Clinical, genetic, and cognitive correlates of seizure occurrences in Phelan-McDermid syndrome.

BACKGROUND: Phelan-McDermid syndrome (PMS) is a genetic neurodevelopmental disorder caused by SHANK3 haploinsufficiency and is associated with an increased risk for seizures. Previous literature indicates that around one third of individuals with PMS also have epilepsy or seizures, with a wide range of types and ages of onset. Investigating the impact of seizures on intellectual and adaptive functioning for PMS is a primary concern for caregivers and is important to understanding the natural history of this syndrome. METHODS: We report on results from 98 individuals enrolled in a prospective, longitudinal study. We detailed seizure frequency, type, and age of onset, and we analyzed seizure occurrence with best estimate IQ, adaptive functioning, clinical features, and genotype. We conducted multiple linear regression analyses to assess the relationship between the presence of seizures and the Vineland Adaptive Behavior Scale, Second Edition (VABS-II) Adaptive Behavior Composite score and the best estimate full-scale IQ. We also performed Chi-square tests to explore associations between seizure prevalence and genetic groupings. Finally, we performed Chi-square tests and t-tests to explore the relationship between seizures and demographic features, features that manifest in infancy, and medical features. RESULTS: Seizures were present in 41% of the cohort, and age of onset was widely variable. The presence of seizures was associated with significantly lower adaptive and intellectual functioning. Genotype-phenotype analyses were discrepant, with no differences in seizure prevalence across genetic classes, but with more genes included in deletions of participants with 22q13 deletions and seizures compared to those with 22q13 deletions and no seizures. No clinical associations were found between the presence of seizures and sex, history of pre- or neonatal complications, early infancy, or medical features. In this cohort, generalized seizures were associated with developmental regression, which is a top concern for PMS caregivers. CONCLUSIONS: These results begin to eludicate correlates of seizures in individuals with PMS and highlight the importance of early seizure management. Importantly, presence of seizures was associated with adaptive and cognitive functioning. A larger cohort might be able to identify additional associations with medical features. Genetic findings suggest an increased capability to realize genotype-phenotype relationships when deletion size is taken into account.

Altered Hippocampal Activation in Seizure-Prone CACNA2D2 Knock-out Mice.

The voltage-gated calcium channel subunit α2δ-2 controls calcium-dependent signaling in neurons, and loss of this subunit causes epilepsy in both mice and humans. To determine whether mice without α2δ-2 demonstrate hippocampal activation or histopathological changes associated with seizure activity, we measured expression of the activity-dependent gene c-fos and various histopathological correlates of temporal lobe epilepsy (TLE) in hippocampal tissue from wild-type (WT) and α2δ-2 knock-out (CACNA2D2 KO) mice using immunohistochemical staining and confocal microscopy. Both genotypes demonstrated similarly sparse c-fos and ΔFosB expressions within the hippocampal dentate granule cell layer (GCL) at baseline, consistent with no difference in basal activity of granule cells between genotypes. Surprisingly, when mice were assayed 1 h after handling-associated convulsions, KO mice had fewer c-fos-positive cells but dramatically increased ΔFosB expression in the dentate gyrus compared with WT mice. After administration of a subthreshold pentylenetetrazol dose, however, KO mice dentate had significantly more c-fos expression compared with WT mice. Other histopathological markers of TLE in these mice, including markers of neurogenesis, glial activation, and mossy fiber sprouting, were similar between WT and KO mice, apart from a small but statistically significant increase in hilar mossy cell density, opposite to what is typically found in mice with TLE. This suggests that the differences in seizure-associated dentate gyrus function in the absence of α2δ-2 protein are likely due to altered functional properties of the network without associated structural changes in the hippocampus at the typical age of seizure onset.

Aneuploidy is Linked to Neurological Phenotypes Through Oxidative Stress.

Aneuploidy, having an aberrant genome, is gaining increasing attention in neurodegenerative diseases. It gives rise to proteotoxic stress as well as a stereotypical oxidative shift which makes these cells sensitive to internal and environmental stresses. A growing body of research from numerous laboratories suggests that many neurodegenerative disorders, especially Alzheimer's disease and frontotemporal dementia, are characterised by neuronal aneuploidy and the ensuing apoptosis, which may contribute to neuronal loss. Using Drosophila as a model, we investigated the effect of induced aneuploidy in GABAergic neurons. We found an increased proportion of aneuploidy due to Mad2 depletion in the third-instar larval brain and increased cell death. Depletion of Mad2 in GABAergic neurons also gave a defective climbing and seizure phenotype. Feeding animals an antioxidant rescued the climbing and seizure phenotype. These findings suggest that increased aneuploidy leads to higher oxidative stress in GABAergic neurons which causes cell death, climbing defects, and seizure phenotype. Antioxidant feeding represents a potential therapy to reduce the aneuploidy-driven neurological phenotype.

[Correlation between clinical phenotypes and genotypes among 46 children with SCN1A-related developmental epileptic encephalopathy].

OBJECTIVE: To explore the correlation between clinical phenotypes and genotypes among 46 children with SCN1A-related developmental epileptic encephalopathy (DEE). METHODS: Clinical data of 46 children with DEE and SCN1A variants identified at the Guangzhou Women and Children's Medical Center between January 2018 and June 2022 were collected. The children were grouped based on their age of onset, clinical manifestations, neurodevelopmental status, and results of genetic testing. The correlation between SCN1A genotypes and clinical phenotypes was analyzed. RESULTS: Among the 46 patients, 2 children (4.35%) had developed the symptoms before 3 months of age, 42 (91.30%) were between 3 to 9 months, and 2 cases (4.35%) were after 10 months. Two cases (4.35%) presented with epilepsy of infancy with migrating focal seizures (EIMFS), while 44 (95.7%) had presented with Dravet syndrome (DS), including 28 cases (63.6%) with focal onset (DS-F), 13 cases (29.5%) with myoclonic type (DS-M), 1 case (2.27%) with generalized type (DS-G), and 2 cases (4.55%) with status epilepticus type (DS-SE). Both of the two EIMFS children had severe developmental delay, and among the DS patients, 7 cases had normal development, while the remaining had developmental delay. A total of 44 variants were identified through genetic sequencing, which included 16 missense variants and 28 truncating variants. All EIMFS children had carried the c.677C>T (p.Thr226Met) missense variant. In the DS group, there was a significant difference in the age of onset between the missense variants group and the truncating variants group (P < 0.05). Missense variants were more common in D1 (7/15, 46.7%) and pore regions (8/15, 53.3%), while truncating variants were more common in D1 (12/28, 42.9%). Children with variants outside the pore region were more likely to develop myoclonic seizures. CONCLUSION: The clinical phenotypes of DEE are diverse. There is a difference in the age of onset between individuals with truncating and missense variants in the SCN1A gene. Missense variants outside the pore region are associated with a higher incidence of myoclonic seizures.

Expression of GCaMP6s in the dentate gyrus induces tonic-clonic seizures.

GCaMP is a genetically encoded calcium indicator (GECI) widely used in neuroscience research. It measures intracellular Ca2+ level by fluorescence changes as it directly binds to Ca2+. In this process, the effect of this calcium buffer on the intracellular calcium signaling and cell physiology is often not taken into consideration. However, growing evidence from calcium imaging studies shows GCaMP expression under certain conditions can generate aberrant activity, such as seizures. In this study, we examined the effect of GCaMP6 expression in the dentate gyrus (DG) on epileptogenesis. We found that viral expression of GCaMP6s but not GCaMP6f in the DG induces tonic-clonic seizures several weeks after viral injection. Cell-type specific expression of GCaMP6s revealed the granule cells (GCs) as the key player in GCaMP6s-induced epilepsy. Finally, by using slice electrophysiology, we demonstrated that GCaMP6s expression increases neuronal excitability in the GCs. Together, this study highlights the ability of GCaMP6s in DG-associated epileptogenesis.

A high seizure burden increases several prostaglandin species in the hippocampus of a Scn1a+/- mouse model of Dravet syndrome.

Dravet syndrome is an intractable epilepsy with a high seizure burden that is resistant to current anti-seizure medications. There is evidence that neuroinflammation plays a role in epilepsy and seizures, however few studies have specifically examined neuroinflammation in Dravet syndrome under conditions of a higher seizure burden. Here we used an established genetic mouse model of Dravet syndrome (Scn1a+/- mice), to examine whether a higher seizure burden impacts the number and morphology of microglia in the hippocampus. Moreover, we examined whether a high seizure burden influences classical inflammatory mediators in this brain region. Scn1a+/- mice with a high seizure burden induced by thermal priming displayed a localised reduction in microglial cell density in the granule cell layer and subgranular zone of the dentate gyrus, regions important to postnatal neurogenesis. However, microglial cell number and morphology remained unchanged in other hippocampal subfields. The high seizure burden in Scn1a+/- mice did not affect hippocampal mRNA expression of classical inflammatory mediators such as interleukin 1ß and tumour necrosis factor α, but increased cyclooxygenase 2 (COX-2) expression. We then quantified hippocampal levels of prostanoids that arise from COX-2 mediated metabolism of fatty acids and found that Scn1a+/- mice with a high seizure burden displayed increased hippocampal concentrations of numerous prostaglandins, notably PGF2α, PGE2, PGD2, and 6-K-PGF1A, compared to Scn1a+/- mice with a low seizure burden. In conclusion, a high seizure burden increased hippocampal concentrations of various prostaglandin mediators in a mouse model of Dravet syndrome. Future studies could interrogate the prostaglandin pathways to further better understand their role in the pathophysiology of Dravet syndrome.

The homeostatic effects of the RE-1 silencing transcription factor on cortical networks are altered under ictogenic conditions in the mouse.

AIM: The Repressor Element-1 Silencing Transcription Factor (REST) is an epigenetic master regulator playing a crucial role in the nervous system. In early developmental stages, REST downregulation promotes neuronal differentiation and the acquisition of the neuronal phenotype. In addition, postnatal fluctuations in REST expression contribute to shaping neuronal networks and maintaining network homeostasis. Here we investigate the role of the early postnatal deletion of neuronal REST in the assembly and strength of excitatory and inhibitory synaptic connections. METHODS: We investigated excitatory and inhibitory synaptic transmission by patch-clamp recordings in acute neocortical slices in a conditional knockout mouse model (RestGTi) in which Rest was deleted by delivering PHP.eB adeno-associated viruses encoding CRE recombinase under the control of the human synapsin I promoter in the lateral ventricles of P0-P1 pups. RESULTS: We show that, under physiological conditions, Rest deletion increased the intrinsic excitability of principal cortical neurons in the primary visual cortex and the density and strength of excitatory synaptic connections impinging on them, without affecting inhibitory transmission. Conversely, in the presence of a pathological excitation/inhibition imbalance induced by pentylenetetrazol, Rest deletion prevented the increase in synaptic excitation and decreased seizure severity. CONCLUSION: The data indicate that REST exerts distinct effects on the excitability of cortical circuits depending on whether it acts under physiological conditions or in the presence of pathologic network hyperexcitability. In the former case, REST preserves a correct excitatory/inhibitory balance in cortical circuits, while in the latter REST loses its homeostatic activity and may become pro-epileptogenic.

Genotype and phenotype features and prognostic factors of neonatal-onset pyridoxine-dependent epilepsy: A systematic review.

Pyridoxine-dependent epilepsy (PDE-ALDH7A1) is a rare autosomal recessive disorder due to a deficiency of α-aminoadipic semialdehyde dehydrogenase. This study aimed to systematically explore genotypic and phenotypic features and prognostic factors of neonatal-onset PDE. A literature search covering PubMed, Elsevier, and Web of Science was conducted from January 2006 to August 2023. We identified 56 eligible studies involving 169 patients and 334 alleles. The c.1279 G>C variant was the most common variant of neonatal-onset PDE (25.7 %). All patients were treated with pyridoxine; forty patients received dietary intervention therapy. 63.9 % of the patients were completely seizure-free; however, 68.6 % of the patients had neurodevelopmental delays. Additionally, homozygous c.1279 G>C variants were significantly associated with ventriculomegaly, abnormal white matter signal, and cysts (P<0.05). In contrast, homozygous c.1364 T>C was associated with clonic seizure (P=0.031). Pyridoxine used immediately at seizure onset was an independent protective factor for developmental delay (P=0.035; odds ratio [OR]: 3.14). Besides, pyridoxine used early in the neonatal period was a protective factor for language delay (P=0.044; OR: 4.59). In contrast, neonatal respiratory distress (P=0.001; OR: 127.44) and abnormal brain magnetic resonance imaging (P=0.049; OR: 3.64) were risk factors. Prenatal movement abnormality (P=0.041; OR: 20.56) and abnormal white matter signal (P=0.012; OR: 24.30) were risk factors for motor delay. Myoclonic seizure (P=0.023; OR: 7.13) and status epilepticus (P=0.000; OR: 9.93) were risk factors for breakthrough seizures. In conclusion, our study indicated that pyridoxine should be started immediately when unexplained neonatal seizures occur and not later than the neonatal period to prevent poor neurodevelopmental outcomes.

Association of Clinical, Tumor, and Treatment Characteristics With Seizure Control in Patients With IDH1/2-Mutant Lower-Grade Glioma.

BACKGROUND AND OBJECTIVES: Patients with IDH1/2-mutant lower-grade glioma have a high frequency of seizures. We aimed to investigate the correlations between seizures and tumor/patient characteristics and the impact of surgery and adjuvant treatments (AT) on seizure control along the disease trajectory. METHODS: We retrospectively included patients with IDH1/2-mutant lower-grade glioma who underwent surgery at the neurosurgery divisions of the University of Turin and Milan and were treated at the Division of Neuro-Oncology of Turin. Inclusion criteria were a diagnosis according to the 2021 WHO Classification and presentation with seizures; exclusion criteria were presence of CDKN2A/B homozygous deletion, intense/ring contrast enhancement on MRI at presentation, and small tissue biopsy. We evaluated seizure freedom for 2 months after surgery, 6 months from starting observation or AT, at recurrence, and for 6 months after treatments of recurrence. RESULTS: We included 150 patients. There were 77 (51%) and 31 (21%) patients with IDH-mutant/1p19q-codeleted grade 2 and 3 oligodendroglioma and 30 (20%) and 12 (8%) with IDH-mutant grade 2 and 3 astrocytoma, respectively. Total resection was accomplished in 68 (45%). Seventy-five patients (50%) received AT while the remaining 75 were observed with MRI. After 6 months after AT, 28 of 29 patients (96.5%) displayed seizure reduction, 5 of 28 (18%) being seizure-free. 66 of 124 patients (53%) had seizures at recurrence. After 6 months after second-line treatments, 60 of 66 patients (91%) had seizure reduction, 11 (17%) being seizure-free. In multivariable analyses, grade 3 histology positively correlated with seizure freedom at 2 months after surgery (OR 3.5, 1.4-8.9, p = 0.008), 6 months after AT (OR 9.0, 1.5-54.9, p = 0.017), and 6 months after treatment of recurrence (OR 4.9, 1.5-16.5, p = 0.009). Adjuvant radiotherapy reduced seizures at recurrence in a univariate analysis (OR 0.14, 0.03-0.7, p = 0.020). Patients with seizure freedom after surgery and AT displayed longer progression-free survival (PFS) (65, 24.5-105, vs 48 months, 32-63.5, p = 0.037). DISCUSSION: This study analyzed seizure control in patients with IDH1/2-mutant lower-grade glioma across multiple time points. Grade 3 correlated with better seizure control throughout the entire disease trajectory, and seizure freedom after surgery and AT correlated with a longer PFS regardless of tumor grade. These results could serve as an external control arm in clinical trials evaluating the efficacy on seizures of antitumor agents in patients with IDH-mutant lower-grade glioma.

[Genetic analysis of a child with Multiple congenital anomalies-hypotonia-seizures syndrome 1 due to variant of PIGN gene].

OBJECTIVE: To analyze the clinical phenotype and genetic etiology of a child with Multiple congenital anomalies-hypotonia-seizures syndrome 1 (MCAHS1). METHODS: Clinical data of a 2-year-old boy who had presented at the Affiliated Hospital of Qingdao University in March 2023 for "intermittent limb twitching for 2 years" was collected. Peripheral blood samples were collected from the child and his parents for whole-exome sequencing (WES). Candidate variants were verified by Sanger sequencing and bioinformatic analysis based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). RESULTS: The child had manifested with distinctive facial features, limb deformities, hypotonia, motor and intellectual delays, and epileptic seizures. WES revealed that he has harbored compound heterozygous variants of the PIGN gene, namely c.963G>A (p.Q321=) and c.994A>T (p.I332F), which were inherited from his phenotypically normal mother and father, respectively. Based on the ACMG guidelines, the c.963G>A was classified as a pathogenic variant (PVS1+PM2_Supporting+PM3), whilst the c.994A>T was classified as a variant of uncertain significance (PM2_Supporting+PP3). CONCLUSION: Above discovery has expanded the mutational spectrum of the PIGN gene variants associated with MCAHS1, which may facilitate delineation of its genotype-phenotype correlation.

Inhibition of Granule Cell Dispersion and Seizure Development by Astrocyte Elevated Gene-1 in a Mouse Model of Temporal Lobe Epilepsy.

Although granule cell dispersion (GCD) in the hippocampus is known to be an important feature associated with epileptic seizures in temporal lobe epilepsy (TLE), the endogenous molecules that regulate GCD are largely unknown. In the present study, we have examined whether there is any change in AEG-1 expression in the hippocampus of a kainic acid (KA)-induced mouse model of TLE. In addition, we have investigated whether the modulation of astrocyte elevated gene-1 (AEG-1) expression in the dentate gyrus (DG) by intracranial injection of adeno-associated virus 1 (AAV1) influences pathological phenotypes such as GCD formation and seizure susceptibility in a KA-treated mouse. We have identified that the protein expression of AEG-1 is upregulated in the DG of a KA-induced mouse model of TLE. We further demonstrated that AEG-1 upregulation by AAV1 delivery in the DG-induced anticonvulsant activities such as the delay of seizure onset and inhibition of spontaneous recurrent seizures (SRS) through GCD suppression in the mouse model of TLE, while the inhibition of AEG-1 expression increased susceptibility to seizures. The present observations suggest that AEG-1 is a potent regulator of GCD formation and seizure development associated with TLE, and the significant induction of AEG-1 in the DG may have therapeutic potential against epilepsy.

Radiprodil, a selective GluN2B negative allosteric modulator, rescues audiogenic seizures in mice carrying the GluN2A(N615S) mutation.

BACKGROUND AND PURPOSE: GRIN-related disorders are neurodevelopmental disorders caused by mutations in N-methyl-D-aspartate receptor (NMDAR) subunit genes. A large fraction of these mutations lead to a 'gain of function' (GoF) of the NMDAR. Patients present with a range of symptoms including epilepsy, intellectual disability, behavioural and motor. Controlling seizures is a significant unmet medical need in most patients with GRIN-related disorders. Although several hundred GRIN mutations have been identified in humans, until recently none of the mouse models carrying Grin mutations/deletions showed an epileptic phenotype. The two recent exceptions both carry mutations of GluN2A. The aim of this study was to assess the efficacy of radiprodil, a selective negative allosteric modulator of GluN2B-containing NMDARs, in counteracting audiogenic seizures (AGS) in a murine model carrying the GluN2A(N615S) homozygous mutation (Grin2aS/S mice). EXPERIMENTAL APPROACH: Grin2aS/S mice were acutely treated with radiprodil at different doses before the presentation of a high-frequency acoustic stimulus commonly used for AGS induction. KEY RESULTS: Radiprodil significantly and dose-dependently reduced the onset and severity of AGS in Grin2aS/S mice. Surprisingly, the results revealed a sex-dependent difference in AGS susceptibility and in the dose-dependent protection of radiprodil in the two genders. Specifically, radiprodil was more effective in female versus male mice. CONCLUSION AND IMPLICATIONS: Overall, our data clearly show that radiprodil, a GluN2B selective negative allosteric modulator, may have the potential to control seizures in patients with GRIN2A GoF mutations. Further studies are warranted to better understand the sex-dependent effects observed in this study.

Yield and Utility of Routine Epilepsy Panel Genetic Testing Among Young Patients With Seizures.

Objective: We examined the yield of routine epilepsy panel genetic testing in pediatric patients. Methods: We retrospectively reviewed epilepsy genetic panel results routinely performed in the hospital or clinic on patients <8 years old from July 2021 to July 2023. We evaluated demographics, family history, seizure type, severity, and frequency, development, tone and movement abnormalities, dysmorphism, and electroencephalography (EEG) or magnetic resonance imaging (MRI) results as predictors of results. Results: 65 patients were included with mean age 4.5 years. Sixty percent of patients were male; 11 patients had pathogenic variants (16.9%), 7 were carriers for autosomal recessive conditions (10.8%), 36 had variants of uncertain significance (55.4%), and 11 tested negative (16.9%). Pathogenic variants and variants of uncertain significance were unassociated with demographics, clinical features, imaging, or family history. Conclusion: Variants identified have potential implications for treatment (SCN1), comorbidity screening (TSC1), reproduction (ATAD1, PSAT1, and CLN8), and prognostication (FOXG1). Patients not routinely screened for a genetic cause of epilepsy by our standard practices had clinically relevant results.

Emergence of lingual dystonia and strabismus in early-onset SCN8A self-limiting familial infantile epilepsy.

Pathogenic variants in SCN8A are associated with a broad phenotypic spectrum, including Self-Limiting Familial Infantile Epilepsy (SeLFIE), characterized by infancy-onset age-related seizures with normal development and cognition. Movement disorders, particularly paroxysmal kinesigenic dyskinesia typically arising after puberty, may represent another core symptom. We present the case of a 1-year-old girl with a familial disposition to self-limiting focal seizures from the maternal side and early-onset orofacial movement disorders associated with SCN8A-SeLFIE. Brain MRI was normal. Genetic testing revealed a maternally inherited SCN8A variant [c.4447G > A; p.(Glu1483Lys)]. After the introduction of valproic acid, she promptly achieved seizure control as well as complete remission of strabismus and a significant decrease in episodes of tongue deviation. Family history, genetic findings, and epilepsy phenotype are consistent with SCN8A-SeLFIE. Movement disorders are an important part of the SCN8A phenotypic spectrum, and this case highlights the novel early-onset orofacial movement disorders associated with this condition. The episodes of tongue deviation and protrusion suggest focal oromandibular (lingual) dystonia. Additionally, while infantile strabismus or esophoria is a common finding in healthy individuals, our case raises the possibility of an ictal origin of the strabismus. This study underscores the importance of recognizing and addressing movement disorders in SCN8A-SeLFIE patients, particularly the rare early-onset orofacial manifestations. It adds to the growing body of knowledge regarding the diverse clinical presentations of SCN8A-associated disorders and suggests potential avenues for clinical management and further research.

A de novo mutation in CACNA1A is associated with autosomal dominant bovine familial convulsions and ataxia in Angus cattle.

Bovine familial convulsions and ataxia (BFCA) is considered an autosomal dominant syndrome with incomplete penetrance. Nine Angus calves from the same herd were diagnosed with BFCA within days of birth. Necropsy revealed cerebellar and spinal cord lesions associated with the condition. Parentage testing confirmed that all affected calves had a common sire. The sire was then bred to 36 cows across two herds using artificial insemination, producing an additional 14 affected calves. The objective of this investigation was to identify hypothesized dominant genetic variation underlying the condition. Whole-genome sequencing was performed on the sire, six affected and seven unaffected paternal half-sibling calves and combined with data from 135 unrelated controls. The sire and five of the six affected calves were heterozygous for a nonsense variant (Chr7 g.12367906C>T, c.5073C>T, p.Arg1681*) in CACNA1A. The other affected calves (N = 8) were heterozygous for the variant but it was absent in the other unaffected calves (N = 7) and parents of the sire. This variant was also absent in sequence data from over 6500 other cattle obtained via public repositories and collaborator projects. The variant in CACNA1A is expressed in the cerebellum of the ataxic calves as detected in the transcriptome and was not differentially expressed compared with controls. The CACNA1A protein is part of a highly expressed cerebellar calcium voltage gated channel. The nonsense variant is proposed to cause haploinsufficiency, preventing proper transmission of neuronal signals through the channel and resulting in BFCA.

[Clinical and genetic spectrum of 6 cases with asparagine synthetase deficiency].

Objective: To explore the clinical and genetic characteristics of asparagine synthase deficiency. Methods: Case series studies. Retrospective analysis and summary of the clinical data of 6 cases with asparagine synthase deficiency who were diagnosed by genetic testing and admitted to the Third Affiliated Hospital of Zhengzhou University from May 2017 to April 2023 were analyzed retrospectively. The main clinical features, laboratory and imaging examination characteristics of the 6 cases were summarized, and the gene variation sites of them were analyzed. Results: All of the 6 cases were male, with onset ages ranging from 1 month to 1 year and 4 months. All of the 6 cases had cognitive and motor developmental delay, with 3 cases starting with developmental delay, 3 cases starting with convulsions and later experiencing developmental arrest or even regression. All of 6 cases had epilepsy, in whom 2 cases with severe microcephaly developed epileptic encephalopathy in the early stages of infancy with spasms as the main form of convulsions, 4 cases with mild or no microcephaly gradually evolved into convulsions with no fever after multiple febrile convulsions with focal seizures, tonic clonic seizures and tonic seizure as the main forms of convulsions. Three cases of 4 gradually developed into stagnation or even regression of development and ataxia after multiple convulsions with no fever. There were normal cranial imaging in 2 cases, dysplasia of the brains in 1 cases, frontal lobe apex accompanied by abnormal white matter signal in the frontal lobe and thin corpus callosum in 1 case, thin corpus callosum and abnormal lateral ventricular morphology in 1 case, and normal in early stage, but gradually developing into cerebellar atrophy at the age of 5 years and 9 months in 1 case. Two cases underwent visual evoked potential tests, the results of which were both abnormal. Three cases underwent auditory evoked potential examination, with 1 being normal and 2 being abnormal. All of 6 cases had variations in the asparagine synthase gene, with 2 deletion variations and 7 missense variations. The variations of 2 cases had not been reported so far, including c.1341_1343del and c.1283A>G, c.1165_1167del and c.1075G>A. The follow-up time ranged from 3 months to 53 months. Two cases who had severe microcephaly died in infancy, while the other 4 cases with mild or no microcephaly were in survival states until the follow-up days but the control of epilepsy was poor. Conclusions: Asparagine synthase deficiency has a certain degree of heterogeneity in clinical phenotype. Children with obvious microcephaly often present as severe cases, while children with mild or no microcephaly have relatively mild clinical manifestations. The variation of asparagine synthetase gene is mainly missense variation.

Hypomagnesaemia with varying degrees of extrarenal symptoms as a consequence of heterozygous CNNM2 variants.

Variants in the CNNM2 gene are causative for hypomagnesaemia, seizures and intellectual disability, although the phenotypes can be variable. This study aims to understand the genotype-phenotype relationship in affected individuals with CNNM2 variants by phenotypic, functional and structural analysis of new as well as previously reported variants. This results in the identification of seven variants that significantly affect CNNM2-mediated Mg2+ transport. Pathogenicity of these variants is further supported by structural modelling, which predicts CNNM2 structure to be affected by all of them. Strikingly, seizures and intellectual disability are absent in 4 out of 7 cases, indicating these phenotypes are caused either by specific CNNM2 variant only or by additional risk factors. Moreover, in line with sporadic observations from previous reports, CNNM2 variants might be associated with disturbances in parathyroid hormone and Ca2+ homeostasis.

Genomic evidence for the suitability of Göttingen Minipigs with a rare seizure phenotype as a model for human epilepsy.

Epilepsy is a complex genetic disorder that affects about 2% of the global population. Although the frequency and severity of epileptic seizures can be reduced by a range of pharmacological interventions, there are no disease-modifying treatments for epilepsy. The development of new and more effective drugs is hindered by a lack of suitable animal models. Available rodent models may not recapitulate all key aspects of the disease. Spontaneous epileptic convulsions were observed in few Göttingen Minipigs (GMPs), which may provide a valuable alternative animal model for the characterisation of epilepsy-type diseases and for testing new treatments. We have characterised affected GMPs at the genome level and have taken advantage of primary fibroblast cultures to validate the functional impact of fixed genetic variants on the transcriptome level. We found numerous genes connected to calcium metabolism that have not been associated with epilepsy before, such as ADORA2B, CAMK1D, ITPKB, MCOLN2, MYLK, NFATC3, PDGFD, and PHKB. Our results have identified two transcription factor genes, EGR3 and HOXB6, as potential key regulators of CACNA1H, which was previously linked to epilepsy-type disorders in humans. Our findings provide the first set of conclusive results to support the use of affected subsets of GMPs as an alternative and more reliable model system to study human epilepsy. Further neurological and pharmacological validation of the suitability of GMPs as an epilepsy model is therefore warranted.

Loss of TBC1D2B causes a progressive neurological disorder with gingival overgrowth.

Biallelic loss-of-function variants in TBC1D2B have been reported in five subjects with cognitive impairment and seizures with or without gingival overgrowth. TBC1D2B belongs to the family of Tre2-Bub2-Cdc16 (TBC)-domain containing RAB-specific GTPase activating proteins (TBC/RABGAPs). Here, we report five new subjects with biallelic TBC1D2B variants, including two siblings, and delineate the molecular and clinical features in the ten subjects known to date. One of the newly reported subjects was compound heterozygous for the TBC1D2B variants c.2584C>T; p.(Arg862Cys) and c.2758C>T; p.(Arg920*). In subject-derived fibroblasts, TBC1D2B mRNA level was similar to control cells, while the TBC1D2B protein amount was reduced by about half. In one of two siblings with a novel c.360+1G>T splice site variant, TBC1D2B transcript analysis revealed aberrantly spliced mRNAs and a drastically reduced TBC1D2B mRNA level in leukocytes. The molecular spectrum included 12 different TBC1D2B variants: seven nonsense, three frameshifts, one splice site, and one missense variant. Out of ten subjects, three had fibrous dysplasia of the mandible, two of which were diagnosed as cherubism. Most subjects developed gingival overgrowth. Half of the subjects had developmental delay. Seizures occurred in 80% of the subjects. Six subjects showed a progressive disease with mental deterioration. Brain imaging revealed cerebral and/or cerebellar atrophy with or without lateral ventricle dilatation. The TBC1D2B disorder is a progressive neurological disease with gingival overgrowth and abnormal mandible morphology. As TBC1D2B has been shown to positively regulate autophagy, defects in autophagy and the endolysosomal system could be associated with neuronal dysfunction and the neurodegenerative disease in the affected individuals.