Progesterone may be a regulator and B12 could be an indicator of the proximal D4Z4 repeat methylation status on 4q35ter.

Facioscapulohumeral dystrophy (FSHD) has a hypomethylation-related epigenetic background and exhibits a different course in male and female patients. The differences between males and females have been linked to the levels of sex hormones. This study is the first to investigate the possible effect of these hormones on methylation status. We hypothesized that the levels of sex-related hormones, estradiol, testosterone, progesterone, and prolactin might be associated with the methylation status of the proximal part of the D4Z4. We also investigated the effect of fT3, folic acid, and vitamin B12 levels. We collected blood from 28 FSHD patients and 28 controls. DNA was extracted from each individual for bisulfite methylation analysis and serum was separated for biochemical analysis of estradiol, testosterone, progesterone, prolactin, fT3, folic acid, and B12 analysis. Methylation analysis was specified to the DR1, 5P regions and the proximal region covering both DR1 and 5P. Methylation levels were compared between FSHD patients and controls. The correlation of methylation levels with estradiol, testosterone, progesterone, prolactin, fT3, folic acid, and B12 was investigated. We found that the 5P region and the proximal region were significantly hypomethylated in FSHD patients compared to the controls, but not the DR1 region. Male patients exhibited a significant reduction in DNA methylation compared to male controls. Older FSHD patients exhibited a notable decrease in fT3 levels and hypomethylation of the 5P region. Analyses of each CpG revealed seven hypomethylated positions that were significantly different from the control group. Two of the positions demonstrated a correlation with progesterone in the control group. With the exception of one position, the methylation levels were inversely correlated with vitamin B12 in FSHD patients. The results of our study indicate that the methylation of the proximal D4Z4 region, particularly at specific positions, may be associated with progesterone. In addition, vitamin B12 may be an indicator of hypomethylation. We suggest that examining position-specific methylations may be a useful approach for the development of epigenetic treatment modalities.

Oligonucleotide Therapies for Facioscapulohumeral Muscular Dystrophy: Current Preclinical Landscape.

Facioscapulohumeral muscular dystrophy (FSHD) is an inherited myopathy, characterized by progressive and asymmetric muscle atrophy, primarily affecting muscles of the face, shoulder girdle, and upper arms before affecting muscles of the lower extremities with age and greater disease severity. FSHD is a disabling condition, and patients may also present with various extramuscular symptoms. FSHD is caused by the aberrant expression of double homeobox 4 (DUX4) in skeletal muscle, arising from compromised epigenetic repression of the D4Z4 array. DUX4 encodes the DUX4 protein, a transcription factor that activates myotoxic gene programs to produce the FSHD pathology. Therefore, sequence-specific oligonucleotides aimed at reducing DUX4 levels in patients is a compelling therapeutic approach, and one that has received considerable research interest over the last decade. This review aims to describe the current preclinical landscape of oligonucleotide therapies for FSHD. This includes outlining the mechanism of action of each therapy and summarizing the preclinical results obtained regarding their efficacy in cellular and/or murine disease models. The scope of this review is limited to oligonucleotide-based therapies that inhibit the DUX4 gene, mRNA, or protein in a way that does not involve gene editing.

French National Protocol for diagnosis and care of facioscapulohumeral muscular dystrophy (FSHD).

Facioscapulohumeral muscular dystrophy (FSHD) is one of the most common genetically inherited myopathies in adults. It is characterized by incomplete penetrance and variable expressivity. Typically, FSHD patients display asymmetric weakness of facial, scapular, and humeral muscles that may progress to other muscle groups, particularly the abdominal and lower limb muscles. Early-onset patients display more severe muscle weakness and atrophy, resulting in a higher frequency of associated skeletal abnormalities. In these patients, multisystem involvement, including respiratory, ocular, and auditory, is more frequent and severe and may include the central nervous system. Adult-onset FSHD patients may also display some degree of multisystem involvement which mainly remains subclinical. In 95% of cases, FSHD patients carry a pathogenic contraction of the D4Z4 repeat units (RUs) in the subtelomeric region of chromosome 4 (4q35), which leads to the expression of DUX4 retrogene, toxic for muscles (FSHD1). Five percent of patients display the same clinical phenotype in association with a mutation in the SMCHD1 gene located in chromosome 18, inducing epigenetic modifications of the 4q D4Z4 repeated region and expression of DUX4 retrogene. This review highlights the complexities and challenges of diagnosing and managing FSHD, underscoring the importance of standardized approaches for optimal patient outcomes. It emphasizes the critical role of multidisciplinary care in addressing the diverse manifestations of FSHD across different age groups, from skeletal abnormalities in early-onset cases to the often-subclinical multisystem involvement in adults. With no current cure, the focus on alleviating symptoms and slowing disease progression through coordinated care is paramount.

Muscle diffusion tensor imaging in facioscapulohumeral muscular dystrophy.

INTRODUCTION/AIMS: Muscle diffusion tensor imaging has not yet been explored in facioscapulohumeral muscular dystrophy (FSHD). We assessed diffusivity parameters in FSHD subjects compared with healthy controls (HCs), with regard to their ability to precede any fat replacement or edema. METHODS: Fat fraction (FF), water T2 (wT2), mean, radial, axial diffusivity (MD, RD, AD), and fractional anisotropy (FA) of thigh muscles were calculated in 10 FSHD subjects and 15 HCs. All parameters were compared between FSHD and controls, also exploring their gradient along the main axis of the muscle. Diffusivity parameters were tested in a subgroup analysis as predictors of disease involvement in muscle compartments with different degrees of FF and wT2 and were also correlated with clinical severity scores. RESULTS: We found that MD, RD, and AD were significantly lower in FSHD subjects than in controls, whereas we failed to find a difference for FA. In contrast, we found a significant positive correlation between FF and FA and a negative correlation between MD, RD, and AD and FF. No correlation was found with wT2. In our subgroup analysis we found that muscle compartments with no significant fat replacement or edema (FF < 10% and wT2 < 41 ms) showed a reduced AD and FA compared with controls. Less involved compartments showed different diffusivity parameters than more involved compartments. DISCUSSION: Our exploratory study was able to demonstrate diffusivity parameter abnormalities even in muscles with no significant fat replacement or edema. Larger cohorts are needed to confirm these preliminary findings.

Clinical Application of Optical Genome Mapping for Molecular Diagnosis of Facioscapulohumeral Muscular Dystrophy.

Background: Facioscapulohumeral muscular dystrophy (FSHD) is a common form of muscular dystrophy that mainly affects skeletal muscle. FSHD1 accounts for 95% of all FSHD cases and can be diagnosed based on the pathogenic contraction of the D4Z4-repeat array on chromosome 4q35. Genetic diagnosis of FSHD1 is challenging because of the large size and repetitive nature of the D4Z4 region. We evaluated the clinical applicability of optical genome mapping (OGM) for the genetic diagnosis of FSHD1. Methods: We included 25 individuals with clinically confirmed or suspected/probable FSHD and their families. Ultra-high-molecular-weight DNA from peripheral blood was labeled, stained, and imaged using a single-molecule OGM platform (Bionano Genomics Saphyr system). D4Z4 repeat size and haplotype information were analyzed using the manufacturer's dedicated pipeline. We also compared the workflow and test time between Southern blot analysis and OGM. Results: We obtained concordant OGM and Southern blot results with 10 samples from patients with clinically confirmed FSHD. The D4Z4 repeat size differed within 1 unit between the Southern blot analysis and OGM. Among nine patients with clinically suspected or probable FSHD, six patients were confirmed to have pathogenic contractions by OGM. In our cohort, one de novo mosaic FSHD1 patient was successfully diagnosed with OGM. Moreover, OGM has a more straightforward and less time-consuming workflow than Southern blot analysis. Conclusions: OGM enables accurate and reliable detection of pathogenic contraction of the D4Z4-repeat array and is a valuable tool for the genetic diagnosis of FSHD1.

Scapulothoracic tenodesis using hamstring tendon graft for treatment of problematic scapula winging: A new surgical technique.

Introduction: Winging of the scapula occurs due to dysfunction of its stabilising muscles, most commonly serratus anterior and/or trapezius, for example in facioscapulohumeral muscular dystrophy. Resultant loss of scapular control and abnormal kinematics can decrease shoulder function due to glenohumeral joint instability, loss of range of motion and pain. Previously described treatment for cases resistant to physiotherapy includes scapulothoracic arthrodesis which involves risk of non-union and metalwork failure, as well as reduced respiratory function due to immobilisation of a segment of the adjacent chest wall. Technique: We present a novel surgical approach to the management of problematic scapular winging by using hamstring graft to achieve a scapulothoracic tenodesis. Discussion: We believe this technique provides an adequately stable scapula for improved shoulder movement and function, a sufficiently mobile chest wall for improved lung function and avoidance of complications specifically associated with arthrodesis.

Comprehensive genetic analysis of facioscapulohumeral muscular dystrophy by Nanopore long-read whole-genome sequencing.

BACKGROUND: Facioscapulohumeral muscular dystrophy (FSHD) is a high-prevalence autosomal dominant neuromuscular disease characterized by significant clinical and genetic heterogeneity. Genetic diagnosis of FSHD remains a challenge because it cannot be detected by standard sequencing methods and requires a complex diagnosis workflow. METHODS: We developed a comprehensive genetic FSHD detection method based on Oxford Nanopore Technologies (ONT) whole-genome sequencing. Using a case-control design, we applied this procedure to 29 samples and compared the results with those from optical genome mapping (OGM), bisulfite sequencing (BSS), and whole-exome sequencing (WES). RESULTS: Using our ONT-based method, we identified 59 haplotypes (35 4qA and 24 4qB) among the 29 samples (including a mosaic sample), as well as the number of D4Z4 repeat units (RUs). The pathogenetic D4Z4 RU contraction identified by our ONT-based method showed 100% concordance with OGM results. The methylation levels of the most distal D4Z4 RU and the double homeobox 4 gene (DUX4) detected by ONT sequencing are highly consistent with the BSS results and showed excellent diagnostic efficiency. Additionally, our ONT-based method provided an independent methylation profile analysis of two permissive 4qA alleles, reflecting a more accurate scenario than traditional BSS. The ONT-based method detected 17 variations in three FSHD2-related genes from nine samples, showing 100% concordance with WES. CONCLUSIONS: Our ONT-based FSHD detection method is a comprehensive method for identifying pathogenetic D4Z4 RU contractions, methylation level alterations, allele-specific methylation of two 4qA haplotypes, and variations in FSHD2-related genes, which will all greatly improve genetic testing for FSHD.

Best practice guidelines on genetic diagnostics of facioscapulohumeral muscular dystrophy: Update of the 2012 guidelines.

The gold standard for facioscapulohumeral muscular dystrophy (FSHD) genetic diagnostic procedures was published in 2012. With the increasing complexity of the genetics of FSHD1 and 2, the increase of genetic testing centers, and the start of clinical trials for FSHD, it is crucial to provide an update on our knowledge of the genetic features of the FSHD loci and renew the international consensus on the molecular testing recommendations. To this end, members of the FSHD European Trial Network summarized the evidence presented during the 2022 ENMC meeting on Genetic diagnosis, clinical outcome measures, and biomarkers. The working group additionally invited genetic and clinical experts from the USA, India, Japan, Australia, South-Africa, and Brazil to provide a global perspective. Six virtual meetings were organized to reach consensus on the minimal requirements for genetic confirmation of FSHD1 and FSHD2. Here, we present the clinical and genetic features of FSHD, specific features of FSHD1 and FSHD2, pros and cons of established and new technologies (Southern blot in combination with either linear or pulsed-field gel electrophoresis, molecular combing, optical genome mapping, FSHD2 methylation analysis and FSHD2 genotyping), the possibilities and challenges of prenatal testing, including pre-implantation genetic testing, and the minimal requirements and recommendations for genetic confirmation of FSHD1 and FSHD2. This consensus is expected to contribute to current clinical management and trial-readiness for FSHD.

Muscle strength, quantity and quality and muscle fat quantity and their association with oxidative stress in patients with facioscapulohumeral muscular dystrophy: Effect of antioxidant supplementation.

The purpose of this study was to identify causes of quadriceps muscle weakness in facioscapulohumeral muscular dystrophy (FSHD). To this aim, we evaluated quadriceps muscle and fat volumes by magnetic resonance imaging and their relationships with muscle strength and oxidative stress markers in adult patients with FSHD (n = 32) and healthy controls (n = 7), and the effect of antioxidant supplementation in 20 of the 32 patients with FSHD (n = 10 supplementation and n = 10 placebo) (NCT01596803). Compared with healthy controls, the dominant quadriceps strength and quality (muscle strength per unit of muscle volume) were decreased in patients with FSHD. In addition, fat volume was increased, without changes in total muscle volume. Moreover, in patients with FSHD, the lower strength of the non-dominant quadriceps was associated with lower muscle quality compared with the dominant muscle. Antioxidant supplementation significantly changed muscle and fat volumes in the non-dominant quadriceps, and muscle quality in the dominant quadriceps. This was associated with improved muscle strength (both quadriceps) and antioxidant response. These findings suggest that quadriceps muscle strength decline may not be simply explained by atrophy and may be influenced also by the muscle intrinsic characteristics. As FSHD is associated with increased oxidative stress, supplementation might reduce oxidative stress and increase antioxidant defenses, promoting changes in muscle function.

The complementary use of muscle ultrasound and MRI in FSHD: Early versus later disease stage follow-up.

OBJECTIVES: Muscle MRI and ultrasound provide complementary techniques for characterizing muscle changes and tracking disease progression in facioscapulohumeral muscular dystrophy (FSHD). In this cohort study, we provide longitudinal data that compares both imaging modalities head-to-head. METHODS: FSHD patients were assessed at baseline and after five years. Standardized muscle MRI and ultrasound images of five leg muscles were assessed bilaterally. Fat replacement was quantified using MRI fat-fraction (FF) and ultrasound Heckmatt and echogenicity z-scores (EZ-score). Muscle edema was evaluated using T2-weighted turbo inversion recovery magnitude (TIRM) MRI. RESULTS: Twenty FSHD patients were included. Muscles with normal baseline imaging showed increases in ultrasound EZ-scores (≥1; in 17%) more often than MRI FF increases (≥10%; in 7%) over time. Muscles with only baseline ultrasound abnormalities often showed considerable FF increases (in 22%), and TIRM positivity at follow-up (44%). Muscles with increased FF at baseline showed stable (80%) or increasing FF (20%) over time. EZ-scores of those muscles either increased (23%), decreased (33%) or remained stable (44%). CONCLUSIONS: Muscle ultrasound may capture accelerated pathological muscle changes in FSHD in early disease, while muscle MRI appears better-suited to detecting and monitoring pathology in later stages. SIGNIFICANCE: Our results help establish each techniques' optimal use as imaging biomarker.

Facioscapulohumeral muscular dystrophy Health Index: Japanese translation and validation study.

PURPOSE: The Facioscapulohumeral Muscular Dystrophy Health Index (FSHD-HI) is a patient-reported outcome measure developed for patients with FSHD. This study aimed to translate the FSHD-HI into Japanese (FSHD-HI-J), evaluate cultural adaptation, and examine its psychometric properties. MATERIALS AND METHODS: We created two forward translations, integrated them into a single Japanese version, and evaluated the back-translated version of the FSHD-HI. After finalizing the translation and cultural adaptation, we conducted a survey of 66 patients with FSHD to examine the reliability and validity of the FSHD-HI-J. For psychometric evaluations, we used Cronbach's alpha to assess internal consistency, the intraclass correlation coefficient (ICC) for test-retest reliability, and assessed validity based on the associations between FSHD-HI-J, clinical variables, and quality of life measures. RESULTS: The FSHD-HI-J was found to be clinically relevant, indicating high internal consistency and test-retest reliability (ICC = 0.92 [95% confidence interval: 0.86-0.95] for the total score), as well as significant associations with clinical variables (D4Z4 repeats and functional impairment) and other quality of life measures (|rho| = 0.25-0.73). CONCLUSIONS: The FSHD-HI-J is a valid and reliable patient-reported outcome measure for Japanese patients with FSHD. This validated, disease-specific patient-reported outcome is essential for future clinical practice and clinical trials.

Facioscapulohumeral muscular dystrophy (FSHD) affects not only a patient's physical abilities but also their social activities, participation, and overall quality of life.The FSHD-Health Index (FSHD-HI) is an instrument developed as a disease-specific patient-reported outcome measure to evaluate the burden experienced by patients.The Japanese version of the FSHD-HI has been established as a reliable and validated measure for Japanese-speaking patients with FSHD.The Japanese version of the FSHD-HI can serve as a useful instrument for evaluating the effectiveness of interventions in future trials.

First-trimester noninvasive prenatal diagnosis of seven facioscapulohumeral muscular dystrophy type 1 families using SNP-based amplicon sequencing: An earlier, rapid and safer way.

The study is to explore the feasibility and value of SNP-based noninvasive prenatal diagnosis (NIPD) for facioscapulohumeral muscular dystrophy type 1 (FSHD1) in early pregnancy weeks. We prospectively collected seven FSHD1 families, with an average gestational age of 8+6. Among these seven couples, there were three affected FSHD1 mothers and four affected fathers. A multiplex-PCR panel comprising 402 amplicons was designed to selective enrich for highly heterozygous SNPs upstream of the DUX4 gene. Risk haplotype was constructed based on familial linkage analysis. Fetal genotypes were accurately inferred through relative haplotype dosage analysis using Bayes Factor. All tests were successfully completed in a single attempt, and no recombination events were detected. NIPD results were provided within a week, which is 4 weeks earlier than karyomapping and 7 weeks earlier than Bionano single-molecule optical mapping (BOM). Ultimately, five FSHD1 fetuses and two normal fetuses were successfully identified, with a 100% concordance rate with karyomapping and BOM. Therefore, SNP-based NIPD for FSHD1 was demonstrated to be feasible and accurate in early weeks of gestation, although the risk of recombination events cannot be completely eliminated. In the future, testing of more cases is still necessary to fully determine the clinical utility.

Case report: Identification of facioscapulohumeral muscular dystrophy 1 in two siblings with normal phenotypic parents using optical genome mapping.

Objective: Facioscapulohumeral muscular dystrophy type 1 (FSHD1) is one of the most common forms of autosomal-dominant muscular dystrophies characterized by variable disease penetrance due to shortened D4Z4 repeat units on 4q35. The molecular diagnosis of FSHD1 is usually made by Southern blotting, which is complex, time-consuming, and lacks clinical practicality. Therefore, in this study, optical genome mapping (OGM) is employed for the genetic diagnosis of FSHD1. Furthermore, epigenetic heterogeneity is determined from methylation analysis. Methods: Genomic DNA samples from four members of the same family were subjected to whole-exome sequencing. OGM was used to identify structural variations in D4Z4, while sodium bisulfite sequencing helped identify the methylation levels of CpG sites in a region located distally to the D4Z4 array. A multidisciplinary team collected the clinical data, and comprehensive family analyses aided in the assessment of phenotypes and genotypes. Results: Whole-exome sequencing did not reveal variants related to clinical phenotypes in the patients. OGM showed that the proband was a compound heterozygote for the 4qA allele with four and eight D4Z4 repeat units, whereas the affected younger brother had only one 4qA allele with four D4Z4 repeat units. Both the proband and her younger brother were found to display asymmetric weakness predominantly involving the facial, shoulder girdle, and upper arm muscles, whereas the younger brother had more severe clinical symptoms. The proband's father, who was found to be normal after a neurological examination, also carried the 4qA allele with eight D4Z4 repeat units. The unaffected mother exhibited 49 D4Z4 repeat units of the 4qA allele and a minor mosaic pattern with four D4Z4 repeat units of the 4qA allele. Consequently, the presence of the 4qA allele in the four D4Z4 repeat units strongly pointed to the occurrence of maternal germline mosaicism. The CpG6 methylation levels were lower in symptomatic patients compared to those in the asymptomatic parents. The older sister had lower clinical scores and ACSS and higher CpG6 methylation levels than that of her younger brother. Conclusions: In this study, two siblings with FSHD1 with phenotypically normal parents were identified by OGM. Our findings suggest that the 4qA allele of four D4Z4 repeats was inherited through maternal germline mosaicism. The clinical phenotype heterogeneity is influenced by the CpG6 methylation levels. The results of this study greatly aid in the molecular diagnosis of FSHD1 and in also understanding the clinical phenotypic variability underlying the disease.

Combined Lumbar-Sacral Plexus Block in Facioscapulohumeral Muscular Dystrophy for Hip Fracture Surgery: A Case Report.

Facioscapulohumeral muscular dystrophy (FSHD) is a muscular dystrophy that can affect individuals of all age groups. Its prevalence is reported to be 0.4-1 in 10,000 people. Because of the low occurrence of FSHD, anaesthetic management is primarily based on expert opinions, case reviews, or brief series. Here, we present the case of a 72-year-old woman with FSHD who underwent hip fracture (HF) surgery. To prevent respiratory compromise due to FSHD, we opted for lumbar-sacral plexus block. To the best of our knowledge, there is no information in the literature regarding the use of combined lumbar-sacral plexus block in patients with FSHD undergoing HF surgery.

IL-6 and TNF are Potential Inflammatory Biomarkers in Facioscapulohumeral Muscular Dystrophy.

Background: FSHD is a highly prevalent inherited myopathy with a still poorly understood pathology. Objective: To investigate whether proinflammatory cytokines are associated with FSHD and which specific innate immune cells are involved in its pathology. Methods: First, we measured circulating cytokines in serum samples: IL-6 (FSHD, n = 150; HC, n = 98); TNF (FSHD, n = 150; HC, n = 59); IL-1α (FSHD, n = 150; HC, n = 66); IL-1ß (FSHD, n = 150; HC, n = 98); MCP-1 (FSHD, n = 14; HC, n = 14); VEGF-A (FSHD, n = 14; HC, n = 14). Second, we tested trained immunity in monocytes (FSHD, n = 15; HC, n = 15) and NK cells (FSHD, n = 11; HC, n = 11). Next, we explored the cytokine production capacity of NK cells in response to different stimuli (FSHD, n = 39; HC, n = 22). Lastly, we evaluated the cytokine production of ex vivo stimulated MRI guided inflamed (TIRM+) and paired MRI guided non inflamed (TIRM-) muscle biopsies of 21 patients and of 8 HC muscle biopsies. Results: We included a total of 190 FSHD patients (N = 190, 48±14 years, 49% men) and of 135 HC (N = 135, 44±15 years, 47% men). We found that FSHD patients had higher concentrations of IL-6 and TNF measured (a) in the circulation, (b) after ex-vivo stimulation of NK cells, and (c) in muscle specimens. Besides, IL-6 circulating concentrations, as well as its production by NK cells and IL-6 content of FSHD muscle specimens, showed a mild correlation with disease duration, disease severity, and muscle weakness. Conclusion: These results show that IL-6 and TNF may contribute to FSHD pathology and suggest novel therapeutic targets. Additionally, the activation of NK cells in FSHD may be a novel pathway contributing to FSHD pathology.

Living with Dysphagia: A Survey Exploring the Experiences of Adults Living with Neuromuscular Disease and their Caregivers in the United Kingdom.

Background: Dysphagia is common in adults living with neuromuscular disease (NMD). Increased life expectancy, secondary to improvements in standards of care, requires the recognition and treatment of dysphagia with an increased priority. Evidence to support the establishment of healthcare pathways is, however, lacking. The experiences of people living with NMD (pplwNMD) and their caregivers are valuable to guide targeted, value-based healthcare. Objective: To generate preliminary considerations for neuromuscular dysphagia care and future research in the United Kingdom, based on the experiences of those living with, or caring for, people with NMD. Methods: Two surveys (one for adults living with NMD and dysphagia, and a second for caregivers) were co-designed with an advisory group of people living with NMD. Surveys were electronically distributed to adults living with NMD and their caregivers between 18th May and 26th July 2020. Distribution was through UK disease registries, charity websites, newsletters, and social media. Results: Adults living with NMD receive little information or education that they are likely to develop swallowing difficulties. Most respondents report wanting this information prior to developing these difficulties. Difficulties with swallowing food and medication are common in this group, and instrumental assessment is considered a helpful assessment tool. Both adults living with NMD and caregivers want earlier access to neuromuscular swallowing specialists and training in how best to manage their difficulties. Conclusions: Improvement is needed in the dysphagia healthcare pathway for adults living with NMD to help mitigate any profound physical and psychological consequences that may be caused by dysphagia. Education about swallowing difficulties and early referral to a neuromuscular swallowing specialist are important to pplwNMD and their caregivers. Further research is required to better understand the experiences of pplwNMD and their caregivers to inform the development of dysphagia healthcare pathways.

Facioscapulohumeral Muscular Dystrophy European Patient Survey: Assessing Patient Reported Disease Burden and Preferences in Clinical Trial Participation.

Background: Facioscapulohumeral muscular dystrophy (FSHD) is a genetic disorder characterized by progressive muscle weakness leading to permanent disability. There are no curative treatments, however, there are several upcoming clinical trials testing new therapies in FSHD. Objective: This study aimed to explore the disease burden and patient preferences of people with FSHD to ensure that clinical trials can be designed to include outcome measures that are relevant and important to patients. Methods: A survey was developed with a steering committee clinicians and physiotherapists with relevant experience in the disease, patient representatives, a registry expert and industry consultants. Themes of the survey included; participant demographics, disease progression and impact on function, factors encouraging or discouraging clinical trial participation, and positive outcomes of a clinical trial. Results: 1147 participants responded to the online survey, representing 26 countries across Europe and a range of disease severities. The study highlighted the key symptoms causing concern for FSHD patients - muscle weakness and mobility issues - reflecting what participants want targeted for future therapies. The need for clear information and communication throughout clinical trials was emphasised. Factors most encouraging trial participation included access to new investigational therapies, access to trial results and benefits for the FSHD community. Factors most discouraging trial participation included travel related issues and fear of side effects. Conclusions: The results from this study identify the patient reported burden of FSHD and should provide researchers and industry with areas of therapeutic research that would be meaningful to patients, as well as supporting the development of patient centric outcome measures in clinical trials.

The lived experience of facioscapulohumeral muscular dystrophy: A systematic review and synthesis of the qualitative literature.

INTRODUCTION: In this review we sought to characterize the lived experience of people living with FSHD (pwFSHD) to help clinicians to orient their services to the needs of these individuals. METHODS: Five electronic databases were systematically searched for qualitative research studies containing quotations from pwFSHD. ENhancing Transparency in REporting the Synthesis of Qualitative research and Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines informed the methodology. Study quality was assessed using the Critical Appraisal Skills Programme Checklist tool, which measures the methodological quality of qualitative research. Data extracted from included studies were analyzed using thematic synthesis. RESULTS: Ninety-nine pwFSHD took part in the six studies included in this review - from research teams based in two countries. Five descriptive themes emerged: "Engaging with life as symptoms progress"; "The emotional journey"; "A family burden to bear"; "Social connection and disconnection"; and "Tension between visibility and invisibility." From these, two analytical themes were derived: "The emotional challenge of continuing and intensifying adaptation" and "The relational burden of rare disease." DISCUSSION: The lived experience of pwFSHD is characterized by physical, emotional, and social challenges that impact on engagement with life, particularly as symptoms progress. Further research is needed to provide a fuller understanding of the experience of pain in FSHD and of the lived experience of FSHD across cultures.

Autosomal dominant in cis D4Z4 repeat array duplication alleles in facioscapulohumeral dystrophy.

Facioscapulohumeral dystrophy (FSHD) has a unique genetic aetiology resulting in partial chromatin relaxation of the D4Z4 macrosatellite repeat array on 4qter. This D4Z4 chromatin relaxation facilitates inappropriate expression of the transcription factor DUX4 in skeletal muscle. DUX4 is encoded by a retrogene that is embedded within the distal region of the D4Z4 repeat array. In the European population, the D4Z4 repeat array is usually organized in a single array that ranges between 8 and 100 units. D4Z4 chromatin relaxation and DUX4 derepression in FSHD is most often caused by repeat array contraction to 1-10 units (FSHD1) or by a digenic mechanism requiring pathogenic variants in a D4Z4 chromatin repressor like SMCHD1, combined with a repeat array between 8 and 20 units (FSHD2). With a prevalence of 1.5% in the European population, in cis duplications of the D4Z4 repeat array, where two adjacent D4Z4 arrays are interrupted by a spacer sequence, are relatively common but their relationship to FSHD is not well understood. In cis duplication alleles were shown to be pathogenic in FSHD2 patients; however, there is inconsistent evidence for the necessity of an SMCHD1 mutation for disease development. To explore the pathogenic nature of these alleles we compared in cis duplication alleles in FSHD patients with or without pathogenic SMCHD1 variant. For both groups we showed duplication-allele-specific DUX4 expression. We studied these alleles in detail using pulsed-field gel electrophoresis-based Southern blotting and molecular combing, emphasizing the challenges in the characterization of these rearrangements. Nanopore sequencing was instrumental to study the composition and methylation of the duplicated D4Z4 repeat arrays and to identify the breakpoints and the spacer sequence between the arrays. By comparing the composition of the D4Z4 repeat array of in cis duplication alleles in both groups, we found that specific combinations of proximal and distal repeat array sizes determine their pathogenicity. Supported by our algorithm to predict pathogenicity, diagnostic laboratories should now be furnished to accurately interpret these in cis D4Z4 repeat array duplications, alleles that can easily be missed in routine settings.