Rachel Kuperman

De novo mutations in epileptic encephalopathies

Epileptic encephalopathies are a devastating group of severe childhood epilepsy disorders for which the cause is often unknown. Here we report a screen for de novo mutations in patients with two classical epileptic encephalopathies: infantile spasms (n = 149) and Lennox–Gastaut syndrome (n = 115). We sequenced the exomes of 264 probands, and their parents, and confirmed 329 de novo mutations. A likelihood analysis showed a significant excess of de novo mutations in the ∼4,000 genes that are the most intolerant to functional genetic variation in the human population (P = 2.9 × 10−3). Among these are GABRB3, with de novo mutations in four patients, and ALG13, with the same de novo mutation in two patients; both genes show clear statistical evidence of association with epileptic encephalopathy. Given the relevant site-specific mutation rates, the probabilities of these outcomes occurring by chance are P = 4.1 × 10−10 and P = 7.8 × 10−12, respectively. Other genes with de novo mutations in this cohort include CACNA1A, CHD2, FLNA, GABRA1, GRIN1, GRIN2B, HNRNPU, IQSEC2, MTOR and NEDD4L. Finally, we show that the de novo mutations observed are enriched in specific gene sets including genes regulated by the fragile X protein (P < 10−8), as has been reported previously for autism spectrum disorders.

Efficacy and safety of everolimus for subependymal giant cell astrocytomas associated with tuberous sclerosis complex (EXIST-1): a multicentre, randomised, placebo-controlled phase 3 trial

BACKGROUND Tuberous sclerosis complex is a genetic disorder leading to constitutive activation of mammalian target of rapamycin (mTOR) and growth of benign tumours in several organs. In the brain, growth of subependymal giant cell astrocytomas can cause life-threatening symptoms--eg, hydrocephalus, requiring surgery. In an open-label, phase 1/2 study, the mTOR inhibitor everolimus substantially and significantly reduced the volume of subependymal giant cell astrocytomas. We assessed the efficacy and safety of everolimus in patients with subependymal giant cell astrocytomas associated with tuberous sclerosis complex. METHODS In this double-blind, placebo-controlled, phase 3 trial, patients (aged 0-65 years) in 24 centres in Australia, Belgium, Canada, Germany, the UK, Italy, the Netherlands, Poland, Russian Federation, and the USA were randomly assigned, with an interactive internet-response system, in a 2:1 ratio to oral everolimus 4·5 mg/m(2) per day (titrated to achieve blood trough concentrations of 5-15 ng/mL) or placebo. Eligible patients had a definite diagnosis of tuberous sclerosis complex and at least one lesion with a diameter of 1 cm or greater, and either serial growth of a subependymal giant cell astrocytoma, a new lesion of 1 cm or greater, or new or worsening hydrocephalus. The primary endpoint was the proportion of patients with confirmed response--ie, reduction in target volume of 50% or greater relative to baseline in subependymal giant cell astrocytomas. Analysis was by intention to treat. This study is registered with ClinicalTrials.gov, number NCT00789828. FINDINGS 117 patients were randomly assigned to everolimus (n=78) or placebo (n=39). 27 (35%) patients in the everolimus group had at least 50% reduction in the volume of subependymal giant cell astrocytomas versus none in the placebo group (difference 35%, 95% CI 15-52; one-sided exact Cochran-Mantel-Haenszel test, p<0·0001). Adverse events were mostly grade 1 or 2; no patients discontinued treatment because of adverse events. The most common adverse events were mouth ulceration (25 [32%] in the everolimus group vs two [5%] in the placebo group), stomatitis (24 [31%] vs eight [21%]), convulsion (18 [23%] vs ten [26%]), and pyrexia (17 [22%] vs six [15%]). INTERPRETATION These results support the use of everolimus for subependymal giant cell astrocytomas associated with tuberous sclerosis. Additionally, everolimus might represent a disease-modifying treatment for other aspects of tuberous sclerosis. FUNDING Novartis Pharmaceuticals.

Everolimus for subependymal giant cell astrocytoma in patients with tuberous sclerosis complex: 2-year open-label extension of the randomised EXIST-1 study

BACKGROUND In the EXIST-1 trial, initiated on Aug 10, 2009, more than 35% of patients with subependymal giant cell astrocytoma (SEGA) associated with tuberous sclerosis complex had at least 50% reduction in SEGA volume after 9·6 months of treatment with everolimus. In this Article, we report interim data (up to Jan 11, 2013) to support longer-term tolerability and efficacy of everolimus from the continuing 4-year extension phase of EXIST-1. METHODS We assessed data from a prospective, open-label extension of a multicentre, phase 3, randomised, double-blind, placebo-controlled study in patients with tuberous sclerosis complex who had SEGA that was growing and needed treatment. In this extension study, we included all patients who had been assigned everolimus during the double-blind, randomised phase of the trial and those patients who crossed over from the placebo group to receive everolimus during the randomised phase or at the start of the extension phase. All patients received oral everolimus at a starting dose of 4·5 mg/m(2) per day. Everolimus dose was subsequently adjusted subject to tolerability to attain blood trough concentrations of 5-15 ng/mL. An independent central radiology review team assessed SEGA response (at least a 50% reduction from baseline in total volume of all target SEGAs; the primary endpoint) by MRI at 12, 24, and 48 weeks, then every year thereafter in all patients who received at least one dose of everolimus. This study was registered with ClinicalTrials.gov, number NCT00789828. FINDINGS Of the original 117 randomly assigned patients, 111 were given everolimus between Aug 20, 2009, and Jan 11, 2013 (date of data cutoff); we included these patients in our longer-term analysis. Median duration of everolimus exposure was 29·3 months (IQR 19·4-33·8). Median follow-up was 28·3 months (IQR 19·3-33·0). 54 (49%) patients had a response of 50% or greater reduction in SEGA volume (95% CI 39·0-58·3), and duration of response was between 2·1 and 31·1 months (median not reached). SEGA volume was reduced by 50% or more in 39 (37%) of 105 patients at 24 weeks, 48 (46%) of 104 patients at 48 weeks, 36 (47%) of 76 patients at 96 weeks, and 11 (38%) of 29 patients at 144 weeks. Stomatitis (48 [43%] patients) and mouth ulceration (33 [30%] patients) were the most frequent treatment-related adverse events; infections were the most commonly reported treatment-related serious adverse event, occurring in 15 (14%) patients. 35 (32%) patients reported treatment-related grade 3 or 4 adverse events, the most common of which were stomatitis (nine [8%]) and pneumonia (nine [8%]). 18 (16%) patients had treatment-related serious adverse events. Six (5%) patients withdrew because of adverse events. INTERPRETATION These results support the longer-term use of everolimus in patients who have few treatment options and who need continued treatment for tuberous sclerosis complex and its varied manifestations. Reduction or stabilisation of tumour volume with everolimus will hopefully provide long-term clinical benefit in patients with SEGA. FUNDING Novartis Pharmaceuticals.

Dynamic Changes in Phase-Amplitude Coupling Facilitate Spatial Attention Control in Fronto-Parietal Cortex

Electrocorticography reveals how coupling between two frequencies of neuronal oscillation allows the frontal and parietal areas of the cortex to control visual attention from moment to moment in the human brain.

Ultra-rare genetic variation in common epilepsies: a case-control sequencing study

BACKGROUND Despite progress in understanding the genetics of rare epilepsies, the more common epilepsies have proven less amenable to traditional gene-discovery analyses. We aimed to assess the contribution of ultra-rare genetic variation to common epilepsies. METHODS We did a case-control sequencing study with exome sequence data from unrelated individuals clinically evaluated for one of the two most common epilepsy syndromes: familial genetic generalised epilepsy, or familial or sporadic non-acquired focal epilepsy. Individuals of any age were recruited between Nov 26, 2007, and Aug 2, 2013, through the multicentre Epilepsy Phenome/Genome Project and Epi4K collaborations, and samples were sequenced at the Institute for Genomic Medicine (New York, USA) between Feb 6, 2013, and Aug 18, 2015. To identify epilepsy risk signals, we tested all protein-coding genes for an excess of ultra-rare genetic variation among the cases, compared with control samples with no known epilepsy or epilepsy comorbidity sequenced through unrelated studies. FINDINGS We separately compared the sequence data from 640 individuals with familial genetic generalised epilepsy and 525 individuals with familial non-acquired focal epilepsy to the same group of 3877 controls, and found significantly higher rates of ultra-rare deleterious variation in genes established as causative for dominant epilepsy disorders (familial genetic generalised epilepsy: odd ratio [OR] 2·3, 95% CI 1·7-3·2, p=9·1 × 10-8; familial non-acquired focal epilepsy 3·6, 2·7-4·9, p=1·1 × 10-17). Comparison of an additional cohort of 662 individuals with sporadic non-acquired focal epilepsy to controls did not identify study-wide significant signals. For the individuals with familial non-acquired focal epilepsy, we found that five known epilepsy genes ranked as the top five genes enriched for ultra-rare deleterious variation. After accounting for the control carrier rate, we estimate that these five genes contribute to the risk of epilepsy in approximately 8% of individuals with familial non-acquired focal epilepsy. Our analyses showed that no individual gene was significantly associated with familial genetic generalised epilepsy; however, known epilepsy genes had lower p values relative to the rest of the protein-coding genes (p=5·8 × 10-8) that were lower than expected from a random sampling of genes. INTERPRETATION We identified excess ultra-rare variation in known epilepsy genes, which establishes a clear connection between the genetics of common and rare, severe epilepsies, and shows that the variants responsible for epilepsy risk are exceptionally rare in the general population. Our results suggest that the emerging paradigm of targeting of treatments to the genetic cause in rare devastating epilepsies might also extend to a proportion of common epilepsies. These findings might allow clinicians to broadly explain the cause of these syndromes to patients, and lay the foundation for possible precision treatments in the future. FUNDING National Institute of Neurological Disorders and Stroke (NINDS), and Epilepsy Research UK.

The effect of everolimus on renal angiomyolipoma in patients with tuberous sclerosis complex being treated for subependymal giant cell astrocytoma: subgroup results from the randomized, placebo-controlled, Phase 3 trial EXIST-1

BACKGROUND Tuberous sclerosis complex (TSC) is characterized by benign tumours in multiple organs, including the brain, kidneys, skin, lungs and heart. Our objective was to evaluate everolimus, an mTOR inhibitor, in the treatment of angiomyolipoma in patients with subependymal giant cell astrocytoma (SEGA) associated with TSC. METHODS EXamining everolimus In a Study of Tuberous Sclerosis Complex-1 (NCT00789828), a prospective, double-blind, randomized, placebo-controlled, Phase 3 study, examined everolimus in treating SEGA associated with TSC. Patients with serial SEGA growth from pre-baseline to baseline scans were randomly assigned (2:1) to receive 4.5 mg/m(2)/day everolimus (target blood trough: 5-15 ng/mL; n = 78) or placebo (n = 39). Angiomyolipoma response rates were analysed in patients (n = 44) with target baseline angiomyolipoma lesions (≥1 angiomyolipoma; longest diameter ≥1.0 cm). An angiomyolipoma response rate, defined as the proportion of patients with confirmed angiomyolipoma response, was assessed by kidney CT or MRI screening at baseline, at 12, 24 and 48 weeks and annually. RESULTS Angiomyolipoma response rates were 53.3% (16/30) and 0% (0/14) for everolimus- and placebo-treated patients, respectively. Angiomyolipoma reductions ≥50% in the sum of volumes of all target lesions were seen only in everolimus-treated patients (56.5, 78.3 and 80.0%) compared with placebo-treated patients (0% at each time point) at Weeks 12, 24 and 48, respectively. Greater percentages of everolimus-treated patients had angiomyolipoma reductions ≥30% at these same time points (82.6, 100 and 100% versus 8.3, 18.2 and 16.7% for everolimus versus placebo, respectively). CONCLUSIONS Everolimus showed efficacy in reducing angiomyolipoma lesion volume in patients with SEGA associated with TSC.The trial is registered with ClinicalTrials.gov, number NCT00789828; http://clinicaltrials.gov/ct2/show/NCT00789828?term=EXIST-1&rank=1.

Long-Term Use of Everolimus in Patients with Tuberous Sclerosis Complex: Final Results from the EXIST-1 Study

Background Everolimus, a mammalian target of rapamycin (mTOR) inhibitor, has demonstrated efficacy in treating subependymal giant cell astrocytomas (SEGAs) and other manifestations of tuberous sclerosis complex (TSC). However, long-term use of mTOR inhibitors might be necessary. This analysis explored long-term efficacy and safety of everolimus from the conclusion of the EXIST-1 study (NCT00789828). Methods and Findings EXIST-1 was an international, prospective, double-blind, placebo-controlled phase 3 trial examining everolimus in patients with new or growing TSC-related SEGA. After a double-blind core phase, all remaining patients could receive everolimus in a long-term, open-label extension. Everolimus was initiated at a dose (4.5 mg/m2/day) titrated to a target blood trough of 5–15 ng/mL. SEGA response rate (primary end point) was defined as the proportion of patients achieving confirmed ≥50% reduction in the sum volume of target SEGA lesions from baseline in the absence of worsening nontarget SEGA lesions, new target SEGA lesions, and new or worsening hydrocephalus. Of 111 patients (median age, 9.5 years) who received ≥1 dose of everolimus (median duration, 47.1 months), 57.7% (95% confidence interval [CI], 47.9–67.0) achieved SEGA response. Of 41 patients with target renal angiomyolipomas at baseline, 30 (73.2%) achieved renal angiomyolipoma response. In 105 patients with ≥1 skin lesion at baseline, skin lesion response rate was 58.1%. Incidence of adverse events (AEs) was comparable with that of previous reports, and occurrence of emergent AEs generally decreased over time. The most common AEs (≥30% incidence) suspected to be treatment-related were stomatitis (43.2%) and mouth ulceration (32.4%). Conclusions Everolimus use led to sustained reduction in tumor volume, and new responses were observed for SEGA and renal angiomyolipoma from the blinded core phase of the study. These findings support the hypothesis that everolimus can safely reverse multisystem manifestations of TSC in a significant proportion of patients. Trial Registration ClinicalTrials.gov NCT00789828

Copy number variant analysis from exome data in 349 patients with epileptic encephalopathy

Infantile spasms (IS) and Lennox–Gastaut syndrome (LGS) are epileptic encephalopathies characterized by early onset, intractable seizures, and poor developmental outcomes. De novo sequence mutations and copy number variants (CNVs) are causative in a subset of cases. We used exome sequence data in 349 trios with IS or LGS to identify putative de novo CNVs. We confirm 18 de novo CNVs in 17 patients (4.8%), 10 of which are likely pathogenic, giving a firm genetic diagnosis for 2.9% of patients. Confirmation of exome‐predicted CNVs by array‐based methods is still required due to false‐positive rates of prediction algorithms. Our exome‐based results are consistent with recent array‐based studies in similar cohorts and highlight novel candidate genes for IS and LGS. Ann Neurol 2015;78:323–328

Epilepsy surgery failure in children: a quantitative and qualitative analysis.

OBJECT Resection is a safe and effective treatment option for children with pharmacoresistant focal epilepsy, but some patients continue experience seizures after surgery. While most studies of pediatric epilepsy surgery focus on predictors of postoperative seizure outcome, these factors are often not modifiable, and the reasons for surgical failure may remain unclear. METHODS The authors performed a retrospective cohort study of children and adolescents who received focal resective surgery for pharmacoresistant epilepsy. Both quantitative and qualitative analyses of factors associated with persistent postoperative seizures were conducted. RESULTS Records were reviewed from 110 patients, ranging in age from 6 months to 19 years at the time of surgery, who underwent a total of 115 resections. At a mean 3.1-year follow-up, 76% of patients were free of disabling seizures (Engel Class I outcome). Seizure freedom was predicted by temporal lobe surgery compared with extratemporal resection, tumor or mesial temporal sclerosis compared with cortical dysplasia or other pathologies, and by a lower preoperative seizure frequency. Factors associated with persistent seizures (Engel Class II-IV outcome) included residual epileptogenic tissue adjacent to the resection cavity (40%), an additional epileptogenic zone distant from the resection cavity (32%), and the presence of a hemispheric epilepsy syndrome (28%). CONCLUSIONS While seizure outcomes in pediatric epilepsy surgery may be improved by the use of high-resolution neuroimaging and invasive electrographic studies, a more aggressive resection should be considered in certain patients, including hemispherectomy if a hemispheric epilepsy syndrome is suspected. Family counseling regarding treatment expectations is critical, and reoperation may be warranted in select cases.

Intractable episodic bradycardia resulting from progressive lead traction in an epileptic child with a vagus nerve stimulator: a delayed complication.

Vagus nerve stimulation (VNS) is used as palliation for adult and pediatric patients with intractable epilepsy who are not candidates for curative resection. Although the treatment is generally safe, complications can occur intraoperatively, perioperatively, and in a delayed time frame. In the literature, there are 2 reports of pediatric patients with implanted VNS units who had refractory bradycardia that resolved after the stimulation was turned off. The authors report the case of a 13-year-old boy with a history of vagus nerve stimulator placement at 2 years of age, who developed intractable episodic bradycardia that persisted despite the cessation of VNS and whose imaging results suggested vagus nerve tethering by the leads. He was subsequently taken to the operating room for exploration, where it was confirmed that the stimulator lead was exerting traction on the vagus nerve, which was displaced from the carotid sheath. After the vagus nerve was untethered and the leads were replaced, the bradycardia eventually resolved with continual effective VNS therapy. When placing a VNS unit in a very young child, accommodations must be made for years of expected growth. Delayed intractable bradycardia can result from a vagus nerve under traction by tethered stimulator leads.