Genetic Risk Factors for Neurological Disorders in Children with Adverse Events Following Immunization: A Descriptive Study of a Polish Case Series.

Studies conducted on large populations show a lack of connection between vaccination and serious neurological symptoms. However, there are isolated cases that indicate such a relationship. These reports on adverse effects following immunization (AEFI) reduce social confidence in vaccination; however, their background may be rare genetic defects. The aim of the presented study was to examine if neurological AEFI in children may be associated with variants in genes related to neurodevelopment. To identify such possible associations, a descriptive study of the Polish case series was conducted. We performed next-generation sequencing in patients who, up to 4 weeks of injection of any vaccine, manifested neurological AEFI. We included 23 previously normally developing children with first seizures that occurred after vaccination. We identified pathogenic/likely pathogenic variants in genes engaged in neurodevelopment in nine patients and variants of uncertain significance in another nine patients. The mutated genes belonged to the group of genes related to epilepsy syndromes/epileptic encephalopathy. We showed that AEFI might have a genetic background. We hypothesized that in some AEFI patients, the vaccine might only trigger neurological symptoms that would have been manifested anyway as a result of a pathogenic variant in a gene engaged in neurodevelopment.

Hypertrophic olivary degeneration in a 16-year-old girl after subtotal surgery of a brainstem pilocytic astrocytoma - a case report.

PURPOSE: Hypertrophic olivary degeneration (HOD) is a unique neurological condition caused by interruption of the dentato-rubro-olivary pathway, also known as the triangle of Guillain and Mollaret. Magnetic resonance (MR) imaging is the best modality to diagnose both the degeneration of the inferior olivary nucleus and the underlying cause. CASE REPORT: We describe a case of a unilateral HOD in a 16-year-old girl several months after a subtotal excision of a brainstem pilocytic astrocytoma. Taking into account the patient's history, tumour recurrence must have been considered, but the typical location and MR morphology, as well as the time of occurrence after brainstem surgery, contributed to the diagnosis of HOD. The causative factor was the interruption of the central tegmental tract, which forms one arm of the Guillain and Mollaret triangle. Additionally, this is an interesting case of a child, who stayed in a coma for several months following brainstem surgery, but finally was discharged home with only minor neurological defects and returned to normal life. CONCLUSIONS: Hypertrophic olivary degeneration is an infrequent neurological condition, especially in the paediatric population. Nevertheless, it should be considered when interpreting late postoperative scans of children with a history of a brain tumour.

Neuroimaging results, short-term assessment of psychomotor development and the risk of autism spectrum disorder in extremely premature infants (≤28 GA) - a prospective cohort study (preliminary report).

OBJECTIVE: Infants ≤28 GA are at particular risk of psychomotor and neurological developmental disorder. They also remain at a higher risk of developing autism spectrum disorder (ASD), characterized by persistent deficits in communication/social interactions and restricted, repetitive behaviors, activities and interests. Monitoring their development by a team of specialists (a neurologist, psychologist, psychiatrist) allows us to make an early diagnosis and to implement appropriate therapy. Neuroimaging studies during the neonatal period may be helpful in clarifying diagnosis and prognosis. Objective: The aim of the study was to search for the interrelation between the results of neuroimaging and the neurological, psychological and psychiatric evaluation at the age of 2. PATIENTS AND METHODS: Material and methods: Neonates born at ≤28 weeks between 01.06.2013 and 31.12.2015 and hospitalized at NICU were enrolled. We present the results of the first 12 children who have attained 2 years of corrected age and have undergone both neuroimaging, and neurological, psychological and psychiatric assessments. Transfontanel ultrasound was performed according to general standards, MRI between 38 and 42 weeks of corrected age. Neurological examination based on the Denver scale, ASD screening with use of the STAT test and psychological DSR assessment were performed at 2 years of corrected age. RESULTS: Results: Median GA was 26 weeks and median weight 795 g. The ultrasound examination was normal in 9 cases (75%) and MRI in 4 (33%). Abnormalities in the cerebellum were the main additional information found in MRI as compared to US. Neurological examination was normal in 8 infants (67#37;), in 4 of whom neuroimaging was normal. In 4 (33%) infants the neurological examination was abnormal. Psychomotor development at an average level or above was found in seven (58#37;) children. In 4 of them neuroimaging was normal, whereas 3 had ventricular dilatation and haemorrhagic infarct. There were no abnormalities within the cerebellum in this group. In the remaining 5 children (42#37;) psychomotor development was rated as delayed. All of them had cerebellar haemorrhage. An increased risk of ASD was observed in 4 children who developed cerebellar hemorrhage. CONCLUSION: Conclusions: 1. The use of MRI at a term-equivalent age may contribute to the prognosis of neurodevelopmental outcomes in extremely premature infants, allowing risk stratification and thus enhancing early monitoring of a child's development and functional status 2. There is a clear tendency towards abnormal psychomotor development and positive screening for ASD to co-occur with abnormal MRI findings in the cerebellum.

From focal epilepsy to Dravet syndrome--Heterogeneity of the phenotype due to SCN1A mutations of the p.Arg1596 amino acid residue in the Nav1.1 subunit.

OBJECTIVE: The aim of this study was to analyze the intra-/interfamilial phenotypic heterogeneity due to variants at the highly evolutionary conservative p.Arg1596 residue in the Nav1.1 subunit. MATERIALS/PARTICIPANTS: Among patients referred for analysis of the SCN1A gene one recurrent, heritable mutation was found in families enrolled into the study. Probands from those families even clinically diagnosed with atypical Dravet syndrome (DS), generalized epilepsy with febrile seizures plus (GEFS+), and focal epilepsy, had heterozygous p.Arg1596 His/Cys missense substitutions, c.4787G>T and c.4786C>T in the SCN1A gene. METHOD: Full clinical evaluation, including cognitive development, neurological examination, EEGs, MRI was performed in probands and affected family members in developmental age. The whole SCN1A gene sequencing was performed for all probands. The exon 25, where the identified missense substitutions are localized, was directly analyzed for the other family members. RESULTS: Mutation of the SCN1A p.1596Arg was identified in three families, in one case substitution p.Arg1596Cys and in two cases p.Arg1596His. Both mutations were previously described as pathogenic and causative for DS, GEFS+ and focal epilepsy. Spectrum of phenotypes among presented families with p.Arg1596 mutations shows heterogeneity ranged from asymptomatic cases, through FS and FS+ to GEFS+/Panayiotopoulos syndrome and epilepsies with and without febrile seizures, and epileptic encephalopathy such as DS. Phenotypes differ among patients displaying both focal and generalized epilepsies. Some patients demonstrated additionally Asperger syndrome and ataxia. CONCLUSION: Clinical picture heterogeneity of the patients carrying mutation of the same residue indicates the involvement of the other factors influencing the SCN1A gene mutations' penetrance.

Glucose transporter type 1 deficiency due to SLC2A1 gene mutations--a rare but treatable cause of metabolic epilepsy and extrapyramidal movement disorder; own experience and literature review.

THE AIM: To present the molecular and clinical characteristics of three children with glucose deficiency syndrome, an inborn rare metabolic disease, caused by mutations in the SLC2A1 gene. MATERIAL AND METHODS: The investigation was carried out in three children: two girls and one boy showing symptoms of GLUT1 deficiency syndrome (GLUT1-DS). They were referred for SLC2A1 gene analysis. RESULTS: The presence of mutations in all of them was confirmed. Only point mutations were identified, two missenses p.Gly132Ser, p.Arg212Cys and amino acid insertion p.Ser_Val227insValProPro. In two cases the mutations arose de novo, one was heritable of paternal origin. CONCLUSIONS: GLUT1-DS shows high clinical variability. It should be suspected in children of any age presenting with single features or a combination of any form of intractable epilepsy with seizures of various types, movement disorders and paroxysmal events, especially triggered by exercise, exertion, or fasting, and any unexplainable neurological deterioration. The basic diagnostic hallmarks of GLUT1-DS are CSF hypoglycorrhachia and lowered CSF/Blood serum glucose ratio. This is why lumbar punction should be considered more frequently than it is in practice being performed nowadays. Antiepileptic drug treatment may be ineffective or even potentially detrimental. Early identification and molecular confirmation of GLUT1-DS is important, because this is a metabolic disorder and patients should as soon as possible primarily be treated with a ketogenic diet.

[Patterns of the EEG records in children with West Syndrome]. / Wzorce zapisu EEG u dzieci z zespolem Westa.

INTRODUCTION: West Syndrome (WS) (infantile myoclonic encephalopathy with hypsarrhythmia--IMEH) belongs to the infantile epileptic encephalopathies and is characterized by infantile spasms, hypsarrythmia in EEG, and abnormal psychomotor development of children. AIM: Evaluation of the EEG patterns of patients with WS, correlation of the EEG patterns with the cause of epilepsy and an assessment of the influence of antiepileptic drugs (AEDs). CASE REPORTS: EEG patterns of four children with symptomatic WS of different etiology (tuberous sclerosis, brain defects, autoimmune) were analyzed before and during treatment with various antiepileptic drugs. SUMMARY: The basic pattern of EEG in children with WS is hypsarrhythmia. Variabilities of the patterns are the results of degree of development of the child's brain, the etiology of disease, as well as the effects of administration of different antiepileptic drugs..

Genetic epilepsies with febrile seizures plus: clinical spectrum of Polish patients with SCN1A mutation - preliminary report.

UNLABELLED: Diseases caused by mutations in SCN1A are currently named Genetic Epilepsies with Febrile Seizures Plus, and this term stands for expanded spectrum of syndrome previously called as GEFS+ (Generalized Epilepsy with Febrile Seizures Plus). SCN1A is the uniquely identified gene directly linked to specific type of epilepsy, and its testing has been included in the screening processes. THE AIM: To diagnose and describe epileptic syndromes caused by SCN1A mutations. MATERIAL AND METHODS: 203 patients were included in the screening process with suspected SCN1A mutation, based on clinical features and family history. Study group was selected based on inclusion and exclusion criteria and then preliminary epilepsy diagnosis was verified using ILAE classification. Molecular testing to screen SCN1A mutations was performed in the study group. RESULTS: Mutations were detected in 57 cases. Majority of patients (50 cases - 87.5%) suffered from Dravet syndrome, 8.8% (5 cases) were diagnosed as GEFS+, 3% as vaccines encephalopathy and Panayotopoulous syndrome. Mutations were not detected in children with isolated febrile seizures, family febrile seizures nor in patients with myoclonic - astatic epilepsy. CONCLUSIONS: Frequency of mutations in SCN1A in Dravet syndrome and GEFS+ in Polish populations are similar to other countries. Diagnostic clinical criteria are currently insufficient to draw precise diagnosis. There is a strong need to establish clinical criteria for molecular testing and this topic will be investigated in the future.

Use of Fibrin Glue in the Treatment of Persistent Pneumothorax in Premature Infants at the Limit of Viability: Ethical Issues and Two and A Half Years Follow-Up.

INTRODUCTION: Due to the extreme immaturity of many internal organs, including lungs, infants at the limit of viability are more predisposed to a pneumothorax (PTX). In some cases, PTX becomes persistent. Previously, only a few attempts of PTX treatment with fibrin glue were reported. However, its impact on further lung development is unknown. CASE REPORT: We present a case of an extremely preterm infant with persistent PTX who was successfully treated with fibrin glue. In addition, we present a two-and-a-half-year corrected age follow-up focusing on respiratory problems, motor development and sensory organs. Furthermore, we touch upon the related ethical issues. CONCLUSIONS: Fibrin glue should be used to treat persistent PTX even in an extremely preterm infant. No adverse effects were observed. At the two-and-a-half-year corrected age follow-up, despite severe bronchopulmonary dysplasia development, no serious pulmonary problems were observed. However, the child's development is uncertain. This situation raises important ethical issues concerning saving the lives of infants at the limit of viability.

Application of array comparative genomic hybridization in 102 patients with epilepsy and additional neurodevelopmental disorders.

Copy-number variants (CNVs) collectively represent an important cause of neurodevelopmental disorders such as developmental delay (DD)/intellectual disability (ID), autism, and epilepsy. In contrast to DD/ID, for which the application of microarray techniques enables detection of pathogenic CNVs in -10-20% of patients, there are only few studies of the role of CNVs in epilepsy and genetic etiology in the vast majority of cases remains unknown. We have applied whole-genome exon-targeted oligonucleotide array comparative genomic hybridization (array CGH) to a cohort of 102 patients with various types of epilepsy with or without additional neurodevelopmental abnormalities. Chromosomal microarray analysis revealed 24 non-polymorphic CNVs in 23 patients, among which 10 CNVs are known to be clinically relevant. Two rare deletions in 2q24.1q24.3, including KCNJ3 and 9q21.13 are novel pathogenic genetic loci and 12 CNVs are of unknown clinical significance. Our results further support the notion that rare CNVs can cause different types of epilepsy, emphasize the efficiency of detecting novel candidate genes by whole-genome array CGH, and suggest that the clinical application of array CGH should be extended to patients with unexplained epilepsies.

Decoding working memory-related information from repeated psychophysiological EEG experiments using convolutional and contrastive neural networks.

Objective.Extracting reliable information from electroencephalogram (EEG) is difficult because the low signal-to-noise ratio and significant intersubject variability seriously hinder statistical analyses. However, recent advances in explainable machine learning open a new strategy to address this problem.Approach.The current study evaluates this approach using results from the classification and decoding of electrical brain activity associated with information retention. We designed four neural network models differing in architecture, training strategies, and input representation to classify single experimental trials of a working memory task.Main results.Our best models achieved an accuracy (ACC) of 65.29 ± 0.76 and Matthews correlation coefficient of 0.288 ± 0.018, outperforming the reference model trained on the same data. The highest correlation between classification score and behavioral performance was 0.36 (p= 0.0007). Using analysis of input perturbation, we estimated the importance of EEG channels and frequency bands in the task at hand. The set of essential features identified for each network varies. We identified a subset of features common to all models that identified brain regions and frequency bands consistent with current neurophysiological knowledge of the processes critical to attention and working memory. Finally, we proposed sanity checks to examine further the robustness of each model's set of features.Significance.Our results indicate that explainable deep learning is a powerful tool for decoding information from EEG signals. It is crucial to train and analyze a range of models to identify stable and reliable features. Our results highlight the need for explainable modeling as the model with the highest ACC appeared to use residual artifactual activity.

Early-onset seizures due to mosaic exonic deletions of CDKL5 in a male and two females.

PURPOSE: Mutations in the CDKL5 gene have been associated with an X-linked dominant early infantile epileptic encephalopathy-2. The clinical presentation is usually of severe encephalopathy with refractory seizures and Rett syndrome (RTT)-like phenotype. We attempted to assess the role of mosaic intragenic copy number variation in CDKL5. METHODS: We have used comparative genomic hybridization with a custom-designed clinical oligonucleotide array targeting exons of selected disease and candidate genes, including CDKL5. RESULTS: We have identified mosaic exonic deletions of CDKL5 in one male and two females with developmental delay and medically intractable seizures. These three mosaic changes represent 60% of all deletions detected in 12,000 patients analyzed by array comparative genomic hybridization and involving the exonic portion of CDKL5. CONCLUSION: We report the first case of an exonic deletion of CDKL5 in a male and emphasize the importance of underappreciated mosaic exonic copy number variation in patients with early-onset seizures and RTT-like features of both genders.

[Clinical usefulness of serial EEG examinations in the diagnostic of hereditary epileptic encephalopathies case of severe epileptic encephalopathy type 2]. / Przydatnosc kliniczna seryjnych badan EEG w diagnostyce genetycznie uwarunkowanych encefalopatii padaczkowych na przykladzie przypadku ciezkiej encefalopatii padaczkowej typu 2.

BACKGROUND: Molecular studies allow to study background of many epilepsy types but qualification for genotyping is not easy, especially in initial stages of severe encephalopathy in newborns and infants. One of them is a type 2 severe epileptic encephalopathy (EIEE2), caused by dominant mutation in CDKL5 gene (Xp22.3). In this type of encephalopathy appearing mainly in females, treatment resistant epileptic seizures occur in the first two months of life, they are polymorphic-generalized or focal. Psychomotor development is significantly impaired and in course of time phenotype shows similarities to an Angelman syndrome or an atypical severe form of Rett syndrome. Correlation between clinical status and repeating EEG might be a diagnostic indicator for looking for CDKL5 gene mutation. AIM: We report a case of 3.5 years old girl with refractory epilepsy and dysmorphia like in Angelman syndrome. Mutation in CDKL5 gene was supposed during clinical observation and EEG examination and finally was confirmed. CASE REPORT: Family history and fetal & perinatal history were negative. The patient suffered from treatment resistant polymorphic epileptic seizures appearing from 3 months of life. Psychomotor impairment, significant flaccidity and microcephaly were observed from early infant period. Additionally, pale complexion, always opened mouth, protruding tongue, prognathia, wide-spaced teeth, frequent laughter/smiling were seen. Brain MRI (including MRS) was normal. Repeating EEG showed evolution from normal during infantile spasms to multifocal discharges and loss of sleep spindles. Metabolic disorders, Angelman and Rett syndromes were excluded. Finally, mutation in CDKL5 gene was confirmed at the age of 2.5 into genetic counseling. CONCLUSIONS: There is a need to correlate phenotype features and sequential EEG and epileptic seizures evolution to determine indication for genotyping. Genetic testing looking for CDKL5 mutation is indicated in each female child with impaired psychomotor development, refractory epilepsy with early onset polymorphic seizures and clinical & EEG phenotype of atypical Rett/Angelman syndrome.

Neurosurgical treatment of drug-resistant epilepsy on the basis of a fusion of MRI and SPECT images - case report.

BACKGROUND: Epilepsy concerns at least 0.5% of population and in most of the cases (approx. 70%) can be treated pharmacologically, which helps to prevent seizures. In all other patients, such a treatment does not produce the desired results. Their condition may require neurosurgical management. The aim of this work was to fuse anatomical MRI images and functional SPECT images in patients with drug-resistant epilepsy, without structural changes on MRI or with changes so severe that it would be impossible to establish which ones are responsible for seizures. The authors presented a case of a child subjected to a neurosurgical procedure carried out on the basis of the fused MRI and SPECT images. CASE REPORT: A seven-year-old boy with an extensive defect of the right hemisphere (cortical dysplasia with multiple balloon-like cells) operated on three times due to a history of treatment-resistant seizures present since the age of one. A subsequent MRI examination was performed with magnetic field intensity of 1.5 T, within a routine epilepsy protocol applying volumetric thin-slice T1-weighted images. Next, in the interictal period, a SPECT examination was performed with the use of the (99m)Tc-labelled ethyl cysteinate dimer ((99m)TcECD). For fusion and postprocessing, the following software was used: PMOD (Biomedical Image Quantification PMOD Technologies) with PFUS (Flexible Image Matching and Fusion Tool) and a program for a quantitative analysis of counts in the region of interest, so called VOI Constructor (Volume of Interest Constructor). On the basis of the fusion of images, the boy was subjected to the next operation procedure. The remaining fragments of the right frontal and parietal lobe adjacent to the occipital lobe were removed. Seizure remission was obtained and it was already 31 months long when we were writing this article. CONCLUSIONS: Owing to this multi-stage procedure, it was possible to avoid a total anatomical and functional hemispherectomy. This allowed for a resection limited to regions indicated by integrated imaging. Removal of cortical areas including lesions was advantageous in this presented case, as it allowed for saving active regions of the brain.

Intrathecal Infusion of Autologous Adipose-Derived Regenerative Cells in Autoimmune Refractory Epilepsy: Evaluation of Safety and Efficacy.

Objective/Purpose. Evaluation of efficacy and safety of autologous adipose-derived regenerative cells (ADRCs) treatment in autoimmune refractory epilepsy. Patients. Six patients with proven or probable autoimmune refractory epilepsy (2 with Rasmussen encephalitis, 2 with antineuronal autoantibodies in serum, and 2 with possible FIRES) were included in the project with approval of the Bioethics Committee. METHOD: Intrathecal injection of autologous ADRC acquired through liposuction followed by enzymatic isolation was performed. The procedure was repeated 3 times every 3 months with each patient. Neurological status, brain MRI, cognitive function, and antiepileptic effect were monitored during 12 months. RESULTS: Immediately after the procedure, all patients were in good condition. In some cases, transient mildly elevated body temperature, pain in regions of liposuction, and slight increasing number of seizures during 24 hours were observed. During the next months, some improvements in school, social functioning, and manual performance were observed in all patients. One patient has been seizure free up to the end of trial. In other patients, frequency of seizures was different: from reduced number to the lack of improvement (3-year follow-up). CONCLUSION: Autologous ADRC therapy may emerge as a promising option for some patients with autoimmune refractory epilepsy. Based on our trial and other clinical data, the therapy appears to be safe and feasible. Antiepileptic efficacy proved to be various; however, some abilities improved in all children. No signs of psychomotor regression were observed during the first year following the treatment.

Comprehensive genomic analysis of patients with disorders of cerebral cortical development.

Malformations of cortical development (MCDs) manifest with structural brain anomalies that lead to neurologic sequelae, including epilepsy, cerebral palsy, developmental delay, and intellectual disability. To investigate the underlying genetic architecture of patients with disorders of cerebral cortical development, a cohort of 54 patients demonstrating neuroradiologic signs of MCDs was investigated. Individual genomes were interrogated for single-nucleotide variants (SNV) and copy number variants (CNV) with whole-exome sequencing and chromosomal microarray studies. Variation affecting known MCDs-associated genes was found in 16/54 cases, including 11 patients with SNV, 2 patients with CNV, and 3 patients with both CNV and SNV, at distinct loci. Diagnostic pathogenic SNV and potentially damaging variants of unknown significance (VUS) were identified in two groups of seven individuals each. We demonstrated that de novo variants are important among patients with MCDs as they were identified in 10/16 individuals with a molecular diagnosis. Three patients showed changes in known MCDs genes  and a clinical phenotype beyond the usual characteristics observed, i.e., phenotypic expansion, for a particular known disease gene clinical entity. We also discovered 2 likely candidate genes, CDH4, and ASTN1, with human and animal studies supporting their roles in brain development, and 5 potential candidate genes. Our findings emphasize genetic heterogeneity of MCDs disorders and postulate potential novel candidate genes involved in cerebral cortical development.

Magnetic resonance imaging of neonates in the magnetic resonance compatible incubator.

INTRODUCTION: The authors present the first experience in neonatal magnetic resonance imaging (MRI) examinations using an MR compatible incubator (INC) at the Institute of Mother and Child. MATERIAL AND METHODS: Forty-nine examinations of 47 newborns (20 girls, 27 boys) were performed using the GE Signa HDxt 1.5T system and INC Nomag IC 1.5. Demographic data, anesthetic methods and MRI findings in the INC in comparison with previously performed imaging were analyzed. RESULTS: Thirty-two neonates were prematurely born (68.1%) at gestational age 23-37 weeks, mean: 29.9 weeks. They were examined at 26 weeks postmenstrual age to 1 month corrected age, mean: 37.5 weeks. Body weight of newborns on the study day was 600-4300 g, mean: 2724 g. Seventeen (34.7%) children were examined in physiological sleep, 32 (65.3%) anesthetized. In none of them did anesthesiological complications or disease worsening occur. In 43 (91.5%) children brain MRI was performed, in 4 (8.5%) MRI of the spinal cord and canal and of the abdomen/pelvis. In children prenatally examined by MRI, the INC provided new diagnostic information in 5 (83.3%) cases, in neonates studied after birth by ultrasound in 32 (82%). Magnetic resonance imaging in the INC did not entail additional knowledge in 9 (18.7%) cases. CONCLUSIONS: The INC enables MRI in preterm newborns and those with low/extremely low body weight. These studies are necessary to assess the extent of changes in the central nervous system and other organs. Incubator coils, designed specifically for neonates, allow more accurate diagnosis than previously used coils for adults. MRI results allow one to determine prognosis, for more accurate planning of diagnostics, helping to make appropriate therapeutic decisions.

Epilepsy and mental retardation restricted to females: X-linked epileptic infantile encephalopathy of unusual inheritance.

Epilepsy in females with mental retardation (EFMR) is a rare early infantile epileptic encephalopathy (EIEE), phenotypically resembling Dravet syndrome (DS). It is characterised by a variable degree of intellectual deficits and epilepsy. EFMR is caused by heterozygous mutations in the PCDH19 gene (locus Xq22.1) encoding protocadherin-19, a protein that is highly expressed during brain development. The protein is involved in cell adhesion and probably plays an important role in neuronal migration and formation of synaptic connections. EFMR is considered X-linked of variable mutations' penetrance. Mutations in the PCDH19 gene mainly arise de novo, but if inherited, they show a unique pattern of transmission. Females with heterozygous mutations are affected, while hemizygous males are not, regardless of mutation carriage. This singular mode might be explained by cell interference as a pathogenic molecular mechanism leading to neuronal dysfunction. Recently, PCDH19-related EIEE turned out to be more frequent than initially thought, contributing to around 16% of cases (25% in female groups) in the SCN1A-negative DS-like patients. Therefore, the PCDH19 gene is now estimated to be the second, after SCN1A, most clinically relevant gene in epilepsy.