Mendelian randomization and colocalization analysis reveal novel drug targets for myasthenia gravis.

OBJECTIVE: Myasthenia gravis (MG) is a complex autoimmune disease affecting the neuromuscular junction with limited drug options, but the field of MG treatment recently benefits from novel biological agents. We performed a drug-targeted Mendelian randomization (MR) study to identify novel therapeutic targets of MG. METHODS: Cis-expression quantitative loci (cis-eQTL), which proxy expression levels for 2176 druggable genes, were used for MR analysis. Causal relationships between genes and disease, identified by eQTL MR analysis, were verified by comprehensive sensitivity, colocalization, and protein quantitative loci (pQTL) MR analyses. The protein-protein interaction (PPI) analysis was also performed to extend targets, followed by enzyme-linked immunosorbent assay (ELISA) to explore the serum level of drug targets in MG patients. A phenome-wide MR analysis was then performed to assess side effects with a clinical trial review assessing druggability. RESULTS: The eQTL MR analysis has identified eight potential targets for MG, one for early-onset MG and seven for late-onset MG. Further colocalization analyses indicated that CD226, CDC42BPB, PRSS36, and TNFSF12 possess evidence for colocalization with MG or late-onset MG. pQTL MR analyses identified the causal relations of TNFSF12 and CD226 with MG and late-onset MG. Furthermore, PPI analysis has revealed the protein interaction between TNFSF12-TNFSF13(APRIL) and TNFSF12-TNFSF13B(BLyS). Elevated TNFSF13 serum level of MG patients was also identified by ELISA experiments. This study has ultimately proposed three promising therapeutic targets (TNFSF12, TNFSF13, TNFSF13B) of MG. CONCLUSIONS: Three drug targets associated with the BLyS/APRIL pathway have been identified. Multiple biological agents, including telitacicept and belimumab, are promising for MG therapy.

Paraneoplastic myopathies.

This chapter reviews the association between cancer and the idiopathic inflammatory myopathies (IIM), which includes dermatomyositis (DM), antisynthetase syndrome (ASyS), immune-mediated necrotizing myopathy (IMNM), and inclusion body myositis (IBM). Accumulating evidence shows that the risk of a coexisting malignancy is high in patients with DM, especially among those with anti-Tif1γ autoantibodies. Patients with IMNM and no defined autoantibodies also have an increased risk of malignancy. Recent evidence demonstrates that many IBM patients have increased numbers of circulating CD57+ CD8+ T cells, consistent with a diagnosis of large granular lymphocytic leukemia. In contrast, IMNM patients with anti-SRP or anti-HMGCR autoantibodies as well as patients with ASyS syndrome do not have a definitively increased risk of cancer. Patients who have a cancer treated with one of the immune checkpoint inhibitors can develop myositis (ICI-myositis), sometimes along with myasthenia gravis and/or myocarditis.

Unexpected fibrous mediastinitis in a patient with myasthenia gravis - a case report.

BACKGROUND: Fibrous mediastinitis (FM) is a rare mediastinal lesion characterized by proliferation of fibrous tissue within the mediastinum. Previous reports have shown that this lesion can be caused by histoplasmosis and tuberculosis. In extremely rare cases, FM can also be caused by autoimmune diseases such as antineutrophil cytoplasmic antibody-associated vasculitis and large-vessel arteritis. CASE PRESENTATION: In our case, we report unexpected fibrous mediastinitis found after robotic thymectomy in a patient with myasthenia gravis (MG). The preoperative imaging indicated no obvious lesion in the mediastinum and the patient denied histories of both histoplasmosis and tuberculosis. After the operation, both proliferation of fibrous tissue and ectopic germinal centres (GCs) could be found in the thymus. CONCLUSION: This rare case might enrich our knowledge of the relationship between FM and autoimmune diseases.

Outcomes of extended resection for locally advanced thymic malignancies.

BACKGROUND: Thymic malignancies are rare tumors about which data are limited. Our objective here was to evaluate the outcomes and risk factors for complications and death in patients who underwent extended surgery to remove thymic malignancies. METHODS: We retrospectively included patients who underwent extended resection of locally advanced, nonmetastatic thymic malignancies at our institution. Patients were deemed eligible for resection by a multidisciplinary team. During surgery, priority was given to achieving complete resection rather than to sparing organs. RESULTS: The 108 patients had a mean age of 53 ± 15 years (range, 9-83); among them, 91 had thymoma, 12 thymic carcinoma, and 5 neuroendocrine tumor. The Masaoka stage was III or higher in 86 patients; examination of operative specimens resulted in downstaging of 22 patients. Tumor-free resection margins were achieved in 98 patients. Overall 5- and 10-year survival rates were 80% and 68%, respectively. Myasthenia gravis, present in 36 patients, was the only independent significant risk factor for major postoperative complications. Age older than 70 years, thymic carcinoma or neuroendocrine tumor, pT3 or pT4 stage, and R1 or R2 resection margins independently predicted death. The number of resected structures was not associated with survival. Thymic carcinoma or neuroendocrine tumor was independently associated with shorter disease-free survival. CONCLUSION: In an expert center, extended resection targeting complete resection rather than organ preservation provided good outcomes in patients with locally advanced thymic malignancies. The risk/benefit ratio of surgery should be assessed with special care in patients who are elderly or have myasthenia gravis.

The value of postoperative radiotherapy in thymoma patients with myasthenia gravis.

INTRODUCTION: Surgery is the first-line treatment for patients with thymoma associated with myasthenia gravis (MG); however, the value of radiotherapy among these patients remains debatable. Herein, we examined the impact of postoperative radiotherapy (PORT) on the efficacy and prognosis of patients with thymoma and MG. METHODS: This retrospective cohort study included 126 patients with thymoma and MG who were enrolled in the Xiangya Hospital clinical database between 2011 and 2021. Demographic and clinical data were collected including sex, age, histologic subtype, Masaoka-Koga staging, primary tumor, lymph node, metastasis (TNM) staging, and therapeutic modalities. To evaluate short-term MG symptom improvement following PORT, we examined changes in the quantitative myasthenia gravis (QMG) scores within 3 months post-treatment. Minimal manifestation status (MMS) was the main endpoint for assessing long-term improvement in MG symptoms. Overall survival (OS) and disease-free survival (DFS) were primary endpoints to determine the impact of PORT on prognosis. RESULTS: Effects of PORT on MG symptoms: QMG scores significantly differed between the non-PORT and PORT groups (χ2 = 6.300, p = 0.012). The median time to achieve MMS was significantly shorter in the PORT group than that in the non-PORT group (2.0 years vs. 4.4 years; p = 0.031). Multivariate analysis revealed that radiotherapy was associated with a reduced time to achieve MMS (hazard ratio [HR] 1.971, 95% confidence interval [CI]:1.102-3.525, p = 0.022). Effects of PORT on DFS and OS: The 10-year OS rate of the entire cohort was 90.5%, whereas OS rates for the PORT and non-PORT groups were 94.4 and 85.1%, respectively. The 5-year DFS rates for the whole cohort, PORT group, and non-PORT group were 89.7, 95.8, and 81.5%, respectively. PORT was associated with improved DFS (HR 0.139, 95% CI: 0.037-0.533, p = 0.004). In the high-risk histologic subgroup (type B2, B3), patients who received PORT had better OS (p = 0.015) and DFS (p = 0.0053) than those who did not receive PORT. PORT was associated with improved DFS (HR 0.232, 95% CI: 0.069-0.782, p = 0.018) in Masaoka-Koga stages II, III, and IV disease. CONCLUSIONS: Overall, our findings indicate that PORT positively impacts thymoma patients with MG, particularly those with a higher histologic subtype and Masaoka-Koga staging.

Comparison of Short-Term Post-Thymectomy Outcomes by Time-Weighted Dosages of Drug Requirements between Thymoma and Non-Thymoma Myasthenia Gravis Patients.

(1) Background: Early thymectomy is suggested in all clinically indicated myasthenia gravis (MG) patients. However, short-term clinical response after thymectomy in MG patients has been limitedly described in the literature. This study aimed to compare the 5-year post-thymectomy outcomes between thymoma (Th) and non-thymoma (non-Th) MG patients. (2) Methods: MG patients aged ≥18 years who underwent transsternal thymectomy and had tissue histopathology reports in Songklanagarind Hospital between 2002 and 2020 were enrolled in a retrospective review. The differences in the baseline demographics and clinical characteristics between ThMG and non-Th MG patients were studied. We compared the time-weighted averages (TWAs) of daily required dosages of pyridostigmine, prednisolone or azathioprine to efficiently maintain daily living activities and earnings between the MG patient groups during 5 consecutive years following thymectomy. Post-thymectomy clinical status, exacerbations or crises were followed. Descriptive statistics were used for analysis with statistical significance set at p < 0.05. (3) Results: ThMG patients had significantly older ages of onset and shorter times from the MG diagnosis to thymectomy. Male gender was the only significant factor associated with ThMG. TWAs of the daily MG treatment drug dosages required showed no differences between the groups. Additionally, the rates of exacerbations and crises were not different, but decremental trends were shown in both groups after the thymectomies. (4) Conclusions: The daily dosage requirements of MG treatment drugs were not different. There was a trend of decreasing adverse event rates despite no statistically significant differences during the first 5 years after thymectomy in ThMG and non-ThMG patients.

Expression of Programmed Cell Death 1 Ligand 2 in Patients With Thymoma and Thymomatous Myasthenia Gravis.

OBJECTIVES: This study aimed to examine the expression of programmed cell death 1 ligand 2 (PD-L2) in thymoma and thymomatous myasthenia gravis (MG). METHODS: The records of 70 patients with thymoma receiving surgical resection between January 2017 and December 2018 were retrospectively reviewed. Thymoma PD-L2 expression was evaluated by immunohistochemistry staining. Associations between PD-L2 expression and clinicopathologic features were examined. RESULTS: PD-L2 expression was positive in 41 patients (58.6%) and negative in 29 patients (41.4%). Of them, 33 had thymomatous MG. Patients with MG were more likely to be 50 years of age or younger (69.70% vs 35.14%); have more World Health Organization (WHO) type B thymomas (84.85% vs 64.86%); have tumors of smaller size (4.09 ± 2.33 cm vs 6.47 ± 2.42 cm); have positive PD-L2 expression (78.79% vs 40.54%); and have a higher percentage of PD-L2-positive cells, higher PD-L2 expression intensity, and score (all P < .05). Positive PD-L2 expression was associated with more type B thymomas, higher Masaoka-Koga stage, smaller tumor size, ectopic thymus, and MG (all P < .05). Factors significantly associated with MG were age under 50 years, tumor size less than 5 cm, and positive PD-L2 expression (all P < .05). CONCLUSIONS: Thymoma PD-L2 expression is significantly associated with thymomatous MG and WHO histologic types B2 and B3.

Receptor clustering and pathogenic complement activation in myasthenia gravis depend on synergy between antibodies with multiple subunit specificities.

Myasthenia gravis is an autoimmune disorder defined by muscle weakness and fatigability associated with antibodies against proteins of the neuromuscular junction (NMJ). The most common autoantibody target is the acetylcholine receptor (AChR). Three mechanisms have been postulated by which autoantibodies might interfere with neurotransmission: direct antagonism of the receptor, complement-mediated destruction of the postsynaptic membrane, and enhanced internalization of the receptor. It is very likely that more than one of these mechanisms act in parallel. Dissecting the mechanisms of autoantibody-mediated pathology requires patient-derived, monoclonal antibodies. Using membrane antigen capture activated cell sorting (MACACS), we isolated AChR-specific B cells from patients with myasthenia gravis, and produced six recombinant antibodies. All AChR-specific antibodies were hypermutated, including isotypes IgG1, IgG3, and IgG4, and recognized different subunits of the AChR. Despite clear binding, none of the individual antibodies showed significant antagonism of the AChR measured in an in vitro neuromuscular synapse model, or AChR-dependent complement activation, and they did not induce myasthenic signs in vivo. However, combinations of antibodies induced strong complement activation in vitro, and severe weakness in a passive transfer myasthenia gravis rat model, associated with NMJ destruction and complement activation in muscle. The strongest complement activation was mediated by combinations of antibodies targeting disparate subunits of the AChR, and such combinations also induced the formation of large clusters of AChR on the surface of live cells in vitro. We propose that synergy between antibodies of different epitope specificities is a fundamental feature of this disease, and possibly a general feature of complement-mediated autoimmune diseases. The importance of synergistic interaction between antibodies targeting different subunits of the receptor can explain the well-known discrepancy between serum anti-AChR titers and clinical severity, and has implications for therapeutic strategies currently under investigation.

Retrospective histopathology audit of thymectomy specimens: A clinicopathological study of 303 cases spanning the non-neoplastic, benign and malignant spectrum.

Aims: Thymectomy specimens are uncommon in routine histopathology practice. However, awareness of various pathologic entities and definite reporting of these specimens is paramount to optimal patient management. Our objective was to determine the histomorphologic spectrum of thymectomy specimens spanning the non-neoplastic, benign to malignant spectrum. Methods and Results: Thymectomies received over an 8-year period were retrospectively analyzed by reviewing clinical details and histologic findings in detail, incorporating the latest World Health Organization (WHO) 2015 histologic classification. A total of 303 thymectomy specimens (179 males/124 females, mean age 45.3 years [3-84 years]) were included. Around 51.2% (n = 155) patients had associated myasthenia gravis (MG), while 17.5% (n = 53) had incidentally detected anterior mediastinal mass (AMM). Non-neoplastic and benign pathologies comprised 31% (n = 94) cases and showed stronger association with MG (P = 0.009). Thymic follicular hyperplasia (TFH) was the commonest non-neoplastic pathology (n = 32), while the benign tumor group included thymic hemangioma/lymphangioma, thymolipoma, and ectopic parathyroid adenoma. Thymic epithelial tumors (TETs) comprised 64.7% cases, with majority being thymomas (185/303; 61.1%). Thymoma type B2 was the commonest histologic subtype and Stage I/T1 was the most frequent stage. Type A and AB thymomas affected older patients (P = 0.005) and were in lower stage (both Masaoka and American Joint Committee on Cancer [AJCC]) than type B thymomas (P = 0.007). No significant association between MG and thymoma subtype, patient sex or Masaoka stage was seen (P > 0.05). Thymic carcinomas comprised 11 cases and showed no association with MG (0/11, P < 0.001); squamous cell carcinoma was the commonest histologic type (8/11; 72.7%). Conclusion: TETs are the commonest thymic lesions; however, a diverse spectrum of pathologic processes can affect the thymus.

Ectopic germinal centers in the thymus accurately predict prognosis of myasthenia gravis after thymectomy.

The ability of thymic histopathology to predict the long-term impact of thymectomy in non-thymomatous myasthenia gravis (NTMG) is mainly uncharted. We applied digital pathology to quantitatively characterize differences of thymic histology between early-onset (EOMG) and late-onset MG (LOMG) and to investigate the role of thymic changes for thymectomy outcomes in MG. We analyzed 83 thymic H&E slides from thymectomized NTMG patients, of which 69 had EOMG and 14 LOMG, using digital pathology open-access software QuPath. We compared the results to the retrospectively assessed clinical outcome at two years after thymectomy and at the last follow-up visit where complete stable remission and minimal use of medication were primary outcomes. The automated annotation pipeline was an effective and reliable way to analyze thymic H&E samples compared to manual annotation with mean intraclass correlation of 0.80. The ratio of thymic tissue to stroma and fat was increased in EOMG compared to LOMG (p = 8.7e-07), whereas no difference was observed in the ratio of medulla to cortex between these subtypes. AChRAb seropositivity correlated with the number of ectopic germinal centers (eGC; p = 0.00067) but not with other histological areas. Patients with an increased number of eGCs had better post-thymectomy outcomes at two years after thymectomy (p = 0.0035) and at the last follow-up (p = 0.0267). ROC analysis showed that eGC area predicts thymectomy outcome in EOMG with an AUC of 0.79. Digital pathology can thus help in providing a predictive tool to the clinician, the eGC number, to guide the post-thymectomy treatment decisions in EOMG patients.

Partial Versus Complete Thymectomy in Non-Myasthenic Patients With Thymoma: A Systematic Review and Meta-Analysis of Clinical Outcomes.

OBJECTIVE: The optimal extent of surgical resection for non-myasthenic patients with thymoma is controversial. The objective of this meta-analysis was to compare complete to partial thymectomy in non-myasthenic patients for oncological and postoperative clinical outcomes. METHODS: We performed a PubMed and EMBASE search (from inception to January 2020) for English-language studies directly comparing partial thymectomy (thymomectomy) to complete thymectomy for thymoma resection. Clinical endpoints studied included overall and disease-free survival, Masaoka and World Health Organization staging, adjuvant therapy, postoperative complications, postoperative drainage, length of hospital stay, thymoma-related deaths, postresection development of myasthenia gravis, incomplete resection, and recurrence. Random effects meta-analyses across all clinical endpoints was done. RESULTS: There was no statistically significant difference between the two approaches with regard to recurrence (odds ratio [OR], 1.22; 95% confidence interval [CI], 0.78-1.92), completeness of resection (OR, 1.17; 95% CI, 0.66-2.10), adjuvant therapy (OR, 0.71; 95% CI, 0.40-1.26), or thymoma-related deaths (OR, 0.76; 95% CI, 0.12-4.66). There was a statistically significant decrease in postoperative complications (OR, 0.61; 95% CI, 0.39-0.97), drainage (mean difference [MD], -0.99; 95% CI, -1.98 to -0.01), and length of hospital length (MD, -1.88; 95% CI, -3.39 to -0.36) with partial thymectomy. CONCLUSIONS: The evidence appeared to suggest that partial thymectomy is oncologically equivalent to complete thymectomy for non-myasthenic patients with early-stage thymoma. There is an additional advantage of reduced postoperative complications and decreased length of hospital stay with partial thymectomy.

A Review on the Role of Non-Coding RNAs in the Pathogenesis of Myasthenia Gravis.

Myasthenia gravis (MG) is an autoimmune condition related to autoantibodies against certain proteins in the postsynaptic membranes in the neuromuscular junction. This disorder has a multifactorial inheritance. The connection between environmental and genetic factors can be established by epigenetic factors, such as microRNAs (miRNAs) and long non-coding RNAs (lncRNAs). XLOC_003810, SNHG16, IFNG-AS1, and MALAT-1 are among the lncRNAs with a possible role in the pathoetiology of MG. Moreover, miR-150-5p, miR-155, miR-146a-5p, miR-20b, miR-21-5p, miR-126, let-7a-5p, and let-7f-5p are among miRNAs whose roles in the pathogenesis of MG has been assessed. In the current review, we summarize the impact of miRNAs and lncRNAs in the development or progression of MG.

To Be or Not To Be Vaccinated: That Is a Question in Myasthenia Gravis.

Myasthenia gravis (MG) is an autoimmune disease characterized by muscle weakness and abnormal fatigability due to the antibodies against postsynaptic receptors. Despite the individual discrepancy, patients with MG share common muscle weakness, autoimmune dysfunction, and immunosuppressive treatment, which predispose them to infections that can trigger or exacerbate MG. Vaccination, as a mainstay of prophylaxis, is a major management strategy. However, the past years have seen growth in vaccine hesitancy, owing to safety and efficacy concerns. Ironically, vaccines, serving as an essential and effective means of defense, may induce similar immune cross-reactivity to what they are meant to prevent. Herein, we outline the progress in vaccination, review the current status, and postulate the clinical association among MG, vaccination, and immunosuppression. We also address safety and efficacy concerns of vaccination in MG, in relation to COVID-19. Since only a handful of studies have reported vaccination in individuals with MG, we further review the current clinical studies and guidelines in rheumatic diseases. Overall, our reviews offer a reference to guide future vaccine clinical decision-making and improve the management of MG patients.

GTF2I Mutation in Thymomas: Independence From Racial-Ethnic Backgrounds. An Indian/German Comparative Study.

Thymomas are the most frequent adult mediastinal cancers. Their etiology is unknown and their pathogenesis poorly understood. Racial, ethnic and environmental factors influence tumorigenesis in many cancers, but their role in thymomas remains unclear to date. In this study that included pretreatment thymoma cases from India and Germany (n = 37 and n = 77, respectively) we compared i) the prevalence of the thymoma-specific chromosome 7 c.74146970T > A mutation of the GTF2I gene in type A and AB thymomas; ii) epidemiological features; and iii) the frequency of myasthenia gravis (MG). Due to a known predominance of GTF2I mutation in A and AB histotypes, we included only a marginal number of type B thymomas as a control group in both cohorts. While the distribution of histological types between the cohorts was similar (p = 0.1622), Indian patients were strikingly younger (p < 0.0001; median age 50 vs. 65 years) and showed significantly lower tumour stage (Masaoka-Koga stage I) at primary diagnosis (p = 0.0005) than the German patients. In patients with known MG status (n = 17 in Indian and n = 25 in German cohort), a clear trend towards more frequent MG was observed in the Indian group (p = 0.0504; 48 vs. 82%). The prevalence of the GTF2I mutation (analysed in n = 34 Indian and n = 77 German patients) was identical in the two cohorts. We conclude that racial-ethnic and environmental factors do not significantly influence the most common molecular feature of thymomas but may have an impact on the timing of clinical presentation.

NKT and NKT-like Cells in Autoimmune Neuroinflammatory Diseases-Multiple Sclerosis, Myasthenia Gravis and Guillain-Barre Syndrome.

NKT cells comprise three subsets-type I (invariant, iNKT), type II, and NKT-like cells, of which iNKT cells are the most studied subset. They are capable of rapid cytokine production after the initial stimulus, thus they may be important for polarisation of Th cells. Due to this, they may be an important cell subset in autoimmune diseases. In the current review, we are summarising results of NKT-oriented studies in major neurological autoimmune diseases-multiple sclerosis, myasthenia gravis, and Guillain-Barre syndrome and their corresponding animal models.

Whole-exome sequencing and human leukocyte antigen analysis in familial myasthenia gravis with thymoma: Case report and literature review.

Myasthenia gravis (MG) is an autoimmune disease characterized by impaired neurotransmission at the neuromuscular junction. MG is generally non-inherited but is rarely inherited. Here, we report two patients with MG in the same pedigree: a 62-year-old Japanese man and his 46-year-old daughter who were positive for anti-acetylcholine receptor antibodies and had thymoma. We performed whole-exome sequencing (WES) and human leukocyte antigen (HLA) analyses to investigate the genetic contribution to familial onset. WES analysis of both patients showed no known variations in candidate genes for familial MG, and HLA analysis failed to detect HLA haplotypes seen in early-onset and late-onset MG. These findings suggest the presence of an unknown genetic background. Previous genetic studies on familial MG have identified ENOX1 and IFNGR1 as candidate genes in patients without thymoma, whereas no studies have identified candidate genes in patients with thymoma. To explore causative genes, it may be necessary to consider whether the genetic background differs between patients with and without thymoma in familial autoimmune MG.

Myositis/Myasthenia after Pembrolizumab in a Bladder Cancer Patient with an Autoimmunity-Associated HLA: Immune-Biological Evaluation and Case Report.

Pembrolizumab (mAb to PD-1) has been recently approved for the therapy of pretreated urothelial cancer. Despite the efficacy, it is often accompanied by unpredictable and sometime severe immune-related (ir) adverse events (AEs). Here, we report the clinical and immune-biological characterization of a patient with a metastatic bladder cancer who developed myositis signs (M) and a myasthenia-like syndrome (MLS) during treatment with pembrolizumab. The patient presented an autoimmunity-associated HLA haplotype (HLA-A*02/HLA-B*08/HLA-C*07/HLA-DRB1*03) and experienced an increase in activated CD8 T-cells along the treatment. The symptomatology regressed after pembrolizumab discontinuation and a pyridostigmine and steroids-based therapy. This is the first report of concurrent M and MLS appearance in cancer patients receiving pembrolizumab. More efforts are needed to define early the risk and the clinical meaning of irAEs in this setting.

Natural killer cells promote the differentiation of follicular helper T cells instead of inducing apoptosis in myasthenia gravis.

Recent evidence has shown that natural killer (NK) cells have an immunoregulatory function in the pathogenesis of myasthenia gravis (MG). In this study, the phenotype and function of NK cell subsets in peripheral blood of new-onset MG (N-MG) and stable MG (S-MG) patients were explored. Circulating CD56dim and CD56bright NK cells were increased and decreased, respectively, in patients with N-MG and S-MG compared with healthy control (HC). Moreover, all circulating NK cell subsets from N-MG patients showed significantly lower expression of activating receptor NKG2D and production of Interferon (IFN) -γ than that from HC. The killing effects of NK cells on CD4+ T cells and Tfh cells were impaired in MG patients, whereas, they promoted the differentiation and activation of Tfh cells. These data indicated that the immune-regulation of NK cells on CD4+ T cells and Tfh cells in MG patients was abnormal, which may contribute to the immune-pathological mechanism of MG.