This thesis describes a set of excitability measurements -transcranial magnetic stimulation (TMS) combined with electromyography (EMG) and electroencephalography (EEG), nerve excitability threshold... Show moreThis thesis describes a set of excitability measurements -transcranial magnetic stimulation (TMS) combined with electromyography (EMG) and electroencephalography (EEG), nerve excitability threshold tracking (NETT), and muscle velocity recovery cycles (MVRC)- and the applicability of these tools in early phase clinical drug development. We validated the biomarkers in healthy subjects with registered drugs and showed that the measurements are all repeatable and sensitive to pharmacological effects, even in a small number of subjects. Furthermore, we have evaluated effects of a novel AMPA-positive allosteric modulator with TMS-EMG/EEG, and a first-in-class skeletal muscle-specific chloride channel (ClC-1) inhibitor with MVRC, and the findings helped us to confirm proof-of-mechanism of these compounds in healthy subjects. In conclusion, these measurements proved to be valuable pharmacodynamic biomarkers in two drug development programs, encouraging their further use in clinical development of other future drug candidates targeting cortical-, neuronal-, and muscle cell excitability. The use of such clinical pharmacodynamic biomarkers could improve the quality and efficiency of the development process of drugs for e.g. amyotrophic lateral sclerosis, chronic pain, depression, treatment-resistant epilepsy, and neuromuscular diseases. Show less
Paardekooper, L.M.; Fillié-Grijpma, Y.E.; Sluijs-gelling, A.J. van der; Zlei, M.; Doorn, R. van; Vermeer, M.H.; ... ; T2B Consortium 2023
A subset of autoimmune diseases is characterized by predominant pathogenic IgG4 autoantibodies (IgG4-AID). Why IgG4 predominates in these disorders is unknown. We hypothesized that dysregulated B... Show moreA subset of autoimmune diseases is characterized by predominant pathogenic IgG4 autoantibodies (IgG4-AID). Why IgG4 predominates in these disorders is unknown. We hypothesized that dysregulated B cell maturation or aberrant class switching causes overrepresentation of IgG4+ B cells and plasma cells. Therefore, we compared the B cell compartment of patients from four different IgG4-AID with two IgG1-3-AID and healthy donors, using flow cytometry. Relative subset abundance at all maturation stages was normal, except for a, possibly treatment-related, reduction in immature and naïve CD5+ cells. IgG4+ B cell and plasma cell numbers were normal in IgG4-AID patients, however they had a (sub)class-independent 8-fold increase in circulating CD20-CD138+ cells. No autoreactivity was found in this subset. These results argue against aberrant B cell development and rather suggest the autoantibody subclass predominance to be antigen-driven. The similarities between IgG4-AID suggest that, despite displaying variable clinical phenotypes, they share a similar underlying immune profile. Show less
Myasthenia gravis (MG) is a neuromuscular disorder in which patients experience fluctuating and fatigable muscle weakness. Ocular muscle weakness and bulbar weakness are most common, with ocular... Show moreMyasthenia gravis (MG) is a neuromuscular disorder in which patients experience fluctuating and fatigable muscle weakness. Ocular muscle weakness and bulbar weakness are most common, with ocular symptoms such as double vision (diplopia) and drooping eyelids (ptosis) often being prominent. The aim was to develop new methods to improve clinical care for ocular MG patients. In this thesis, I investigated the eye muscles using muscle electrophysiology, orthoptic measurements, and quantitative MRI. I have made several advancements in the areas of diagnostics, understanding refractory ocular symptoms, and the development of objective measurements of ocular muscle weakness in MG. Show less
Marcuse, F.; Hoeijmakers, J.G.J.; Hochstenbag, M.; Hamid, M.A.; Keijzers, M.; Mané-Damas, M.; ... ; Baets, M.H.V. de 2023
The aim of this study was to investigate the surgical and long-term neurological outcomes of patients with acetylcholine-receptor-antibody-associated myasthenia gravis (AChR-MG) who underwent... Show moreThe aim of this study was to investigate the surgical and long-term neurological outcomes of patients with acetylcholine-receptor-antibody-associated myasthenia gravis (AChR-MG) who underwent robotic thymectomy (RATS). We retrospectively analyzed the clinical-pathological data of all patients with AChR-MG who underwent RATS using the DaVinci® Robotic System at the MUMC+ between April 2004 and December 2018. Follow-up data were collected from 60 referring Dutch hospitals. In total, 230 myasthenic patients including 76 patients with a thymoma (33.0%) were enrolled in this study. Mean follow-up time, procedure time and hospitalization were, respectively 65.7 ± 43.1 months, 111±52.5 min and 3.3 ± 2.2 days. Thymomatous patients had significantly more frequently and more severe complications than nonthymomatous patients (18.4% vs. 3.9%, p<0.001). Follow up data was available in 71.7% of the included patients. The Myasthenia Gravis Foundation of America postintervention score showed any kind of improvement of MG-symptoms after RATS in 82.4% of the patients. Complete stable remission (CSR) or pharmacological remission (PR) of MG was observed in 8.4% and 39.4% of the patients, respectively. Mean time till CSR/PR remission after thymectomy was 26.2 ± 29.2 months. No statistical difference was found in remission or improvement in MGFA scale between thymomatous and nonthymomatous patients. RATS is safe and feasible in patients with MG. The majority of the patients (82.4%) improved after thymectomy. CSR and PR were observed in 8.4% and 39.4% of the patients, respectively, with a mean of 26.2 months after thymectomy. Thymomatous patients had more frequently and more severe complications compared to nonthymomatous patients. Show less
Muscle-specific kinase (MuSK) myasthenia gravis (MG) is a neuromuscular autoimmune disorder caused by predominantly IgG4 antibodies targeting the MuSK protein. IgG4 has the unique ability to... Show moreMuscle-specific kinase (MuSK) myasthenia gravis (MG) is a neuromuscular autoimmune disorder caused by predominantly IgG4 antibodies targeting the MuSK protein. IgG4 has the unique ability to exchange half-molecules with other IgG4s, resulting in monovalent binding to their antigen. To investigate if MuSK-antibody valency influences its pathogenicity, recombinant bivalent and functionally monovalent MuSK antibodies were generated from B-cell receptor sequences isolated from MuSK MG patients. Passive transfer studies revealed that monovalency amplifies MuSK antibody pathogenicity in vivo. This may be because monovalent MuSK antibodies inhibit MuSK signaling (antagonist), while bivalent MuSK antibodies activate MuSK signaling (agonist) in vitro. The binding epitope on MuSK further influences the consequences and pathogenicity of MuSK antibodies. Collectively, these results suggest that the pathophysiology in individual patients depends on their unique antibody composition and that class-switching to IgG4 is a critical step in developing MuSK MG. Furthermore, the IgG4 response against MuSK does not appear to result from a global increase in IgG4 responses, as MuSK MG patients only had mildly elevated serum IgG4. Instead, it is thought to be driven by the antigen itself. Importantly, MUSK and other MG associated genes are also expressed outside skeletal muscle. These locations are at risk for non-motor symptoms caused by autoantibodies or mutations, or for side-effects of targeted therapeutic strategies. Show less
Introduction:MRI of extra-ocular muscles (EOM) in patients with myasthenia gravis (MG) could aid in diagnosis and provide insights in therapy-resistant ophthalmoplegia. We used quantitative MRI to... Show moreIntroduction:MRI of extra-ocular muscles (EOM) in patients with myasthenia gravis (MG) could aid in diagnosis and provide insights in therapy-resistant ophthalmoplegia. We used quantitative MRI to study the EOM in MG, healthy and disease controls, including Graves’ ophthalmopathy (GO), oculopharyngeal muscular dystrophy (OPMD) and chronic progressive external ophthalmoplegia (CPEO).Methods:Twenty recently diagnosed MG (59±19yrs), nineteen chronic MG (51±16yrs), fourteen seronegative MG (57±9yrs) and sixteen healthy controls (54±13yrs) were included. Six CPEO (49±14yrs), OPMD (62±10yrs) and GO patients (44±12yrs) served as disease controls. We quantified muscle fat fraction (FF), T2water and volume. Eye ductions and gaze deviations were assessed by synoptophore and Hess-charting.Results:Chronic, but not recent onset, MG patients showed volume increases (e.g. superior rectus and levator palpebrae [SR+LPS] 985±155 mm3 compared to 884±269 mm3 for healthy controls, p < 0.05). As expected, in CPEO volume was decreased (e.g. SR+LPS 602±193 mm3, p < 0.0001), and in GO volume was increased (e.g. SR+LPS 1419±457 mm3, p < 0.0001). FF was increased in chronic MG (e.g. medial rectus increased 0.017, p < 0.05). In CPEO and OPMD the FF was more severely increased. The severity of ophthalmoplegia did not correlate with EOM volume in MG, but did in CPEO and OPMD. No differences in T2water were found.Interpretation:We observed small increases in EOM volume and FF in chronic MG compared to healthy controls. Surprisingly, we found no atrophy in MG, even in patients with long-term ophthalmoplegia. This implies that even long-term ophthalmoplegia in MG does not lead to secondary structural myopathic changes precluding functional recovery. Show less
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... Show moreMuscle-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 antiinflammatory 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. Show less
Objective: To determine the effect of tetanus toxoid (TT) revaccination on circulating B-, T-and NK-cell com-partments in myasthenia gravis (MG) patients.Methods: Lymphocyte (sub)populations and... Show moreObjective: To determine the effect of tetanus toxoid (TT) revaccination on circulating B-, T-and NK-cell com-partments in myasthenia gravis (MG) patients.Methods: Lymphocyte (sub)populations and differentiation stages were assessed by flow cytometry in 50 TT revaccinated MG patients. TT-specific proliferative responses were explored in PBMC cultures. Results: In patients treated with azathioprine B-and NK cell numbers were strongly decreased. Lymphocyte (sub) populations remained unaffected upon TT revaccination. t All patients showed a significant TT-induced prolif-erative response.Conclusion: TT revaccination is effective in MG patients with stable disease irrespective of their thymectomy status and medication and does not alter the composition of the lymphocyte compartment. Show less
Plomp, J.J.; Huijbers, M.G.M.; Verschuuren, J.J.G.M.; Borodovsky, A. 2022
Background: Myasthenia gravis (MG) is an autoimmune neuromuscular disorder hallmarked by fluctuating fatigable muscle weakness. Most patients have autoantibodies against acetylcholine receptors ... Show moreBackground: Myasthenia gravis (MG) is an autoimmune neuromuscular disorder hallmarked by fluctuating fatigable muscle weakness. Most patients have autoantibodies against acetylcholine receptors (AChRs) at the neuromuscular junction (NMJ). These are thought to have three possible pathogenic mode-of-actions: 1) cross linking and endocytosis of AChRs, 2) direct block of AChRs and 3) complement activation. The relative contributions of these mechanisms to synaptic block and muscle weakness of individual patients cannot be determined. It likely varies between patients and perhaps also with disease course, depending on the nature of the circulating AChR antibodies.New method: We developed a new bioassay which specifically enables functional characterization and quantification of complement-mediated synaptic damage at NMJs, without interference of the other pathogenic mechanisms. To this end, we pre-incubated mouse hemi-diaphragm muscle-nerve preparations with mAb35-hG1, a humanized rat AChR monoclonal and subsequently exposed the preparation to normal human serum as a complement source. NMJ-restricted effects were studied.Results: Clearly NMJ-restricted damage occurred. With immunohistology we showed complement deposition at NMJs, and synaptic electrophysiological measurements demonstrated transmission block. In whole-muscle contraction experiments we quantified the effect and characterized its onset and progression during the incubation with normal human serum. Comparison with existing methods: With this new assay the complement-mediated component of myasthenic NMJ pathology can be studied separately. Conclusions: Our assay will be of importance in detailed mechanistic studies of local complement activation at NMJs, investigations of new complement inhibitors, and laboratory pre-screening of therapeutic efficacy for individual MG patients to optimize care with clinically approved complement inhibitors. Show less
This thesis addresses several pathophysiological and clinical aspects of both the Lambert-Eaton myasthenic syndrome (LEMS, introduced in chapter 1) and myasthenia gravis (MG).In chapter 2, the... Show moreThis thesis addresses several pathophysiological and clinical aspects of both the Lambert-Eaton myasthenic syndrome (LEMS, introduced in chapter 1) and myasthenia gravis (MG).In chapter 2, the search for new LEMS associated small cell lung cancer (SCLC) markers is described, leading to the discovery of ERC1 as a new antigen associated with LEMS.In chapter 3, a literature review of screening methods for SCLC in LEMS is presented, aswell as the role of associated SOX1 antibodies in SCLC-LEMS patients.In chapter 4 long-term follow-up, functional impairment and quality of life in LEMSpatients are described. We also report a normal survival in LEMS patients without associated tumour as well as an improved tumour survival in SCLC-LEMS patients.In chapter 5, we report that lowering the cut-off value for increment from 100% to 60% increases sensitivity for diagnosis of LEMS, without a negative effect on the specificity.Chapter 6 reports that a lower initial compound muscle action potential amplitude is a marker of more severe disease in AChR MG.In chapter 7, we show a small symptomatic effect of ephedrine as a symptomatic treatment in MG patients, in a placebo-controlled, double-blind, randomised, multiple crossover series of n-of-1 trials. Show less
Disorders of the neuromuscular junction (NMJ) comprise a spectrum of rare diseases causing muscle fatigability and weakness, leading to life-long effects on quality of life. We established the... Show moreDisorders of the neuromuscular junction (NMJ) comprise a spectrum of rare diseases causing muscle fatigability and weakness, leading to life-long effects on quality of life. We established the Dutch-Belgian registry for NMJ disorders, based on a unique combination of patient -and physician-reported information. Information on natural course, disease burden, prevalence of complications and comorbidity is collected through patient-reported standardized questionnaires and verified using medical documentation. Currently, the registry contains information of 565 Myasthenia Gravis (MG) patients and 38 Lambert-Eaton myasthenic syndrome (LEMS) patients, constituting approximately 25% (MG) and 80% (LEMS) of patients in the Netherlands. This is a very large registry, with the highest participation rate per capita. In addition to confirming many disease characteristics previously described in the literature, this registry provides several novel insights. The reported rate of potentially corticosteroid-related comorbidity, including hypertension, heart disease, osteoporosis and type 2 diabetes was high, emphasizing the need to commence corticosteroid-sparing immune suppressive treatment as soon as possible. The reported rate of other auto-immune diseases is far higher than previously expected: 27% of MG and 38% of LEMS patients, and a surprisingly high number of MG patients (47%) is unaware of their antibody status. In conclusion, this registry provides a valuable collection of information regarding MG and LEMS disease course. Continuous collection of annual follow-up data will provide further longitudinal insights in disease burden, course and treatment effect. (c) 2021 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY license ( http://creativecommons.org/licenses/by/4.0/ ) Show less
Fatigue is usually defined as a subjective perception of lacking energy, mentally or physically, with a difficulty sustaining voluntary activities. It is a common symptom of many diseases and most... Show moreFatigue is usually defined as a subjective perception of lacking energy, mentally or physically, with a difficulty sustaining voluntary activities. It is a common symptom of many diseases and most likely has a multifactorial cause. In myasthenia gravis (MG), fatigue has a high prevalence and is correlated with female sex and disease severity. However, no large scale studies have been performed. Therefore, we aimed to evaluate fatigue in the Dutch participants (n = 420) of the Dutch-Belgian Myasthenia Patient Registry using an online survey. Additional information was obtained on mood, sleep, coping, quality of life, disease severity, physical activities and medication. Severe fatigue was present in 62% with a mean score of 37.1 +/- 13.2 points. Fatigue severity and prevalence increased significantly with disease severity. A positive correlation was found for female gender, BMI, disease severity and depressive symptoms. A negative correlation was found for strenuous physical activities and older age. The strong association with disease severity suggests that fatigue should be recognized as an element of the symptomatology of MG. The observed association between strenuous activity and fatigue and differences in coping style between fatigued and non-fatigued patients warrant future clinical trials on exercise and cognitive behavioral therapy. (C) 2021 The Authors. Published by Elsevier B.V. Show less
This thesis describes two studies on the efficacy and safety of vaccinations in patients stable autoimmune myasthenia gravis, one with tetanus revaccination and one with influenza vaccination. Of... Show moreThis thesis describes two studies on the efficacy and safety of vaccinations in patients stable autoimmune myasthenia gravis, one with tetanus revaccination and one with influenza vaccination. Of both vaccinations, the humoral response and clinical parameters of the disease are described. For tetanus revaccination, also the cellular response is described. Furthermore, the validation of a disease specific quality of life questionnaire is described. Show less
Myasthenia gravis is an autoimmune disease characterized by dysfunction of the neuromuscular junction. Current treatment is based on lifestyle advice, symptomatic treatment, immunosuppressive drugs... Show moreMyasthenia gravis is an autoimmune disease characterized by dysfunction of the neuromuscular junction. Current treatment is based on lifestyle advice, symptomatic treatment, immunosuppressive drugs and thymectomy. Corticosteroids remain the cornerstone of treatment beside symptomatic medication due to their low cost, wide availability and fast mode of action. However, long term steroid use carries substantial risks of severe adverse side effects. Therefore, non-steroidal immunosuppressive drugs are commonly added to the treatment. Unfortunately, they have a delayed-onset effect and evidence of their efficacy appears to be difficult to obtain. Several trials using drugs that have had clear positive results in other immunological disorders have failed in myasthenia. This failure may in part be related to difficulties in the design of clinical trial for myasthenia, which has a fluctuating disease course involving weakness that may be difficult to assess quantitively. This problem is exacerbated by the tendency of most clinical trials to select patients with a stable, but severe disease. Future trials should: select patients with weakness and fatigability that is completely explained by their myasthenia gravis, use a design that avoids the exclusion of patients with recent changes in medication, and explore the possibilities to completely avoid the use of corticosteroids. (C) 2019 The Author(s). Published by Elsevier B.V. Show less
The distribution of muscle weakness in MG is highly heterogeneous and frequently shifts in patients. In 5% of patients MG was restricted to ocular weakness, whereas 7% of patients never had any... Show moreThe distribution of muscle weakness in MG is highly heterogeneous and frequently shifts in patients. In 5% of patients MG was restricted to ocular weakness, whereas 7% of patients never had any form of ocular weakness throughout their disease course. In 83% of MG patients at least one other extra-ocular muscle was involved at the second visit.Patients with late-onset MG and the presence of additional autoimmune diseases had more exacerbations (OR = 47) and emergency treatments (OR = 26) than early-onset MG patients without other autoimmune diseases.MG-ADL (patient questionnaire on ADL) was found to be relatively insensitive to changes in generalized weakness. This led to the absence of a significant treatment response on MG-ADL in the presence of a significant response on QMG (an objective measure of muscle weakness) in the eculizumab study. A new ADL outcome measure, MGII, was shown to have a higher sensitivity for generalized weakness than MG-ADL.RoVEMP, a novel neurophysiological test, was shown to be a specific measure for myasthenic extra-ocular muscle weakness. In addition, we found a significant correlation between magnitude of decrement and the time since the last intake of pyridostigmine, supporting that RoVEMP decrement reflects reversible neuromuscular transmission failure. Show less
Introduction: The recently developed Myasthenia Gravis Impairment Index (MGII) is a promising measure as it has less floor effects and a higher relative efficiency in its responsiveness to... Show moreIntroduction: The recently developed Myasthenia Gravis Impairment Index (MGII) is a promising measure as it has less floor effects and a higher relative efficiency in its responsiveness to treatment effect compared to other MG measures. This study aimed at validating the MGII in a Dutch cohort of MG patients and analyzing the sensitivity of MGII compared to MG-ADL for changes in generalized weakness.Methods: We analyzed (generalized items of; -gen) MGII, quantitative myasthenia gravis (QMG), Myasthenia Gravis Activities of Daily Living (MG-ADL), EQ-5D visual analog, Myasthenia Gravis Composite (MGC) and ACTIVLIM (an ADL questionnaire focusing on generalized weakness) scores in a prospective cohort of 99 MG patients. We investigated correlations between MGII and other outcome measures. We used a generalized linear model to assess whether MGIIgen had an additional sensitivity on top of MG-ADLgen for changes (A) in QMGgen in individual patients.Results: MGII had a lower floor effect (4%) compared to QMG (6%), MG-ADL (11%) and MGC (16%). MGII correlated well with QMG (r = 0.68), MG-ADL (r = 0.83) and MGC (r = 0.74). As expected, the correlations with EQ visual analog and ACTIVLIM were lower (r = -0.57 and -0.48). AMGIIgen had an additional value on top of AMG-ADLgen in the prediction of Delta QMGgen (B = 0.54, p = 0.01).Discussion: The MGII score was cross-culturally validated in a Dutch cohort of MG patients. MGII had a higher sensitivity for generalized weakness than MG-ADL. Show less
