The impact of immune markers on thymectomy prognosis in thymoma-myasthenia gravis

Introduction

Myasthenia gravis (MG) is an autoimmune disorder characterized by antibodies attacking the postsynaptic components at the neuromuscular junction (1). Thymomas, being the most common tumors of the anterior mediastinum, are associated with various autoimmune diseases (2), especially exhibiting a close relationship with MG. Approximately 15% of patients with thymomas also suffer from MG (3). Hence, thymectomy is considered a critical therapeutic measure for such patients (4), and the comprehensive removal of the thymus along with surrounding tissues, as proposed by Masaoka et al., has become the accepted standard of treatment (5). This surgery not only effectively removes the tumor but also plays a significant role in alleviating the symptoms of MG (6), thereby substantially improving the patient’s quality of life.

Research has unveiled the intricate link between thymomas and MG, particularly highlighting significant changes in immune markers such as T helper (Th)17 cells, regulatory T (Treg) cells, immunoglobulin (Ig)G, and C-reactive protein (CRP) in patients with thymomas. Notably, elevated levels of CRP have been closely associated with poor prognosis in patients with thymomas (7). Further, in patients with thymomas who also have MG, significant alterations in immune indicators including Th17 cells, Treg cells, and IgG, as well as complement components C3, C4, and CRP, have been observed in both thymic tissue and peripheral blood (8-11). These changes underscore the importance of immune marker variations in the development of thymomas and MG. However, current research on how these immune markers affect the prognosis of patients with thymoma-associated MG undergoing complete thymectomy is still relatively sparse. Understanding the variations in these immune-related indicators is crucial for improving treatment strategies and prognoses for patients.

The aim of this study was to evaluate the predictive role of various immune markers in the peripheral blood preoperatively (including CD4⁺ T cells, CD8⁺ T cells, Th17 cells, Treg cells, various Igs, CRP, complement C3, and complement C4) on disease progression post-surgery in patients with MG associated with thymomas. This research intends to identify key immune indicators closely related to sustained remission postoperatively. The findings of this study will support the development of personalized treatment strategies, thereby enhancing treatment efficacy and improving patient outcomes. We present this article in accordance with the STROBE reporting checklist (available at https://jtd.amegroups.com/article/view/10.21037/jtd-24-824/rc).

Methods

Patient enrollment and definition

We conducted a retrospective analysis on 163 patients with thymoma-associated MG who underwent complete thymectomy at the Department of Cardiothoracic Surgery, Tianjin Medical University General Hospital, from January 2011 to December 2022. The diagnosis of MG was based on typical clinical manifestations, including fluctuating weakness and fatigue that improves in the morning and worsens at night, exacerbation after physical activity, and alleviation after rest. A positive test in any of the following three criteria confirmed the diagnosis: (I) anti-acetylcholine receptor antibodies (anti-AChR Abs); (II) repetitive nerve stimulation (RNS) test; or (III) positive response to edrophonium chloride (Tensilon test) or neostigmine test.

The main outcome measures of the study were changes in myasthenic symptoms, fluctuations in clinical presentation, and the time span of symptom changes (calculated from the postoperative period).

Inclusion criteria were age greater than or equal to 18 years, having at least 3 months of corresponding myasthenic symptoms, and a follow-up period of no less than 1 year. Patients with missing follow-up data or incomplete clinical information were excluded. Additionally, patients with thymic carcinomas and thymic cysts, which have not been definitively linked to the pathogenesis of MG, were also excluded (12).

All thymoma patients were classified and staged according to the World Health Organization (WHO) histological classification (13) and the Masaoka-Koga clinical staging system (14). We recorded patient demographics, including age, gender, onset time of symptoms, severity of symptoms, WHO histological classification, Masaoka-Koga staging of thymoma, follow-up duration, postoperative changes in patient condition and medication adjustments (patients with severe symptoms generally take both pyridostigmine bromide and methylprednisolone, while those with milder symptoms take pyridostigmine bromide alone; patients whose symptoms have completely resolved do not continue medication), and characteristics of various immune markers.

Diagnosis, classification, and staging of thymomas were determined based on postoperative pathological results, while the postoperative assessment of the disease was based on regular follow-up examinations. The diagnosis of MG was based on clinical symptoms and corresponding auxiliary examinations. The evaluation of disease progression was defined according to clinical manifestations and the response to medication therapy (clinical improvement was defined as a significant reduction in clinical presentations compared to pre-treatment or a sustained significant reduction in the usage of MG medications, while exacerbation was defined as the worsening of clinical symptoms or an increase in MG medication dosage or the addition of steroid treatment).

The patient cohort was categorized into two groups: the sustained remission group and the non-sustained remission group. The sustained remission group comprised patients who demonstrated continuous complete remission of MG symptoms or a consistent reduction in related medication postoperatively, and had not taken corticosteroids up until the end of the follow-up period. Conversely, patients who experienced postoperative worsening of MG symptoms or had a history of symptom exacerbation (even if in remission at the follow-up endpoint), those currently on corticosteroids, and those who died from MG were classified into the non-sustained remission group.

In this study, all cases underwent comprehensive thymectomy, including the removal of the entire thymus and the mediastinal fat tissue between the bilateral phrenic nerves. This surgical strategy is primarily aimed at patients with thymoma-associated MG, particularly those with thymic abnormalities or suspected thymic hyperplasia, especially applicable to patients with poor response to acetylcholinesterase inhibitor therapy, resistance to immunosuppressive treatment, contraindications to immunosuppressive drugs, or drug resistance (15). Thymectomy was recommended even in patients where imaging studies could not definitively distinguish between thymoma and thymic hyperplasia, to exclude the possibility of thymoma (16).

The study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by the ethics committee of Tianjin Medical University General Hospital (ethical No. ZYY-IRB2018-KY-018). Due to the nature of the research, obtaining patient informed consent was not required.

After applying stringent inclusion and exclusion criteria, 53 patients were excluded, leaving a total of 163 patients included in the study. The inclusion and exclusion processes are detailed in Figure 1.

Figure 1 Flowchart of the recruitment and exclusion process for patients with thymoma-associated MG following thymectomy. MG, myasthenia gravis.

Statistical analysis

To analyze categorical variables, we employed the Chi-square test and Fisher’s exact test. Continuous variables were analyzed using the two-tailed t-test or Mann-Whitney U test. The cumulative incidence of sustained remission in patients with MG post-thymectomy was calculated using the Kaplan-Meier method, and factors influencing sustained remission of MG symptoms postoperatively during the follow-up period were determined through univariate and multivariate Cox proportional hazards regression analysis. All analyses were considered statistically significant at a P value of less than 0.05. Data analysis was performed using SPSS Statistics for Mac, version 27.0 (IBM, Armonk, NY, USA).

Results

Baseline characteristics

Table 1 summarizes the perioperative clinical characteristics, relevant laboratory indices, and outcome measures of all patients participating in the study, with all immune-related indices based on analyses of peripheral blood samples. The average age at onset was 52.6 years (range, 23 to 81 years), while the average age at the time of thymectomy was 53 years (range, 24 to 82 years). The participants included 75 males (46.0%) and 88 females (54.0%). According to the clinical classification by the Myasthenia Gravis Foundation of America (MGFA), the distribution was as follows: type I, 64 cases (39.3%); type IIa, 18 cases (11.0%); type IIb, 31 cases (19.0%); type IIIa, 12 cases (7.4%); type IIIb, 19 cases (11.7%); type IVa, 7 cases (4.3%); type IVb, 11 cases (6.7%); and type V, 1 case (0.6%). The pathological types of thymomas were classified according to the WHO histological classification, with type B3 (36.8%) being the most common, followed by type B2 (34.4%), type AB (14.1%), and type B1 (12.9%).

The overall average follow-up duration was 68.2 months (range, 13.7 to 133.6 months). Of the patients, 77 (47.2%) achieved sustained remission postoperatively, 49 patients (30.1%) experienced no change in symptoms, 20 patients (12.3%) had a relapse or exacerbation after surgery, 10 patients (6.1%) currently exhibit worsened symptoms, and 7 patients (4.3%) died due to aggravated MG.

Comparison of clinical features between groups

In Table 2, we compared the clinical characteristics between the 88 patients who achieved postoperative sustained remission and the 75 patients who did not achieve postoperative sustained remission. There were no statistically significant differences between the two groups in terms of levels of IgG, IgA, IgM, CRP, complement C4, Th17 cells, Treg cells, CD4⁺ T cells, and CD8⁺ T cells. However, a significantly higher proportion of patients in the sustained remission group had serum IgE (P=0.04) and complement C3 (P=0.004) within the normal range compared to the non-sustained remission group, indicating statistically significant differences.

Cumulative probability of sustained remission

Figure 2 illustrates the cumulative probability of sustained remission for patients in the sustained remission group compared to the non-sustained remission group postoperatively. At the 3-, 6-, and 9-year marks, the cumulative probabilities of sustained remission were approximately 88.6%, 63.1%, and 37.2%, respectively. Over time, Figure 2A demonstrates that all patients may eventually experience a relapse or worsening of symptoms. By conducting a Kaplan-Meier analysis of the status of complement C3, IgG, IgA, IgM, IgE, CRP, and complement C4, we calculated the cumulative probability of postoperative sustained remission among the 163 patient samples. The results indicate that patients with normal levels of complement C3 have a higher likelihood of sustained remission postoperatively compared to those with decreased levels of complement C3. Other indicators did not significantly affect the cumulative rate of postoperative sustained remission.

Figure 2 Kaplan-Meier curves for the cumulative probability of sustained remission after thymectomy in patients with thymoma-associated generalized MG. (A) Kaplan-Meier curves for the cumulative probability of sustained remission after thymectomy in all patients with thymoma-associated generalized MG. Kaplan-Meier curves for cumulative probability of sustained remission after thymectomy in different patient groups: (B) IgG within normal range vs. abnormal range (P=0.67); (C) IgA within normal range vs. abnormal range (P=0.74); (D) IgM within normal range vs. abnormal range (P=0.14); (E) complement C3 within normal range vs. decline (P<0.001); (F) complement C4 within normal range vs. decline (P=0.22); (G) CRP within normal range vs. elevated (P=0.59); (H) IgE within normal range vs. abnormal range (P=0.74). Ig, immunoglobulin; CRP, C-reactive protein; MG, myasthenia gravis.

Predictors of postoperative sustained remission

In Table 3, univariate and multivariate analyses conducted using the Cox proportional hazards model identified potential prognostic factors associated with postoperative sustained remission. In these analyses, a decrease in complement C3 was confirmed as a significant negative predictor of postoperative sustained remission [hazard ratio (HR) =3.250; 95% confidence interval (CI): 1.931–5.469; P<0.001], while other factors did not show a clear predictive effect on postoperative sustained remission.

Discussion

In this retrospective single-center study, we evaluated immune-related indicators that could predict sustained remission in patients with MG post-thymectomy. Comparing the sustained remission group to the non-sustained remission group, significant statistical differences were found in IgE (P=0.04) and complement C3 (P=0.004) levels. However, in Kaplan-Meier survival analysis, IgE and other immune indicators did not significantly differ in predicting the cumulative rate of postoperative sustained remission. In contrast, a reduction in complement C3 was associated with a faster decline in the cumulative rate of sustained remission postoperatively. By the end of follow-up (approximately 133 months), the cumulative rate of sustained remission for patients with normal levels of complement C3 dropped to 0, indicating that even patients with normal complement C3 levels could experience a recurrence or progression of MG symptoms postoperatively. Univariate and multivariate regression analyses revealed that, apart from complement C3 (P<0.001) significantly acting as a negative predictor for postoperative sustained remission, other immune indicators did not demonstrate predictive capabilities for the progression of MG postoperatively. The Kaplan-Meier analysis overall showed that, by the end of follow-up, the cumulative rate of sustained remission for all patients dropped to 0, suggesting that all patients might encounter MG symptom recurrence or worsening after a certain period postoperatively.

When exploring the pathophysiological mechanisms of autoimmune diseases, the role of the complement system, particularly complement C3, cannot be overlooked. As the most abundant complement component in serum, complement C3 plays a critical role in regulating inflammatory responses and facilitating the clearance of immune complexes. Studies have suggested that a decrease in complement C3 levels not only indicates increased disease activity but may also results in an imbalance in immune regulation, exacerbating autoimmune responses (17); for example, in autoimmune diseases such as systemic lupus erythematosus, reduced complement C3 is closely associated with disease activity and prognosis (18). Our study further corroborates this, finding that a reduction in complement C3 is associated with a decrease in sustained remission in patients with MG postoperatively, highlighting the importance of complement in the pathogenesis of MG. Additionally, the significant elevation of IgE levels in patients with thymoma-associated MG suggests its potential role in the autoimmune process, including activating autoreactive T cells and causing B lymphocytes and plasma cells to remain in lymphoid tissues, thus amplifying the autoimmune response (19). However, despite significant statistical differences in the number of patients with elevated IgE between the two groups, our study indicates that it does not significantly predict patient prognosis, which could be due to the small sample size or that elevated IgE levels do not directly affect prognosis. Other study has shown significant differences in the levels of complement C4 and CRP in the peripheral blood of thymoma patients with MG compared to those without MG (10). In terms of immune regulation, the proportion of Th17 cells is higher in thymomas unrelated to MG than in those related to MG (20), while the proportion of Treg is higher in MG-related thymomas. Although these findings reveal different expressions of immune-related indicators in the disease, they did not show significant correlations statistically or in prognostic predictions in this study, which could be attributed to the small sample size or no direct association between these indicator abnormalities and prognosis.

As a single-center retrospective design, this study primarily explored the impact of immune indicators on prognosis, while other studies at our center have also identified the influence of age and thymoma staging on the progression of MG. Furthermore, literature has shown that factors such as anti-AChR Ab levels (21), thymic hyperplasia (22), and thymoma histology (23) are also closely related to prognosis. In this study, a total of 21 patients received short-term corticosteroid therapy before surgery (with treatment durations of no more than 1 week). Post-surgery, the use of corticosteroids or immunomodulators varied depending on the patients’ conditions. Previous studies have indicated that hormones and immunomodulators might affect immune markers like complements C3, C4, and IgE (24-26). All patients included in this study were diagnosed with mediastinal tumors through chest computed tomography (CT) scans after presenting symptoms of MG. Even those with severe symptoms underwent thymectomy after short-term symptom control. To further explore the impact of corticosteroids or immunomodulators on immune markers and prognosis, we plan to conduct a prospective study to systematically monitor the specific effects of these drug treatments on immune indicators and to conduct an in-depth analysis of patients’ long-term prognosis. This research will help clarify the role of hormones and immunomodulators in the treatment of thymoma with concomitant severe MG and their potential impact on treatment outcomes. Therefore, for patients with thymoma-associated MG, considering the multiple factors influencing prognosis post-thymectomy, a comprehensive judgment is required. However, limitations due to the small sample size and potential recall bias regarding the timing of postoperative recurrence could impact study results. Moreover, changes in postoperative immune indicators and their potential impact on prognosis should not be overlooked. Hence, future studies should expand to multi-center designs, increase sample sizes to reduce recall bias, and regularly monitor changes in postoperative immune indicators to more deeply understand their specific impact on patient prognosis, thereby obtaining more accurate study outcomes.

Conclusions

Complement C3 level is a crucial predictor for long-term remission in thymoma-associated MG post-thymectomy. Patients with lower C3 levels are more likely to experience MG symptom progression post-surgery compared to those with normal levels, indicating its potential as a key prognostic marker. For patients with lower C3 levels, a more comprehensive preoperative assessment of the patient’s overall condition should be conducted. Postoperatively, regular monitoring of complement C3 levels is necessary to enable timely adjustments to the treatment plan.

Acknowledgments

Funding: None.

Footnote

Reporting Checklist: The authors have completed the STROBE reporting checklist. Available at https://jtd.amegroups.com/article/view/10.21037/jtd-24-824/rc

Data Sharing Statement: Available at https://jtd.amegroups.com/article/view/10.21037/jtd-24-824/dss

Peer Review File: Available at https://jtd.amegroups.com/article/view/10.21037/jtd-24-824/prf

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://jtd.amegroups.com/article/view/10.21037/jtd-24-824/coif). The authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. The study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by the ethics committee of Tianjin Medical University General Hospital (ethical No. ZYY-IRB2018-KY-018). Due to the nature of the research, obtaining patient informed consent was not required.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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