by hyperandrogenism. Residual enzymatic activity ranges from 20% to 50% and normal cortisol production is maintained by excess ACTH.

• #34 Clinical Update on Congenital Adrenal Hyperplasia: Recommendations from a Multidisciplinary Adrenal Program

  https://www.mdpi.com/2077-0383/12/9/3128

  As explained, alterations in the CYP21A2 gene translate into enzymatic deficiency of the 21-hydroxylase (21OHD) activity causing a decrease in cortisol biosynthesis. However, in certain severe cases, CYP21A2 mutations can also affect aldosterone production. Then, the lack of cortisol feedback results in increased ACTH, which promotes the accumulation of 17-hydroxyprogesterone (17OHP) and other steroids that serve as substrates for androgen excess. […] Clinical CAH phenotypes are characterized by decreased cortisol synthesis and increased androgen secretion and depend on both the age at presentation and the severity of the CYP21A2 mutation. Hence, CAH cases can be divided into three categories: (a) Salt-wasting (SW) represents 65–75% of the classic CAH cases. These are manifested in infancy and are characterized by a marked cortisol and aldosterone deficiency along with hyperandrogenism. Residual enzymatic activity of SW-CAH is typically < 1%. (b) Simple virilizing CAH comprises 25–35% of the classic CAH. Unlike SW-CAH, this form manifests later in life and is characterized by a severe cortisol deficit, but unaltered aldosterone. Residual enzymatic activity of this form of CAH is 1–2%; and (c) Non-classic CAH is the most frequently seen in the clinic. It usually manifests during puberty and is characterized by hyperandrogenism. Residual enzymatic activity ranges from 20% to 50% and normal cortisol production is maintained by excess ACTH.

• #35 21-hydroxylase deficiency: MedlinePlus GeneticsLock

  https://medlineplus.gov/genetics/condition/21-hydroxylase-deficiency/

  The amount of functional 21-hydroxylase enzyme determines the severity of the disorder. Individuals with the salt-wasting type have CYP21A2 mutations that result in a completely nonfunctional enzyme. People with the simple virilizing type of this condition have CYP21A2 gene mutations that allow the production of low levels of functional enzyme. Individuals with the non-classic type of this disorder have CYP21A2 mutations that result in the production of reduced amounts of the enzyme, but more enzyme than either of the other types.

• #36 Clinical manifestations and diagnosis of classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency in infants and children – UpToDate

  https://www.uptodate.com/contents/clinical-manifestations-and-diagnosis-of-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-in-infants-and-children

  Defective conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol due to 21-hydroxylase deficiency (21OHD) accounts for more than 95 percent of cases of congenital adrenal hyperplasia (CAH). […] The defective conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol in patients with 21-hydroxylase deficiency (21OHD) results in decreased cortisol synthesis, loss of negative feedback, and therefore increased corticotropin (ACTH) secretion. The resulting adrenal stimulation leads to increased production of adrenal-derived androgens and a variable degree of aldosterone deficiency. The severity of disease relates to the degree to which the mutations compromise enzyme activity.

• #37 21-hydroxylase deficiency: MedlinePlus GeneticsLock

  https://medlineplus.gov/genetics/condition/21-hydroxylase-deficiency/

  21-hydroxylase deficiency is one of a group of disorders known as congenital adrenal hyperplasias that impair hormone production and disrupt sexual development. 21-hydroxylase deficiency is responsible for about 95 percent of all cases of congenital adrenal hyperplasia. […] Mutations in the CYP21A2 gene cause 21-hydroxylase deficiency. The CYP21A2 gene provides instructions for making an enzyme called 21-hydroxylase. This enzyme is found in the adrenal glands, where it plays a role in producing hormones called cortisol and aldosterone. […] 21-hydroxylase deficiency is caused by a shortage (deficiency) of the 21-hydroxylase enzyme. When 21-hydroxylase is lacking, substances that are usually used to form cortisol and aldosterone instead build up in the adrenal glands and are converted to androgens. The excess production of androgens leads to abnormalities of sexual development in people with 21-hydroxylase deficiency. A lack of aldosterone production contributes to the salt loss in people with the salt-wasting form of this condition.

• #38 Congenital Adrenal Hyperplasia | Texas DSHS

  https://www.dshs.texas.gov/newborn-screening-program/newborn-screening-parent-resources/congenital-adrenal-hyperplasia-cah/congenital-adrenal-hyperplasia-a

  Unfortunately, ACTH signals the entire adrenal gland to work harder, not just the part that makes cortisol. The faulty enzyme causes a bottleneck in the chemical assembly line that makes cortisol, resulting in a piling up of „raw products” for making the final adrenal hormones. These substances spill over into other production lines which do not use the faulty enzyme: the production lines for androgens (male hormones). […] High levels of male-like hormones continue to affect the adolescent. The girl with untreated CAH will not have normal menstrual periods because the male-like hormones interfere with the work of the ovary. The testicles of the boy cannot function well and will not make sperm normally. […] There are two forms of CAH. The classical and non-classical forms. Non-classical CAH refers to a condition in which the enzyme defect is less severe. Individuals with this form of CAH are typically normal at birth but begin to suffer mild effects of too much male hormone in childhood or adolescence. This is also called „late onset” CAH.

• #39 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #40 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #41 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #42 Neurocrine Biosciences Announces Classic Congenital Adrenal Hyperplasia Supplement Published Today in The Journal of Clinical Endocrinology & Metabolism | Neurocrine Biosciences

  https://neurocrine.gcs-web.com/news-releases/news-release-details/neurocrine-biosciences-announces-classic-congenital-adrenal

  „Approximately 95% of CAH cases are caused by variants of the CYP21A2 gene that leads to deficiency of the enzyme 21-hydroxylase.” […] „Severe deficiency of this enzyme leads to an inability of the adrenal glands to produce enough cortisol and, in approximately 75% of cases, aldosterone.” […] „Because individuals with CAH are still able to produce androgens, the unused precursors that would normally be used to make cortisol instead result in the production of excess amounts of androgens.” […] „If left untreated, CAH can result in salt wasting, dehydration and even death.” […] „Antagonism of CRF1 receptors in the pituitary has been shown to decrease ACTH levels, which in turn decreases the production of adrenal androgens and potentially the symptoms associated with CAH.” […] „The robust clinical study data demonstrate that lowering adrenal androgen levels with CRENESSITY enables lower, more physiologic dosing of GCs to replace missing cortisol.”

• #43 Neurocrine Biosciences Announces Classic Congenital Adrenal Hyperplasia Supplement Published Today in The Journal of Clinical Endocrinology & Metabolism | Neurocrine Biosciences

  https://neurocrine.gcs-web.com/news-releases/news-release-details/neurocrine-biosciences-announces-classic-congenital-adrenal

  „Approximately 95% of CAH cases are caused by variants of the CYP21A2 gene that leads to deficiency of the enzyme 21-hydroxylase.” […] „Severe deficiency of this enzyme leads to an inability of the adrenal glands to produce enough cortisol and, in approximately 75% of cases, aldosterone.” […] „Because individuals with CAH are still able to produce androgens, the unused precursors that would normally be used to make cortisol instead result in the production of excess amounts of androgens.” […] „If left untreated, CAH can result in salt wasting, dehydration and even death.” […] „Antagonism of CRF1 receptors in the pituitary has been shown to decrease ACTH levels, which in turn decreases the production of adrenal androgens and potentially the symptoms associated with CAH.” […] „The robust clinical study data demonstrate that lowering adrenal androgen levels with CRENESSITY enables lower, more physiologic dosing of GCs to replace missing cortisol.”

• #44 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #45 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #46 Congenital adrenal hyperplasia – Symptoms and causes – Mayo Clinic

  https://www.mayoclinic.org/diseases-conditions/congenital-adrenal-hyperplasia/symptoms-causes/syc-20355205

  Congenital adrenal hyperplasia (CAH) is the medical name for a group of genetic conditions that affect the adrenal glands. […] In people with CAH, a gene change results in a lack of one of the enzyme proteins needed to make these hormones. […] The most common cause of CAH is the lack of the enzyme protein known as 21-hydroxylase. Sometimes, CAH is called 21-hydroxylase deficiency. The body needs this enzyme to make proper amounts of hormones. […] CAH is a genetic condition. That means it’s passed from parents to children. It’s present at birth. Children with the condition have two parents who both carry the genetic change that causes CAH. Or they have two parents who have CAH themselves. This is known as the autosomal recessive inheritance pattern.

• #47 Congenital Adrenal Hyperplasia: Practice Essentials, Background, Pathophysiology

  https://emedicine.medscape.com/article/919218-overview

  The clinical manifestations of each form of congenital adrenal hyperplasia are related to the degree of cortisol deficiency and/or the degree of aldosterone deficiency. In some cases, these manifestations reflect the accumulation of precursor adrenocortical hormones. When present in supraphysiologic concentrations, these precursors lead to excess androgen production with resultant virilization, or because of mineralocorticoid properties, cause sodium retention and hypertension. […] The phenotype depends on the degree or type of gene deletion or mutation and the resultant deficiency of the steroidogenic enzyme. The most common form of adrenal hyperplasia (due to a deficiency of 21-hydroxylase activity) is clinically divided into 3 phenotypes: salt wasting, simple virilizing, and nonclassic.

• #48 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #49 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #50 Congenital Adrenal Hyperplasia – National Adrenal Diseases Foundation

  https://www.nadf.us/congenital-adrenal-hyperplasia-cah.html

  Nonclassic CAH is mild and not life threatening. Signs and symptoms might not appear until childhood or adulthood. […] Some patients have no symptoms and require no treatment. Others need low-dose glucocorticoids but might not need life-long treatment. Nonclassic CAH is not associated with genital abnormalities at birth and is not detected in most newborn screening programs. […] With proper care, people with either type of CAH can live long and healthy lives.

• #51 Congenital Adrenal Hyperplasia – National Adrenal Diseases Foundation

  https://www.nadf.us/congenital-adrenal-hyperplasia-cah.html

  Nonclassic CAH is mild and not life threatening. Signs and symptoms might not appear until childhood or adulthood. […] Some patients have no symptoms and require no treatment. Others need low-dose glucocorticoids but might not need life-long treatment. Nonclassic CAH is not associated with genital abnormalities at birth and is not detected in most newborn screening programs. […] With proper care, people with either type of CAH can live long and healthy lives.

• #52 Congenital Adrenal Hyperplasia (CAH) Panel Test – PreventionGenetics

  https://www.preventiongenetics.com/testInfo?val=Congenital-Adrenal-Hyperplasia-%28CAH%29-Panel

  Other less common forms of CAH include 3-hydroxysteroid dehydrogenase type 2 (HSD3B2) deficiency, 17-hydroxylase (CYP17A1) deficiency, congenital lipoid adrenal hyperplasia (STAR), side-chain cleavage enzyme (CYP11A1) deficiency, and cytochrome P450 oxidoreductase (POR) deficiency. […] Founder pathogenic variants in different geographic regions worldwide are evident in all of these genes. […] Since our test utilizes an integrative strategy via Sanger sequencing to perform a comprehensive evaluation for CYP21A2, the detection rate of pathogenic CYP21A2 variants is expected to be approximately 98%. […] Other genes in this panel have no paralogous sequences in the genome. Therefore, this panel is able to detect over 98% of pathogenic variants for congenital adrenal hyperplasia patients overall.

• #53 Orphanet: Congenital adrenal hyperplasia

  https://www.orpha.net/en/disease/detail/418

  A group of rare inherited endocrine disorders caused by a steroidogenic enzyme deficiency and characterized by adrenal insufficiency and variable degrees of hyper- or hypoandrogenism manifestations, depending on disease type and severity. […] In 90-95% of cases, CAH is caused by a mutation in the CYP21A2 gene located on chromosome 6p21.3 which encodes for an enzyme that controls cortisol and aldosterone production. Other genes are less frequently involved and result in the following variants of CAH: CAH due to 17-alpha-hydroxylase deficiency, 3-beta-hydroxysteroid dehydrogenase deficiency, 11-beta-hydroxylase deficiency, cytochrome P450 oxidoreductase deficiency and congenital lipoid adrenal hyperplasia.

• #54 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Cortisol deficiency triggers feedback stimulation of ACTH release from the pituitary gland. Elevated ACTH levels can cause adrenal cortex hyperplasia, accumulation of cortisol precursors, and diversion of steroidogenic pathways, resulting in excess production of androgens or mineralocorticoid precursors. […] Inadequate sex steroid production may occur depending on the extent of the enzyme defect in steroidogenesis (eg, 17-OH, POR, StAR, and SCC). This disruption can lead to clinical manifestations in infants, including hyperpigmentation from elevated ACTH, failure to thrive, electrolyte imbalances, acidosis or alkalosis, and shock due to adrenal insufficiency. […] A defect in StAR (STARD1) impairs the transport of cholesterol to the inner mitochondrial membrane, disrupting the production of all steroid hormones.

• #55 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Severe StAR defects result in classic lipoid congenital adrenal hyperplasia (LCAH), which is characterized by lipid accumulation in steroidogenic cells of the adrenal glands and gonads. […] This condition presents with adrenal insufficiency during the neonatal period or early infancy and female or near-female genitalia, regardless of chromosomal sex.

• #56 Congenital Adrenal Hyperplasia – StatPearls – NCBI Bookshelf

  https://www.ncbi.nlm.nih.gov/books/NBK448098/

  Severe StAR defects result in classic lipoid congenital adrenal hyperplasia (LCAH), which is characterized by lipid accumulation in steroidogenic cells of the adrenal glands and gonads. […] This condition presents with adrenal insufficiency during the neonatal period or early infancy and female or near-female genitalia, regardless of chromosomal sex.

• #57 Congenital Adrenal Hyperplasia | Treatment & Management | Point of Care

  https://www.statpearls.com/point-of-care/17229

  Inadequate sex steroid production may occur depending on the extent of the enzyme defect in steroidogenesis (eg, 17-OH, POR, StAR, and SCC). […] A defect in StAR (STARD1) impairs the transport of cholesterol to the inner mitochondrial membrane, disrupting the production of all steroid hormones. […] Severe StAR defects result in classic lipoid congenital adrenal hyperplasia (LCAH), which is characterized by lipid accumulation in steroidogenic cells of the adrenal glands and gonads. […] This condition presents with adrenal insufficiency during the neonatal period or early infancy and female or near-female genitalia, regardless of chromosomal sex. […] A milder form, known as nonclassic LCAH, occurs when approximately 20% to 30% of enzyme activity is preserved. […] Defects in 3-HSD-2 (HSD3B2) impair the conversion of pregnenolone, 17-hydroxypregnenolone, and DHEA to progesterone, 17-OHP, and androstenedione, respectively.

• #58 Nationwide carrier screening for congenital adrenal hyperplasia: integrated approach of CYP21A2 pathogenic variant genotyping and comprehensive large gene deletion analysis | BMC Medical Genomics | Full Text

  https://bmcmedgenomics.biomedcentral.com/articles/10.1186/s12920-025-02089-5

  Technically, genomic approaches for deciphering the gene responsible for 21-OHD CAH are intricate due to the multifaceted etiology. The presence of more than one variant in cis complicates the molecular diagnosis of affected cases. While small-scale gene conversions, constituting the majority of variants, can be identified through straightforward sequence analysis, gene deletion detection necessitates a more intricate gene-targeted deletion/duplication panel. Various panels have been developed for detecting gene deletion/duplication, including Southern blot analysis (SB), PCR-based restriction fragment length polymorphism (PCR-based RFLP), quantitative real-time PCR (qPCR), and multiplex ligation-dependent probe amplification (MLPA). […] Our investigation revealed that both qPCR and PCR-based RFLP were effective and congruent. However, qPCR had the added advantage of detecting rare gene duplications, a capability lacking in PCR-based RFLP. The detection of gene duplications (3 N copy number) in carrier screening helps distinguish two scenarios: (1) A heterozygous mutation on one allele and a normal duplication on the other, indicating the individual is a carrier; or (2) A 2 N copy with one normal and one mutated gene on one allele and a normal gene on the other, suggesting the individual is a non-carrier. Thus, qPCR’s ability to detect duplications improves carrier assessment and reduces false positives. […] In summary, for national genetic screening tests, we recommend a combination of either ARMS-PCR or mass spectrometry for detecting common point mutations, paired with PCR-based RFLP for CNV detection. The major trade-off to consider is the reduced accuracy.

• #59 Nationwide carrier screening for congenital adrenal hyperplasia: integrated approach of CYP21A2 pathogenic variant genotyping and comprehensive large gene deletion analysis | BMC Medical Genomics | Full Text

  https://bmcmedgenomics.biomedcentral.com/articles/10.1186/s12920-025-02089-5

  Technically, genomic approaches for deciphering the gene responsible for 21-OHD CAH are intricate due to the multifaceted etiology. The presence of more than one variant in cis complicates the molecular diagnosis of affected cases. While small-scale gene conversions, constituting the majority of variants, can be identified through straightforward sequence analysis, gene deletion detection necessitates a more intricate gene-targeted deletion/duplication panel. Various panels have been developed for detecting gene deletion/duplication, including Southern blot analysis (SB), PCR-based restriction fragment length polymorphism (PCR-based RFLP), quantitative real-time PCR (qPCR), and multiplex ligation-dependent probe amplification (MLPA). […] Our investigation revealed that both qPCR and PCR-based RFLP were effective and congruent. However, qPCR had the added advantage of detecting rare gene duplications, a capability lacking in PCR-based RFLP. The detection of gene duplications (3 N copy number) in carrier screening helps distinguish two scenarios: (1) A heterozygous mutation on one allele and a normal duplication on the other, indicating the individual is a carrier; or (2) A 2 N copy with one normal and one mutated gene on one allele and a normal gene on the other, suggesting the individual is a non-carrier. Thus, qPCR’s ability to detect duplications improves carrier assessment and reduces false positives. […] In summary, for national genetic screening tests, we recommend a combination of either ARMS-PCR or mass spectrometry for detecting common point mutations, paired with PCR-based RFLP for CNV detection. The major trade-off to consider is the reduced accuracy.

• #60 Nationwide carrier screening for congenital adrenal hyperplasia: integrated approach of CYP21A2 pathogenic variant genotyping and comprehensive large gene deletion analysis | BMC Medical Genomics | Full Text

  https://bmcmedgenomics.biomedcentral.com/articles/10.1186/s12920-025-02089-5

  Technically, genomic approaches for deciphering the gene responsible for 21-OHD CAH are intricate due to the multifaceted etiology. The presence of more than one variant in cis complicates the molecular diagnosis of affected cases. While small-scale gene conversions, constituting the majority of variants, can be identified through straightforward sequence analysis, gene deletion detection necessitates a more intricate gene-targeted deletion/duplication panel. Various panels have been developed for detecting gene deletion/duplication, including Southern blot analysis (SB), PCR-based restriction fragment length polymorphism (PCR-based RFLP), quantitative real-time PCR (qPCR), and multiplex ligation-dependent probe amplification (MLPA). […] Our investigation revealed that both qPCR and PCR-based RFLP were effective and congruent. However, qPCR had the added advantage of detecting rare gene duplications, a capability lacking in PCR-based RFLP. The detection of gene duplications (3 N copy number) in carrier screening helps distinguish two scenarios: (1) A heterozygous mutation on one allele and a normal duplication on the other, indicating the individual is a carrier; or (2) A 2 N copy with one normal and one mutated gene on one allele and a normal gene on the other, suggesting the individual is a non-carrier. Thus, qPCR’s ability to detect duplications improves carrier assessment and reduces false positives. […] In summary, for national genetic screening tests, we recommend a combination of either ARMS-PCR or mass spectrometry for detecting common point mutations, paired with PCR-based RFLP for CNV detection. The major trade-off to consider is the reduced accuracy.

• #61 Management challenges and therapeutic advances in congenital adrenal hyperplasia | Nature Reviews Endocrinology

  https://www.nature.com/articles/s41574-022-00655-w

  Adrenal-derived 11-oxygenated androgens have emerged as potential new biomarkers for CAH, as traditional biomarkers are subject to variability and are not adrenal-specific, contributing to management challenges. […] Multiple alternative treatment approaches are being developed with the aim of tailoring therapy for improved patient outcomes. […] This Review focuses on challenges and advances in the management and treatment of CAH due to 21-hydroxylase deficiency, the most common type of CAH. […] Furthermore, we examine new therapeutic developments, including treatments designed to replace cortisol in a physiological manner and adjunct agents intended to control excess androgens and thereby enable reductions in glucocorticoid doses. […] Challenges in the management of congenital adrenal hyperplasia (CAH) arise from multiple hormonal imbalances, the intrinsic tendency of the CAH-affected adrenal gland to overproduce androgens and limited treatment options, which often necessitate glucocorticoid excess.

• #62 Neurocrine Biosciences Announces Classic Congenital Adrenal Hyperplasia Supplement Published Today in The Journal of Clinical Endocrinology & Metabolism | Neurocrine Biosciences

  https://neurocrine.gcs-web.com/news-releases/news-release-details/neurocrine-biosciences-announces-classic-congenital-adrenal

  „Approximately 95% of CAH cases are caused by variants of the CYP21A2 gene that leads to deficiency of the enzyme 21-hydroxylase.” […] „Severe deficiency of this enzyme leads to an inability of the adrenal glands to produce enough cortisol and, in approximately 75% of cases, aldosterone.” […] „Because individuals with CAH are still able to produce androgens, the unused precursors that would normally be used to make cortisol instead result in the production of excess amounts of androgens.” […] „If left untreated, CAH can result in salt wasting, dehydration and even death.” […] „Antagonism of CRF1 receptors in the pituitary has been shown to decrease ACTH levels, which in turn decreases the production of adrenal androgens and potentially the symptoms associated with CAH.” […] „The robust clinical study data demonstrate that lowering adrenal androgen levels with CRENESSITY enables lower, more physiologic dosing of GCs to replace missing cortisol.”

• #63 Management challenges and therapeutic advances in congenital adrenal hyperplasia | Nature Reviews Endocrinology

  https://www.nature.com/articles/s41574-022-00655-w

  Adrenal-derived 11-oxygenated androgens have emerged as potential new biomarkers for CAH, as traditional biomarkers are subject to variability and are not adrenal-specific, contributing to management challenges. […] Multiple alternative treatment approaches are being developed with the aim of tailoring therapy for improved patient outcomes. […] This Review focuses on challenges and advances in the management and treatment of CAH due to 21-hydroxylase deficiency, the most common type of CAH. […] Furthermore, we examine new therapeutic developments, including treatments designed to replace cortisol in a physiological manner and adjunct agents intended to control excess androgens and thereby enable reductions in glucocorticoid doses. […] Challenges in the management of congenital adrenal hyperplasia (CAH) arise from multiple hormonal imbalances, the intrinsic tendency of the CAH-affected adrenal gland to overproduce androgens and limited treatment options, which often necessitate glucocorticoid excess.

• #64 Management challenges and therapeutic advances in congenital adrenal hyperplasia | Nature Reviews Endocrinology

  https://www.nature.com/articles/s41574-022-00655-w

  Adrenal-derived 11-oxygenated androgens have emerged as potential new biomarkers for CAH, as traditional biomarkers are subject to variability and are not adrenal-specific, contributing to management challenges. […] Multiple alternative treatment approaches are being developed with the aim of tailoring therapy for improved patient outcomes. […] This Review focuses on challenges and advances in the management and treatment of CAH due to 21-hydroxylase deficiency, the most common type of CAH. […] Furthermore, we examine new therapeutic developments, including treatments designed to replace cortisol in a physiological manner and adjunct agents intended to control excess androgens and thereby enable reductions in glucocorticoid doses. […] Challenges in the management of congenital adrenal hyperplasia (CAH) arise from multiple hormonal imbalances, the intrinsic tendency of the CAH-affected adrenal gland to overproduce androgens and limited treatment options, which often necessitate glucocorticoid excess.

• #65 Management challenges and therapeutic advances in congenital adrenal hyperplasia | Nature Reviews Endocrinology

  https://www.nature.com/articles/s41574-022-00655-w

  Traditional biomarkers vary with glucocorticoid dose or time of day and are not adrenal-specific, reflecting the need for new biomarkers; for example, the biologically active 11-oxygenated androgens, which are elevated in CAH. […] Circadian glucocorticoid replacement and adjunct non-glucocorticoid therapies promise to enable glucocorticoid dose reduction; furthermore, the development of personalized gene and cellular therapies is under way.

• #66 Management challenges and therapeutic advances in congenital adrenal hyperplasia | Nature Reviews Endocrinology

  https://www.nature.com/articles/s41574-022-00655-w

  Treatment for congenital adrenal hyperplasia (CAH) was introduced in the 1950s following the discovery of the structure and function of adrenocortical hormones. […] Although major advances in molecular biology have delineated steroidogenic mechanisms and the genetics of CAH, management and treatment of this condition continue to present challenges. […] Management is complicated by a combination of comorbidities that arise from disease-related hormonal derangements and treatment-related adverse effects. […] The clinical outcomes of CAH can include life-threatening adrenal crises, altered growth and early puberty, and adverse effects on metabolic, cardiovascular, bone and reproductive health. […] Standard-of-care glucocorticoid formulations fall short of replicating the circadian rhythm of cortisol and controlling efficient adrenocorticotrophic hormone-driven adrenal androgen production.

• #67 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #68 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

  https://www.termedia.pl/A-recent-overview-of-non-classic-congenital-adrenal-hyperplasia-due-to-21-hydroxylase-deficiency-pathophysiology-recognition-and-management,127,51988,1,1.html

  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

• #69 A recent overview of non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: pathophysiology, recognition, and management

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  Steroid hormone synthesis is a multistep process that takes place in the gonads, the cortex of the adrenal glands, and some peripheral tissues. The adrenal gland cortex consists of 3 layers. The outer zona glomerulosa is responsible for the synthesis of mineralocorticoids with the most important representative, aldosterone. In the largest, middle layer (zona fasciculata), glucocorticoids are synthesized with the most important representative, cortisol. The inner zona reticularis is responsible for the adrenal androgens production, mainly dehydroepiandrosterone (DHEA) and DHEA sulphate. It should be also noted that the biosynthesis of steroid hormones is regulated by many factors, such as the transcription and post-translational modification of steroidogenic enzymes, the mechanisms of negative feedback loop of the hypothalamic-pituitary-adrenal axis, the ratio between free and bound circulating steroid compounds, and peripheral metabolism of steroids. In the process of steroidogenesis, 2 types of enzymes are involved: cytochrome P450 (CYP450) and steroid dehydrogenases (HSDs). Dysfunction of the enzyme involved in one of the intermediate stages of steroid biosynthesis in congenital adrenal hyperplasia (CAH) causes insufficient cortisol secretion. Lower level of blood cortisol, the most potent natural glucocorticoid, induces the hypothalamus and pituitary gland to secrete corticoliberin (CRH) and adrenocorticotropin (ACTH), respectively, hormones that indirectly (CRH) and directly (ACTH) stimulate the secretory activity of the adrenal cortex. This vicious circle results in a persistent elevated ACTH level, leading to adrenal hyperplasia, accumulation of steroid hormone precursors prior the action of dysfunctional enzymes, and an undesired shift in hormone synthesis toward other steroidogenic pathways. If there is excessive androgen production, some symptoms of hyperandrogenism become visible. However, non-classic CAH (NCAH) patients typically have normal basal levels of ACTH, cortisol, and mineralocorticoids, but mildly elevated adrenal androgens. Nevertheless, the lack of sufficient cortisol rise after adrenocorticotropic hormone (ACTH) stimulation are reported in one-third of NCAH individuals. Congenital adrenal hyperplasia was described for the first time by an Italian physician almost 150 years ago. He published an autopsy report on a 44-year-old man who had external male genitals, internal female reproductive organs, significantly enlarged adrenal glands, and died during repeated episodes of vomiting and diarrhoea due to an apparent Addisonian crisis. Since then, many types of CAH associated with impaired function of enzymes involved in the synthesis of steroids have been described. The most frequent cause of CAH is 21-hydroxylase deficiency, approximately 8% of CAH cases are associated with 11-hydroxylase deficiency, and further with 17-hydroxylase deficiency, P450 oxidoreductase deficiency, or lipoid congenital adrenal hyperplasia due to StAR protein deficiency. The first case of NCAH was reported in 1957 in a 26-year-old married woman who had regular menstruation periods, properly developed genitalia, and manifested signs of hirsutism and acne. Within CAH, on the basis of clinical symptoms and diagnostic tests, we distinguish 3 forms of the disease: classic salt-wasting (SW CAH), classic without salt loss simple virilizing form (SV CAH), and NCAH. 21-hydroxylase deficiency alters the conversion of 17-hydroxyprogesterone (17OHP) to 11-deoxycortisol, and progesterone to 11-deoxycorticosterone, key precursors in the cortisol and aldosterone synthesis pathway. The accumulated excess of 17-hydroxyprogesterone is metabolized into 21-deoxycortisol or incorporated into a properly functioning androgen production pathway in which 21-hydroxylase plays no role. SW CAH is the most severe form of the disease, which accounts for 75% of all classic CAH cases. It manifests itself when mutations in the CYP21A2 gene are so extensive that 21-hydroxylase almost completely loses its enzymatic activity. A slight increase in the enzymatic activity of 21-hydroxylase compared to SW CAH leads to the development of SV CAH, which is 25% of the classic form of CAH. As in the case of SW CAH, the accumulation of steroid precursors causes an overproduction of adrenal androgen precursors, resulting in the development of ambiguous genitalia in girls of varying severity. Aldosterone synthesis is sufficient to prevent the onset of salt loss and adrenal crisis; therefore, mineralocorticoid replacement therapy is not necessary in normal conditions. It should also be noted that male patients, when screening tests are not performed, are often diagnosed with a delay of several years, after the manifestation of long-term hyperandrogenism symptoms. NCAH is one of the most common autosomal recessive genetic disorders, which very often, especially in men, is never diagnosed. The frequency in the population is estimated at 1:200 to 1:2000 cases, which is up to 50 times higher compared to the classic form of CAH. Among women with signs and symptoms of androgen excess, the prevalence of NCAH is 4.2%. NCAH occurs when the enzymatic function of 21-hydroxylase is affected to a lesser extent (20-70% of normal enzyme function). Aldosterone and cortisol levels are sufficient to support vital functions. Androgens produced by the adrenal cortex, such as DHEA and androstenedione, are relatively weak agonists of androgen receptors, but they may be transformed peripherally into more potent androgens testosterone and dihydrotestosterone. Hyperandrogenic NCAH with similar clinical presentation can also be caused by a defect in other enzymes, such as impairment of 11-hydroxylase activity or 3-hydroxysteroid dehydrogenase deficiency. However, the vast majority of NCAH cases are associated with mutations in the CYP21A2 gene, which encodes 21-steroid hydroxylase. The remaining cases are very rare; therefore, in practice, it can be assumed that NCAH is caused by 21-hydroxylase deficiency. Improperly treated or untreated NCAH causes postnatal hyperandrogenism leading to progressive virilization in boys and girls, including premature pubic and axillary hair development, acne, advanced skeletal age, rapid somatic development, and premature puberty. It should be emphasized that the classification of CAH described above is intended to systematize the possible variants of the disease related to genetic background, and in clinical practice, distinguishing a single pure form of CAH based on clinical symptoms is often difficult, due to the intermingling equivocal symptoms of the disease. This is particularly difficult when distinguishing between SV CAH and NCAH, especially in males with one severe mutation and one non-classic allele. Furthermore, NCAH symptoms may be indistinguishable from those associated with polycystic ovary syndrome (PCOS) or premature adrenarche. The genetic cause of 21-hydroxylase deficiency is quite well understood. The 21-hydroxylase gene (CYP21A2) is a complex gene located on the short arm of chromosome 6, within the HLA locus and adjacent to the genes of the fourth component of the complement, a region where genetic recombination occurs frequently. About 95% of the mutations identified in CAH patients are accounted for by 17 mutations, mainly deletions and insertions, which result from the crossing or conversion of the pseudogene CYP21A2P with the CYP21A2 gene. For this reason, the diversity of genetic mutations in patients with CAH is relatively small. The genotype-phenotype correlation in CAH is generally good, especially for the genotypes for SW CAH (100%) and SV CAH (95%) but only 70% for NCAH. The genetic cause of NCAH has a biallelic nature, and most patients are heterozygotes with different mutations in the 2 alleles. NCAH patients might harbour one severe mutation (classic) and one non-classic allele or 2 non-classic alleles, and the phenotype is considered to be determined by the milder mutation of the 2 affected alleles. The most common genetic cause of NCAH is V281L, which comprises 73-87% of cases. Other NCAH-associated missense mutations include P453S, P30L, R339H, or R369W, and combinations of mutations (heterozygous vs. homozygous) can yield different phenotypes. The clinical symptoms of NCAH are related to increased concentrations of androgens (mainly excessive production of androstenedione) and progestogens (17OHP and progesterone). These hormones produced by the adrenal cortex can exert direct and indirect effects or can be precursors for more potent androgens, especially testosterone and 5-dihydrotestosterone. Recent studies have described adrenal cortex as being able to secrete small amounts of testosterone directly through the action of 17-hydroxysteroid dehydrogenase type 5 on androstenedione, but the clinical importance of these small quantities of adrenal testosterone is largely unknown. In patients with 21OHD, ACTH-dependent androgen excess comes mainly from the 4 pathway, which has been found to be the predominant source of elevated androgen level, and excess 17OHP is converted directly to androstenedione. 17OHP can also be converted through a multiple-step cycle called the alternative or backdoor pathway to 5-dihydrotestosterone, bypassing DHEA and testosterone. Adrenal androgens can also derive from androstenedione or 21-deoxycortisol via the 11-oxygenated androgen pathway, for which the initial steps depend on 11-hydroxylase activity. These 2 metabolites can be converted to 11-hydroxyandrostenedione, and then further to 11-ketoandrostenedione, 11-ketotestosterone, and finally to 11-ketodihydrotestosterone. The last 2 hormones have comparable affinity to the androgen receptor as testosterone and 5-dihydrotestosterone, respectively. NCAH patients with an increased ACTH level have the same pathophysiological mechanisms of androgen excess as classic CAH, but generally NCAH individuals do not present elevated levels of ACTH or CRH or reduced cortisol. In such cases, androgen excess could be attributed to altered enzyme kinetics due to 21OHD. The less efficient conversion ability of 21-hydroxylase results in an increased precursor to product ratio and accumulation of 17OHP, regardless of ACTH levels. Another cause of androgen excess in NCAH patients may be ovarian hypersecretion and peripheral conversion from steroid precursors primarily through the backdoor pathway. Excessive progesterone and androgens by the adrenal glands can cause alterations in hypothalamic-pituitary-ovarian function that promote rapid pulses of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) hypersecretion contributing to androgen excess in the ovaries. Furthermore, overexpression of 5-reductase in the ovary is often observed. LH hypersecretion can initiate and sustain a vicious cycle in which LH stimulates androgen overproduction by theca cells of the ovary, exacerbating the consequences of excessive androgen secretion by the adrenal glands. At birth, children with NCAH have normally developed genitalia consistent with the karyotype of the subject and the 17OHP level is normal or only slightly elevated; therefore, the diagnosis is often made many years later, at puberty or even in adulthood, during treatment of other diseases. Newborn screening most often fails to detect children affected by NCAH. The first symptoms appear no sooner than 60 months after birth, but they can manifest at any age of a person with the burden. In a 2000 multicentre study involving 220 women with NCAH, only 11% were diagnosed before the age of 10 years, while the majority, 80%, of NCAH were diagnosed between the ages of 10 and 40 years. In childhood, symptoms indicative of NCAH are early adrenarche and peripheral precocious puberty, oily skin, premature development of pubic hair, acne, advanced bone age, accelerated growth, and ultimately reduced height compared to predicted. Recent studies have shown that 5-10% of all cases with premature pubic hair and 4-25% of premature puberty cases are patients with NCAH. Early diagnosis and initiation of treatment before the age of 9 years allow the target height calculated from the parents height to be reached. However, data on final height are inconclusive, and it has been reported that in untreated patients with NCAH, the target height is within the normal range in most studies, but contradictory results have also been described. These might suggest that elevated androgen levels in NCAH patients do not lead to significant growth reduction, which is probably more affected by the severity of the phenotype. In adults, the most common symptoms are: hirsutism, acne, menstrual disorders, and fertility disorders. These clinical manifestations are mainly the result of elevated androgen levels and are more pronounced in women. Another well-known complication of CAH, especially in men, is an increased risk of adrenal rest tumour formation, especial testicular adrenal rest tumour (TART), which develops when adrenal rest cells are stimulated by permanently elevated ACTH levels. TARTs are diagnosed in up to 94% of men with CAH and have been reported at all ages, even in 6-year-old boys. However, the incidence of TARTs in patients with NCAH is much lower, but it remains an important cause of male infertility. In order to reduce tumour size and infertility, glucocorticoid therapy should be introduced or intensified. The Endocrine Society Clinical Practice Guideline describes all issues related to the correct diagnosis and management of patients with congenital adrenal hyperplasia. In countries where newborn screening is carried out, the diagnosis is based on the determination of plasma levels of 17-hydroxyprogesterone, a substrate for the dysfunctional 21-hydroxylase enzyme, which can prevent serious infant morbidity and mortality. 17OHP 30 nmol/l in a random blood sample is diagnostic of classic CAH; the cut-off value is below 6 nmol/l at 3 days in full-term infants. The immunoassays use dried blood spots on the same filter paper card taken from the babys heel, which is also used for other newborn screening tests. This allows an unequivocal diagnosis of SW CAH, with a lower probability of SV CAH, whereas NCAH is not usually identified. It should be noted that immunoenzymatic techniques can give false positive results due to cross-reactivity of steroid conjugates and insufficient specificity of the antibodies used. These phenomena are particularly common in preterm infants, sick children, children with low birth weight, and in relation to delayed functional maturation of 11-hydroxylase. On the other hand, glucocorticoid treatment during pregnancy may result in false negative results. Therefore, the reference values of the 17OHP concentration should be stratified by gestational age or birth weight. A positive result should be confirmed by another, more advanced analytical method, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) or gas chromatography-mass spectrometry (GC-MS). If an LC-MS/MS assay is not available, a cosyntropin stimulation test should be performed to confirm the diagnosis before initiating corticosteroid treatment. Despite the lower probability of obtaining false positive results, which are unnecessary concerns for parents, performing the analysis by chromatographic methods is not common because it is an expensive and time-consuming technique and requires specialized knowledge. Chromatographic techniques enable the simultaneous measurement of several analytes in a sample and the determination of the precursor/product ratio, which significantly reduces the possibility of a false positive result, also in preterm and stressed neonates. Obtaining a complete steroid profile is especially recommended in patients with borderline 17OHP levels after a cosyntropin stimulation test to differentiate 21-OHD from other enzyme defects. The diagnosis of NCAH is not as simple as that of classic CAH. Newborn screening is highly beneficial for early detection of classic form, but most screening programs fail to distinguish those individuals affected by NCAH. Many people with NCAH do not show any symptoms of the disease for a long time. The parameters of growth and maturation remain normal, reproductive function is preserved, and the diagnosis is made only as a result of related tests, most often when looking for causes of increased androgen levels. The diagnosis of NCAH is possible using basal and/or dynamic hormone measurements. An effective screening test for NCAH is the measurement of 17OHP in the early morning hours in the follicular phase of the cycle. Early morning measurement is very important because the concentration of 17OHP decreases rapidly during the day. In the screening, 17-OHP concentrations of less than 25 nmol/l in children and less than 6 nmol/l in adults do not exclude the diagnosis of NCAH. The genetic diagnosis of CAH due to 21-OHD is recommended only if the results of the adrenocortical profile after an ACTH stimulation test are inconclusive, or the stimulation test cannot be accurately performed, or for genetic counselling purposes. As mentioned above, most mutations in the CYP21A2 gene encoding 21-hydroxylase result from the exchange of genetic material during meiotic recombination or conversion between this gene and the 98% homologous inactive CYP21A1P pseudogene. CYP21A2 genotyping allows the detection of heterozygosity for mutations of the gene, which may cause CAH in the offspring of these patients. Most NCAH patients carry one allele with a severe mutation for CYP21A2, so there is a 1 in 240 risk for a parent with NCAH to have an offspring with classical CAH. However, the prevalence of CAH among children of women with NCAH was higher than predicted and was 2.5% for CAH and at least 15% for NCAH, presumably due to the fact that affected individuals tend to marry within their own ethnic subpopulations. With normal baseline and after cosyntropin stimulation, 17OHP levels do not exclude carrier status for mild or severe CYP21A2 mutations. The study by Guarnotta et al. confirmed previous reports that 17OHP values after the stimulation test were in the normal range in approximately 50% of heterozygous CYP21A2 mutation carriers, indicating the unsuitability of a simple ACTH stimulation test to detect heterozygosity for 21OHD. The 17OHP/cortisol ratio can be a good and simple tool to identify heterozygous CAH carriers before proceeding to genetic analysis. The study found significantly higher 17OHP/cortisol ratios compared to controls in both heterozygous carriers of the classical CAH and NCAH genes. Thus, the value of the 17OHP/cortisol ratio may be an independent biochemical predictor of gene heterozygosity. The cut-off value of this ratio is 0.03 in heterozygous patients, to differentiate from controls. 21-deoxycortisol has also been shown to be a more sensitive marker than 17OHP, capable of detecting more than 90% of heterozygous CYP21A2 mutation carriers. 21-deoxycortisol is produced by 11-hydroxylation of 17OHP and is generally not excreted in large amounts, and elevated levels are highly specific for 21OHD. However, this marker must be measured by advanced diagnostic methods as LC-MS/MS; therefore, its use in diagnostics is limited for the time being. Conversely, tests that evaluate the ratio of 17OHP/cortisol are readily available and simple to calculate. The objective of treatment in patients with NCAH is to inhibit excessive adrenal androgen synthesis and related complications. In patients diagnosed with NCAH, pharmacological treatment is initiated only in those who manifest clinical symptoms. Most men with NCAH do not need pharmacological therapy, and in women it is indicated when patients have significant symptoms of hyperandrogenism and/or fertility problems. Glucocorticoid therapy in NCAH is also recommended in children and adolescents with premature onset and rapid progression of puberty or accelerated growth velocity with bone maturation significantly advanced, and in adolescents with overt virilization. Treatment is also implemented in children identified on the basis of neonatal screening tests, who develop symptoms relatively early, which is a sign of a more severe phenotype. In NCAH treatment, glucocorticoids are most commonly used to inhibit excessive ACTH secretion, normalizing the cortisol level and excessive adrenal androgen production resulting not so much from cortisol deficiency but from altered enzyme kinetics. During treatment, androgen levels may even be below normal values in both sexes. The steroids most commonly used in NCAH are hydrocortisone, prednisolone, and dexamethasone. Hydrocortisone is preferred in children due to its lower growth inhibition compared to long-acting preparations. The recommended basal dose of hydrocortisone in classic CAH is 10-15 mg/m2 of body surface area, divided into 3 or 4 daily doses. Higher doses may have disadvantageous effects on growth and result in the development of iatrogenic Cushings syndrome. Bone mineral density should also be routinely assessed. In patients with NCAH, the doses required to normalize androgen levels are often much lower. Prednisolone is often preferred in adults due to its simpler dosing and longer action. In the case of dexamethasone, some clinicians are cautious and avoid its use altogether because of its poorer metabolic and bone profile. Therefore, dexamethasone as well as other long-acting glucocorticoids should be shunned in childhood. At the same time, as mentioned earlier, dexamethasone crosses the placenta and can be used to treat a foetus with genetically confirmed CAH. This effect may be undesirable in women with CAH/NCAH with a healthy embryo. However, some physicians choose dexamethasone as the preferred treatment option in NCAH, especially in the case of fertility problems related to TART. Daily glucocorticoid supplementation is recommended in patients diagnosed with NCAH, whose stimulated cortisol level is less than 500 nmol/L. This applies to approximately one-third of NCAH patients. However, it is important to note that when glucocorticoid treatment is initiated, the hypothalamic-pituitary-adrenal axis is inhibited, increasing the risk of adrenal crisis after a severe stress stimulus. Therefore, in situations generating high stress, it may be necessary to increase the doses of glucocorticoid, and sometimes to use intravenous administration. In this case, proper education of patients and their families is important. More recently, modified-release hydrocortisone formulations have been introduced and experimental studies have been conducted with subcutaneous infusion systems, which have been shown to better mimic the circadian rhythm in classic CAH. However, there is no evidence yet of their more beneficial effects on NCAH. In children with increased growth velocity and whose bone age is significantly higher than chronological age, GnRH analogues are helpful in treating secondary GnRH-dependent precocious puberty. However, such therapy should be considered an experimental treatment and is not recommended routinely except if the predicted height SD is 2.25 or below their target height. Most adult men generally do not receive daily glucocorticoid therapy. The exceptions are patients with infertility, adrenal tumours, testicular adrenal rest tumours, and in the case of phenotypes intermediate between classic and non-classic phenotypes. In patients with NCAH, the inclusion of mineralocorticoids (fludrocortisone) is rare and usually implemented to reduce the doses of glucocorticoids. Side effects, such as hypertension and swelling, hinder its use in the elderly, especially women over 50 years of age. Treatment can be terminated during early to intermediate puberty for boys and 2-3 years after menstruation in girls. In girls with persistent symptoms, prolonged glucocorticoid treatment should be avoided. Recent studies have shown that lowering excessive androgen production in NCAH could be achieved not only by glucocorticoid administration. Other treatment options include blocking androgen action with antiandrogens or reducing ovarian androgen secretion with oral contraceptive pills (OCP) or GnRH agonists. A 40-60% decrease in testosterone levels has been reported after oral contraceptive pills containing the oestrogen-progestin combination, due to the inhibitory effect on androgen production in the ovaries, as well as increased synthesis of the sex hormone binding protein (SHBG) in the liver. Currently, the administration of OCP or antiandrogens in adult women with NCAH is becoming the most popular treatment strategy. Among the antiandrogenic drugs that minimize the symptoms of hirsutism, cyproterone acetate and spironolactone, and flutamide and finasteride are the most commonly used. However, these drugs should not be used during pregnancy due to teratogenic effects and for the long term, especially in monotherapy, without including contraceptives due to the risk of disruption of the menstrual cycle frequency and liver damage. Preliminary data suggest that the prostate cancer drug (abiraterone acetate), a steroid 17a-hydroxylase inhibitor, may be a useful therapeutic agent in adult women with NCAH. In mild cases of excessive hair, cosmetic treatments such as shaving, waxing, plucking, electrolysis, and laser therapy can be used to complement medical therapy. This nonpharmacological approach is especially preferred in children and adolescents. In children, parameters of growth rate, body weight, bone age, and pubertal development are used to evaluate the need for inclusion and effectiveness of glucocorticoid therapy. Therefore, children should be regularly monitored clinically for weight, height, signs of androgen excess, puberty, and bone age advancement, while in adults there are no uniform guidelines. Frequent genital examinations should be avoided in girls, unless there are worrisome clinical or laboratory symptoms. The Endocrine Society recommends at least an annual physical examination and determination of hormones (in the morning 17OHP and androstenedione). However, there are reports indicating that a single morning measurement of 17OHP levels has limited utility in adjusting the glucocorticoid dose. Furthermore, normalization of 17OHP may indicate glucocorticoid overtreatment, so moderate elevation of 17OHP should be acceptable. The therapeutic goal should be to maintain androstenedione and testosterone levels within the appropriate age range, and in women the testosterone to SHBG ratio at a level lower than 0.05. More recently, it has been found that the assessment of 11-oxyandrogens, such as 11-ketotestosterone, whose elevated levels have been reported in both NCAH and PCOS, may be useful in monitoring NCAH. However, further studies are needed to clarify the clinical utility of 11-oxyandrogens as biomarkers of adrenal and gonadal overactivity. Several studies have reported that adult patients with NCAH demonstrated an increased risk of metabolic and cardiovascular morbidities. Obesity and type 2 diabetes are significantly more common in NCAH patients than in the general population, as well as cardiovascular disease (mainly stroke). These disorders might be related to prolonged use of glucocorticoids that can improve symptoms of androgen excess and fertility, but they can lead to long-term complications. Therefore, the balance of benefits and harms should always be assessed before implementing glucocorticoid therapy, and regular clinical monitoring is recommended to avoid risk factors and to improve clinical outcomes. NCAH patients have also been shown to have a higher ratio of adrenal incidentalomas and psychiatric diseases such as increased psychotic disorders in males and anxiety disorders in women, which should be reflected in the proper management of patients. Nonclassical congenital adrenal hyperplasia is a difficult endocrine disorder to diagnose and treat due to the direct and indirect effects of the disease on steroidogenesis pathways and ambiguous symptoms. It may manifest in childhood, adolescence, or adulthood. The early diagnosis of the classic form of CAH is crucial for saving newborn lives, while the diagnosis and treatment of NCAH can significantly improve patient comfort and reduce the degree of complaints. NCAH is not a fatal disease, and in many cases it remains undiagnosed, but it can be a burden to many patients affecting their appearance, self-confidence, and therefore quality of life. Advances in metabolomics, genetics, and treatment strategies offer the opportunity to better understand this complex disease and choose the appropriate therapy. Determination of the 17OHP level is used primarily in the screening of 21OHD, while the gold standard of NCAH diagnosis is the ACTH stimulation test with a measurement of raised 17OHP. The introduction of more sensitive and selective analytical techniques, such as LC-MS/MS or GC-MS, can be useful to confirm the diagnosis and targeted analysis of steroid hormones. Identifying mutations within the CYP21A2 gene is important to understand the extent of the mutation and the severity of the disease because there is a wide variation in 17OHP levels among patients. The prevalence of PCOS far exceeds that of NCAH, and despite the overlap in management, infertility treatment and pre-conception considerations warrant careful diagnosis. Therapy should be tailored individually and should aim to minimize symptoms to achieve normal sexual development, fertility, and a generally understood good quality of life. Although there have been many advances in recent years, there is still much to learn about the optimal treatment and management of individuals with both classic and non-classic forms of CAH.

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