The mechanistic target of rapamycin (mTOR) signaling pathway is an essential regulator of numerous cellular activities such as metabolism, growth, proliferation, and survival. The mTOR cascade ...recently emerged as a critical player in the pathogenesis of focal epilepsies and cortical malformations. The ‘mTORopathies’ comprise a spectrum of cortical malformations that range from whole brain (megalencephaly) and hemispheric (hemimegalencephaly) abnormalities to focal abnormalities, such as focal cortical dysplasia type II (FCDII), which manifest with drug-resistant epilepsies. The spectrum of cortical dysplasia results from somatic brain mutations in the mTOR pathway activators AKT3, MTOR, PIK3CA, and RHEB and from germline and somatic mutations in mTOR pathway repressors, DEPDC5, NPRL2, NPRL3, TSC1 and TSC2. The mTORopathies are characterized by excessive mTOR pathway activation, leading to a broad range of structural and functional impairments. Here, we provide a comprehensive literature review of somatic mTOR-activating mutations linked to epilepsy and cortical malformations in 292 patients and discuss the perspectives of targeted therapeutics for personalized medicine.
Rare multiplex families with autosomal dominant focal epilepsies have been described with specific age-related and electroclinical syndromes: autosomal dominant nocturnal frontal lobe epilepsy ...(ADNFLE), familial temporal lobe epilepsy (FTLE), and familial focal epilepsy with variable foci (FFEVF). Molecular genetic advances in inherited focal epilepsies have pinpointed their genetic heterogeneity and the fact that they are mediated by different biological pathways: ion channel subunit genes have been linked to ADNFLE (CHRNA4, CHRNA2, CHRNB2, and KCNT1, encoding, respectively, the α4, α2, and β2 subunits of the neuronal nicotinic acetylcholine receptor, and a potassium channel subunit); neuronal secreted protein (LGI1-encoding epitempin) has been linked to autosomal dominant epilepsy with auditory features; and mTORC1-repressor DEPDC5 (DEP domain-containing protein 5) gene has recently been reported in a broad spectrum of inherited focal epilepsies (ADNFLE, FTLE, FFEVF). This chapter focuses on DEPDC5, a newly identified gene.
Ongoing challenges in diagnosing focal cortical dysplasia (FCD) mandate continuous research and consensus agreement to improve disease definition and classification. An International League Against ...Epilepsy (ILAE) Task Force (TF) reviewed the FCD classification of 2011 to identify existing gaps and provide a timely update. The following methodology was applied to achieve this goal: a survey of published literature indexed with ((Focal Cortical Dysplasia) AND (epilepsy)) between 01/01/2012 and 06/30/2021 (n = 1349) in PubMed identified the knowledge gained since 2012 and new developments in the field. An online survey consulted the ILAE community about the current use of the FCD classification scheme with 367 people answering. The TF performed an iterative clinico‐pathological and genetic agreement study to objectively measure the diagnostic gap in blood/brain samples from 22 patients suspicious for FCD and submitted to epilepsy surgery. The literature confirmed new molecular‐genetic characterizations involving the mechanistic Target Of Rapamycin (mTOR) pathway in FCD type II (FCDII), and SLC35A2 in mild malformations of cortical development (mMCDs) with oligodendroglial hyperplasia (MOGHE). The electro‐clinical‐imaging phenotypes and surgical outcomes were better defined and validated for FCDII. Little new information was acquired on clinical, histopathological, or genetic characteristics of FCD type I (FCDI) and FCD type III (FCDIII). The survey identified mMCDs, FCDI, and genetic characterization as fields for improvement in an updated classification. Our iterative clinico‐pathological and genetic agreement study confirmed the importance of immunohistochemical staining, neuroimaging, and genetic tests to improve the diagnostic yield. The TF proposes to include mMCDs, MOGHE, and “no definite FCD on histopathology” as new categories in the updated FCD classification. The histopathological classification can be further augmented by advanced neuroimaging and genetic studies to comprehensively diagnose FCD subtypes; these different levels should then be integrated into a multi‐layered diagnostic scheme. This update may help to foster multidisciplinary efforts toward a better understanding of FCD and the development of novel targeted treatment options.
Genetic models of focal epilepsies Boillot, Morgane; Baulac, Stéphanie
Journal of neuroscience methods,
02/2016, Letnik:
260
Journal Article
Recenzirano
Odprti dostop
•Several focal epilepsy syndromes are proven to be monogenic disorders.•Mutations in CHRNA4, CHRNB2, CHRNA2 and KCNT1 cause autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE).•Mutations in ...LGI1 cause autosomal dominant epilepsy with auditory features (ADEAF).•This review provides an update of the mutational spectrum in these genes.•We review cellular and genetic animal models generated for autosomal dominant focal epilepsies.
Focal epilepsies were for a long time thought to be acquired disorders secondary to cerebral lesions. However, the important role of genetic factors in focal epilepsies is now well established. Several focal epilepsy syndromes are now proven to be monogenic disorders. While earlier genetic studies suggested a strong contribution of ion channel and neurotransmitter receptor genes, later work has revealed alternative pathways, among which the mammalian target of rapamycin (mTOR) signal transduction pathway with DEPDC5. In this article, we provide an update on the mutational spectrum of neuronal nicotinic acetylcholine receptor genes (CHRNA4, CHRNB2, CHRNA2) and KCNT1 causing autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), and of LGI1 in autosomal dominant epilepsy with auditory features (ADEAF). We also emphasize, through a review of the current literature, the contribution of in vitro and in vivo models developed to unveil the pathogenic mechanisms underlying these two epileptic syndromes.
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.
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.
The identification of lesion‐specific somatic variants, gene fusion, and epigenetic profiles has helped to rationalize disease classification in cancer and more recently also in epilepsy. Many structural brain lesions which are commonly associated with epilepsy including low‐grade epilepsy‐associated tumors (LEAT) and the broad spectrum of malformations of cortical development (MCD) have proven difficult to diagnose based on their histopathology alone. Here, molecular diagnostic approaches can be a game‐changer and help provide more accurate diagnoses as well as identify new clinically meaningful disease entities.
Abstract DEP-domain containing 5 ( DEPDC5 ), encoding a repressor of the mechanistic target of rapamycin complex 1 (mTORC1) signaling pathway, has recently emerged as a major gene mutated in familial ...focal epilepsies and focal cortical dysplasia. Here we established a global knockout rat using TALEN technology to investigate in vivo the impact of Depdc5 -deficiency. Homozygous Depdc5−/− embryos died from embryonic day 14.5 due to a global growth delay. Constitutive mTORC1 hyperactivation was evidenced in the brains and in cultured fibroblasts of Depdc5−/− embryos, as reflected by enhanced phosphorylation of its downstream effectors S6K1 and rpS6. Consistently, prenatal treatment with mTORC1 inhibitor rapamycin rescued the phenotype of Depdc5−/− embryos. Heterozygous Depdc5+/− rats developed normally and exhibited no spontaneous electroclinical seizures, but had altered cortical neuron excitability and firing patterns. Depdc5+/− rats displayed cortical cytomegalic dysmorphic neurons and balloon-like cells strongly expressing phosphorylated rpS6, indicative of mTORC1 upregulation, and not observed after prenatal rapamycin treatment. These neuropathological abnormalities are reminiscent of the hallmark brain pathology of human focal cortical dysplasia. Altogether, Depdc5 knockout rats exhibit multiple features of rodent models of mTORopathies, and thus, stand as a relevant model to study their underlying pathogenic mechanisms.
Febrile seizures affect 2-4% of all children and have a strong genetic component. Recurrent mutations in three main genes (SCN1A, SCN1B and GABRG2) have been identified that cause febrile seizures ...with or without epilepsy. Here we report the identification of mutations in STX1B, encoding syntaxin-1B, that are associated with both febrile seizures and epilepsy. Whole-exome sequencing in independent large pedigrees identified cosegregating STX1B mutations predicted to cause an early truncation or an in-frame insertion or deletion. Three additional nonsense or missense mutations and a de novo microdeletion encompassing STX1B were then identified in 449 familial or sporadic cases. Video and local field potential analyses of zebrafish larvae with antisense knockdown of stx1b showed seizure-like behavior and epileptiform discharges that were highly sensitive to increased temperature. Wild-type human syntaxin-1B but not a mutated protein rescued the effects of stx1b knockdown in zebrafish. Our results thus implicate STX1B and the presynaptic release machinery in fever-associated epilepsy syndromes.
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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