Muscle-specific Kinase Myasthenia Gravis Research Articles

A retrospective multicenter study on clinical and serological parameters in patients with MuSK myasthenia gravis with and without general immunosuppression.

Muscle-specific kinase (MuSK)- myasthenia gravis (MG) is caused by pathogenic autoantibodies against MuSK that correlate with disease severity and are predominantly of the IgG4 subclass. The first-line treatment for MuSK-MG is general immunosuppression with corticosteroids, but the effect of treatment on IgG4 and MuSK IgG4 levels has not been studied. We analyzed the clinical data and sera from 52 MuSK-MG patients (45female, 7 male, median age 49 (range 17-79) years) from Italy, the Netherlands, Greece and Belgium, and 43 AChR-MG patients (22 female, 21 male, median age 63 (range 2-82) years) from Italy, receiving different types of immunosuppression, and sera from 46 age- and sex-matched non-disease controls (with no diagnosed diseases, 38 female, 8 male, median age 51.5 (range 20-68) years) from the Netherlands. We analyzed the disease severity (assessed by MGFA or QMG score), and measured concentrations of MuSK IgG4, MuSK IgG, total IgG4 and total IgG in the sera by ELISA, RIA and nephelometry. We observed that MuSK-MG patients showed a robust clinical improvement and reduction of MuSK IgG after therapy, and that MuSK IgG4 concentrations, but not total IgG4 concentrations, correlated with clinical severity. MuSK IgG and MuSK IgG4 concentrations were reduced after immunosuppression in 4/5 individuals with before-after data, but data from non-linked patient samples showed no difference. Total serum IgG4 levels were within the normal range, with IgG4 levels above threshold (1.35g/L) in 1/52 MuSK-MG, 2/43 AChR-MG patients and 1/45 non-disease controls. MuSK-MG patients improved within the first four years after disease onset, but no further clinical improvement or reduction of MuSK IgG4 were observed four years later, and only 14/52 (26.92%) patients in total, of which 13 (93.3%) received general immunosuppression, reached clinical remission. We conclude that MuSK-MG patients improve clinically with general immunosuppression but may require further treatment to reach remission. Longitudinal testing of individual patients may be clinically more useful than single measurements of MuSK IgG4. No significant differences in the serum IgG4 concentrations and IgG4/IgG ratio between AChR- and MuSK-MG patients were found during follow-up. Further studies with larger patient and control cohorts are necessary to validate the findings.

Change of voltage-gated sodium channel repertoire in skeletal muscle of a MuSK myasthenia gravis mouse model.

Muscle-specific kinase myasthenia gravis (MuSK MG) is caused by autoantibodies against MuSK in the neuromuscular junction (NMJ). MuSK MG patients have fluctuating, fatigable skeletal muscle weakness, in particular of bulbar muscles. Severity differs greatly between patients, in spite of comparable autoantibody levels. One explanation for inter-patient and inter-muscle variability in sensitivity might be variations in compensatory muscle responses. Previously, we developed a passive transfer mouse model for MuSK MG. In preliminary ex vivo experiments, we observed that muscle contraction of some mice, in particular those with milder myasthenia, had become partially insensitive to inhibition by μ-Conotoxin-GIIIB, a blocker of skeletal muscle NaV1.4 voltage-gated sodium channels. We hypothesised that changes in NaV channel expression profile, possibly co-expression of (μ-Conotoxin-GIIIB insensitive) NaV1.5 type channels, might lower the muscle fibre's firing threshold and facilitate neuromuscular synaptic transmission. To test this hypothesis, we here performed passive transfer in immuno-compromised mice, using 'high', 'intermediate' and 'low' dosing regimens of purified MuSK MG patient IgG4. We compared myasthenia levels, μ-Conotoxin-GIIIB resistance and muscle fibre action potential characteristics and firing thresholds. High- and intermediate-dosed mice showed clear, progressive myasthenia, not seen in low-dosed animals. However, diaphragm NMJ electrophysiology demonstrated almost equal myasthenic severities amongst all regimens. Nonetheless, low-dosed mouse diaphragms showed a much higher degree of μ-Conotoxin-GIIIB resistance. This was not explained by upregulation of Scn5a (the NaV1.5 gene), lowered muscle fibre firing thresholds or histologically detectable upregulated NaV1.5 channels. It remains to be established which factors are responsible for the observed μ-Conotoxin-GIIIB insensitivity and whether the NaV repertoire change is compensatory beneficial or a bystander effect.

MuSK-CAART: A novel precision cellular therapy for muscle-specific tyrosine kinase myasthenia gravis

Chimeric autoantibody receptor (CAAR) T cells are a novel genetically-engineered T cell immunotherapy that aims to durably eliminate antigen-specific B cells while sparing healthy B cells, ideally leading to safe and lasting remission of B cell-mediated autoimmune diseases with a one-time infusion. We describe the preclinical development of muscle-specific tyrosine kinase CAAR T cells (MuSK-CAART) for the treatment of MuSK myasthenia gravis, a debilitating autoantibody-mediated disease that causes potentially life-threatening muscle weakness.

Development and characterization of agonistic antibodies targeting the Ig-like 1 domain of MuSK

Muscle-specific kinase (MuSK) is crucial for acetylcholine receptor (AChR) clustering and thereby neuromuscular junction (NMJ) function. NMJ dysfunction is a hallmark of several neuromuscular diseases, including MuSK myasthenia gravis. Aiming to restore NMJ function, we generated several agonist monoclonal antibodies targeting the MuSK Ig-like 1 domain. These activated MuSK and induced AChR clustering in cultured myotubes. The most potent agonists partially rescued myasthenic effects of MuSK myasthenia gravis patient IgG autoantibodies in vitro. In an IgG4 passive transfer MuSK myasthenia model in NOD/SCID mice, MuSK agonists caused accelerated weight loss and no rescue of myasthenic features. The MuSK Ig-like 1 domain agonists unexpectedly caused sudden death in a large proportion of male C57BL/6 mice (but not female or NOD/SCID mice), likely caused by a urologic syndrome. In conclusion, these agonists rescued pathogenic effects in myasthenia models in vitro, but not in vivo. The sudden death in male mice of one of the tested mouse strains revealed an unexpected and unexplained role for MuSK outside skeletal muscle, thereby hampering further (pre-) clinical development of these clones. Future research should investigate whether other Ig-like 1 domain MuSK antibodies, binding different epitopes, do hold a safe therapeutic promise.

DaT Scan Positive Parkinson’s Disease in a Patient with Anti-MuSK Positive Myasthenia Gravis (P5-11.007)

<h3>Objective:</h3> We present a unique clinical case of antibody positive Muscle Specific Kinase (MuSK) Myasthenia Gravis and DaT scan positive Parkinson’s Disease to highlight the overlapping nature of clinical symptoms and challenges seen in clinical diagnosis. <h3>Background:</h3> The concomitant diagnosis of Myasthenia Gravis (MG), an autoimmune condition that affects the neuromuscular junction, and Parkinson’s disease (PD), a neurodegenerative disease that affects the basal ganglia, is remarkably exceptional. Furthermore, there are very few case reports in the literature that have recognized muscle-specific kinase (MuSK), a subtype of MG with overlapping features of PD. We present a unique case of MuSK MG and DaT scan positive Parkinson’s Disease, which to our knowledge has not been previously reported. <h3>Design/Methods:</h3> NA <h3>Results:</h3> 49-year-old male initially presented to neurology clinic with six months of waxing and waning nasal quality dysarthria. After thorough workup with serologic and electrophysiologic studies, patient was diagnosed with seropositive bulbar predominant MuSK Myasthenia Gravis for which steroid sparing immunosuppressive treatment was initiated with mycophenolate mofetil. He subsequently developed clinical exam findings of hypomimia, bilateral bradykinesia, asymmetrical limb rigidity, right arm and jaw tremor that raised concern for the possibility of parkinsonism. Review of systems for premotor symptoms was positive for constipation and dream enactment behavior. There was no previous exposure to anti-dopaminergic therapies and MRI Brain was unrevealing. DaT scan was subsequently obtained, which demonstrated objective evidence of bilateral striatal dopaminergic deficiency, indicative of an inherent basal ganglia disorder. In the absence of any clinical findings of atypical parkinsonism, patient met clinical diagnostic criteria for Idiopathic Parkinson’s Disease. <h3>Conclusions:</h3> This case demonstrates the clinical challenges that present in this overlap of neuromuscular and neurodegenerative disease in a single individual. We highlight the importance of a thorough history and physical examination, supported with objective evidence by a positive DaT scan, to elucidate the appropriate etiology of symptoms. <b>Disclosure:</b> Dr. Andraos has nothing to disclose. Dr. Veerappan has nothing to disclose.

Precision targeting of autoantigen-specific B cells in muscle-specific tyrosine kinase myasthenia gravis with chimeric autoantibody receptor T cells.

Muscle-specific tyrosine kinase myasthenia gravis (MuSK MG) is an autoimmune disease that causes life-threatening muscle weakness due to anti-MuSK autoantibodies that disrupt neuromuscular junction signaling. To avoid chronic immunosuppression from current therapies, we engineered T cells to express a MuSK chimeric autoantibody receptor with CD137-CD3ζ signaling domains (MuSK-CAART) for precision targeting of B cells expressing anti-MuSK autoantibodies. MuSK-CAART demonstrated similar efficacy as anti-CD19 chimeric antigen receptor T cells for depletion of anti-MuSK B cells and retained cytolytic activity in the presence of soluble anti-MuSK antibodies. In an experimental autoimmune MG mouse model, MuSK-CAART reduced anti-MuSK IgG without decreasing B cells or total IgG levels, reflecting MuSK-specific B cell depletion. Specific off-target interactions of MuSK-CAART were not identified in vivo, in primary human cell screens or by high-throughput human membrane proteome array. These data contributed to an investigational new drug application and phase 1 clinical study design for MuSK-CAART for the treatment of MuSK autoantibody-positive MG.

Plasma Myokine Profiles in Patients With AChR- and MuSK-Ab-Positive Myasthenia Gravis.

Myokines include cytokines secreted by muscle fibers, which are the final targets of myasthenia gravis (MG). This pilot study investigated whether myokine plasma concentrations are altered in patients with MG and assessed the association between the concentration of each myokine and disease severity. We compared the plasma concentrations of 15 myokines in 63 patients with acetylcholine receptor antibody (Ab)-positive MG and 14 with muscle-specific tyrosine kinase Ab-positive MG (MuSK MG) with those in 15 healthy controls. Plasma myokine concentrations were measured using a Luminex multiplex assay kit with magnetic beads that contained Abs for 15 myokines. Correlations between myokine concentration and clinical scale results were analyzed. The concentration of fractalkine in plasma was higher in MG (median [interquartile range]=419.6 [38.7-732.5] pg/mL) than in controls (158.5 [0.0-313.2] pg/mL, p=0.034). The leukemia inhibitory factor concentration was also found to be higher in MuSK MG (29.9 [8.7-40.1] pg/mL) than in healthy controls (7.6 [0.0-15.6] pg/mL, p=0.013). Fatty-acid-binding protein 3 (FABP3) concentrations in plasma were positively associated with clinical parameters for MG severity, including scores on the Quantitative Myasthenia Gravis score (p=0.008), Myasthenia Gravis Activities of Daily Living (p=0.003), and Myasthenia Gravis Composite (p=0.024) scales. FABP3 concentration in plasma tended to decrease after treatment in patients without additional relapse but increased in those with further relapse. The plasma myokine profile was significantly altered in patients with MG. FABP3 concentration may be useful in assessing disease severity and predicting the treatment response.

Clinical pitfalls and serological diagnostics of MuSK myasthenia gravis

BackgroundWe aimed to evaluate the diagnostic accuracy of enzyme-linked immunosorbent assay (ELISA) for anti-muscle specific tyrosine kinase (MuSK) antibody (Ab) in a large cohort of anti-acetylcholine receptor (AChR) Ab-negative generalized myasthenia gravis (MG), and also to investigate clinical contexts for the diagnosis of MuSK MG.MethodsA retrospective study of 160 patients with a clinical suspicion of AChR Ab-negative generalized MG was performed. The serum samples were tested for anti-clustered AChR Ab by cell-based assay (CBA), anti-MuSK Ab by ELISA, CBA and/or radioimmunoprecipitation assay (RIPA). Clinical data were compared between anti-MuSK Ab-positive MG and double seronegative (AChR and MuSK) MG groups.ResultsAfter excluding non-MG and clustered AChR Ab-positive patients, we identified 89 patients as a cohort of AChR Ab-negative generalized MG. Anti-MuSK Ab was positive by ELISA in 22 (24.7%) patients. While CBA identified five additional anti-MuSK Ab-positive patients, the results of ELISA were mostly consistent with CBA and RIPA with Cohen’s kappa of 0.80 and 0.90, respectively (p < 0.001). The most frequent differential diagnosis was motor neuron disease particularly of bulbar onset which showed remarkably overlapping clinical and electrophysiological features with MuSK MG at presentation.ConclusionWhile confirming the highest sensitivity of CBA for detecting anti-MuSK Ab, our results highlight the clinical pitfalls in making a diagnosis of MuSK MG and may support a diagnostic utility of MuSK-ELISA in clinical practice.

Reemergence of pathogenic, autoantibody-producing B cell clones in myasthenia gravis following B cell depletion therapy

Myasthenia gravis (MG) is an autoantibody-mediated autoimmune disorder of the neuromuscular junction. A small subset of patients (<10%) with MG, have autoantibodies targeting muscle-specific tyrosine kinase (MuSK). MuSK MG patients respond well to CD20-mediated B cell depletion therapy (BCDT); most achieve complete stable remission. However, relapse often occurs. To further understand the immunomechanisms underlying relapse, we studied autoantibody-producing B cells over the course of BCDT. We developed a fluorescently labeled antigen to enrich for MuSK-specific B cells, which was validated with a novel Nalm6 cell line engineered to express a human MuSK-specific B cell receptor. B cells (≅ 2.6 million) from 12 different samples collected from nine MuSK MG patients were screened for MuSK specificity. We successfully isolated two MuSK-specific IgG4 subclass-expressing plasmablasts from two of these patients, who were experiencing a relapse after a BCDT-induced remission. Human recombinant MuSK mAbs were then generated to validate binding specificity and characterize their molecular properties. Both mAbs were strong MuSK binders, they recognized the Ig1-like domain of MuSK, and showed pathogenic capacity when tested in an acetylcholine receptor (AChR) clustering assay. The presence of persistent clonal relatives of these MuSK-specific B cell clones was investigated through B cell receptor repertoire tracing of 63,977 unique clones derived from longitudinal samples collected from these two patients. Clonal variants were detected at multiple timepoints spanning more than five years and reemerged after BCDT-mediated remission, predating disease relapse by several months. These findings demonstrate that a reservoir of rare pathogenic MuSK autoantibody-expressing B cell clones survive BCDT and reemerge into circulation prior to manifestation of clinical relapse. Overall, this study provides both a mechanistic understanding of MuSK MG relapse and a valuable candidate biomarker for relapse prediction.

Enrichment of serum IgG4 in MuSK myasthenia gravis patients

Muscle-specific kinase (MuSK) myasthenia gravis (MG) is a neuromuscular autoimmune disease belonging to a growing group of IgG4 autoimmune diseases (IgG4-AIDs), in which the majority of pathogenic autoantibodies are of the IgG4 subclass. The more prevalent form of MG with acetylcholine receptor (AChR) antibodies is caused by IgG1–3 autoantibodies. A dominant role for IgG4 in autoimmune disease is intriguing due to its anti-inflammatory characteristics. It is unclear why MuSK autoantibodies are predominantly IgG4. We hypothesized that MuSK MG patients have a general predisposition to generate IgG4 responses, therefore resulting in high levels of circulating IgG4. To investigate this, we quantified serum Ig isotypes and IgG subclasses using nephelometric and turbidimetric assays in MuSK MG and AChR MG patients not under influence of immunosuppressive treatment. Absolute serum IgG1 was increased in both MuSK and AChR MG patients compared to healthy donors. In addition, only MuSK MG patients on average had significantly increased and enriched serum IgG4. Although more MuSK MG patients had elevated serum IgG4, for most the IgG4 serum levels fell within the normal range. Correlation analyses suggest MuSK-specific antibodies do not solely explain the variation in IgG4 levels. In conclusion, although serum IgG4 levels are slightly increased, the levels do not support ubiquitous IgG4 responses in MuSK MG patients as the underlying cause of dominant IgG4 MuSK antibodies.

Novel pathophysiological insights in autoimmune myasthenia gravis.

This review summarizes recent insights into the immunopathogenesis of autoimmune myasthenia gravis (MG). Mechanistic understanding is presented according to MG disease subtypes and by leveraging the knowledge gained through the use of immunomodulating biological therapeutics. The past two years of research on MG have led to a more accurate definition of the mechanisms through which muscle-specific tyrosine kinase (MuSK) autoantibodies induce pathology. Novel insights have also emerged from the collection of stronger evidence on the pathogenic capacity of low-density lipoprotein receptor-related protein 4 autoantibodies. Clinical observations have revealed a new MG phenotype triggered by cancer immunotherapy, but the underlying immunobiology remains undetermined. From a therapeutic perspective, MG patients can now benefit from a wider spectrum of treatment options. Such therapies have uncovered profound differences in clinical responses between and within the acetylcholine receptor and MuSK MG subtypes. Diverse mechanisms of immunopathology between the two subtypes, as well as qualitative nuances in the autoantibody repertoire of each patient, likely underpin the variability in therapeutic outcomes. Although predictive biomarkers of clinical response are lacking, these observations have ignited the development of assays that might assist clinicians in the choice of specific therapeutic strategies. Recent advances in the understanding of autoantibody functionalities are bringing neuroimmunologists closer to a more detailed appreciation of the mechanisms that govern MG pathology. Future investigations on the immunological heterogeneity among MG patients will be key to developing effective, individually tailored therapies.

A systematic review and meta-analysis of HLA class II associations in patients with IgG4 autoimmunity

Autoimmune diseases caused by pathogenic IgG4 subclass autoantibodies (IgG4-AID) include diseases like MuSK myasthenia gravis, pemphigus vulgaris or thrombotic thrombocytopenic purpura. Their etiology is still unknown. Polymorphisms in the human leukocyte antigen (HLA) gene locus, particularly in HLA-DRB1, are known genetic susceptibility factors for autoimmune diseases. We hypothesized a similar role for HLA polymorphisms in IgG4-AID and conducted a systematic review and meta-analysis with case–control studies on IgG4-AID based on MOOSE/ HuGENet guidelines. Genotype (G) and allele (A) frequencies of HLA-DQB1*05 (G: OR 3.8; 95% CI 2.44–5.9; p < 0.00001; A: OR 2.54; 95% CI 1.82–3.55; p < 0.00001) and HLA-DRB1*14 (G: OR 4.31; 95% CI 2.82–6.59; p < 0.00001; A: OR 4.78; 95% CI 3.52–6.49; p < 0.00001) and the HLA-DRB1*14-DQB1*05 haplotype (OR 6.3; 95% CI 3.28–12.09; p < 0.00001/OR 4.98; 95% CI 3.8–6.53; p < 0.00001) were increased while HLA-DRB1*13 (G: OR 0.48; 95% CI 0.34–0.68; p < 0.0001; A: OR 0.46; 95% CI 0.34–0.62; p < 0.00001) was decreased in IgG4-AID patients. In conclusion, the HLA-DQB1*05, HLA-DRB1*14 alleles and the HLA-DQB1*05-DRB1*14 haplotype could be genetic risk factors that predispose for the production of pathogenic IgG4 autoantibodies and the HLA-DRB1*13 allele may protect from IgG4 autoimmunity.

Isolated vocal cord paralysis mimicking respiratory weakness in MuSK myasthenia

Vocal cord paralysis is a rare manifestation of myasthenia gravis and challenging to diagnose with presentations other than stridor. We discuss a patient with MuSK myasthenia gravis who had respiratory difficulty due to vocal cord palsy and the diagnostic difficulties encountered in establishing the etiology for the dyspnea. The diagnosis was captured by spirometry flow volume loop showing flattening of the inspiratory portion suggestive of extrathoracic upper airway obstruction. Early diagnosis can prevent the need for tracheostomy and other emergent measures.

Safety and tolerability of SARS-Cov-2 vaccination in patients with myasthenia gravis: A multicenter experience.

Background and purposeDuring the COVID‐19 pandemic, myasthenia gravis (MG) patients have been identified as subjects at high risk of developing severe COVID‐19, and thus were offered vaccination with priority. The lack of direct data on the safety and tolerability of SARS‐CoV‐2 vaccines in MG have contributed to vaccine hesitancy. To address this issue, the safety and tolerability of SARS‐CoV‐2 vaccines were assessed in a large cohort of MG patients from two referral centers.MethodsPatients with confirmed MG diagnosis, consecutively seen between October and December 2021 at two MG centers, were enrolled. Demographics, clinical characteristics, and information regarding SARS‐CoV‐2 infection/vaccination were extracted from medical reports and/or collected throughout telephonic or in‐person interviews.ResultsNinety‐eight (94.2%) of 104 patients included were administered at least two vaccine doses 4 weeks before the interview or earlier, and among them, 63 of 98 (64.2%) have already received the “booster” dose. The most frequently used vaccines were BNT162b2‐Pfizer‐BioNTech and mRNA‐1273‐Moderna. Overall, only minor side effects were reported, most commonly local pain and fever. MG worsening after vaccination was observed in eight of 104 (7.7%) cases. The frequency of worsening among muscle‐specific tyrosine kinase MG cases (3/9, 33.3%) was significantly higher compared to other serological subgroups. Spontaneous symptom regression was observed in six of eight cases. Twelve of 104 (11.5%) patients had SARS‐CoV‐2 infection, and none of the SARS‐CoV‐2‐infected MG patients worsened after vaccination.ConclusionsOur data support the safety and tolerability of mRNA COVID‐19 vaccines, which should be strongly recommended in MG patients, who could be at higher risk of complications if exposed to SARS‐CoV‐2 infection.

Seropositive Muscle-Specific Tyrosine Kinase Myasthenia Gravis Presenting as a Late-Onset Isolated Sixth Nerve Palsy: A Case Report and a Brief Review of Subtypes of Myasthenia Gravis

Autoimmune myasthenia gravis (MG) is a well-characterized post-synaptic disorder of neuromuscular transmission. Immunologically, there is complement activation with autoantibodies binding to the acetylcholine receptor (AChR), leading to cross-linking and internalization of the receptor. The diminished functional clustering leads to impaired folding of the post-synaptic membrane. The antibodies generated by the autoimmune process are directed at the various components of the post-synaptic membrane and its scaffolding, including the AChR, muscle-specific tyrosine kinase (MuSK), low-density lipoprotein receptor-related protein 4 (LRP4), and the other recently described epitopes including the extracellular membrane proteins agrin and collagen Q (ColQ). MuSK MG is phenotypically different from classic AChR-antibody-mediated MG by a more frequent presentation of bulbar weakness, less responsiveness to symptomatic therapy with acetylcholinesterase inhibitors, the absence of a thymoma, and a better therapeutic response to a cluster of differentiation (CD-20) B-cell therapy such as rituximab. The pleiotropic ocular findings of ocular MG include ptosis, fluctuating and variable involvement of cranial nerves III, IV, and VI, pseudo-internuclear ophthalmoplegia (INO), near-complete or complete ophthalmoplegia, and variable gaze palsies. To our knowledge, we present one of the very few reported cases of MuSK MG presenting as isolated sixth nerve palsy. The localization of a sixth nerve palsy with lateral rectus muscle weakness can be due to disease anywhere along its path from the abducens nucleus, coursing at the skull base through Dorello's canal, through the cavernous sinus, and along its path through the superior orbital fissure and into the orbits. A painless sixth nerve palsy should alert the clinician to MuSK-MG as we outline in this case report.

To measure musk ABS in ACHR AB negative sera: RIPA is more specific, and SPR is more sensitive

Muscle-specific kinase (MuSK) myasthenia gravis (MG) is often a more severe form of MG that presents with a rapid onset and prominent bulbar symptoms. Today, the diagnosis of MuSK MG is heavily reliant on the antibody (Ab) status of the patient. In most laboratories worldwide, the Abs are measured by radio immune precipitation assay (RIPA). We have developed another testing platform using surface plasmon resonance (SPR) technology. Our aim was to measure the clinical sensitivity and specificity for two different MuSK Ab assays.

WS3.7. Repetitive Nerve Stimulation Testing

Repetitive nerve stimulation (RNS) is the most frequently used electrodiagnostic test to diagnose neuromuscular junction disorders. Although seemingly simple, attention to technique is essential to avoid technical errors. Stimulation at low rates (2–5 Hz) (LF-RNS) depletes acetylcholine stores at diseased motor endplates, causing failure of neuromuscular transmission. A decrement of at least 10% to trains of 2-3 Hz stimulation at rest or immediately after a period of maximal muscle contraction (“activation”) is the usual criterion of abnormality; 5% can be used if technical artifacts can be excluded. Take care to deliver adequate stimulation and prevent movement of the tested muscle throughout nerve stimulation; peripheral muscles should be kept warm during testing. RNS is reportedly abnormal in up to 90% of generalized myasthenia gravis (MG) patients and in up to 60% of ocular myasthenia – sensitivity is strongly dependent on the muscles selected for testing. RNS is more often abnormal in proximal, facial and clinically weak muscles; even so, RNS is frequently normal in mild or purely ocular disease. For maximal diagnostic yield, test several muscles, including those that are weak. The characteristic findings in Lambert-Eaton myasthenia (LEM) are: small CMAPs at rest, a decrement to LF-RNS, and marked facilitation. In MG, activation for 30–60 seconds is used to unmask a mild decrement. In LEM, activation is used to demonstrate facilitation; the simplest procedure is to elicit a single CMAP from the rested muscle, have the patient contract the muscle for 10 seconds, then immediately elicit a single CMAP. Facilitation of at least 60% is evidence of a pre-synaptic disorder, but may not be seen in all muscles. A decrement in at least one hand muscle is the most frequent RNS abnormality in LEM and may be seen even when CMAPs are not small and diagnostic facilitation cannot be demonstrated. In MG the decrement is usually greatest in the 4th or 5th response and is less during subsequent responses, producing a so-called ‘‘U-shaped” pattern. A continually progressive decrement is usually seen in LEM. The pattern in MuSK MG resembles that in LEM.

Evaluating an In-House Cell-Based Assay for Detecting Antibodies Against Muscle-Specific Tyrosine Kinase in Myasthenia Gravis.

Background and PurposeDetecting antibodies against muscle-specific tyrosine kinase (MuSK Abs) is essential for diagnosing myasthenia gravis (MG). We applied an in-house cell-based assay (CBA) to detect MuSK Abs.MethodsA stable cell line was generated using a lentiviral vector, which allowed the expression of MuSK tagged with green fluorescent protein in human embryonic kidney 293 (HEK293) cells. Serum and anti-human IgG antibody conjugated with red fluorescence were added. The presence of MuSK Abs was determined based on the fluorescence intensity and their colocalization in fluorescence microscopy. Totals of 218 serum samples collected from 177 patients with MG, 31 with other neuromuscular diseases, and 10 healthy controls were analyzed. The CBA results were compared with those of a radioimmunoprecipitation assay (RIPA) and an enzyme-linked immunosorbent assay (ELISA).ResultsThe MuSK-HEK293 cell line stably expressed MuSK protein. The CBA detected MuSK Abs in 34 (19.2%) of 177 samples obtained from patients with MG and in none of the participants having other neuromuscular diseases or in the healthy controls. The clinical characteristics of the patients with MuSK MG determined based on the CBA were strongly correlated with known clinical features of MuSK MG. There was an almost perfect agreement between the results of the CBA and those of the RIPA (Cohen's kappa=0.880, p<0.001) and ELISA (Cohen's kappa=0.982, p<0.001).ConclusionsThe results of the in-house CBA showed excellent agreement with both the RIPA and ELISA. Our in-house CBA can be considered a reliable method for detecting MuSK Abs.

Myasthenia gravis: MuSK MG, late-onset MG and ocular MG.

Myasthenia gravis (MG) is an autoimmune disease of the neuromuscular junction which affects all striated muscles, resulting in fluctuating weakness. Approaching MG as a disease with subgroups having different clinical, serological and genetic features is crucial in predicting the progression and planning treatment. Three relatively less frequently seen subtypes of MG are the subject of this review: MG with anti-MuSK antibodies (MuSK MG), non-thymomatous late-onset MG (LOMG), and ocular MG (OMG). In addition to reviewing the literature, mainly from a clinical point of view, our experience in each of the subgroups, based on close to 600 patients seen over a 10 year period, is related. MuSK MG is a severe disease with predominant bulbar involvement. It is more common in women and in early-onset patients. With the use of high dose corticosteroids, azathioprine and more recently rituximab, outcome is favorable, though the patients usually require higher maintenance doses of immunosuppressives. LOMG with onset ≥ 50 years of age is more common in men and ocular onset is common. Frequency of anti-AChR and anti-titin antibodies are high. Although it can be severe in some patients, response to treatment is usually very good. OMG is reported to be more frequent in men in whom the disease has a later onset. Anti-AChR antibodies are present in about half of the patients. Generalization is less likely when symptoms remain confined to ocular muscles for 2 years. Low dose corticosteroids are usually sufficient. Thyroid disease is the most common autoimmune disease accompanying all three subgroups.

Improving accuracy of myasthenia gravis autoantibody testing by reflex algorithm.

To improve myasthenia gravis (MG) autoantibody testing. MG serologic tests with confirmatory or refuting clinical-electrodiagnostic (EDX) testing and cancer evaluations were reviewed over 4 years (2012-2015). All patients had acetylcholine receptor-binding (AChR-Bi), modulating (AChR-Mo), and striational (STR) autoantibody testing, and negatives reflexed to muscle-specific kinase (MuSK). Thymoma and cancer occurrences were correlated with STR and reflexed glutamic acid decarboxylase 65 (GAD65), ganglionic acetylcholine receptor (α3), collapsin response mediating protein-5, and voltage-gated potassium channel complex autoantibodies. Of 433 samples tested, 133 (31%) met clinical-EDX criteria for MG. Best sensitivity (90%) occurred at AChR-Bi >0.02 nmol/L, leaving 14 negative (6 ocular MG, 7 generalized MG, 1 MuSK MG) with specificity 90% (31 false-positives). Using AChR-Mo antibodies (>20% loss), specificity was better (92%, 24 false-positives), but sensitivity dropped (85%). Specificity improved (95%) by testing AChR-Mo when AChR-Bi are positive, resulting in 45% reduction of false-positives (31-17), maintaining AChR-Bi 90% sensitivity. Cutoff values recommended by area under the curve analysis did not outperform this approach. AChR-Bi and AChR-Mo values were significantly higher in true-positives. CT evaluations in 121 MG samples revealed 16 thymomas. Historical or subsequent cancers occurred in 22. STR and reflexed autoantibodies were not more common in MG with thymoma or other cancers. Full-body CT (n = 34) was performed in those with STR and reflex autoantibody positivity, but without additional cancers found. Accuracy of MG serologic testing is improved by reflexing AChR-Bi-positive cases to AChR-Mo. STR and other reflexed cancer evaluation autoantibodies did not provide value beyond standard CT chest imaging at the time of MG diagnosis. Diagnostic certainty is informed by AChR-Bi and AChR-Mo with higher values increasing specificity.