Sun YQ, Su J, Ye KN, Chen QX, Kang EZ, Lv CB, Lian MQ, Zeng WM, Zhang YB, Cai LS. High cyclin-dependent kinase 5 expression promotes tumor progression in gastric neuroendocrine carcinoma. World J Gastroenterol 2026; 32(26): 119574 [DOI: 10.3748/wjg.v32.i26.119574]
Corresponding Author of This Article
Li-Sheng Cai, Department of Gastrointestinal Surgery, Zhangzhou Affiliated Hospital of Fujian Medical University, No. 59 West Shengli Road, Zhangzhou 363000, Fujian Province, China. wxccls2024@163.com
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Gastroenterology & Hepatology
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research-article
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Sun YQ, Su J, Ye KN, Chen QX, Kang EZ, Lv CB, Lian MQ, Zeng WM, Zhang YB, Cai LS. High cyclin-dependent kinase 5 expression promotes tumor progression in gastric neuroendocrine carcinoma. World J Gastroenterol 2026; 32(26): 119574 [DOI: 10.3748/wjg.v32.i26.119574]
Yu-Qin Sun, Qiu-Xian Chen, Chen-Bin Lv, Ming-Qiao Lian, Wei-Ming Zeng, Yong-Bin Zhang, Li-Sheng Cai, Department of Gastrointestinal Surgery, Zhangzhou Affiliated Hospital of Fujian Medical University, Zhangzhou 363000, Fujian Province, China
Jie Su, Department of Ultrasonography, Zhangpu Hospital, Zhangzhou 363000, Fujian Province, China
Kai-Ning Ye, En-Zhou Kang, Department of Pathology, Zhangzhou Affiliated Hospital of Fujian Medical University, Zhangzhou 363000, Fujian Province, China
Author contributions: Sun YQ and Su J made equal contributions as co-first authors; Sun YQ, Su J, and Cai LS contributed to concept and design, drafting of the manuscript, and supervision; Ye KN, Chen QX, Kang EZ, Lv CB, Lian MQ, Zeng WM, and Zhang YB contributed to acquisition, analysis, or interpretation of data; Sun YQ, Su J, Zhang YB, and Zeng WM contributed to statistical analysis; Sun YQ, Su J, Ye KN, and Cai LS contributed to administrative, technical, or material support. All authors approved the final version to publish.
AI contribution statement: This article was translated using DeepL.
Supported by Natural Science Foundation of Fujian Province, No. 2025J011630; and the Climbing Fund of PhD Workstation, Zhangzhou Affiliated Hospital of Fujian Medical University, No. PDA202101.
Institutional review board statement: The study was approved by the Ethics Committee of the Zhangzhou Affiliated Hospital of Fujian Medical University, No. 2026LWB036.
Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article.
Data sharing statement: The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.
Corresponding author: Li-Sheng Cai, Department of Gastrointestinal Surgery, Zhangzhou Affiliated Hospital of Fujian Medical University, No. 59 West Shengli Road, Zhangzhou 363000, Fujian Province, China. wxccls2024@163.com
Received: February 2, 2026 Revised: February 23, 2026 Accepted: March 23, 2026 Published online: July 14, 2026 Processing time: 151 Days and 6.1 Hours
Abstract
BACKGROUND
The function of cyclin-dependent kinase 5 (CDK5) in gastric neuroendocrine carcinoma (GNEC) is not yet fully understood.
AIM
To investigate the expression and function of CDK5 in GNEC.
METHODS
Immunohistochemistry was performed to measure CDK5, programmed death ligand-1, human epidermal growth factor receptor 2 (HER2), DNA mismatch repair, and Ki-67 antigen levels in paraffin-embedded samples. Epstein-Barr virus was detected via in situ hybridization, and HER2 was further analyzed with fluorescence in situ hybridization. The association between CDK5 expression, clinicopathological features, and patient outcomes was analyzed using univariate and multivariate statistics. CDK5’s role in GNEC was investigated through cell proliferation and colony formation assays.
RESULTS
This study included 65 patients diagnosed with GNEC, of whom 39 (60.0%) demonstrated elevated CDK5 expression and 26 (40.0%) exhibited reduced CDK5 expression. The observed expression pattern significantly correlated with tumor size and tumor-node-metastasis staging. Survival analyses indicated that patients with elevated CDK5 expression had lower overall survival and disease-free survival rates. Multivariate analysis further identified CDK5 as an independent prognostic risk factor for GNEC (hazard ratio = 2.13, 95% confidence interval: 1.03-4.41, P = 0.041). Additional analyses revealed a correlation between high CDK5 expression and increased Ki-67 indices, although no significant associations were found with programmed death ligand-1, HER2, Epstein-Barr virus, or mismatch repair status. Elevated CDK5 expression was also detected in GNEC cell lines ECC10 and ECC12. The knockdown of CDK5 significantly reduced the clonogenic and proliferative capacity of GNEC cells, whereas CDK5 overexpression did not significantly affect clonogenicity or cell proliferation.
CONCLUSION
These findings indicate that CDK5 is an independent prognostic risk factor and may play a crucial role in the proliferation of GNEC cells.
Core Tip: This study aims to investigate the expression and function of cyclin-dependent kinase 5 (CDK5) in gastric neuroendocrine carcinoma (GNEC). The findings indicate that patients with elevated CDK5 expression in GNEC have a poor prognosis. Furthermore, CDK5 is identified as an independent prognostic risk factor and may play a critical role in the proliferation of GNEC.
Citation: Sun YQ, Su J, Ye KN, Chen QX, Kang EZ, Lv CB, Lian MQ, Zeng WM, Zhang YB, Cai LS. High cyclin-dependent kinase 5 expression promotes tumor progression in gastric neuroendocrine carcinoma. World J Gastroenterol 2026; 32(26): 119574
Gastric cancer is the fifth most common malignancy worldwide in terms of both incidence and cancer-related mortality[1]. Unlike gastric adenocarcinoma, gastric neuroendocrine carcinoma (GNEC) constitutes a distinct pathological subtype of gastric cancer originating from neuroendocrine cells within the gastric mucosa[2]. This subtype exhibits a different biological behavior compared to adenocarcinoma, with a higher propensity for metastasis to the perigastric lymph nodes and liver, and demonstrates reduced sensitivity to chemotherapy, often leading to a poorer prognosis[3]. Therefore, there is an urgent need to elucidate the mechanisms underlying the initiation and progression of GNEC and identify the key factors influencing its advancement. The aim of this study was to identify novel therapeutic targets for the treatment of this disease.
Cyclin-dependent kinase 5 (CDK5) is a member of the cyclin-dependent kinase family. Unlike other cyclin-dependent kinases, CDK5 possesses unique binding partners and regulators that control various cellular functions. CDK5 was initially shown to play a critical role in the progression of neurodegenerative diseases[4]. Recent evidence suggests that CDK5 is a pivotal regulator of various oncogenic processes such as tumorigenesis, metastasis, and angiogenesis. Aberrant expression of CDK5 and its activators, through amplification, messenger RNA upregulation, or both, has been observed in numerous human cancers, including colorectal[5], breast[6], lung[7], prostate[8], and pancreatic cancers[9], and melanoma[10]. Emerging studies have proposed that the role of CDK5 as either an oncogene or a tumor suppressor is contingent on its subcellular localization[11]. Previous studies have indicated that CDK5 may act as a tumor suppressor in gastric adenocarcinoma and that its activity is intricately associated with cellular localization[12]. In thyroid[13] and prostate neuroendocrine carcinomas (NECs)[14], activated CDK5 facilitates tumor progression by targeting the Rb1 gene. Nonetheless, the role of CDK5 in GNEC remains unclear. Consequently, this study aimed to elucidate the function and clinical significance of CDK5 in GNEC by analyzing clinical tissue samples, follow-up data, and cellular experiments.
MATERIALS AND METHODS
Patients and GNEC tissues
Clinicopathological data and tumor tissue specimens were collected from 65 patients with GNEC who were admitted to the Zhangzhou Affiliated Hospital of Fujian Medical University between January, 2013, and October, 2021. The diagnosis of GNEC was confirmed by an experienced pathologist according to the 2019 World Health Organization classification of tumors of the digestive system[15]. Pathological tumor-node-metastasis (pTNM) staging was reassessed based on the eighth edition of the American Joint Committee on Cancer staging manual[16,17]. The inclusion criteria were: (1) Pathologically confirmed GNEC; and (2) Having undergone radical gastrectomy with complete clinical and follow-up data. The exclusion criteria were: (1) Neuroendocrine tumors G1, G2, and G3 or mixed neuroendocrine–non-neuroendocrine neoplasms; (2) Preoperative anticancer therapy; and (3) Concurrent or prior history of other malignancies. Patients with advanced disease received postoperative adjuvant chemotherapy comprising etoposide and cisplatin. The study was approved by the Ethics Committee of the Zhangzhou Affiliated Hospital of Fujian Medical University, No. 2026LWB036, and written informed consent was obtained from all participating patients.
Cell culture
GNEC cell lines ECC10 and ECC12 were purchased from the RIKEN BRC Cell Bank (Japan)[18]. These cells, along with gastric adenocarcinoma cell lines MKN-74, HGC-27, NUGC-3, and MGC-803, were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (Thermo Fisher Scientific, MA, United States). An AGS cell line was maintained in DMEM/F12 medium containing 10% fetal bovine serum. All cells were incubated at 37 °C in a humidified atmosphere containing 5% CO2.
Immunohistochemistry analysis
Formalin-fixed, paraffin-embedded tumor tissues were sectioned at 4 μm and mounted on silane-coated slides. After deparaffinization in xylene and rehydration through a graded ethanol series, antigen retrieval was performed by autoclaving in sodium citrate buffer (pH 6.0) at 121 °C. Endogenous peroxidase activity was blocked with 3% H2O2, and nonspecific binding was reduced by incubation with 10% normal goat serum. The sections were then incubated overnight at 4 °C with the following primary antibodies: CDK5 (cat. no. 2506; 1:150, Cell Signaling Technology, Inc., Danvers, MA, United States), Ki-67 (cat. no. QCS-PM-1126; 1:150), MLH1 (cat. no. QCS-PM-1181; 1:100), MSH2 (cat. no. QCS-PM-1137; 1:100), MSH6 (cat. no. QCS-PM-1011; 1:100), PMS2 (cat. no. QCS-PM-1107; 1:100), human epidermal growth factor receptor 2 (HER2, cat. no. N12091; 1:150; Roche; Switzerland), and programmed death ligand-1 (PD-L1, cat. no. 2506; 22C3 Dako, Denmark; 1:100). After washing, a horseradish peroxidase-conjugated secondary antibody was applied, and immunoreactivity was visualized with 3,3′-diaminobenzidine. The sections were counterstained with hematoxylin, dehydrated, and mounted. Negative controls were prepared by replacing the primary antibody with antibody diluent.
Two independent pathologists, blinded to the clinical data, evaluated the stained sections. The expression of CDK5 in GNEC did not demonstrate intratumoral heterogeneity, consistent with previous studies on gastric adenocarcinoma[19]. The scoring method and criteria used in this study were based on those established for gastric adenocarcinoma in earlier research[20]. CDK5 expression was observed as yellowish to brown staining in the cytoplasm and/or nuclei of tumor cells. The staining intensity was scored as: 0 (none), 1 (weak, light yellow), 2 (moderate, yellow-brown), or 3 (strong, brown). CDK5 expression was categorized as low (score ≤ 1) or high (score ≥ 2). DNA mismatch repair (MMR) status was assessed by evaluating the expression of MLH1, PMS2, MSH2, and MSH6. Loss of nuclear expression of one or both proteins in a functional pair (MLH1/PMS2 or MSH2/MSH6) with appropriate internal positive controls was defined as MMR deficiency (dMMR). PD-L1 expression was assessed using the combined positive score (CPS)[21]. For HER2 immunohistochemistry (IHC) scoring, the 4-tiered Hoffmann scoring criteria were used. When complete or basolateral membranous reactivity was observed in > 10% of tumor cells, scores of 1+, 2+, and 3+ were assigned according to the intensity of membranous reactivity (faint, moderate, or intense, respectively). Patients with a score of 3+ were classified as positive.
In situ hybridization for Epstein-Barr virus-encoded RNA
Epstein-Barr virus (EBV) status was detected by in situ hybridization of EBV-encoded small RNA (EBER) in formalin-fixed, paraffin-embedded tissue sections. The assay was performed on a BenchMark ULTRA automated staining system (Ventana Medical Systems, AZ, United States) using fluorescein-conjugated EBER probes (Ventana Medical Systems, AZ, United States) according to the manufacturer’s protocol, which included enzymatic digestion with ISH protease 3. Signals were detected using the iViewBlue detection kit. In each experiment, EBV-positive tonsil tissue was used as a positive control and EBV-negative tissue as a negative control. Nuclear staining in tumor cells was interpreted as positive for EBER.
Fluorescence in situ hybridization
To evaluate the HER2 status, IHC scores of 2+ (equivocal) were further analyzed using fluorescence in situ hybridization. The PathVysion HER-2 DNA Probe Kit (Abbott Molecular, IL, United States) was used according to the manufacturer’s instructions. HER2 gene amplification was defined as a HER2/CEP17 ratio ≥ 2.0. Those with IHC 2+ and fluorescence in situ hybridization-positive results were classified as positive, whereas all others were classified as negative.
Western blot analysis
Cells were seeded into 60-mm dishes at a concentration of 5 × 104 cells/well (20%-30% confluence) and cultured to 80% confluence. The cells were then scraped and lysed in radioimmunoprecipitation assay buffer containing protease and phosphatase inhibitors. Lysates were centrifuged at 10000 × g for 10 minutes at 4 °C. Protein concentrations were determined using the bicinchoninic acid Protein Assay kit (Thermo Fisher Scientific, MA, United States). A total of 40 μg of protein from each sample was denatured, loaded into each well, separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and transferred to a polyvinylidene difluoride membrane (EMD Millipore, Billerica, MA, United States). The membranes were blocked with 5% non-fat milk in Tris-buffered saline with 0.1% Tween-20 (TBST) at room temperature for 1 hour, and then incubated overnight at 4 °C with primary antibodies against CDK5 (1:1000) and glyceraldehyde-3-phosphate dehydrogenase (1:5000; as a loading control) in TBST. After washing with TBST, the membranes were incubated for 1 hour at room temperature with the corresponding horseradish peroxidase-conjugated secondary antibody. Protein bands were visualized using an enhanced chemiluminescence detection system.
In vitro cell function experiments
Stable cell lines with CDK5 knockdown and overexpression were established according to previously described methodology[12]. Colony formation assays were conducted to evaluate the cells’ capacity to form colonies. Cell viability and proliferation were assessed using cell counting kit-8 (CCK-8; Dojindo Molecular Technologies, Inc., Kumamoto, Japan) and sulforhodamine B (SRB) Assay (Sigma-Aldrich, MO, United States; Merck KGaA, Germany; catalog no. S9012). In brief, cells were seeded at a density of 1000 cells per well in 96-well tissue culture plates and incubated at 37 °C with 5% CO2 in a humidified incubator for 24, 72, 120, or 168 hours. Post-treatment and incubation, 20 μL of CCK-8 was added to each well, followed by an additional 3-hour incubation. Absorbance was measured using a Bio-Rad ELISA plate reader (Biotek Instruments, Inc., Winooski, VT, United States) at a wavelength of 450 nm. Data from at least three independent experiments were analyzed in duplicate. The relative cell proliferation ratios were plotted against the untreated controls, which were set as the 100% activity baseline. For the SRB colorimetric assay, cells were seeded at 1000 cells per well and allowed to adhere for 7 days. After 24, 72, 120, or 168 hours, cells were harvested and processed according to the SRB protocol as described previously[22].
Statistical analysis
Statistical analyses were performed using the IBM SPSS Statistics software (version 26.0; IBM Corp., NY, United States). Categorical variables were evaluated using the χ2 test or Fisher’s exact test, whereas continuous variables were examined using either the Student’s t-test or the Mann-Whitney U test, contingent upon their distribution. Survival analysis was conducted using the log-rank test, and the determinants of tumor recurrence were investigated using both univariate and multivariate Cox regression analyses. Statistical significance was established at a two-sided P < 0.05.
RESULTS
Patient characteristics
Based on the inclusion and exclusion criteria, a total of 65 patients diagnosed with GNEC were included in this study. The clinicopathological data of all the enrolled patients are presented in Supplementary Table 1. The mean age of the patients was 63.2 ± 9.3 years. Of the participants, 48 (73.8%) were male and 17 (26.2%) were female. All patients underwent radical gastrectomy, with 46 patients (70.8%) undergoing total gastrectomy and 19 (29.2%) undergoing partial gastrectomy. The mean number of dissected lymph nodes was 32.9 ± 11.7, and the mean number of metastatic lymph nodes was 4.2 ± 4.9. Postoperative pTNM staging revealed that 7 patients (10.8%) were classified as stage I, 23 (35.4%) as stage II, and 35 (53.8%) as stage III GNEC. Additionally, 49 of the cases (75.4%) were identified as having large cell NEC, while the rest (16 cases; 24.6%) were identified as having small cell NEC.
Expression of CDK5 in GNEC and its correlation with clinicopathologic parameters
In this study, the expression of CDK5 in GNEC was assessed by IHC. In the patient cohort, 39 individuals exhibited high CDK5 expression, while the rest (26 individuals) exhibited low CDK5 expression. Notably, CDK5 was predominantly localized in the cytoplasm and nucleus of tumor cells (Figure 1). Further analysis of the correlation between CDK5 expression and clinicopathological parameters revealed significant associations between CDK5 expression and clinical T stage, clinical N stage, tumor length, metastatic lymph node involvement, pathological T stage, pathological N stage, and pTNM stage. Conversely, no significant correlations were observed between CDK5 expression and sex, age, body mass index, tumor location, tumor resection status, number of dissected lymph nodes, nerve invasion, vascular invasion, or cell type (Table 1).
High expression of CDK5 is associated with a poor prognosis
The median follow-up time for all patients was 54 months. Survival analysis demonstrated that patients in the high CDK5 expression group exhibited significantly lower overall survival rates than those in the low CDK5 expression group at 1 year (96.2% vs 82.1%), 3 years (73.1% vs 37.2%), and 5 years (60.3% vs 12.6%). Additionally, patients in the high CDK5 expression group had significantly lower disease-free survival rates at 1 year (88.5% vs 79.5%), 3 years (64.9% vs 30.8%), and 5 years (60.3% vs 12.3%). These differences were statistically significant (overall survival: P = 0.004; disease-free survival: P = 0.002; Figure 2). Univariate and multivariate Cox regression analyses identified elevated CDK5 expression as an independent prognostic factor associated with poor outcomes in patients with GNEC (hazard ratio = 2.13, 95% confidence interval: 1.03-4.41, P = 0.041; Table 2).
Figure 2 Overall survival and disease-free survival of different cyclin-dependent kinase 5 expression.
A: Kaplan-Meier curves for overall survival; B: Kaplan-Meier curves for disease-free survival; C: Tabular summary of overall survival and disease-free survival.
Table 2 Univariable and multivariable Cox regression analyses of risk factors for overall survival in patients with gastric neuroendocrine carcinoma.
CDK5 expression is positively correlated with the expression of Ki-67
The expression levels of Ki-67, PD-L1, HER2, EBV, and MMR were examined in GNEC tissues. The findings revealed that 48 patients (73.8%) exhibited a Ki-67 expression greater than 50%, 23 (35.4%) had a PD-L1 CPS > 1, 1 (1.5%) was positive for HER2 expression, 3 (4.6%) tested positive for EBV, and 2 (3.1%) were identified as having dMMR. Analysis of the correlation between CDK5 expression and the molecular phenotype of NEC demonstrated a significant association between CDK5 and Ki-67 expression (P = 0.016), with 68.8% of patients exhibiting Ki-67 > 50% also showing elevated CDK5 expression. Conversely, no significant correlation was observed between CDK5 expression and the expression status of PD-L1, HER2, EBV, or MMR. The initial pathological molecular characteristics are shown in Figure 3.
Figure 3 The relationship between cyclin-dependent kinase 5 expression and the markers Ki-67, programmed death ligand-1, human epidermal growth factor receptor 2, Epstein-Barr virus, and mismatch repair status.
A: Representative images; B: Oncoprint depicting survival status, pathological tumor-node-metastasis stage, and molecular marker profiles across individual cases; C-G: Bar graphs illustrating the distribution of cyclin-dependent kinase 5-low and high groups. PD-L1: Programmed death ligand-1; CPS: Combined positive score; HER2: Human epidermal growth factor receptor 2; EBV: Epstein-Barr virus; MMR: Mismatch repair; pTNM: Pathological tumor-node-metastasis; CDK5: Cyclin-dependent kinase 5.
Downregulation of CDK5 in GNEC cells inhibits both cellular proliferation and colony formation
To further elucidate the expression and functional role of CDK5 in GNEC cell lines, ECC10 and ECC12 cells were used in this study (Supplementary Figure 1). Western blot analysis demonstrated that ECC10 and ECC12 cells exhibit significantly elevated levels of CDK5 expression compared to gastric adenocarcinoma cell lines MKN-74, HGC-27, AGS, NUGC-3, and MGC-803 (Figure 4A). To investigate the effect of CDK5 on GNEC progression, ECC10 and ECC12 cell lines with stable CDK5 knockdown as well as control cells were established (Figure 4B). Subsequently, treatment with the CDK5 inhibitor roscovitine significantly inhibited the proliferation of GNEC cells (Figure 4C-E). Similarly, CCK-8 and SRB assays demonstrated a significant reduction in cell proliferation (Figure 4D and E) and colony formation (Figure 4F and G) following CDK5 knockdown. These findings indicate that, in vitro, CDK5 downregulation suppresses the proliferation and colony formation of ECC10 and ECC12 cells.
Figure 4 Downregulation of cyclin-dependent kinase 5 in gastric neuroendocrine carcinoma cells inhibits both cellular proliferation and colony formation.
A: The expression levels of cyclin-dependent kinase 5 (CDK5) were analyzed in gastric neuroendocrine carcinoma and gastric cancer cell lines; B: ECC10 and ECC12 cell lines with stable CDK5 knockdown, alongside control cell lines, were established; C-E: The proliferation rates of cells with shCDK5 knockdown or treated with the CDK5 inhibitor roscovitine, as well as control cells, were evaluated using cell counting kit-8 and sulforhodamine B assays; F and G: Colony formation assays were conducted to evaluate the colony-forming capabilities of shCDK5 knockdown and control cells. aP < 0.05. GAPDH: Glyceraldehyde-3-phosphate dehydrogenase; CCK8: Cell count kit-8; SRB: Sulforhodamine B; CDK5: Cyclin-dependent kinase 5; NS: Not significant.
Overexpression of CDK5 in GNEC cells does not inhibit proliferation or colony formation
To investigate the role of CDK5 overexpression in GNEC cells, we initially overexpressed EGFP-CDK5 and p-TRE-EGFP in the ECC10 and ECC12 GNEC cell lines (Figure 5A and B). In vitro analyses revealed no significant differences in cell proliferation (Figure 5C) or colony formation (Figure 5D) between the CDK5-overexpressing cells and control cells.
Figure 5 Overexpression of cyclin-dependent kinase 5 in gastric neuroendocrine carcinoma cells does not inhibit proliferation or colony formation.
A and B: ECC12 cell lines with stable cyclin-dependent kinase 5 (CDK5) overexpression and control cell lines were established; C: The proliferation rates of cells with CDK5 overexpression and control cells, were evaluated using CCK-8 assays; D: Colony formation assays were conducted to evaluate the colony-forming capabilities of CDK5 overexpression and control cells. CDK5: Cyclin-dependent kinase 5; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase; NS: Not significant.
DISCUSSION
GNEC demonstrates distinct biological behavior and molecular characteristics compared to gastric adenocarcinomas. This study identified that CDK5 is markedly overexpressed in GNEC, with elevated CDK5 expression correlating with a poorer patient prognosis. Furthermore, CDK5 expression has been established as an independent prognostic risk factor for patients with GNEC. Analysis of tissue samples revealed that elevated CDK5 expression is associated with increased Ki-67 indices, but does not exhibit a significant relationship with PD-L1, HER2, EBV, or MMR status. Similarly, high levels of CDK5 expression were observed in GNEC cell lines ECC10 and ECC12. The knockdown of CDK5 significantly inhibited the clonogenic and proliferative capacity of GNEC cells, whereas CDK5 overexpression did not alter these cellular functions.
CDK5 is expressed in various tumors and is associated with tumor prognosis. In tongue squamous cell carcinoma, CDK5 modulates microRNA 513c-5p, thereby promoting tumor proliferation and correlating with poor prognosis[23]. Similarly, elevated CDK5 expression has been linked to adverse prognostic outcomes in several malignancies, including salivary gland carcinoma[24], hepatocellular carcinoma[25], and prostate cancer[25]. Our previous study indicated a potential tumor-suppressive role of CDK5 in gastric adenocarcinoma, where patients with elevated CDK5 expression demonstrated improved prognoses[20]. This phenomenon may be related to the subcellular localization of CDK5 in gastric cancer. The current study revealed that CDK5 was highly expressed in the majority of patients with GNEC. Nevertheless, individuals with elevated CDK5 expression exhibited higher rates of lymph node metastasis, advanced TNM staging, and, consequently, poorer prognosis, establishing CDK5 as an independent prognostic risk factor for GNEC. These findings suggest a possible oncogenic role of CDK5 in GNEC, although the precise mechanisms underlying this role remain to be elucidated.
In the context of gastric cancer, molecular subtyping plays a crucial role in guiding treatment decisions with key clinical markers, including PD-L1, HER2, EBV, and MMR status. This study found that approximately 35.4% of patients with GNEC demonstrated PD-L1 expression with CPS > 1. HER2 positivity was observed in 1.5% of the patients, while 3.1% exhibited dMMR, which is a slightly lower prevalence than that in patients with gastric adenocarcinoma. EBV positivity was noted in 4.6% of the cases, a proportion similar to that found in gastric adenocarcinoma. Immunotherapy has become a pivotal strategy in cancer treatment, involving multiple molecules and signaling pathways in the antitumor immune response[26]. CDK5 has been implicated in the regulation of PD-L1 expression and immune evasion[27]. CDK5 disrupts PD-L1 stability through autophagy mediated by chaperone molecules. Targeting CDK5 in conjunction with PD-1 blockade has been shown to synergistically inhibit HCC growth[28]. Nonetheless, this study did not establish a significant association between CDK5 and PD-L1 expression in tissue samples. Further investigation is warranted to determine whether the relatively lower PD-L1 expression levels in GNEC compared with those in gastric adenocarcinoma are linked to elevated CDK5 expression. Additionally, we found that patients with GNEC and high CDK5 expression exhibited increased Ki-67 indices, indicating a positive correlation between CDK5 and Ki-67 expression levels and suggesting that CDK5 may be associated with tumor cell proliferation and stemness. The CDK5-SOX2 signaling axis regulates the self-renewal of glioblastoma stem cells[29]. Furthermore, activation of the Wnt and nuclear factor-κB (NF-κB) signaling pathways is crucial for maintaining tumor cell stemness[30,31]. In colorectal cancer, HOXC8 enhances tumor cell stemness by activating the TRIM22/NF-κB pathway[32]. CDK5 is involved in various biological processes by modulating the Wnt and NF-κB pathways[33,34], suggesting a potential role in regulating stemness in GNEC through these pathways. Mutations in the TP53, APC, and PIK3CA genes are commonly observed during the progression of colorectal cancer[35], and extensive research has identified CDK5 as playing an oncogenic role in this disease[36]. The potential involvement of CDK5 in gene mutation processes has emerged as an important area of research.
CDK5 has been implicated in the development and progression of various tumors. Recent investigations have highlighted the critical role of the LIM kinase 1/CDK5-Wnt/β-catenin signaling pathway in the metastasis of esophageal squamous cell carcinoma, where elevated expression levels of LIM kinase 1/CDK5 are associated with poorer patient prognosis[37]. In medullary thyroid carcinoma, the RET proto-oncogene modulates tumor proliferation by activating CDK5 and signal transducer and activator of transcription 3[38]. Similarly, in hepatocellular carcinoma, the CDK5-PRMT1-WDR24 pathway serves as a critical driver of tumor proliferation[39]. The present study demonstrated that silencing CDK5 expression in GNEC cell lines, such as ECC10 and ECC12, markedly inhibited cell proliferation and clonogenic capacity, whereas CDK5 overexpression did not enhance these properties. This phenomenon may be attributed to the saturation of CDK5 expression in GNEC, rendering further increases ineffective. Alternatively, CDK5 activation may require ligand binding, suggesting that increased expression alone is insufficient to modify its functional state. In contrast, previous studies have suggested that CDK5 functions as a tumor suppressor in gastric adenocarcinoma, and its role is potentially influenced by its nuclear-cytoplasmic localization[12]. CDK5 exhibits diverse functions across different tumor types within the same organ, possibly reflecting their distinct cellular origins. Gastric adenocarcinoma cells are derived from the gastric glandular epithelium, whereas GNEC cells are thought to originate from chromaffin cells within the gastric epithelium. Unlike other cyclin-dependent kinases, CDK5 has unique functions that depend on binding to different activators, thereby influencing various downstream molecules.
Nonetheless, the oncogenic role of CDK5 suggests its potential as a therapeutic target. In colorectal cancer, studies have shown that the small-molecule peptide TP5 selectively inhibits aberrantly activated CDK5/p25 complexes while preserving the physiological function of CDK5/p35. This inhibition reduces cell viability and clonogenicity, induces apoptosis, increases DNA damage, and leads to G1-phase cell cycle arrest, thereby exerting antitumor effects[40]. Currently, there are limited effective targeted therapies for GNEC. Advancing our understanding of the oncogenic mechanisms of CDK5 will aid in the development of antitumor vaccines and targeted therapies[41,42]. In neuroendocrine cancers, the CDK5-Rb1-E2F1 signaling pathway is essential for the development of medullary thyroid and prostate carcinomas[13]. Furthermore, research on gastric adenocarcinoma suggests that CDK5 facilitates apoptosis and decreases chemotherapy resistance by modulating the E2F1 signaling pathway[43]. The role of CDK5 in promoting proliferation in GNEC via the CDK5-Rb1-E2F1 signaling pathway or through CDK5-associated cellular mechanisms requires further investigation.
This study has some limitations. First, the patient cohort was derived from a single center; however, considering the rarity of GNEC, it constitutes a relatively large single-center cohort. Second, the analyzed tissue samples were collected over an extended period; nonetheless, all pathological diagnoses were reassessed by pathologists in accordance with the latest standards, ensuring diagnostic and staging accuracy. Lastly, although this study identified a pivotal role for CDK5 in the proliferation of GNEC, the detailed molecular mechanisms remain to be fully elucidated. Future research utilizing more sophisticated models, such as patient-derived organoids, will be instrumental in unraveling the specific pathways involved.
CONCLUSION
In conclusion, this study demonstrated that high CDK5 expression in GNEC is associated with a poorer prognosis and that CDK5 expression serves as an independent risk factor affecting the prognosis of patients with GNEC. In vitro experiments suggested that CDK5 may be a critical factor in GNEC cell proliferation.
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