Genetic landscape of pediatric acute liver failure of indeterminate origin.

BACKGROUND AND AIMS: Pediatric acute liver failure (PALF) is a life-threatening condition. In Europe, the main causes are viral infections (12%-16%) and inherited metabolic diseases (14%-28%). Yet, in up to 50% of cases the underlying etiology remains elusive, challenging clinical management, including liver transplantation. We systematically studied indeterminate PALF cases referred for genetic evaluation by whole-exome sequencing (WES), and analyzed phenotypic and biochemical markers, and the diagnostic yield of WES in this condition. APPROACH AND RESULTS: With this international, multicenter observational study, patients (0-18 y) with indeterminate PALF were analyzed by WES. Data on the clinical and biochemical phenotype were retrieved and systematically analyzed. RESULTS: In total, 260 indeterminate PALF patients from 19 countries were recruited between 2011 and 2022, of whom 59 had recurrent PALF. WES established a genetic diagnosis in 37% of cases (97/260). Diagnostic yield was highest in children with PALF in the first year of life (41%), and in children with recurrent acute liver failure (64%). Thirty-six distinct disease genes were identified. Defects in NBAS (n=20), MPV17 (n=8), and DGUOK (n=7) were the most frequent findings. When categorizing, the most frequent were mitochondrial diseases (45%), disorders of vesicular trafficking (28%), and cytosolic aminoacyl-tRNA synthetase deficiencies (10%). One-third of patients had a fatal outcome. Fifty-six patients received liver transplantation. CONCLUSIONS: This study elucidates a large contribution of genetic causes in PALF of indeterminate origin with an increasing spectrum of disease entities. The high proportion of diagnosed cases and potential treatment implications argue for exome or in future rapid genome sequencing in PALF diagnostics.

Complement and MHC patterns can provide the diagnostic framework for inflammatory neuromuscular diseases.

Histopathological analysis stands as the gold standard for the identification and differentiation of inflammatory neuromuscular diseases. These disorders continue to constitute a diagnostic challenge due to their clinical heterogeneity, rarity and overlapping features. To establish standardized protocols for the diagnosis of inflammatory neuromuscular diseases, the development of cost-effective and widely applicable tools is crucial, especially in settings constrained by limited resources. The focus of this review is to emphasize the diagnostic value of major histocompatibility complex (MHC) and complement patterns in the immunohistochemical analysis of these diseases. We explore the immunological background of MHC and complement signatures that characterize inflammatory features, with a specific focus on idiopathic inflammatory myopathies. With this approach, we aim to provide a diagnostic algorithm that may improve and simplify the diagnostic workup based on a limited panel of stainings. Our approach acknowledges the current limitations in the field of inflammatory neuromuscular diseases, particularly the scarcity of large-scale, prospective studies that validate the diagnostic potential of these markers. Further efforts are needed to establish a consensus on the diagnostic protocol to effectively distinguish these diseases.

Alteration of LARGE1 abundance in patients and a mouse model of 5q-associated spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a neuromuscular disorder caused by recessive pathogenic variants affecting the survival of motor neuron (SMN1) gene (localized on 5q). In consequence, cells lack expression of the corresponding protein. This pathophysiological condition is clinically associated with motor neuron (MN) degeneration leading to severe muscular atrophy. Additionally, vulnerability of other cellular populations and tissues including skeletal muscle has been demonstrated. Although the therapeutic options for SMA have considerably changed, treatment responses may differ thus underlining the persistent need for validated biomarkers. To address this need and to identify novel marker proteins for SMA, we performed unbiased proteomic profiling on cerebrospinal fluid derived (CSF) from genetically proven SMA type 1-3 cases and afterwards performed ELISA studies on CSF and serum samples to validate the potential of a novel biomarker candidates in both body fluids. To further decipher the pathophysiological impact of this biomarker, immunofluorescence studies were carried out on spinal cord and skeletal muscle derived from a 5q-SMA mouse model. Proteomics revealed increase of LARGE1 in CSF derived from adult patients showing a clinical response upon treatment with nusinersen. Moreover, LARGE1 levels were validated in CSF samples of further SMA patients (type 1-3) by ELISA. These studies also unveiled a distinguishment between groups in improvement of motor skills: adult patients do present with lowered level per se at baseline visit while no elevation upon treatment in the pediatric cohort can be observed. ELISA-based studies of serum samples showed no changes in the pediatric cohort but unraveled elevated level in adult patients responding to future intervention with nusinersen, while non-responders did not show a significant increase. Additional immunofluorescence studies of LARGE1 in MN and skeletal muscle of a SMA type 3 mouse model revealed an increase of LARGE1 during disease progression. Our combined data unraveled LARGE1 as a protein dysregulated in serum and CSF of SMA-patients (and in MN and skeletal muscle of SMA mice) holding the potential to serve as a disease marker for SMA and enabling to differentiate between patients responding and non-responding to therapy with nusinersen.

Effect of nusinersen on motor, respiratory and bulbar function in early-onset spinal muscular atrophy.

5q-associated spinal muscular atrophy is a rare neuromuscular disorder with the leading symptom of a proximal muscle weakness. Three different drugs have been approved by the European Medicines Agency and Food and Drug Administration for the treatment of spinal muscular atrophy patients, however, long-term experience is still scarce. In contrast to clinical trial data with restricted patient populations and short observation periods, we report here real-world evidence on a broad spectrum of patients with early-onset spinal muscular atrophy treated with nusinersen focusing on effects regarding motor milestones, and respiratory and bulbar insufficiency during the first years of treatment. Within the SMArtCARE registry, all patients under treatment with nusinersen who never had the ability to sit independently before the start of treatment were identified for data analysis. The primary outcome of this analysis was the change in motor function evaluated with the Children's Hospital of Philadelphia Infant Test of Neuromuscular Disorders and motor milestones considering World Health Organization criteria. Further, we evaluated data on the need for ventilator support and tube feeding, and mortality. In total, 143 patients with early-onset spinal muscular atrophy were included in the data analysis with a follow-up period of up to 38 months. We observed major improvements in motor function evaluated with the Children's Hospital of Philadelphia Infant Test of Neuromuscular Disorders. Improvements were greater in children >2 years of age at start of treatment than in older children. 24.5% of children gained the ability to sit independently. Major improvements were observed during the first 14 months of treatment. The need for intermittent ventilator support and tube feeding increased despite treatment with nusinersen. Our findings confirm the increasing real-world evidence that treatment with nusinersen has a dramatic influence on disease progression and survival in patients with early-onset spinal muscular atrophy. Major improvements in motor function are seen in children younger than 2 years at the start of treatment. Bulbar and respiratory function needs to be closely monitored, as these functions do not improve equivalent to motor function.

Bi-allelic variants of FILIP1 cause congenital myopathy, dysmorphism and neurological defects.

Filamin-A-interacting protein 1 (FILIP1) is a structural protein that is involved in neuronal and muscle function and integrity and interacts with FLNa and FLNc. Pathogenic variants in filamin-encoding genes have been linked to neurological disorders (FLNA) and muscle diseases characterized by myofibrillar perturbations (FLNC), but human diseases associated with FILIP1 variants have not yet been described. Here, we report on five patients from four unrelated consanguineous families with homozygous FILIP1 variants (two nonsense and two missense). Functional studies indicated altered stability of the FILIP1 protein carrying the p.[Pro1133Leu] variant. Patients exhibit a broad spectrum of neurological symptoms including brain malformations, neurodevelopmental delay, muscle weakness and pathology and dysmorphic features. Electron and immunofluorescence microscopy on the muscle biopsy derived from the patient harbouring the homozygous p.[Pro1133Leu] missense variant revealed core-like zones of myofibrillar disintegration, autophagic vacuoles and accumulation of FLNc. Proteomic studies on the fibroblasts derived from the same patient showed dysregulation of a variety of proteins including FLNc and alpha-B-crystallin, a finding (confirmed by immunofluorescence) which is in line with the manifestation of symptoms associated with the syndromic phenotype of FILIP1opathy. The combined findings of this study show that the loss of functional FILIP1 leads to a recessive disorder characterized by neurological and muscular manifestations as well as dysmorphic features accompanied by perturbed proteostasis and myopathology.

The emerging spectrum of fetal acetylcholine receptor antibody-related disorders (FARAD).

In utero exposure to maternal antibodies targeting the fetal acetylcholine receptor isoform (fAChR) can impair fetal movement, leading to arthrogryposis multiplex congenita (AMC). Fetal AChR antibodies have also been implicated in apparently rare, milder myopathic presentations termed fetal acetylcholine receptor inactivation syndrome (FARIS). The full spectrum associated with fAChR antibodies is still poorly understood. Moreover, since some mothers have no myasthenic symptoms, the condition is likely underreported, resulting in failure to implement effective preventive strategies. Here we report clinical and immunological data from a multicentre cohort (n = 46 cases) associated with maternal fAChR antibodies, including 29 novel and 17 previously reported with novel follow-up data. Remarkably, in 50% of mothers there was no previously established myasthenia gravis (MG) diagnosis. All mothers (n = 30) had AChR antibodies and, when tested, binding to fAChR was often much greater than that to the adult AChR isoform. Offspring death occurred in 11/46 (23.9%) cases, mainly antenatally due to termination of pregnancy prompted by severe AMC (7/46, 15.2%), or during early infancy, mainly from respiratory failure (4/46, 8.7%). Weakness, contractures, bulbar and respiratory involvement were prominent early in life, but improved gradually over time. Facial (25/34; 73.5%) and variable peripheral weakness (14/32; 43.8%), velopharyngeal insufficiency (18/24; 75%) and feeding difficulties (16/36; 44.4%) were the most common sequelae in long-term survivors. Other unexpected features included hearing loss (12/32; 37.5%), diaphragmatic paresis (5/35; 14.3%), CNS involvement (7/40; 17.5%) and pyloric stenosis (3/37; 8.1%). Oral salbutamol used empirically in 16/37 (43.2%) offspring resulted in symptom improvement in 13/16 (81.3%). Combining our series with all previously published cases, we identified 21/85 mothers treated with variable combinations of immunotherapies (corticosteroids/intravenous immunoglobulin/plasmapheresis) during pregnancy either for maternal MG symptom control (12/21 cases) or for fetal protection (9/21 cases). Compared to untreated pregnancies (64/85), maternal treatment resulted in a significant reduction in offspring deaths (P < 0.05) and other complications, with treatment approaches involving intravenous immunoglobulin/ plasmapheresis administered early in pregnancy most effective. We conclude that presentations due to in utero exposure to maternal (fetal) AChR antibodies are more common than currently recognized and may mimic a wide range of neuromuscular disorders. Considering the wide clinical spectrum and likely diversity of underlying mechanisms, we propose 'fetal acetylcholine receptor antibody-related disorders' (FARAD) as the most accurate term for these presentations. FARAD is vitally important to recognize, to institute appropriate management strategies for affected offspring and to improve outcomes in future pregnancies. Oral salbutamol is a symptomatic treatment option in survivors.

Homozygous WASHC4 variant in two sisters causes a syndromic phenotype defined by dysmorphisms, intellectual disability, profound developmental disorder, and skeletal muscle involvement.

Recessive variants in WASHC4 are linked to intellectual disability complicated by poor language skills, short stature, and dysmorphic features. The protein encoded by WASHC4 is part of the Wiskott-Aldrich syndrome protein and SCAR homolog family, co-localizes with actin in cells, and promotes Arp2/3-dependent actin polymerization in vitro. Functional studies in a zebrafish model suggested that WASHC4 knockdown may also affect skeletal muscles by perturbing protein clearance. However, skeletal muscle involvement has not been reported so far in patients, and precise biochemical studies allowing a deeper understanding of the molecular etiology of the disease are still lacking. Here, we report two siblings with a homozygous WASHC4 variant expanding the clinical spectrum of the disease and provide a phenotypical comparison with cases reported in the literature. Proteomic profiling of fibroblasts of the WASHC4-deficient patient revealed dysregulation of proteins relevant for the maintenance of the neuromuscular axis. Immunostaining on a muscle biopsy derived from the same patient confirmed dysregulation of proteins relevant for proper muscle function, thus highlighting an affliction of muscle cells upon loss of functional WASHC4. The results of histological and coherent anti-Stokes Raman scattering microscopic studies support the concept of a functional role of the WASHC4 protein in humans by altering protein processing and clearance. The proteomic analysis confirmed key molecular players in vitro and highlighted, for the first time, the involvement of skeletal muscle in patients. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

[Update on drug treatment of spinal muscular atrophy]. / Update zur medikamentösen Behandlung der spinalen Muskelatrophie.

BACKGROUND: The 5q-associated spinal muscular atrophy (SMA) is a hereditary motor neuron disease leading to progressive tetraplegia, often involving the bulbopharyngeal and respiratory muscle groups. The disease usually manifests in early childhood and, if untreated, is progressive throughout life and associated with numerous complications depending on the severity. Since 2017, genetically based therapeutic mechanisms are now available that correct the causative deficiency of survival motor neuron (SMN) protein and lead to significant modifications in disease progression. As the number of treatment options increases, the question of which patient is suitable for which treatment also arises. OBJECTIVE: This review article provides an update on the current treatment strategies for SMA in children and adults.

[Essen transition model for neuromuscular diseases]. / "Essener Transitionsmodell" bei neuromuskulären Erkrankungen.

BACKGROUND: Advances in healthcare systems with new therapeutic options improve the life expectancy of patients with neuromuscular diseases. With this, a shift in the phenotype of the diseases from the neuromuscular system towards other organs is more frequently observed, requiring closer interdisciplinary cooperation in caring for these young adults. Therefore, the transition to the adult caring system is nowadays a multilayered transfer with the need for complex care of these patients. OBJECTIVE: How can the transitional process be efficiently structured to combine the therapeutic effort of each specialist discipline involved and improve the healthcare process and quality of life in young adults with neuromuscular diseases? MATERIAL AND METHOD: The Departments of Neuropediatrics and Neurology of the University Medicine Essen established the Essen transition model for a structured transitional process. A concept of care was developed for the late onset Pompe's disease, Duchenne muscular dystrophy and juvenile myasthenia gravis representatively for neuromuscular diseases. It consists of four components: 1) In a standardized operational procedure (SOP), general processes, clinical diagnostic steps and guidance of treatment between the two departments are harmonized and specified. 2) The young adults and their relatives are seen in a joint consultation of both disciplines allowing a comprehensive handover for healthcare professionals. 3) In a quarterly meeting, transition conference representatives from the different specialized disciplines from pediatric and adult medicine get together for a case-related interdisciplinary exchange. 4) An interdepartmental transitional database was created to integrate all diagnostic results and parameters as a common information platform and data basis. CONCLUSION: The Essen transition model aims to close a gap in the transition of patients with neuromuscular diseases and improve healthcare in these patients with their complex needs.

A de novo CSDE1 variant causing neurodevelopmental delay, intellectual disability, neurologic and psychiatric symptoms in a child of consanguineous parents.

CSDE1 encodes the cytoplasmic cold shock domain-containing protein E1 (CSDE1), which is highly conserved across species and functions as an RNA-binding protein involved in translationally coupled mRNA turnover. CSDE1 displays a bidirectional role: promoting and repressing the translation of RNAs but also increasing and decreasing the abundance of RNAs. Preclinical studies highlighted an involvement of CSDE1 in different forms of cancer. Moreover, CSDE1 is highly expressed in human embryonic stem cells and plays a role in neuronal migration and differentiation. A genome-wide association study suggested CSDE1 as a potential autism-spectrum disorder risk gene. A multicenter next generation sequencing approach unraveled likely causative heterozygous variants in CSDE1 in 18 patients, identifying a new autism spectrum disorder-related syndrome consisting of autism, intellectual disability, and neurodevelopmental delay. Since then, no further patients with CSDE1 variants have been reported in the literature. Here, we report a 9.5-year-old girl from a consanguineous family of Turkish origin suffering from profound delayed speech and motor development, moderate intellectual disability, neurologic and psychiatric symptoms as well as hypoplasia of corpus callosum and mildly reduced brain volume on brain magnetic resonance imaging associated with a recurrent de novo mutation in CSDE1 (c.367C > T; p.R123*) expanding the phenotypical spectrum associated with pathogenic CSDE1 variants.

Cathepsin D as biomarker in cerebrospinal fluid of nusinersen-treated patients with spinal muscular atrophy.

BACKGROUND AND PURPOSE: The therapeutic landscape of spinal muscular atrophy (SMA) has changed dramatically during the past 4 years, but treatment responses differ remarkably between individuals, and therapeutic decision-making remains challenging, underlining the persistent need for validated biomarkers. METHODS: We applied untargeted proteomic analyses to determine biomarkers in cerebrospinal fluid (CSF) samples of SMA patients under treatment with nusinersen. Identified candidate proteins were validated in CSF samples of SMA patients by Western blot and enzyme-linked immunosorbent assay. Furthermore, levels of peripheral neurofilament heavy and light chain were determined. RESULTS: Untargeted proteomic analysis of CSF samples of three SMA type 1 patients revealed the lysosomal protease cathepsin D as a candidate biomarker. Subsequent validation analysis in a larger cohort of 31 pediatric SMA patients (type 1, n = 12; type 2, n = 9; type 3, n = 6; presymptomatically treated, n = 4; age = 0-16 years) revealed a significant decline of cathepsin D levels in SMA patients aged ≥2 months at the start of treatment. Although evident in all older age categories, this decline was only significant in the group of patients who showed a positive motor response. Moreover, downregulation of cathepsin D was evident in muscle biopsies of SMA patients. CONCLUSIONS: We identified a decline of cathepsin D levels in CSF samples of SMA patients under nusinersen treatment that was more pronounced in the group of "treatment responders" than in "nonresponders." We believe that our results indicate a suitability of cathepsin D levels as a possible biomarker in SMA also in older patients, in combination with analysis of peripheral neurofilament light chain in adolescents or alone in adult patients.

[Newborn screening program for spinal muscular atrophy]. / Neugeborenenscreeningprogramm für die spinale Muskelatrophie.

BACKGROUND: The introduction of a comprehensive newborn screening program for spinal muscular atrophy (SMA), specifically for 5q-SMA, is planned for the end of 2021 in Germany. Several targeted treatment options have become available for all patients with SMA. MATERIAL AND METHODS: Newborn screening for 5q-SMA is based on the detection of a homozygous deletion of exon 7 in the SMN1 gene by molecular genetic analysis from the dried blood card. In all cases a second blood sample must be drawn as a part of confirmation diagnostics including the determination of the SMN2 copy numbers. RESULTS: Insights from pilot projects performed in parts of Germany are presented. Advantages and disadvantages of the screening project are discussed. CONCLUSION: Consultation and treatment should be carried out in a department of neuropediatrics with experience in the treatment of children with 5q-SMA, which is able to provide all current treatment options for the child, so that, when necessary, the treatment can be started within the first month of life.

Inflammation, fibrosis and skeletal muscle regeneration in LGMDR9 are orchestrated by macrophages.

AIMS: Variable degrees of inflammation, necrosis, regeneration and fibrofatty replacement are part of the pathological spectrum of the dystrophic process in alpha dystroglycanopathy LGMDR9 (FKRP-related, OMIM #607155), one of the most prevailing types of LGMDs worldwide. Inflammatory processes and their complex interplay with vascular, myogenic and mesenchymal cells may have a major impact on disease development. The purpose of our study is to describe the specific immune morphological features in muscle tissue of patients with LGMDR9 to enable a better understanding of the phenotype of muscle damage leading to disease progression. METHODS: We have analysed skeletal muscle biopsies of 17 patients genetically confirmed as having LGMDR9 by histopathological and molecular techniques. RESULTS: We identified CD206+ MHC class II+ and STAT6+ immune-repressed macrophages dominating the endomysial infiltrate in areas of myofibre regeneration and fibrosis. Additionally, PDGFRß+ pericytes were located around MHC class II+ activated capillaries residing in close proximity to areas of fibrosis and regenerating fibres. Expression of VEGF was found on many regenerating neonatal myosin+ fibres, myofibres and CD206+ macrophages also co-expressed VEGF. CONCLUSION: Our results show characteristic immune inflammatory features in LGMDR9 and more specifically shed light on the predominant role of macrophages and their function in vascular organisation, fibrosis and myogenesis. Understanding disease-specific immune phenomena potentially inform about possibilities for anti-fibrotic and anti-inflammatory therapeutic strategies, which may complement Ribitol replacement and gene therapies for LGMDR9 that may be available in the future.

Intellectual disability associated with craniofacial dysmorphism, cleft palate, and congenital heart defect due to a de novo MEIS2 mutation: A clinical longitudinal study.

Intellectual disability (ID) has an estimated prevalence of 1.5%-2%. Whole exome sequencing (WES) studies have identified a multitude of novel causative gene defects and have shown that sporadic ID cases result from de novo mutations in genes associated with ID. Here, we report on a 10-year-old girl, who has been regularly presented in our neuropediatric and genetic outpatient clinic. A median cleft palate and a heart defect were surgically corrected in infancy. Apart from ID, she has behavioral anomalies, muscular hypotonia, scoliosis, and hypermobile joints. The facial phenotype is characterized by arched eyebrows, mildly upslanting long palpebral fissures, prominent nasal tip, and large, protruding ears. Trio WES revealed a de novo missense variant in MEIS2 (c.998G>A; p.Arg333Lys). Haploinsufficiency of MEIS2 had been discussed as the most likely mechanism of the microdeletion 5q14-associated complex phenotype with ID, cleft palate, and heart defect. Recently, four studies including in total 17 individuals with intragenic MEIS2 variants were reported. Here we present the evolution of the clinical phenotype and compare with the data of known individuals.

Expanding the clinical and molecular spectrum of ATP6V1A related metabolic cutis laxa.

Several inborn errors of metabolism show cutis laxa as a highly recognizable feature. One group of these metabolic cutis laxa conditions is autosomal recessive cutis laxa type 2 caused by defects in v-ATPase components or the mitochondrial proline cycle. Besides cutis laxa, muscular hypotonia and cardiac abnormalities are hallmarks of autosomal recessive cutis laxa type 2D (ARCL2D) due to pathogenic variants in ATP6V1A encoding subunit A of the v-ATPase. Here, we report on three affected individuals from two families with ARCL2D in whom we performed whole exome and Sanger sequencing. We performed functional studies in fibroblasts from one individual, summarized all known probands' clinical, molecular, and biochemical features and compared them, also to other metabolic forms of cutis laxa. We identified novel missense and the first nonsense variant strongly affecting ATP6V1A expression. All six ARCL2D affected individuals show equally severe cutis laxa and dysmorphism at birth. While for one no information was available, two died in infancy and three are now adolescents with mild or absent intellectual disability. Muscular weakness, ptosis, contractures, and elevated muscle enzymes indicated a persistent myopathy. In cellular studies, a fragmented Golgi compartment, a delayed Brefeldin A-induced retrograde transport and glycosylation abnormalities were present in fibroblasts from two individuals. This is the second and confirmatory report on pathogenic variants in ATP6V1A as the cause of this extremely rare condition and the first to describe a nonsense allele. Our data highlight the tremendous clinical variability of ATP6V1A related phenotypes even within the same family.

Muscular and Molecular Pathology Associated with SPATA5 Deficiency in a Child with EHLMRS.

Mutations in the SPATA5 gene are associated with epilepsy, hearing loss and mental retardation syndrome (EHLMRS). While SPATA5 is ubiquitously expressed and is attributed a role within mitochondrial morphogenesis during spermatogenesis, there is only limited knowledge about the associated muscular and molecular pathology. This study reports on a comprehensive workup of muscular pathology, including proteomic profiling and microscopic studies, performed on an 8-year-old girl with typical clinical presentation of EHLMRS, where exome analysis revealed two clinically relevant, compound-heterozygous variants in SPATA5. Proteomic profiling of a quadriceps biopsy showed the dysregulation of 82 proteins, out of which 15 were localized in the mitochondrion, while 19 were associated with diseases presenting with phenotypical overlap to EHLMRS. Histological staining of our patient's muscle biopsy hints towards mitochondrial pathology, while the identification of dysregulated proteins attested to the vulnerability of the cell beyond the mitochondria. Through our study we provide insights into the molecular etiology of EHLMRS and provide further evidence for a muscle pathology associated with SPATA5 deficiency, including a pathological histochemical pattern accompanied by dysregulated protein expression.

First clinical and myopathological description of a myofibrillar myopathy with congenital onset and homozygous mutation in FLNC.

Filamin C (encoded by the FLNC gene) is a large actin-cross-linking protein involved in shaping the actin cytoskeleton in response to signaling events both at the sarcolemma and at myofibrillar Z-discs of cross-striated muscle cells. Multiple mutations in FLNC are associated with myofibrillar myopathies of autosomal-dominant inheritance. Here, we describe for the first time a boy with congenital onset of generalized muscular hypotonia and muscular weakness, delayed motor development but no cardiac involvement associated with a homozygous FLNC mutation c.1325C>G (p.Pro442Arg). We performed ultramorphological, proteomic, and functional investigations as well as immunological studies of known marker proteins for dominant filaminopathies. We show that the mutant protein is expressed in similar quantities as the wild-type variant in control skeletal muscle fibers. The proteomic signature of quadriceps muscle is altered and ultrastructural perturbations are evident. Moreover, filaminopathy marker proteins are comparable both in our homozygous and a dominant control case (c.5161delG). Biochemical investigations demonstrate that the recombinant mutant protein is less stable and more prone to degradation by proteolytic enzymes than the wild-type variant. The unusual congenital presentation of the disease clearly demonstrates that homozygosity for mutations in FLNC severely aggravates the phenotype.

Diagnostic utility of small fiber analysis in skin biopsies from children with chronic pain.

INTRODUCTION: Small fiber neuropathies (SFN) are associated with a reduction in quality of life. In adults, epidermal nerve fiber density (END) analysis is recommended for the diagnosis of SFN. In children, END assessment is not often performed. We analyzed small nerve fiber innervation to elucidate the potential diagnostic role of skin biopsies in young patients with pain. METHODS: Epidermal nerve fiber density and sudomotor neurite density (SND) were assessed in skin biopsies from 26 patients aged 7 to 20 years (15 female patients) with unexplained chronic pain. The results were compared with clinical data. RESULTS: Epidermal nerve fiber density was abnormal in 50% and borderline in 35% of patients. An underlying medical condition was found in 42% of patients, including metabolic, autoimmune, and genetic disorders. DISCUSSION: Reduction of epidermal nerve fibers can be associated with treatable conditions. Therefore, the analysis of END in children with pain may help to uncover a possible cause and guide potential treatment options.

HADHA and HADHB gene associated phenotypes - Identification of rare variants in a patient cohort by Next Generation Sequencing.

The heterooctameric mitochondrial trifunctional protein (MTP), composed of four α- and ß-subunits harbours three enzymes that each perform a different function in mitochondrial fatty acid ß-oxidation. Pathogenic variants in the MTP genes (HADHA and HADHB) cause MTP deficiency, a rare autosomal recessive metabolic disorder characterized by phenotypic heterogeneity ranging from severe, early-onset, cardiac disease to milder, later-onset, myopathy and neuropathy. Since metabolic myopathies and neuropathies are a group of rare genetic disorders and their associated muscle symptoms may be subtle, the diagnosis is often delayed. Here we evaluated data of 161 patients with myopathy and 242 patients with neuropathy via next generation sequencing (NGS) and report the diagnostic yield in three patients of this cohort by the detection of disease-causing variants in the HADHA or HADHB gene. The mitigated phenotypes of this treatable disease were missed by the newborn screening, highlighting the importance of phenotype-based NGS analysis in patients with rare and clinically very variable disorders such as MTP deficiency.