The recurrent deep intronic pseudoexon-inducing variant COL6A1 c.930+189C>T results in a consistently severe phenotype of COL6-related dystrophy: Towards clinical trial readiness for splice-modulating therapy.

Collagen VI-related dystrophies (COL6-RDs) manifest with a spectrum of clinical phenotypes, ranging from Ullrich congenital muscular dystrophy (UCMD), presenting with prominent congenital symptoms and characterised by progressive muscle weakness, joint contractures and respiratory insufficiency, to Bethlem muscular dystrophy, with milder symptoms typically recognised later and at times resembling a limb girdle muscular dystrophy, and intermediate phenotypes falling between UCMD and Bethlem muscular dystrophy. Despite clinical and immunohistochemical features highly suggestive of COL6-RD, some patients had remained without an identified causative variant in COL6A1, COL6A2 or COL6A3. With combined muscle RNA-sequencing and whole-genome sequencing we uncovered a recurrent, de novo deep intronic variant in intron 11 of COL6A1 (c.930+189C>T) that leads to a dominantly acting in-frame pseudoexon insertion. We subsequently identified and have characterised an international cohort of forty-four patients with this COL6A1 intron 11 causative variant, one of the most common recurrent causative variants in the collagen VI genes. Patients manifest a consistently severe phenotype characterised by a paucity of early symptoms followed by an accelerated progression to a severe form of UCMD, except for one patient with somatic mosaicism for this COL6A1 intron 11 variant who manifests a milder phenotype consistent with Bethlem muscular dystrophy. Characterisation of this individual provides a robust validation for the development of our pseudoexon skipping therapy. We have previously shown that splice-modulating antisense oligomers applied in vitro effectively decreased the abundance of the mutant pseudoexon-containing COL6A1 transcripts to levels comparable to the in vivo scenario of the somatic mosaicism shown here, indicating that this therapeutic approach carries significant translational promise for ameliorating the severe form of UCMD caused by this common recurrent COL6A1 causative variant to a Bethlem muscular dystrophy phenotype.

Biochemical characterization of two novel mutations in the human high-affinity choline transporter 1 identified in a patient with congenital myasthenic syndrome.

Congenital myasthenic syndrome (CMS) is a heterogeneous condition associated with 34 different genes, including SLC5A7, which encodes the high-affinity choline transporter 1 (CHT1). CHT1 is expressed in presynaptic neurons of the neuromuscular junction where it uses the inward sodium gradient to reuptake choline. Biallelic CHT1 mutations often lead to neonatal lethality, and less commonly to non-lethal motor weakness and developmental delays. Here, we report detailed biochemical characterization of two novel mutations in CHT1, p.I294T and p.D349N, which we identified in an 11-year-old patient with a history of neonatal respiratory distress, and subsequent hypotonia and global developmental delay. Heterologous expression of each CHT1 mutant in human embryonic kidney cells showed two different mechanisms of reduced protein function. The p.I294T CHT1 mutant transporter function was detectable, but its abundance and half-life were significantly reduced. In contrast, the p.D349N CHT1 mutant was abundantly expressed at the cell membrane, but transporter function was absent. The residual function of the p.I294T CHT1 mutant may explain the non-lethal form of CMS in this patient, and the divergent mechanisms of reduced CHT1 function that we identified may guide future functional studies of the CHT1 myasthenic syndrome. Based on these in vitro studies that provided a diagnosis, treatment with cholinesterase inhibitor together with physical and occupational therapy significantly improved the patient's strength and quality of life.

MRM2 variants in families with complex dystonic syndromes: evidence for phenotypic heterogeneity.

BACKGROUND: Dystonia involves repetitive movements and muscle contractions leading to abnormal postures. We investigated patients in two families, DYAF11 and M, exhibiting dystonic or involuntary movement disorders. METHODS: Clinical investigations were performed for all patients. Genetic analyses included genome-wide linkage analysis and exome sequencing followed by Sanger sequencing validation. MRM2-specific transcripts were analysed from participants' blood samples in Family DYAF11 after cloning of gene-specific cDNA. RESULTS: Four affected siblings in Family DYAF11 had progressive dystonic features. Two patients in Family M exhibited a neurodevelopmental disorder accompanied by involuntary movements. In Family DYAF11, linkage was detected to the telomere at chromosome 7p22.3, spanning <2 Mb. Exome sequencing identified a donor splice-site variant, c.8+1G>T in MRM2, which segregated with the phenotype, corresponding to the linkage data since all affected individuals were homozygous while the obligate unaffected carriers were heterozygous for the variant. In the MRM2 c.8+1G>T allele, an aberrant alternative acceptor splice-site located within exon 2 was used in a subset of the transcripts, creating a frameshift in the open reading frame. Exome sequencing in Family M revealed a rare missense variant c.242C>T, p.(Ala81Val), which affected a conserved amino acid. CONCLUSIONS: Our results expand the clinical and allelic spectrum of MRM2 variants. Previously, these descriptions were based on observations in a single patient, diagnosed with mitochondrial DNA depletion syndrome 17, in whom movement disorder was accompanied by recurrent strokes and epilepsy. We also demonstrate a subset of correctly spliced tt-ag MRM2 transcripts, raising the possibility to develop treatment by understanding the disease mechanism.

Measuring ambulation with wrist-based and hip-based activity trackers for children with cerebral palsy.

AIM: To assess the accuracy of consumer available wrist-based and hip-based activity trackers in quantitatively measuring ambulation in children with cerebral palsy (CP). METHOD: Thirty-nine children (23 males, 16 females; mean age [SD] 9y 7mo [3y 5mo]; range 4-15y) with CP were fitted with trackers both on their wrist and hip. Each participant stood for 3 minutes, ambulated in a hallway, and sat for 3 minutes. The number of steps and distance were recorded on trackers and compared to manually counted steps and distance. Pearson correlation coefficients were determined for the number of steps during ambulation from each tracker and a manual count. Mean absolute error (MAE) and range of errors were calculated for steps during ambulation for each tracker and a manual count and for distance for each tracker and hallway distance. RESULTS: For the number of steps, a weak inverse relationship (r=-0.033) was found for the wrist-based tracker and a strong positive relationship (r=0.991) for the hip-based tracker. The MAE was 88 steps for the wrist-based and seven steps for the hip-based tracker. The MAE for distance was 0.06 miles for the wrist-based and 0.07 miles for the hip-based tracker. INTERPRETATION: Only the hip-based tracker provided an accurate step count; neither tracker was accurate for distance. Thus, ambulation of children with CP can be accurately quantified with readily available trackers. WHAT THIS PAPER ADDS: Consumer available activity trackers accurately measure ambulation in children with cerebral palsy (CP). The hip-based tracker is more accurate than the wrist-based tracker for children with CP. The hip-based Fitbit activity tracker accurately measures step counts of children with CP during ambulation.

MEDICIÓN DE LA AMBULACIÓN CON RASTREADORES DE ACTIVIDAD DE MUÑECA Y CADERA PARA NIÑOS CON PARÁLISIS CEREBRAL: OBJETIVO: Evaluar la precisión de los rastreadores de actividad basados ​​en la muñeca y en la cadera disponibles para el consumidor en la medición cuantitativa de la ambulación en niños con parálisis cerebral (PC) METODO: Treinta y nueve niños (23 varones, 16 mujeres; edad media [DS] 9 años y 7 meses [3 años y 5 meses]; rango 4-15 años) con PC fueron equipados con rastreadores en su muñeca y cadera. Cada participante se paró durante 3 minutos, caminó por un pasillo y se sentó durante 3 minutos. La cantidad de pasos y la distancia se registraron en los rastreadores y se compararon con los pasos y la distancia contados manualmente. Los coeficientes de correlación de Pearson se determinaron para el número de pasos durante la ambulación de cada rastreador y un conteo manual. El error absoluto medio (MAE) y el rango de errores se calcularon para los pasos durante la ambulación y la distancia del pasillo para cada rastreador y para el conteo manual. RESULTADOS: Para el número de pasos, se encontró una relación inversa débil (r = -0.033) para el rastreador ubicado en la muñeca y una relación positiva fuerte (r = 0.991) para el rastreador ubicado en la cadera. El MAE fue de 88 pasos para la muñeca y siete pasos para el rastreador de la cadera. El MAE para la distancia fue de 0.06 millas para la muñeca y 0.07 millas para el rastreador ubicado en la cadera. INTERPRETACIÓN: Solo el rastreador ubicado en la cadera proporcionó un conteo de pasos preciso; ninguno de los rastreadores era preciso para la distancia. Por lo tanto, la deambulación de los niños con PC se puede cuantificar con precisión con rastreadores fácilmente disponibles.

MEDINDO A DEAMBULAÇÃO COM RASTREADOR DE ATIVIDADE POSICIONADO NO PUNHO E QUADRIL COMERCIALMENTE DISPONÍVEL EM CRIANÇAS COM PARALISIA CEREBRAL: OBJETIVO: Avaliar a precisão de rastreadores de atividade posicionados no punho e quadril, disponíveis para o consumidor, para mensurar qualitativamente a deambulação em crianças com paralisia cerebral (PC). MÉTODO: Trinta e nove crianças (23 meninos, 16 meninas; média da idade [DP] 9 anos e 7 meses [3 anos e 5 meses]; amplitude 4-15 anos) com PC foram equipados com rastreadores em punho e quadril. Cada participante permaneceu em pé durante 3 minutos, andando em um corredor, e sentado por 3 minutos. O número de passos e distância foram registrados nos rastreadores e comparados com os passos e distância medidos manualmente. Coeficientes de correlação de Pearson foram determinados para o número de passos durante a deambulação para cada rastreador e a contagem manual. O Erro Médio Absoluto (EMA) e variância de erros foram calculados para os passos durante a deambulação para cada rastreador e a contagem manual e para a distância de cada rastreador e a distância do corredor. RESULTADOS: Para o número de passos, uma relação inversa fraca (r=-0,033) foi encontrada entre o rastreador do punho e uma relação positiva forte (r=0,991) para o reastreador do quadril. A EMA foi de 88 passos para o rastreador do punho e sete passos para o rastreador do quadril. A EMA para a distância foi de 0,06 milhas (9,66km) para o rastreador do punho e 0,07 milhas (11,26km) para o rastreador do quadril. INTERPRETAÇÃO: Somente o rastreador do quadril forneceu a contagem precisa dos passos; nenhum rastreador foi preciso para a distância. Assim, a deambulação em crianças com PC pode ser quantificada com precisão com os rastreadores atualmente disponíveis.

The Limb-Girdle Muscular Dystrophies: Is Treatment on the Horizon?

There has been an ever-expanding list of the Limb-Girdle Muscular Dystrophies (LGMD). There are currently 8 subtypes of autosomal dominant (AD) and 26 subtypes of autosomal recessive (AR) LGMD. Despite continued research efforts to conquer this group of genetic neuromuscular disease, patients continue to be treated symptomatically with the aim of prevention or addressing complications. Mouse models have been helpful in clarifying disease pathogenesis as well as strategizing pathways for treatment. Discoveries in translational research as well as molecular therapeutic approaches have kept clinicians optimistic that more promising clinical trials will lead the way to finding the cure for these devastating disorders. It is well known that the challenge for these rare diseases is the ability to assemble adequate numbers of patients for a clinically meaningful trial, but current efforts in developing patient registries have been encouraging. Natural history studies will be essential in establishing and interpreting the appropriate outcome measures for clinical trials. Nevertheless, animal studies continue to be key in providing proof of concept that will be necessary in moving research along. This review will briefly discuss each type of LGMD, highlighting their distinguishing features, then focus on research efforts that have been published in the literature for the past few years, many of which are still in the preclinical trial stage.

A Founder Mutation in VPS11 Causes an Autosomal Recessive Leukoencephalopathy Linked to Autophagic Defects.

Genetic leukoencephalopathies (gLEs) are a group of heterogeneous disorders with white matter abnormalities affecting the central nervous system (CNS). The causative mutation in ~50% of gLEs is unknown. Using whole exome sequencing (WES), we identified homozygosity for a missense variant, VPS11: c.2536T>G (p.C846G), as the genetic cause of a leukoencephalopathy syndrome in five individuals from three unrelated Ashkenazi Jewish (AJ) families. All five patients exhibited highly concordant disease progression characterized by infantile onset leukoencephalopathy with brain white matter abnormalities, severe motor impairment, cortical blindness, intellectual disability, and seizures. The carrier frequency of the VPS11: c.2536T>G variant is 1:250 in the AJ population (n = 2,026). VPS11 protein is a core component of HOPS (homotypic fusion and protein sorting) and CORVET (class C core vacuole/endosome tethering) protein complexes involved in membrane trafficking and fusion of the lysosomes and endosomes. The cysteine 846 resides in an evolutionarily conserved cysteine-rich RING-H2 domain in carboxyl terminal regions of VPS11 proteins. Our data shows that the C846G mutation causes aberrant ubiquitination and accelerated turnover of VPS11 protein as well as compromised VPS11-VPS18 complex assembly, suggesting a loss of function in the mutant protein. Reduced VPS11 expression leads to an impaired autophagic activity in human cells. Importantly, zebrafish harboring a vps11 mutation with truncated RING-H2 domain demonstrated a significant reduction in CNS myelination following extensive neuronal death in the hindbrain and midbrain. Thus, our study reveals a defect in VPS11 as the underlying etiology for an autosomal recessive leukoencephalopathy disorder associated with a dysfunctional autophagy-lysosome trafficking pathway.

Fidelity of gamma-glutamyl transferase (GGT) in differentiating skeletal muscle from liver damage.

This study tested the hypothesis that gamma-glutamyl transferase (GGT) can be used as a reliable biomarker to distinguish skeletal muscle from liver damage. Twenty-eight Duchenne muscular dystrophy subjects with proven dystrophin gene mutations were enrolled. Included were 14 ambulatory and 14 nonambulatory patients with approximately half of each cohort taking corticosteroids. Twenty normal males served as controls. Initial blood samples for serum GGT and creatine kinase were taken between 8AM and 9AM and redrawn 8 hours later to test for variability. Between blood draws, subjects resumed normal activities in a play environment or could leave the clinic. Not a single duchenne muscular dystrophy patient showed a GGT outside the control range at any time point, while creatine kinase levels were 14 to 200 times normal. Validation of this finding is essential for management of patients with muscle disorders exposed to potentially hepatotoxic drugs for clinical management or monitoring subjects participating in clinical trials.

The use of tiagabine in pediatric spasticity management.

Tiagabine, developed as an anti-epileptic medication, has the potential to reduce spasticity. The purpose of the present study was to assess the effectiveness of tiagabine in decreasing spasticity and improving the functional abilities of children with spastic cerebral palsy (CP). Nine children (seven females, two males) with CP (six spastic quadriplegia, three moderate to severe spastic diplegia) were treated with tiagabine for a mean of 7.2 months. Median age was 4y 5mo (range 3y 2mo-10y). All children were non-ambulatory. According to the Gross Motor Function Classification System, six were Level IV and three were Level V. Only one child showed a median decrease >or=1.0 grade on the modified Ashworth scale in upper extremities, lower extremities, and overall. Another child had significant improvement in the Pediatric Evaluation of Disability Inventory Self-care score and improved feeding. None of the participants was found to have a significant improvement in motor function or a decrease in the number of motions (passive range of motion and muscle length test) that were limited. Reduction of nocturnal awakenings from painful spasms was reported in one child. Eight of the nine children experienced adverse side-effects during treatment. Although tiagabine was not found to be effective in decreasing children's spasticity or improving their function, its potential use in the relief of painful spasms associated with neurological conditions in the pediatric population warrants further investigation.

Demyelinating focal motor neuropathy of the ulnar nerve masquerading as compression in Guyon's canal: a case report.

Ulnar nerve-innervated intrinsic muscle weakness, in the absence of sensory complaints or deficits, usually is the result of compression at the ulnar nerve in zone II of Guyon's canal. In rare instances the problem is not caused by a compressive neuropathy but by a demyelinating focal motor neuropathy. Demyelinating neuropathies have been well documented in the neurologic literature but they have received little attention in the hand surgery literature. We report on one such case and the importance of differentiating the 2 neuropathies. Although surgery often is necessary for a compressive neuropathy it is contraindicated for a demyelinating neuropathy.