Dissecting genetics of spectrum of epilepsies with eyelid myoclonia by exome sequencing.

OBJECTIVE: Epilepsy with eyelid myoclonia (EEM) spectrum is a generalized form of epilepsy characterized by eyelid myoclonia with or without absences, eye closure-induced seizures with electroencephalographic paroxysms, and photosensitivity. Based on the specific clinical features, age at onset, and familial occurrence, a genetic cause has been postulated. Pathogenic variants in CHD2, SYNGAP1, NEXMIF, RORB, and GABRA1 have been reported in individuals with photosensitivity and eyelid myoclonia, but whether other genes are also involved, or a single gene is uniquely linked with EEM, or its subtypes, is not yet known. We aimed to dissect the genetic etiology of EEM. METHODS: We studied a cohort of 105 individuals by using whole exome sequencing. Individuals were divided into two groups: EEM- (isolated EEM) and EEM+ (EEM accompanied by intellectual disability [ID] or any other neurodevelopmental/psychiatric disorder). RESULTS: We identified nine variants classified as pathogenic/likely pathogenic in the entire cohort (8.57%); among these, eight (five in CHD2, one in NEXMIF, one in SYNGAP1, and one in TRIM8) were found in the EEM+ subcohort (28.57%). Only one variant (IFIH1) was found in the EEM- subcohort (1.29%); however, because the phenotype of the proband did not fit with published data, additional evidence is needed before considering IFIH1 variants and EEM- an established association. Burden analysis did not identify any single burdened gene or gene set. SIGNIFICANCE: Our results suggest that for EEM, as for many other epilepsies, the identification of a genetic cause is more likely with comorbid ID and/or other neurodevelopmental disorders. Pathogenic variants were mostly found in CHD2, and the association of CHD2 with EEM+ can now be considered a reasonable gene-disease association. We provide further evidence to strengthen the association of EEM+ with NEXMIF and SYNGAP1. Possible new associations between EEM+ and TRIM8, and EEM- and IFIH1, are also reported. Although we provide robust evidence for gene variants associated with EEM+, the core genetic etiology of EEM- remains to be elucidated.

Randomized Double-Blind Crossover Study for Evaluating a Probiotic Mixture on Gastrointestinal and Behavioral Symptoms of Autistic Children.

Autism spectrum disorders (ASDs) represent a diagnostic challenge with a still partially uncertain etiology, in which genetic and environmental factors have now been assessed. Among the hypotheses underlying the involvement of biological and environmental factors, the gut-brain axis is of particular interest in autism spectrum disorders. Several studies have highlighted the related incidence of particular gastrointestinal symptoms (GISs) in children suffering from ASDs. Probiotics have shown success in treating several gastrointestinal dysbiotic disorders; therefore, it is plausible to investigate whether they can alleviate behavioral symptoms as well. On these bases, a randomized double-blind crossover study with a placebo was conducted, evaluating the effects of a mixture of probiotics in a group of 61 subjects aged between 24 months and 16 years old with a diagnosis of ASD. Behavioral evaluation was performed through the administration of a questionnaire including a Parenting Stress Index (PSI) test and the Vineland Adaptive Behavior Scale (VABS). The Psycho-Educational Profile and the Autism Spectrum Rating Scale (ASRS) were also evaluated. Microbial composition analyses of fecal samples of the two groups was also performed. The study showed significant improvements in GISs, communication skills, maladaptive behaviors, and perceived parental stress level after the administration of probiotics. Microbiome alpha diversity was comparable between treatment arms and no significant differences were found, although beta diversity results were significantly different in the treatment group between T0 and T1 time points. Streptococcus thermophilus, Bifidobacterium longum, Limosilactobacillus fermentum, and Ligilactobacillus salivarius species were identified as some of the most discriminant taxa positively associated with T1 samples. This preliminary study corroborates the relationship between intestinal microbiota and ASD recently described in the literature.

EEG at onset and MRI predict long-term clinical outcome in Aicardi syndrome.

OBJECTIVE: Descriptions of electroencephalographic (EEG) patterns in Aicardi syndrome (AIC) have to date referred to small cohorts of up to six cases and indicated severe derangement of electrical activity in all cases. The present study was conducted to describe the long-term EEG evolution in a larger AIC cohort, followed for up to 23 years, and identify possible early predictors of the clinical and EEG outcomes. METHODS: In a retrospective study, two experienced clinical neurophysiologists systematically reviewed all EEG traces recorded in 12 AIC cases throughout their follow-up, from epilepsy onset to the present. Clinical outcome was assessed with standardized clinical outcome scales. RESULTS: Analysis of the data revealed two distinct AIC phenotypes. In addition to the "classical severe phenotype" already described in the literature, we identified a new "mild phenotype". The two phenotypes show completely different EEG features at onset of epilepsy and during its evolution, which correspond to different clinical outcomes. CONCLUSIONS: Data from our long-term EEG and clinical-neuroradiological study allowed us to describe two different phenotypes of AIC, with different imaging severity and, in particular, different EEG at onset, which tend to remain constant over time. SIGNIFICANCE: Together, these findings might help to predict long-term clinical outcomes.

The Usefulness of a Targeted Next Generation Sequencing Gene Panel in Providing Molecular Diagnosis to Patients With a Broad Spectrum of Neurodevelopmental Disorders.

Background: Neurodevelopmental disorders comprise a clinically and genetically heterogeneous group of conditions that affect 2%-5% of children and represents a public health challenge due to complexity of the etiology. Only few patients with unexplained syndromic and non-syndromic NDDs receive a diagnosis through first-tier genetic tests as array-CGH and the search for FMR1 CGG expansion. The aim of this study was to evaluate the clinical performance of a targeted next-generation sequencing (NGS) gene panel as a second-tier test in a group of undiagnosed patients with NDDs. Method: A 221-gene next-generation sequencing custom panel was designed and used to analyze a cohort of 338 patients with a broad spectrum of NDDs (202 males and 136 females) including Intellectual Disability (ID), Autism Spectrum Disorders (ASD), Epilepsy, language and motor disorders. Results: A molecular diagnosis was established in 71 patients (21%) and a de novo origin was present in 38 (64.4%) of the available trios. The diagnostic yield was significantly higher in females than in males (29.4% vs. 15.3%; p = 0.0019) in particular in ASD (36.8% vs. 7.6%; p = 0.0026) and Epilepsy (38.9% vs. 14.4% p = 0.001). The most involved genes were SLC2A1, SCN1A, ANKRD11, ATP1A2, CACNA1A, FOXP1, and GNAS altered in more than two patients and accounting for the 19.7% of the diagnosis. Conclusion: Our findings showed that this NGS panel represents a powerful and affordable clinical tool, significantly increasing the diagnostic yield in patients with different form of NDDs in a cost- and time-effective manner without the need of large investments in data storage and bioinformatic analysis.

Impact and management of drooling in children with neurological disorders: an Italian Delphi consensus.

BACKGROUND: The rate of chronic drooling in children older than 4 years is 0.5%, but it rises to 60% in those with neurological disorders. Physical and psychosocial consequences lead to a reduction in the quality of Life (QoL) of affected patients; however, the problem remains under-recognized and under-treated. We conducted an Italian consensus through a modified Delphi survey to discuss the current treatment paradigm of drooling in pediatric patients with neurological disorders. METHODS: After reviewing the literature, a board of 10 experts defined some statements to be administered to a multidisciplinary panel through an online encrypted platform. The answers to the questions were based on a 1-5 Likert scale (1 = strongly disagree; 5 = strongly agree). The scores were grouped into 1-2 (disagreement) and 4-5 (agreement), while 3 was discarded. The consensus was reached when the sum of the disagreement or agreement was ≥75%. RESULTS: Fifteen statements covered three main topics, namely clinical manifestations and QoL, quantification of drooling, and treatment strategies. All statements reached consensus (≥75% agreement). The 55 Italian experts agreed that drooling should be assessed in all children with complex needs, having a major impact on the QoL. Attention should be paid to investigating posterior hypersalivation, which is often neglected but may lead to important clinical consequences. Given that the severity of drooling fluctuates over time, its management should be guided by the patients' current needs. Furthermore, the relative lack of validated and universal scales for drooling quantification limits the evaluation of the response to treatment. Finally, the shared therapeutic paradigm is progressive, with conservative treatments preceding the pharmacological ones and reserving surgery only for selected cases. CONCLUSION: This study demonstrates the pivotal importance of a multidisciplinary approach to the management of drooling. National experts agree that progressive treatment can reduce the incidence of complications, improve the QoL of patients and caregivers, and save healthcare resources. Finally, this study highlights how the therapeutic strategy should be reconsidered over time according to the available drugs on the market, the progression of symptoms, and the patients' needs.

A nationwide study on Sydenham's chorea: Clinical features, treatment and prognostic factors.

OBJECTIVES: Sydenham's Chorea (SC) is a neuropsychiatric disorder and a major manifestation of acute rheumatic fever. The erroneous assumption that SC is a benign and self-limiting disease, has led to a lack of high-quality scientific evidence of the therapeutical and prognostic features of SC. STUDY DESIGN: We retrospectively analyzed the medical records of patients <18-years old with SC in 17 Italian pediatric centers. Recorded data included clinical, instrumental and laboratory parameters. Prognostic risk factors including treatment regimens were assessed with univariate and multivariate sub-analysis. RESULTS: We included 171 patients with SC. 66% had generalized chorea, and 34% hemichorea. 81% had carditis (subclinical in 65%). Additional neurological symptoms were reported in 60% of the patients, mainly dysarthria and dysgraphia. 51% had neuropsychiatric symptoms at onset, which persisted after 12 months in 10%. Among psychiatric manifestations, the most common was anxiety disorder/depression (77%). Neurological remission was reached by 93% of the patients at 6 months; 9% relapsed. Patients were treated as follows: 11% penicillin alone, 37% immunomodulatory therapy, 16% symptomatic drugs (i.e. anti-seizure medication, dopamine antagonists) and 37% both symptomatic and immunomodulatory treatment. Neurological outcome did not differ between groups. Patients receiving symptomatic drugs had a higher risk of relapse on multivariate analysis (p = 0.045). CONCLUSIONS: Treatment of SC was largely heterogeneous. Based on our results, immunomodulatory therapy did not show higher efficacy at medium term, although it was associated to a slightly lower risk of relapse compared to symptomatic therapy. Longitudinal studies are needed to assess specific risk factors and best treatment options.

Video game-induced reflex seizures via a smartphone.

Reflex seizures are consistently evoked by a specific afferent stimulus or by patient activity. Patients experiencing reflex seizures when playing a game on a mobile phone are rarely reported. We describe a boy with reflex seizures after prolonged exposure to the game, Cut the rope, on his mobile phone. The video-EEG documented electroclinical events characterized by distal myoclonic jerks of the upper limbs, in combination with irregular, diffuse spike-and-wave and polyspike-and-wave discharges on EEG, followed by a tonic-clonic seizure. Playing video games on mobile phones may potentially induce reflex seizures, similar to other commonly used platforms such as docking stations connected to video screens.

The epileptology of Aicardi-Goutières syndrome: electro-clinical-radiological findings.

OBJECTIVE: Although epileptic seizures occur in approximately a quarter of patients with Aicardi-Goutières syndrome (AGS), their phenotypic and electrophysiological characterization remains elusive. The aim of our study was to characterize epilepsy phenotypes and electroencephalographic (EEG) patterns in AGS and look for possible correlations with clinical, genetic and neuroradiological features. METHODS: We selected patients with an established AGS diagnosis followed at three Italian reference centers. Medical records, EEGs and MRI/CT findings were reviewed. EEGs were independently and blindly reviewed by three board-certified pediatric epileptologists. Chi square and Fisher's exact tests were used to test associations between epilepsy and EEG feature categories and clinical, radiological and genetic variables. RESULTS: Twenty-seven patients were enrolled. We reviewed 63 EEGs and at least one brain MRI scan per patient. Epilepsy, mainly in the form of epileptic spasms and focal seizures, was present in 37 % of the cohort; mean age at epilepsy onset was 9.5 months (range 1-36). The presence of epilepsy was associated with calcification severity (p = 0.016) and startle reactions (p = 0.05). Organization of EEG electrical activity appeared to be disrupted or markedly disrupted in 73 % of cases. Severe EEG disorganization correlated with microcephaly (p < 0.001) and highly abnormal MRI T2-weighted signal intensity in white matter (p = 0.022). Physiological organization of the EEG was found to be better preserved during sleep (87 %) than wakefulness (38 %). Focal slow activity was recorded in more than one third of cases. Fast activity, either diffuse or with frontal location, was more frequent in the awake state (78 %) than in sleep (50 %). Interictal epileptiform discharges (IEDs) were present in 33 % of awake and 45 % of sleep recordings. IEDs during sleep were associated with a higher risk of a epileptic seizures (p = 0.008). SIGNIFICANCE: The hallmarks of EEG recordings in AGS were found to be: disruption of electrical organization, the presence of focal slow and fast activity, and the presence of IEDs, both in patients with and in those without epilepsy. The associations between epilepsy and calcification and between EEG pattern and the finding of a highly abnormal white matter T2 signal intensity suggest a common anatomical correlate. However, the complex anatomical-electroclinical basis of AGS-related epilepsy still requires further elucidation.

Anakinra usage in febrile infection related epilepsy syndrome: an international cohort.

Febrile-infection related epilepsy syndrome (FIRES) is a devastating neurological condition characterized by a febrile illness preceding new onset refractory status epilepticus (NORSE). Increasing evidence suggests innate immune dysfunction as a potential pathological mechanism. We report an international retrospective cohort of 25 children treated with anakinra, a recombinant interleukin-1 receptor antagonist, as an immunomodulator for FIRES. Anakinra was potentially safe with only one child discontinuing therapy due to infection. Earlier anakinra initiation was associated with shorter duration of mechanical ventilation, ICU and hospital length of stay. Our retrospective data lay the groundwork for prospective consensus-driven cohort studies of anakinra in FIRES.

STXBP1 encephalopathy: A neurodevelopmental disorder including epilepsy.

OBJECTIVE: To give a comprehensive overview of the phenotypic and genetic spectrum of STXBP1 encephalopathy (STXBP1-E) by systematically reviewing newly diagnosed and previously reported patients. METHODS: We recruited newly diagnosed patients with STXBP1 mutations through an international network of clinicians and geneticists. Furthermore, we performed a systematic literature search to review the phenotypes of all previously reported patients. RESULTS: We describe the phenotypic features of 147 patients with STXBP1-E including 45 previously unreported patients with 33 novel STXBP1 mutations. All patients have intellectual disability (ID), which is mostly severe to profound (88%). Ninety-five percent of patients have epilepsy. While one-third of patients presented with Ohtahara syndrome (21%) or West syndrome (9.5%), the majority has a nonsyndromic early-onset epilepsy and encephalopathy (53%) with epileptic spasms or tonic seizures as main seizure type. We found no correlation between severity of seizures and severity of ID or between mutation type and seizure characteristics or cognitive outcome. Neurologic comorbidities including autistic features and movement disorders are frequent. We also report 2 previously unreported adult patients with prominent extrapyramidal features. CONCLUSION: De novo STXBP1 mutations are among the most frequent causes of epilepsy and encephalopathy. Most patients have severe to profound ID with little correlation among seizure onset, seizure severity, and the degree of ID. Accordingly, we hypothesize that seizure severity and ID present 2 independent dimensions of the STXBP1-E phenotype. STXBP1-E may be conceptualized as a complex neurodevelopmental disorder rather than a primary epileptic encephalopathy.

Dramatic effect of levetiracetam in early-onset epileptic encephalopathy due to STXBP1 mutation.

BACKGROUND: Syntaxin Binding Protein 1 (STXBP1) mutations determine a central neurotransmission dysfunction through impairment of the synaptic vesicle release, thus causing a spectrum of phenotypes varying from syndromic and non-syndromic epilepsy to intellectual disability of variable degree. Among the antiepileptic drugs, levetiracetam has a unique mechanism of action binding SV2A, a glycoprotein of the synaptic vesicle release machinery. PATIENT DESCRIPTION: We report a 1-month-old boy manifesting an epileptic encephalopathy with clonic seizures refractory to phenobarbital, pyridoxine and phenytoin that presented a dramatic response to levetiracetam with full epilepsy control and EEG normalization. Genetic analysis identified a novel de novo heterozygous mutation (c.[922A>T]p.[Lys308(∗)]) in the STXBP1 gene that severely affects the protein. CONCLUSIONS: The observation of a dramatic efficacy of levetiracetam in a case of STXBP1 epileptic encephalopathy refractory to other antiepileptic drugs and considerations regarding the specific mechanism of action of levetiracetam modulating the same system affected by STXBP1 mutations support the hypothesis that this drug may be able to reverse specifically the disease epileptogenic abnormalities. Further clinical observations and laboratory studies are needed to confirm this hypothesis and eventually lead to consider levetiracetam as the first choice treatment of patients with suspected or confirmed STXBP1-related epilepsies.

Mild Lafora disease: clinical, neurophysiologic, and genetic findings.

We report clinical, neurophysiologic, and genetic features of an Italian series of patients with Lafora disease (LD) to identify distinguishing features of those with a slowly progressive course. Twenty-three patients with LD (17 female; 6 male) were recruited. Mean age (± SD) at the disease onset was 14.5 ± 3.9 years and mean follow-up duration was 13.2 ± 8.0 years. NHLRC1 mutations were detected in 18 patients; EPM2A mutations were identified in 5. Patients who maintained >10 years gait autonomy were labeled as "mild" and were compared with the remaining LD patients with a typical course. Six of 23 patients were mild and presented significantly delay in the age at onset, lower neurologic disability score at 4 years after the onset, less severe seizure phenotype, lower probability of showing both photoparoxysmal response on electroencephalography (EEG) and giant somatosensory evoked potentials, as compared to patients with typical LD. However, in both mild and typical LD patients, EEG showed disorganization of background activity and frequent epileptiform abnormalities. Mild LD patients had NHLRC1 mutations and five of six carried homozygous or compound heterozygous D146N mutation. This mutation was found in none of the patients with typical LD. The occurrence of specific NHLRC1 mutations in patients with mild LD should be taken into account in clinical practice for appropriate management and counseling.

Does the co-occurrence of FGFR3 gene mutation in hypochondroplasia, medial temporal lobe dysgenesis, and focal epilepsy suggest a syndrome?

BACKGROUND: Hypochondroplasia is a rare skeletal dysplasia characterized by disproportionately short stature, lumbar lordosis, and limited extension of the elbow caused by mutations in the fibroblast growth factor receptor 3 (FGFR3) gene that plays a role in controlling nervous system development. Hypochondroplasia with FGFR3 mutation associated with bilateral medial temporal lobe anomalies and focal epilepsy was previously reported in several patients. PATIENT: We report clinical, electroclinical, and neuroradiological findings of one patient affected by hypochondroplasia. RESULTS: Clinical diagnosis was confirmed by molecular analysis of the FGFR3 gene, which showed a N540 K mutation. The patient had normal psychomotor development and showed early-onset focal seizures with left temporal localization on interictal and ictal electroencephalograph. The seizures were well controlled, and the patient has been seizure-free since infancy. Magnetic resonance imaging showed abnormal anteriorly posteriorly infolding in the hippocampus and abnormally oriented parahippocampus sulci, and additional cortical rim dysplasia with gray-white matter junction blurring in the hippocampus. CONCLUSIONS: The present case of hypochondroplasia and FGFR3 mutation in Asn540Lys associated with characteristic abnormalities involving bilaterally medial temporal lobe structures, probable hippocampal cortex focal dysplasia, and early onset of focal epilepsy underscores the possibility of a rare syndrome.

Vaccination and occurrence of seizures in SCN1A mutation-positive patients: a multicenter Italian study.

BACKGROUND: The relation between epileptic seizures and vaccinations is sometimes debated. In the present work, the impact of vaccination on seizure onset and clinical outcome of SCN1A mutation-positive patients is addressed. METHODS: Seventy-two patients diagnosed with Dravet syndrome or generalized epilepsy with febrile seizure plus, carrying SCN1A mutations or not, were included. Details on vaccination type, temporal relationship between vaccination and seizure occurrence, seizure type at onset and during development, cognitive functioning, and vaccination completion was obtained by reviewing clinical records. Patients were divided into two groups based on the temporal window between vaccination and seizure onset (proximate group: <48 hours; distant group: >48 hours). RESULTS: Vaccination-related seizures occurred in 25% of patients with SCN1A mutation and 18% of patients without the mutation (no significant difference). The proximate group showed an earlier age at seizure onset and a higher frequency of status epilepticus during development than did the distant group. No other significant differences were found. Subsequent vaccinations did not significantly alter the evolution of the disease. CONCLUSIONS: Results from this relatively small series provide evidence that vaccinations do not significantly affect clinical and cognitive evolution of Dravet syndrome and generalized epilepsy with febrile seizure plus patients even if they carry SCN1A mutations.

Lacosamide in pediatric and adult patients: comparison of efficacy and safety.

PURPOSE: This multicenter, prospective study investigates the efficacy and safety of lacosamide adjunctive therapy in pediatric and adult patients with uncontrolled epilepsy. METHOD: This study was carried out between September 2010 and December 2011 at 16 Italian and 1 German neurologic centers. Lacosamide was added to the baseline therapy at a starting dose of 1 mg/kg/day in patients aged <16 years (group A) and 100 mg daily in subjects aged 16 and older (group B), and titrated to the target dose, ranging from 3 to 12 mg/kg/day or from 100 to 600 mg daily, respectively. After completing the titration period, patients entered a 12-month maintenance period and they were followed up at 3, 6 and 12 months. The primary assessment of efficacy was based on the change from baseline in seizure frequency per 28 days and was evaluated at 3, 6 and 12 months as follows: number and proportion of 100% responders, 50% responders, non-responders and worsening patients. Safety evaluation was also performed at 3, 6 and 12 months. RESULTS: A total of 118 patients (59 group A, 59 group B) with uncontrolled generalized and focal epilepsy were enrolled. Patient mean±SD age was 15.9±6.80 years and the age range was 4-38 years. At 3-month evaluation, of 118 treated patients 56 subjects (47.4% group A; 47.4% group B; p=0.8537) experienced at least a 50% reduction in seizure frequency. At 6 and 12-month follow-up, the 50% responders were 57 (52.5% group A; 44.1% group B; p=0.4612) and 51 (47.4% group A; 39% group B; p=0.4573), respectively. Thirty-five subjects (30.5% group A; 28.8% group B; p=1) experienced side effects during the treatment period. The most common adverse events were dyspepsia for group A and dizziness for group B. CONCLUSION: Lacosamide may be a useful and safe pharmacological treatment option for both pediatric and adult patients with uncontrolled seizures.

Genetic testing in benign familial epilepsies of the first year of life: clinical and diagnostic significance.

PURPOSE: To dissect the genetics of benign familial epilepsies of the first year of life and to assess the extent of the genetic overlap between benign familial neonatal seizures (BFNS), benign familial neonatal-infantile seizures (BFNIS), and benign familial infantile seizures (BFIS). METHODS: Families with at least two first-degree relatives affected by focal seizures starting within the first year of life and normal development before seizure onset were included. Families were classified as BFNS when all family members experienced neonatal seizures, BFNIS when the onset of seizures in family members was between 1 and 4 months of age or showed both neonatal and infantile seizures, and BFIS when the onset of seizures was after 4 months of age in all family members. SCN2A, KCNQ2, KCNQ3, PPRT2 point mutations were analyzed by direct sequencing of amplified genomic DNA. Genomic deletions involving KCNQ2 and KCNQ3 were analyzed by multiple-dependent probe amplification method. KEY FINDINGS: A total of 46 families including 165 affected members were collected. Eight families were classified as BFNS, 9 as BFNIS, and 29 as BFIS. Genetic analysis led to the identification of 41 mutations, 14 affecting KCNQ2, 1 affecting KCNQ3, 5 affecting SCN2A, and 21 affecting PRRT2. The detection rate of mutations in the entire cohort was 89%. In BFNS, mutations specifically involve KCNQ2. In BFNIS two genes are involved (KCNQ2, six families; SCN2A, two families). BFIS families are the most genetically heterogeneous, with all four genes involved, although about 70% of them carry a PRRT2 mutation. SIGNIFICANCE: Our data highlight the important role of KCNQ2 in the entire spectrum of disorders, although progressively decreasing as the age of onset advances. The occurrence of afebrile seizures during follow-up is associated with KCNQ2 mutations and may represent a predictive factor. In addition, we showed that KCNQ3 mutations might be also involved in families with infantile seizures. Taken together our data indicate an important role of K-channel genes beyond the typical neonatal epilepsies. The identification of a novel SCN2A mutation in a family with infantile seizures with onset between 6 and 8 months provides further confirmation that this gene is not specifically associated with BFNIS and is also involved in families with a delayed age of onset. Our data indicate that PRRT2 mutations are clustered in families with BFIS. Paroxysmal kinesigenic dyskinesia emerges as a distinctive feature of PRRT2 families, although uncommon in our series. We showed that the age of onset of seizures is significantly correlated with underlying genetics, as about 90% of the typical BFNS families are linked to KCNQ2 compared to only 3% of the BFIS families, for which PRRT2 represents the major gene.

Electroclinical presentation and genotype-phenotype relationships in patients with Unverricht-Lundborg disease carrying compound heterozygous CSTB point and indel mutations.

PURPOSE: Unverricht-Lundborg disease (EPM1A) is frequently due to an unstable expansion of a dodecamer repeat in the CSTB gene, whereas other types of mutations are rare. EPM1A due to homozygous expansion has a rather stereotyped presentation with prominent action myoclonus. We describe eight patients with five different compound heterozygous CSTB point or indel mutations in order to highlight their particular phenotypical presentations and evaluate their genotype-phenotype relationships. METHODS: We screened CSTB mutations by means of Southern blotting and the sequencing of the genomic DNA of each proband. CSTB messenger RNA (mRNA) aberrations were characterized by sequencing the complementary DNA (cDNA) of lymphoblastoid cells, and assessing the protein concentrations in the lymphoblasts. The patient evaluations included the use of a simplified myoclonus severity rating scale, multiple neurophysiologic tests, and electroencephalography (EEG)-polygraphic recordings. To highlight the particular clinical features and disease time-course in compound heterozygous patients, we compared some of their characteristics with those observed in a series of 40 patients carrying the common homozygous expansion mutation observed at the C. Besta Foundation, Milan, Italy. KEY FINDINGS: The eight compound heterozygous patients belong to six EPM1A families (out of 52; 11.5%) diagnosed at the Laboratory of Genetics of the Galliera Hospitals in Genoa, Italy. They segregated five different heterozygous point or indel mutations in association with the common dodecamer expansion. Four patients from three families had previously reported CSTB mutations (c.67-1G>C and c.168+1_18del); one had a novel nonsense mutation at the first exon (c.133C>T) leading to a premature stop codon predicting a short peptide; the other three patients from two families had a complex novel indel mutation involving the donor splice site of intron 2 (c.168+2_169+21delinsAA) and leading to an aberrant transcript with a partially retained intron. The protein dose (cystatin B/ß-actin) in our heterozygous patients was 0.24 ± 0.02, which is not different from that assessed in patients bearing the homozygous dodecamer expansion. The compound heterozygous patients had a significantly earlier disease onset (7.4 ± 1.7 years) than the homozygous patients, and their disease presentations included frequent myoclonic seizures and absences, often occurring in clusters throughout the course of the disease. The seizures were resistant to the pharmacologic treatments that usually lead to complete seizure control in homozygous patients. EEG-polygraphy allowed repeated seizures to be recorded. Action myoclonus progressively worsened and all of the heterozygous patients older than 30 years were in wheelchairs. Most of the patients showed moderate to severe cognitive impairment, and six had psychiatric symptoms. SIGNIFICANCE: EPM1A due to compound heterozygous CSTB mutations presents with variable but often markedly severe and particular phenotypes. Most of our patients presented with the electroclinical features of severe epilepsy, which is unexpected in homozygous patients, and showed frequent seizures resistant to pharmacologic treatment. The presence of variable phenotypes (even in siblings) suggests interactions with other genetic factors influencing the final disease presentation.

Homozygous c.649dupC mutation in PRRT2 worsens the BFIS/PKD phenotype with mental retardation, episodic ataxia, and absences.

Heterozygous mutations of PRRT2, which encodes proline-rich transmembrane protein 2, are associated with heterogeneous phenotypes including benign familial infantile seizures (BFIS), or familial paroxysmal kinesigenic dystonia (PKD). We report a consanguineous Italian family with BFIS/PKD phenotype that contained 14 living members with 6 affected individuals (four men, ranging in age from 6-44 years). We identified the reported c.649dupC (p.Arg217ProfsX8) mutation of PRRT2 gene that cosegregated with the disease and was not observed in 100 controls of matched ancestry. Four patients with BFIS phenotype were heterozygous for this mutation, including the consanguineous parents of the two affected brothers with more severe phenotypes of BFIS/PKD--mental retardation, episodic ataxia, and absences--who were the only individuals to carry a homozygous c.649dupC mutation. This family provides strong evidence that homozygous PRRT2 mutations give rise to more severe clinical disease of mental retardation, episodic ataxia, and absences, and, thus, enlarges the clinical spectrum related to PRRT2 mutations. Moreover, it suggests an additive effect of double dose of the genetic mutation and underscores the complexity of the phenotypic consequences of mutations in this gene.