De novo variants in GABRA4 are associated with a neurological phenotype including developmental delay, behavioral abnormalities and epilepsy.

Nine out of 19 genes encoding GABAA receptor subunits have been linked to monogenic syndromes characterized by seizures and developmental disorders. Previously, we reported the de novo variant p.(Thr300Ile) in GABRA4 in a patient with epilepsy and neurodevelopmental abnormalities. However, no new cases have been reported since then. Through an international collaboration, we collected molecular and phenotype data of individuals carrying de novo variants in GABRA4. Patients and their parents were investigated either by exome or genome sequencing, followed by targeted Sanger sequencing in some cases. All variants within the transmembrane domain, including the previously reported p.(Thr300Ile) variant, were characterized in silico and analyzed by molecular dynamics (MD) simulation studies. We identified three novel de novo missense variants in GABRA4 (NM_000809.4): c.797 C > T, p.(Pro266Leu), c.899 C > A, p.(Thr300Asn), and c.634 G > A, p.(Val212Ile). The p.(Thr300Asn) variant impacts the same codon as the previously reported variant p.(Thr300Ile) and likely arose post-zygotically as evidenced by sequencing oral mucosal cells. Overlapping phenotypes among affected individuals included developmental delay (4/4), epileptiform EEG abnormalities (3/4), attention deficits (3/4), seizures (2/4), autistic features (2/4) and structural brain abnormalities (2/4). MD simulations of the three variants within the transmembrane domain of the receptor indicate that sub-microsecond scale dynamics differ between wild-type and mutated subunits. Taken together, our findings further corroborate an association between GABRA4 and a neurological phenotype including variable neurodevelopmental, behavioral and epileptic abnormalities.

Pathogenic variants in SMARCA1 cause an X-linked neurodevelopmental disorder modulated by NURF complex composition.

Pathogenic variants in ATP-dependent chromatin remodeling proteins are a recurrent cause of neurodevelopmental disorders (NDDs). The NURF complex consists of BPTF and either the SNF2H (SMARCA5) or SNF2L (SMARCA1) ISWI-chromatin remodeling enzyme. Pathogenic variants in BPTF and SMARCA5 were previously implicated in NDDs. Here, we describe 40 individuals from 30 families with de novo or maternally inherited pathogenic variants in SMARCA1. This novel NDD was associated with mild to severe ID/DD, delayed or regressive speech development, and some recurrent facial dysmorphisms. Individuals carrying SMARCA1 loss-of-function variants exhibited a mild genome-wide DNA methylation profile and a high penetrance of macrocephaly. Genetic dissection of the NURF complex using Smarca1, Smarca5, and Bptfsingle and double mouse knockouts revealed the importance of NURF composition and dosage for proper forebrain development. Finally, we propose that genetic alterations affecting different NURF components result in a NDD with a broad clinical spectrum.

Report of two cases of Schaaf-Yang syndrome: Same genotype and different phenotype.

We report two, genotypically identical but phenotypically distinct cases of Schaaf-Yang syndrome and propose the early use of Genome Sequencing in patients with nonspecific presentations to facilitate the early diagnosis of children with rare genetic diseases and improve overall health care outcomes.

Hemizygous variants in protein phosphatase 1 regulatory subunit 3F (PPP1R3F) are associated with a neurodevelopmental disorder characterized by developmental delay, intellectual disability and autistic features.

Protein phosphatase 1 regulatory subunit 3F (PPP1R3F) is a member of the glycogen targeting subunits (GTSs), which belong to the large group of regulatory subunits of protein phosphatase 1 (PP1), a major eukaryotic serine/threonine protein phosphatase that regulates diverse cellular processes. Here, we describe the identification of hemizygous variants in PPP1R3F associated with a novel X-linked recessive neurodevelopmental disorder in 13 unrelated individuals. This disorder is characterized by developmental delay, mild intellectual disability, neurobehavioral issues such as autism spectrum disorder, seizures and other neurological findings including tone, gait and cerebellar abnormalities. PPP1R3F variants segregated with disease in affected hemizygous males that inherited the variants from their heterozygous carrier mothers. We show that PPP1R3F is predominantly expressed in brain astrocytes and localizes to the endoplasmic reticulum in cells. Glycogen content in PPP1R3F knockout astrocytoma cells appears to be more sensitive to fluxes in extracellular glucose levels than in wild-type cells, suggesting that PPP1R3F functions in maintaining steady brain glycogen levels under changing glucose conditions. We performed functional studies on nine of the identified variants and observed defects in PP1 binding, protein stability, subcellular localization and regulation of glycogen metabolism in most of them. Collectively, the genetic and molecular data indicate that deleterious variants in PPP1R3F are associated with a new X-linked disorder of glycogen metabolism, highlighting the critical role of GTSs in neurological development. This research expands our understanding of neurodevelopmental disorders and the role of PP1 in brain development and proper function.

Clinical, biochemical and molecular characterization of 12 patients with pyruvate carboxylase deficiency treated with triheptanoin.

Pyruvate carboxylase (PC) deficiency is a rare autosomal recessive mitochondrial neurometabolic disorder of energy deficit resulting in high morbidity and mortality, with limited therapeutic options. The PC homotetramer has a critical role in gluconeogenesis, anaplerosis, neurotransmitter synthesis, and lipogenesis. The main biochemical and clinical findings in PC deficiency (PCD) include lactic acidosis, ketonuria, failure to thrive, and neurological dysfunction. Use of the anaplerotic agent triheptanoin on a limited number of individuals with PCD has had mixed results. We expand on the potential utility of triheptanoin in PCD by examining the clinical, biochemical, molecular, and health-related quality-of-life (HRQoL) findings in a cohort of 12 individuals with PCD (eight with Type A and two each with Types B and C) treated with triheptanoin ranging for 6 days to about 7 years. The main endpoints were changes in blood lactate and HRQoL scores, but collection of useful data was limited to about half of subjects. An overall trend of lactate reduction with time on triheptanoin was noted, but with significant variability among subjects and only one subject reaching close to statistical significance for this endpoint. Parent reported HRQoL assessments with treatment showed mixed results, with some subjects showing no change, some improvement, and some worsening of overall scores. Subjects with buried amino acids in the pyruvate carboxyltransferase domain of PC that undergo destabilizing replacements may be more likely to respond (with lactate reduction or HRQoL improvement) to triheptanoin compared to those with replacements that disrupt tetramerization or subunit-subunit interface contacts. The reason for this difference is unclear and requires further validation. We observed significant variability but an overall trend of lactate reduction with time on triheptanoin and mixed parent reported outcome changes by HRQoL assessments for subjects with PCD on long-term triheptanoin. The mixed results noted with triheptanoin therapy in this study could be due to endpoint data limitation, variability of disease severity between subjects, limitation of the parent reported HRQoL tool, or subject genotype variability. Alternative designed trials and more study subjects with PCD will be needed to validate important observations from this work.

Erratum: Discovery of a neuromuscular syndrome caused by biallelic variants in ASCC3.

[This corrects the article DOI: 10.1016/j.xhgg.2021.100024.].

Real-world economic evaluation of prospective rapid whole-genome sequencing compared to a matched retrospective cohort of critically ill pediatric patients in the United States.

There is an increasing demand for supporting the adoption of rapid whole-genome sequencing (rWGS) by demonstrating its real-world value. We aimed to assess the cost-effectiveness of rWGS in critically ill pediatric patients with diseases of unknown cause. Data were collected prospectively of patients admitted to the Nicklaus Children's Hospital's intensive care units from March 2018 to September 2020, with rWGS (N = 65). Comparative data were collected in a matched retrospective cohort with standard diagnostic genetic testing. We determined total costs, diagnostic yield (DY), and incremental cost-effectiveness ratio (ICER) adjusted for selection bias and right censoring. Sensitivity analyses explored the robustness of ICER through bootstrapping. rWGS resulted in a diagnosis in 39.8% while standard testing in 13.5% (p = 0.026). rWGS resulted in a mean saving per person of $100,440 (SE = 26,497, p < 0.001) and a total of $6.53 M for 65 patients. rWGS in critically ill pediatric patients is cost-effective, cost-saving, shortens diagnostic odyssey, and triples the DY of traditional approaches.

Clinico-radiological features, molecular spectrum, and identification of prognostic factors in developmental and epileptic encephalopathy due to inosine triphosphate pyrophosphatase (ITPase) deficiency.

Developmental and epileptic encephalopathy 35 (DEE 35) is a severe neurological condition caused by biallelic variants in ITPA, encoding inosine triphosphate pyrophosphatase, an essential enzyme in purine metabolism. We delineate the genotypic and phenotypic spectrum of DEE 35, analyzing possible predictors for adverse clinical outcomes. We investigated a cohort of 28 new patients and reviewed previously described cases, providing a comprehensive characterization of 40 subjects. Exome sequencing was performed to identify underlying ITPA pathogenic variants. Brain MRI (magnetic resonance imaging) scans were systematically analyzed to delineate the neuroradiological spectrum. Survival curves according to the Kaplan-Meier method and log-rank test were used to investigate outcome predictors in different subgroups of patients. We identified 18 distinct ITPA pathogenic variants, including 14 novel variants, and two deletions. All subjects showed profound developmental delay, microcephaly, and refractory epilepsy followed by neurodevelopmental regression. Brain MRI revision revealed a recurrent pattern of delayed myelination and restricted diffusion of early myelinating structures. Congenital microcephaly and cardiac involvement were statistically significant novel clinical predictors of adverse outcomes. We refined the molecular, clinical, and neuroradiological characterization of ITPase deficiency, and identified new clinical predictors which may have a potentially important impact on diagnosis, counseling, and follow-up of affected individuals.

Clinical and molecular features of 66 patients with musculocontractural Ehlers-Danlos syndrome caused by pathogenic variants in CHST14 (mcEDS-CHST14).

BACKGROUND: Musculocontractural Ehlers-Danlos syndrome is caused by biallelic loss-of-function variants in CHST14 (mcEDS-CHST14) or DSE (mcEDS-DSE). Although 48 patients in 33 families with mcEDS-CHST14 have been reported, the spectrum of pathogenic variants, accurate prevalence of various manifestations and detailed natural history have not been systematically investigated. METHODS: We collected detailed and comprehensive clinical and molecular information regarding previously reported and newly identified patients with mcEDS-CHST14 through international collaborations. RESULTS: Sixty-six patients in 48 families (33 males/females; 0-59 years), including 18 newly reported patients, were evaluated. Japanese was the predominant ethnicity (27 families), associated with three recurrent variants. No apparent genotype-phenotype correlation was noted. Specific craniofacial (large fontanelle with delayed closure, downslanting palpebral fissures and hypertelorism), skeletal (characteristic finger morphologies, joint hypermobility, multiple congenital contractures, progressive talipes deformities and recurrent joint dislocation), cutaneous (hyperextensibility, fine/acrogeria-like/wrinkling palmar creases and bruisability) and ocular (refractive errors) features were observed in most patients (>90%). Large subcutaneous haematomas, constipation, cryptorchidism, hypotonia and motor developmental delay were also common (>80%). Median ages at the initial episode of dislocation or large subcutaneous haematoma were both 6 years. Nine patients died; their median age was 12 years. Several features, including joint and skin characteristics (hypermobility/extensibility and fragility), were significantly more frequent in patients with mcEDS-CHST14 than in eight reported patients with mcEDS-DSE. CONCLUSION: This first international collaborative study of mcEDS-CHST14 demonstrated that the subtype represents a multisystem disorder with unique set of clinical phenotypes consisting of multiple malformations and progressive fragility-related manifestations; these require lifelong, multidisciplinary healthcare approaches.

ITSN1: a novel candidate gene involved in autosomal dominant neurodevelopmental disorder spectrum.

ITSN1 plays an important role in brain development. Recent studies in large cohorts of subjects with neurodevelopmental disorders have identified de novo variants in ITSN1 gene thereby suggesting that this gene is involved in the development of such disorders. The aim of this study is to provide further proof of such a link. We performed trio exome sequencing in a patient presenting autism, intellectual disability, and severe behavioral difficulties. Additional affected patients with a neurodevelopmental disorder harboring a heterozygous variant in ITSN1 (NM_003024.2) were collected through a worldwide collaboration. All patients underwent detailed phenotypic and genetic assessment and data was collected and shared by healthcare givers. We identified ten novel patients from eight families with heterozygous truncating or missense variants in ITSN1 gene. In addition, four previously published patients from large meta-analysis studies were included. In total, 7/14 patients presented a de novo variant in ITSN1. All patients showed neurodevelopmental disorders from autism spectrum disorders (90%), intellectual disability (86%), and epilepsy (30%). We demonstrated that truncating variants are in the first half of ITSN1 whereas missense variants are clustered in C-terminal region. We suggest ITSN1 gene is involved in development of an autism spectrum disorder with variable additional neurodevelopmental deficiency, thus confirming the hypothesis that ITSN1 is important for brain development.

Case Report: The Association of Wilson Disease in a Patient With Ataxia and GLUT-1 Deficiency.

Background: Wilson disease (WD) and glucose transporter type 1 (GLUT1) deficiency syndrome are two syndromes with different modes of inheritance but share certain similarities on neurological presentation. To date we have not found previous reports of an association between these two disorders. Case Presentation: Here we describe a 9-year-old male with global developmental delay that presented with intermittent and sudden onset weakness that first occurred at age 3. He was diagnosed with a mutation in the SLC2A1 (Solute Carrier Family 2 Member 1) gene, which results in GLUT1 deficiency. A ketogenic diet could not be started because of unexplained elevated liver enzymes. Due to his liver enzymes' persistent elevation, further investigations demonstrated mildly decreased ceruloplasmin levels, high basal 24-h urinary copper excretion, and an elevated hepatic parenchymal copper concentration on liver biopsy, consistent with WD. Genetic testing revealed two separate mutations in the ATP7B (ATPase Copper Transporting Beta) gene, consistent with WD. The patient was treated with a low copper diet, zinc acetate, and trientine hydrochloride. When liver enzymes normalized, he was subsequently started on a ketogenic diet with improvement in neurological symptoms. His neurological symptoms were most likely secondary to GLUT1 deficiency syndrome, as WD's neurological symptoms are primarily observed in the second decade of life. Conclusion: Recent studies have demonstrated the importance of genetic testing upon unexplained persistent elevation of liver enzymes. This case highlights the importance of carefully evaluating a patient with an unexplained liver disorder, even in the presence of primary neurological disease, as it can have significant therapeutic implications.

UBA2 variants underlie a recognizable syndrome with variable aplasia cutis congenita and ectrodactyly.

PURPOSE: The human chromosome 19q13.11 deletion syndrome is associated with a variable phenotype that includes aplasia cutis congenita (ACC) and ectrodactyly as specific features. UBA2 (ubiquitin-like modifier-activating enzyme 2) lies adjacent to the minimal deletion overlap region. We aimed to define the UBA2-related phenotypic spectrum in humans and zebrafish due to sequence variants and to establish the mechanism of disease. METHODS: Exome sequencing was used to detect UBA2 sequence variants in 16 subjects in 7 unrelated families. uba2 loss of function was modeled in zebrafish. Effects of human missense variants were assessed in zebrafish rescue experiments. RESULTS: Seven human UBA2 loss-of-function and missense sequence variants were detected. UBA2-phenotypes included ACC, ectrodactyly, neurodevelopmental abnormalities, ectodermal, skeletal, craniofacial, cardiac, renal, and genital anomalies. uba2 was expressed in zebrafish eye, brain, and pectoral fins; uba2-null fish showed deficient growth, microcephaly, microphthalmia, mandibular hypoplasia, and abnormal fins. uba2-mRNAs with human missense variants failed to rescue nullizygous zebrafish phenotypes. CONCLUSION: UBA2 variants cause a recognizable syndrome with a wide phenotypic spectrum. Our data suggest that loss of UBA2 function underlies the human UBA2 monogenic disorder and highlights the importance of SUMOylation in the development of affected tissues.

Clinical, neuroimaging, and molecular spectrum of TECPR2-associated hereditary sensory and autonomic neuropathy with intellectual disability.

Bi-allelic TECPR2 variants have been associated with a complex syndrome with features of both a neurodevelopmental and neurodegenerative disorder. Here, we provide a comprehensive clinical description and variant interpretation framework for this genetic locus. Through international collaboration, we identified 17 individuals from 15 families with bi-allelic TECPR2-variants. We systemically reviewed clinical and molecular data from this cohort and 11 cases previously reported. Phenotypes were standardized using Human Phenotype Ontology terms. A cross-sectional analysis revealed global developmental delay/intellectual disability, muscular hypotonia, ataxia, hyporeflexia, respiratory infections, and central/nocturnal hypopnea as core manifestations. A review of brain magnetic resonance imaging scans demonstrated a thin corpus callosum in 52%. We evaluated 17 distinct variants. Missense variants in TECPR2 are predominantly located in the N- and C-terminal regions containing ß-propeller repeats. Despite constituting nearly half of disease-associated TECPR2 variants, classifying missense variants as (likely) pathogenic according to ACMG criteria remains challenging. We estimate a pathogenic variant carrier frequency of 1/1221 in the general and 1/155 in the Jewish Ashkenazi populations. Based on clinical, neuroimaging, and genetic data, we provide recommendations for variant reporting, clinical assessment, and surveillance/treatment of individuals with TECPR2-associated disorder. This sets the stage for future prospective natural history studies.

Chromosomal microarray detects genetic risks of neurodevelopmental disorders in newborns with congenital heart disease.

OBJECTIVE: To compare the genetic testing results of neonates with CHD by chromosomal microarray to karyotyping and fluorescence in situ hybridisation analysis. METHODS: This was a single-centre retrospective comparative study of patients with CHD and available genetic testing results admitted to the cardiac ICU between January, 2004 and December, 2017. Patients from 2004 to 2010 were tested by karyotyping and fluorescence in situ hybridisation analysis, while patients from 2012 to 2017 were analysed by chromosomal microarray. RESULTS: Eight-hundred and forty-nine neonates with CHD underwent genetic testing, 482 by karyotyping and fluorescence in situ hybridization, and 367 by chromosomal microarray. In the karyotyping and fluorescence in situ hybridisation analysis group, 86/482 (17.8%) had genetic abnormalities detected, while in the chromosomal microarray group, 135/367 (36.8%) had genetic abnormalities detected (p < 0.00001). Of patients with abnormal chromosomal microarray results, 41/135 (30.4%) had genetic abnormality associated with neurodevelopmental disorders that were exclusively identified by chromosomal microarray. Conotruncal abnormalities were the most common diagnosis in both groups, with karyotyping and fluorescence in situ hybridisation analysis detecting genetic abnormalities in 26/160 (16.3%) patients and chromosomal microarray detecting abnormalities in 41/135 (30.4%) patients (p = 0.004). In patients with d-transposition of the great arteries, 0/68 (0%) were found to have genetic abnormalities by karyotyping and fluorescence in situ hybridisation compared to 7/54 (13.0%) by chromosomal microarray. CONCLUSIONS: Chromosomal microarray identified patients with CHD at genetic risk of neurodevelopmental disorders, allowing earlier intervention with multidisciplinary care and more accurate pre-surgical prognostic counselling.

Uridine-responsive epileptic encephalopathy due to inherited variants in CAD: A Tale of Two Siblings.

We report two siblings with intractable epilepsy, developmental regression, and progressive cerebellar atrophy due to biallelic variants in the gene CAD. For the affected girl, uridine started at age 5 resulted in dramatic improvements in seizure control and development, cessation of cerebellar atrophy, and resolution of hematological abnormalities. Her older brother had a more severe course and only modest response to uridine started at 14 years old. Treatment of this progressive condition via uridine supplementation provides an example of precision diagnosis and treatment using clear outcome measures and biomarkers to monitor efficacy.

Discovery of a neuromuscular syndrome caused by biallelic variants in ASCC3.

Activating Signal Cointegrator 1 Complex, Subunit 3 (ASCC3) is part of the four-part ASC-1 transcriptional cointegrator complex. This complex includes ASCC1 (associated with spinal muscular atrophy with congenital bone fractures 2), TRIP4 (associated with spinal muscular atrophy with congenital bone fractures 1), and ASCC2 (not yet associated with human disease.) ASCC3 encodes a DNA helicase responsible for generating single-stranded DNA as part of the DNA damage response. Interestingly, ASCC3 expresses coding and non-coding isoforms, which act in opposition to balance the recovery of gene transcription after UV-induced DNA damage. Here we report the discovery of ASCC3 as the cause of a neuromuscular syndrome in seven unreported individuals from six unrelated families and updates on the one previously reported family. All the individuals share a neurologic phenotype that ranges from severe developmental delay to muscle fatigue. There appears to be genotype-phenotype correlation, as the most mildly affected individual is homozygous for a rare missense variant, while the more severely affected individuals are compound heterozygotes for a missense and a presumed loss-of-function (LOF) variant. There are no individuals with biallelic presumed LOF variants in our cohort or in gnomAD, as this genotype may not be compatible with life. In summary we report a syndrome in these eleven individuals from seven families with biallelic variants in ASCC3.

A Case of UDP-Galactose 4'-Epimerase Deficiency Associated with Dyshematopoiesis and Atrioventricular Valve Malformations: An Exceptional Clinical Phenotype Explained by Altered N-Glycosylation with Relative Preservation of the Leloir Pathway.

The generalized form of UDP-galactose-4'-epimerase (GALE) deficiency causes hypotonia, failure to thrive, cataracts, and liver failure. Individuals with non-generalized forms may remain asymptomatic with uncertain long-term outcomes. We report a 2-year-old child compound heterozygous for GALE p.R51W/p.G237D who never developed symptoms of classic galactosemia but has a history of congenital combined mitral and tricuspid valve malformation and pyloric stenosis, and presented with pancytopenia. Variant pathogenicity was supported by predictive computational tools and decreased GALE activity measured in erythrocytes. GALE function extends to the biosynthesis of glycans by epimerization of UDP-N-acetyl-galactosamine and -glucosamine. Interrogation of the Gene Ontology consortium database revealed several putative proteins involved in normal hematopoiesis and atrioventricular valve morphogenesis, requiring N-glycosylation for adequate functionality. We hypothesize that by limiting substrate supply due to GALE deficiency, alterations in N-linked protein glycosylation can explain the patient's phenotype.

De novo substitutions of TRPM3 cause intellectual disability and epilepsy.

The developmental and epileptic encephalopathies (DEE) are a heterogeneous group of chronic encephalopathies frequently associated with rare de novo nonsynonymous coding variants in neuronally expressed genes. Here, we describe eight probands with a DEE phenotype comprising intellectual disability, epilepsy, and hypotonia. Exome trio analysis showed de novo variants in TRPM3, encoding a brain-expressed transient receptor potential channel, in each. Seven probands were identically heterozygous for a recurrent substitution, p.(Val837Met), in TRPM3's S4-S5 linker region, a conserved domain proposed to undergo conformational change during gated channel opening. The eighth individual was heterozygous for a proline substitution, p.(Pro937Gln), at the boundary between TRPM3's flexible pore-forming loop and an adjacent alpha-helix. General-population truncating variants and microdeletions occur throughout TRPM3, suggesting a pathomechanism other than simple haploinsufficiency. We conclude that de novo variants in TRPM3 are a cause of intellectual disability and epilepsy.

Variants in TCF20 in neurodevelopmental disability: description of 27 new patients and review of literature.

PURPOSE: To define the clinical characteristics of patients with variants in TCF20, we describe 27 patients, 26 of whom were identified via exome sequencing. We compare detailed clinical data with 17 previously reported patients. METHODS: Patients were ascertained through molecular testing laboratories performing exome sequencing (and other testing) with orthogonal confirmation; collaborating referring clinicians provided detailed clinical information. RESULTS: The cohort of 27 patients all had novel variants, and ranged in age from 2 to 68 years. All had developmental delay/intellectual disability. Autism spectrum disorders/autistic features were reported in 69%, attention disorders or hyperactivity in 67%, craniofacial features (no recognizable facial gestalt) in 67%, structural brain anomalies in 24%, and seizures in 12%. Additional features affecting various organ systems were described in 93%. In a majority of patients, we did not observe previously reported findings of postnatal overgrowth or craniosynostosis, in comparison with earlier reports. CONCLUSION: We provide valuable data regarding the prognosis and clinical manifestations of patients with variants in TCF20.

Novel Loss of Function in the AGK Gene: Rare Cause of End-Stage Heart Failure.

The authors present a case of mitochondrial cardiomyopathy due to a novel mutation of AGK gene that led to progressive heart failure. The cardiac magnetic resonance image findings of diffusely elevated relaxation time and increase in extracellular volume in the myocardium without early or late gadolinium enhancement may suggest mitochondrial cardiomyopathy. The authors emphasized the multidisciplinary team approach in the care of patients with mitochondrial cardiomyopathies. (Level of Difficulty: Advanced.).