Form Of 21-hydroxylase Deficiency Research Articles

Diagnosis, treatment and preventive measures in congenital adrenal hyperplasia due to 21-hydroxylase deficiency in patients in adulthood

In the article there are presented main approaches to the diagnosis, treatment and follow-up of patients with classical and nonclassical forms of 21-hydroxylase deficiency. The techniques of diagnostic tests and parameters of hormonal indices for the assessment of the adequacy of replacement therapy with glucocorticoids and mineralocorticoids are considered.

A de novo mutation in CYP21A2 gene in a case of in vitro fertilization

Congenital adrenal hyperplasia, one of the most frequent autosome recessive disorders, is caused by defects in steroidogenic enzymes involved in the cortisol biosynthesis. Approximately 95% of the cases are caused by abnormal function of the 21-hydroxylase enzyme. This deficiency leads to androgen excess, consequently, to virilization and rapid somatic growth with accelerated skeletal maturation. Mutations in CYP21A2 are responsible for different forms of 21-hydroxylase deficiency. Mild impairment in the enzymatic activity causes the non-classic or late-onset congenital adrenal hyperplasia that is observed with a prevalence of 1 in 1000 subjects in different populations. The present paper describes a de novo mutation that occurred in the paternal meiosis. The child, who was conceived by in vitro fertilization, presented with precocious puberty and diagnosed with non-classical 21-hydroxylase deficiency. DNA sequencing showed the compound heterozygosis for a de novo CYP21A1P/A2 chimeric gene and the p.Val281Leu mutation inherited from her mother, who was heterozygous for the mutation. The chimeric gene showed pseudogene-derived sequence from 5′-end to intron 3 and CYP21A2 sequences from intron 3 to 3′-end of the gene. Sequencing analysis of the father did not show any mutation. The multiplex ligation-dependent probe amplification (MLPA) assay did not indicate loss of DNA discarding gene deletion but confirmed the chimeric gene. In addition, supernumerary copies of CYP21A1P were observed for both parents and for the affect child. Since paternity has been confirmed, those results suggest that a de novo large gene conversion in the paternal meiosis could have occurred by misalignment of alleles bearing different copy numbers of genes in CYP21 locus.

Prevalence and Clinical Outcome of CYP21A2 Gene Mutations in Patients with Nonfunctional Adrenal Incidentalomas.

Adrenal tumors, discovered incidentally in approximately 4.5% of imaging procedures, are known as adrenal incidentalomas. Nonclassic congenital adrenal hyperplasia, mild form of 21-hydroxylase deficiency, may lead to the development of adrenocortical tumors. The aim of the study was to evaluate prevalence of the most common nonclassic mutations of CYP21A2 gene in patients with adrenal incidentalomas and investigate possible relationship with clinical outcome. One hundred adult patients with such lesions were enrolled. Clinical, imaging and biochemical evaluation were performed to rule out hormonal overproduction or potential malignancy. All subjects and a control group of 100 neonates were genotyped for P30L, P453S, and V281L mutations of CYP21A2 gene using direct sequencing. Clinical and imaging features as well as hormone levels were analyzed. Heterozygous CYP21A2 gene mutations were detected in 8 subjects but not in the neonates. Thus, the risk of carrying mutant allele was significantly higher in subjects with adrenal tumors (OR=8.7; 95% CI=2.23-389.56; p=0.003). Mean concentrations of renin, basal, and stimulated 17-hydroxyprogesterone were higher and ACTH was lower in the carriers than in the remaining subjects. Furthermore, the carriers had higher incidence of hypertension (100 vs. 52.1%, p=0.008) and diabetes (50 vs. 11.9%, p=0.003). ACTH-stimulated 17-hydroxyprogesterone levels varied widely among the carriers. In summary, prevalence of P30L, P453S, and V281L mutations of CYP21A2 gene is increased in patients with adrenocortical tumors. In these subjects, carrying the analyzed mutant alleles may increase the risk of diabetes and hypertension. ACTH-stimulation test does not satisfactorily predict presence of heterozygous CYP21A2 mutations in patients with adrenal tumors.

Déficits en 21-hydroxylase et infertilité féminine : de la physiopathologie à la prise en charge thérapeutique

Steroid 21-hydroxylase deficiency is the most common adrenal genetic disease and is also named congenital adrenal hyperplasia. Depending on the severity of CYP21A2 gene mutations, there are severe or “classical” forms and moderate or “nonclassical” forms of 21-hydroxylase deficiency. The enzyme deficiency causes a disruption of adrenal steroidogenesis, which induces hyperandrogenism and elevated plasma levels of progesterone and 17-hydroxyprogesterone, the two substrates of 21-hydroxylase. These endocrine abnormalities will disrupt gonadal axis, endometrial growth and maturation and finally secretion of cervical mucus. All these phenomena contribute to a female hypofertility. Infertility is more severe in classical forms. When to become pregnant, treatment with hydrocortisone or dexamethasone can limit the production of adrenal androgens and progesterone and improves spontaneous pregnancy rates while minimizing the risk of miscarriage, which is usually relatively high in this disease. When planning pregnancy in patients with a 21-hydroxylase deficiency, genotyping the partner is required to screen for heterozygozity (1/50) and to assess the risk of transmission of a classical form in the progeny.

The prevalence of the non-classical form of congenital adrenal hyperplasia as exemplified (by the population of the Moscow region)

The non-classical form of 21-hydroxylase deficiency (NC21OH) is one of the most common monogenic diseases inherited in the autosomal-recessive pattern. The incidence of this condition in the Russian population, unlike that of its classical variant, remains to be elucidated. Aim. The objective of the present study was to estimate the true incidence of NC21OH based on the prevalence of the two most frequent mutations associated with this disease. A total of 998 randomly selected blood spots were obtained in the course of neonatal screening of the children born within one calendar year at the territory of the Moscow region. The incidence of the disease was calculated with the use of Hardi-Weinberg equation. The minimal prevalence rate of NC21OH in the population of the Moscow region was estimated to be 1:2206. The level of 17-hydroxyprogesterone (17-OHP) calculated based on the results of the screening studies can not be used to identify the carriers of the pathology of interest whereas neonatal screening allows to diagnose no more than 90% of the cases of NC21OH.

Isolated p.H62L Mutation in theCYP21A2Gene in a Simple Virilizing 21-Hydroxylase Deficient Patient

Congenital adrenal hyperplasia due to 21-hydroxylase deficiency accounts for 90%–95% of cases. This autosomal recessive disorder has a broad spectrum of clinical forms, ranging from severe or classical, which includes the salt-wasting and simple virilizing forms, to the mild late onset or nonclassical form. Most of the disease-causing mutations described are likely to be the consequence of nonhomologous recombination or gene conversion events between the active CYP21A2 gene and its homologous CYP21A1P pseudogene. Nevertheless, an increasing number of naturally occurring mutations have been found. The change p.H62L is one of the most frequent rare mutations of the CYP21A2 gene. It was suggested that the p.H62L represents a mild mutation that may be responsible for a more severe enzymatic impairment when presented with another mild mutation on the same allele. In this report, a 20-year-old woman carrying an isolated p.H62L mutation in compound heterozygosity with c.283-13A/C>G mutation is described. Although a mildly nonclassical phenotype was expected, clinical signs and hormonal profile of the patient are consistent with a more severe simple virilizing form of 21-hydroxylase deficiency. The study of genotype-phenotype correlation in additional patients would help in defining the role of p.H62L in disease manifestation.

Zona fasciculata 21-hydroxysteroids and precursor-to-product ratios in 21-hydroxylase deficiency: further characterization of classic and non-classic patients and heterozygote carriers.

Although much is known about the increased levels of the 21-hydroxylase substrates 17-hydroxyprogesterone (17OHP) and 21-deoxycortisol (21DF) - the biochemical markers of all forms of 21-hydroxylase deficiency (21OHD), only limited information is available on the zona fasciculata (ZF) products distal to the enzymatic block: 11-deoxycortisol (S), 11-deoxycorticosterone (DOC), and corticosterone (B). To investigate whether basal and post-ACTH levels of S, DOC, and B and the 21-hydroxylase precursor-to-product ratios determined by tandem mass spectrometry preceded by high-performance liquid chromatography separation (liquid chromatography-tandem mass spectrometry) could disclose distinct profiles in genotypically confirmed classic (no.=14) and non-classic (NC) (no.=18) patients, heterozygote carriers (no.=61) and wildtypes (WT) (no.=27) for 21OHD. Salt wasting (SW) and simple virilizing (SV) had higher basal levels of DOC with no further increase in response to ACTH. Stimulated DOC was similar in 21OHD patients and carriers but was reduced as compared to WT. ACTH-stimulated B increased gradually from SW and SV through WT. The post-ACTH 21DF/B ratio was able to detect 92% of the carriers among WT. All NC patients could be detected by post-ACTH 17OHP/DOC and 21DF/B, with no overlap with 21OHD carriers. Although 21-hydroxylase is a key enzymatic step in both 17-hydroxy and 17-deoxy pathways of ZF, the reaction is mostly affected in the latter pathway, leading to a significant impairment of B production, which may further characterize the 21OHD subtypes. Also, the precursor-to-product ratios, particularly 21DF/B, can demonstrate the distinctive outline of 21OHD subtypes, including carriers and normal subjects.

Low-dose ACTH test for the diagnosis of non-classical form of congenital adrenocortical dysfunction

The possibilities to diagnose the non-classical form of 21-hydroxylase deficiency using the low-dose (5 mcg) 1-24 ACTH stimulation test are considered.

Klinik und Diagnostik bei Männern mit leichtem 3β-Hydroxysteroiddehydrogenase-Mangel

3 beta-hydroxysteroid dehydrogenase (HSD) deficiency was demonstrated in six males, aged between 18 and 24 years, who had gynaecomastia, hypogonadism or infertility. The predominant laboratory finding was a striking elevation of dehydroepiandrosterone sulphate (DHEAS) levels. The diagnosis of HSD deficiency was confirmed by finding a marked rise in dehydroepiandrosterone (DHEA) and 17-hydroxypregnenolone levels. In contrast to these findings in late-onset enzyme deficiency, in four males with the classical form of 21-hydroxylase deficiency the only sign was a reduction in adult height. The prevalence of late-onset HSD deficiency in men is not known and may be more relevant in patients with gynaecomastia or abnormal gonadal function than has hitherto been realized.

Late Diagnosis of Congenital Adrenal Hyperplasia Due to 21-Hydroxylase Deficiency

Congenital adrenal hyperplasia results from 21-hydroxylase deficiency in more than ninety percent of cases. The classical form of 21-hydroxylase deficiency presents in the neonatal period with virilization or adrenal insufficiency, with or without concurrent salt wasting. We report on a rare case of classic 21-hydroxylase deficiency diagnosed in late adulthood. A 39-year-old male patient presented for workup of infertility. Urologic investigation revealed small testes, bilateral testicular masses, and asthenozoospermia. The patient's steroid metabolism showed markedly increased levels of adrenal androgens, in particular of 17-hydroxyprogesterone amd 21-deoxycortisol. The gas chromatographic-mass spectrometric (GC-MS) urinary steroid profile was dominated by metabolites of 17-hydroxyprogesterone, while the endogenous glucocorticoid production was subnormally low. ACTH levels in plasma were elevated. These hormonal findings were consistent with 21-hydroxylase deficiency. Therapy with dexamethasone was initiated. The CTP21A2 gene analysis revealed the mutation I172N (ATC --> AAC) in exon 4 of allele 1 and a large gene deletion in allele 2. Cases of 21-hydroxylase deficiency diagnosed in late adulthood are rare; however, clinicians should be alert of this possibility.

Nonclassic congenital adrenal hyperplasia resulting from 21-hydroxylase deficiency

Nonclassic CAH, also termed as late onset of CAH, is a very mild form of 21-hydroxylase deficiency. The incidence of disease is estimated at 0.1% of population. Nonclassic CAH is usually diagnosed in the childhood before the age of 6 to 8 years as premature pubarche. The disease is not common in the infants and usually not before 6 to 8 months. This is a case report of 7-month female infant who was suspected of mild hyperandrogenism because of premature pubarche. The diagnosis was confirmed by mild basal elevation of 17-OHP (5.55 ng/ml) and characteristic hyper-response to ACTH, reaching values of 21 ng/ml, as well as accelerated bone maturation. The conventional treatment of NCAH was initiated, with glucocorticoid therapy (hydrocortisone) for one year and a half. After that period, our decision was to discontinue the hormonal therapy because of the impression that hyperandrogenism was mild (mild deficiency of the enzymes for steroid hormone synthesis). Child's growth, development and maturation are under constant control.

MASSIVE ADRENOCORTICAL ADENOMA FOLLOWING LONG-TERM TREATMENT OF 21-HYDROXYLASE DEFICIENCY

Recent reports have revealed a high prevalence of small tumors in patients with untreated congenital adrenal hyperplasia due to 21-hydroxylase deficiency.1, 2 However, to our knowledge no case of long treated 21-hydroxylase deficiency complicated by a large adenoma has been reported in the literature. We report on a female patient with the simple virilizing form of 21-hydroxylase deficiency in whom a massive adrenal adenoma developed following 21 years of steroid treatment. CASE REPORT

Variable ACTH-stimulated 17-hydroxyprogesterone values in 21-hydroxylase deficiency carriers are not related to the different CYP21 gene mutations.

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

H28+C Insertion in the CYP21 Gene: A Novel Frameshift Mutation in a Brazilian Patient with the Classical Form of 21-Hydroxylase Deficiency

H28+C Insertion in the CYP21 Gene: A Novel Frameshift Mutation in a Brazilian Patient with the Classical Form of 21-Hydroxylase Deficiency

Molecular genotyping in Brazilian patients with the classical and nonclassical forms of 21-hydroxylase deficiency.

The aim of our study was to determine, by allele-specific PCR, the frequency of point mutations in 130 Brazilian patients with the classical and nonclassical forms of 21-hydroxylase deficiency and to correlate genotype with phenotype. The most frequent mutations were 12 splice (41.8% in salt wasting), I172N (32.6% in simple virilizing), and V281L (40.2% in late onset form). The frequency of the 9 most common point mutations was similar to that reported for other countries, except for Del 8 nt and Cluster, which were less frequent in the classical form. Rarer mutations such as P453S, G291S, I7 splice, W405X, R483P, and R483-->frameshift were rarely found or were absent. The 93 fully genotyped patients were classified into 3 mutation groups, based on the degree of enzymatic activity (group A, <2%; group B, approximately 2%, and group C, >18%). In group A, 62% of the cases presented the salt wasting form; in group B, 96% the simple virilizing form; and in group C, 88% the late onset form. We diagnosed 80% of the affected alleles after screening for large rearrangements and 15 point mutations. The absence of previously described mutations in 20% of the affected alleles suggests the presence of new mutations in our population.

Nonclassical 21-Hydroxylase Deficiency Detected in Newborn Mass Screening Program for Congenital Adrenal Hyperplasia (CAH): Report of a Case

We identified a patient with a nonclassical (NC) form of 21-hydroxylase deficiency (21-OHD). A 6-month-old boy was referred to the Shinshu University Hospital because of increased concentration of serum 17-OH progesterone (17-OHP). The high 17-OHP concentration was detected in mass screening program for CAH. He did not manifest any signs or symptoms of adrenal insufficiency or androgen excess at birth. Molecular analysis of the CYP21 gene revealed that he carried a compound heterozygote with the I172N mutation in one allele and the V281L mutation in the other allele. This suggests that the newborn mass screening can detect some NC 21-OHD cases in which both alleles contain only NC-associated mutations. He is the first patient with the NC phenotype which shares the same mutation as caucasian NC patients.

21-hydroxylase deficiency.

A deficiency of 21-hydroxylase in the adrenal cortex results in insufficient cortisol production. The salt-wasting form of21-hydroxylase deficiency is characterized by inadequate aldosterone production, as well. Because the hypothalamic-adr enal negative feedback system is broken, excess adrenal androgens are produced. This disordered corticosteroid production causes hormonal and clinical symptoms, including pseudohermaphrod itism in genetic females and disordered puberty in both males and females. There is a spectrum of the time of onset and the severity of these symptoms. This disorder is inherited in an autosomal recessive manner. The 21-hydroxylase deficiency is genetically linked to the human leukocyte antigen (HLA)

Altered CYP21 genes in HLA-haplotypes associated with congenital adrenal hyperplasia (CAH): a family study

Disorders of the CYP21 gene, which is located within the major histocompatibility complex on the short arm of chromosome 6, are the leading causes of congenital adrenal hyperplasia (CAH). The coding gene and a highly homologous pseudogene are tandemly arranged with the two genes for the fourth component of complement (C4A and C4B). To analyse the prevalence rates of mutations of the CYP21 genes and the segregation of the CYP21 genes with their corresponding human leucocyte antigen (HLA)-haplotypes, 21 families with one or two children with the severe form of 21-hydroxylase deficiency were studied. Mutations of the CYP21 gene on their corresponding HLA-haplotype were detected by hybridisation of polymerase chain reaction (PCR)-amplified genomic DNA with sequence-specific oligonucleotides and solid phase direct sequencing. Our study has shown the following. (1) A single basepair mutation (A-->G or C-->G) within the second intron is the most frequent mutation leading to impaired 21-hydroxylase activity. This mutation is only detected in HLA-haplotypes associated with the salt-wasting form of CAH. (2) A large deletion of part or all of the CYP21 gene is associated with the HLA-haplotype A3, BW47, C6, DR7, DR53, DQ2 but is also observed in other HLA-haplotypes and can be detected by a simple rapid PCR restriction fragment length polymorphism method. (3) Two alleles of the coding CYP21 gene differing in a leucine codon within the first exon, (formerly described as a mutation associated with 21-hydroxylase deficiency) have been found with an equal distribution in patients with 21-hydroxylase deficiency, non-disease HLA-haplotypes and the local healthy controls.

Genetic basis of endocrine disease 2: congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

Most cases of congenital adrenal hyperplasia, the inherited inability to synthesize cortisol, are caused by a deficiency of the 21-hydroxylase activity required to convert 17-hydroxyprogesterone to 11-deoxycortisol. Poor synthesis of cortisol results in chronic stimulation of the adrenal cortex by corticotropin with consequent overproduction of cortisol precursors. Some of these precursors are shunted into the androgen biosynthetic pathway causing the signs and symptoms of androgen excess seen in this disorder. The disease occurs in a wide spectrum of clinical variants, including a severely affected form with a concurrent defect in aldosterone biosynthesis (“salt wasting” type), a form with apparently normal aldosterone biosynthesis (“simple virilizing” type) and a mild “nonclassic” form that may be asymptomatic or may be associated with symptoms of androgen excess developing during childhood or at puberty (1). The disorder is inherited as a monogenic autosomal recessive trait located within the HLA major histocompatibility complex on chromosome 6~21.3. There are a number of associations between the different forms of 21-hydroxylase deficiency and specific combinations of HLA antigens, or haplotypes. In particular, the unusual haplotype A3;Bw47;DR7 is associated with about 10% of classic (i.e. salt wasting or simple virilizing) 21-hydroxylase deficiency alleles, whereas HLA-B14;DRl is associated with 70% of alleles for nonclassic disease. Based on these associations and on studies of pedigrees containing patients with the different forms of the disease, it appears that these forms are inherited as allelic variants. Classic disease occurs in approximately l/10,000 births, whereas nonclassic disease occurs in approximately 1% of the general population (2). The molecular genetic basis of 21-hydroxylase deficiency has been thoroughly studied. The 21-hydroxylase enzyme is a microsomal cytochrome P450 termed P45OXXI or P45Oc21. The structural gene encoding

Changes in Size and Sonographic Characteristics of the Adrenal Glands During the First Year of Life and the Sonographic Diagnosis of Adrenal Hyperplasia in Infants with 21‐Hydroxylase Deficiency

The sonographic representation of developmental changes occurring in the normal adrenal gland during the first year of life was examined in 84 healthy newborns and infants. As a measure of adrenal size we chose a defined part of standardized adrenal sections, the adrenal size index (ASI). In newborns, ASI was high and decreased during the first year of life. Three different echo types were delineated, each one being predominant in a defined age group. ASI was markedly elevated in 5 patients with the salt-wasting variety of 21-hydroxylase deficiency, whereas in 3 patients without a severe salt loss, ASI was in the upper normal range. Although adrenal enlargement due to salt-wasting 21-hydroxylase deficiency can be recognized by determining ASI, other forms of 21-hydroxylase deficiency do exist in which adrenal enlargement, detectable by ASI measurements, may not be present.