Aralar Sequesters GABA into Hyperactive Mitochondria, Causing Social Behavior Deficits.

Social impairment is frequently associated with mitochondrial dysfunction and altered neurotransmission. Although mitochondrial function is crucial for brain homeostasis, it remains unknown whether mitochondrial disruption contributes to social behavioral deficits. Here, we show that Drosophila mutants in the homolog of the human CYFIP1, a gene linked to autism and schizophrenia, exhibit mitochondrial hyperactivity and altered group behavior. We identify the regulation of GABA availability by mitochondrial activity as a biologically relevant mechanism and demonstrate its contribution to social behavior. Specifically, increased mitochondrial activity causes gamma aminobutyric acid (GABA) sequestration in the mitochondria, reducing GABAergic signaling and resulting in social deficits. Pharmacological and genetic manipulation of mitochondrial activity or GABA signaling corrects the observed abnormalities. We identify Aralar as the mitochondrial transporter that sequesters GABA upon increased mitochondrial activity. This study increases our understanding of how mitochondria modulate neuronal homeostasis and social behavior under physiopathological conditions.

Congenital myasthenic syndromes in adults: clinical features, diagnosis and long-term prognosis.

Congenital myasthenic syndromes (CMS) are clinically and genetically heterogeneous diseases caused by mutations affecting neuromuscular transmission. Even if the first symptoms mainly occur during childhood, adult neurologists must confront this challenging diagnosis and manage these patients throughout their adulthood. However, long-term follow-up data from large cohorts of CMS patients are lacking and the long-term prognosis of these patients is largely unknown. We report the clinical features, diagnostic difficulties, and long-term prognosis of a French nationwide cohort of 235 adult patients with genetically confirmed CMS followed in 23 specialized neuromuscular centres. Data were retrospectively analysed. Of the 235 patients, 123 were female (52.3%). The diagnosis was made in adulthood in 139 patients, 110 of whom presented their first symptoms before the age of 18. Mean follow-up time between first symptoms and last visit was 34 years (SD = 15.1). Pathogenic variants were found in 19 disease-related genes. CHRNE-low expressor variants were the most common (23.8%), followed by variants in DOK7 (18.7%) and RAPSN (14%). Genotypes were clustered into four groups according to the initial presentation: ocular group (CHRNE-LE, CHRND, FCCMS), distal group (SCCMS), limb-girdle group (RAPSN, COLQ, DOK7, GMPPB, GFPT1), and a variable-phenotype group (MUSK, AGRN). The phenotypical features of CMS did not change throughout life. Only four genotypes had a proportion of patients requiring intensive care unit (ICU) admission that exceeded 20%: RAPSN (54.8%), MUSK (50%), DOK7 (38.6%) and AGRN (25.0%). In RAPSN and MUSK patients most ICU admissions occurred before age 18 years and in DOK7 and AGRN patients at or after 18 years of age. Different patterns of disease course (stability, improvement and progressive worsening) may succeed one another in the same patient throughout life, particularly in AGRN, DOK7 and COLQ. At the last visit, 55% of SCCMS and 36.3% of DOK7 patients required ventilation; 36.3% of DOK7 patients, 25% of GMPPB patients and 20% of GFPT1 patients were wheelchair-bound; most of the patients who were both wheelchair-bound and ventilated were DOK7 patients. Six patients died in this cohort. The positive impact of therapy was striking, even in severely affected patients. In conclusion, even if motor and/or respiratory deterioration could occur in patients with initially moderate disease, particularly in DOK7, SCCMS and GFPT1 patients, the long-term prognosis for most CMS patients was favourable, with neither ventilation nor wheelchair needed at last visit. CHRNE patients did not worsen during adulthood and RAPSN patients, often severely affected in early childhood, subsequently improved.

Long-term prognosis of fatty-acid oxidation disorders in adults: Optimism despite the limited effective therapies available.

INTRODUCTION: Fatty-acid oxidation disorders (FAODs) are recessive genetic diseases. MATERIALS AND METHODS: We report here clinical and paraclinical data from a retrospective study of 44 adults with muscular FAODs from six French reference centers for neuromuscular or metabolic diseases. RESULTS: The study cohort consisted of 44 adult patients: 14 with carnitine palmitoyl transferase 2 deficiency (32%), nine with multiple acyl-CoA deficiency (20%), 13 with very long-chain acyl-CoA dehydrogenase deficiency (30%), three with long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (7%), and five with short-chain acyl-CoA dehydrogenase deficiency (11%). Disease onset occurred during childhood in the majority of patients (59%), with a mean age at onset of 15 years (range = 0.5-35) and a mean of 12.6 years (range = 0-58) from disease onset to diagnosis. The principal symptoms were acute muscle manifestations (rhabdomyolysis, exercise intolerance, myalgia), sometimes associated with permanent muscle weakness. Episodes of rhabdomyolysis were frequent (84%), with a mean creatinine kinase level of 68,958 U/L (range = 660-300,000). General metabolic complications were observed in 58% of patients, respiratory manifestations in 18% of cases, and cardiological manifestations in 9% of cases. Fasting acylcarnitine profile was used to orient genetic explorations in 65% of cases. After a mean follow-up of 10 years, 33% of patients were asymptomatic and 56% continued to display symptoms after exercise. The frequency of rhabdomyolysis decreased after diagnosis in 64% of cases. CONCLUSION: A standardized register would complete this cohort description of muscular forms of FAODs with exhaustive data, making it possible to assess the efficacy of therapeutic protocols in real-life conditions and during the long-term follow-up of patients.

Real-life effectiveness 1 year after switching to avalglucosidase alfa in late-onset Pompe disease patients worsening on alglucosidase alfa therapy: A French cohort study.

INTRODUCTION: Late-onset Pompe disease (LOPD) is characterized by a progressive myopathy resulting from a deficiency of acid α-glucosidase enzyme activity. Enzyme replacement therapy has been shown to be effective, but long-term treatment results vary. Avalglucosidase alfa demonstrated non-inferiority to alglucosidase alfa in a phase 3 study, allowing in France compassionate access for advanced LOPD patients unresponsive to alglucosidase alfa. METHODS: Data from the French Pompe registry were analyzed for patients who benefited from a switch to avalglucosidase alfa with at least 1 year of follow-up. Respiratory (forced vital capacity [FVC]) and motor functions (Six-Minute Walk Test [6MWT]) were assessed before and 1 year after switching. Individual changes in FVC and 6MWT were expressed as slopes and statistical analyses were performed to compare values. RESULTS: Twenty-nine patients were included (mean age 56 years, 11 years of prior treatment). The FVC and 6MWT values remained stable. The individual analyses showed a stabilization of motor worsening: -1 m/year on the 6MWT after the switch versus -63 m/year the year before the switch (i.e., a worsening of 33%/year before vs. an improvement of 3%/year later). Respiratory data were not statistically different. DISCUSSION: At the group level, gait parameters improved slightly with a stabilization of previous worsening, but respiratory parameters showed limited changes. At the individual level, results were discordant, with some patients with a good motor or respiratory response and some with further worsening. CONCLUSION: Switching to avalglucosidase alfa demonstrated varied responses in advanced LOPD patients with failing alglucosidase alfa therapy, with a general improvement in motor stabilization.

CHCHD2 harboring Parkinson's disease-linked T61I mutation precipitates inside mitochondria and induces precipitation of wild-type CHCHD2.

The T61I mutation in coiled-coil-helix-coiled-coil-helix domain containing 2 (CHCHD2), a protein residing in the mitochondrial intermembrane space (IMS), causes an autosomal dominant form of Parkinson's disease (PD), but the underlying pathogenic mechanisms are not well understood. Here, we compared the subcellular localization and solubility of wild-type (WT) and T61I mutant CHCHD2 in human cells. We found that mitochondrial targeting of both WT and T61I CHCHD2 depended on the four cysteine residues in the C-terminal coiled-coil-helix-coiled-coil-helix (CHCH) domain but not on the N-terminal predicted mitochondrial targeting sequence. The T61I mutation did not interfere with mitochondrial targeting of the mutant protein but induced its precipitation in the IMS. Moreover, T61I CHCHD2 induced increased mitochondrial production of reactive oxygen species and apoptosis, which was prevented by treatment with anti-oxidants. Retention of T61I CHCHD2 in the cytosol through mutation of the cysteine residues in the CHCH domain prevented its precipitation as well as its apoptosis-inducing effect. Importantly, T61I CHCHD2 potently impaired the solubility of WT CHCHD2. In conclusion, our data show that the T61I mutation renders mutant CHCHD2 insoluble inside mitochondria, suggesting loss of function of the mutant protein. In addition, T61I CHCHD2 exerts a dominant-negative effect on the solubility of WT CHCHD2, explaining the dominant inheritance of this form of PD.

Motor and respiratory decline in patients with late onset Pompe disease after cessation of enzyme replacement therapy during COVID-19 pandemic.

BACKGROUND AND PURPOSE: Data on interruption of enzyme replacement therapy (ERT) are scarce in late onset Pompe disease. Due to the COVID-19 crisis, eight neuromuscular reference centers in France were obligated to stop the treatment for 31 patients. METHODS: We collected the motor and respiratory data from our French registry, before COVID-19 and at treatment restart. RESULTS: In 2.2 months (mean), patients showed a significant deterioration of 37 m (mean) in the 6-min walk test and a loss of 210 ml (mean) of forced vital capacity, without ad integrum restoration after 3 months of ERT restart. CONCLUSIONS: This national study based on data from the French Pompe Registry shows that the interruption of ERT, even as short as a few months, worsens Pompe patients' motor and respiratory function.

Melanoma addiction to the long non-coding RNA SAMMSON.

Focal amplifications of chromosome 3p13-3p14 occur in about 10% of melanomas and are associated with a poor prognosis. The melanoma-specific oncogene MITF resides at the epicentre of this amplicon. However, whether other loci present in this amplicon also contribute to melanomagenesis is unknown. Here we show that the recently annotated long non-coding RNA (lncRNA) gene SAMMSON is consistently co-gained with MITF. In addition, SAMMSON is a target of the lineage-specific transcription factor SOX10 and its expression is detectable in more than 90% of human melanomas. Whereas exogenous SAMMSON increases the clonogenic potential in trans, SAMMSON knockdown drastically decreases the viability of melanoma cells irrespective of their transcriptional cell state and BRAF, NRAS or TP53 mutational status. Moreover, SAMMSON targeting sensitizes melanoma to MAPK-targeting therapeutics both in vitro and in patient-derived xenograft models. Mechanistically, SAMMSON interacts with p32, a master regulator of mitochondrial homeostasis and metabolism, to increase its mitochondrial targeting and pro-oncogenic function. Our results indicate that silencing of the lineage addiction oncogene SAMMSON disrupts vital mitochondrial functions in a cancer-cell-specific manner; this silencing is therefore expected to deliver highly effective and tissue-restricted anti-melanoma therapeutic responses.

PARL deficiency in mouse causes Complex III defects, coenzyme Q depletion, and Leigh-like syndrome.

The mitochondrial intramembrane rhomboid protease PARL has been implicated in diverse functions in vitro, but its physiological role in vivo remains unclear. Here we show that Parl ablation in mouse causes a necrotizing encephalomyelopathy similar to Leigh syndrome, a mitochondrial disease characterized by disrupted energy production. Mice with conditional PARL deficiency in the nervous system, but not in muscle, develop a similar phenotype as germline Parl KOs, demonstrating the vital role of PARL in neurological homeostasis. Genetic modification of two major PARL substrates, PINK1 and PGAM5, do not modify this severe neurological phenotype. Parl-/- brain mitochondria are affected by progressive ultrastructural changes and by defects in Complex III (CIII) activity, coenzyme Q (CoQ) biosynthesis, and mitochondrial calcium metabolism. PARL is necessary for the stable expression of TTC19, which is required for CIII activity, and of COQ4, which is essential in CoQ biosynthesis. Thus, PARL plays a previously overlooked constitutive role in the maintenance of the respiratory chain in the nervous system, and its deficiency causes progressive mitochondrial dysfunction and structural abnormalities leading to neuronal necrosis and Leigh-like syndrome.

Improved detection of mitochondrial DNA instability in mitochondrial genome maintenance disorders.

PURPOSE: Diseases caused by defects in mitochondrial DNA (mtDNA) maintenance machinery, leading to mtDNA deletions, form a specific group of disorders. However, mtDNA deletions also appear during aging, interfering with those resulting from mitochondrial disorders. METHODS: Here, using next-generation sequencing (NGS) data processed by eKLIPse and data mining, we established criteria distinguishing age-related mtDNA rearrangements from those due to mtDNA maintenance defects. MtDNA deletion profiles from muscle and urine patient samples carrying pathogenic variants in nuclear genes involved in mtDNA maintenance (n = 40) were compared with age-matched controls (n = 90). Seventeen additional patient samples were used to validate the data mining model. RESULTS: Overall, deletion number, heteroplasmy level, deletion locations, and the presence of repeats at deletion breakpoints were significantly different between patients and controls, especially in muscle samples. The deletion number was significantly relevant in adults, while breakpoint repeat lengths surrounding deletions were discriminant in young subjects. CONCLUSION: Altogether, eKLIPse analysis is a powerful tool for measuring the accumulation of mtDNA deletions between patients of different ages, as well as in prioritizing novel variants in genes involved in mtDNA stability.

Deep phenotyping of an international series of patients with late-onset dysferlinopathy.

BACKGROUND: To describe the clinical, pathological, and molecular characteristics of late-onset (LO) dysferlinopathy patients. METHODS: Retrospective series of patients with LO dysferlinopathy, defined by an age at onset of symptoms ≥30 years, from neuromuscular centers in France and the International Clinical Outcome Study for dysferlinopathy (COS). Patients with early-onset (EO) dysferlinopathy (<30 years) were randomly selected from the COS study as a control group, and the North Star Assessment for Dysferlinopathy (NSAD) and Activity Limitation (ACTIVLIM) scores were used to assess functionality. Muscle biopsies obtained from 11 LO and 11 EO patients were revisited. RESULTS: Forty-eight patients with LO dysferlinopathy were included (28 females). Median age at onset of symptoms was 37 (range 30-57) years and most patients showed a limb-girdle (n = 26) or distal (n = 10) phenotype. However, compared with EO dysferlinopathy patients (n = 48), LO patients more frequently showed atypical phenotypes (7 vs. 1; p = 0.014), including camptocormia, lower creatine kinase levels (2855 vs. 4394 U/L; p = 0.01), and higher NSAD (p = 0.008) and ACTIVLIM scores (p = 0.016). Loss of ambulation in LO patients tended to occur later (23 ± 4.4 years after disease onset vs. 16.3 ± 6.8 years; p = 0.064). Muscle biopsy of LO patients more frequently showed an atypical pattern (unspecific myopathic changes) as well as significantly less necrosis regeneration and inflammation. Although LO patients more frequently showed missense variants (39.8% vs. 23.9%; p = 0.021), no differences in dysferlin protein expression were found on Western blot. CONCLUSIONS: Late-onset dysferlinopathy patients show a higher frequency of atypical presentations, are less severely affected, and show milder dystrophic changes in muscle biopsy.

Restricted Sensitivity of FJ-C Staining to Assess Neuronal Degeneration and Death in Preclinical Mouse Studies.

Fluorescein-derived fluorochromes and anionic dyes such as Fluoro-Jade (FJ) stains have been introduced to facilitate recognition of dying neurons in tissue sections. However, the definition of what is really detected by FJ-based stains and its sensitivity in the detection of neuronal cell death is unclear. In our work, we evaluated the outcome of FJ-C staining in mouse brains from 4 different well-characterized models of neurodegeneration. Neuronal degeneration and loss were highlighted with high sensitivity by FJ-C stain in mice with dysfunctional γ-secretase in the glutamatergic neurons and in mice affected by acute cerebral ischemia. Histopathologically, acute eosinophilic necrosis or "red dead" neurons were associated with FJ-C staining in both settings. Conversely, in mice affected by chronic cerebral microinfarcts due to tumor lysis syndrome as well as in a model of mitochondrial encephalopathy, FJ-C staining failed to detect neuronal death. Histopathologically, these models were characterized by extensive neuronal vacuolation associated with fading neurons ("ghost cells"). Therefore, contrary to the widespread belief that FJ-C stain has high affinity for all degenerating neurons regardless of the underlying cell death mechanism, we observed restricted sensitivity of the technique to specific conditions of neuronal cell death. As such, complementary techniques are essential to evaluate the presence of neurodegeneration in the absence of a positive FJ-C signal.

Spontaneous Incidental Brain Lesions in C57BL/6J Mice.

Genetically engineered mouse lines on a C57BL/6J background are widely employed as preclinical models to study neurodegenerative human disorders and brain tumors. However, because of the lack of comprehensive data on the spontaneous background neuropathology of the C57BL/6J strain, discriminating between naturally occurring changes and lesions caused by experimental mutations can be challenging. In this context, this study aims at defining the spectrum and frequency of spontaneous brain changes in a large cohort of C57BL/6J mice and their association with specific biological variables, including age and sex. Brains from 203 experimentally naive and clinically unremarkable C57BL/6J mice were collected and analyzed by means of histopathology and immunohistochemistry. Mice ranged in age from 3 to 110 weeks with 89 females, 111 males, and 3 unknowns. Sixteen different spontaneous lesion categories were described in this cohort. Age-related neurodegenerative and/or neuroinflammatory findings represented the most common pathologic changes and included (1) Hirano-like inclusions in the thalamic neurons, (2) neuroaxonal dystrophy in the medulla oblongata, (3) periodic acid-Schiff-positive granular deposits in the neuropil of the hippocampus, and (4) progressive neuroinflammation characterized by microgliosis and astrogliosis. Neoplastic conditions, developmental abnormalities, and circulatory disorders were rarely observed incidental findings. In conclusion, this study describes spontaneous age-related brain lesions of the C57BL/6J mouse and provides a reference for evaluating and interpreting the neuropathological phenotype in genetically engineered mouse models developed and maintained on this congenic background.

Increased mitophagy in the skeletal muscle of spinal and bulbar muscular atrophy patients.

Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular disorder caused by polyglutamine expansion in the androgen receptor (AR) and characterized by the loss of lower motor neurons. Here we investigated pathological processes occurring in muscle biopsy specimens derived from SBMA patients and, as controls, age-matched healthy subjects and patients suffering from amyotrophic lateral sclerosis (ALS) and neurogenic atrophy. We detected atrophic fibers in the muscle of SBMA, ALS and neurogenic atrophy patients. In addition, SBMA muscle was characterized by the presence of a large number of hypertrophic fibers, with oxidative fibers having a larger size compared with glycolytic fibers. Polyglutamine-expanded AR expression was decreased in whole muscle, yet enriched in the nucleus, and localized to mitochondria. Ultrastructural analysis revealed myofibrillar disorganization and streaming in zones lacking mitochondria and degenerating mitochondria. Using molecular (mtDNA copy number), biochemical (citrate synthase and respiratory chain enzymes) and morphological (dark blue area in nicotinamide adenine dinucleotide-stained muscle cross-sections) analyses, we found a depletion of the mitochondria associated with enhanced mitophagy. Mass spectrometry analysis revealed an increase of phosphatidylethanolamines and phosphatidylserines in mitochondria isolated from SBMA muscles, as well as a 50% depletion of cardiolipin associated with decreased expression of the cardiolipin synthase gene. These observations suggest a causative link between nuclear polyglutamine-expanded AR accumulation, depletion of mitochondrial mass, increased mitophagy and altered mitochondrial membrane composition in SBMA muscle patients. Given the central role of mitochondria in cell bioenergetics, therapeutic approaches toward improving the mitochondrial network are worth considering to support SBMA patients.

PARL: The mitochondrial rhomboid protease.

The rhomboid family comprises evolutionary conserved intramembrane proteases involved in a wide spectrum of biologically relevant activities. A mitochondrion-localized rhomboid, called PARL in mammals, and conserved in yeast and Drosophila as RBD1/PCP1 and rho-7, respectively, plays an indispensable role in cell homeostasis as illustrated by the severe phenotypes caused by its genetic ablation in the various investigated species. Although several substrates of PARL have been proposed to explain these phenotypes, there remains a lot of controversy in this important area of research. We review here the putative functions and substrates of PARL and its orthologues in different species, highlighting areas of uncertainty, and discuss its potential involvement in some prevalent diseases such as type II diabetes and Parkinson's disease.

Deficiency of the miR-29a/b-1 cluster leads to ataxic features and cerebellar alterations in mice.

miR-29 is expressed strongly in the brain and alterations in expression have been linked to several neurological disorders. To further explore the function of this miRNA in the brain, we generated miR-29a/b-1 knockout animals. Knockout mice develop a progressive disorder characterized by locomotor impairment and ataxia. The different members of the miR-29 family are strongly expressed in neurons of the olfactory bulb, the hippocampus and in the Purkinje cells of the cerebellum. Morphological analysis showed that Purkinje cells are smaller and display less dendritic arborisation compared to their wildtype littermates. In addition, a decreased number of parallel fibers form synapses on the Purkinje cells. We identified several mRNAs significantly up-regulated in the absence of the miR-29a/b-1 cluster. At the protein level, however, the voltage-gated potassium channel Kcnc3 (Kv3.3) was significantly up-regulated in the cerebella of the miR-29a/b knockout mice. Dysregulation of KCNC3 expression may contribute to the ataxic phenotype.

Impaired copper and iron metabolism in blood cells and muscles of patients affected by copper deficiency myeloneuropathy.

AIMS: Severe copper deficiency leads in humans to a treatable multisystem disease characterized by anaemia and degeneration of spinal cord and nerves, but its mechanisms have not been investigated. We tested whether copper deficit leads to alterations in fundamental copper-dependent proteins and in iron metabolism in blood and muscles of patients affected by copper deficiency myeloneuropathy, and if these metabolic abnormalities are associated with compensatory mechanisms for copper maintenance. METHODS: We evaluated the expression of critical copper enzymes, of iron-related proteins, and copper chaperones and transporters in blood and muscles from five copper-deficient patients presenting with subacute sensory ataxia, muscle paralysis, liver steatosis and variable anaemia. Severe copper deficiency was caused by chronic zinc intoxication in all of the patients, with an additional history of gastrectomy in two cases. RESULTS: The antioxidant enzyme SOD1 and subunit 2 of cytochrome c oxidase were significantly decreased in blood cells and in muscles of copper-deficient patients compared with controls. In muscle, the iron storage protein ferritin was dramatically reduced despite normal serum ferritin, and the expression of the haem-proteins cytochrome c and myoglobin was impaired. Muscle expression of the copper transporter CTR1 and of the copper chaperone CCS, was strikingly increased, while antioxidant protein 1 was diminished. CONCLUSIONS: copper-dependent enzymes with critical functions in antioxidant defences, in mitochondrial energy production, and in iron metabolism are affected in blood and muscles of patients with profound copper deficiency leading to myeloneuropathy. Homeostatic mechanisms are strongly activated to increase intracellular copper retention.

Local mitochondrial replication in the periphery of neurons requires the eEF1A1 protein and thetranslation of nuclear-encoded proteins.

In neurons, it is commonly assumed that mitochondrial replication only occurs in the cell body, after which the mitochondria must travel to the neuron's periphery. However, while mitochondrial DNA replication has been observed to occur away from the cell body, the specific mechanisms involved remain elusive. Using EdU-labelling in mouse primary neurons, we developed a tool to determine the mitochondrial replication rate. Taking of advantage of microfluidic devices, we confirmed that mitochondrial replication also occurs locally in the periphery of neurons. To achieve this, mitochondria require de novo nuclear-encoded, but not mitochondrial-encoded protein translation. Following a proteomic screen comparing synaptic with non-synaptic mitochondria, we identified two elongation factors - eEF1A1 and TUFM - that were upregulated in synaptic mitochondria. We found that mitochondrial replication is impaired upon the downregulation of eEF1A1, and this is particularly relevant in the periphery of neurons.

Phenotype variability and natural history of X-linked myopathy with excessive autophagy.

OBJECTIVE: X-linked myopathy with excessive autophagy (XMEA) linked to the VMA21 gene leads to autophagy failure with progressive vacuolation and atrophy of skeletal muscles. Current knowledge of this rare disease is limited. Our objective was to define the clinical, radiological, and natural history of XMEA. METHODS: We conducted a retrospective study collecting clinical, genetic, muscle imaging, and biopsy data of XMEA patients followed in France and reviewed the literature for additional cases. RESULTS: Eighteen males had genetically confirmed XMEA in France, carrying four different VMA21 variants. Mean age at disease onset was 9.4 ± 9.9 (range 1-40) years. In 14/18 patients (77.8%), onset occurred during childhood (< 15 years); however in four patients, the disease started in adulthood. Patients had anterior and medial compartment thigh muscle weakness, distal contractures (56.3%), elevated CK levels (1287.9 ± 757.8 U/l) and autophagic vacuoles with sarcolemmal features on muscle histopathology. Muscle MRI (n = 10) showed a characteristic pattern of lower limb muscle involvement. In 11 patients, outcome measures were available for an average follow-up period of 10.6 ± 9.8 years and six of them show disease progression. Mean change of functional outcomes was 0.5 ± 1.2 points for Brooke and 2.2 ± 2.5 points for Vignos score, 7/16 patients (43.8%) needed a walking aid and 3/16 (18.8%) were wheelchair-bound (median age of 40 years old, range 39-48). The variant c.164-7 T > G was associated with a later onset of symptoms. Respiratory insufficiency was common (57.1%) but cardiac involvement rare (12.5%). INTERPRETATION: XMEA has variable age of onset, but a characteristic clinical, histopathological, and muscle imaging presentation, guiding the diagnosis. Although slowly, motor disability progresses with time, and relevant genotype-phenotype correlations will help design future clinical trials.

Multidisciplinary team meetings in treatment of spinal muscular atrophy adult patients: a real-life observatory for innovative treatments.

BACKGROUND: In 2017, a new treatment by nusinersen, an antisense oligonucleotide delivered by repeated intrathecal injections, became available for patients with spinal muscular atrophy (SMA), whereas clinical trials had mainly involved children. Since 2020, the oral, selective SMN2-splicing modifier risdiplam has been available with restrictions evolving with time. In this peculiar context of lack of data regarding adult patients, many questions were raised to define the indications of treatment and the appropriate follow-up in this population. To homogenize access to treatment in France, a national multidisciplinary team meeting dedicated to adult SMA patients, named SMA multidisciplinary team meeting, (SMDTs) was created in 2018. Our objective was to analyze the value of SMDTs in the decision-making process in SMA adult patients and to provide guidelines about treatment. METHODS: From October 2020 to September 2021, data extracted from the SMDT reports were collected. The primary outcome was the percentage of cases in which recommendations on validating treatment plans were given. The secondary outcomes were type of treatment requested, description of expectations regarding treatment and description of recommendations or follow-up and discontinuation. Data were analyzed using descriptive statistics. Comparisons between the type of treatment requested were performed using Mann-Whitney test or the Student t test for quantitative data and the Fisher's exact test or the χ2 test for qualitative data. RESULTS: Cases of 107 patients were discussed at the SMDTs with a mean age of 35.3 (16-62). Forty-seven were SMA type 2, and 57 SMA type 3. Twelve cases were presented twice. Out of 122 presentations to the SMDTs, most of requests related to the initiation of a treatment (nusinersen (n = 46), risdiplam (n = 54), treatment without mentioning preferred choice (n = 5)) or a switch of treatment (n = 12). Risdiplam requests concerned significantly older patients (p = 0.002), mostly SMA type 2 (p < 0.0001), with greater disease severity in terms of motor and respiratory function compared to requests for nusinersen. In the year prior to presentation to the SMDTs, most of the patients experienced worsening of motor weakness assessed by functional tests as MFM32 or other meaningful scales for the most severe patients. Only 12% of the patients discussed had a stable condition. Only 49/122 patients (40.1%) expressed clear expectations regarding treatment. The treatment requested was approved by the SMDTs in 72 patients (67.2%). The most common reasons to decline treatment were lack of objective data on the disease course prior discussion to the SMDTs or inappropriate patient's expectations. Treatment requests were more likely to be validated by the SMDTs if sufficient pre-therapeutic functional assessment had been performed to assess the natural history (55% vs. 32%) and if the patient had worsening rather than stable motor function (p = 0.029). In patients with approved treatment, a-priori criteria to define a further ineffectiveness of treatment (usually after 14 months of treatment) were proposed for 67/72 patients. CONCLUSIONS: In the context of costly treatments with few controlled studies in adults with SMA, in whom assessment of efficacy can be complex, SMDTs are 'real-world observatories' of great interest to establish national recommendations about indications of treatment and follow-up.

Unravelling undiagnosed rare disease cases by HiFi long-read genome sequencing.

Solve-RD is a pan-European rare disease (RD) research program that aims to identify disease-causing genetic variants in previously undiagnosed RD families. We utilised 10-fold coverage HiFi long-read sequencing (LRS) for detecting causative structural variants (SVs), single nucleotide variants (SNVs), insertion-deletions (InDels), and short tandem repeat (STR) expansions in extensively studied RD families without clear molecular diagnoses. Our cohort includes 293 individuals from 114 genetically undiagnosed RD families selected by European Rare Disease Network (ERN) experts. Of these, 21 families were affected by so-called 'unsolvable' syndromes for which genetic causes remain unknown, and 93 families with at least one individual affected by a rare neurological, neuromuscular, or epilepsy disorder without genetic diagnosis despite extensive prior testing. Clinical interpretation and orthogonal validation of variants in known disease genes yielded thirteen novel genetic diagnoses due to de novo and rare inherited SNVs, InDels, SVs, and STR expansions. In an additional four families, we identified a candidate disease-causing SV affecting several genes including an MCF2 / FGF13 fusion and PSMA3 deletion. However, no common genetic cause was identified in any of the 'unsolvable' syndromes. Taken together, we found (likely) disease-causing genetic variants in 13.0% of previously unsolved families and additional candidate disease-causing SVs in another 4.3% of these families. In conclusion, our results demonstrate the added value of HiFi long-read genome sequencing in undiagnosed rare diseases.