Germline pathogenic variants in chromatin-modifying enzymes are a common cause of pediatric developmental disorders. These enzymes catalyze reactions that regulate epigenetic inheritance via histone ...post-translational modifications and DNA methylation. Cytosine methylation (5-methylcytosine 5mC) of DNA is the quintessential epigenetic mark, yet no human Mendelian disorder of DNA demethylation has yet been delineated. Here, we describe in detail a Mendelian disorder caused by the disruption of DNA demethylation. TET3 is a methylcytosine dioxygenase that initiates DNA demethylation during early zygote formation, embryogenesis, and neuronal differentiation and is intolerant to haploinsufficiency in mice and humans. We identify and characterize 11 cases of human TET3 deficiency in eight families with the common phenotypic features of intellectual disability and/or global developmental delay; hypotonia; autistic traits; movement disorders; growth abnormalities; and facial dysmorphism. Mono-allelic frameshift and nonsense variants in TET3 occur throughout the coding region. Mono-allelic and bi-allelic missense variants localize to conserved residues; all but one such variant occur within the catalytic domain, and most display hypomorphic function in an assay of catalytic activity. TET3 deficiency and other Mendelian disorders of the epigenetic machinery show substantial phenotypic overlap, including features of intellectual disability and abnormal growth, underscoring shared disease mechanisms.
Mutations that perturb normal pre-mRNA splicing are significant contributors to human disease. We used exome sequencing data from 7833 probands with developmental disorders (DDs) and their unaffected ...parents, as well as more than 60,000 aggregated exomes from the Exome Aggregation Consortium, to investigate selection around the splice sites and quantify the contribution of splicing mutations to DDs. Patterns of purifying selection, a deficit of variants in highly constrained genes in healthy subjects, and excess de novo mutations in patients highlighted particular positions within and around the consensus splice site of greater functional relevance. By using mutational burden analyses in this large cohort of proband-parent trios, we could estimate in an unbiased manner the relative contributions of mutations at canonical dinucleotides (73%) and flanking noncanonical positions (27%), and calculate the positive predictive value of pathogenicity for different classes of mutations. We identified 18 patients with likely diagnostic de novo mutations in dominant DD-associated genes at noncanonical positions in splice sites. We estimate 35%-40% of pathogenic variants in noncanonical splice site positions are missing from public databases.
Thirty percent of all inherited retinal disease (IRD) is accounted for by conditions with extra-ocular features. This study aimed to establish the genetic diagnostic pick-up rate for IRD patients ...with one or more extra-ocular features undergoing panel-based screening in a clinical setting. One hundred and six participants, tested on a gene panel which contained both isolated and syndromic IRD genes, were retrospectively ascertained from the Manchester Genomic Diagnostics Laboratory database spanning 6 years (2012-2017). Phenotypic features were extracted from the clinical notes and classified according to Human Phenotype Ontology; all identified genetic variants were interpreted in accordance to the American College of Medical Genetics and Genomics guidelines. Overall, 49% (n = 52) of patients received a probable genetic diagnosis. A further 6% (n = 6) had a single disease-associated variant in an autosomal recessive disease-relevant gene. Fifty-two percent (n = 55) of patients had a clinical diagnosis at the time of testing. Of these, 71% (n = 39) received a probable genetic diagnosis. By contrast, for those without a provisional clinical diagnosis (n = 51), only 25% (n = 13) received a probable genetic diagnosis. The clinical diagnosis of Usher (n = 33) and Bardet-Biedl syndrome (n = 10) was confirmed in 67% (n = 22) and 80% (n = 8), respectively. The testing diagnostic rate in patients with clinically diagnosed multisystemic IRD conditions was significantly higher than those without one (71% versus 25%; p value < 0.001). The lower pick-up rate in patients without a clinical diagnosis suggests that panel-based approaches are unlikely to be the most effective means of achieving a molecular diagnosis for this group. Here, we suggest that genome-wide approaches (whole exome or genome) are more appropriate.
A key property to consider in all genetic tests is clinical utility, the ability of the test to influence patient management and health outcomes. Here we assess the current clinical utility of ...genetic testing in diverse pediatric inherited eye disorders (IEDs).
Two hundred one unrelated children (0–5 years old) with IEDs were ascertained through the database of the North West Genomic Laboratory Hub, Manchester, UK. The cohort was collected over a 7-year period (2011–2018) and included 74 children with bilateral cataracts, 8 with bilateral ectopia lentis, 28 with bilateral anterior segment dysgenesis, 32 with albinism, and 59 with inherited retinal disorders. All participants underwent panel-based genetic testing.
The diagnostic yield of genetic testing for the cohort was 64% (ranging from 39% to 91% depending on the condition). The test result led to altered management (including preventing additional investigations or resulting in the introduction of personalized surveillance measures) in 33% of probands (75% for ectopia lentis, 50% for cataracts, 33% for inherited retinal disorders, 7% for anterior segment dysgenesis, 3% for albinism).
Genetic testing helped identify an etiological diagnosis in the majority of preschool children with IEDs. This prevented additional unnecessary testing and provided the opportunity for anticipatory guidance in significant subsets of patients.
Variants in the chromodomain helicase DNA‐binding protein 8 (CHD8) have been associated with intellectual disability (ID), autism spectrum disorders (ASDs) and overgrowth and CHD8 is one of the ...causative genes for OGID (overgrowth and ID). We investigated 25 individuals with CHD8 protein truncating variants (PTVs), including 10 previously unreported patients and found a male to female ratio of 2.7:1 (19:7) and a pattern of common features: macrocephaly (62.5%), tall stature (47%), developmental delay and/or intellectual disability (81%), ASDs (84%), sleep difficulties (50%), gastrointestinal problems (40%), and distinct facial features. Most of the individuals in this cohort had moderate‐to‐severe ID, some had regression of speech (37%), seizures (27%) and hypotonia (27%) and two individuals were non‐ambulant. Our study shows that haploinsufficiency of CHD8 is associated with a distinctive OGID syndrome with pronounced autistic traits and supports a sex‐dependent penetrance of CHD8 PTVs in humans.
Variants in the chromodomain helicase DNA‐binding protein 8 (CHD8) have been associated with intellectual disability (ID), autism spectrum disorders (ASD) and overgrowth and CHD8 is one of the causative genes for OGID (overgrowth and ID). We investigated 25 individuals with CHD8 protein truncating variants (PTVs), including 10 previously unreported patients and found a male to female ratio of 2.7:1 (19:7) and a pattern of common features: macrocephaly (62.5%), tall stature (47%), developmental delay and/or intellectual disability (DD/ID, 81%), autism spectrum disorders (ASD, 84%), sleep difficulties (50%), gastrointestinal problems (40%), and distinct facial features. Most of the individuals in this cohort had moderate‐to‐severe intellectual disability, some had regression of speech (37%), seizures (27%) and hypotonia (27%) and two individuals were also non‐ambulant. Our study shows that haploinsufficiency of CHD8 is associated with a distinctive OGID syndrome with pronounced autistic traits and supports a sex‐dependent penetrance of CHD8 PTVs in humans.
The craniofacial developmental disorder Burn-McKeown Syndrome (BMKS) is caused by biallelic variants in the pre-messenger RNA splicing factor gene TXNL4A/DIB1. The majority of affected individuals ...with BMKS have a 34 base pair deletion in the promoter region of one allele of TXNL4A combined with a loss-of-function variant on the other allele, resulting in reduced TXNL4A expression. However, it is unclear how reduced expression of this ubiquitously expressed spliceosome protein results in craniofacial defects during development. Here we reprogrammed peripheral mononuclear blood cells from a BMKS patient and her unaffected mother into induced pluripotent stem cells (iPSCs) and differentiated the iPSCs into induced neural crest cells (iNCCs), the key cell type required for correct craniofacial development. BMKS patient-derived iPSCs proliferated more slowly than both mother- and unrelated control-derived iPSCs, and RNA-Seq analysis revealed significant differences in gene expression and alternative splicing. Patient iPSCs displayed defective differentiation into iNCCs compared to maternal and unrelated control iPSCs, in particular a delay in undergoing an epithelial-to-mesenchymal transition (EMT). RNA-Seq analysis of differentiated iNCCs revealed widespread gene expression changes and mis-splicing in genes relevant to craniofacial and embryonic development that highlight a dampened response to WNT signalling, the key pathway activated during iNCC differentiation. Furthermore, we identified the mis-splicing of TCF7L2 exon 4, a key gene in the WNT pathway, as a potential cause of the downregulated WNT response in patient cells. Additionally, mis-spliced genes shared common sequence properties such as length, branch point to 3' splice site (BPS-3'SS) distance and splice site strengths, suggesting that splicing of particular subsets of genes is particularly sensitive to changes in TXNL4A expression. Together, these data provide the first insight into how reduced TXNL4A expression in BMKS patients might compromise splicing and NCC function, resulting in defective craniofacial development in the embryo.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
To assess the clinical usefulness of genetic testing in a pediatric population with inherited retinal disease (IRD).
Single-center retrospective case series.
Eighty-five unrelated children with a ...diagnosis of isolated or syndromic IRD who were referred for clinical genetic testing between January 2014 and July 2016.
Participants underwent a detailed ophthalmic examination, accompanied by electrodiagnostic testing (EDT) and dysmorphologic assessment where appropriate. Ocular and extraocular features were recorded using Human Phenotype Ontology terms. Subsequently, multigene panel testing (105 or 177 IRD-associated genes) was performed in an accredited diagnostic laboratory, followed by clinical variant interpretation.
Diagnostic yield and clinical usefulness of genetic testing.
Overall, 78.8% of patients (n = 67) received a probable molecular diagnosis; 7.5% (n = 5) of these had autosomal dominant disease, 25.4% (n = 17) had X-linked disease, and 67.2% (n = 45) had autosomal recessive disease. In a further 5.9% of patients (n = 5), a single heterozygous ABCA4 variant was identified; all these participants had a spectrum of clinical features consistent with ABCA4 retinopathy. Most participants (84.7%; n = 72) had undergone EDT and 81.9% (n = 59) of these patients received a probable molecular diagnosis. The genes most frequently mutated in the present cohort were CACNA1F and ABCA4, accounting for 14.9% (n = 10) and 11.9% (n = 8) of diagnoses respectively. Notably, in many cases, genetic testing helped to distinguish stationary from progressive IRD subtypes and to establish a precise diagnosis in a timely fashion.
Multigene panel testing pointed to a molecular diagnosis in 84.7% of children with IRD. The diagnostic yield in the study population was significantly higher compared with that in previously reported unselected IRD cohorts. Approaches similar to the one described herein are expected to become a standard component of care in pediatric ophthalmology. We propose the introduction of genetic testing early in the diagnostic pathway in children with clinical and/or electrophysiologic findings, suggestive of IRD.
The European Reference Networks, ERNs, are virtual networks for healthcare providers across Europe to collaborate and share expertise on complex or rare diseases and conditions. As part of the ERNs, ...the Clinical Patient Management System, CPMS, a secure digital platform, was developed to allow and facilitate web-based, clinical consultations between submitting clinicians and relevant international experts. The European Reference Network on Intellectual Disability, TeleHealth and Congenital Anomalies, ERN ITHACA, was formed to harness the clinical and diagnostic expertise in the sector of rare, multiple anomaly and/or intellectual disability syndromes, chromosome disorders and undiagnosed syndromic disorders. We present the first year results of CPMS use by ERN ITHACA as an example of a telemedicine strategy for the diagnosis and management of patients with rare developmental disorders.
ERN ITHACA ranked third in telemedicine activity amongst 24 European networks after 12 months of using the CPMS. Information about 28 very rare cases from 13 different centres across 7 countries was shared on the platform, with diagnostic or other management queries. Early interaction with patient support groups identified data protection as of primary importance in adopting digital platforms for patient diagnosis and care. The first launch of the CPMS was built to accommodate the needs of all ERNs. The ERN ITHACA telemedicine process highlighted a need to customise the CPMS with network-specific requirements. The results of this effort should enhance the CPMS utility for telemedicine services and ERN-specific care outcomes.
We present the results of a long and fruitful process of interaction between the ERN ITHACA network lead team and EU officials, software developers and members of 38 EU clinical genetics centres to organise and coordinate direct e-healthcare through a secure, digital platform. The variability of the queries in just the first 28 cases submitted to the ERN ITHACA CPMS is a fair representation of the complexity and rarity of the patients referred, but also proof of the sophisticated and variable service that could be provided through a structured telemedicine approach for patients and families with rare developmental disorders. Web-based approaches are likely to result in increased accessibility to clinical genomic services.
Inherited vitreoretinopathies arise as a consequence of congenital retinal vascularisation abnormalities. They represent a phenotypically and genetically heterogeneous group of disorders that can ...have a major impact on vision. Several genes encoding proteins and effectors of the canonical Wnt/β-catenin pathway have been associated and precise diagnosis, although difficult, is essential for proper clinical management including syndrome specific management where appropriate. This work aimed to investigate the molecular basis of disease in a single proband born to consanguineous parents, who presented with microphthalmia, persistent foetal vasculature, posterior lens vacuoles, vitreoretinal dysplasia, microcephaly, hypotelorism and global developmental delay, and was registered severely visually impaired by 5 months of age.
Extensive genomic pre-screening, including microarray comparative genomic hybridisation and sequencing of a 114 gene panel associated with cataract and congenital ophthalmic disorders was conducted by an accredited clinical laboratory. Whole exome sequencing (WES) was undertaken on a research basis and in vitro TOPflash transcriptional reporter assay was utilised to assess the impact of the putative causal variant.
In the proband, WES revealed a novel, likely pathogenic homozygous mutation in the cadherin-associated protein beta-1 gene (CTNNB1), c.884C>G; p.(Ala295Gly), which encodes a co-effector molecule of the Wnt/β-catenin pathway. The proband's parents were shown to be heterozygous carriers but ophthalmic examination did not detect any abnormalities. Functional assessment of the missense variant demonstrated significant reduction of β-catenin activity.
This is the first report of a biallelic disease-causing variation in CTNNB1. We conclude that this biallelic, transcriptional inactivating mutation of CTNNB1 causes a severe, syndromic form of microphthalmia, persistent foetal vasculature and vitreoretinal dysplasia that results in serious visual loss in infancy.
CHD8 has been reported as an autism susceptibility/intellectual disability gene but emerging evidence suggests that it additionally causes an overgrowth phenotype. This study reports 27 unrelated ...patients with pathogenic or likely pathogenic CHD8 variants (25 null variants, two missense variants) and a male:female ratio of 21:6 (3.5:1, p < .01). All patients presented with intellectual disability, with 85% in the mild or moderate range, and 85% had a height and/or head circumference ≥2 standard deviations above the mean, meeting our clinical criteria for overgrowth. Behavioral problems were reported in the majority of patients (78%), with over half (56%) either formally diagnosed with an autistic spectrum disorder or described as having autistic traits. Additional clinical features included neonatal hypotonia (33%), and less frequently seizures, pes planus, scoliosis, fifth finger clinodactyly, umbilical hernia, and glabellar hemangioma (≤15% each). These results suggest that, in addition to its established link with autism and intellectual disability, CHD8 causes an overgrowth phenotype, and should be considered in the differential diagnosis of patients presenting with increased height and/or head circumference in association with intellectual disability.
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