The value of chromosome 8 heteroploid tumor cells in the peripheral blood and pleural effusion in evaluating treatment efficacy and the prognosis of metastatic non-small cell lung cancer patients

Introduction

Non-small cell lung cancer (NSCLC) is a leading cause of cancer-related illness and mortality, and thus represents a significant global health challenge (1). The incidence of NSCLC has been increasing, which has serious implications for public health and economic resources (2). The World Health Organization reports that lung cancer is the most prevalent type of cancer, and NSCLC accounts for approximately 80% of all lung cancer cases (3). Despite advancements in treatments, including surgery, chemotherapy, targeted therapy, and immunotherapy, the prognosis of patients with advanced NSCLC remains poor. This is primarily due to issues such as treatment resistance and insufficient therapeutic responses that complicate effective disease management.

Recent research has highlighted the potential of circulating tumor cells (CTCs) as biomarkers for assessing disease prognosis in NSCLC (4,5). CTCs are cancer cells that detach from the primary tumor and enter the bloodstream, and can serve as indicators of disease progression and metastasis (6). Zheng et al. showed that the presence and characteristics of CTCs are closely linked to the treatment responses and overall survival (OS) of various cancers, including gastric cancer and lung cancer (7). The advantages of CTC detection in lung cancer include its non-invasive nature, real-time monitoring capabilities, reflection of tumor heterogeneity, and advancements in detection technologies. In clinical practice, acquiring sufficient tumor biopsy material and conducting repeated biopsies in lung cancer patients is frequently challenging. Aneuploidy is a feature of most cancer cells accompanied by an elevated rate of mis-segregation of chromosomes, called chromosome instability (CIN). Aneuploidy contributes to tumor heterogeneity, drug resistance, and treatment failure, which are considered predictors of poor prognosis (8). Previous researches have explored the correlation between CTCs and novel prognostic biomarkers (9). They indicated that the quantification of chromosome ploidy for CTCs might be a useful tool to predict and evaluate therapeutic efficacy as well as to monitoring disease progression (10). However, the relationship between CTCs and specific chromosomal abnormalities, such as aneuploidy, in NSCLC remains poorly understood, and there is a significant gap in our understanding of the biological mechanisms that contribute to tumor progression and treatment resistance.

To address this gap in the literature, this study employed a combination of differential enrichment techniques and fluorescence in situ hybridization (iFISH) to isolate and characterize CTCs from the peripheral blood and pleural effusions of patients diagnosed with NSCLC. This approach significantly enhances the sensitivity and specificity of methods used to quantify CTCs and assess chromosomal aberrations (11). By focusing on the aneuploidy of chromosome 8, we aimed to clarify the relationship between the characteristics of CTCs and patient responses to chemotherapy, which may help identify new prognostic indicators for NSCLC.

The primary aim of this research was to analyze the correlation between CTC counts and chromosomal abnormalities on chromosome 8 in relation to the chemotherapy responses and survival outcomes of NSCLC patients. Aneuploidy of chromosome 8 is often associated with the malignancy, prognosis, and treatment response of tumors. In lung cancer, abnormalities of chromosome 8 may affect the biological characteristics of tumor cells, promoting tumor progression and chemotherapy resistance. Aneuploidy of chromosome 8 is also somewhat correlated with patient survival, suggesting that the status of chromosome 8 could be used as a potential prognostic biomarker in clinical practice. We hypothesized that specific patterns of CTCs and chromosomal changes could serve as reliable biomarkers for predicting treatment efficacy and patient prognosis. By shedding light on the dynamics of CTCs and their chromosomal characteristics, this study sought to contribute to the development of more personalized treatment strategies in NSCLC, which could ultimately improve patient outcomes and survival rates (12).

In summary, given the increasing burden of NSCLC, ongoing research needs to be conducted to discover effective biomarkers for prognosis and the treatment response. This study, which focused on CTCs and their chromosomal features, represents a promising direction, and may enhance our understanding of the biology of NSCLC and improve clinical management strategies. By conducting a thorough analysis and employing innovative methodologies, we aimed to deepen our understanding of the relationship between CTCs and patient outcomes, thereby facilitating advancements in personalized treatments for NSCLC patients. We present this article in accordance with the REMARK reporting checklist (available at https://jtd.amegroups.com/article/view/10.21037/jtd-2025-723/rc).

Methods

Clinical data and sample collection

This study was approved by the Ethics Committee of Northern Jiangsu People’s Hospital Affiliated to Yangzhou University (No. 2013010), and was conducted in accordance with the principles of the Declaration of Helsinki and its subsequent amendments. All patients signed a written informed consent form. The study included 26 newly diagnosed NSCLC patients with malignant pleural effusion but without distant metastasis (stage IVA). All patients were recruited from department of respiratory and critical care medicine, Northern Jiangsu People’s Hospital Affiliated to Yangzhou University between September 1, 2017, and August 31, 2018. Of the 26 patients, 16 were male and 10 were female. The age of the patients ranged from 41 to 74 years (median: 61.5 years). In terms of the histopathological classification, 21 patients had adenocarcinoma and five had squamous cell carcinoma. Malignant pleural effusion was confirmed via medical thoracoscopy biopsy or pleural fluid cytology.

The following patient clinical data were collected: gender, age, smoking history, histopathological type, Eastern Cooperative Oncology Group performance status (ECOG-PS) score, imaging data, first-line treatment regimen, progression-free survival (PFS), and OS. The clinical staging of NSCLC was performed according to the International Union Against Cancer/American Joint Committee on Cancer 8th edition TNM staging criteria. All the enrolled patients were negative for driver genes. The adenocarcinoma patients received first-line chemotherapy with an AP (pemetrexed + platinum) regimen of pemetrexed combined with carboplatin; the squamous cell carcinoma patients received first-line chemotherapy with a TP (paclitaxel + platinum) regimen of paclitaxel combined with cisplatin. Pleural effusion was treated by placing a drainage tube and extracting the fluid, and changes in the pleural fluid and the amount drained were monitored. Peripheral blood was collected from the patients before treatment, and after two cycles of chemotherapy. Regular follow-ups were conducted to determine PFS and OS. Clinical response was assessed according to the Response Evaluation Criteria in Solid Tumors (RECIST) (version 1.1). Each patient’s peripheral blood (6 mL) was collected in an ethylenediaminetetraacetic acid anticoagulant tube, and all samples were processed within 5 hours of collection. Under sterile conditions, 10 mL of pleural fluid was extracted by closed thoracic drainage, and all samples were stored at room temperature, and processed within 24 hours of collection.

Differential enrichment technology [separation enrichment (SE)] for CTC isolation

After the centrifugation of 6 mL of peripheral anticoagulated blood or 10 mL of pleural fluid at 600 g, the supernatant was discarded, and 3 mL of Cytelligen cell separation solution was added, followed by centrifugation at 350 g for 6 minutes. After centrifugation, all liquid from the bottom layer of the sedimented red blood cells was transferred to a 50-mL centrifuge tube, and magnetic beads buffer was slowly added while shaking the centrifuge tube. An immune magnetic bead mixture with anti-cytokeratin 18 antibodies was added and incubated at room temperature for 15 minutes, then placed on a magnetic rack for magnetic separation. After 4 minutes, the liquid in the centrifuge tube was carefully transferred to a new 15 mL centrifuge tube. Cleaning solution was added twice and centrifuged. The supernatant was discarded, and tissue fixing solution was added. The final pellet cells were then counted. The specimen solution was added to the specimen frame of the iFISH slide (Cytointelligen, China) (1 person/slide). Specimens were dried and stored for tumor-labeled iFISH.

iFISH detection

A mixture of tissue fixation and sample dilution solutions were added to cover the specimen box on the CTC slide, and left in the dark at room temperature for 10 minutes; the slide was then placed in an ethanol bath for 2 minutes, removed, and dried. Ten microliters of chromosome 8 centromere probes (CEP8) was added to the center of the specimen box, and a coverslip was placed on top to allow the liquid to spread throughout the specimen box; after sealing the edges of the coverslip with rubber cement, the slide was put in a hybridization instrument for hybridization, and then underwent denaturation at 76 ℃ for 10 minutes, followed by hybridization at 37 ℃ for 3 hours; after removing the coverslip, the slide was dried, and 10 µL of 4',6-diamidino-2-phenylindole (DAPI) staining solution for blood cell analysis was added. A coverslip was then placed on the slide, and it was sealed and stored in the dark at 2–8 ℃.

The CTCs were examined using an Axio Imager.Z2 fluorescence microscope, and the automatic imaging scanning and analysis were performed using the Metafer 5 system.

Efficacy evaluation criteria

The changes in tumor size and clinical responses of all patients were objectively assessed using the RECIST (version 1.1) after completing two cycles (6–8 weeks) of chemotherapy and before the start of the third cycle of chemotherapy. The evaluation criteria included complete response (CR), partial response (PR), stable disease (SD), and progressive disease (PD).

Survival analysis: the patient follow-up continued until December 31, 2021; complete follow-up data were available for all 26 enrolled patients. Patients who had not progressed or were still alive at follow-up were counted as censored data, and all variables were based on patients’ medical records at the time of diagnosis.

Identification of CTCs

The staining data of cluster of differentiation 45 (CD45), DAPI, cytokeratin 18 (CK18) and CEP8 were combined and observed under a fluorescence microscope to identify the CTCs. CD45 immunostaining was used to exclude blood-derived leukocytes; DAPI staining was used to observe the cell nuclei; CK18 is a tumor marker of CTC and CEP8 was used to count the chromosome 8 copy number. Based on the chromosome 8 copy number, the CTCs were classified as mononuclear CTCs, triploid CTCs, tetraploid CTCs, or multiploid CTCs (≥5 copies). To identify the CTCs, the following criteria were used: DAPI⁺/CD45^–/CK18⁺ or CEP8⁺ (monosomic, triploid, tetraploid, polyploid, or diploid).

Statistical analysis

IBM SPSS 26.0 software was used for the statistical analysis. The χ² test was used to analyze the correlation of the positive rates of CTCs and tumor cells in the pleural effusion with the clinical pathological features. The Mann-Whitney U and Kruskal-Wallis tests were used to analyze the correlation of the number of CTCs and tumor cells in the pleural effusion with the clinical pathological features. The Mann-Whitney U test was used to analyze the correlation between different ploidy thresholds of the chromosome 8 CTCs, and PFS and prognosis (OS). A P value <0.05 was considered statistically significant, and all P values were two-sided.

Results

Correlation of peripheral blood CTCs, the positive rate, and the quantity of pleural tumor cells with the clinical characteristics

CTCs in peripheral blood and tumor cells in pleural effusion were detected using SE-iFISH in 26 treatment-naïve patients with newly diagnosed NSCLC accompanied by malignant pleural effusion. CTCs were detected in 24 of the 26 patients. A total of 272 CTCs were detected, averaging 11.3 CTCs per patient (272/24); the CTC positive detection rate was 92.3% (24/26). No significant correlation was found between the CTC positive rate, CTC quantity, and the clinical characteristics of the patients (i.e., gender, age, smoking status, ECOG-PS score, and histological type) (P>0.05) (see Tables 1,2).

Pleural tumor cells were detected in 25 of the 26 patients. A total of 109 pleural tumor cells were detected, averaging 4.4 tumor cells per patient (109/25); the detection rate of pleural tumor cells was 96.2% (25/26). No significant correlation was found between the positive rate of tumor cells in pleural effusion and the patients’ clinical characteristics (i.e., gender, age, smoking status, and ECOG-PS score); however, a correlation with the patients’ histological type was found (P<0.05) (see Table 1). No statistically significant correlation was found between the quantity of tumor cells in pleural effusion and the patients’ clinical characteristics (i.e., gender, age, smoking status, ECOG-PS score, and histological type) (P>0.05) (see Table 2).

Correlation between aneuploid CTC quantity on chromosome 8 and chemotherapy efficacy

This study analyzed the changes in the average quantity of aneuploid CTCs (including monosomy, trisomy, tetrasomy, and polyploidy) on chromosome 8 before and after two cycles of standard first-line chemotherapy in the 26 patients, and evaluated their correlation with chemotherapy efficacy. Among the patients, no CRs were observed, while 14 (53.8%) achieved PR, 5 (19.2%) had stable disease (SD), and 7 (26.9%) experienced progressive disease (PD). After two cycles of first-line chemotherapy, the quantity of aneuploid CTCs decreased from 5.5 (25–75th percentile, 2.75–10.25) to 3 (25–75th percentile, 1.00–5.00), and the difference was statistically significant (P=0.004). The 26 patients were divided into the following three groups based on treatment response: the PR group (14 patients); the SD group (5 patients), and the PD group (7 patients). The quantity of aneuploid CTCs before and after two cycles of chemotherapy showed in Tables 3,4. The results in Table 3 indicate that the changes in aneuploid CTC quantity were related to the treatment response; thus, monitoring the trend of changes in aneuploid CTC quantity before and after chemotherapy could predict treatment efficacy. The results in Table 4 indicate that polyploid CTCs may be more sensitive to chemotherapy than other aneuploid CTCs.

Correlation between CTC chromosome 8 ploidy and chemotherapy efficacy

This study investigated the correlation between changes in chromosome 8 ploidy of aneuploid CTCs and the efficacy of standard first-line chemotherapy, before and after two treatment cycles, in 26 patients. We observed the presence of heterologous polyclonal chromosome 8 in CTCs of advanced NSCLC patients, including monosomy, triploid, tetraploid, and polyploid (≥5 copies). The quantity of different aneuploid CTCs before and after two cycles of chemotherapy showed in Table 5. Before treatment, the occurrence rates of patients with monosomy, trisomy, tetraploidy, and polyploidy were 15.38%, 50.00%, 57.69%, and 80.77%, respectively. After treatment, the occurrence rates of patients with monosomy, trisomy, tetraploidy, and polyploidy were 3.85%, 65.38%, 57.69%, and 30.77%, respectively. Thus, after two cycles of chemotherapy, the frequencies of the monosomy and polyploid CTCs decreased, such that the monosomy CTCs decreased by 11.53% and the polyploidy CTCs decreased by 50.00%. While the frequency of the patients with trisomy CTCs increased from 50.00% to 65.38%. This suggests that polyploid CTCs are more sensitive to chemotherapy, while trisomy CTCs have a higher rate of chemotherapy resistance.

This study also explored the changes in different ploidy CTCs of chromosome 8 after chemotherapy among the patients in the different treatment response groups. The proportion of cases with different aneuploid CTCs before and after two cycles of chemotherapy in PR group and SD + PD group showed in Table 6. Thus, trisomy CTCs emerged, and tetrasomy and polyploidy CTCs disappeared after treatment, indicating effective chemotherapy. From the above results, no decrease in the proportion of advanced NSCLC patients with trisomy CTCs was observed after two cycles of first-line AP chemotherapy, nor was there any increase in the proportion of patients with polyploid CTCs. Conversely, the opposite trend was observed between the different treatment response groups before and after chemotherapy in terms of the proportion of tetrasomy CTCs, suggesting that tetrasomy CTCs may develop acquired resistance after chemotherapy.

Correlation of CTC chromosome 8 ploidy with PFS and OS

This study further explored the correlation between the aneuploid CTC count and patient prognosis after two cycles of first-line chemotherapy. We compared the correlation of different counts of trisomy, tetrasomy, and polyploid CTCs with patient PFS and OS (see Table 7). After treatment, the patients’ peripheral blood had 0 (9 cases), 1 (12 cases), and ≥2 (5 cases) trisomy CTCs. No statistically significant difference was found in the PFS and OS of the patients with a trisomy CTC count ≥1 and those with a trisomy CTC count of 0 (P=0.18, P=0.33); nor was any statistically significant difference found in the PFS and OS of patients with a trisomy CTC count ≥2 or a trisomy CTC count <2 (P=0.43, P=0.72).

After treatment, the patients’ peripheral blood had 0–1 (12 cases), 2 (5 cases), and ≥3 (9 cases) tetrasomy CTCs. The results showed that there was a statistically significant difference in the PFS of patients with a tetrasomy CTC count ≥2 and those with a tetrasomy CTC count <2 (P=0.03) (see Figure 1A), but no statistically significant difference was found in terms of OS (P=0.27). Further, a statistically significant difference was found in the PFS of patients with a tetrasomy CTC count ≥3 and those with a tetrasomy CTC count <3 (P=0.01) (see Figure 1B), but no statistically significant difference was found in terms of OS (P=0.16).

Figure 1 Correlation between PFS and different tetrasomy CTC count. (A) Tetrasomy CTC count <2 and ≥2. (B) Tetrasomy CTC count <3 and ≥3. CTC, circulating tumor cell; PFS, progression-free survival.

After treatment, the patients’ peripheral blood had 0 (18 cases) and ≥1 (8 cases) polyploid CTCs. The results showed that there was no statistically significant difference in the PFS and OS of the patients with a polyploid CTC count ≥1 and those with a polyploid CTC count of 0 (P=0.11, P=0.29).

Thus, after two cycles of chemotherapy, the patients with a tetrasomy CTC count ≥2 had a significantly shorter PFS than those with a tetrasomy CTC count <2 {3.5 [3–8] vs. 8 [5–10.5] months, P=0.03} (see Table 7). Therefore, unfavorable (≥2) tetrasomy CTCs after treatment predicted worse PFS in advanced NSCLC patients.

Correlation between tumor cell count in pleural effusion chromosome 8 ploidy, and PFS and OS

This study analyzed the relationship between the tumor cell count in pleural effusion and patient prognosis. We examined the correlation between trisomy, tetrasomy, and polyploid tumor cell count in pleural effusion on chromosome 8, and patient PFS and OS (see Table 8). Before treatment, there were 0 (9 cases) and ≥1 (17 cases) trisomy tumor cells in patients’ pleural effusion. The results revealed a statistically significant difference in the PFS and OS of patients with a trisomy tumor cell count ≥1 and those with a count of 0 (P=0.02, P=0.046) (see Figures 2,3).

Figure 2 Correlation between PFS and trisomy tumor cell count in pleural effusion ≥1 and 0. PFS, progression-free survival.

Figure 3 Correlation between OS and trisomy tumor cell count in pleural effusion ≥1 and 0. OS, overall survival.

Before treatment, there were 0 (13 cases) and ≥1 (13 cases) tetrasomy tumor cells in patients’ pleural effusion. The results revealed a statistically significant difference in the PFS and OS of patients with a tetrasomy tumor cell count ≥1 and those with a count of 0 (P=0.001, P=0.002) (see Figures 4,5).

Figure 4 Correlation between PFS and tetrasomy tumor cell count in pleural effusion ≥1 and 0. PFS, progression-free survival.

Figure 5 Correlation between OS and tetrasomy tumor cell count in pleural effusion ≥1 and 0. OS, overall survival.

Before treatment, there were 0 (14 cases) and ≥1 (12 cases) polyploid tumor cells in patients’ pleural effusion. The results revealed a statistically significant difference in the PFS and OS of patients with a polyploid tumor cell count ≥1 and those with a count of 0 (P=0.03, P=0.02) (see Figures 6,7).

Figure 6 Correlation between PFS and polyploid tumor cell count in pleural effusion ≥1 and 0. PFS, progression-free survival.

Figure 7 Correlation between OS and polyploid tumor cell count in pleural effusion ≥1 and 0. OS, overall survival.

Before treatment, there were ≤2 (12 cases) and >2 (14 cases) tumor cells in patients’ pleural effusion. The results revealed no statistically significant difference in the PFS of patients with a tumor cell count ≤2 and those with a count >2 (P=0.07), but a statistically significant difference was observed in terms of OS (P=0.04) (see Figure 8).

Figure 8 Correlation between OS and tumor cell count in pleural effusion. OS, overall survival.

Before treatment, there were ≤3 (15 cases) and >3 (11 cases) tumor cells in patients’ pleural effusion. The results revealed no statistically significant difference in the PFS and OS of patients with a tumor cell count ≤3 and those with a count >3 (P=0.13, P=0.08).

Additionally, among the 26 patients, only two had monosomy tumor cells in their pleural effusion, and these two patients had a PFS of 2 and 3 months, and an OS of 12 and 8 months, respectively, indicating a poor prognosis.

Thus, advanced NSCLC patients with pleural tumor cells, whether trisomy, tetrasomy, polyploidy, or monosomy, have significantly shortened PFS and OS. Therefore, pleural tumor cells may be a predictive factor for a poor prognosis, and patients with tumor cell counts >2 have shorter OS.

Discussion

NSCLC is the predominant subtype of lung cancer, accounting for approximately 85% of all cases (13). The complexity of NSCLC is underscored by its diverse histological types, which include adenocarcinoma and squamous cell carcinoma, which exhibit varying responses to treatment. Current therapeutic options primarily consist of surgery, chemotherapy, radiotherapy, targeted therapy, and immunotherapy. Unfortunately, the prognosis of patients with advanced NSCLC is often poor due to issues such as treatment resistance and disease progression (14). Thus, it is essential that we better understand the underlying biological mechanisms of NSCLC, and identify reliable biomarkers of prognosis and the treatment response (6,15).

This study sought to investigate the role of CTCs and their chromosomal abnormalities, specifically the aneuploidy of chromosome 8, in predicting the treatment response and OS of NSCLC patients. Using advanced techniques such as SE and iFISH, we analyzed the presence and characteristics of CTCs in patients with malignant pleural effusion. Our findings revealed a significant correlation between CTC counts, chromosomal abnormalities, and treatment outcomes, indicating that CTCs may serve as crucial prognostic biomarkers in the management of NSCLC. These insights could enhance clinical decision making and contribute to the development of personalized treatment strategies (16).

Ploidy increase has been shown to occur in different type of tumors and participate in tumor initiation and resistance to the treatment (17). Polyploid cancer cells contribute significantly to cellular heterogeneity and can arise from various stressors, including radiation, chemotherapy, hypoxia, and environmental factors. The formation of polyploid cancer cells can occur through mechanisms such as endoreplication, cell fusion, cytokinesis failure, mitotic slippage, or cell cannibalism (18).

The present study provides novel insights into the implications of CTCs and chromosomal abnormalities, including the role of aneuploidy in patients with NSCLC. This research filled a notable gap in the existing literature by establishing a direct connection between the presence of CTCs and the chromosomal characteristics of these cells, as well as their influence on the clinical outcomes of the patients receiving chemotherapy. While Zhou et al. focused on the general role of CTCs in cancer metastasis and prognosis (19), our findings specifically emphasize that certain chromosomal abnormalities, particularly those involving chromosome 8, can serve as significant predictors of the treatment response and OS in NSCLC patients. Lu et al. highlighted the prognostic importance of CTC counts but did not delve into the genetic aspects of these cells. Thus, the originality of our contributions to the field should be noted (16).

The implications of our findings go far beyond mere academic interest; they open up significant opportunities for clinical practice and policy development. By identifying CTCs with chromosomal alterations as reliable biomarkers for predicting responses to chemotherapy, healthcare providers can tailor treatment plans more effectively, ultimately improving patient outcomes. Our results indicate that patients with a higher count of four ploidy CTCs have a notably shorter PFS than those with lower counts. This relationship underscores the importance of incorporating CTC analysis into standard clinical evaluations, which could lead to more personalized treatment strategies and better resource allocation in healthcare systems (3). Further, these findings could inform healthcare policies aimed at optimizing patient stratification and management approaches for NSCLC (6).

Another contribution of our research is the comparative analysis of CTCs detected in both peripheral blood and pleural effusions, which fills a critical knowledge gap regarding the sources of CTCs in NSCLC. Previous studies have predominantly focused on CTC detection in peripheral blood, often overlooking the potential insights provided by pleural effusions, which can harbor a higher concentration of CTCs in advanced disease stages. Our findings indicate that the presence of CTCs in pleural effusions correlates significantly with worse prognosis and treatment outcomes, thus highlighting the importance of utilizing multiple fluid sources for comprehensive tumor monitoring (20,21).

The study had several limitations. First, the sample size of the study was relatively small, which, coupled with the absence of long-term follow-up data, might restrict the generalizability and replicability of our findings. Second, as the research was conducted at a single center, there is a potential for biases related to the examination of a localized patient population. These limitations could affect the robustness and reproducibility of our results. Thus, in the future, larger, multi-center studies should be conducted to validate our findings, and explore the biological mechanisms underlying CTC aneuploidy in NSCLC more thoroughly. Moreover, longitudinal studies need to be conducted to assess the long-term implications of CTC characteristics on patient survival and treatment efficacy. This strategy will provide a more profound understanding of the intricate dynamics involved in NSCLC progression and treatment responses (12). Third, the lack of comprehensive biological validation via wet laboratory experiments limits our ability to draw definitive conclusions about the mechanisms underlying the CTC characteristics and the chemotherapy response. Given these limitations, our results should be interpreted cautiously, and further larger studies with extended follow-up periods and further experimental validation are needed to establish the clinical significance of CTCs as prognostic biomarkers for NSCLC.

In summary, this study found a significant correlation between CTC counts and chromosomal aneuploidy, and the prognosis of NSCLC patients, which suggests that these elements could serve as important prognostic indicators. Given the relationship between CTC dynamics and the treatment response, CTC monitoring could be used to personalize treatment strategies (22). To confirm our findings and explore the clinical application of CTCs as dependable biomarkers for guiding treatment decisions in advanced NSCLC, future studies with larger patient cohorts need to be conducted. Moreover, as the field of liquid biopsy continues to evolve, future research should explore the integration of CTCs with other biomarkers, such as circulating tumor DNA (ctDNA), to enhance the accuracy of tumor monitoring and optimize personalized treatment strategies. This combined approach holds promise for improving patient outcomes in the context of NSCLC.

Conclusions

A significant decrease in the aneuploid CTC counts of the patients who exhibited PRs to chemotherapy. The detection of ≥2 trisomy or tetrasomy CTCs post-treatment was associated with shorter PFS. The presence of aneuploid tumor cells in pleural effusion was correlated with poorer PFS and OS outcomes. So, CTC counts and their chromosomal features could serve as significant prognostic indicators for predicting treatment responses in NSCLC. Future studies with larger patient cohorts need to be conducted to validate these findings and assess the feasibility of using CTC analyses as routine clinical instruments.

Acknowledgments

We would like to thank all participating patients for enrolling in this study. We also would like to thank all research nurses for their help with patient enrolment and peripheral venous blood collection.

Footnote

Reporting Checklist: The authors have completed the REMARK reporting checklist. Available at https://jtd.amegroups.com/article/view/10.21037/jtd-2025-723/rc

Data Sharing Statement: Available at https://jtd.amegroups.com/article/view/10.21037/jtd-2025-723/dss

Peer Review File: Available at https://jtd.amegroups.com/article/view/10.21037/jtd-2025-723/prf

Funding: This work was supported by Clinical Medicine Science and Technology Special Project of Jiangsu Province–New Clinical Diagnosis and Treatment Technology Tackling (No. BL2012054).

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://jtd.amegroups.com/article/view/10.21037/jtd-2025-723/coif). A.T. serves as an unpaid editorial board member of Journal of Thoracic Disease from August 2023 to July 2025. The other 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 and its subsequent amendments, and approved by the Ethics Committee of Northern Jiangsu People’s Hospital Affiliated to Yangzhou University (No. 2013010). All patients signed a written informed consent form.

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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(English Language Editor: L. Huleatt)