Genetic Diversity in Early Infantile Epileptic Encephalopathy: A Three-Year Cohort Study.

Kcnq2 R213 knock-in mice reveal variant- and region-specific mechanisms underlying self-limited familial neonatal-infantile epilepsy and early infantile developmental and epileptic encephalopathy.

Background and ObjectivesPathogenic variants in the SCN2A gene, encoding the α-subunit type 2 of the voltage-gated sodium channel NaV1.2, cause a phenotypic spectrum including 4 major disorders as benign familial infantile seizures, developmental and epileptic encephalopathy, intellectual disability, and autism. Gain-of-function variants resulting phenotypes may be treated with sodium channel blockers, while loss-of-function (LoF) conditions are non-respondent. We focused on the effects of the pathogenic SCN2A variant c.4976C>T (p.A1659V) found in heterozygosity in 3 patients affected by DEE non responsive to SCB. We functionally investigated this previously uncharacterized SCN2A variant.MethodsThree individuals with the SCN2A c.4976C>T (p.A1659V) variant were studied. This variant was detected by next-generation sequencing (NGS). The nucleotide substitution was inserted by site-directed mutagenesis in a stabilized SCN2A plasmid encoding NaV1.2. Expression and functional characterization of the NaV1.2 A1659V variant was performed in HEK293 cells by western blotting, confocal microscopy, and patch clamp electrophysiology.ResultsThe same de novo pathogenic SCN2A variant was detected in 3 patients with DEE characterized by early onset, severe ID, and seizures unresponsive to SCB. In 2 patients, the variant is in a mosaic state. The NaV1.2 A1659V variant did not affect channel protein expression while exhibiting significant effects on its function as shown by the reduced Na+ currents, a shift of the activation curve toward more negative potentials, a shift of the inactivation curve to more negative voltages, and slower kinetics of inactivation compared with native NaV1.2 in HEK293 cells. Simulations suggested that the variant increases excitability in neurons.DiscussionThese results revealed the multifaceted functional effect of A1659V variant on channel activity and highlighted the complex genotype-phenotype correlation underlying significant clinical and pharmacological variability in SCN2A-related encephalopathies.

Cannabidiol (CBD), one of the compounds found in cannabis sativa, has drawn a lot of interest in the study and treatment of epilepsy. The antiepileptic qualities of CBD are being investigated for their ability to reduce seizure frequency and intensity in people with rare epilepsies, including West syndrome, Ohtahara's syndrome, Dravet syndrome, Lennox-Gastaut syndrome, and Tuberous Sclerosis. This review attempts to analyze the antiepileptic effects of cannabidiol against voltage-gated calcium channel T-type (CaV), GammaAminobutyric Acid A (GABAA), voltage-gated potassium channel of the Q family (KCNQ2), voltage-gated sodium channel (NaV), and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) using in silico techniques. Studies were conducted to investigate Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) parameters, and subsequently, molecular docking was performed. CBD demonstrated good oral absorption and the ability to cross the blood-brain barrier, as indicated by its pharmacokinetic parameters. The CBD may lead to potential drug interactions and increased bioavailability of the molecule due to metabolic interactions with the cytochrome P450 enzymatic system. CBD did not present toxicity parameters evaluated in this work. The molecular docking of CBD showed good interactions with NMDA and Nav. It also demonstrated good binding energy.

Abstract Multisystem inflammatory syndrome in children (MIS-C) is a severe complication of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) infection. While children are commonly affected by MIS-C, reports have described MIS in neonates (MIS-N) following maternal coronavirus disease 2019 (COVID-19) infection. We describe a case of early infantile developmental and epileptic encephalopathy (EIDEE) secondary to MIS-N in a day 18 old neonate secondary to maternal COVID-19. A term-born male neonate with a history of maternal COVID-19 at 35 weeks of gestation presented on day 18 of life with seizures, encephalopathy, pneumonitis, transaminitis, and elevated inflammatory markers. Magnetic resonance imaging (MRI) brain showed extensive cortical laminar necrosis. He was managed for MIS-N with antiseizure medications, antibiotics, and intravenous immunoglobulin. COVID-19 reverse transcriptase-polymerase chain reaction was negative, and anti-COVID immunoglobulin G antibody was positive. Near continuous multifocal myoclonic seizures were associated with a suppression burst pattern in the EEG. Follow-up MRI showed cystic encephalomalacia and loss of periventricular white matter. The EIDEE remained drug refractory with poor neurological outcome in follow-up. Cytotoxin-mediated neuronal injury in MIS-N can lead to a catastrophic complication of EIDEE, resulting in drug-refractory epilepsy, microcephaly, and adverse neurological outcome.

To systematically summarize the clinical phenotypes, treatment responses, prognosis, and genetic characteristics of STXBP1-encephalopathy in Chinese pediatric patients, and to explore the clinical value of genetic testing in this disease. We retrospectively analyzed the clinical data, gene variant information, and treatment outcomes of 19 children with STXBP1-encephalopathy admitted to the Department of Pediatrics, Second Affiliated Hospital of Zhejiang University, between January 2020 and January 2024. Whole-exome sequencing was performed for genetic diagnosis, and Sanger sequencing was used to verify variants and confirm their parental origin. Pathogenicity of variants was classified according to the American College of Medical Genetics and Genomics guidelines. STXBP1-encephalopathy showed early onset, with 15 cases (78.9%) occurring within the 1st month of life. Five patients were diagnosed with Ohtahara syndrome, 5 with West syndrome, and 9 with unclassifiable early-onset epileptic encephalopathy. All patients had abnormal electro-encephalogram findings, mainly burst suppression (68.4%) and hypsarrhythmia (63.2%). Among the 19 patients, 1 achieved seizure freedom and discontinued antiepileptic drugs, and 4 achieved seizure control with levetiracetam. A total of 18 de novo pathogenic/likely pathogenic variants in STXBP1 were identified, including 7 novel variants: c.326-3(IVS5)delC, c.656del(p.Met219Argfs*13), c.746-747del(p.F249fs*6), c.798T > G(p.Y266*), c.1155delC(p.D385fs), c.1249G > A(p.G417S), and c.1250-2A > G. STXBP1 pathogenic variants are an important cause of early-onset developmental epileptic encephalopathy in Chinese children. Genetic testing is crucial for early diagnosis of STXBP1-encephalopathy. Levetiracetam shows good efficacy in controlling seizures in these patients, and early use is recommended. The 7 novel variants identified in this study expand the STXBP1 mutation spectrum.

Genetically determined effects on the development of epilepsy are diverse. First of all, they are aimed at changing the activity of ion channels and functional proteins (including synaptic proteins). Epigenetic regulation of genome activity is also considered one of the most important factors in the development of both single paroxysms and epilepsy as a disease. To date, numerous associations between gene disorders and epilepsy have been described. They are detected in genes of very different determinant activity, and identical clinical phenotypes, such as West, Dravet, Ohtahara, and Lennox-Gastaut syndromes, can be observed when different genes are affected. Within a single gene-associated condition, multiple mutations of different types can occur, and distinct clinical features of the disease correspond to distinct types of mutations. Moreover, involvement of a single gene can induce several mechanisms of epileptogenesis (impaired receptor regulation, permeability of the blood-brain barrier, impaired neuronal differentiation, etc.). Epigenetic mechanisms provide genome functional lability by changing the accessibility of individual DNA parts without altering their structure. The genome-altering mechanisms during epileptogenesis include the methylation of genes encoding voltage-gated calcium and potassium channels, as well as genes that affect neuronal plasticity, signaling pathway function, and neuroinflammation processes. Histone acetylation can be considered both as a maintenance and progression factor of the epileptic process and as a «rapid response agent» in status epilepticus. Non-coding RNAs regulate neuroinflammation, apoptosis, synaptogenesis, synaptic plasticity, abnormal sprouting, and neural network remodeling. The circle is closed by pathogenic variants of genes that determine the synthesis of epigenetic effectors. Expanding knowledge about the genome lability mechanisms is one of the prerequisites for success in finding new ways to treat epilepsy.

Developmental epileptic encephalopathy is characterized by a malignant course of refractory epilepsy and a growing defect in mental and motor development. To date, determinant genes have been identified for most of these conditions. This condition is also caused by mutations in the SCN8A gene of a voltage-gated sodium channel, which is detected in about 1% of children with early infantile epileptic encephalopathies. SCN8A gene mutations result in both an increase (gain-of-function, GOF mutation) and a decrease (loss-of-function, LOF mutation) in NaV1.6 sodium channel function, leading to a decrease in current amplitude and a depolarizing shift. With GOF mutation, hypotension, cerebral visual dysfunction, dyskinesia, ataxia, and epileptic seizures are mainly present as bilateral tonic and focal seizures and, in most cases, lead to epileptic encephalopathy. With the LOF mutation, seizures (absences, bilateral tonic-clonic myoclonic seizures) may not appear, while cognitive impairment, movement disorders, and autistic traits are present. Anticonvulsant regimens include non-selective sodium channel blockers, GABA agonists, Na+ and Ca2+ channel blockers, and other agents. A specific feature of the phenotype is a relatively high probability of exacerbation of the epilepsy during anticonvulsant selection, which requires special care and caution from the doctor. As an illustration of the most typical features of the condition, a clinical case of severe refractory epilepsy with a developmental disorder in an infant is presented. The disease progressed despite all attempts to select therapy. Treatment with corticosteroids gave a temporary effect, and when the drug was discontinued, the frequency and severity of seizures increased, suggesting the GOF scenario; however, genetic confirmation was inaccessible. The authors believe that the prospects for treating such conditions are determined primarily by progress in the development of new drugs, including oligonucleotides and selective inhibitors of NaV1.6 channels.

According to the World Health Organization, more than 50 million people worldwide have epilepsy, and about 5 million new cases are reported annually. According to other data, the number of patients exceeds 70.000.000, highlighting the relevance of this condition. Studies of developmental epileptic encephalopathies leading to early disability are particularly relevant. About 900 gene mutations associated with different types of epilepsy have been identified so far. One of the prominent «channelopathy genes» representatives is KCNT1, coding the largest subunit of the sodium-activated potassium channel, KNa1.1. Its mutations are associated with many neurological disorders, such as leukodystrophy, leukoencephalopathy, West syndrome, Ohtahara syndrome, early myoclonic encephalopathy, focal epilepsy, and multifocal epilepsy. A clinical case of a typical course of KCNT1-associated epilepsy with early onset (in the first half of life), focal and asynchronous paroxysms, psychomotor retardation, and drug resistance is presented. The effectiveness of the anticonvulsants used and the nature of pharmacological resistance were analyzed. The applicability of four «Drug Resistance Hypotheses» to this case is shown: genetic, since the pathogenesis of this particular case is based on a change in the properties of potassium channels; a change in the sensitivity of the anticonvulsant targets explains the intermittent (according to Schmidt and Löscher) nature of resistance. The «Neural Networks Hypothesis» is illustrated by the change of encephalographic indicators from multifocal epileptiform activity with an intact background to pronounced dysrhythmia with periods of the «burst-suppression» pattern after several months. All these features, combined, refer to the «Intrinsic Severity» hypothesis by Rogawski and Johnson, since the child's disease was initially severe due to the intensity and frequency of epileptic seizures. As a result, a conclusion was made about the complexity of the refractoriness formation mechanisms, even in cases with a simple and understandable pathogenesis of channelopathy.

Beyond traditional therapies: CRISPR-Cas9 targets the genetic core of ohtahara syndrome for lasting impact

Increasing evidence supports the human sodium-coupled citrate transporter (hNaCT) as a potential therapeutic target for metabolic syndrome and early infantile epileptic encephalopathy type 25 (EIEE25). While isotopic tracers remain the reference method for evaluating citrate transport, the need for specialized equipment and regulatory approval restricts their widespread use. Here, we report the development and validation of a robust, fluorescent-based assay to evaluate hNaCT-mediated citrate transport in live cells at both single-cell and high-throughput levels. This method utilizes a baculoviral vector to modify HEK293 cells to co-express a genetically encoded citrate sensor (Citron1) and the hNaCT. This cell-based platform enabled real-time monitoring of citrate transport using fluorescent microscopy and a standard multiwell plate reader. A key strength of this approach is its ability to assess citrate transport in the same cells before and after experimental interventions. Accordingly, this approach enables the functional characterization of hNaCT, including its pharmacological inhibitors and genetic variants with altered activity. Overall, the method provides a reliable assessment of citrate transport and offers a versatile platform suitable for identifying novel lead compounds for the therapeutic modulation of hNaCT.

Background: Developmental and Epileptic Encephalopathy (DEE) is a severe and heterogeneous neurological disorder in infancy/early childhood. DEE’s genetic and phenotypic variability complicates diagnosis and treatment. This retrospective study aimed to identify genetic variants and explore genotype–phenotype correlations in children with DEE using a targeted epilepsy gene panel (TGP) and Whole Exome Sequencing (WES). Patients and Methods: Medical records of children who underwent custom-designed 55-gene TGP and WES were reviewed. The diagnostic yield of each method was determined based on the detection of pathogenic (P) and likely pathogenic (LP) variants. Results: A total of 129 patients (66 males, 63 females) underwent TGP, which identified P/LP variants in 29 cases (22.48%). Variants were detected in SCN1A, KCNQ2, STXBP1, CDKL5, PCDH19, PLCB1, WWOX, SCN2A, FGF12, HCN1, SCN8A, and SLC35A2. WES further identified several variants in children with West syndrome. A TSC1 variant was detected in a patient without cutaneous stigmata of tuberous sclerosis complex. The NALCN variant in a patient was linked to Infantile Hypotonia with Psychomotor Retardation and Characteristic Facies 1. A CTBP1 variant associated with extremely rare Hypotonia, Ataxia, Developmental Delay, and Tooth Enamel Defect Syndrome was detected in another patient. A PIEZO2 variant—associated with Marden–Walker syndrome—was found in a child with Early Infantile Developmental and Epileptic Encephalopathy. Conclusions: These findings highlight the extensive genetic heterogeneity and phenotypic variability of DEE. WES demonstrates substantial value in identifying novel gene-disease associations and may be considered as a first-tier diagnostic tool in epilepsy and DEE.

Rapid Electroclinical Evolution in HECW2-Related Developmental and Epileptic Encephalopathy: Report of a Likely Splicing Variant With Familial Transmission.

Developmental and epileptic encephalopathy (DEE) caused by a mutation in the KCNT1 gene (KCNT1-DEE) is registered in the Online Mendelian Inheritance in Man (OMIM) catalogue inder code number 614959. Alternative names: DEE 14, early infantile epileptic encephalopathy 14. KCNT1-DEE most often manifests as a syndrome of “epilepsy in infancy with migrating focal seizures”. However, the course of epilepsies due to mutant KCNT1 gene is characterized by a broad clinical polymorphism. The article describes a child with severe KCNT1-DEE with clinical picture dominated by drug-resistant focal seizures, profound mental retardation and spastic tetraparesis, as well as microcephaly and microsomia.

The sodium-citrate cotransporter (NaCT) plays a crucial role in citrate transport during amelogenesis. Mutations in the SLC13A5 gene, which encodes the NaCT, cause early infantile epileptic encephalopathy 25 and amelogenesis imperfecta. We analyzed developing pig molars and determined that the citrate concentrations in secretory- and maturation-stage enamel are both 5.3 µmol/g, with about 95% of the citrate being bound to mineral. To better understand how citrate might enter developing enamel, we developed Slc13a5Flag reporter mice that express NaCT with a C-terminal Flag-tag (DYKDDDDK) that can be specifically and accurately recognized by commercially available anti-Flag antibodies. The 24-base Flag coding sequence was located immediately upstream of the natural translation termination codon (TAG) and was validated by Sanger sequencing. The general development, physical activities, and reproductive outcomes of this mouse strain were comparable to those of the C57BL/6 mice. No differences were detected between the Slc13a5Flag and wild-type mice. Tooth development was extensively characterized using dissection microscopy, bSEM, light microscopy, in situ hybridization, and immunohistochemistry. Tooth formation was not altered in any detectable way by the introduction of the Flag. The Slc13a5Flag citrate transporter was observed on all outer membranes of secretory ameloblasts (distal, lateral, and proximal), with the strongest signal on the Tomes process, and was detectable in all but the distal membrane of maturation-stage ameloblasts. The papillary layer also showed positive immunostaining for Flag. The outer membrane of odontoblasts stained stronger than ameloblasts, except for the odontoblastic processes, which did not immunostain. As NaCT is thought to only facilitate citrate entry into the cell, we performed in situ hybridization that showed Ank is not expressed by secretory- or maturation-stage ameloblasts, ruling out that ANK can transport citrate into enamel. In conclusion, we developed Slc13a5Flag reporter mice that provide specific and sensitive localization of a fully functional NaCT-Flag protein. The localization of the Slc13a5Flag citrate transporter throughout the ameloblast membrane suggests that either citrate enters enamel by a paracellular route or NaCT can transport citrate bidirectionally (into or out of ameloblasts) depending upon local conditions.

Developmental and epileptic encephalopathy (DEE) is epilepsy related to developmental impairment that may be caused by both the underlying etiology (developmental encephalopathy) and superimposed epileptic activity (epileptic encephalopathy). The origin of DEE and the causes of its variations remain unknown. Owing the lack of clarity regarding the role of genetic variables in DEE, we conducted a scoping review to qualitatively identify the genes most important in the development of DEE to provide an up-to-date review. We searched all published studies related to the genetic factors of DEE. The identified publications were screened and selected by the authors on basis of on inclusion and exclusion criteria and assessed for methodological quality. Eighteen articles were included. The extracted data included age of onset, sex, gene mutations and inheritance (e.g. nucleotide change, protein change, and family testing), clinical manifestation, electroencephalogram, imaging, medication, and outcomes. A total of 18 studies were included in this scoping review. The most frequently reported gene variants were STXBP1 in Ohtahara Syndrome, SLC1A2 in Early Myoclonic Encephalopathy (EME), CDKL5 in West Syndrome, SCN1A in Dravet Syndrome, and KCNT1 in Epilepsy of Infancy with Migrating Focal Seizures (EIMFS). Each gene was associated with distinct electroclinical features, including differences in age of onset, seizure type, EEG patterns, and developmental outcomes. While genotype and phenotype associations were heterogeneous, certain variants showed consistent patterns indicative of more severe disease courses. This review identified key gene variants commonly associated with early-onset DEE in infants, particularly STXBP1, SLC1A2, CDKL5, SCN1A, and KCNT1, each linked to unique clinical presentations and outcomes. These findings support the clinical utility of next-generation sequencing (NGS) for early diagnosis and tailored treatment planning in DEE. Understanding genotype-phenotype correlations may enhance prognostication and highlight potential avenues for targeted therapy in future research.

Seizure reduction following ventriculoperitoneal shunt surgery in an Ohtahara syndrome patient who developed hydrocephalus

<p>Background: Developmental and epileptic encephalopathies (DEEs) are severe disorders marked by refractory seizures and developmental delay. Pathogenic variants in the syntaxin-binding protein 1 (STXBP1) gene are among the top five genetic causes of DEE and impair neurotransmitter release, especially in GABAergic interneurons. The clinical presentation is highly variable, and diagnosis depends on molecular genetic testing. Precision medicine is key for diagnosis, treatment, follow-up, prognosis, and hereditary risk reduction.</p> <p>Case Presentation: We present a male patient with drug-resistant epilepsy, polypharmacy, and cognitive impairment, without significant family or perinatal risk factors. Given the clinical complexity and strong suspicion of genetic cause, a molecular study using next-generation sequencing of epilepsy-related genes and copy number variation analysis was performed. A heterozygous pathogenic variant in STXBP1, c.1652G>A (p.Arg551His), was identified, associated with early infantile epileptic encephalopathy type type 4, also known as STXBP1-DEE (MONDO:0012812 - orphanet identifier (ORPHA):599373], with autosomal dominant inheritance.</p> <p>Conclusion: STXBP1-DEE represents a heterogeneous spectrum of neurodevelopmental disorders with refractory epilepsy, developmental delay, and intellectual disability. Diagnosis requires clinical suspicion, imaging, laboratory tests, and molecular confirmation. While current treatments are limited, promising approaches are under investigation. The lack of genotype-phenotype correlation and wide phenotypic variability complicate management, but advances in precision medicine support more individualized treatment strategies. Although most variants are de novo, genetic counseling remains crucial to assess recurrence risk. Preclinical studies show potential for novel therapies, yet clinical trials are needed to confirm their efficacy in affected individuals.</p>

Pathogenic variants in γ-aminobutyric acid type A (GABAA) receptor subunit genes are increasingly associated with epilepsy and neurodevelopmental disorders. Pathogenic variants in GABRA2, encoding the α-2 subunit of GABAA receptors, have been recently reported. This study aims to better delineate the phenotypic spectrum of GABRA2 pathogenic variants. We conducted a retrospective multicenter study, analyzing six new patients with GABRA2 pathogenic variants identified through a French national collaboration. Clinical, electroencephalographic (EEG), and genetic data were reviewed alongside a literature analysis of eight previously reported cases. Two distinct electroclinical phenotypes were identified. The most severe, in four of six patients, featured early infantile developmental and epileptic encephalopathy with an EEG pattern of rapid rhythms suggestive of GABAergic hyperactivity. The milder phenotype, in two of six patients, included later onset, drug-responsive epilepsy with moderate developmental delay. A literature review confirmed these phenotypes and supported genotype-phenotype correlations, with transmembrane domain variants more frequently associated with severe phenotypes. This study refines the phenotypic spectrum of GABRA2-related disorders, highlighting two distinct electroclinical phenotypes. The identification of a recognizable EEG pattern of unusual rapid rhythms for age may be a biomarker for early diagnosis of a severe phenotype and suggests a potential underlying gain-of-function mechanism, to be confirmed by functional studies.

A large number of cases with Dravet syndrome (DS) has been attributed to SCN1A loss of function (LOF), whereas SCN1A gain‐of‐function (GOF) causes early infantile developmental and epileptic encephalopathy (EIDEE) and familial hemiplegic migraine 3. We retrospectively analyzed 37 individuals with SCN1A pathogenic variants at our institute between January 2012 and October 2024 to investigate phenotype–function correlations. Variant functions were classified as LOF, GOF, or mixed, based on existing patch‐clamp data, paralog sodium channel experimental findings, and in silico prediction tools. Clinical characteristics, antiseizure medication (ASM) responses, and variant location were compared. Nine variants were novel. One variant with insufficient data for functional prediction was excluded. Of the 36 cases with predictable functions, five cases (14%) were classified as GOF/mixed (DS = 4, EIDEE = 1) and 31 (86%) as LOF (DS = 31). GOF/mixed‐DS had earlier epilepsy onset but otherwise resembled LOF‐DS. Sodium channel blocking ASMs (SCB‐ASMs) did not exacerbate seizures in GOF/mixed DS cases, with carbamazepine reducing seizures in one case. GOF/mixed variants clustered in the intracellular S6 segment, whereas LOF variants clustered in the S5–S6 pore loop. These findings highlight a potential GOF effect for certain DS cases, suggesting that SCB‐ASMs may be effective for GOF/mixed DS. This underscores the importance of functional characterization for tailored therapy, warranting further research to confirm and extend these results.Plain Language SummaryDravet syndrome is a severe epilepsy that usually begins in infancy and is linked to changes in a gene called SCN1A. Most cases are caused by gene changes that reduce function, but in some cases, the gene may become overactive. In this study, we found that some patients with Dravet syndrome had these overactive changes and still showed typical symptoms. We found that people with overactive SCN1A function might respond differently to certain medications.