Genetic malformations of cortical development (MCDs), such as mild MCDs (mMCD), focal cortical dysplasia (FCD), and hemimegalencephaly (HME), are major causes of severe pediatric refractory epilepsies subjected to neurosurgery. FCD2 are characterized by neuropathological hallmarks that include enlarged dysmorphic neurons (DNs) and balloon cells (BCs). Here, we provide a comprehensive assessment of the contribution of germline and somatic variants in a large cohort of surgical MCD cases. We enrolled in a monocentric study 80 children with drug-resistant epilepsy and a postsurgical neuropathological diagnosis of mMCD, FCD1, FCD2, or HME. We performed targeted gene sequencing (≥ 2000X read depth) on matched blood-brain samples to search for low-allele frequency variants in mTOR pathway and FCD genes. We were able to elucidate 29% of mMCD/FCD1 patients and 63% of FCD2/HME patients. Somatic loss-of-function variants in the N-glycosylation pathway-associated SLC35A2 gene were found in mMCD/FCD1 cases. Somatic gain-of-function variants in MTOR and its activators (AKT3, PIK3CA, RHEB), as well as germline, somatic and two-hit loss-of-function variants in its repressors (DEPDC5, TSC1, TSC2) were found exclusively in FCD2/HME cases. We show that panel-negative FCD2 cases display strong pS6-immunostaining, stressing that all FCD2 are mTORopathies. Analysis of microdissected cells demonstrated that DNs and BCs carry the pathogenic variants. We further observed a correlation between the density of pathological cells and the variant-detection likelihood. Single-cell microdissection followed by sequencing of enriched pools of DNs unveiled a somatic second-hit loss-of-heterozygosity in a DEPDC5 germline case. In conclusion, this study indicates that mMCD/ FCD1 and FCD2/HME are two distinct genetic entities: while all FCD2/HME are mosaic mTORopathies, mMCD/FCD1 are not caused by mTOR-pathway-hyperactivating variants, and ~ 30% of the cases are related to glycosylation defects. We provide a framework for efficient genetic testing in FCD/HME, linking neuropathology to genetic findings and emphasizing the usefulness of molecular evaluation in the pediatric epileptic neurosurgical population.
A full list of authors and affiliations appears at the end of the paper.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 drugresistant (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.Genetics in Medicine (2018) https://doi
DEP domain-containing 5 protein (DEPDC5) is a repressor of the recently recognized amino acid-sensing branch of the mTORC1 pathway. So far, its function in the brain remains largely unknown. Germline loss-of-function mutations in DEPDC5 have emerged as a major cause of familial refractory focal epilepsies, with case reports of sudden unexpected death in epilepsy (SUDEP). Remarkably, a fraction of patients also develop focal cortical dysplasia (FCD), a neurodevelopmental cortical malformation. We therefore hypothesized that a somatic second-hit mutation arising during brain development may support the focal nature of the dysplasia. Here, using postoperative human tissue, we provide the proof of concept that a biallelic 2-hit - brain somatic and germline - mutational mechanism in DEPDC5 causes focal epilepsy with FCD. We discovered a mutation gradient with a higher rate of mosaicism in the seizure-onset zone than in the surrounding epileptogenic zone. Furthermore, we demonstrate the causality of a Depdc5 brain mosaic inactivation using CRISPR-Cas9 editing and in utero electroporation in a mouse model recapitulating focal epilepsy with FCD and SUDEP-like events. We further unveil a key role of Depdc5 in shaping dendrite and spine morphology of excitatory neurons. This study reveals promising therapeutic avenues for treating drug-resistant focal epilepsies with mTORC1-targeting molecules.
BackgroundDravet syndrome (DS) is currently considered as an epileptic encephalopathy, a condition in which epilepsy causes deterioration or developmental delay but preliminary data suggested that cognitive course may worsen independently from epilepsy. Our objective was to prospectively analyze the neuropsychological features in a large cohort of DS patients and its relationships with epilepsy and SCN1A mutation.Methods81 examinations were performed in 67 patients with typical DS (9m-24y, 15 longitudinally studied) using Brunet-Lezine (developmental/intelligence quotient [DQ/IQ] and DQ sub-scores), Achenbach, Conners, and a semi-quantitative psychomotor score (SQPS). We studied the correlation between DQ/IQ/SQPS and age, epilepsy characteristics, and whether patients presented SCN1A mutation.ResultsDQ/IQ significantly decreased with age (r = −.53, p < .001), from normal before 2y (mean 80, range 64–105) to low after 3y (mean 48, range 30–69), with hyperactivity and attention disorders hampering learning abilities especially up to 6y. However, raw (not age-adjusted) DQ sub-scores increased with age during the first decade, showing that there is no regression. We did not find any significant correlation between DQ/IQ at last evaluation and epilepsy data, i.e. first seizure (age, type, duration, fever), seizures during the course (type, fever sensitivity), status epilepticus (age of onset, number, fever), photosensitivity, and treatment, except for myoclonus and focal seizures which were associated with a lower QD/IQ after 3y. SCN1A mutated patients (n = 58) seemed to exhibit worse psychomotor course than non-mutated ones (n = 9) (severe SQPS in 26% vs 0%), although their epilepsy tended to be less severe (tonic seizures in 12% vs 44% [p = 0.04], first status epilepticus before 6 m in 26% vs 67% [p = .02], mean number of SE 2.5 vs 4.5 [p = .09]). DQ sub-scores were dissociated throughout the whole course: from onset hand-eye coordination was significantly lower than language, posture and sociability (p < .01). Dissociation seemed to be more frequent in mutated than in non-mutated patients (motor SQPS was normal for in 77% vs 44% [p = 0.017] whereas language SQPS was normal for 47% vs 100%).ConclusionsAlthough psychomotor/cognitive delay declines with age, there is no regression. In addition, encephalopathy is not a pure consequence of epilepsy but SCN1A mutation seems to play an additional, direct role.
Objective:To assess the prevalence of somatic MTOR mutations in focal cortical dysplasia (FCD) and of germline MTOR mutations in a broad range of epilepsies.Methods:We collected 20 blood-brain paired samples from patients with FCD and searched for somatic variants using deep-targeted gene panel sequencing. Germline mutations in MTOR were assessed in a French research cohort of 93 probands with focal epilepsies and in a diagnostic Danish cohort of 245 patients with a broad range of epilepsies. Data sharing among collaborators allowed us to ascertain additional germline variants in MTOR.Results:We detected recurrent somatic variants (p.Ser2215Phe, p.Ser2215Tyr, and p.Leu1460Pro) in the MTOR gene in 37% of participants with FCD II and showed histologic evidence for activation of the mTORC1 signaling cascade in brain tissue. We further identified 5 novel de novo germline missense MTOR variants in 6 individuals with a variable phenotype from focal, and less frequently generalized, epilepsies without brain malformations, to macrocephaly, with or without moderate intellectual disability. In addition, an inherited variant was found in a mother–daughter pair with nonlesional autosomal dominant nocturnal frontal lobe epilepsy.Conclusions:Our data illustrate the increasingly important role of somatic mutations of the MTOR gene in FCD and germline mutations in the pathogenesis of focal epilepsy syndromes with and without brain malformation or macrocephaly.
SEEG is feasible and useful in children with drug-resistant insular epilepsy which is often characterized by autonomic symptoms as the initial symptoms and should be suspected in cases with HMS, asymmetric tonic seizures and even asymmetric spasms. Early propagation is mostly frontal and central. Analysis of a larger population is required to refine these findings.
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