Publications by authors named "Stephanie Baulac"

73 Publications

Involvement of Gene in Familial Forms of Genetic Generalized Epilepsy.

Front Neurol 2021 21;12:738272. Epub 2021 Oct 21.

Sorbonne Université, Institut du Cerveau - Paris Brain Institute - ICM, Inserm, CNRS, Paris, France.

Genetic generalized epilepsies (GGE) including childhood absence epilepsy (CAE), juvenile absence epilepsy (JAE), juvenile myoclonic epilepsy (JME), and GGE with tonic-clonic seizures alone (GGE-TCS), are common types of epilepsy mostly determined by a polygenic mode of inheritance. Recent studies showed that susceptibility genes for GGE are numerous, and their variants rare, challenging their identification. In this study, we aimed to assess GGE genetic etiology in a Sudanese population. We performed whole-exome sequencing (WES) on DNA of 40 patients from 20 Sudanese families with GGE searching for candidate susceptibility variants, which were prioritized by CADD software and functional features of the corresponding gene. We assessed their segregation in 138 individuals and performed genotype-phenotype correlations. In a family including three sibs with GGE-TCS, we identified a rare missense variant in encoding an adhesion G protein-coupled receptor V1, which was already involved in the autosomal recessive Usher type C syndrome. In addition, five other rare missense variants were identified in four additional families and absent from 119 Sudanese controls. In one of these families, an variant was found at a homozygous state, in a female more severely affected than her heterozygous brother, suggesting a gene dosage effect. In the five families, GGE phenotype was statistically associated with variants (0R = 0.9 10). This study highly supports, for the first time, the involvement of missense variants in familial GGE and that is a susceptibility gene for CAE/JAE and GGE-TCS phenotypes.
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http://dx.doi.org/10.3389/fneur.2021.738272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8567843PMC
October 2021

Cardiac Investigations in Sudden Unexpected Death in DEPDC5-Related Epilepsy.

Ann Neurol 2021 Oct 24. Epub 2021 Oct 24.

Sorbonne University, Paris Brain Institute (ICM), Inserm, CNRS, AP-HP, Pitié-Salpêtrière Hospital, Paris, France.

Objective: Germline loss-of-function mutations in DEPDC5, and in its binding partners (NPRL2/3) of the mammalian target of rapamycin (mTOR) repressor GATOR1 complex, cause focal epilepsies and increase the risk of sudden unexpected death in epilepsy (SUDEP). Here, we asked whether DEPDC5 haploinsufficiency predisposes to primary cardiac defects that could contribute to SUDEP and therefore impact the clinical management of patients at high risk of SUDEP.

Methods: Clinical cardiac investigations were performed in 16 patients with pathogenic variants in DEPDC5, NPRL2, or NPRL3. Two novel Depdc5 mouse strains, a human HA-tagged Depdc5 strain and a Depdc5 heterozygous knockout with a neuron-specific deletion of the second allele (Depdc5 ), were generated to investigate the role of Depdc5 in SUDEP and cardiac activity during seizures.

Results: Holter, echocardiographic, and electrocardiographic (ECG) examinations provided no evidence for altered clinical cardiac function in the patient cohort, of whom 3 DEPDC5 patients succumbed to SUDEP and 6 had a family history of SUDEP. There was no cardiac injury at autopsy in a postmortem DEPDC5 SUDEP case. The HA-tagged Depdc5 mouse revealed expression of Depdc5 in the brain, heart, and lungs. Simultaneous electroencephalographic-ECG records on Depdc5 mice showed that spontaneous epileptic seizures resulting in a SUDEP-like event are not preceded by cardiac arrhythmia.

Interpretation: Mouse and human data show neither structural nor functional cardiac damage that might underlie a primary contribution to SUDEP in the spectrum of DEPDC5-related epilepsies. ANN NEUROL 2021.
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http://dx.doi.org/10.1002/ana.26256DOI Listing
October 2021

Neocortical development and epilepsy: insights from focal cortical dysplasia and brain tumours.

Lancet Neurol 2021 Nov;20(11):943-955

Sorbonne Université, Institut du Cerveau - Paris Brain Institute (ICM), Inserm, CNRS, Paris, France.

During the past decade, there have been considerable advances in understanding of the genetic and morphogenic processes underlying cortical malformations and developmental brain tumours. Focal malformations are caused by somatic (postzygotic) variants in genes related to cell growth (ie, in the mTOR pathway in focal cortical dysplasia type 2), which are acquired in neuronal progenitors during neurodevelopment. In comparison, developmental brain tumours result from somatic variants in genes related to cell proliferation (eg, in the MAP-kinase pathway in ganglioglioma), which affect proliferating glioneuronal precursors. The timing of the genetic event and the specific gene involved during neurodevelopment will drive the nature and size of the lesion, whether it is a developmental malformation or a brain tumour. There is also emerging evidence that epigenetic processes underlie a molecular memory in epileptogenesis. This knowledge will together facilitate understanding of why and how patients with these lesions have epilepsy, and could form a basis for a move towards precision medicine for this challenging cohort of patients.
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http://dx.doi.org/10.1016/S1474-4422(21)00265-9DOI Listing
November 2021

Reply to "Improving Specificity of CSF Liquid Biopsy for Genetic Testing".

Ann Neurol 2021 Oct 20;90(4):694-695. Epub 2021 Aug 20.

Graduate School of Medical Science and Engineering, KAIST, Daejeon, South Korea.

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http://dx.doi.org/10.1002/ana.26188DOI Listing
October 2021

Molecular diagnostics in drug-resistant focal epilepsy define new disease entities.

Brain Pathol 2021 07;31(4):e12963

Graduate School of Medical Science and Engineering, KAIST, Daejeon, Korea.

Structural brain lesions, including the broad range of malformations of cortical development (MCD) and glioneuronal tumors, are among the most common causes of drug-resistant focal epilepsy. Epilepsy surgery can provide a curative treatment option in respective patients. The currently available pre-surgical multi-modal diagnostic armamentarium includes high- and ultra-high resolution magnetic resonance imaging (MRI) and intracerebral EEG to identify a focal structural brain lesion as epilepsy underlying etiology. However, specificity and accuracy in diagnosing the type of lesion have proven to be limited. Moreover, the diagnostic process does not stop with the decision for surgery. The neuropathological diagnosis remains the gold standard for disease classification and patient stratification, but is particularly complex with high inter-observer variability. Here, the identification of lesion-specific mosaic variants together with epigenetic profiling of lesional brain tissue became new tools to more reliably identify disease entities. In this review, we will discuss how the paradigm shifts from histopathology toward an integrated diagnostic approach in cancer and the more recent development of the DNA methylation-based brain tumor classifier have started to influence epilepsy diagnostics. Some examples will be highlighted showing how the diagnosis and our mechanistic understanding of difficult to classify structural brain lesions associated with focal epilepsy has improved with molecular genetic data being considered in decision making.
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http://dx.doi.org/10.1111/bpa.12963DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8412082PMC
July 2021

KCNT1-related epilepsies and epileptic encephalopathies: phenotypic and mutational spectrum.

Brain 2021 Jun 11. Epub 2021 Jun 11.

Pediatric Neurology Department, Lyon University Hospital, 69500 Bron, France.

Variants in KCNT1, encoding a sodium-gated potassium channel (subfamily T member 1), have been associated with a spectrum of epilepsies and neurodevelopmental disorders. These range from familial autosomal dominant or sporadic sleep-related hypermotor epilepsy ((AD)SHE) to epilepsy of infancy with migrating focal seizures (EIMFS) and include developmental and epileptic encephalopathies (DEE). This study aims to provide a comprehensive overview of the phenotypic and genotypic spectrum of KCNT1 mutation-related epileptic disorders in 248 individuals, including 66 unpreviously published and 182 published cases, the largest cohort reported so far. Four phenotypic groups emerged from our analysis: i) EIMFS (152 individuals, 33 previously unpublished); ii) DEE other than EIMFS (non-EIMFS DEE) (37 individuals, 17 unpublished); iii) (AD)SHE (53 patients, 14 unpublished); iv) other phenotypes (6 individuals, 2 unpublished). In our cohort of 66 new cases, the most common phenotypic features were: a) in EIMFS, heterogeneity of seizure types, including epileptic spasms, epilepsy improvement over time, no epilepsy-related deaths; b) in non-EIMFS DEE, possible onset with West syndrome, occurrence of atypical absences, possible evolution to DEE with SHE features; one case of sudden unexplained death in epilepsy (SUDEP); c) in (AD)SHE, we observed a high prevalence of drug-resistance, although seizure frequency improved with age in some individuals, appearance of cognitive regression after seizure onset in all patients, no reported severe psychiatric disorders, although behavioural/psychiatric comorbidities were reported in about 50% of the patients, SUDEP in one individual; d) other phenotypes in individuals with mutation of KCNT1 included temporal lobe epilepsy, and epilepsy with tonic-clonic seizures and cognitive regression. Genotypic analysis of the whole cohort of 248 individuals showed only missense mutations and one inframe deletion in KCNT1. Although the KCNT1 mutations in affected individuals were seen to be distributed among the different domains of the KCNT1 protein, genotype-phenotype considerations showed many of the (AD)SHE-associated mutations to be clustered around the RCK2 domain in the C-terminus, distal to the NADP domain. Mutations associated with EIMFS/non-EIMFS DEE did not show a particular pattern of distribution in the KCNT1 protein. Recurrent KCNT1 mutations were seen to be associated with both severe and less severe phenotypes. Our study further defines and broadens the phenotypic and genotypic spectrums of KCNT1-related epileptic conditions and emphasizes the increasingly important role of this gene in the pathogenesis of early onset DEEs as well as in focal epilepsies, namely (AD)SHE.
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http://dx.doi.org/10.1093/brain/awab219DOI Listing
June 2021

Toward a better definition of focal cortical dysplasia: An iterative histopathological and genetic agreement trial.

Epilepsia 2021 06 5;62(6):1416-1428. Epub 2021 May 5.

Department of Neuropathology, Institute of Neurology, University College London, London, UK.

Objective: Focal cortical dysplasia (FCD) is a major cause of difficult-to-treat epilepsy in children and young adults, and the diagnosis is currently based on microscopic review of surgical brain tissue using the International League Against Epilepsy classification scheme of 2011. We developed an iterative histopathological agreement trial with genetic testing to identify areas of diagnostic challenges in this widely used classification scheme.

Methods: Four web-based digital pathology trials were completed by 20 neuropathologists from 15 countries using a consecutive series of 196 surgical tissue blocks obtained from 22 epilepsy patients at a single center. Five independent genetic laboratories performed screening or validation sequencing of FCD-relevant genes in paired brain and blood samples from the same 22 epilepsy patients.

Results: Histopathology agreement based solely on hematoxylin and eosin stainings was low in Round 1, and gradually increased by adding a panel of immunostainings in Round 2 and the Delphi consensus method in Round 3. Interobserver agreement was good in Round 4 (kappa = .65), when the results of genetic tests were disclosed, namely, MTOR, AKT3, and SLC35A2 brain somatic mutations in five cases and germline mutations in DEPDC5 and NPRL3 in two cases.

Significance: The diagnoses of FCD 1 and 3 subtypes remained most challenging and were often difficult to differentiate from a normal homotypic or heterotypic cortical architecture. Immunohistochemistry was helpful, however, to confirm the diagnosis of FCD or no lesion. We observed a genotype-phenotype association for brain somatic mutations in SLC35A2 in two cases with mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy. Our results suggest that the current FCD classification should recognize a panel of immunohistochemical stainings for a better histopathological workup and definition of FCD subtypes. We also propose adding the level of genetic findings to obtain a comprehensive, reliable, and integrative genotype-phenotype diagnosis in the near future.
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http://dx.doi.org/10.1111/epi.16899DOI Listing
June 2021

Detection of Brain Somatic Mutations in Cerebrospinal Fluid from Refractory Epilepsy Patients.

Ann Neurol 2021 06 20;89(6):1248-1252. Epub 2021 Apr 20.

Sorbonne University, Paris Brain Institute (ICM), National Institute of Health and Medical Research (INSERM), National Center for Scientific Research (CNRS), Paris, France.

Brain mosaic mutations are a major cause of refractory focal epilepsies with cortical malformations such as focal cortical dysplasia, hemimegalencephaly, malformation of cortical development with oligodendroglial hyperplasia in epilepsy, and ganglioglioma. Here, we collected cerebrospinal fluid (CSF) during epilepsy surgery to search for somatic variants in cell-free DNA (cfDNA) using targeted droplet digital polymerase chain reaction. In 3 of 12 epileptic patients with known somatic mutations previously identified in brain tissue, we here provide evidence that brain mosaicism can be detected in the CSF-derived cfDNA. These findings suggest future opportunities for detecting the mutant allele driving epilepsy in CSF. ANN NEUROL 2021;89:1248-1252.
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http://dx.doi.org/10.1002/ana.26080DOI Listing
June 2021

Gradient of brain mosaic RHEB variants causes a continuum of cortical dysplasia.

Ann Clin Transl Neurol 2021 02 12;8(2):485-490. Epub 2021 Jan 12.

Department of Paediatrics, The University of Melbourne, Parkville, 3052, Australia.

Focal cortical dysplasia (FCD) and hemimegalencephaly (HME) are related malformations with shared etiologies. We report three patients with a spectrum of cortical malformations associated with pathogenic brain-specific somatic Ras homolog enriched in brain (RHEB) variants. The somatic variant load directly correlated with the size of the malformation, with upregulated mTOR activity confirmed in dysplastic tissues. Laser capture microdissection showed enrichment of RHEB variants in dysmorphic neurons and balloon cells. Our findings support the role of RHEB in a spectrum of cortical malformations confirming that FCD and HME represent a disease continuum, with the extent of dysplastic brain directly correlated with the somatic variant load.
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http://dx.doi.org/10.1002/acn3.51286DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7886042PMC
February 2021

Frequent SLC35A2 brain mosaicism in mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE).

Acta Neuropathol Commun 2021 01 6;9(1). Epub 2021 Jan 6.

Institut du Cerveau - Paris Brain Institute - ICM, Inserm, CNRS, Sorbonne Université, Hôpital Pitié-Salpêtrière - 47, bd de l'hôpital, 75013, Paris, France.

Focal malformations of cortical development (MCD) are linked to somatic brain mutations occurring during neurodevelopment. Mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE) is a newly recognized clinico-pathological entity associated with pediatric drug-resistant focal epilepsy, and amenable to neurosurgical treatment. MOGHE is histopathologically characterized by clusters of increased oligodendroglial cell densities, patchy zones of hypomyelination, and heterotopic neurons in the white matter. The molecular etiology of MOGHE remained unknown so far. We hypothesized a contribution of mosaic brain variants and performed deep targeted gene sequencing on 20 surgical MOGHE brain samples from a single-center cohort of pediatric patients. We identified somatic pathogenic SLC35A2 variants in 9/20 (45%) patients with mosaic rates ranging from 7 to 52%. SLC35A2 encodes a UDP-galactose transporter, previously implicated in other malformations of cortical development (MCD) and a rare type of congenital disorder of glycosylation. To further clarify the histological features of SLC35A2-brain tissues, we then collected 17 samples with pathogenic SLC35A2 variants from a multicenter cohort of MCD cases. Histopathological reassessment including anti-Olig2 staining confirmed a MOGHE diagnosis in all cases. Analysis by droplet digital PCR of pools of microdissected cells from one MOGHE tissue revealed a variant enrichment in clustered oligodendroglial cells and heterotopic neurons. Through an international consortium, we assembled an unprecedented series of 26 SLC35A2-MOGHE cases providing evidence that mosaic SLC35A2 variants, likely occurred in a neuroglial progenitor cell during brain development, are a genetic marker for MOGHE.
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http://dx.doi.org/10.1186/s40478-020-01085-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7788938PMC
January 2021

Distinctive binding properties of human monoclonal LGI1 autoantibodies determine pathogenic mechanisms.

Brain 2020 06;143(6):1731-1745

Oxford Autoimmune Neurology Group, Nuffield Department of Clinical Neurosciences, University of Oxford, Oxford, UK.

Autoantibodies against leucine-rich glioma inactivated 1 (LGI1) are found in patients with limbic encephalitis and focal seizures. Here, we generate patient-derived monoclonal antibodies (mAbs) against LGI1. We explore their sequences and binding characteristics, plus their pathogenic potential using transfected HEK293T cells, rodent neuronal preparations, and behavioural and electrophysiological assessments in vivo after mAb injections into the rodent hippocampus. In live cell-based assays, LGI1 epitope recognition was examined with patient sera (n = 31), CSFs (n = 11), longitudinal serum samples (n = 15), and using mAbs (n = 14) generated from peripheral B cells of two patients. All sera and 9/11 CSFs bound both the leucine-rich repeat (LRR) and the epitempin repeat (EPTP) domains of LGI1, with stable ratios of LRR:EPTP antibody levels over time. By contrast, the mAbs derived from both patients recognized either the LRR or EPTP domain. mAbs against both domain specificities showed varied binding strengths, and marked genetic heterogeneity, with high mutation frequencies. LRR-specific mAbs recognized LGI1 docked to its interaction partners, ADAM22 and ADAM23, bound to rodent brain sections, and induced internalization of the LGI1-ADAM22/23 complex in both HEK293T cells and live hippocampal neurons. By contrast, few EPTP-specific mAbs bound to rodent brain sections or ADAM22/23-docked LGI1, but all inhibited the docking of LGI1 to ADAM22/23. After intrahippocampal injection, and by contrast to the LRR-directed mAbs, the EPTP-directed mAbs showed far less avid binding to brain tissue and were consistently detected in the serum. Post-injection, both domain-specific mAbs abrogated long-term potentiation induction, and LRR-directed antibodies with higher binding strengths induced memory impairment. Taken together, two largely dichotomous populations of LGI1 mAbs with distinct domain binding characteristics exist in the affinity matured peripheral autoantigen-specific memory pools of individuals, both of which have pathogenic potential. In human autoantibody-mediated diseases, the detailed characterization of patient mAbs provides a valuable method to dissect the molecular mechanisms within polyclonal populations.
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http://dx.doi.org/10.1093/brain/awaa104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7296845PMC
June 2020

Acute knockdown of Depdc5 leads to synaptic defects in mTOR-related epileptogenesis.

Neurobiol Dis 2020 06 27;139:104822. Epub 2020 Feb 27.

Center for Synaptic Neuroscience and Technology, Istituto Italiano di Tecnologia, Largo Rosanna Benzi 10, 16132 Genova, Italy; IRCSS, Ospedale Policlinico San Martino, Largo Rosanna Benzi 10, 16132 Genova, Italy. Electronic address:

DEP-domain containing 5 (DEPDC5) is part of the GATOR1 complex that functions as key inhibitor of the mechanistic target of rapamycin complex 1 (mTORC1). Loss-of-function mutations in DEPDC5 leading to mTOR hyperactivation have been identified as the most common cause of either lesional or non-lesional focal epilepsy. However, the precise mechanisms by which DEPDC5 loss-of-function triggers neuronal and network hyperexcitability are still unclear. In this study, we investigated the cellular mechanisms of hyperexcitability by comparing the constitutive heterozygous Depdc5 knockout mouse versus different levels of acute Depdc5 deletion (≈40% and ≈80% neuronal knockdown of Depdc5 protein) by RNA interference in primary cortical cultures. While heterozygous Depdc5 neurons have only a subtle phenotype, acutely knocked-down neurons exhibit a strong dose-dependent phenotype characterized by mTOR hyperactivation, increased soma size, dendritic arborization, excitatory synaptic transmission and intrinsic excitability. The robust synaptic phenotype resulting from the acute knockdown Depdc5 deficiency highlights the importance of the temporal dynamics of Depdc5 knockdown in triggering the phenotypic changes, reminiscent of the somatic second-hit mechanism in patients with focal cortical dysplasia. These findings uncover a novel synaptic phenotype that is causally linked to Depdc5 knockdown, highlighting the developmental role of Depdc5. Interestingly, the synaptic defect appears to affect only excitatory synapses, while inhibitory synapses develop normally. The increased frequency and amplitude of mEPSCs, paralleled by increased density of excitatory synapses and expression of glutamate receptors, may generate an excitation/inhibition imbalance that triggers epileptogenesis.
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http://dx.doi.org/10.1016/j.nbd.2020.104822DOI Listing
June 2020

Dissecting the genetic basis of focal cortical dysplasia: a large cohort study.

Acta Neuropathol 2019 12 23;138(6):885-900. Epub 2019 Aug 23.

Sorbonne Université, UPMC Univ Paris 06, UMR S 1127, Paris, France.

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.
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http://dx.doi.org/10.1007/s00401-019-02061-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6851393PMC
December 2019

Juvenile myoclonic epilepsy phenotype in a family with Unverricht-Lundborg disease.

Epileptic Disord 2019 Aug;21(4):359-365

Razi Hospital, Department of Neurology, LR 18SP03, Tunis, Université de Tunis El Manar, Faculté de Médecine de Tunis, Tunis, Tunisia.

Unverricht-Lundborg disease (ULD), an autosomal recessive progressive myoclonus epilepsy, is due to an expansion, or less commonly a mutation, of the cystatin B (CSTB) gene. We report a clinical and molecular study of a Tunisian ULD family with five affected members presenting with a juvenile myoclonic epilepsy (JME)-like phenotype. The expansion of dodecamers was detected by a deamination/PCR assay. The expression profiles of CSTB and other candidate modifying genes, cathepsin B and cystatin C, were established by quantitative RT-PCR, and their respective transcription levels were compared with those from patients with a classic picture of ULD. Three patients had a fixed phenotype mimicking JME after 29 years of evolution. Only a discrete dysarthria was noticed in the two other patients. No correlation was observed between transcription level and severity of disease. Genetic screening should be performed in patients with a JME-like phenotype, when careful examination reveals discrete atypical signs of JME. This particular phenotype may be due to modifying genes and/or gene-environment interactions which require further clarification.
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http://dx.doi.org/10.1684/epd.2019.1078DOI Listing
August 2019

Treatment Responsiveness in KCNT1-Related Epilepsy.

Neurotherapeutics 2019 07;16(3):848-857

Division of Child Neurology, Department of Neurology, University of Rochester School of Medicine, Rochester, NY, USA.

Pathogenic variants in KCNT1 represent an important cause of treatment-resistant epilepsy, for which an effective therapy has been elusive. Reports about the effectiveness of quinidine, a candidate precision therapy, have been mixed. We sought to evaluate the treatment responsiveness of patients with KCNT1-related epilepsy. We performed an observational study of 43 patients using a collaborative KCNT1 patient registry. We assessed treatment efficacy based upon clinical seizure reduction, side effects of quinidine therapy, and variant-specific responsiveness to treatment. Quinidine treatment resulted in a > 50% seizure reduction in 20% of patients, with rare patients achieving transient seizure freedom. Multiple other therapies demonstrated some success in reducing seizure frequency, including the ketogenic diet and vigabatrin, the latter particularly in patients with epileptic spasms. Patients with the best quinidine response had variants that clustered distal to the NADP domain within the RCK2 domain of the protein. Half of patients did not receive a quinidine trial. In those who did, nearly half did not achieve therapeutic blood levels. More favorable response to quinidine in patients with KCNT1 variants distal to the NADP domain within the RCK2 domain may suggest a variant-specific response.
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http://dx.doi.org/10.1007/s13311-019-00739-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6694367PMC
July 2019

[A second-hit somatic mutation drives neurodevelopmental epilepsy].

Med Sci (Paris) 2019 Apr 30;35(4):289-291. Epub 2019 Apr 30.

Institut du cerveau et de la moelle épinière (ICM), Inserm U1127, CNRS UMR 7225, Sorbonne Université, Hôpital Pitié-Salpêtrière, 47, boulevard de l'Hôpital, 75013 Paris, France.

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http://dx.doi.org/10.1051/medsci/2019058DOI Listing
April 2019

Mild malformations of cortical development in sleep-related hypermotor epilepsy due to mutations.

Ann Clin Transl Neurol 2019 02 25;6(2):386-391. Epub 2018 Dec 25.

Danish Epilepsy Centre, Filadelfia Dianalund Denmark.

Mutations in the sodium-activated potassium channel gene have been associated with nonlesional sleep-related hypermotor epilepsy (SHE). We report the co-occurrence of mild malformation of cortical development (mMCD) and mutations in four patients with SHE. Focal cortical dysplasia type I was neuropathologically diagnosed after epilepsy surgery in three unrelated MRI-negative patients, periventricular nodular heterotopia was detected in one patient by MRI. Our findings suggest that epileptogenicity may result not only from dysregulated excitability by controlling Na+K+ transport, but also from mMCD. Therefore, pathogenic variants in may encompass both lesional and nonlesional epilepsies.
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http://dx.doi.org/10.1002/acn3.708DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6389734PMC
February 2019

The Nogo Receptor Ligand LGI1 Regulates Synapse Number and Synaptic Activity in Hippocampal and Cortical Neurons.

eNeuro 2018 Jul-Aug;5(4). Epub 2018 Sep 7.

Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec H3A 2B4, Canada.

Leucine-rich glioma-inactivated protein 1 (LGI1) is a secreted neuronal protein and a Nogo receptor 1 (NgR1) ligand. Mutations in LGI1 in humans causes autosomal dominant lateral temporal lobe epilepsy and homozygous deletion of LGI1 in mice results in severe epileptic seizures that cause early postnatal death. NgR1 plays an important role in the development of CNS synapses and circuitry by limiting plasticity in the adult cortex via the activation of RhoA. These relationships and functions prompted us to examine the effect of LGI1 on synapse formation and . We report that application of LGI1 increases synaptic density in neuronal culture and that LGI1 null hippocampus has fewer dendritic mushroom spines than in wild-type (WT) littermates. Further, our electrophysiological investigations demonstrate that LGI1 null hippocampal neurons possess fewer and weaker synapses. RhoA activity is significantly increased in cortical cultures derived from LGI1 null mice and using a reconstituted system; we show directly that LGI1 antagonizes NgR1-tumor necrosis factor receptor orphan Y (TROY) signaling. Our data suggests that LGI1 enhances synapse formation in cortical and hippocampal neurons by reducing NgR1 signaling.
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http://dx.doi.org/10.1523/ENEURO.0185-18.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6140115PMC
March 2019

The landscape of epilepsy-related GATOR1 variants.

Genet Med 2019 02 10;21(2):398-408. Epub 2018 Aug 10.

Stichting Epilepsie Instellingen Nederland, Zwolle/Heemstede, The Netherlands.

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.
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http://dx.doi.org/10.1038/s41436-018-0060-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6292495PMC
February 2019

Rare coding variants in genes encoding GABA receptors in genetic generalised epilepsies: an exome-based case-control study.

Lancet Neurol 2018 08 17;17(8):699-708. Epub 2018 Jul 17.

Background: Genetic generalised epilepsy is the most common type of inherited epilepsy. Despite a high concordance rate of 80% in monozygotic twins, the genetic background is still poorly understood. We aimed to investigate the burden of rare genetic variants in genetic generalised epilepsy.

Methods: For this exome-based case-control study, we used three different genetic generalised epilepsy case cohorts and three independent control cohorts, all of European descent. Cases included in the study were clinically evaluated for genetic generalised epilepsy. Whole-exome sequencing was done for the discovery case cohort, a validation case cohort, and two independent control cohorts. The replication case cohort underwent targeted next-generation sequencing of the 19 known genes encoding subunits of GABA receptors and was compared to the respective GABA receptor variants of a third independent control cohort. Functional investigations were done with automated two-microelectrode voltage clamping in Xenopus laevis oocytes.

Findings: Statistical comparison of 152 familial index cases with genetic generalised epilepsy in the discovery cohort to 549 ethnically matched controls suggested an enrichment of rare missense (Nonsyn) variants in the ensemble of 19 genes encoding GABA receptors in cases (odds ratio [OR] 2·40 [95% CI 1·41-4·10]; p=0·0014, adjusted p=0·019). Enrichment for these genes was validated in a whole-exome sequencing cohort of 357 sporadic and familial genetic generalised epilepsy cases and 1485 independent controls (OR 1·46 [95% CI 1·05-2·03]; p=0·0081, adjusted p=0·016). Comparison of genes encoding GABA receptors in the independent replication cohort of 583 familial and sporadic genetic generalised epilepsy index cases, based on candidate-gene panel sequencing, with a third independent control cohort of 635 controls confirmed the overall enrichment of rare missense variants for 15 GABA receptor genes in cases compared with controls (OR 1·46 [95% CI 1·02-2·08]; p=0·013, adjusted p=0·027). Functional studies for two selected genes (GABRB2 and GABRA5) showed significant loss-of-function effects with reduced current amplitudes in four of seven tested variants compared with wild-type receptors.

Interpretation: Functionally relevant variants in genes encoding GABA receptor subunits constitute a significant risk factor for genetic generalised epilepsy. Examination of the role of specific gene groups and pathways can disentangle the complex genetic architecture of genetic generalised epilepsy.

Funding: EuroEPINOMICS (European Science Foundation through national funding organisations), Epicure and EpiPGX (Sixth Framework Programme and Seventh Framework Programme of the European Commission), Research Unit FOR2715 (German Research Foundation and Luxembourg National Research Fund).
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http://dx.doi.org/10.1016/S1474-4422(18)30215-1DOI Listing
August 2018

Clinical and genetic study of Tunisian families with genetic generalized epilepsy: contribution of CACNA1H and MAST4 genes.

Neurogenetics 2018 08 12;19(3):165-178. Epub 2018 Jun 12.

UPMC Univ Paris 06, Inserm, CNRS, APHP, Institut du Cerveau et la Moelle (ICM), Hôpital Pitié-Salpêtrière, Sorbonne Universités, Paris, France.

Genetic generalized epilepsies (GGE) (childhood absence epilepsy (CAE), juvenile myoclonic epilepsy (JME) and epilepsy with generalized tonic-clonic seizures (GTCS)) are mainly determined by genetic factors. Since few mutations were identified in rare families with autosomal dominant GGE, a polygenic inheritance was suspected in most patients. Recent studies on large American or European cohorts of sporadic cases showed that susceptibility genes were numerous although their variants were rare, making their identification difficult. Here, we reported clinical and genetic characteristics of 30 Tunisian GGE families, including 71 GGE patients. The phenotype was close to that in sporadic cases. Nineteen pedigrees had a homogeneous type of GGE (JME-CAE-CGTS), and 11 combined these epileptic syndromes. Rare non-synonymous variants were selected in probands using a targeted panel of 30 candidate genes and their segregation was determined in families. Molecular studies incriminated different genes, mainly CACNA1H and MAST4. The segregation of at least two variants in different genes in some pedigrees was compatible with the hypothesis of an oligogenic inheritance, which was in accordance with the relatively low frequency of consanguineous probands. Since at least 2 susceptibility genes were likely shared by different populations, genetic factors involved in the majority of Tunisian GGE families remain to be discovered. Their identification should be easier in families with a homogeneous type of GGE, in which an intra-familial genetic homogeneity could be suspected.
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http://dx.doi.org/10.1007/s10048-018-0550-zDOI Listing
August 2018

Depdc5 knockdown causes mTOR-dependent motor hyperactivity in zebrafish.

Ann Clin Transl Neurol 2018 May 6;5(5):510-523. Epub 2018 Apr 6.

Sorbonne Universités Paris VI UMR CNRS 1127 UPMC INSERM U 1127 CNRS UMR 7225 Institut du Cerveau et de la Moelle épinière - ICM Paris France.

Objective: was identified as a major genetic cause of focal epilepsy with deleterious mutations found in a wide range of inherited forms of focal epilepsy, associated with malformation of cortical development in certain cases. Identification of frameshift, truncation, and deletion mutations implicates haploinsufficiency of in the etiology of focal epilepsy. DEPDC5 is a component of the GATOR1 complex, acting as a negative regulator of mTOR signaling.

Methods: Zebrafish represents a vertebrate model suitable for genetic analysis and drug screening in epilepsy-related disorders. In this study, we defined the expression of during development and established an epilepsy model with reduced Depdc5 expression.

Results: Here we report a zebrafish model of Depdc5 loss-of-function that displays a measurable behavioral phenotype, including hyperkinesia, circular swimming, and increased neuronal activity. These phenotypic features persisted throughout embryonic development and were significantly reduced upon treatment with the mTORC1 inhibitor, rapamycin, as well as overexpression of human WT transcript. No phenotypic rescue was obtained upon expression of epilepsy-associated mutations (p.Arg487* and p.Arg485Gln), indicating that these mutations cause a loss of function of the protein.

Interpretation: This study demonstrates that Depdc5 knockdown leads to early-onset phenotypic features related to motor and neuronal hyperactivity. Restoration of phenotypic features by WT but not epilepsy-associated Depdc5 mutants, as well as by mTORC1 inhibition confirm the role of Depdc5 in the mTORC1-dependent molecular cascades, defining this pathway as a potential therapeutic target for -inherited forms of focal epilepsy.
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http://dx.doi.org/10.1002/acn3.542DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5945968PMC
May 2018

Second-hit mosaic mutation in mTORC1 repressor DEPDC5 causes focal cortical dysplasia-associated epilepsy.

J Clin Invest 2018 06 30;128(6):2452-2458. Epub 2018 Apr 30.

Institut du Cerveau et de la Moelle épinière (ICM), INSERM U 1127, CNRS UMR 7225, Sorbonne Université, Paris, France.

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.
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http://dx.doi.org/10.1172/JCI99384DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5983335PMC
June 2018

LGI1 tunes intrinsic excitability by regulating the density of axonal Kv1 channels.

Proc Natl Acad Sci U S A 2017 07 3;114(29):7719-7724. Epub 2017 Jul 3.

Unité de Neurobiologie des Canaux Ioniques et de la Synapse, INSERM UMR 1072, Aix Marseille Université, 13015 Marseille, France;

Autosomal dominant epilepsy with auditory features results from mutations in leucine-rich glioma-inactivated 1 (LGI1), a soluble glycoprotein secreted by neurons. Animal models of LGI1 depletion display spontaneous seizures, however, the function of LGI1 and the mechanisms by which deficiency leads to epilepsy are unknown. We investigated the effects of pure recombinant LGI1 and genetic depletion on intrinsic excitability, in the absence of synaptic input, in hippocampal CA3 neurons, a classical focus for epileptogenesis. Our data indicate that LGI1 is expressed at the axonal initial segment and regulates action potential firing by setting the density of the axonal Kv1.1 channels that underlie dendrotoxin-sensitive D-type potassium current. LGI1 deficiency incurs a >50% down-regulation of the expression of Kv1.1 and Kv1.2 via a posttranscriptional mechanism, resulting in a reduction in the capacity of axonal D-type current to limit glutamate release, thus contributing to epileptogenesis.
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http://dx.doi.org/10.1073/pnas.1618656114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5530646PMC
July 2017

mTOR pathway in familial focal epilepsies.

Oncotarget 2017 Jan;8(4):5674-5675

Sorbonne Universités, UPMC Univ Paris 06, Inserm, CNRS, APHP, Institut du Cerveau et la Moelle (ICM), Hôpital Pitié-Salpêtrière, Paris, France.

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http://dx.doi.org/10.18632/oncotarget.14234DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5351575PMC
January 2017

Germline and somatic mutations in the gene in focal cortical dysplasia and epilepsy.

Neurol Genet 2016 Dec 31;2(6):e118. Epub 2016 Oct 31.

The Danish Epilepsy Centre Filadelfia (R.S.M., G.R.), Dianalund, Denmark; Institute for Regional Health Services (R.S.M.), University of Southern Denmark, Odense; Sorbonne Universités (S.W., E.M., V.L., E.L., S.B.), UPMC Univ Paris 06 UMR S 1127, Inserm U1127, CNRS UMR 7225, AP-HP, Institut du cerveau et la moelle (ICM)-Hôpital Pitié-Salpêtrière, Paris, France; Epilepsy Unit (S.W., V.L.), AP-HP Groupe hospitalier Pitié-Salpêtrière, Paris, France; Neurogenetics Group (S.W.), VIB-Department of Molecular Genetics; Laboratory of Neurogenetics (S.W.), Institute Born-Bunge, University of Antwerp, Belgium; Department of Neurology (S.W.), University Hospital Antwerp, Belgium; Department of Pediatric Neurosurgery (M.C., S.F.-S., G.D.), Fondation Rothschild, Paris, France; Université Paris Sorbonne Cité (V.T.), INSERM UMR-S1147 MEPPOT, CNRS SNC5014, Centre Universitaire des Saints-Pères, Paris, France; Department of Neurology (E.M.B.), University of Alabama at Birmingham; HudsonAlpha Institute for Biotechnology (S.M.H., J.W.P., K.M.B., G.M.C.), Huntsville, AL; Pediatric Neurology, Neurogenetics and Neurobiology Unit and Laboratories (D.M., V.C., R.G.), Neuroscience Department, A Meyer Children's Hospital, University of Florence, Florence, Italy; Genosplice (P.d.l.G.), Institut du Cerveau et de la Moelle épinière, ICM, Paris, France; Amplexa Genetics (L.H.G.L.), Odense, Denmark; Department of Genetics and Cytogenetics (E.L., S.B.), AP-HP Groupe hospitalier Pitié-Salpêtrière, Paris, France; and University of Copenhagen (G.R.), Denmark.

Objective: To assess the prevalence of somatic mutations in focal cortical dysplasia (FCD) and of germline 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 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 .

Results: We detected recurrent somatic variants (p.Ser2215Phe, p.Ser2215Tyr, and p.Leu1460Pro) in the 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 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 gene in FCD and germline mutations in the pathogenesis of focal epilepsy syndromes with and without brain malformation or macrocephaly.
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http://dx.doi.org/10.1212/NXG.0000000000000118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5089441PMC
December 2016

Involvement of GATOR complex genes in familial focal epilepsies and focal cortical dysplasia.

Epilepsia 2016 06 13;57(6):994-1003. Epub 2016 May 13.

INSERM, U1127, ICM, Paris, France.

Objective: The discovery of mutations in DEPDC5 in familial focal epilepsies has introduced a novel pathomechanism to a field so far dominated by ion channelopathies. DEPDC5 is part of a complex named GAP activity toward RAGs (GATOR) complex 1 (GATOR1), together with the proteins NPRL2 and NPRL3, and acts to inhibit the mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) pathway. GATOR1 is in turn inhibited by the GATOR2 complex. The mTORC1 pathway is a major signaling cascade regulating cell growth, proliferation, and migration. We aimed to study the contribution of GATOR complex genes to the etiology of focal epilepsies and to describe the associated phenotypical spectrum.

Methods: We performed targeted sequencing of the genes encoding the components of the GATOR1 (DEPDC5, NPRL2, and NPRL3) and GATOR2 (MIOS, SEC13, SEH1L, WDR24, and WDR59) complex in 93 European probands with focal epilepsy with or without focal cortical dysplasia. Phospho-S6 immunoreactivity was used as evidence of mTORC1 pathway activation in resected brain tissue of patients carrying pathogenic variants.

Results: We identified four pathogenic variants in DEPDC5, two in NPRL2, and one in NPRL3. We showed hyperactivation of the mTORC1 pathway in brain tissue from patients with NPRL2 and NPRL3 mutations. Collectively, inactivating mutations in GATOR1 complex genes explained 11% of cases of focal epilepsy, whereas no pathogenic mutations were found in GATOR2 complex genes. GATOR1-related focal epilepsies differ clinically from focal epilepsies due to mutations in ion channel genes by their association with focal cortical dysplasia and seizures emerging from variable foci, and might confer an increased risk of sudden unexplained death in epilepsy (SUDEP).

Significance: GATOR1 complex gene mutations leading to mTORC1 pathway upregulation is an important cause of focal epilepsy with cortical malformations and represents a potential target for novel therapeutic approaches.
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http://dx.doi.org/10.1111/epi.13391DOI Listing
June 2016
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