Noninvasive prenatal diagnosis of genetic diseases induced by triplet repeat expansion by linked read haplotyping and Bayesian approach.

The field of noninvasive prenatal diagnosis (NIPD) has undergone significant progress over the last decade. Direct haplotyping has been successfully applied for NIPD of few single-gene disorders. However, technical issues remain for triplet-repeat expansions. The objective of this study was to develop an NIPD approach for couples at risk of transmitting dynamic mutations. This method includes targeted enrichment for linked-read libraries and targeted maternal plasma DNA sequencing. We also developed an innovative Bayesian procedure to integrate the Hoobari fetal genotyping model for inferring the fetal haplotype and the targeted gene variant status. Our method of directly resolving parental haplotypes through targeted linked-read sequencing was smoothly performed using blood samples from families with Huntington's disease or myotonic dystrophy type 1. The Bayesian analysis of transmission of parental haplotypes allowed defining the genotype of five fetuses. The predicted variant status of four of these fetuses was in agreement with the invasive prenatal diagnosis findings. Conversely, no conclusive result was obtained for the NIPD of fragile X syndrome. Although improvements should be made to achieve clinically acceptable accuracy, our study shows that linked-read sequencing and parental haplotype phasing can be successfully used for NIPD of triplet-repeat expansion diseases.Trial registration: NCT04698551_date of first registration: 07/01/2021.

Molecular diagnosis and genetic counseling for spinal muscular atrophy (SMA).

Spinal muscular atrophy (SMA) is a neuromuscular autosomal recessive disorder caused by bi-allelic pathogenic variants in the SMN1 gene. 95% of SMA patients have a SMN1 homozygous deletion. In the 5% remaining affected patients, a heterozygous SMN1 deletion is associated with an intragenic SMN1 rare inactivating pathogenic variant on the other allele. The clinical phenotype of SMA is heterogeneous and severity is inversely correlated with the number of SMN2 copies, a non-functional SMN1 copy. The development of new treatments leads to the generalization of carrier and newborn screening in many countries and new robust and low cost methods for large population-based screening have been developed. It is important that all diagnosed patients and relatives receive appropriate genetic counseling, taking into account the great complexity of SMA region to avoid pitfalls. © 2020 French Society of Pediatrics. Published by Elsevier Masson SAS. All rights reserved.

Mutation update and uncommon phenotypes in a French cohort of 96 patients with WFS1-related disorders.

WFS1 mutations are responsible for Wolfram syndrome (WS) characterized by juvenile-onset diabetes mellitus and optic atrophy, and for low-frequency sensorineural hearing loss (LFSNHL). Our aim was to analyze the French cohort of 96 patients with WFS1-related disorders in order (i) to update clinical and molecular data with 37 novel affected individuals, (ii) to describe uncommon phenotypes and, (iii) to precise the frequency of large-scale rearrangements in WFS1. We performed quantitative polymerase chain reaction (PCR) in 13 patients, carrying only one heterozygous variant, to identify large-scale rearrangements in WFS1. Among the 37 novel patients, 15 carried 15 novel deleterious putative mutations, including one large deletion of 17,444 base pairs. The analysis of the cohort revealed unexpected phenotypes including (i) late-onset symptoms in 13.8% of patients with a probable autosomal recessive transmission; (ii) two siblings with recessive optic atrophy without diabetes mellitus and, (iii) six patients from four families with dominantly-inherited deafness and optic atrophy. We highlight the expanding spectrum of WFS1-related disorders and we show that, even if large deletions are rare events, they have to be searched in patients with classical WS carrying only one WFS1 mutation after sequencing.

An overview of neurological and neuromuscular signs in mitochondrial diseases.

Mitochondrial disorders have a broad clinical spectrum and are genetically heterogeneous, involving two genomes. These disorders may be develop at any age, with isolated or multiple system involvement, and any pattern of inheritance. Neurological involvement is the most frequent, and concerns muscular, peripheral and central nervous system. Among these diverse signs, some are suggestive of mitochondrial disease, such as progressive external ophthalmoplegia, exercise intolerance, psychomotor regression, stroke-like episodes, refractory epilepsy and Epilepsia Partialis Continua. Others are less specific and mitochondrial hypothesis may be evocated because of either association of different neuromuscular signs or a multisystemic involvement. This review describes the wealth of this neurological and neuromuscular symptomatology through different syndromes reported in the literature, according to preponderant signs and to modes of inheritance, as key elements to guide genetics testing.

[Bladder-sphincter disorders associated with Wolfram syndrome]. / Troubles vésicosphinctériens au cours du syndrome de Wolfram.

OBJECTIVES: To identify and describe functional urinary symptoms, uro-nephrological complications and their impact on quality of life in a cohort of patients diagnosed with the Wolfram syndrome (SW). PATIENTS AND METHODS: A transversal descriptive patient's cohort study was performed. The Urinary Symptom Profile (USP) and the International Consultation Incontinence Questionnaire - Female Lower Urinary Tracts Symptoms (ICIQ-FLUTS) were used to evaluate urinary symptoms and their impact on quality of life through direct interviews conducted by telephone. A less than 6-month old renal ultrasound and serum creatinine results were asked to the patient or physician. RESULTS: Thirty-three patients have been contacted and 22 (73%) agreed to participate in this study. Eighteen patients over 22 had spontaneous micturition when four of them had an urinary diversion (two definitive, two temporaries) before being included in this study. Seventy-three percent of patients with spontaneous micturition had urinary symptoms. A severe or moderate symptoms score was noted in 67% (12/18 patients) and 11% (2/18 patients) respectively. CONCLUSION: Functional urinary dysfunctions were frequent and impacted quality of life in more than one half of patients diagnosed with SW in this study. Early diagnostic and regular urological follow-up can improve the quality of life and prevent severe urinary complications.

[Epilepsy and mitochondrial diseases: retrospective study on 53 epileptic children]. / Épilepsie et cytopathies mitochondriales : étude rétrospective de 53 enfants épileptiques.

AIM: Mitochondrial disease is a heterogeneous disorder entity induced by defects in the mitochondrial respiratory chain complex. Neurological symptoms, including epilepsy, are common in children. The aim of this study was to research the clinical signs indicating mitochondrial disease. METHODS: We retrospectively studied epileptic children who underwent a muscle and/or hepatic biopsy between 1995 and 2010 searching for a mitochondrial disease. Patients were separated into 2 groups depending on the biopsy result: group 1 (presence of mitochondrial disease) and group 2 (absence of mitochondrial disease). Epileptic phenotypes were compared between these 2 groups. In group 1, we specified the clinical phenotype and characterized mitochondrial disease. RESULTS: Fifty-three children were included: 29 in group 1 and 24 in group 2. The average age at onset of epilepsy was 39.6 months in group 1 versus 11.8 months in group 2. In the 1st group, epilepsy was less refractory and associated with other clinical symptoms. CONCLUSIONS: In this study, epilepsy did not appear to be a unique sign of mitochondrial disease. It most often appeared during the 2nd year of life and is correlated with multiorgan involvement, notably ophthalmologic, such as oculomotor apraxia, optic atrophy, and retinitis pigmentosa, as well as auditory (deafness) and hepatic (hepatic failure, hepatomegaly). On the other hand, in children who did not have mitochondrial disease, epilepsy often began earlier (before 3 months of age), it was refractory, isolated without multiorgan involvement, and seems to be due to genetic anomalies in developmental genes, a finding that requires further research.

The severity of phenotype linked to SUCLG1 mutations could be correlated with residual amount of SUCLG1 protein.

BACKGROUND: Succinate-CoA ligase deficiency is responsible for encephalomyopathy with mitochondrial DNA depletion and mild methylmalonic aciduria. Mutations in SUCLA2, the gene encoding a ß subunit of succinate-CoA ligase, have been reported in 17 patients until now. Mutations in SUCLG1, encoding the α subunit of the enzyme, have been described in two pedigrees only. METHODS AND FINDINGS: In this study, two unrelated patients harbouring three novel pathogenic mutations in SUCLG1 were reported. The first patient had a severe disease at birth. He was compound heterozygous for a missense mutation (p.Pro170Arg) and a c.97+3G>C mutation, which leads to the complete skipping of exon 1 in a minigene expression system. The involvement of SUCLG1 was confirmed by western blot analysis, which showed absence of SUCLG1 protein in fibroblasts. The second patient has a milder phenotype, similar to that of patients with SUCLA2 mutations, and is still alive at 12 years of age. Western blot analysis showed some residual SUCLG1 protein in patient's fibroblasts. CONCLUSIONS: Our results suggest that SUCLG1 mutations that lead to complete absence of SUCLG1 protein are responsible for a very severe disorder with antenatal manifestations, whereas a SUCLA2-like phenotype is found in patients with residual SUCLG1 protein. Furthermore, it is shown that in the absence of SUCLG1 protein, no SUCLA2 protein is found in fibroblasts by western blot analysis. This result is consistent with a degradation of SUCLA2 when its heterodimer partner, SUCLG1, is absent.

[Chorea-acanthocytosis without acanthocytes]. / Choréo-acanthocytose sans acanthocytes.

INTRODUCTION: Chorea-acanthocytosis (ChAc) is one of the neuroacanthocytosis syndromes which form a group of disorders characterized by the association of neurological abnormalities and spiculated red blood cells called acanthocytes. ChAc patients exhibit involuntary movements, psychiatric abnormalities and progressive cognitive deterioration. We report a case of ChAc in which blood smears failed to demonstrate acanthocytes. CASE REPORT: A 26-year-old man presented since two years with hyperkinetic movements. The family history was non contributive, parents were consanguineous. Neurological examination revealed choreatic hyperkinesia and dystonia, predominant in the orofacial region. Mild cognitive decline and behavior abnormalities were noted with repetitive activities. Brain MRI showed striatal atrophy. Molecular testing for Huntington's disease was negative. Routine biological screening was normal except for elevated CPK and LDH. Copper and ceruloplasmin blood levels were normal, as well as purine metabolism and lipoproteins. Further screening for metabolic diseases showed no significant abnormality. Expression of Kell antigens was normal. In several blood smears no acanthocytes were seen. Electromyographic studies showed slight neuropathic changes. Despite the absence of acanthocytes, chorein western blot was performed on blood samples which revealed an absent or markedly reduced level of chorein in erythrocyte membranes. A mutation of the ChAc gene was thus likely so the diagnosis of ChAc was retained. Genetic studies for VPS13A are pending. DISCUSSION: ChAc is an autosomal recessive disorder due to mutations of the VPS13A gene coding for chorein. Absence or late appearance of acanthocytes in ChAc has been described in a few case reports. In conclusion ChAc is a rare disorder in which the presence of acanthocytes is not mandatory. In case of doubt, chorein western blot can be useful.

[Late cerebellar ataxia associated with fragile X premutation]. / Ataxie cérébelleuse tardive due à la prémutation de l'X fragile.

INTRODUCTION: The fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurodegenerative disorder that affects carriers, principally males after 50 years of age, of premutation alleles of the fragile X mental retardation 1 (FMR1) gene. Clinical features of FXTAS are an intention tremor and/or a cerebellar ataxia, with or without parkinsonism, cognitive disorders, neuropathy and autonomic dysfunction. MRI shows symmetrical signal abnormalities in the middle cerebellar peduncles and deep cerebellar white matter, typically sparing the dentate nucleus, associated with generalized atrophy. We report a case of FXTAS with isolated cerebellar ataxia. CASE REPORT: We report the case of a 60-year-old right-handed man with an uneventful personal and familial history. Since 2002, he progressively developed gait ataxia associated with paresthesia in the lower limbs. The physical examination revealed static and kinetic cerebellar ataxia with dysarthria. Exhaustive screening tests (inflammatory, immunological, metabolic, infectious and neoplasic) and the cerebrospinal fluid analysis were normal. The brain MRI T2-weighted sequences showed diffuse increased signal in both cerebellar white matter and middle peduncles suggestive of FXTAS. Functional explorations (evoked somesthesic and visual potentials, electromyogram, and cerebral scintigraphy) confirmed the isolated cerebellar involvement. The FXTAS was suggested and the genotype was explored. Southern Blot revealed an expansion of trinucleotide CGG with 107 repetitions, confirming the diagnosis of FXTAS. DISCUSSION: In patients with a fragile X premutation, FMR1 protein levels are gradually reduced with increased repeat number, despite elevated FMR1 transcripts levels. Neuropathologic examination reveals eosinophilic intranuclear inclusions in neurons and astrocytes throughout the cortex, subcortical regions, and brain stem. The pathogenic hypothesis would be related to a gain of function with toxic effects of FMR1 messenger RNA on cellular metabolism. Cerebellar ataxia, which may be isolated or not, is the most frequent neurologic manifestation, like many studies showed. CONCLUSION: FXTAS should be suspected in patients with unexplained late-onset cerebellar ataxia. Typical presentation includes a male in his fifties with progressive ataxia and tremor. Brain MRI with symmetrical cerebellar abnormalities is very suggestive of this diagnosis. Genetic screening and advice for the patient and his family must be proposed in order to detect the fragile X syndrome.

[Neurosarcoidosis treated with mycophenolate mofetil: two cases]. / Neurosarcoïdose et mycophénolate mofétil.

INTRODUCTION: Neurosarcoidosis is a rare (5 cases for one million) immune-mediated disease generally observed in young adults. Neurological symptoms are present in the half of patients, and symptoms remain limited to neurological system in 10p.cent. Histological criteria are mandatory to prove the diagnosis. The sensitivity and complications of biopsy are variable. The best sensitivity appears to be achieved with muscle biopsies which in addition have a lower risk of complications. Neurosarcoidosis is usually treated with corticosteroid therapy and immunosuppressive drugs (cyclophosphamide, cyclosporine, aziathoprine, methotrexate), but frequently resists standard schedules. In addition the many contraindications, side effects and cumulated toxicities of immunosuppressive drugs compromises their use. Knowledge of the effectiveness of other treatments would therefore be useful. Mycophenolate mofetil (MMF) has been used for treatment of many immune-mediated neurological diseases, like polymyositis, multifocal motor neuropathy, myasthenia or chronic inflammatory demyelinating polyradiculoneuropathy. MMF is efficient and well tolerated, but there is no case-report about neurosarcoidosis. CASE REPORT: We report two observations of young patients (14 and 27 years) with a diagnosis of resistant neurosarcoidosis treated with MMF (2 g/j) and corticosteroids. A significant and rapid effectiveness was clinically and radiologically observed, with good clinical and hematologic tolerance. CONCLUSION: The MMF seems to be an interesting rescue treatment for neurosarcoidosis. Further evaluation is needed.