Somatic variants as a cause of drug-resistant epilepsy including mesial temporal lobe epilepsy with hippocampal sclerosis.

OBJECTIVE: The contribution of somatic variants to epilepsy has recently been demonstrated, particularly in the etiology of malformations of cortical development. The aim of this study was to determine the diagnostic yield of somatic variants in genes that have been previously associated with a somatic or germline epilepsy model, ascertained from resected brain tissue from patients with multidrug-resistant focal epilepsy. METHODS: Forty-two patients were recruited across three categories: (1) malformations of cortical development, (2) mesial temporal lobe epilepsy with hippocampal sclerosis, and (3) nonlesional focal epilepsy. Participants were subdivided based on histopathology of the resected brain. Paired blood- and brain-derived DNA samples were sequenced using high-coverage targeted next generation sequencing to high depth (585× and 1360×, respectively). Variants were identified using Genome Analysis ToolKit (GATK4) MuTect-2 and confirmed using high-coverage Amplicon-EZ sequencing. RESULTS: Sequence data on 41 patients passed quality control. Four somatic variants were validated following amplicon sequencing: within CBL, ALG13, MTOR, and FLNA. The diagnostic yield across 41 patients was 10%, 9% in mesial temporal lobe epilepsy with hippocampal sclerosis and 20% in malformations of cortical development. SIGNIFICANCE: This study provides novel insights into the etiology of mesial temporal lobe epilepsy with hippocampal sclerosis, highlighting a potential pathogenic role of somatic variants in CBL and ALG13. We also report candidate diagnostic somatic variants in FLNA in focal cortical dysplasia, while providing further insight into the importance of MTOR and related genes in focal cortical dysplasia. This work demonstrates the potential molecular diagnostic value of variants in both germline and somatic epilepsy genes.

Improving the clinical interpretation of missense variants in X linked genes using structural analysis.

BACKGROUND: Improving the clinical interpretation of missense variants can increase the diagnostic yield of genomic testing and lead to personalised management strategies. Currently, due to the imprecision of bioinformatic tools that aim to predict variant pathogenicity, their role in clinical guidelines remains limited. There is a clear need for more accurate prediction algorithms and this study aims to improve performance by harnessing structural biology insights. The focus of this work is missense variants in a subset of genes associated with X linked disorders. METHODS: We have developed a protein-specific variant interpreter (ProSper) that combines genetic and protein structural data. This algorithm predicts missense variant pathogenicity by applying machine learning approaches to the sequence and structural characteristics of variants. RESULTS: ProSper outperformed seven previously described tools, including meta-predictors, in correctly evaluating whether or not variants are pathogenic; this was the case for 11 of the 21 genes associated with X linked disorders that met the inclusion criteria for this study. We also determined gene-specific pathogenicity thresholds that improved the performance of VEST4, REVEL and ClinPred, the three best-performing tools out of the seven that were evaluated; this was the case in 11, 11 and 12 different genes, respectively. CONCLUSION: ProSper can form the basis of a molecule-specific prediction tool that can be implemented into diagnostic strategies. It can allow the accurate prioritisation of missense variants associated with X linked disorders, aiding precise and timely diagnosis. In addition, we demonstrate that gene-specific pathogenicity thresholds for a range of missense prioritisation tools can lead to an increase in prediction accuracy.

A comparison of genomic diagnostics in adults and children with epilepsy and comorbid intellectual disability.

Next generation sequencing provides an important opportunity for improved diagnosis in epilepsy. To date, the majority of diagnostic genetic testing is conducted in the paediatric arena, while the utility of such testing is less well understood in adults with epilepsy. We conducted whole exome sequencing (WES) and copy number variant analyses in an Irish cohort of 101 people with epilepsy and co-morbid intellectual disability to compare the diagnostic yield of genomic testing between adult and paediatric patients. Variant interpretation followed American College of Medical Genetics and Genomics (ACMG) guidelines. We demonstrate that WES, in combination with array-comparative genomic hybridisation, provides a diagnostic rate of 27% in unrelated adult epilepsy patients and 42% in unrelated paediatric patients. We observe a 2.7% rate of ACMG-defined incidental findings. Our findings indicate that WES has similar utility in both adult and paediatric cohorts and is appropriate for diagnostic testing in both epilepsy patient groups.

Re-annotation of 191 developmental and epileptic encephalopathy-associated genes unmasks de novo variants in SCN1A.

The developmental and epileptic encephalopathies (DEE) are a group of rare, severe neurodevelopmental disorders, where even the most thorough sequencing studies leave 60-65% of patients without a molecular diagnosis. Here, we explore the incompleteness of transcript models used for exome and genome analysis as one potential explanation for a lack of current diagnoses. Therefore, we have updated the GENCODE gene annotation for 191 epilepsy-associated genes, using human brain-derived transcriptomic libraries and other data to build 3,550 putative transcript models. Our annotations increase the transcriptional 'footprint' of these genes by over 674 kb. Using SCN1A as a case study, due to its close phenotype/genotype correlation with Dravet syndrome, we screened 122 people with Dravet syndrome or a similar phenotype with a panel of exon sequences representing eight established genes and identified two de novo SCN1A variants that now - through improved gene annotation - are ascribed to residing among our exons. These two (from 122 screened people, 1.6%) molecular diagnoses carry significant clinical implications. Furthermore, we identified a previously classified SCN1A intronic Dravet syndrome-associated variant that now lies within a deeply conserved exon. Our findings illustrate the potential gains of thorough gene annotation in improving diagnostic yields for genetic disorders.

Diagnostic implications of genetic copy number variation in epilepsy plus.

OBJECTIVE: Copy number variations (CNVs) represent a significant genetic risk for several neurodevelopmental disorders including epilepsy. As knowledge increases, reanalysis of existing data is essential. Reliable estimates of the contribution of CNVs to epilepsies from sizeable populations are not available. METHODS: We assembled a cohort of 1255 patients with preexisting array comparative genomic hybridization or single nucleotide polymorphism array based CNV data. All patients had "epilepsy plus," defined as epilepsy with comorbid features, including intellectual disability, psychiatric symptoms, and other neurological and nonneurological features. CNV classification was conducted using a systematic filtering workflow adapted to epilepsy. RESULTS: Of 1097 patients remaining after genetic data quality control, 120 individuals (10.9%) carried at least one autosomal CNV classified as pathogenic; 19 individuals (1.7%) carried at least one autosomal CNV classified as possibly pathogenic. Eleven patients (1%) carried more than one (possibly) pathogenic CNV. We identified CNVs covering recently reported (HNRNPU) or emerging (RORB) epilepsy genes, and further delineated the phenotype associated with mutations of these genes. Additional novel epilepsy candidate genes emerge from our study. Comparing phenotypic features of pathogenic CNV carriers to those of noncarriers of pathogenic CNVs, we show that patients with nonneurological comorbidities, especially dysmorphism, were more likely to carry pathogenic CNVs (odds ratio = 4.09, confidence interval = 2.51-6.68; P = 2.34 × 10-9 ). Meta-analysis including data from published control groups showed that the presence or absence of epilepsy did not affect the detected frequency of CNVs. SIGNIFICANCE: The use of a specifically adapted workflow enabled identification of pathogenic autosomal CNVs in 10.9% of patients with epilepsy plus, which rose to 12.7% when we also considered possibly pathogenic CNVs. Our data indicate that epilepsy with comorbid features should be considered an indication for patients to be selected for a diagnostic algorithm including CNV detection. Collaborative large-scale CNV reanalysis leads to novel declaration of pathogenicity in unexplained cases and can promote discovery of promising candidate epilepsy genes.

Exome sequencing for prenatal diagnosis of fetuses with sonographic abnormalities.

OBJECTIVE: In the absence of aneuploidy or other pathogenic cytogenetic abnormality, fetuses with increased nuchal translucency (NT ≥ 3.5 mm) and/or other sonographic abnormalities have a greater incidence of genetic syndromes, but defining the underlying pathology can be challenging. Here, we investigate the value of whole exome sequencing in fetuses with sonographic abnormalities but normal microarray analysis. METHOD: Whole exome sequencing was performed on DNA extracted from chorionic villi or amniocytes in 24 fetuses with unexplained ultrasound findings. In the first 14 cases sequencing was initially performed on fetal DNA only. For the remaining 10, the trio of fetus, mother and father was sequenced simultaneously. RESULTS: In 21% (5/24) cases, exome sequencing provided definitive diagnoses (Milroy disease, hypophosphatasia, achondrogenesis type 2, Freeman-Sheldon syndrome and Baraitser-Winter Syndrome). In a further case, a plausible diagnosis of orofaciodigital syndrome type 6 was made. In two others, a single mutation in an autosomal recessive gene was identified, but incomplete sequencing coverage precluded exclusion of the presence of a second mutation. CONCLUSION: Whole exome sequencing improves prenatal diagnosis in euploid fetuses with abnormal ultrasound scans. In order to expedite interpretation of results, trio sequencing should be employed, but interpretation can still be compromised by incomplete coverage of relevant genes.

Favourable response to ketogenic dietary therapies: undiagnosed glucose 1 transporter deficiency syndrome is only one factor.

AIM: We aimed to determine whether response to ketogenic dietary therapies (KDT) was due to undiagnosed glucose transporter type 1 deficiency syndrome (GLUT1-DS). METHOD: Targeted resequencing of the SLC2A1 gene was completed in individuals without previously known GLUT1-DS who received KDT for their epilepsy. Hospital records were used to obtain demographic and clinical data. Response to KDT at various follow-up points was defined as seizure reduction of at least 50%. Seizure freedom achieved at any follow-up point was also documented. Fisher's exact and gene-burden association tests were conducted using the PLINK/SEQ open-source genetics library. RESULTS: Of the 246 participants, one was shown to have a novel variant in SLC2A1 that was predicted to be deleterious. This individual was seizure-free on KDT. Rates of seizure freedom in cases without GLUT1-DS were below 8% at each follow-up point. Two cases without SLC2A1 mutations were seizure-free at every follow-up point recorded. No significant results were obtained from Fisher's exact or gene-burden association tests. INTERPRETATION: A favourable response to KDT is not solely explained by mutations in SLC2A1. Other genetic factors should be sought to identify those who are most likely to benefit from dietary treatment for epilepsy, particularly those who may achieve seizure freedom.

CHD2 variants are a risk factor for photosensitivity in epilepsy.

Photosensitivity is a heritable abnormal cortical response to flickering light, manifesting as particular electroencephalographic changes, with or without seizures. Photosensitivity is prominent in a very rare epileptic encephalopathy due to de novo CHD2 mutations, but is also seen in epileptic encephalopathies due to other gene mutations. We determined whether CHD2 variation underlies photosensitivity in common epilepsies, specific photosensitive epilepsies and individuals with photosensitivity without seizures. We studied 580 individuals with epilepsy and either photosensitive seizures or abnormal photoparoxysmal response on electroencephalography, or both, and 55 individuals with photoparoxysmal response but no seizures. We compared CHD2 sequence data to publicly available data from 34 427 individuals, not enriched for epilepsy. We investigated the role of unique variants seen only once in the entire data set. We sought CHD2 variants in 238 exomes from familial genetic generalized epilepsies, and in other public exome data sets. We identified 11 unique variants in the 580 individuals with photosensitive epilepsies and 128 unique variants in the 34 427 controls: unique CHD2 variation is over-represented in cases overall (P = 2.17 × 10(-5)). Among epilepsy syndromes, there was over-representation of unique CHD2 variants (3/36 cases) in the archetypal photosensitive epilepsy syndrome, eyelid myoclonia with absences (P = 3.50 × 10(-4)). CHD2 variation was not over-represented in photoparoxysmal response without seizures. Zebrafish larvae with chd2 knockdown were tested for photosensitivity. Chd2 knockdown markedly enhanced mild innate zebrafish larval photosensitivity. CHD2 mutation is the first identified cause of the archetypal generalized photosensitive epilepsy syndrome, eyelid myoclonia with absences. Unique CHD2 variants are also associated with photosensitivity in common epilepsies. CHD2 does not encode an ion channel, opening new avenues for research into human cortical excitability.

Non-invasive prenatal diagnosis of achondroplasia and thanatophoric dysplasia: next-generation sequencing allows for a safer, more accurate, and comprehensive approach.

OBJECTIVE: Accurate prenatal diagnosis of genetic conditions can be challenging and usually requires invasive testing. Here, we demonstrate the potential of next-generation sequencing (NGS) for the analysis of cell-free DNA in maternal blood to transform prenatal diagnosis of monogenic disorders. METHODS: Analysis of cell-free DNA using a PCR and restriction enzyme digest (PCR-RED) was compared with a novel NGS assay in pregnancies at risk of achondroplasia and thanatophoric dysplasia. RESULTS: PCR-RED was performed in 72 cases and was correct in 88.6%, inconclusive in 7% with one false negative. NGS was performed in 47 cases and was accurate in 96.2% with no inconclusives. Both approaches were used in 27 cases, with NGS giving the correct result in the two cases inconclusive with PCR-RED. CONCLUSION: NGS provides an accurate, flexible approach to non-invasive prenatal diagnosis of de novo and paternally inherited mutations. It is more sensitive than PCR-RED and is ideal when screening a gene with multiple potential pathogenic mutations. These findings highlight the value of NGS in the development of non-invasive prenatal diagnosis for other monogenic disorders.

Targeted gene panel sequencing in children with very early onset inflammatory bowel disease--evaluation and prospective analysis.

BACKGROUND: Multiple monogenetic conditions with partially overlapping phenotypes can present with inflammatory bowel disease (IBD)-like intestinal inflammation. With novel genotype-specific therapies emerging, establishing a molecular diagnosis is becoming increasingly important. DESIGN: We have introduced targeted next-generation sequencing (NGS) technology as a prospective screening tool in children with very early onset IBD (VEOIBD). We evaluated the coverage of 40 VEOIBD genes in two separate cohorts undergoing targeted gene panel sequencing (TGPS) (n=25) and whole exome sequencing (WES) (n=20). RESULTS: TGPS revealed causative mutations in four genes (IL10RA, EPCAM, TTC37 and SKIV2L) discovered unexpected phenotypes and directly influenced clinical decision making by supporting as well as avoiding haematopoietic stem cell transplantation. TGPS resulted in significantly higher median coverage when compared with WES, fewer coverage deficiencies and improved variant detection across established VEOIBD genes. CONCLUSIONS: Excluding or confirming known VEOIBD genotypes should be considered early in the disease course in all cases of therapy-refractory VEOIBD, as it can have a direct impact on patient management. To combine both described NGS technologies would compensate for the limitations of WES for disease-specific application while offering the opportunity for novel gene discovery in the research setting.

Evaluation of non-invasive prenatal testing (NIPT) for aneuploidy in an NHS setting: a reliable accurate prenatal non-invasive diagnosis (RAPID) protocol.

BACKGROUND: Non-invasive prenatal testing (NIPT) for aneuploidies is now available through commercial companies in many countries, including through private practice in the United Kingdom (UK). Thorough evaluation of service delivery requirements are needed to facilitate NIPT being offered more widely within state funded healthcare systems such as the UK's National Health Service (NHS). Successful implementation will require the development of laboratory standards, consideration of stakeholder views, an analysis of costs and development of patient and health professional educational materials. METHODS/DESIGN: NIPT will be offered in an NHS setting as a contingent screening test. Pregnant woman will be recruited through six maternity units in England and Scotland. Women eligible for Down's syndrome screening (DSS) will be informed about the study at the time of booking. Women that choose routine DSS will be offered NIPT if they have a screening risk ≥ 1:1000. NIPT results for trisomy 21, 18, 13 will be reported within 7-10 working days. Data on DSS, NIPT and invasive testing uptake, pregnancy outcomes and test efficacy will be collected. Additional data will be gathered though questionnaires to a) determine acceptability to patients and health professionals, b) evaluate patient and health professional education, c) assess informed choice in women accepting or declining testing and d) gauge family expenses. Qualitative interviews will also be conducted with a sub-set of participating women and health professionals. DISCUSSION: The results of this study will make a significant contribution to policy decisions around the implementation of NIPT for aneuploidies within the UK NHS. The laboratory standards for testing and reporting, education materials and counselling strategies developed as part of the study are likely to underpin the introduction of NIPT into NHS practice. NIHR PORTFOLIO NUMBER: 13865.

Whole exome sequencing of familial hypercholesterolaemia patients negative for LDLR/APOB/PCSK9 mutations.

BACKGROUND: Familial hypercholesterolaemia (FH) is an autosomal dominant disease of lipid metabolism, which leads to early coronary heart disease. Mutations in LDLR, APOB and PCSK9 can be detected in 80% of definite FH (DFH) patients. This study aimed to identify novel FH-causing genetic variants in patients with no detectable mutation. METHODS AND RESULTS: Exomes of 125 unrelated DFH patients were sequenced, as part of the UK10K project. First, analysis of known FH genes identified 23 LDLR and two APOB mutations, and patients with explained causes of FH were excluded from further analysis. Second, common and rare variants in genes associated with low-density lipoprotein cholesterol (LDL-C) levels in genome-wide association study (GWAS) meta-analysis were examined. There was no clear rare variant association in LDL-C GWAS hits; however, there were 29 patients with a high LDL-C SNP score suggestive of polygenic hypercholesterolaemia. Finally, a gene-based burden test for an excess of rare (frequency <0.005) or novel variants in cases versus 1926 controls was performed, with variants with an unlikely functional effect (intronic, synonymous) filtered out. CONCLUSIONS: No major novel locus for FH was detected, with no gene having a functional variant in more than three patients; however, an excess of novel variants was found in 18 genes, of which the strongest candidates included CH25H and INSIG2 (p<4.3×10(-4) and p<3.7×10(-3), respectively). This suggests that the genetic cause of FH in these unexplained cases is likely to be very heterogeneous, which complicates the diagnostic and novel gene discovery process.

Clinico-pathological correlations of congenital and infantile nephrotic syndrome over twenty years.

BACKGROUND: Nephrotic syndrome (NS) presenting early in life is caused by heterogeneous glomerular diseases. We retrospectively evaluated whether histological diagnosis in children presenting with NS in the first year of life predicts remission or progression to end-stage kidney disease (ESKD). METHODS: This is a single centre retrospective review of all children diagnosed with NS before one year of age between 1990 and 2009. All subjects had a renal biopsy, which was independently blindly reviewed by a single renal pathologist for the purpose of this study. RESULTS: Forty-nine children (25 female) who presented at 0.1-11.6 (median 1.6) months were included with 31 presenting within the first three months of life. Histopathological review diagnostic categories were; 13 Mesangial proliferative glomerulopathy (MesGN), 12 Focal and segmental glomerulosclerosis (FSGS), 11 Finnish type changes, eight Diffuse Mesangial Sclerosis (DMS), three Minimal change disease (MCD) and one each of Dense Deposit Disease (DDD) and Membranous nephropathy. Two children died from haemorrhagic complications of the biopsy. Eight children achieved remission (four MesGN, one Finnish type changes, one FSGS, one MCD and one membranous) with patient and renal survival of 73 % and 43 %, respectively, at follow-up duration of 5-222 (median 73) months (with five lost to follow-up). All children with Finnish-type histopathological changes presented within five months of age. Due to the historical nature of the cohort, genetic testing was only available for 14 children, nine of whom had an identifiable genetic basis (seven NPHS1, one PLCE1 and one ITGA3) with none of these nine children achieving remission. All of them had presented within four months of age and required renal replacement therapy, and two died. CONCLUSIONS: Histopathological findings are varied in children presenting with NS early in life. Whilst groups of histological patterns of disease are associated with differing outcomes, accurate prediction of disease course in a specific case is difficult and more widespread genetic testing may improve the understanding of this group of diseases and their optimal management.

A novel homozygous ERCC5 truncating mutation in a family with prenatal arthrogryposis--further evidence of genotype-phenotype correlation.

We report on a family with five fetuses conceived to first cousin parents presenting with abnormal ultrasound findings including contractures and microcephaly. Cerebellar hypoplasia and ventriculomegaly were also present in two and fetal edema developed in the one fetus that survived beyond 24 weeks of gestation. Linkage studies of 15 members of the family, including four affecteds, were undertaken followed by exome sequencing of one affected individual and their parents. Analysis of exome data was restricted to the 9.3 Mb largest shared region of homozygosity identified by linkage; a single novel homozygous mutation in the proband that was heterozygous in the parents (ERCC5 c.2766dupA, p.Leu923ThrfsX7) was identified. This segregated with disease. ERCC5 is a component of the nucleotide excision repair machinery and biallelic mutations in the gene have previously been associated with xeroderma pigmentosum (group G), Cockayne syndrome and the more severe cerebrooculofacioskeletal syndrome. The phenotype in the family we report on is consistent with a severe manifestation of cerebrooculofacioskeletal syndrome. Our data broaden the reported clinical spectrum of ERCC5 mutations and provide further evidence of genotype-phenotype correlation with truncating mutations being associated with severe phenotypes. They also demonstrate the molecular diagnostic power of a combined approach of linkage studies and exome sequencing in families with rare, genetically heterogeneous disorders and a well described pedigree.

A novel heterozygous SOX2 mutation causing congenital bilateral anophthalmia, hypogonadotropic hypogonadism and growth hormone deficiency.

Heterozygous de novo mutations in SOX2 have been reported in approximately 10-20% of patients with unilateral or bilateral anophthalmia or microphthalmia. An additional phenotype of hypopituitarism, with anterior pituitary hypoplasia and hypogonadotropic hypogonadism, has been reported in patients carrying SOX2 alterations. We report a novel heterozygous mutation in the SOX2 gene in a male affected with congenital bilateral anophthalmia, hypogonadotrophic hypogonadism and growth hormone deficiency. The mutation we describe is a cytosine deletion in position 905 (c905delC) which causes frameshift and an aberrant C-terminal domain. Our report highlights the fact that subjects affected with eye anomalies and harboring SOX2 mutations are at high risk for gonadotropin deficiency, which has important implications for their clinical management.

KCNJ10 mutations display differential sensitivity to heteromerisation with KCNJ16.

BACKGROUND/AIMS: Mutations in the inwardly-rectifying K(+)-channel KCNJ10/Kir4.1 cause autosomal recessive EAST syndrome (epilepsy, ataxia, sensorineural deafness and tubulopathy). KCNJ10 is expressed in the distal convoluted tubule of the kidney, stria vascularis of the inner ear and brain glial cells. Patients diagnosed clinically with EAST syndrome were genotyped and mutations in KCNJ10 were studied functionally. METHODS: Patient DNA was amplified and sequenced, and new mutations were identified. Mutant and wild-type KCNJ10 constructs were cloned and heterologously expressed in Xenopus oocytes. Whole-cell K(+) currents were measured by 2-electrode voltage clamping and channel expression was analysed by Western blotting. RESULTS: We identified 3 homozygous mutations in KCNJ10 (p.F75C, p.A167V and p.V91fs197X), with mutation p.A167V previously reported in a compound heterozygous state. Oocytes expressing wild-type human KCNJ10 showed inwardly rectified currents, which were significantly reduced in all of the mutants (p < 0.001). Specific inhibition of KCNJ10 currents by Ba(2+) demonstrated a large residual function in p.A167V only, which was not compatible with causing disease. However, co-expression with KCNJ16 abolished function in these heteromeric channels almost completely. CONCLUSION: This study provides an explanation for the pathophysiology of the p.A167V KCNJ10 mutation, which had previously not been considered pathogenic on its own. These findings provide evidence for the functional cooperation of KCNJ10 and KCNJ16. Thus, in vitro ascertainment of KCNJ10 function may necessitate co-expression with KCNJ16.

Evaluation of real-time quantitative PCR as a standard cytogenetic diagnostic tool for confirmation of microarray (aCGH) results and determination of inheritance.

AIM: To evaluate the use of real-time quantitative PCR (qPCR) as a diagnostic tool for follow up of abnormal microarray (aCGH) results. METHOD: qPCR was performed on 207 samples with known aCGH results to detect chromosomal abnormality and determine the capability of qPCR. Eighty-four samples were processed and the results compared with the original aCGH result and with one or more of the alternative follow-up methods: aCGH, fluorescence in situ hybridization (FISH), or karyotyping. A separate cohort of 107 samples was used to determine critical threshold values for qPCR. A further 16 samples were assessed in reproducibility and sensitivity studies. RESULTS: All qPCR findings were consistent with the original aCGH results, and qPCR was found to be a superior follow-up method compared to FISH and karyotyping. Critical threshold values were also determined from this study. CONCLUSION: In this study, qPCR analysis identified all copy number changes. qPCR is an accurate, rapid, reliable, and inexpensive technique for confirming copy number changes, and for determining the inheritance of such abnormalities to aid interpretation of results. We also present the critical threshold values required for qPCR as a practical tool. This technique has now been successfully implemented as part of the clinical diagnostic service within our laboratory.

Neurological features of epilepsy, ataxia, sensorineural deafness, tubulopathy syndrome.

AIM: Recently, we reported a previously unrecognized symptom constellation comprising epilepsy, ataxia, sensorineural deafness, and tubulopathy (EAST syndrome) associated with recessive mutations in the KCNJ10 gene. Here, we provide a detailed characterization of the clinical features of the syndrome to aid patient management with respect to diagnosis, prognostic counselling, and identification of best treatment modalities. METHOD: We conducted a retrospective review of the detailed neurological and neuroradiological features of nine children (four females, five males; age range at last examination 6-20y) with genetically proven EAST syndrome. RESULTS: All children presented with tonic-clonic seizures in infancy. Later, non-progressive, cerebellar ataxia and hearing loss were noted. Whilst seizures mostly responded well to treatment, ataxia proved to be the most debilitating feature, with three patients non-ambulant. All available magnetic resonance imaging (MRI) revealed subtle symmetrical signal changes in the cerebellar dentate nuclei. Moreover, four patients had a small corpus callosum and brainstem hypoplasia, and three had a small spinal cord. Regional quantitative volumetric analysis of the images confirmed the corpus callosum and brainstem hypoplasia and showed further patterns of variation from the norm. INTERPRETATION: The neurological features of EAST syndrome appear to be non-progressive, which is important for prognostic counselling. The spectrum of EAST syndrome includes consistent abnormalities on brain MRI, which may aid diagnosis. Further longitudinal documentation is required to determine the true natural history of the disorder.