Video-EEG-monitoring to guide antiseizure medication withdrawal.

BACKGROUND: Discontinuing anti-seizure medication (ASM) should be considered in persons with epilepsy with long-term seizure freedom. Clinicians should also pursue ASM withdrawal in persons with one-time seizures without increased recurrence risk and those with suspected non-epileptic events. However, ASM withdrawal is associated with the risk of recurring seizures. Monitored ASM withdrawal in an epilepsy monitoring unit (EMU) could help better evaluate the risk of seizure recurrence. Here, we investigate the practice of EMU-guided ASM withdrawal, assess its indications, and aim to determine positive and negative predictors for successful withdrawal. METHODS: We screened the medical records of all patients admitted to our EMU between November 1, 2019, and October 31, 2021, and included patients of at least 18 years admitted with the aim of permanent ASM withdrawal. We defined four groups of withdrawal indications: (1) long-term seizure freedom; (2) suspected non-epileptic events; (3) history of epileptic seizures but not fulfilling diagnostic criteria of epilepsy; and (4) seizure-freedom after epilepsy surgery. Successful withdrawal was defined according to the following criteria: no recoding of (sub)clinical seizure activity during VEM (groups 1, 2, and 3), patients did not meet the International League Against Epilepsy (ILAE) definition of epilepsy (groups 2 and 3) [14], and patients were discharged without ongoing ASM treatment (all groups). We also evaluated the prediction model by Lamberink et al. (LPM) for the risk of seizure recurrence in groups 1 and 3. RESULTS: 55/651 (8.6%) patients fulfilled the inclusion criteria. Withdrawal indications were distributed as follows; group 1: 2/55 (3.6%); group 2: 44/55 (80%); group 3: 9/55 (16,4%); group 4: 0/55. Overall, ASM withdrawal was successful in 90.9%. The sensitivity of the LPM for a 2-year 50% relapse risk threshold was 75%, the specificity 33.3%; for a 5-year relapse risk respectively 12.5% and 33.3%, suggesting that the model is not suitable for risk assessment in patients with one-time seizures or acute-symptomatic seizures, who constituted most of the evaluated patients. CONCLUSIONS: Our study suggests that EMU-guided ASM withdrawal could be a helpful tool to support clinical decision-making and improve patient safety. Prospective, randomized trials should further evaluate this method in the future.

Drosophila melanogaster as a versatile model organism to study genetic epilepsies: An overview.

Epilepsy is one of the most prevalent neurological disorders, affecting more than 45 million people worldwide. Recent advances in genetic techniques, such as next-generation sequencing, have driven genetic discovery and increased our understanding of the molecular and cellular mechanisms behind many epilepsy syndromes. These insights prompt the development of personalized therapies tailored to the genetic characteristics of an individual patient. However, the surging number of novel genetic variants renders the interpretation of pathogenetic consequences and of potential therapeutic implications ever more challenging. Model organisms can help explore these aspects in vivo. In the last decades, rodent models have significantly contributed to our understanding of genetic epilepsies but their establishment is laborious, expensive, and time-consuming. Additional model organisms to investigate disease variants on a large scale would be desirable. The fruit fly Drosophila melanogaster has been used as a model organism in epilepsy research since the discovery of "bang-sensitive" mutants more than half a century ago. These flies respond to mechanical stimulation, such as a brief vortex, with stereotypic seizures and paralysis. Furthermore, the identification of seizure-suppressor mutations allows to pinpoint novel therapeutic targets. Gene editing techniques, such as CRISPR/Cas9, are a convenient way to generate flies carrying disease-associated variants. These flies can be screened for phenotypic and behavioral abnormalities, shifting of seizure thresholds, and response to anti-seizure medications and other substances. Moreover, modification of neuronal activity and seizure induction can be achieved using optogenetic tools. In combination with calcium and fluorescent imaging, functional alterations caused by mutations in epilepsy genes can be traced. Here, we review Drosophila as a versatile model organism to study genetic epilepsies, especially as 81% of human epilepsy genes have an orthologous gene in Drosophila. Furthermore, we discuss newly established analysis techniques that might be used to further unravel the pathophysiological aspects of genetic epilepsies.

Loss-of-function variants in the KCNQ5 gene are implicated in genetic generalized epilepsies.

BACKGROUND: De novo missense variants in KCNQ5, encoding the voltage-gated K+ channel KV7.5, have been described to cause developmental and epileptic encephalopathy (DEE) or intellectual disability (ID). We set out to identify disease-related KCNQ5 variants in genetic generalized epilepsy (GGE) and their underlying mechanisms. METHODS: 1292 families with GGE were studied by next-generation sequencing. Whole-cell patch-clamp recordings, biotinylation and phospholipid overlay assays were performed in mammalian cells combined with homology modelling. FINDINGS: We identified three deleterious heterozygous missense variants, one truncation and one splice site alteration in five independent families with GGE with predominant absence seizures; two variants were also associated with mild to moderate ID. All missense variants displayed a strongly decreased current density indicating a loss-of-function (LOF). When mutant channels were co-expressed with wild-type (WT) KV7.5 or KV7.5 and KV7.3 channels, three variants also revealed a significant dominant-negative effect on WT channels. Other gating parameters were unchanged. Biotinylation assays indicated a normal surface expression of the variants. The R359C variant altered PI(4,5)P2-interaction. INTERPRETATION: Our study identified deleterious KCNQ5 variants in GGE, partially combined with mild to moderate ID. The disease mechanism is a LOF partially with dominant-negative effects through functional deficits. LOF of KV7.5 channels will reduce the M-current, likely resulting in increased excitability of KV7.5-expressing neurons. Further studies on network level are necessary to understand which circuits are affected and how this induces generalized seizures. FUNDING: DFG/FNR Research Unit FOR-2715 (Germany/Luxemburg), BMBF rare disease network Treat-ION (Germany), foundation 'no epilep' (Germany).

Postictal Psychosis in Epilepsy: A Clinicogenetic Study.

OBJECTIVE: Psychoses affecting people with epilepsy increase disease burden and diminish quality of life. We characterized postictal psychosis, which comprises about one quarter of epilepsy-related psychoses, and has unknown causation. METHODS: We conducted a case-control cohort study including patients diagnosed with postictal psychosis, confirmed by psychiatric assessment, with available data regarding epilepsy, treatment, psychiatric history, psychosis profile, and outcomes. After screening 3,288 epilepsy patients, we identified 83 with psychosis; 49 had postictal psychosis. Controls were 98 adults, matched by age and epilepsy type, with no history of psychosis. Logistic regression was used to investigate clinical factors associated with postictal psychosis; univariate associations with a p value < 0.20 were used to build a multivariate model. Polygenic risk scores for schizophrenia were calculated. RESULTS: Cases were more likely to have seizure clustering (odds ratio [OR] = 7.59, p < 0.001), seizures with a recollected aura (OR = 2.49, p = 0.013), and a family history of psychiatric disease (OR = 5.17, p = 0.022). Cases showed predominance of right temporal epileptiform discharges (OR = 4.87, p = 0.007). There was no difference in epilepsy duration, neuroimaging findings, or antiseizure treatment between cases and controls. Polygenic risk scores for schizophrenia in an extended cohort of postictal psychosis cases (n = 58) were significantly higher than in 1,366 epilepsy controls (R2  = 3%, p = 6 × 10-3 ), but not significantly different from 945 independent patients with schizophrenia (R2  = 0.1%, p = 0.775). INTERPRETATION: Postictal psychosis occurs under particular circumstances in people with epilepsy with a heightened genetic predisposition to schizophrenia, illustrating how disease biology (seizures) and trait susceptibility (schizophrenia) may interact to produce particular outcomes (postictal psychosis) in a common disease. ANN NEUROL 2021;90:464-476.

Real-life survey of pitfalls and successes of precision medicine in genetic epilepsies.

OBJECTIVE: The term 'precision medicine' describes a rational treatment strategy tailored to one person that reverses or modifies the disease pathophysiology. In epilepsy, single case and small cohort reports document nascent precision medicine strategies in specific genetic epilepsies. The aim of this multicentre observational study was to investigate the deeper complexity of precision medicine in epilepsy. METHODS: A systematic survey of patients with epilepsy with a molecular genetic diagnosis was conducted in six tertiary epilepsy centres including children and adults. A standardised questionnaire was used for data collection, including genetic findings and impact on clinical and therapeutic management. RESULTS: We included 293 patients with genetic epilepsies, 137 children and 156 adults, 162 females and 131 males. Treatment changes were undertaken because of the genetic findings in 94 patients (32%), including rational precision medicine treatment and/or a treatment change prompted by the genetic diagnosis, but not directly related to known pathophysiological mechanisms. There was a rational precision medicine treatment for 56 patients (19%), and this was tried in 33/56 (59%) and was successful (ie, >50% seizure reduction) in 10/33 (30%) patients. In 73/293 (25%) patients there was a treatment change prompted by the genetic diagnosis, but not directly related to known pathophysiological mechanisms, and this was successful in 24/73 (33%). SIGNIFICANCE: Our survey of clinical practice in specialised epilepsy centres shows high variability of clinical outcomes following the identification of a genetic cause for an epilepsy. Meaningful change in the treatment paradigm after genetic testing is not yet possible for many people with epilepsy. This systematic survey provides an overview of the current application of precision medicine in the epilepsies, and suggests the adoption of a more considered approach.

Desynchronization of temporal lobe theta-band activity during effective anterior thalamus deep brain stimulation in epilepsy.

BACKGROUND: Bilateral cyclic high frequency deep brain stimulation (DBS) of the anterior nucleus of the thalamus (ANT) reduces the seizure count in a subset of patients with epilepsy. Detecting stimulation-induced alterations of pathological brain networks may help to unravel the underlying physiological mechanisms related to effective stimulation delivery and optimize target engagement. METHODS: We acquired 64-channel electroencephalography during ten ANT-DBS cycles (145 â€‹Hz, 90 â€‹µs, 3-5 â€‹V) of 1-min ON followed by 5-min OFF stimulation to detect changes in cortical activity related to seizure reduction. The study included 14 subjects (three responders, four non-responders, and seven healthy controls). Mixed-model ANOVA tests were used to compare differences in cortical activity between subgroups both ON and OFF stimulation, while investigating frequency-specific effects for the seizure onset zones. RESULTS: ANT-DBS had a widespread desynchronization effect on cortical theta and alpha band activity in responders, but not in non-responders. Time domain analysis showed that the stimulation induced reduction in theta-band activity was temporally linked to the stimulation period. Moreover, stimulation induced theta-band desynchronization in the temporal lobe channels correlated significantly with the therapeutic response. Responders to ANT-DBS and healthy-controls had an overall lower level of theta-band activity compared to non-responders. CONCLUSION: This study demonstrated that temporal lobe channel theta-band desynchronization may be a predictive physiological hallmark of therapeutic response to ANT-DBS and may be used to improve the functional precision of this intervention by verifying implantation sites, calibrating stimulation contacts, and possibly identifying treatment responders prior to implantation.

Phenotypic and genetic spectrum of epilepsy with myoclonic atonic seizures.

OBJECTIVE: We aimed to describe the extent of neurodevelopmental impairments and identify the genetic etiologies in a large cohort of patients with epilepsy with myoclonic atonic seizures (MAE). METHODS: We deeply phenotyped MAE patients for epilepsy features, intellectual disability, autism spectrum disorder, and attention-deficit/hyperactivity disorder using standardized neuropsychological instruments. We performed exome analysis (whole exome sequencing) filtered on epilepsy and neuropsychiatric gene sets to identify genetic etiologies. RESULTS: We analyzed 101 patients with MAE (70% male). The median age of seizure onset was 34 months (range = 6-72 months). The main seizure types were myoclonic atonic or atonic in 100%, generalized tonic-clonic in 72%, myoclonic in 69%, absence in 60%, and tonic seizures in 19% of patients. We observed intellectual disability in 62% of patients, with extremely low adaptive behavioral scores in 69%. In addition, 24% exhibited symptoms of autism and 37% exhibited attention-deficit/hyperactivity symptoms. We discovered pathogenic variants in 12 (14%) of 85 patients, including five previously published patients. These were pathogenic genetic variants in SYNGAP1 (n = 3), KIAA2022 (n = 2), and SLC6A1 (n = 2), as well as KCNA2, SCN2A, STX1B, KCNB1, and MECP2 (n = 1 each). We also identified three new candidate genes, ASH1L, CHD4, and SMARCA2 in one patient each. SIGNIFICANCE: MAE is associated with significant neurodevelopmental impairment. MAE is genetically heterogeneous, and we identified a pathogenic genetic etiology in 14% of this cohort by exome analysis. These findings suggest that MAE is a manifestation of several etiologies rather than a discrete syndromic entity.

Pharmacoresponse in genetic generalized epilepsy: a genome-wide association study.

Aim: Pharmacoresistance is a major burden in epilepsy treatment. We aimed to identify genetic biomarkers in response to specific antiepileptic drugs (AEDs) in genetic generalized epilepsies (GGE). Materials & methods: We conducted a genome-wide association study (GWAS) of 3.3 million autosomal SNPs in 893 European subjects with GGE - responsive or nonresponsive to lamotrigine, levetiracetam and valproic acid. Results: Our GWAS of AED response revealed suggestive evidence for association at 29 genomic loci (p <10-5) but no significant association reflecting its limited power. The suggestive associations highlight candidate genes that are implicated in epileptogenesis and neurodevelopment. Conclusion: This first GWAS of AED response in GGE provides a comprehensive reference of SNP associations for hypothesis-driven candidate gene analyses in upcoming pharmacogenetic studies.

Gene family information facilitates variant interpretation and identification of disease-associated genes in neurodevelopmental disorders.

BACKGROUND: Classifying pathogenicity of missense variants represents a major challenge in clinical practice during the diagnoses of rare and genetic heterogeneous neurodevelopmental disorders (NDDs). While orthologous gene conservation is commonly employed in variant annotation, approximately 80% of known disease-associated genes belong to gene families. The use of gene family information for disease gene discovery and variant interpretation has not yet been investigated on a genome-wide scale. We empirically evaluate whether paralog-conserved or non-conserved sites in human gene families are important in NDDs. METHODS: Gene family information was collected from Ensembl. Paralog-conserved sites were defined based on paralog sequence alignments; 10,068 NDD patients and 2078 controls were statistically evaluated for de novo variant burden in gene families. RESULTS: We demonstrate that disease-associated missense variants are enriched at paralog-conserved sites across all disease groups and inheritance models tested. We developed a gene family de novo enrichment framework that identified 43 exome-wide enriched gene families including 98 de novo variant carrying genes in NDD patients of which 28 represent novel candidate genes for NDD which are brain expressed and under evolutionary constraint. CONCLUSION: This study represents the first method to incorporate gene family information into a statistical framework to interpret variant data for NDDs and to discover new NDD-associated genes.

Testing association of rare genetic variants with resistance to three common antiseizure medications.

OBJECTIVE: Drug resistance is a major concern in the treatment of individuals with epilepsy. No genetic markers for resistance to individual antiseizure medication (ASM) have yet been identified. We aimed to identify the role of rare genetic variants in drug resistance for three common ASMs: levetiracetam (LEV), lamotrigine (LTG), and valproic acid (VPA). METHODS: A cohort of 1622 individuals of European descent with epilepsy was deeply phenotyped and underwent whole exome sequencing (WES), comprising 575 taking LEV, 826 LTG, and 782 VPA. We performed gene- and gene set-based collapsing analyses comparing responders and nonresponders to the three drugs to determine the burden of different categories of rare genetic variants. RESULTS: We observed a marginally significant enrichment of rare missense, truncating, and splice region variants in individuals who were resistant to VPA compared to VPA responders for genes involved in VPA pharmacokinetics. We also found a borderline significant enrichment of truncating and splice region variants in the synaptic vesicle glycoprotein (SV2) gene family in nonresponders compared to responders to LEV. We did not see any significant enrichment using a gene-based approach. SIGNIFICANCE: In our pharmacogenetic study, we identified a slightly increased burden of damaging variants in gene groups related to drug kinetics or targeting in individuals presenting with drug resistance to VPA or LEV. Such variants could thus determine a genetic contribution to drug resistance.

Jumonji domain containing 1C (JMJD1C) sequence variants in seven patients with autism spectrum disorder, intellectual disability and seizures.

The Jumonji domain containing 1C (JMJD1C) gene encodes the Jumonji domain-containing protein 1C (JMJD1C) and is a member of the jmJC domain-containing protein family involved in histone demethylation that is expressed in the brain. We report seven, unrelated patients with developmental delays or intellectual disability and heterozygous, de novo sequence variants in JMJD1C. All patients had developmental delays, but there were no consistent additional findings. Two patients were reported to have seizures for which there was no other identified cause. De novo, deleterious sequence variants in JMJD1C have previously been reported in patients with autism spectrum disorder and a phenotype resembling classical Rett syndrome, but only one JMJD1C variant has undergone functional evaluation. In all of the seven patients in this report, there was a plausible, alternative explanation for the neurocognitive phenotype or a modifying factor, such as an additional potentially pathogenic variant, presence of the variant in a population database, heteroplasmy for a mitochondrial variant or mosaicism for the JMJD1C variant. Although the de novo variants in JMJD1C are likely to be relevant to the developmental phenotypes observed in these patients, we conclude that further data supporting the association of JMJD1C variants with intellectual disability is still needed.

Long-term adult human brain slice cultures as a model system to study human CNS circuitry and disease.

Most of our knowledge on human CNS circuitry and related disorders originates from model organisms. How well such data translate to the human CNS remains largely to be determined. Human brain slice cultures derived from neurosurgical resections may offer novel avenues to approach this translational gap. We now demonstrate robust preservation of the complex neuronal cytoarchitecture and electrophysiological properties of human pyramidal neurons in long-term brain slice cultures. Further experiments delineate the optimal conditions for efficient viral transduction of cultures, enabling 'high throughput' fluorescence-mediated 3D reconstruction of genetically targeted neurons at comparable quality to state-of-the-art biocytin fillings, and demonstrate feasibility of long term live cell imaging of human cells in vitro. This model system has implications toward a broad spectrum of translational studies, regarding the validation of data obtained in non-human model systems, for therapeutic screening and genetic dissection of human CNS circuitry.

Changes in drug load during lacosamide combination therapy: A noninterventional, observational study in German and Austrian clinical practice.

INTRODUCTION: Effects of antiepileptic drug (AED) load changes in patients with focal seizures have not been well evaluated. METHODS: SP1065 (NCT01673282) was a noninterventional, prospective, observational study conducted in a clinical practice setting. Patients (aged ≥18 years) with focal seizures were enrolled within 7 days of being prescribed adjunctive lacosamide. Observation period was ~6 months. Drug load was assessed using percentage change in ratio of actual prescribed dose and World Health Organization defined daily dose (DDD) for concomitant AEDs and all AEDs (including lacosamide). Subgroups were defined for patients with at least one concomitant sodium channel-blocking AED (SCB [+]) and those without (SCB [-]). RESULTS: A total of 311 patients were assessed for safety, 302 for measurement of drug load, and 240 for effectiveness. Ratio of AED dose to DDD decreased for concomitant AEDs (-9.6%) and increased for all AEDs (including lacosamide; 15.5%). Median reduction in focal seizure frequency per 28 days was 100% (range: -100, 2275.8). 70.4% and 61.7% of patients had a ≥50% or ≥75% reduction in seizure frequency, respectively; 50.8% became seizure-free. In the SCB (+) subgroup (n = 149), ratio of AED dose to DDD decreased for concomitant AEDs (-15.0%) and increased for all AEDs (10.7%). In the SCB (-) subgroup (n = 153), ratio of AED dose to DDD decreased for concomitant AEDs (-4.4%) and increased for all AEDs (20.2%). Fifty-seven patients (18.3%) reported ADRs, most commonly dose >400 mg/d (7.1%). Seventeen patients (5.5%) had ADRs leading to discontinuation. SIGNIFICANCE: Addition of lacosamide resulted in reduction of concomitant AED drug load regardless of whether concomitant AEDs were SCB (+) or SCB (-). These results indicate that addition of lacosamide in patients with focal seizures could allow clinicians to withdraw or reduce the dose of less well-tolerated or less effective AEDs.

Assessment of genetic variant burden in epilepsy-associated brain lesions.

It is challenging to estimate genetic variant burden across different subtypes of epilepsy. Herein, we used a comparative approach to assess the genetic variant burden and genotype-phenotype correlations in four most common brain lesions in patients with drug-resistant focal epilepsy. Targeted sequencing analysis was performed for a panel of 161 genes with a mean coverage of >400×. Lesional tissue was histopathologically reviewed and dissected from hippocampal sclerosis (n = 15), ganglioglioma (n = 16), dysembryoplastic neuroepithelial tumors (n = 8), and focal cortical dysplasia type II (n = 15). Peripheral blood (n = 12) or surgical tissue samples histopathologically classified as lesion-free (n = 42) were available for comparison. Variants were classified as pathogenic or likely pathogenic according to American College of Medical Genetics and Genomics guidelines. Overall, we identified pathogenic and likely pathogenic variants in 25.9% of patients with a mean coverage of 383×. The highest number of pathogenic/likely pathogenic variants was observed in patients with ganglioglioma (43.75%; all somatic) and dysembryoplastic neuroepithelial tumors (37.5%; all somatic), and in 20% of cases with focal cortical dysplasia type II (13.33% somatic, 6.67% germline). Pathogenic/likely pathogenic positive genes were disorder specific and BRAF V600E the only recurrent pathogenic variant. This study represents a reference for the genetic variant burden across the four most common lesion entities in patients with drug-resistant focal epilepsy. The observed large variability in variant burden by epileptic lesion type calls for whole exome sequencing of histopathologically well-characterized tissue in a diagnostic setting and in research to discover novel disease-associated genes.

A Recurrent Missense Variant in AP2M1 Impairs Clathrin-Mediated Endocytosis and Causes Developmental and Epileptic Encephalopathy.

The developmental and epileptic encephalopathies (DEEs) are heterogeneous disorders with a strong genetic contribution, but the underlying genetic etiology remains unknown in a significant proportion of individuals. To explore whether statistical support for genetic etiologies can be generated on the basis of phenotypic features, we analyzed whole-exome sequencing data and phenotypic similarities by using Human Phenotype Ontology (HPO) in 314 individuals with DEEs. We identified a de novo c.508C>T (p.Arg170Trp) variant in AP2M1 in two individuals with a phenotypic similarity that was higher than expected by chance (p = 0.003) and a phenotype related to epilepsy with myoclonic-atonic seizures. We subsequently found the same de novo variant in two individuals with neurodevelopmental disorders and generalized epilepsy in a cohort of 2,310 individuals who underwent diagnostic whole-exome sequencing. AP2M1 encodes the µ-subunit of the adaptor protein complex 2 (AP-2), which is involved in clathrin-mediated endocytosis (CME) and synaptic vesicle recycling. Modeling of protein dynamics indicated that the p.Arg170Trp variant impairs the conformational activation and thermodynamic entropy of the AP-2 complex. Functional complementation of both the µ-subunit carrying the p.Arg170Trp variant in human cells and astrocytes derived from AP-2µ conditional knockout mice revealed a significant impairment of CME of transferrin. In contrast, stability, expression levels, membrane recruitment, and localization were not impaired, suggesting a functional alteration of the AP-2 complex as the underlying disease mechanism. We establish a recurrent pathogenic variant in AP2M1 as a cause of DEEs with distinct phenotypic features, and we implicate dysfunction of the early steps of endocytosis as a disease mechanism in epilepsy.

Targeted gene sequencing in 6994 individuals with neurodevelopmental disorder with epilepsy.

PURPOSE: We aimed to gain insight into frequencies of genetic variants in genes implicated in neurodevelopmental disorder with epilepsy (NDD+E) by investigating large cohorts of patients in a diagnostic setting. METHODS: We analyzed variants in NDD+E using epilepsy gene panel sequencing performed between 2013 and 2017 by two large diagnostic companies. We compared variant frequencies in 6994 panels with another 8588 recently published panels as well as exome-wide de novo variants in 1942 individuals with NDD+E and 10,937 controls. RESULTS: Genes with highest frequencies of ultrarare variants in NDD+E comprised SCN1A, KCNQ2, SCN2A, CDKL5, SCN8A, and STXBP1, concordant with the two other epilepsy cohorts we investigated. In only 46% of the analyzed 262 dominant and X-linked panel genes ultrarare variants in patients were reported. Among genes with contradictory evidence of association with epilepsy, CACNB4, CLCN2, EFHC1, GABRD, MAGI2, and SRPX2 showed equal frequencies in cases and controls. CONCLUSION: We show that improvement of panel design increased diagnostic yield over time, but panels still display genes with low or no diagnostic yield. With our data, we hope to improve current diagnostic NDD+E panel design and provide a resource of ultrarare variants in individuals with NDD+E to the community.

Clinical spectrum of STX1B-related epileptic disorders.

OBJECTIVE: The aim of this study was to expand the spectrum of epilepsy syndromes related to STX1B, encoding the presynaptic protein syntaxin-1B, and establish genotype-phenotype correlations by identifying further disease-related variants. METHODS: We used next-generation sequencing in the framework of research projects and diagnostic testing. Clinical data and EEGs were reviewed, including already published cases. To estimate the pathogenicity of the variants, we used established and newly developed in silico prediction tools. RESULTS: We describe 17 new variants in STX1B, which are distributed across the whole gene. We discerned 4 different phenotypic groups across the newly identified and previously published patients (49 patients in 23 families): (1) 6 sporadic patients or families (31 affected individuals) with febrile and afebrile seizures with a benign course, generally good drug response, normal development, and without permanent neurologic deficits; (2) 2 patients with genetic generalized epilepsy without febrile seizures and cognitive deficits; (3) 13 patients or families with intractable seizures, developmental regression after seizure onset and additional neuropsychiatric symptoms; (4) 2 patients with focal epilepsy. More often, we found loss-of-function mutations in benign syndromes, whereas missense variants in the SNARE motif of syntaxin-1B were associated with more severe phenotypes. CONCLUSION: These data expand the genetic and phenotypic spectrum of STX1B-related epilepsies to a diverse range of epilepsies that span the International League Against Epilepsy classification. Variants in STX1B are protean and contribute to many different epilepsy phenotypes, similar to SCN1A, the most important gene associated with fever-associated epilepsies.

No evidence for a BRD2 promoter hypermethylation in blood leukocytes of Europeans with juvenile myoclonic epilepsy.

Juvenile myoclonic epilepsy (JME) is a common syndrome of genetic generalized epilepsies (GGEs). Linkage and association studies suggest that the gene encoding the bromodomain-containing protein 2 (BRD2) may increase risk of JME. The present methylation and association study followed up a recent report highlighting that the BRD2 promoter CpG island (CpG76) is differentially hypermethylated in lymphoblastoid cells from Caucasian patients with JME compared to patients with other GGE subtypes and unaffected relatives. In contrast, we found a uniform low average percentage of methylation (<4.5%) for 13 CpG76-CpGs in whole blood cells from 782 unrelated European Caucasians, including 116 JME patients, 196 patients with genetic absence epilepsies, and 470 control subjects. We also failed to confirm an allelic association of the BRD2 promoter single nucleotide polymorphism (SNP) rs3918149 with JME (Armitage trend test, P = 0.98), and we did not detect a substantial impact of SNP rs3918149 on CpG76 methylation in either 116 JME patients (methylation quantitative trait loci [meQTL], P = 0.29) or 470 German control subjects (meQTL, P = 0.55). Our results do not support the previous observation that a high DNA methylation level of the BRD2 promoter CpG76 island is a prevalent epigenetic motif associated with JME in Caucasians.

Neuronal mechanisms of mutations in SCN8A causing epilepsy or intellectual disability.

Ion channel mutations can cause distinct neuropsychiatric diseases. We first studied the biophysical and neurophysiological consequences of four mutations in the human Na+ channel gene SCN8A causing either mild (E1483K) or severe epilepsy (R1872W), or intellectual disability and autism without epilepsy (R1620L, A1622D). Only combined electrophysiological recordings of transfected wild-type or mutant channels in both neuroblastoma cells and primary cultured neurons revealed clear genotype-phenotype correlations. The E1483K mutation causing mild epilepsy showed no significant biophysical changes, whereas the R1872W mutation causing severe epilepsy induced clear gain-of-function biophysical changes in neuroblastoma cells. However, both mutations increased neuronal firing in primary neuronal cultures. In contrast, the R1620L mutation associated with intellectual disability and autism-but not epilepsy-reduced Na+ current density in neuroblastoma cells and expectedly decreased neuronal firing. Interestingly, for the fourth mutation, A1622D, causing severe intellectual disability and autism without epilepsy, we observed a dramatic slowing of fast inactivation in neuroblastoma cells, which induced a depolarization block in neurons with a reduction of neuronal firing. This latter finding was corroborated by computational modelling. In a second series of experiments, we recorded three more mutations (G1475R, M1760I, G964R, causing intermediate or severe epilepsy, or intellectual disability without epilepsy, respectively) that revealed similar results confirming clear genotype-phenotype relationships. We found intermediate or severe gain-of-function biophysical changes and increases in neuronal firing for the two epilepsy-causing mutations and decreased firing for the loss-of-function mutation causing intellectual disability. We conclude that studies in neurons are crucial to understand disease mechanisms, which here indicate that increased or decreased neuronal firing is responsible for distinct clinical phenotypes.

The glucose transporter type 1 (Glut1) syndromes.

The glucose transporter type 1 (Glut1) is the most important energy carrier of the brain across the blood-brain barrier. In the early nineties, the first genetic defect of Glut1 was described and known as the Glut1 deficiency syndrome (Glut1-DS). It is characterized by early infantile seizures, developmental delay, microcephaly, and ataxia. Recently, milder variants have also been described. The clinical picture of Glut1 defects and the understanding of the pathophysiology of this disease have significantly grown. A special form of transient movement disorders, the paroxysmal exertion-induced dyskinesia (PED), absence epilepsies particularly with an early onset absence epilepsy (EOAE) and childhood absence epilepsy (CAE), myoclonic astatic epilepsy (MAE), episodic choreoathetosis and spasticity (CSE), and focal epilepsy can be based on a Glut1 defect. Despite the rarity of these diseases, the Glut1 syndromes are of high clinical interest since a very effective therapy, the ketogenic diet, can improve or reverse symptoms especially if it is started as early as possible. The present article summarizes the clinical features of Glut1 syndromes and discusses the underlying genetic mutations, including the available data on functional tests as well as the genotype-phenotype correlations. This article is part of the Special Issue "Individualized Epilepsy Management: Medicines, Surgery and Beyond".