X-linked congenital adrenal hypoplasia: Report of long clinical follow-up and description of a new complex variant in the NR0B1 gene.

Adrenal hypoplasia congenita, attributed to NR0B1 pathogenic variants, accounts for more than 50% of the incidence of primary adrenal insufficiency in children. Although more than 250 different deleterious variations have been described, no genotype-phenotype correlation has been defined to date. We report a case of an adopted boy who reported the onset of an adrenal crisis at 2 weeks of age, requiring replacement therapy with mineralocorticoids and glucocorticoids for 4 months. For 3 years, he did well without treatment. At almost 4 years of age, the disorder was restarted. A long follow-up showed the evolution of hypogonadotropic hypogonadism. Molecular studies on NR0B1 revealed a novel and deleterious deletion-insertion-inversion-deletion complex rearrangement sorted in the 5'-3' direction, which is described as follows: (1) deletion of the intergenic region (between TASL and NR0B1 genes) and 5' region, (2) insertion of a sequence containing 37 bp at the junction of the intergenic region of the TASL gene and a part of exon 1 of the NR0B1 gene, (3) inversion of a part of exon 1, (4) deletion of the final portion of exon 1 and exon 2 and beginning of the 3'UTR region, (5) maintenance of part of the intergenic sequence (between genes MAGEB1 and NR0B1, telomeric sense), (6) large posterior deletion, in the same sense. The path to molecular diagnosis was challenging and involved several molecular biology techniques. Evaluating the breakpoints in our patient, we assumed that it was a nonrecurrent rearrangement that had not yet been described. It may involve a repair mechanism known as nonhomologous end-joining (NHEJ), which joins two ends of DNA in an imprecise manner, generating an "information scar," represented herein by the 37 bp insertion. In addition, the local Xp21 chromosome architecture with sequences capable of modifying the DNA structure could impact the formation of complex rearrangements.

SOX3 duplication in a boy with 46,XX ovotesticular disorder of sex development and his 46,XX sister with atypical genitalia: Probable germline mosaicism.

Ovotesticular disorders of sex development (OT-DSD) are characterized by ovarian follicles and seminiferous tubules in the same individual, with a wide range of atypical genitalia. We report on two sibs with atypical genitalia and SRY-negative 46,XX DSD, OT-DSD was confirmed only in the boy, while the girl had bilateral ovaries. Chromosome microarray analysis (CMA) showed a 737-kb duplication at Xq27.1 including the entire SOX3 gene in both sibs, which was confirmed by quantitative real time PCR. Also, X chromosome inactivation assay showed random inactivation in both sibs. Whole exome sequencing revealed no pathogenic or likely pathogenic variant. CMA of the parents showed normal results for both, suggesting that germline mosaicism could be the reason of recurrence of this duplication in the siblings. Our results support a pathogenic role of SOX3 overexpression in 46,XX subjects leading to variable DSD phenotypes.

APOL1 in an ethnically diverse pediatric population with nephrotic syndrome: implications in focal segmental glomerulosclerosis and other diagnoses.

BACKGROUND: APOL1 high-risk genotypes (HRG) are associated with increased risk of kidney disease in individuals of African ancestry. We analyzed the effects of APOL1 risk variants on an ethnically diverse Brazilian pediatric nephrotic syndrome (NS) cohort. METHODS: Multicenter study including 318 NS patients, categorized as progressors to advanced CKD [estimated glomerular filtration rate (eGFR)] < 30 mL/min/1.73 m2] and slow/non-progressors (eGFR > 30 mL/min/1.73 m2 through the study). We employed Cox regression with progression time as the outcome and APOL1 genotype as the independent variable. We tested this association in the entire cohort and three subgroups; (1) focal segmental glomerulosclerosis (FSGS), (2) steroid-resistant NS (SRNS), and (3) those who underwent kidney biopsy. RESULTS: Nineteen patients (6%) had an HRG. Of these, 47% were self-reported White. Patients with HRG manifested NS at older ages and presented higher frequencies of FSGS and SRNS. HRG patients progressed to advanced CKD more often than low-risk-genotype (LRG) children in the whole NS cohort (p = 0.001) and the three subgroups. In SRNS and biopsied patients, a single risk variant was associated with trends of higher CKD progression risk. CONCLUSIONS: Novel discoveries include a substantial prevalence of HRG among patients self-reported White, worse kidney outcomes in HRG versus LRG children in the FSGS subgroup, and a trend of higher CKD progression risk associated with a single risk variant in the SRNS cohort. These findings suggest APOL1-associated NS extends beyond patients self-reported non-White, the HRG effect is independent of FSGS, and a single risk variant may have a detrimental impact in children with NS.

Mutation update for the NR5A1 gene involved in DSD and infertility.

Nuclear receptor subfamily 5 group A member 1 (NR5A1), also named steroidogenic factor 1, is an essential transcription factor that regulates a number of target genes crucial for normal reproductive physiology and endocrine function. It is encoded by NR5A1 gene and is expressed in high doses mainly in steroidogenic tissues, where it controls several steps of adrenal and gonadal development. NR5A1 mutations are associated with a wide phenotypic spectrum of disorders/differences of sex development (DSD), a group of conditions in which development of chromosomal, gonadal, or anatomic sex is atypical. Here, we reviewed 188 NR5A1 mutations from 238 cases reported in literature so far. Additionally, we report the variations p.Ser4*, p.(Cys55Ser), p.(Met78Leu), and p.Met98Glyfs*45, which have not been annotated for NR5A1 before and were identified in some of the 205 46,XY patients of our own cohort. This is the first NR5A1 mutation review which includes both 46,XX and 46,XY karyotype, with the purpose of discussing the complexity of genotype-phenotype correlations among DSD and infertile male patients and also females with primary ovarian failure.

Promises and pitfalls of whole-exome sequencing exemplified by a nephrotic syndrome family.

High-throughput techniques such as whole-exome sequencing (WES) show promise for the identification of candidate genes that underlie Mendelian diseases such as nephrotic syndrome (NS). These techniques have enabled the identification of a proportion of the approximately 54 genes associated with NS. However, the main pitfall of using WES in clinical and research practice is the identification of multiple variants, which hampers interpretation during downstream analysis. One useful strategy is to evaluate the co-inheritance of rare variants in affected family members. Here, we performed WES of a patient with steroid-resistant NS (SRNS) and intermittent microhematuria. Currently, 15 years after kidney transplantation, this patient presents normal kidney function. The patient was found to be homozygous for a rare MYO1E stop-gain variant, and was heterozygous for rare variants in NS-associated genes, COL4A4, KANK1, LAMB2, ANLN, E2F3, and APOL1. We evaluated the presence or absence of these variants in both parents and 11 siblings, three of whom exhibited a milder phenotype of the kidney disease. Analysis of variant segregation in the family, indicated the MYO1E stop-gain variant as the putative causal variant underlying the kidney disease in the patient and two of her affected sisters. Two secondary variants in COL4A4-identified in some other affected family members-require further functional studies to determine whether they play a role in the development of microhematuria in affected family members. Our data illustrate the difficulties in distinguishing the causal pathogenic variants from incidental findings after WES-based variant analysis, especially in heterogenous genetic conditions, such as NS.

Functional characterization of five NR5A1 gene mutations found in patients with 46,XY disorders of sex development.

Steroidogenic factor-1 (SF1), encoded by the NR5A1 gene, is a key regulator of steroidogenesis and reproductive development. NR5A1 mutations described in 46,XY patients with disorders of sex development (DSD) can be associated with a range of conditions of phenotypes; however, the genotype-phenotype correlation remains elusive in many cases. In the present study, we describe the impact of five NR5A1 variants (three novel: p.Arg39Cys, p.Ser32Asn, and p.Lys396Argfs*34; and two previously described: p.Cys65Tyr and p.Cys247*) on protein function, identified in seven patients with 46,XY DSD. In vitro functional analyses demonstrate that NR5A1 mutations impair protein functions and result in the DSD phenotype observed in our patients. Missense mutations in the DNA binding domain and the frameshift mutation p.Lys396Argfs*34 lead to both, markedly affected transactivation assays, and loss of DNA binding, whereas the mutation p.Cys247* retained partial transactivation capacity and the ability to bind a consensus SF1 responsive element. SF1 acts in a dose-dependent manner and regulates a cascade of genes involved in the sex determination and steroidogenesis, but in most cases reported so far, still lead to a sufficient adrenal steroidogenesis and function, just like in our cases, in which heterozygous mutations are associated to 46,XY DSD with intact adrenal steroid biosynthesis.

Dopamine D2 receptor gene polymorphisms and externalizing behaviors in children and adolescents.

BACKGROUND: Dopamine is involved in several cerebral physiological processes, and single nucleotide polymorphisms (SNP) in the dopamine D2 receptor gene (DRD2) have been associated with numerous neurological and mental disorders, including those involving alterations in cognitive and emotional processes. METHODS: The aim of this study was to evaluate the association between the SNPs c.957C > T (rs6277) and c.-585A > G (rs1799978) in the DRD2 gene and behavioral characteristics of children and adolescents based on an inventory of the Child Behavior Checklist (CBCL). Children and adolescents between 8 and 20 years old who were clinically followed-up were genotyped for the SNPs c.957C > T and c.-585A > G, and related to data of the CBCL/6-18 scale assessment performed with the help of caregivers. The chi-squared test was used to assess the differences in the frequencies of the C and T alleles in the polymorphism c.957C > T and of the A and G alleles in the polymorphism c.-585A > G with respect to the grouped CBCL scores at a significance level of 5%. Multiple logistic regression models were performed, to control whether sex and/or ethnicity could influence the results. RESULTS: Eighty-five patients were assessed overall, and the presence of the T allele (C/T and T/T) of DRD2 c.957C > T polymorphism was found to be significantly associated with the occurrence of defiant and oppositional problems and with attention and hyperactivity problems. There were no associations detected with polymorphism DRD2 c.-585A > G polymorphism. Both SNPs were in Hardy-Weinberg-equilibrium. CONCLUSIONS: Although the findings of this study are preliminary, due to its small number of participants, the presence of T allele (C/T, T/T) in c.957C > T SNP was associated with difficulty in impulse control, self-control of emotions, and conduct adjustment, which can contribute to improving the identification of mental and behavioral phenotypes associated with gene expression.

In vitro functional studies of rare CYP21A2 mutations and establishment of an activity gradient for nonclassic mutations improve phenotype predictions in congenital adrenal hyperplasia.

BACKGROUND: A detailed genotype-phenotype evaluation is presented by studying the enzyme activities of five rare amino acid substitutions (Arg233Gly, Ala265Ser, Arg341Trp, Arg366Cys and Met473Ile) identified in the CYP21A2 gene in patients investigated for Congenital adrenal hyperplasia (CAH). OBJECTIVE: To investigate whether the mutations identified in the CYP21A2 gene are disease causing and to establish a gradient for the degree of enzyme impairment to improve prediction of patient phenotype. DESIGN AND PATIENTS: The CYP21A2 genes of seven patients investigated for CAH were sequenced and five mutations were identified. The mutant proteins were expressed in vitro in COS-1 cells, and the enzyme activities towards the two natural substrates were determined to verify the disease-causing state of the mutations. The in vitro activities of these rare mutations were also compared with the activities of four mutations known to cause nonclassic CAH (Pro30Leu, Val281Leu, Pro453Ser and Pro482Ser) in addition to an in silico structural evaluation of the novel mutants. MAIN OUTCOME MEASURE: To verify the disease-causing state of novel mutations. RESULTS: Five CYP21A2 mutations were identified (Arg233Gly, Ala265Ser, Arg341Trp, Arg366Cys and Met473Ile). All mutant proteins exhibited enzyme activities above 5%, and four mutations were classified as nonclassic and one as a normal variant. By comparing the investigated protein changes with four common mutations causing nonclassic CAH, a gradient for the degree of enzyme impairment could be established. Studying rare mutations in CAH increases our knowledge regarding the molecular mechanisms that render a mutation pathogenic. It also improves phenotype predictions and genetic counselling for future generations.

The novel p.Cys65Tyr mutation in NR5A1 gene in three 46,XY siblings with normal testosterone levels and their mother with primary ovarian insufficiency.

BACKGROUND: Disorders of sex development (DSD) is the term used for congenital conditions in which development of chromosomal, gonadal, or phenotypic sex is atypical. Nuclear receptor subfamily 5, group A, member 1 gene (NR5A1) encodes steroidogenic factor 1 (SF1), a transcription factor that is involved in gonadal development and regulates adrenal steroidogenesis. Mutations in the NR5A1 gene may lead to different 46,XX or 46,XY DSD phenotypes with or without adrenal failure. We report a Brazilian family with a novel NR5A1 mutation causing ambiguous genitalia in 46,XY affected individuals without Müllerian derivatives and apparently normal Leydig function after birth and at puberty, respectively. Their mother, who is also heterozygous for the mutation, presents evidence of primary ovarian insufficiency. CASE PRESENTATION: Three siblings with 46,XY DSD, ambiguous genitalia and normal testosterone production were included in the study. Molecular analyses for AR, SRD5A2 genes did not reveal any mutation. However, NR5A2 sequence analysis indicated that all three siblings were heterozygous for the p.Cys65Tyr mutation which was inherited from their mother. In silico analysis was carried out to elucidate the role of the amino acid change on the protein function. After the mutation was identified, all sibs and the mother had been reevaluated. Basal hormone concentrations were normal except that ACTH levels were slightly elevated. After 1 mcg ACTH stimulation test, only the older sib showed subnormal cortisol response. CONCLUSION: The p.Cys65Tyr mutation located within the second zinc finger of DNA binding domain was considered deleterious upon analysis with predictive algorithms. The identification of heterozygous individuals with this novel mutation may bring additional knowledge on structural modifications that may influence NR5A1 DNA-binding ability, and may also contribute to genotype-phenotype correlations in DSD. The slightly elevated ACTH basal levels in all three patients with 46,XY DSD and the subnormal cortisol response after 1 mcg ACTH stimulation in the older sib indicate that a long-term follow-up for adrenal function is important for these patients. Our data reinforce that NR5A1 analysis must also be performed in 46,XY DSD patients with normal testosterone levels without AR mutations.

Screening of Y chromosome microdeletions in 46,XY partial gonadal dysgenesis and in patients with a 45,X/46,XY karyotype or its variants.

BACKGROUND: Partial and mixed gonadal dysgenesis (PGD and MGD) are characterized by genital ambiguity and the finding of either a streak gonad and a dysgenetic testis or two dysgenetic testes. The karyotype in PGD is 46,XY, whereas a 45,X/46,XY mosaicism or its variants (more than two lineages and/or structural abnormalities of the Y chromosome) is generally found in MGD. Such mosaics are also compatible with female phenotype and Turner syndrome, ovotesticular disorder of sex development, and infertility in men with normal external genitalia. During the last few years, evidences of a linkage between Y microdeletions and 45,X mosaicism have been reported. There are also indications that the instability caused by such deletions might be more significant in germ cells. The aim of this work was to investigate the presence of Y chromosome microdeletions in individuals with PGD and in those with 45,X/46,XY mosaicism or its variants and variable phenotypes. METHODS: Our sample comprised 13 individuals with PGD and 15 with mosaicism, most of them with a MGD phenotype (n = 11). Thirty-six sequence tagged sites (STS) spanning the male specific region (MSY) on the Y chromosome (Yp, centromere and Yq) were analyzed by multiplex PCR and some individual reactions. RESULTS: All STS showed positive amplifications in the PGD group. Conversely, in the group with mosaicism, six individuals with MGD had been identified with Yq microdeletions, two of them without structural abnormalities of the Y chromosome by routine cytogenetic analysis. The deleted STSs were located within AZFb and AZFc (Azoospermia Factor) regions, which harbor several genes responsible for spermatogenesis. CONCLUSIONS: Absence of deletions in individuals with PGD does not confirm the hypothesis that instability of the Y chromosome in the gonads could be one of the causes of such condition. However, deletions identified in the second group indicate that mosaicism may be associated with Y chromosome abnormalities detectable only at the molecular level. If patients with mosaicism and Y microdeletions reared as males decide to undergo in vitro fertilization, Y chromosomes which tend to be unstable during cell division may be transmitted to offspring.

DHX37 and the implications in Disorders of Sex Development: an update review.

BACKGROUND: DHX37 is an autosomal gene responsible for encoding a helicase from the DExD/H-box family that plays an essential role in ribosome biogenesis. Variants in this gene were previously reported in two different phenotypes: neurodevelopmental disorders and disorders/differences of sex development (DSD). Particularly for the DSD group, variants were mainly reported associated with gonadal dysgenesis and testicular regression syndrome. SUMMARY: Focusing specific in the DSD group, we revised the 21 DHX37 variants described across a total of 55 cases published in the literature so far. We summarized the most important clinical and molecular features of all cases, trying to have a better comprehensiveness about this gene in the sexual development. KEY MESSAGES: The trick question regarding DHX37 is how a helicase involved in basic cell function could have a specific role in testis development. Little is known about the impact of DHX37 variants in DSD individuals. Nevertheless, current research strongly suggests that DHX37 is involved in the male sex development pathway, particularly in testis determination and maintenance. This is evidenced by the predominant assignment of affected individuals as males and the presence of Wolffian structures in most of the cases. Advancements in molecular techniques, such as the generation of induced pluripotent stem cells, and the digenic inheritance for DHX37 cases, are also addressed in this paper. This represents the first comprehensive review of all DHX37 variants published in the literature to date.

DHX37 and NR5A1 Variants Identified in Patients with 46,XY Partial Gonadal Dysgenesis.

The group of disorders known as 46,XY gonadal dysgenesis (GD) is characterized by anomalies in testis determination, including complete and partial GD (PGD) and testicular regression syndrome (TRS). Several genes are known to be involved in sex development pathways, however approximately 50% of all cases remain elusive. Recent studies have identified variants in DHX37, a gene encoding a putative RNA helicase essential in ribosome biogenesis and previously associated with neurodevelopmental disorders, as a cause of PGD and TRS. To investigate the potential role of DHX37 in disorders of sexual development (DSD), 25 individuals with 46,XY DSD were analyzed and putative pathogenic variants were found in four of them. WES analyses were performed on these patients. In DHX37, the variant p.(Arg308Gln), recurrent associated with DSD, was identified in one patient; the p.(Leu467Val), predicted to be deleterious, was found together with an NR5A1 loss-of-function variant in patient 2; and, the p.(Val999Met) was identified in two unrelated patients, one of whom (patient 3) also carried a pathogenic NR5A1 variant. For both patients carrying DHX37 and NR5A1 pathogenic variants, a digenic inheritance is suggested. Our findings support the importance of DHX37 variants as a cause of disorders of sex development, implying a role in testis development.

So, and if it is not congenital adrenal hyperplasia? Addressing an undiagnosed case of genital ambiguity.

BACKGROUND: The Congenital Adrenal Hyperplasia due to 21 hydroxylase deficiency is the most common cause of genital ambiguity in persons with XX sexual chromosomes. Genital ambiguity among persons with XY sexual chromosomes comprises diverse and rare etiologies. The deficiency of 17-beta-hydroxysteroid dehydrogenase type 3 enzyme (HSD17B3) is a rare autosomal recessive disorder due to functionally altered variants of the HSD17B3 gene. In this disorder/difference of sex development, the conversion of androstenedione into testosterone is impaired. The appearance of external genitalia of 46,XY individuals varies from typically male to almost female. CASE PRESENTATION: We report on a child presenting severe ambiguous genitalia. Due to access constraints, specialized care did not start until the child was 10 months old. Parents are consanguineous and were born in an area of high isonymy that is a cluster for rare recessive diseases. A new homozygous missense variant c.785G > T was found in exon 10 of the HSD17B3 gene. CONCLUSIONS: Researchers-clinicians and researchers-researchers collaborative efforts to elucidate the genetic basis of this disease were critical since this etiologic investigation is not available through the public health system. This case exemplifies the families' pilgrimage in cases of genital ambiguity due to a rare genetic condition. Recognizing the etiology was the baseline to provide information on prognosis and treatment options, and to shelter family and child doubts and hopes in order to better support their decisions.

Molecular diagnosis of 5α-reductase type II deficiency in Brazilian siblings with 46,XY disorder of sex development.

The steroid 5α-reductase type II enzyme catalyzes the conversion of testosterone (T) to dihydrotestosterone (DHT), and its deficiency leads to undervirilization in 46,XY individuals, due to an impairment of this conversion in genital tissues. Molecular analysis in the steroid 5α-reductase type II gene (SRD5A2) was performed in two 46,XY female siblings. SRD5A2 gene sequencing revealed that the patients were homozygous for p.Gln126Arg missense mutation, which results from the CGA > CAA nucleotide substitution. The molecular result confirmed clinical diagnosis of 46,XY disorder of sex development (DSD) for the older sister and directed the investigation to other family members. Studies on SRD5A2 protein structure showed severe changes at NADPH binding region indicating that structural modeling analysis can be useful to evaluate the deleterious role of a mutation as causing 5α-reductase type II enzyme deficiency.

The novel WT1 gene mutation p.H377N associated to Denys-Drash syndrome.

SUMMARY: Denys-Drash syndrome (DDS, Online Mendelian Inheritance in Man number 194080) is a rare human developmental disease generally occurring in 46,XY individuals characterized by the combination of disorder of sex development, early onset nephropathy, and Wilms' tumor (WT). DDS is mainly caused by mutations in the WT1 gene. This report describes a novel WT1 gene mutation in a DDS patient. Sequencing the WT1 gene revealed a heterozygous transversion CAT>AAT within exon 8, causing the substitution of an asparagine for a histidine at residue 377. The p.H377N mutation is predicted to diminish the WT1 protein DNA-binding affinity as it might disrupt the normal zinc finger 2 conformation.

Androgens by immunoassay and mass spectrometry in children with 46,XY disorder of sex development.

OBJECTIVE: Steroid measurement is a challenge in pediatric endocrinology. Currently, liquid chromatography with tandem mass spectrometry (LC-MS/MS) is considered a gold standard for this purpose. The aim of this study was to compare both LC-MS/MS and immunoassay (IA) for androgens before and after human recombinant chorionic gonadotropin (rhCG) stimulus in children with 46,XY disorders of sex development (DSD). METHODS: Nineteen patients with 46,XY DSD were evaluated; all of them were prepubertal and non-gonadectomized. Testosterone, dihydrotestosterone (DHT), DHEA and androstenedione were measured by IA and LC-MS/MS before and 7 days after rhCG injection. The correlation between IA and LC-MS/MS was analyzed by the intraclass correlation coefficient (ICC) and Spearman's rank correlation coefficient (SCC). For concordance analysis the Passing and Bablok (PB) regression and the Bland and Altman (BA) method were used. RESULTS: Testosterone showed excellent correlation (ICC = 0.960 and SCC = 0.964); DHT showed insignificant and moderate correlations as indicated by ICC (0.222) and SCC (0.631), respectively; DHEA showed moderate correlation (ICC = 0.585 and SCC = 0.716); and androstenedione had poor and moderate correlations in ICC (0.363) and SCC (0.735), respectively. Using the PB method, all hormones showed a linear correlation, but proportional and systematic concordance errors were detected for androstenedione, systematic errors for testosterone and no errors for DHEA and DHT. By the BA method, there was a trend of IA to overestimate testosterone and androstenedione and underestimate DHEA and DHT when compared to LC-MS/MS. CONCLUSION: Traditional IA should be replaced by LC-MS/MS for the androgens measurement in prepubertal children whenever is possible.

Impairment in anthropometric parameters and body composition in females with classical 21-hydroxylase deficiency.

The aim of this study was to evaluate the physical measurements and body composition of female patients with the classic form of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency. Twenty-eight girls with CAH were classified according to both hormonal control (well or not well controlled) and the clinical form of the disease (simple virilizing or salt-wasting). In the control group, 112 healthy individuals were included, divided into two subgroups (male and female). Both patients and controls were subdivided by age into three groups according to pubertal stage: < or =10 years (prepubertal), 11-15 years (pubertal), and > or =15 years (postpubertal). Anthropometrical evaluations and bioelectrical impedance were used to obtain the physical measurement and body composition data. The patients with the simple virilizing form presented higher values for BMI, waist, arm fat area, and fat mass percentage. The not well controlled group presented shorter leg length. Values obtained for BMI as well as for arm fat area, brachial circumference, waist, hip, bi-iliac diameter and fat mass percentage were significantly higher in the patients than in the controls, whereas leg length, hand size and the percentages of water and lean mass were lower. Alterations in body composition were observed in all age groups, mainly by increase of fat mass with age. After puberty, impairments in limb measurements (leg, hand and foot) were more evident. Patients with CAH presented differences in anthropometric parameters but mainly in body composition. Hence, more comprehensive and careful anthropometric evaluation during monitoring of patients with CAH is recommended.

XX Maleness and XX true hermaphroditism in SRY-negative monozygotic twins: additional evidence for a common origin.

CONTEXT: Differentiation of testicular tissue in 46,XX individuals is seen either in XX males, the majority of them with SRY gene, or in individuals, usually SRY(-), with ovotesticular disorder of sex development (OT-DSD). Although they are sporadic cases, there are some reports on familial recurrence, including coexistence of XX maleness and OT-DSD in the same family. OBJECTIVE: We report on a case of SRY(-) 46,XX monozygotic twins with genital ambiguity. METHODS: Hormonal evaluation included testosterone, FSH, and LH measurements. SRY gene was investigated by PCR and two-step PCR in peripheral leukocytes and gonadal tissues, respectively. Direct DNA sequencing of the DAX-1 coding sequence was performed. Real-time PCR for SOX9 region on chromosome 17 was obtained. RESULTS: Both twins had a 46,XX karyotype. Twin A had a 1-cm phallus with chordee, penoscrotal hypospadias, and palpable gonads. Serum levels of FSH (2.34 mIU/ml), LH (8.8 mIU/ml), and testosterone (1.6 ng/ml) were normal, and biopsies revealed bilateral testes. Twin B had a 0.5-cm phallus, perineal hypospadias, no palpable gonad on the right, and a left inguinal hernia. Hormonal evaluation revealed high FSH (8.2 mIU/ml) and LH (15 mIU/ml) and low testosterone (0.12 ng/ml). Upon herniotomy, a right testis (crossed ectopia) and a small left ovotestis were found. SRY gene was absent in both peripheral leukocytes and gonadal tissue samples. Neither DAX-1 mutations nor SOX9 duplication was identified. CONCLUSIONS: This case provides evidence that both XX maleness and XX OT-DSD are different manifestations of the same disorder of gonadal development.

Three new Brazilian cases of 17α-hydroxylase deficiency: clinical, molecular, hormonal, and treatment features.

BACKGROUND: Deficiency of 17α-hydroxylase (17OHD) is a rare form of adrenal hyperplasia. Diagnosis is generally delayed, impairing appropriate treatment. CASE PRESENTATION: Here, we report the clinical, molecular, hormonal, and treatment data of three unrelated 17OHD patients, aged 14-16 years with hypergonadotrophic hypogonadism; uncontrolled hypertension; primary adrenal insufficiency; and high progesterone, low to normal potassium, and low dehydroepiandrosterone, androstenedione, and testosterone levels. Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) at baseline and after an adrenocorticotropic hormone test showed low cortisol and cortisone and high deoxycorticosterone (DOC) and corticosterone levels; both DOC/21-deoxycortisol and costicosterone/cortisol ratios were very high. Patient 2 had 46,XX karyotype and patients 1 and 3, had 46,XY. A molecular analysis showed that two of the patients were homozygous for p.W406R mutation and the other patient was compound heterozygous for p.W406R and p.P428L. Hypertension was controlled only after the administration of both prednisone and mineralocorticoid antagonist. CONCLUSIONS: Hypertension in young women must lead to diagnostic suspicion, even in the pre-pubertal period. The basal level of progesterone is an indicator of 17OHD. Mineral and glucocorticoid ratios obtained from LC-MS/MS can reinforce the diagnosis. Hypertension can be controlled using glucocorticoid replacement therapy and mineralocorticoid antagonist.

Comparison between two inhibin B ELISA assays in 46,XY testicular disorders of sex development (DSD) with normal testosterone secretion.

BACKGROUND: Inhibin B is a hormone produced by the Sertoli cells that can provide important information for the investigation of disorders of sex development (DSD) with 46,XY karyotype. The aim of this study is to compare two enzyme-linked immunosorbent assay (ELISA) assays for dosage of serum inhibin B in patients with 46,XY DSD with normal testosterone secretion. METHODS: Twenty-nine patients with 46,XY DSD and normal testosterone secretion (partial androgen insensitivity syndrome [PAIS] [n=8]; 5α-reductase deficiency [n=7] and idiopathic 46,XY DSD [n=14]) were included. Molecular analysis of the AR and SRD5A2 genes were performed in all patients and the NR5A1 gene analysis in the idiopathic group. Measurements of inhibin B were performed by two second-generation ELISA assays (Beckman-Coulter and AnshLabs). Assays were compared using the interclass correlation coefficient (ICC) and the Bland-Altman method. RESULTS: ICC was 0.915 [95% confidence interval (CI): 0.828-0.959], however, a discrepancy was observed between trials, which is more evident among higher values when analyzed by the Bland-Altman method. CONCLUSIONS: It is recommended to perform the inhibin B measurement always using the same ELISA kit when several evaluations are required for a specific patient.