A Humanized and Viable Animal Model for Congenital Adrenal Hyperplasia-CYP21A2-R484Q Mutant Mouse.

Congenital Adrenal Hyperplasia (CAH) is an autosomal recessive disorder impairing cortisol synthesis due to reduced enzymatic activity. This leads to persistent adrenocortical overstimulation and the accumulation of precursors before the blocked enzymatic step. The predominant form of CAH arises from mutations in CYP21A2, causing 21-hydroxylase deficiency (21-OHD). Despite emerging treatment options for CAH, it is not always possible to physiologically replace cortisol levels and counteract hyperandrogenism. Moreover, there is a notable absence of an effective in vivo model for pre-clinical testing. In this work, we developed an animal model for CAH with the clinically relevant point mutation p.R484Q in the previously humanized CYP21A2 mouse strain. Mutant mice showed hyperplastic adrenals and exhibited reduced levels of corticosterone and 11-deoxycorticosterone and an increase in progesterone. Female mutants presented with higher aldosterone concentrations, but blood pressure remained similar between wildtype and mutant mice in both sexes. Male mutant mice have normal fertility with a typical testicular appearance, whereas female mutants are infertile, exhibit an abnormal ovarian structure, and remain in a consistent diestrus phase. Conclusively, we show that the animal model has the potential to contribute to testing new treatment options and to prevent comorbidities that result from hormone-related derangements and treatment-related side effects in CAH patients.

A novel homozygous nonsense mutation of VPS13B associated with previously unreported features of Cohen syndrome.

Cohen syndrome (CS) is a rare autosomal recessive disorder associated with mutations in the vacuolar protein sorting 13 homolog B (VPS13B; formerly COH1) gene. The core clinical phenotype comprises a characteristic facial gestalt, marked developmental delay, and myopia. Additional, nonobligatory features include obesity, microcephaly, short stature, muscular hypotonia, scoliosis, narrow hands and feet, progressive retinopathy, as well as neutropenia. Here we report a novel homozygous nonsense mutation in the VPS13B gene and previously undescribed clinical features in a 19-year-old woman with developmental delay, intellectual disability, and a particular facial appearance. The patient showed several features consistent with CS. In addition, the parents observed congenital alacrima and anhidrosis persisting until onset of puberty. The diagnosis was not established based on the clinical phenotype. We performed whole-genome sequencing and identified a novel homozygous nonsense mutation c.62T>G (NM_152564.4), p.(Leu21*) in the VPS13B gene. Our findings extended the previously reported phenotype of CS. We conclude that transient, prepubertal alacrima and anhidrosis are part of the phenotypic spectrum of CS associated with a novel homozygous nonsense mutation in the VPS13B gene.

Biochemical testing for neuroblastoma using plasma free 3-O-methyldopa, 3-methoxytyramine, and normetanephrine.

BACKGROUND: Neuroblastoma, the most common extracranial solid tumor of childhood, produces catecholamines that are metabolized within tumor cells. Homovanillic acid (HVA) and vanillylmandelic acid (VMA), the end products of catecholamine metabolism, have limited accuracy for testing of the tumors. This study assessed whether metabolites produced in earlier steps of catecholamine metabolism might offer improved diagnostic accuracy over urinary HVA and VMA. PROCEDURE: Plasma concentrations of 3-methoxytyramine, normetanephrine, and metanephrine were measured in two pediatric cohorts: (i) 96 children with confirmed neuroblastoma and (ii) 41 children with signs and symptoms of a catecholamine-producing tumor or other neoplasms and in whom neuroblastoma was excluded. Additional measurements of plasma 3-O-methyldopa and relationships of metabolites to MYCN amplification were examined in patient subgroups. RESULTS: Overall, 94 of the 96 patients with neuroblastoma had concentrations of 3-methoxytyramine or normetanephrine above age-specific upper limits of reference intervals, providing a diagnostic sensitivity of 97.9% that was higher (P < 0.0001) than that of 82.2% for HVA and VMA. One of the two patients with normal plasma results showed an elevation of plasma 3-O-methyldopa. Diagnostic specificities were, respectively, 95.1% and 84.8%. Areas under receiver-operating characteristic curves confirmed the superior diagnostic power of the plasma than the urinary test (0.994 vs 0.945; P = 0.0095). Ratios of plasma 3-methoxytyramine to normetanephrine were 7.2-fold higher (P < 0.0001) for patients who had neuroblastomas with MYCN amplification than without MYCN amplification. CONCLUSIONS: Measurements of plasma 3-methoxytyramine and normetanephrine provide a highly accurate diagnostic test for neuroblastoma and also offer potential for prognostic risk stratification.

Just Expect It: Compound Heterozygous Variants of POMT1 in a Consanguineous Family-The Role of Next Generation Sequencing in Neuromuscular Disorders.

Muscular dystrophy-dystroglycanopathies (MDDG) are a group of genetically heterogeneous autosomal recessive disorders characterized by hypoglycosylation of α-dystroglycan. Here, we report on two female patients from a consanguineous Lebanese family that presented in early infancy with generalized muscle hypotonia and primary microcephaly. Brain magnetic resonance imaging (MRI) showed different degrees of hypoplasia of the cerebellar vermis and hypoplasia of corpus callosum. Muscle biopsy analyses revealed a muscular dystrophy with reduced expression of α-dystroglycan and merosin in immunoblot analyses. Homozygosity mapping failed to elucidate the causal mutation due to the accepted notion that, in consanguineous families, homozygote mutations cause disease. However, by applying whole exome sequencing, we identified a novel compound heterozygous POMT1 mutation that segregates with the phenotype and is in line with the clinical presentation. This underscores that a less expected compound heterozygous instead of homozygous mutation in a consanguineous marriage results in a recessive disorder and highlights the growing role of next generation sequencing in neuromuscular disorder diagnostics.

Two patients with MIRAGE syndrome lacking haematological features: role of somatic second-site reversion SAMD9 mutations.

BACKGROUND: Myelodysplasia, infection, restriction of growth, adrenal hypoplasia, genital phenotypes and enteropathy (MIRAGE) syndrome is a recently described congenital disorder caused by heterozygous SAMD9 mutations. The phenotypic spectrum of the syndrome remains to be elucidated. METHODS AND RESULTS: We describe two unrelated patients who showed manifestations compatible with MIRAGE syndrome, with the exception of haematological features. Leucocyte genomic DNA samples were analysed with next-generation sequencing and Sanger sequencing, revealing the patients to have two de novoSAMD9 mutations on the same allele (patient 1 p.[Gln695*; Ala722Glu] and patient 2 p.[Gln39*; Asp769Gly]). In patient 1, p.Gln695* was absent in genomic DNA extracted from hair follicles, implying that the non-sense mutation was acquired somatically. In patient 2, with the 46,XX karyotype, skewed X chromosome inactivation pattern was found in leucocyte DNA, suggesting monoclonality of cells in the haematopoietic system. In vitro expression experiments confirmed the growth-restricting capacity of the two missense mutant SAMD9 proteins that is a characteristic of MIRAGE-associated SAMD9 mutations. CONCLUSIONS: Acquisition of a somatic nonsense SAMD9 mutation in the cells of the haematopoietic system might revert the cellular growth repression caused by the germline SAMD9 mutations (ie, second-site reversion mutations). Unexpected lack of haematological features in the two patients would be explained by the reversion mutations.

A novel TRAPPC11 mutation in two Turkish families associated with cerebral atrophy, global retardation, scoliosis, achalasia and alacrima.

BACKGROUND: Triple A syndrome (MIM #231550) is associated with mutations in the AAAS gene. However, about 30% of patients with triple A syndrome symptoms but an unresolved diagnosis do not harbour mutations in AAAS. OBJECTIVE: Search for novel genetic defects in families with a triple A-like phenotype in whom AAAS mutations are not detected. METHODS: Genome-wide linkage analysis, whole-exome sequencing and functional analyses were used to discover and verify a novel genetic defect in two families with achalasia, alacrima, myopathy and further symptoms. Effect and pathogenicity of the mutation were verified by cell biological studies. RESULTS: We identified a homozygous splice mutation in TRAPPC11 (c.1893+3A>G, [NM_021942.5], g.4:184,607,904A>G [hg19]) in four patients from two unrelated families leading to incomplete exon skipping and reduction in full-length mRNA levels. TRAPPC11 encodes for trafficking protein particle complex subunit 11 (TRAPPC11), a protein of the transport protein particle (TRAPP) complex. Western blot analysis revealed a dramatic decrease in full-length TRAPPC11 protein levels and hypoglycosylation of LAMP1. Trafficking experiments in patient fibroblasts revealed a delayed arrival of marker proteins in the Golgi and a delay in their release from the Golgi to the plasma membrane. Mutations in TRAPPC11 have previously been described to cause limb-girdle muscular dystrophy type 2S (MIM #615356). Indeed, muscle histology of our patients also revealed mild dystrophic changes. Immunohistochemically, ß-sarcoglycan was absent from focal patches. CONCLUSIONS: The identified novel TRAPPC11 mutation represents an expansion of the myopathy phenotype described before and is characterised particularly by achalasia, alacrima, neurological and muscular phenotypes.

"Crying without tears" as an early diagnostic sign-post of triple A (Allgrove) syndrome: two case reports.

BACKGROUND: Triple A syndrome (or Allgrove syndrome) is a rare autosomal recessive disorder characterized by alacrima, achalasia, adrenal insufficiency and autonomic/neurological abnormalities. The majority of cases are caused by mutations in the AAAS gene located on chromosome 12q13. However, the clinical picture as well as genetic testing may be complex since symptomatology is variable and mutations cannot be identified in all clinically diagnosed patients. We present two unrelated patients with triple-A syndrome illustrating the importance of alacrima as an early clinical sign. CASE PRESENTATION: A 3.5 year old girl presented with repeated hypoglycaemic myoclonic events. Adrenal insufficiency was diagnosed. In addition, alacrima, obvious since early infancy, was incidentally reported by the mother and finally lead to the clinical diagnosis of triple A syndrome. This was confirmed by positive mutation analysis of the AAAS gene. The second patient, an 8 months old boy was presented because of anisocoria and unilateral optic atrophy. MRI revealed cerebellar vermis hypotrophy. Psychomotor retardation, failure to thrive, and frequent vomiting lead to further diagnostic work-up. Achalasia was diagnosed radiologically. In addition, the mother mentioned absence of tears since birth leading to the clinical diagnosis of triple A syndrome. In contrast to the first cases genetic testing was negative. CONCLUSION: These two patients illustrate the heterogeneity of triple A syndrome in both terms, clinical expression and genetic testing. We particularly aim to stress the importance of alacrima, which should be considered as a red flag symptom. Further differential diagnosis is required in every child affected by alacrima.

Mutations in GMPPA cause a glycosylation disorder characterized by intellectual disability and autonomic dysfunction.

In guanosine diphosphate (GDP)-mannose pyrophosphorylase A (GMPPA), we identified a homozygous nonsense mutation that segregated with achalasia and alacrima, delayed developmental milestones, and gait abnormalities in a consanguineous Pakistani pedigree. Mutations in GMPPA were subsequently found in ten additional individuals from eight independent families affected by the combination of achalasia, alacrima, and neurological deficits. This autosomal-recessive disorder shows many similarities with triple A syndrome, which is characterized by achalasia, alacrima, and variable neurological deficits in combination with adrenal insufficiency. GMPPA is a largely uncharacterized homolog of GMPPB. GMPPB catalyzes the formation of GDP-mannose, which is an essential precursor of glycan moieties of glycoproteins and glycolipids and is associated with congenital and limb-girdle muscular dystrophies with hypoglycosylation of α-dystroglycan. Surprisingly, GDP-mannose pyrophosphorylase activity was unchanged and GDP-mannose levels were strongly increased in lymphoblasts of individuals with GMPPA mutations. This suggests that GMPPA might serve as a GMPPB regulatory subunit mediating feedback inhibition of GMPPB instead of displaying catalytic enzyme activity itself. Thus, a triple-A-like syndrome can be added to the growing list of congenital disorders of glycosylation, in which dysregulation rather than mere enzyme deficiency is the basal pathophysiological mechanism.

Dysferlinopathy in Switzerland: clinical phenotypes and potential founder effects.

BACKGROUND: Dysferlin is reduced in patients with limb girdle muscular dystrophy type 2B, Miyoshi myopathy, distal anterior compartment myopathy, and in certain Ethnic clusters. METHODS: We evaluated clinical and genetic patient data from three different Swiss Neuromuscular Centers. RESULTS: Thirteen patients from 6 non-related families were included. Age of onset was 18.8 ± 4.3 years. In all patients, diallelic disease-causing mutations were identified in the DYSF gene. Nine patients from 3 non-related families from Central Switzerland carried the identical homozygous mutation, c.3031 + 2 T>C. A possible founder effect was confirmed by haplotype analysis. Three patients from two different families carried the heterozygous mutation, c.1064_1065delAA. Two novel mutations were identified (c.2869 C>T (p.Gln957Stop), c.5928 G>A (p.Trp1976Stop)). CONCLUSIONS: Our study confirms the phenotypic heterogeneity associated with DYSF mutations. Two mutations (c.3031 + 2 T>C, c.1064_1065delAA) appear common in Switzerland. Haplotype analysis performed on one case (c. 3031 + 2 T>C) suggested a possible founder effect.

Mutations in MRAP, encoding a new interacting partner of the ACTH receptor, cause familial glucocorticoid deficiency type 2.

Familial glucocorticoid deficiency (FGD), or hereditary unresponsiveness to adrenocorticotropin (ACTH; OMIM 202200), is an autosomal recessive disorder resulting from resistance to the action of ACTH on the adrenal cortex, which stimulates glucocorticoid production. Affected individuals are deficient in cortisol and, if untreated, are likely to succumb to hypoglycemia or overwhelming infection in infancy or childhood. Mutations of the ACTH receptor (melanocortin 2 receptor, MC2R) account for approximately 25% of cases of FGD. FGD without mutations of MC2R is called FGD type 2. Using SNP array genotyping, we mapped a locus involved in FGD type 2 to chromosome 21q22.1. We identified mutations in a gene encoding a 19-kDa single-transmembrane domain protein, now known as melanocortin 2 receptor accessory protein (MRAP). We show that MRAP interacts with MC2R and may have a role in the trafficking of MC2R from the endoplasmic reticulum to the cell surface.

Fertility and sexual activity in patients with Triple A syndrome.

Objective: Triple A syndrome, caused by autosomal recessively inherited mutations in the AAAS gene is characterized by alacrima, achalasia, adrenal insufficiency, and neurological impairment. To the best of our knowledge, no patients of both sexes have been reported to have offspring. Our aim was to assess the causes of infertility in male patients with this multisystemic syndrome, and to present a female patient that spontaneously conceived a child. Design: Cross-sectional study. Methods: Six males aged 19-48 years were included. Gonadotropins, testosterone, DHEAS, androstenedione, inhibin B, anti-Mullerian hormone measurements and testicular ultrasound were performed. Results: All six male patients had impaired general health and neurological symptoms including erectile and ejaculatory dysfunction. None of them had an offspring. The only demonstrated cause of infertility in our male patients was erectile and ejaculatory dysfunction which precludes sexual intercourse. Our patients had normal libido but were sexually abstinent. Except for low adrenal androgen levels, the concentrations of all measured hormones as well as testicular ultrasound were normal which may indicate the possibility of spermatogenesis in male patients with triple A syndrome. Little is known about fertility in female patients, but based on our observations spontaneous pregnancies seem to be possible. Conclusion: Our results contribute to still scarce knowledge on fertility in patients with Triple A syndrome and as well represents a foundation for further research on causes of infertility and possible treatment options.

Congenital myasthenic syndromes: achievements and limitations of phenotype-guided gene-after-gene sequencing in diagnostic practice: a study of 680 patients.

Congenital myasthenic syndromes (CMSs) are clinically and genetically heterogeneous disorders characterized by a neuromuscular transmission defect. Even though CMSs are genetic disorders, they are highly treatable, and the appropriate drug treatment depends on the underlying genetic defect. This highlights the importance of genetic testing in CMS. In recent years, the molecular basis of CMS has constantly broadened and disease-associated mutations have been identified in 14 genes encoding proteins of the neuromuscular junction. In the dawn of novel sequencing strategies, we report on our 14-year experience in traditional Sanger-based mutation screening of a large cohort of 680 independent patients with suspected CMS. In total, we identified disease-causing mutations in 299 patients (44%) of patients in various known CMS genes, confirming the high degree of genetic heterogeneity associated with the disease. Apart from four known founder mutations, and a few additional recurrent mutations, the majority of variants are private, found in single families. The impact of previously reported genotype-phenotype correlations on efficiency of genetic testing was analyzed in our population. Taking our experiment into account, we present our algorithm for genetic testing in CMS.

A founder mutation in Anoctamin 5 is a major cause of limb-girdle muscular dystrophy.

The limb-girdle muscular dystrophies are a group of disorders with wide genetic and clinical heterogeneity. Recently, mutations in the ANO5 gene, which encodes a putative calcium-activated chloride channel belonging to the Anoctamin family of proteins, were identified in five families with one of two previously identified disorders, limb-girdle muscular dystrophy 2L and non-dysferlin Miyoshi muscular dystrophy. We screened a candidate group of 64 patients from 59 British and German kindreds and found the truncating mutation, c.191dupA in exon 5 of ANO5 in 20 patients, homozygously in 15 and in compound heterozygosity with other ANO5 variants in the rest. An intragenic single nucleotide polymorphism and an extragenic microsatellite marker are in linkage disequilibrium with the mutation, suggesting a founder effect in the Northern European population. We have further defined the clinical phenotype of ANO5-associated muscular dystrophy. Patients show adult onset proximal lower limb weakness with highly raised serum creatine kinase values (average 4500 IU/l) and frequent muscle atrophy and asymmetry of muscle involvement. Onset varies from the early 20 s to 50 s and the weakness is generally slowly progressive, with most patients remaining ambulant for several decades. Distal presentation is much less common but a milder degree of distal lower limb weakness is often observed. Upper limb strength is only mildly affected and cardiac and respiratory function is normal. Females appear less frequently affected. In the North of England population we have identified eight patients with ANO5 mutations, suggesting a minimum prevalence of 0.27/100,000, twice as common as dysferlinopathy. We suggest that mutations in ANO5 represent a relatively common cause of adult onset muscular dystrophy with high serum creatine kinase and that mutation screening, particularly of the common mutation c.191dupA, should be an early step in the diagnostic algorithm of adult limb-girdle muscular dystrophy patients.

Long-term clinical follow-up and molecular genetic findings in eight patients with triple A syndrome.

UNLABELLED: The triple A syndrome (Allgrove syndrome, OMIM #231550) is caused by autosomal recessively inherited mutations in the AAAS gene on chromosome 12q13 encoding the nuclear pore protein ALADIN. This multisystemic disease is characterised by achalasia, alacrima, adrenal insufficiency and neurological impairment. We analyse long-term clinical follow-up and results of sequencing of the AAAS gene in eight patients with triple A syndrome aged from 2 to 35 years. At the time of diagnosis, all patients presented with alacrima, neurological dysfunction, dermatological abnormalities, seven of them with adrenal insufficiency and five of them with achalasia. Sequencing of the AAAS gene identified the p.S263P mutation in five of eight patients, supporting the hypothesis that this mutation is a founder mutation in Slavic population. One of the patients is homozygous for the p.S263P mutation, two are compound heterozygous for the p.S263P and the p.G14fs mutation, two are compound heterozygous for the p.S263Pro mutation and p.S296Y mutation, two are compound heterozygous for the p.G14fs and the p.Q387X mutations and one is homozygous for the p.Q387X mutation. In the course of the follow-up time of 4-29 years, progression of existing and appearance of new symptoms developed. Although severe, many of these symptoms presented in all six young adult patients are often overlooked or neglected: postural hypotension with blurred vision and syncope, hyposalivation resulting with complete edentulosis, talocrular contractures with permanent walking difficulties and erectile dysfunction in male patients. Triple A syndrome is a progressive debilitating disorder which may seriously affect quality of life and even be life-threatening in patients with severe neurological impairment. CONCLUSION: Long-term follow-up of patients with triple A syndrome revealed a variety of the clinical features involving many systems. Progressive natural course of the disease may seriously affect quality of life and even be life-threatening in patients with severe neurological impairment.

Biochemical Diagnosis of Catecholamine-Producing Tumors of Childhood: Neuroblastoma, Pheochromocytoma and Paraganglioma.

Catecholamine-producing tumors of childhood include most notably neuroblastoma, but also pheochromocytoma and paraganglioma (PPGL). Diagnosis of the former depends largely on biopsy-dependent histopathology, but this is contraindicated in PPGL where diagnosis depends crucially on biochemical tests of catecholamine excess. Such tests retain some importance in neuroblastoma though continue to largely rely on measurements of homovanillic acid (HVA) and vanillylmandelic acid (VMA), which are no longer recommended for PPGL. For PPGL, urinary or plasma metanephrines are the recommended most accurate tests. Addition of methoxytyramine to the plasma panel is particularly useful to identify dopamine-producing tumors and combined with normetanephrine also shows superior diagnostic performance over HVA and VMA for neuroblastoma. While use of metanephrines and methoxytyramine for diagnosis of PPGL in adults is established, there are numerous pitfalls for use of these tests in children. The establishment of pediatric reference intervals is particularly difficult and complicated by dynamic changes in metabolites during childhood, especially in infants for both plasma and urinary measurements, and extending to adolescence for urinary measurements. Interpretation of test results is further complicated in children by difficulties in following recommended preanalytical precautions. Due to this, the slow growing nature of PPGL and neglected consideration of the tumors in childhood the true pediatric prevalence of PPGL is likely underappreciated. Earlier identification of disease, as facilitated by surveillance programs, may uncover the true prevalence and improve therapeutic outcomes of childhood PPGL. For neuroblastoma there remain considerable obstacles in moving from entrenched to more accurate tests of catecholamine excess.

Acromicric dysplasia due to a novel missense mutation in the fibrillin 1 gene in a three-generation family.

OBJECTIVES: Short stature is one of the most common reasons for consulting a paediatric endocrinologist. Targeted diagnosis of familial short stature can be challenging due to a broad spectrum of differential diagnoses. CASE PRESENTATION: Here we report a novel mutation in the fibrillin 1 gene (FBN1) in six family members causing a mild phenotype of acromicric dysplasia. Additionally, we present the effects of growth hormone therapy in one of the affected children. CONCLUSIONS: Acromicric dysplasia is a very rare skeletal dysplasia with a prevalence of <1 of 1.000.000 with only about 60 cases being reported worldwide. It is characterized by short stature, acromelia, mild facial dysmorphy but normal intelligence. This study aims to exemplify the clinical and molecular features of FBN1-related acromicric dysplasia and illustrates its pleiotropy by presenting a new, mild phenotype.

CYP21A2 Gene Expression in a Humanized 21-Hydroxylase Mouse Model Does Not Affect Adrenocortical Morphology and Function.

Steroid 21-hydroxylase is an enzyme of the steroid pathway that is involved in the biosynthesis of cortisol and aldosterone by hydroxylation of 17α-hydroxyprogesterone and progesterone at the C21 position. Mutations in CYP21A2, the gene encoding 21-hydroxylase, cause the most frequent form of the autosomal recessive disorder congenital adrenal hyperplasia (CAH). In this study, we generated a humanized 21-hydroxylase mouse model as the first step to the generation of mutant mice with different CAH-causing mutations. We replaced the mouse Cyp21a1 gene with the human CYP21A2 gene using homologous recombination in combination with CRISPR/Cas9 technique. The aim of this study was to characterize the new humanized mouse model. All results described are related to the homozygous animals in comparison with wild-type mice. We show analogous expression patterns of human 21-hydroxylase by the murine promoter and regulatory elements in comparison to murine 21-hydroxylase in wild-type animals. As expected, no Cyp21a1 transcript was detected in homozygous CYP21A2 adrenal glands. Alterations in adrenal gene expression were observed for Cyp11a1, Star, and Cyb11b1. These differences, however, were not pathological. Outward appearance, viability, growth, and fertility were not affected in the humanized CYP21A2 mice. Plasma steroid levels of corticosterone and aldosterone showed no pathological reduction. In addition, adrenal gland morphology and zonation were similar in both the humanized and the wild-type mice. In conclusion, humanized homozygous CYP21A2 mice developed normally and showed no differences in histological analyses, no reduction in adrenal and gonadal gene expression, or in plasma steroids in comparison with wild-type littermates.

Congenital Adrenal Hyperplasia-Current Insights in Pathophysiology, Diagnostics, and Management.

Congenital adrenal hyperplasia (CAH) is a group of autosomal recessive disorders affecting cortisol biosynthesis. Reduced activity of an enzyme required for cortisol production leads to chronic overstimulation of the adrenal cortex and accumulation of precursors proximal to the blocked enzymatic step. The most common form of CAH is caused by steroid 21-hydroxylase deficiency due to mutations in CYP21A2. Since the last publication summarizing CAH in Endocrine Reviews in 2000, there have been numerous new developments. These include more detailed understanding of steroidogenic pathways, refinements in neonatal screening, improved diagnostic measurements utilizing chromatography and mass spectrometry coupled with steroid profiling, and improved genotyping methods. Clinical trials of alternative medications and modes of delivery have been recently completed or are under way. Genetic and cell-based treatments are being explored. A large body of data concerning long-term outcomes in patients affected by CAH, including psychosexual well-being, has been enhanced by the establishment of disease registries. This review provides the reader with current insights in CAH with special attention to these new developments.