Genome sequencing and comprehensive rare-variant analysis of 465 families with neurodevelopmental disorders.

Despite significant progress in unraveling the genetic causes of neurodevelopmental disorders (NDDs), a substantial proportion of individuals with NDDs remain without a genetic diagnosis after microarray and/or exome sequencing. Here, we aimed to assess the power of short-read genome sequencing (GS), complemented with long-read GS, to identify causal variants in participants with NDD from the National Institute for Health and Care Research (NIHR) BioResource project. Short-read GS was conducted on 692 individuals (489 affected and 203 unaffected relatives) from 465 families. Additionally, long-read GS was performed on five affected individuals who had structural variants (SVs) in technically challenging regions, had complex SVs, or required distal variant phasing. Causal variants were identified in 36% of affected individuals (177/489), and a further 23% (112/489) had a variant of uncertain significance after multiple rounds of re-analysis. Among all reported variants, 88% (333/380) were coding nuclear SNVs or insertions and deletions (indels), and the remainder were SVs, non-coding variants, and mitochondrial variants. Furthermore, long-read GS facilitated the resolution of challenging SVs and invalidated variants of difficult interpretation from short-read GS. This study demonstrates the value of short-read GS, complemented with long-read GS, in investigating the genetic causes of NDDs. GS provides a comprehensive and unbiased method of identifying all types of variants throughout the nuclear and mitochondrial genomes in individuals with NDD.

Structural mapping of GABRB3 variants reveals genotype-phenotype correlations.

PURPOSE: Pathogenic variants in GABRB3 have been associated with a spectrum of phenotypes from severe developmental disorders and epileptic encephalopathies to milder epilepsy syndromes and mild intellectual disability (ID). In this study, we analyzed a large cohort of individuals with GABRB3 variants to deepen the phenotypic understanding and investigate genotype-phenotype correlations. METHODS: Through an international collaboration, we analyzed electro-clinical data of unpublished individuals with variants in GABRB3, and we reviewed previously published cases. All missense variants were mapped onto the 3-dimensional structure of the GABRB3 subunit, and clinical phenotypes associated with the different key structural domains were investigated. RESULTS: We characterized 71 individuals with GABRB3 variants, including 22 novel subjects, expressing a wide spectrum of phenotypes. Interestingly, phenotypes correlated with structural locations of the variants. Generalized epilepsy, with a median age at onset of 12 months, and mild-to-moderate ID were associated with variants in the extracellular domain. Focal epilepsy with earlier onset (median: age 4 months) and severe ID were associated with variants in both the pore-lining helical transmembrane domain and the extracellular domain. CONCLUSION: These genotype-phenotype correlations will aid the genetic counseling and treatment of individuals affected by GABRB3-related disorders. Future studies may reveal whether functional differences underlie the phenotypic differences.

Fifteen-minute consultation: The efficient investigation of infantile and childhood epileptic encephalopathies in the era of modern genomics.

The investigation of children presenting with infantile and childhood epileptic encephalopathies (ICEE) is challenging due to diverse aetiologies, overlapping phenotypes and the relatively low diagnostic yield of MRI, electroencephalography (EEG) and biochemical investigations. Careful history and thorough examination remain essential as these may identify an acquired cause or indicate more targeted investigation for a genetic disorder. Whole exome sequencing (WES) with analysis of a panel of candidate epilepsy genes has increased the diagnostic yield. Whole genome sequencing (WGS), particularly as a trio with both parents' DNA, is likely to supersede WES. Modern genomic investigation impacts on the timing and necessity of other testing. We propose a structured approach for children presenting with ICEE where there is diagnostic uncertainty, emphasising the importance of WGS or, if unavailable, WES early in the investigative process. We note the importance of expert review of all investigations, including radiology, neurophysiology and biochemistry, to confirm the technique used was appropriate as well as the results. It is essential to counsel families on the risks associated with the procedures, the yield of the procedures, findings that are difficult to interpret and implication of 'negative' results. Where children remain without a diagnosis despite comprehensive investigation, we note the importance of ongoing multidisciplinary care.

Post-traumatic seizure with an unexpected finding.

A 5-year-old boy of non-consanguineous Indian descent presented to the emergency department (ED) following a prolonged seizure, preceded by a minor head injury from a low-level unwitnessed fall. The seizure was described as focal with head and neck version to the right. There was urinary and faecal incontinence and foaming at the mouth. The seizure lasted for 30 min, following which the child made a rapid recovery and had no neurological deficit when examined in the ED. His initial observations were unremarkable, with him being afebrile, normotensive and having a blood sugar of 4.6 mmol/L. QUESTION 1: Which imaging modality, if any, should be performed? QUESTION 2: Initial imaging showed two parenchymal lesions, one within the left frontal lobe and the other in the right parietal lobe with possible areas of blood or calcification. MRI (figure 1A,B) was urgently undertaken and reported as likely to be either haemorrhage into cavernomatous malformations, haemorrhagic metastases or haemorrhage within infected lesions such as tuberculomas.What would be the most useful next step in view of the differential diagnosis?CT chest and abdomenPerform a C-reactive proteinPerform a Mantoux testRefer to neurosurgeryTake a detailed family history QUESTION 3: As the patient remained stable on the ward, with no signs of focal neurology and no mass effects on MRI, a lumbar puncture was performed the following day to exclude tuberculosis (TB) (table 1). What cell appearance would indicate TB infection? QUESTION 4: A review of the mother's medical records and MRI scans revealed her to have multiple discrete cerebral cavernous malformations (CCMs) (figure 2).In view of this new information, what is the next step in investigating this child?

Fifteen minute consultation: an approach to the child with chronic headaches.

Children with chronic headache are a common referral to paediatric outpatients. This article suggests an approach to the assessment and management of chronic headaches, offering practical strategies for management as there is limited literature in paediatrics for this.

Mosaicism for a pathogenic MFN2 mutation causes minimal clinical features of CMT2A in the parent of a severely affected child.

Charcot-Marie-Tooth disease (CMT) refers to a genetically heterogeneous group of disorders which cause a peripheral motor and sensory neuropathy. The overall prevalence is 1 in 2500 individuals. Mutations in the MFN2 gene are the commonest cause for the axonal (CMT2) type. We describe a Caucasian 5-year old girl affected by CMT2A since the age of 2 years. She presented with unsteady gait, in-turning of the feet and progressive foot deformities. Nerve conduction studies suggested an axonal neuropathy and molecular testing identified a previously reported pathogenic variant c.1090C > T, p.(Arg364Trp) in the MFN2 gene. This variant was also detected in a mosaic state in blood and saliva by Sanger sequencing in her subjectively healthy father. Next generation sequencing showed that the level of mosaicism was 21% in blood and 24% in saliva. A high recurrence risk was given because the father had proven somatic mosaicism and an affected child implying gonadal mosaicism. The parents were referred for pre-implantation genetic diagnosis. To the best of our knowledge, this is the first reported case of somatic mosaicism for MFN2. This study has important implications for genetic counselling in families with CMT2A.

GABRB3 mutations: a new and emerging cause of early infantile epileptic encephalopathy.

The gamma-aminobutyric acid type A receptor ß3 gene (GABRB3) encodes the ß3-subunit of the gamma-aminobutyric acid type A (GABAA ) receptor, which mediates inhibitory signalling within the central nervous system. Recently, GABRB3 mutations have been identified in a few patients with infantile spasms and Lennox-Gastaut syndrome. We report the clinical and electrographic features of a novel case of GABRB3-related early-onset epileptic encephalopathy. Our patient presented with neonatal hypotonia and feeding difficulties, then developed pharmacoresistant epileptic encephalopathy, characterized by multiple seizure types from 3 months of age. Electroencephalography demonstrated ictal generalized and interictal multifocal epileptiform abnormalities. Using a SureSelectXT custom multiple gene panel covering 48 early infantile epileptic encephalopathy/developmental delay genes, a novel de novo GABRB3 heterozygous missense mutation, c.860C>T (p.Thr287Ile), was identified and confirmed on Sanger sequencing. GABRB3 is an emerging cause of early-onset epilepsy. Novel genetic technologies, such as whole-exome/genome sequencing and multiple gene panels, will undoubtedly identify further cases, allowing more detailed electroclinical delineation of the GABRB3-related genotypic and phenotypic spectra.

Risdiplam in Spinal Muscular Atrophy: Safety Profile and Use Through The Early Access to Medicine Scheme for the Paediatric Cohort in Great Britain.

Background: Spinal muscular atrophy (SMA) is a progressive neuromuscular disease caused by mutations in Survival motor neuron 1 (SMN1) gene, leading to reduction in survival motor neuron protein (SMN), key for motor neuron survival and function in the brainstem and spinal cord. Risdiplam is an orally administered SMN2-splicing modifier which increases production of functional SMN protein. Risdiplam was offered in the UK under early access to medicines scheme (EAMS) to SMA type 1 and 2 patients aged 2 months and older, not suitable for authorised treatments from September 2020 to December 2021. Objective: To describe the largest paediatric European real-world set of data on patients' characteristics and short-term safety for risdiplam in Great Britain through EAMS. Methods: We collated data from SMA REACH UK a national clinical and research network for all patients enrolled onto EAMS and assessed all submitted adverse events. Results: Of the 92 patients; 78% were Type 2 SMA, mean age 10.9 years, range 0-17 years. 56 were treatment naïve, 33 previously treated; of these 25 had received nusinersen, 3 previous treatment unknown. Sixty adverse events (AEs) were reported occurring in 34 patients. The commonest were respiratory tract infections and gastrointestinal disturbance. Four life-threatening events were reported with 2 deaths and permanent cessation of risdiplam in 3 patients.Overall, 38/60 AEs were considered unrelated to risdiplam, 10/60 related to risdiplam and for 12/60 causality not specified. Conclusions: This study found a safety profile similar to clinical trials with no new safety concerns identified. With the restricted eligibility of onasemnogene abeparvovec and complications of nusinersen administration, EAMS allowed access or continued treatment to naïve patients or patients no longer suitable for approved medications. Collection of longitudinal data for this complex population is needed, to provide greater insights into risdiplam's role in addressing patients' needs into the future.

Heterozygous frameshift variants in HNRNPA2B1 cause early-onset oculopharyngeal muscular dystrophy.

Missense variants in RNA-binding proteins (RBPs) underlie a spectrum of disease phenotypes, including amyotrophic lateral sclerosis, frontotemporal dementia, and inclusion body myopathy. Here, we present ten independent families with a severe, progressive muscular dystrophy, reminiscent of oculopharyngeal muscular dystrophy (OPMD) but of much earlier onset, caused by heterozygous frameshift variants in the RBP hnRNPA2/B1. All disease-causing frameshift mutations abolish the native stop codon and extend the reading frame, creating novel transcripts that escape nonsense-mediated decay and are translated to produce hnRNPA2/B1 protein with the same neomorphic C-terminal sequence. In contrast to previously reported disease-causing missense variants in HNRNPA2B1, these frameshift variants do not increase the propensity of hnRNPA2 protein to fibrillize. Rather, the frameshift variants have reduced affinity for the nuclear import receptor karyopherin ß2, resulting in cytoplasmic accumulation of hnRNPA2 protein in cells and in animal models that recapitulate the human pathology. Thus, we expand the phenotypes associated with HNRNPA2B1 to include an early-onset form of OPMD caused by frameshift variants that alter its nucleocytoplasmic transport dynamics.

Effectiveness and safety of the tri-iodothyronine analogue Triac in children and adults with MCT8 deficiency: an international, single-arm, open-label, phase 2 trial.

BACKGROUND: Deficiency of the thyroid hormone transporter monocarboxylate transporter 8 (MCT8) causes severe intellectual and motor disability and high serum tri-iodothyronine (T3) concentrations (Allan-Herndon-Dudley syndrome). This chronic thyrotoxicosis leads to progressive deterioration in bodyweight, tachycardia, and muscle wasting, predisposing affected individuals to substantial morbidity and mortality. Treatment that safely alleviates peripheral thyrotoxicosis and reverses cerebral hypothyroidism is not yet available. We aimed to investigate the effects of treatment with the T3 analogue Triac (3,3',5-tri-iodothyroacetic acid, or tiratricol), in patients with MCT8 deficiency. METHODS: In this investigator-initiated, multicentre, open-label, single-arm, phase 2, pragmatic trial, we investigated the effectiveness and safety of oral Triac in male paediatric and adult patients with MCT8 deficiency in eight countries in Europe and one site in South Africa. Triac was administered in a predefined escalating dose schedule-after the initial dose of once-daily 350 µg Triac, the daily dose was increased progressively in 350 µg increments, with the goal of attaining serum total T3 concentrations within the target range of 1·4-2·5 nmol/L. We assessed changes in several clinical and biochemical signs of hyperthyroidism between baseline and 12 months of treatment. The prespecified primary endpoint was the change in serum T3 concentrations from baseline to month 12. The co-primary endpoints were changes in concentrations of serum thyroid-stimulating hormone (TSH), free and total thyroxine (T4), and total reverse T3 from baseline to month 12. These analyses were done in patients who received at least one dose of Triac and had at least one post-baseline evaluation of serum throid function. This trial is registered with ClinicalTrials.gov, number NCT02060474. FINDINGS: Between Oct 15, 2014, and June 1, 2017, we screened 50 patients, all of whom were eligible. Of these patients, four (8%) patients decided not to participate because of travel commitments. 46 (92%) patients were therefore enrolled in the trial to receive Triac (median age 7·1 years [range 0·8-66·8]). 45 (98%) participants received Triac and had at least one follow-up measurement of thyroid function and thus were included in the analyses of the primary endpoints. Of these 45 patients, five did not complete the trial (two patients withdrew [travel burden, severe pre-existing comorbidity], one was lost to follow-up, one developed of Graves disease, and one died of sepsis). Patients required a mean dose of 38.3 µg/kg of bodyweight (range 6·4-84·3) to attain T3 concentrations within the target range. Serum T3 concentration decreased from 4·97 nmol/L (SD 1·55) at baseline to 1·82 nmol/L (0·69) at month 12 (mean decrease 3·15 nmol/L, 95% CI 2·68-3·62; p<0·0001), while serum TSH concentrations decreased from 2·91 mU/L (SD 1·68) to 1·02 mU/L (1·14; mean decrease 1·89 mU/L, 1·39-2·39; p<0·0001) and serum free T4 concentrations decreased from 9·5 pmol/L (SD 2·5) to 3·4 (1·6; mean decrease 6·1 pmol/L (5·4-6·8; p<0·0001). Additionally, serum total T4 concentrations decreased by 31·6 nmol/L (28·0-35·2; p<0·0001) and reverse T3 by 0·08 nmol/L (0·05-0·10; p<0·0001). Seven treatment-related adverse events (transiently increased perspiration or irritability) occurred in six (13%) patients. 26 serious adverse events that were considered unrelated to treatment occurred in 18 (39%) patients (mostly hospital admissions because of infections). One patient died from pulmonary sepsis leading to multi-organ failure, which was unrelated to Triac treatment. INTERPRETATION: Key features of peripheral thyrotoxicosis were alleviated in paediatric and adult patients with MCT8 deficiency who were treated with Triac. Triac seems a reasonable treatment strategy to ameliorate the consequences of untreated peripheral thyrotoxicosis in patients with MCT8 deficiency. FUNDING: Dutch Scientific Organization, Sherman Foundation, NeMO Foundation, Wellcome Trust, UK National Institute for Health Research Cambridge Biomedical Centre, Toulouse University Hospital, and Una Vita Rara ONLUS.

Clinical and molecular characterization of KCNT1-related severe early-onset epilepsy.

OBJECTIVE: To characterize the phenotypic spectrum, molecular genetic findings, and functional consequences of pathogenic variants in early-onset KCNT1 epilepsy. METHODS: We identified a cohort of 31 patients with epilepsy of infancy with migrating focal seizures (EIMFS) and screened for variants in KCNT1 using direct Sanger sequencing, a multiple-gene next-generation sequencing panel, and whole-exome sequencing. Additional patients with non-EIMFS early-onset epilepsy in whom we identified KCNT1 variants on local diagnostic multiple gene panel testing were also included. When possible, we performed homology modeling to predict the putative effects of variants on protein structure and function. We undertook electrophysiologic assessment of mutant KCNT1 channels in a xenopus oocyte model system. RESULTS: We identified pathogenic variants in KCNT1 in 12 patients, 4 of which are novel. Most variants occurred de novo. Ten patients had a clinical diagnosis of EIMFS, and the other 2 presented with early-onset severe nocturnal frontal lobe seizures. Three patients had a trial of quinidine with good clinical response in 1 patient. Computational modeling analysis implicates abnormal pore function (F346L) and impaired tetramer formation (F502V) as putative disease mechanisms. All evaluated KCNT1 variants resulted in marked gain of function with significantly increased channel amplitude and variable blockade by quinidine. CONCLUSIONS: Gain-of-function KCNT1 pathogenic variants cause a spectrum of severe focal epilepsies with onset in early infancy. Currently, genotype-phenotype correlations are unclear, although clinical outcome is poor for the majority of cases. Further elucidation of disease mechanisms may facilitate the development of targeted treatments, much needed for this pharmacoresistant genetic epilepsy.

Glycine receptor antibody-associated epilepsy in a boy aged 4†years.

Our patient was a previously normal boy who presented to his local hospital with an explosive onset of prolonged seizures and encephalopathy. He was treated for a presumed central nervous system infection and initial neuroimaging was normal. Despite treatment with antibiotics and antiepileptic drugs (AEDs), he remained encephalopathic and became ataxic over the next 48 hours, not related to medication. The seizures also proved resistant to treatment despite polytherapy with AEDs, and he required immune-modulatory treatment, intravenous methylprednisolone and intravenous immunoglobulin, in addition to the AEDs to achieve seizure control. The ataxia also improved following treatment. The initial EEG was normal but subsequent EEGs, separated by a week each, were abnormal and revealed subtle atypical 'delta-brush-like waves'. The patient's serum and cerebrospinal fluid were tested for autoantibodies, and he was found to be positive for glycine receptor antibodies that are neuronal antibodies.

LOWER EXTREMITY HYPERMOBILITY, BUT NOT CORE MUSCLE ENDURANCE INFLUENCES BALANCE IN FEMALE COLLEGIATE DANCERS.

BACKGROUND: Dance is a physically demanding activity, with almost 70% of all injuries in dancers occurring in the lower extremity (LE). Prior researchers report that muscle function (e.g. muscle endurance) and anatomical factors (e.g. hypermobility) affect physical performance (e.g. balance) and can subsequently influence LE injury risk. Specifically, lesser core muscle endurance, balance deficits, and greater hypermobility are related to increased LE injury risk. However, the potentials interrelationships among these factors in dancers remain unclear. PURPOSE: The purposes of this study were to examine the relationships among core muscle endurance, balance, and LE hypermobility, and determine the relative contributions of core muscle endurance and LE hypermobility as predictors of balance in female collegiate dancers. STUDY DESIGN: Cross-sectional. METHODS: Core muscle endurance was evaluated using the combined average anterior, left, and right lateral plank test time scores(s). LE hypermobility was measured using the LE-specific Beighton hypermobility measure, defining hypermobility if both legs had greater than 10 ° knee hyperextension. Balance was measured via the composite anterior, posterolateral, and posteromedial Star Excursion Balance Test (SEBT) reach distances (normalized to leg length) in 15 female healthy collegiate dancers (18.3 + 0.5yrs, 165.5 + 6.9cm, 63.7 + 12.1kg). Point-biserial-correlation-coefficients examined relationships and a linear regression examined whether core endurance and hypermobility predicted balance (p<.05). RESULTS: LE hypermobility (Yes; n = 3, No; n = 12) and balance (87.2 + 8.3% leg length) were positively correlated r(14)=.67, (p=.01). However, core endurance (103.9 + 50.6 s) and balance were not correlated r(14)=.32, (p=.26). LE hypermobility status predicted 36.9% of the variance in balance scores (p=.01). CONCLUSION: LE hypermobility, but not core muscle endurance may be related to balance in female collegiate dancers. While LE hypermobility status influenced balance in the female collegiate dancers, how this LE hypermobility status affects their longitudinal injury risk as their careers progress needs further study. Overall, the current findings suggest that rather than using isolated core endurance-centric training, clinicians may encourage dancers to use training programs that incorporate multiple muscles - in order to improve their balance, and possibly reduce their LE injury risk. LEVEL OF EVIDENCE: 2b.

The multiple phenotypes of arthrogryposis multiplex congenita with reference to the neurogenic variant.

Arthrogryposis can occur in isolation or as part of a syndrome. Amyoplasia, the commonest type of arthrogryposis, has been well described in literature, but neurogenic arthrogryposis, a rarer but significantly heterogeneous variant, has not. We conducted a single-centre, 10-year retrospective study of all children with arthrogryposis at the Dubowitz Neuromuscular Centre, London, UK to describe the various phenotypes of arthrogryposis with special reference to the neurogenic variant including presentation, associated features and long-term outcome. Twenty-seven children with arthrogryposis were identified (13 males) and 25 survivors followed-up for 6.4 ± 2.32 yrs. Perinatal history, presenting clinical features, investigations, final diagnosis and long-term outcomes were recorded. All four limbs were involved in 19 (ankles>wrists>elbows) whilst 8 had isolated upper (UL) or lower limb (LL) involvement. Twelve children had neurogenic arthrogryposis confirmed by a combination of clinical examination, EMG and/or muscle pathology. CK was normal in all children with neurogenic arthrogryposis. Three children in this cohort had abnormal brain MRIs and global developmental delay. Long-term follow up did not show deterioration of muscle power in any of our 12 children with neurogenic arthrogryposis, although contractures lead to temporary worsening of function ability in some (n = 3). Most children improved with physiotherapy and well-fitted orthoses. Our study suggests that neurogenic arthrogryposis is usually a non-progressive disorder and in the absence of concomitant brain abnormalities, allows the clinician to offer an optimistic prognosis to the family.