Defining the Genetic Landscape of Congenital Mirror Movements in 80 Affected Individuals.

BACKGROUND: Congenital mirror movements (CMM) is a rare neurodevelopmental disorder characterized by involuntary movements from one side of the body that mirror voluntary movements on the opposite side. To date, five genes have been associated with CMM, namely DCC, RAD51, NTN1, ARHGEF7, and DNAL4. OBJECTIVE: The aim of this study is to characterize the genetic landscape of CMM in a large group of 80 affected individuals. METHODS: We screened 80 individuals with CMM from 43 families for pathogenic variants in CMM genes. In large CMM families, we tested for presence of pathogenic variants in multiple affected and unaffected individuals. In addition, we evaluated the impact of three missense DCC variants on binding between DCC and Netrin-1 in vitro. RESULTS: Causal pathogenic/likely pathogenic variants were found in 35% of probands overall, and 70% with familial CMM. The most common causal gene was DCC, responsible for 28% of CMM probands and 80% of solved cases. RAD51, NTN1, and ARHGEF7 were rare causes of CMM, responsible for 2% each. Penetrance of CMM in DCC pathogenic variant carriers was 68% and higher in males than females (74% vs. 54%). The three tested missense variants (p.Ile164Thr; p.Asn176Ser; and p.Arg1343His) bind Netrin-1 similarly to wild type DCC. CONCLUSIONS: A genetic etiology can be identified in one third of CMM individuals, with DCC being the most common gene involved. Two thirds of CMM individuals were unsolved, highlighting that CMM is genetically heterogeneous and other CMM genes are yet to be discovered. © 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

mTOR Pathway Somatic Pathogenic Variants in Focal Malformations of Cortical Development: Novel Variants, Topographic Mapping, and Clinical Outcomes.

Background and Objectives: Somatic and germline pathogenic variants in genes of the mammalian target of rapamycin (mTOR) signaling pathway are a common mechanism underlying a subset of focal malformations of cortical development (FMCDs) referred to as mTORopathies, which include focal cortical dysplasia (FCD) type II, subtypes of polymicrogyria, and hemimegalencephaly. Our objective is to screen resected FMCD specimens with mTORopathy features on histology for causal somatic variants in mTOR pathway genes, describe novel pathogenic variants, and examine the variant distribution in relation to neuroimaging, histopathologic classification, and clinical outcomes. Methods: We performed ultra-deep sequencing using a custom HaloPlexHS Target Enrichment kit in DNA from 21 resected fresh-frozen histologically confirmed FCD type II, tuberous sclerosis complex, or hemimegalencephaly specimens. We mapped the variant alternative allele frequency (AAF) across the resected brain using targeted ultra-deep sequencing in multiple formalin-fixed paraffin-embedded tissue blocks. We also functionally validated 2 candidate somatic MTOR variants and performed targeted RNA sequencing to validate a splicing defect associated with a novel DEPDC5 variant. Results: We identified causal mTOR pathway gene variants in 66.7% (14/21) of patients, of which 13 were somatic with AAF ranging between 0.6% and 12.0%. Moreover, the AAF did not predict balloon cell presence. Favorable seizure outcomes were associated with genetically clear resection borders. Individuals in whom a causal somatic variant was undetected had excellent postsurgical outcomes. In addition, we demonstrate pathogenicity of the novel c.4373_4375dupATG and candidate c.7499T>A MTOR variants in vitro. We also identified a novel germline aberrant splice site variant in DEPDC5 (c.2802-1G>C). Discussion: The AAF of somatic pathogenic variants correlated with the topographic distribution, histopathology, and postsurgical outcomes. Moreover, cortical regions with absent histologic FCD features had negligible or undetectable pathogenic variant loads. By contrast, specimens with frank histologic abnormalities had detectable pathogenic variant loads, which raises important questions as to whether there is a tolerable variant threshold and whether surgical margins should be clean, as performed in tumor resections. In addition, we describe 2 novel pathogenic variants, expanding the mTORopathy genetic spectrum. Although most pathogenic somatic variants are located at mutation hotspots, screening the full-coding gene sequence remains necessary in a subset of patients.

De novo mutations in the X-linked TFE3 gene cause intellectual disability with pigmentary mosaicism and storage disorder-like features.

INTRODUCTION: Pigmentary mosaicism (PM) manifests by pigmentation anomalies along Blaschko's lines and represents a clue toward the molecular diagnosis of syndromic intellectual disability (ID). Together with new insights on the role for lysosomal signalling in embryonic stem cell differentiation, mutations in the X-linked transcription factor 3 (TFE3) have recently been reported in five patients. Functional analysis suggested these mutations to result in ectopic nuclear gain of functions. MATERIALS AND METHODS: Subsequent data sharing allowed the clustering of de novo TFE3 variants identified by exome sequencing on DNA extracted from leucocytes in patients referred for syndromic ID with or without PM. RESULTS: We describe the detailed clinical and molecular data of 17 individuals harbouring a de novo TFE3 variant, including the patients that initially allowed reporting TFE3 as a new disease-causing gene. The 12 females and 5 males presented with pigmentation anomalies on Blaschko's lines, severe ID, epilepsy, storage disorder-like features, growth retardation and recognisable facial dysmorphism. The variant was at a mosaic state in at least two male patients. All variants were missense except one splice variant. Eleven of the 13 variants were localised in exon 4, 2 in exon 3, and 3 were recurrent variants. CONCLUSION: This series further delineates the specific storage disorder-like phenotype with PM ascribed to de novo TFE3 mutation in exons 3 and 4. It confirms the identification of a novel X-linked human condition associated with mosaicism and dysregulation within the mechanistic target of rapamycin (mTOR) pathway, as well as a link between lysosomal signalling and human development.

Excess of de novo variants in genes involved in chromatin remodelling in patients with marfanoid habitus and intellectual disability.

PURPOSE: Marfanoid habitus (MH) combined with intellectual disability (ID) (MHID) is a clinically and genetically heterogeneous presentation. The combination of array CGH and targeted sequencing of genes responsible for Marfan or Lujan-Fryns syndrome explain no more than 20% of subjects. METHODS: To further decipher the genetic basis of MHID, we performed exome sequencing on a combination of trio-based (33 subjects) or single probands (31 subjects), of which 61 were sporadic. RESULTS: We identified eight genes with de novo variants (DNVs) in at least two unrelated individuals (ARID1B, ATP1A1, DLG4, EHMT1, NFIX, NSD1, NUP205 and ZEB2). Using simulation models, we showed that five genes (DLG4, NFIX, EHMT1, ZEB2 and ATP1A1) met conservative Bonferroni genomewide significance for an excess of the observed de novo point variants. Overall, at least one pathogenic or likely pathogenic variant was identified in 54.7% of subjects (35/64). These variants fell within 27 genes previously associated with Mendelian disorders, including NSD1 and NFIX, which are known to be mutated in overgrowth syndromes. CONCLUSION: We demonstrated that DNVs were enriched in chromatin remodelling (p=2×10-4) and genes regulated by the fragile X mental retardation protein (p=3×10-8), highlighting overlapping genetic mechanisms between MHID and related neurodevelopmental disorders.

Author Correction: Postzygotic inactivating mutations of RHOA cause a mosaic neuroectodermal syndrome.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Heterozygous Missense Pathogenic Variants Within the Second Spectrin Repeat of SPTBN2 Lead to Infantile-Onset Cerebellar Ataxia.

The term spinocerebellar ataxia encompasses a heterogeneous group of neurodegenerative disorders due to pathogenic variants in more than 100 genes, underlying 2 major groups of ataxia: autosomal dominant cerebellar ataxias (ADCA, also known as spinocerebellar ataxias [SCAs]) due to heterozygous variants or polyglutamine triplet expansions leading to adult-onset ataxia, and autosomal recessive spinocerebellar ataxias (ARCAs, also known as SCARs) due to biallelic variants, usually resulting in more severe and earlier-onset cerebellar ataxia. Certain ataxia genes, including SPTBN2 which encodes ß-III spectrin, are responsible for both SCA and SCAR, depending on whether the pathogenic variant occurs in a monoallelic or biallelic state, respectively. Accordingly, 2 major phenotypes have been linked to SPTBN2: pathogenic heterozygous in-frame deletions and missense variants result in an adult-onset, slowly progressive ADCA (SCA5) through a dominant negative effect, whereas biallelic loss-of-function variants cause SCAR14, an allelic disorder characterized by infantile-onset cerebellar ataxia and cognitive impairment. Of note, 2 heterozygous missense variants (c.1438C>T, p.R480 W; c.1309C>G, p.R437G), both lying in the second spectrin repeat of SPTBN2, have been linked to infantile-onset cerebellar ataxia, similar to SCAR14. Here, we report a novel de novo heterozygous pathogenic missense variant (c.1310G>A) in SPTBN2 in a child with infantile-onset cerebellar ataxia and mild cognitive impairment. This variant affects the same R437 residue of the second spectrin repeat but results in a different amino acid change (p.R437Q). We review previously reported cases and discuss possible pathomechanisms responsible for the early-onset cerebellar phenotype due to disease-causing variants in the second spectrin repeat.

Postzygotic inactivating mutations of RHOA cause a mosaic neuroectodermal syndrome.

Hypopigmentation along Blaschko's lines is a hallmark of a poorly defined group of mosaic syndromes whose genetic causes are unknown. Here we show that postzygotic inactivating mutations of RHOA cause a neuroectodermal syndrome combining linear hypopigmentation, alopecia, apparently asymptomatic leukoencephalopathy, and facial, ocular, dental and acral anomalies. Our findings pave the way toward elucidating the etiology of pigmentary mosaicism and highlight the role of RHOA in human development and disease.

Author Correction: Postzygotic inactivating mutations of RHOA cause a mosaic neuroectodermal syndrome.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

De Novo Pathogenic Variants in N-cadherin Cause a Syndromic Neurodevelopmental Disorder with Corpus Collosum, Axon, Cardiac, Ocular, and Genital Defects.

Cadherins constitute a family of transmembrane proteins that mediate calcium-dependent cell-cell adhesion. The extracellular domain of cadherins consists of extracellular cadherin (EC) domains, separated by calcium binding sites. The EC interacts with other cadherin molecules in cis and in trans to mechanically hold apposing cell surfaces together. CDH2 encodes N-cadherin, whose essential roles in neural development include neuronal migration and axon pathfinding. However, CDH2 has not yet been linked to a Mendelian neurodevelopmental disorder. Here, we report de novo heterozygous pathogenic variants (seven missense, two frameshift) in CDH2 in nine individuals with a syndromic neurodevelopmental disorder characterized by global developmental delay and/or intellectual disability, variable axon pathfinding defects (corpus callosum agenesis or hypoplasia, mirror movements, Duane anomaly), and ocular, cardiac, and genital anomalies. All seven missense variants (c.1057G>A [p.Asp353Asn]; c.1789G>A [p.Asp597Asn]; c.1789G>T [p.Asp597Tyr]; c.1802A>C [p.Asn601Thr]; c.1839C>G [p.Cys613Trp]; c.1880A>G [p.Asp627Gly]; c.2027A>G [p.Tyr676Cys]) result in substitution of highly conserved residues, and six of seven cluster within EC domains 4 and 5. Four of the substitutions affect the calcium-binding site in the EC4-EC5 interdomain. We show that cells expressing these variants in the EC4-EC5 domains have a defect in cell-cell adhesion; this defect includes impaired binding in trans with N-cadherin-WT expressed on apposing cells. The two frameshift variants (c.2563_2564delCT [p.Leu855Valfs∗4]; c.2564_2567dupTGTT [p.Leu856Phefs∗5]) are predicted to lead to a truncated cytoplasmic domain. Our study demonstrates that de novo heterozygous variants in CDH2 impair the adhesive activity of N-cadherin, resulting in a multisystemic developmental disorder, that could be named ACOG syndrome (agenesis of corpus callosum, axon pathfinding, cardiac, ocular, and genital defects).

Pathogenic variants in AIMP1 cause pontocerebellar hypoplasia.

Aminoacyl-tRNA synthetase-interacting multifunctional protein 1 (AIMP1) is a non-catalytic component of the multi-tRNA synthetase complex which catalyzes the ligation of amino acids to the correct tRNAs. Pathogenic variants in several aminoacyl-tRNA synthetases genes have been linked to various neurological disorders, including leukodystrophies and pontocerebellar hypoplasias (PCH). To date, loss-of-function variants in AIMP1 have been associated with hypomyelinating leukodystrophy-3 (MIM 260600). Here, we report a novel frameshift AIMP1 homozygous variant (c.160delA,p.Lys54Asnfs) in a child with pontocerebellar hypoplasia and simplified gyral pattern, a phenotype not been previously described with AIMP1 variants, thus expanding the phenotypic spectrum. AIMP1 should be included in diagnostic PCH gene panels.

Lysosomal Signaling Licenses Embryonic Stem Cell Differentiation via Inactivation of Tfe3.

Self-renewal and differentiation of pluripotent murine embryonic stem cells (ESCs) is regulated by extrinsic signaling pathways. It is less clear whether cellular metabolism instructs developmental progression. In an unbiased genome-wide CRISPR/Cas9 screen, we identified components of a conserved amino-acid-sensing pathway as critical drivers of ESC differentiation. Functional analysis revealed that lysosome activity, the Ragulator protein complex, and the tumor-suppressor protein Folliculin enable the Rag GTPases C and D to bind and seclude the bHLH transcription factor Tfe3 in the cytoplasm. In contrast, ectopic nuclear Tfe3 represses specific developmental and metabolic transcriptional programs that are associated with peri-implantation development. We show differentiation-specific and non-canonical regulation of Rag GTPase in ESCs and, importantly, identify point mutations in a Tfe3 domain required for cytoplasmic inactivation as potentially causal for a human developmental disorder. Our work reveals an instructive and biomedically relevant role of metabolic signaling in licensing embryonic cell fate transitions.

Helping Bridge the Great Divides: Supporting Nurse Communication at End of Life.

Many, especially newer, nurses lack the experience for nuanced skillful communication at end of life. This study incorporated the End-of-Life Nursing Education Consortium (ELNEC) Core Curriculum Communication module into a nurse residency program and tested for change in attitudes toward end-of-life (EOL) care. Researchers found significant differences in attitudes toward care of the dying before and after the ELNEC training, and between nurses with greater than and less than 5 years experience. Findings support that ELNEC education can significantly impact nurse attitudes about EOL care.

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

PurposeCongenital 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.

Expanding the clinical spectrum of recessive truncating mutations of KLHL7 to a Bohring-Opitz-like phenotype.

BACKGROUND: Bohring-Opitz syndrome (BOS) is a rare genetic disorder characterised by a recognisable craniofacial appearance and a typical 'BOS' posture. BOS is caused by sporadic mutations ofASXL1. However, several typical patients with BOS have no molecular diagnosis, suggesting clinical and genetic heterogeneity. OBJECTIVES: To expand the phenotypical spectrum of autosomal recessive variants of KLHL7, reported as causing Crisponi syndrome/cold-induced sweating syndrome type 1 (CS/CISS1)-like syndrome. METHODS: We performed whole-exome sequencing in two families with a suspected recessive mode of inheritance. We used the Matchmaker Exchange initiative to identify additional patients. RESULTS: Here, we report six patients with microcephaly, facial dysmorphism, including exophthalmos, nevus flammeus of the glabella and joint contractures with a suspected BOS posture in five out of six patients. We identified autosomal recessive truncating mutations in the KLHL7 gene. KLHL7 encodes a BTB-kelch protein implicated in the cell cycle and in protein degradation by the ubiquitin-proteasome pathway. Recently, biallelic mutations in the KLHL7 gene were reported in four families and associated with CS/CISS1, characterised by clinical features overlapping with our patients. CONCLUSION: We have expanded the clinical spectrum of KLHL7 autosomal recessive variants by describing a syndrome with features overlapping CS/CISS1 and BOS.

Reducing diagnostic turnaround times of exome sequencing for families requiring timely diagnoses.

BACKGROUND AND OBJECTIVE: Whole-exome sequencing (WES) has now entered medical practice with powerful applications in the diagnosis of rare Mendelian disorders. Although the usefulness and cost-effectiveness of WES have been widely demonstrated, it is essential to reduce the diagnostic turnaround time to make WES a first-line procedure. Since 2011, the automation of laboratory procedures and advances in sequencing chemistry have made it possible to carry out diagnostic whole genome sequencing from the blood sample to molecular diagnosis of suspected genetic disorders within 50 h. Taking advantage of these advances, the main objective of the study was to improve turnaround times for sequencing results. METHODS: WES was proposed to 29 patients with severe undiagnosed disorders with developmental abnormalities and faced with medical situations requiring rapid diagnosis. Each family gave consent. The extracted DNA was sequenced on a NextSeq500 (Illumina) instrument. Data were analyzed following standard procedures. Variants were interpreted using in-house software. Each rare variant affecting protein sequences with clinical relevance was tested for familial segregation. RESULTS: The diagnostic rate was 45% (13/29), with a mean turnaround time of 40 days from reception of the specimen to delivery of results to the referring physician. Besides permitting genetic counseling, the rapid diagnosis for positive families led to two pre-natal diagnoses and two inclusions in clinical trials. CONCLUSIONS: This pilot study demonstrated the feasibility of rapid diagnostic WES in our primary genetics center. It reduced the diagnostic odyssey and helped provide support to families.

Fifteen years of research on oral-facial-digital syndromes: from 1 to 16 causal genes.

Oral-facial-digital syndromes (OFDS) gather rare genetic disorders characterised by facial, oral and digital abnormalities associated with a wide range of additional features (polycystic kidney disease, cerebral malformations and several others) to delineate a growing list of OFDS subtypes. The most frequent, OFD type I, is caused by a heterozygous mutation in the OFD1 gene encoding a centrosomal protein. The wide clinical heterogeneity of OFDS suggests the involvement of other ciliary genes. For 15 years, we have aimed to identify the molecular bases of OFDS. This effort has been greatly helped by the recent development of whole-exome sequencing (WES). Here, we present all our published and unpublished results for WES in 24 cases with OFDS. We identified causal variants in five new genes (C2CD3, TMEM107, INTU, KIAA0753 and IFT57) and related the clinical spectrum of four genes in other ciliopathies (C5orf42, TMEM138, TMEM231 and WDPCP) to OFDS. Mutations were also detected in two genes previously implicated in OFDS. Functional studies revealed the involvement of centriole elongation, transition zone and intraflagellar transport defects in OFDS, thus characterising three ciliary protein modules: the complex KIAA0753-FOPNL-OFD1, a regulator of centriole elongation; the Meckel-Gruber syndrome module, a major component of the transition zone; and the CPLANE complex necessary for IFT-A assembly. OFDS now appear to be a distinct subgroup of ciliopathies with wide heterogeneity, which makes the initial classification obsolete. A clinical classification restricted to the three frequent/well-delineated subtypes could be proposed, and for patients who do not fit one of these three main subtypes, a further classification could be based on the genotype.

Molecular diagnosis of PIK3CA-related overgrowth spectrum (PROS) in 162 patients and recommendations for genetic testing.

PURPOSE: Postzygotic activating mutations of PIK3CA cause a wide range of mosaic disorders collectively referred to as PIK3CA-related overgrowth spectrum (PROS). We describe the diagnostic yield and characteristics of PIK3CA sequencing in PROS. METHODS: We performed ultradeep next-generation sequencing (NGS) of PIK3CA in various tissues from 162 patients referred to our clinical laboratory and assessed diagnostic yield by phenotype and tissue tested. RESULTS: We identified disease-causing mutations in 66.7% (108/162) of patients, with mutant allele levels as low as 1%. The diagnostic rate was higher (74%) in syndromic than in isolated cases (35.5%; P = 9.03 × 10-5). We identified 40 different mutations and found strong oncogenic mutations more frequently in patients without brain overgrowth (50.6%) than in those with brain overgrowth (15.2%; P = 0.00055). Mutant allele levels were higher in skin and overgrown tissues than in blood and buccal samples (P = 3.9 × 10-25), regardless of the phenotype. CONCLUSION: Our data demonstrate the value of ultradeep NGS for molecular diagnosis of PROS, highlight its substantial allelic heterogeneity, and confirm that optimal diagnosis requires fresh skin or surgical samples from affected regions. Our findings may be of value in guiding future recommendations for genetic testing in PROS and other mosaic conditions.Genet Med advance online publication 02 February 2017.

Biallelic SCN10A mutations in neuromuscular disease and epileptic encephalopathy.

OBJECTIVES: Two consanguineous families, one of Sudanese ethnicity presenting progressive neuromuscular disease, severe cognitive impairment, muscle weakness, upper motor neuron lesion, anhydrosis, facial dysmorphism, and recurrent seizures and the other of Egyptian ethnicity presenting with neonatal hypotonia, bradycardia, and recurrent seizures, were evaluated for the causative gene mutation. METHODS AND RESULTS: Homozygosity mapping and whole exome sequencing (WES) identified damaging homozygous variants in SCN10A, namely c.4514C>T; p.Thr1505Met in the first family and c.4735C>T; p.Arg1579* in the second family. A third family, of Western European descent, included a child with febrile infection-related epilepsy syndrome (FIRES) who also had compound heterozygous missense mutations in SCN10A, namely, c.3482T>C; p.Met1161Thr and c.4709C>A; p.Thr1570Lys. A search for SCN10A variants in three consortia datasets (EuroEPINOMICS, Epi4K/EPGP, Autism/dbGaP) identified an additional five individuals with compound heterozygous variants. A Hispanic male with infantile spasms [c.2842G>C; p.Val948Leu and c.1453C>T; p.Arg485Cys], and a Caucasian female with Lennox-Gastaut syndrome [c.1529C>T; p.Pro510Leu and c.4984G>A; p.Gly1662Ser] in the epilepsy databases and three in the autism databases with [c.4009T>A; p.Ser1337Thr and c.1141A>G; p.Ile381Val], [c.2972C>T; p.Pro991Leu and c.2470C>T; p.His824Tyr], and [c.4009T>A; p.Ser1337Thr and c.2052G>A; p.Met684Ile]. INTERPRETATION: SCN10A is a member of the voltage-gated sodium channel (VGSC) gene family. Sodium channels are responsible for the instigation and proliferation of action potentials in central and peripheral nervous systems. Heterozygous mutations in VGSC genes cause a wide range of epileptic and peripheral nervous system disorders. This report presents autosomal recessive mutations in SCN10A that may be linked to epilepsy-related phenotypes, Lennox-Gastaut syndrome, infantile spasms, and Autism Spectrum Disorder.