Genotype and phenotype correlation of PHACTR1-related neurological disorders.

BACKGROUND: PHACTR1 (phosphatase and actin regulators) plays a key role in cortical migration and synaptic activity by binding and regulating G-actin and PPP1CA. This study aimed to expand the genotype and phenotype of patients with de novo variants in PHACTR1 and analyse the impact of variants on protein-protein interaction. METHODS: We identified seven patients with PHACTR1 variants by trio-based whole-exome sequencing. Additional two subjects were ascertained from two centres through GeneMatcher. The genotype-phenotype correlation was determined, and AlphaFold-Multimer was used to predict protein-protein interactions and interfaces. RESULTS: Eight individuals carried missense variants and one had CNV in the PHACTR1. Infantile epileptic spasms syndrome (IESS) was the unifying phenotype in eight patients with missense variants of PHACTR1. They could present with other types of seizures and often exhibit drug-resistant epilepsy with a poor prognosis. One patient with CNV displayed a developmental encephalopathy phenotype. Using AlphaFold-Multimer, our findings indicate that PHACTR1 and G-actin-binding sequences overlap with PPP1CA at the RPEL3 domain, which suggests possible competition between PPP1CA and G-actin for binding to PHACTR1 through a similar polymerisation interface. In addition, patients carrying missense variants located at the PHACTR1-PPP1CA or PHACTR1-G-actin interfaces consistently exhibit the IESS phenotype. These missense variants are mostly concentrated in the overlapping sequence (RPEL3 domain). CONCLUSIONS: Patients with variants in PHACTR1 can have a phenotype of developmental encephalopathy in addition to IESS. Moreover, our study confirmed that the variants affect the binding of PHACTR1 to G-actin or PPP1CA, resulting in neurological disorders in patients.

SCN8A self-limited infantile epilepsy: Does epilepsy resolve?

SCN8A variants cause a spectrum of epilepsy phenotypes ranging from self-limited infantile epilepsy (SeLIE) to developmental and epileptic encephalopathy. SeLIE is an infantile onset focal epilepsy, occurring in developmentally normal infants, which often resolves by 3 years. Our aim was to ascertain when epilepsy resolves in SCN8A-SeLIE. We identified unpublished individuals with SCN8A-SeLIE and performed detailed phenotyping. Literature was searched for published SCN8A-SeLIE cases. Nine unpublished individuals from four families were identified (age at study = 3.5-66 years). Six had their last seizure after 3 years (range = 4-21 years); although drug-responsive and despite multiple weaning attempts (1-5), five of six remain on antiseizure medications (carbamazepine, n = 3; lamotrigine, n = 2). We identified 29 published individuals with SCN8A-SeLIE who had data on seizure progression. Of the 22 individuals aged at least 10 years, reported here or in the literature, nine of 22 (41%) had seizure offset prior to 3 years, five of 22 (23%) had seizure offset between 3 and 10 years, and eight of 22 (36%) had seizures after 10 years. Our data highlight that more than half of individuals with SCN8A-SeLIE continue to have seizures into late childhood. In contrast to SeLIE due to other etiologies, many individuals have a more persistent, albeit drug-responsive, form of epilepsy.

PIGN encephalopathy: Characterizing the epileptology.

OBJECTIVE: Epilepsy is common in patients with PIGN diseases due to biallelic variants; however, limited epilepsy phenotyping data have been reported. We describe the epileptology of PIGN encephalopathy. METHODS: We recruited patients with epilepsy due to biallelic PIGN variants and obtained clinical data regarding age at seizure onset/offset and semiology, development, medical history, examination, electroencephalogram, neuroimaging, and treatment. Seizure and epilepsy types were classified. RESULTS: Twenty six patients (13 female) from 26 families were identified, with mean age 7 years (range = 1 month to 21 years; three deceased). Abnormal development at seizure onset was present in 25 of 26. Developmental outcome was most frequently profound (14/26) or severe (11/26). Patients presented with focal motor (12/26), unknown onset motor (5/26), focal impaired awareness (1/26), absence (2/26), myoclonic (2/26), myoclonic-atonic (1/26), and generalized tonic-clonic (2/26) seizures. Twenty of 26 were classified as developmental and epileptic encephalopathy (DEE): 55% (11/20) focal DEE, 30% (6/20) generalized DEE, and 15% (3/20) combined DEE. Six had intellectual disability and epilepsy (ID+E): two generalized and four focal epilepsy. Mean age at seizure onset was 13 months (birth to 10 years), with a lower mean onset in DEE (7 months) compared with ID+E (33 months). Patients with DEE had drug-resistant epilepsy, compared to 4/6 ID+E patients, who were seizure-free. Hyperkinetic movement disorder occurred in 13 of 26 patients. Twenty-seven of 34 variants were novel. Variants were truncating (n = 7), intronic and predicted to affect splicing (n = 7), and missense or inframe indels (n = 20, of which 11 were predicted to affect splicing). Seven variants were recurrent, including p.Leu311Trp in 10 unrelated patients, nine with generalized seizures, accounting for nine of the 11 patients in this cohort with generalized seizures. SIGNIFICANCE: PIGN encephalopathy is a complex autosomal recessive disorder associated with a wide spectrum of epilepsy phenotypes, typically with substantial profound to severe developmental impairment.

Inherited RORB pathogenic variants: Overlap of photosensitive genetic generalized and occipital lobe epilepsy.

Variants in RORB have been reported in eight individuals with epilepsy, with phenotypes ranging from eyelid myoclonia with absence epilepsy to developmental and epileptic encephalopathies. We identified novel RORB variants in 11 affected individuals from four families. One was from whole genome sequencing and three were from RORB screening of three epilepsy cohorts: developmental and epileptic encephalopathies (n = 1021), overlap of generalized and occipital epilepsy (n = 84), and photosensitivity (n = 123). Following interviews and review of medical records, individuals' seizure and epilepsy syndromes were classified. Three novel missense variants and one exon 3 deletion were predicted to be pathogenic by in silico tools, not found in population databases, and located in key evolutionary conserved domains. Median age at seizure onset was 3.5 years (0.5-10 years). Generalized, predominantly absence and myoclonic, and occipital seizures were seen in all families, often within the same individual (6/11). All individuals with epilepsy were photosensitive, and seven of 11 had cognitive abnormalities. Electroencephalograms showed generalized spike and wave and/or polyspike and wave. Here we show a striking RORB phenotype of overlap of photosensitive generalized and occipital epilepsy in both individuals and families. This is the first report of a gene associated with this overlap of epilepsy syndromes.

The epileptology of GNB5 encephalopathy.

Pathogenic variants in GNB5 cause an autosomal recessive neurodevelopmental disorder with neonatal sinus bradycardia. Seizures or epilepsy occurred in 10 of 22 previously reported cases, including 6 children from one family. We delineate the epileptology of GNB5 encephalopathy. Our nine patients, including five new patients, were from seven families. Epileptic spasms were the most frequent seizure type, occurring in eight of nine patients, and began at a median age of 3 months (2 months to 3 years). Focal seizures preceded spasms in three children, with onset at 7 days, 11 days, and 4 months. One child presented with convulsive status epilepticus at 6 months. Three children had burst suppression on electroencephalography (EEG), three had hypsarrhythmia, and one evolved from burst suppression to hypsarrhythmia. Background slowing was present in all after age 3 years. Magnetic resonance imaging (MRI) showed cerebral atrophy in one child and cerebellar atrophy in another. All nine had abnormal development prior to seizure onset and ultimately had profound impairment without regression. Hypotonia was present in all, with contractures developing in two older patients. All individuals had biallelic pathogenic variants in GNB5, predicted by in silico tools to result in protein truncation and loss-of-function. GNB5 developmental and epileptic encephalopathy is characterized by epileptic spasms, focal seizures, and profound impairment.

Leveraging multiple approaches for the detection of pathogenic deep intronic variants in developmental and epileptic encephalopathies: A case report.

About 50% of individuals with developmental and epileptic encephalopathies (DEEs) are unsolved following genetic testing. Deep intronic variants, defined as >100 bp from exon-intron junctions, contribute to disease by affecting the splicing of mRNAs in clinically relevant genes. Identifying deep intronic pathogenic variants is challenging and resource intensive, and interpretation is difficult due to limited functional annotations. We aimed to identify deep intronic variants in individuals suspected to have unsolved single gene DEEs. In a research cohort of unsolved cases of DEEs, we searched for children with a DEE syndrome predominantly caused by variants in specific genes in >80% of described cases. We identified two children with Dravet syndrome and one individual with classic lissencephaly. Multiple sequencing and bioinformatics strategies were employed to interrogate intronic regions in SCN1A and PAFAH1B1. A novel de novo deep intronic 12 kb deletion in PAFAH1B1 was identified in the individual with lissencephaly. We showed experimentally that the deletion disrupts mRNA splicing, which results in partial intron retention after exon 2 and disruption of the highly conserved LisH motif. We demonstrate that targeted interrogation of deep intronic regions using multiple genomics technologies, coupled with functional analysis, can reveal hidden causes of unsolved monogenic DEE syndromes. PLAIN LANGUAGE SUMMARY: Deep intronic variants can cause disease by affecting the splicing of mRNAs in clinically relevant genes. A deep intronic deletion that caused abnormal splicing of the PAFAH1B1 gene was identified in a patient with classic lissencephaly. Our findings reinforce that targeted interrogation of deep intronic regions and functional analysis can reveal hidden causes of unsolved epilepsy syndromes.

Neurodevelopmental and Epilepsy Phenotypes in Individuals With Missense Variants in the Voltage-Sensing and Pore Domains of KCNH5.

BACKGROUND AND OBJECTIVES: KCNH5 encodes the voltage-gated potassium channel EAG2/Kv10.2. We aimed to delineate the neurodevelopmental and epilepsy phenotypic spectrum associated with de novo KCNH5 variants. METHODS: We screened 893 individuals with developmental and epileptic encephalopathies for KCNH5 variants using targeted or exome sequencing. Additional individuals with KCNH5 variants were identified through an international collaboration. Clinical history, EEG, and imaging data were analyzed; seizure types and epilepsy syndromes were classified. We included 3 previously published individuals including additional phenotypic details. RESULTS: We report a cohort of 17 patients, including 9 with a recurrent de novo missense variant p.Arg327His, 4 with a recurrent missense variant p.Arg333His, and 4 additional novel missense variants. All variants were located in or near the functionally critical voltage-sensing or pore domains, absent in the general population, and classified as pathogenic or likely pathogenic using the American College of Medical Genetics and Genomics criteria. All individuals presented with epilepsy with a median seizure onset at 6 months. They had a wide range of seizure types, including focal and generalized seizures. Cognitive outcomes ranged from normal intellect to profound impairment. Individuals with the recurrent p.Arg333His variant had a self-limited drug-responsive focal or generalized epilepsy and normal intellect, whereas the recurrent p.Arg327His variant was associated with infantile-onset DEE. Two individuals with variants in the pore domain were more severely affected, with a neonatal-onset movement disorder, early-infantile DEE, profound disability, and childhood death. DISCUSSION: We describe a cohort of 17 individuals with pathogenic or likely pathogenic missense variants in the voltage-sensing and pore domains of Kv10.2, including 14 previously unreported individuals. We present evidence for a putative emerging genotype-phenotype correlation with a spectrum of epilepsy and cognitive outcomes. Overall, we expand the role of EAG proteins in human disease and establish KCNH5 as implicated in a spectrum of neurodevelopmental disorders and epilepsy.

Infantile-onset myoclonic developmental and epileptic encephalopathy: A new RARS2 phenotype.

Recessive variants in RARS2, a nuclear gene encoding a mitochondrial protein, were initially reported in pontocerebellar hypoplasia. Subsequently, a recessive RARS2 early-infantile (<12 weeks) developmental and epileptic encephalopathy was described with hypoglycaemia and lactic acidosis. Here, we describe two unrelated patients with a novel RARS2 phenotype and reanalyse the published RARS2 epilepsy phenotypes and variants. Our novel cases had infantile-onset myoclonic developmental and epileptic encephalopathy, presenting with a progressive movement disorder from 9 months on a background of normal development. Development plateaued and regressed thereafter, with mild to profound impairment. Multiple drug-resistant generalized and focal seizures occurred with episodes of non-convulsive status epilepticus. Seizure types included absence, atonic, myoclonic, and focal seizures. Electroencephalograms showed diffuse slowing, multifocal, and generalised spike-wave activity, activated by sleep. Both patients had compound heterozygous RARS2 variants with likely impact on splicing and transcription. Remarkably, of the now 52 RARS2 variants reported in 54 patients, our reanalysis found that 44 (85%) have been shown to or are predicted to affect splicing or gene expression leading to protein truncation or nonsense-mediated decay. We expand the RARS2 phenotypic spectrum to include infantile encephalopathy and suggest this gene is enriched for pathogenic variants that disrupt splicing.

Evaluating the Genetics of Common Variable Immunodeficiency: Monogenetic Model and Beyond.

Common variable immunodeficiency (CVID) is the most frequent symptomatic primary immunodeficiency characterized by recurrent infections, hypogammaglobulinemia and poor response to vaccines. Its diagnosis is made based on clinical and immunological criteria, after exclusion of other diseases that can cause similar phenotypes. Currently, less than 20% of cases of CVID have a known underlying genetic cause. We have analyzed whole-exome sequencing and copy number variants data of 36 children and adolescents diagnosed with CVID and healthy relatives to estimate the proportion of monogenic cases. We have replicated an association of CVID to p.C104R in TNFRSF13B and reported the second case of homozygous patient to date. Our results also identify five causative genetic variants in LRBA, CTLA4, NFKB1, and PIK3R1, as well as other very likely causative variants in PRKCD, MAPK8, or DOCK8 among others. We experimentally validate the effect of the LRBA stop-gain mutation which abolishes protein production and downregulates the expression of CTLA4, and of the frameshift indel in CTLA4 producing expression downregulation of the protein. Our results indicate a monogenic origin of at least 15-24% of the CVID cases included in the study. The proportion of monogenic patients seems to be lower in CVID than in other PID that have also been analyzed by whole exome or targeted gene panels sequencing. Regardless of the exact proportion of CVID monogenic cases, other genetic models have to be considered for CVID. We propose that because of its prevalence and other features as intermediate penetrancies and phenotypic variation within families, CVID could fit with other more complex genetic scenarios. In particular, in this work, we explore the possibility of CVID being originated by an oligogenic model with the presence of heterozygous mutations in interacting proteins or by the accumulation of detrimental variants in particular immunological pathways, as well as perform association tests to detect association with rare genetic functional variation in the CVID cohort compared to healthy controls.

Genetic Load of Loss-of-Function Polymorphic Variants in Great Apes.

Loss of function (LoF) genetic variants are predicted to disrupt gene function, and are therefore expected to substantially reduce individual's viability. Knowing the genetic burden of LoF variants in endangered species is of interest for a better understanding of the effects of declining population sizes on species viability. In this study, we have estimated the number of LoF polymorphic variants in six great ape populations, based on whole-genome sequencing data in 79 individuals. Our results show that although the number of functional variants per individual is conditioned by the effective population size, the number of variants with a drastic phenotypic effect is very similar across species. We hypothesize that for those variants with high selection coefficients, differences in effective population size are not important enough to affect the efficiency of natural selection to remove them. We also describe that mostly CpG LoF mutations are shared across species, and an accumulation of LoF variants at olfactory receptor genes in agreement with its pseudogenization in humans and other primate species.