Yang J, Li ZX, Song MJ, Han SJ, Yang AJ, Zhang ZP, Sui CS, Qiao JL, Huang WH, He JQ. Prognostic value and therapeutic efficacy of interstitial circulating tumor cells in patients with advanced gastric cancer. World J Clin Oncol 2025; 16(5): 101762 [DOI: 10.5306/wjco.v16.i5.101762]
Corresponding Author of This Article
Wen-Hua Huang, PhD, Professor, Guangdong Engineering Research Center for Translation of Medical 3D Printing Application, Guangdong Provincial Key Laboratory of Medical Biomechanics, National Key Discipline of Human Anatomy, School of Basic Medical Sciences, Southern Medical University, No. 1023 Shatai North Road, Guangzhou 510000, Guangdong Province, China. huangwenhua2009@139.com
Research Domain of This Article
Surgery
Article-Type of This Article
Retrospective Study
Open-Access Policy of This Article
This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Jing Yang, Wen-Hua Huang, Guangdong Engineering Research Center for Translation of Medical 3D Printing Application, Guangdong Provincial Key Laboratory of Medical Biomechanics, National Key Discipline of Human Anatomy, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510000, Guangdong Province, China
Jing Yang, Department of General Surgery, Gansu Provincial Hospital, Lanzhou 730030, Gansu Province, China
Jing Yang, Key Laboratory of Molecular Diagnostics and Precision Medicine for Surgical Oncology in Gansu Province, Gansu Provincial Hospital, Lanzhou 730030, Gansu Province, China
Zu-Xi Li, Department of Peripheral Vascular Intervention, Gansu Provincial Hospital of Traditional Chinese Medicine, Lanzhou 730060, Gansu Province, China
Mei-Juan Song, Shang-Jun Han, Ai-Jia Yang, Ze-Ping Zhang, Chang-Sheng Sui, Ji-Lin Qiao, The First Clinical Medical College of Gansu University of Chinese Medicine, Lanzhou 730030, Gansu Province, China
Jun-Qiang He, Department of General Surgery, Xinhui People’s Hospital of Southern Medical University, Jiangmen 529000, Guangdong Province, China
Co-corresponding authors: Wen-Hua Huang and Jun-Qiang He.
Author contributions: Yang J and Li ZX contributed to study design, and contributed equally as co-first authors; Li ZX contributed to data analysis and manuscript preparation; Song MJ and Han SJ contributed to data acquisition and analysis; Yang AJ, Zhang ZP, and Sui CS contributed to data visualization and design enhancement; Qiao JL contributed to academic literature research; Huang WH and He JQ contributed to manuscript evaluation and supervision, and contributed equally as co-corresponding authors; and all authors contributed to the article and approved the submitted version.
Supported by China Postdoctoral Science Foundation, No. 2024M751334.
Institutional review board statement: This study was approved by the Ethics Committee of Gansu Provincial People’s Hospital (No.2020-208).
Informed consent statement: All study participants, or their legal guardian, provided informed written consent prior to study enrollment.
Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article.
Data sharing statement:sharing statement: All authors agree that all data related to this study will be publicly available.
Open Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: https://creativecommons.org/Licenses/by-nc/4.0/
Corresponding author: Wen-Hua Huang, PhD, Professor, Guangdong Engineering Research Center for Translation of Medical 3D Printing Application, Guangdong Provincial Key Laboratory of Medical Biomechanics, National Key Discipline of Human Anatomy, School of Basic Medical Sciences, Southern Medical University, No. 1023 Shatai North Road, Guangzhou 510000, Guangdong Province, China. huangwenhua2009@139.com
Received: September 25, 2024 Revised: March 8, 2025 Accepted: April 8, 2025 Published online: May 24, 2025 Processing time: 236 Days and 3.7 Hours
Abstract
BACKGROUND
The high mortality rate and recurrence/metastasis remain major challenges in the clinical management of gastric cancer (GC) patients. To optimize treatment stratification and management, there is an urgent need for efficient and non-invasive biomarkers. A meta-analysis on the prognostic role of circulating tumor cells (CTCs) in GC revealed a strong association between CTCs and patient prognosis. Among CTC subtypes, Interstitial CTCs (I-CTCs) exhibited the strongest invasiveness. This study innovatively investigated the expression profile of I-CTCs in advanced GC patients to evaluate their clinical utility.
AIM
To evaluate the clinical utility of I-CTCs as a non-invasive prognostic biomarker in advanced GC. To investigate the correlation between I-CTC count thresholds and chemotherapy efficacy in advanced GC patients. To establish the potential of preoperative I-CTC profiling for optimizing treatment stratification and postoperative surveillance.
METHODS
This study retrospectively analyzed 59 patients with advanced GC treated at the General Surgery Clinical Medical Center of Gansu Provincial Hospital between October 2019 and October 2020. The expression levels of I-CTCs were measured, and patient survival was monitored. The receiver operating characteristic curve was plotted to determine the optimal cut-off value for I-CTCs expression levels. Based on this cut-off value, 59 GC patients were grouped into positive and negative groups. The differences in clinicopathological characteristics between the two groups were analyzed. Patient survival was follow-up and recorded until October 2022. Plotting survival curves and performing univariate and multifactorial analyses of patient prognostic factors. The Kaplan-Meier method and Cox regression model were used, respectively.
RESULTS
A total of 59 patients were included in this study, and receiver operating characteristic curve analysis showed that the best cut-off value for I-CTCs was 5, with an area under the curve of 0.8356 (95%CI: 0.7122-0.9590). The I-CTC count of ≥ 5 defines the positive group, while counts < 5 are classified as the negative group. Positive I-CTCs correlated with the degree of tumor differentiation and disease progression (P < 0.05). 16 of 59 patients received neoadjuvant chemotherapy. There were divided into progressive disease and disease control groups based on response to neoadjuvant chemotherapy. Patients in the I-CTCs-negative group had longer overall survival and disease-free survival than those in the positive group (P < 0.05). Multifactorial analysis revealed that I-CTCs positivity (HR = 13.323, 95%CI: 1.675-105.962, P = 0.014) was an independent risk factor for survival in patients with advanced GC.
CONCLUSION
In patients with advanced GC, an I-CTC count of ≥ 5 is associated with both poor prognosis and reduced chemotherapy efficacy. I-CTCs may serve as a valuable preoperative biomarker for predicting the prognosis of advanced GC.
Core Tip: This study builds upon prior evidence-based research to innovatively assess the expression of interstitial circulating tumor cell (I-CTCs) in patients with advanced gastric cancer, demonstrating that I-CTCs can serve as valid indicators for evaluating chemotherapy efficacy and prognosis in this patient population. In patients with advanced gastric cancer, I-CTC count of ≥ 5 is associated with poor prognosis and reduced chemotherapy efficacy.
Citation: Yang J, Li ZX, Song MJ, Han SJ, Yang AJ, Zhang ZP, Sui CS, Qiao JL, Huang WH, He JQ. Prognostic value and therapeutic efficacy of interstitial circulating tumor cells in patients with advanced gastric cancer. World J Clin Oncol 2025; 16(5): 101762
Gastric cancer (GC) is one of the most common malignancies worldwide[1] and has the third-highest mortality rate in China[2]. The treatment of GC includes surgery, chemotherapy, radiotherapy, targeted therapy, immunotherapy, as well as supportive care and clinical trials, while emphasizing the importance of personalized treatment and a multidisciplinary approach[3]. It is characterized by a multifactorial pathogenesis and a heterogeneous geographic distribution[1-2,4]. The high mortality rate of malignant tumors is closely associated with metastasis, and GC is no exception[5]. Many GC patients experience recurrent metastases following surgical treatment, leading to a poor prognosis[6]. Timely disease management through prognostic surveillance can improve clinical outcomes. Conventional tumor monitoring methods for GC including imaging techniques, tissue biopsy, and various clinical indicators, have limited accuracy owing to time delays and low sensitivity and specificity. Currently, an effective biomarker for identifying early relapse and assessing chemotherapy response in GC patients remains underexplored[7]. Therefore, there is an urgent need for a more precise biomarker to predict patient outcomes and stratify them into different risk groups for appropriate postoperative interventions.
Circulating tumor cells (CTCs) were first observed in the blood of patients with metastatic cancer by Ashworth in 1869[8]. Subsequently, numerous studies have identified CTCs as key contributors to tumor recurrence and metastasis[9]. They are defined as tumor cells that enter the circulation either owing to medical intervention or spontaneous detachment from the primary or metastatic lesion[10]. CTCs are classified into three main categories based on their phenotype: Epithelial, mesenchymal, and mixed. Castro-Giner et al[11] found that the vast majority of CTCs undergo epithelial-mesenchymal transition (EMT) during tumor metastasis. This transition enhances their resistance to immune system attacks and mechanical forces in the circulatory system. In addition, interstitial CTCs (I-CTCs), which undergo EMT, exhibit greater resilience than epithelial CTCs[12]. Experimental evidence suggests that mesenchymal CTCs are significantly more resistant than epithelial CTCs and have a much higher chance of survival in the circulatory system[13]. Owing to the heterogeneity of CTCs, they exhibit more mesenchymal characteristics than cancer cells from primary or metastatic tumors. The expression of mesenchymal markers has been observed in CTCs from patients with various cancers, including breast[14], hepatocellular carcinoma[15], nasopharyngeal[16], bladder[17], pancreatic[18], non-small cell lung[19], colon[20], and GC[21]. In studies examining the relationship between breast cancer and CTC subtypes, only I-CTCs are significantly associated with shorter progression-free survival and overall survival (OS) in breast cancer patients, which indicates that I-CTCs are the most aggressive subtype[22,23]. The clinical validity of CTCs in breast cancer has been well established, which demonstrates their value in prognostic assessment and in predicting treatment efficacy for metastatic breast cancer[24-26]. Consequently, CTC testing has been incorporated into the Chinese Society of Clinical Oncology Breast Cancer Guidelines[27]. In recent years, CTCs have been increasingly studied in GC. Research has shown that CTCs not only help assess prognosis and treatment efficacy in GC patients[28-31] but are also closely associated with recurrent metastasis[21,32]. Our study aims to investigate the relationship between I-CTCs and both prognosis and chemotherapy efficacy in GC patients. We seek to clarify the clinical significance of I-CTCs in disease progression and treatment response via the detection of pre-treatment I-CTC expression levels in patients with locally advanced GC.
MATERIALS AND METHODS
Research subjects and ethics statement
GC patients treated at the General Surgery Clinical Medical Center of Gansu Provincial Hospital between October 2019 and October 2020 were retrospectively analyzed. Inclusion criteria were: (1) Age 18 years or older; (2) Pre-treatment endoscopic biopsy confirming a pathological diagnosis of gastric adenocarcinoma with clinical stage II-III; (3) I-CTC testing and quantification performed before treatment; (4) Postoperative chemotherapy with SOX or XELOX regimens in accordance with the Chinese Society of Clinical Oncology guidelines for GC[31,33]; (5) No prior therapeutic interventions before enrollment; (6) Stage III patients receiving neoadjuvant chemotherapy were treated with the SOX regimen; (7) Patients who underwent surgery received radical gastrectomy with D2 lymph node dissection; and (8) Complete clinical information available. Exclusion criteria were: (1) Patients with severe cardiopulmonary insufficiency who are unable to tolerate surgery; (2) Patients with other malignancies or serious diseases; (3) Stage IV GC patients receiving palliative care; (4) Patients with immunological diseases or those undergoing immunotherapy; and (5) Patients with incomplete clinicopathological data or missed follow-up visits. According to these criteria, 59 patients with locally advanced GC were ultimately included in the study (Figure 1). The study was approved by the Medical Ethics Committee of Gansu Provincial Hospital (Approval No. 2020-208). Patient identities remained anonymous, and the requirement for informed consent was waived owing to the retrospective study design.
Figure 1 Screening flow chart of patients with advanced gastric cancer.
I-CTC: Interstitial circulating tumor cells.
CTC detection method
The I-CTC assay was performed using third-generation microfluidic chip technology (nano-microfluidic chip + immunofluorescence staining). The CytoSorter® system, along with the CytoSorter™ I-CTC Kit (Watson Biotech, China), was used for detection. The I-CTC test kit was provided by Hangzhou Huadeisen Biotechnology Co. A 7.5 mL peripheral blood sample, collected within one week before treatment, was diluted at a 1:1 ratio with phosphate buffered saline to a final volume of 15 mL. The sample was then evenly distributed into two separate tubes, each containing 3 mL of density-gradient centrifugal buffer. The assay works by first washing the peripheral blood sample to isolate a layer of peripheral blood mononuclear cells. The cell capture agent was encapsulated on a nano-microfluidic chip (CytoNanoChip). Peripheral blood mononuclear cell samples were introduced into the CytoNanoChip through the spiral channel with X design spiral sample chamber to enrich CTCs. Immunofluorescence staining was performed using CSV-FITC, CD45-PE, and Hoechst to detect I-CTCs. Finally, I-CTCs were isolated and identified. Biotin-labeled epithelial adhesion protein (EpCAM) antibodies were used to capture CTCs. Anti-CSV (FITC)-Ab and anti-CD45 (PE)-Ab was used to identify I-CTCs and white blood cells, respectively, while nuclei were stained with Hoechst 33342. The experimental results were visualized using fluorescence markers of different colors (Figure 2).
Figure 2 Immunofluorescent staining of interstitial circulating tumor cells in gastric cancer patients.
Interstitial circulating tumor cells are identified as DAPI (blue) positive, CSV (FITC, green) positive, and CD45 (PE, orange) negative cells. Scale bar represents 10 μm, immunofluorescent staining, 20 ×.
Data collection and follow-up
Clinical information was collected from patients, including age, gender, pre-treatment I-CTC count, tumor markers [carcinoembryonic antigen (CEA), carbohydrate antigen 19-9 (CA19-9), CA125, and CA72-4], tumor differentiation grade, clinical stage, T stage, lymph node metastasis, postoperative chemotherapy regimen, and response to chemotherapy. The efficacy of preoperative neoadjuvant chemotherapy was assessed based on the RECIST 1.1 (response evaluation criteria in solid tumors). Tumor response, as measured by imaging, was classified into four categories: Complete response, partial response, stable disease, and progressive disease (PD). Patients were categorized into two groups: PD and disease control (DC), with DC defined as complete response + partial response + stable disease. Patient survival status and disease progression were monitored through outpatient reviews, phone calls, and WeChat. Follow-up was conducted every 1 to 3 months during the first year after surgery and every 3 months in the second year. OS was measured from the date of surgery to the follow-up endpoint, which was defined as either patient death or the follow-up cut-off date. Disease-free survival (DFS) was defined as the time from surgical resection to local recurrence.
Statistical analysis
Statistical analyses were performed using SPSS version 25.0 and GraphPad Prism version 8. Categorical data were expressed as n (%) and analyzed using the χ2-test to compare differences between groups. Receiver operating characteristic (ROC) curves (Figure 3) were constructed based on I-CTC expression levels and patient survival outcomes[34]. The optimal cut-off value for I-CTCs was determined by selecting the threshold that maximized the Youden index and effectively stratified patients into two groups with statistically significant differences in relapse rates. Cox regression models were applied for univariate and multivariate analyses. Survival curves were plotted using GraphPad Prism 8. A P value of < 0.05 was considered statistically significant.
Figure 3 Relationship between interstitial circulating tumor cells and gastric cancer.
A: Interstitial circulating tumor cells (I-CTCs) expression status before treatment; B: I-CTCs and prognostic receiver operating characteristic curves of patients with advanced gastric cancer (GC); C: Two-year overall survival curves of I-CTCs-positive and negative groups of patients with advanced GC; D: Two-year disease-free survival curves of I-CTCs-positive and negative groups of patients with advanced GC. I-CTC: Interstitial circulating tumor cell; AUC: Area under the curve; OS: Overall survival; DFS: Disease-free survival.
RESULTS
Relationship between I-CTCs and clinicopathological characteristics in patients with advanced GC
A total of 59 patients with GC were included in this study, which comprised 46 (78%) male and 13 (22%) female patients. The ages of the patients ranged from 31 to 82 years, with a mean age of 57.9 years. Tumor-node-metastasis (TNM) staging was determined according to the eighth edition of the American Joint Committee on Cancer staging system[35], with 30 patients classified as stage II and 29 as stage III. According to ROC curve analysis, the optimal cut-off value for I-CTCs was determined to be 5, with an area under the curve of 0.8356 (95%CI: 0.7122-0.9590) (Figure 4). I-CTC counts < 5 were classified as negative, while counts ≥ 5 were considered positive based on the determined cut-off value. Among the 59 patients, 33 (55.9%) were in the I-CTC-positive group, and 26 (44.1%) were in the I-CTC-negative group. I-CTC positivity showed no significant correlation with patient gender, age, depth of infiltration, TNM stage, lymph node metastasis, or tumor markers (CA125, CA19-9, CA72-4, and CEA). However, a statistically significant association was observed between I-CTC positivity and the degree of tumor differentiation (P < 0.05) (Table 1).
Table 1 Relationship between interstitial circulating tumor cells and clinicopathological characteristics of patients with advanced gastric cancer, n (%).
Clinicopathological features
I-CTCs negative (n = 26)
I-CTCs positive (n = 33)
χ2
P value
Gender
Male
20 (43.5)
26 (56.5)
Female
6 (46.2)
7 (53.8)
0.029
0.864
Age, years
< 60
18 (52.9)
16 (47.1)
≥ 60
8 (32)
17 (68.0)
2.563
0.109
TNM staging
II
16 (53.3)
14 (46.7)
III
10 (34.5)
19 (65.5)
2.126
0.145
Infiltration depth
T1-T2
10 (58.8)
7 (41.2)
T3-T4
16 (38.1)
26 (61.9)
2.110
0.146
Lymph node metastasis
N0-N1
17 (54.8)
14 (45.2)
N2-N3
9 (32.1)
19 (67.9)
3.075
0.080
Degree of differentiation
Well
19 (59.4)
13 (40.6)
Poor
7 (25.9)
20 (74.1)
6.647
0.010
CEA
< 5
23 (47.9)
25 (52.1)
≥ 5
3 (27.3)
8 (72.7)
1.547
0.214
CA125
< 35
22 (42.3)
30 (57.7)
≥ 35
4 (57.1)
3 (42.9)
0.551
0.458
CA19-9
< 37
20 (41.7)
28 (58.3)
≥ 37
6 (54.5)
5 (45.5)
0.602
0.438
CA72-4
< 10
21 (41.2)
30 (58.8)
≥ 10
5 (62.5)
3 (37.5)
1.276
0.259
Relationship between I-CTCs and the efficacy of chemotherapy in patients with advanced GC
A total of 16 patients with locally progressive GC underwent neoadjuvant chemotherapy before radical gastrectomy. The chemotherapy regimens included SOX and XELOX, with 13 patients receiving SOX and 3 patients receiving XELOX. Among these patients, 6 (46.2%) in the SOX group and 1 (33.3%) in the XELOX group were I-CTC-positive, with no statistically significant difference between the two groups (P > 0.05). Chemotherapy efficacy was assessed based on RECIST criteria version 1.1, which classified patients into the PD and DC groups. Six patients were in the PD group, and 10 were in the DC group. I-CTC positivity was observed in 5 (83.3%) cases in the PD group and 2 (20%) cases in the DC group, with a statistically significant difference between the groups (P < 0.05) (Table 2).
Table 2 Relationship between interstitial circulating tumor cells and the efficacy of chemotherapy in patients with advanced gastric cancer, n (%).
I-CTCs negative (n = 9)
I-CTCs positive (n = 7)
χ2
P value
Chemotherapy regimens
SOX
7 (53.8)
6 (46.2)
XELOX
2 (66.7)
1 (33.3)
0.163
0.687
Chemotherapy efficacy
DC
8 (80)
2 (20)
PD
1 (16.7)
5 (83.3)
6.112
0.013
Relationship between I-CTCs and survival time in patients with advanced GC
This study included 59 patients with complete follow-up data. The median follow-up time for OS and DFS was 27.3 months and 18.8 months, respectively. During the follow-up period, 15 patients died. Survival curve analysis showed that patients in the I-CTCs-negative group had significantly longer OS and DFS compared with those in the I-CTCs-positive group (P < 0.05) (Figure 5). Univariate analysis indicated that gender, age, chemotherapy regimen, TNM stage, depth of infiltration, lymph node metastasis, and tumor markers (CEA, CA125, CA19-9, and CA72-4) were not significantly associated with OS (all P > 0.05). However, the degree of tumor differentiation and I-CTC count were significantly correlated with OS in patients with GC (P < 0.05) (Table 3). Multivariate analysis further identified I-CTC positivity as an independent risk factor for survival in patients with advanced GC (P < 0.05) (Table 4).
Table 3 Results of univariate analysis affecting the prognosis of patients with advanced gastric cancer.
Prognostic factors
HR
95%CI
P value
Gender, male vs female
0.975
0.271-0.501
0.969
Age, < 60 years vs ≥ 60 years
2.427
0.859-6.851
0.094
Degree of differentiation, well vs poor
4.076
1.291-12.863
0.017
TNM staging, I-II vs III
1.041
0.365-2.968
0.941
Infiltration depth, T1-T2 vs T3-T4
3.676
0.821-16.462
0.089
Lymph node metastasis, N0-1 vs N2-3
1.020
0.357-2.912
0.970
I-CTCs, negative vs positive
17.158
2.227-132.196
0.006
CEA, < 5 vs ≥ 5
1.732
0.531-5.646
0.362
CA125, < 35 vs ≥ 35
0.041
0-66.291
0.397
CA19-9, < 37 vs ≥ 37
0.348
0.045-2.663
0.309
CA72-4, < 10 vs ≥ 10
0.936
0.208-4.205
0.931
Table 4 Results of multifactorial analysis affecting the prognosis of patients with advanced gastric cancer.
Prognostic factors
HR
95%CI
P value
Degree of differentiation, well vs poor
2.357
0.725-7.662
0.154
I-CTCs, negative vs positive
13.323
1.675-105.962
0.014
DISCUSSION
CTC testing, as one of the “liquid biopsy” techniques, has broad application prospects in tumor efficacy assessment. CTC testing is applied in guiding postoperative precision interventions, monitoring immunotherapy, and surveilling tumor recurrence and metastasis. Its ease of operation and non-invasive nature make it particularly valuable for prognostic evaluation in cancer patients[36]. Currently, traditional diagnostic methods for GC have significant limitations in prognosis assessment and efficacy monitoring. While imaging can delineate the relationship between the tumor and surrounding tissues, it cannot fully characterize the tumor and it also involves radiation exposure. The sensitivity and specificity of serum markers such as CEA, CA19-9, and CA72-4 are insufficient for reliable disease monitoring. In addition, a significant proportion of patients with advanced disease lack access to multiple tissue samples, which makes dynamic disease monitoring challenging. The temporal and spatial heterogeneity of tumor characteristics further limits the effectiveness of pathological examinations. CTC assays offer several advantages, including the ability to be performed using blood samples, minimal invasiveness, repeatability, and continuous sampling for dynamic monitoring. This technique helps overcome the spatial and temporal heterogeneity of tumors and makes it a valuable tool for disease assessment. The vast majority of CTCs perish in circulation, likely owing to oxidative stress, a lack of growth factors and cytokines, and the body’s immune response. However, a small subset of surviving CTCs plays a crucial role in tumor recurrence and metastasis[37]. Some of these CTCs exit the bloodstream, proliferate, and establish metastatic colonies, while others enter a dormant state and retain the potential to develop into metastatic lesions over time.
CTC detection involves three key steps: Enrichment, detection, and analysis. Enrichment methods are classified into two main categories: Physical enrichment and bioenrichment. Physical enrichment is based on differences in size, density, and charge and allows CTC isolation without the need for immunolabeling. In contrast, bioenrichment relies on immune antibodies to specifically capture CTCs and is further divided into positive and negative selection methods[38]. Negative selection mainly utilizes the leukocyte surface marker CD45 to remove leukocytes, while positive selection isolates CTCs based on specific markers they express, such as EpCAM. The CellSearch system, the first assay platform approved by the United States Food and Drug Administration for clinical use[39], remains one of the most widely recognized CTC detection methods. However, relying solely on EpCAM enrichment presents a notable limitation of immunocapture techniques, including the CellSearch system[40]. In response, numerous new methods have been developed in recent years to improve CTC detection. Examples include AdnaTest, epithelial tumor cell size separation, density gradient centrifugation, microfiltration, microfluidics, and size-based immunocapture microarrays[41,42]. Techniques for CTC detection and analysis include polymerase chain reaction and cellular protein detection methods such as immunofluorescence, immunohistochemistry, and fluorescence in situ hybridization, though these are considered more conventional approaches. More advanced detection methods include high-throughput fiber optic array scanning technology, sequencing, and histological techniques. Nagrath et al[43] developed the “CTC chip”, a microfluidic-based device for detecting CTCs, which offers higher detection efficiency. With the continuous advancement of CTC detection technology, the third-generation microfluidic chip technology - based on physical or biochemical properties - is currently the mainstream method used in clinical applications, including in our experiments.
Our study demonstrates that I-CTCs play a significant role in the process of CTC-driven tumor metastasis[44]. CTC detection shows significant potential in tumor treatment efficacy assessment, chemotherapy guidance, immunotherapy monitoring, and recurrence/metastasis surveillance[32,37]. This research focuses on evaluating the clinical value of I-CTCs. Therefore, the detection method utilized is based on specific antibody nano-microfluidic immunocapture combined with immunofluorescent staining. Following EMT, I-CTCs acquire the ability to resist external forces and evade immune surveillance[45], which facilitates tumor metastasis. Studies have found that CTCs can be detected in a patient’s blood at the early stages of tumor development and provide crucial evidence for their application in early diagnosis and disease monitoring[46]. In recent years, CTC testing has demonstrated significant value in early diagnosis, treatment monitoring, prognosis assessment, and the evaluation of recurrence and metastasis in solid tumors. In China, CTC testing has been incorporated into the breast cancer treatment guidelines[47] and the expert consensus for colorectal cancer treatment[48]. Furthermore, Helicobacter pylori (H. pylori) is closely associated with gastric carcinogenesis and metastasis[49]. The role of H. pylori in the EMT of tumor cells remains an intriguing topic. Ou et al[50] investigated the impact of H. pylori infection on the survival and migration of GC cells. Their study revealed that H. pylori infection enhances the migratory capacity of gastric malignancies, a process regulated by related genes. However, whether H. pylori infection is linked to the development of I-CTCs remains unexplored and represents a critical direction for future research. To date, no definitive conclusions have been reached.
In this study, the optimal cut-off value for I-CTCs was determined to be 5 based on ROC curve analysis, following a two-year follow-up of 59 patients with gastric adenocarcinoma. I-CTC positivity (I-CTC ≥ 5) was strongly correlated with the degree of tumor differentiation, while no significant associations were found with gender, age, TNM stage, depth of infiltration, lymph node metastasis, or tumor markers. A subgroup analysis of neoadjuvant chemotherapy efficacy in 16 patients showed that I-CTC positivity was significantly associated with disease progression and poor chemotherapy response. Survival curve analysis indicated that patients in the I-CTCs-negative group had longer OS and DFS than those in the I-CTCs-positive group. Cox regression analysis indicated that the degree of tumor differentiation and I-CTC count were associated with OS in patients with GC. I-CTC positivity was identified as an independent risk factor for the prognosis of patients with advanced GC. Yu et al[51] reported that the positive rate of I-CTCs was positively correlated with tumor infiltration depth and lymph node metastasis and that patients with low I-CTCs expression demonstrated a better response to chemotherapy. Similarly, Ning et al[52] analyzed the detection rates of CTCs in GC patients across different tumor stages and found that those with advanced-stage disease exhibited higher CTC levels. Our meta-analysis revealed that CTC-positive patients had more advanced TNM staging, poorer tumor differentiation, and a higher likelihood of early distant metastasis. GC patients with CTC-positive tumors may have a poorer prognosis compared with those with CTC-negative tumors[53]. However, our study found a strong correlation only between tumor differentiation and CTC positivity. The exclusion of stage IV GC patients and the limited sample size may have contributed to discrepancies between our findings and the meta-analysis results.
Positive I-CTCs were significantly associated with shorter survival in GC patients. In this study, after the follow-up period, the overall mortality rate among the 59 GC patients was 25.4%. However, in the I-CTCs-positive group, the mortality rate was as high as 42.4%. Ishiguro et al[54] found that high preoperative I-CTCs expression and increased postoperative levels were significantly associated with a poorer prognosis. These findings suggest that I-CTCs can serve as a prognostic marker for assessing survival risk and stratifying patients accordingly. Zhao et al[55] tested 32 GC patients for CTC and followed them for 30 months. The study found that patients with CTC ≥ 2 had a mortality rate of 66.7% and a worse prognosis compared with those with CTC < 2. Similarly, Wang et al[56] analyzed 242 GC patients via CTC testing and reported that the 3-year OS was 49.0% in CTC-positive patients compared with 72.5% in CTC-negative patients, which indicated a poorer prognosis for those with CTC positivity. The study also analyzed differences among the three types of CTCs in CTC-positive patients. Among I-CTCs, epithelial CTCs, mixed CTCs, and patients with I-CTCs-positive GC had the worst prognosis. Zhang et al[57] investigated the correlation between preoperative CTC counts and postoperative recurrence in GC patients and found that the 3-year DFS was significantly lower in patients with preoperative CTC ≥ 5 compared with those with CTC < 5. These findings strongly supported the association between CTCs and GC prognosis and highlighted their potential as a prognostic indicator.
There was also a significant correlation between CTCs and the efficacy of chemotherapy in patients with progressive GC. We found that the positive rate of I-CTCs in the PD group was as high as 83.3%, compared with only 20% in the DC group. Chen et al[58] investigated the relationship between CTCs and chemotherapy resistance in 114 GC patients and found that higher CTC counts were positively correlated with resistance to chemotherapy and poorer prognosis. Similarly, Lee et al[59] examined CTCs in 100 patients with metastatic GC and found that CTC presence was associated with a poor response to chemotherapy. These findings highlight the important role of I-CTCs in assessing chemotherapy efficacy. The present study confirmed that the expression level of I-CTCs was negatively correlated with chemotherapy efficacy and prognosis. We analyzed 16 patients with progressive GC who received neoadjuvant chemotherapy. The results showed that patients with I-CTC ≥ 5 before chemotherapy had a poorer prognosis and lower postoperative chemotherapy efficacy than those with I-CTC < 5. Our findings indicated that I-CTCs were an independent risk factor for GC patients and could serve as a valuable marker for prognosis and treatment efficacy assessment. Multifactorial analysis demonstrated that I-CTCs positivity is a strong prognostic factor; however, the wide confidence interval may reflect limitations such as a small sample size or residual confounding. Additionally, tumor burden, timing of I-CTC detection, and methodological variability in detection are potential contributing factors. We can stratify patients and predict the efficacy and prognosis of chemotherapy to guide the development of individualized treatment strategies via the detection of I-CTC expression levels for pre-treatment in patients with advanced GC. Although CTC is not yet recommended as a standard test in GC treatment guidelines, I-CTCs, as an emerging tumor marker, have the potential to play a crucial role in diagnosis, prognosis assessment, recurrence and metastasis monitoring, and chemotherapy resistance evaluation.
CONCLUSION
In patients with advanced GC, an I-CTC count of ≥ 5 is associated with both poor prognosis and reduced chemotherapy efficacy. I-CTCs may serve as a valuable preoperative biomarker for predicting the prognosis of advanced GC.
Footnotes
Provenance and peer review: Unsolicited article; Externally peer reviewed.
Peer-review model: Single blind
Specialty type: Oncology
Country of origin: China
Peer-review report’s classification
Scientific Quality: Grade A, Grade B, Grade C
Novelty: Grade A, Grade B, Grade B
Creativity or Innovation: Grade A, Grade B, Grade B
Scientific Significance: Grade A, Grade A, Grade B
P-Reviewer: Mousa NH; Zhang L S-Editor: Wei YF L-Editor: A P-Editor: Zhang XD
Papadaki MA, Messaritakis I, Fiste O, Souglakos J, Politaki E, Kotsakis A, Georgoulias V, Mavroudis D, Agelaki S. Assessment of the Efficacy and Clinical Utility of Different Circulating Tumor Cell (CTC) Detection Assays in Patients with Chemotherapy-Naïve Advanced or Metastatic Non-Small Cell Lung Cancer (NSCLC).Int J Mol Sci. 2021;22:925.
[RCA] [PubMed] [DOI] [Full Text] [Full Text (PDF)][Cited by in Crossref: 6][Cited by in RCA: 7][Article Influence: 1.8][Reference Citation Analysis (0)]
Marcuello M, Vymetalkova V, Neves RPL, Duran-Sanchon S, Vedeld HM, Tham E, van Dalum G, Flügen G, Garcia-Barberan V, Fijneman RJ, Castells A, Vodicka P, Lind GE, Stoecklein NH, Heitzer E, Gironella M. Circulating biomarkers for early detection and clinical management of colorectal cancer.Mol Aspects Med. 2019;69:107-122.
[RCA] [PubMed] [DOI] [Full Text][Cited by in Crossref: 127][Cited by in RCA: 186][Article Influence: 31.0][Reference Citation Analysis (0)]
Bulfoni M, Gerratana L, Del Ben F, Marzinotto S, Sorrentino M, Turetta M, Scoles G, Toffoletto B, Isola M, Beltrami CA, Di Loreto C, Beltrami AP, Puglisi F, Cesselli D. In patients with metastatic breast cancer the identification of circulating tumor cells in epithelial-to-mesenchymal transition is associated with a poor prognosis.Breast Cancer Res. 2016;18:30.
[RCA] [PubMed] [DOI] [Full Text] [Full Text (PDF)][Cited by in Crossref: 113][Cited by in RCA: 116][Article Influence: 12.9][Reference Citation Analysis (0)]
Hu ZY, Li XJ. [Frontier advances in the clinical application of circulating tumor cell technology].Shiyan Yu Jianyan Yixue. 2021;39:1-5+18.
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Bahnassy AA, Salem SE, Mohanad M, Abulezz NZ, Abdellateif MS, Hussein M, Zekri CAN, Zekri AN, Allahloubi NMA. Prognostic significance of circulating tumor cells (CTCs) in Egyptian non-metastatic colorectal cancer patients: A comparative study for four different techniques of detection (Flowcytometry, CellSearch, Quantitative Real-time PCR and Cytomorphology).Exp Mol Pathol. 2019;106:90-101.
[RCA] [PubMed] [DOI] [Full Text][Cited by in Crossref: 17][Cited by in RCA: 23][Article Influence: 3.3][Reference Citation Analysis (0)]
Nagrath S, Sequist LV, Maheswaran S, Bell DW, Irimia D, Ulkus L, Smith MR, Kwak EL, Digumarthy S, Muzikansky A, Ryan P, Balis UJ, Tompkins RG, Haber DA, Toner M. Isolation of rare circulating tumour cells in cancer patients by microchip technology.Nature. 2007;450:1235-1239.
[RCA] [PubMed] [DOI] [Full Text][Cited by in Crossref: 2735][Cited by in RCA: 2564][Article Influence: 150.8][Reference Citation Analysis (0)]
Orrapin S, Udomruk S, Lapisatepun W, Moonmuang S, Phanphaisarn A, Phinyo P, Pruksakorn D, Chaiyawat P. Clinical Implication of Circulating Tumor Cells Expressing Epithelial Mesenchymal Transition (EMT) and Cancer Stem Cell (CSC) Markers and Their Perspective in HCC: A Systematic Review.Cancers (Basel). 2022;14:3373.
[RCA] [PubMed] [DOI] [Full Text] [Full Text (PDF)][Cited by in RCA: 21][Reference Citation Analysis (0)]
Committee on Minimally Invasive Surgery; Chinese Research Hospital Association. [Circulating tumor cell assay Expert Consensus on the Application in Colorectal Cancer (2018)].Fuqiangjing Waike Zazhi. 2019;24:74-80.
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Ou L, Liu H, Peng C, Zou Y, Jia J, Li H, Feng Z, Zhang G, Yao M. Helicobacter pylori infection facilitates cell migration and potentially impact clinical outcomes in gastric cancer.Heliyon. 2024;10:e37046.
[RCA] [PubMed] [DOI] [Full Text] [Full Text (PDF)][Cited by in Crossref: 19][Reference Citation Analysis (0)]
Zhao L, Liu YH, Dong CH, Zhang WW, Duan XQ, Liu XZ, Chen L. [Difference of CTC detection in the prognosis of gastric cancer and colorectal cancer].J Xian Jiaotong University (Med Sci). 2020;41:263-267.
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Wang C, Liang WQ, Lu YX, Gao YH, Zhuang ZW, Wang LL, Xi HQ, Chen L. [The prognostic value of circulating tumor cells in patients undergoing gastric cancer surgery].Zhongguo Puwai Jichu Yu Linchuang Zazhi. 2022;29:17-23.
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