Whole-exome sequencing (WES) has revolutionized Mendelian diagnostics, however, there is no consensus on the timing of data review in undiagnosed individuals and only preliminary data on the ...cost-effectiveness of this technology. We aimed to assess the utility of WES data reanalysis for diagnosis in Mendelian disorders and to analyze the cost-effectiveness of this technology compared with a traditional diagnostic pathway.
WES was applied to a cohort of 54 patients from 37 families with a variety of Mendelian disorders to identify the genetic etiology. Reanalysis was performed after 12 months with an improved WES diagnostic pipeline. A comparison was made between costs of a modeled WES pathway and a traditional diagnostic pathway in a cohort with intellectual disability (ID).
Reanalysis of WES data at 12 months improved diagnostic success from 30 to 41% due to interim publication of disease genes, expanded phenotype data from referrer, and an improved bioinformatics pipeline. Cost analysis on the ID cohort showed average cost savings of US$586 (AU$782) for each additional diagnosis.
Early application of WES in Mendelian disorders is cost-effective and reanalysis of an undiagnosed individual at a 12-month time point increases total diagnoses by 11%.
Certain mutations can cause proteins to accumulate in neurons, leading to neurodegeneration. We recently showed, however, that upregulation of a wild-type protein, Ataxin1, caused by ...haploinsufficiency of its repressor, the RNA-binding protein Pumilio1 (PUM1), also causes neurodegeneration in mice. We therefore searched for human patients with PUM1 mutations. We identified eleven individuals with either PUM1 deletions or de novo missense variants who suffer a developmental syndrome (Pumilio1-associated developmental disability, ataxia, and seizure; PADDAS). We also identified a milder missense mutation in a family with adult-onset ataxia with incomplete penetrance (Pumilio1-related cerebellar ataxia, PRCA). Studies in patient-derived cells revealed that the missense mutations reduced PUM1 protein levels by ∼25% in the adult-onset cases and by ∼50% in the infantile-onset cases; levels of known PUM1 targets increased accordingly. Changes in protein levels thus track with phenotypic severity, and identifying posttranscriptional modulators of protein expression should identify new candidate disease genes.
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•The brain is sensitive to levels of PUM1 and some of its targets•PUM1 haploinsufficiency causes developmental delay, ataxia, and other problems•Mutations that reduce PUM1 levels by 25% are associated with adult-onset ataxia•Regulators of disease-driving proteins are a pool of new candidate disease genes
Different dosages of an RNA-binding protein result in human neurological diseases of corresponding severities.
Genetic sequencing is identifying an expanding number of variants of GABAA receptors associated with human epilepsies. We identified a new de novo variant of the β2 subunit (β2L51M) of the inhibitory ...GABAA receptor associated with seizures. Our analysis determined the pathogenicity of the variant and the effects of anti-seizure medications. Our data demonstrates that the variant reduced cell surface trafficking and peak GABA-gated currents. Synaptic currents mediated by variant-containing receptors decayed faster than wild-type and single receptor currents showed that the variant shortened the duration of receptor activity by decreasing receptor open times. We tested the effects of the anti-seizure medications, midazolam, carbamazepine and valproate and found that all three enhance variant receptor surface expression. Additionally, midazolam restored receptor function by increasing single receptor active periods and synaptic current decay times towards wild-type levels. By contrast, valproate increased synaptic peak currents, event frequency and promoted synaptic bursting. Our study identifies a new disease-causing variant to the GABAA receptor, profiles its pathogenic effects and demonstrates how anti-seizure drugs correct its functional deficits.
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•Through genetic screening we have discovered a new variant to the gene (GABRB2) that encodes the β2 subunit (p.Leu51Met) of the GABAA receptor.•The variant reduced cell surface expression, peak currents and synaptic current decay.•Midazolam, carbamazepine and valproate enhanced surface expression of this variant.•Midazolam restored the synaptic decay times of the variant by prolonging the active duration of single GABAA receptors.•Valproate enhanced synaptic currents of the variant and promoted synaptic bursting, thus strengthening GABAergic input.
•The majority (93 %) of families had a positive molecular genetic diagnosis.•Ten people (20 %) had recurrent seizures, 12–13 years from last childhood seizure.•Seven children had global developmental ...delay (14 %), 5 (10 %) had Autism.•The majority (82 %) have average adaptive functioning.•Predictors of adverse outcomes were high seizure burden & longer epilepsy duration.
To determine: i) seizure recurrence; ii) developmental disability; iii) co-morbidities and risk factors in self-limited familial neonatal and/or infantile epilepsy (SeLFE) in a multigenerational study.
Families were retrospectively recruited from epilepsy databases (2021–2022) in 2 paediatric hospitals, Sydney, Australia. Eligible families had 2 first degree relatives with seizures and underwent genetic testing. Demographics/clinical data were collected from interviews and medical records. Vineland Adaptive Behaviour Scales-Third Edition measured adaptive function.
Fifteen families participated. Fourteen had a genetic diagnosis (93%): 11 pathogenic; PRRT2 (n=4), KCNQ2 (n=3), SCN2A (n=4), 3 likely pathogenic; KCNQ2 (n=1), SCN8A (n=2). Seizures affected 73 individuals (ages 1–76 years); 30 children and 20 adults had in-depth phenotyping. Ten of 50 individuals (20%) had seizure recurrence, aged 8–65 years. Median time from last neonatal/infantile seizure was 11.8/12.8 years. Predictors of recurrence were high seizure number (p=0.05) and longer treatment duration (p=0.03). Seven children had global developmental delay (GDD): mild (n=4), moderate (n=1) and severe (n=2). Vineland-3 identified 3 had low-average and 3 had mild-moderately impaired functioning. The majority (82%) were average. GDD was associated with older age at last seizure (p=0.03), longer epilepsy duration (p=0.02), and higher number of anti-seizure medications (p=0.05). Four children had speech delay, 5 (10%) had Autism Spectrum Disorder. Paroxysmal kinesiogenic dyskinesia (n=5) occurred in 4 families and hemiplegic migraine (n=8) in 3 families.
Individuals with SeLFE have a small risk of recurrent seizures (20%) and neurodevelopmental disability. Significant predictors are higher seizure number and longer epilepsy duration. Developmental surveillance is imperative.
Early infantile epileptic encephalopathy 38 (EIEE38, MIM #617020) is caused by biallelic variants in ARV1, encoding a transmembrane protein of the endoplasmic reticulum with a pivotal role in ...glycosylphosphatidylinositol (GPI) biosynthesis. We ascertained seven new patients from six unrelated families harboring biallelic variants in ARV1, including five novel variants. Affected individuals showed psychomotor delay, hypotonia, early onset refractory seizures followed by regression and specific neuroimaging features. Flow cytometric analysis on patient fibroblasts showed a decrease in GPI‐anchored proteins on the cell surface, supporting a lower residual activity of the mutant ARV1 as compared to the wildtype. A rescue assay through the transduction of lentivirus expressing wild type ARV1 cDNA effectively rescued these alterations. This study expands the clinical and molecular spectrum of the ARV1‐related encephalopathy, confirming the essential role of ARV1 in GPI biosynthesis and brain function.
IntroductionDevelopmental and epileptic encephalopathies (DEEs) are rare epilepsy conditions that collectively impact 1 in 2000 children. They are highly genetically heterogeneous, resulting in ...significant barriers to accurate and adequate information for caregivers. This can lead to increased distress and dissatisfaction with the healthcare system. To address this gap, we developed ‘GenE Compass’ to provide caregivers with the highest-quality possible, understandable and relevant information in response to specific questions about their child’s DEE. Using a mixed-method design, we will now pilot GenE Compass to evaluate the acceptability to caregivers and clinicians, feasibility and impact to caregivers.Methods and analysisWe will recruit 88 caregivers (estimated final sample of 50 at follow-up) who have a child under 18 years of age with a suspected or confirmed DEE diagnosis. Following consent and a baseline questionnaire (questionnaire 1 (Q1)), participants will be able to submit questions to GenE Compass over a 3-month period. After 3 months, participants will complete a follow-up questionnaire (Q2) and an optional telephone interview to answer the research questions. Primary outcomes are acceptability of GenE Compass and feasibility of delivering the intervention (eg, cost of the intervention, number of questions submitted and time taken to respond to questions). Secondary outcomes include the impact of GenE Compass on caregivers’ quality of life, information searching behaviours, perceptions of their child’s illness and activation.Ethics and discussionThe study protocol (V.2, dated 16 September 2021) has been approved by the Sydney Children’s Hospitals Network Human Research Ethics Committee (ETH11277). The results will be disseminated in peer-reviewed journals and at scientific conferences. A lay summary will be disseminated to all participants.Trial registration numberACTRN12621001544864.
Background
Epileptic encephalopathies are a devastating group of neurological conditions in which etiological diagnosis can alter management and clinical outcome. Exome sequencing and gene panel ...testing can improve diagnostic yield but there is no cost‐effectiveness analysis of their use or consensus on how to best integrate these tests into clinical diagnostic pathways.
Methods
We conducted a retrospective cost‐effectiveness study comparing trio exome sequencing with a standard diagnostic approach, for a well‐phenotyped cohort of 32 patients with epileptic encephalopathy, who remained undiagnosed after “first‐tier” testing. Sensitivity analysis was included with a range of commercial exome and multigene panels.
Results
The diagnostic yield was higher for the exome sequencing (16/32; 50%) than the standard arm (2/32; 6.2%). The trio exome sequencing pathway was cost‐effective compared to the standard diagnostic pathway with a cost saving of AU$5,236 (95% confidence intervals $2,482; $9,784) per additional diagnosis; the standard pathway cost approximately 10 times more per diagnosis. Sensitivity analysis demonstrated that the majority of commercial exome sequencing and multigene panels studied were also cost‐effective. The clinical utility of all diagnoses was reported.
Conclusion
Our study supports the integration of exome sequencing and gene panel testing into the diagnostic pathway for epileptic encephalopathy, both in terms of cost effectiveness and clinical utility. We propose a diagnostic pathway that integrates initial rapid screening for treatable causes and comprehensive genomic screening. This study has important implications for health policy and public funding for epileptic encephalopathy and other neurological conditions.
This study uniquely compares the diagnostic yield, cost effectiveness, and clinical utility of the exome diagnostic model over the standard diagnostic model for patients with epileptic encephalopathy, who remain undiagnosed after first‐tier assessment. Our findings strongly support the exome diagnostic model, which resulted in a cost saving of AU$5,236 per additional diagnosis: approximately 10 times less expensive per diagnosis than the standard diagnostic model. We thus propose an optimal approach for the investigation of epileptic encephalopathy that benefits patients and their families by reducing the length and invasiveness of the “diagnostic odyssey” and provides a cost saving for the health budget.
Early infantile epileptic encephalopathies (EOEE) are a debilitating spectrum of disorders associated with cognitive impairments. We present a clinical report of a KCNT2 mutation in an EOEE patient. ...The de novo heterozygous variant Phe240Leu SLICK was identified by exome sequencing and confirmed by Sanger sequencing. Phe240Leu rSlick and hSLICK channels were electrophysiologically, heterologously characterized to reveal three significant alterations to channel function. First, Cl−i sensitivity was reversed in Phe240Leu channels. Second, predominantly K+-selective WT channels were made to favor Na+ over K+ by Phe240Leu. Third, and consequent to altered ion selectivity, Phe240Leu channels had larger inward conductance. Further, rSlick channels induced membrane hyperexcitability when expressed in primary neurons, resembling the cellular seizure phenotype. Taken together, our results confirm that Phe240Leu is a “change-of-function” KCNT2 mutation, demonstrating unusual altered selectivity in KNa channels. These findings establish pathogenicity of the Phe240Leu KCNT2 mutation in the reported EOEE patient.
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•We report a variant in the KNa channel KCNT2 linked to a human phenotype•Phe240Leu channels are no longer predominantly permissive to only K+•Expressing Phe240Leu in primary neurons induces membrane hyperexcitability•“Change-of-function” establishes pathogenicity of mutation in epileptic encephalopathy
Gururaj et al. report a KCNT2 mutation in a patient with epileptic encephalopathy and employ electrophysiological analyses to establish channel properties that could underlie epileptogenesis: namely, inhibition by high Cl−i and loss of exclusive selectivity to K+. Furthermore, primary neurons expressing Ph240Leu display a hyperexcitable phenotype.
We report an individual who presented with severe neurodevelopmental delay and an intractable infantile-onset seizure disorder. Exome sequencing identified a homozygous single nucleotide change that ...abolishes a splice donor site in the ARV1 gene (c.294 + 1G > A homozygous). This variant completely prevented splicing in minigene assays, and resulted in exon skipping and an in-frame deletion of 40 amino acids in primary human fibroblasts (NP_073623.1: p.(Lys59_Asn98del). The p.(Lys59_Asn98del) and previously reported p.(Gly189Arg) ARV1 variants were evaluated for protein expression and function. The p.(Gly189Arg) variant partially rescued the temperature-dependent growth defect in arv1Δ yeast, while p.(Lys59-Asn98del) completely failed to rescue at restrictive temperature. In contrast to wild type human ARV1, neither variant expressed detectable levels of protein in mammalian cells. Mice with a neuronal deletion of Arv1 recapitulated the human phenotype, exhibiting seizures and a severe survival defect in adulthood. Our data support ARV1 deficiency as a cause of autosomal recessive epileptic encephalopathy.
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