Type I Interferonopathy due to a Homozygous Loss-of-Inhibitory Function Mutation in STAT2.

PURPOSE: STAT2 is both an effector and negative regulator of type I interferon (IFN-I) signalling. We describe the characterization of a novel homozygous missense STAT2 substitution in a patient with a type I interferonopathy. METHODS: Whole-genome sequencing (WGS) was used to identify the genetic basis of disease in a patient with features of enhanced IFN-I signalling. After stable lentiviral reconstitution of STAT2-null human fibrosarcoma U6A cells with STAT2 wild type or p.(A219V), we performed quantitative polymerase chain reaction, western blotting, immunofluorescence, and co-immunoprecipitation to functionally characterize the p.(A219V) variant. RESULTS: WGS identified a rare homozygous single nucleotide transition in STAT2 (c.656C > T), resulting in a p.(A219V) substitution, in a patient displaying developmental delay, intracranial calcification, and up-regulation of interferon-stimulated gene (ISG) expression in blood. In vitro studies revealed that the STAT2 p.(A219V) variant retained the ability to transduce an IFN-I stimulus. Notably, STAT2 p.(A219V) failed to support receptor desensitization, resulting in sustained STAT2 phosphorylation and ISG up-regulation. Mechanistically, STAT2 p.(A219V) showed defective binding to ubiquitin specific protease 18 (USP18), providing a possible explanation for the chronic IFN-I pathway activation seen in the patient. CONCLUSION: Our data indicate an impaired negative regulatory role of STAT2 p.(A219V) in IFN-I signalling and that mutations in STAT2 resulting in a type I interferonopathy state are not limited to the previously reported R148 residue. Indeed, structural modelling highlights at least 3 further residues critical to mediating a STAT2-USP18 interaction, in which mutations might be expected to result in defective negative feedback regulation of IFN-I signalling.

Remarkable effect of transdermal nicotine in children with CHRNA4-related autosomal dominant sleep-related hypermotor epilepsy.

OBJECTIVE: Autosomal dominant sleep-related hypermotor epilepsy (ADSHE) is characterized by hypermotor seizures and may be caused by gain-of-function mutations affecting the nicotinic acetylcholine receptor (nAChR). Benefit from nicotine consumption has been reported in adult patients with this disorder. For the first time, the effect of transdermal nicotine is evaluated in children. METHODS: Transdermal nicotine was applied to three boys, two aged 10 years (7 mg/24 h) and one six years (3.5 mg/24 h). Autosomal dominant sleep-related hypermotor epilepsy was caused by the p.S280F-CHRNA4 (cholinergic receptor, nicotinic, alpha polypeptide 4) mutation. The children suffered from frequent, persistent nocturnal seizures and had developed educational and psychosocial problems. Seizure frequency and cognitive and behavioral parameters were assessed before and after treatment. RESULTS: A striking seizure reduction was reported soon after treatment onset. Hypermotor seizures disappeared; only sporadic arousals, sometimes with minor motor elements, were observed. Psychometric testing documented improvement in cognitive domains such as visuospatial ability, processing speed, memory, and some areas of executive functions. SIGNIFICANCE: Nicotine appears to be a mechanistic treatment for this specific disorder, probably because of desensitization of the mutated receptors. It may control seizures resistant to conventional drugs for epilepsy and impact socioeducational function in children. This mode of precision therapy should receive more attention and should be available to more patients with uncontrolled CHRNA4-related ADSHE across the age span.

KCNT1 epilepsy with migrating focal seizures shows a temporal sequence with poor outcome, high mortality and SUDEP.

Epilepsy of infancy with migrating focal seizures was first described in 1995. Fifteen years later, KCNT1 gene mutations were identified as the major disease-causing gene of this disease. Currently, the data on epilepsy of infancy with migrating focal seizures associated with KCNT1 mutations are heterogeneous and many questions remain unanswered including the prognosis and the long-term outcome especially regarding epilepsy, neurological and developmental status and the presence of microcephaly. The aim of this study was to assess data from patients with epilepsy in infancy with migrating focal seizures with KCNT1 mutations to refine the phenotype spectrum and the outcome. We used mind maps based on medical reports of children followed in the network of the French reference centre for rare epilepsies and we developed family surveys to assess the long-term outcome. Seventeen patients were included [age: median (25th-75th percentile): 4 (2-15) years, sex ratio: 1.4, length of follow-up: 4 (2-15) years]. Seventy-one per cent started at 6 (1-52) days with sporadic motor seizures (n = 12), increasing up to a stormy phase with long lasting migrating seizures at 57 (30-89) days. The others entered this stormy phase directly at 1 (1-23) day. Ten patients entered a consecutive phase at 1.3 (1-2.8) years where seizures persisted at least daily (n = 8), but presented different semiology: brief and hypertonic with a nocturnal (n = 6) and clustered (n = 6) aspects. Suppression interictal patterns were identified on the EEG in 71% of patients (n = 12) sometimes from the first EEG (n = 6). Three patients received quinidine without reported efficacy. Long-term outcome was poor with neurological sequelae and active epilepsy except for one patient who had an early and long-lasting seizure-free period. Extracerebral symptoms probably linked with KCNT1 mutation were present, including arteriovenous fistula, dilated cardiomyopathy and precocious puberty. Eight patients (47%) had died at 3 (1.5-15.4) years including three from suspected sudden unexpected death in epilepsy. Refining the electro-clinical characteristics and the temporal sequence of epilepsy in infancy with migrating focal seizures should help diagnosis of this epilepsy. A better knowledge of the outcome allows one to advise families and to define the appropriate follow-up and therapies. Extracerebral involvement should be investigated, in particular the cardiac system, as it may be involved in the high prevalence of sudden unexpected death in epilepsy in these cases.

De novo loss-of-function KCNMA1 variants are associated with a new multiple malformation syndrome and a broad spectrum of developmental and neurological phenotypes.

KCNMA1 encodes the large-conductance Ca2+- and voltage-activated K+ (BK) potassium channel α-subunit, and pathogenic gain-of-function variants in this gene have been associated with a dominant form of generalized epilepsy and paroxysmal dyskinesia. Here, we genetically and functionally characterize eight novel loss-of-function (LoF) variants of KCNMA1. Genome or exome sequencing and the participation in the international Matchmaker Exchange effort allowed for the identification of novel KCNMA1 variants. Patch clamping was used to assess functionality of mutant BK channels. The KCNMA1 variants p.(Ser351Tyr), p.(Gly356Arg), p.(Gly375Arg), p.(Asn449fs) and p.(Ile663Val) abolished the BK current, whereas p.(Cys413Tyr) and p.(Pro805Leu) reduced the BK current amplitude and shifted the activation curves toward positive potentials. The p.(Asp984Asn) variant reduced the current amplitude without affecting kinetics. A phenotypic analysis of the patients carrying the recurrent p.(Gly375Arg) de novo missense LoF variant revealed a novel syndromic neurodevelopmental disorder associated with severe developmental delay, visceral and cardiac malformations, connective tissue presentations with arterial involvement, bone dysplasia and characteristic dysmorphic features. Patients with other LoF variants presented with neurological and developmental symptoms including developmental delay, intellectual disability, ataxia, axial hypotonia, cerebral atrophy and speech delay/apraxia/dysarthria. Therefore, LoF KCNMA1 variants are associated with a new syndrome characterized by a broad spectrum of neurological phenotypes and developmental disorders. LoF variants of KCNMA1 cause a new syndrome distinctly different from gain-of-function variants in the same gene.

The landscape of epilepsy-related GATOR1 variants.

PURPOSE: To define the phenotypic and mutational spectrum of epilepsies related to DEPDC5, NPRL2 and NPRL3 genes encoding the GATOR1 complex, a negative regulator of the mTORC1 pathway METHODS: We analyzed clinical and genetic data of 73 novel probands (familial and sporadic) with epilepsy-related variants in GATOR1-encoding genes and proposed new guidelines for clinical interpretation of GATOR1 variants. RESULTS: The GATOR1 seizure phenotype consisted mostly in focal seizures (e.g., hypermotor or frontal lobe seizures in 50%), with a mean age at onset of 4.4 years, often sleep-related and drug-resistant (54%), and associated with focal cortical dysplasia (20%). Infantile spasms were reported in 10% of the probands. Sudden unexpected death in epilepsy (SUDEP) occurred in 10% of the families. Novel classification framework of all 140 epilepsy-related GATOR1 variants (including the variants of this study) revealed that 68% are loss-of-function pathogenic, 14% are likely pathogenic, 15% are variants of uncertain significance and 3% are likely benign. CONCLUSION: Our data emphasize the increasingly important role of GATOR1 genes in the pathogenesis of focal epilepsies (>180 probands to date). The GATOR1 phenotypic spectrum ranges from sporadic early-onset epilepsies with cognitive impairment comorbidities to familial focal epilepsies, and SUDEP.

Quantitative analysis and EEG markers of KCNT1 epilepsy of infancy with migrating focal seizures.

OBJECTIVE: We aimed to characterize epilepsy of infancy with migrating focal seizures (EIMFS), a rare, severe early onset developmental epilepsy related to KCNT1 mutation, and to define specific electroencephalography (EEG) markers using EEG quantitative analysis. The ultimate goal would be to improve early diagnosis and to better understand seizure onset and propagation of EIMFS as compared to other early onset developmental epilepsy. METHODS: EEG of 7 EIMFS patients with KCNT1 mutations (115 seizures) and 17 patients with other early onset epilepsies (30 seizures) was included in this study. After detection of seizure onset and termination, spatiotemporal characteristics were quantified. Seizure propagation dynamics were analyzed using chronograms and phase coherence. RESULTS: In patients with EIMFS, seizures started and were localized predominantly in temporal and occipital areas, and evolved with a stable frequency (4-10 Hz). Inter- and intrahemispheric migrations were present in 60% of EIMFS seizures with high intraindividual reproducibility of temporospatial dynamics. Interhemispheric migrating seizures spread in 71% from temporal or occipital channels to the homologous contralateral ones, whereas intrahemispheric seizures involved mainly frontotemporal, temporal, and occipital channels. Causality links were present between ictal activities detected under different channels during migrating seizures. Finally, time delay index (based on delays between the different ictal onsets) and phase correlation index (based on coherence of ictal activities) allowed discrimination of EIMFS and non-EIMFS seizures with a specificity of 91.2% and a sensitivity of 84.4%. SIGNIFICANCE: We showed that the migrating pattern in EIMFS is not a random process, as suggested previously, and that it is a particular propagation pattern that follows the classical propagation pathways. It is notable that this study reveals specific EEG markers (time delay and phase correlation) accessible to visual evaluation, which will improve EIMFS diagnosis.

Homozygous Truncating Variants in TBC1D23 Cause Pontocerebellar Hypoplasia and Alter Cortical Development.

Pontocerebellar hypoplasia (PCH) is a heterogeneous group of rare recessive disorders with prenatal onset, characterized by hypoplasia of pons and cerebellum. Mutations in a small number of genes have been reported to cause PCH, and the vast majority of PCH cases are explained by mutations in TSEN54, which encodes a subunit of the tRNA splicing endonuclease complex. Here we report three families with homozygous truncating mutations in TBC1D23 who display moderate to severe intellectual disability and microcephaly. MRI data from available affected subjects revealed PCH, small normally proportioned cerebellum, and corpus callosum anomalies. Furthermore, through in utero electroporation, we show that downregulation of TBC1D23 affects cortical neuron positioning. TBC1D23 is a member of the Tre2-Bub2-Cdc16 (TBC) domain-containing RAB-specific GTPase-activating proteins (TBC/RABGAPs). Members of this protein family negatively regulate RAB proteins and modulate the signaling between RABs and other small GTPases, some of which have a crucial role in the trafficking of intracellular vesicles and are involved in neurological disorders. Here, we demonstrate that dense core vesicles and lysosomal trafficking dynamics are affected in fibroblasts harboring TBC1D23 mutation. We propose that mutations in TBC1D23 are responsible for a form of PCH with small, normally proportioned cerebellum and should be screened in individuals with syndromic pontocereballar hypoplasia.

Mutations in MDH2, Encoding a Krebs Cycle Enzyme, Cause Early-Onset Severe Encephalopathy.

MDH2 encodes mitochondrial malate dehydrogenase (MDH), which is essential for the conversion of malate to oxaloacetate as part of the proper functioning of the Krebs cycle. We report bi-allelic pathogenic mutations in MDH2 in three unrelated subjects presenting with early-onset generalized hypotonia, psychomotor delay, refractory epilepsy, and elevated lactate in the blood and cerebrospinal fluid. Functional studies in fibroblasts from affected subjects showed both an apparently complete loss of MDH2 levels and MDH2 enzymatic activity close to null. Metabolomics analyses demonstrated a significant concomitant accumulation of the MDH substrate, malate, and fumarate, its immediate precursor in the Krebs cycle, in affected subjects' fibroblasts. Lentiviral complementation with wild-type MDH2 cDNA restored MDH2 levels and mitochondrial MDH activity. Additionally, introduction of the three missense mutations from the affected subjects into Saccharomyces cerevisiae provided functional evidence to support their pathogenicity. Disruption of the Krebs cycle is a hallmark of cancer, and MDH2 has been recently identified as a novel pheochromocytoma and paraganglioma susceptibility gene. We show that loss-of-function mutations in MDH2 are also associated with severe neurological clinical presentations in children.

Epilepsy in young Tsc1(+/-) mice exhibits age-dependent expression that mimics that of human tuberous sclerosis complex.

OBJECTIVE: To describe the epileptic phenotype of Tsc1(+/-) mice pups in comparison with age-related seizures in human tuberous sclerosis complex (TSC). METHODS: Tsc1(+/-) and control mice underwent intracranial electroencephalography (EEG) recording at postnatal ages (P)8 to P33, with linear silicon probe implanted in the somatosensory cortex of one or both hemispheres for 8-24 h. Ictal events were classified visually by independent analyzers; distinct EEG patterns were related to age and analyzed to quantify field potential characteristics and signal dynamics between hemispheres. We collected retrospectively 20 infants with prenatally diagnosed TSC and EEG before seizure onset, and analyzed the electroclinical course of epilepsy, taking into account a first-line treatment by vigabatrin. RESULTS: Spontaneous seizures were disclosed in 55% of Tsc1(+/-) mice at P9-18. Three ictal patterns were identified: from P9 to P12 "spike clusters" consisted of recurring large spikes without clinical correlate; "spasm-like" discharges dominated from P13 to P16 consisting of high amplitude large field potential superimposed with or followed by fast activity repeated every 2-10 s for at least 20 s, accompanied by rhythmic limb contractions; from P14 to P18 a "tonic-clonic like" pattern comprised rhythmic spikes of increasing amplitude with tonic-clonic movements. Early onset "spike clusters" were mainly unilateral, whereas "spasm-like" and "tonic-clonic like" patterns were bilateral. Interhemispheric propagation was significantly faster for "tonic-clonic like" than for "spasm-like" events. In infants diagnosed prenatally with TSC, clusters of sharp waves or spikes preceded the first seizure, and vigabatrin prevented the development of seizures. Patients treated after seizure onset developed spasms or focal seizures that were pharmacoresistant in 66.7% of cases. SIGNIFICANCE: Tsc1(+/-) mice pups exhibit an age-dependent seizure pattern sequence mimicking early human TSC epilepsy features. Spike clusters before seizure onset in TSC should be considered as a first stage of epilepsy reinforcing the concept of preventive antiepileptic therapy.

Intelligence quotient improves after antiepileptic drug withdrawal following pediatric epilepsy surgery.

OBJECTIVE: Antiepileptic drugs (AEDs) have cognitive side effects that, particularly in children, may affect intellectual functioning. With the TimeToStop (TTS) study, we showed that timing of AED withdrawal does not majorly influence long-term seizure outcomes. We now aimed to evaluate the effect of AED withdrawal on postoperative intelligence quotient (IQ), and change in IQ (delta IQ) following pediatric epilepsy surgery. METHODS: We collected IQ scores of children from the TTS cohort with both pre- and postoperative neuropsychological assessments (NPAs; n = 301) and analyzed whether reduction of AEDs prior to the latest NPA was related to postoperative IQ and delta IQ, using linear regression analyses. Factors previously identified as independently relating to (delta) IQ, and currently identified predictors of (delta) IQ, were considered possible confounders and used for adjustment. Additionally, we adjusted for a compound propensity score that contained previously identified determinants of timing of AED withdrawal. RESULTS: Mean interval to the latest NPA was 19.8 ± 18.9 months. Reduction of AEDs at the latest NPA significantly improved postoperative IQ and delta IQ (adjusted regression coefficient [RC] = 3.4, 95% confidence interval [CI] = 0.6-6.2, p = 0.018 and RC = 4.5, 95% CI = 1.7-7.4, p = 0.002), as did complete withdrawal (RC = 4.8, 95% CI = 1.4-8.3, p = 0.006 and RC = 5.1, 95% CI = 1.5-8.7, p = 0.006). AED reduction also predicted ≥ 10-point IQ increase (p = 0.019). The higher the number of AEDs reduced, the higher was the IQ (gain) after surgery (RC = 2.2, 95% CI = 0.6-3.7, p = 0.007 and RC = 2.6, 95% CI = 1.0-4.2, p = 0.001, IQ points per AED reduced). INTERPRETATION: Start of AED withdrawal, number of AEDs reduced, and complete AED withdrawal were associated with improved postoperative IQ scores and gain in IQ, independent of other determinants of cognitive outcome.

ARX polyalanine expansions are highly implicated in familial cases of mental retardation with infantile epilepsy and/or hand dystonia.

Mutations in the ARX gene cause both nonsyndromic and several forms of syndromic mental retardation (MR). Two polyalanine (polyA) expansions of ARX are recurrent mutations. The most common one, the c.428_451dup, is associated with a wide spectrum of phenotypes, ranging from the most severe West syndrome to Partington syndrome (MR and hand dystonia), and even nonsyndromic X-linked mental retardation (NS-XLMR). Studies of patients not selected for specific clinical signs showed that the c.428_451dup is relatively frequent in families harboring X-linked MR (7.5%), but less common in familial cases compatible with X-linked NR (1%), and very rare in sporadic cases (0.1%). The c.333_334ins(GCG)7 expansion is less frequent and mainly associated with West syndrome. We screened for both ARX polyA expansions in 98 unrelated patients selected for the presence of NR associated with different types of epilepsy and/or with hand dystonia. We also studied two families with an initial diagnosis of NS-XLMR, one of which was identified as showing linkage to the ARX locus. The c.428_451dup was identified in three patients and the c.333_334ins(GCG)7 in one; all of the patients were from families with two affected brothers. We also found the c.428_451dup in the family linked to ARX, and clinical re-evaluation showed subtle, previously undetected signs. Our study illustrates that ARX polyA expansions are primarily associated with syndromic MR and shows a higher yield (18% in our cohort) when these mutations are screened in familial cases of MR with epilepsy and/or dystonia.

SRPX2 mutations in disorders of language cortex and cognition.

The rolandic and sylvian fissures divide the human cerebral hemispheres and the adjacent areas participate in speech processing. The relationship of rolandic (sylvian) seizure disorders with speech and cognitive impairments is well known, albeit poorly understood. We have identified the Xq22 gene SRPX2 as being responsible for rolandic seizures (RSs) associated with oral and speech dyspraxia and mental retardation (MR). SRPX2 is a secreted sushi-repeat containing protein expressed in neurons of the human adult brain, including the rolandic area. The disease-causing mutation (N327S) resulted in gain-of-glycosylation of the secreted mutant protein. A second mutation (Y72S) was identified within the first sushi domain of SRPX2 in a male with RSs and bilateral perisylvian polymicrogyria and his female relatives with mild MR or unaffected carrier status. In cultured cells, both mutations were associated with altered patterns of intracellular processing, suggesting protein misfolding. In the murine brain, Srpx2 protein expression appeared in neurons at birth. The involvement of SRPX2 in these disorders suggests an important role for SRPX2 in the perisylvian region critical for language and cognitive development.

Antiglial cell autoantibodies and childhood epilepsy: a case report.

We report the case of a patient with severe partial epilepsy associated with a focal rolandic, pathologically proven, cortical dysplasia. By measuring intracellular calcium concentrations, functional antiglial non-glutamate receptor 3 (GluR3) autoantibodies were identified in the serum of this patient. Antineuronal autoantibodies, which interfere with GluR3-receptor function, also were detected. This observation provides new clues about the involvement of immunologic mechanisms in epileptic disorders.

Antiepileptic effect of high-frequency stimulation of the subthalamic nucleus (corpus luysi) in a case of medically intractable epilepsy caused by focal dysplasia: a 30-month follow-up: technical case report.

OBJECTIVE AND IMPORTANCE: Currently, some forms of epilepsy are resistant to both pharmacological and surgical interventions. As a result, there is a need for new therapeutic strategies. Because the nigral system modulates neuronal excitability in animal models of epilepsy, we considered therapeutic high-frequency stimulation of the subthalamic nucleus (STN). We were encouraged by the known relationship between the STN and the nigral system, as well as by our experience with high-frequency stimulation of the STN in Parkinsonian patients. CLINICAL PRESENTATION: A 5-year-old girl with pharmacologically resistant, inoperable epilepsy caused by focal centroparietal dysplasia underwent implantation with a permanent electrode in the left STN and was chronically stimulated. To date, we have followed up this patient for 30 months postoperatively. TECHNIQUE: High-frequency stimulation of the STN induced a significant voltage-dependent reduction (by 80%) in the number and severity of seizures. In addition, consistent improvement in both motor and cognitive functions was noted as a result of reduced postictal states. The effect was more prominent for seizures occurring in clusters (89% reduction) and during the day (88% reduction) than for those that occurred during sleep (53% reduction). CONCLUSION: This is the first report of epilepsy control using chronic high-frequency stimulation of the STN. Preliminary observations in three other operated patients (at 2, 12, and 18 mo) confirm these data. We think that high-frequency stimulation of the STN may hold significant future potential as a treatment for epilepsy, similar to its established role in the treatment of Parkinson's disease. This finding opens completely new experimental and therapeutic avenues for the treatment of surgically and medically intractable epilepsy.