The Ohio State University College of Medicine and the Research Institute at Nationwide Children’s Hospital, Columbus, Ohio, USA; Armatus Bio
Facioscapulohumeral muscular dystrophy (FSHD) is a potentially devastating autosomal dominant or digenic disorder caused by abnormal de-repression of the DUX4 gene in skeletal muscle. DUX4 expression causes muscle cell dysfunction and death, eventually leading to muscular dystrophy. We hypothesize that DUX4 silencing is the most direct approach to FSHD therapy, but there are currently no approved disease-modifying treatments for FSHD. To accomplish this, our research group developed a DUX4-targeted RNAi-based gene therapy for FSHD using an engineered miRNA, called mi405, delivered by AAV vectors and have extensively tested various safety and efficacy parameters in FSHD cells and animal models. First generation AAVs like AAVrh74 or AAV9 require high doses exceeding 1E14 vg/kg to achieve significant global muscle transduction and disease-modifying benefits in mice and humans. Such high doses have been associated with serious adverse events in humans, including death. To potentially reduce dose, we tested several, next generation AAV vectors engineered for myotropism and liver detargeting, including Solid Biosciences’ next-generation capsid, POLARIS-101TM (AAV-SLB101), which is currently being evaluated in the Phase 1/2 INSPIRE DUCHENNE clinical trial for Duchenne muscular dystrophy (NCT06138639). We found that POLARIS-101TM-delivered mi405 completely protected TIC-DUX4 functional, histopathological, and molecular deficits at doses one log lower than first generation vectors, thereby establishing a potential minimal effective dose for human trials in the low- to mid-E13 vg/kg range. In particular, POLARIS-101TM.mi405 normalized mouse activity in open field and rotarod tests in an aggressive FSHD mouse model at a dose of 6E13 vg/kg, and significantly or completely reduced FSHD-associated biomarkers in mice at doses as low as 9E12 vg/kg. Our data support that combining POLARIS-101TM with the highly efficacious mi405 product will enable lower and potentially safer AAV doses while maintaining or improving the therapeutic efficacy achieved using high-dose first generation vectors.
S02Allosteric activation of DNA polymerase gamma as a therapeutic strategy in POLG-related mitochondrial disease
Professor Maria Falkenberg
University of Gothenburg, Institute of Biomedicine
Mitochondrial DNA (mtDNA) replication is carried out by DNA polymerase gamma (POLγ), a heterotrimer consisting of the catalytic POLγA subunit and two accessory POLγB subunits. Mutations in POLG, encoding POLγA, are among the most common causes of inherited mitochondrial disease and currently lack effective treatment options. Restoring POLγ function therefore represents an attractive therapeutic strategy.
In collaboration with Pretzel Therapeutics, we have identified PZL-A, a small-molecule activator that restores enzymatic activity in four common disease-associated POLG variants. We now extend this work by analysing approximately 50 additional pathogenic POLG mutations to assess the broader therapeutic potential of allosteric activation. We also present structural and biochemical data that define the mechanism underlying POLγ activation by PZL-A. Using single-particle cryo-EM in combination with enzymatic assays, we map the binding site of the compound to the POLγA–POLγB interface and show that this interaction stabilizes an active conformation of the holoenzyme.
Together, these findings strengthen the rationale for therapeutic activation of POLγ and broaden the range of genotypes that may benefit from treatment with PZL-A.
University of Manchester and Manchester University NHS Foundation Trust
Why some individuals experience severe neuropathy following infection is unknown. Nucleocytoplasmic trafficking (NCT) is an essential process in nucleated cells, and its disruption has been implicated in many neurodegenerative conditions including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia.
We performed initial genomic and clinical studies in 24 children (age 6 months – 11 years) from 12 families with variable phenotypes ranging from a rapidly progressive, fatal axonal neuropathy with encephalopathy to a mild motor neuropathy resulting in impaired walking. In most patients, neurological presentation was secondary to infection, resulting in prior diagnosis of Guillain-Barré syndrome (GBS). By genome or exome sequencing, we identified deleterious biallelic variants in human RCC1, encoding a GTP exchange factor essential in maintaining Ran GTPase-dependent NCT function. The efficiency of cellular Ran GDP-GTP exchange and the thermal stability of Rcc1 protein was reduced by disease-associated variants. Heat shock or oxidative stress revealed defects in Ran nuclear localization, impaired NCT, and TDP-43 mislocalization in patient fibroblasts. Disease associated variants were unable to rescue the thermosensitive phenotype of a rcc1 deficient hamster cell line. RCC1 Drosophila models revealed a fatal intolerance to oxidative stress. Subsequent studies have identified 10 further affected children with biallelic RCC1 variants from 5 unrelated families with overlapping clinical features to the original series.
We describe a novel autosomal recessive acute onset axonal neuropathy triggered by infection caused by biallelic RCC1 variants, which mimics GBS and has important mechanistic overlap with ALS. The age to onset and development of newborn genome sequencing create a window of opportunity for therapeutic intervention.
S04Bridging the translational divide: evaluating therapeutic efficacy in the DE50-MD canine model of Duchenne Muscular Dystrophy
Professor Richard Piercy
Royal Veterinary College, London, UK
Duchenne Muscular Dystrophy (DMD) is a universally fatal, X-linked, genetic disorder affecting 1 in 5000 male births. Patients lack dystrophin, a critical protein that maintains integrity and signalling within muscle cell membranes: they display weakness as toddlers, require wheelchairs by their teens and most die of heart failure, usually in their twenties or thirties. Animal models remain crucial for investigations of pathophysiology and treatments for rare diseases in particular, given that human trials in these fields often suffer from logistic constraints, including poor availability of suitable patients, absence of blinding and randomisation, and the requisite use of historical natural history (control) data, rather than appropriately matched untreated controls. In 2012, The Wellcome Trust awarded a Biomedical Resource Award to the RVC to develop the unique DE50-MD canine model of DMD. Our work has created one of the best characterised, phenotypically relevant models of this devastating disorder; DE50-MD dogs offer unique translational potential, enabling controlled, tractable and robust treatment trials that are either inappropriate or unfeasible in other species or that are unethical in human patients. In this presentation I will introduce the DE50-MD model and its phenotype and describe treatment approaches we have evaluated within the colony, including use of gene editing and gene therapy approaches. Our work is helping bridge the gap between use of less phenotypically-relevant human models (such as with mice) and trials in humans. Our data shine lights on several critical elements related to recent historical or ongoing trials in humans that have received minimal attention and emphasise the requirement for appropriate preclinical testing in relevant animal species.
S05Programmed axon death and human disease
Professor Michael Coleman
Department of Clinical Neurosciences, University of Cambridge
Programmed axon death is a widespread and completely preventable mechanism in injury and disease. Its central players are SARM1, an enzyme that degrades NAD, and its axonal regulator NMNAT2, which synthesises NAD. NMNAT2 is essential in healthy axons to maintain a low level of its substrate, NMN, which is important because NMN is a potent activator of SARM1. If anything lowers the supply of active NMNAT2, a surprisingly unstable protein, in axons, NMN accumulates, activating SARM1 and killing axons. Such triggers include impairment of axonal transport, protein synthesis or mitochondrial respiration, axon injury, some neurotropic viruses, toxins and loss-of-function mutation of the NMNAT2 gene. Gain-of-function variants of SARM1 also exist in humans and are enriched in sporadic ALS.
Inhibitors for SARM1 are currently in clinical trials for ALS and chemotherapy-induced peripheral neuropathy. However, animal data suggest much wider involvement of programmed axon death in disease. Blocking programmed axon death alleviates models of inherited, diabetic or chemotherapy-induced peripheral neuropathy, some motor neuron disease models, traumatic brain injury, glaucoma, and viral neuropathies. The lethal phenotype of NMNAT2 null mice, which represents a rare human neuropathy, is rescued completely and permanently. However, since all models have limitations it is essential also to study these in human.
I will present our recent work using human genetics and hiPSC-derived neurons to identify human diseases and individual patients where programmed axon death is involved. This implicates NMNAT2 loss in polyneuropathy and SARM1 gain-of-function in ALS and more complex neurological conditions. Gene-environment interactions are also important, where pyridine-based neurotoxins or neurotropic viruses may combine with genetic variants to cause disease. I will discuss proposals for how SARM1 blocking drugs can be targeted where they are likely to provide greatest benefit.
S06Learning from the last 5 years of AAV clinical trials in NMD
Professor Carsten G Bönnemann
National Institute of Neurological Disorders and Stroke, MD, USA
Abstract not supplied
S07CAR-T cell therapies applied to neuromuscular diseases
S. Armando Villalta, Ph.D.1,2,3,4
1Physiology and Biophysics, University of California Irvine, Irvine, Ca, USA
2Muscle Biology and Disease and Research Center, University of California Irvine, Irvine, Ca, USA
3Institute for Immunology, University of California Irvine, Irvine, Ca, USA
4Neurology, University of California Irvine, Irvine, Ca, USA
Chimeric antigen receptor (CAR)-T cell therapy has transformed the treatment landscape for hematological malignancies over the past decade. Following the landmark clinical success of CD19-directed CAR-T cells in B cell leukemias and lymphomas, and subsequent regulatory approvals, this platform has emerged as a powerful strategy for precision immunotherapy. This lecture will provide a historical overview of CAR-T cell development, highlighting key advances in receptor design and clinical translation that established its efficacy in oncology, and will discuss the repurposing of CD19-directed CAR-T therapies for neuromuscular and autoimmune conditions. The application of CAR technology to generate antigen-specific regulatory T cells (CAR-Tregs) will then be introduced as an evolution of the platform aimed at inducing immune tolerance rather than cytotoxicity. Growing preclinical and early translational data demonstrate that CAR-Tregs exert potent tolerogenic activity in models of autoimmunity and transplantation, positioning them as a promising modality for controlling pathogenic immune responses in muscle disease and transgene immunity in gene therapy. In support of this concept, preliminary data from a mouse model of Duchenne muscular dystrophy (DMD) show that polyclonal Treg expansion suppresses the induction of dystrophin-specific T cells, providing a rationale for developing muscle-specific, antigen-directed Treg therapies to enhance potency and specificity. Finally, early efforts to engineer muscle-specific CAR-Tregs designed to promote transgene tolerance during gene therapy for DMD will be presented, with the goal of mitigating anti-transgene immunity and improving the durability and safety of gene replacement approaches.
S08Novel Treatments for Autoimmune Myasthenia Gravis
Professor Tobias Ruck
Ruhr-University Bochum, Department of Neurology, BG University Hospital Bergmannsheil Bochum, Germany
Autoimmune myasthenia gravis (MG) represents a prototypical antibody-mediated neurological disorder in which advances in immunopathological understanding have directly translated into novel therapeutic strategies. Over the past decade, treatment concepts have shifted from broad immunosuppression toward targeted, mechanism-based immunotherapies aimed at achieving rapid and sustained disease control while minimizing long-term treatment toxicity.
This lecture provides an overview of the evolving therapeutic landscape in MG within a modern treat-to-target framework. Recently approved therapies, particularly complement inhibitors and neonatal Fc receptor (FcRn) antagonists, have significantly expanded treatment options for patients with generalized MG. These agents enable rapid amelioration of autoantibody-mediated pathology and improved clinical stabilization, allowing more individualized treatment decisions.
Emerging therapeutic approaches targeting B cells and plasma cells, cytokine signaling pathways, and complement activation are currently under clinical investigation and may further refine treatment algorithms. In addition, innovative strategies such as antigen-specific tolerance induction and cellular therapies, including CAR-T–based approaches, hold promise for long-term immune modulation in refractory disease.
The increasing availability of targeted therapies necessitates dynamic and personalized treatment strategies supported by regular assessment of disease activity using validated clinical outcome measures and emerging digital monitoring tools. Remaining challenges include the management of seronegative MG, ocular manifestations, and prevention of myasthenic crisis.
Overall, the expanding spectrum of therapeutic options marks a paradigm shift toward precision medicine in MG, aiming to achieve optimal disease control, reduce corticosteroid exposure, and improve long-term functional outcomes and quality of life for affected patients.
S09Mitochondrial Donation in Clinical Practice: Insights from the UK Mitochondrial Reproductive Care Pathway
Professor Bobby McFarland
Action Medical Research Professor of Neuromuscular Disease, Newcastle University and Hon Consultant Paediatric Neurologist, Newcastle upon Tyne Hospitals NHS Foundation Trust, UK
Mitochondrial diseases can be some of the most devastating, debilitating and life-limiting conditions. Although there have been some developments with regard to treatment, progress remains slow. Against this background, many couples with a family or personal history of inherited mitochondrial disease have chosen reproductive options to avoid transmission of disease to their children. Preimplantation Genetic Testing (PGT) was established for nuclear genetic disorders over 35 years ago and has been utilised successfully by many couples for autosomal recessive mitochondrial disease. Women with mtDNA related disease are also able to benefit from PGT though only when there is a realistic prospect of having eggs with low level heteroplasmy. Mitochondrial Donation, or Mitochondrial Replacement Therapy, emerged as a potential strategy for human reproductive choice in the early part of this century. This IVF process involves the use of mitochondria from the egg of a healthy female donor – the third person in what has become known colloquially as the ‘three parent baby’ technique. I shall discuss the scientific, clinical, legislative and regulatory aspects of implementing this reproductive option in the UK as well as our preliminary results from the follow-up of children conceived using this technique.
Saturday 18th April 2026
S10Serum autoantibodies (as biomarkers) of myositis- can we trust them?
Professor Werner Stenzel
Charité University Hospital, Berlin, Germany
Myositis is considered an inflammatory myopathy of skeletal muscles with a presumed autoimmune pathogenesis.
The spectrum of different entities summarized under the general term ‘myositis’ is huge and our understanding of pathogenetic mechanisms initiating, driving and sustaining these increased considerably during the past years. To set the stage, recent advances in classifying different types of myositis, based on morpho-molecular patterns, are presented.
Those ‘patterns’ have been linked to clinical key features of the disease, such as severe lung disease or cancer association as well as skin manifestations, informing about characteristic associations.
Similarly, autoantibodies typically occurring associated with myositis were linked to clinical, morphological and molecular features as well.
These autoantibodies have been detected by different techniques since the 80’s but new ones are continuously detected nowadays and seem to fit well with some of the presumed pathogenic aspects of the respective disease entities.
However, the autoantibodies in myositis are quite diverse and show different patterns, some target nuclear, some cytoplasmic autoantigens, and, conversely, they target diverse subcellular structures even in a given entity such as e.g. dermatomyositis. Hence, it has long been strongly debated whether these autoantibodies should be considered bystanders of the diseases or involved in pathogenesis. Of note, they have of course been discussed to be useful as biomarkers for diagnosis, prognosis and even therapeutic guidance and decisions.
The most recent discoveries in the field of autoantibody-relevance will be addressed during the talk.
S11Friedreich Ataxia: New Treatments, New Mechanisms
David Lynch, MD, PhD.
Children's Hospital of Philadelphia / University of Pennsylvania
Friedreich ataxia (FA) is an autosomal recessive neurodegenerative disorder that affects 1 in 50,000-100,000 individuals of European descent leading to an estimated prevalence of 4-5000 people in US and 15K worldwide. The usual presentation involves neurological features, which are 100% penetrant, but FA is a multi-system disease, also affecting cardiac, endocrine, and skeletal systems. GAA expansions in the FXN gene provide the causative mutations; such expansions silence the FXN gene, leading to relative lack of frataxin protein. Frataxin deficiency leads to mitochondrial dysfunctions including loss if Fe-S containing enzymes (aconitase etc.) that produce clinical manifestations including neurological features. All of the therapeutic approaches thus involve either amelioration of mitochondrial dysfunction or augmentation of frataxin levels. One agent, the NRF2 activator omaveloxolone, is approved for treatment of individuals 16 years or older and is in clinical trials for children. It improves neurological function for several years, but its effect wanes over time. Other agents facilitating mitochondrial function have had minimally positive results and are not approved. The ideal therapy would be augmentation of frataxin levels. This can be accomplished in theory by overcoming the silencing of the FXN gene with epigenetic approaches, replacement of frataxin protein with cell permeable frataxin, or with gene therapy. However, issues arise based on the limited distribution of such agents, off target effects, as well as technical details such as measurement of frataxin restoration and definition of the ideal target tissue. Resolution of such issues is needed to provide successful restoration of sufficient frataxin levels and thus more effective clinical therapy.
S12Advances in mouse models – from generation to characterisation
Dr Sara Wells
The Mary Lyon Centre at MRC Harwell, The Centre for Macaques at Porton Down and the Francis Crick Institute
There has been considerable publicity around advances in technologies that support non-animal preclinical models. Molecular and cellular studies are increasingly well served by alternative methods such as organoids, microfluidic systems, and advanced data science. However, for complex diseases such as neuromuscular disorders, many investigations still require—and are likely to continue requiring for the foreseeable future—the use of mammalian model organisms.
When the use of an animal model such as the mouse is the only viable option, we must ensure that the model is as relevant as possible to the human disease state. This includes not only genetic relevance and the ability to generate the correct genotype and biological complexity, but also the ways in which we characterise the animals, expose them to meaningful environmental cues, administer drugs, and disseminate the resulting data.
In this talk, I will provide an overview of recent advances in genome editing and whole-animal phenotyping that are enabling the development of more precisely disease-positioned preclinical mouse models for neuromuscular disorders.
S13The role of snRNAs in syndromic neurodevelopmental disorders
Dr Nicola Whiffin
Big Data Institute and Centre for Human Genetics, University of Oxford
Small nuclear RNAs (snRNAs) are key components of the spliceosome complex. In 2024, we showed that de novo variants in an 18-nucleotide region of RNU4-2, which is transcribed into the U4 snRNA, cause one of the most prevalent neurodevelopmental disorders (NDD), predicted to affect ∼100,000 individuals globally. Through collaboration with families the disorder was named ReNU syndrome. Subsequent to this discovery, we performed saturation genome editing of RNU4-2 and identified a recessive NDD associated with biallelic variants in other regions of RNU4-2. Further, other groups discovered dominant disorders associated with RNU2-2 and RNU5B-1 and a recessive NDD caused by biallelic variants in RNU2-2. Together, these ‘RNU-opathies’ explain over 1.5% of previous undiagnosed NDDs. In this talk I will describe these discoveries in detail as well as outlining progress in understanding these disorders and into developing therapies.
S14Deciphering Developmental and Neuromuscular Disorders: Insights from the DDD study
Professor Helen Firth
Consultant Clinical Geneticist, Cambridge University Hospitals and Hon Faculty, Wellcome Sanger Institute, Hinxton, UK
The Deciphering Developmental Disorders (DDD) study* was initiated in 2010. From 2011-2015 the DDD study recruited 13,500 young patients from across the UK who had severe developmental disorders with a high suspicion of a monogenic cause in whom full investigation had been unable to identify a diagnosis. The DDD study had two aims: firstly, to transform the diagnostic approach to severe developmental disorders using genome-wide assays and secondly, to research and understand the genomic architecture of severe developmental disorders. To fulfil these dual aims, the study was delivered as a partnership between the 23 NHS Clinical Genetics services in the UK together with the Eire Clinical Genetics service, and the Wellcome Sanger Institute.
In order to deliver at scale, the project used the DECIPHER platform (www.deciphergenomics.org) for recruitment, high-quality phenotyping and delivery of results, thus building a knowledge legacy to support future diagnosis and research. Recruitment was based in the clinical genetics’ services with high-quality phenotyping by the patient’s consultant. Salivary DNA was sent by the family to the Sanger Institute from the child and both parents and, in parallel, blood-derived DNA from the child was sent by the submitting clinical service. Exon-resolution array for the child and trio exome sequencing for the child and both parents was undertaken by the Sanger Institute resulting in 33,500 exomes (including 10,000 trios).
Research outputs from the DDD study include discovery of >67 new disease genes, publication of >350 papers and diagnoses for >5,500 rare disease patients.
This talk will illustrate key principles essential to the success of the DDD study
A dynamic platform to share candidate diagnostic variants and phenotype (www.deciphergenomics.org )
A custom gene panel (DDG2P) delineating allelic requirement, mechanism, phenotype and evidence ( www.ebi.ac.uk/gene2phenotype )
An iterative reporting strategy enabling new diagnoses to be made from old data
A scalable mechanism to identify approved therapies and those under development
Acknowledgement that there is still much to discover
and will invite you to use DECIPHER (hosted by EMBL-EBI and free to use) for your own clinical and research practice in neuromuscular disorders.
*Wright CF et al. Genomic Diagnosis of Rare Pediatric Disease in the United Kingdom and Ireland. N Engl J Med. 2023 Apr 27;388(17):1559-1571. doi: 10.1056/NEJMoa2209046. PMID: 37043637
19th UK Neuromuscular Translational Research Conference
17th and 18th April 2026
Posters, Flash and Platform Presentations
‡indicates a platform or flash presentation
Motor Nerve Disorders
001Cardiovascular Safety and Efficacy of Nusinersen in Spinal Muscular Atrophy: A Propensity Score-Matched Real-World Cohort Study
Samiha Zaman Akhter1, Ibrahim Khalil2, Imran Hossain3, Pallab Sarker4
1University College London, London, United Kingdom
2Dhaka Medical College and Hospital, Dhaka, Bangladesh
3Manikganj Medical College and Hospital, Manikganj, Bangladesh
4Reading Hospital, West Reading, Pennsylvania, USA
samiha.akhter.25@ucl.ac.uk
Background: Spinal muscular atrophy (SMA) is a progressive neuromuscular disorder associated with motor neuron loss, muscle weakness, and increased risks of mortality, hospitalization, and cardiovascular complications due to respiratory and autonomic involvement. While nusinersen improves motor function, its cardiovascular safety and efficacy in real-world SMA patients remains underexplored.
Aims: To assess the cardiovascular safety and efficacy of nusinersen in SMA patients using propensity score-matched real-world data, focusing on all-cause mortality, hospitalization, major adverse cardiovascular events (MACE), and cardiac arrest at 3-year follow-up.
Methods: This real-world retrospective cohort study based on TriNetX US Collaborative Network utilized propensity score matching to evaluate the cardiovascular safety and efficacy of nusinersen in patients with SMA. Two cohorts were identified: Cohort 1 (nusinersen-treated) included patients with confirmed SMA who received nusinersen, while Cohort 2 (non-nusinersen) included SMA patients without nusinersen. Propensity score matching (1:1, caliper 0.1) balanced >50 covariates baseline characteristics, including demographics, comorbidities (e.g., hypertensive diseases, cardiomyopathy), medications (e.g., antiarrhythmics, anticoagulants), and clinical parameters (e.g., blood pressure, heart rate, relevant laboratory values). Matching resulted in balanced cohorts of 929 patients each, with post-matching standardized mean differences generally <0.1, indicating excellent balance.
Results: At 3-year follow-up, nusinersen-treated patients showed superior outcomes compared to matched controls, analyzed via Kaplan-Meier estimation and Cox proportional hazards models. All-cause mortality was significantly lower (21 vs. 72 events; Kaplan-Meier event-free survival 97.40% vs. 89.81%; hazard ratio [HR] 0.23, 95% CI 0.142–0.375, p<0.0001). All-cause hospitalization rates were reduced (187 vs. 232 events; 77.76% vs. 70.45%; HR 0.64, 95% CI 0.527–0.776, p<0.0001). MACE were fewer (13 vs. 31 events; 98.48% vs. 95.36%; HR 0.336, 95% CI 0.176–0.644, p=0.0006). Cardiac arrest showed no significant difference (13 vs. 34 events; HR 0.746, 95% CI 0.35–1.59, p=0.181).
Conclusion: Nusinersen is associated with substantial improvements in survival, reduced hospitalizations, and lower cardiovascular event risk in SMA, with no increased cardiac arrest, supporting its favorable cardiovascular profile in real-world practice.
002Adult SMA REACH: A comparative UK real world data study to assess the safety and efficacy profiles of Nusinersen and Risdiplam in adults with SMA
Aleks Carver1, Elena Karkkainen1, Jess Page1, Laura Simms1 , Jose Verdu-Diaz1, Grecia Alvarez1, Stephanie Tanner1, Dionne Moat1, Robert Muni Lofra1, Chiara Marini Bettolo1.
1The John Walton Muscular Dystrophy Research Centre, Newcastle University and Newcastle Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK.
aleks.carver@newcastle.ac.uk
Background: Advancements in treatment options aimed at modifying disease progression in SMA in recent years have been made, with the introduction of drug treatments Nusinersen and Risdiplam. There has been a reported improvement in motor and pulmonary function tests in SMA patients both post-Nusinersen and post-Risdiplam treatment based on real-world observational data. Most studies have focused on evaluating each drug individually in clinical trials or real-world settings, but comparative analyses are limited in adults. Adult SMA REACH is a longitudinal observational data collection study that collects RWD during routine clinical visits across 18 different sites in the UK. The study includes patients aged ≥16 years with genetically confirmed 5q SMA.
Aims: Utilising data collected from the 460 patients enrolled in this study, we aim to perform an analysis comparing the efficacy and safety profiles of both interventions within the UK adult SMA population, while also identifying correlations with clinical phenotype, non-SMA medications, and patient-perceived benefits as well as identifying differences in the treated cohorts. The primary objective is to assess the efficacy and safety of Nusinersen and Risdiplam in adults, focusing on the following areas:
Changes in functional assessments will be derived from functional scales such as RHS, ATEND, HFMSE, EK2, RULM, WHO and 6MWT.
Changes in respiratory outcomes including FVC and PCF.
Analysis of safety profiles based on the frequency of adverse events recorded during treatments
Methods: Baseline and follow-up visit scores will be summarised using descriptive statistics for safety and efficacy profiles. Repeated measures ANOVA will be used to evaluate changes in functional assessments and respiratory outcome over time. Subgroup analysis will be performed to compare outcomes by SMA type, SMN2 copy number, and functional status. Chi squared tests will be used to compare frequencies of adverse events between subgroups.
Results and Conclusion: The data presented will represent the first direct comparison of these treatments in the adult UK population using real-world evidence, and aims to compare the efficacy and safety profiles of both interventions within the UK adult SMA population providing critical insights that will support improved patient outcomes and inform future treatment decisions.
003UK SMA Patient Registry: The integration of PROMs and real-world data: a holistic approach to characterise disease burden and treatment impact in spinal muscular atrophy
Aleks Carver1, Jess Page1, Lindsay Murphy1, Elena Karkkainen1, Robert Muni-Lofra1, Grecia Benesperi1, Dionne Moat1, Stephanie Tanner1, Laura Simms1, Kate Adcock2, Maria Elena Farrugia3, Gennadiy Ilyashenko4, James B. Lilleker5, John McConville6, Andria Merrison7, Matthew Parton8, Portia Thorman9, Giovanni Baranello10, Annemarie Rohwer10, Liz Ryburn9, Andi Thornton4, Anne-Marie Childs11, Channa Hewamadduma12, 13, Clare M. Galtrey14, Francesco Muntoni10, Mariacristina Scoto10, and Chiara Marini-Bettolo1, on behalf of the Adult SMA REACH clinical network and study team1
1. John Walton Muscular Dystrophy Research Centre, Newcastle University & Newcastle upon Tyne Hospitals NHS Foundation Trust; NIHR Newcastle Biomedical Research Centre (BRC), Newcastle upon Tyne, UK
2. Muscular Dystrophy UK, London, UK
3. Institute of Neurological Sciences, Queen Elizabeth University Hospital, Glasgow, UK
4. TreatSMA, London, UK
5. Manchester Centre for Clinical Neurosciences, Northern Care Alliance NHS Foundation Trust, Manchester Academic Health Science Centre, Salford, UK
6. Ulster Hospital, Belfast, UK
7. South West Neuromuscular Operational Delivery Network, North Bristol NHS Trust, Bristol, UK
8. National Hospital for Neurology and Neurosurgery, University College London Hospitals NHS Foundation Trust, London, UK
9. SMA UK, Stratford-upon-Avon, UK
10. Dubowitz Neuromuscular Centre, Great Ormond Street Hospital, London, UK
11. Leeds Teaching Hospitals NHS Trust, Leeds, UK
12. Sheffield Institute for Translational Neuroscience (SITRAN), School of Medicine and Population Health, University of Sheffield, Sheffield, UK
13. Academic Neuromuscular Unit, Sheffield Teaching Hospitals NHS Foundation Trust, Sheffield, UK
14. Atkinson Morley Neurosciences Centre, Department of Neurology, St George’s Hospital, London, UK
aleks.carver@newcastle.ac.uk
Background: The UK SMA Patient Registry, established in 2008, represents a well-defined cohort of individuals living with Spinal muscular atrophy (SMA) in the United Kingdom and Ireland. The registry has 688 participants: 474 adults (≥16 years); 214 paediatric (<16 years). The registry is a valuable tool for the collection of SMA data through patient-reported outcome measures (PROMs). PROMs capture the SMA patients’ perspectives about their quality of life and the impact of their condition.
Aims: In April 2022, the registry implemented a range of PROMs in order to contribute to Managed Access Agreement (MAA) data collection supporting the national regulatory review of recently emerged SMA treatments Nusinersen and Risdiplam, aiming to collect PROMs data from 100 patients receiving each respective treatment.
Methods: In April 2022, the registry implemented the following PROMs:
EQ-5D-5L
EQ-5D-Y-3L
Patient Global Impression of Severity (PGI-S)
Patient Global Impression of Improvement (PGI-I)
SMA Independence Scale – Upper Limb Module (SMAIS-ULM)
Free-text box
The registry simultaneously launched a pilot study in collaboration with the national Adult SMA REACH and SMA REACH UK clinical networks to support the collection of PROMs data through the registry.
Results: Since their implementation, PROMs questionnaires have been completed by 257 adults and by the caregivers of 106 paediatric patients in the registry. In total, the registry has collected 3764 entries to PROMs questionnaires.
In collaboration with the UK’s national SMA REACH clinical networks, PROMs data collected from patients receiving SMA treatment has been aligned with SMA REACH clinical data and submitted to UK regulatory authorities for consideration as part of the treatment reviews.
Participants’ first submission of PROMs data post treatment initiation will be presented in comparison to their most recent submission of PROMs data post treatment initiation.
Conclusion: The PROMs data presented indicates that participants reported stabilised or improved scores when comparing their first PROMs entry post treatment initiation to their most recent PROMs entry post treatment initiation.
The registry’s presented data proves that PROMs add value to clinician-reported outcomes and offer a different perspective, demonstrating that it is worth continuing the effort of collecting the patient voice.
004Prenatal intervention in an SMA mouse model rescues neurodevelopmental brain defects associated with spinal muscular atrophy
Charalambos Demetriou1,2, Karl Frontzek1, Annika Arora1, Chaobo Wang1, Qiang Zhang2, Sachin Deshpande2, Mariacristina Scoto1, 3, Francesco Muntoni1, Haiyan Zhou2, Naresh K. Hanchate2*, Giovanni Baranello1*
1The Dubowitz Neuromuscular Centre, Developmental Neurosciences Department, UCL Great Ormond Street Institute of Child Health, University College London, London, UK
2Genetics and Genomic Medicine Research and Teaching Department, UCL Great Ormond Street Institute of Child Health, University College London, London, UK
3Great Ormond Street Hospital for Children NHS Trust, London UK
*These authors jointly supervised work
c.demetriou@ucl.ac.uk
Background: Recent studies have shown that SMN-associated neurodevelopmental disorders (SAND) affect a proportion of children with early-onset Spinal Muscular Atrophy (SMA).
Aims: We aim to characterise SAND in the Taiwanese mouse model of SMA using a range of behavioural tests assessing social behaviour and cognitive functioning.
Methods/Materials: We explored rescuing SAND in the Taiwanese mice using either SMN-splicing modifier R07021707, an analogue of risdiplam, which enhances production of SMN protein levels from SMN2 gene, or PMO25, an antisense oligonucleotide that induces SMN2 exon 7 inclusion. Our goal was to test the efficacy of prenatal treatment (administrated at E13.5) versus postnatal (at P0) in rescuing SAND due to the developmental impact of SMN deficiency on brain development and function.
Results: Our findings reveal that the severe SMAI mice treated at P0 with either PMO25 or RO7021707, as well as the untreated mild SMAIII mice exhibited severe abnormalities in behavioral assays compared to healthy controls. These included the resident-intruder test, the marble burying test and the 3-chamber test. Motor function assessed with the grip strength test was normal. Importantly, SMAI mice treated prenatally with risdiplam exhibited behaviours similar to their control mice, suggesting a full neurobehavioral rescue with prenatal treatment.
No significant differences in the anatomy of cerebellum (H&E staining) and the number of neurons (NeuN+ cells) were observed between the different groups at 12 weeks. Protein analysis using WES revealed that treated severe SMAI mice had significantly lower SMN protein levels in cerebellum compared to WT or mild SMAIII mice.
Conclusion: Our findings suggest that SMN deficiency can cause developmental defects in the brain. We show that postnatal restoration of SMN is sufficient to rescue the neuromuscular phenotype but it is not sufficient to fully restore the behavioural phenotypes, which can only be rescued with prenatal treatment. Collectively, our studies indicate that SMN deficiency during development in the severe SMA mouse model is associated with neurodevelopmental defects that require prenatal treatment to be fully rescued. Further studies will investigate the molecular and cellular correlates of these abnormal behavioural phenotypes.
005Hedgehog Signaling as a Novel Regulator of Muscle Health in Neuromuscular Diseases
Kiara E. Fierman1,2, Alessandra M. Norris1,2, Lan Wei-LaPierre1,3, Daniel Kopinke1,2
1Myology Institute, University of Florida, Gainesville, Florida, United States
2Department of Pharmacology & Therapeutics, College of Medicine, University of Florida, Gainesville, Florida, United States
3Department of Applied Physiology and Kinesiology, College of Health and Human Performance, University of Florida, Gainesville, Florida, United States
kfierman@ufl.edu
Background: Muscle weakness and atrophy are debilitating symptoms that significantly impair quality of life and clinical outcomes in patients with nerve injuries, Amyotrophic Lateral Sclerosis (ALS), and other neuromuscular disorders. A shared pathological hallmark across these conditions is the accumulation of intramuscular adipose tissue (IMAT), which we and others have shown replaces healthy muscle, contributes to reduced mobility, and correlates to poorer health outcomes. Despite its prevalence, there are currently no therapies to prevent IMAT formation. We have previously identified the Hedgehog (Hh) pathway as a potent anti-adipogenic and pro-myogenic signal after muscle injury. While we have shown that of the three Hh ligands, Desert Hh (DHH), is the sole ligand responsible for Hh pathway activation after muscle damage, the role of DHH after neuromuscular damage is unknown.
Aims: This study investigates IMAT formation and Hh activity in two contexts, a physical nerve insult model using sciatic nerve cut, and a genetic model of ALS, to understand how Hh signaling contributes to neuromuscular disease pathology.
Methods/Materials: Dhh null mice and wildtype littermate controls underwent unilateral sciatic nerve cut, with the contralateral limb as a control. Additionally, the hSOD1-G96A transgenic mouse model, which recapitulates key pathological features of ALS, were analyzed at 18 and 22 weeks of age. Muscle tissues were collected for histological and molecular analyses to assess IMAT formation, myofiber size, and Hh pathway activity.
Results: Hh signaling was significantly downregulated following sciatic nerve cut at 3-, 5-, 7-, and 21-days post-injury, as well as in hSOD1-G96A mice at 18 and 22 weeks of age. Both ALS mice at 22 weeks and Dhh null mice 21 days post injury exhibited a significant increase in IMAT and reduced myofiber cross sectional area compared to wild-type controls. These changes phenocopied the Dhh null phenotype after acute muscle damage, indicating that loss of Hh signaling exacerbates muscle degeneration and fat infiltration following nerve insult.
Conclusion: Hh signaling acts as an anti-adipogenic cue that is lost in neuromuscular diseases, thereby allowing IMAT to form. Targeting the Hh pathway offers a promising therapeutic target for blocking IMAT and preserving muscle mobility in neuromuscular disorders.
006‡The Honeycomb Synapse: Characterisation of a novel Synapse in the mouse lumbar spinal cord
Carlotta Löer1, Emma Butler1, Ani Ayvazian-Hancock1, Molly Roberts1, Seth Grant2, David Hughes2, Simon Sharples1, Gareth Miles1, and Matthew Broadhead1
1 School of Psychology & Neuroscience, University of St Andrews, UK
2 Centre for Clinical Brain Sciences, University of Edinburgh, UK
3 School of Psychology & Neuroscience, University of Glasgow, UK
carlotta.loeer@stcatz.ox.ac.uk
Background: Synaptic diversity underlies the formation of functionally distinct neural circuits and arises from both molecular composition and synaptic nanostructural organisation. Disruption of specific synapse populations is a hallmark of Amyotrophic Lateral Sclerosis (ALS), a progressive and fatal neurodegenerative disease affecting upper and lower motoneurons (MNs). We have identified a novel synaptic subtype in the mammalian lumbar spinal cord that is selectively vulnerable in a mouse model of ALS. This synapse, termed the Honeycomb Synapse, is defined by its distinctively large, perforated postsynaptic density (PSD). However, the circuit identity, molecular composition, and anatomical distribution of this synapse remain unknown, limiting understanding of its potential contribution to ALS pathogenesis.
Aims: Therefore, this study aims to provide the first anatomical, molecular, and functional characterisation of the novel Honeycomb Synapse.
Methods/Materials: We employed immunohistochemical labelling, 3-D high-resolution microscopy, and supervised machine learning-based image analysis of Honeycomb Synapses in the ventral horn of the mouse lumbar spinal cord.
Results: We report that Honeycomb Synapses are predominantly associated with VGLUT1/Parvalbumin+ presynaptic afferents, located on fast-fatigable α-motoneurons within the lateral motor column of both the upper and lower lumbar spinal cord. We conclude that Honeycomb Synapses are a distinct synapse subtype of the Monosynaptic Stretch Reflex (MSR) pathway and thus may modulate proprioceptive input to motoneurons innervating distal limb muscles. Notably, we identified a subpopulation of Honeycomb Synapses in the medial motor column that are not associated with the MSR pathway and have a smaller PSD.
Conclusion: In summary, we have determined the circuit and anatomical distribution of this novel synapse, revealing never-before-seen diversity in the MSR pathway. Our findings provide a framework for future investigations into Honeycomb Synapses in health and disease.
007Novel substrate identification of the ALS associated kinase TBK1
Gabriel Morel1,2, Helen Flynn1, Simeon R. Mihaylov1, Sila Ultanir1
1The Francis Crick Institute, London, United Kingdom
2University College London, London, United Kingdom
gabriel.morel@crick.ac.uk
Background: Tank Binding Kinase 1 (TBK1) has been genetically associated to the ALS-FTD spectrum with over 90 unique loss of function (LoF) mutations now identified across all functional protein domains. Characterization of TBK1’s cellular roles has been driven by the identification of its downstream substrates, leading to its association to autophagy pathways, through phosphorylation of key autophagy adapters and neuroinflammation, through phosphorylation of IRF3. CNS specific substrates of TBK1 however, are still poorly characterized, resulting in an incomplete understanding of TBK1’s cellular functions in the CNS.
Aims: A more comprehensive list of TBK1’s CNS specific substrates will provide a better understanding of its cellular functions and how ALS-linked TBK1 mutations are contributing to ALS pathology.
Methods/Materials: Using an established chemical genetic approach for kinase substrate identification, a list of novel TBK1 candidate substrates was generated. Purified analogue specific (AS) TBK1, capable of utilizing modified bulky ATP was incubated with mouse brain lysates to thiophosphorylate direct TBK1 substrates. Reactions were then analyzed using LC-MS/MS, to identify direct TBK1 phosphorylation sites in vitro.
Results: After applying various filtering criteria, a final list of 278 phosphosites, mapping to 178 proteins were identified as TBK1 candidate substrates. Our results are supported by the identification of previously reported TBK1 phosphosites pS187-Optn and pS72-Rab7. Further analysis of candidate substrates showed an enrichment for proteins associated to neurodegenerative pathways and in particular ALS. KEGG pathway analysis of candidate substrates revealed an association between TBK1 activity and various cellular pathways linked to ALS pathology including: autophagy pathways, ubiquitin proteasome system, cytoskeletal functions and mitochondrial functions.
Conclusion: This data functionally links TBK1 to various ALS pathomechanisms in an in vitro model, highlighting the importance of a better understanding of TBK1’s cellular functions. Additional functional studies should be carried out to link specific TBK1 mediated phosphorylation to neuronal function and downstream ALS pathology. These findings will provide the basis for safer and more efficacious therapeutic targeting of TBK1 in ALS.
008Exploring the impact of Bridging Treatment in Spinal Muscular Atrophy type 1 (SMA1) with two SMN2 copies- Implications for future Newborn screening
P Munot1, A Murugan2, G McCullagh3, Min Tsui Ong4, V Gowda5, A Y Manzur1, D Ridout6, G Baranello1,8, L , Abbott1, A Childs7, F Muntoni1.8, M. Scoto1 and SMA REACH UK
1Dubowitz Neuromuscular Centre, UCL Great Ormond Street Institute of Child Health & Great Ormond Street Hospital, London, UK.
2 Department of Paediatric Neurology, University Hospital Bristol, Bristol, United Kingdom
3 Department of Neurology, Manchester Children’s Hospital, Manchester
4 Sheffield Children's NHS Foundation Trust, Sheffield, United Kingdom
5 Evelina London Children's Hospital, Guy's and St Thomas' NHS Foundation Trust, London, United Kingdom
6Population, Policy & Practice Department, UCL Great Ormond Street Institute of Child Health, London, United Kingdom
7The Leeds Teaching Hospitals NHS Trust, Leeds, United Kingdom
8National Institute for Health Research Great Ormond Street Hospital Biomedical Research Centre, UCL Great Ormond Street Institute of Child Health, London, United Kingdom
Background: Three disease modifying treatments are approved for SMA1. Of these, Onasemnogene abeparvovec (OA) is often the treatment of choice but requires adequate patient preparation. Clinical outcomes in symptomatic and some presymptomatic SMA1 infants with two SMN2 copies remain suboptimal due to earlier onset and rapid ongoing motor neuron loss until OA is administered. Initiation of Risdiplam, an oral SMN2 splice modifier has increasingly been used for bridging whilst awaiting OA administration. National guidance recommending bridging from diagnosis until OA administration is advocated in United Kingdom, but real-world impact of this strategy is unknown.
Aims and Methods: Retrospective cohort study to compare the clinical outcomes in SMA1(two SMN2 copies) infants bridged with risdiplam with those who received only OA in the pre-bridging era. Due to the impact of disease duration and severity on outcomes, infants were grouped into presymptomatic, age<42 days and >42 days at first intervention.
Results: Three presymptomatic infants, all siblings of known SMA1 patients, were bridged from mean age of 8 days (range 7-9), followed by OA administration at 61 days (range 54-82). Mean baseline CHOP was 49 (range 43-56) and was maintained (n=1) or improved by 10 points (mean 5) before OA.
Twenty-five symptomatic infants were bridged.
22 infants (age>42 days) were bridged from mean age of 112 days (Range 44-279) with a mean baseline CHOP of 23 (Range 14-33) for a mean duration of 48 days (Range 6-123). OA was administered at a mean age of 160 days (Range71-381 days). With bridging, CHOP improved by an average of 8 points (range 0-39) before OA administration.
Remaining 3 (<42D) with severe onset (mean CHOP 9) bridged from mean age of 28 days (range 23-38) for a mean duration of 5.2 months (range 4.7 -5.5 months) showed an average gain of 31 points (range 29-34) prior to OA administration.
Comparison of outcomes in presymptomatic and symptomatic infants <42D with matched unbridged cohort is underway.
Conclusions: After diagnosis, families with SMA1 need time to prepare for OA administration. Early bridging treatment with Risdiplam is crucial and can stabilise function while gene therapy is planned and this strategy could further improve outcomes in SMA1. These results will have implications for UK Newborn screening.
009Profile time course of motor unit regeneration and stability following administration of Smn-upregulating therapeutics
I Partlova1,2 L H Comley1,2, L M Murray1,2
1 Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh, United Kingdom
2 Euan McDonald Centre for Motor Neuron Disease Research, University of Edinburgh, Edinburgh, United Kingdom
I.Partlova@sms.ed.ac.uk
Background: Despite the availability of therapeutic options for SMA, impactful deficits persist in affected patients even after pre-symptomatic treatment. A hallmark of motor unit pathology in SMA involves the loss of motor neurons and disruption of neuromuscular junctions (NMJs).
Aims: To determine what deficits remain following treatment with an Smn up-regulating drugs over time and whether a dual therapy approach, combining Nusinersen with additional SMN therapeutics could ameliorate these deficts.
Methods: Here, we analyzed NMJ recovery following early (PD2) administration of Nusinersen in the SMNΔ7 mouse model. Using the mouse cranial muscles, which allow for whole mount analysis of a group of differentially vulnerable motor units, we assessed recovery at ages P6, P12, and P18. The parameters assessed included axon and endplate number, motor unit size, as well as sprouting and polyinnervation, enabling a comprehensive evaluation of NMJ regeneration and stability following early Nusinersen treatment in the SMNΔ7 model.
A key aspect of our study was the inclusion of a dual therapy approach, combining Nusinersen with additional SMN therapeutics. This enabled us to assess the potential synergistic effects on motor unit recovery.
Results: Treatment with Nusinersen significantly improved innervation of endplates at the NMJ even in the most vulnerable muscles. The vast majority of endplates were fully occupied at all time points analysed following treatment. However, number of intramuscular axons innervating the most vulnerable muscles were reduced from the P6 time point onwards. The loss of axons coupled with stabilisation in endplate innervation gave rise to a profound increase in motor unit size, which could have implications for the health of the remaining axons over time. This work suggests that Nusinersen treatment rescues some but not all parameters of neuromuscular pathology in this mouse model of SMA, and reveals important deficits in motor unit recovery following an increase in SMN levels.
Conclusion: This study emphasizes the regenerative potential of motor neurons following Smn restoration but stresses that recovery with a Smn upregulating treatments is incomplete. This is particularly highlighted in the progressive decline of axon number with one treatment alone, which increases the motor unit size over time.
010‡Prenatal treatment for severe spinal muscular atrophy (SMA): the UK experience
Mariacristina Scoto 1, 5, Salma Jabak2, Kypros Nicolaides 2, Lize-Marie Wium 2, Giovanni Baranello 1, 5, Ineke Van Herwijnen 3 and Marjorie Anne Illingworth 4
1 Great Ormond Street Hospital for Children Trust., London
2 Fetal medicine King's College Hospital NHS Trust, London
3 Fetal medicine Unit at the Princess Anne Hospital, University Hospital Southampton
4 Department of paediatric neurology, University Hospital Southampton.
5 UCL Great Ormond Street Institute of Child Health, London
mariacristina.scoto@gosh.nhs.uk
Background: Spinal muscular atrophy (SMA) is a motor neuron disease caused by the deficiency of survival of motor neuron 1 (SMN1) protein arising from the loss or mutations in SMN1 gene. With the approval of lifesaving SMN-enhancing drugs, SMA children achieve significant motor milestones and survival. However, mounting data indicate that SMA infants with two copies of SMN2, have already lost substantial motor neurons before term birth with >50% showing SMA signs/symptoms at the time of initial assessment. Risdiplam is one of the approved SMN-enhancing drugs, licenced for treatment after birth; it is a small molecule, orally available. A published case report of transplacental treatment of a SMA fetus and two copies of SMN2, by oral administration of risdiplam to the mother in the third trimester of pregnancy, has prompted collaborations between neurologists and maternal-fetal medicine communities leading to few additional cases treated worldwide. We present the experience of prenatal treatment of two SMA babies in two centers in the UK.
Methods: Both babies were diagnosed during the 1st trimester of pregnancy via NIPD, as had sibling(s) with severe SMA. After obtaining local ethical approval, both mothers received oral Risdiplam, via compassionate use, from 32 weeks and 5 days of gestation at the dosage of 5mg/day.
Regular follow-ups included maternal weekly medical review and fetal Ultrasound.
Results: Growth scans were reassuring with no maternal side-effects reported. Labour was induced at 38 weeks in both cases. At birth, clinical examination and electrophysiological assessment were normal in both cases. Both infants had two SMN2 copies, confirmed after birth, and received bridging therapy with risdiplam from 3 and 2 days of life respectively, prior to gene-replacement treatment, which occurred at the age of 3 months in one baby and has been planned in the second one.
At the last assessment, at the age of five and three months respectively, the two infants showed no obvious signs or symptoms of SMA.
Conclusion: Our initial experience support consideration of prenatal risdiplam therapy to maximize therapeutic benefit. However, larger studies and long-term follow up are still required to establish safety and efficacy during early development
011Quantitative neck strength assessment in 5q SMA using handheld dynamometer – reliability, validity and clinical relevance
Jon Street 1,2, Alexandra Botha 2, Katie Nevin 1, Lindsay Maidment 1, Channa Hewamadduma 1,2
1 Academic Unit of Neuromuscular Disorders, Sheffield Teaching Hospitals NHS Foundation Trust
2 Sheffield Institute of Translational Neurosciences, University of Sheffield
Introduction: 5q Spinal muscular atrophy (SMA) leads to progressive, muscle weakness affecting the trunk, limbs, respiratory, and bulbar muscles. Currently used outcome measures may be significantly impacted by severe weakness, joint contracture, pain and injury. Neck movements are relatively unaffected by these processes and rarely routinely assessed. We studied the reliability, validity and clinical relevance of quantitative neck strength assessment using MicroFET2 handheld dynamometer (HHD), performed by a single evaluator during clinic visits.
Methods: HHD assessment of neck flexion and extension was performed, alongside standard of care assessments of physical function, and lung function.
Results: Fifty eight SMA patients (Non-sitter = 5, Sitter = 41, Walker = 11) underwent HHD assessment at baseline. All patients tolerated assessments well, and assessments took < 3 minutes to complete. Test-retest reliability was excellent for both neck flexion and neck extension (ICC >0.95). Significant differences were observed in neck flexor strength between different functional states (Kruskal Wallis test, p = 0.002). Neck flexor strength correlated strongly with FVC (r = 0.75), RULM (r = 0.77), and EK2 (r = -0.78). Neck flexor strength was highly predictive of an FVC <1.5l, placing patients at high risk of respiratory insufficiency (AUC 0.93) – as well as ambulatory status (AUC = 0.95).
Conclusion: Neck strength assessment with HHD was feasible and well tolerated across SMA functional states. Results correlated strongly with measures of motor function, and FVC.
Neuromuscular Junction Disorders and Channelopathies
012‡Clinical and genetic spectrum and diagnostic options for congenital myasthenic syndrome across an international cohort
Setareh Alabaf1 Seena Vengalil2, Ajmal Roshan2, Sanjana Angadi2, Milena B. Levignali3, Pedro J. Tomaselli3, Rodrigo S. S. Frezatti3, Nimita Rani4, Kireshnee Naidu5,6, Özlem Yayıcı Köken7, Ipek Polat8,9, Maryke Schoonen10, Michelle Bisschoff11, Musambo M. Kapapa12, Sharika Raga13,14, Catarina Olimpio1, Katherine Schon1, Banitz Rajmund1, Lindsay A Wilson15, William L Macken15, Natalia Dominik15, Stephanie Efthymiou15, Kumarasamy Thangaraj16, Henry Houlden15, Robert D S Pitceathly15, Michael G Hanna15, Jo M. Wilmshurst13,14, David R. Bearden17,18, Michelle P. Kvalsund 18,19, Francois van der Westhuizen10, Izelle Smuts20, Franclo Henning6, Ahmet Cevdet Ceylan21,22, Semra Hiz8,23 Yavuz Oktay9,23, Uluç Yiş8, Haluk Topaloğlu24, Jeannine M. Heckmann5,13, Sireesha Yareeda25, Venugopalan Y. Vishnu4, Wilson Marques Jr3, Claudia Ferreira da Rosa Sobreira3, Atchayaram Nalini2, Rita Horvath1
1. Department of Clinical Neurosciences, University of Cambridge, Cambridge CB2 0QQ, UK
2. Department of Neurology, National Institute of Mental Health and Neurosciences (NIMHANS), Bengaluru, India
3. Department of Neurosciences, Division of Neurology, Ribeirão Preto Medical School, University of São Paulo, Brazil
4. Department of Neurology, All India Institute of Medical Sciences (AIIMS), Delhi, India
5. Neurology Research Group, Division of Neurology, Department of Medicine, University of Cape Town, Cape Town, South Africa
6. Division of Neurology, Department of Medicine, Stellenbosch University, Cape Town, South Africa
7. Faculty of Medicine, Department of Pediatric Neurology, Akdeniz University, Antalya, Turkiye
8. Faculty of Medicine, Pediatric Neurology Department, Dokuz Eylül University, Izmir, Turkiye
9. Izmir International Biomedicine and Genome Institute, Dokuz Eylül University, Izmir, Turkiye
10. Centre for Human Metabolomics, Desmond Tutu School of Medicine, Faculty of Health Sciences, North-West University, Potchefstroom, South Africa
11. Focus Area for Human Metabolomics, North-West University, Potchefstroom, South Africa
12. Department of Physiotherapy, University of Zambia School of Health Sciences & University Teaching Hospital Neurology Research Office, Lusaka, Zambia
13. Neuroscience Institute, University of Cape Town, Cape Town, South Africa
14. Division of Paediatric Neurology, Department of Paediatrics and Child Health, Red Cross War Memorial Children’s Hospital, Cape Town, South Africa
15. Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology and The National Hospital for Neurology and Neurosurgery, London WC1N 3BG, UK
16. CSIR—Centre for Cellular and Molecular Biology (CCMB), Hyderabad, Telangana, India
17. University of Zambia Department of Educational Psychology, Lusaka, Zambia
18. Department of Neurology, School of Medicine and Dentistry, University of Rochester Medical Center, Rochester, NY 14642, USA
19. Department of Internal Medicine, University of Zambia School of Medicine, Lusaka, Zambia
20. Department of Paediatrics, Steve Biko Academic Hospital, University of Pretoria, Pretoria, South Africa
21. Department of Medical Genetics, Ankara Bilkent City Hospital, Ankara, Turkiye
22. Faculty of Medicine, Department of Medical Genetics, Ankara Yıldırım Beyazıt University, Ankara, Turkiye
23. Izmir Biomedicine and Genome Center (IBG), Izmir, Turkiye
24. Yeditepe University Hospitals, Istanbul, Turkiye
25. Department of Neurology, Nizam’s Institute of Medical Sciences (NIMS), Hyderabad, Telangana, India
Background: Congenital myasthenic syndrome (CMS) is a neuromuscular disorder caused by genetic defects in neuromuscular transmission. Symptoms include fatigability and characteristic muscle weakness. Genetically diagnosing CMS can inform treatment.
Aims: To grow knowledge of the global genetic causes of CMS to inform applicability of existing and emerging therapies, and to survey international diagnostic pathways.
Methods/Materials: We used the ICGNMD pipeline (Wilson, Perry, Macken et al, Brain 2023) to generate and analyse whole exome sequencing with reference to participant phenotype and history. Participants were defined as genetically solved if a pathogenic/likely pathogenic variant in a relevant gene and zygosity was present.
Preliminary results: Data from 96 participants with clinically suspected CMS were reviewed (50% male, 50% female). The majority (56; 58%) were recruited in India, followed by Brazil (32; 33%) South Africa (5; 5%) and Turkiye (3; 3%). No CMS participants could be recruited from other South African sites (714 families with suspected inherited NMD), nor from the Zambian site (89 families). Mean age at symptom onset was 7.3± 8.2 years and age at study recruitment was 25.1±14.4 years. 64% (N=61) were genetically confirmed as CMS, 3% (N=3) as non-CMS while (33%, N=32) remained genetically unsolved after review. Primary AChR deficiency was the most prevalent subtype (29%, N=28) followed by AChR clustering defect (15%, N=14), glycosylation defects (12%, N=11), endplate acetylcholinesterase deficiency (5%, N=5) and slow channel CMS (3%, N=3).
Most ICGNMD partners are in middle-income countries and reported access to relevant tests including repetitive nerve stimulation (RNS) and Anti-AChR antibody tests, but lack of affordable genetic testing was a common barrier to diagnosis.
Preliminary conclusion: CMS constituted a small percentage of inherited neuromuscular disease participants recruited to ICGNMD. However, multiple factors impacted, and many partners felt CMS under-referred to their clinics. Among the clinically suspected CMS cases, (61; 64%) had a confirmed genetic CMS confirmed. Our cohort is too small to reach general conclusions, however data from Southern India aligned with an earlier publication, while more GFPT1 variants were noted in Northern India (New Delhi). We did not find any RAPSN-CMS nor any fast channel CMS within this specific study population.
013Repurposing a Clinical Technique to Evaluate Muscle Ion Channel Drug Responses across the translational spectrum
Jethro Choi1,2, Jo Ng3, Roope Mannikko1, Stuart Baker4, Karen Suetterlin5,6
1 Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, London, UK
2 Translational and Clinical Research Institute, Newcastle University, Newcastle upon Tyne, UK
3 Genetic Therapy Accelerator Centre, UCL Queen Square Institute of Neurology, London, UK
4 Institute of Neuroscience, Newcastle University, Newcastle upon Tyne, UK
5 AGE Research Group, Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University; Department of Clinical Neurophysiology, Newcastle upon Tyne Hospitals NHS Foundation Trust and NIHR Newcastle Biomedical Research Centre, Newcastle upon Tyne, UK
6 John Walton Centre for Muscular Dystrophy, Newcastle upon Tyne Hospitals NHS Foundation Trust and Newcastle University, Newcastle upon Tyne, UK
jethro.choi.22@ucl.ac.uk
Background: Muscle velocity recovery cycle (MVRC) analysis characterises muscle membrane excitability and is clinically used to investigate muscle channelopathies, complementing electromyography and genetic testing. Previous bedside-to-bench studies applied MVRC in vivo in mice treated with ion channel blockers. Here, we reverse-translated MVRC to isolated muscle, enabling controlled pharmacological assessment prior to in vivo or clinical testing. MVRC therefore represents a unified biomarker spanning isolated muscle to clinical application.
Aims:
To elucidate physiological mechanisms underlying individual MVRC components, and their diagnostic relevance.
To assess MVRC as a translational tool for characterising pharmacological effects of ion channel–targeting drugs and as a biomarker for validating genetic therapies.
Methods: Whole soleus and extensor digitorum longus (EDL) muscles were mounted under two EMG needle electrodes with silver wire reference anode and silver pellet ground in a 35 °C chamber. One, two, or five conditioning prepulses were followed by a test pulse at progressively shorter interstimulus intervals. Latency changes in the test response reflected ion channel contributions to muscle activation and recovery. A reduction in latency (increased conduction velocity) is supernormality, while an increase in latency is subormality. The point at which conditioned and unconditioned test responses have equal latency is the Muscle Relative Refractory Period (MRRP).
Pharmacological experiments examined sodium channel blocker mexiletine, potassium channel blocker barium, late sodium current inhibitor ranolazine, and fast-twitch–selective myosin inhibitor N-benzyl-p-toluenesulfonamide (BTS).
Results: Mexiletine has activity-dependent increase in subnormality at 10 and 50 µM, with reduced supernormality at 10 µM and complete loss at 50 µM. Barium had minimal effect with one pulse, but five pulses (30 µM) eliminated supernormality in EDL and reduced it in soleus. Ranolazine produced similar stimulation-dependent effects in EDL, with five pulses (50 µM) abolishing supernormality and inducing residual subnormality. BTS abolished supernormality in EDL and reduction in soleus, suggesting a correlation between supernormality and fibre contraction.
Conclusion: This adapted in vitro MVRC approach allows mechanistic understanding of MVRC signal and showed potential as a translational platform for screening pharmacological agents targeting muscle channelopathies. MVRC changes reflected known drug mechanisms, supporting its application as a functional biomarker in therapeutic development.
014Congenital myasthenic syndrome: A multi-centric cohort from India
Amlan Kusum Datta1, Jasodhara Chaudhuri2
1 Narayana Health, Jessore Road, Kolkata, India
2 Nil Ratan Sircar medical college and hospital, Kolkata, India
amlankd@gmail.com
Background: Congenital myasthenic syndromes (CMS) are a diverse group of phenotypically miscellaneous, inherited neuromuscular disorders, resulting from pathogenic variants of various genes encoding proteins, perturbations of which disrupt the structural integrity of the neuromuscular junction (NMJ).
Aims: The authors aim to assemble and analyze clinical records of CMS patients, and review existing literature, particularly with respect to the Indian subcontinent.
Methods/Materials: A registry based retrospective study was conducted in the neuromuscular clinics of two tertiary care centers in Kolkata, India, between February 2018 and January 2023. All cases of suspected CMS, with decremental response on repetitive nerve stimulation (RNST) study, and genetically confirmed cases (at least one or more pathogenic variant or familial cases) were included in the study. Genetic testing information was reviewed. Sequencing of protein coding regions (∼30 Mb of human genome) and complete mitochondrial genome sequencing was performed using Illumina next generation sequencing (NGS). American college of medical genetics and genomics (ACMG) criteria was used for classification.
Results: Nine (n=9) patients, all of Bengali ethnicity were included for analysis. Majority of patients were female (6; 66.6%). Mean age of onset of symptoms was 7.22 years (S.D.=7.01; Range 7 to 288 months). Mean age at diagnosis was 10.38 years (S.D.= 10. 24), with a mean delay of 3.16 years. Ptosis was the most common presentation (77.8%), followed by limb girdle pattern of weakness (55.5%%). A total of 11 mutations were documented. Most common genetic etiology was CHRNE [45.45%; (5/11)]. Most (8/11; 72.7%) of the mutations identified were novel mutations. Only one mutation (9.9%) was pathogenic. Mean dose for salbutamol was 9.71mg (6-16mg). Majority of patients (6;85.71%) improved after therapy with a mean improvement in QGMS of 34.28% (SD 19.88). Mean lag between onset of therapy and improvement was 38.16 months (SD 36.16)
Conclusion: Factors such as lack of physician awareness and unavailability of molecular diagnostic techniques in large parts of the country have contributed to a dearth of clinical and genetic profiling of CMS in India. Phenotyping and genetic analysis is crucial for the characterization, and prognostication of CMS, and thereby for formulating a treatment plan
015Positive allosteric modulator selective for adult muscle nicotinic acetylcholine receptor
YY Dong1, RG Webster1, S Alabaf1, A Li1, R Beerli2, S Siehler2, P Imbert2, L Lee1, J Cossins1, M Koziczak-Holbro2, L Flotte2, L Gaudet2, N Gerwin2, D Beeson1,
1 Nuffield Department of Clinical Neurosciences, University of Oxford, UK
2 Novartis Biomedical Research, Basel, Switzerland
yin.dong@ndcn.ox.ac.uk
Background: The muscle nicotinic acetylcholine receptor (AChR) is the key mediator of neuromuscular signal transmission, and its dysfunction is the most common cause of myasthenic syndromes. Moreover, impaired neuromuscular signaling is involved in the pathology of other neuromuscular diseases such as amyotrophic lateral sclerosis, spinal muscular atrophy, and sarcopenia. Consequently, therapies that improve AChR function have potential for treating a range of neuromuscular diseases.
Aims: The aim of this study was to find a positive allosteric modulator (PAM) that is selective for the muscle type AChR.
Methods/Materials: Ca2+ FLIPR assays used to identify PAM and counter screen against neuronal AChR. Single channel electrophysiology for PAM functional characterization. Ex vivo mouse diaphragm neurophysiology to determine effects on NMJ. Nerve-muscle contractility to determine effect on muscle traction.
Results: We identified DC-98-LC74 as a PAM for the adult skeletal muscle-type AChR using Ca2+ FLIPR assays, and demonstrate that it is selective for the adult skeletal muscle AChR over neuronal subtypes. Neurophysiological recordings from ex vivo mouse diaphragm preparations revealed that DC-98-LC74 elongates the endplate currents of wildtype (WT) adult but not fetal channel containing diaphragms. Single channel studies on chimeric channels of the adult and fetal receptor, and in saturating concentrations of choline, suggest that the PAM does not bind at either orthosteric site, but works by increasing the unliganded open probability via a mechanism that involves the CHRNE M2-M3 loop. We also show that DC-98-LC74 increases the burst duration of multiple fast channel mutant AChR to WT levels, suggesting that positive allosteric modulation could be a therapeutic strategy for this difficult to treat sub-type of congenital myasthenia. Improved nerve-induced muscle force was also observed in isolated sarcopenic mouse muscles in preliminary nerve-muscle contractility studies.
Conclusions: Positive allosteric modulation of the muscle type nAChR has potential benefits not only in myasthenia, but also other neuromuscular disorders involving the neuromuscular junction.
016Novel NaV1.4 variant with severe defect in channel inactivation in a patient with severe myotonic myopathy
Roope Männikkö1, Rotem Orbach2, Sandra Donkervoort2, Mark Levin3, Michael G. Hanna1, Carsten G. Bönnemann2, A. Reghan Foley2
1 UCL Queen Square Institute of Neurology, Department of Neuromuscular disease, UCL, London
2 Neuromuscular and Neurogenetic Disorders of Childhood Section, NINDS, National Institutes of Health, Bethesda, Maryland, United States
3 Translational Vascular Medicine Branch, NHLBI, National Institutes of Health, Bethesda, Maryland, United States
r.mannikko@ucl.ac.ukreghan.foley@nih.gov
Background: Channelopathies of skeletal muscle voltage-gated sodium channel NaV1.4 include dominantly inherited myotonias and periodic paralyses. Both are caused by increased excitatory currents through the channel that in case of PP drive the muscle into a stable depolarised state, leading to sodium channel inactivation and muscle paralysis. More recently, very rare recessive Nav1.4 conditions caused by biallelic loss of function variants have been reported in patients with congenital myopathy with fixed muscle weakness. Heterozygous carriers of LoF variants are asymptomatic.
Aims: We report phenotypic data notable for both myotonic and myopathies features and assess functional consequences of an ion channel variant identified.
Methods/Materials: Routine clinical and genetic analysis was performed. Ion channel properties were analysed using patch clamp methodology.
Results: The proband presented congenitally with severe muscle weakness and respiratory insufficiency. Frequent laryngospasms started at age 2 months and strikingly improved with carbamazepine (stopped due to Stevens-Johnson syndrome) and mexiletine. Muscle histology was myopathic, and EMG demonstrated myopathic features and spontaneous activity suggestive of myotonia. CK has ranged 1,049 – 6,446 U/L. The severe muscle weakness is suggestive of a LoF condition while the responsiveness to sodium channel blockers and CK elevation are suggestive of a GoF condition. A clinical NGS panel revealed de novo SCN4A: c.1328T>C; (p.L443P); this variant introduces a kink-forming amino acid into a pore-lining α-helix. Research-based whole-genome sequencing has not revealed a second SCN4A variant or causative variants in other genes. Functional analysis of L443P channel revealed drastically attenuated inactivation: voltage dependence and extent of fast inactivation were attenuated, and voltage dependence of slow inactivation was massively shifted towards more depolarised voltages. These GoF features are large in amplitude.
Conclusion: Our patient’s surprising degree of myopathy due to a dominantly acting allele is likely an ‘impaired slow inactivation’ phenotype with enhanced sodium currents due to severely defective inactivation of L443P channel driving the muscle into a persistently depolarised state, reducing channel availability to a degree that results in persistent weakness or extreme fatiguability. NaV blocker mexiletine may prevent excess depolarising currents thereby recovering sodium channel availability whilst also preventing further myotonic overactivity.
Dystrophy (Pre-clinical)
017‡The role of extracellular matrix in muscular dystrophies
James Clark1, Elisa Villalobos1, Esther Fernández-Simón1, Andrew Galloway1, Dan Cox1, Alexandra Monceau1, Lidia Matias Valiente1, José Verdu-Díaz1, Priyanka Mehra1, Rasya Gokul Nath1, Panos Katsikis1, Andrew Porter2, Pawel Palmowski2, Jordi Diaz Manera1.
1 The John Walton Muscular Dystrophy Research Centre, Translational and Clinical Research Institute, Newcastle University and Newcastle Hospitals NHS Foundation Trust, UK
2 Proteomic Core Facility, Newcastle University, UK
James.clark4@newcastle.ac.uk
Background: Sarcoglycanopathies are genetic muscular dystrophies caused by mutations affecting the sarcoglycan complex, a component of the dystrophin–glycoprotein complex that maintains sarcolemmal stability and protects muscle fibres from damage. Characterised by progressive muscle weakness, predominantly affecting the limb-girdle muscles. Sarcoglycanopathies significantly impair quality of life and reduce life expectancy. In healthy muscle, extracellular matrix (ECM) plays an essential role in structural support, protection and repair. In disease, excessive accumulation of ECM proteins impair normal repair mechanisms. The deposition of fibrotic tissue leads to increased muscle stiffness, contractures, and functional decline. Understanding how ECM composition changes and how muscle cells respond to these alterations is crucial for identifying mechanisms driving fibrosis in muscular dystrophies.
Aims: To characterise changes in ECM protein composition in sarcoglycanopathies and to develop a cell culture model to investigate the effects of these alterations on muscle cells.
Methods/Materials: Muscle samples from 35 patients with sarcoglycanopathy and 10 controls were analysed using mass spectrometry. Both whole muscle and decellularised samples were compared. Significantly differentially expressed ECM proteins were identified for further investigation. Decellularised ECM was repopulated both as a 3D hydrogel and by direct seeding on tissue sections.
Results: A total of 504 proteins were significantly differentially expressed between patients and controls. 443 proteins were upregulated in disease samples, 61 were upregulated in controls. Key components of the dystrophin–glycoprotein complex were significantly downregulated in sarcoglycanopathy, whereas numerous ECM proteins were markedly overrepresented. Comparisons between severely and mildly affected patients revealed significant differences in ECM composition, indicating that ECM remodelling is a dynamic process that precedes the end-stage fibro-fatty replacement in advanced disease. Cell culture experiments showed fibro-adipogenic progenitor cells differentiated to fibrotic phenotypes more readily when grown on dystrophic ECM.
Conclusion: Sarcoglycanopathies are associated with increased expression of ECM proteins and significant alterations in ECM composition. These changes between patients and controls but also with disease severity. Changes appear to have a real manifestation on the behaviour of muscle cells. As fibrosis is a major driver of disease progression, improved understanding of the transition of ECM from reparative to pathological state may reveal novel therapeutic targets.
018‡Evaluating drug potential of macrolide azithromycin in Myotonic Dystrophy 1 (DM1)
Tushar K. Ghosh1, Junting wang2, Anjani Kumari1, Pavel Gershkovich2, Rebecca Trueman1 and J. David Brook1
1School of Life Sciences, Queen’s Medical Centre, University of Nottingham, Nottingham NG7 2UH, UK
2School of Pharmacy, Biodiscovery Institute, University of Nottingham, Nottingham, NG7 2RD, UK
Tushar.Ghosh1@nottingham.ac.uk
Background: Myotonic dystrophy type 1 (DM1) is a neuromuscular disease with clinical symptoms of that include myotonia, muscle wasting, cardiac conduction defects, cataracts and insulin resistance. It is the most common adult form of muscular dystrophy affecting 1 in 8000 individuals and as of now there is no cure. DM1 is caused by an expanded CTG repeat at the 3’ untranslated region of the dystrophia myotonica protein kinase gene (DMPK). The expanded CUG repeat RNAs form secondary structures that sequester regulatory RNA-binding proteins including MBNL1, forming toxic nuclear foci and affecting normal cellular functions.
Aim: We have investigated a macrolide antibiotic azithromycin for its potential as a drug treatment for DM1 and compared its effect to that of another macrolide erythromycin.
Methods: We employed a cell-based foci assay to analyse foci elimination following drug treatment. We looked for the tissue distribution of the macrolides in HSALR mice by LC-MS. Using RT-PCR we will be analysing splicing correction in mouse muscle tissue and in DM1 cells. For molecular mechanism of the macrolides, we will carry out RNA sequencing.
Results: Azithromycin (AZM) was found to be more potent than erythromycin (ERM) in our foci assay. Wash out experiments suggested that the AZM was more stable than ERM inside DM1 cells. Tissue distribution studies in the HSALR mouse found that AZM accumulated more in muscle and heart and least in the brain. Currently we are examining the efficacy of AZM via splicing correction of key genes in DM1 cells and HSALR mouse muscle tissues. Also, we are doing direct RNA sequencing from the treated HSALR mice and DM1 cells to understand the molecular mechanism of the drug.
Conclusion: The macrolide azithromycin is more potent than erythromycin in foci assay and more accessible to muscle and heart tissues in mouse.
019Utrophin upregulation as a therapeutic route to control myopathy in Duchenne muscular dystrophy
Dr Hannah Gleneadie1, Dr Beatriz Fernandez-Ruiz1, Dr Thomas Francis2, Professor Stephen Harridge2, Professor Amanda Fisher1,3
1. MRC Laboratory of Medical Sciences (MRC LMS), London
2. Centre for Human & Applied Physiological Sciences, Kings College London
3. Biochemistry Department, University of Oxford
hannah.eadie@lms.mrc.ac.uk
Background: Duchenne muscular dystrophy (DMD) is a progressive and ultimately fatal disorder, caused by loss of function mutations in the dystrophin (DMD, Dmd) gene. In animal and cellular models of DMD, loss of utrophin (UTRN, Utrn), a homologue of dystrophin, exasperates the dystrophic phenotype, while overexpression can compensate for dystrophin loss. Therefore, utrophin upregulation could be a therapeutic option for DMD. The search for small molecules to upregulate utrophin is an ongoing challenge for the DMD community, and while some therapies have progressed to clinical trials, none have made it to the clinic.
Aims: A limitation in this search is the complex regulation of utrophin expression; with control at both the transcriptional and post-transcriptional level. Previous screening platforms have not represented utrophin in its full genomic context. Therefore, we aimed to generate mouse lines to report on endogenous dystrophin and utrophin expression in vivo and, using myoblasts derived from these mice, create a screening platform to search for utrophin-upregulating compounds.
Methods/Materials: A green click beetle luciferase (CBG99Luc) and a red-shifted firefly luciferase (RFluc) gene were inserted into the 3’UTR of endogenous Dmd and Utrn respectively, separated from the target genes by T2A sites. In this design, bioluminescent imaging reports on transcription and translation without impacting protein function.
Results: Spectrally distinct luciferase enzymes allow simultaneous visualisation of both dystrophin and utrophin expression throughout development in live mice. Immortalised myoblasts generated from Utrn-RFluc mice were used to perform a pilot screen for drugs that increase Utrn expression. We demonstrate that inhibition of ERK1/2 and the epigenetic regulator EZH2, increase Utrn, both alone and in combination, and confirm these drugs increase UTRN expression in human myoblasts from healthy young adults and DMD patients, validating the efficacy of the screening protocol.
Conclusion: Here we describe bioluminescent reporters for utrophin and dystrophin expression, to screen for utrophin upregulation in vitro and noninvasively visualise endogenous expression of both genes in vivo.
020Of muscle, mice and dogs: comparative RNAseq analysis of dystrophin expression and dysregulation in animal models of Duchenne Muscular Dystrophy
John C.W. Hildyard1, Liberty E. Roskrow1, Richard J. Piercy1
Comparative Neuromuscular Diseases Laboratory, Royal Veterinary College, Royal College Street, Camden, London
jhildyard@rvc.ac.uk
Background: The dystrophin gene is essential for muscle fibre integrity, and mutations resulting in loss of muscle dystrophin protein result in the fatal muscle wasting disease Duchenne muscular dystrophy (DMD). The gene is unusual, with an enormous genomic locus (∼2.3Mb): production of a single transcript takes ∼16 hours, ostensibly precluding responsive transcriptional behaviour. Dystrophin mRNA exhibits a phenomenon termed “transcript imbalance” (found in humans, mice and dogs), whereby 5’ sequence is detected at levels substantially greater than 3’. We proposed that this behaviour is not pathological, but instead reflects normal expression for this long gene: continuous initiation, long transcription time, and short mature transcript lifespan. This “pay in advance” model supplies mRNA at high levels continuously regardless of demand, with mature levels being controlled via post-transcriptional degradation (circumventing the unavoidable delay). 5’ sequence is more abundant than 3,’ simply because at steady state, most dystrophin transcripts have not yet been completed. This model has, to date, been indirectly inferred rather than empirically confirmed.
Aims and Methods: We generated both polyA purified (mature mRNA only) and ribodepleted (nascent and mature mRNA) RNAseq datasets using skeletal muscle RNA from healthy and dystrophic mice (mdx) and dogs (DE50-MD). Comparison of these datasets allowed detailed assessment of dystrophin expression specifically, and dystrophy-associated transcriptomic changes more broadly.
Results: Our data confirm our transcriptional model: ribodepleted data reveal high levels of 5’ sequence, declining progressively toward the 3’ terminus, while polyA does not. Dystrophin mRNA is indeed chiefly immature, in healthy and dystrophic muscle, in both mice and dogs. Further, we show that while mature transcripts are reduced in both mdx and DE50-MD models as expected, transcriptional initiation is also impaired in mouse (but not dog). More broadly, we evaluate sensitivity of dystrophic gene expression analysis to sequencing pipeline, and identify differences in disease-associated transcriptional programmes between the mildly affected mdx mice and the more severely affected DE50-MD dogs.
Conclusion: Our findings build on our previous work, revealing how dystrophin transcript imbalance likely occurs, and supporting a predominantly post-transcriptional model of control. Determining how mature dystrophin transcript stability is regulated might provide avenues for therapeutic interventions.
021Investigating DNA damage response, oxidative stress and autophagy in astrocytes from Duchenne Muscular Dystrophy patients
James A. K. Lee1, Yasmin B Orandi1, Aahnaf Newaz2, Reem Al Kharji1, Francesco Muntoni3, Jenny Lange4, Patrizia Ferretti1
1Department of Developmental Biology and Cancer, Stem Cells and Regenerative Medicine Section, UCL Great Ormond Street Institute of Child Health, University College London, London, UK; 2Institute of Opthalmology, University College London, London, UK; 3Dubowitz Neuromuscular Centre, UCL Great Ormond Street Institute of Child Health, University College London, London, UK; 4UCL UK Dementia Research Institute, University College London, London, UK
James.a.lee@ucl.ac.uk
Background: Duchenne Muscular Dystrophy (DMD) is an X-linked progressive muscle wasting disease caused by mutations in the dystrophin gene (DMD). DMD patients can also present neuropsychiatric disorders, learning disabilities, autism and epilepsy, consistent with dystrophin isoform expression in the central nervous system, including in astrocytes. It has been suggested that cell autonomous dysfunction in human astrocytes may play a significant role in the development and maintenance of DMD neural pathology, but studies on the underlying mechanisms are still limited.
Aims: We aimed to test the hypothesis that DNA damage response, oxidative stress and autophagy are altered in DMD patient-derived astrocytes, as suggested by changes in gene expression detected by our previous RNA-seq experiments.
Methods/Materials: Astrocytes were generated from DMD patient-derived iPSC’s and healthy controls. Isogenic iPSC lines recreating the same DMD mutations have also been generated and used in this study.
Results: We have found variable changes across healthy and DMD astrocytes in the expression of key genes involved in cellular responses to DNA damage by RT-qPCR. However, when assessing nuclear accumulation of gH2AX in response to DNA damage, DMD astrocytes displayed higher basal gH2AX accumulation and greater vulnerability to DNA damage induction by camptothecin, a DNA topoisomerase inhibitor, and hydrogen peroxide, an inducer of oxidative stress. We have also observed some changes in DMD astrocytes, as compared to healthy ones, in the expression of genes involved in cellular antioxidant responses, which may underlie some of this vulnerability. Furthermore, we observed increased autophagic flux in DMD astrocytes relative to controls, suggesting cells may be increasing autophagy to cope with increased cell damage.
Conclusion: Our data support the hypothesis that DMD astrocytes are vulnerable to oxidative stress which subsequently impacts DNA damage burden and leads to increases in autophagic flux. We are currently assessing ROS production in DMD astrocytes and lysosome activity to explore these as potential therapeutic targets.
022Application of PASTE for the development of Duchenne muscular dystrophy therapies: Optimisation of atgRNAs to DMD intron 1
David Liddell1, Linda Popplewell1, Marc Moore1
1Royal Holloway of University London
David.Liddell.2026@live.rhul.ac.uk
Background: Duchenne muscular dystrophy (DMD) is an X-linked muscle-wasting disease caused by loss of dystrophin. Many genetic therapies are mutation-specific and/or require repeat dosing, while double-strand break (DSB) genome editing raises safety concerns and homology-directed repair (HDR) is inefficient in skeletal and cardiac muscle. PASTE (Programmable Addition by Site-Specific Targeting Elements) (Yarnall et al., 2022) enables targeted, DSB-free integration through an attachment-site containing guide RNA (atgRNA) which installs an attB landing site, which supports Bxb1-mediated integration of an attP-bearing payload. We aim to target DMD intron 1 to integrate a sequence-optimised microdystrophin cDNA (exon 2 onwards) as a durable, broadly applicable replacement strategy.
Aims: To develop a broadly applicable DMD editing approach using PASTE by (i) enabling targeted intronic integration without DSBs; (ii) optimising attB landing-site installation and microdystrophin knock-in efficiency; and (iii) establishing suitability for non-dividing skeletal and cardiac muscle. Optimisation will be performed in vitro (HEK293T and cultured patient muscle cells), followed by in vivo proof-of-concept testing in hDMD(del45) mdxD2 mice with functional assessment of muscle performance.
Methods/Materials: Candidate atgRNA sets were designed in silico using ML-guided ranking and clustered across a ∼2 kb target window in DMD intron 1. Initial analysis generated 278,031 putative atgRNA candidates; these were filtered using predefined selection criteria (Fell et al, 2024) to prioritise 24 top-ranked atgRNAs (atgRNA_rank). Selected atgRNAs and control atgRNAs were cloned into appropriate plasmids and transiently transfected into HEK293T cells alongside PASTE plasmids (Yarnall lab). attB installation was screened by junction PCR (nested PCR where required). Top-performing guides will be confirmed by targeted next-generation sequencing, and insertion efficiency/junction fidelity quantified by ddPCR across integration boundaries.
Results: Twenty-four intron-1 atgRNAs have been prioritised and cloned. A design-and-screen workflow has been established to compare attB insertion efficiencies across guide combinations in HEK293T cells, using published atgRNAs to other targets as positive controls.
Conclusion: This work establishes a streamlined pipeline for PASTE guide design and benchmarking at the DMD locus, enabling prioritisation of high-performing guide architectures for subsequent dystrophin cargo knock-in and functional testing in patient cells and humanised DMD mouse models.
023Longitudinal Assessment of Arterial Blood Pressures during General Anaesthesia in the DE50-MD Canine Model of Duchenne Muscular Dystrophy
Livingstone TS1, Stathopoulou T1,2, Piercy RJ2
1Department of Anaesthesia and Analgesia, Department of Clinical Sciences and Services, Royal Veterinary College, Hatfield AL9 7TA, United Kingdom
2Comparative Neuromuscular Diseases Laboratory, Department of Clinical Science and Services, Royal Veterinary College, London, NW10TU, United Kingdom
tlivingstone5@rvc.ac.uk
Background: Duchenne muscular dystrophy (DMD) is a fatal muscle wasting disease caused by mutations in the dystrophin gene and absence of dystrophin protein expression: most patients die due to progressive cardiovascular disease. However, exploring cardiovascular changes in DMD is hampered by poor sample availability, concomitant treatments (corticosteroids and blood-pressure modifying therapies), by co-morbidities (e.g. obesity) and difficulties accessing suitable controls. In contrast, large animal DMD models provide unrivalled opportunity for relevant pathophysiological investigation: of these, the DE50-MD canine model carries a splice-site mutation that induces exon 50 deletion (at the centre of the DMD mutational hotspot), sharing a phenotype that closely resembles young DMD patients. Here we hypothesised that DE50-MD dogs would display aberrant blood pressures in the controlled environment of general anaesthesia (GA).
Aims: To longitudinally investigate changes in arterial blood pressures in the DE50-MD model of DMD during GA compared to a control group.
Methods/Materials: Untreated 18 DE50-MD and 12 wild-type (WT) control male dogs underwent GA for non-invasive MRI imaging at the ages of 3, 6, 9 and 12 months with identical protocols according to Association of Veterinary Anaesthetist guidelines; relevant parameters were recorded at 5 minutes intervals for (154 minutes +/- 54) and data averaged for each animal. Linear mixed models with post-hoc analysis were used to evaluate effects of genotype and age.
Results: Compared to WT dogs, DE50-MD dogs had significantly higher mean (67mmHg +- 9.8 vs 59mmHg +- 6.6, P=0.003) and diastolic (49mmHg +- 10.0 vs 42mmHg +- 7.3, P=0.006) blood pressures from the ages of 6m and 9m onwards respectively; no changes were detected in systolic blood pressures. End-tidal sevoflurane (EtSevo) was not significantly different between genotypes or age groups (P=0.58). Heart rates were significantly higher in DE50-MD dogs compared to WT, but did not correlate with mean (R2=0.06, P=0.08) or diastolic (R2=1.16e-006, P=0.99) blood pressure.
Conclusion: Higher blood pressures detected during GA in DE50-MD dogs compared to controls, might reflect a critical, previously under-recognised early DMD disease feature. Our future goal is to determine whether blood pressure differences are accentuated in conscious, exercising
024Enhancing antisense oligonucleotide efficacy in Duchenne muscular dystrophy by histone deacetylase inhibitors drug repurposing
Ramla Omar1, Rachele Rossi2, Francesco Muntoni1
1UCL Great Ormond Street Institute of Child Health
2Vesalic Limited
ramla.omar@ucl.ac.uk
Background: Duchenne muscular dystrophy (DMD) is a severe neuromuscular disorder caused by mutations in the DMD gene, most commonly deletions that disrupt the reading frame and prevent production of functional dystrophin protein. Mutations that preserve the reading frame result in the milder Becker muscular dystrophy (BMD). Antisense oligonucleotides (ASOs) have emerged as a promising therapeutic strategy for DMD, using exon skipping to restore the dystrophin reading frame and produce truncated but functional protein similar to BMD. Currently, four ASOs, eteplirsen, golodirsen, viltepso, and casimersen, are FDA-approved for DMD. However, a major limitation of ASO therapy is the reduced dystrophin transcript levels in DMD patients, secondary to epigenetic changes at the DMD locus, which limits the availability of pre-mRNA for exon skipping and reduces dystrophin production.
Aims: This study aimed to enhance dystrophin expression in DMD patient-derived myotube cells by using histone deacetylase inhibitors (HDACi), specifically valproic acid (VPA) and givinostat, to promote an open chromatin structure and increase transcription of dystrophin. By increasing dystrophin transcript levels, we hypothesized that the substrate for ASO-induced exon skipping would be amplified, resulting in higher dystrophin production.
Methods/Materials: DMD patient myotubes amenable to exon 53 skipping were treated with 3 mM valproic acid (VPA) or 200 nM givinostat to induce HDAC inhibition followed by treatment with ASO golodirsen (N3). Dystrophin transcript levels were quantified using qRT-PCR, and dystrophin protein expression was analysed using Wes protein detection.
Results: Combinatorial treatment with HDACi (VPA or givinostat) and golodirsen significantly increased dystrophin transcript levels. Protein analysis showed an approximate two-fold increase in VPA+ golodirsen treated cells and a ∼1.5-fold increase in givinostat+ golodirsen treated cells compared to golodirsen only treated cells.
Conclusion: These results demonstrate that HDAC inhibition can enhance dystrophin transcript availability and dystrophin protein production in DMD patient cells. We are currently extending this combinational approach to DMD patients with deletions eligible to casimersen and eteplirsen.
025Characterisation of skeletal muscle immune cell infiltrates in the DE50-MD canine model of Duchenne muscular dystrophy
Priskila Sophiana1, Donald Palmer2, Richard J. Piercy1
1Comparative Neuromuscular Disease Laboratory, Department of Clinical Science and Services, Royal Veterinary College, London, NW10TU, UK
2Comparative Physiology and Clinical Sciences, Department of Comparative Biomedical Sciences, Royal Veterinary College, London, NW10TU, UK
psophiana@rvc.ac.uk
Background: Duchenne muscular dystrophy (DMD) results from dystrophin deficiency and is characterised by progressive skeletal muscle inflammation and fibrosis. Although histopathological studies demonstrate that focal inflammation precedes fibrosis, the cellular identity and temporal kinetics of immune cell infiltration remain poorly understood, since muscle biopsy is infrequent in DMD patients and most are treated from a young age with corticosteroids. Further, obtaining age-matched control human tissue is challenging. Consequently, the immune mechanisms driving DMD pathological changes are incompletely characterised. Here, we provide a longitudinal characterisation of immune cell infiltrates in DMD muscle to elucidate the cellular and temporal dynamics of inflammation.
Aim: We utilised the DE50-MD canine model, which recapitulates human DMD pathology including progressive skeletal muscle degeneration, functional decline, and robust inflammatory responses, to comprehensively and longitudinally characterise and define the cellular composition of immune infiltrates in DMD muscle.
Methods/Materials: We analysed vastus lateralis muscle tissue from male DE50-MD and control (WT) dogs at 3, 6, 9, 12, 15, and 18 months of age, supplemented by post-mortem sampling of multiple skeletal muscle regions. Immune cell populations—including macrophages, T lymphocytes, and B lymphocytes—were characterised using complementary histopathological approaches: conventional histology (haematoxylin and eosin; H&E) and acid phosphatase (AP) staining, coupled with immunohistochemistry and immunofluorescence microscopy.
Results: Immunofluorescence microscopy using anti-CD3 and anti-CD20 markers revealed early T and B lymphocyte infiltration in DE50-MD muscle from 3 months of age, spatially localised to regions of myofibre degeneration and regeneration. Both lymphocyte markers displayed characteristic perinuclear immunoreactivity with substantial CD3/CD20 co-expression and overlapping pan-leukocyte (CD18) signals, confirmed by confocal analysis. Notably, immunoreactivity was markedly attenuated in age-matched wild-type controls across all timepoints. T cell subset profiling identified significant CD4+ and CD8+ co-expression within CD3+ populations, whilst macrophage infiltration correlated with sites of acute myonecrosis.
Conclusion: We demonstrate that mixed T and B lymphocyte infiltration occurs early and spatially correlates with regions of muscle damage, establishing lymphocyte-driven inflammation as a central mechanism in DMD progression.
026Dynamic 31P NMR spectroscopy in skeletal and cardiac muscle in the canine Duchenne muscular dystrophy (DE50-MD) model
Thaleia Stathopoulou1, Victoria Watts1, Ladislav Valkovič2, Richard J. Piercy1
Department of Clinical Sciences and Services, Royal Veterinary College, London, UK
Division of Cardiovascular Medicine, Oxford Centre for Clinical Magnetic Resonance Research
tstathopoulou@rvc.ac.uk
Background: Duchenne muscular dystrophy (DMD) is an X-linked, progressive, fatal neuromuscular disease caused by null mutations in the dystrophin gene, a gene that is normally expressed in skeletal and cardiac muscle. Mitochondrial dysfunction is an early consequence of dystrophin’s absence: it precedes muscle fibre degeneration. Dysfunctional mitochondria are normally eliminated by mitophagy, an important quality control process. In DMD, mitophagy is impaired providing an early indicator of disease progression. 31P Nuclear Magnetic Resonance spectroscopy (MRS) is a non-invasive technique that uses high field magnetic scanners to measure cellular energetics in small molecules within the intra- and extracellular space as markers of mitochondrial function. The DE50-MD canine model of DMD carries a mutation in the 3’ splice donor site of exon 50. In contrast with mouse models, it faithfully represents the early human DMD phenotype. Here we hypothesised that 31P MRS in DE50-MD dogs would reveal early metabolic compromise within cardiac and post stimulation in skeletal muscle.
Aims: To optimise the use of static and dynamic 31P MRS in skeletal and cardiac muscle in DE50-MD and control dogs
Methods/Materials: Five DE50-MD and two, age-matched unaffected wild type (WT) male dogs, aged 22 to 42 weeks, underwent 31P MRS of the cardiac muscle and of the cranial tibial muscle before and after isometric muscle contractions. A 3T clinical MRI scanner and a 28cm 31P/1H Flex surface coil 3T were used to acquire images. Two MRI-compatible subdermal unipolar stimulation needle electrodes (13mm X 0.40mm) were positioned subcutaneously on the latero-distal aspect of the stifle, to stimulate the fibular nerve. The muscle protocol consisted of 50Hz stimulations of 1 second each, repeated every 3 seconds for 30 repetitions.
Results: Cardiac and tibial cranial muscle 31P -MRS spectra were successfully acquired from all dogs. No muscle fatigue was observed. Early pilot data reveal that DE50-MD dogs have reduced muscle phosphocreatine (PCr) levels compared to WT controls.
Conclusion: Static and dynamic 31P MRS is feasible in DE50-MD dogs and might offer early, in vivo demonstration of metabolic biomarkers for disease progression and treatment efficacy.
027‡Amelioration of emotional reactivity behaviours and restoration of dystrophin interactors in mdx52 mice with neuronal targeted AAV micro-dystrophin gene therapy
Konstantina Tetorou1,2, Monica Rebeca Gil Garzon1, Alex Kavanagh2, Nicha Songsliph2, Shashwat Guha1, Ayana Withana2, Jenny Vartianen1, Wing Sum Chu1, Camila Vallve Maine1, Riccardo Privolizzi1, Simon Waddington3,4, Simon Beggs5,6, Joanne Ng1, Francesco Muntoni1,2
1Genetic Therapy Accelerator Centre, UCL Queen Square Institute of Neurology, London WC1N 3BG
2Dubowitz Neuromuscular Centre, UCL Great Ormond Street Institute of Child Health, London WC1N 3JH
3Gene Transfer Technology Group, EGA Institute for Women's Health, University College London, London, UK
4Wits/SAMRC Antiviral Gene Therapy Research Unit, Faculty of Health Sciences, University of the Witwatersrand, 2193 Johannesburg, South Africa
5Neuroscience, Physiology and Pharmacology, UCL, London, WC1E 6BT United Kingdom
6Developmental Neurosciences, UCL Great Ormond Street Institute of Child Health, London, WC1N 1EH United Kingdom
konstantina.tetorou.18@ucl.ac.uk
Background: Duchenne muscular dystrophy (DMD) is a severe neuromuscular disorder caused by mutations in X-linked DMD gene, resulting in disruption of functional dystrophin protein production. DMD patients exhibit progressive muscle weakness and 44% DMD individuals also experience intellectual disability and/or neurobehavioral complications, such as autism spectrum disorder, attention deficit hyperactivity disorder, and anxiety. These latter complications are due to deficiency of different dystrophin isoforms in the brain differentially affected by the location of the DMD gene mutation. The brain involvement is recapitulated in DMD mouse models with mutations differentially affecting Dp427, Dp140 dystrophin isoforms and displaying enhanced fear response with increased anxiety- and depressive-like behaviours.
Previous preclinical studies have shown partial restoration of dystrophin in the brain with antisense oligonucleotide (ASO) therapy delivered intracerebroventricular (ICV) resulted in partial amelioration of neurobehavioural phenotypes but low levels of Dp427 restoration.
Aims: To further address therapeutic approaches for brain dystrophin deficiency we generated Adeno-associated virus (AAV) gene therapy vector with neuronal targeted micro-dystrophin and evaluated neurobehavioural efficacy in mdx52 mice lacking both Dp427 and Dp140 isoforms.
Methods/Materials: We treated neonatal male mdx52 at P1 with ICV or intravenous (IV) 5x10vg/pup and 4x11vg/pup respectively of AAV.micro-dystrophin, controls were untreated male mdx52 and wildtype littermates n=15 per group). We performed a battery of neurobehavioural tests between 8-12 weeks including fear response, elevated zero maze, open field, light-dark box, balance beam and Catwalk XT. We also used simple western (WES) to quantify micro-dystrophin and dystrophin’s interacting proteins in the brain.
Results: We demonstrated statistically significant improvement of emotional reactivity related behaviours in treated compared to untreated mdx52 littermates. Some behaviours were restored to wildtype levels showing improved efficacy compared to previous ICV ASO studies. Micro-dystrophin protein levels showed a rostro-caudal gradient with 60% wild-type levels in hippocampus and midbrain, with 10% in the cerebellum after ICV delivery and between 10-40% in all brain areas with IV delivery.
Conclusion: This is the first preclinical therapeutic study demonstrating brain targeted micro-dystrophin improves the neurobehavioural deficits observed in DMD mouse model with ICV and IV delivery.
028Screening of novel exon-skipping antisense oligonucleotide conjugates for Duchenne muscular dystrophy
Laia Torres-Masjoan1,*, Arta Aghaeipour1, David Coulson2, Jennifer Frommer2, Sara Aguti1, Haiyan Zhou3,4, Francesco Muntoni1,4,5, and TransNAT consortium
1Neurodegenerative Diseases, UCL Queen Square Institute of Neurology, London, UK
2Department of paediatrics, Oxford University, Oxford, UK
3Genetics and Genomic Medicine Research and Teaching Department, UCL Great Ormond Street Institute of Child Health, London, UK
4National Institute for Health Research, UCL Great Ormond Street Institute of Child Health, London, UK
5The Dubowitz Neuromuscular Centre, Molecular Neuroscience Section, Developmental Neurosciences Research and Teaching Department, UCL Great Ormond Street Institute of Child Health, London, UK
l.masjoan@ucl.ac.uk
Background: Duchenne muscular dystrophy (DMD) is an X-linked neuromuscular disease caused by the absence of dystrophin, a protein crucial for muscle function and integrity, and most cases result from reading frame-shifting exon deletions of the DMD gene. One therapeutical approach is the use of exon-skipping antisense oligonucleotides (ASOs) to restore the reading frame of the DMD mRNA, allowing the production of a shorter but functional form of dystrophin. While four exon-skipping ASOs to treat DMD have been approved by the US FDA, they fail to achieve dystrophin protein restoration levels higher than 5%, and there is a current need to develop new ASOs with improved cellular uptake, endosomal escape and target engagement by modifications of their structure or by conjugations to different moieties.
Aims: As part of the UKRI-funded TransNAT consortium, this study aims at developing and testing new ASOs conjugated to different moieties, on various DMD models.
Methods/Materials: The novel exon-skipping ASO-conjugates, all synthetized on a PS-2’MOE backbone and targeting exon 53 of the DMD gene, were screened on immortalized human skeletal muscle cells and hiPSCs-derived cardiac muscle cells, all derived from patients carrying a deletion of exon 52. Exon-skipping and protein restoration were analysed with RT-qPCR and Simple Western Blot, respectively. In addition, the intracellular localization of the ASOs inside the muscle cells was assessed using and antibody targeting the PS-bond of the ASOs in combination with antibodies targeting endosomal and lysosomal markers.
Results: When delivered gymnotically, we observed exon-skipping with all the conjugates tested, although this was not significantly higher than the exon-skipping levels achieved with the unconjugated PS-2’MOE counterpart. A PS-bond immunostaining showed that most of the ASOs remain in the cytoplasm, with limited access to the nucleus. Further analyses in which the ASOs were colocalized with lysosomal markers suggest that the conjugates tested remain largely entrapped in the endo-lysosomal system.
Conclusion: The exclusion of the PS-2’MOE ASO-conjugates from the nucleus and their co-localization with in endolysosomal markers could explain the low levels of exon-skipping observed.
029Mechanoregulation of Muscle Degeneration in Duchenne Muscular Dystrophy
1 John Walton Muscular Dystrophy Research Centre (JWMDRC), Clinical and Translational Research Institute, Newcastle University, Newcastle upon Tyne, UK
2 Dubowitz Neuromuscular Centre, Great Ormond Street Institute of Child Health, University College London, London, UK
3 Preclinical In Vivo Imaging, Translational and Clinical Research Institute, Newcastle University, Newcastle upon Tyne, UK
4Centre de Biologie Structurale, INSERM, CNRS, University of Montpellier, Montpellier, France
E.Villalobos@ucl.ac.uk
Background: In Duchenne Muscular Dystrophy (DMD), mutations in the dystrophin gene lead to muscle degeneration and progressive fibro-fatty tissue remodelling. Fibro/adipogenic progenitors (FAPs) are central to this process through their differentiation into fibroblasts and adipocytes that reshape the muscle microenvironment. While tissue remodelling is accompanied by changes in mechanical properties, including increased stiffness, the impact of these cues on FAPs differentiation in DMD is not yet defined. This study aims to dissect how stiffness-driven remodelling of the muscle microenvironment in DMD regulates FAPs cell fate and to identify key mechanisms driving this process.
Methods: FAPs and preadipocytes (SGBS) were cultured on substrates of varying stiffness. (2 – 106 kPa). Physiological muscle stiffness was assessed in control individuals’ muscle (gastrocnemius) using shear wave elastography (SWE) and a palpation device. We induced FAP differentiation into fibroblast and adipocytes with TGF-ß1(3d) and adipogenic cocktail (1-10d), respectively. Differentiation was assessed by western blot (WB) analysis of fibrotic (Collagen I, Fibronectin, α-SMA) and adipogenic markers (FABP4 and PPARγ). To evaluate changes of stiffness in DMD, single-cell mechanical properties were measured using atomic force microscopy (AFM). Mechanistic pathways of adipogenesis were investigated using targeted protein kinase profiling, immunofluorescence, and WB.
Results: Cell morphology and survival (SGBS and FAPs) was impaired at soft surfaces. This effect was greater after adipogenic differentiation. Measurements of muscle stiffness enabled the selection of physiologically relevant stiffness (8 kPa) for in vitro experiments. After differentiation, FAPs and SGBS cells cultured on stiffer substrates displayed increased levels of adipogenic markers (FABP4 and PPARγ). The effect of stiffness on extracellular matrix proteins induced by TGF-β1 was less clear. Under basal conditions, FAPs mechanical properties were comparable in DMD and control cells. DMD FAPs exhibited reduced levels of FAK (Tyr397) and an altered vinculin distribution under basal conditions. Proteomic analysis further identified WNK1 and PRAS40 as key kinases involved in early adipogenic differentiation.
Conclusions: These findings indicate that stiffness of the muscle microenvironment regulates FAPs adipogenic differentiation, with stiffer substrates promoting adipogenesis. Altered mechanotransduction in DMD FAPs, including reduced FAK activation, vinculin redistribution, and involvement of WNK1 and PRAS40, may contribute to pathological fibro-fatty remodelling.
030A novel human 3D culture platform for disease and therapy modelling in Duchenne Muscular Dystrophy
Mariam Zouhair1,2, Eugenia Carraro2,3, Sumitava Dastidar1,2, Sungwoo Choi1,2, Enrica Cristiano1,4, Annamaria De Luca4, Eduard Ayuso5, Andrea Serio2,3, Francesco Saverio Tedesco1,2,6*
1 Department of Cell and Developmental Biology, University College London, London, UK
2 The Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK
3 United Kingdom Dementia Research Institute Centre, Institute of Psychiatry, Psychology and Neuroscience, King’s College London, London, UK
4 Department of Pharmacy Drug Science, University of Bari Aldo Moro, Bari, Italy
5 DINAMIQS, Wagistrasse 18, 8952 Schlieren, Switzerland
6 Dubowitz Neuromuscular Centre, Great Ormond Street Hospital for Children & UCL Great Ormond Street Institute of Child Health, London, UK
presenting author: mariam.zouhair@crick.ac.uk
Corresponding author: f.s.tedesco@ucl.ac.uk
Background: Duchenne muscular dystrophy (DMD) is a severe X-linked neuromuscular disorder caused by mutations in the dystrophin gene, resulting in progressive muscle degeneration, functional decline, and reduced life expectancy. Although gene therapies hold considerable promise, their clinical translation has been hindered by variable efficacy and safety concerns that are often poorly predicted by conventional animal models. In addition, species-specific differences limit the direct translatability of preclinical findings to human patients, underscoring the need for improved human-relevant in vitro models.
Aims: To develop and validate a human-specific three-dimensional (3D) skeletal muscle model that recapitulates key pathological features of DMD and enables in vitro evaluation of DMD-targeted gene therapy strategies.
Methods/Materials: We engineered 3D human skeletal muscle constructs using tissue-derived or induced pluripotent stem cell (iPSC)-derived myogenic cells combined with microfabricated devices featuring defined topographical cues to promote myofibre alignment and functional maturation. This platform integrates expertise in human stem cell biology, muscle regeneration, and bioengineering. As an extension of the model, a controlled bilineage configuration was implemented to assess adeno-associated virus (AAV) transduction in a multicellular context.
Results: Engineered human muscle constructs reproduced hallmark morphological and functional features of DMD, including dystrophic muscle architecture and impaired sarcolemmal integrity, reflecting disease-associated phenotypes observed in vivo. Within the framework of the international and multi-disciplinary MAGIC consortium (www.magic-horizon.eu), this platform was used to evaluate and compare transduction efficiency of different gene therapy vector serotypes. In this context, incorporation of supporting cell types revealed differences in AAV transduction efficiency and specificity that are not captured in myofibre-only systems.
Conclusion: This human 3D skeletal muscle platform provides a robust and physiologically relevant model for studying DMD pathology and evaluating gene therapy strategies. The multi-lineage model improves the predictive assessment of AAV specificity by introducing controlled cellular heterogeneity relevant to dystrophic muscle, thereby enhancing translational relevance beyond traditional animal models and supporting the development of more effective genetic therapies for muscular dystrophies.
Dystrophy Clinical
031The Newcastle Distal and Myofibrillar Myopathy Cohort
Ariele Barreto Haagsma1, Doaa Salman1, Carla Bolano Diaz1, Marianela Schiava1, Tara Reeves1, Pietro Riguzzi1, Goknur Kocak1, Maha Elseed1, Elizabeth Harris1 , Ana Töpf1, Judith Hudson2, Robert Muni Lofra1, Emma Grover1, Emma-Jayne Robinson1, Michelle McCallum1, Jassi Michell-Sodhi1, Dionne Moat1, Karen Wong1, Peter Waldock1, Julie Walsh1, Stephanie Tanner1, Jordi Diaz-Manera1 , Michela Guglieri1 , Chiara Marini Bettolo1 , Volker Straub1, Giorgio Tasca1
1The John Walton Muscular Dystrophy Research Centre, Translational and Clinical Research Institute, Newcastle University and Newcastle Hospitals NHS Foundation Trust, UK
2Northern Molecular Genetics Service, Biomedicine East Wing, Newcastle upon Tyne, UK
ariele.barreto-haagsma@newcastle.ac.uk
Background: Distal and myofibrillar myopathies (DM and MFM) are umbrella terms for rare diseases showing significant overlap.
Aims: to describe the features of the MFM and DM population followed at the John Walton Muscular Dystrophy Research Centre.
Methods/Materials: This retrospective single-centre study included patients with either a myopathological diagnosis of MFM or a clinical diagnosis of DM. Demographic, genetic, clinical, and respiratory data were collected from July 1995 to April 2024. Variables assessed included disease causing variants, age at onset, first symptoms, and ambulatory status.
Results: The cohort included 127 patients. Sixty-four patients fulfilled the criteria for MFM (mean age 49.26 ± 14.62 years, 68.8% male) and eighty-four for DM (mean age 47.32 ± 15.39 years, 63.1% male). Twenty-one patients (16.5% of the whole cohort) matched the criteria for both. Seventy-eight percent of the patients were molecularly diagnosed. In DM, the most frequent causative gene was GNE (29.8%), while variants in TTN or DES were found in 59.4% of MFM patients. Age at onset did not significantly differ between the MFM and DM subgroups (mean 40.89 ± 14.64 years versus 37.26 ± 14.41 years, p = 0.169). TTN variants were significantly associated with respiratory symptoms and DES variants with cardiac symptoms at onset. Both these genes were also significantly associated with lower sitting forced vital capacity values at initial assessment.
Conclusion: This service evaluation provides an initial cross sectional characterization of the DM/MFM cohort followed at the John Walton Muscular Dystrophy Research Centre. The next steps will be the collection of retrospective longitudinal, muscle biopsy and MRI data.
032COS-PREPARED: Clinical Outcome Study in PRe- and Early-symptomatic PAtients with REcessive Dysferlinopathy
Carla Bolano Diaz1, Ariele Barreto Haagsma1, Heather Hilsden1, Sarah Emmons2, Meredith K James1, Heather Gordish Dressman,3,4, Ian Wilson5, Laura Rufibach, Lisa Alcock6, Volker Straub1
1 John Walton Muscular Dystrophy Research Centre, Translational and Clinical Research Institute, The Newcastle upon Tyne Hospitals NHS Foundation Trust and Newcastle University, UK;
2 Jain Foundation, Seattle, USA
3 Center for Translational Science, Division of Biostatistics and Study Methodology, Children’s National Health System, Washington, DC, USA;
4 Pediatrics, Epidemiology and Biostatistics, George Washington University, Washington, DC, USA;
5 NMR Centre, Translational and Clinical Research Institute, Newcastle University, UK,
6 Gait Laboratory, Translational and Clinical Research Institute, The Newcastle upon Tyne Hospitals NHS Foundation Trust and Newcastle University, UK;
Background: The International Clinical Outcome Study in Dysferlinopathy (Jain COS) recruited 328 participants with LGMDR2 dysferlinopathy between 2012 and 2022. Almost all participants were already symptomatic by the time of enrolment.
Pre-symptomatic diagnoses or diagnoses at very early disease stages are increasingly generated through incidental CK results, screening for segregation analysis, or the diagnosis of a symptomatic family member leads to sibling diagnoses. An established pre- or early symptomatic diagnosis provides the opportunity to expand current knowledge of the early natural history and disease onset.
Aims:
1) To define the early natural history of dysferlin-deficient limb girdle muscular dystrophy (dysferlinopathy/LGMDR2) by monitoring muscle function, strength, and biomarkers of muscle pathology.
2) To identify early indicators of disease onset that will inform recognition, diagnosis, care considerations, clinical trial design and treatment indications.
Methods: Participants can be included if they have a confirmed diagnosis of dysferlin-deficient LGMDR2 proven by two (predicted) pathogenic in the DYSF gene, can stand on Tiptoes on both feet at the same time and can hop clearing foot off floor.
Participants are assessed using quantitative MRI (T2, Dixon), instrumented gait assessment and functional outcome measures over a five year period. Patient reported outcome measures (ACTIVLIM, McGill Pain, PROMIS Fatigue and Fatigue Severity score, IPAQ and RAPA) and age-appropriate quality of life measures will be completed.
Due to the rarity of pre symptomatic diagnoses, we seek the support of UK Neuromuscular Centres to identify eligible participants. The recruitment target is 20. All participants will be seen at Newcastle upon Tyne Hospitals.
Results: 5 participants (aged 13-19, 3 male, 2 female) have been recruited at the point of abstract submission. Baseline visit data will be presented
Conclusion: The study of symptomatic and early symptomatic patients will inform gene replacement therapy approaches which are likely to be most effective in early stages of disease progression. The PrePARED model could be applied to other neuromuscular diseases.
COS PRePARED is funded by the Jain Foundation. We are grateful to the young people and families who have committed to participate in this study over the next 5 years
033Clinical and phenotypic spectrum of caveolinopathies: an expanded family-based cohort study
Carla F Bolano-Diaz1, Marianela Schiava1, Ariele Barreto Haagsma1, Doaa Salma1, Pietro Riguzzi1, Karen Wong1, Emma Robinson1, Emma Grover1, Emmanuel Adu1, Pete Waldock1, Michelle McCallum1, Joseph Mason1, Stephanie Tanner1, Julie Walsh1, Jassi Michell Sodhi1, Dionne Moat1, Tiago Bernardino Gomes1,2, Karen Suetterlin1,3, Maha Elseed1, Meredith K James1, Robert Muni Lofra1, Yolande Parkhurst1,4, Elizabeth Harris1, Michela Guglieri1, Giorgio Tasca1, Jordi Diaz Manera1, Volker Straub1, Chiara Marini Bettolo1.
1The John Walton Muscular Dystrophy Research Centre, Translational and Clinical Research Institute, Newcastle University and Newcastle Hospitals NHS Foundation Trust, Central Parkway, Newcastle upon Tyne, UK.
2NHS England Highly Specialised Service for Rare Mitochondrial Disorders, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK.
3AGE Research Group, NIHR Newcastle Biomedical Research Centre, Newcastle Upon Tyne Hospitals NHS Foundation Trust, Cumbria, Northumberland, Tyne and Wear NHS Foundation Trust and Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK.
4NHS England HSS for Rare Neuromuscular Diseases, Muscle Immunoanalysis Unit, Newcastle upon Tyne, UK.
carla.bolano-diaz@newcastle.ac.uk
Introduction: Caveolinopathies are rare muscle disorders caused by pathogenic variants in CAV3, which encodes caveolin-3, a muscle-specific protein essential for sarcolemmal integrity, vesicular trafficking, and signal transduction within caveolae. Most CAV3 mutations act through a dominant-negative mechanism, leading to reduced or absent caveolin-3 at the plasma membrane. Clinically, caveolinopathies show a wide phenotypic spectrum, ranging from asymptomatic hyperCKaemia to muscle pain, stiffness, hypertrophy, and progressive weakness, often with marked intrafamilial variability.
Aims: To describe the clinical, genetic, functional, imaging, and pathological characteristics of a cohort of patients with CAV3-related disease and to further characterise phenotypic variability within and between families.
Methods: A retrospective review was performed of patients with CAV3 variants identified in a tertiary neuromuscular centre database. Available data included clinical features, family history, serum creatine kinase (CK), genetic results, electrophysiology, muscle imaging, and muscle biopsy findings.
Results: Thirty-nine patients from 15 families were identified, with detailed clinical data available for 25 patients from 13 families. Ten patients from five families had been previously reported in a UK caveolinopathy cohort1, while the remaining cases represent newly identified patients or additional affected relatives. The cohort was predominantly female (72%), and 84% had another affected family member. Median CK was 1035 U/L (IQR 754–1681). Five patients were asymptomatic at last follow-up, commonly identified through family screening or incidental findings. All patients were ambulant, and one required walking aids. Median age at assessment was 32.4 years, with a median symptom duration of 9.1 years. Muscle weakness was present in 15 patients, most commonly with a proximal pattern. No severe respiratory involvement was observed. Muscle biopsies frequently demonstrated markedly reduced or absent caveolin-3 expression with variable myopathic changes. Electrophysiological findings ranged from mild myopathic changes to spontaneous activity. Muscle MRI showed mild, selective fatty replacement with negative STIR sequences.
Conclusions: Caveolinopathies are characterised by substantial clinical and pathological heterogeneity, even within families. Despite significant biochemical and histopathological abnormalities, many patients remain mildly affected or asymptomatic. This expanded cohort highlights the value of comprehensive family-based assessment and longitudinal follow-up to better understand disease expression and progression.
References:1Aboumousa A, Hoogendijk J, Charlton R, et al. Caveolinopathy--new mutations and additional symptoms. Neuromuscul Disord 2008;18:572–578.
034‡Longitudinal behaviour of NSAD and muscle fat fraction as monitoring biomarkers in LGMD R2-DYSF related
Carla F Bolano-Diaz1, Heather Hilsden1, Meredith James1, Jain COS Consortium2, Giorgio Tasca1, Volker Straub1, Jordi Diaz Manera1.
1The John Walton Muscular Dystrophy Research Centre, Translational and Clinical Research Institute, Newcastle University and Newcastle Hospitals NHS Foundation Trust, Central Parkway, Newcastle upon Tyne, UK.
2The Jain Foundation, Seattle, Washington, USA
carla.bolano-diaz@newcastle.ac.uk
Introduction: Measuring disease progression and clinically meaningful change in limb girdle muscular dystrophies (LGMD) is challenging for multiple reasons. In the era of disease-modifying therapies, this challenge becomes even more. LGMD are inherited, progressive muscle-wasting disorders in which skeletal muscle is gradually replaced by fat and fibrotic tissue; therefore, sustained clinical improvement is not expected in the natural disease course.
Aims: The aim of this study was to better characterise two frequently used outcome measures, muscle fat fraction (FF) and the North Star Assessment for Dysferlinopathy (NSAD), as monitoring biomarkers.
Methods: We analysed NSAD and FF data from up to 193 participants enrolled in the Clinical Outcome Study for Dysferlinopathy (COS 1). Changes in both measures were assessed over one- and three-year intervals, with a focus on changes indicating clinical improvement. Descriptive analyses were performed, followed by comparative analyses using McNemar’s test. A reliability analysis was conducted for NSAD only, due to data availability.
Results: At one year, approximately 20% of both NSAD and FF assessments indicated improvement in function and muscle structure, respectively. For NSAD, 85.7% of improvements were ≤4 points, while mean FF change was −2.43% (95% CI −3.42 to −1.44). At three years, the proportion of apparent improvement decreased to 9% for both measures. NSAD improvements were ≤3 points in 83.3% of cases, and mean FF reduction was −4.50% (95% CI −6.72 to −2.26). FF improvements were observed in a relatively homogeneous subgroup—non-ambulant patients with baseline lower-limb FF >70%—whereas no consistent pattern was seen for NSAD improvements. No significant differences were found between outcome measures at either time point. The NSAD intraclass correlation coefficient was 0.985, with a minimal detectable change at 95% confidence of 5.07 points.
Conclusions: Although FF and NSAD show similar rates of unexpected improvement and comparable temporal behaviour, FF improvements occur within a more homogeneous patient subgroup, potentially facilitating interpretation. Most NSAD improvements fall below the calculated MDC95, highlighting the importance of interpreting small changes with caution and supporting further investigation of FF reliability thresholds.
Riccardo Curro1, Liedewei Van de Vondel2,3, Stefano Facchini1, Isaac Xu2, Jonathan De Winter3,4, Ilaria Quartesan1, Alice Monticelli3, Alicia Alonso-Jimenez4, Willem De Ridder4, Mark Roberts5, Stefen Brady6, Ashirwad Merve7, Kornelia Neveling8, Alexander Hoischen8, Solve-RD Consortium, Peter de Jonghe4, Henry Houlden1, Michael G Hanna1, Enrico Bugiardini1, Stephan Züchner2, Jonathan Baets3,4, Andrea Cortese1,9
r.curro@ucl.ac.uk
1Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, London, UK
2Dr. John T. Macdonald Foundation Department of Human Genetics and John P. Hussman Institute for Human Genomics, University of Miami Miller School of Medicine, USA
3Translational Neurosciences, University of Antwerp, Belgium
4Neuromuscular Reference Centre, Antwerp University Hospital, Belgium
5Department of Neurology, Salford Royal NHS Foundation Trust, Manchester, United Kingdom.
6Oxford Neuromuscular Centre, Department of Neurology, John Radcliffe Hospital, Oxford, UK.
7Division of Neuropathology, UCL Institute of Neurology, National Hospital for Neurology and Neurosurgery, London, UK.
8Department of Human Genetics, Radboud University Medical Center, Nijmegen 6525 GA, the Netherlands.
9Department Medical Biotechnology and Translational Medicine, Università Degli Studi Di Milano, Milan, Italy and Unit of Rare Neurological Diseases, Department of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy
Background: Oculopharyngodistal myopathy (OPDM) is a rare inherited myopathy characterized by a combination of progressive ocular, bulbar, and distal limb weakness. A hallmark pathological feature of OPDM is the presence of rimmed vacuoles and intranuclear inclusions on muscle biopsy. Recently, CCG/CGG repeat expansions in the 5’ UTR of six different genes have been associated with OPDM. However, a large proportion of OPDM patients remain genetically unsolved.
Aims: In this study, we identified a novel CCG repeat expansion in the 5' UTR of TBC1D7 as a cause of OPDM.
Methods/Materials: A combination of short and long-read sequencing was employed for the repeat identification and for profiling the repeat motif and size across large datasets, including a cohort of 940 neuromuscular cases from the 100,000 Genome Project. Long-read sequencing also provided information about the methylation status at the repeat locus in patients and unaffected family members. Repeat-primed PCR was performed for segregation analysis. TBC1D7 transcript expression in patients-derived fibroblasts was quantified using quantitative PCR. Routine histopathology and p62-staining were performed on the muscle biopsy from two patients.
Results: CCG expansions in the 5' UTR of TBC1D7 were identified in five individuals from three unrelated OPDM families. Median age at onset was 19 years (range: 14-58) and median repeat size in affected cases was 130 units (range: 83-264; reference sequence (CCGCTG3)(CCG 4). The largest repeat expansion (270 units) was detected in the unaffected father of one of the index cases; however, methylation profiling in this individual showed that the expanded allele was fully methylated. TBC1D7 transcript levels were upregulated in patient-derived fibroblasts compared to controls, and p62-positive intranuclear inclusions were detected on muscle biopsy, suggesting a toxic gain-of-function mechanism.
Conclusion: This study broadens the genetic landscape of OPDM and strengthens the link between the CCG/CGG repeat motif and a characteristic pattern of muscle weakness. Furthermore, it provides further evidence for the role of epigenetic factors as modifiers of the disease penetrance. Finally, it underscores the emerging role of non-coding repeat expansions in unsolved neurogenetic and neuromuscular disorders.
Affiliations: (1) Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, London, UK
036Two brothers with Morimoto-Ryu-Malicdan neuromuscular syndrome and review of phenotype
Elizabeth Harris1,2, Yolande Parkhurst 1,2, Adrian Miller3, Margaret Phillips4, Volker Straub1,2, Chiara Marini Bettolo1,2
1Highly Specialised Service for Rare Neuromuscular Disorders, Newcastle Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK
2The John Walton Muscular Dystrophy Research Centre, Translational and Clinical Research Institute, Newcastle University and Newcastle Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK
3University Hospital Southampton NHS Foundation Trust, Southampton, UK
4University Hospitals of Derby, Derby, UK
Elizabeth.harris20@nhs.net
Background: Biallelic variants in the RFC4 gene, which is implicated in DNA replication, are a recently described and so far rare cause of a disorder known as Morimoto-Ryu-Malicdan neuromuscular syndrome. Features of this condition are variable and include myopathy, respiratory insufficiency, cerebellar signs and cerebellar atrophy. Of the few reported cases age of symptom onset from the neonatal period, to childhood or adulthood. Some affected individuals have died prematurely. There is a need to better characterise this disease in order to provide useful information to affected individuals and families and to tailor medical care.
Aims: To describe genetic data and compare the phenotypes of two adult brothers with Morimoto-Ryu-Malicdan neuromuscular syndrome to previously reported cases.
Methods/Materials: Review of clinical data including genetic reports, clinical examination, muscle MRI and muscle biopsy of the patients and review of the literature.
Results: These brothers had symptom onset in their 20s or 30s with weakness, ophthalmoplegia and for one individual dysarthria and respiratory insufficiency. CK was 500-4000 units/l. Muscle biopsy showed perifascicular adipose tissue, rimmed vacuoles and cytoplasmic aggregates. Through the NHS Diagnostic Discovery service compound heterozygous variants in RFC4 were identified. Combining these two new cases with those reported in the literature we provide summaries of the 13 individuals with this diagnosis. In common with previously reported cases these brothers have predominantly proximal and axial weakness and cerebellar signs (dysarthria).
Conclusion: The two brothers described here add to the awareness of adult-onset Morimoto-Ryu-Malicdan neuromuscular syndrome, which can present from birth to adulthood with myopathy often together with cerebellar signs.
037Protein Intake is Associated with Body Composition and Quality of Life in Adults with Becker Muscular Dystrophy
1School of Sport, Rehabilitation and Exercise Sciences, University of Birmingham, UK
2Department of Sport and Exercise Sciences, Institute of Sport, Manchester
3The Neuromuscular Centre, Winsford, UK
4Faculty of Kinesiology and Physical Education, University of Toronto, Canada
ikh553@student.bham.ac.uk
Background: Becker Muscular Dystrophy (BMD) is a rare X-linked genetic condition characterised by reduced dystrophin expression, causing progressive skeletal muscle and functional decline, and poor quality of life (QoL). With no current pharmacological cure, practical easy-to-implement strategies are needed to improve QoL. Dietary interventions can positively influence lean body mass (LBM) and physical function [1, 2], both reduced in BMD, in other clinical conditions [3, 4]. However, little is known about how habitual diet is associated with these outcomes and QoL in BMD.
Aims: To characterise habitual dietary practises in adults with BMD and examine associations with muscle size, strength, physical function and self-reported QoL.
Methods/Materials: Adults with BMD (n=20; 46±12 years) and age-matched controls (n=12; 47±13 years) completed two weighed food diaries ∼6 weeks apart. Participants completed assessments of body composition, muscle size, strength and function as well as validated questionnaires on perceived physical function, pain/fatigue and QoL. Between-group differences were assessed using independent t-tests or Mann-Whitney U tests and correlations with Pearson’s R or Spearman’s Rho depending on normality (p<0.05). Ethical approval was obtained from institutional research ethics committee and procedures conformed to Declaration of Helsinki.
Results: BMD consumed 29% less energy than controls (1602 vs. 2229kcal, p<0.001), with 25-30% lower carbohydrate and fat intake (p<0.01). Relative protein intake was 21% lower (0.86±0.28 vs. 1.08±0.24g/kg/day, p=0.028), and only 10% of individuals met recommendations for disease-related catabolism (1.2g/kg/day). BMD also exhibited higher body fat (31.1±4.7 vs. 22.9±6.9%, p=0.002), fatigue and pain alongside reduced LBM, strength, physical function and self-reported QoL (all p<0.05).
Within BMD, relative protein intake was correlated positively with LBM (r=0.619; p=0.018) and QoL (r=0.596; p=0.006). LBM was also inversely associated with self-reported fatigue (r=-0.591, p=0.026) whilst BMI correlated positively with pain (r=0.691, p<0.001).
Conclusion: Here, we observe adults with BMD report lower energy consumption, yet display elevated body fat. Relative protein intake was also lower in BMD and below guidelines for similar clinical conditions (1.2g/kg/day). Moderate-to-strong correlations between protein intake, LBM and QoL in BMD suggest interventions which elevate protein consumption could benefit this population, to not only improve LBM but reduce pain/fatigue and elevate QoL.
Wirth, J., E. Hillesheim, and L. Brennan, The role of protein intake and its timing on body composition and muscle function in healthy adults: a systematic review and meta-analysis of randomized controlled trials. The Journal of nutrition, 2020. 150(6): p. 1443-1460.
Noh, H.-M., et al., Association between dietary protein intake, regular exercise, and low back pain among middle-aged and older korean adults without osteoarthritis of the lumbar spine. Journal of Clinical Medicine, 2022. 11(5): p. 1220.
Damanti, S., et al., Efficacy of nutritional interventions as stand-alone or synergistic treatments with exercise for the management of sarcopenia. Nutrients, 2019. 11(9): p. 1991.
Van de Worp, W.R., et al., Nutritional interventions in cancer cachexia: evidence and perspectives from experimental models. Frontiers in nutrition, 2020. 7: p. 601329.
038Protein Intake Positively Associates with Body Composition and Quality of Life Independent of Mobility Status in Adults with Muscular Dystrophy
Leaver, Meg.1,2, Haslam, Isobel.1,2, Morgan Paul. 2, Orme, Paul.3, Flannery, Orla2., Bowden-Davies, Kelly. 2, Morse, I. Christopher2. Hodson, Nathan1,2,4.
1 School of Sport, Exercise and Rehabilitation Science, University of Birmingham, Birmingham
2 Department of Sport and Exercise Sciences, Institute of Sport, Manchester Metropolitan University, Manchester, UK
3Neuromuscular Centre, Winsford, Cheshire, UK
4 Faculty of Kinesiology and Physical Education, University of Toronto, Ontario, Canada
mnl538@student.bham.ac.uk
Background: In recent years, advancements in clinical care have substantially increased longevity in those living Muscular Dystrophy (MD), yet ambulatory loss still occurs in many individuals (1-3). We have recently observed that adults with MD consume less dietary protein than non-dystrophic individuals and, in MD, protein intake was positively associated with lean mass, grip strength and quality of life. In similar chronic conditions, mobility is a known barrier to nutritional adequacy(4), however the impact of mobility on habitual diet in MD remains unexamined.
Aims: To evaluate the impact of mobility on nutritional status in MD adults and probe associations between nutrition, skeletal muscle parameters and quality of life (QoL) while considering mobility as a cofounding factor.
Methods: Secondary analysis from two adult MD cohorts (FSHD, LGMD, BMD, DM1), stratified by mobility: ambulatory (AB, n=37; 56±2 yrs; 27.4±4.5 kg/m2) and non-ambulatory (NAB, n=22; 54±2 yrs; 27.8±5.5 kg/m2) was conducted. Participants completed two 3-day food diaries, validated functional/QoL questionnaires, alongside assessments of body composition, forearm muscle thickness, and upper-limb strength. Group differences were analysed using t-tests or Mann–Whitney U tests, correlations with Pearson’s r or Spearman’s ρ, and protein intake distribution with linear mixed models with Bonferroni- corrected pairwise t-tests performed post hoc. Significance was set at p<0.05, corrected for multiple comparisons where appropriate.
Results: Groups did not differ in BMI, lean mass percentage or self-reported physical activity (p>0.05). NAB reported lower absolute (g) (-16%, p=0.003) but not relative (g/kg) (-13%, p=0.088) protein intake, with no difference in total energy intake. Energy and protein intake (g/kg) were both highest at dinner (+30% vs. lunch; +62% vs. breakfast; all p<0.001) with no effect of ambulatory status. NAB exhibited 49–54% lower strength (all p<0.05) but similar forearm muscle thickness. Adjusted for mobility, relative protein intake (g/kg) correlated positively with lean mass (r=0.34, p=0.029), lower limb function (r= 0.386, p = 0.006), and QoL (r=0.353, p=0.013).
Conclusion: Non-ambulatory MD adults consume less protein despite comparable energy intake. Given the observed associations with function and QoL, ensuring adequate protein intake may be critical for maintaining muscle and QoL in MD, independent of mobility status.
Audhya IF, Cheung A, Szabo SM, Flint E, Weihl CC, Gooch KL. Progression to Loss of Ambulation Among Patients with Autosomal Recessive Limb-girdle Muscular Dystrophy: A Systematic Review. Journal of Neuromuscular Diseases. 2022;9(4):477-92.
Tawil R, Kissel JT, Heatwole C, Pandya S, Gronseth G, Benatar M. Evidence-based guideline summary: Evaluation, diagnosis, and management of facioscapulohumeral muscular dystrophy: Report of the Guideline Development, Dissemination, and Implementation Subcommittee of the American Academy of Neurology and the Practice Issues Review Panel of the American Association of Neuromuscular & Electrodiagnostic Medicine. Neurology. 2015;85(4):357-64.
Stimpson G, Ridout D, Sarkozy A, Manzur A, Muntoni F, Baranello G. Relationship between growth and ambulation loss in Duchenne muscular dystrophy boys on steroids. Eur J Neurol. 2024;31(12):e16415.
Wetherill MS, Duncan AR, Bowman H, Collins R, Santa-Pinter N, Jackson M, et al. Promoting nutrition equity for individuals with physical challenges: A systematic review of barriers and facilitators to healthy eating. Preventive Medicine. 2021;153:106723.
039A Telomeric Translocation Causing Facioscapulohumeral Muscular Dystrophy in a Family with Normal D4Z4 Size
Ilaria Quartesan1, Stefano Facchini1, Richard J.L.F. Lemmers2, Stephanie Efthymiou1, Natalia Dominik1, Patrick J. van der Vliet2, Giorgio Tasca3, Andrea Cortese1, Silvère M. van der Maarel2, Volker Straub3, Enrico Bugiardini1
i.quartesan@ucl.ac.uk
1UCL Queen Square Institute of Neurology, University College London, London, UK
2Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands
3John Walton Muscular Dystrophy Research Centre, Newcastle University, Newcastle upon Tyne, UK
Background: Facioscapulohumeral muscular dystrophy (FSHD) results from aberrant DUX4 expression associated with D4Z4 chromatin relaxation, caused by either repeat contraction (FSHD1) or mutations in D4Z4 chromatin repressors such as SMCHD1 (FSHD2). While standard testing confirms diagnosis in the vast majority cases, a small subset of patients with clinically FSHD remains unsolved, suggesting that pathogenic mechanisms not detected by conventional diagnostics may exist, and require advanced technologies to be uncovered.
Aims: To identify the genetic cause in a familial case with highly suggestive FSHD features but negative extensive genetic workup.
Methods: We investigated two affected siblings presenting with asymmetric shoulder girdle and distal leg weakness, scapular winging, and variable facial involvement. Muscle MRI showed a characteristic FSHD pattern. Standard testing included Southern blotting, whole genome sequencing (WGS), and methylation analysis. Optical genome mapping (OGM) was performed in both affected siblings and one unaffected sibling. Long-read sequencing (LRS) was used to better characterise the 4q35 region and methylation status. DUX4 expression was assessed in fibroblasts from both affected individuals.
Results: Southern blotting and methylation analysis were negative for both FSHD type 1 and 2 and neuromuscular gene panels from WGS were also unremarkable. However, OGM detected a structural rearrangement involving chromosomes 4q and 7 on the 4qA-17 allele, present in both affected siblings but absent in the unaffected sibling. LRS demonstrated that this translocation results in a ∼115 kb insertion within the telomeric region, shortening the (TTAGGG)n sequence from ∼3 kb to <1 kb. Methylation-aware analysis revealed focal hypomethylation of distal D4Z4 units. Aberrant DUX4 expression was confirmed in fibroblasts from both affected siblings.
Conclusions We report a novel FSHD mechanism whereby a telomeric translocation causes focal D4Z4 hypomethylation and aberrant DUX4 expression, independent of both repeat contraction and chromatin modifier mutations. Importantly, this mechanism escapes gold-standard diagnostics, underscoring the utility of advanced genomic technologies in genetically unsolved case where clinical suspicion of FSHD is high.
040A qualitative study of the applicability of the United Kingdom Duchenne Muscular Dystrophy guidelines to the practice of Physiotherapists working in Zambia
Kapapa MM1, Kvalsund MP2 3, Banda M 4 , Bearden D 2, ICGNMD Consortium5, Ramdharry G 5.
1University Teaching Hospital Zambian Institute for Neurologic Care, Research & Education Office
2Department of Neurology, University of Rochester Medical Center, Rochester, New York, USA
3Department of Internal Medicine, University of Zambia, Lusaka, Zambia
4Centre for Research in Disability, Rehabilitation and Policy Development (CR-DRPD)
5Centre for Neuromuscular Diseases. 8-11 Queen Square. London. WC1N 3BG UK
mutihams1@gmail.com
Background information: Advanced care pathways in the management of Duchenne Muscular Dystrophy (DMD) in corticosteroids, other disease-modifying drugs and the use of guidelines is a growing topic. Care guidelines for Physiotherapists working with people with DMD were developed in the UK in 2025 https://doi.org/10.1016/j.ejpn.2025.03.011. Assessing the knowledge and applicability of existing guidelines, especially in low- & middle-income countries, is crucial to improving implementation of evidence-based recommendations and patient outcomes. No previous studies have reported the applicability of these guidelines in Zambia.
Aim: Examine how physiotherapists view the applicability of United Kingdom developed guidelines for DMD in a resource-constrained country.
Method: An online qualitative survey was conducted among physiotherapists working in different clinical settings. Purposive sampling was employed, and the data was collected online. Thematic analysis was used to code data using the Social Ecological Model to group themes.
Results:Intra and interpersonal level: Regular functional assessments, exercise therapy, early contracture prevention, and education were seen as highly applicable by physiotherapists. Physiotherapists had limited practical knowledge in respiratory management and reported poor referral networks to specialist departments. Organizational level: Use of low-cost interventions was viewed as more implementable than interventions involving expensive technology and equipment. Participants described an absence of a focused DMD system and inconsistent cardiopulmonary testing access as well as a lack of neuromuscular occupational and speech therapists outside tertiary referral centers.
Community level: lack of funding for non-invasive respiratory equipment, community visits, education and follow-up programs.
Policy level: No local DMD registry to track patients' needs, service gaps, including lack of ministerial awareness/education programs at provincial, district, and hospital levels.
Conclusion: Resource allocations, including focused physiotherapy training, are needed to optimise the applicability of the UK DMD guideline among Zambian physiotherapists in clinical settings, and the inclusion of community-based rehabilitation programs which are currently unavailable.
Additional studies are required to evaluate if and how possible adaptations and health service re-organisation may improve implementation and perceived applicability of the guidelines in this and similar settings.
Mitochondrial Disease
041A Homozygous DCLK2 Variant as a Potential Modifier of Disease Severity in PDHA1-Associated Primary Pyruvate Dehydrogenase Complex Deficiency
Dalal A. Al-Mutairi1*, Fahad K. Al-Mutairi2, Ahmad Al-Shami2, Nawal Makhseed2, William L. Macken3,4, Robert D Pitceathly5,6
1 Department of Pathology, Faculty of Medicine, Kuwait University, , Kuwait City 13110, Kuwait
2 Paediatric Department, Al-Jahra Hospital, , Al-Jahra 00042, Kuwait
3 Department of Neuromuscular Diseases, University College London Queen Square Institute of Neurology, London, UK.
4 NHS Highly Specialised Service for Rare Mitochondrial Disorders, Queen Square Centre for Neuromuscular Diseases, The National Hospital for Neurology and Neurosurgery, London, UK.
5 Department of Neuromuscular Diseases, University College London Queen Square Institute of Neurology, London, UK.
6 NHS Highly Specialised Service for Rare Mitochondrial Disorders, Queen Square Centre for Neuromuscular Diseases, The National Hospital for Neurology and Neurosurgery, London, UK.
Introduction: Primary pyruvate dehydrogenase complex deficiency (PDCD) is a genetic mitochondrial disorder characterized by lactic acidosis, progressive neurological and neuromuscular degeneration, and a high risk of early mortality, including sudden unexpected infant death (SUID).
Methods: A consanguineous multiplex family with two affected male siblings suspected of PDCD was recruited for genetic investigation. Genomic DNA was isolated from affected family members using standard procedures. Whole-exome sequencing (WES) was performed on DNA samples from the two affected individuals. Clinical evaluations included biochemical analyses and neuroimaging studies.
Results: Both affected siblings presented with neonatal lactic acidosis, Leigh syndrome, and brain abnormalities on MRI, followed by later manifestations including adult-onset myopathy and ataxia. Linkage analysis identified a small shared identical-by-descent (IBD) interval on chromosome 4, and exome data revealed a novel homozygous missense variant in DCLK2 (c.101G>A; p.Ser34Asn). However, given the clinical diagnosis of PDCD and the occurrence of disease in two male siblings, an X-linked etiology was strongly suspected. Subsequent analysis of the X chromosome identified a novel hemizygous missense variant in exon 6 of PDHA1 (c.754C>G; p.Leu252Val; rs1555934383), consistent with X-linked inheritance.
Conclusions: We report a novel hemizygous missense variant in PDHA1 and a novel homozygous missense variant in DCLK2 identified in two affected male siblings from a consanguineous family of Arabian origin. Our findings suggest that DCLK2 modulates the severity of PDCD. These results expand the mutational spectrum associated with PDCD and highlight the importance of comprehensive genomic analysis for X-linked disorders in genetically complex families.
042Systematic prioritisation of noncoding regulatory variants in unresolved adult-onset mitochondrial disease
Eszter S Arany1,2,3, Hannah Maude1, Matthew Doyle1, William Macken2,4, Tamara Hill2,4, Renata Kabiljo2,4, Robert D S Pitceathly2,4, Inês Cebola1
1Section of Genetics and Genomics, Department of Metabolism, Digestion and Reproduction, Imperial College London
2University College London, Department of Neuromuscular Disease
3Imperial College Healthcare NHS Trust
4University College London Hospitals NHS Foundation Trust
eszter.arany@nhs.net
Background: Despite advances in genomic diagnostics, a substantial proportion of adults with clinically confirmed mitochondrial disease remain genetically unresolved. This suggests that routinely used pipelines that focus on coding variants of genes may not capture all disease-contributing variants.
Aims: To systematically investigate noncoding regulatory variation as a potential contributor to unexplained adult-onset mitochondrial disease, and to develop an integrated computational framework for prioritising candidate regulatory variants with plausible functional impact.
Methods / Materials: Our cohort included four probands with long-read (PacBio) and one additional patient with short-read whole-genome sequencing (WGS) data, each harbouring a previously identified heterozygous pathogenic variant. All individuals demonstrated Chronic Progressive External Ophthalmoplegia, ragged-red fibres, and multiple mtDNA deletions. Rare coding and splicing variants were prioritised using established pathogenicity filters. We extended the analysis to noncoding regions, interrogating single-nucleotide variants and structural variants overlapping experimentally supported cis-regulatory elements (CREs). Regulatory annotations were derived from ENCODE (SCREEN) and GeneHancer databases. Candidate variants were prioritised based on chromatin accessibility (ATAC-seq/DNase), histone marks (H3K27ac), transcription factor binding density, motif disruption analyses, and phasing relative to known pathogenic coding variants where possible.
Results: We identified rare candidate non-coding single nucleotide variants in trans with known pathogenic coding variants that overlapped regulatory elements with evidence of chromatin accessibility and transcription factor occupancy. Computational modelling predicted disruption of important transcription factor binding sites, including factors implicated in the regulation of nuclear-encoded mitochondrial genes. Structural variants overlapping regulatory elements were also identified and prioritised. These findings suggest plausible regulatory mechanisms arising from non-coding regions that may contribute to reduced gene expression or altered transcriptional control in the absence of biallelic pathogenic variants in protein coding regions.
Conclusion: Our results demonstrate that systematic regulatory variant analysis could reveal candidate pathogenic mechanisms in cases unresolved by conventional coding and splicing variant analysis. This integrative framework highlights the potential contribution of noncoding regulatory variation to mitochondrial disease and supports the inclusion of regulatory region analysis in future diagnostic and research pipelines. These findings are based on integrative computational analyses and require experimental validation to confirm the variants’ functional effects.
043A Systematic Review of In Vitro Neuronal Models of Mitochondrial Diseases
1School of Clinical Medicine, University of Cambridge, Cambridge, UK
2Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK
*NAB and MZM contributed equally to this work
nihal.basha@esneft.nhs.uk
Background: Primary Mitochondrial Diseases (PMDs) can be considered disorders of the nuclear or mitochondrial genome that perturb the structure or function of mitochondria, particularly oxidative phosphorylation. The metabolically active nature of neuronal cells and their reliance on oxidative phosphorylation renders them vulnerable and PMDs often have severe neurological features. Brain tissue is typically only available post-mortem and animal models are hampered by difficulty altering mtDNA and unreliable recapitulation of phenotypes.
Aims: Our study aimed to provide a comprehensive overview of the available in vitro neuronal models of PMDs, key findings obtained through their use, and their relative strengths and limitations.
Methods/Materials: PubMed was searched on 17/01/2023. Peer reviewed, primary research investigating a specific PMD or strongly linked mutation principally affecting mitochondrial structure or function, through one or more in vitro models of human neurons and glia or their precursors, neuroblastoma cells, or inner-ear hair cells were included. Papers not in English, retracted, or exclusively using RGCs or retinal/optic nerve tissue were excluded. Blinded inter-rater agreement on 102 records was 100% and the remaining were divided between the two.
Results: The search yielded 1936 records. 2 duplicates and 3 retractions were removed. 58 passed title/abstract screening and a further 8 were removed on full text screening. The three main categories of models were neuroblastoma and related cell lines (SH-SY5Y, SK-N-AS, and NT2); induced-pluripotent stem cell (iPSC) derived neuronal progenitors, neurons (dopaminergic, excitatory, motor, and spinal), inner-ear hair cells, and spinal and forebrain organoids; and cybrids. Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) was the most frequently modelled PMD and evidenced unique models like blood-brain barrier and olfactory neurospheres.
Conclusion: Neuroblastoma cell lines enable high throughput screening, but nuclear-mitochondrial genome interactions are typically lost. iPSCs overcome this but lack isogenic controls and face possible heteroplasmy shifts and mutations after reprogramming. Cerebral organoids enable more detailed investigation but lack vascularisation and immune elements. They all also have only limited developmental age. Interestingly, all models of neuronal pathology observed issues with maturation and differentiation. These emerging techniques represent an exciting frontier that will hopefully facilitate greater understanding of PMDs in years to come.
044‡Amino acid supplementation during early development of ears2-/- zebrafish to treat mitochondrial translation deficiencies
Emily Cross1, Oliver Podmanicky1, Benjamin Munro1, Rita Horvath1
1Department of Clinical Neuroscience, University of Cambridge
Ec912@cam.ac.uk
Background: Mitochondrial translation deficiencies refer to a group of inherited defects affecting the formation of mitochondrial proteins, which can lead to impaired activity of the mitochondrial complexes. These disorders can be caused by a variety of mutations in either the mitochondrial DNA (mtDNA) or nuclear DNA (nDNA). Of the deficiencies affecting the nuclear DNA, mitochondrial aminoacyl-tRNA synthetase (mt-ARS) disorders are highly prevalent and affect the formation of mitochondrial tRNAs required for protein translation. A subset of this group, the EARS2 gene encodes for the glutamyl-tRNA synthetase. Variants in EARS2 have been identified to cause a variety of symptoms including leukoencephalopathy and psychomotor developmental delay.
Previous work in patients with mutations in tRNA synthetases suggested that supplementation with cognate amino acid may have a beneficial effect on the phenotype. It has been also hypothesized that cysteine may improve mitochondrial translation in reversible infantile diseases. Based on these data, we decided to use zebrafish as a model to test amino acid supplementation in vivo.
Aims: To assess the effects of amino acid supplementation (glutamine or cysteine) in developing zebrafish embryos with the ears2-/- mutation.
Methods: A ears2+/- zebrafish line was generated using CRISPR/Cas9 and was used to produce ears-/- embryos. Eggs were dechorionated and treated with amino acid-supplemented E3 media on Day 1 and maintained at 28°C and pH 7-7.4. Fish media was changed daily until Day 4, when embryos were assessed and euthanized with MS-222 and frozen at -20C. Embryo remains were stored long-term at -80°C for further analysis.
Results:Ears2-/- zebrafish embryos exhibit a distinct phenotype compared to wild-types, featuring deflated swim bladders and shorter length. Wild-type zebrafish tolerate both N-acetyl-cysteine (NAC) and glutamine well at 5 mM concentrations. Amino acid supplementation of NAC, but not glutamine appears to have a beneficial effect on the development of ears2-/- zebrafish, partially restoring their phenotype including size and swimming ability.
Conclusions: This work highlights the benefit of cysteine supplementation in early development in zebrafish with the ears2-/- mutation, identifying a potential novel therapeutic for patients with impaired EARS2 function.
045Resistance exercise training and ubiquinone treatment reveal patient-specific molecular signatures in mitochondrial myopathy
Valeria Di Leo1,2, Jane Newman1,3, Tiago B Gomes1,2,3,4, Conor Lawless1, Charlotte Warren1,2, Atif Khan1,2, Jordan Child1, George O. T. Merces5,6, Fiona Robertson1, Yotam Levy9, Millicent Rice1, Julien Ochala10, Chun Chen9, Michael F. Marusich10, Sarah J. Pickett9, Gavin Hudson2,9, Helen A.L. Tuppen1, Amy E. Vincent1,2,4,* & Oliver M. Russell1,2
1Mitochondrial Research Group, Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK;
2NIHR Newcastle Biomedical Research Centre, Newcastle upon Tyne Hospitals NHS, Foundation Trust and Newcastle University, Newcastle upon Tyne, UK;
3NHS Highly Specialised Service for Rare Mitochondrial Disorders, Royal Victoria Infirmary, Newcastle upon Tyne, UK;
4John Walton Muscular Dystrophy Research Centre, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK.
5Biosciences Institute, Innovation, Methodology and Application (IMA) research Theme, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK;
6Image Analysis Unit, Faculty of Medical Sciences, Newcastle University, upon Tyne, UK;
7Entrepreneurship Institute, King’s College London, London, UK;
8Exercise Laboratories, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark;
9Mitochondrial Research Group, Biosciences Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK.
10mAbDx, Inc., Eugene, OR, United States
valeria.di-leo@newcastle.ac.uk
Introduction: Mitochondrial myopathy, a hallmark of mitochondrial disorders, causes progressive skeletal muscle (SKM) weakness driven by mitochondrial dysfunction. Although resistance exercise training benefits affected individuals, the molecular mechanisms underlying these adaptations remain incompletely understood.
Aims: This study examined SKM biopsies from participants with single, large-scale mitochondrial DNA deletions collected before and after a 16-week resistance exercise training intervention. Two participants in the mitochondrial myopathy exercise cohort (n=5) received adjunct ubiquinone treatment.
Methods/Materials: Functional performance was assessed pre- and post-intervention. Fibre type distribution, size, and contractile force were assessed using Myh7, Myh2, and Myh1 staining. Mitochondrial DNA heteroplasmy was quantified by real-time PCR targeting MT-ND1 and MT-ND4. Mitochondrial respiratory chain content was assessed using imaging mass cytometry for dystrophin, VDAC1, NDUFB8 and ND4 (complex I), SDHA (complex II), CytB (complex III), CO1 and CO4 (complex IV), ATPB (complex V), and mtTFA as a marker of mitochondrial biogenesis. Differential gene expression and corresponding protein abundance were evaluated using RNA sequencing and imaging mass cytometry, respectively.
Results: Three participants were classified as responders based on improvements in concentric and eccentric peak torque. Fibre hypertrophy of either type I or type2a/2x, or both, was observed in all participants. Fibre type distribution changed after exercise inconsistently. Partial restoration of oxidative phosphorylation capacity was consistently improved in complexes II and IV across participants and associated with disease severity. Transcriptomic analyses revealed marked inter-individual variability, including differential responses to adjunct ubiquinone supplementation, with enrichment of pathways related to extracellular matrix remodelling, satellite cell activation, and metabolic reprogramming. Proteomic analyses confirmed increased abundance of selected transcriptomics-derived targets associated with hypertrophy-related signalling pathways, with more pronounced adaptations in participants classified as responders. Classical mitochondrial markers showed minimal responsiveness to exercise; instead, Stearoyl-CoA desaturase and Nicotinamide N-methyltransferase emerged as candidate biomarkers of resistance exercise–mediated therapeutic efficacy.
Conclusion: These findings highlight the heterogeneous molecular responses to resistance exercise training in mitochondrial myopathy, reflecting both disease severity and inter-individual variability in adaptive capacity. Collectively, they support a precision-medicine approach to exercise-based therapies and emphasise the need for robust biomarkers to stratify patients and monitor therapeutic responses in future clinical studies.
046MT-ATP8, a mitochondrial disease gene?
Carl Fratter1, Conrad Smith1, Philip Hodsdon1, Stefen Brady2, Louisa Kent3, Loukia Stavrakaki-Kallergi4, Mary O’Driscoll5, Philip Campbell5, Amitav Parida6, Saiju Jacob7, Emma Rees5, Langping He8, Robert W Taylor8,9, Amanda Lam10, Padraig J Flannery10,11, Victoria Nesbitt3, Kate Sergeant1
1Oxford Genetics Laboratories, Oxford University Hospitals NHS Foundation Trust, Oxford
2Oxford Muscle Service, Oxford University Hospitals NHS Foundation Trust, Oxford
3NHS Highly Specialised Services for Rare Mitochondrial Disorders, Oxford University Hospitals NHS Foundation Trust, Oxford
4Department of Paediatric Neurology, University Hospitals Bristol NHS Foundation Trust, Bristol
5West Midlands Regional Clinical Genetics Service, Birmingham Women's and Children’s NHS Foundation Trust, Birmingham
6Department of Paediatric Neurology, Birmingham Women's and Children’s NHS Foundation Trust, Birmingham
7Department of Neurology, University Hospitals Birmingham NHS Foundation Trust, Birmingham
8NHS Highly Specialised Service for Rare Mitochondrial Disorders, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne
9Mitochondrial Research Group, Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne
10Neurometabolic Unit, National Hospital for Neurology and Neurosurgery, London
11Neurogenetics Unit, Rare and Inherited Disease Laboratory, North Thames Genomic Laboratory Hub, London
carl.fratter@ouh.nhs.uk
Background: Assigning pathogenicity to genetic variants detected through clinical diagnostic testing is challenging and particularly so for mitochondrial DNA (mtDNA) variants since the available guidelines for variant classification are mostly aimed at interpreting variants in nuclear genes. Moreover, many mtDNA variants previously considered to be the cause of rare mitochondrial disease do not fulfil the more stringent criteria to enable classification as pathogenic or likely pathogenic by current standards. The MT-ATP8 gene, which partially overlaps MT-ATP6, encodes a small 68 amino acid subunit of ATP synthase (subunit 8 within the membrane F0 domain), which is essential for generation of ATP via oxidative phosphorylation. However, no variants that are only located in MT-ATP8 have been classed as pathogenic or likely pathogenic by the ClinGen Mitochondrial Variant Curation Expert Panel and the ClinGen Gene-Disease Validity classification for MT-ATP8 states that there is currently only limited evidence to support a gene-disease relationship.
Aims: We aimed to review the evidence for and against pathogenicity of MT-ATP8 variants detected in a cohort of individuals that underwent genomic analysis for suspected mitochondrial disease, to further evaluate the role of MT-ATP8 as a mitochondrial disease gene.
Methods/Materials: Mitochondrial genome sequencing and/or whole genome sequencing data was reviewed to identify individuals with variants in MT-ATP8. Evaluation of phenotype, family studies, heteroplasmy assessment in available tissues, and review of other available evidence was undertaken to assess pathogenicity of the detected MT-ATP8 variants.
Results: Four cases with predicted loss of function variants in MT-ATP8 were identified. Available evidence at the time of reporting the four variants only allowed classification as variants of uncertain significance. Additional studies, in combination with recent reports in the literature, indicated that the MT-ATP8 variants may be responsible for the individuals’ disease symptoms in at least two of the four cases, and have enabled re-evaluation of pathogenicity classification.
Conclusion: The cases presented, plus recent reports in the literature, provide further evidence to support MT-ATP8 as a causative mitochondrial disease gene.
047Quantifying longitudinal change in skeletal muscle OXPHOS status in mitochondrial myopathy
T. Bernardino Gomes1,2,3, V. Di Leo1,4, C. Warren1,4, A. Khan1,5, I. Barrow4, G. Hudson6, D. M. Turnbull1,2, C. Lawless1,4, A. E. Vincent1,3,4
1Mitochondrial Research Group, Translational and Clinical Research Institute, Newcastle University, UK
2NHS Highly Specialised Service for Rare Mitochondrial Disorders of Adults and Children, Directorate of Neurosciences, Newcastle upon Tyne Hospitals NHS Foundation Trust, UK
3John Walton Muscular Dystrophy Research Centre, Newcastle University, Centre for Life, Newcastle upon Tyne, UK
4NIHR Biomedical Research Centre, Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK
5Centre for Doctoral Training in Cloud Computing and Big Data, Newcastle University, UK
6Mitochondrial Research Group, Biosciences Institute, Newcastle University, Newcastle upon Tyne, UK
tiago.gomes@newcastle.ac.uk
Background: Mitochondrial myopathy is a prevalent and disabling feature of mitochondrial disease. While skeletal muscle oxidative phosphorylation (OXPHOS) markers are increasingly used as quantitative endpoints for longitudinal studies, their ability to quantify disease progression remains poorly defined. We previously characterised the cross-sectional intra-individual variability of OXPHOS protein markers using antibody-based imaging (PMID:41419999). However, the absence of systematic longitudinal natural history data on such markers still complicates their interpretability in clinical research.
Aims: To evaluate the performance of antibody-based protein imaging in capturing longitudinal shifts in muscle OXPHOS deficiency in mitochondrial myopathy.
Methods/Materials: We utilised imaging mass cytometry to quantify proportions of myofibres deficient in commonly used protein markers for OXPHOS complexes I (CI), CIII, CIV and CV. Repeat muscle biopsies from a genetically confirmed mitochondrial myopathy cohort were analysed using our previously established workflows. Rates of change were derived using per-patient bootstrapped differences between timepoints.
Results: Rates of change in proportions of OXPHOS deficiency were generally small and heterogeneous, including over time increases, stability, and even reductions. CI, CIV and CV deficiencies typically shifted in tandem, preserving baseline inter-marker relationships. While CI and CIV were the most common deficits, with a predominant combined CI-IV-V deficiency profile, CIII deficiency was rare and the most variable. The magnitude and direction of these changes were patient-specific and showed no clear relationship with major genotype or phenotype grouping, demographics, or biopsy-related factors in this analysis.
Conclusion: Antibody-based protein imaging reliably captures differences in muscle OXPHOS deficiency between timepoint biopsies. The observed low overall rates of change align with the slowly progressive nature of mitochondrial myopathy. While inter-individual variability reflects the known genotype-phenotype heterogeneity of these diseases, the declining deficiency in some patients was an unexpected finding in a progressive condition. Such patterns likely reflect combined compensatory responses to endogenous or exogenous stimuli (e.g., exercise) not addressed by this study, while sampling and assay-specific factors cannot be fully excluded. These findings provide critical insights into the natural progression of OXPHOS deficiency and suggest that to use OXPHOS protein markers as reliable endpoints, studies must incorporate extended follow-up periods or target interventions with large effect sizes.
048Validation of Targeted Mitochondrial Proteomics for ISO15189:2022 in an NHS Diagnostic Laboratory
Robert Winter1, Kathryn Oprych2, William Macken2, Robert D S Pitceathly2, Wendy Heywood1,4 Francesca Robertson1,4 Mike Hanna2 Simon Heales1,3, Amanda Lam1,2
1 Neurometabolic Unit, Queen Square Division, University College London Hospitals, London, UK
2 Department of Neuromuscular Diseases, University College London Queen Square Institute of Neurology, London, UK
3 Department of Genetics and Genomics Medicine, University College London Great Ormond Street Institute of Child Health, London, UK
4 Guilford Street Laboratories, London, UK
E-mail of presenting author: amanda.lam@nhs.net
Background: The Neurometabolic Unit is a Highly Specialised NHS diagnostic laboratory providing UKAS-accredited testing for mitochondrial respiratory chain enzyme function, commissioned by the NHS Rare Mitochondrial Disorders Service. While omics-based approaches such as untargeted proteomics and metabolomics offer powerful tools for mitochondrial diagnostics in research settings, their reliance on complex data processing and relative quantitation poses significant challenges for translation into routine clinical practice. In particular, demonstrating compliance with ISO15189:2022 and achieving UKAS accreditation for these methods remains a major barrier.
Aims: This study aims to develop a validation workflow for targeted mitochondrial proteomics that will ultimately meet ISO15189:2022 requirements. As an initial step, we focused on identifying optimal peptides and transitions to ensure analyte specificity within complex biological matrices. Using the mitochondrial complex I subunit NDUFV1 as a model protein, we evaluated predicted peptides for suitability in human muscle homogenate, thereby facilitating the integration of proteomics into an NHS diagnostic service.
Methods: Candidate peptides specific to NDUFV1 were identified using open-source tools including UniProt BLAST, PeptideAtlas and Skyline. LC-MS analysis was performed on a Waters Xevo TQ-XS mass spectrometer with a Cortecs C18 column. Method validation employed remnant human muscle homogenate obtained with consent for diagnostic mitochondrial studies. Objective evidence required for UKAS accreditation was generated by confirming that predicted peptides and transitions provided adequate signal for reliable quantitation.
Results: Using recombinant human NDUFV1 protein, we demonstrated robust signal for three peptides (YLVVNADEGEPGTCK, GPDWILGEIK, and LLEGCLVGGR). These same peptides were subsequently detected in human muscle homogenate, validating their specificity and suitability for further analytical validation.
Conclusion: In conclusion, this work establishes a critical foundation for translating mitochondrial proteomics into an accredited NHS laboratory setting. By validating peptide selection for NDUFV1, we have now identified a pathway towards ISO15189:2022 compliance and UKAS accreditation for targeted mitochondrial proteomics methods. This approach represents an important step toward expanding diagnostic capabilities for rare mitochondrial disorders within the NHS.
049(Protocol) Assessing organ-specific non-invasive biomarkers in mitochondrial disease
Shamini Saravanabavan1, Rita Horvath1, Jelle van den Ameele1, William Watson2, Heather Biggs1, Patrick Yu Wai Man1, Patrick Chinnery1
1 Department of Clinical Neurosciences, University of Cambridge, UK
2 Department of Interventional Cardiology, Papworth Hospital, Cambridge, UK
ss2698@cam.ac.uk
Background: Primary mitochondrial diseases (PMD) are among the most common inherited neurological disorders. Their clinical heterogeneity and slowly progressive course have limited the development of reliable, non-invasive biomarkers to monitor disease progression or evaluate therapeutic response. Current gold-standard assessments often rely on invasive tissue biopsies, underlining the need for validated in vivo biomarkers of mitochondrial dysfunction.
Aims: This study aims to develop and validate a portfolio of non-invasive, organ-specific imaging biomarkers of mitochondrial dysfunction in the brain and heart. We will evaluate longitudinal change in these measures over 24 months and assess associations with clinical scales, circulating biomarkers, and participant-reported outcome measures (PROMs).
Methods: We developed a protocol for a prospective, longitudinal, observational study enrolling 20 participants with genetically confirmed PMD. Participants will be recruited into parallel neurological (n=10) and cardiac (n=10) arms, with optional overlap, and followed over 24 months. Neurological imaging will use deuterium metabolic imaging (DMI) at 7 Tesla after oral administration of deuterated glucose to quantify regional cerebral glucose metabolism and downstream metabolites. Cardiac assessments will combine cine cardiac magnetic resonance imaging (MRI) and phosphorus-31 magnetic resonance spectroscopy (31P-MRS) at 3 Tesla to measure ventricular volumes, ejection fraction, myocardial wall thickness, strain-derived contractility, myocardial fibrosis (T1 mapping and late gadolinium enhancement), and myocardial energetics (phosphocreatine-to-adenosine triphosphate ratio and creatine kinase flux). Multimodal clinical assessments (e.g. NMDAS, MOCA & SARA, serial blood biomarkers (including markers of mitochondrial function and immune activity), and validated PROMs capturing fatigue, pain, quality of life, global severity, and most bothersome symptoms will be collected.
Results: Neurological outcomes include feasibility, reproducibility, and longitudinal sensitivity of DMI-derived measures of cerebral glucose metabolism and downstream metabolites (lactate and glutamate/glutamine) as biomarkers of neurological disease severity. Cardiac outcomes include longitudinal changes in myocardial energetics, structure, and global function, assessed using cine MRI and 31P-MRS (including phosphocreatine-to-ATP ratio). Blood-based molecular biomarkers will be correlated with imaging measures and clinical phenotypes.
Conclusions: This study will establish the feasibility and longitudinal utility of advanced metabolic MRI and spectroscopy as non-invasive biomarkers of mitochondrial dysfunction in PMD, supporting future clinical trials through objective monitoring of disease progression and treatment response.
050‡De novo truncation variants in XRN1 cause a novel dominant form of lethal infantile mitochondrial cardiomyopathy
Lucie S. Taylor1,2, Liana N. Semcesen3, Leah E. Frajman4,5, Marisa W. Friederich6,7, Andrew M. Frey1,8, Stefan J. Siira9,10, Daniella H. Hock3,4,5, Tegan Stait4, Sila Hopton2, Yoshihito Kishita11,12, Queenie K-G. Tan13, Vandana Shashi13, Muriel Holder-Espinasse14, Hugh Lemonde15, Kay Metcalfe16, Lisette Curnow5, Rebecca C. Spillman13, Kelly Schoch13, Karen Stals17, Agata Oliwa1, MitoMDT Diagnostic Network for Genomics and Omics, Undiagnosed Diseases Network, Matthias Trost8, Kei Murayama11,18, Yasushi Okazaki11,19, Akira Ohtake20, Aleksandra Filipovska9,10, Charlotte L. Alston1,2, John Christodoulou4,5,21, David R. Thorburn4,5,21, Johan L.K. Van Hove6,7, Zornitza Stark5,21,22, David A. Stroud3,4,5, Alison G. Compton4,5,21, Robert W. Taylor1,2
1 Mitochondrial Research Group, Newcastle University, Newcastle upon Tyne, UK
2 NHS Highly Specialised Mitochondrial Diagnostic Laboratory, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK
3 The University of Melbourne, Parkville, Australia
4 Murdoch Childrens Research Institute, Melbourne, Australia
5 Victorian Clinical Genetic Services, Melbourne, Australia
6 Department of Pediatrics, University of Colorado, Aurora, USA
7 Mitochondrial Laboratory, Childrens Hospital Colorado, Aurora, USA
8 Biosciences Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK
9 ARC Centre of Excellence in Synthetic Biology, University of Western Australia, Crawley, Australia
10 Perth Children’s Hospital, Nedlands, Australia
11 Intractable Disease Research Center, Juntendo University, Tokyo, Japan
12 Kindai University, Osaka, Japan
13 Duke University School of Medicine, North Carolina, USA
14 Guy’s Hospital, London, UK
15 Evelina London Children’s Hospital, London, UK
16 St. Mary’s Hospital, Manchester, UK
17 Royal Devon and Exeter NHS Foundation Trust, Exeter, UK
18 Chiba Children's Hospital, Chiba, Japan
19 RIKEN Center for Integrative Medical Sciences, Kanagawa, Japan
20 Saitama Medical University, Saitama, Japan
21 Department of Paediatrics, University of Melbourne, Melbourne, Australia
22 Australian Genomics, Melbourne, Australia
lucie.taylor@newcastle.ac.uk
Background: XRN1 is a highly conserved cytoplasmic 5’-3’ exoribonuclease involved in mRNA decay, quality control, gene silencing, rRNA maturation, transcription termination and viral infection responses. Here we characterise an association between XRN1 genetic variants and human disease in a cohort of infants with a lethal infantile form of mitochondrial cardiomyopathy.
Aims: We present heterozygous de novo XRN1 variants as the pathological origin of lethal cardiomyopathy in a cohort of infants, despite no prior human disease association reported. Previous complete knockout of XRN1 in multicellular organisms has proven lethal, most likely due to its central role in mRNA homeostasis, however, whilst XRN1 has no established mitochondrial function in mammals, both clinical and molecular findings across the cohort were compellingly consistent with mitochondrial disease.
Methods/Materials: Using GeneMatcher (https://genematcher.org/), we collated a cohort of individuals with similar heterozygous de novo truncation variants clustering in the C-terminal region of XRN1, each predicted to evade nonsense-mediated mRNA decay. Individuals were identified independently through clinical and research-based whole genome sequencing (WGS) strategies. Functional studies, including quantitative proteomics were performed using patient-derived tissues and cell lines.
Results: Seven individuals from six unrelated families presented similarly with lethal perinatal cardiomyopathy, global developmental delay, moderate bilateral sensorineural hearing loss and failure to thrive following an acute viral infection. WGS identified unique de novo XRN1 variants resulting in a near-identical XRN1 C-terminal sequence predicted to alter a characterised binding domain that interacts with the mRNA decapping enhancer EDC4. Quantitative proteomics conducted on patient-derived skeletal muscle revealed consistent mitochondrial disease phenotypes, characterised by a combined abundance defect in oxidative phosphorylation (OXPHOS) complexes. These findings were supported by the demonstration of impaired OXPHOS assembly assessed by Blue-Native PAGE, marked histochemical defects and decreased respiratory chain enzyme activities in clinically-relevant tissues of affected individuals. RNA-sequencing of patient skeletal muscle additionally determined an increase in mitochondrial genome transcription, indicating a response to mitochondrial stress.
Conclusion: We propose XRN1 as a candidate gene associated with a novel form of autosomal dominant lethal perinatal mitochondrial cardiomyopathy. Functional studies on patient-derived cells are continuing to provide insight into the precise disease mechanism.
1Department of Neurology, Derriford Hospital, University Hospitals Plymouth NHS Foundation Trust
2 Department of Neurology, Derriford Hospital, University Hospitals Plymouth NHS Foundation Trust
zeeshan.yousuf@nhs.net
Background: Chronic progressive external ophthalmoplegia (CPEO) is a mitochondrial DNA maintenance disorder characterized by slowly progressive ptosis and ophthalmoplegia. Its diagnosis can be challenging due to overlap with neuromuscular, autoimmune, and structural causes of ocular dysfunction.
Case Presentation: A 73-year-old male presented with a 20-year history of diplopia and progressive bilateral ptosis over five years. Despite repeated spectacle adjustments, prism correction reached its limit. He had undergone ptosis surgery five years prior. Brain MRI in 2022 identified a pituitary macroadenoma compressing the optic chiasm, though this did not explain his chronic ophthalmoplegia. Acetylcholine receptor and MUSK antibodies were negative. His mother had late onset ptosis without diplopia. Examination revealed mild bilateral ptosis with otherwise normal neurological findings. Further investigations including CT angiography, nerve conduction studies, EMG, and mitochondrial myopathy screening were normal.
Results: Genetic testing revealed an autosomal dominant TWNK related mitochondrial DNA maintenance disorder, confirming the diagnosis of chronic progressive external ophthalmoplegia (CPEO).
Conclusion: This case highlights the diagnostic complexity of long-standing ptosis and diplopia in older adults and emphasizes the importance of genetic testing when routine investigations are inconclusive. Recognition of TWNK-related CPEO is essential for accurate diagnosis, counselling, and management.
052‡Modelling POLG mutations in mice unravels a critical role of POLγΒ in regulating phenotypic severity
Carlo Viscomi1,2, Alessandro Zuppardo1, Samantha Corrà1, Raffaele Cerutti1, Christian Santangeli1,3, Francesco Chemello1,3
1 Veneto Institute of Molecular Medicine (VIMM), Padova, Italy
2 Department of Biosciences, University of Milan, Milano, Italy
3 Department of Pharmacy and Biotechnology (FaBiT), University of Bologna, Bologna, Italy
carlo.viscomi@unimi.it
Background: Mutations in the POLG gene, encoding the catalytic subunit of mitochondrial DNA polymerase γ (POLγ), are the most common cause of inherited mitochondrial disease due to defects in a single nuclear gene. These variants impair mtDNA replication and maintenance, leading to mtDNA depletion, or deletions respiratory chain dysfunction, and progressive multisystem disease. However, robust in vivo models and mutation-specific mechanistic and therapeutic insights remain limited.
Aims: Here, we aimed to (i) generate and characterize murine models carrying the most common recessive POLG mutations (A467T, W748S, G848S) and the major dominant mutation (Y955C), (ii) define the molecular, structural, and tissue-specific consequences of these variants, and (iii) establish a platform for developing precise gene-editing strategies to correct the dominant Y955C and recessive A467T mutations, with the goal of identifying efficient and translatable therapeutic approaches.
Methods/Materials: We generated homozygous knock-in mice, compound heterozygous mice carrying one mutant and one Polg null allele, and compound heterozygous mice combining different pathogenic variants. mtDNA content and integrity and respiratory chain activities were assessed in high-energy–demand tissues, alongside histological, histochemical, and ultrastructural analyses of brain, liver, and skeletal muscle.
For therapeutic development, we designed base-editing strategies targeting Y955C (c.2864A>G) using cytosine base editors and A467T (c.1399G>A) using adenine base editors.
Results: Of nine allelic combinations, four were embryonically lethal, mirroring clinical severity. The remaining viable models enabled detailed disease analysis. Polg+/Y955C mice showed normal functional performance and preserved mtDNA, but developed late-onset pathological changes in brain, liver, and skeletal muscle, including white matter vacuolization, gliosis, and mitochondrial abnormalities. Structural and in vitro analyses revealed mutation-specific effects on polymerase activity and species-specific differences between human and murine POLγ. Base-editing screens identified efficient and precise mutation-correcting strategies.
Conclusion: This work provides mechanistic insight into POLG-related disease, establishes robust preclinical models, and supports precise base editing as a promising therapeutic approach for POLG-associated mitochondrial disorders.
Peripheral Neuropathy
053Loss of ARHGAP19 function disrupts RhoA regulation in Charcot-Marie-Tooth disease: mechanisms and therapeutic targets
NataliaDominik1#, Stephanie Efthymiou1#§, Christopher J. Record1, Xinyu Miao2,51, Renee Lin1, Jevin M. Parmar3, Annarita Scardamaglia1, Reza Maroofian1, Gabriel Aughey4, Abigail D. Wilson4, Simon Lowe4, Riccardo Curro1,5, Ricardo P. Schnekenberg1 , Shahryar Alavi1, Leif Leclaire2,51, Yi He2,51, Kristina Zhelcheska1, Yohanns Bellaiche6, Isabelle Gaugué6, Mariola Skorupinska1, Liedewei Van de Vondel7,8, Sahar I. Da'as9, Valentina Turchetti1, Serdal Güngör10, Gavin Monahan3, Ehsan Ghayoor Karimiani,1,11, Yalda Jamshidi11, Phillipa J. Lamont12, Camila Armirola Ricaurte13,14, Haluk Topaloglu15, Albena Jordanova13,14,16, Mashaya Zaman17, Selina H. Banu17, Wilson Marques18, Pedro José Tomaselli19, Busra Aynekin1, Ali Cansu20, Huseyin Per21, Ayten Güleç21, Javeria Raza Alvi22, Tipu Sultan22, Arif Khan23,24, Giovanni Zifarelli25, Shahnaz Ibrahim26, Grazia M. S. Mancini27, M. Mahdi Motazacker28, Esther Brusse27, Vincenzo Lupo29, Teresa Sevilla30,31, A. Nazlı Başak32, Seyma Tekgul32, Robin J. Palvadeau32, Jonathan Baets7,8,33, Yesim Parman34, Arman Çakar34, Rita Horvath35,36, Tobias B. Haack37,38, Jan-Hendrik Stahl39,40, Kathrin Grundmann-Hauser37,38, Joohyun Park37,38, Stephan Züchner41,42, Nigel G. Laing3, Lindsay Wilson1, Alexander M. Rossor43, James Polke44, Fernanda Barbosa Figueiredo45, André Luiz Pessoa46, Fernando Kok45, Antônio Rodrigues Coimbra-Neto47, Marcondes C. França Jr47, Gianina Ravenscroft3, Sherifa Ahmed Hamed48, Wendy K. Chung49, Daniel P. Osborn50, Michael Hanna1, Andrea Cortese1,5, Mary M. Reilly1#, James E. C. Jepson4#, Nathalie Lamarche-Vane2,51#, Henry Houlden1#
1. Department of Neuromuscular Disease, UCL Queen Square Institute of Neurology, London, WC1N 3BG, UK.
2. Department of Anatomy and Cell Biology, McGill University, Montréal, QC H3A 0C7, Canada.
3. Harry Perkins Institute of Medical Research, Centre for Medical Research, University of Western Australia, Perth, Western Australia, Australia.
4. Department of Clinical and Experimental Epilepsy, UCL Queen Square Institute of Neurology, London, UK.
5. Department of Brain and Behavioral Sciences, University of Pavia, 27100 Pavia, Italy
6. Institut Curie, Université PSL, Sorbonne Université, CNRS UMR3215, INSERM U934, Genetics and Developmental Biology, 75005 Paris, France.
7. Translational Neurosciences, Faculty of Medicine and Health Sciences, University of Antwerp, Antwerp, Belgium.
8. Laboratory of Neuromuscular Pathology, Institute Born-Bunge, University of Antwerp, Antwerp, Belgium.
9. Department of Human Genetics, Sidra Medicine, Doha, Qatar.
10. Inonu University, Faculty of Medicine, Turgut Ozal Research Center, Department of Pediatric Neurology, Malatya, Turkey.
11. Molecular and Clinical Sciences Institute, St. George's, University of London, SW17 0RE, UK.
12. Royal Perth Hospital, Perth, WA, 6000, Australia
13. Molecular Neurogenomics group, VIB Center for Molecular Neurology, VIB, Antwerp, Belgium.
14. Molecular Neurogenomics group, Department of Biomedical Sciences, University of Antwerp, Antwerp, Belgium.
15. Department of Pediatric Neurology, Hacettepe University, Ankara, Turkey.
16. Department of Medical Chemistry and Biochemistry, Medical University-Sofia, 1431, Sofia, Bulgaria.
17. Department of Pediatric Neurology, Dr. M.R. Khan Shishu (Children) Hospital and ICH, Mirpur, Dhaka, Bangladesh.
18. Department of Neurosciences, School of Medicine of Ribeirão Preto, University of São Paulo, São Paulo, Brazil.
19. Clinical Hospital of Ribeirão Preto, Department of Neurosciences and Behaviour Sciences, University of São Paulo, Ribeirão Preto, Brazil.
20. Department of Pediatric Neurology, Faculty of Medicine, Farabi Hospital, Karadeniz Technical University, Trabzon, Turkey.
21. Department of Pediatric Neurology, Erciyes University, Kayseri, Turkey.
22. Children's Hospital & the Institute of Child Health, Lahore, Pakistan.
23. Pediatric Neurology, Neuropedia Hospital, Dubai, ARE.
24. Pediatric Neurology, Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai, ARE.
25. CENTOGENE GmbH, Am Strande 7, 18055 Rostock, Germany.
26. Department of Pediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
27. Department of Clinical Genetics, ErasmusMC University Medical Center, Dr.Molewaterplein 40, 3015GD Rotterdam, The Netherlands.
28. Laboratory of Genome Diagnostics, Department of Human Genetics, Amsterdam UMC, University of Amsterdam, Meibergdreef 9, 1105 AZ, Amsterdam, the Netherlands.
29. Rare Neurodegenerative Diseases Laboratory, Centro de Investigación Príncipe Felipe (CIPF), 46012 Valencia, Spain.
30. Hospital Universitari i Politècnic La Fe & IIS La Fe, Neuromuscular Diseases Unit, Department of Neurology, Valencia, Spain.
31. Universitat de València, Valencia, Spain. Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Spain.
32. Suna and İnan Kıraç Foundation, Neurodegeneration Research Laboratory (NDAL), Research Center for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey.
33. Neuromuscular Reference Centre, Department of Neurology, Antwerp University Hospital, Antwerp, Belgium.
34. Department of Neurology, Istanbul Medical School, Istanbul University, Istanbul, Turkey.
35. Department of Clinical Neurosciences, School of Clinical Medicine, University of Cambridge, Cambridge Biomedical Campus, Cambridge, UK.
36. Department of Clinical Neurosciences, John Van Geest Centre for Brain Repair, School of Clinical Medicine, University of Cambridge, Cambridge, UK.
37. Center for Rare Disease, University of Tübingen, Tübingen, Germany.
38. Institute of Medical Genetics and Applied Genomics, University of Tübingen, Tübingen, Germany
39. Department of Epileptology, Center of Neurology, University of Tübingen, Germany.
40. Hertie Institute for Clinical Brain Research, University of Tübingen, Tübingen, Germany.
41. John P. Hussman Institute for Human Genomics, University of Miami, Miami, FL, USA.
42. Dr. John T. Macdonald Foundation Department of Human Genetics, University of Miami, Miami, FL, USA.
43. Centre for Neuromuscular Diseases, Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, London, UK.
44. Neurogenetics Laboratory, The National Hospital for Neurology and Neurosurgery and the North Thames Genomics Laboratory Hub, London, UK.
45. Mendelics Genomic Analysis, São Paulo, SP, Brazil.
46. Universidade Federal Do Ceara - UFC and Hospital Infantil Albert Sabin, Fortaleza, Brazil.
47. Department of Neurology, School of Medical Sciences, University of Campinas (UNICAMP), Campinas, SP, Brazil.
48. Department of Neurology and Psychiatry, Faculty of Medicine, Assiut University, Assiut, Egypt.
49. Department of Pediatrics, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA.
50. Genetics Sections, Molecular and Clinical Sciences Institute, St George's University of London, Cranmer Terrace, London, SW17 0RE, UK.
51. Cancer Research Program, Research Institute of the McGill University Health Centre, Montréal, QC H4A 3J1, Canada.
#Contributed equally
Introduction: Charcot-Marie-Tooth Disease is a clinically and genetically heterogeneous group of hereditary neuropathies. Despite the progress of genetic diagnosis, there are still no drug therapies for any type of CMT.
Aims: To describe ARHGAP19-neuropathy and to address the therapeutic potential of ARHGAP19-RhoA signalling modulation in CMT.
Methods: By using next-generation-sequencing, we identified 16 recessive variants in the RhoGTPase activating protein 19 gene (ARHGAP19) causing motor-predominant neuropathy in 25 individuals from 20 unrelated families. The ARHGAP19 protein acts as a negative regulator of the RhoA GTPase and we used various approaches to model these variants; in-vitro GTPase-activating-protein (GAP) assays to assess if the GAP activity is affected by overexpression of proteins carrying ARHGAP19, complemented by an in-vivo Drosophila melanogaster model to test for movement, lifespan and neuromuscular junction integrity, an in-vivo Danio rerio model to test for locomotion and morphological change; and patient derived fibroblasts and iPSC motor neurons to assess their morphology and behaviour.
To address the limited therapeutic options, using a Drosophila model of ARHGAP19 ortholog, RhoGAP54D, loss-of-function, we established an in-vivo movement-based drug repurposing screen to identify novel therapies, and we have acutely fed the flies ∼120 drugs from a library of FDA-approved compounds and screened for drugs that rescued locomotor dysfunction. Potential candidate compounds are being validated in patient-derived tissues to assess their clinical potential.
Results: In-vitro biochemical and cellular assays revealed that patient variants impair the GTPase-activating protein (GAP) activity of ARHGAP19 and reduce ARHGAP19 protein levels. Parallel in-vivo studies in Drosophila and zebrafish models demonstrate a conserved role for ARHGAP19 orthologs in regulating locomotor capacity, the length and branching of motoneuron axons in zebrafish. Transcriptomics provided evidence that pathways associated with motor proteins and cell cycle are impacted.
Conclusions: Our findings reveal ARHGAP19 loss-of-function as a novel driver of inherited neuropathy and support integrated Drosophila and cell-based studies as a means to enable drug repurposing in CMT.
054Age-dependent synaptic vulnerability in NARS1 deficiency despite preserved early neurodevelopment
Stephanie Efthymiou1, Annarita Scardamaglia1, Estefania Rojo Bustamante2, Busra Aynekin3, Michelle Stewart4, Sara E Wells4, Ellie Rhymes1, Louisa Snape1, Luiz Almeida Silva5, Dimitri Kullmann5, James Sleigh1, Henry Houlden1
1. Department of Neuromuscular Disorders, UCL Queen Square Institute of Neurology, London, United Kingdom.
2. Neurology Department, Hospital Universitario Fundación Santa Fe de Bogotá, Cra. 7 #117 -15, Bogotá, Colombia; Universidad El Bosque, Ak. 9 #131a-2, Bogotá, Colombia.
3. Department of Molecular Biology and Genetics, Biruni University, Istanbul, 34015, Türkiye.
4. Mary Lyon Centre, Medical Research Council Harwell, Oxfordshire OX11 0RD, UK.
5. Department of Clinical and Experimental Epilepsy, UCL Queen Square Institute of Neurology, London, United Kingdom.
s.efthymiou@ucl.ac.uk
Background: Pathogenic variants in NARS1, encoding asparaginyl-tRNA synthetase, cause a severe neurodevelopmental disorder with progressive neurological deterioration. While impaired protein synthesis is presumed to underlie disease, the temporal and cellular mechanisms linking NARS1 dysfunction to neuronal pathology remain poorly defined. Using patient-derived iPSCs and genetically engineered mice we are seeking to disentangle early developmental effects from later neurodegenerative processes. Here, we combine in vivo and hiPSC-based models to investigate how NARS1 deficiency impacts neural development, synaptic integrity and age-dependent vulnerability.
Methods: We analysed a Nars1 R545C knock-in mouse using SHIRPA behavioural screening and confocal imaging of neuromuscular junctions (NMJs) at 1 and 3 months of age. NMJ architecture was quantitatively assessed using the NMJ_Morph platform. In parallel, patient-derived R656C iPSCs were differentiated into neural progenitor cells (NPCs) and spinal motor neurons (SMNs). NPC proliferation was quantified, while neuronal identity, axonal outgrowth and branching were assessed by immunocytochemistry and Sholl analysis. Bulk protein synthesis, transcriptomic profiling and stress-response gene expression were evaluated to assess translational integrity.
Results: Nars1-mutant mice exhibited ataxia, craniofacial abnormalities, hydrocephalus and seizure susceptibility. NMJs were normally innervated at 1 month but showed subtle partial denervation by 3 months. Quantitative analysis identified reduced acetylcholine receptor (AChR) cluster area as the primary abnormality, indicating a postsynaptic defect. In human models, patient-derived NPCs displayed markedly reduced proliferative capacity despite preserved progenitor identity. Differentiated SMNs showed largely normal axonal outgrowth with occasional axonal swellings. Bulk protein synthesis remained largely intact, while RNA-seq revealed mild induction of ATF4-responsive stress genes and reduced expression of axon-stabilizing transcripts, consistent with translational stress.
Conclusion: NARS1 deficiency permits near-normal early neuronal development but leads to progressive synaptic and postsynaptic instability. Chronic, subtle impairments in aminoacylation and protein homeostasis may compromise long-term synaptic maintenance, providing a mechanistic basis for age-dependent disease progression and a rationale for therapeutic strategies targeting translational stress.
055Protocol: Developing A Model Of Balance Impairment For People Living With CANVAS
Louie Lee1,2, Diego Kaski3, Matthew J Bancroft3, Nehzat Koohi3, Riccardo Curro1.2, Andrea Cortese1,2, Mary M Reilly1,2, Gita Ramdharry1,2,
1 Queen Square Centre for Neuromuscular Diseases, National Hospital for Neurology and Neurosurgery, University College London Hospitals NHS Foundation Trust, London, UK
2 Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, London, UK
3 Department of Clinical and Movement Neurosciences, UCL Queen Square Institute of Neurology, London, UK
Louie.lee@ucl.ac.uk. g.ramdharry@ucl.ac.uk
Background: Cerebellar ataxia, neuropathy and vestibular areflexia syndrome (CANVAS) is characterised by combined cerebellar, somatosensory and vestibular dysfunction, frequently leading to marked postural instability and falls. Rehabilitation is constrained by limited condition-specific evidence and uncertainty about how these multi-system impairments interact to drive balance problems. This study will characterise balance impairment in people living with CANVAS to inform a pragmatic model of multi-modal balance dysfunction and prioritise potential rehabilitation targets.
Methods: This observational study will recruit adults with CANVAS from specialist clinics (target n≈30). Guided by the WHO International Classification of Functioning, Disability and Health (ICF), assessments will capture impairments in Body Functions and limitations in Activity. Body Functions will be quantified through measures of proprioceptive impairment (vibration thresholds). Vestibular function will be characterised across peripheral and central domains using vestibulo-ocular reflex (VOR) measures (e.g., vHIT VOR gain and compensatory saccade parameters) together with central oculomotor measures relevant to vestibular–cerebellar dysfunction (e.g., smooth pursuit gain and gaze-evoked nystagmus metrics). Postural control will be assessed using gold-standard posturography during static standing under altered sensory conditions and during dynamic tasks (forward and lateral reach). Activity will be assessed using standardised functional balance measures alongside patient-reported balance confidence and perceived walking ability. A 6-month retrospective falls history will be collected at baseline, with falls prospectively monitored for the subsequent 6 months. Analyses will explore associations between impairment, postural control, functional performance and falls to identify candidate contributors to postural instability.
Results: The study will generate an integrated profile linking sensory, vestibular and cerebellar measures to posturography, functional outcomes, patient-reported measures and falls, to identify candidate mechanisms and phenotypic patterns relevant to rehabilitation targeting.
Conclusion: This study will provide CANVAS-specific observational evidence to underpin a model of balance impairment and to prioritise modifiable targets for subsequent intervention development and feasibility testing.
056‡Phenotypic Characterisation and Gene Therapy Development in a mouse model for SORD neuropathy
Sijiang Liu1,2, Sara Negri3, David Villarroel-Campos1,2,4, Elena R. Rhymes1,2, Aurélie Paulo-Ramos1,2, Lona Kroese5, Michael Groves6, Zoe Windsor7, Roberto Bellanti1,8, Amy F. Geard9, Riccardo Curro1, Ilaria Quartesan1, Annalucia Darbey1,2, Elena Veleva4, Bárbara F. Gomes4, Rhiannon Laban4, Amanda Heslegrave4,10, Henrik Zetterberg4,10,11, Carlo Gaetano3,12, Michael P. Lunn1, Paul Krimpenfort5, Andrea Cortese1,13*, James N. Sleigh1,2,4*
1 Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, University College London, London, UK.
2 UCL Queen Square Motor Neuron Disease Centre, UCL Queen Square Institute of Neurology, University College London, London, UK.
3 Laboratorio di Epigenetica, Dipartimento Medicina Riabilitativa NeuroMotoria - MeRiNM, Istituti Clinici Scientifici Maugeri IRCCS, Pavia, Italy.
4 UK Dementia Research Institute, University College London, London, UK.
5 Animal Modeling Facility, Netherlands Cancer Institute, Amsterdam, The Netherlands.
6 UCL Queen Square Institute of Neurology, University College London, Queen Square, London, UK.
7 Department of Epilepsy, UCL Queen Square Institute of Neurology and Biological Services, University College London, London, United Kingdom.
8 Nuffield Department of Clinical Neurosciences, University of Oxford, Oxford, UK.
9 Department of Pharmacology, UCL School of Pharmacy, 29-39 Brunswick Square, London, UK.
10 Department of Neurodegenerative Disease, UCL Institute of Neurology, London, UK.
11 Institute of Neuroscience and Physiology, Goteborgs Universitet, Goteborg, Sweden.
12 Departmental Faculty of Medicine, Saint Camillus International University of Health and Medical Sciences, Via di Sant'Alessandro 8, 00131 Rome, Italy.
13 Department of Medical Biotechnology and Translational Medicine, University of Milan, Milan, Italy.
* Corresponding authors. Email: andrea.cortese@ucl.ac.uk and j.sleigh@ucl.ac.uk
Presenting author. Email: sijiang.liu.22@ucl.ac.uk
Background: Biallelic loss-of-function mutations in sorbitol dehydrogenase (SORD) are the most common cause of recessive peripheral neuropathy, with the nonsense c.757delG variant (p.Ala253GlnfsTer27) being the most frequently identified. Sorbitol dehydrogenase is a key enzyme of the polyol pathway that converts sorbitol into fructose, and its absence leads to neurotoxic polyol accumulation.
Aims: Existing animal models have not captured the spectrum of sensory phenotypes in SORD neuropathy. Here, we generated and conducted extensive motor and sensory phenotyping of Sordc.757delG mice and developed a SORD-enhancing adeno-associated virus (AAV) gene therapy.
Methods/Materials: SORD-related carbohydrates were measured by UPLC-MS/MS, while neurofilament light chain (NfL) and periaxin were assessed using Simoa technology. Sensory function assessments (i.e., beam-walking, von Frey, pinprick, and Hargreaves tests) and footprint analysis were performed at 3 and 12 months, respectively. The AAV9-CAG-SORD gene therapy virus was first tested via intramuscular (in adults) and intraperitoneal (in pups) injections.
Results: Similar to sorbitol, there is an early and persistent accumulation of xylitol in plasma of Sordc.757delG/c.757delG mice, which is significantly higher in males than females. Conversely, fructose levels are consistently decreased in homozygous males and females but remain relatively stable over time. Sordc.757delG/c.757delG mice exhibit a robust deficit in thermal nociception by 3 months and clear gait disturbances by 12 months, while motor function is preserved. Additionally, plasma periaxin levels are altered in Sordc.757delG mice at both early (increased) and late (decreased) stages, whereas NfL remains unaffected. AAV9-CAG-SORD administration causes clear increases in SORD production. We are currently longitudinally assessing sensory function and peripheral nerve pathology up to 12 months, and evaluating the efficacy of AAV9-CAG-SORD in Sordc.757delG mice.
Conclusion: We present a new mouse model for SORD-CMT that replicates the blood biomarker profile of patients and displays a clear neuropathy phenotype, which will facilitate the ongoing testing of our AAV9-CAG-SORD gene therapy.
057‡Recurrent Laryngeal Neuropathy: a highly prevalent, naturally occurring, length-dependent motor neuropathy of horses
Victoria O’Hara1, Stephen D Cahalan1, Abdulaziz Almuhanna1, Justin D Perkins2, Tom Gillingwater3, David Goodwin1, Diego Robledo3, Androniki Psifidi2, Richard J Piercy1
1 Comparative Neuromuscular Disease Laboratory, Royal Veterinary College, London, UK
2Royal Veterinary College, London, UK
3 The University of Edinburgh, UK
vohara4@rvc.ac.uk
Background: Modelling length-dependent neuropathies is challenging due to the short axon lengths of standard laboratory species and poor availability of human samples. In horses, the left recurrent laryngeal nerve (RLn) is approximately 250 cm long and is predisposed to a highly prevalent distal axonopathy known as recurrent laryngeal neuropathy (RLN) making the horse a valuable model for length-dependent human axonopathies. However, despite its prevalence, the pathomechanisms underlying RLN remain incompletely understood.
Aims: Present an overview of recent histological and bioinformatic results evaluating RLN pathophysiology, using morphological, immunohistochemical and bioinformatic approaches.
Methods/Materials: Histopathological analysis of left and right side RLns from 15 adult horses, with varying degrees of RLN severity, was performed using resin-embedded sections and immunohistochemical labelling of axon subtypes. Neuromuscular junction (NMJ) morphology was examined in laryngeal and distal limb muscles from 9 horses via bungarotoxin and immunofluorescence labelling . Transcriptional profiling of the left and right nucleus ambiguus (nAmb) (where RLn cell bodies reside) was undertaken in 5 horses of varying disease severity using single-nucleus RNA sequencing.
Results: Nerve histopathology demonstrated that the RLn contains mixed motor, sensory, and sympathetic fibres, however, there is a selective loss of large-diameter, myelinated ChAT-positive motor axons (left>right), with preservation of unmyelinated fibres. NMJ analysis revealed extensive compensatory remodelling, including increased nerve terminal branching, enlarged endplates, and fragmentation (left>right). Similar changes in distal limb muscle NMJs from severely affected horses indicated a more generalised motor polyneuropathy. Transcriptional nAmb analysis showed no consistent left vs right neuronal signature. Instead, patterns associated with severity emerged, with neuron homeostasis disrupted between sides in mild disease, and glial stress and dysregulation in severe disease. Neurons exhibited enrichment of axonal degeneration and regeneration pathways in horses across severity groups.
Conclusion: These studies demonstrate selective vulnerability of motor axons in RLN, implicate RLN as a polyneuropathy, indicate glia involvement with disease progression, and support a length dependent metabolic pathomechanism. These findings highlight the horse as a valuable model for studying length-dependent axonopathies, relevant to both equine and human disease. Study of this common equine disorder could reveal pathways involved in the normal homeostasis of long mammalian nerves.
058Serum Polyols as Disease Biomarkers in SORD Neuropathy
Ilaria Quartesan1, Sara Negri2, Maike F Dohrn3, Riccardo Currò1, Saif Haddad1, Sijiang Liu1, Christian Laurini4, Stefano Tozza5, Adriana Rebelo6, Danique Beijer3,7, Jacquelyn Raposo6, Mario Saporta6, Pascal Achenbach8, Ulrike Schöne9, Marcus Deschauer10, Isabell Cordts10, Stefania Magri11, Paola Saveri12, Elisa Vegezzi13, Paolo Ripellino14,15, Carlo Gaetano2, Ricardo Rojas-Garcia16,17, Beate Schlotter-Weigel18, Hülya Kayserili19, Fiore Manganelli5, David N Herrmann20, Steven S Scherer21, Isabella Moroni12, Chiara Pisciotta12, Stefano C Previtali4, Davide Pareyson12, Michael E Shy22, Stephan Züchner6, James N Sleigh1,23, Mary M Reilly1, Andrea Cortese1,24
1Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, London, UK
2Laboratorio di Epigenetica, Istituti Clinici Scientifici Maugeri IRCCS, Pavia, Italy
3Department of Neurology, Medical Faculty, RWTH Aachen University, Aachen, Germany
4Neuromuscular Repair Unit, Institute of Experimental Neurology (InSpe), Division of Neuroscience, IRCCS Ospedale San Raffaele, Milan, Italy
5Department of Neurosciences, Reproductive Sciences and Odontostomatology, University of Naples Federico II, Naples, Italy
6Dr. John T. Macdonald Foundation, John P. Hussman Institute for Human Genomics, University of Miami, Miami, FL, USA
7Translational Genomics of Neurodegenerative Diseases, Hertie-Institute for Clinical Brain Research, University of Tübingen, Tübingen, Germany
8Institute of Neuropathology, Medical Faculty, RWTH Aachen University, Aachen, Germany
9Department of Neurology, Maria Hilf Hospital Mönchengladbach, Mönchengladbach, Germany
10Department of Neurology, University Hospital rechts der Isar, Technical University Munich, Munich, Germany
11Unit of Medical Genetics and Neurogenetics, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy
12Unit of Rare Neurological Diseases, Department of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy
13IRCCS Mondino Foundation, Pavia, Italy
14Department of Neurology, Neurocenter of Southern Switzerland EOC, Lugano, Switzerland
15Faculty of Biomedical Sciences, Università della Svizzera Italiana, Lugano, Switzerland
16Department of Neurology, Neuromuscular Diseases Unit, Hospital de la Santa Creu i Sant Pau, Universitat Autònoma de Barcelona, Barcelona, Spain
17Center for Networked Biomedical Research into Rare Diseases (CIBERER), Madrid, Spain
18Friedrich Baur Institute at the Department of Neurology, LMU University Hospital, LMU Munich, Munich, Germany
19Department of Medical Genetics, Koç University School of Medicine (KUSOM), Istanbul, Turkey
20Department of Neurology, University of Rochester, Rochester, NY, USA
21Department of Neurology, The Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA, USA
22Department of Neurology, Carver College of Medicine, University of Iowa, Iowa City, IA, USA
23UK Dementia Research Institute, University College London, London, UK
24Department of Medical Biotechnology and Translational Medicine, University of Milan, Milan, Italy
i.quartesan@ucl.ac.uk
Background: Biallelic loss-of-function variants in the sorbitol dehydrogenase (SORD) gene are a common cause of axonal Charcot-Marie-Tooth (CMT) disease. Loss of sorbitol dehydrogenase (SDH) activity impairs sorbitol metabolism, leading to its accumulation. However, because SDH has broad substrate specificity, its deficiency may also cause accumulation of other polyols that could contribute to disease pathophysiology. Serum polyol profiling may therefore help clarify associations with disease severity, progression, and demographic or genetic factors, and assess their potential as complementary biomarkers.
Aims: To measure serum polyols in CMT-SORD patients and assess their potential as disease biomarkers.
Methods: Serum sorbitol and three additional predicted substrates of SDH (xylitol, adonitol, and threitol) were measured by UPLC-MS/MS in 47 CMT-SORD patients from an international multicentre cohort and 43 healthy controls. Patients' clinical, genetic, and demographic data were collected from each participating centre.
Results: Of the 47 CMT-SORD patients, 30 were male (64%). Mean age of disease onset was 19±8 years (11-50) and mean age at serum sampling was 43±14 years (range 17-79). Sorbitol was 79-fold elevated in SORD compared to HC (13.3±3.1 vs 0.17±0.09 mg/L; p<0.001) and xylitol 37-fold (23.3±5.7 vs 0.63±0.20 mg/L; p<0.001), adonitol was 2.7-fold (0.22±0.05 vs 0.08±0.02 p<0.001), while threitol showed no significant difference. No differences in polyol levels or disease severity were observed between missense and truncating mutation carriers. Both sorbitol and xylitol showed a steady increase with age: in patients xylitol increased by 0.21 mg/L/year (95%CI 0.10-0.31; p<0.001) and sorbitol by 0.09 mg/L/year (95%CI 0.03-0.15; p=0.003). Males showed a non-significant trend toward higher age-adjusted sorbitol and xylitol levels compared to females. Foot dorsiflexion weakness, used as a proxy for disease severity, was independently associated with xylitol levels (β=-0.11; p=0.008) and male sex (p=0.019).
Conclusions: Serum sorbitol and xylitol are excellent diagnostic biomarkers for CMT-SORD with complete discrimination from controls. Both sugars show progressive age-dependent increase and xylitol showed a promising association with motor impairment, suggesting a potential role as severity biomarkers in therapeutic trials.
059Neurofilament light chain in all-cause Charcot-Marie-Tooth disease; preliminary data
Mariola Skorupinska1, Christopher J. Record1, Alexander M. Rossor1, Matilde Laurá1, Mary M. Reilly1
mariola.skorupinska@nhs.net
1Centre for Neuromuscular Diseases, Dept of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, London UK
Background: Charcot–Marie–Tooth disease (CMT) comprises a heterogeneous group of inherited peripheral neuropathies characterized by chronic axonal degeneration. Neurofilament light chain (NfL) is a biomarker of neuroaxonal injury that can be measured in blood using ultrasensitive assays and is elevated in several neurodegenerative and peripheral neuropathic disorders. However, its relevance in slowly progressive inherited neuropathies such as CMT remains unclear.
Aims: This study aimed to assess serum NfL levels in patients with CMT and explore associations with clinical features.
Methods: Serum NfL levels were measured in 59 adult patients with genetically or clinically diagnosed CMT. Abnormal NfL values were defined according to established age-adjusted reference ranges and percentage of the upper limit of normal (%ULN). Demographic and clinical data were collected.
Results: The cohort included 39 males and 19 females, with a mean age of 56.3 years (range 17–88). The mean CMT Neuropathy Score (CMTNS) was 6.6 (range 0–24), reflecting variable disease severity. Twelve patients had CMT1 (9 CMT1A, 1 CMT4F, and 1 genetically undiagnosed), 30 had CMT2 (17 genetically undiagnosed; 13 genetically confirmed). Two patients had intermediate CMT associated with variants in GJB1 and PLEKHG5. The remaining participants had other inherited neuropathies, including hereditary motor neuropathy (HMN, n=10), hereditary sensory neuropathy (HSN, n=1), hereditary neuropathy with liability to pressure palsies (HNPP, n=1), and a Hirayama-like syndrome (n=1). Serum NfL values ranged from 0.14 to 1.50.
Overall, 6 patients (10.2%) demonstrated abnormal serum NfL levels (range 1.01-1.50), including three with CMT1A and three with CMT2 (one with an LRSAM1 variant and two without a molecular diagnosis). Elevated NfL levels were equally distributed between sexes.
Conclusion: Elevated serum NfL levels were detected in a small subset of patients with CMT, suggesting that significant neuroaxonal injury may occur very slowly in most patients with CMT and/or at specific disease stages. Overall, normal NfL levels in most CMT patients are consistent with the slowly progressive nature of CMT. We have previously shown that NfL levels are stable over 6 years in patients with CMT1A1. Longitudinal studies are needed to clarify the role of NfL as a biomarker of disease activity and progression in other forms of CMT and as a potential biomarker of treatment response in CMT.
1Rossor AM, Kapoor M, Wellington H, Spaulding E, Sleigh JN, Burgess RW, Laura M, Zetterberg H, Bacha A, Wu X, Heslegrave A, Shy ME, Reilly MM.J Peripher Nerv Syst. 2022 Mar;27(1):50-57.
060Better Balance-CMT: Protocol for a trial of efficacy of a home-based, balance training intervention for people living with Charcot-Marie-Tooth disease
Gita Ramdharry1,2, Louie Lee1,2, Magdalena Dudziec1, Mary Reilly1,2
1Department of Neuromuscular Diseases, Queen Square UCL Institute of Neurology, London, United Kingdom,
2National Hospital for Neurology and Neurosurgery, University College Hospitals, NHS Trust, London, United Kingdom
Introduction: Multi-sensory rehabilitation has shown promising effects in people with sensory loss, and resistance training can improve proximal lower limb muscle strength. Studies of multi-sensory rehabilitation and resistance training can improve balance in people with CMT in specialist clinics. We developed a pragmatic, home-based balance rehabilitation program. Our proof-of-concept study found it to be safe and acceptable for people with CMT, with excellent engagement. This will now be tested at scale through the Better Balance-CMT (BB-CMT) trial.
Methods: We are partnering with patients and stakeholder to co-produce web-based resources for the BB-CMT intervention. It will be delivered at home by trained physiotherapists, through 3 face-to-face sessions, using self-management principles, digital materials and remote support.
A randomised, single blinded, two arm trial will ascertain efficacy of the BB-CMT intervention compared to treatment as usual. A hybrid-1 trial design is planned, to include the secondary aim of exploring potential barriers and facilitators to "real-world" implementation into practice. The program will last 12 weeks and compared to a 12-week control period. A 12-week open label is included to assess continued engagement and carry over.
The primary outcome measure is the Berg Balance Scale (BBS), calculating mean difference in BBS score between the BB-CMT and the control group using a linear regression, adjusted for baseline BBS score. Target sample is 84 participants based on previous studies of balance training in CMT, with a detectable standardised effect size of 0.66, at 80% power and 5% 1-sided alpha, allowing for a 10% drop out. Participants will be recruited from 6 NHS hospitals in England.
Results: The Better Balance-CMT digital development is currently in progress.
Conclusions: Funding for this work has been acquired through a National Institute for Health Research award. The co-production work started in late 2025, with the BB-CMT trial to commence in autumn 2026.
Other Diseases
061‡Utilising 3D human iPSC-derived skeletal muscle models to advance genetic treatment strategies for LMNA-related congenital muscular dystrophy
Daniel Moore1,2, Lucia Rossi1,2, Arta Aghaeipour11,2,8, Eugenia Carraro2,3, Heather Steele-Stallard1, Cherry Tsz Yan Wong1,2, Valentina Lionello1,2, Sungwoo Choi1,2, Luca Pinton1,2, Salma Jalal1,2, Jean-Marie Cuisset5, Gisèle Bonne6, Andrea Serio2,3, Peter Zammit4 and Francesco Saverio Tedesco1,2,7*
1Department of Cell & Developmental Biology, University College London, London WC1E 6DE, UK.
2The Francis Crick Institute, London NW1 1AT, UK.
3Centre for Craniofacial & Regenerative Biology, King’s College London, London, SE1 1UL, UK.
4Randall Centre for Cell & Molecular Biophysics, King’s College London, SE1 1UL London, UK.
5Centre de Référence des maladies neuromusculaires Nord/Est/Ile de France, Service de Neuropédiatrie, Hôpital Roger Salengro, CHRU Lille, Lille, France.
6Sorbonne Université, INSERM, Institut de Myologie, Centre de Recherche en Myologie, Paris, France.
7Dubowitz Neuromuscular Centre, Great Ormond Street Hospital for Children & UCL Great Ormond Street Institute of Child Health, London, UK.
8Infection, Immunity & Inflammation Dept, UCL GOS Institute of Child Health Zayed Centre for Research into Rare Disease in Children, London, UK.
Presenting author: a.dehkaei@ucl.ac.uk
*Correspondence: f.s.tedesco@ucl.ac.uk
Background:LMNA-related congenital muscular dystrophy (L-CMD) is a severe and incurable laminopathy. It is caused by mutations in the LMNA gene, which encodes the nuclear lamin A/C proteins. These proteins, together with LAMIN B1 and B2, construct the nuclear lamina, a critical meshwork structure providing structural stabilisation to the nucleus and regulating both chromatin organisation and gene expression. However, L-CMD research is frequently hindered by the lack of humanised, tissue-specific disease models that can accurately recapitulate the complex L-CMD phenotypes.
Aims: Our laboratory has pioneered utilising advanced 3D engineered skeletal muscle models using transgene-free based iPSC differentiation protocols. In this study, we aimed to employ this 3D platform to facilitate the screening of new therapeutic approaches. We sought to validate a transgene-free differentiation protocol and expand the number of phenotypic readouts as well as including a broader range of mutations, specifically a pathogenic variant in LMNA exon 5.
Methods/Materials: We applied our refined 3D platform to investigate L-CMD cellular phenotypes. This included assessing myogenic differentiation, and nuclear morphological abnormalities. Furthermore, we employed this platform to evaluate mutation-specific gene therapy strategies, specifically focusing on CRISPR-based gene editing to target the LMNA mutation.
Results: Our results indicate that there are no significant defects in developmental myogenesis in these models. Our platform successfully recapitulated hallmark L-CMD features, including nuclear morphological abnormalities. Notably, the precise CRISPR-based correction of the LMNA mutation resulted in the normalisation of nuclear morphology and protein localisation, without any negative impact on the myogenic process.
Conclusion: We have successfully established an improved 3D humanised model for L-CMD that accurately reflects patient-specific cellular defects. The ability to reverse these phenotypes through CRISPR editing demonstrates the potential of this platform for future therapeutic development. Ongoing research is currently focused on examining transcriptional and functional readouts in both LMNA-mutant and CRISPR-corrected 3D cultures to further validate these findings.
062‡N-of-few ASO Therapeutic Development for Patients with X-Linked Myotubular Myopathy Caused by Deep Intronic Mutations in MTM1 Gene
Sara Aguti1, Yasin Shafi2, Julienne Mueller1, Chloe Flett3, Giovanni Baranello3, Haiyan Zhou2,4, Francesco Muntoni1,3,4.
1Neurodegenerative Diseases, Queen Square Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK
2Genetics and Genomic Medicine Research and Teaching Department, Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
3The Dubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Research and Teaching Department, Great Ormond Street Institute of Child Health, University College London, 30 Guilford Street, London WC1N 1EH, UK
4NIHR Great Ormond Street Hospital Biomedical Research Centre, London WC1N 1EH, UK
sara.aguti@ucl.ac.uk
Background: X-linked myotubular myopathy (XLMTM) is a severe congenital neuromuscular disorder caused by MTM1 loss-of-function mutations. Affected infants present with profound hypotonia and respiratory failure, with ∼50% mortality in the first year. Although historically viewed as a muscle disease, recent clinical and preclinical studies demonstrate that MTM1 is also essential for hepatobiliary function. The ASPIRO AAV gene therapy trial was halted after four boys died from cholestatic liver failure because the therapy restored MTM1 only in muscle, not liver. This highlights the critical requirement for therapeutic strategies capable of correcting MTM1 deficiency in both tissues. No disease-modifying treatments currently exist.
Aims: Development of personalised antisense oligonucleotide (ASO) therapies for three deep intronic MTM1 variants and evaluation of their therapeutic potential across both muscle and liver models.
Methods/Materials: Through the Highly Specialised Congenital Myopathy Service at Great Ormond Street Hospital, we have identified three ultra-rare deep intronic MTM1 mutations: one previously reported (c.1468-577A>G) and two novels (c.342+575C>G, c.445-220A>G). All introduce pseudo-exons that disrupt the reading frame and abolish MTM1 protein expression. Mutation-specific exon-skipping ASOs were designed to block aberrant splice sites and restore the wild-type transcript. ASOs were synthesised in 2′-OMe and 2’-MOE chemistries and screened in patient-derived fibroblasts.
Result: Ten candidate ASOs were tested across the two variants. For c.1468-577A>G, two lead ASOs achieved near-complete pseudo-exon removal and robust MTM1 rescue at RNA and protein levels. For the novel c.342+575C>G variant, four lead ASOs restored the wild-type transcript and substantially increased MTM1 protein expression.
Conclusion: We have established strong proof-of-concept for ASO-mediated correction of deep intronic MTM1 mutations in patient fibroblasts. We are now validating lead ASOs in patient-derived myogenic cells and, in parallel, generating the human liver organoid model of XLMTM using iPSCs from the patient with the most severe phenotype (c.1468-577A>G). This organoid platform will enable rigorous assessment of MTM1 rescue and ASO safety in liver, a critical requirement given the multisystem nature of XLMTM and the liver-related failures of prior clinical trials.
063Phage Display for Fibroblast Specific Peptide Discovery: Improving ASO Targeted Delivery in Fibroblasts & COL6-Related Congenital Muscular Dystrophies
Sean Briggs1, Shuzhi Cheng1, Sara Aguti2, Francesco Muntoni2-3-4 and Haiyan Zhou1,4,
1Genetics and Genomic Medicine Research and Teaching Department, Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
2Neurodegenerative disease department, UCL Queen Square Institute of Neurology, London WC1N 3BG.
3 Developmental Neurosciences Research and Teaching Department, Great Ormond Street Institute of Child Health, University College London, 30 Guilford Street, London WC1N 1EH, UK
4 NIHR Great Ormond Street Hospital Biomedical Research Centre, London WC1N 1EH, UK
sean.briggs@ucl.ac.uk
Background: Collagen VI (COL6) is a major component of the skeletal muscle extracellular matrix. COL6-related congenital muscular dystrophies (COL6-CMDs) are genetic disorders caused by mutations that impair COL6 subunit tetramer formation, leading to progressive muscle degeneration and weakness, with early mortality in severe cases. Currently, no curative treatments are available; however, antisense oligonucleotide (ASO) therapies show therapeutic promise. ASOs are nucleic acid–based therapeutics that target RNA through Watson–Crick base pairing, enabling restoration of functional protein. We have previously demonstrated strong in vitro proof-of-concept supporting ASOs as a therapeutic strategy for COL6-CMDs. A major barrier to clinical translation, however, is inefficient ASO uptake by the target cells, muscle interstitial fibroblasts (MIFs). In vivo studies indicate poor uptake of naked ASOs in MIFs. To overcome this limitation, we are developing targeted ASO delivery using ASO–peptide conjugates designed to preferentially target fibroblasts while minimising uptake by non-target cell types.
Aims: To identify peptides capable of preferentially targeting MIFs to enable targeted ASO delivery while minimising uptake by off-target cell types.
Methods/Materials: Peptide phage display combined with next-generation sequencing (NGS) was used to identify fibroblast-targeting peptides. Three rounds of biopanning were performed across human fibroblasts, myoblasts, hepatocytes, and collecting duct cells to enrich for peptides with selective binding and internalisation. Candidate peptides identified by NGS were fluorescently labelled and screened across all cell types to assess specificity. Selected peptides were subsequently conjugated to ASOs, and their efficacy was evaluated in mutant cells.
Results: NGS analysis identified ten candidate peptides with preferential association to fibroblasts. Fluorescence-based screening revealed two peptides that bound fibroblasts while avoiding hepatocytes and collecting duct cells, although both also showed binding to myoblasts. Conjugation of these peptides to ASOs did not impair transcript correction, and improved efficacy compared to naked ASOs was observed in vitro.
Conclusion: Phage display coupled with NGS is a viable approach for identifying fibroblast-targeting peptides for ASO delivery. Although specificity challenges remain, this strategy provides a strong foundation for the development of targeted ASO therapeutics for COL6-CMD. Further in vivo studies are required to assess biodistribution and therapeutic efficacy.
064Assessing The Role of CDK13 In Mutant DMPK Transcription and RNA Foci Formation
Jessie Brown1
University of Nottingham
Jessie.brown@nottingham.ac.uk
Background: Myotonic Dystrophy (DM) is a complex autosomal dominant genetic disorder that affects 1 in 8000 people worldwide. The disorder is characterised by multisystemic manifestations including progressive myotonia, myopathy, muscle atrophy, and cataracts, with further abnormalities presenting in the cardiovascular, respiratory, endocrinal and gastrointestinal systems. DM is classified as a repeat expansion disorder. DM type 1 is caused by a trinucleotide repeat expansion of CTG in the 3’untranslated region of the dystrophia myotonica protein kinase gene (DMPK). The mutant expansion RNA is primarily retained in the nucleus and sequesters the RNA binding protein, muscle blind-like 1 (MBNL1), to form nuclear RNA foci resulting in downstream cellular consequences. Cyclin dependent kinases 12 and 13 (CDK12/13) are transcriptional kinases which promote and maintain RNA polymerase II transcriptional elongation and processivity. Knockdown of either kinase has been shown to negatively impact transcription, DNA repair mechanisms, R-loop resolution and cell viability with greater effects seen upon dual knockdown. Previous reports show CDK12 knockdown in DM1 cell lines and mouse models results in reduced nuclear RNA foci and improved myotonia.
Aims: We aim to understand the transcriptional mechanisms underlying myotonic dystrophy and whether the transcriptional kinase CDK13 behaves like its ortholog CDK12 and influences RNA foci dynamics.
Methods: We screened online databases to investigate and compare CDK12/CDK13 expression in healthy and DM1 patients. We used siRNAs and shRNA lentiviruses to knockdown CDK13 in the DM1 cell line. Protein levels were quantified using western blotting techniques and RNA foci dynamics (abundance, intensity and area) were measured using fluorescence in-situ hybridisation.
Results: A 30% knockdown in CDK13 resulted in a visible but not statistically significant decrease in RNA foci abundance and area. The CDK13 knockdown did result in a significant decrease in RNA foci intensity.
Conclusion: We have shown preliminary evidence that reduced CDK13 levels, just like CDK12, influences RNA foci dynamics. Further replicates and experiments are needed in order to understand how transcriptional elongation influences the metabolism of the mutant DMPK transcript.
065Disease-specific longitudinal muscle ultrasound patterns in Paediatric TTN-related congenital myopathy
Gianpaolo Cicala1,2, Giovanni Baranello1,2,5, Maria Vanegas3, Luke Perry1,2, Francesco Muntoni1,2,5, Heinz Jungbluth3,4, and Anna Sarkozy1,2
1Great Ormond Street Hospital for Children NHS Foundation Trust, London, UK.
2Dubowitz Neuromuscular Centre, Institute of Child Health, University College London, London, UK.
3Department of Paediatric Neurology – Neuromuscular Service, Evelina London Children’s Hospital, Guy’s & St Thomas’ NHS Foundation Trust, London, UK.
4Randall Centre for Cell and Molecular Biophysics, Muscle Signalling Section, Faculty of Life Sciences and Medicine (FoLSM), King’s College London
5UCL NIHR GOSH Biomedical Research Centre
Corresponding author:
Gianpaolo Cicala (Gianpaolo.cicala@gosh.nhs.uk)
Great Ormond Street Hospital for Children NHS Foundation Trust, London, UK.
Background: Biallelic pathogenic TTN gene variants cause a form of congenital myopathy (TTN-Cmyo) with mostly neonatal/paediatric onset. Natural history and markers of disease progression are not fully known. A natural history study for TTN-Cmyo (TREATIN) has been initiated at Great Ormond Street and the Evelina London Children’s Hospitals in January 2024, with one aim to investigate the role of imaging and more specifically muscle ultrasound (US) to aid diagnosis and monitor progression.
Aims: To describe 12-month longitudinal muscle US findings performed in Paediatric TTN-Cmyo patients, focusing on increases in echogenicity and disease-specific patterns of muscle involvement over time.
Methods/Materials: Ultrasounds (Siemens Acuson X300 machine, with a 11.4 MHz linear array probe) were performed wherever possible on the right upper and lower limb in 16 muscles at baseline and at 12-month. Muscle echogenicity was scored with the semi-quantitative Heckmatt scale (grade 1-4). Scoring was completed by the examiner and subsequently by minimum three additional raters, with final consensus score. Median echogenicity scores were created for each muscle and each patient to study global involvement. Longitudinal change was defined as worsening by ≥1 Heckmatt grade in at least one muscle.
Results: 15 patients completed scans at 12 months interval. At baseline, higher median echogenicity scores were observed in the semitendinosus, semimembranosus, tibialis anterior and peroneal muscles, while other muscles (in particular vastus intermedius, biceps femoris, soleus and deltoid) showed relative sparing. This pattern was similar at Month 12. Median muscle echogenicity per patient remained largely stable over the follow-up period (median change=0). Longitudinal changes were observed in 14/15 patients (93%), affecting 5 muscles/patient on average (range 2-9).
Conclusion: This study demonstrates a selective and disease-specific pattern of muscle involvement on ultrasound in paediatric TTN-Cmyo, with overall stability of median echogenicity scores over a 12-month period and only mild, focal changes at the individual muscle level. Importantly, more advanced and quantitative ultrasound techniques may improve sensitivity to subtle disease-related changes and enable detection of progression over shorter follow-up intervals. Overall, these findings support muscle ultrasound as a feasible and informative imaging tool for natural history studies and future clinical trials in paediatric TTN-Cmyo.
066Evolution of pharmacological management for non-dystrophic myotonias (NDM): a UK cohort study (1998-2024)
E. Leone1, M. Asad1, I. Skorupinska1, N. James1, S. Holmes1, V. Vivekanandam1, M. G. Hanna1, D. Jayaseelan1
1Queen Square Centre for Neuromuscular Diseases (CNMD), London, WC1N 3BG
e.leone@nhs.net
Background: Non-dystrophic myotonias (NDM) are skeletal muscle channelopathies caused by ion channel dysfunction, including myotonia congenita (MC), sodium channel myotonia (SCM), and paramyotonia congenita (PMC). They are characterised by muscle stiffness due to myotonia. Though incurable, symptoms are treatable pharmacologically.
Aims: To analyse efficacy of pharmacological management in NDM.
Methods/Materials: We analysed genetically confirmed adult NDM patients referred to the UK Highly Specialised Service (HSS) between 1998-2024 from our skeletal muscle channelopathy cohort database.
Results: Of 96 adults (CLCN1: 54.2%; SCN4A: 45.8%; female: 36.5%; median age: 51), 93.8% initiated medications. Patients declined treatments because of mild symptoms, contraindications and side effect concerns. Mexiletine was first-line in 75.5%, Lamotrigine second-line (55.2%). At follow-up, 67.4% remained on medications (Mexiletine: 53.9%; Lamotrigine: 34.9%). Medication changes were common (83.4% switched ≤3 times; max 9), driven by inefficacy (45.0%) and side effects (33.5.0%; of which 38.3% were gastrointestinal-related). Males switched mainly due to inefficacy (55.7%), females due to side effects (42.3%). Myotonia symptoms persisted in 48.3% of medication users at last visit, more in females (54.3%) than males (37.7%), and in CLCN1 (50.0%) than in SCN4A (41.5%). 36% of patients prescribed Mexiletine reached the maximum recommended dose of 600mg; among them 85% were men and 15% were female. Most patients required doses of Mexiletine between 400mg (19.7%) and 600mg (21.1%) for symptom control.
Conclusion: Drug therapy is common in NDM, but myotonia symptoms often persist, leading to multiple medication changes. Females remained more symptomatic and less likely to reach higher mexiletine doses. These findings highlight the need for more effective, targeted treatments.
067Advanced modelling of X-linked myotubular myopathy using human myogenic stem cells and 3D tissue engineering
Valentina Maria Lionello1,2, Lucia Rossi1,2, Leah Zerlin1,2, Daniel Moore1,2, Mariam Zouhair1,2, Owen Li Cheuk Kwan1, Rocco D’Antuono2, Kamel Mamchaoui3, Anne Bigot3, Ok-Ryul Song2, Francesco Muntoni4, Francesco Saverio Tedesco1,2,4,*
1Department of Cell & Developmental Biology, University College London, London WC1E 6DE, UK.
2The Francis Crick Institute, London NW1 1AT, UK.
3Center for Research in Myology UMRS974, Sorbonne Université, INSERM, Myology Institute AIM, Paris, France.
4Dubowitz Neuromuscular Centre, Great Ormond Street Hospital for Children & UCL GOS Institute of Child Health, London, UK.
*Correspondence: f.s.tedesco@ucl.ac.ukPresenting author:valentina.lionello@crick.ac.uk
Background: X-linked myotubular myopathy (XLMTM) is a congenital, untreatable muscular disease featuring muscle weakness from birth. Patient muscle biopsies reveal small fibres and abnormally positioned organelles, including nuclei. XLMTM is caused by pathogenic variants in the MTM1 gene, coding for a crucial enzyme for cellular membrane remodelling. Conventional in vitro models rely on cells obtained via invasive sampling, which are unsuitable for extended experimental timeframes. Animal models raise ethical concerns and fail to replicate the complexity of human phenotypes, as evidenced by the recent termination of therapeutic trials due to poor predictive outcomes in patients.
Aims: We aim to model XLMTM human muscle tissue pathology across different developmental stages and cellular maturation.
Methods/Materials: We developed a human XLMTM platform by generating 3D bioengineered muscle tissues from both human immortalized myoblasts and induced pluripotent stem cells (iPSCs) harbouring different MTM1 variants.
Results: Bioengineered XLMTM muscles exhibited aligned and hypertrophic fibres with abnormal nuclear distribution, mirroring patterns seen in patient tissue samples. Functional assessment demonstrated significant contractile impairment and calcium regulation defects in the XLMTM bioengineered muscles compared to the healthy control lines. To examine the early associated phenotype of XLMTM, we generated an iPSC line harbouring MTM1 mutation. iPSC-derived XLMTM muscles showed reduced fibre size compared to unaffected samples, validating our previous observations. Genetic correction via CRISPR-Cas9 improved fibre size, proving these abnormalities originate during early muscle development and are consistent with alterations identified by transcriptomic analyses.
Conclusion: Our platform faithfully recapitulates XLMTM disease phenotypes and their developmental trajectory, providing the foundations for new human models that enable evaluation of advanced therapies.
068Evaluation of the GSL30 transgenic mouse to model equine polysaccharide storage myopathy (PSSM1)
Claire Massey1, Dominic Wells1, Richard J. Piercy1
1 Royal Veterinary College, London, NW1 0TU
cmassey@rvc.ac.uk
Background: Skeletal muscle polyglucosan accumulation occurs in several human and mammalian glycogen storage myopathies, including the highly prevalent equine disorder polysaccharide storage myopathy (PSSM1). PSSM1 is associated with a missense, gain of function mutation in the glycogen synthase (GYS1) gene resulting in constitutive glycogen synthase activation. Similarly, the GSL30 mouse expresses a muscle-specific constitutively active glycogen synthase enzyme. Recent reports reveal that gain of function GYS1 mutations are associated with myopathies in humans.
Aims: Explore the suitability of the GSL30 mouse to model the equine PSSM1 histological phenotype.
Methods/Materials: Cryosections from gastrocnemius muscle samples were obtained from GSL30 and WT mice, euthanased at discreet time points from 6 weeks to 18 months of age. These were examined alongside cryosections from semimembranosus muscle samples from horses over the age of 6 years diagnosed with PSSM1, following submission of a muscle biopsy sample to the RVC Comparative Neuromuscular Diseases Laboratory, following staining with haematoxylin and eosin, periodic acid Schiff (with and without prior amylase digestion) and anti-mouse or anti-horse IgG antibodies. Slides were evaluated for pathological features including the presence of internalised nuclei, amylase-resistant polyglucosan and sarcoplasmic staining within myofibres.
Results: Gastrocnemius muscle of GSL30 mice, particularly at 6 months of age and older, showed occasional intracellular IgG staining within myofibres, internalised nuclei and infiltrating mononuclear cells including macrophages- suggesting low-level degeneration and regeneration. Additionally, there was progressive accumulation of polyglucosan within the sarcoplasm from 6 weeks of age. The number of fibres containing internalised nuclei or amylase-resistant polyglucosan were both significantly increased in comparison to WT controls at 8 months of age and 3 months of age respectively. Semimembranosus muscle of PSSM1-affected horses similarly showed the presence of occasional intracellular IgG staining, internalised nuclei, infiltrating mononuclear cells and sarcoplasmic amylase-resistant polyglucosan.
Conclusion: The myopathy observed histologically in the GSL30 transgenic mouse is similar to that seen in equine PSSM1 and therefore the GSL30 mouse could provide a valuable tool in which to examine exercise, dietary or therapeutic strategies for use in PSSM1-affected horses of relevance to humans with the similar disorder.
069A severe case of congenital myopathy caused by a novel homozygous intronic MYL1 variant
Juliane S Müller1, Darren Chambers1, Hannah Robinson2, Anna-Marie Johnson2, Deena-Shefali Patel3, Kayal Vijayakumar3, Luke Mills3, Jan Cobben4, Rahul Phadke1, Anna Sarkozy1, Francesco Muntoni1
1The Dubowitz Neuromuscular Centre, University College London, and Great Ormond Street Hospital, London, UK
2Exeter Genomics Laboratory, Royal Devon and Exeter Hospital, Exeter, UK
3Chelsea and Westminster Hospital, London, UK
4Imperial College Healthcare Trust and Imperial College, Section of Genetics and Genomics, Faculty of Medicine, London, UK
j.mueller@ucl.ac.uk
Background:MYL1 related congenital myopathy (CMYO) is an ultra-rare, severe condition, associated with a deficiency of the fast skeletal muscle-specific essential myosin light chain and impaired development of fast-twitch muscle fibres. To date, only four individuals with MYL1 related CMYO have been described in the literature.
Aims: To report a novel homozygous intronic MYL1 variant in two siblings from a consanguineous family.
Methods/Materials: The index patient is a female baby born prematurely at 32+3 weeks in poor condition, with marked hypotonia, only occasional distal antigravity movements of hands and pulmonary hypoplasia with pneumothorax on the right side. The baby died at the age of three weeks on the neonatal intensive care unit ward. Two years prior, a sibling also born prematurely at 29 weeks presented with congenital hypotonia with a similar movement pattern, dependence on the ventilator and died at the age of 10 days. Rapid trio whole genome sequencing (WGS) was performed at the Exeter Genomics Laboratory in 2023 for the first and in 2025 for the second child. RNA and protein analysis was performed on a muscle biopsy of the index patient.
Results: A quadriceps muscle biopsy showed ‘floret-like’ arrangement of a distinct two-fibre population comprising few hypertrophic fibres expressing slow myosin, each surrounded by hypotrophic/atrophic fibres predominantly expressing fast myosin, with extensive foetal/embryonic myosin coexpression. WGS analysis revealed a novel homozygous intronic variant in the MYL1 gene in both affected children. SpliceAI predicted aberrant splicing with a score of 0.4. RNA analysis from the muscle biopsy confirmed the insertion of 19 intronic nucleotides and subsequent frameshift as predicted by SpliceAI. A low amount of wild type MYL1 transcript was also present, suggesting that the variant did not cause a complete loss of MYL1 expression.
Conclusion: We report a new case of MYL1 related CMYO associated with a very severe clinical picture, similar to previously published patients. The characteristic ‘floret pattern’ of fast-fibre hypotrophy/atrophy is an important diagnostic clue in the muscle biopsy. Our case also demonstrates how the improvement of AI splice prediction tools can support variant interpretation and prioritisation for WGS analysis.
070Spatial Transcriptomic Profiling Reveals Compensatory and Degenerative Pathways in VCP-MSP Muscle Across Histological Severity
Marianela Schiava1,2, Sandra de Haan3, Pietro Spitali3, Yolande Parkhurst2, Dan Cox1, Andrew Galloway1, James Clark1, Jordi Diaz Manera1,2.
1 John Walton Muscular Dystrophy Research Centre, Newcastle University, Newcastle upon Tyne, UK.
2 Newcastle Hospitals NHS Foundation Trusts, Newcastle upon Tyne, UK.
3 Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands.
marianela.schiava@newcastle.ac.ukmarianela.schiava@nhs.net
Background: Valosin-containing protein multisystem proteinopathy (VCP-MSP) is an autosomal dominant, young adult-onset, progressive, and lethal neurodegenerative disorder caused by variants in VCP. VCP-MSP affects the peripheral and central nervous systems and bone, resulting in a heterogeneous phenotype including inclusion body myopathy, Paget disease of bone, frontotemporal dementia, and neuropathy, dying due to respiratory insufficiency or end-stage dementia. No disease-modifying treatments exist. Identifying therapeutic targets requires first uncovering the intracellular molecular pathways disrupted and their association with disease severity.
Aims: Building on our previous work identifying local degenerative niches in VCP muscle, we used spatial transcriptomics to compare gene expression in VCP-MSP patient muscles and healthy controls, stratified by histological severity and degenerative niche sites, to uncover intracellular pathways driving disease.
Methods: Spatial transcriptomic analysis (Xenium) was performed on 11 muscle biopsies from VCP-MSP patients (histology damage: mild 3, moderate 5, severe 3; females 3, males 8; VCP variants: c.464G>A p.Arg155His 4, c.463C>T p.Arg155Cys 3, c.475C>T p.Arg159Cys 2, c.277C>T p.Arg93Cys 2) and 4 healthy controls. R-based differential expression and Metascape pathway analysis was done.
Results: The pseudo-bulk gene expression of myofibers analysis revealed that pathways upregulated in VCP-MSP (MAP4, RNF187, NFE2L2, FXR2, UBTF, LARP4B, NF2) samples versus controls included cellular homeostasis, protein and organelle modification and localization, mitophagy, cell signalling, cytoskeleton organization, cell junctions, and angiogenesis. Principal component pseudotime analysis showed higher expression of these genes in mild and moderate VCP-MSP samples, with reduced expression in severe histology cases. Genes involved in intracellular energy production (SDHA, MFN2, PGM1) were downregulated in VCP-MSP samples compared to controls, with downregulation progressively increasing from mild to severe cases. No significant differences in gene expression were observed by gender or VCP variant.
Conclusion: To our knowledge, this is the first study reporting spatial transcriptomic data in VCP-MSP. Reduced expression of genes involved in cellular regulation in severely affected samples, combined with downregulation of energy production genes, may indicate cellular senescence. The next step will explore gene expression in localized areas of muscle damage (“degenerative niches”) compared to the rest of the muscle to understand inter-fiber communication and site of disease onset.
071Seronegative Immune-Mediated Necrotizing Myopathy: A Clinically Challenging Presentation and Its Management
Zeeshan Yousuf1, Azlisham Mohd Nor2
1Department of Neurology, Derriford Hospital, University Hospitals Plymouth NHS Foundation Trust
2 Department of Neurology, Derriford Hospital, University Hospitals Plymouth NHS Foundation Trust
zeeshan.yousuf@nhs.net
Background: Immune Mediated Necrotizing Myopathy (IMNM) is a rare autoimmune disorder that can attack skeletal muscles, leading to progressive weakness. Seronegative forms, lacking detectable autoantibodies, are particularly challenging to diagnose and may masquerade as more common conditions, delaying timely treatment.
Case Presentation: We report a 52-year-old male with a complex medical history hypertension, type 2 diabetes, chronic smoking, and alcoholic liver disease who presented with gradually worsening limb weakness, difficulty swallowing, and slurred speech. Initial laboratory tests revealed strikingly elevated creatine kinase levels, yet the myositis antibody panel was negative, leaving the cause uncertain. Muscle biopsy ultimately revealed necrotizing myopathy without inflammatory infiltrates, clinching the diagnosis of seronegative IMNM.
Results: Prompt initiation of high-dose intravenous methylprednisolone, transitioned to oral prednisolone and mycophenolate mofetil, led to a dramatic improvement in muscle strength and swallowing, illustrating the profound impact of early intervention even in antibody-negative cases.
Conclusion: This case underscores the diagnostic puzzle posed by seronegative IMNM and highlights the critical role of muscle biopsy and early immunosuppressive therapy. Clinicians should maintain a high index of suspicion in patients with unexplained progressive weakness, as timely recognition can transform outcomes. Further research is essential to refine diagnostic criteria and treatment strategies for this elusive but treatable condition.
Diagnostics and Cross-cutting Therapies
072‡Using untargeted proteomics in the diagnosis of Rare Genetic Conditions: the paradigm of mitochondrial disease
Andrew M. Frey1,2,3, Jamie Leighton1,2, Sarah J. Smith8, Abeer Dannoura1, Lucie Taylor1,8, Daniella H. Hock4,5,6, Nikeisha Caruana4, Charlotte L. Alston1,2, Andrew Morris17,18, Samira Saadi15, Véronique Paquis-Flucklinger16, Taghreed Shuaib13,14, Naif A. M. Almontashiri9,10, David R. Thorburn5,6,7, David A. Stroud4,5,6, Matthias Trost3,11, Robert W. Taylor,1,2,8
1. Translational and Clinical Research Institute, Newcastle University, UK
2. Mitochondrial Research Group, Newcastle University, UK
3. Biosciences Institute, Newcastle University, UK
4. Department of Biochemistry & Pharmacology, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Australia
5. Murdoch Children’s Research Institute, Melbourne, Australia
6. Victorian Clinical Genetics Services, Melbourne, Australia
7. Department of Paediatrics, University of Melbourne, Australia
8. NHS Highly Specialised Service for Rare Mitochondrial Disorders, Newcastle Upon Tyne Hospitals NHS Foundation Trust, UK
9. Center for Genetics and Inherited Diseases (CGID), Taibah University, Madinah, Saudi Arabia
10. Research Department, King Khaled Eye Specialist Hospital and Research Center, Riyadh 11462, Saudi Arabia
11. Faculty of Biology, Medicine and Health, Manchester University, UK
13. Princess Al-Jawhara Centre of Excellence in Research of Hereditary Disorders (PACER-HD), Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia
14. Pediatrics Department, King Abdulaziz University, Jeddah, Saudi Arabia.
15. Nice Teaching Hospital (CHU Nice), Nice, France
16. Institute for Research on Cancer and Aging, Université Côte d'Azur, Nice, France
17. Manchester Centre for Genomic Medicine, Manchester University Hospitals NHS Trust, UK
18. Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK
andrew.frey1@ncl.ac.uk
Background: Similarly to the cumulative burden of rare diseases, mitochondrial diseases are common, and display clinical heterogeneity and multisystem involvement. Diagnosis is driven by genomics, but only diagnoses a proportion of cases. Additional ‘omic strategies may leverage further diagnoses and provide evidence assessing pathogenicity of variants of unknown significance. We have recently shown that untargeted proteomics outperforms the measurement of respiratory chain enzyme activities in providing supportive evidence of a genetic diagnosis in patient samples with confirmed mitochondrial disease (PMID: 40400026). Furthermore, the untargeted nature of proteomics means that thousands of potentially clinically relevant proteins can be quantified in a single assay.
Aims: To further establish a high-throughput untargeted proteomics workflow capable of deep quantitation of the proteome, assessing its utility as a diagnostic tool for mitochondrial and other rare genetic conditions.
Methods/Materials: Dermal fibroblasts were cultured and then entirely processed in a 96-well format enabling high-throughput sample preparation. High throughput LC-MS/MS was performed using an EvoSep One LC and timsToF-HT mass spectrometer. Several samples were derived from patients harbouring well-characterised pathogenic variants in a number of mitochondrial disease genes, as well as from families with candidate variants in UQCC1, a complex III assembly factor with no currently reported disease-gene association.
Results: The high throughput workflow delivered deep proteome coverage: ∼7000 cellular and ∼1000/1131 mitochondrial proteins were quantified per sample. High coverage of OXPHOS complex subunits enabled relative complex abundance (RCA) analysis, and this could be used to observe predictable OXPHOS proteome deficiencies arising in UQCC1 patients. Crucially, total hands-on time for a single operator to process ∼40 samples—from lysis through LC-MS/MS and data analysis—was ∼60–72 hours, with scope for further optimisation. While this study focused on dermal fibroblasts, we show that the workflow is compatible with other cell types and tissues.
Conclusion: The proteomics workflow demonstrated rapid turnaround, scalability, and deep quantification of clinically relevant proteins, highlighting its potential to complement genomic and other clinical testing. This study contributes to the evidence base for integrating untargeted proteomics into clinical diagnostics for mitochondrial and other rare diseases with complex aetiology.
073Allied Health Professionals Neuromuscular Hub: Empowering Allied Health Professionals to support adults living with neuromuscular conditions
1University College Hospitals NHS Trust, London, United Kingdom,
2King's College Hospital NHS Trust, London, United Kingdom,
3St George's University Hospitals NHS Foundation Trust, London, United Kingdom,
4Peacocks Medical Group, Newcastle, United Kingdom,
5Muscular Dystrophy UK, London, United Kingdom,
6North Bristol NHS Trust, Bristol, United Kingdom,
7Self-employed Wheelchair Specialist, London, United Kingdom,
8University College London, London, United Kingdom,
9University Hospital Southampton NHS Trust,
10Oxford University Hospitals NHS Trust, Oxford, United Kingdom,
11Treat-NMD, Newcastle, United Kingdom,
12Newcastle University, Newcastle, United Kingdom,
13Newcastle Hospitals NHS Foundation Trust
Background: Providing allied health professionals (AHPs) with current knowledge around assessment and management of neuromuscular conditions is critical to delivery of best care. When conditions are very rare and complex, access to experts is limited and an easily accessible platform for gaining knowledge can be extremely valuable. Muscular Dystrophy UK recognised this need and supported the development of an online education module for physiotherapists in 2016. We now acknowledge the need to expand this successful resource to wider AHP practice and update with new knowledge.
Aims:
Deliver a digital suite of curated, evidence-based educational resources for AHPs, providing best management strategies for their adult patients.
Raise awareness of the additional care needs and concerns among adult patients and identify strategies that support patient independence with dignity.
Provide greater visibility of therapy training achievements and understanding of user profiles via platform analytics.
Methods: An expert panel of AHPs working in Neuromuscular Diseases were invited to update and expand the learning resource to 15 modules. The work is funded by Muscular Dystrophy UK, in partnership with Treat-NMD. The AHP Neuromuscular Hub is situated on POD-LMS and can be accessed directly or via pod-nmd.org.
Results: The work is now complete, and the 15 modules have been reviewed by colleagues not involved in the development. They include recorded presentations, a list of available resources, references for each module, useful materials which can be downloaded, and links to related podcasts. Downloadable certificates will be available on successful completion of a quizzes for each module.
The module topics are: Introduction to Neuromuscular Diseases, Assessment & outcomes; Exercise; Fatigue Management; Contracture Management; Splints and Orthoses; Postural Management; Wheelchairs; Dietetics for Weight Loss; Dietetics for Weight Gain; Speech Therapy: Communication; Speech Therapy: Chewing and Swallowing; Respiratory Management; Occupational Therapy; Self-management.
Conclusion: The modules will be launched on 2nd February 2026 through Muscular Dystrophy UK communications. Registrations and completion of the modules will be monitored.
074DECIPHER: Clinical interpretation of nuclear and mitochondrial DNA variation and sharing of rare disease patient data
K. R. Schon1,2, J. Foreman3, J. Lecarpentier-Guillou-Keredan3, B. Ashimi3, Y. Haider3, M. Eladawy3, S.E. Hunt3, M.E. Hurles4, M. A. Freeberg3 and H.V. Firth1,2,4
1Department of Genomic Medicine, University of Cambridge, Cambridge Biomedical Campus, Hills Road, Cambridge CB2 0QQ
2.Cambridge University Hospitals NHS Foundation Trust, Cambridge Biomedical Campus, Hills Road, Cambridge CB2 0QQ
3.European Molecular Biology Laboratory, European Bioinformatics Institute, Wellcome Genome Campus, Hinxton, Cambridge, CB10 1SD, United Kingdom
4.Wellcome Sanger Institute, Wellcome Genome Campus, Cambridge, Hinxton CB10 1SA
ks339@cam.ac.uk
Background: DECIPHER (https://deciphergenomics.org) is a free web-based platform which fuels rare disease research by enabling clinicians and researchers to share phenotype-linked candidate diagnostic variants. Frequent updates incorporate curated new resources for interpretation of genetic variants.
Aims: To improve DECIPHER resources for interpretation of mitochondrial DNA (mtDNA) variants.
Methods/Materials: gnomAD population data and gene predictive scores for mtDNA
Results: DECIPHER supports interpretation of variants in the nuclear genome and mtDNA in a single user-friendly interface, including sequence variants, copy-number-variants and structural variants. Heteroplasmy in different tissues can now be recorded for mtDNA variants.
DECIPHER provides variant interpretation interfaces which summarize and contextualize genotypic and phenotypic data. Gene pages provide gene/disease association information from resources such as Gene2Phenotype and the ClinGen Gene Curation working group. Gene predictive scores, including scores from the mitochondrial genome constraint model developed by Lake et al., are provided which indicate which type of variants are depleted/enriched in gnomAD. Protein predictive scores which predict if a protein is associated with a specific disease mechanism are also displayed.
An interactive genome browser and protein browser allows users to visualize their patient’s variant on a 2D protein view and 3D protein structure. Mitochondrial DNA variants observed in gnomAD are displayed in the genome browser. Homoplasmic/ heteroplasmic counts and allele frequencies for each haplotype are displayed alongside lineage information from MITOMAP and heteroplasmy distribution.
Annotation of variant pathogenicity using ACMG/ClinGen technical standards for CNVs and ACMG guidelines for sequence variants is supported. ClinGen Variant Curation Expert Panel Recommendations are also displayed (e.g. Mitochondrial Diseases Variant Curation Expert Panel).
DECIPHER currently hosts >51,000 open-access records including >65,000 variants and >205,000 phenotypes and enables contact with the depositing clinician for patients of interest. This facilitates research into new gene/disease associations, mechanisms of disease and new phenotypes associated with a disease.
Conclusions: Data sharing via DECIPHER fuels rare disease research and collaboration, including identifying potential participants for clinical trials. DECIPHER has facilitated over 4,000 publications over the last 20 years. A therapies interface is currently under development which will display known treatments, late-stage clinical trials and therapies under development, including nucleic acid therapies.
AUTHOR INDEX
Abbott, L (008) S12
Achenbach, Pascal (058) S45
Adcock, Kate (003) S9
Adu, Emmanuel (033) S27
Aghaeipour, Arta (028) S24, (061‡) S47
Aguti, Sara (028) S24, (062‡) S48, (063) S48
Akhter, Samiha Zaman (001) S8
Al Kharji, Reem (021) S20
Alabaf, Setareh (012‡) S15
Alabaf, S (015) S17
Alavi, Shahryar (053) S41
Alcock, Lisa (032) S27
Allen, Jodi (073) S54
Almontashiri, Naif A. M. (072‡) S54
Almuhanna, Abdulaziz (057‡) S44
Al-Mutairi, Dalal A. (041) S33
Al-Mutairi, Fahad K. (041) S33
Alonso-Jimenez, Alicia (035‡) S29
Al-Shami, Ahmad (041) S33
Alston, Charlotte L. (050‡) S39,
(072‡) S54
Alvarez, Grecia (002) S9
Alvi, Javeria Raza (053) S41
Angadi, Sanjana (012‡) S15
Appleton, Pamela (073) S54
Arany, Eszter S (042) S34
Armirola Ricaurte, Camila (053) S41
Arora, Annika (004) S10
Asad, M. (066) S50
Ashimi, B. (074) S55
Aughey, Gabriel (053) S41
Aynekin, Busra (053) S41, (054) S42
Ayuso, Eduard (030) S26
Ayvazian-Hancock, Ani (006‡) S11
Başak, A. Nazlı (053) S41
Baets, Jonathan (035‡) S29, (053) S41
Baker, Stuart (013) S16
Bancroft, Matthew J (055) S43
Banda, M (040) S32
Banu, Selina H. (053) S41
Baranello, Giovanni (003) S9, (004) S10
Baranello, G (008) S12
Baranello, Giovanni (010‡) S14, (062‡) S48, (065) S50
