Recognition and epileptology of protracted CLN3 disease.

OBJECTIVE: This study was undertaken to analyze phenotypic features of a cohort of patients with protracted CLN3 disease to improve recognition of the disorder. METHODS: We analyzed phenotypic data of 10 patients from six families with protracted CLN3 disease. Haplotype analysis was performed in three reportedly unrelated families. RESULTS: Visual impairment was the initial symptom, with onset at 5-9 years, similar to classic CLN3 disease. Mean time from onset of visual impairment to seizures was 12 years (range = 6-41 years). Various seizure types were reported, most commonly generalized tonic-clonic seizures; focal seizures were present in four patients. Progressive myoclonus epilepsy was not seen. Interictal electroencephalogram revealed mild background slowing and 2.5-3.5-Hz spontaneous generalized spike-wave discharges. Additional interictal focal epileptiform discharges were noted in some patients. Age at death for the three deceased patients was 31, 31, and 52 years. Molecular testing revealed five individuals were homozygous for c.461-280_677 + 382del966, the "common 1-kb" CLN3 deletion. The remaining individuals were compound heterozygous for various combinations of recurrent pathogenic CLN3 variants. Haplotype analysis demonstrated evidence of a common founder for the common 1-kb deletion. Dating analysis suggested the deletion arose approximately 1500 years ago and thus did not represent cryptic familial relationship in this Australian cohort. SIGNIFICANCE: We highlight the protracted phenotype of a disease generally associated with death in adolescence, which is a combined focal and generalized epilepsy syndrome with progressive neurological deterioration. The disorder should be suspected in an adolescent or adult patient presenting with generalized or focal seizures preceded by progressive visual loss. The common 1-kb deletion has been typically associated with classic CLN3 disease, and the protracted phenotype has not previously been reported with this genotype. This suggests that modifying genetic factors may be important in determining this somewhat milder phenotype and identification of these factors should be the subject of future research.

Defective lipid signalling caused by mutations in PIK3C2B underlies focal epilepsy.

Epilepsy is one of the most frequent neurological diseases, with focal epilepsy accounting for the largest number of cases. The genetic alterations involved in focal epilepsy are far from being fully elucidated. Here, we show that defective lipid signalling caused by heterozygous ultra-rare variants in PIK3C2B, encoding for the class II phosphatidylinositol 3-kinase PI3K-C2ß, underlie focal epilepsy in humans. We demonstrate that patients' variants act as loss-of-function alleles, leading to impaired synthesis of the rare signalling lipid phosphatidylinositol 3,4-bisphosphate, resulting in mTORC1 hyperactivation. In vivo, mutant Pik3c2b alleles caused dose-dependent neuronal hyperexcitability and increased seizure susceptibility, indicating haploinsufficiency as a key driver of disease. Moreover, acute mTORC1 inhibition in mutant mice prevented experimentally induced seizures, providing a potential therapeutic option for a selective group of patients with focal epilepsy. Our findings reveal an unexpected role for class II PI3K-mediated lipid signalling in regulating mTORC1-dependent neuronal excitability in mice and humans.

Sporadic hypothalamic hamartoma is a ciliopathy with somatic and bi-allelic contributions.

Hypothalamic hamartoma with gelastic seizures is a well-established cause of drug-resistant epilepsy in early life. The development of novel surgical techniques has permitted the genomic interrogation of hypothalamic hamartoma tissue. This has revealed causative mosaic variants within GLI3, OFD1 and other key regulators of the sonic-hedgehog pathway in a minority of cases. Sonic-hedgehog signalling proteins localize to the cellular organelle primary cilia. We therefore explored the hypothesis that cilia gene variants may underlie hitherto unsolved cases of sporadic hypothalamic hamartoma. We performed high-depth exome sequencing and chromosomal microarray on surgically resected hypothalamic hamartoma tissue and paired leukocyte-derived DNA from 27 patients. We searched for both germline and somatic variants under both dominant and bi-allelic genetic models. In hamartoma-derived DNA of seven patients we identified bi-allelic (one germline, one somatic) variants within one of four cilia genes-DYNC2I1, DYNC2H1, IFT140 or SMO. In eight patients, we identified single somatic variants in the previously established hypothalamic hamartoma disease genes GLI3 or OFD1. Overall, we established a plausible molecular cause for 15/27 (56%) patients. Here, we expand the genetic architecture beyond single variants within dominant disease genes that cause sporadic hypothalamic hamartoma to bi-allelic (one germline/one somatic) variants, implicate three novel cilia genes and reconceptualize the disorder as a ciliopathy.

Evidence for a Dual-Pathway, 2-Hit Genetic Model for Focal Cortical Dysplasia and Epilepsy.

BACKGROUND AND OBJECTIVES: The 2-hit model of genetic disease is well established in cancer, yet has only recently been reported to cause brain malformations associated with epilepsy. Pathogenic germline and somatic variants in genes in the mechanistic target of rapamycin (mTOR) pathway have been implicated in several malformations of cortical development. We investigated the 2-hit model by performing genetic analysis and searching for germline and somatic variants in genes in the mTOR and related pathways. METHODS: We searched for germline and somatic pathogenic variants in 2 brothers with drug-resistant focal epilepsy and surgically resected focal cortical dysplasia (FCD) type IIA. Exome sequencing was performed on blood- and brain-derived DNA to identify pathogenic variants, which were validated by droplet digital PCR. In vitro functional assays of a somatic variant were performed. RESULTS: Exome analysis revealed a novel, maternally inherited, germline pathogenic truncation variant (c.48delG; p.Ser17Alafs*70) in NPRL3 in both brothers. NPRL3 is a known FCD gene that encodes a negative regulator of the mTOR pathway. Somatic variant calling in brain-derived DNA from both brothers revealed a low allele fraction somatic variant (c.338C>T; p.Ala113Val) in the WNT2 gene in 1 brother, confirmed by droplet digital PCR. In vitro functional studies suggested a loss of WNT2 function as a consequence of this variant. A second somatic variant has not yet been found in the other brother. DISCUSSION: We identify a pathogenic germline mTOR pathway variant (NPRL3) and a somatic variant (WNT2) in the intersecting WNT signaling pathway, potentially implicating the WNT2 gene in FCD and supporting a dual-pathway 2-hit model. If confirmed in other cases, this would extend the 2-hit model to pathogenic variants in different genes in critical, intersecting pathways in a malformation of cortical development. Detection of low allele fraction somatic second hits is challenging but promises to unravel the molecular architecture of FCDs.

Genome-wide association study of febrile seizures implicates fever response and neuronal excitability genes.

Febrile seizures represent the most common type of pathological brain activity in young children and are influenced by genetic, environmental and developmental factors. In a minority of cases, febrile seizures precede later development of epilepsy. We conducted a genome-wide association study of febrile seizures in 7635 cases and 83 966 controls identifying and replicating seven new loci, all with P < 5 × 10-10. Variants at two loci were functionally related to altered expression of the fever response genes PTGER3 and IL10, and four other loci harboured genes (BSN, ERC2, GABRG2, HERC1) influencing neuronal excitability by regulating neurotransmitter release and binding, vesicular transport or membrane trafficking at the synapse. Four previously reported loci (SCN1A, SCN2A, ANO3 and 12q21.33) were all confirmed. Collectively, the seven novel and four previously reported loci explained 2.8% of the variance in liability to febrile seizures, and the single nucleotide polymorphism heritability based on all common autosomal single nucleotide polymorphisms was 10.8%. GABRG2, SCN1A and SCN2A are well-established epilepsy genes and, overall, we found positive genetic correlations with epilepsies (rg = 0.39, P = 1.68 × 10-4). Further, we found that higher polygenic risk scores for febrile seizures were associated with epilepsy and with history of hospital admission for febrile seizures. Finally, we found that polygenic risk of febrile seizures was lower in febrile seizure patients with neuropsychiatric disease compared to febrile seizure patients in a general population sample. In conclusion, this largest genetic investigation of febrile seizures to date implicates central fever response genes as well as genes affecting neuronal excitability, including several known epilepsy genes. Further functional and genetic studies based on these findings will provide important insights into the complex pathophysiological processes of seizures with and without fever.

Progressive myoclonus epilepsies-Residual unsolved cases have marked genetic heterogeneity including dolichol-dependent protein glycosylation pathway genes.

Progressive myoclonus epilepsies (PMEs) comprise a group of clinically and genetically heterogeneous rare diseases. Over 70% of PME cases can now be molecularly solved. Known PME genes encode a variety of proteins, many involved in lysosomal and endosomal function. We performed whole-exome sequencing (WES) in 84 (78 unrelated) unsolved PME-affected individuals, with or without additional family members, to discover novel causes. We identified likely disease-causing variants in 24 out of 78 (31%) unrelated individuals, despite previous genetic analyses. The diagnostic yield was significantly higher for individuals studied as trios or families (14/28) versus singletons (10/50) (OR = 3.9, p value = 0.01, Fisher's exact test). The 24 likely solved cases of PME involved 18 genes. First, we found and functionally validated five heterozygous variants in NUS1 and DHDDS and a homozygous variant in ALG10, with no previous disease associations. All three genes are involved in dolichol-dependent protein glycosylation, a pathway not previously implicated in PME. Second, we independently validate SEMA6B as a dominant PME gene in two unrelated individuals. Third, in five families, we identified variants in established PME genes; three with intronic or copy-number changes (CLN6, GBA, NEU1) and two very rare causes (ASAH1, CERS1). Fourth, we found a group of genes usually associated with developmental and epileptic encephalopathies, but here, remarkably, presenting as PME, with or without prior developmental delay. Our systematic analysis of these cases suggests that the small residuum of unsolved cases will most likely be a collection of very rare, genetically heterogeneous etiologies.

Cerebrospinal fluid liquid biopsy for detecting somatic mosaicism in brain.

Brain somatic mutations are an increasingly recognized cause of epilepsy, brain malformations and autism spectrum disorders and may be a hidden cause of other neurodevelopmental and neurodegenerative disorders. At present, brain mosaicism can be detected only in the rare situations of autopsy or brain biopsy. Liquid biopsy using cell-free DNA derived from cerebrospinal fluid has detected somatic mutations in malignant brain tumours. Here, we asked if cerebrospinal fluid liquid biopsy can be used to detect somatic mosaicism in non-malignant brain diseases. First, we reliably quantified cerebrospinal fluid cell-free DNA in 28 patients with focal epilepsy and 28 controls using droplet digital PCR. Then, in three patients we identified somatic mutations in cerebrospinal fluid: in one patient with subcortical band heterotopia the LIS1 p. Lys64* variant at 9.4% frequency; in a second patient with focal cortical dysplasia the TSC1 p. Phe581His*6 variant at 7.8% frequency; and in a third patient with ganglioglioma the BRAF p. Val600Glu variant at 3.2% frequency. To determine if cerebrospinal fluid cell-free DNA was brain-derived, whole-genome bisulphite sequencing was performed and brain-specific DNA methylation patterns were found to be significantly enriched (P = 0.03). Our proof of principle study shows that cerebrospinal fluid liquid biopsy is valuable in investigating mosaic neurological disorders where brain tissue is unavailable.

Contribution of rare genetic variants to drug response in absence epilepsy.

OBJECTIVE: We investigated the possible significance of rare genetic variants to response to valproic acid (VPA) and ethosuximide (ETX) in patients with absence epilepsy. Our primary hypothesis was that rare CACNA1H variants are more frequent in ETX-non-responsive patients compared to ETX-responsive. Our secondary hypothesis was that rare variants in GABA-receptor genes are more frequent in VPA-non-responsive patients compared to VPA-responsive. METHODS: We recruited patients with absence epilepsy treated with both VPA and ETX, and performed whole exome sequencing in order to investigate the potential role of rare variants in CACNA1H, other voltage-gated calcium channel (VGCC) genes, or GABA-receptor genes in predicting response to ETX or VPA. RESULTS: Sixty-two patients were included; 12 were ETX-responsive, 14 VPA-responsive, and 36 did not have a clear positive response to either medication. We did not find significant enrichment inCACNA1H rare variants in ETX-responsive patients (odds ratio 3.43; 0.43-27.65; p = 0.20), nor was there enrichment for other VGCC genes. No significant enrichment of GABA-receptor gene rare variants was seen for VPA-non-responsive patients versus VPA-responsive. We found enrichment of rare GABA-receptor variants in our absence cohort compared to controls (odds ratio 3.82; 1.68-8.69). There was no difference in frequency of CACNA1H rs61734410 and CACNA1I rs3747178 polymorphisms between ETX-responsive and ETX-non-responsive groups; these polymorphisms have previously been reported to predict lack of response to ETX in absence epilepsy. SIGNIFICANCE: We conclude that if CACNA1H rare variants predict lack of response to ETX, a larger sample is necessary to test this with sufficient power. Increased GABA-receptor gene rare variant frequency in absence epilepsy patients who fail initial anti-seizure therapy suggests subtle GABA receptor dysfunction may contribute to the underlying pathophysiology.

Pathogenic Variants in CEP85L Cause Sporadic and Familial Posterior Predominant Lissencephaly.

Lissencephaly (LIS), denoting a "smooth brain," is characterized by the absence of normal cerebral convolutions with abnormalities of cortical thickness. Pathogenic variants in over 20 genes are associated with LIS. The majority of posterior predominant LIS is caused by pathogenic variants in LIS1 (also known as PAFAH1B1), although a significant fraction remains without a known genetic etiology. We now implicate CEP85L as an important cause of posterior predominant LIS, identifying 13 individuals with rare, heterozygous CEP85L variants, including 2 families with autosomal dominant inheritance. We show that CEP85L is a centrosome protein localizing to the pericentriolar material, and knockdown of Cep85l causes a neuronal migration defect in mice. LIS1 also localizes to the centrosome, suggesting that this organelle is key to the mechanism of posterior predominant LIS.

SCN1A Variants in vaccine-related febrile seizures: A prospective study.

OBJECTIVE: Febrile seizures may follow vaccination. Common variants in the sodium channel gene, SCN1A, are associated with febrile seizures, and rare pathogenic variants in SCN1A cause the severe developmental and epileptic encephalopathy Dravet syndrome. Following vaccination, febrile seizures may raise the specter of poor outcome and inappropriately implicate vaccination as the cause. We aimed to determine the prevalence of SCN1A variants in children having their first febrile seizure either proximal to vaccination or unrelated to vaccination compared to controls. METHODS: We performed SCN1A sequencing, blind to clinical category, in a prospective cohort of children presenting with their first febrile seizure as vaccine proximate (n = 69) or as non-vaccine proximate (n = 75), and children with no history of seizures (n = 90) recruited in Australian pediatric hospitals. RESULTS: We detected 2 pathogenic variants in vaccine-proximate cases (p.R568X and p.W932R), both of whom developed Dravet syndrome, and 1 in a non-vaccine-proximate case (p.V947L) who had febrile seizures plus from 9 months. All had generalized tonic-clonic seizures lasting >15 minutes. We also found enrichment of a reported risk allele, rs6432860-T, in children with febrile seizures compared to controls (odds ratio = 1.91, 95% confidence interval = 1.31-2.81). INTERPRETATION: Pathogenic SCN1A variants may be identified in infants with vaccine-proximate febrile seizures. As early diagnosis of Dravet syndrome is essential for optimal management and outcome, SCN1A sequencing in infants with prolonged febrile seizures, proximate to vaccination, should become routine. ANN NEUROL 2020;87:281-288.

Epidemiology and etiology of infantile developmental and epileptic encephalopathies in Tasmania.

We sought to determine incidence, etiologies, and yield of genetic testing in infantile onset developmental and epileptic encephalopathies (DEEs) in a population isolate, with an intensive multistage approach. Infants born in Tasmania between 2011 and 2016, with seizure onset <2 years of age, epileptiform EEG, frequent seizures, and developmental impairment, were included. Following review of EEG databases, medical records, brain MRIs, and other investigations, clinical genetic testing was undertaken with subsequent research interrogation of whole exome sequencing (WES) in unsolved cases. The incidence of infantile DEEs was 0.44/1000 per year (95% confidence interval 0.25 to 0.71), with 16 cases ascertained. The etiology was structural in 5/16 cases. A genetic basis was identified in 6 of the remaining 11 cases (3 gene panel, 3 WES). In two further cases, WES identified novel variants with strong in silico data; however, paternal DNA was not available to support pathogenicity. The etiology was not determined in 3/16 (19%) cases, with a candidate gene identified in one of these. Pursuing clinical imaging and genetic testing followed by WES at an intensive research level can give a high diagnostic yield in the infantile DEEs, providing a solid base for prognostic and genetic counseling.

Kufs disease due to mutation of CLN6: clinical, pathological and molecular genetic features.

Kufs disease is the major adult form of neuronal ceroid lipofuscinosis, but is rare and difficult to diagnose. Diagnosis was traditionally dependent on the demonstration of characteristic storage material, but distinction from normal age-related accumulation of lipofuscin can be challenging. Mutation of CLN6 has emerged as the most important cause of recessive Kufs disease but, remarkably, is also responsible for variant late infantile ceroid lipofuscinosis. Here we provide a detailed description of Kufs disease due to CLN6 pathogenic variants. We studied 20 cases of Kufs disease with CLN6 pathogenic variants from 13 unrelated families. Mean age of onset was 28 years (range 12-51) with bimodal peaks in teenage and early adult life. The typical presentation was of progressive myoclonus epilepsy with debilitating myoclonic seizures and relatively infrequent tonic-clonic seizures. Patients became wheelchair-bound with a mean 12 years post-onset. Ataxia was the most prominent motor feature. Dementia appeared to be an invariable accompaniment, although it could take a number of years to manifest and occasionally cognitive impairment preceded myoclonic seizures. Patients were usually highly photosensitive on EEG. MRI showed progressive cerebral and cerebellar atrophy. The median survival time was 26 years from disease onset. Ultrastructural examination of the pathology revealed fingerprint profiles as the characteristic inclusions, but they were not reliably seen in tissues other than brain. Curvilinear profiles, which are seen in the late infantile form, were not a feature. Of the 13 unrelated families we observed homozygous CLN6 pathogenic variants in four and compound heterozygous variants in nine. Compared to the variant late infantile form, there was a lower proportion of variants that predicted protein truncation. Certain heterozygous missense variants in the same amino acid position were found in both variant late infantile and Kufs disease. There was a predominance of cases from Italy and surrounding regions; this was partially explained by the discovery of three founder pathogenic variants. Clinical distinction of type A (progressive myoclonus epilepsy) and type B (dementia with motor disturbance) Kufs disease was supported by molecular diagnoses. Type A is usually caused by recessive pathogenic variants in CLN6 or dominant variants in DNAJC5. Type B Kufs is usually associated with recessive CTSF pathogenic variants. The diagnosis of Kufs remains challenging but, with the availability of genetic diagnosis, this will largely supersede the use of diagnostic biopsies, particularly as biopsies of peripheral tissues has unsatisfactory sensitivity and specificity.

Evidence of linkage to chromosome 5p13.2-q11.1 in a large inbred family with genetic generalized epilepsy.

The clinical genetics of genetic generalized epilepsy suggests complex inheritance; large pedigrees, with multiple affected individuals, are rare exceptions. We studied a large consanguineous family from Turkey where extensive electroclinical phenotyping revealed a familial phenotype most closely resembling juvenile myoclonic epilepsy. For a subject to be considered affected (n = 14), a diagnostic electroencephalogram was required. Seizure onset ranged between 6 and 19 years (mean = 12 years). Thirteen of 14 experienced myoclonic jerks; in 11, this was associated with eyelid blinking, and in 10 it was interspersed with absences. Generalized tonic-clonic seizures were seen in 11. One individual had generalized tonic-clonic seizures alone. Electroencephalograms demonstrated generalized polyspike and wave discharges that were not associated with photoparoxysmal response. Intellect was normal. Nineteen family members were subsequently chosen for nonparametric multipoint linkage analyses, which identified a 39.5 Mb region on chromosome 5 (P < 0.0001). Iterative analysis, including discovery of a subtly affected individual, narrowed the critical region to 15.4 Mb and possibly to 5.5 Mb. Homozygous versus heterozygous state of the refined 5p13.2-q11.1 haplotype was not associated with phenotypic severity or onset age, suggesting that one versus two pathogenic variants may result in similar phenotypes. Whole exome sequencing (n = 3) failed to detect any rare, protein-coding variants within the highly significant linkage region that includes HCN1 as a promising candidate.

Somatic GNAQ mutation in the forme fruste of Sturge-Weber syndrome.

OBJECTIVE: To determine whether the GNAQ R183Q mutation is present in the forme fruste cases of Sturge-Weber syndrome (SWS) to establish a definitive molecular diagnosis. METHODS: We used sensitive droplet digital PCR (ddPCR) to detect and quantify the GNAQ mutation in tissues from epilepsy surgery in 4 patients with leptomeningeal angiomatosis; none had ocular or cutaneous manifestations. RESULTS: Low levels of the GNAQ mutation were detected in the brain tissue of all 4 cases-ranging from 0.42% to 7.1% frequency-but not in blood-derived DNA. Molecular evaluation confirmed the diagnosis in 1 case in which the radiologic and pathologic data were equivocal. CONCLUSIONS: We detected the mutation at low levels, consistent with mosaicism in the brain or skin (1.0%-18.1%) of classic cases. Our data confirm that the forme fruste is part of the spectrum of SWS, with the same molecular mechanism as the classic disease and that ddPCR is helpful where conventional diagnosis is uncertain.

Sensitive quantitative detection of somatic mosaic mutation in "double cortex" syndrome.

Somatic mutation of the lissencephaly-1 gene is a cause of subcortical band heterotopia ("double cortex"). The severity of the phenotype depends on the level of mutation in brain tissue. Detecting and quantifying low-level somatic mosaic mutations is challenging. Here, we utilized droplet digital PCR, a sensitive method to detect low-level mutation. Droplet digital PCR was used in concert with classic genotyping techniques (SNaPshot assays and pyrosequencing) to detect and characterize the tissue mosaicism of a somatic mutation (LIS1 c.190A>T; p.K64X) in a patient with posterior bilateral SBH and refractory epilepsy. The high sensitivity of droplet digital PCR and the ability to target individual DNA molecules allowed us to detect the mutation at low level in the brain, despite the low quality of the DNA sample derived from formalin-fixed paraffin-embedded tissue. This low mutation frequency in the brain was consistent with the relatively subtle malformation resolved by magnetic resonance imaging. The presence of the mutation in other tissues from the patient permitted us to predict the timing of mutagenesis. This sensitive methodology will have utility for a variety of other brain malformation syndromes associated with epilepsy for which historical pathological specimens are available and specific somatic mosaic mutations are predicted.

Genetic epilepsy with febrile seizures plus: Refining the spectrum.

OBJECTIVE: Following our original description of generalized epilepsy with febrile seizures plus (GEFS+) in 1997, we analyze the phenotypic spectrum in 409 affected individuals in 60 families (31 new families) and expand the GEFS+ spectrum. METHODS: We performed detailed electroclinical phenotyping on all available affected family members. Genetic analysis of known GEFS+ genes was carried out where possible. We compared our phenotypic and genetic data to those published in the literature over the last 19 years. RESULTS: We identified new phenotypes within the GEFS+ spectrum: focal seizures without preceding febrile seizures (16/409 [4%]), classic genetic generalized epilepsies (22/409 [5%]), and afebrile generalized tonic-clonic seizures (9/409 [2%]). Febrile seizures remains the most frequent phenotype in GEFS+ (178/409 [44%]), followed by febrile seizures plus (111/409 [27%]). One third (50/163 [31%]) of GEFS+ families tested have a pathogenic variant in a known GEFS+ gene. CONCLUSION: As 37/409 (9%) affected individuals have focal epilepsies, we suggest that GEFS+ be renamed genetic epilepsy with febrile seizures plus rather than generalized epilepsy with febrile seizures plus. The phenotypic overlap between GEFS+ and the classic generalized epilepsies is considerably greater than first thought. The clinical and molecular data suggest that the 2 major groups of generalized epilepsies share genetic determinants.

Evaluation of GLUT1 variation in non-acquired focal epilepsy.

Brain glucose transport is dependent on glucose transporter 1 (GLUT1), encoded by the solute carrier family 2 member 1 (SLC2A1) gene. Mutations in SLC2A1 cause GLUT1 deficiency which is characterized by a broad spectrum of neurological phenotypes including generalized epilepsy, motor disorders, developmental delay and microcephaly. Recent case reports suggest SLC2A1 mutations can contribute to non-acquired focal epilepsy (NAFE) but interrogation of a large patient cohort has not been reported. We studied 200 patients with NAFE (126 with temporal lobe epilepsy) comprising 104 females and 96 males with a mean age of onset of 18 years. Polymerase chain reaction (PCR) and Sanger sequencing was performed to detect variants in all 10 coding exons and splice site regions of the SLC2A1 gene. We did not detect any pathogenic mutations in SLC2A1 in this cohort. Our data suggests that the frequency of GLUT1 mutations in NAFE is low. Limitations of this study include the mean age of onset and cohort size. Future research should focus on subpopulations of focal epilepsy with lower age of seizure onset particularly with co-existent movement disorders in which GLUT1 mutations may play a more important role.

Real-world utility of whole exome sequencing with targeted gene analysis for focal epilepsy.

OBJECTIVE: Driven by advances in genomic technology and reduction in costs, next-generation sequencing (NGS) is venturing into routine clinical care. The 'real-world' clinical utility of NGS remains to be determined in focal epilepsies, which account for 60% of all epilepsies and for which the importance of genetic factors is just beginning to emerge. We investigated the diagnostic yield and management implications of whole exome sequencing (WES)-based screening of selected genes in the routine care of common focal epilepsies suspected to have a genetic basis. METHODS: We performed WES, followed by targeted analysis of 64 epilepsy genes, on 40 consecutive children and adults enrolled prospectively from routine clinical practice who had MRI-negative focal epilepsy and a family history of febrile seizures or any type of epilepsy in at least one first- or second-degree relative. Exclusion criteria were previous genetic testing, severe intellectual disability and benign focal epilepsies of childhood. RESULTS: 5/40 (12.5%) patients had a pathogenic or likely pathogenic variant, detected in SCN1A, DEPDC5, PCDH19, GABRG2 or NPRL2. Identifying a pathogenic SCN1A variant in a patient with drug-resistant epilepsy prompted to halt presurgical investigations due to concern of unfavorable post-surgical outcome. It also led in the same patient to discontinue long-standing carbamazepine therapy (a potentially aggravating drug in epilepsies due to SCN1A mutations), resulting in complete seizure control. Patients with pathogenic or likely pathogenic variants had a younger median age of seizure onset (range) compared to those without [18 months (8 months-18 years) vs 18 years (18 months-70 years), p=0.02]. SIGNIFICANCE: Our data demonstrate that WES with targeted gene analysis is an effective diagnostic tool for patients with common focal epilepsies in whom a genetic etiology is suspected. It can also influence clinical decision-making, including antiepileptic drug selection and consideration of epilepsy surgery, hence supporting its incorporation in the routine clinical care of this patient group.

SCN1A clinical spectrum includes the self-limited focal epilepsies of childhood.

INTRODUCTION: Amongst autosomal dominant genetic epilepsy with febrile seizures plus (GEFS+) families, SCN1A variants are the most common genetic cause. Initially regarded as a generalized form of epilepsy, the GEFS+ spectrum is now known to include some focal epilepsies, but it is generally not conceptualized as extending to the self-limited focal epilepsies of childhood, such as Panayiotopoulos syndrome. There are, however, three reports of SCN1A variants in Panayiotopoulos syndrome. We describe the variable clinical phenotypes that include the self-limited focal epilepsies of childhood, present in a large GEFS+ family, segregating a heterozygous SCN1A missense variant. MATERIAL AND METHODS: Electro-clinical details on all putatively affected family members were sought and blood samples were taken for genetic analysis. Two individuals were chosen for SCN1A testing. All 26 exons and exon-intron junctions were amplified, sequenced and analyzed. This was followed by pedigree segregation analysis of the variant identified. RESULTS: A pathogenic heterozygous SCN1A (c.2624C>A; p.Thr875Lys) variant was identified. Sixteen of the 18 variant positive family members were affected (88% penetrance): 8 with febrile seizures, 2 febrile seizures plus, 1 unclassified seizures and 5 with self-limited focal epilepsy of childhood. Of these, one was diagnosed with atypical childhood epilepsy with centrotemporal spikes and four with Panayiotopoulos syndrome. DISCUSSION: By characterizing the heterogeneous clinical phenotypes in a large, SCN1A mutation positive GEFS+ family, we conclude that the GEFS+ spectrum can extend to the self-limited focal epilepsies of childhood, including Panayiotopoulos syndrome, and in turn highlight the complex genotype-phenotype correlations associated with SCN1A-related epilepsies.

De novo SCN1A pathogenic variants in the GEFS+ spectrum: Not always a familial syndrome.

Genetic epilepsy with febrile seizures plus (GEFS+) is a familial epilepsy syndrome characterized by heterogeneous phenotypes ranging from mild disorders such as febrile seizures to epileptic encephalopathies (EEs) such as Dravet syndrome (DS). Although DS often occurs with de novo SCN1A pathogenic variants, milder GEFS+ spectrum phenotypes are associated with inherited pathogenic variants. We identified seven cases with non-EE GEFS+ phenotypes and de novo SCN1A pathogenic variants, including a monozygotic twin pair. Febrile seizures plus (FS+) occurred in six patients, five of whom had additional seizure types. The remaining case had childhood-onset temporal lobe epilepsy without known febrile seizures. Although early development was normal in all individuals, three later had learning difficulties, and the twin girls had language impairment and working memory deficits. All cases had SCN1A missense pathogenic variants that were not found in either parent. One pathogenic variant had been reported previously in a case of DS, and the remainder were novel. Our finding of de novo pathogenic variants in mild phenotypes within the GEFS+ spectrum shows that mild GEFS+ is not always inherited. SCN1A screening should be considered in patients with GEFS+ phenotypes because identification of pathogenic variants will influence antiepileptic therapy, and prognostic and genetic counseling.