Towards improved genetic diagnosis of human differences of sex development.

Despite being collectively among the most frequent congenital developmental conditions worldwide, differences of sex development (DSD) lack recognition and research funding. As a result, what constitutes optimal management remains uncertain. Identification of the individual conditions under the DSD umbrella is challenging and molecular genetic diagnosis is frequently not achieved, which has psychosocial and health-related repercussions for patients and their families. New genomic approaches have the potential to resolve this impasse through better detection of protein-coding variants and ascertainment of under-recognized aetiology, such as mosaic, structural, non-coding or epigenetic variants. Ultimately, it is hoped that better outcomes data, improved understanding of the molecular causes and greater public awareness will bring an end to the stigma often associated with DSD

Genome-wide neonatal epigenetic changes associated with maternal exposure to the COVID-19 pandemic

Abstract Background During gestation, stressors to the fetus, including viral exposure or maternal psychological distress, can fundamentally alter the neonatal epigenome, and may be associated with long-term impaired developmental outcomes. The impact of in utero exposure to the COVID-19 pandemic on the newborn epigenome has yet to be described. Methods This study aimed to determine whether there are unique epigenetic signatures in newborns who experienced otherwise healthy pregnancies that occurred during the COVID-19 pandemic (Project RESCUE). The pre-pandemic control and pandemic cohorts (Project RESCUE) included in this study are part of a prospective observational and longitudinal cohort study that evaluates the impact of elevated prenatal maternal stress during the COVID-19 pandemic on early childhood neurodevelopment. Using buccal swabs collected at birth, differential DNA methylation analysis was performed using the Infinium MethylationEPIC arrays and linear regression analysis. Pathway analysis and gene ontology enrichment were performed on resultant gene lists. Results Widespread differential methylation was found between neonates exposed in utero to the pandemic and pre-pandemic neonates. In contrast, there were no apparent epigenetic differences associated with maternal COVID-19 infection during pregnancy. Differential methylation was observed among genomic sites that underpin important neurological pathways that have been previously reported in the literature to be differentially methylated because of prenatal stress, such as NR3C1. Conclusions The present study reveals potential associations between exposure to the COVID-19 pandemic during pregnancy and subsequent changes in the newborn epigenome. While this finding warrants further investigation, it is a point that should be considered in any study assessing newborn DNA methylation studies obtained during this period, even in otherwise healthy pregnancies

Newborn screening protocols and positive predictive value for congenital adrenal hyperplasia vary across the United States

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). Newborn screening for congenital adrenal hyperplasia (CAH) caused by 21-hydroxylase deficiency is mandated throughout the US. Filter paper blood specimens are assayed for 17-hydroxyprogesterone (17OHP). Prematurity, low birth weight, or critical illness cause falsely elevated results. The purpose of this report is to highlight differences in protocols among US state laboratories. We circulated a survey to state laboratory directors requesting qualitative and quantitative information about individual screening programs. Qualitative and quantitative information provided by 17 state programs were available for analysis. Disease prevalence ranged from 1:9941 to 1:28,661 live births. Four state laboratories mandated a second screen regardless of the initial screening results; most others did so for infants in intensive care units. All but one program utilized birthweight cut-points, but cutoffs varied widely: 17OHP values of 25 to 75 ng/mL for birthweights \u3e2250–2500 g. The positive predictive values for normal birthweight infants varied from 0.7% to 50%, with the highest predictive values based in two of the states with a mandatory second screen. Data were unavailable for negative predictive values. These data imply differences in sensitivity and specificity in CAH screening in the US. Standardization of newborn screening protocols could improve the positive predictive value

Identification of novel candidate genes for 46,XY disorders of sex development (DSD) using a C57BL/6J-Y POS mouse model

Abstract Background Disorders of sex development (DSD) have an estimated frequency of 0.5% of live births encompassing a variety of urogenital anomalies ranging from mild hypospadias to a discrepancy between sex chromosomes and external genitalia. In order to identify the underlying genetic etiology, we had performed exome sequencing in a subset of DSD cases with 46,XY karyotype and were able to identify the causative genetic variant in 35% of cases. While the genetic etiology was not ascertained in more than half of the cases, a large number of variants of unknown clinical significance (VUS) were identified in those exomes. Methods To investigate the relevance of these VUS in regards to the patient’s phenotype, we utilized a mouse model in which the presence of a Y chromosome from the poschiavinus strain (Y POS ) on a C57BL/6J (B6) background results in XY undervirilization and sex reversal, a phenotype characteristic to a large subset of human 46,XY DSD cases. We assessed gene expression differences between B6-Y B6 and undervirilized B6-Y POS gonads at E11.5 and identified 515 differentially expressed genes (308 underexpressed and 207 overexpressed in B6-Y POS males). Results We identified 15 novel candidate genes potentially involved in 46,XY DSD pathogenesis by filtering the list of human VUS-carrying genes provided by exome sequencing with the list of differentially expressed genes from B6-Y POS mouse model. Additionally, we identified that 7 of the 15 candidate genes were significantly underexpressed in the XY gonads of mice with suppressed Sox9 expression in Sertoli cells suggesting that some of the candidate genes may be downstream of a well-known sex determining gene, Sox9. Conclusion The use of a DSD-specific animal model improves variant interpretation by correlating human sequence variants with transcriptome variation

Beyond the exome: What\u27s next in diagnostic testing for Mendelian conditions

Despite advances in clinical genetic testing, including the introduction of exome sequencing (ES), more than 50% of individuals with a suspected Mendelian condition lack a precise molecular diagnosis. Clinical evaluation is increasingly undertaken by specialists outside of clinical genetics, often occurring in a tiered fashion and typically ending after ES. The current diagnostic rate reflects multiple factors, including technical limitations, incomplete understanding of variant pathogenicity, missing genotype-phenotype associations, complex gene-environment interactions, and reporting differences between clinical labs. Maintaining a clear understanding of the rapidly evolving landscape of diagnostic tests beyond ES, and their limitations, presents a challenge for non-genetics professionals. Newer tests, such as short-read genome or RNA sequencing, can be challenging to order, and emerging technologies, such as optical genome mapping and long-read DNA sequencing, are not available clinically. Furthermore, there is no clear guidance on the next best steps after inconclusive evaluation. Here, we review why a clinical genetic evaluation may be negative, discuss questions to be asked in this setting, and provide a framework for further investigation, including the advantages and disadvantages of new approaches that are nascent in the clinical sphere. We present a guide for the next best steps after inconclusive molecular testing based upon phenotype and prior evaluation, including when to consider referral to research consortia focused on elucidating the underlying cause of rare unsolved genetic disorders

Mutations in the PCNA-binding domain of CDKN1C cause IMAGe syndrome.

IMAGe syndrome (intrauterine growth restriction, metaphyseal dysplasia, adrenal hypoplasia congenita and genital anomalies) is an undergrowth developmental disorder with life-threatening consequences. An identity-by-descent analysis in a family with IMAGe syndrome identified a 17.2-Mb locus on chromosome 11p15 that segregated in the affected family members. Targeted exon array capture of the disease locus, followed by high-throughput genomic sequencing and validation by dideoxy sequencing, identified missense mutations in the imprinted gene CDKN1C (also known as P57KIP2) in two familial and four unrelated patients. A familial analysis showed an imprinted mode of inheritance in which only maternal transmission of the mutation resulted in IMAGe syndrome. CDKN1C inhibits cell-cycle progression, and we found that targeted expression of IMAGe-associated CDKN1C mutations in Drosophila caused severe eye growth defects compared to wild-type CDKN1C, suggesting a gain-of-function mechanism. All IMAGe-associated mutations clustered in the PCNA-binding domain of CDKN1C and resulted in loss of PCNA binding, distinguishing them from the mutations of CDKN1C that cause Beckwith-Wiedemann syndrome, an overgrowth syndrome

Precocious Puberty in a Boy With Bilateral Leydig Cell Tumors due to a Somatic Gain-of-Function Variant

Context: Autosomal dominant and rarely de novo gain-of-function variants in the gene are associated with precocious male puberty, while somatic variants have been found in isolated Leydig cell adenomas and Leydig cell hyperplasia. Bilateral diffuse Leydig cell tumor formation in peripheral precocious male puberty has not been reported. Case Description: We present a boy with gonadotropin-independent precocious puberty and rapid virilization beginning in infancy resistant to standard therapy. Treatment with abiraterone in addition to letrozole and bicalutamide proved effective. Bilateral diffuse Leydig cell tumors were identified at age 5 years. Results: Whole-genome sequencing of tumor and blood samples was performed. The patient was confirmed to have bilateral, diffuse Leydig cell tumors harboring the somatic, gain-of-function p.Asp578His variant in the gene. Digital droplet polymerase chain reaction of the variant performed in tumor and blood samples detected low levels of this same variant in the blood. Conclusion: We report a young boy with severe gonadotropin-independent precocious puberty beginning in infancy who developed bilateral diffuse Leydig cell tumors at age 5 years due to a somatic gain-of-function p.Asp578His variant in . The gain-of-function nature of the variant and the developmental timing of the somatic mutation likely play a role in the risk of tumor formation. Abiraterone (a CYP17A1 inhibitor), in combination with an antiandrogen, aromatase inhibitor, and glucocorticoid, appears to be an effective therapy for severe peripheral precocious puberty in boys