Identifying phenotypic expansions for congenital diaphragmatic hernia plus (CDH+) using DECIPHER data.

Congenital diaphragmatic hernia (CDH) can occur in isolation or in conjunction with other birth defects (CDH+). A molecular etiology can only be identified in a subset of CDH cases. This is due, in part, to an incomplete understanding of the genes that contribute to diaphragm development. Here, we used clinical and molecular data from 36 individuals with CDH+ who are cataloged in the DECIPHER database to identify genes that may play a role in diaphragm development and to discover new phenotypic expansions. Among this group, we identified individuals who carried putatively deleterious sequence or copy number variants affecting CREBBP, SMARCA4, UBA2, and USP9X. The role of these genes in diaphragm development was supported by their expression in the developing mouse diaphragm, their similarity to known CDH genes using data from a previously published and validated machine learning algorithm, and/or the presence of CDH in other individuals with their associated genetic disorders. Our results demonstrate how data from DECIPHER, and other public databases, can be used to identify new phenotypic expansions and suggest that CREBBP, SMARCA4, UBA2, and USP9X play a role in diaphragm development.

Implementing a rapid fetal exome sequencing service: What do parents and health professionals think?

OBJECTIVES: Prenatal exome sequencing (pES) for the diagnosis of fetal abnormalities is being introduced more widely in clinical practice. Here we explore parents' and professionals' views and experiences of pES, to identify perceived benefits, concerns, and support needs. METHODS: Semi-structured interviews were conducted with 11 parents and 20 health professionals (fetal medicine and clinical genetics) with experience of rapid pES prior to implementation in the English National Health Service. Interviews were transcribed verbatim and analysed thematically. RESULTS: Parents and professionals were largely positive about pES, emphasising clinical and psychosocial benefits of a timely, definitive diagnosis in pregnancy. Concerns included parental anxiety related to the timing of pES results or uncertain findings, a need for guidelines for case selection and reporting, and ensuring sufficient capacity for counselling, phenotyping and variant interpretation. Professionals were concerned non-genetics professionals may not be equipped to counsel parents on the complexities of pES. CONCLUSION: These findings highlight important issues for clinical implementation of pES. Expert counselling is required to enable parents to make informed decisions during a stressful time. To achieve this, professionals need further education and training, and fetal medicine and genetics services must work closely together to ensure parental understanding and appropriate support.

Evaluating the Feasibility of Screening Relatives of Patients Affected by Nonsyndromic Thoracic Aortic Diseases: The REST Study.

Background Diseases of the thoracic aorta are characterized by a familial etiology in up to 30% of the cases. Nonsyndromic thoracic aorta diseases (NS-TADs) lack overt clinical signs and systemic features, which hinder early detection and prompt surgical intervention. We hypothesize that tailored genetic testing and imaging of first-degree and second-degree relatives of patients affected by NS-TADs may enable early diagnosis and allow appropriate surveillance or intervention. Methods and Results We conducted a feasibility study involving probands affected by familial or sporadic NS-TADs who had undergone surgery, which also offered screening to their relatives. Each participant underwent a combined imaging (echocardiogram and magnetic resonance imaging) and genetic (whole exome sequencing) evaluation, together with physical examination and psychological assessment. The study population included 16 probands (8 sporadic, 8 familial) and 54 relatives (41 first-degree and 13 second-degree relatives) with median age 48 years (range: 18-85 years). No syndromic physical features were observed. Imaging revealed mild-to-moderate aortic dilation in 24% of relatives. A genetic variant of uncertain significance was identified in 3 families. Imaging, further phenotyping, or a form of secondary prevention was indicated in 68% of the relatives in the familial group and 54% in the sporadic group. No participants fulfilled criteria for aortic surgery. No differences between baseline and 3-month follow-up scores for depression, anxiety, and self-reported quality of life were observed. Conclusions In NS-TADs, imaging tests, genetic counseling, and family screening yielded positive results in up to 1 out of 4 screened relatives, including those in the sporadic NS-TAD group. Registration URL: https://www.clinicaltrials.gov; Unique identifier: NCT03861741.

Pathogenic variants causing ABL1 malformation syndrome cluster in a myristoyl-binding pocket and increase tyrosine kinase activity.

ABL1 is a proto-oncogene encoding a nonreceptor tyrosine kinase, best known in the somatic BCR-ABL fusion gene associated with chronic myeloid leukaemia. Recently, germline missense variants in ABL1 have been found to cause an autosomal dominant developmental syndrome with congenital heart disease, skeletal malformations and characteristic facies. Here, we describe a series of six new unrelated individuals with heterozygous missense variants in ABL1 (including four novel variants) identified via whole exome sequencing. All the affected individuals in this series recapitulate the phenotype of the ABL1 developmental syndrome and additionally we affirm that hearing impairment is a common feature of the condition. Four of the variants cluster in the myristoyl-binding pocket of ABL1, a region critical for auto-inhibitory regulation of the kinase domain. Bio-informatic analysis of transcript-wide conservation and germline/somatic variation reveals that this pocket region is subject to high missense constraint and evolutionary conservation. Functional work to investigate ABL1 kinase activity in vitro by transient transfection of HEK293T cells with variant ABL1 plasmid constructs revealed increased phosphorylation of ABL1-specific substrates compared to wild-type. The increased tyrosine kinase activity was suppressed by imatinib treatment. This case series of six new patients with germline heterozygous ABL1 missense variants further delineates the phenotypic spectrum of this condition and recognises microcephaly as a common finding. Our analysis supports an ABL1 gain-of-function mechanism due to loss of auto-inhibition, and demonstrates the potential for pharmacological inhibition using imatinib.

BICRA, a SWI/SNF Complex Member, Is Associated with BAF-Disorder Related Phenotypes in Humans and Model Organisms.

SWI/SNF-related intellectual disability disorders (SSRIDDs) are rare neurodevelopmental disorders characterized by developmental disability, coarse facial features, and fifth digit/nail hypoplasia that are caused by pathogenic variants in genes that encode for members of the SWI/SNF (or BAF) family of chromatin remodeling complexes. We have identified 12 individuals with rare variants (10 loss-of-function, 2 missense) in the BICRA (BRD4 interacting chromatin remodeling complex-associated protein) gene, also known as GLTSCR1, which encodes a subunit of the non-canonical BAF (ncBAF) complex. These individuals exhibited neurodevelopmental phenotypes that include developmental delay, intellectual disability, autism spectrum disorder, and behavioral abnormalities as well as dysmorphic features. Notably, the majority of individuals lack the fifth digit/nail hypoplasia phenotype, a hallmark of most SSRIDDs. To confirm the role of BICRA in the development of these phenotypes, we performed functional characterization of the zebrafish and Drosophila orthologs of BICRA. In zebrafish, a mutation of bicra that mimics one of the loss-of-function variants leads to craniofacial defects possibly akin to the dysmorphic facial features seen in individuals harboring putatively pathogenic BICRA variants. We further show that Bicra physically binds to other non-canonical ncBAF complex members, including the BRD9/7 ortholog, CG7154, and is the defining member of the ncBAF complex in flies. Like other SWI/SNF complex members, loss of Bicra function in flies acts as a dominant enhancer of position effect variegation but in a more context-specific manner. We conclude that haploinsufficiency of BICRA leads to a unique SSRIDD in humans whose phenotypes overlap with those previously reported.

Exome sequencing in patients with antiepileptic drug exposure and complex phenotypes.

INTRODUCTION: Fetal anticonvulsant syndrome (FACS) describes the pattern of physical and developmental problems seen in those children exposed to certain antiepileptic drugs (AEDs) in utero. The diagnosis of FACS is a clinical one and so excluding alternative diagnoses such as genetic disorders is essential. METHODS: We reviewed the pathogenicity of reported variants identified on exome sequencing in the Deciphering Developmental Disorders (DDD) Study in 42 children exposed to AEDs in utero, but where a diagnosis other than FACS was suspected. In addition, we analysed chromosome microarray data from 10 patients with FACS seen in a Regional Genetics Service. RESULTS: Seven children (17%) from the DDD Study had a copy number variant or pathogenic variant in a developmental disorder gene which was considered to explain or partially explain their phenotype. Across the AED exposure types, variants were found in 2/15 (13%) valproate exposed cases and 3/14 (21%) carbamazepine exposed cases. No pathogenic copy number variants were identified in our local sample (n=10). CONCLUSIONS: This study is the first of its kind to analyse the exomes of children with developmental disorders who were exposed to AEDs in utero. Though we acknowledge that the results are subject to bias, a significant number of children were identified with alternate diagnoses which had an impact on counselling and management. We suggest that consideration is given to performing whole exome sequencing as part of the diagnostic work-up for children exposed to AEDs in utero.

Diagnosis of lethal or prenatal-onset autosomal recessive disorders by parental exome sequencing.

OBJECTIVE: Rare genetic disorders resulting in prenatal or neonatal death are genetically heterogeneous, but testing is often limited by the availability of fetal DNA, leaving couples without a potential prenatal test for future pregnancies. We describe our novel strategy of exome sequencing parental DNA samples to diagnose recessive monogenic disorders in an audit of the first 50 couples referred. METHOD: Exome sequencing was carried out in a consecutive series of 50 couples who had 1 or more pregnancies affected with a lethal or prenatal-onset disorder. In all cases, there was insufficient DNA for exome sequencing of the affected fetus. Heterozygous rare variants (MAF < 0.001) in the same gene in both parents were selected for analysis. Likely, disease-causing variants were tested in fetal DNA to confirm co-segregation. RESULTS: Parental exome analysis identified heterozygous pathogenic (or likely pathogenic) variants in 24 different genes in 26/50 couples (52%). Where 2 or more fetuses were affected, a genetic diagnosis was obtained in 18/29 cases (62%). In most cases, the clinical features were typical of the disorder, but in others, they result from a hypomorphic variant or represent the most severe form of a variable phenotypic spectrum. CONCLUSION: We conclude that exome sequencing of parental samples is a powerful strategy with high clinical utility for the genetic diagnosis of lethal or prenatal-onset recessive disorders. © 2017 The Authors Prenatal Diagnosis published by John Wiley & Sons Ltd.

A cohort of 17 patients with kyphoscoliotic Ehlers-Danlos syndrome caused by biallelic mutations in FKBP14: expansion of the clinical and mutational spectrum and description of the natural history.

PurposeIn 2012 we reported in six individuals a clinical condition almost indistinguishable from PLOD1-kyphoscoliotic Ehlers-Danlos syndrome (PLOD1-kEDS), caused by biallelic mutations in FKBP14, and characterized by progressive kyphoscoliosis, myopathy, and hearing loss in addition to connective tissue abnormalities such as joint hypermobility and hyperelastic skin. FKBP14 is an ER-resident protein belonging to the family of FK506-binding peptidyl-prolyl cis-trans isomerases (PPIases); it catalyzes the folding of type III collagen and interacts with type III, type VI, and type X collagens. Only nine affected individuals have been reported to date.MethodsWe report on a cohort of 17 individuals with FKBP14-kEDS and the follow-up of three previously reported patients, and provide an extensive overview of the disorder and its natural history based on clinical, biochemical, and molecular genetics data.ResultsBased on the frequency of the clinical features of 23 patients from the present and previous cohorts, we define major and minor features of FKBP14-kEDS. We show that myopathy is confirmed by histology and muscle imaging only in some patients, and that hearing impairment is predominantly sensorineural and may not be present in all individuals.ConclusionOur data further support the extensive clinical overlap with PLOD1-kEDS and show that vascular complications are rare manifestations of FKBP14-kEDS.

WAC loss-of-function mutations cause a recognisable syndrome characterised by dysmorphic features, developmental delay and hypotonia and recapitulate 10p11.23 microdeletion syndrome.

BACKGROUND: Rare de novo mutations have been implicated as a significant cause of idiopathic intellectual disability. Large deletions encompassing 10p11.23 have been implicated in developmental delay, behavioural abnormalities and dysmorphic features, but the genotype-phenotype correlation was not delineated. Mutations in WAC have been recently reported in large screening cohorts of patients with intellectual disability or autism, but no full phenotypic characterisation was described. METHODS: Clinical and molecular characterisation of six patients with loss-of-function WAC mutations identified by whole exome sequencing was performed. Clinical data were obtained by retrospective chart review, parental interviews, direct patient interaction and formal neuropsychological evaluation. RESULTS: Five heterozygous de novo WAC mutations were identified in six patients. Three of the mutations were nonsense, and two were frameshift; all are predicted to cause loss of function either through nonsense-mediated mRNA decay or protein truncation. Clinical findings included developmental delay (6/6), hypotonia (6/6), behavioural problems (5/6), eye abnormalities (5/6), constipation (5/6), feeding difficulties (4/6), seizures (2/6) and sleep problems (2/6). All patients exhibited common dysmorphic features, including broad/prominent forehead, synophrys and/or bushy eyebrows, depressed nasal bridge and bulbous nasal tip. Posteriorly rotated ears, hirsutism, deep-set eyes, thin upper lip, inverted nipples, hearing loss and branchial cleft anomalies were also noted. CONCLUSIONS: Our case series show that loss-of-function mutations in WAC cause a recognisable genetic syndrome characterised by a neurocognitive phenotype and facial dysmorphism. Our data highly suggest that WAC haploinsufficiency is responsible for most of the phenotypic features associated with deletions encompassing 10p11.23.

Mainzer-Saldino syndrome is a ciliopathy caused by IFT140 mutations.

Mainzer-Saldino syndrome (MSS) is a rare disorder characterized by phalangeal cone-shaped epiphyses, chronic renal failure, and early-onset, severe retinal dystrophy. Through a combination of ciliome resequencing and Sanger sequencing, we identified IFT140 mutations in six MSS families and in a family with the clinically overlapping Jeune syndrome. IFT140 is one of the six currently known components of the intraflagellar transport complex A (IFT-A) that regulates retrograde protein transport in ciliated cells. Ciliary abundance and localization of anterograde IFTs were altered in fibroblasts of affected individuals, a result that supports the pivotal role of IFT140 in proper development and function of ciliated cells.

Heterozygous missense mutations in SMARCA2 cause Nicolaides-Baraitser syndrome.

Nicolaides-Baraitser syndrome (NBS) is characterized by sparse hair, distinctive facial morphology, distal-limb anomalies and intellectual disability. We sequenced the exomes of ten individuals with NBS and identified heterozygous variants in SMARCA2 in eight of them. Extended molecular screening identified nonsynonymous SMARCA2 mutations in 36 of 44 individuals with NBS; these mutations were confirmed to be de novo when parental samples were available. SMARCA2 encodes the core catalytic unit of the SWI/SNF ATP-dependent chromatin remodeling complex that is involved in the regulation of gene transcription. The mutations cluster within sequences that encode ultra-conserved motifs in the catalytic ATPase region of the protein. These alterations likely do not impair SWI/SNF complex assembly but may be associated with disrupted ATPase activity. The identification of SMARCA2 mutations in humans provides insight into the function of the Snf2 helicase family.

Mutations in CEP57 cause mosaic variegated aneuploidy syndrome.

Using exome sequencing and a variant prioritization strategy that focuses on loss-of-function variants, we identified biallelic, loss-of-function CEP57 mutations as a cause of constitutional mosaic aneuploidies. CEP57 is a centrosomal protein and is involved in nucleating and stabilizing microtubules. Our findings indicate that these and/or additional functions of CEP57 are crucial for maintaining correct chromosomal number during cell division.

Nicolaides-Baraitser syndrome: Delineation of the phenotype.

Nicolaides-Baraitser syndrome (NBS) is an infrequently described condition, thus far reported in five cases. In order to delineate the phenotype and its natural history in more detail, we gathered data on 18 hitherto unreported patients through a multi-center collaborative study, and follow-up data of the earlier reported patients. A detailed comparison of the 23 patients is provided. NBS is a distinct and recognizable entity, and probably has been underdiagnosed until now. Main clinical features are severe mental retardation with absent or limited speech, seizures, short stature, sparse hair, typical facial characteristics, brachydactyly, prominent finger joints and broad distal phalanges. Some of the features are progressive with time. The main differential diagnosis is Coffin-Siris syndrome. There is no important gender difference in occurrence and frequency of the syndrome, and all cases have been sporadic thus far. Microarray analysis performed in 14 of the patients gave normal results. Except for the progressive nature there are no clues to the cause.

Microcephaly-lymphoedema-chorioretinal dysplasia: three cases to delineate the facial phenotype and review of the literature.

Microcephaly-lymphoedema-chorioretinal dysplasia (MIM 152950) has been described as a distinct clinical entity. The mode of inheritance is uncertain, but male to male transmission has been observed supporting autosomal dominant inheritance. A characteristic facial phenotype has recently been suggested. We report three isolated male patients with this condition who have all of the major features and share a distinct facial appearance with upslanting palpebral fissures, a broad nose with rounded tip, anteverted nares, long philtrum with thin upper lip, pointed chin and prominent ears. The clinical features in our patients support the hypothesis of a characteristic facial phenotype in this syndrome.

Constitutional aneuploidy and cancer predisposition caused by biallelic mutations in BUB1B.

Mosaic variegated aneuploidy is a rare recessive condition characterized by growth retardation, microcephaly, childhood cancer and constitutional mosaicism for chromosomal gains and losses. In five families with mosaic variegated aneuploidy, including two with embryonal rhabdomyosarcoma, we identified truncating and missense mutations of BUB1B, which encodes BUBR1, a key protein in the mitotic spindle checkpoint. These data are the first to relate germline mutations in a spindle checkpoint gene with a human disorder and strongly support a causal link between aneuploidy and cancer development.