Evaluation of N-Acetylmannosamine Administration to Restore Sialylation in GNE-Deficient Human Embryonal Kidney Cells.

BACKGROUND: A key mechanism in the neuromuscular disease GNE myopathy (GNEM) is believed to be that point mutations in the GNE gene impair sialic acid synthesis - maybe due to UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) activity restrictions - and resulting in muscle tissue loss. N-acetylmannosamine (ManNAc) is the first product of the bifunctional GNE enzyme and can therefore be regarded as a precursor of sialic acids. This study investigates whether this is also a suitable substance for restoring the sialic acid content in GNE-deficient cells. METHODS: A HEK-293 GNE-knockout cell line was generated using CRISPR-Cas9 and analyzed for its ability to synthesize sialic acids. The cells were then supplemented with ManNAc to compensate for possible GNE inactivity and thereby restore sialic acid synthesis. Sialic acid levels were monitored by immunoblot and high performance liquid chromatography (HPLC). RESULTS: The HEK-293 GNE-knockout cells showed almost no polysialylation signal (immunoblot) and a reduced overall (-71%) N-acetylneuraminic acid (Neu5Ac) level (HPLC) relative to total protein and normalized to wild type level. Supplementation of GNE-deficient HEK-293 cells with 2 mM ManNAc can restore polysialylation and free intracellular sialic acid levels to wild type levels. The addition of 1 mM ManNAc is sufficient to restore the membrane-bound sialic acid level. CONCLUSIONS: Although the mechanism behind this needs further investigation and although it remains unclear why adding ManNAc to GNE-deficient cells is sufficient to elevate polysialylation back to wild type levels - since this substance is also converted by the GNE, all of this might yet prove helpful in the development of an appropriate therapy for GNEM.

Targeting neuromuscular junction to treat neuromuscular disorders.

The integrity and preservation of the neuromuscular junction (NMJ), the interface between the motor neuron and skeletal muscle, is critical for maintaining a healthy skeletal muscle. The structural and/or functional defects in the three cellular components of NMJ, namely the pre-synaptic terminal, synaptic cleft, and post-synaptic region, negatively affect skeletal muscle mass and/or strength. Therefore, NMJ repair appears to be an appropriate therapy for muscle disorders. Mouse models provide a detailed molecular characterization of various cellular components of NMJ with relevance to human diseases. This review discusses different molecular targets on the three cellular components of NMJ for treating muscle diseases. The potential effects of these therapies on NMJ morphology and motor performance, their therapeutic efficacy, and clinical relevance are discussed. Collectively, the available data supports targeting NMJ alone or as an adjunct therapy in treating muscle disorders. However, the potential impact of such interventions on human patients with muscle disorders requires further investigation.

First-line immunosuppression in neuromuscular diseases.

Autoimmune neuromuscular diseases are common and often treatable causes for peripheral nervous system dysfunction. If not optimally managed, they result in meaningful impairments and disability. The treating neurologist should aim to maximise clinical recovery with minimal iatrogenic risk. This requires careful patient and medication selection, appropriate counselling and close monitoring of clinical efficacy and safety. Here, we summarise our consensus departmental approach to first-line immunosuppression in neuromuscular diseases. We combine multispecialty evidence and expertise with a focus on autoimmune neuromuscular diseases to create guidance on starting, dosing and monitoring for toxic effects of the commonly used drugs. These include corticosteroids, steroid-sparing agents and cyclophosphamide. We also provide efficacy monitoring advice, as clinical response informs dosage and drug choice. The principles of this approach could be applied across much of the spectrum of immune-mediated neurological disorders where there is significant therapeutic crossover.

Systematic review of colchicine neuromyopathy: Risk factors, duration and resolution.

OBJECTIVE: To identify reports of colchicine-induced neuropathy and myopathy and ascertain risk factors associated with this toxicity at commonly used doses. METHODS: A systematic review of case reports was conducted in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA methodology). PubMed and EMBASE were searched through October 2021 for case reports of neuropathy and/or myopathy associated with the use of colchicine at therapeutic doses. RESULTS: A total of 143 cases of neuromyopathy from 99 articles were identified as having a "definite" or "probable" association with colchicine usage, as assessed by the Naranjo algorithm. Most of these cases presented with features of both neuropathy and myopathy (n=72, 51%) but symptoms of myopathy were predominant. The mean total daily dose was 1.25±0.60 mg and 48% had been taking colchicine for more than 12 months before presenting with neuromyopathy. A total of 117 (82%) of all reports had either a significant co-morbidity or possible colchicine drug-drug interaction, while 57 (40%) had both risk factors. A total of 26 cases (18%) had no significant risk factor but only 15 of these reports contained complete descriptions of the patient's co-morbidities and co-medications. Cessation of colchicine generally led to complete resolution of symptoms in 70% of cases within a median of 21 days. There were 3 deaths reported which were due to multi-organ failure despite cessation of colchicine and medical management. Colchicine was restarted at reduced doses in 15 cases and 73% had no symptom recurrence. CONCLUSION: Neuromyopathy is an uncommon but reported adverse effect of colchicine. Cases generally present with proximal myopathy symptoms. Cases of colchicine neuromyopathy are largely reported in patients on commonly used doses. Renal and hepatic dysfunction and medications that inhibit cytochrome P450 3A4 isozyme (CYP3A4) and P-glycoprotein (P-gp) appear to be the most significant risk factors. Fortunately, cessation of colchicine generally leads to complete resolution of symptoms. Recommencement of colchicine at reduced doses appeared to be usually safe.

Cannabinoids, Endocannabinoids, and Synthetic Cannabimimetic Molecules in Neuromuscular Disorders.

Neuromuscular disorders (NMDs) encompass a large heterogeneous group of hereditary and acquired diseases primarily affecting motor neurons, peripheral nerves, and the skeletal muscle system. The symptoms of NMDs may vary depending on the specific condition, but some of the most common ones include muscle weakness, pain, paresthesias, and hyporeflexia, as well as difficulties with swallowing and breathing. NMDs are currently untreatable. Therapeutic options include symptomatic and experimental medications aimed at delaying and alleviating symptoms, in some cases supplemented by surgical and physical interventions. To address this unmet medical need, ongoing research is being conducted on new treatments, including studies on medical cannabis, endocannabinoids, and related molecules with cannabimimetic properties. In this context, a significant amount of knowledge about the safety and effectiveness of cannabinoids in NMDs has been obtained from studies involving patients with multiple sclerosis experiencing pain and spasticity. In recent decades, numerous other preclinical and clinical studies have been conducted to determine the potential benefits of cannabinoids in NMDs. This review article aims to summarize and provide an unbiased point of view on the current knowledge about the use of cannabinoids, endocannabinoids, and synthetic analogs in NMDs, drawing from an array of compelling studies.

[SARS-CoV-2 antibody response to the second COVID-19 vaccination in neuromuscular disease patients under immune modulating treatment]. / SARS-CoV-2-Antikörper-Antwort auf die zweite COVID-19-Impfung bei neuromuskulären Patienten unter immunmodulierender Therapie.

Successful vaccination (adequate elevation of anti-spike protein antibodies) is attributed with sufficient protection against a severe course of coronavirus disease 2019 (COVID-19). For patients with chronic inflammatory diseases (CID) and immunosuppression the success of vaccination is an ongoing scientific discourse. Therefore, we evaluated the antibody titer against the S1 antigen of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 2 weeks after complete immunization in patients with an underlying neuromuscular disease (NMD), who presented to our neurological day clinic and outpatient department for regular infusions of immunoglobulins. The data show that patients with chronic autoimmune NMD and simultaneous immunosuppressive or immune modulating treatment show an antibody response after vaccination with both mRNA and vector vaccines. In comparison to healthy subjects there is a comparable number of seroconversions due to the vaccination. A correlation between immunoglobulin dose and vaccination response could not be found; however, in contrast, there was a significant reduction of specific antibody synthesis, especially for the combination of mycophenolate mofetil (MMF) and prednisolone.

Classical and Unexpected Effects of Ultra-Micronized PEA in Neuromuscular Function.

Recently, the endocannabinoid system has attracted growing attention from the scientific community for its involvement in homeostatic and pathological processes as they pertains to human physiology. Among the constituents of the endocannabinoid system, the molecule palmitoyl ethanolamide has particularly been studied for its ability to reduce several inflammatory processes involving the central nervous system. Here, we reviewed published literature and summarized the main targets of the palmitoyl ethanolamide, along with its unique possible mechanisms for restoring correct functioning of the central nervous system. Moreover, we have highlighted a less-known characteristic of palmitoyl ethanolamide, namely its ability to modulate the function of the neuromuscular junction by binding to acetylcholine receptors in different experimental conditions. Indeed, there are several studies that have highlighted how ultra-micronized palmitoyl ethanolamide is an interesting nutraceutical support for the treatment of pathological neuromuscular conditions, specifically when the normal activity of the acetylcholine receptor is altered. Although further multicentric clinical trials are needed to confirm the efficacy of ultra-micronized palmitoyl ethanolamide in improving symptoms of neuromuscular diseases, all the literature reviewed here strongly supports the ability of this endocannabinoid-like molecule to modulate the acetylcholine receptors thus resulting as a valid support for the treatment of human neuromuscular diseases.

Clinical advances of RNA therapeutics for treatment of neurological and neuromuscular diseases.

RNA therapeutics comprise a diverse group of oligonucleotide-based drugs such as antisense oligonucleotides (ASOs), small interfering RNAs (siRNAs), and short hairpin RNAs (shRNAs) that can be designed to selectively interact with drug targets currently undruggable with small molecule-based drugs or monoclonal antibodies. Furthermore, RNA-based therapeutics have the potential to modulate entire disease pathways, and thereby represent a new modality with unprecedented potential for generating disease-modifying drugs for a wide variety of human diseases, including central nervous system (CNS) disorders. Here, we describe different strategies for delivering RNA drugs to the CNS and review recent advances in clinical development of ASO drugs and siRNA-based therapeutics for the treatment of neurological diseases and neuromuscular disorders.Abbreviations 2'-MOE: 2'-O-(2-methoxyethyl); 2'-O-Me: 2'-O-methyl; 2'-F: 2'-fluoro; AD: Alzheimer's disease; ALS: Amyotrophic lateral sclerosis; ALSFRS-R: Revised Amyotrophic Lateral Sclerosis Functional Rating Scale; ARC: Antibody siRNA Conjugate; AS: Angelman Syndrome; ASGRP: Asialoglycoprotein receptor; ASO: Antisense oligonucleotide; AxD: Alexander Disease; BBB: Blood brain barrier; Bp: Basepair; CNM: Centronuclear myopathies; CNS: Central Nervous System; CPP: Cell-penetrating Peptide; CSF: Cerebrospinal fluid; DMD: Duchenne muscular dystrophy; DNA: Deoxyribonucleic acid; FAP: Familial amyloid polyneuropathy; FALS: Familial amyotrophic lateral sclerosis; FDA: The United States Food and Drug Administration; GalNAc: N-acetylgalactosamine; GoF: Gain of function; hATTR: Hereditary transthyretin amyloidosis; HD: Huntington's disease; HRQOL: health-related quality of life; ICV: Intracerebroventricular; IT: Intrathecal; LNA: Locked nucleic acid; LoF: Loss of function; mRNA: Messenger RNA; MS: Multiple Sclerosis; MSA: Multiple System Atrophy; NBE: New Biological Entity; NCE: New Chemical Entity; NHP: Nonhuman primate; nt: Nucleotide; PD: Parkinson's disease; PNP: Polyneuropathy; PNS: Peripheral nervous system; PS: Phosphorothioate; RISC: RNA-Induced Silencing Complex; RNA: Ribonucleic acid; RNAi: RNA interference; s.c.: Subcutaneous; siRNA: Small interfering RNA; SMA: Spinal muscular atrophy; SMN: Survival motor neuron; TTR: Transthyretin.

The Evolution of Complex Muscle Cell In Vitro Models to Study Pathomechanisms and Drug Development of Neuromuscular Disease.

Many neuromuscular disease entities possess a significant disease burden and therapeutic options remain limited. Innovative human preclinical models may help to uncover relevant disease mechanisms and enhance the translation of therapeutic findings to strengthen neuromuscular disease precision medicine. By concentrating on idiopathic inflammatory muscle disorders, we summarize the recent evolution of the novel in vitro models to study disease mechanisms and therapeutic strategies. A particular focus is laid on the integration and simulation of multicellular interactions of muscle tissue in disease phenotypes in vitro. Finally, the requirements of a neuromuscular disease drug development workflow are discussed with a particular emphasis on cell sources, co-culture systems (including organoids), functionality, and throughput.

Distinct mechanisms underlying therapeutic potentials of CD20 in neurological and neuromuscular disease.

Cluster of differentiation 20 (CD20) is an integral membrane protein expressed mainly on different developmental stages of B lymphocytes and rarely on T lymphocytes, and it functions as a link to B cell antigen receptor (BCR) and immune microenvironment via regulating calcium ion influx, cell cycle progression and interaction between isotypic BCRs and their co-receptors. Diverse therapeutic monoclonal antibodies (mAbs) targeting CD20 are generated and grouped into two types based on the ability to redistribute CD20 into lipid rafts, which results in huge differences in response. Currently, multiple anti-CD20 mAbs have been approved as drugs for neurological and neuromuscular diseases with promising clinical efficacy. This review aims to summarize the potential mechanisms, development and current evidence for anti-CD20 therapy in neurological and neuromuscular diseases.

[Intravenous immunoglobulin-induced eczematous eruption in autoimmune neuromuscular diseases].

BACKGROUND: Intravenous immunoglobulin (IVIg) have been administrated for the long time in patients with several autoimmune neuromuscular diseases. Eczematous eruption has been described as IVIg-induced adverse effect. OBJECTIVE: The purpose of this study is to clarify the incidence and characteristic of IVIg-induced eczematous eruption in autoimmune neuromuscular disease. METHODS: We retrospectively collected the data from 92 patients with autoimmune neuromuscular diseases, including 35 patients with chronic inflammatory demyelinating polyneuropathy (CIDP), 8 patients with multifocal motor neuropathy (MMN), 25 patients with myositis, 15 patients with Guillain-Barré syndrome (GBS), and 9 patients with myasthenia gravis (MG), who have administrated IVIg in Yamaguchi University Hospital. RESULTS: There are 10 patients (6 CIDP/4 MMN), who had an eczematous skin reaction after IVIg infusion. The frequencies of IVIg-induced eczematous eruption were significantly higher in patients with multifocal acquired demyelinating sensory and motor (MADSAM) and MMN than in patients with GBS, myositis, and MG. In addition, corticosteroids or immunosuppressive drugs had been administrated before IVIg treatment more frequently in patients with myositis and MG than in those with MADSAM and MMN. CONCLUSION: MADSAM or MMN patients had more frequently IVIg-induced eczematous eruption than other autoimmune neuromuscular diseases. Pathophysiology of MADAM and MMN is considered to be cell-mediated immunity against the peripheral nerve and the accumulation of IgG in both epidermis and dermis of the hand after IVIg may induce the infiltration of inflammatory cells around the vessels in the skin, causing eczematous eruption in MADSAM and MMN.

mRNA intramuscular vaccination produces a robust IgG antibody response in advanced neuromuscular disease.

SARS-CoV-2 vaccines protect against symptomatic and severe COVID-19. The BNT162b2/Pfizer and mRNA-1273/Moderna vaccines represent new vaccine technology relying on administration of mRNA encoding SARS-CoV-2 viral spike protein encased in lipid nanoparticles. The vaccines are administered as two doses into muscle, which elicits a strong response, typically within 14 days after the second dose. Neuromuscular diseases are characterized by the progressive loss of muscle and are often treated with chronic glucocorticoid steroids, both of which may contribute to a blunted immune response to vaccination. Here, we measured IgG antibody content and neutralizing antibody response after mRNA COVID-19 vaccination in non-ambulatory neuromuscular disease patients. After two doses of mRNA COVID-19 vaccine, median anti-receptor binding domain IgG and percent surrogate viral neutralization in non-ambulatory neuromuscular disease samples were significantly elevated similar to healthy vaccinated controls. As in healthy controls, COVID-19 vaccines produce greater antibody levels compared to those with a history of outpatient COVID-19 infection. This data documents that non-ambulatory neuromuscular disease patients respond well to two doses of mRNA COVID-19 vaccine despite low muscle mass and even chronic steroid use.

Practical Aspects of Transitioning from Intravenous to Subcutaneous Immunoglobulin Therapy in Neuromuscular Disorders.

Recent evidence shows that subcutaneous immunoglobulin (SCIG) is as efficacious as intravenous immunoglobulin (IVIG) and has a better safety profile and acceptance rate among patients with neuromuscular disorders who require maintenance IVIG treatment. Awareness of the practical aspects of patient selection, enrollment, dose calculation, administration, and follow-up would help physicians coordinate a smooth and seamless transition from IVIG to SCIG. SCIG is ideally offered to patients having intolerable side effects during IVIG or wearing-off effect and in those keen for treatment autonomy. The weekly dose of SCIG is calculated by multiplying the maintenance dose of IVIG by the dose adjustment factor and dividing by the interval between IVIG in weeks and is initiated 1 week after the last dose of IVIG. The physician places the order for the SCIG and the clinic nurse or the physician refers the patient to the home care nursing program for further education and training. The necessary supplies are dispatched to the patient who would also collect the SCIG from the transfusion center of the nearest hospital. The patient is educated on assembling and administering the infusion, and home visits are continued until the patient or caregiver is confident. Regular follow-up with the patient is maintained to assess treatment response and side effects if any. With a smooth transition, most patients have excellent tolerance to SCIG and in our experience seldom request switching back to IVIG. Transitioning patients from IVIG to SCIG offers several advantages and thus, in general, is preferable for multiple stakeholders.

Immunosuppression and immunization: Vaccination in pediatric patients with neuromuscular diseases treated with steroids or immune-modulating drugs.

BACKGROUND: Modulating the immune response has proven to be beneficial in different neurologic diseases, even in those not perceived thus far to be autoimmune. METHODS: Extensive literature search has been done for available data on vaccine safety, efficacy and immunization recommendations in patients with neuromuscular disease in general and when receiving immune-modulating treatments. RESULTS: Vaccinations have been associated with some neuromuscular diseases, but these occurrences are very rare and should not influence the general vaccination recommendations for the pediatric population and for the especially vulnerable patient populations, such as neuromuscular disease patients. Specific guidelines for the immunization of children with neuromuscular diseases in general and those on immune-suppressive treatments were not found, but most guidelines and standards of care for specific neuromuscular diseases recognize and stress the importance of vaccinations, some giving more specific instructions. CONCLUSION: With just a few exceptions, vaccines are safe in this group of patients and they should receive the same immunizations and according to the same schedule, as all children. Live vaccines should not be administered in patients receiving high dose steroid or immune-modulating drugs such as anti-B cell treatments (rituximab), high dose methotrexate, azathioprine or 6-mercaptopurine. Whenever possible, all live vaccines should be administered prior to long term immune-suppressant treatments. Additional vaccines are recommended in this risk population of children (influenza, pneumococcal, varicella).

The emerging role of complement in neuromuscular disorders.

The complement cascade is a key arm of the immune system that protects the host from exogenous and endogenous toxic stimuli through its ability to potently regulate inflammation, phagocytosis, and cell lysis. Due to recent clinical trial successes and drug approvals for complement inhibitors, there is a resurgence in targeting complement as a therapeutic approach to prevent ongoing tissue destruction in several diseases. In particular, neuromuscular diseases are undergoing a recent focus, with demonstrated links between complement activation and disease pathology. This review aims to provide a comprehensive overview of complement activation and its role during the initiation and progression of neuromuscular disorders including myasthenia gravis, amyotrophic lateral sclerosis, and Duchenne muscular dystrophy. We will review the preclinical and clinical evidence for complement in these diseases, with an emphasis on the complement-targeting drugs in clinical trials for these indications.

A Retrospective Observational Study of Adverse Reactions Associated With Intravenous Immunoglobulin Infusion.

Background: Although intravenous immunoglobulin (IVIG) therapy is generally safe and well tolerated, adverse reactions (ARs) do occur. The majority of these ARs are mild and transient. Risk factors for ARs associate with IVIG infusions are not well established. This study investigated possible risk factors influencing the occurrence of IVIG-associated ARs. Study Design and Methods: This was a retrospective observational analysis of data accumulated over 5 years, including patient demographics, clinical condition, IVIG dosing regimens, number of IVIG infusions, and any ARs. Results: ARs were associated with IVIG in 4.9% of patients and 2.5% of infusions. By univariate analyses, ARs correlated with female sex, adult age, high dose IVIG, and autoimmune disease. Multivariate logistic regression identified three statistically significant of risk factors: on a per-patient basis, being female (p=0.0018), having neuromuscular disease (p=0.0002), and receiving higher doses of IVIG per patient body weight (p<0.001), on a per-infusion basis, being female (p < 0.001), being adolescents to middle age (p < 0.001), and having neuromuscular disease (p < 0.001). Conclusion: Neuromuscular disease emerged as one of the significant factors for ARs to IVIG.

Botulinum Toxin Services for Neurorehabiliation: Recommendations for Challenges and Opportunities during the COVID-19 Pandemic.

The COVID-19 pandemic severely impacted the function of medical facilities and rehabilitation services worldwide, including toxin services delivering Botulinum toxin treatments for neuromuscular conditions such as spasticity, dystonia, and sialorrhea. The aim of this paper is to understand how toxin services have dealt with the situation and what strategies have been adopted to continue services. The recommendations are based on a virtual round table held with toxin services experts from different European countries who shared their experiences and discussed the best practices. The challenges for toxin services were reviewed based on the experts' experiences and on relevant literature from 2020 and 2021. A set of recommendations and best practices were compiled, focusing firstly on guidance for clinical practice, including assessing patients' health and risk status and the urgency of their treatment. Secondly, it was discussed how patients on botulinum toxin therapy can be cared for and supported during the pandemic, and how modern technology and tele-medicine platforms can be generally used to optimize effectiveness and safety of toxin treatments. The technological advances prompted by the COVID-19 crisis can result in better and more modern patient care in the future.

Enterovirus A71 causing meningoencephalitis and acute flaccid myelitis in a patient receiving rituximab.

We present the case of a young woman being treated with rituximab for rheumatoid arthritis who developed a severe enteroviral meningoencephalitis and acute flaccid myelitis (AFM). Cerebrospinal fluid (CSF) and stool reverse transcription-polymerase chain reaction (RT-PCR) testing confirmed the diagnosis and additional sequencing studies performed at the CDC further characterized the enterovirus as enterovirus A71 (EV-A71). After treatment with intravenous immunoglobulin (IVIg) and fluoxetine (based on previous reports of possible efficacy) the patient experienced a remarkable improvement over time. This case highlights the importance of considering enteroviral infection in patients treated with rituximab, depicts a possible clinical course of enteroviral meningoencephalitis and AFM, and illustrates the importance of testing multiple sites for enterovirus infection (CSF, stool, nasopharyngeal swab, blood). Here we present the case with a brief review of the literature pertaining to EV-A71.

Contribution of Single-Fiber Evaluation on Monitoring Outcomes Following Injection of Botulinum Toxin-A: A Narrative Review of the Literature.

Botulinum toxin-A (BoNT-A) blocks acetylcholine release at the neuromuscular junction (NMJ) and is widely used for neuromuscular disorders (involuntary spasms, dystonic disorders and spasticity). However, its therapeutic effects are usually measured by clinical scales of questionable validity. Single-fiber electromyography (SFEMG) is a sensitive, validated diagnostic technique for NMJ impairment such as myasthenia. The jitter parameter (µs) represents the variability of interpotential intervals of two muscle fibers from the same motor unit. This narrative review reports SFEMG use in BoNT-A treatment. Twenty-four articles were selected from 175 eligible articles searched in Medline/Pubmed and Cochrane Library from their creation until May 2020. The results showed that jitter is sensitive to early NMJ modifications following BoNT-A injection, with an increase in the early days' post-injection and a peak between Day 15 and 30, when symptoms diminish or disappear. The reappearance of symptoms accompanies a tendency for a decrease in jitter, but always precedes its normalization, either delayed or nonexistent. Increased jitter is observed in distant muscles from the injection site. No dose effect relationship was demonstrated. SFEMG could help physicians in their therapeutic evaluation according to the pathology considered. More data are needed to consider jitter as a predictor of BoNT-A clinical efficacy.