Large-scale discovery of novel genetic causes of developmental disorders.

Despite three decades of successful, predominantly phenotype-driven discovery of the genetic causes of monogenic disorders, up to half of children with severe developmental disorders of probable genetic origin remain without a genetic diagnosis. Particularly challenging are those disorders rare enough to have eluded recognition as a discrete clinical entity, those with highly variable clinical manifestations, and those that are difficult to distinguish from other, very similar, disorders. Here we demonstrate the power of using an unbiased genotype-driven approach to identify subsets of patients with similar disorders. By studying 1,133 children with severe, undiagnosed developmental disorders, and their parents, using a combination of exome sequencing and array-based detection of chromosomal rearrangements, we discovered 12 novel genes associated with developmental disorders. These newly implicated genes increase by 10% (from 28% to 31%) the proportion of children that could be diagnosed. Clustering of missense mutations in six of these newly implicated genes suggests that normal development is being perturbed by an activating or dominant-negative mechanism. Our findings demonstrate the value of adopting a comprehensive strategy, both genome-wide and nationwide, to elucidate the underlying causes of rare genetic disorders

A New, Atypical Case of Cobalamin F Disorder Diagnosed by Whole Exome Sequencing

The Deciphering Developmental Disorders Study presents independent research commissioned by the Health Innovation Challenge Fund (HICF-1009-003), a parallel funding partnership between the Wellcome Trust and the Department of Health, and the Wellcome Trust Sanger Institute (WT098051). The views expressed in this publication are those of the author(s) and not necessarily those of the Wellcome Trust or the Department of Health. The research team acknowledges the support of the National Institute for Health Research, through the Comprehensive Clinical Research Network.Peer reviewedPublisher PD

Detection of structural mosaicism from targeted and whole-genome sequencing data.

Structural mosaic abnormalities are large post-zygotic mutations present in a subset of cells and have been implicated in developmental disorders and cancer. Such mutations have been conventionally assessed in clinical diagnostics using cytogenetic or microarray testing. Modern disease studies rely heavily on exome sequencing, yet an adequate method for the detection of structural mosaicism using targeted sequencing data is lacking. Here, we present a method, called MrMosaic, to detect structural mosaic abnormalities using deviations in allele fraction and read coverage from next-generation sequencing data. Whole-exome sequencing (WES) and whole-genome sequencing (WGS) simulations were used to calculate detection performance across a range of mosaic event sizes, types, clonalities, and sequencing depths. The tool was applied to 4911 patients with undiagnosed developmental disorders, and 11 events among nine patients were detected. For eight of these 11 events, mosaicism was observed in saliva but not blood, suggesting that assaying blood alone would miss a large fraction, possibly >50%, of mosaic diagnostic chromosomal rearrangements

Integrating population variation and protein structural analysis to improve clinical interpretation of missense variation: application to the WD40 domain

We present a generic, multidisciplinary approach for improving our understanding of novel missense variants in recently discovered disease genes exhibiting genetic heterogeneity, by combining clinical and population genetics with protein structural analysis. Using six new de novo missense diagnoses in TBL1XR1 from the Deciphering Developmental Disorders study, together with population variation data, we show that the β-propeller structure of the ubiquitous WD40 domain provides a convincing way to discriminate between pathogenic and benign variation. Children with likely pathogenic mutations in this gene have severely delayed language development, often accompanied by intellectual disability, autism, dysmorphology and gastrointestinal problems. Amino acids affected by likely pathogenic missense mutations are either crucial for the stability of the fold, forming part of a highly conserved symmetrically repeating hydrogen-bonded tetrad, or located at the top face of the β-propeller, where ‘hotspot’ residues affect the binding of β-catenin to the TBLR1 protein. In contrast, those altered by population variation are significantly less likely to be spatially clustered towards the top face or to be at buried or highly conserved residues. This result is useful not only for interpreting benign and pathogenic missense variants in this gene, but also in other WD40 domains, many of which are associated with disease

Arid1b haploinsufficient mice reveal neuropsychiatric phenotypes and reversible causes of growth impairment.

Sequencing studies have implicated haploinsufficiency of ARID1B, a SWI/SNF chromatin-remodeling subunit, in short stature (Yu et al., 2015), autism spectrum disorder (O'Roak et al., 2012), intellectual disability (Deciphering Developmental Disorders Study, 2015), and corpus callosum agenesis (Halgren et al., 2012). In addition, ARID1B is the most common cause of Coffin-Siris syndrome, a developmental delay syndrome characterized by some of the above abnormalities (Santen et al., 2012; Tsurusaki et al., 2012; Wieczorek et al., 2013). We generated Arid1b heterozygous mice, which showed social behavior impairment, altered vocalization, anxiety-like behavior, neuroanatomical abnormalities, and growth impairment. In the brain, Arid1b haploinsufficiency resulted in changes in the expression of SWI/SNF-regulated genes implicated in neuropsychiatric disorders. A focus on reversible mechanisms identified Insulin-like growth factor (IGF1) deficiency with inadequate compensation by Growth hormone-releasing hormone (GHRH) and Growth hormone (GH), underappreciated findings in ARID1B patients. Therapeutically, GH supplementation was able to correct growth retardation and muscle weakness. This model functionally validates the involvement of ARID1B in human disorders, and allows mechanistic dissection of neurodevelopmental diseases linked to chromatin-remodeling

Making new genetic diagnoses with old data:iterative reanalysis and reporting from genome-wide data in 1,133 families with developmental disorders

Purpose Given the rapid pace of discovery in rare disease genomics, it is likely that improvements in diagnostic yield can be made by systematically reanalyzing previously generated genomic sequence data in light of new knowledge. Methods We tested this hypothesis in the United Kingdom–wide Deciphering Developmental Disorders study, where in 2014 we reported a diagnostic yield of 27% through whole-exome sequencing of 1,133 children with severe developmental disorders and their parents. We reanalyzed existing data using improved variant calling methodologies, novel variant detection algorithms, updated variant annotation, evidence-based filtering strategies, and newly discovered disease-associated genes. Results We are now able to diagnose an additional 182 individuals, taking our overall diagnostic yield to 454/1,133 (40%), and another 43 (4%) have a finding of uncertain clinical significance. The majority of these new diagnoses are due to novel developmental disorder–associated genes discovered since our original publication. Conclusion This study highlights the importance of coupling large-scale research with clinical practice, and of discussing the possibility of iterative reanalysis and recontact with patients and health professionals at an early stage. We estimate that implementing parent–offspring whole-exome sequencing as a first-line diagnostic test for developmental disorders would diagnose >50% of patients.</p

Mutations in genes encoding condensin complex proteins cause microcephaly through decatenation failure at mitosis

Correction to Martin et al. available at: Genes & Development 30 (19): 2158 (http://genesdev.cshlp.org/content/31/9/953.full.pdf+html).Compaction of chromosomes is essential for accurate segregation of the genome duringmitosis. In vertebrates, two condensin complexes ensure timely chromosome condensation, sister chromatid disentanglement, and maintenance of mitotic chromosome structure. Here,we report that biallelic mutations inNCAPD2,NCAPH, orNCAPD3, encoding subunits of these complexes, cause microcephaly. In addition, hypomorphic Ncaph2 mice have significantly reduced brain size, with frequent anaphase chromatin bridge formation observed in apical neural progenitors during neurogenesis. Such DNA bridges also arise in condensin-deficient patient cells, where they are the consequence of failed sister chromatid disentanglement during chromosome compaction. This results in chromosome segregation errors, leading to micronucleus formation and increased aneuploidy in daughter cells. These findings establish “condensinopathies” as microcephalic disorders, with decatenation failure as an additional disease mechanism for microcephaly, implicating mitotic chromosome condensation as a key process ensuring mammalian cerebral cortex size.This work was supported by funding from the Medical Research Council, the Lister Institute for Preventative Medicine, and the European Research Council (ERC; 281847 to A.P.J.); a Biotechnology and Biological Sciences Research Council grant (BB/ K017632/1 to P.V); a Sir Henry Dale Fellowship (grant 102560/ Z/13/Z to A.J.W.); Medical Research Scotland (to L.S.B.); the Potentials Foundation (to C.A.W.); and the Indian Council of Medical Research (BMS 54/2/2013 to S.R.P). The Deciphering Developmental Disorders Study presents independent research commissioned by the Health Innovation Challenge Fund (grant no. HICF-1009-003), a parallel funding partnership between the Wellcome Trust and the Department of Health, and the Wellcome Trust Sanger Institute (grant no. WT098051). The views expressed here are those of the authors and not necessarily those of the Wellcome Trust or the Department of Health. The study has UK Research Ethics Committee approval (10/H0305/83) granted by the Cambridge South Research Ethics Committee, and GEN/ 284/12 granted by the Republic of Ireland. We acknowledge the support of the National Institute for Health Research through the Comprehensive Clinical Research Network

Heterozygous truncation mutations of the SMC1A gene cause a severe early onset epilepsy with cluster seizures in females: Detailed phenotyping of 10 new cases

OBJECTIVE: The phenotype of seizure clustering with febrile illnesses in infancy/early childhood is well recognized. To date the only genetic epilepsy consistently associated with this phenotype is PCDH19, an X-linked disorder restricted to females, and males with mosaicism. The SMC1A gene, which encodes a structural component of the cohesin complex is also located on the X chromosome. Missense variants and small in-frame deletions of SMC1A cause approximately 5% of Cornelia de Lange Syndrome (CdLS). Recently, protein truncating mutations in SMC1A have been reported in five females, all of whom have been affected by a drug-resistant epilepsy, and severe developmental impairment. Our objective was to further delineate the phenotype of SMC1A truncation. METHOD: Female cases with de novo truncation mutations in SMC1A were identified from the Deciphering Developmental Disorders (DDD) study (n = 8), from postmortem testing of an affected twin (n = 1), and from clinical testing with an epilepsy gene panel (n = 1). Detailed information on the phenotype in each case was obtained. RESULTS: Ten cases with heterozygous de novo mutations in the SMC1A gene are presented. All 10 mutations identified are predicted to result in premature truncation of the SMC1A protein. All cases are female, and none had a clinical diagnosis of CdLS. They presented with onset of epileptic seizures between <4 weeks and 28 months of age. In the majority of cases, a marked preponderance for seizures to occur in clusters was noted. Seizure clusters were associated with developmental regression. Moderate or severe developmental impairment was apparent in all cases. SIGNIFICANCE: Truncation mutations in SMC1A cause a severe epilepsy phenotype with cluster seizures in females. These mutations are likely to be nonviable in males

Compound heterozygous variants in NBAS as a cause of atypical osteogenesis imperfecta

Background Osteogenesis imperfecta (OI), the commonest inherited bone fragility disorder, affects 1 in 15,000 live births resulting in frequent fractures and reduced mobility, with significant impact on quality of life. Early diagnosis is important, as therapeutic advances can lead to improved clinical outcome and patient benefit. Report Whole exome sequencing in patients with OI identified, in two patients with a multi-system phenotype, compound heterozygous variants in NBAS (neuroblastoma amplified sequence). Patient 1: NBAS c.5741G > A p.(Arg1914His); c.3010C > T p.(Arg1004*) in a 10-year old boy with significant short stature, bone fragility requiring treatment with bisphosphonates, developmental delay and immunodeficiency. Patient 2: NBAS c.5741G > A p.(Arg1914His); c.2032C > T p.(Gln678*) in a 5-year old boy with similar presenting features, bone fragility, mild developmental delay, abnormal liver function tests and immunodeficiency. Discussion Homozygous missense NBAS variants cause SOPH syndrome (short stature; optic atrophy; Pelger-Huet anomaly), the same missense variant was found in our patients on one allele and a nonsense variant in the other allele. Recent literature suggests a multi-system phenotype. In this study, patient fibroblasts have shown reduced collagen expression, compared to control cells and RNAseq studies, in bone cells show that NBAS is expressed in osteoblasts and osteocytes of rodents and primates. These findings provide proof-of-concept that NBAS mutations have mechanistic effects in bone, and that NBAS variants are a novel cause of bone fragility, which is distinguishable from ‘Classical’ OI. Conclusions Here we report on variants in NBAS, as a cause of bone fragility in humans, and expand the phenotypic spectrum associated with NBAS. We explore the mechanism underlying NBAS and the striking skeletal phenotype in our patients

HACE1 deficiency causes an autosomal recessive neurodevelopmental syndrome

Background: The genetic etiology of neurodevelopmental defects is extremely diverse, and the lack of distinctive phenotypic features means that genetic criteria are often required for accurate diagnostic classification. We aimed to identify the causative genetic lesions in two families in which eight affected individuals displayed variable learning disability, spasticity and abnormal gait. Methods: Autosomal recessive inheritance was suggested by consanguinity in one family and by sibling recurrences with normal parents in the second. Autozygosity mapping and exome sequencing, respectively, were used to identify the causative gene. Results: In both families, biallelic loss-of-function mutations in HACE1 were identified. HACE1 is an E3 ubiquitin ligase that regulates the activity of cellular GTPases, including Rac1 and members of the Rab family. In the consanguineous family, a homozygous mutation p.R219* predicted a truncated protein entirely lacking its catalytic domain. In the other family, compound heterozygosity for nonsense mutation p.R748* and a 20-nt insertion interrupting the catalytic HECT domain was present; Western analysis of patient cells revealed an absence of detectable HACE1 protein. Conclusion: HACE1 mutations underlie a new autosomal recessive neurodevelopmental disorder. Previous studies have implicated HACE1 as a tumour suppressor gene; however, since cancer predisposition was not observed either in homozygous or heterozygous mutation carriers, this concept may require re-evaluation