Publications by authors named "Renzo Guerrini"

448 Publications

Multicenter Validation of a Deep Learning Detection Algorithm for Focal Cortical Dysplasia.

Neurology 2021 Sep 14. Epub 2021 Sep 14.

Neuroimaging of Epilepsy Laboratory, Montreal Neurological Institute, McGill University, Montreal, Canada

Objective: To test the hypothesis that a multicenter-validated computer deep learning algorithm detects MRI-negative focal cortical dysplasia (FCD).

Methods: We used clinically-acquired 3D T1-weighted and 3D FLAIR MRI of 148 patients (median age, 23 years [range, 2-55]; 47% female) with histologically-verified FCD at nine centers to train a deep convolutional neural network (CNN) classifier. Images were initially deemed as MRI-negative in 51% of cases, in whom intracranial EEG determined the focus. For risk stratification, the CNN incorporated Bayesian uncertainty estimation as a measure of confidence. To evaluate performance, detection maps were compared to expert FCD manual labels. Sensitivity was tested in an independent cohort of 23 FCD cases (13±10 years). Applying the algorithm to 42 healthy and 89 temporal lobe epilepsy disease controls tested specificity.

Results: Overall sensitivity was 93% (137/148 FCD detected) using a leave-one-site-out cross-validation, with an average of six false positives per patient. Sensitivity in MRI-negative FCD was 85%. In 73% of patients, the FCD was among the clusters with the highest confidence; in half it ranked the highest. Sensitivity in the independent cohort was 83% (19/23; average of five false positives per patient). Specificity was 89% in healthy and disease controls.

Conclusions: This first multicenter-validated deep learning detection algorithm yields the highest sensitivity to date in MRI-negative FCD. By pairing predictions with risk stratification this classifier may assist clinicians to adjust hypotheses relative to other tests, increasing diagnostic confidence. Moreover, generalizability across age and MRI hardware makes this approach ideal for pre-surgical evaluation of MRI-negative epilepsy.

Classification Of Evidence: This study provides Class III evidence that deep learning on multimodal MRI accurately identifies FCD in epilepsy patients initially diagnosed as MRI-negative.
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http://dx.doi.org/10.1212/WNL.0000000000012698DOI Listing
September 2021

Monogenic Epilepsies: Disease Mechanisms, Clinical Phenotypes, and Targeted Therapies.

Neurology 2021 Sep 7. Epub 2021 Sep 7.

Department of Clinical and Experimental Epilepsy, UCL Queen Square Institute of Neurology, London, UK.

A monogenic aetiology can be identified in up to 40% of people with severe epilepsy. To address earlier and more appropriate treatment strategies, clinicians are required to know the implications that specific genetic causes might have on pathophysiology, natural history, comorbidities and treatment choices. In this narrative review, we summarise concepts on the genetic epilepsies based on the underlying pathophysiological mechanisms and present the current knowledge on treatment options based on evidence provided by controlled trials or studies with lower classification of evidence. Overall, evidence robust enough to guide antiseizure medication (ASM) choices in genetic epilepsies remains limited to the more frequent conditions for which controlled trials and observational studies have been possible. Most monogenic disorders are very rare and ASM choices for them are still based on inferences drawn from observational studies and early, often anecdotal, experiences with precision therapies. Precision medicine remains applicable to only a narrow number of patients with monogenic epilepsies and may target only part of the actual functional defects. Phenotypic heterogeneity is remarkable, and some genetic mutations activate epileptogenesis through their developmental effects, which may not be reversed postnatally. Other genes seem to have pure functional consequences on excitability, acting through either loss- or gain-of-function effects, and these may have opposite treatment implications. In addition, the functional consequences of missense mutations may be difficult to predict, making precision treatment approaches considerably more complex than estimated by deterministic interpretations. Knowledge of genetic aetiologies can influence the approach to surgical treatment of focal epilepsies. Identification of germline mutations in specific genes contraindicates surgery while mutations in other genes do not. Identification, quantification and functional characterization of specific somatic mutations before surgery using cerebrospinal fluid liquid biopsy or after surgery in brain specimens, will likely be integrated in planning surgical strategies and re-intervention after a first unsuccessful surgery as initial evidence suggests that mutational load may correlate with the epileptogenic zone. Promising future directions include gene manipulation by DNA or mRNA targeting; although most are still far from clinical use, some are in early phase clinical development.
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http://dx.doi.org/10.1212/WNL.0000000000012744DOI Listing
September 2021

Monoallelic KIF1A-related disorders: a multicenter cross sectional study and systematic literature review.

J Neurol 2021 Sep 6. Epub 2021 Sep 6.

Neurofisiopathology Unit, Fondazione Policlinico Universitario Agostino Gemelli IRCCS, 00168, Rome, Italy.

Background: Monoallelic variants in the KIF1A gene are associated with a large set of clinical phenotypes including neurodevelopmental and neurodegenerative disorders, underpinned by a broad spectrum of central and peripheral nervous system involvement.

Methods: In a multicenter study conducted in patients presenting spastic gait or complex neurodevelopmental disorders, we analyzed the clinical, genetic and neuroradiological features of 28 index cases harboring heterozygous variants in KIF1A. We conducted a literature systematic review with the aim to comparing our findings with previously reported KIF1A-related phenotypes.

Results: Among 28 patients, we identified nine novel monoallelic variants, and one a copy number variation encompassing KIF1A. Mutations arose de novo in most patients and were prevalently located in the motor domain. Most patients presented features of a continuum ataxia-spasticity spectrum with only five cases showing a prevalently pure spastic phenotype and six presenting congenital ataxias. Seventeen mutations occurred in the motor domain of the Kinesin-1A protein, but location of mutation did not correlate with neurological and imaging presentations. When tested in 15 patients, muscle biopsy showed oxidative metabolism alterations (6 cases), impaired respiratory chain complexes II + III activity (3/6) and low CoQ10 levels (6/9). Ubiquinol supplementation (1gr/die) was used in 6 patients with subjective benefit.

Conclusions: This study broadened our clinical, genetic, and neuroimaging knowledge of KIF1A-related disorders. Although highly heterogeneous, it seems that manifestations of ataxia-spasticity spectrum disorders seem to occur in most patients. Some patients also present secondary impairment of oxidative metabolism; in this subset, ubiquinol supplementation therapy might be appropriate.
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http://dx.doi.org/10.1007/s00415-021-10792-3DOI Listing
September 2021

Posterior Lissencephaly Associated with Subcortical Band Heterotopia Due to a Variation in the Gene: A Case Report and Refining of the Phenotypic Spectrum.

Genes (Basel) 2021 Aug 5;12(8). Epub 2021 Aug 5.

Medical Genetics Unit, Azienda USL-IRCCS di Reggio Emilia, 42123 Reggio Emilia, Italy.

Lissencephaly describes a group of conditions characterized by the absence of normal cerebral convolutions and abnormalities of cortical development. To date, at least 20 genes have been identified as involved in the pathogenesis of this condition. Variants in , encoding a protein involved in the regulation of neuronal migration, have been recently described as causative of lissencephaly with a posterior-prevalent involvement of the cerebral cortex and an autosomal dominant pattern of inheritance. Here, we describe a 3-year-old boy with slightly delayed psychomotor development and mild dysmorphic features, including bitemporal narrowing, protruding ears with up-lifted lobes and posterior plagiocephaly. Brain MRI at birth identified type 1 lissencephaly, prevalently in the temporo-occipito-parietal regions of both hemispheres with "double-cortex" (Dobyns' 1-2 degree) periventricular band alterations. Whole-exome sequencing revealed a previously unreported de novo pathogenic variant in the gene (NM_001042475.3:c.232+1del). Only 20 patients have been reported as carriers of pathogenic variants to date. They show lissencephaly with prevalent posterior involvement, variable cognitive deficits and epilepsy. The present case report indicates the clinical variability associated with variants that are not invariantly associated with severe phenotypes and poor outcome, and underscores the importance of including this gene in diagnostic panels for lissencephaly.
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http://dx.doi.org/10.3390/genes12081208DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8391275PMC
August 2021

Genotype-phenotype correlations in SCN8A-related disorders reveal prognostic and therapeutic implications.

Brain 2021 Aug 25. Epub 2021 Aug 25.

National Centre for Rare Epilepsy-Related Disorders, Oslo University Hospital, 0001 Oslo, Norway.

We report detailed functional analyses and genotype-phenotype correlations in 392 individuals carrying disease-causing variants in SCN8A, encoding the voltage-gated Na+ channel NaV1.6, with the aim of describing clinical phenotypes related to functional effects. Six different clinical subgroups could be identified: 1) Benign familial infantile epilepsy (BFIE) (n = 15, normal cognition, treatable seizures), 2) intermediate epilepsy (n = 33, mild ID, partially pharmaco-responsive), 3) developmental and epileptic encephalopathy (DEE, n = 177, severe ID, majority pharmaco-resistant), 4) generalized epilepsy (n = 20, mild to moderate ID, frequently with absence seizures), 5) unclassifiable epilepsy (n = 127), and 6) neurodevelopmental disorder without epilepsy (n = 20, mild to moderate ID). Groups 1-3 presented with focal or multifocal seizures (median age of onset: four months) and focal epileptiform discharges, whereas the onset of seizures in group 4 was later (median: 42 months) with generalized epileptiform discharges. We performed functional studies expressing missense variants in ND7/23 neuroblastoma cells and primary neuronal cultures using recombinant tetrodotoxin-insensitive human NaV1.6 channels and whole-cell patch-clamping. Two variants causing DEE showed a strong gain-of-function (GOF, hyperpolarising shift of steady-state activation, strongly increased neuronal firing rate), and one variant causing BFIE or intermediate epilepsy showed a mild GOF (defective fast inactivation, less increased firing). In contrast, all three variants causing generalized epilepsy induced a loss-of-function (LOF, reduced current amplitudes, depolarising shift of steady-state activation, reduced neuronal firing). Including previous studies, functional effects were known for 170 individuals. All 136 individuals carrying a functionally tested GOF variant had either focal (97, groups 1-3), or unclassifiable epilepsy (39), whereas 34 with a LOF variant had either generalized (14), no (11) or unclassifiable (6) epilepsy; only three had DEE. Computational modeling in the GOF group revealed a significant correlation between the severity of the electrophysiological and clinical phenotypes. GOF variant carriers responded significantly better to sodium channel blockers (SCBs) than to other anti-seizure medications, and the same applied for all individuals of groups 1-3. In conclusion, our data reveal clear genotype-phenotype correlations between age at seizure onset, type of epilepsy and gain- or loss-of-function effects of SCN8A variants. Generalized epilepsy with absence seizures is the main epilepsy phenotype of LOF variant carriers and the extent of the electrophysiological dysfunction of the GOF variants is a main determinant of the severity of the clinical phenotype in focal epilepsies. Our pharmacological data indicate that SCBs present a treatment option in SCN8A-related focal epilepsy with onset in the first year of life.
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http://dx.doi.org/10.1093/brain/awab321DOI Listing
August 2021

Clinical and molecular delineation of PUS3-associated neurodevelopmental disorders.

Clin Genet 2021 Aug 20. Epub 2021 Aug 20.

Kennedy Center, Department of Clinical Genetics, Copenhagen University Hospital, Copenhagen, Denmark.

Biallelic variants in PUS3 have recently been recognized as a rare cause of neurodevelopmental disorders. Pseudouridine synthase-3 encoded by PUS3 is an enzyme important for modification of various RNAs, including transfer RNA (tRNA). Here we present the clinical and genetic features of 21 individuals with biallelic PUS3 variants: seven new and 14 previously reported individuals, where clinical features of two were updated. The clinical and genetic information were collected through collaborations or by literature search. All individuals were characterized by the local clinicians and the gene variants were identified by next generation sequencing (NGS) based methodologies. The clinical picture was dominated by global developmental delay, epilepsy, hypotonia and microcephaly. Gray sclera, which has previously been suggested to be a characteristic feature of PUS3-associated phenotypes, was reported in only seven individuals. The patients had some dysmorphic facial features, but a recognizable gestalt was not observed. In conclusion, homozygous and compound heterozygous PUS3 variants lead to a rare neurodevelopmental disorder. Further functional studies are necessary to understand involvement of PUS3 and tRNA biogenesis in normal and abnormal brain development.
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http://dx.doi.org/10.1111/cge.14051DOI Listing
August 2021

A systems-level analysis highlights microglial activation as a modifying factor in common epilepsies.

Neuropathol Appl Neurobiol 2021 Aug 13. Epub 2021 Aug 13.

Neuroscience Research Center, Department of Medical and Surgical Sciences, University "Magna Graecia" of Catanzaro, Catanzaro, Italy.

Aims: The causes of distinct patterns of reduced cortical thickness in the common human epilepsies, detectable on neuroimaging and with important clinical consequences, are unknown. We investigated the underlying mechanisms of cortical thinning using a systems-level analysis.

Methods: Imaging-based cortical structural maps from a large-scale epilepsy neuroimaging study were overlaid with highly spatially resolved human brain gene expression data from the Allen Human Brain Atlas. Cell-type deconvolution, differential expression analysis and cell-type enrichment analyses were used to identify differences in cell-type distribution. These differences were followed up in post-mortem brain tissue from humans with epilepsy using Iba1 immunolabelling. Furthermore, to investigate a causal effect in cortical thinning, cell-type-specific depletion was used in a murine model of acquired epilepsy.

Results: We identified elevated fractions of microglia and endothelial cells in regions of reduced cortical thickness. Differentially expressed genes showed enrichment for microglial markers and, in particular, activated microglial states. Analysis of post-mortem brain tissue from humans with epilepsy confirmed excess activated microglia. In the murine model, transient depletion of activated microglia during the early phase of the disease development prevented cortical thinning and neuronal cell loss in the temporal cortex. Although the development of chronic seizures was unaffected, the epileptic mice with early depletion of activated microglia did not develop deficits in a non-spatial memory test seen in epileptic mice not depleted of microglia.

Conclusions: These convergent data strongly implicate activated microglia in cortical thinning, representing a new dimension for concern and disease modification in the epilepsies, potentially distinct from seizure control.
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http://dx.doi.org/10.1111/nan.12758DOI Listing
August 2021

De novo DHDDS variants cause a neurodevelopmental and neurodegenerative disorder with myoclonus.

Brain 2021 Aug 11. Epub 2021 Aug 11.

Undiagnosed Diseases Program, National Institutes of Health, Bethesda, MD 20892-2152, USA.

Subcellular membrane systems are highly enriched in dolichol, whose role in organelle homeostasis and endosomal-lysosomal pathway remains largely unclear besides being involved in protein glycosylation. DHDDS encodes for the catalytic subunit (DHDDS) of the enzyme cis-prenyltransferase (cis-PTase), involved in dolichol biosynthesis and dolichol-dependent protein glycosylation in the endoplasmic reticulum. An autosomal recessive form of retinitis pigmentosa (retinitis pigmentosa 59) has been associated with a recurrent DHDDS variant. Moreover, two recurring de novo substitutions were detected in a few cases presenting with neurodevelopmental disorder, epilepsy, and movement disorder. We evaluated a large cohort of patients (n=25) with de novo pathogenic variants in DHDDS and provided the first systematic description of the clinical features and long-term outcome of this new neurodevelopmental and neurodegenerative disorder. The functional impact of the identified variants was explored by yeast complementation system and enzymatic assay. Patients presented during infancy or childhood with a variable association of neurodevelopmental disorder, generalized epilepsy, action myoclonus/cortical tremor, and ataxia. Later in the disease course they experienced a slow neurological decline with the emergence of hyperkinetic and/or hypokinetic movement disorder, cognitive deterioration, and psychiatric disturbances. Storage of lipidic material and altered lysosomes were detected in myelinated fibers and fibroblasts, suggesting a dysfunction of the lysosomal enzymatic scavenger machinery. Serum glycoprotein hypoglycosylation was not detected and, in contrast to retinitis pigmentosa and other congenital disorders of glycosylation involving dolichol metabolism, the urinary dolichol D18/D19 ratio was normal. Mapping the disease-causing variants into the protein structure revealed that most of them clustered around the active site of the DHDDS subunit. Functional studies using yeast complementation assay and in vitro activity measurements confirmed that these changes affected the catalytic activity of the cis-PTase and showed growth defect in yeast complementation system as compared with the wild-type enzyme and retinitis pigmentosa-associated protein. In conclusion, we characterized a distinctive neurodegenerative disorder due to de novo DHDDS variants, which clinically belongs to the spectrum of genetic progressive encephalopathies with myoclonus. Clinical and biochemical data from this cohort depicted a condition at the intersection of congenital disorders of glycosylation and inherited storage diseases with several features akin to of progressive myoclonus epilepsy such as neuronal ceroid lipofuscinosis and other lysosomal disorders.
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http://dx.doi.org/10.1093/brain/awab299DOI Listing
August 2021

The Diagnostic Approach to Mitochondrial Disorders in Children in the Era of Next-Generation Sequencing: A 4-Year Cohort Study.

J Clin Med 2021 Jul 22;10(15). Epub 2021 Jul 22.

Department of Molecular Medicine, IRCCS Stella Maris Foundation, 56128 Pisa, Italy.

Mitochondrial diseases (MDs) are a large group of genetically determined multisystem disorders, characterized by extreme phenotypic heterogeneity, attributable in part to the dual genomic control (nuclear and mitochondrial DNA) of the mitochondrial proteome. Advances in next-generation sequencing technologies over the past two decades have presented clinicians with a challenge: to select the candidate disease-causing variants among the huge number of data provided. Unfortunately, the clinical tools available to support genetic interpretations still lack specificity and sensitivity. For this reason, the diagnosis of MDs continues to be difficult, with the new "genotype first" approach still failing to diagnose a large group of patients. With the aim of investigating possible relationships between clinical and/or biochemical phenotypes and definitive molecular diagnoses, we performed a retrospective multicenter study of 111 pediatric patients with clinical suspicion of MD. In this cohort, the strongest predictor of a molecular (in particular an mtDNA-related) diagnosis of MD was neuroimaging evidence of basal ganglia (BG) involvement. Regression analysis confirmed that normal BG imaging predicted negative genetic studies for MD. Psychomotor regression was confirmed as an independent predictor of a definitive diagnosis of MD. The findings of this study corroborate previous data supporting a role for neuroimaging in the diagnostic approach to MDs and reinforce the idea that mtDNA sequencing should be considered for first-line testing, at least in specific groups of children.
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http://dx.doi.org/10.3390/jcm10153222DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8348083PMC
July 2021

Delineating the molecular and phenotypic spectrum of the SETD1B-related syndrome.

Genet Med 2021 Aug 3. Epub 2021 Aug 3.

Department of Clinical and Experimental Epilepsy, UCL Queen Square Institute of Neurology, London, UK.

Purpose: Pathogenic variants in SETD1B have been associated with a syndromic neurodevelopmental disorder including intellectual disability, language delay, and seizures. To date, clinical features have been described for 11 patients with (likely) pathogenic SETD1B sequence variants. This study aims to further delineate the spectrum of the SETD1B-related syndrome based on characterizing an expanded patient cohort.

Methods: We perform an in-depth clinical characterization of a cohort of 36 unpublished individuals with SETD1B sequence variants, describing their molecular and phenotypic spectrum. Selected variants were functionally tested using in vitro and genome-wide methylation assays.

Results: Our data present evidence for a loss-of-function mechanism of SETD1B variants, resulting in a core clinical phenotype of global developmental delay, language delay including regression, intellectual disability, autism and other behavioral issues, and variable epilepsy phenotypes. Developmental delay appeared to precede seizure onset, suggesting SETD1B dysfunction impacts physiological neurodevelopment even in the absence of epileptic activity. Males are significantly overrepresented and more severely affected, and we speculate that sex-linked traits could affect susceptibility to penetrance and the clinical spectrum of SETD1B variants.

Conclusion: Insights from this extensive cohort will facilitate the counseling regarding the molecular and phenotypic landscape of newly diagnosed patients with the SETD1B-related syndrome.
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http://dx.doi.org/10.1038/s41436-021-01246-2DOI Listing
August 2021

Artificial intelligence for classification of temporal lobe epilepsy with ROI-level MRI data: A worldwide ENIGMA-Epilepsy study.

Neuroimage Clin 2021 24;31:102765. Epub 2021 Jul 24.

Institute for Diagnostic Radiology and Neuroradiology, University Medicine Greifswald, Greifswald, Germany; Institute for Diagnostic and Interventional Radiology, Pediatric and Neuroradiology, University Medical Centre Rostock, Rostock, Germany.

Artificial intelligence has recently gained popularity across different medical fields to aid in the detection of diseases based on pathology samples or medical imaging findings. Brain magnetic resonance imaging (MRI) is a key assessment tool for patients with temporal lobe epilepsy (TLE). The role of machine learning and artificial intelligence to increase detection of brain abnormalities in TLE remains inconclusive. We used support vector machine (SV) and deep learning (DL) models based on region of interest (ROI-based) structural (n = 336) and diffusion (n = 863) brain MRI data from patients with TLE with ("lesional") and without ("non-lesional") radiographic features suggestive of underlying hippocampal sclerosis from the multinational (multi-center) ENIGMA-Epilepsy consortium. Our data showed that models to identify TLE performed better or similar (68-75%) compared to models to lateralize the side of TLE (56-73%, except structural-based) based on diffusion data with the opposite pattern seen for structural data (67-75% to diagnose vs. 83% to lateralize). In other aspects, structural and diffusion-based models showed similar classification accuracies. Our classification models for patients with hippocampal sclerosis were more accurate (68-76%) than models that stratified non-lesional patients (53-62%). Overall, SV and DL models performed similarly with several instances in which SV mildly outperformed DL. We discuss the relative performance of these models with ROI-level data and the implications for future applications of machine learning and artificial intelligence in epilepsy care.
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http://dx.doi.org/10.1016/j.nicl.2021.102765DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8346685PMC
September 2021

Large-scale, cell-resolution volumetric mapping allows layer-specific investigation of human brain cytoarchitecture.

Biomed Opt Express 2021 Jun 27;12(6):3684-3699. Epub 2021 May 27.

European Laboratory for Non-Linear Spectroscopy (LENS), University of Florence, Sesto Fiorentino, Italy.

Although neuronal density analysis on human brain slices is available from stereological studies, data on the spatial distribution of neurons in 3D are still missing. Since the neuronal organization is very inhomogeneous in the cerebral cortex, it is critical to map all neurons in a given volume rather than relying on sparse sampling methods. To achieve this goal, we implement a new tissue transformation protocol to clear and label human brain tissues and we exploit the high-resolution optical sectioning of two-photon fluorescence microscopy to perform 3D mesoscopic reconstruction. We perform neuronal mapping of 100mm human brain samples and evaluate the volume and density distribution of neurons from various areas of the cortex originating from different subjects (young, adult, and elderly, both healthy and pathological). The quantitative evaluation of the density in combination with the mean volume of the thousands of neurons identified within the specimens, allow us to determine the layer-specific organization of the cerebral architecture.
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http://dx.doi.org/10.1364/BOE.415555DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8221968PMC
June 2021

PRDX1 gene-related epi-cblC disease is a common type of inborn error of cobalamin metabolism with mono- or bi-allelic MMACHC epimutations.

Clin Epigenetics 2021 Jul 2;13(1):137. Epub 2021 Jul 2.

Molecular and Cell Biology Laboratory of Neurometabolic Diseases, Paediatric Neurology Unit and Laboratories, Meyer Children's Hospital, Viale Pieraccini 24, 50139, Florence, Italy.

Background: The role of epigenetics in inborn errors of metabolism (IEMs) is poorly investigated. Epigenetic changes can contribute to clinical heterogeneity of affected patients but could also be underestimated determining factors in the occurrence of IEMs. An epigenetic cause of IEMs has been recently described for the autosomal recessive methylmalonic aciduria and homocystinuria, cblC type (cblC disease), and it has been named epi-cblC. Epi-cblC has been reported in association with compound heterozygosity for a genetic variant and an epimutation at the MMACHC locus, which is secondary to a splicing variant (c.515-1G > T or c.515-2A > T) at the adjacent PRDX1 gene. Both these variants cause aberrant antisense transcription and cis-hypermethylation of the MMACHC gene promotor with subsequent silencing. Until now, only nine epi-cblC patients have been reported.

Methods: We report clinical/biochemical assessment, MMACHC/PRDX1 gene sequencing and genome-wide DNA methylation profiling in 11 cblC patients who had an inconclusive MMACHC gene testing. We also compare clinical phenotype of epi-cblC patients with that of canonical cblC patients.

Results: All patients turned out to have the epi-cblC disease. One patient had a bi-allelic MMACHC epimutation due to the homozygous PRDX1:c.515-1G > T variant transmitted by both parents. We found that the bi-allelic epimutation produces the complete silencing of MMACHC in the patient's fibroblasts. The remaining ten patients had a mono-allelic MMACHC epimutation, due to the heterozygous PRDX1:c.515-1G > T, in association with a mono-allelic MMACHC genetic variant. Epi-cblC disease has accounted for about 13% of cblC cases diagnosed by newborn screening in the Tuscany and Umbria regions since November 2001. Comparative analysis showed that clinical phenotype of epi-cblC patients is similar to that of canonical cblC patients.

Conclusions: We provide evidence that epi-cblC is an underestimated cause of inborn errors of cobalamin metabolism and describe the first instance of epi-cblC due to a bi-allelic MMACHC epimutation. MMACHC epimutation/PRDX1 mutation analyses should be part of routine genetic testing for all patients presenting with a metabolic phenotype that combines methylmalonic aciduria and homocystinuria.
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http://dx.doi.org/10.1186/s13148-021-01117-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8254308PMC
July 2021

Whole Exome Sequencing Is the Minimal Technological Approach in Probands Born to Consanguineous Couples.

Genes (Basel) 2021 Jun 24;12(7). Epub 2021 Jun 24.

Medical Genetics Unit, Azienda USL-IRCCS di Reggio Emilia, 42123 Reggio Emilia, Italy.

We report on two siblings suffering from different pathogenic conditions, born to consanguineous parents. A multigene panel for brain malformations and microcephaly identified the homozygous splicing variant NM_005886.3:c.1416+1del in the gene in the older sister. On the other hand, exome sequencing revealed the homozygous frameshift variant NM_005245.4:c.9729del in the gene in the younger sister, who had a more complex phenotype: in addition to bilateral anophthalmia and heart defects, she showed a right split foot with 4 toes, 5 metacarpals, second toe duplication and preaxial polydactyly on the right hand. These features have been never reported before in patients with pathogenic variants and support the role of this gene in the development of limb buds. Notably, each parent was heterozygous for both of these variants, which were ultra-rare and rare, respectively. This study raises awareness about the value of using whole exome/genome sequencing rather than targeted gene panels when testing affected offspring born to consanguineous couples. In this way, exomic data from the parents are also made available for carrier screening, to identify heterozygous pathogenetic and likely pathogenetic variants in genes responsible for other recessive conditions, which may pose a risk for subsequent pregnancies.
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http://dx.doi.org/10.3390/genes12070962DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8303193PMC
June 2021

Pathogenic MAST3 Variants in the STK Domain Are Associated with Epilepsy.

Ann Neurol 2021 Aug 13;90(2):274-284. Epub 2021 Jul 13.

Epilepsy Research Centre, Department of Medicine, Austin Health, The University of Melbourne, Heidelberg, VIC, Australia.

Objective: The MAST family of microtubule-associated serine-threonine kinases (STKs) have distinct expression patterns in the developing and mature human and mouse brain. To date, only MAST1 has been conclusively associated with neurological disease, with de novo variants in individuals with a neurodevelopmental disorder, including a mega corpus callosum.

Methods: Using exome sequencing, we identify MAST3 missense variants in individuals with epilepsy. We also assess the effect of these variants on the ability of MAST3 to phosphorylate the target gene product ARPP-16 in HEK293T cells.

Results: We identify de novo missense variants in the STK domain in 11 individuals, including 2 recurrent variants p.G510S (n = 5) and p.G515S (n = 3). All 11 individuals had developmental and epileptic encephalopathy, with 8 having normal development prior to seizure onset at <2 years of age. All patients developed multiple seizure types, 9 of 11 patients had seizures triggered by fever and 9 of 11 patients had drug-resistant seizures. In vitro analysis of HEK293T cells transfected with MAST3 cDNA carrying a subset of these patient-specific missense variants demonstrated variable but generally lower expression, with concomitant increased phosphorylation of the MAST3 target, ARPP-16, compared to wild-type. These findings suggest the patient-specific variants may confer MAST3 gain-of-function. Moreover, single-nuclei RNA sequencing and immunohistochemistry shows that MAST3 expression is restricted to excitatory neurons in the cortex late in prenatal development and postnatally.

Interpretation: In summary, we describe MAST3 as a novel epilepsy-associated gene with a potential gain-of-function pathogenic mechanism that may be primarily restricted to excitatory neurons in the cortex. ANN NEUROL 2021;90:274-284.
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http://dx.doi.org/10.1002/ana.26147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8324566PMC
August 2021

Bi-allelic variants in MTMR5/SBF1 cause Charcot-Marie-Tooth type 4B3 featuring mitochondrial dysfunction.

BMC Med Genomics 2021 Jun 12;14(1):157. Epub 2021 Jun 12.

Department of Medical Basic Sciences, Neurosciences and Sense Organs, University of Bari Aldo Moro, Piazza G. Cesare, 11, 70124, Bari, Italy.

Background: Charcot-Marie-Tooth disease (CMT) type 4B3 (CMT4B3) is a rare form of genetic neuropathy associated with variants in the MTMR5/SBF1 gene. MTMR5/SBF1 is a pseudophosphatase predicted to regulate endo-lysosomal trafficking in tandem with other MTMRs. Although almost ubiquitously expressed, pathogenic variants primarily impact on the peripheral nervous system, corroborating the involvement of MTMR5/SBF1 and its molecular partners in Schwann cells-mediated myelinization.

Case Presentation: We report a case of severe CMT4B3 characterized by early-onset motor and axonal polyneuropathy in an Italian child in absence of any evidence of brain and spine MRI abnormalities or intellectual disability and with a biochemical profile suggestive of mitochondrial disease. Using an integrated approach combining both NGS gene panels and WES analysis, we identified two novel compound heterozygous missense variants in MTMR5/SBF1 gene, p.R763H (c.2291G > A) and p.G1064E (c.3194G > A). Studies in muscle identified partial defects of oxidative metabolism.

Conclusion: We describe the first case of an early onset severe polyneuropathy with motor and axonal involvement, due to recessive variants in the MTMR5/SBF1 gene, with no evidence of brain and spine MRI abnormalities, intellectual disability, no clinical and neurophysiological evidences of distal sensory impairment, and rapid neuromuscular deterioration. This report suggests that MTMR5/SBF1 should be considered in cases of infantile-onset CMT with secondary mitochondrial dysfunction.
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http://dx.doi.org/10.1186/s12920-021-01001-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8199524PMC
June 2021

Results From an Italian Expanded Access Program on Cannabidiol Treatment in Highly Refractory Dravet Syndrome and Lennox-Gastaut Syndrome.

Front Neurol 2021 20;12:673135. Epub 2021 May 20.

Pediatric Neurology and Epileptology Unit, Brotzu Hospital Trust, Cagliari, Italy.

Purified cannabidiol (CBD) was administered to highly refractory patients with Dravet (DS) or Lennox-Gastaut (LGS) syndromes in an ongoing expanded access program (EAP). Herein, we report interim results on CBD safety and seizure outcomes in patients treated for a 12-month period. Thirty centers were enrolled from December 2018 to December 2019 within the open-label prospective EAP up to a maximum of 25 mg/kg per day. Adverse effects and liver function tests were assessed after 2 weeks; 1, 3, and 6 months of treatment; and periodically thereafter. Seizure endpoints were the percentage of patients with ≥50 and 100% reduction in seizures compared to baseline. A total of 93 patients were enrolled and included in the safety analysis. Eighty-two patients [27 (32.9%) DS, 55 (67.1%) LGS] with at least 3 months of treatment have been included in the effectiveness analysis; median previously failed antiseizure medications was eight. Pediatric and adult patients were uniformly represented in the cohort. At 3-month follow-up, compared to the 28-day baseline period, the percentage of patients with at least a 50% reduction in seizure frequency was 40.2% (plus 1.2% seizure-free). Retention rate was similar according to diagnosis, while we found an increased number of patients remaining under treatment in the adult group. CBD was mostly coadministered with valproic acid (62.2%) and clobazam (41.5%). In the safety dataset, 29 (31.2%) dropped out: reasons were lack of efficacy [16 (17.2%)] and adverse events (AEs) [12 (12.9%)], and one met withdrawal criteria (1.1%). Most reported AEs were somnolence (22.6%) and diarrhea (11.9%), followed by transaminase elevation and loss of appetite. CBD is associated with improved seizure control also in a considerable proportion of highly refractory patients with DS and LGS independently from clobazam use. Overall, CBD safety and effectiveness are not dose-related in this cohort.
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http://dx.doi.org/10.3389/fneur.2021.673135DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8173151PMC
May 2021

Biallelic and monoallelic variants in PLXNA1 are implicated in a novel neurodevelopmental disorder with variable cerebral and eye anomalies.

Genet Med 2021 Sep 30;23(9):1715-1725. Epub 2021 May 30.

Department of Medical Genetics, Centre for Applied Neurogenetics, University of British Columbia, Vancouver, BC, Canada.

Purpose: To investigate the effect of PLXNA1 variants on the phenotype of patients with autosomal dominant and recessive inheritance patterns and to functionally characterize the zebrafish homologs plxna1a and plxna1b during development.

Methods: We assembled ten patients from seven families with biallelic or de novo PLXNA1 variants. We describe genotype-phenotype correlations, investigated the variants by structural modeling, and used Morpholino knockdown experiments in zebrafish to characterize the embryonic role of plxna1a and plxna1b.

Results: Shared phenotypic features among patients include global developmental delay (9/10), brain anomalies (6/10), and eye anomalies (7/10). Notably, seizures were predominantly reported in patients with monoallelic variants. Structural modeling of missense variants in PLXNA1 suggests distortion in the native protein. Our zebrafish studies enforce an embryonic role of plxna1a and plxna1b in the development of the central nervous system and the eye.

Conclusion: We propose that different biallelic and monoallelic variants in PLXNA1 result in a novel neurodevelopmental syndrome mainly comprising developmental delay, brain, and eye anomalies. We hypothesize that biallelic variants in the extracellular Plexin-A1 domains lead to impaired dimerization or lack of receptor molecules, whereas monoallelic variants in the intracellular Plexin-A1 domains might impair downstream signaling through a dominant-negative effect.
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http://dx.doi.org/10.1038/s41436-021-01196-9DOI Listing
September 2021

: One More Gene Contributing to the Solution of the Genetic Generalized Epilepsies Mystery.

Neurology 2021 05 23;96(18):831-832. Epub 2021 Mar 23.

From the Neuroscience Department (R.G.), Children's Hospital Anna Meyer-University of Florence; Neuropsichiatria Infantile (C.M.), Centro Regionale per la Diagnosi e Cura Epilessia Infantile, Presidio Ospedaliero G. Salesi; and Ospedali Riuniti Ancona (C.M.), Italy.

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http://dx.doi.org/10.1212/WNL.0000000000011854DOI Listing
May 2021

Migrating Focal Seizures and Myoclonic Status in Related Encephalopathy.

Neurol Genet 2021 Jun 14;7(3):e593. Epub 2021 May 14.

Child Neuropsychiatry Unit(F.D.), Department of Surgical Sciences, Dentistry, Gynecology and Pediatrics University of Verona; Child Neuropsychiatry Unit(T.L.B.), Department of Surgical Sciences, Dentistry, Gynecology and Pediatrics University of Verona; PhD Program in Clinical and Experimental Medicine (T.L.B.), University of Modena and Reggio Emilia; Pediatric Unit (R.O.), Department of Surgical Sciences, Dentistry, Gynecology and Pediatrics University of Verona; Pediatric Neurology (E.P.), Neurogenetics and Neurobiology Unit and Laboratories, Meyer Children's Hospital, University of Florence; Pediatric Neurology (C.B.), Neurogenetics and Neurobiology Unit and Laboratories, Meyer Children's Hospital, University of Florence; Child Neuropsychiatry Unit (E.F.), Azienda Ospedaliera Universitaria Integrata di Verona; Neurology (Child Neurology and Neuropathology) (A.S.), Department of Neuroscience, Biomedicine and Movement, University of Verona; CREP (Research Center for Pediatric Epilepsies) (B.D.B.), Azienda Ospedaliera Universitaria Integrata di Verona; and Pediatric Neurology (R.G.), Neurogenetics and Neurobiology Unit and Laboratories, Meyer Children's Hospital, University of Florence, Italy.

Objective: To report longitudinal clinical, EEG, and MRI findings in 2 sisters carrying compound heterozygous mutations and exhibiting a peculiar form of developmental and epileptic encephalopathy (DEE). Neuropathologic features are also described in one of the sisters.

Methods: Clinical course description, video-EEG polygraphic recordings, brain MRI, skin and muscle biopsies, whole-exome sequencing (WES), and brain neuropathology.

Results: Since their first months of life, both girls exhibited severe axial hypotonia, visual inattention, dyskinetic movements, severe developmental delay, and slow background EEG activity. Intractable nonmotor seizures started in both at the eighth month of life, exhibiting the electroclinical characteristics of epilepsy of infancy with migrating focal seizures (EIMFS). In the second year of life, continuous epileptiform EEG activity of extremely high amplitude appeared in association with myoclonic status, leading to severely impaired alertness and responsiveness. Repeated brain MRI revealed progressive atrophic changes and severe hypomyelination. WES identified a compound heterozygous in the gene [(p.Ser122Glnfs*7) and (p.Trp163*)] in one patient and was subsequently confirmed in the other. Both sisters died prematurely during respiratory infections. Postmortem neuropathologic examination of the brain, performed in one, revealed atrophic brain changes, mainly involving the cerebellum.

Conclusions: This report confirms that biallelic mutations cause a severe form of DEE and adds epilepsy with migrating focal seizures and myoclonic status to the spectrum of epilepsy phenotypes. Considering the potential role of human in glycosylphosphatidylinositol (GPI) anchor biosynthesis, this severe syndrome can be assigned to the group of inherited GPI deficiency disorders, with which it shares remarkably similar clinical and neuroimaging features. should be considered in the genetic screening of individuals with EIMFS.
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http://dx.doi.org/10.1212/NXG.0000000000000593DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8131096PMC
June 2021

Cerebral folate transporter deficiency syndrome in three siblings: Why genetic testing for developmental and epileptic encephalopathies should be performed early and include the FOLR1 gene.

Am J Med Genet A 2021 08 19;185(8):2526-2531. Epub 2021 May 19.

Child and Adolescent Neurology and Psychiatry Unit, Children Hospital, ASST Spedali Civili of Brescia, Brescia, Italy.

Cerebral folate transporter deficiency syndrome, caused by FOLR-1 mutations is characterized by late infantile onset, severe developmental regression, epilepsy, and leukodystrophy. An extremely low concentration of 5-methyltetrahydrofolate in the cerebrospinal fluid provides a crucial clue to its diagnosis and is a treatment target. Oral or intravenous folinic acid (5-formyltetrahydrofolate) administration improves clinical symptoms and brain magnetic resonance imaging (MRI) findings. We describe three siblings carrying a novel homozygous FOLR1 nonsense mutation, that were referred due to intractable epilepsy and progressive neurological decline. Brain MRI showed hypomyelination and cerebellar atrophy. Folinic acid (oral and intravenous) supplementation, initiated after over 15 years illness, has failed to result in any sizeable clinical or neurophysiological improvement. Cerebral folate transport deficiency bears overlapping clinical features with many severe developmental encephalopathies. It is crucial to recognize FOLR1 signs and establish an early clinical and molecular diagnosis in order to provide timely folinic acid treatment and improve outcome.
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http://dx.doi.org/10.1002/ajmg.a.62345DOI Listing
August 2021

Phenotypic Spectrum of Seizure Disorders in MBD5-Associated Neurodevelopmental Disorder.

Neurol Genet 2021 Apr 18;7(2):e579. Epub 2021 Mar 18.

Research Institute of the McGill University Health Centre (K.M.), Montreal, PQ; Division of Child Neurology (K.M.), Department of Pediatrics, Montreal Children's Hospital, McGill University, Montreal, PQ; Department of Neurology & Neurosurgery (K.M.), Montreal Children's Hospital, McGill University, Montreal, PQ; Child Neurology and Psychiatry (C.M.), Salesi Pediatric Hospital, United Hospitals of Ancona, Ancona, Italy; Division of Genetic Medicine (G.L.C., J.N., H.C.M.), Department of Pediatrics, University of Washington, Seattle, WA; Department of Neurology (A.M.), Great Ormond Street Hospital for Children, London, UK; Developmental Neurosciences Programme (A.M.), UCL Great Ormond Street Institute of Child Health, London, UK; Neurology Network Melbourne (J.P.), Melbourne, Victoria, Australia; Murdoch Children's Research Institute (C.S., I.E.S.), Parkville, Victoria, Australia; Department of Paediatrics and Child Health (T.S.), School of Medicine and Health Sciences, University of Otago, Wellington, New Zealand; Division of Neurology (S.M.), Department of Pediatrics, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, Manitoba, Canada; Neurology Unit and Neurogenetic Laboratories (C.B., A.R., R.G.), Meyer Children's Hospital, Florence, Italy; Department of Clinical Genetics (R.H.S.), Great Ormond Street Hospital, London, UK; Epilepsy Research Centre (I.E.S.), Department of Medicine, The University of Melbourne, Austin Health, Heidelberg, Victoria, Australia; Department of Paediatrics (I.E.S.), Royal Children's Hospital, The University of Melbourne, Parkville, Victoria, Australia; and The Florey Institute of Neuroscience and Mental Health (I.E.S.), Heidelberg, Victoria, Australia.

Objective: To describe the phenotypic spectrum in patients with MBD5-associated neurodevelopmental disorder (MAND) and seizures; features of MAND include intellectual disability, epilepsy, psychiatric features of aggression and hyperactivity, and dysmorphic features including short stature and microcephaly, sleep disturbance, and ataxia.

Methods: We performed phenotyping on patients with deletions, duplications, or point mutations and a history of seizures.

Results: Twenty-three patients with MAND and seizures were included. Median seizure onset age was 2.9 years (range 3 days-13 years). The most common seizure type was generalized tonic-clonic; focal, atypical absence, tonic, drop attacks, and myoclonic seizures occurred frequently. Seven children had convulsive status epilepticus and 3 nonconvulsive status epilepticus. Fever, viral illnesses, and hot weather provoked seizures. EEG studies in 17/21 patients were abnormal, typically showing slow generalized spike-wave and background slowing. Nine had drug-resistant epilepsy, although 3 eventually became seizure-free. All but one had moderate-to-severe developmental impairment. Epilepsy syndromes included Lennox-Gastaut syndrome, myoclonic-atonic epilepsy, and infantile spasms syndrome. Behavioral problems in 20/23 included aggression, self-injurious behavior, and sleep disturbance.

Conclusions: disruption may be associated with severe early childhood-onset developmental and epileptic encephalopathy. Because neuropsychiatric dysfunction is common and severe, it should be an important focus of clinical management.
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http://dx.doi.org/10.1212/NXG.0000000000000579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8075573PMC
April 2021

Real-life survey of pitfalls and successes of precision medicine in genetic epilepsies.

J Neurol Neurosurg Psychiatry 2021 Oct 26;92(10):1044-1052. Epub 2021 Apr 26.

Department of Clinical and Experimental Epilepsy, UCL Queen Square Institute of Neurology, London, and Chalfont Centre for Epilepsy, Gerrard Cross, UK

Objective: The term 'precision medicine' describes a rational treatment strategy tailored to one person that reverses or modifies the disease pathophysiology. In epilepsy, single case and small cohort reports document nascent precision medicine strategies in specific genetic epilepsies. The aim of this multicentre observational study was to investigate the deeper complexity of precision medicine in epilepsy.

Methods: A systematic survey of patients with epilepsy with a molecular genetic diagnosis was conducted in six tertiary epilepsy centres including children and adults. A standardised questionnaire was used for data collection, including genetic findings and impact on clinical and therapeutic management.

Results: We included 293 patients with genetic epilepsies, 137 children and 156 adults, 162 females and 131 males. Treatment changes were undertaken because of the genetic findings in 94 patients (32%), including rational precision medicine treatment and/or a treatment change prompted by the genetic diagnosis, but not directly related to known pathophysiological mechanisms. There was a rational precision medicine treatment for 56 patients (19%), and this was tried in 33/56 (59%) and was successful (ie, >50% seizure reduction) in 10/33 (30%) patients. In 73/293 (25%) patients there was a treatment change prompted by the genetic diagnosis, but not directly related to known pathophysiological mechanisms, and this was successful in 24/73 (33%).

Significance: Our survey of clinical practice in specialised epilepsy centres shows high variability of clinical outcomes following the identification of a genetic cause for an epilepsy. Meaningful change in the treatment paradigm after genetic testing is not yet possible for many people with epilepsy. This systematic survey provides an overview of the current application of precision medicine in the epilepsies, and suggests the adoption of a more considered approach.
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http://dx.doi.org/10.1136/jnnp-2020-325932DOI Listing
October 2021

ATP1A2- and ATP1A3-associated early profound epileptic encephalopathy and polymicrogyria.

Brain 2021 06;144(5):1435-1450

Pediatric Neurology, Neurogenetics and Neurobiology Unit and Laboratories, Meyer Children's Hospital, University of Florence, Florence, Italy.

Constitutional heterozygous mutations of ATP1A2 and ATP1A3, encoding for two distinct isoforms of the Na+/K+-ATPase (NKA) alpha-subunit, have been associated with familial hemiplegic migraine (ATP1A2), alternating hemiplegia of childhood (ATP1A2/A3), rapid-onset dystonia-parkinsonism, cerebellar ataxia-areflexia-progressive optic atrophy, and relapsing encephalopathy with cerebellar ataxia (all ATP1A3). A few reports have described single individuals with heterozygous mutations of ATP1A2/A3 associated with severe childhood epilepsies. Early lethal hydrops fetalis, arthrogryposis, microcephaly, and polymicrogyria have been associated with homozygous truncating mutations in ATP1A2. We investigated the genetic causes of developmental and epileptic encephalopathies variably associated with malformations of cortical development in a large cohort and identified 22 patients with de novo or inherited heterozygous ATP1A2/A3 mutations. We characterized clinical, neuroimaging and neuropathological findings, performed in silico and in vitro assays of the mutations' effects on the NKA-pump function, and studied genotype-phenotype correlations. Twenty-two patients harboured 19 distinct heterozygous mutations of ATP1A2 (six patients, five mutations) and ATP1A3 (16 patients, 14 mutations, including a mosaic individual). Polymicrogyria occurred in 10 (45%) patients, showing a mainly bilateral perisylvian pattern. Most patients manifested early, often neonatal, onset seizures with a multifocal or migrating pattern. A distinctive, 'profound' phenotype, featuring polymicrogyria or progressive brain atrophy and epilepsy, resulted in early lethality in seven patients (32%). In silico evaluation predicted all mutations to be detrimental. We tested 14 mutations in transfected COS-1 cells and demonstrated impaired NKA-pump activity, consistent with severe loss of function. Genotype-phenotype analysis suggested a link between the most severe phenotypes and lack of COS-1 cell survival, and also revealed a wide continuum of severity distributed across mutations that variably impair NKA-pump activity. We performed neuropathological analysis of the whole brain in two individuals with polymicrogyria respectively related to a heterozygous ATP1A3 mutation and a homozygous ATP1A2 mutation and found close similarities with findings suggesting a mainly neural pathogenesis, compounded by vascular and leptomeningeal abnormalities. Combining our report with other studies, we estimate that ∼5% of mutations in ATP1A2 and 12% in ATP1A3 can be associated with the severe and novel phenotypes that we describe here. Notably, a few of these mutations were associated with more than one phenotype. These findings assign novel, 'profound' and early lethal phenotypes of developmental and epileptic encephalopathies and polymicrogyria to the phenotypic spectrum associated with heterozygous ATP1A2/A3 mutations and indicate that severely impaired NKA pump function can disrupt brain morphogenesis.
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http://dx.doi.org/10.1093/brain/awab052DOI Listing
June 2021

Consensus statements on the information to deliver after a febrile seizure.

Eur J Pediatr 2021 Sep 17;180(9):2993-2999. Epub 2021 Apr 17.

APHP. Service de Neurologie Pédiatrique, Hôpital Robert Debré, Paris, France.

Febrile seizures (FS) are usually self-limiting and cause no morbidity. Nevertheless they represent very traumatic events for families. There is a need to identify key messages that reassure carers and help to prevent inappropriate, anxiety-driven behaviors associated with "fever phobia." No recommendations have been proposed to date regarding the content of such messages. Using a Delphi process, we have established a consensus regarding the information to be shared with families following a FS. Twenty physicians (child neurologists and pediatricians) from five European countries participated in a three-step Delphi process between May 2018 and October 2019. In the first step, each expert was asked to give 10 to 15 free statements about FS. In the second and third steps, statements were scored and selected according to the expert ranking of importance. A list of key messages for families has emerged from this process, which offer reassurance about FS based on epidemiology, underlying mechanisms, and the emergency management of FS should they recur. Interestingly, there was a high level of agreement between child neurologists and general pediatricians.Conclusion: We propose key messages to be communicated with families in the post-FS clinic setting. What is Known: • Febrile seizures (FS) are traumatic events for families. • No guidelines exist on what information to share with parents following a FS. What is New: • A Delphi process involving child neurologists and pediatricians provides consensual statement about information to deliver after a febrile seizure. • We propose key messages to be communicated with families in the post-FS clinic setting.
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http://dx.doi.org/10.1007/s00431-021-04067-2DOI Listing
September 2021

Focal cortical dysplasia: an update on diagnosis and treatment.

Expert Rev Neurother 2021 Apr 25:1-12. Epub 2021 Apr 25.

Neuroscience Department, Children's Hospital Meyer-University of Florence, Florence, Italy.

Introduction: Focal cortical dysplasias (FCDs) represent the most common etiology in pediatric drug-resistant focal epilepsies undergoing surgical treatment. The localization, extent and histopathological features of FCDs are considerably variable. Somatic mosaic mutations of genes that encode proteins in the PI3K-AKTmTOR pathway, which also includes the tuberous sclerosis associated genes and , have been implicated in FCD type II in a substantial subset of patients. Surgery is the principal therapeutic option for FCD-related epilepsy. Advanced neurophysiological and neuroimaging techniques have improved surgical outcome and reduced the risk of postsurgical deficits. Pharmacological MTOR inhibitors are being tested in clinical trials and might represent an example of personalized treatment of epilepsy based on the known mechanisms of disease, used alone or in combination with surgery.

Areas Covered: This review will critically analyze the advances in the diagnosis and treatment of FCDs, with a special focus on the novel therapeutic options prompted by a better understanding of their pathophysiology.

Expert Opinion: Focal cortical dysplasia is a main cause of drug-resistant epilepsy, especially in children. Novel, personalized approaches are needed to more effectively treat FCD-related epilepsy and its cognitive consequences.
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http://dx.doi.org/10.1080/14737175.2021.1915135DOI Listing
April 2021

A survey of the European Reference Network EpiCARE on clinical practice for selected rare epilepsies.

Epilepsia Open 2021 03 13;6(1):160-170. Epub 2021 Jan 13.

IRCCS Mondino Foundation Pavia Italy.

Objective: Clinical care of rare and complex epilepsies is challenging, because evidence-based treatment guidelines are scarce, the experience of many physicians is limited, and interdisciplinary treatment of comorbidities is required. The pathomechanisms of rare epilepsies are, however, increasingly understood, which potentially fosters novel targeted therapies. The objectives of our survey were to obtain an overview of the clinical practice in European tertiary epilepsy centers treating patients with 5 arbitrarily selected rare epilepsies and to get an estimate of potentially available patients for future studies.

Methods: Members of the ( were invited to participate in a web-based survey on clinical practice of patients with Dravet syndrome, tuberous sclerosis complex (TSC), autoimmune encephalitis, and progressive myoclonic epilepsies including Unverricht Lundborg and Unverricht-like diseases. A consensus-based questionnaire was generated for each disease.

Results: Twenty-six of 30 invited epilepsy centers participated. Cohorts were present in most responding centers for TSC (87%), Dravet syndrome (85%), and autoimmune encephalitis (71%). Patients with TSC and Dravet syndrome represented the largest cohorts in these centers. The antiseizure drug treatments were rather consistent across the centers especially with regard to Dravet syndrome, infantile spasms in TSC, and Unverricht Lundborg / Unverricht-like disease. Available, widely used targeted therapies included everolimus in TSC and immunosuppressive therapies in autoimmune encephalitis. Screening for comorbidities was routinely done, but specific treatment protocols were lacking in most centers.

Significance: The survey summarizes the current clinical practice for selected rare epilepsies in tertiary European epilepsy centers and demonstrates consistency as well as heterogeneity in the treatment, underscoring the need for controlled trials and recommendations. The survey also provides estimates for potential participants of clinical trials recruited via EpiCARE, emphasizing the great potential of Reference Networks for future studies to evaluate new targeted therapies and to identify novel biomarkers.
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http://dx.doi.org/10.1002/epi4.12459DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7918306PMC
March 2021

Angiocentric glioma-associated seizures: The possible role of EATT2, pyruvate carboxylase and glutamine synthetase.

Seizure 2021 Mar 16;86:152-154. Epub 2021 Feb 16.

Pediatric Neurology, Neurogenetics and Neurobiology Unit and Laboratories, Children's Hospital A. Meyer-University of Florence, Italy.

Purpose: Our purpose was to better understand the pathogenesis of seizures associated with angiocentric glioma. Angiocentric glioma is an indolent and rare low-grade glioma. Its typical clinical presentation is with epileptic seizures. The pathogenesis of tumor-associated seizures is poorly understood. Among the possible pathomechanisms, the increased neurotoxic concentrations of the glutamate has been proposed. Glutamate transporters, pyruvate carboxylase and glutamine synthetase are involved in maintaining the physiological concentration of glutamate in the inter synaptic spaces.

Methods: We evaluated the immunohistochemical expression of EAAT2 (the most important glutamate transporter), pyruvate carboxylase and glutamine synthetase in 17 angiocentric gliomas.

Results: EAAT2 was never expressed (0%) in the neoplastic cells in none of the cases studied. Pyruvate carboxylase was expressed in the cytoplasm of the neoplastic cells in 16/17 cases (94 %). Glutamine synthetase was expressed in the cytoplasm of the neoplastic cells in 15/17 cases (88 %).

Conclusion: The net result of this enzymatic expression, in particular considering the loss of EAAT2, could be an increased glutamate concentration in the synaptic clef, which might increase local network excitability initially involving intratumoral neurons. The observation that the angiocentric glioma-associated epilepsy might be at least in part related to EAAT2 deficiency opens up interesting therapeutic perspectives.
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http://dx.doi.org/10.1016/j.seizure.2021.02.014DOI Listing
March 2021

Morquio B disease: From pathophysiology towards diagnosis.

Mol Genet Metab 2021 03 1;132(3):180-188. Epub 2021 Feb 1.

Molecular and Cell Biology Laboratory, Paediatric Neurology Unit and Laboratories, Neuroscience Department, A. Meyer Children's Hospital, Florence, Italy; Department of Neurosciences, Psychology, Pharmacology and Child Health, University of Florence, Italy. Electronic address:

Morquio B disease is an attenuated phenotype within the spectrum of beta galactosidase (GLB1) deficiencies. It is characterised by dysostosis multiplex, ligament laxity, mildly coarse facies and heart valve defects due to keratan sulphate accumulation, predominantly in the cartilage. Morquio B patients have normal neurological development, setting them apart from those with the more severe GM1 gangliosidosis. Morquio B disease, with an incidence of 1:250.000 to 1:1.000.000 live births, is very rare. Here we report the clinical-biochemical data of nine patients. High amounts of keratan sulfate were detected using LC-MS/MS in the patients' urinary samples, while electrophoresis, the standard procedure of qualitative glycosaminoglycans analysis, failed to identify this metabolite in any of the patients' samples. We performed molecular analyses at gene, gene expression and protein expression levels, for both isoforms of the GLB1 gene, lysosomal GLB1, and the cell-surface expressed Elastin Binding Protein. We characterised three novel GLB1 mutations [c.75 + 2 T > G, c.575A > G (p.Tyr192Cys) and c.2030 T > G (p.Val677Gly)] identified in three heterozygous patients. We also set up a copy number variation assay by quantitative PCR to evaluate the presence of deletions/ insertions in the GLB1 gene. We propose a diagnostic plan, setting out the specific clinical- biochemical and molecular features of Morquio B, in order to avoid misdiagnoses and improve patients' management.
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http://dx.doi.org/10.1016/j.ymgme.2021.01.008DOI Listing
March 2021
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