Case report: Development of central precocious puberty in a girl with late-diagnosed simple virilizing congenital adrenal hyperplasia complicated with Williams syndrome.

Congenital adrenal hyperplasia (CAH) and Williams Syndrome (WS; MIM # 194050) are distinct genetic conditions characterized by unique clinical features. 21-Hydroxylase deficiency (21-OHD; MIM #201910), the most common form of CAH, arises from mutations in the CYP21A2 gene, resulting in virilization of the external genitalia in affected females, early puberty in males, and short stature. Williams syndrome, caused by a microdeletion of 7q11.23, presents with distinctive facial features, intellectual disability, unique personality traits, early puberty, and short stature. This case report describe the clinical features of a 4-year-old girl referred due to progressive virilization and developmental delay. Genetic analysis confirmed concurrent CAH and WS, identifying a novel mutation in the CYP21A2 gene (c.1442T>C). Following corticosteroid therapy initiation, the patient developed central precocious puberty. This case report delves into the pubertal change patterns in a patient affected by overlapping genetic conditions, providing valuable insights in to the intricate clinical manifestation and management of these rare complex disorders.

Genetic diagnosis of endocrine disorders in Cyprus through the Cyprus Institute of Neurology and Genetics: an ENDO-ERN Reference Center.

The report covers the current and past activities of the department Molecular Genetics-Function and Therapy (MGFT) at the Cyprus Institute of Neurology and Genetics (CING), an affiliated Reference Center for the European Reference Network on Rare Endocrine Conditions (Endo-ERN).The presented data is the outcome of > 15 years long standing collaboration between MGFT and endocrine specialists from the local government hospitals and the private sector. Up-to-date > 2000 genetic tests have been performed for the diagnosis of inherited rare endocrine disorders. The major clinical entities included Congenital Adrenal Hyperplasia (CAH) due to pathogenic variants in CYP21A2 gene and Multiple Endocrine Neoplasia (MEN) type 2 due to pathogenic variants in the RET proto-oncogene. Other rare and novel pathogenic variants in ANOS1, WDR11, FGFR1, RNF216, and CHD7 genes were also found in patients with Congenital Hypogonadotropic Hypogonadism. Interestingly, a few patients with Disorders of Sexual Differentiation (DSD) shared rare pathogenic variants in the SRD5A2, HSD17B3 and HSD3B2 while patients with Glucose and Insulin Homeostasis carried theirs in GCK and HNF1A genes. Lastly, MGFT over the last few years has established an esteemed diagnostic and research program on premature puberty with emphasis on the implication of MKRN3 gene on the onset of the disease and the identification of other prognosis biomarkers.As an Endo-ERN member MGFT department belongs to this large European network and holds the same humanistic ideals which aim toward the improvements of health care for patients with rare endocrine conditions in respect to improved and faster diagnosis.

Giant Bilateral Adrenal Myelolipomas in a Non-Compliant Patient with Congenital Adrenal Hyperplasia.

BACKGROUND 21-hydroxylase deficiency, an essential enzyme for glucocorticoid and mineralocorticoid synthesis, is the cause of congenital adrenal hyperplasia (CAH) in more than 95% of cases. It is an autosomal recessive disorder encoded by the CYP21A2 gene, categorized into classical forms, which encompass the salt-wasting (SW) and simple virilizing (SV) forms, as well as the nonclassical form (NC). The aim of medical treatment is to replace missing glucocorticoids and, if necessary, mineralocorticoids, while also reducing elevated adrenal androgens. CASE REPORT We present the case of a 42-year-old woman with CAH who discontinued therapy during adolescence and was admitted to hospital with fatigue, nausea, and severe abdominal pain. A CT scan showed an extreme enlargement of the adrenal glands. Laboratory tests revealed elevated levels of 17-hydroxyprogesterone and other adrenal androgens, along with normal plasma metanephrine levels. Decreased morning cortisol levels suggested partial adrenal insufficiency requiring glucocorticoid replacement therapy. Due to the development of several serious complications and clinical deterioration, the multidisciplinary team recommended bilateral removal of masses measuring 300×250×200 mm on the right side and 250×200×200 mm on the left side. Histological and immunochemical examination confirmed the presence of giant myelolipomas with adrenal cortex hyperplasia. CONCLUSIONS Adrenal tumors, particularly myelolipomas, have a higher prevalence in patients with CAH. Our case report provides further evidence of the suspected link between non-compliant CAH therapy and the development of myelolipomas, along with promotion of their pronounced growth.

Testicular tumours in adrenogenital syndrome.

Congenital adrenal hyperplasia (CAH) arises from genetic enzyme defects, often in CYP21A2, causing primary adrenal insufficiency. In this case report, a man in his late 20s with lifelong CAH faced challenges in adhering to medication. Suboptimal treatment led to the development of testicular adrenal rest tumours, diagnosed by ultrasound, and hypogonadism. Enhanced adherence restored hormone levels, promoting eugonadism. Adherence plays a crucial role in diminishing tumour size and preventing complications, potentially necessitating orchiectomy in severe cases.

Evaluating the efficacy of a long-read sequencing-based approach in the clinical diagnosis of neonatal congenital adrenocortical hyperplasia.

Congenital adrenal hyperplasia (CAH) is a group of autosomal recessive disorders predominantly characterized by impaired corticosteroid synthesis. Clinical phenotypes include hypoadrenocorticism, electrolyte disturbances, abnormal gonadal development, and short stature, of which severe hyponadrenocorticism and salt wasting can be life-threatening. Genetic analysis can help in the clinical diagnosis of CAH. However, the 21-OHD-causing gene CYP21A2 is arranged in tandem with the highly homologous CYP21A1P pseudogene, making it difficult to determine the exact genotypes using the traditional method of multiplex ligation-dependent probe amplification (MLPA) plus Sanger sequencing or next-generation sequencing (NGS). We applied a long-read sequencing-based approach termed comprehensive analysis of CAH (CACAH) to 48 newborns with CAH that were diagnosed by clinical features and the traditional MLPA plus Sanger sequencing method for retrospective analysis, to evaluate its efficacy in the clinical diagnosis of neonatal CAH. Compared with the MLPA plus Sanger sequencing method, CACAH showed 100 % consistency in detecting SNV/indel variants located in exons and exon-intron boundary regions of CAH-related genes. It can directly determine the cis-trans relationship without the need to analyze parental genotypes, which reduces the time to diagnosis. Moreover, CACAH was able to distinguish different CYP21A1P/CYP21A2 and TNXA/TNXB chimeras, and detect additional variants (CYP21A2 variants c.-121C > T, c.*13G > A, c.*52C > T, c.*440C > T, c.*443 T > C, and TNXB variants c.12463 + 2 T > C, c.12204 + 5G > A). We also identified the TNXB variant c.11435_11524 + 30del alone instead of as a part of the TNXA/TNXB-CH-1 chimera in two newborns, which might be introduced by gene conversion. All of these characteristics enabled clinicians to better explain the phenotype of subjects and manage them more effectively. CACAH has a great advantage over the traditional MLPA and Sanger sequencing methods, showing substantial potential in the genetic diagnosis and screening of neonatal CAH.

The intrafollicular concentrations of biologically active cortisol in women rise abruptly shortly before ovulation and follicular rupture.

STUDY QUESTION: What is the temporal activity and the concentration in follicular fluid (FF) of the anti-inflammatory steroid cortisol during the ovulatory process in humans? SUMMARY ANSWER: Intrafollicular concentrations of cortisol become massively upregulated close to ovulation concomitant with an exceptionally high biological activity securing a timely and efficient termination of inflammatory processes. WHAT IS KNOWN ALREADY: Ovulation has been described as a local, controlled inflammatory process resulting in the degeneration of the follicle wall which facilitate oocyte extrusion. Ovulation also affects the glucocorticoid metabolism of granulosa cells (GCs) and although de novo synthesis of cortisol only occurs in the adrenal cortex, the mid-cycle surge has been shown to induce a change from high expression of HSD11B2, inactivating cortisol to cortisone, to high expression of HSD11B1 which reversibly catalyses cortisol production from cortisone. Furthermore, high concentrations of progesterone and 17OH-progesterone within follicles may cause dislodging of cortisol from cortisol binding protein (CBP) thereby activating the biological activity of cortisol. STUDY DESIGN, SIZE, DURATION: This prospective cohort study included 50 women undergoing fertility treatment according to a standard antagonist protocol at a university hospital-affiliated fertility clinic in Denmark. PARTICIPANTS/MATERIALS, SETTING, METHODS: Women donated FF and GCs from one follicle for research purpose aspirated at one of four time points during the process of final maturation of follicles: T = 0 h, T = 12 h, T = 17 h, T = 32 h. A second sample was collected at oocyte pick up at T = 36 h. The concentration of cortisol and cortisone together with a range of sex steroids was measured by LC-MS/MS in FF collected at the five time points mentioned above. Whole genome microarray data, validated by q-PCR analysis, was used to evaluate gene expression of CYP11B1, CYP21A2, HSD11B1, HSD11B2, and NR3C1 in GCs at the same time points. MAIN RESULTS AND THE ROLE OF CHANCE: The concentration of cortisol was significantly increased from a few nM at 0 h to around 100-140 nM (P ≤ 0.0001) at 32-36 h, whilst cortisone was almost constant from 0 to 17 h at a concentration of between 90 and 100 nM being significantly reduced to 25-40 nM (P ≤ 0.0001) at 32-36 h. This was paralleled by a 690-fold upregulation of HSD11B1 from 0 to 12 h increasing to a more than 20.000-fold change at 36 h. HSD11B2 was quickly downregulated 15- to 20-fold after ovulation induction. Concentrations of progesterone and 17OH-progesterone increased during the ovulatory process to high levels which in essence displaces cortisol from its binding protein CBP due to similar binding affinities. Furthermore, a significant decrease in 11-deoxycortisol expression was seen, but CYP11B1 expression was below detection limit in GCs. LIMITATIONS, REASONS FOR CAUTION: The study included women undergoing ovarian stimulation and results may differ from the natural cycle. More observations at each specific time point may have strengthened the conclusions. Furthermore, we have not been able to measure the actual active biological concentration of cortisol. WIDER IMPLICATIONS OF THE FINDINGS: For the first time, this study collectively evaluated the temporal pattern of cortisol and cortisone concentrations during human ovulation, rendering a physiological framework for understanding potential dysregulations in the inflammatory reaction of ovulation. STUDY FUNDING/COMPETING INTEREST(S): This research was supported by the University Hospital of Copenhagen, Rigshospitalet, and Novo Nordisk Foundation grant number NNF21OC00700556. Interreg V ÔKS through ReproUnion (www.reprounion.eu); Region Zealand Research Foundation. The funders had no role in study design, collection of data, analyses, writing of the article, or the decision to submit it for publication. The authors have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: N/A.

Familial ApoB-specific familial hypobetalipoproteinemia in a patient with non-classical congenital adrenal hyperplasia. / Hipobetalipoproteinemia familiar ApoB específica en una paciente con hiperplasia suprarrenal congénita no clásica.

Familial hypobetalipoproteinaemia is a disorder of lipid metabolism characterized by low levels of total cholesterol, low-density lipoprotein cholesterol and apolipoprotein B. ApoB-related familial hypolipoproteinemia is an autosomal condition with a codominance inheritance pattern. Non-classical congenital adrenal hyperplasia is an autosomal recessive disorder due to mutations in the CYP21A2, a gene encoding for the enzyme 21-hydroxylase, which results in an androgen excess production from adrenal source. We here present the case of a 25-year-old woman with NCAH showing decreased levels of total-cholesterol, low-density lipoprotein cholesterol and triglycerides. Her parent had digestive symptoms and severe hepatic steatosis with elevated liver enzymes, as well as decreased levels of total and low-density lipoprotein cholesterol. A genetic-molecular study of the proband identified a mutation in the APOB gene, which allowed a diagnosis of heterozygous ApoB-related hypolipoproteinaemia to be made.

Genetic prediction of antihyperglycemic drug targets and risk of epilepsy: a mendelian randomisation study.

A connection between diabetes and an increased risk of epilepsy has been suggested by observational studies. Animal studies have also shown that antihyperglycemic drugs can improve seizures. However, it is unclear whether antihyperglycemic drugs have a causal role in epilepsy in humans. To investigate this potential causal relationship, a Mendelian randomisation study was conducted using International League Against Epilepsy data as the discovery set and FinnGen data as the replication set. It was discovered that three antidiabetic drug target genes, ETFDH, CYP21A2 and CYP2D6, were involved in the occurrence of epilepsy. In particular, ETFDH was identified as a target gene in both the discovery set (inverse variance weighting [IVW], odds ratio [OR] = 1.018, 95% confidence interval [CI], 1.004-1.033, p = 0.009) and replication set (IVW, OR = 1.074, 95% CI, 1.034-1.114, p = 0.00016), and CYP21A2 was identified in the discovery set (IVW, OR = 1.029, 95% CI, 1.005-1.053, p = 0.016) and replication set (IVW, OR = 1.057, 95% CI, 1.001-1.116, p = 0.045) as having a causal association with an increased risk of epilepsy. Conversely, the CYP2D6 gene was found to be a protective factor for epilepsy in both the discovery set (IVW, OR = 0.0984, 95% CI, 0.969-0.998, p = 0.025) and replication set (IVW, OR = 0.977, 95% CI, 0.955-1.000, p = 0.046). A search of DrugBank revealed that metformin, an anti-glucose drug, is an inhibitor of the ETFDH gene and may have a potential therapeutic effect on epilepsy.

Quantitative Characterization of Ectopic Adrenal Gene Expression in Fetal Testes in 21-Hydroxylase Deficient Mice.

Testicular adrenal rest tumors (TART) are a frequent and fertility impairing long-term complication in males with classic congenital adrenal hyperplasia. Due to lack of clear experimental data on their origin, they are hypothesized to be derived from ectopic adrenocortical cells within testicular tissue mainly growing upon stimulation by chronically elevated levels of adrenocorticotropin (ACTH). Alternatively, a more totipotent embryological origin has been discussed as the potential source of these tumors. The aim of this study was to quantify alterations of ectopic expression of adrenocortical genes (CYP11B1, CYP11B2, CYP21, MC2R) and the Leydig cell specific marker (INSL3) in testicular tissue of fetal 21-hydroxylase deficient (21OHD) mice. Timed-pregnancy studies were performed using H-2aw18 (aw18)-mice. Testes and adrenals of E15.5 and E18.5 mouse fetuses were used for real-time PCR and immunohistochemistry. Gene expression levels were analyzed for genotype-dependent alterations and compared with immunohistochemistry. While enzymes of steroidogenesis showed a significant increased expression in adrenals of 21OHD mice at both E15.5 and E18.5 compared to wild-type (WT) mice, expression levels were unaltered in testes of 21OHD mice. When compared to WT adrenals a significant increase of INSL3 expression in adrenals of 21OHD mice at E15.5 and E18.5 was detected. Cells with adrenocortical properties in mice fetal testis differ from in situ adrenocortical cells in gene expression and growth at E15.5 and E18.5. These findings suggest that the different local regulation and different local niche in adrenals and testes influence growth of aberrant adrenal cells.

A MinION-based Long-Read Sequencing Application With One-Step PCR for the Genetic Diagnosis of 21-Hydroxylase Deficiency.

CONTEXT: Recently developed long-read sequencing (LRS) technology has been considered an option for CYP21A2 analysis. However, the clinical use of LRS for CYP21A2 analysis is limited. OBJECTIVE: This study's objective is to develop an efficient and low-cost LRS system for CYP21A2 screening. METHODS: A DNA fragment library was prepared in a single polymerase chain reaction (PCR) that covers the entire CYP21A2 gene and all known junctions caused by TNXB gene structural rearrangements, yielding a single 8-kb product of CYP21A2 or CYP21A1P/CYP21A2 chimera. After barcoding, the PCR products were sequenced on a MinION-based platform with Flongle Flow Cell R9.4.1 and R10.4.1. RESULTS: The reference genotypes of 55 patients with 21-hydroxylase deficiency (21OHD) were established using the conventional method with multiplex ligation-dependent probe amplification (MLPA) and nested PCR. LRS using Flongle Flow Cell R9.4.1 yielded consistent results. Additionally, the recently updated LRS "duplex" analysis with Flongle flow cell R10.4.1 was tested to reveal an advantage of accurately sequencing a variant located on the homopolymer region. By introducing a barcode system, the cost was reduced to be comparable to that of conventional analysis. A novel single-nucleotide variation was discovered at the acceptor site of intron 7, c.940-1G > C. We also identified a subtype of the classical chimeric junction CH2, "CH2a," in the region from the latter part of intron 5 to exon 6. CONCLUSION: We successfully established a novel low-cost and highly accurate LRS system for 21OHD genetic analysis. Our study provides insight into the feasibility of LRS for diagnosing 21OHD and other genetic diseases caused by structural rearrangements.

Targeted long-read sequencing for comprehensive detection of CYP21A2 mutations in patients with 21-hydroxylase deficiency.

BACKGROUND: 21-Hydroxylase deficiency (21-OHD) is caused by pathogenic CYP21A2 variations. CYP21A2 is arranged in tandem with its highly homologous pseudogene CYP21A1P; therefore, it is prone to mismatch and rearrangement, producing different types of complex variations. There were few reports on using only one method to detect different CYP21A2 variants simultaneously. AIMS: Targeted long-read sequencing method was used to detect all types of CYP21A2 variants in a series of patients with 21-OHD. METHODS: A total of 59 patients with 21-OHD were enrolled from Peking Union Medical College Hospital. Long-range locus-specific PCR and long-read sequencing (LRS) were performed to detect the pathogenic variants in CYP21A2. RESULTS: Copy-number variants of CYP21A2 were found in 25.4% of patients, including 5.1% with 3 copies of CYP21A2, 16.9% with 1 copy of CYP21A2, and 3.4% with 0 copy of CYP21A2. The remaining 74.6% of patients had 2 copies of CYP21A2. Pathogenic variants were identified in all 121 alleles of 59 patients. Specifically, single-nucleotide variants and small insertions/deletions (< 50 bp) were detected in 79 alleles, of which conversed from CYP21A1P were detected in 63 alleles, and rare variants were found in the other 16 alleles. Large gene conversions (> 50 bp) from pseudogene were detected in 10 alleles, and different chimeric genes (CYP21A1P/CYP21A2 or TNXA/TNXB) formed by large deletions were detected in 32 alleles. Of all variants, p.I173N was the most common variant (19.0%). CONCLUSIONS: Our study demonstrated that targeted long-read sequencing is a comprehensive method for detecting CYP21A2 variations, which is helpful for genetic diagnosis in 21-OHD patients.

Genotype-Specific Cortisol Reserve in a Cohort of Subjects With Nonclassic Congenital Adrenal Hyperplasia (NCCAH).

CONTEXT: Recent guidelines suggest that patients with nonclassic congenital adrenal hyperplasia (NCCAH) stop glucocorticoid therapy after achieving adult height. However, these guidelines do not differentiate between NCCAH genotype groups. OBJECTIVE: Compare ACTH-stimulated cortisol and 17-hydroxyprogesterone (17OHP) levels, and the rate of partial cortisol insufficiency in subjects with NCCAH carrying one mild and one severe (mild/severe) mutation vs subjects with biallelic mild (mild/mild) mutations. METHODS: Retrospective evaluation of the medical records of 122 patients who presented with postnatal virilization and were diagnosed with NCCAH. Patients underwent standard intravenous 0.25 mg/m2 ACTH stimulation testing. Those with stimulated 17OHP level ≥40 nmol/L were screened for the 9 most frequent CYP21A2 gene mutations followed by multiplex ligation-dependent probe amplification. A stimulated cortisol level below 500 nmol/L was defined as partial cortisol deficiency. RESULTS: Patients were subdivided into 3 genotype groups: 77 carried the mild/mild genotype, mainly homozygous for p.V281L mutation; 29 were compound heterozygous for mild/severe mutation, mainly p.V281L/p.I2Splice, and 16 were heterozygous for p.V281L, and were excluded from statistical evaluation. Stimulated cortisol levels were significantly lower in the mild/severe than in the mild/mild group (mean ± SD, 480 ± 90 vs 570 ± 125 nmol/L, P < .001). The mild/severe group exhibited a significantly higher rate of partial cortisol insufficiency (21/28, 75% vs 28/71, 39%, P = .004). Peak 17OHP was significantly higher in the mild/severe group (198 ± 92 vs 118 ± 50 nmol/L, P < .001). CONCLUSION: The high rate of partial adrenal insufficiency in the mild/severe group underscores the need to carefully consider the value of glucocorticoid therapy cessation and the importance of stress coverage in this group.

Predictors of Cardiovascular Morbidities in Adults With 21-Hydroxylase Deficiency Congenital Adrenal Hyperplasia.

CONTEXT: The prevalence of cardiovascular and metabolic complications among adults with 21-hydroxylase deficiency (21OHD) is unknown. OBJECTIVE: We sought to determine the prevalence of cardiovascular and metabolic morbidities among adults with 21OHD and to identify clinical factors and biomarkers associated with cardiovascular outcomes. METHODS: A 10-year retrospective cross-sectional analysis was conducted on adult patients with confirmed 21OHD, aged 18 to 70 years, who had at least one clinical visit for assessment at the University of Michigan. The presence of cardiovascular diseases (CVDs) and other metabolic comorbidities was extracted from medical records based on International Classification of Diseases (ICD) codes. Medical treatments, glucocorticoid (GC) and mineralocorticoid doses, as well as specific biomarkers of disease control since age 18, were collected for analysis. RESULTS: A total of 254 patients with 21OHD, median age of 35 years (interquartile range, 28.25-46 y), were included in the analysis. The prevalence of CVDs in the entire cohort was 7.5%. An increase in prevalence was seen from early adulthood, reaching 25% in patients older than 60 years. Increasing age (adjusted odds ratio [OR], 1.05; 95% CI, 1.01-1.09), hypertension (OR, 4.27; 95% CI, 1.41-12.92), and higher GC doses (OR, 1.51; 95% CI, 1.11-2.06) were significantly associated with prevalent CVDs. Higher plasma renin activity was significantly associated with CVDs (OR, 1.07; 95% CI, 1.01-1.15) but not other biochemical markers of disease. CONCLUSION: Cardiometabolic morbidities are prevalent among adults with 21OHD. Hypertension, age, and GC exposure are the main predictive factors of established CVDs in our cohort.

A novel SMOC1 pathogenic homozygous variant in a fetus with mesomelia of the lower limbs, micrognathia and hypertelorism and an incidental finding of CYP21A2-related congenital adrenal hyperplasia.

Trio exome sequencing was performed on a fetus with bilateral mesomelia of the lower limbs with significant angulation of the tibial bones, micrognathia and hypertelorism detected on ultrasound scan at 19 + 0 weeks gestation. The couple is consanguineous. A homozygous pathogenic frameshift variant in the SMOC1 gene (c.339_340del p.(Phe114Cysfs*40)) was detected and both parents were shown to be heterozygous. Pathogenic variants in the SMOC1 gene are associated with microphthalmia with limb anomalies which multidisciplinary team discussion determined to be causal of the scan anomalies detected. The fetus was also a compound heterozygote for CYP21A2 pathogenic variants, confirming a second diagnosis of non-classical congenital adrenal hyperplasia, which was felt incidental to the scan findings. The risk that this couple's next pregnancy would be affected by either of these disorders is 1 in 4 (25%) and demonstrates the importance of genetic diagnoses for the family and implications for future pregnancies.

[Clinical, genetic, and pathological analysis in 165 children with disorders of sex development]. / 165ä¾‹å„¿ç«¥æ€§å‘è‚²å¼‚å¸¸ç–¾ç— çš„ä¸´åºŠã€é—ä¼ å­¦ä¸Žç— ç†å­¦åˆ†æž.

OBJECTIVES: To investigate the clinical phenotypes, genetic characteristics, and pathological features of children with disorders of sex development (DSD). METHODS: A retrospective analysis was conducted on epidemiological, clinical phenotype, chromosomal karyotype, gonadal pathology, and genotype data of 165 hospitalized children with DSD at Children's Hospital of Hebei Province and Tangshan Maternal and Child Health Hospital from August 2008 to December 2022. RESULTS: Among the 165 children with DSD, common presenting symptoms were short stature (62/165, 37.6%), clitoromegaly (33/165, 20.0%), cryptorchidism (28/165, 17.0%), hypospadias (24/165, 14.5%), and skin pigmentation abnormalities/exteriorized pigmented labia majora (19/165, 11.5%). Chromosomal karyotype analysis was performed on 127 cases, revealing 36 cases (28.3%) of 46,XX DSD, 34 cases (26.8%) of 46,XY DSD, and 57 cases (44.9%) of sex chromosome abnormalities. Among the sex chromosome abnormal karyotypes, the 45,X karyotype (11/57, 19%) and 45,X/other karyotype mosaicism (36/57, 63%) were more common. Sixteen children underwent histopathological biopsy of gonadal tissues, resulting in retrieval of 25 gonadal tissues. The gonadal tissue biopsies revealed 3 cases of testes, 3 cases of dysplastic testes, 6 cases of ovaries, 11 cases of ovotestes, and 1 case each of streak gonad and agenesis of gonads. Genetic testing identified pathogenic/likely pathogenic variants in 23 cases (23/36, 64%), including 12 cases of 21-hydroxylase deficiency congenital adrenal hyperplasia caused by CYP21A2 pathogenic variants. CONCLUSIONS: Short stature, clitoromegaly, cryptorchidism, hypospadias, and skin pigmentation abnormalities are common phenotypes in children with DSD. 45,X/other karyotype mosaicism and CYP21A2 compound heterozygous variants are major etiological factors in children with DSD. The most commonly observed gonadal histopathology in children with DSD includes ovotestes, ovaries, and testes/dysgenetic testes.

Determination of Carrier Frequency of Actionable Pathogenic Variants in Autosomal Recessive Genetic Diseases in the Turkish Cypriot Population.

Whole-exome DNA sequencing is a rich source of clinically useful information for specialists, patients, and their families, as well as elucidating the genetic basis of monogenic and complex diseases in clinical diagnosis. However, interpreting and reporting variants encompassing exome and genome sequence analysis outcome data are one of the greatest challenges of the genomic era. In this study, we aimed to investigate the frequency and allele frequency spectrum of single nucleotide variants accepted as recessive disease carrier status in Turkish Cypriot exomes. The same sequencing platform and data processing line were used for the analysis of data from 100 Turkish Cypriot whole-exome sequence analysis. Identified variants were classified according to ACMG guidelines, and pathogenic variants were confirmed in other databases such as ClinVar, HGMD, Varsome, etc. Pathogenic variants were detected in 68 genes out of 100 whole-exome sequence data. The carriage rate was the highest in the CYP21A2 gene, causing 21-hydroxylase deficiency (14.70%), 11.76% in the HBB gene causing ß-thalassemia, 10.29% in the BTD gene causing biotinidase deficiency, 8.82% in the CFTR gene causing cystic fibrosis, 8.82% in the RBM8A gene causing thrombocytopenia-absent radius syndrome, which is an ultra-rare disease, and 5.88% in the GAA gene causing glycogen storage disease II. The carriage of pathogenic variants in other genes causing the disease (GJB2, PAH, GALC, CYP11B2, COL4A3, HBA1, etc.) was determined as less than 5.00%. Also, the identified variations in the mentioned gene within the examined population were reported. The most prevalent mutation in North Cyprus was a missense variant (c.1360 C>T, p.Pro454Ser) detected in the CYP21A2 gene (rs6445), and the most frequently seen variant in the HBB gene was c.93-21G>A (rs35004220). We investigated reported pathogenic variants by estimating the lower and upper limits of carrier and population frequencies for autosomal recessive diseases, for which exome sequencing may reveal additional medically relevant information. Determining the lower and upper limits of these frequencies will shed light on preventive medicine practices and governmental actions.

Relation between HLA and copy number variation of steroid 21-hydroxylase in a Swedish cohort of patients with autoimmune Addison's disease.

OBJECTIVE: Autoantibodies against the adrenal enzyme 21-hydroxylase is a hallmark manifestation in autoimmune Addison's disease (AAD). Steroid 21-hydroxylase is encoded by CYP21A2, which is located in the human leucocyte antigen (HLA) region together with the highly similar pseudogene CYP21A1P. A high level of copy number variation is seen for the 2 genes, and therefore, we asked whether genetic variation of the CYP21 genes is associated with AAD. DESIGN: Case-control study on patients with AAD and healthy controls. METHODS: Using next-generation DNA sequencing, we estimated the copy number of CYP21A2 and CYP21A1P, together with HLA alleles, in 479 Swedish patients with AAD and autoantibodies against 21-hydroxylase and in 1393 healthy controls. RESULTS: With 95% of individuals carrying 2 functional 21-hydroxylase genes, no difference in CYP21A2 copy number was found when comparing patients and controls. In contrast, we discovered a lower copy number of the pseudogene CYP21A1P among AAD patients (P = 5 × 10-44), together with associations of additional nucleotide variants, in the CYP21 region. However, the strongest association was found for HLA-DQB1*02:01 (P = 9 × 10-63), which, in combination with the DRB1*04:04-DQB1*03:02 haplotype, imposed the greatest risk of AAD. CONCLUSIONS: We identified strong associations between copy number variants in the CYP21 region and risk of AAD, although these associations most likely are due to linkage disequilibrium with disease-associated HLA class II alleles.

Adrenal steroid metabolites and bone status in patients with adrenal incidentalomas and hypercortisolism.

BACKGROUND: Autonomous cortisol secretion (ACS), resulting from cortisol-producing adenomas (CPA), causes endogenous steroid-induced osteoporosis (SIOP). However, the risk of endogenous SIOP cannot be explained by cortisol excess alone, and how other steroid metabolites affect bone status is unclear. METHODS: ACS was diagnosed as serum cortisol ≥1.8 µg/dL after the 1-mg dexamethasone suppression test (DST-cortisol). Using liquid chromatography tandem mass spectrometry, 21 plasma steroid metabolites were measured in 73 patients with ACS and 85 patients with non-functioning adrenal tumors (NFAT). Expression of steroidogenic enzymes and relevant steroid metabolites were analyzed in some of CPA tissues. FINDINGS: Discriminant and principal component analyses distinguished steroid profiles between the ACS and NFAT groups in premenopausal women. Premenopausal women with ACS exhibited higher levels of a mineralocorticoid metabolite, 11-deoxycorticosterone (11-DOC), and lower levels of androgen metabolites, dehydroepiandrosterone-sulfate, and androsterone-glucuronide. In premenopausal women with ACS, DST-cortisol negatively correlated with trabecular bone score (TBS). Additionally, 11-DOC negatively correlated with lumbar spine-bone mineral density, whereas androsterone-glucuronide positively correlated with TBS. The CPA tissues showed increased 11-DOC levels with increased expression of CYP21A2, essential for 11-DOC synthesis. Adrenal non-tumor tissues were atrophied with reduced expression of CYB5A, required for androgen synthesis. INTERPRETATION: This study demonstrates that unbalanced production of adrenal steroid metabolites, derived from both adrenal tumor and non-tumor tissues, contributes to the pathogenesis of endogenous SIOP in premenopausal women with ACS. FUNDING: JSPS KAKENHI, Secom Science and Technology Foundation, Takeda Science Foundation, Japan Foundation for Applied Enzymology, AMED-CREST, JSTA-STEP, JST-Moonshot, and Ono Medical Research Foundation.