Risdiplam for the treatment of adults with spinal muscular atrophy: Experience of the Northern Ireland neuromuscular service.

INTRODUCTION/AIMS: Risdiplam is the newest available treatment for patients with spinal muscular atrophy (SMA). There is little information on its use in adults. We present the clinical experience of adults with SMA treated with risdiplam through the Early Access to Medicines Scheme (EAMS) in Northern Ireland. METHODS: All adults with Type 2 SMA attending the regional neuromuscular clinic were offered risdiplam treatment. Patients had assessments of respiratory function, the Epworth Sleepiness Scale (ESS), Quality of Life Measure for People with Slowly Progressive and Genetic Neuromuscular Disease (QOLM), and Egen Klassifikation 2 (EK2) every 3 mo and the Revised Upper Limb Module for SMA (RULM) at baseline and 6 mo. All assessments other than the RULM were carried out virtually. RESULTS: Six of seven patients who were offered risdiplam consented to treatment through the EAMS (five female, one male, mean age 33.7 y). It was generally well tolerated other than skin photosensitivity in all patients. All patients remained on therapy at 9 mo. All reported meaningful improvements in overall strength, sense of wellbeing, and speech quality. There was no change in respiratory function, daytime hypersomnolence, or upper limb function (all p > .05). There was improvement in the QOLM (p = .027) and EK2 (p = .009). DISCUSSION: Our study raises hopes that risdiplam may be efficacious in adults; however, more systematic studies in larger cohorts are needed before drawing any definitive conclusions. This study also demonstrated the feasibility of virtual assessments.

Recently Graduated Medical Students Lack Exposure to and Comfort with Chronic Liver Diseases.

BACKGROUND: The prevalence of chronic liver disease (CLD) is rising, but it remains unclear if medical school curricula are emphasizing CLD to reflect its growing epidemiology. AIMS: To assess comfort levels and knowledge of CLD among recently graduated medical students METHODS: An anonymous survey was distributed to incoming categorical Internal Medicine (IM) interns at a single academic institution during a 2-year period. The survey consisted of 38 Likert-like questions evaluating comfort levels and self-assessed knowledge for several general medicine and liver diseases, as well as 12 multiple-choice questions to objectively test knowledge. Wilcoxon ranked sum and Fisher's exact test were then used. RESULTS: There was a 100% (n = 65) completion rate. Only 14 (22%) of those surveyed reported exposure to a hepatology rotation in medical school. Highest mean comfort levels (1 = not at all comfortable, 5 = very comfortable) were for managing congestive heart failure (3.59) and chronic obstructive pulmonary disease (3.77). Mean comfort levels for various liver diseases were significantly lower (2.22-3.03, all p < 0.01). Mean self-rated knowledge (1 = no knowledge, 5 = strong knowledge) was also low (2.14-3.13). Although 98% agreed that hepatology is critical to IM training, only 42% agreed that their hepatology education during medical school was adequate. CONCLUSIONS: Recently graduated medical students are less comfortable managing liver diseases compared to other general medical conditions. Only a minority report satisfaction with hepatology education during medical school. These findings suggest that medical curricula need to be modified to better emphasize CLD.

Pentraxin 3 deletion aggravates allergic inflammation through a TH17-dominant phenotype and enhanced CD4 T-cell survival.

BACKGROUND: Pentraxin 3 (PTX3) is a multifunctional molecule that plays a nonredundant role at the crossroads between pathogen clearance, innate immune system, matrix deposition, female fertility, and vascular biology. It is produced at sites of infection and inflammation by both structural and inflammatory cells. However, its role in allergen-induced inflammation remains to be tested. OBJECTIVE: We sought to determine the effect of Ptx3 deletion on ovalbumin (OVA)-induced allergic inflammation in a murine model of asthma. METHODS: Bronchoalveolar lavage fluid was collected from patients with severe asthma and healthy subjects, and the level of PTX3 was determined by using ELISA. Ptx3+/+ and Ptx3-/- mice were sensitized and challenged with OVA and bronchoalveolar lavage fluid, and the lungs were collected for assessing inflammation. Lung tissue inflammation and mucus production were assessed by means of flow cytometry and hematoxylin and eosin and periodic acid-Schiff staining, respectively. flexiVent was used to determine airway resistance to methacholine in these mice. RESULTS: Here we report that mice with severe asthma and OVA-sensitized/challenged mice had increased PTX3 levels in the lungs compared with healthy control mice. Mice lacking PTX3 have exaggerated neutrophilic/eosinophilic lung inflammation, mucus production, and airway hyperresponsiveness in an experimental model of OVA-induced asthma. Furthermore, OVA-exposed lung Ptx3-/- CD4 T cells exhibit an increased production of IL-17A, an effect that is accompanied by an increased signal transducer and activator of transcription 3 phosphorylation, reduced IL-2 production, and enhanced activation and survival. Also, we observed an increase in numbers of IL-6- and IL-23-producing dendritic cells in OVA-exposed Ptx3-/- mice compared with those in wild-type control mice. CONCLUSION: Altogether, PTX3 deficiency results in augmented airway hyperresponsiveness, mucus production, and IL-17A-dominant pulmonary inflammation, suggesting a regulatory role of PTX3 in the development of allergic inflammation.

Novel educational interventions in residency increase knowledge of chronic liver disease and career interest in hepatology.

There is an increasing burden of chronic liver disease (CLD) in the United States but a significant shortage of hepatologists. Thus, it is necessary to develop new recruitment strategies to the field of hepatology as well as ensure that non-gastroenterology-trained physicians are able to capably assist in the care of CLD. We established a novel, nonelective, inpatient hepatology rotation that uses required modules in the American Association for the Study of Liver Diseases Curriculum and Training-First Hepatitis B and C curriculums as well as in LiverLearning. A paper-based anonymous assessment was distributed to the inaugural 25 postgraduate years 2 and 3 internal medicine residents before and after the 2-week rotation over the course of 1 year. Both the prerotation and postrotation assessments included validated multiple-choice questions and Likert-type questions, which evaluated self-perceived knowledge and comfort with managing CLD. The mean comfort level (1 = not at all comfortable/strongly disagree, 5 = very comfortable/strongly agree) of managing several common liver diseases increased significantly after completion of the rotation (i.e., cirrhosis 2.8 versus 3.8, P < 0.001; hepatitis B 2.4 versus 3.4, P = 0.001; hepatitis C 2.6 versus 3.7, P = 0.002; nonalcoholic steatohepatitis 3.0 versus 4.0, P < 0.001; liver transplant care 2.1 versus 3.4, P < 0.001). There was also a significantly increased interest in hepatology as a career (2.6 versus 3.0, P = 0.03). Finally, the mean percentage of multiple-choice questions answered correctly on the pretest was 62% and posttest was 77% (P = 0.02). CONCLUSION: Our novel curriculum and nonelective hepatology rotation has effectively demonstrated improvement in internal medicine residents' comfort with and knowledge of CLD, and increased career interest in hepatology was also observed after completion of the curriculum, which suggests that more exposure to CLD could positively impact recruitment to the workforce; larger, multicenter studies are needed to validate these results. (Hepatology 2016;64:2210-2218).

Mutations in the SPG7 gene cause chronic progressive external ophthalmoplegia through disordered mitochondrial DNA maintenance.

Despite being a canonical presenting feature of mitochondrial disease, the genetic basis of progressive external ophthalmoplegia remains unknown in a large proportion of patients. Here we show that mutations in SPG7 are a novel cause of progressive external ophthalmoplegia associated with multiple mitochondrial DNA deletions. After excluding known causes, whole exome sequencing, targeted Sanger sequencing and multiplex ligation-dependent probe amplification analysis were used to study 68 adult patients with progressive external ophthalmoplegia either with or without multiple mitochondrial DNA deletions in skeletal muscle. Nine patients (eight probands) were found to carry compound heterozygous SPG7 mutations, including three novel mutations: two missense mutations c.2221G>A; p.(Glu741Lys), c.2224G>A; p.(Asp742Asn), a truncating mutation c.861dupT; p.Asn288*, and seven previously reported mutations. We identified a further six patients with single heterozygous mutations in SPG7, including two further novel mutations: c.184-3C>T (predicted to remove a splice site before exon 2) and c.1067C>T; p.(Thr356Met). The clinical phenotype typically developed in mid-adult life with either progressive external ophthalmoplegia/ptosis and spastic ataxia, or a progressive ataxic disorder. Dysphagia and proximal myopathy were common, but urinary symptoms were rare, despite the spasticity. Functional studies included transcript analysis, proteomics, mitochondrial network analysis, single fibre mitochondrial DNA analysis and deep re-sequencing of mitochondrial DNA. SPG7 mutations caused increased mitochondrial biogenesis in patient muscle, and mitochondrial fusion in patient fibroblasts associated with the clonal expansion of mitochondrial DNA mutations. In conclusion, the SPG7 gene should be screened in patients in whom a disorder of mitochondrial DNA maintenance is suspected when spastic ataxia is prominent. The complex neurological phenotype is likely a result of the clonal expansion of secondary mitochondrial DNA mutations modulating the phenotype, driven by compensatory mitochondrial biogenesis.

Two recurrent mutations are associated with GNE myopathy in the North of Britain.

OBJECTIVE: GNE myopathy is a rare recessive myopathy associated with inclusion bodies on muscle biopsy. The clinical phenotype is associated with distal muscle weakness with quadriceps sparing. Most of the current information on GNE myopathy has been obtained through studies of Jewish and Japanese patient cohorts carrying founder mutations in the GNE gene. However, little is known about GNE myopathy in Europe where the prevalence is thought to be very low. METHODS: Patients were referred through the National Specialist Commissioning Team service for limb-girdle muscular dystrophies at Newcastle (UK). All patients harbouring mutations in the GNE gene were recruited for our study. Detailed clinical and genetic data as well as muscle MRIs and muscle biopsies were reviewed. RESULTS: We identified 26 patients harbouring mutations in the GNE gene. Two previously reported mutations (c.1985C>T, p.Ala662Val and c.1225G>T, p.Asp409Tyr) were prevalent in the Scottish, Northern Irish and Northern English populations; with 90% of these patients carrying at least one of the two mutations. Clinically, we confirmed the homogenous pattern of selective quadriceps sparing but noted additional features like asymmetry of weakness at disease onset. CONCLUSIONS: GNE myopathy is an important diagnosis to consider in patients presenting with distal leg muscle weakness. We report, for the first time, two common mutations in the north of Britain and highlight the broader spectrum of clinical phenotypes. We also propose that the prevalence of GNE myopathy may be underestimated due to the frequent absence of rimmed vacuoles in the muscle biopsy.

Maternal asthma and microRNA regulation of soluble HLA-G in the airway.

BACKGROUND: We previously reported an interaction between maternal asthma and the child's HLA-G genotype on the child's subsequent risk for asthma. The implicated single nucleotide polymorphism at +3142 disrupted a target site for the microRNA (miR)-152 family. We hypothesized that the interaction effect might be mediated by these miRs. OBJECTIVE: The objective of this study was to test this hypothesis in adults with asthma who are a subset of the same subjects who participated in our earlier family-based studies. METHODS: We measured soluble HLA-G (sHLA-G) concentrations in bronchoalveolar lavage fluid (n = 36) and plasma (n = 57) from adult asthmatic subjects with and without a mother with asthma, and HLA-G and miR-152 family (miR-148a, miR-148b, and miR-152) transcript levels in airway epithelial cells from the same subjects. RESULTS: miR-148b levels were significantly increased in airway epithelial cells from asthmatic subjects with an asthmatic mother compared with those seen in asthmatic subjects without an asthmatic mother, and +3142 genotypes were associated with sHLA-G concentrations in bronchoalveolar lavage fluid among asthmatic subjects with an asthmatic mother but not among those with a nonasthmatic mother. Neither effect was observed in the plasma (sHLA-G) or white blood cells (miRNA). CONCLUSION: These combined results are consistent with +3142 allele-specific targeting of HLA-G by the miR-152 family and support our hypothesis that miRNA regulation of sHLA-G in the airway is influenced by both the asthma status of the subject's mother and the subject's genotype. Moreover, we demonstrate that the effects of maternal asthma on the gene regulatory landscape in the airways of the mother's children persist into adulthood.

ANO5 gene analysis in a large cohort of patients with anoctaminopathy: confirmation of male prevalence and high occurrence of the common exon 5 gene mutation.

Limb girdle muscular dystrophy type 2L or anoctaminopathy is a condition mainly characterized by adult onset proximal lower limb muscular weakness and raised CK values, due to recessive ANO5 gene mutations. An exon 5 founder mutation (c.191dupA) has been identified in most of the British and German LGMD2L patients so far reported. We aimed to further investigate the prevalence and spectrum of ANO5 gene mutations and related clinical phenotypes, by screening 205 undiagnosed patients referred to our molecular service with a clinical suspicion of anoctaminopathy. A total of 42 unrelated patients had two ANO5 mutations (21%), whereas 14 carried a single change. We identified 34 pathogenic changes, 15 of which are novel. The c.191dupA mutation represents 61% of mutated alleles and appears to be less prevalent in non-Northern European populations. Retrospective clinical analysis corroborates the prevalently proximal lower limb phenotype, the male predominance and absence of major cardiac or respiratory involvement. Identification of cases with isolated hyperCKaemia and very late symptomatic male and female subjects confirms the extension of the phenotypic spectrum of the disease. Anoctaminopathy appears to be one of the most common adult muscular dystrophies in Northern Europe, with a prevalence of about 20%-25% in unselected undiagnosed cases.

A Neurology Advanced Referral Management System (NARMS) Reduces Face-to-Face Consultations By Over Sixty Percent.

Background: The COVID-19 pandemic has made neurology clinic waiting times longer. To prevent a build-up of patients waiting, we introduced a neurology advanced referral management system (NARMS) to deal with new referrals from GPs, using advice, investigations, or the telephone, as alternatives to face-to-face (FF) assessment. Methods: For six months, electronic referrals from GPs were triaged to the above categories. We recorded the numbers in each category, patient satisfaction, inter-consultant triage variation, re-referrals, and calculated CO2 emissions. Results: There were 573 referrals. Triage destinations were advice 33%, investigations 27%, telephone 17%, and FF 33%. Of patients referred for MRI, 95% were happy not to be seen if their investigation was normal. Less-experienced consultants triaged 20% and 30% respectively, to advice or investigations, compared with 40% by a triage-experienced neurologist. Four percent were re-referred. Numbers on the waiting list did not increase. CO2 emissions were reduced by 50%. Discussion: Two thirds of neurological referrals from GPs did not need to be seen FF and 50% were dealt with without the neurologist meeting the patient. Carbon emission was halved. This system should be employed more, with FF examination reserved for those patients who need a neurological examination for diagnosis and management.

Serum Neurofilaments in Motor Neuron Disease and Their Utility in Differentiating ALS, PMA and PLS.

Neurofilament levels are elevated in many neurodegenerative diseases and have shown promise as diagnostic and prognostic biomarkers in Amyotrophic Lateral Sclerosis (ALS), the most common form of Motor Neuron Disease (MND). This study assesses serum neurofilament light (NFL) and neurofilament heavy (NFH) chain concentrations in patients with ALS, other variants of motor neuron disease such as Progressive Muscular Atrophy (PMA) and Primary Lateral Sclerosis (PLS), and a range of other neurological diseases. It aims to evaluate the use of NFL and NFH to differentiate these conditions and for the prognosis of MND disease progression. NFL and NFH levels were quantified using electrochemiluminescence immunoassays (ECLIA). Both were elevated in 47 patients with MND compared to 34 patients with other neurological diseases and 33 healthy controls. NFL was able to differentiate patients with MND from the other groups with a Receiver Operating Characteristic (ROC) curve area under the curve (AUC) of 0.90 (p < 0.001). NFL correlated with the rate of disease progression in MND (rho 0.758, p < 0.001) and with the ALS Functional Rating Scale (rho -0.335, p = 0.021). NFL levels were higher in patients with ALS compared to both PMA (p = 0.032) and PLS (p = 0.012) and were able to distinguish ALS from both PMA and PLS with a ROC curve AUC of 0.767 (p = 0.005). These findings support the use of serum NFL to help diagnose and differentiate types of MND, in addition to providing prognostic information to patients and their families.

The Role of Sphingomyelin and Ceramide in Motor Neuron Diseases.

Amyotrophic Lateral Sclerosis (ALS), Spinal Bulbar Muscular Atrophy (SBMA), and Spinal Muscular Atrophy (SMA) are motor neuron diseases (MNDs) characterised by progressive motor neuron degeneration, weakness and muscular atrophy. Lipid dysregulation is well recognised in each of these conditions and occurs prior to neurodegeneration. Several lipid markers have been shown to predict prognosis in ALS. Sphingolipids are complex lipids enriched in the central nervous system and are integral to key cellular functions including membrane stability and signalling pathways, as well as being mediators of neuroinflammation and neurodegeneration. This review highlights the metabolism of sphingomyelin (SM), the most abundant sphingolipid, and of its metabolite ceramide, and its role in the pathophysiology of neurodegeneration, focusing on MNDs. We also review published lipidomic studies in MNDs. In the 13 studies of patients with ALS, 12 demonstrated upregulation of multiple SM species and 6 demonstrated upregulation of ceramides. SM species also correlated with markers of clinical progression in five of six studies. These data highlight the potential use of SM and ceramide as biomarkers in ALS. Finally, we review potential therapeutic strategies for targeting sphingolipid metabolism in neurodegeneration.

Real-World Data on Access to Standards of Care for People With Spinal Muscular Atrophy in the UK.

Spinal Muscular Atrophy (SMA) is characterized by muscle atrophy and weakness and has an incidence of 1:11. 000 live births which projects an estimated population in the UK of 650-1,300 affected patients. Standards of Care (SoC) were updated in 2017 and they have been widely adopted as a reference for implementation of care in SMA across the globe. The effectiveness of implementation and adherence to these standards across different countries is unclear. The aim of this study is to describe the experience of individuals with SMA regarding their care in the UK. An online anonymised survey was sent out via patient organizations, the UK SMA Patient Registry, professional networks, and social media to reach across the UK. The survey captured demographic profile, professionals involved in a patient's care, Interventions and access to mobility aids and home adaptations. Participants responded about their access to services and to rate how important each professional and intervention was for their health and wellbeing. One hundred and twenty-eight responses were collected with a median age of 34 years (1-81). Seventy-three percent of participants were adults and 60% men. Overall good access to neurologist (>90%) but limited to nurse specialist (48%) and physiotherapist (57%). Good access to respiratory support was reported but limited for interventions for positioning and bracing and exercise. This survey highlights that access to certain professionals for people with SMA is limited in the UK. Striking differences were noted between pediatric and adult populations. Limited access to care were regularly reported, with half of the study population consistently not accessing full multidisciplinary care. Access to interventions for contracture management were recorded to have significant limitations. Mobility aids and home adaptations are widely available and were also reported as the most valued interventions. Access to nutritional support or speech and language therapy appears only to be available for a small proportion of the participants. Access to respiratory care was good especially in severe forms of SMA. We found pockets of good practice in the UK that align with the SoC. However, access is not equal for adults and children and access to certain professionals is significantly limited.

Genetic defects are common in myopathies with tubular aggregates.

OBJECTIVE: A group of genes have been reported to be associated with myopathies with tubular aggregates (TAs). Many cases with TAs still lack of genetic clarification. This study aims to explore the genetic background of cases with TAs in order to improve our knowledge of the pathogenesis of these rare pathological structures. METHODS: Thirty-three patients including two family members with biopsy confirmed TAs were collected. Whole-exome sequencing was performed on 31 unrelated index patients and a candidate gene search strategy was conducted. The identified variants were confirmed by Sanger sequencing. The wild-type and the mutant p.Ala11Thr of ALG14 were transfected into human embryonic kidney 293 cells (HEK293), and western blot analysis was performed to quantify protein expression levels. RESULTS: Eleven index cases (33%) were found to have pathogenic variant or likely pathogenic variants in STIM1, ORAI1, PGAM2, SCN4A, CASQ1 and ALG14. Among them, the c.764A>T (p.Glu255Val) in STIM1 and the c.1333G>C (p.Val445Leu) in SCN4A were novel. Western blot analysis showed that the expression of ALG14 protein was severely reduced in the mutant ALG14 HEK293 cells (p.Ala11Thr) compared with wild type. The ALG14 variants might be associated with TAs in patients with complex multisystem disorders. INTERPRETATION: This study expands the phenotypic and genotypic spectrums of myopathies with TAs. Our findings further confirm previous hypothesis that genes related with calcium signalling pathway and N-linked glycosylation pathway are the main genetic causes of myopathies with TAs.

Nuclear import of serum response factor in airway smooth muscle.

We have previously shown that the transcription-promoting activity of serum response factor (SRF) is partially regulated by its extranuclear redistribution. In this study, we examined the cellular mechanisms that facilitate SRF nuclear entry in canine tracheal smooth muscle cells. We used in vitro pull-down assays to determine which karyopherin proteins bound SRF and found that SRF binds KPNA1 and KPNB1 through its nuclear localization sequence. Immunoprecipitation studies also demonstrated direct SRF-KPNA1 interaction in HEK293 cells. Import assays demonstrated that KPNA1 and KPNB1 together were sufficient to mediate rapid nuclear import of SRF-GFP. Our studies also suggest that SRF is able to gain nuclear entry through an auxiliary, nuclear localization sequence-independent mechanism.

Akt activation induces hypertrophy without contractile phenotypic maturation in airway smooth muscle.

Airway smooth muscle (ASM) hypertrophy is a cardinal feature of severe asthma, but the underlying molecular mechanisms remain uncertain. Forced protein kinase B/Akt 1 activation is known to induce myocyte hypertrophy in other muscle types, and, since a number of mediators present in asthmatic airways can activate Akt signaling, we hypothesized that Akt activation could contribute to ASM hypertrophy in asthma. To test this hypothesis, we evaluated whether Akt activation occurs naturally within airway myocytes in situ, whether Akt1 activation is sufficient to cause hypertrophy of normal airway myocytes, and whether such hypertrophy is accompanied by excessive accumulation of contractile apparatus proteins (contractile phenotype maturation). Immunostains of human airway sections revealed concordant activation of Akt (reflected in Ser(473) phosphorylation) and of its downstream effector p70(S6Kinase) (reflected in Thr(389) phosphorylation) within airway muscle bundles, but there was no phosphorylation of the alternative Akt downstream target glycogen synthase kinase (GSK) 3ß. Artificial overexpression of constitutively active Akt1 (by plasmid transduction or lentiviral infection) caused a progressive increase in size and protein content of cultured canine tracheal myocytes and increased p70(S6Kinase) phosphorylation but not GSK3ß phosphorylation; however, constitutively active Akt1 did not cause disproportionate overaccumulation of smooth muscle (sm) α-actin and SM22. Furthermore, mRNAs encoding sm-α-actin and SM22 were reduced. These results indicate that forced Akt1 signaling causes hypertrophy of cultured airway myocytes without inducing further contractile phenotypic maturation, possibly because of opposing effects on contractile protein gene transcription and translation, and suggest that natural activation of Akt1 plays a similar role in asthmatic ASM.