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Rationale: In cystic fibrosis (CF), the lung clearance index (LCI), derived from multiple breath washout (MBW), is more sensitive in detecting early lung disease than FEV(1); MBW has been less thoroughly evaluated in young patients with primary ciliary dyskinesia (PCD). Objectives: Our objectives were 1) to evaluate the sensitivity of MBW and spirometry for the detection of mild lung disease in young children with PCD and CF compared with healthy control (HC) subjects and 2) to compare patterns of airway obstruction between disease populations. Methods: We used a multicenter, single-visit, observational study in children with PCD and CF with a forced expiratory volume in 1 second (FEV(1)) greater than 60% predicted and HC subjects, ages 3–12 years. Nitrogen MBW and spirometry were performed and overread for acceptability. χ(2) and Kruskall-Wallis tests compared demographics and lung function measures between groups, linear regression evaluated the effect of disease state, and Spearman’s rank correlation coefficient compared the LCI and spirometric measurements. Results: Twenty-five children with PCD, 49 children with CF, and 80 HC children were enrolled, among whom 17 children with PCD (68%), 36 children with CF (73%), and 53 (66%) HC children performed both acceptable spirometry and MBW; these children made up the analytic cohort. The median age was 9.0 years (interquartile range [IQR], 6.8–11.1). The LCI was abnormal (more than 7.8) in 10 of 17 (59%) patients with PCD and 21 of 36 (58%) patients with CF, whereas FEV(1) was abnormal in three of 17 (18%) patients with PCD and six of 36 (17%) patients with CF. The LCI was significantly elevated in patients with PCD and CF compared with HC subjects (ratio of geometric mean vs. HC: PCD 1.27; 95% confidence interval [CI], 1.15–1.39; and CF 1.24; 95% CI, 1.15–1.33]). Children with PCD had lower midexpiratory-phase forced expiratory flow % predicted compared with children with CF (62% [IQR, 50–78%] vs. 85% [IQR, 68–99%]; P = 0.05). LCI did not correlate with FEV(1). Conclusions: The LCI is more sensitive than FEV(1) in detecting lung disease in young patients with PCD, similar to CF. LCI holds promise as a sensitive endpoint for the assessment of early PCD lung disease.

Background & Aims Serum microRNA (miRNA) levels are known to change in non-alcoholic fatty liver disease (NAFLD) and may serve as useful biomarkers. This study aimed to profile miRNAs comprehensively at all NAFLD stages. Methods We profiled 2,083 serum miRNAs in a discovery cohort (183 cases with NAFLD representing the complete NAFLD spectrum and 10 population controls). miRNA libraries generated by HTG EdgeSeq were sequenced by Illumina NextSeq. Selected serum miRNAs were profiled in 372 additional cases with NAFLD and 15 population controls by quantitative reverse transcriptase PCR. Results Levels of 275 miRNAs differed between cases and population controls. Fewer differences were seen within individual NAFLD stages, but miR-193a-5p consistently showed increased levels in all comparisons. Relative to NAFL/non-alcoholic steatohepatitis (NASH) with mild fibrosis (stage 0/1), 3 miRNAs (miR-193a-5p, miR-378d, and miR378d) were increased in cases with NASH and clinically significant fibrosis (stages 2–4), 7 (miR193a-5p, miR-378d, miR-378e, miR-320b, miR-320c, miR-320d, and miR-320e) increased in cases with NAFLD activity score (NAS) 5–8 compared with lower NAS, and 3 (miR-193a-5p, miR-378d, and miR-378e) increased but 1 (miR-19b-3p) decreased in steatosis, activity, and fibrosis (SAF) activity score 2–4 compared with lower SAF activity. The significant findings for miR-193a-5p were replicated in the additional cohort with NAFLD. Studies in Hep G2 cells showed that following palmitic acid treatment, miR-193a-5p expression decreased significantly. Gene targets for miR-193a-5p were investigated in liver RNAseq data for a case subgroup (n = 80); liver GPX8 levels correlated positively with serum miR-193a-5p. Conclusions Serum miR-193a-5p levels correlate strongly with NAFLD activity grade and fibrosis stage. MiR-193a-5p may have a role in the hepatic response to oxidative stress and is a potential clinically tractable circulating biomarker for progressive NAFLD. Lay summary MicroRNAs (miRNAs) are small pieces of nucleic acid that may turn expression of genes on or off. These molecules can be detected in the blood circulation, and their levels in blood may change in liver disease including non-alcoholic fatty liver disease (NAFLD). To see if we could detect specific miRNA associated with advanced stages of NAFLD, we carried out miRNA sequencing in a group of 183 patients with NAFLD of varying severity together with 10 population controls. We found that a number of miRNAs showed changes, mainly increases, in serum levels but that 1 particular miRNA miR-193a-5p consistently increased. We confirmed this increase in a second group of cases with NAFLD. Measuring this miRNA in a blood sample may be a useful way to determine whether a patient has advanced NAFLD without an invasive liver biopsy., Graphical abstract, Highlights • Serum miRNA was sequenced in 183 NAFLD cases of varying severity and 10 population controls. • Plasma levels of miR-193a-5p were significantly increased in patients with advanced fibrosis, high NAS scores, or high SAF scores. • Other miRNAs including miR378d and miR378e were also significantly increased in certain comparisons. • The findings for miR-193a-5p were replicated in a cohort of 372 additional NAFLD cases.

Objective The goal of this study was to reduce treatment resistance and enhance antidepressant response in patients with treatment-resistant bipolar depression (TRBDD) by modulating inflammatory activation. Methods Forty-seven TRBDD patients completed a randomized, double-blind, placebo (PBO)-controlled two-arm study using celecoxib (CBX), a cyclooxygenase 2 (COX-2) inhibitor, in combination with escitalopram (ESC). The Hamilton Depression (17-Item) and the Hamilton Anxiety Rating Scales were used to quantify symptom severity. Self-rating instruments included BDI, BAI and QLES-Q questioner. An adverse events inventory was used, and the possibility of bleeding diathesis was monitored. Complete blood count (CBC), prothrombin time (PT), and activated partial thromboplastin time (APTT) were determined. Comparison of mood scores between the CBX and PBO groups were conducted using a mixed ANOVA and an Independent Sample Student t-test. Results The CBX adjunctive treatment produced significantly more responders and remitters than the PBO arm. Compared to the PBO group (n = 20), the CBX group (n = 27) experienced lower depression severity through the entire course of the study, showing significant decrease in depression and anxiety scores as early as week 1. Except for initial mild nausea, no adverse events were reported and study medications were well tolerated. Conclusions Modulation of the inflammatory response through targeted inhibition of the enzyme COX-2 by means of CBX reduces TRBDD and augments and accelerates treatment response in an efficacious and safe manner. Future studies should replicate these findings and additionally investigate whether prolonged administration of CBX is required to maintain remission by adding a discontinuation phase to the study design. Trial Registration Clinicaltrials.gov ID: NCT01479829.

OBJECTIVE: The Popeye domain containing 3 (POPDC3) gene encodes a membrane protein involved in cyclic adenosine monophosphate (cAMP) signaling. Besides gastric cancer, no disease association has been described. We describe a new muscular dystrophy associated with this gene.METHODS: We screened 1,500 patients with unclassified limb girdle weakness or hyperCKemia for pathogenic POPDC3 variants. Five patients carrying POPDC3 variants were examined by muscle magnetic resonance imaging (MRI), muscle biopsy, and cardiac examination. We performed functional analyses in a zebrafish popdc3 knockdown model and heterologous expression of the mutant proteins in Xenopus laevis oocytes to measure TREK-1 current.RESULTS: We identified homozygous POPDC3 missense variants (p.Leu155His, p.Leu217Phe, and p.Arg261Gln) in 5 patients from 3 ethnically distinct families. Variants affected highly conserved residues in the Popeye (p.Leu155 and p.Leu217) and carboxy-terminal (p.Arg261) domains. The variants were almost absent from control populations. Probands' muscle biopsies were dystrophic, and serum creatine kinase levels were 1,050 to 9,200U/l. Muscle weakness was proximal with adulthood onset in most patients and affected lower earlier than upper limbs. Muscle MRI revealed fat replacement of paraspinal and proximal leg muscles; cardiac investigations were unremarkable. Knockdown of popdc3 in zebrafish, using 2 different splice-site blocking morpholinos, resulted in larvae with tail curling and dystrophic muscle features. All 3 mutants cloned in Xenopus oocytes caused an aberrant modulation of the mechano-gated potassium channel, TREK-1.INTERPRETATION: Our findings point to an important role of POPDC3 for skeletal muscle function and suggest that pathogenic variants in POPDC3 are responsible for a novel type of autosomal recessive limb girdle muscular dystrophy. ANN NEUROL 2019;86:832-843.

International audience; The mechanisms that drive nonalcoholic fatty liver disease (NAFLD) remain incompletely understood. This large multicenter study characterized the transcriptional changes that occur in liver tissue across the NAFLD spectrum as disease progresses to cirrhosis to identify potential circulating markers. We performed high-throughput RNA sequencing on a discovery cohort comprising histologically characterized NAFLD samples from 206 patients. Unsupervised clustering stratified NAFLD on the basis of disease activity and fibrosis stage with differences in age, aspartate aminotransferase (AST), type 2 diabetes mellitus, and carriage of PNPLA3 rs738409 , a genetic variant associated with NAFLD. Relative to early disease, we consistently identified 25 differentially expressed genes as fibrosing steatohepatitis progressed through stages F2 to F4. This 25-gene signature was independently validated by logistic modeling in a separate replication cohort ( n = 175), and an integrative analysis with publicly available single-cell RNA sequencing data elucidated the likely relative contribution of specific intrahepatic cell populations. Translating these findings to the protein level, SomaScan analysis in more than 300 NAFLD serum samples confirmed that circulating concentrations of proteins AKR1B10 and GDF15 were strongly associated with disease activity and fibrosis stage. Supporting the biological plausibility of these data, in vitro functional studies determined that endoplasmic reticulum stress up-regulated expression of AKR1B10 , GDF15 , and PDGFA , whereas GDF15 supplementation tempered the inflammatory response in macrophages upon lipid loading and lipopolysaccharide stimulation. This study provides insights into the pathophysiology of progressive fibrosing steatohepatitis, and proof of principle that transcriptomic changes represent potentially tractable and clinically relevant markers of disease progression.

Mutations in the cytoplasmic dynein 1 heavy chain gene (DYNC1H1) have been identified in rare neuromuscular (NMD) and neurodevelopmental (NDD) disorders such as spinal muscular atrophy with lower extremity dominance (SMALED) and autosomal dominant mental retardation syndrome 13 (MRD13). Phenotypes and genotypes of ten pediatric patients with pathogenic DYNC1H1 variants were analyzed in a multi-center study. Data mining of large-scale genomic variant databases was used to investigate domain-specific vulnerability and conservation of DYNC1H1. We identified ten patients with nine novel mutations in the DYNC1H1 gene. These patients exhibit a broad spectrum of clinical findings, suggesting an overlapping disease manifestation with intermixed phenotypes ranging from neuropathy (peripheral nervous system, PNS) to severe intellectual disability (central nervous system, CNS). Genomic profiling of healthy and patient variant datasets underlines the domain-specific effects of genetic variation in DYNC1H1, specifically on toleration towards missense variants in the linker domain. A retrospective analysis of all published mutations revealed domain-specific genotype–phenotype correlations, i.e., mutations in the dimerization domain with reductions in lower limb strength in DYNC1H1–NMD and motor domain with cerebral malformations in DYNC1H1–NDD. We highlight that the current classification into distinct disease entities does not sufficiently reflect the clinical disease manifestation that clinicians face in the diagnostic work-up of DYNC1H1-related disorders. We propose a novel clinical classification for DYNC1H1-related disorders encompassing a spectrum from DYNC1H1–NMD with an exclusive PNS phenotype to DYNC1H1–NDD with concomitant CNS involvement.

BackgroundIntellectual disability syndromes (IDSs) with or without developmental delays affect up to 3% of the world population. We sought to clinically and genetically characterise a novel IDS segregating in five unrelated consanguineous families.MethodsClinical analyses were performed for eight patients with intellectual disability (ID). Whole-exome sequencing for selected participants followed by Sanger sequencing for all available family members was completed. Identity-by-descent (IBD) mapping was carried out for patients in two Egyptian families harbouring an identical variant. RNA was extracted from blood cells of Turkish participants, followed by cDNA synthesis and real-time PCR for TTC5.ResultsPhenotype comparisons of patients revealed shared clinical features of moderate-to-severe ID, corpus callosum agenesis, mild ventriculomegaly, simplified gyral pattern, cerebral atrophy, delayed motor and verbal milestones and hypotonia, presenting with an IDS. Four novel homozygous variants in TTC5: c.629A>G;p.(Tyr210Cys), c.692C>T;p.(Ala231Val), c.787C>T;p.(Arg263Ter) and c.1883C>T;p.(Arg395Ter) were identified in the eight patients from participating families. IBD mapping revealed that c.787C>T;p.(Arg263Ter) is a founder variant in Egypt. Missense variants c.629A>G;p.(Tyr210Cys) and c.692C>T;p.(Ala231Val) disrupt highly conserved residues of TTC5 within the fifth and sixth tetratricopeptide repeat motifs which are required for p300 interaction, while the nonsense variants are predicted to decrease TTC5 expression. Functional analysis of variant c.1883C>T;p.(Arg395Ter) showed reduced TTC5 transcript levels in accordance with nonsense-mediated decay.ConclusionCombining our clinical and molecular data with a recent case report, we identify the core and variable clinical features associated with TTC5 loss-of-function variants and reveal the requirement for TTC5 in human brain development and health.

Protein O-glucosyltransferase 1 (POGLUT1) activity is critical for the Notch signaling pathway, being one of the main enzymes responsible for the glycosylation of the extracellular domain of Notch receptors. A biallelic mutation in the POGLUT1 gene has been reported in one family as the cause of an adult-onset limb-girdle muscular dystrophy (LGMD R21; OMIM# 617232). As the result of a collaborative international effort, we have identified the first cohort of 15 patients with LGMD R21, from nine unrelated families coming from different countries, providing a reliable phenotype–genotype and mechanistic insight. Patients carrying novel mutations in POGLUT1 all displayed a clinical picture of limb-girdle muscle weakness. However, the age at onset was broadened from adult to congenital and infantile onset. Moreover, we now report that the unique muscle imaging pattern of “inside-to-outside” fatty degeneration observed in the original cases is indeed a defining feature of POGLUT1 muscular dystrophy. Experiments on muscle biopsies from patients revealed a remarkable and consistent decrease in the level of the NOTCH1 intracellular domain, reduction of the pool of satellite cells (SC), and evidence of α-dystroglycan hypoglycosylation. In vitro biochemical and cell-based assays suggested a pathogenic role of the novel POGLUT1 mutations, leading to reduced enzymatic activity and/or protein stability. The association between the POGLUT1 variants and the muscular phenotype was established by in vivo experiments analyzing the indirect flight muscle development in transgenic Drosophila, showing that the human POGLUT1 mutations reduced its myogenic activity. In line with the well-known role of the Notch pathway in the homeostasis of SC and muscle regeneration, SC-derived myoblasts from patients’ muscle samples showed decreased proliferation and facilitated differentiation. Together, these observations suggest that alterations in SC biology caused by reduced Notch1 signaling result in muscular dystrophy in LGMD R21 patients, likely with additional contribution from α-dystroglycan hypoglycosylation. This study settles the muscular clinical phenotype linked to POGLUT1 mutations and establishes the pathogenic mechanism underlying this muscle disorder. The description of a specific imaging pattern of fatty degeneration and muscle pathology with a decrease of α-dystroglycan glycosylation provides excellent tools which will help diagnose and follow up LGMD R21 patients., This work was supported in part by the Instituto de Salud Carlos III and FEDER (FIS PI16-01843 to C. Paradas and JR15/00042 to M. Cabrera-Serrano), the Consejería de Salud, Junta de Andalucía (PI-0085-2016 and PE-S1275 to C. Paradas, and B-0005-2017 to M. Cabrera-Serrano), NIH/NIGMS (R01GM084135 and R35GM130317 to H. Jafar-Nejad, and R01GM061126 to R.S. Haltiwanger), JSPS KAKENHI Grants-in-Aid for Research Activity Start-up and Scientific Research (B) (JP17H06743 and JP19H03176 to H. Takeuchi), and Takeda Science Foundation and Daiichi Sankyo Foundation of Life Science (to H. Takeuchi). MYO-SEQ has been supported by Sanofi Genzyme, Ultragenyx, the LGMD2I Research Fund, Samantha J Brazzo Foundation, LGMD2D Foundation, Kurt + Peter Foundation, Muscular Dystrophy UK and Coalition to Cure Calpain 3. Work in CGB’s group is supported by NINDS/NIH intramural funds.

BACKGROUND: Late-onset Pompe disease is a rare genetic neuromuscular disorder caused by a primary deficiency of α-glucosidase and the associated accumulation of glycogen in lysosomal vacuoles. The deficiency of α-glucosidase can often be detected using an inexpensive and readily accessible dried blood spot test when Pompe disease is suspected. Like several neuromuscular disorders, Pompe disease typically presents with progressive weakness of limb-girdle muscles and respiratory insufficiency. Due to the phenotypic heterogeneity of these disorders, however, it is often difficult for clinicians to reach a diagnosis for patients with Pompe disease. Six hundred and six patients from a European population were recruited onto our study. Inclusion criteria stipulated that index cases must present with limb-girdle weakness or elevated serum creatine kinase activity. Whole exome sequencing with at least 250 ng DNA was completed using an Illumina exome capture and a 38 Mb baited target. A panel of 169 candidate genes for limb-girdle weakness was analysed for disease-causing variants. RESULTS: A total of 35 variants within GAA were detected. Ten distinct variants in eight unrelated index cases (and four siblings not sequenced in our study) were considered disease-causing, with the patients presenting with heterogeneous phenotypes. The eight unrelated individuals were compound heterozygotes for two variants. Six patients carried the intronic splice site c.-13 T > G transversion and two of the six patients also carried the exonic p.Glu176ArgfsTer45 frameshift. Four of the ten variants were novel in their association with Pompe disease. CONCLUSIONS: Here, we highlight the advantage of using whole exome sequencing as a tool for detecting, diagnosing and treating patients with rare, clinically variable genetic disorders. ispartof: Orphanet Journal of Rare Diseases vol:12 issue:1 pages:173-184 ispartof: location:England status: published

ObjectiveTo assess the clinical, radiologic, myopathologic, and proteomic findings in a patient manifesting a multisystem proteinopathy due to a homozygous valosin-containing protein gene (VCP) mutation previously reported to be pathogenic in the heterozygous state.MethodsWe studied a 36-year-old male index patient and his father, both presenting with progressive limb-girdle weakness. Muscle involvement was assessed by MRI and muscle biopsies. We performed whole-exome sequencing and Sanger sequencing for segregation analysis of the identified p.Arg159His VCP mutation. To dissect biological disease signatures, we applied state-of-the-art quantitative proteomics on muscle tissue of the index case, his father, 3 additional patients with VCP-related myopathy, and 3 control individuals.ResultsThe index patient, homozygous for the known p.Arg159His mutation in VCP, manifested a typical VCP-related myopathy phenotype, although with a markedly high creatine kinase value and a relatively early disease onset, and Paget disease of bone. The father exhibited a myopathy phenotype and discrete parkinsonism, and multiple deceased family members on the maternal side of the pedigree displayed a dementia, parkinsonism, or myopathy phenotype. Bioinformatic analysis of quantitative proteomic data revealed the degenerative nature of the disease, with evidence suggesting selective failure of muscle regeneration and stress granule dyshomeostasis.ConclusionWe report a patient showing a multisystem proteinopathy due to a homozygous VCP mutation. The patient manifests a severe phenotype, yet fundamental disease characteristics are preserved. Proteomic findings provide further insights into VCP-related pathomechanisms.

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

Background Extensive genetic screening results in the identification of thousands of rare variants that are difficult to interpret. Because of its sheer size, rare variants in the titin gene (TTN) are detected frequently in any individual. Unambiguous interpretation of molecular findings is almost impossible in many patients with myopathies or cardiomyopathies. Objective To refine the current classification framework for TTN-associated skeletal muscle disorders and standardize the interpretation of TTN variants. Methods We used the guidelines issued by the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) to re-analyze TTN genetic findings from our patient cohort. Results We identified in the classification guidelines three rules that are not applicable to titin-related skeletal muscle disorders; six rules that require disease-/gene-specific adjustments and four rules requiring quantitative thresholds for a proper use. In three cases, the rule strength need to be modified. Conclusions We suggest adjustments are made to the guidelines. We provide frequency thresholds to facilitate filtering of candidate causative variants and guidance for the use and interpretation of functional data and co-segregation evidence. We expect that the variant classification framework for TTN-related skeletal muscle disorders will be further improved along with a better understanding of these diseases.

Several hundred genetic muscle diseases have been described, all of which are rare. Their clinical and genetic heterogeneity means that a genetic diagnosis is challenging. We established an international consortium, MYO-SEQ, to aid the work-ups of muscle disease patients and to better understand disease etiology., info:eu-repo/semantics/published

BackgroundVariants in the Structural Maintenance of Chromosomes flexible Hinge Domain-containing protein 1 (SMCHD1) can cause facioscapulohumeral muscular dystrophy type 2 (FSHD2) and the unrelated Bosma arhinia microphthalmia syndrome (BAMS). In FSHD2, pathogenic variants are found anywhere in SMCHD1 while in BAMS, pathogenic variants are restricted to the extended ATPase domain. Irrespective of the phenotypic outcome, both FSHD2-associated and BAMS-associated SMCHD1 variants result in quantifiable local DNA hypomethylation. We compared FSHD2, BAMS and non-pathogenic SMCHD1 variants to derive genotype–phenotype relationships.MethodsExamination of SMCHD1 variants and methylation of the SMCHD1-sensitive FSHD locus DUX4 in 187 FSHD2 families, 41 patients with BAMS and in control individuals. Analysis of variants in a three-dimensional model of the ATPase domain of SMCHD1.ResultsDUX4 methylation analysis is essential to establish pathogenicity of SMCHD1 variants. Although the FSHD2 mutation spectrum includes all types of variants covering the entire SMCHD1 locus, missense variants are significantly enriched in the extended ATPase domain. Identification of recurrent variants suggests disease-specific residues for FSHD2 and in BAMS, consistent with a largely disease-specific localisation of variants in SMCHD1.ConclusionsThe localisation of missense variants within the ATPase domain of SMCHD1 may contribute to the differences in phenotypic outcome.

Introduction TRIM32-related myopathies represent a phenotypic spectrum of a rare autosomal recessive muscle disorder. The disease is described as a mild and progressive myopathy without characteristic clinical features. Originally classified as limb-girdle muscular dystrophy (LGMD) 2H (OMIM #254110), the disorder was first identified in the Hutterite population and the homozygous TRIM32 founder mutation, p.Asp487Asn, was identified as the cause of this disease.1 Only seven patients with definite non-Hutterite TRIM32-related myopathy have been reported in the literature. Apart from two missense mutations residing in the NHL repeats of TRIM32, only deletions, frameshift and nonsense mutations have been reported.2 Having applied next generation sequencing technologies to over 1000 patients with suspected genetic muscle disorders, we present nine patients with TRIM32-related myopathies and three patients with a homozygous TRIM32 variant of unknown significance (VUS). Subjects and methods DNA samples from 1000 patients with unexplained limb-girdle muscle weakness and/or elevated serum creatine kinase (CK) levels were gathered through the MYO-SEQ project. Samples were processed and whole exome sequencing (WES) performed as described previously.3 Additional patients with TRIM32-related myopathy were diagnosed by the Northern Molecular Genetics Service (NMGS) diagnostic laboratory through panel sequencing of 32 LGMD genes. Variants were classified according to ACMG guidelines.4 MRI imaging was performed for eight patients on a 1.5T MRI platform. Muscle biopsies for all patients were analysed following standard histological techniques. Results Genetic findings Of the 1000 MYO-SEQ patients, we identified 36 with rare coding variants in TRIM32 (minor allele frequency

LGMD2L is a subtype of limb-girdle muscular dystrophy (LGMD), caused by recessive mutations in ANO5, encoding anoctamin-5 (ANO5). We present the analysis of five patients with skeletal muscle weakness for whom heterozygous mutations within ANO5 were identified by whole exome sequencing (WES). Patients varied in the age of the disease onset (from 22 to 38 years) and severity of the morphological and clinical phenotypes. Out of the nine detected mutations one was novel (missense p.Lys132Met, accompanied by p.His841Asp) and one was not yet characterized in the literature (nonsense, p.Trp401Ter, accompanied by p.Asp81Gly). The p.Asp81Gly mutation was also identified in another patient carrying a p.Arg758Cys mutation as well. Also, a c.191dupA frameshift (p.Asn64LysfsTer15), the first described and common mutation was identified. Mutations were predicted by in silico tools to have damaging effects and are likely pathogenic according to criteria of the American College of Medical Genetics and Genomics (ACMG). Indeed, molecular modeling of mutations revealed substantial changes in ANO5 conformation that could affect the protein structure and function. In addition, variants in other genes associated with muscle pathology were identified, possibly affecting the disease progress. The presented data indicate that the identified ANO5 mutations contribute to the observed muscle pathology and broaden the genetic spectrum of LGMD myopathies.

ObjectiveTo study the genetic and phenotypic spectrum of patients harboring recessive mutations in BVES.MethodsWe performed whole-exome sequencing in a multicenter cohort of 1929 patients with a suspected hereditary myopathy, showing unexplained limb-girdle muscular weakness and/or elevated creatine kinase levels. Immunohistochemistry and mRNA experiments on patients' skeletal muscle tissue were performed to study the pathogenicity of identified loss-of-function (LOF) variants in BVES.ResultsWe identified 4 individuals from 3 families harboring homozygous LOF variants in BVES, the gene that encodes for Popeye domain containing protein 1 (POPDC1). Patients showed skeletal muscle involvement and cardiac conduction abnormalities of varying nature and severity, but all exhibited at least subclinical signs of both skeletal muscle and cardiac disease. All identified mutations lead to a partial or complete loss of function of BVES through nonsense-mediated decay or through functional changes to the POPDC1 protein.ConclusionsWe report the identification of homozygous LOF mutations in BVES, causal in a young adult-onset myopathy with concomitant cardiac conduction disorders in the absence of structural heart disease. These findings underline the role of POPDC1, and by extension, other members of this protein family, in striated muscle physiology and disease. This disorder appears to have a low prevalence, although it is probably underdiagnosed because of its striking phenotypic variability and often subtle yet clinically relevant manifestations, particularly concerning the cardiac conduction abnormalities.

We describe two Finnish siblings in whom an incidentally detected elevated creatine kinase activity eventually led to a diagnosis of limb-girdle muscular dystrophy-dystroglycanopathy (Type C12; MDDGC12). When diagnosed at age 10 and 13 years, they were mildly affected with a slow or non-progressive disease course. The main symptoms comprised infrequent hip cramps triggered by flexion, neck cramps triggered by yawning, transient growing pains, calf hypertrophy and mild proximal muscle weakness. Their cognitive and motor developments were unremarkable and they were physically active. Whole-exome sequencing revealed compound heterozygous mutations, both of which were novel, in the protein O-mannosyl kinase (POMK) gene in both siblings; a missense mutation, p.Pro322Leu (c.965C > T), and a nonsense mutation, p.Arg46Ter (c.136C > T). The results were confirmed by Sanger sequencing, showing that the parents were heterozygous carriers of one mutation each. This report adds to the literature by providing phenotype and genotype data on this ultra-rare POMK-related dystroglycanopathy. (C) 2018 Elsevier B.V. All rights reserved.

PDF Neuromuscular Research paper Limb girdle muscular dystrophy due to mutations in POMT2 Sofie Thurø Østergaard1, Katherine Johnson2, Tanya Stojkovic3, Thomas Krag1, Willem De Ridder4,5,6, Peter De Jonghe4,5,6, Jonathan Baets4,5,6, Kristl G Claeys7,8, Roberto Fernández-Torrón9, Lauren Phillips2, Ana Topf2, Jaume Colomer10, Shahriar Nafissi11, Shirin Jamal-Omidi11, Celine Bouchet-Seraphin12, France Leturcq13, Daniel G MacArthur14,15, Monkol Lek14,15, Liwen Xu14,15, Isabelle Nelson16, Volker Straub2, John Vissing1 Author affiliations Abstract Background Mutations in the gene coding for protein O-mannosyl-transferase 2 (POMT2) are known to cause severe congenital muscular dystrophy, and recently, mutations in POMT2 have also been linked to a milder limb-girdle muscular dystrophy (LGMD) phenotype, named LGMD type 2N (LGMD2N). Only four cases have been reported so far. ClinicalTrials.gov ID: NCT02759302 Methods We report 12 new cases of LGMD2N, aged 1863 years. Muscle involvement was assessed by MRI, muscle strength testing and muscle biopsy analysis. Other clinical features were also recorded. Results Presenting symptoms were difficulties in walking, pain during exercise, delayed motor milestones and learning disabilities at school. All had some degree of cognitive impairment. Brain MRIs were abnormal in 3 of 10 patients, showing ventricular enlargement in one, periventricular hyperintensities in another and frontal atrophy of the left hemisphere in a third patient. Most affected muscle groups were hip and knee flexors and extensors on strength testing. On MRI, most affected muscles were hamstrings followed by paraspinal and gluteal muscles. The 12 patients in our cohort carried 11 alleles with known mutations, whereas 11 novel mutations accounted for the remaining 13 alleles. Conclusion We describe the first cohort of patients with LGMD2N and show that unlike other LGMD types, all patients had cognitive impairment. Primary muscle involvement was found in hamstring, paraspinal and gluteal muscles on MRI, which correlated well with reduced muscle strength in hip and knee flexors and extensors. The study expands the mutational spectrum for LGMD2N, with the description of 11 novel POMT2 mutations in the association with LGMD2N. Clinical trial registration NCT02759302.

ObjectiveTo determine whether congenital arhinia/Bosma arhinia microphthalmia syndrome (BAMS) and facioscapulohumeral muscular dystrophy type 2 (FSHD2), 2 seemingly unrelated disorders both caused by heterozygous pathogenic missense variants in the SMCHD1 gene, might represent different ends of a broad single phenotypic spectrum associated with SMCHD1 dysfunction.MethodsWe examined and/or interviewed 14 patients with FSHD2 and 4 unaffected family members with N-terminal SMCHD1 pathogenic missense variants to identify BAMS subphenotypes.ResultsNone of the patients with FSHD2 or family members demonstrated any congenital defects or dysmorphic features commonly found in patients with BAMS. One patient became anosmic after nasal surgery and one patient was hyposmic; one man was infertile (unknown cause) but reported normal pubertal development.ConclusionThese data suggest that arhinia/BAMS and FSHD2 do not represent one phenotypic spectrum and that SMCHD1 pathogenic variants by themselves are insufficient to cause either of the 2 disorders. More likely, both arhinia/BAMS and FSHD2 are caused by complex oligogenic or multifactorial mechanisms that only partially overlap at the level of SMCHD1.

The authors regret that in the Abstract and in the Molecular Analysis paragraphs of the above paper, the following was incorrect: “a homozygous c.131T > G (p.Leu44Pro) substitution”. This should read “a homozygous c.131T > C (p.Leu44Pro) substitution”. The authors would like to apologise for any inconvenience caused. They would like to thank Sally Heywood, Research Assistant, Human Gene Mutation Database, Institute of Medical Genetics, Cardiff University for bringing this error to their attention.

Background: Dystroglycanopathies are a clinically and genetically heterogeneous group of disorders that are typically characterised by limb-girdle muscle weakness. Mutations in 18 different genes have been associated with dystroglycanopathies, the encoded proteins of which typically modulate the binding of α-dystroglycan to extracellular matrix ligands by altering its glycosylation. This results in a disruption of the structural integrity of the myocyte, ultimately leading to muscle degeneration. Methods: Deep phenotypic information was gathered using the PhenoTips online software for 1001 patients with unexplained limb-girdle muscle weakness from 43 different centres across 21 European and Middle Eastern countries. Whole-exome sequencing with at least 250ng DNA was completed using an Illumina exome capture and a 38Mb baited target. Genes known to be associated with dystroglycanopathies were analysed for disease-causing variants. Results: Suspected pathogenic variants were detected in DPM3, ISPD, POMT1 and FKTN in one patient each, in POMK in two patients, in GMPPB in three patients, in FKRP in eight patients and in POMT2 in ten patients. This indicated a frequency of 2.7% for the disease group within the cohort of 1001 patients with unexplained limb-girdle muscle weakness. The phenotypes of the 27 patients were highly variable, yet with a fundamental presentation of proximal muscle weakness and elevated serum creatine kinase. Conclusions: Overall, we have identified 27 patients with suspected pathogenic variants in dystroglycanopathy-associated genes. We present evidence for the genetic and phenotypic diversity of the dystroglycanopathies as a disease group, while also highlighting the advantage of incorporating next-generation sequencing into the diagnostic pathway of rare diseases., SCOPUS: ar.j, info:eu-repo/semantics/published

Defects of O-linked glycosylation of alpha-dystroglycan cause a wide spectrum of muscular dystrophies ranging from severe congenital muscular dystrophy associated with abnormal brain and eye development to mild limb girdle muscular dystrophy. We report a female patient who developed isolated pelvic girdle muscle weakness and wasting, which became symptomatic at age 42. Exome sequencing uncovered a homozygous c.131T > G (p.Leu44Pro) substitution in DPM3, encoding dolichol-P-mannose (DPM) synthase subunit 3, leading to a 50% reduction of enzymatic activity. Decreased availability of DPM as an essential donor substrate for protein O-mannosyltransferase (POMT) 1 and 2 explains defective skeletal muscle alpha-dystroglycan O-glycosylation. Our findings show that DPM3 mutations may lead to an isolated and mild limb girdle muscular dystrophy phenotype without cardiomyopathy.

BackgroundThere is a dearth of research into the mental health of gay men, lesbians and bisexual men and women in the UK.AimsTo assess rates and possible predictors of mental illness in these groups.MethodA comprehensive assessment was made of the psychological and social well-being of a sample of gay men, lesbians and bisexual men and women, identified using ‘snowball’ sampling.ResultsOf the 1285 gay, lesbian and bisexual respondents who took part, 556 (43%) had mental disorder as defined by the revised Clinical Interview Schedule (CIS – R). Out of the whole sample, 361 (31%) had attempted suicide. This was associated with markers of discrimination such as recent physical attack (OR=l.7, 95% CI 1.3–2.3) and school bullying (OR=l.4, 95% CI 1.1–2.0), but not with higher scores on the CIS-R.ConclusionsGay, lesbian and bisexual men and women have high levels of mental disorder, possibly linked with discrimination.

Families frequently report that they have received the diagnosis of “a little dementia” for a loved one whose memory is impaired. There is no such thing as “a little dementia.” It is like being a “little pregnant”—either you have it or you do not! And having “dementia” is never a good thing—although families who receive that diagnosis will frequently say, “Well at least it is not Alzheimer’s!” This denial keeps them from learning about the disease, planning for a time when the loved one with the diagnosis is unable to survive without 24-h care, and making other decisions that are best made before a situation becomes a crisis.

Another issue that challenges family and paid caregivers alike has to do with the expression of sexuality in older adults with dementia. Contrary to common perceptions, sexual behavior may actually increase in those who have dementia. Even with profound cognitive impairments, many of those with dementia enjoy a high degree of sociability and the capacity to engage in intimate relationships. In fact, to be human is to be sexual—we are “hardwired” for this. Sometimes, the desire to be close to another person manifests itself in positive feelings of being cared for and desirable. Sometimes, this same desire for closeness is manifested in flirtation, affection, passing compliments, proximity to another, as well as sexual intercourse. The expression of sexual feelings can be directed to a spouse, a professional caregiver, to other residents in a facility, as well as toward adult children of the person with dementia. The adult child might closely resemble the person’s spouse when at a similar age. In this case, the sexual overtures are basically a case of “mistaken identity.”

As baby boomers age, the projected number of those who will develop Alzheimer’s is staggering—not only for family caregivers who might still be raising children but also for the health-care system of the USA. Consider these statistics from the 2014 edition of Alzheimer’s disease Facts and Figures, Alzheimer’s and Dementia (Alzheimer’s Association 2014; Auer et al. 1994; Bobinski et al. 1996).

Prostate cancer accounts for close to 30,000 deaths annually in the United States. The most frequent alteration in the prostate oncogenome landscape is loss of chromosome 8p, which is also observed in other epithelial tumors. This region harbors NKX3.1, a prostate specific tumor suppressor, but 8p copy number loss does not correlate with its mRNA expression. The PPP2R2A gene, which encodes a regulatory subunit of protein phosphatase 2 (PP2A) designated B55, is located in 8p21.2 and is hemizygously lost in more that 50% of prostate adenocarcinomas. TCGA data analysis shows that hemizygous loss of PPP2R2A correlates with its reduced expression, poorer prognosis and that loss of PPP2R2A increases with metastasis to more than 80%. Of note, while homozygous loss is less common (4-10%), deletion of other subunits of the B55, but not other PP2A regulatory families, increases dramatically with metastasis despite their independent chromosomal locations. However, functional evidence that PPP2R2A acts as an haploinsufficient tumor suppressor and the potential mechanisms implicated are lacking. We have identified prostate cancer (PrCa) cell lines with reduced expression of B55 and shown that reconstitution of B55 expression reduces proliferation and induces senescence and loss of viability. B55 reconstitution in PC3 cells, an AR-negative model of human castration-resistant prostate cancer (CRPC), also blocks transformation in vitro and tumorigenicity in SCID mice. Reconstitution of B55 in PCa cells with reduced expression (PC3 and DU145 cells) induces cell cycle defects in G1, G2/M and mitosis diminishing and delaying phosphorylation of pRB pocket proteins and mitotic substrates and altering associated kinase networks. Mitogenic signaling is also attenuated. Importantly, the effects of B55 reconstitution are at least partially mimicked via treatment of these cells with PP2A activating drugs, which have anti-tumorigenic activity, indicating that development of specific B55/PP2A activating drugs targeting the unaltered B55 alleles remaining in PCa cells may have therapeutic potential for a high proportion of prostate tumors and perhaps other epithelial tumors such as luminal B breast and ovarian cancers which share this alteration. Citation Format: Ziran Zhao, Alison Kurimchak, Petr Makhov, Katherine Johnson, Vladimir Kolenko, James Duncan, Xavier Graña. Reconstituting B55/protein phosphatase 2A activity in B55/PP2A-defective prostate cancer cells suppresses proliferation and tumorigenicity through pleiotropic mechanisms [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-320. doi:10.1158/1538-7445.AM2017-LB-320

Objectives In pharmacogenetic medicine, we aim to predict responders to therapy and those who will suffer side effects, so we can choose an alternative agent. In this review, we assessed the current evidence regarding pharmacogenetic guided warfarin, clopidogrel, statin and antihypertensive therapy in stroke. We also considered the barriers to implementation of large scale pharmacogenetic guided therapy in Australia, a nation of many ethnicities. Methods A computerised literature search was conducted through to November 2016 for warfarin and statins, and October 2016 for antihypertensives and antiplatelets. We utilised Medline, Premedline, Embase and the Cochrane Database of Systematic Reviews using the search term pharmacogenetics, then overall drug class and individual drug names, such as ‘coumarin’, ‘warfarin’, ‘clopidogrel’. Results We extracted 184 relevant articles for warfarin, 133 for clopidogrel, 89 for statins and 70 for antihypertensives. Overall, the majority of studies were prospective and retrospective observational association studies, with or without controls, which often reported dramatic and contradictory results. We also reviewed a number of randomised controlled trials, narrative and systematic reviews and college guidelines. There is a potential role for pharmacogenetic guided warfarin and clopidogrel therapy. However, neither are ready for prime time, as the utility of genetic testing remains unclear and there are significant barriers to implementation, particularly in a multiethnic population, like Australia. The role for pharmacogenetic guided statin and antihypertensive therapy is less apparent. Conclusions The current evidence does not support pharmacogenetic guided therapy in stroke in Australia. However, a one-size fits all approach does not work and there is a definite dearth of Australian based pharmacogenetic research, which must be addressed.

To identify and summarize available research literature about rehabilitation interventions for multiple sclerosis (MS) fatigue management.Database searches (PubMed, EMBASE, CINAHL and PsychINFO) were conducted in October 2011 and last updated in July 2013. To be included, studies must have been published in a peer-reviewed scientific journal, written in English and included an intervention to manage MS fatigue. Effect size (ES) were calculated for the quantitative studies to estimate intervention effects, and major themes were summarized for the qualitative studies.Thirty-eight studies were included in this review. A variety of exercise and behavior change interventions were prescribed to adults with MS. The two most common interventions were progressive resistive training and fatigue management programs. Three exercise intervention studies and nine behavior change intervention studies with quantitative data presented significant ES. Four studies with qualitative data supported the positive impact of certain exercise and behavior change interventions.This review identified a variety of exercise and behavior change interventions for MS fatigue management. While the findings may provide helpful information to inform practice, future researchers need to develop and evaluate knowledge translation strategies to facilitate the application of this evidence to daily practice to advance MS rehabilitation care.Both exercise and behavior change interventions demonstrate some degree of effectiveness for managing MS fatigue. Effect sizes for exercise and behavior change interventions are similar, although the populations examined are different. Overall, evidence for exercise focuses on people who are less disabled, while evidence for behavior change interventions includes a broader population. Future researchers need to develop and evaluate knowledge translation strategies that facilitate application of evidence in daily practice in order to advance MS rehabilitation.

Receptor tyrosine kinases are involved in multiple cellular processes, and drugs that inhibit their action are used in the clinic to treat several types of cancer. However, the value of receptor tyrosine kinase inhibitors (RTKIs) for treating infectious disease has yet to be explored. Here, we have shown in mice that administration of the broad-spectrum RTKI sunitinib maleate (Sm) blocked the vascular remodeling and progressive splenomegaly associated with experimental visceral leishmaniasis. Furthermore, Sm treatment restored the integrity of the splenic microarchitecture. Although restoration of splenic architecture was accompanied by an increase in the frequency of IFN-gamma+CD4+ T cells, Sm treatment alone was insufficient to cause a reduction in tissue parasite burden. However, preconditioning by short-term Sm treatment proved to be successful as an adjunct therapy, increasing the frequency of IFN-gamma+ and IFN-gamma+TNF+CD4+ T cells, enhancing NO production by splenic macrophages, and providing dose-sparing effects when combined with a first-line immune-dependent anti-leishmanial drug. We propose, therefore, that RTKIs may prove clinically useful as agents to restore immune competence before the administration of chemo- or immunotherapeutic drugs in the treatment of visceral leishmaniasis or other diseases involving lymphoid tissue remodeling, including cancer.

[Background and Aims] Vibration-controlled transient elastography (VCTE), point shear wave elastography (pSWE), 2-dimensional shear wave elastography (2DSWE), magnetic resonance elastography (MRE), and magnetic resonance imaging (MRI) have been proposed as non-invasive tests for patients with non-alcoholic fatty liver disease (NAFLD). This study evaluated their diagnostic accuracy for liver fibrosis and non-alcoholic steatohepatitis (NASH)., [Methods] PubMED/MEDLINE, EMBASE and the Cochrane Library were searched for studies examining the diagnostic accuracy of these index tests, against histology as the reference standard, in adult patients with NAFLD. Two authors independently screened and assessed methodological quality of studies and extracted data. Summary estimates of sensitivity, specificity and area under the curve (sAUC) were calculated for fibrosis stages and NASH, using a random effects bivariate logit-normal model., [Results] We included 82 studies (14,609 patients). Meta-analysis for diagnosing fibrosis stages was possible in 53 VCTE, 11 MRE, 12 pSWE and 4 2DSWE studies, and for diagnosing NASH in 4 MRE studies. sAUC for diagnosis of significant fibrosis were: 0.83 for VCTE, 0.91 for MRE, 0.86 for pSWE and 0.75 for 2DSWE. sAUC for diagnosis of advanced fibrosis were: 0.85 for VCTE, 0.92 for MRE, 0.89 for pSWE and 0.72 for 2DSWE. sAUC for diagnosis of cirrhosis were: 0.89 for VCTE, 0.90 for MRE, 0.90 for pSWE and 0.88 for 2DSWE. MRE had sAUC of 0.83 for diagnosis of NASH. Three (4%) studies reported intention-to-diagnose analyses and 15 (18%) studies reported diagnostic accuracy against pre-specified cut-offs., [Conclusions] When elastography index tests are acquired successfully, they have acceptable diagnostic accuracy for advanced fibrosis and cirrhosis. The potential clinical impact of these index tests cannot be assessed fully as intention-to-diagnose analyses and validation of pre-specified thresholds are lacking., [Lay summary] Non-invasive tests that measure liver stiffness or use magnetic resonance imaging (MRI) have been suggested as alternatives to liver biopsy for assessing the severity of liver scarring (fibrosis) and fatty inflammation (steatohepatitis) in patients with non-alcoholic fatty liver disease (NAFLD). In this study, we summarise the results of previously published studies on how accurately these non-invasive tests can diagnose liver fibrosis and inflammation, using liver biopsy as the reference. We found that some techniques that measure liver stiffness had a good performance for the diagnosis of severe liver scarring., This work has been undertaken as part of the LITMUS (Liver Investigation: Testing Marker Utility in Steatohepatitis) project. The LITMUS project has received funding from the Innovative Medicines Initiative 2 Joint Undertaking under grant agreement No. 777377. This Joint Undertaking receives support from the European Union’s Horizon 2020 research and innovation programme and Europen Federation of Pharmaceutical Industries and Associations (efpia.eu).