Genetic testing in children enrolled in epilepsy surgery program. A real-life study.

OBJECTIVE: Although genetic causes of drug-resistant focal epilepsy and selected focal malformations of cortical development (MCD) have been described, a limited number of studies comprehensively analysed genetic diagnoses in patients undergoing pre-surgical evaluation, their outcomes and the effect of genetic diagnosis on surgical strategy. METHODS: We analysed a prospective cohort of children enrolled in epilepsy surgery program over January 2018-July 2022. The majority of patients underwent germline and/or somatic genetic testing. We searched for predictors of surgical outcome and positive result of germline genetic testing. RESULTS: Ninety-five patients were enrolled in epilepsy surgery program and 64 underwent resective epilepsy surgery. We ascertained germline genetic diagnosis in 13/74 patients having underwent germline gene testing (pathogenic or likely pathogenic variants in CHRNA4, NPRL3, DEPDC5, FGF12, GRIA2, SZT2, STXBP1) and identified three copy number variants. Thirty-five patients underwent somatic gene testing; we detected 10 pathogenic or likely pathogenic variants in genes SLC35A2, PTEN, MTOR, DEPDC5, NPRL3. Germline genetic diagnosis was significantly associated with the diagnosis of focal epilepsy with unknown seizure onset. SIGNIFICANCE: Germline and somatic gene testing can ascertain a definite genetic diagnosis in a significant subgroup of patients in epilepsy surgery programs. Diagnosis of focal genetic epilepsy may tip the scales against the decision to proceed with invasive EEG study or surgical resection; however, selected patients with genetic focal epilepsies associated with MCD may benefit from resective epilepsy surgery and therefore, a genetic diagnosis does not disqualify patients from presurgical evaluation and epilepsy surgery.

Dominant KPNA3 Mutations Cause Infantile-Onset Hereditary Spastic Paraplegia.

OBJECTIVE: Hereditary spastic paraplegia (HSP) is a highly heterogeneous neurologic disorder characterized by lower-extremity spasticity. Here, we set out to determine the genetic basis of an autosomal dominant, pure, and infantile-onset form of HSP in a cohort of 8 patients with a uniform clinical presentation. METHODS: Trio whole-exome sequencing was used in 5 index patients with infantile-onset pure HSP to determine the genetic cause of disease. The functional impact of identified genetic variants was verified using bioinformatics and complementary cellular and biochemical assays. RESULTS: Distinct heterozygous KPNA3 missense variants were found to segregate with the clinical phenotype in 8 patients; in 4 of them KPNA3 variants had occurred de novo. Mutant karyopherin-α3 proteins exhibited a variable pattern of altered expression level, subcellular distribution, and protein interaction. INTERPRETATION: Our genetic findings implicate heterozygous variants in KPNA3 as a novel cause for autosomal dominant, early-onset, and pure HSP. Mutant karyopherin-α3 proteins display varying deficits in molecular and cellular functions, thus, for the first time, implicating dysfunctional nucleocytoplasmic shuttling as a novel pathomechanism causing HSP. ANN NEUROL 2021;90:738-750.

The Cause of Hereditary Hearing Loss in GJB2 Heterozygotes-A Comprehensive Study of the GJB2/DFNB1 Region.

Hearing loss is a genetically heterogeneous sensory defect, and the frequent causes are biallelic pathogenic variants in the GJB2 gene. However, patients carrying only one heterozygous pathogenic (monoallelic) GJB2 variant represent a long-lasting diagnostic problem. Interestingly, previous results showed that individuals with a heterozygous pathogenic GJB2 variant are two times more prevalent among those with hearing loss compared to normal-hearing individuals. This excess among patients led us to hypothesize that there could be another pathogenic variant in the GJB2 region/DFNB1 locus. A hitherto undiscovered variant could, in part, explain the cause of hearing loss in patients and would mean reclassifying them as patients with GJB2 biallelic pathogenic variants. In order to detect an unknown causal variant, we examined 28 patients using NGS with probes that continuously cover the 0.4 Mb in the DFNB1 region. An additional 49 patients were examined by WES to uncover only carriers. We did not reveal a second pathogenic variant in the DFNB1 region. However, in 19% of the WES-examined patients, the cause of hearing loss was found to be in genes other than the GJB2. We present evidence to show that a substantial number of patients are carriers of the GJB2 pathogenic variant, albeit only by chance.

Biallelic variants in HPDL cause pure and complicated hereditary spastic paraplegia.

Human 4-hydroxyphenylpyruvate dioxygenase-like (HPDL) is a putative iron-containing non-heme oxygenase of unknown specificity and biological significance. We report 25 families containing 34 individuals with neurological disease associated with biallelic HPDL variants. Phenotypes ranged from juvenile-onset pure hereditary spastic paraplegia to infantile-onset spasticity and global developmental delays, sometimes complicated by episodes of neurological and respiratory decompensation. Variants included bona fide pathogenic truncating changes, although most were missense substitutions. Functionality of variants could not be determined directly as the enzymatic specificity of HPDL is unknown; however, when HPDL missense substitutions were introduced into 4-hydroxyphenylpyruvate dioxygenase (HPPD, an HPDL orthologue), they impaired the ability of HPPD to convert 4-hydroxyphenylpyruvate into homogentisate. Moreover, three additional sets of experiments provided evidence for a role of HPDL in the nervous system and further supported its link to neurological disease: (i) HPDL was expressed in the nervous system and expression increased during neural differentiation; (ii) knockdown of zebrafish hpdl led to abnormal motor behaviour, replicating aspects of the human disease; and (iii) HPDL localized to mitochondria, consistent with mitochondrial disease that is often associated with neurological manifestations. Our findings suggest that biallelic HPDL variants cause a syndrome varying from juvenile-onset pure hereditary spastic paraplegia to infantile-onset spastic tetraplegia associated with global developmental delays.

GATOR1-related focal cortical dysplasia in epilepsy surgery patients and their families: A possible gradient in severity?

BACKGROUND: Variants of GATOR1-genes represent a recognised cause of focal cortical dysplasia (FCD), the most common structural aetiology in paediatric drug-resistant focal epilepsy. Reports on familial cases of GATOR1-associated FCD are limited, especially with respect to epilepsy surgery outcomes. METHODS: We present phenotypical manifestations of four unrelated patients with drug-resistant focal epilepsy, FCD and a first-degree relative with epilepsy. All patients underwent targeted gene panel sequencing as a part of the presurgical work up. Literature search was performed to compare our findings to previously published cases. RESULTS: The children (probands) had a more severe phenotype than their parents, including drug-resistant epilepsy and developmental delay, and they failed to achieve seizure freedom post-surgically. All patients had histopathologically confirmed FCD (types IIa, IIb, Ia). In Patient 1 and her affected father, we detected a known pathogenic NPRL2 variant. In patients 2 and 3 and their affected parents, we found novel likely pathogenic germline DEPDC5 variants. In family 4, we detected a novel variant in NPRL3. We identified 15 additional cases who underwent epilepsy surgery for GATOR1-associated FCD, with a positive family history of epilepsy in the literature; in 8/13 tested, the variant was inherited from an asymptomatic parent. CONCLUSION: The presented cases displayed a severity gradient in phenotype with children more severely affected than the parents. Although patients with GATOR1-associated FCD are considered good surgical candidates, post-surgical seizure outcome was poor in our familial cases, suggesting that accurate identification of the epileptogenic zone may be more challenging in this subgroup of patients.

Variant c.2158-2A>G in MANBA is an important and frequent cause of hereditary hearing loss and beta-mannosidosis among the Czech and Slovak Roma population- evidence for a new ethnic-specific variant.

BACKGROUND: The Roma are a European ethnic minority threatened by several recessive diseases. Variants in MANBA cause a rare lysosomal storage disorder named beta-mannosidosis whose clinical manifestation includes deafness and mental retardation. Since 1986, only 23 patients with beta-mannosidosis and biallelic MANBA variants have been described worldwide. RESULTS: We now report on further 10 beta-mannosidosis patients of Roma origin from eight families in the Czech and Slovak Republics with hearing loss, mental retardation and homozygous pathogenic variants in MANBA. MANBA variant c.2158-2A>G screening among 345 anonymized normal hearing controls from Roma populations revealed a carrier/heterozygote frequency of 3.77%. This is about 925 times higher than the frequency of this variant in the gnomAD public database and classifies the c.2158-2A>G variant as a prevalent, ethnic-specific variant causing hearing loss and mental retardation in a homozygous state. The frequency of heterozygotes/carriers is similar to another pathogenic variant c.71G>A (p.W24*) in GJB2, regarded as the most frequent variant causing deafness in Roma populations. CONLCUSION: Beta-mannosidosis, due to a homozygous c.2158-2A>G MANBA variant, is an important and previously unknown cause of hearing loss and mental retardation among Central European Roma.

Autosomal recessive hereditary spastic paraplegia type SPG35 due to a novel variant in the FA2H gene in a Czech patient.

Biallelic pathogenic variants in FA2H gene have been repeatedly described as a cause of hereditary spastic paraplegia (HSP) type35 (SPG35). Targeted massive parallel sequencing (MPS) of the HSP genes panel revealed a novel homozygous variant c.130C > T (p.P44S) in the FA2H gene in the 30-year-old patient presenting with spastic paraplegia. The patient originated form the Czech minority in Romania. The patient manifests typical clinical signs for SPG35 (youth onset gait impairment, progressive spastic paraparesis on lower limbs, dysarthria, white matter changes in MRI).