Widening of the genetic and clinical spectrum of Lamb-Shaffer syndrome, a neurodevelopmental disorder due to SOX5 haploinsufficiency

Purpose Lamb-Shaffer syndrome (LAMSHF) is a neurodevelopmental disorder described in just over two dozen patients with heterozygous genetic alterations involving SOX5, a gene encoding a transcription factor regulating cell fate and differentiation in neurogenesis and other discrete developmental processes. The genetic alterations described so far are mainly microdeletions. The present study was aimed at increasing our understanding of LAMSHF, its clinical and genetic spectrum, and the pathophysiological mechanisms involved. Methods Clinical and genetic data were collected through GeneMatcher and clinical or genetic networks for 41 novel patients harboring various types ofSOX5 alterations. Functional consequences of selected substitutions were investigated. Results Microdeletions and truncating variants occurred throughout SOX5. In contrast, most missense variants clustered in the pivotal SOX-specific high-mobility-group domain. The latter variants prevented SOX5 from binding DNA and promoting transactivation in vitro, whereas missense variants located outside the high-mobility-group domain did not. Clinical manifestations and severity varied among patients. No clear genotype-phenotype correlations were found, except that missense variants outside the high-mobility-group domain were generally better tolerated. Conclusions This study extends the clinical and genetic spectrum associated with LAMSHF and consolidates evidence that SOX5 haploinsufficiency leads to variable degrees of intellectual disability, language delay, and other clinical features

A Solve-RD ClinVar-based reanalysis of 1522 index cases from ERN-ITHACA reveals common pitfalls and misinterpretations in exome sequencing

Purpose Within the Solve-RD project (https://solve-rd.eu/), the European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies aimed to investigate whether a reanalysis of exomes from unsolved cases based on ClinVar annotations could establish additional diagnoses. We present the results of the “ClinVar low-hanging fruit” reanalysis, reasons for the failure of previous analyses, and lessons learned. Methods Data from the first 3576 exomes (1522 probands and 2054 relatives) collected from European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies was reanalyzed by the Solve-RD consortium by evaluating for the presence of single-nucleotide variant, and small insertions and deletions already reported as (likely) pathogenic in ClinVar. Variants were filtered according to frequency, genotype, and mode of inheritance and reinterpreted. Results We identified causal variants in 59 cases (3.9%), 50 of them also raised by other approaches and 9 leading to new diagnoses, highlighting interpretation challenges: variants in genes not known to be involved in human disease at the time of the first analysis, misleading genotypes, or variants undetected by local pipelines (variants in off-target regions, low quality filters, low allelic balance, or high frequency). Conclusion The “ClinVar low-hanging fruit” analysis represents an effective, fast, and easy approach to recover causal variants from exome sequencing data, herewith contributing to the reduction of the diagnostic deadlock

Phenotypic and molecular characterization of frontonasal dysplasias

Les dysplasies frontonasales (DFN) sont un groupe de malformations rares de la face résultant d’une anomalie de développement du processus frontonasal, et se manifestant cliniquement par l’association variable d’une fente faciale médiane, d’un hypertélorisme, et d’anomalies nasales. Elles s’intègrent généralement dans un cadre syndromique, la plupart étant peu spécifiques et non caractérisées. Une douzaine d’entités ont été décrites ; les bases moléculaires sont connues pour sept d’entre elles seulement. Pour les autres entités, les hypothèses initiales étaient celles de pathologies autosomiques dominantes liées à des mutations de novo.Ce travail s’est fondé sur une cohorte de 80 patients présentant une DFN, recrutés au niveau national et international, avec l’objectif d’identifier les bases moléculaires de plusieurs entités de DFN. Une caractérisation phénotypique a d’abord été effectuée. Deux stratégies moléculaires ont ensuite été poursuivies en parallèle : une approche « phenotype-first », visant à étudier les patients classés par cohortes homogènes cliniquement, et une approche « genotype-first », visant à réaliser des études moléculaires chez des patients présentant un tableau aspécifique.Lorsque le diagnostic était celui d’un syndrome dont le gène causal était connu, nous avons réalisé un séquençage ciblé dudit gène (EFNB1, ZSWIM6). Nous avons effectué un séquençage haut-débit d’exome (SHD-E) chez 11 patients présentant un syndrome de Pai (5 en trio, 5 en solo, un en profondeur sur tissu atteint en paire), trois patients avec syndrome oculoauriculofrontonasal (en trio), et respectivement un patient avec syndrome oculocérébrocutané et syndrome de Teebi (en trio). Nous avons réalisé un séquençage haut-débit de génome (SHD-G) chez 3 patients avec un syndrome de Pai, ainsi qu’une patiente avec syndrome oculoauriculofrontonasal. Enfin, nous avons séquencé les gènes ALX1, ALX3 et ALX4 chez 13 individus avec DFN aspécifique ; réalisé un SHD-E en profondeur chez une patiente avec DFN et hypomélanose d’Ito, ainsi que trois SHD-E en trio chez des patients avec DFN aspécifique ou non classée.Nous avons réalisé un travail de caractérisation phénotypique des patients de la cohorte, décrivant les diagnostics différentiels principaux et les chevauchements phénotypiques. Nous avons décrit une nouvelle entité de DFN, identifiée chez 4 de nos patients, sans base moléculaire à l’heure actuelle. Nous avons confirmé les diagnostics cliniques lorsque le gène causal était connu, chez 5 patients. Nous avons retrouvé des variants candidats dans deux gènes de la voie du TGFβ chez quatre patients avec syndrome de Pai : un de novo dans le gène TGFBRAP1, deux de novo et un hérité d’un parent asymptomatique dans le gène PCSK7. Nous avons identifié les mutations dans TFE3 comme étant à l’origine d’un syndrome neurocutané de mosaïcisme pigmentaire chez une patiente et 6 patients additionnels. Enfin, nous avons identifié un variant dans le gène POLR2A chez un fœtus avec DFN aspécifique. Au total, 34 individus avec un syndrome connu et 34 avec une DFN aspécifique restent à l’heure actuelle sans piste moléculaire identifiée.Au total, ce travail a permis de démontrer l’intérêt d’une meilleure connaissance clinique de ces syndromes rares, pour le diagnostic et le conseil génétique. Il a permis l’individualisation et la description de deux syndromes pouvant s’accompagner d’une DFN – dont un à l’échelle moléculaire –, et la démonstration du rôle du gène TFE3 dans le développement. Enfin, des hypothèses sont émises concernant les résultats incertains et négatifs : celles d’un oligogénisme, d’une anomalie épigénétique, d’une mutation post-zygotique ou de l’influence de l’environnement.Frontonasal dysplasias (FND) are a group of rare facial malformations due to an abnormal development of the frontonasal process, clinically resulting in the variable association of median facial cleft, hypertelorism and nasal anomalies. Most of them are syndromic, but non-specific and not characterized. A dozen entities have been described; molecular bases are known for only seven of them. Regarding the other entities, the hypothesis of dominant disorders due to de novo mutations had been raised.This work was based on a cohort of 80 patients presenting with FND, nationally and internationaly recruited with the goal of identifying molecular bases of FND entities. We first phenotypically characterized the individuals. Then two molecular strategies were performed in parallel: a “phenotype-first” approach, aiming to study clinically homogeneous cohorts of patients, and a “genotype-first” approach, aiming to perform molecular studies on patients with an aspecific phenotype.When the causative gene for the disorder was known, we performed targeted sequencing of the gene (EFNB1, ZSWIM6). We performed whole-exome sequencing (WES) in 11 patients presenting with Pai syndrome (5 trio WES, 5 solo WES, one deep-sequencing pair-WES on affected tissue), 3 individuals presenting with oculoauriculofrontonasal syndrome (OAFNS) (trio WES), and respectively one patient with oculocerebrocutaneous syndrome and Teebi syndrome (both trio WES). We performed whole-genome sequencing (WGS) on 3 patients with Pai syndrome and one patient with OAFNS. Finally, we performed ALX1, ALX3 and ALX4 genes sequencing in 13 individuals with aspecific FND; deep-WES in one patient with FND and Ito hypomelanosis; as well as 3 trio WES in individuals with aspecific or non characterized FND.We achieved a phenotypic characterization of the patients from the cohort, describing major differential diagnosis and clinical overlaps. We described a new FND entity, identified in 4 individuals, with no molecular basis so far. We confirmed the clinical diagnosis when the causative gene was known, for 5 patients. We identified candidate variants in two genes from the TGFβ pathway in four patients with Pai syndrome: one de novo variant in the TGFBRAP1 gene, two de novo and one inherited from an asymptomatic parent in the PCSK7 gene. We identified mutations in TFE3 as causative for a neurocutaneous syndrome of pigmentary mosaicism in a female patient and six additional individuals. Finally, we identified a variant in the POLR2A gene in a fetus with aspecific FND. A total of 34 individuals with a known syndrome and 34 with an aspecific FND still have no identified molecular basis so far.In conclusion, this work allowed proving the importance of a better clinical knowledge of these rare disorders, in terms of diagnosis and genetics counseling. It allowed the delineation of two syndromes with FND – one at a molecular level –, and the demonstration of a role for TFE3 in development. Finally, four hypotheses are raised regarding the negative or uncertain results: oligogenism, epigenetic anomaly, post-zygotic mutation or environmental influence

Neuroblastome syndromique (revue de la littérature et présentation d une série de 49 patients)

Le neuroblastome est une tumeur maligne du système nerveux sympathique d origine embryonnaire et dérivée des crêtes neurales. Il représente la tumeur solide extra-crânienne la plus fréquente chez l enfant, avec une incidence annuelle d'environ 1/80 000 enfants de moins de 14 ans, et est par ailleurs responsable de 15% des décès pédiatriques par cancer. On observe une grande hétérogénéité clinique et pronostique. L augmentation du risque de survenue de tumeurs, notamment de neuroblastomes, chez les enfants présentant des malformations congénitales est étudiée depuis plusieurs décennies, et corrobore l hypothèse selon laquelle certaines tumeurs embryonnaires s intègrent dans une pathologie plus globale du développement. L étude de ces syndromes de prédisposition aux tumeurs de l enfant est importante pour améliorer le dépistage des sujets à risque et leur prise en charge, et également pour la compréhension des mécanismes de la tumorigenèse, ouvrant la voie à des thérapeutiques ciblées contre le neuroblastome. La première partie de ce travail est une revue de la littérature, présentant les syndromes de prédisposition au neuroblastome décrits, classés selon leur physiopathologie. La seconde partie rapporte une série de 49 patients présentant un neuroblastome et une ou plusieurs malformations associées. Enfin, la dernière partie récapitule brièvement les thérapeutiques récentes développées dans le traitement du neuroblastome. Nous insistons sur l importance d un registre national colligeant les enfants présentant l association d une tumeur et d une anomalie de développement.PARIS6-Bibl.Pitié-Salpêtrie (751132101) / SudocSudocFranceF

Missense and truncating variants in CHD5 in a dominant neurodevelopmental disorder with intellectual disability, behavioral disturbances, and epilepsy

Located in the critical 1p36 microdeletion region, the chromodomain helicase DNA-binding protein 5 (CHD5) gene encodes a subunit of the nucleosome remodeling and deacetylation (NuRD) complex required for neuronal development. Pathogenic variants in six of nine chromodomain (CHD) genes cause autosomal dominant neurodevelopmental disorders, while CHD5-related disorders are still unknown. Thanks to GeneMatcher and international collaborations, we assembled a cohort of 16 unrelated individuals harboring heterozygous CHD5 variants, all identified by exome sequencing. Twelve patients had de novo CHD5 variants, including ten missense and two splice site variants. Three familial cases had nonsense or missense variants segregating with speech delay, learning disabilities, and/or craniosynostosis. One patient carried a frameshift variant of unknown inheritance due to unavailability of the father. The most common clinical features included language deficits (81%), behavioral symptoms (69%), intellectual disability (64%), epilepsy (62%), and motor delay (56%). Epilepsy types were variable, with West syndrome observed in three patients, generalized tonic-clonic seizures in two, and other subtypes observed in one individual each. Our findings suggest that, in line with other CHD-related disorders, heterozygous CHD5 variants are associated with a variable neurodevelopmental syndrome that includes intellectual disability with speech delay, epilepsy, and behavioral problems as main features

2.5 years’ experience of GeneMatcher data-sharing: a powerful tool for identifying new genes responsible for rare diseases

International audienceExome sequencing (ES) powerfully identifies the molecular bases of heterogeneous conditions such as intellectual disability and/or multiple congenital anomalies (ID/MCA). Current ES analysis, combining diagnosis analysis restricted to disease-causing genes reported in OMIM database and subsequent research investigation extended to other genes, indicated causal and candidate genes around 40% and 10%. Nonconclusive results are frequent in such ultrarare conditions that recurrence and genotype-phenotype correlations are limited. International data-sharing permits the gathering of additional patients carrying variants in the same gene to draw definitive conclusions on their implication as disease causing. Several web-based tools have been developed and grouped in Matchmaker Exchange. In this study, we report our current experience as a regional center that has implemented ES as a first-line diagnostic test since 2013, working with a research laboratory devoted to disease gene identification

Clinical whole-exome sequencing for the diagnosis of rare disorders with congenital anomalies and/or intellectual disability: substantial interest of prospective annual reanalysis

International audiencePurposeCongenital anomalies and intellectual disability (CA/ID) are a major diagnostic challenge in medical genetics—50% of patients still have no molecular diagnosis after a long and stressful diagnostic “odyssey.” Solo clinical whole-exome sequencing (WES) was applied in our genetics center to improve diagnosis in patients with CA/ID.MethodsThis retrospective study examined 416 consecutive tests performed over 3 years to demonstrate the effectiveness of periodically reanalyzing WES data. The raw data from each nonpositive test was reanalyzed at 12 months with the most recent pipeline and in the light of new data in the literature. The results of the reanalysis for patients enrolled in the third year are not yet available.ResultsOf the 416 patients included, data for 156 without a diagnosis were reanalyzed. We obtained 24 (15.4%) additional diagnoses: 12 through the usual diagnostic process (7 new publications, 4 initially misclassified, and 1 copy-number variant), and 12 through translational research by international data sharing. The final yield of positive results was 27.9% through a strict diagnostic approach, and 2.9% through an additional research strategy.ConclusionThis article highlights the effectiveness of periodically combining diagnostic reinterpretation of clinical WES data with translational research involving data sharing for candidate genes

Impaired eIF5A function causes a Mendelian disorder that is partially rescued in model systems by spermidine

The structure of proline prevents it from adopting an optimal position for rapid protein synthesis. Poly-proline-tract (PPT) associated ribosomal stalling is resolved by highly conserved eIF5A, the only protein to contain the amino acid hypusine. We show that de novo heterozygous EIF5A variants cause a disorder characterized by variable combinations of developmental delay, microcephaly, micrognathia and dysmorphism. Yeast growth assays, polysome profiling, total/hypusinated eIF5A levels and PPT-reporters studies reveal that the variants impair eIF5A function, reduce eIF5A-ribosome interactions and impair the synthesis of PPT-containing proteins. Supplementation with 1 mM spermidine partially corrects the yeast growth defects, improves the polysome profiles and restores expression of PPT reporters. In zebrafish, knockdown eif5a partly recapitulates the human phenotype that can be rescued with 1 µM spermidine supplementation. In summary, we uncover the role of eIF5A in human development and disease, demonstrate the mechanistic complexity of EIF5A-related disorder and raise possibilities for its treatment.</p

Second-tier trio exome sequencing after negative solo clinical exome sequencing: an efficient strategy to increase diagnostic yield and decipher molecular bases in undiagnosed developmental disorders

International audienceDevelopmental disorders (DD), characterized by malformations/dysmorphism and/or intellectual disability, affecting around 3% of worldwide population, are mostly linked to genetic anomalies. Despite clinical exome sequencing (cES) centered on genes involved in human genetic disorders, the majority of patients affected by DD remain undiagnosed after solo-cES. Trio-based strategy is expected to facilitate variant selection thanks to rapid parental segregation. We performed a second step trio-ES (not only focusing on genes involved in human disorders) analysis in 70 patients with negative results after solo-cES. All candidate variants were shared with a MatchMaking exchange system to identify additional patients carrying variants in the same genes and with similar phenotype. In 18/70 patients (26%), we confirmed causal implication of nine OMIM-morbid genes and identified nine new strong candidate genes (eight de novo and one compound heterozygous variants). These nine new candidate genes were validated through the identification of patients with similar phenotype and genotype thanks to data sharing. Moreover, 11 genes harbored variants of unknown significance in 10/70 patients (14%). In DD, a second step trio-based ES analysis appears an efficient strategy in diagnostic and translational research to identify highly candidate genes and improve diagnostic yield