STXBP1 Mutations Research Articles

CASK aberrations in male patients with Ohtahara syndrome and cerebellar hypoplasia

Ohtahara syndrome (OS) is one of the most severe and earliest forms of epilepsy. STXBP1 and ARX mutations have been reported in patients with OS. In this study, we aimed to identify new genes involved in OS by copy number analysis and whole exome sequencing. Copy number analysis and whole exome sequencing were performed in 34 and 12 patients with OS, respectively. Fluorescence in situ hybridization, quantitative polymerase chain reaction (PCR), and breakpoint-specific and reverse-transcriptase PCR analyses were performed to characterize a deletion. Immunoblotting using lymphoblastoid cells was done to examine expression of CASK protein. Genomic microarray analysis revealed a 111-kb deletion involving exon 2 of CASK at Xp11.4 in a male patient. The deletion was inherited from his mother, who was somatic mosaic for the deletion. Sequencing of the mutant transcript expressed in lymphoblastoid cell lines derived from the patient confirmed the deletion of exon 2 in the mutant transcript with a premature stop codon. Whole exome sequencing identified another male patient who was harboring a c.1A>G mutation in CASK, which occurred de novo. Both patients showed severe cerebellar hypoplasia along with other congenital anomalies such as micrognathia, a high arched palate, and finger anomalies. No CASK protein was detected by immunoblotting in lymphoblastoid cells derived from two patients. The detected mutations are highly likely to cause the loss of function of the CASK protein in male individuals. CASK mutations have been reported in patients with intellectual disability with microcephaly and pontocerebellar hypoplasia or congenital nystagmus, and those with FG syndrome. Our data expand the clinical spectrum of CASK mutations to include OS with cerebellar hypoplasia and congenital anomalies at the most severe end.

Epileptic and nonepileptic features in patients with early onset epileptic encephalopathy and STXBP1 mutations

STXBP1 (MUNC18-1) mutations have been associated with various types of epilepsies, mostly beginning early in life. To refine the phenotype associated with STXBP1 aberrations in early onset epileptic syndromes, we studied this gene in a cohort of patients with early onset epileptic encephalopathy. STXBP1 was screened in a multicenter cohort of 52 patients with early onset epilepsy (first seizure observed before the age of 3 months), no cortical malformation on brain magnetic resonance imaging (MRI), and negative metabolic screening. Three groups of patients could be distinguished in this cohort: (1) Ohtahara syndromes (n = 38); (2) early myoclonic encephalopathies (n = 7); and (3) early onset epileptic encephalopathies that did not match any familiar syndrome (n = 7). None of the patients displayed any cortical malformation on brain MRI and all were screened through multiple video-electroencephalography (EEG) recordings for a time period spanning from birth to their sixth postnatal month. Subsequently, patients had standard EEG or video-EEG recordings. We found five novel STXBP1 mutations in patients for whom video-EEG recordings could be sampled from the beginning of the disease. All patients with a mutation displayed Ohtahara syndrome, since most early seizures could be classified as epileptic spasms and since the silent EEG periods were on average shorter than bursts. However, each patient in addition displayed a particular clinical and EEG feature: In two patients, early seizures were clonic, with very early EEG studies exhibiting relatively low amplitude bursts of activity before progressing into a typical suppression-burst pattern, whereas the three other patients displayed epileptic spasms associated with typical suppression-burst patterns starting from the early recordings. Epilepsy dramatically improved after 6 months and finally disappeared before the end of the first year of life for four patients; the remaining one patient had few seizures until 18 months of age. In parallel, EEG paroxysmal abnormalities disappeared in three patients and decreased in two, giving place to continuous activity with fast rhythms. Each patient displayed frequent nonepileptic movement disorders that could easily be mistaken for epileptic seizures. These movements could be observed as early as the neonatal period and, unlike seizures, persisted during all the follow-up period. We confirm that STXBP1 is a major gene to screen in cases of Ohtahara syndrome, since it is mutated in >10% of the Ohtahara patients within our cohort. This gene should particularly be tested in the case of a surprising evolution of the patient condition if epileptic seizures and EEG paroxysmal activity disappear and are replaced by fast rhythms after the end of the first postnatal year.

Paternal mosaicism of an STXBP1 mutation in OS

Ohtahara syndrome (OS) is one of the most severe and earliest forms of epilepsy. We have recently identified that the de novo mutations of STXBP1 are important causes for OS. Here we report a paternal somatic mosaicism of an STXBP1 mutation. The affected daughter had onset of spasms at 1 month of age, and interictal electroencephalogram showed suppression-burst pattern, leading to the diagnosis of OS. She had a heterozygous c.902+5G>A mutation of STXBP1, which affects donor splicing of exon 10, resulting in 138-bp insertion of intron 10 sequences in the transcript. The mutant transcript had a premature stop codon, and was degraded by nonsense-mediated mRNA decay in lymphoblastoid cells derived from the patient. High-resolution melting analysis of clinically unaffected parental DNAs suggested that the father was somatic mosaic for the mutation, which was also suggested by sequencing. Cloning of PCR products amplified with the paternal DNA samples extracted from blood, saliva, buccal cells, and nails suggested that 5.3%, 8.7%, 11.9%, and 16.9% of alleles harbored the mutation, respectively. This is a first report of somatic mosaicism of an STXBP1 mutation, which has implications in genetic counseling of OS.

STXBP1 mutations cause not only Ohtahara syndrome but also West syndrome—Result of Japanese cohort study

We performed STXBP1 mutation analyses in 86 patients with various types of epilepsies, including 10 patients with OS, 43 with West syndrome, 2 with Lennox-Gastaut syndrome, 12 with symptomatic generalized epilepsy, 14 with symptomatic partial epilepsy, and 5 with other undetermined types of epilepsy. In all patients, the etiology was unknown, but ARX and CDKL5 mutations were negative in all cases. All coding exons of STXBP1 were analyzed by direct-sequencing. Two de novo nucleotide alterations of STXBP1 were identified in two patients with Ohtahara and West syndrome, respectively. No de novo or deleterious mutations in STXBP1 were found in the remaining 84 patients with various types of symptomatic epilepsies. This is the first case report showing that STXBP1 mutations caused West syndrome from the onset of epilepsy. STXBP1 analysis should be considered as an etiology of symptomatic West syndrome without explainable cause.

STXBP1 mutations in early infantile epileptic encephalopathy with suppression‐burst pattern

De novo STXBP1 mutations have been found in individuals with early infantile epileptic encephalopathy with suppression-burst pattern (EIEE). Our aim was to delineate the clinical spectrum of subjects with STXBP1 mutations, and to examine their biologic aspects. STXBP1 was analyzed in 29 and 54 cases of cryptogenic EIEE and West syndrome, respectively, as a second cohort. RNA splicing was analyzed in lymphoblastoid cells from a subject harboring a c.663 + 5G>A mutation. Expression of STXBP1 protein with missense mutations was examined in neuroblastoma2A cells. A total of seven novel STXBP1 mutations were found in nine EIEE cases, but not in West syndrome. The mutations include two frameshift mutations, three nonsense mutations, a splicing mutation, and a recurrent missense mutation in three unrelated cases. Including our previous data, 10 of 14 individuals (71%) with STXBP1 aberrations had the onset of spasms after 1 month, suggesting relatively later onset of epileptic spasms. Nonsense-mediated mRNA decay associated with abnormal splicing was demonstrated. Transient expression revealed that STXBP1 proteins with missense mutations resulted in degradation in neuroblastoma2A cells. Collectively, STXBP1 aberrations can account for about one-third individuals with EIEE (14 of 43). These genetic and biologic data clearly showed that haploinsufficiency of STXBP1 is the important cause for cryptogenic EIEE.

Clinical spectrum of early-onset epileptic encephalopathies associated with STXBP1 mutations

Heterozygous mutations in STXBP1, encoding the syntaxin binding protein 1, have recently been identified in Ohtahara syndrome, an epileptic encephalopathy with very early onset. In order to explore the phenotypic spectrum associated with STXBP1 mutations, we analyzed a cohort of patients with unexplained early-onset epileptic encephalopathies. We collected and clinically characterized 106 patients with early-onset epileptic encephalopathies. Mutation analysis of the STXBP1 gene was done using sequence analysis of the exon and intron-exon boundaries and multiplex amplification quantification to detect copy number variations. We identified 4 truncating mutations and 2 microdeletions partially affecting STXBP1 in 6 of the 106 patients. All mutations are predicted to abolish STXBP1 function and 5 mutations were proven to occur de novo. None of the mutation-carrying patients had Ohtahara syndrome. One patient was diagnosed with West syndrome at disease onset, while the initial phenotype of 5 further patients did not fit into a specific recognized epilepsy syndrome. Three of these patients later evolved to West syndrome. All patients had severe to profound mental retardation, and ataxia or dyskinetic movements were present in 5 patients. This study shows that mutations in STXBP1 are not limited to patients with Ohtahara syndrome, but are also present in 10% (5/49) of patients with an early-onset epileptic encephalopathy that does not fit into either Ohtahara or West syndrome and rarely in typical West syndrome. STXBP1 mutational analysis should be considered in the diagnostic evaluation of this challenging group of patients.

O2-6 Medical and socioeconomic outcome in adulthood of childhood onset epilepsy: Findings from the National Child Development Study

O2-5 The clinical spectrum of STXBP1 mutations in early onset epileptic encephalpathy: between Ohtahara-syndrome and West syndrome? B. Ceulemans1,2,3 *, S. Weckhuysen4,5,6, K. Verhaert1, L. Deprez7,8,9, A. Holmgren7,8,9, A. Janssen10, W. Van Paesschen11, A. Jordanova4,5,12, L. Lagae3,13, P. De Jonghe4,5,14. 1Child Neurology, University Hospital of Antwerp, Antwerp, Belgium; 2Child Neurology, University of Antwerp, Antwerp, Belgium; 3Child Neurology, Epilepsy Centre Pulderbos, Pulderbos, Belgium; 4Neurology, University of Antwerp, Antwerp, Belgium; 5Neurology, Department of Molecular Genetics, VIB, Antwerp, Belgium; 6Neurology, Epilepsy Centre Kempenhaeghe, Oosterhout, The Netherlands; 7Biochemistry, University of Antwerp, Antwerp, Belgium; 8Biochemistry, Department of Molecular Genetics, VIB, Antwerp, Belgium; 9Biochemistry, Institute Born-Bunge, Antwerp, Belgium; 10Child Neurology, University Hospital of Brussels, Brussels, Belgium; 11Neurology, University Hospital Gasthuisberg, KU Leuven, Leuven, Belgium; 12Neurology, Institute Born-Bunge, Antwerp, Belgium; 13Child Neurology, University Hospital Gasthuisberg, KU Leuven, Leuven, Belgium; 14Neurology, University Hospital of Antwerp, Antwerp, Belgium

O2-5 The clinical spectrum of STXBP1 mutations in early onset epileptic encephalpathy: between Ohtahara-syndrome and West syndrome?

O2-5 The clinical spectrum of STXBP1 mutations in early onset epileptic encephalpathy: between Ohtahara-syndrome and West syndrome? B. Ceulemans1,2,3 *, S. Weckhuysen4,5,6, K. Verhaert1, L. Deprez7,8,9, A. Holmgren7,8,9, A. Janssen10, W. Van Paesschen11, A. Jordanova4,5,12, L. Lagae3,13, P. De Jonghe4,5,14. 1Child Neurology, University Hospital of Antwerp, Antwerp, Belgium; 2Child Neurology, University of Antwerp, Antwerp, Belgium; 3Child Neurology, Epilepsy Centre Pulderbos, Pulderbos, Belgium; 4Neurology, University of Antwerp, Antwerp, Belgium; 5Neurology, Department of Molecular Genetics, VIB, Antwerp, Belgium; 6Neurology, Epilepsy Centre Kempenhaeghe, Oosterhout, The Netherlands; 7Biochemistry, University of Antwerp, Antwerp, Belgium; 8Biochemistry, Department of Molecular Genetics, VIB, Antwerp, Belgium; 9Biochemistry, Institute Born-Bunge, Antwerp, Belgium; 10Child Neurology, University Hospital of Brussels, Brussels, Belgium; 11Neurology, University Hospital Gasthuisberg, KU Leuven, Leuven, Belgium; 12Neurology, Institute Born-Bunge, Antwerp, Belgium; 13Child Neurology, University Hospital Gasthuisberg, KU Leuven, Leuven, Belgium; 14Neurology, University Hospital of Antwerp, Antwerp, Belgium