Clinicopathological features and prognoses of HER‑2‑positive gastric cancer

Introduction

Gastric cancer is a notable contributor to cancer-related morbidity and mortality worldwide, ranking fifth in terms of incidence and fourth in terms of mortality among all malignancies (1) Patients with gastric cancer in China represent over one-half of the worldwide total. Recent national data from 2022 reveal that gastric cancer has the third highest incidence and mortality rates among all cancer types in the country (2). Despite advancements in diagnosis and treatment, the overall survival (OS) rate remains low, with a 5-year survival rate <50%, dropping to <30% for advanced stages of the disease. Although trastuzumab has improved survival in human epidermal growth factor receptor-2 (HER-2)-positive disease and established anti-HER-2 therapy as a standard of care (3,4), overall outcomes remain poor. HER-2-positive tumors comprise 12-20% of gastric cancer cases (5) and represent a distinct therapeutic entity defined by the efficacy of anti-HER-2 agents. Although HER-2 upregulation portends inferior survival and heightened metastatic potential relative to HER-2-negative disease, existing evidence largely derives from dichotomous comparisons between these subgroups (6-9). Although research on the specific subtype of disease with low HER-2 expression in gastric cancer has increased (10,11), few studies have further differentiated this subgroup of HER-2-positive gastric cancer, leading to a limited understanding of whether variations in clinical characteristics and prognosis exist among patients with HER-2-positive gastric cancer with differing levels of HER-2 protein expression. Clinical data comparing immunohistochemistry (IHC) 2+ and 3+ HER-2-positive tumors, scored according to the Rüschoff/Hofmann grading system (12,13), remain limited. The present study stratified HER-2-positive gastric cancer by protein expression to evaluate its impact on clinicopathological features and survival.

Patients and methods

Patient enrollment and data collection

The present retrospective study examines clinical data from patients with HER-2-positive gastric cancer who underwent a radical gastrectomy at the Zhangzhou Affiliated Hospital of Fujian Medical University (Zhangzhou, China) between March 2017 and March 2023. All patients were of Asian ethnicity and originated from Zhangzhou, Fujian, China. Inclusion criteria necessitated patients to meet the following conditions: Preoperative clinical and imaging assessments indicating stage I-III gastric cancer according to the 8th edition of the American Joint Committee on Cancer Tumor-Node-Metastasis (TNM) system (14); successful radical resection (D2 lymphadenectomy and negative resection margins); postoperative pathological confirmation of gastric adenocarcinoma (International Classification of Diseases for Oncology no. C16 M8140/3); HER-2 status determined as IHC 3+ or 2+, with positive fluorescence in situ hybridization (FISH) in postoperative pathological specimens; and availability of comprehensive medical records. Exclusion criteria encompassed distant organ metastasis, concurrent malignancies, residual gastric cancer, incomplete medical documentation, or perioperative mortality attributed to surgical complications such as bleeding, septic shock or multiple organ failure. The study was approved by the Institutional Ethics Review Board of Zhangzhou Affiliated Hospital of Fujian Medical University (approval no. 2025LWB426).

Treatment strategy

All patients were treated according to clinical TNM staging. cT1-3N0-1M0 cases proceeded directly to radical gastrectomy, whereas cT3-4N1-3M0 cases received 2-4 cycles of neoadjuvant chemotherapy (fluoropyrimidine- or taxane-based plus platinum: FLOT, FOLFOX, XELOX/SOX or taxane-cisplatin) before surgery. Doses were as follows: Docetaxel, 50 mg/m2; oxaliplatin, 85-130 mg/m2; 5-FU, 2,400-2,600 mg/m2; capecitabine, 1,000 mg/m2 BID; S-1, 40 mg/m2 BID; cisplatin, 70-75 mg/m2; all on day 1, every 2-3 weeks. Adjuvant fluoropyrimidine-based chemotherapy was given to patients with pII (pT1-2N3M0) or pIII (pT3-4aN1-3M0) disease. Perioperative trastuzumab was not routinely used, in line with National Comprehensive Cancer Network, European Society for Medical Oncology, Chinese Society of Clinical Oncology and Japanese Gastric Cancer Association guidelines (15-18).

Detection of HER-2 expression

The expression of HER-2 protein in gastric cancer tissues was assessed using the Elivision two-step immunohistochemical method. The HER-2/neu rabbit monoclonal antibody (4B5; catalog number 790-2991; prediluted) from Roche Diagnostics (Shanghai) Co., Ltd., was applied on a Roche BenchmarkXT automated IHC platform. HER-2 IHC scoring was independently performed by two experienced gastrointestinal pathologists who were blinded to clinical outcomes. Discrepant or equivocal cases were resolved by consensus review using a multi-headed microscope. Immunostaining of HER-2 was categorized as 0, 1+, 2+ or 3+ utilizing the Rüschoff/Hofmann grading system (12,13). Tumor tissues with a score of 2+ underwent fluorescence in situ hybridization (FISH) analysis to confirm that they were indeed HER-2+. Scores of 0 were designated as HER-2-negative; 1+ and FISH-non-amplified 2+ cases were classified as low expression; and 3+ and FISH-amplified 2+ cases were considered HER-2-positive. FISH was performed on 4-µm FFPE sections using the Abbott PathVysion HER-2 DNA Probe Kit. Specimens were fixed in 10% neutral buffered formalin at room temperature for 24-48 h before processing and embedding. Deparaffinized sections were post-fixed in 10% neutral buffered formalin for 10 min at room temperature, then processed per the manufacturer's protocol (pretreatment at 80˚C for 30 min, protease digestion at 37˚C for 10 min, overnight hybridization at 37˚C with HER-2/neu and CEP17 probes). Tumor cells were identified on the corresponding H&E sections. At least 20 consecutive nuclei were scored, and the HER-2/CEP17 ratio was calculated. A HER-2/CEP17 ratio ≥2 indicated a HER-2-positive result, while a ratio <1.8 indicated a HER-2-negative result. Samples with ratios between 1.8 and 2 necessitated the evaluation of an additional 20-40 tumor cells, with a final ratio <1.8 classified as HER-2-negative and ≥2 as HER-2-positive. For cases between 1.8 and 2, a further 80-160 nuclei were counted, and the assessment was independently repeated by at least two observers to reach a consensus.

Statistical analysis

The associations between HER-2 protein expression and clinical characteristics were assessed through χ2 testing. DFS and OS were analyzed using Kaplan-Meier curves with log-rank testing for between-group comparisons, and univariate and multivariate Cox regression analyses were performed to identify the associations between clinicopathological features and survival outcomes. All statistical analyses were conducted using SPSS 18.0 software (SPSS, Inc.), with P<0.05 used to indicate a statistically significant difference.

Results

Characteristics of the overall population

A total of 99 patients with HER-2-positive gastric cancer were included in the present study (Table I). The age of participants ranged from 32 to 86 years, with a median age of 68 years. The majority of individuals (58.6%) were aged ≥65 years, and the cohort was predominantly male (75.8%). The cohort primarily consisted of patients with intestinal-type gastric cancer (79.8%), well-differentiated tumors (57.6%), T3-4 infiltration depth (73.7%), stage III disease (59.6%), tumor size <5 cm (60.6%), lymph node metastasis (60.6%), LVI (76.8%), and PNI (75.8%). All patients exhibited proficient mismatch repair (pMMR), and 20 patients (20.2%) underwent neoadjuvant therapy. As of the last follow-up in February 2025, recurrence was observed in 47 patients (47.5%), and 33 patients (33.3%) passed away.

Table I Clinical characteristics of patients with gastric cancer.

Table I

Clinical characteristics of patients with gastric cancer.

Characteristics	n (%)
Total	99 (100.0)
Sex
Male	75 (75.8)
Female	24 (24.2)
Age, years
<65	41 (41.4)
≥65	58 (58.6)
Differentiation
Poor	42 (42.4)
Well	57 (57.6)
Lymphovascular invasion
No	23 (23.2)
Yes	76 (76.8)
TNM stage
I-II	40 (40.4)
III	59 (59.6)
Tumor invasion
T1-2	26 (26.3)
T3-4	73 (73.7)
HER-2 expression
IHC 2+	38 (38.4)
IHC 3+	61 (61.6)
Lauren type
Intestinal	79 (79.8)
Non-intestinal	20 (20.2)
Tumor size, cm
<5	60 (60.6)
≥5	39 (39.4)
Perineural invasion
No	24 (24.2)
Yes	75 (75.8)
Lymph node metastasis
Negative	39 (39.4)
Positive	60 (60.6)
Neoadjuvant therapy
No	79 (79.8)
Yes	20 (20.2)
Follow-up outcome
Recurrence	47 (47.5)
Death	33 (33.3)

[i] TNM stage, Tumor-Node-Metastasis stage; IHC, immunohistochemistry; HER-2, human epidermal growth factor receptor-2.

Univariate and multivariate analysis of the overall population

Univariate and multivariate analyses were conducted to determine the prognostic risk factors for HER-2-positive gastric cancer. The univariate Cox regression analysis showed that HER-2 expression (P=0.002), tumor size (P=0.003), depth of tumor invasion (P=0.002), presence of lymph node metastasis (P<0.001), TNM stage (P<0.001), PNI (P=0.001) and LVI (P=0.007) were associated with poor DFS. Furthermore, the multivariable Cox regression analysis revealed that HER-2 expression (P=0.039), lymph node metastasis (P=0.030) and PNI (P=0.040) were independent prognostic factors impacting DFS among patients with gastric cancer (Table II). Notably, the univariate Cox regression analysis showed that tumor size (P=0.014), tumor invasion (P=0.010), TNM stage (P=0.002), lymph node metastasis (P=0.009), and PNI (P=0.018) were associated with decreased OS. HER-2 expression (P=0.074) did not exhibit a significant association with OS in patients with gastric cancer. The multivariable Cox regression analysis revealed no independent prognostic factors for OS in these patients (Table III).

Table II Univariate and multivariate Cox regression analysis of disease-free survival.

Table II

Univariate and multivariate Cox regression analysis of disease-free survival.

	Univariate analysis			Multivariate analysis
Variables	HR (95% CI)		P-value	HR (95% CI)		P-value
Sex (male/female)	1.081	0.516-2.006	0.960
Age (<65/≥65 years)	1.030	0.577-1.838	0.921
Differentiation (well/poor)	0.778	0.437-1.384	0.393
TNM stage (I-II/III)	4.392	2.048-9.421	<0.001	0.759	0.219-2.629	0.664
Lauren type (intestinal/non-intestinal)	1.281	0.637-2.578	0.487
Tumor size (<5/≥5 cm)	2.372	1.332-4.228	0.003	1.718	0.934-3.159	0.082
Tumor invasion (T1-2/T3-4)	5.277	1.889-14.74	0.002	1.510	0.416-5.477	0.531
Lymph node metastasis (no/yes)	4.842	2.166-10.82	<0.001	3.796	1.140-12.63	0.030
Perineural invasion (no/yes)	9.978	2.417-41.18	0.001	7.592	1.101-52.36	0.040
Lymphovascular invasion (no/yes)	4.113	1.474-11.47	0.007	0.337	0.081-1.397	0.134
HER-2 expression (IHC 2+/IHC 3+)	3.045	1.504-6.167	0.002	2.167	1.038-4.520	0.039

[i] TNM, Tumor-Node-Metastasis; IHC, immunohistochemistry; HR, hazard ratio; CI, confidence interval; HER-2, human epidermal growth factor receptor-2.

Table III Univariate and multivariate analysis of overall survival.

Table III

Univariate and multivariate analysis of overall survival.

	Univariate analysis			Multivariate analysis
Variables	HR (95% CI)		P-value	HR (95% CI)		P-value
Sex (male/female)	0.948	0.623-1.441	0.801
Age (<65/≥65 years)	0.829	0.581-1.184	0.303
Differentiation (well/poor)	1.645	0.911-1.180	0.154
TNM stage (I-II/III)	4.354	1.680-11.28	0.002	1.699	0.329-8.767	0.526
Lauren type (intestinal/non-intestinal)	1.827	0.848-3.937	0.124
Tumor size (<5/≥5 cm)	2.364	1.188-4.703	0.014	1.657	0.810-3.390	0.167
Tumor invasion (T1-2/T3-4)	6.611	1.581-27.64	0.010	2.443	0.393-15.19	0.338
Lymph node metastasis (no/yes)	3.239	1.337-7.849	0.009	1.340	0.337-5.332	0.678
Perineural invasion (no/yes)	5.630	1.347-23.53	0.018	1.525	0.241-9.646	0.654
Lymphovascular invasion (no/yes)	2.422	0.850-6.896	0.098
HER-2expression (IHC 2+/IHC 3+)	2.015	0.934-4.349	0.074

[i] TNM, Tumor-Node-Metastasis; IHC, immunohistochemistry; HR, hazard ratio; CI, confidence interval; HER-2, human epidermal growth factor receptor-2.

Different characteristics of gastric cancer with different HER-2 protein expression levels

HER-2 positivity was identified in 99 patients: 61 (61.6%) were IHC 3+ and 38 (38.4%) were IHC 2+ with FISH confirmation. These 38 FISH-positive cases represent 22.6% (38/168) of the initial IHC 2+ screening cohort (Fig. 1). Based on an analysis of clinicopathological characteristics according to HER-2 expression, it was found that PNI and LVI were more frequent in HER-2 IHC 3+ patients (51/61 and 51/61, respectively) than in IHC 2+ patients (24/38 and 25/38, respectively), with P=0.021 and P=0.041 for the respective comparisons. However, there were no statistically significant differences in terms of sex, age, TNM stage, Lauren classification, tumor size, tumor invasion or lymph node metastasis between the two groups (Table IV).

Figure 1 Immunohistochemical staining and fluorescence in situ hybridization of HER-2. (A) HER-2 2+ (x200 magnification); (B) HER-2 3+ (x200 magnification); (C) positive HER-2 gene amplification (x1,000 magnification). Arrows indicate HER2 (red) and CEP17 (green) signals. HER-2, human epidermal growth factor receptor-2; CEP17, chromosome enumeration probe 17.

Table IV Characteristics of gastric cancer with different HER-2 protein expression.

Table IV

Characteristics of gastric cancer with different HER-2 protein expression.

Characteristics	HER-2 IHC 2+	HER-2 IHC 3+	P-value
Sex, n (%)			0.919
Male	29	46
Female	9	15
Age, n (%)			0.912
<65 years	16	25
≥65 years	22	36
Differentiation, n (%)			0.713
Poor	17	25
Well	21	36
TNM stage, n (%)
I-II	19	21	0.125
III	19	40
Lauren type, n (%)			0.232
Intestinal	28	51
Non-intestinal	10	10
Tumor size, n (%)			0.093
<5 cm	27	33
≥5 cm	11	28
Tumor invasion, n (%)			0.059
T1-2	14	10
T3-4	24	51
Lymph node metastasis, n (%)			0.088
No	19	20
Yes	19	41
Perineural invasion, n (%)			0.021
No	14	10
Yes	24	51
Lymphovascular invasion, n (%)			0.041
No	13	10
Yes	25	51
Neoadjuvant therapy, n (%)
No	33	46	0.168
Yes	5	15

[i] TNM, Tumor-Node-Metastasis; HER-2, human epidermal growth factor receptor-2; IHC, immunohistochemistry.

Survival outcome

The study involved 99 patients, with a median DFS time of 29 months [95% CI, 21-37) and a median OS time of 39 months (95% CI, 36-50). Kaplan-Meier survival analysis revealed a significantly shorter median DFS time in patients with HER-2 IHC 3+ compared with those in patients with HER-2 IHC 2+ (24 months vs. not reached, P=0.002). The disease-free survival rates at 1, 3 and 5 years post-operation were notably lower in patients with HER-2 IHC 3+ gastric cancer compared with those in patients with HER-2 IHC 2+ gastric cancer (81.96, 43.38 and 28.34% vs. 89.47, 76.02 and 72.02%; P=0.0009). However, the OS rates at 1, 3, and 5 years post-surgery exhibited no differences between patients with HER-2 IHC 3+ gastric cancer and those with HER-2 IHC 2+ gastric cancer (93.84, 63.47 and 52.01% vs. 94.74, 83.59 and 76.43%; P=0.0629) (Fig. 2).

Figure 2 DFS and OS of patients in the HER-2 IHC 2+ and HER-2 IHC 3+ groups. (A) DFS and (B) OS. DFS, disease-free survival; OS, overall survival.

Discussion

The HER-2 gene, located on the long arm of chromosome 17, encodes the ERBB-2 receptor protein situated on the cell membrane (19). This protein belongs to the epidermal growth factor receptor family and regulates various cellular functions, such as differentiation, apoptosis, adhesion, migration and growth, through the tyrosine kinase pathway upon activation (20). Overexpression or amplification of HER-2/ERBB-2 has been observed in breast cancer (21), colorectal cancer (22), lung cancer (23), tumors of the reproductive system (24), and gastric cancer (25).

Upregulation/amplification of HER-2 in breast cancer is associated with an unfavorable prognosis. Similarly, the HER-2 gene is an important prognostic factor for gastric cancer. Aznab et al (6) observed a marked difference in survival and mortality rates between patients with HER-2-positive and HER-2-negative gastric cancer. The analysis of HER-2 status and prognostic indicators in 97 patients with gastric cancer revealed a strong association between HER-2 status and the stage of the disease. Zhang et al (7) reported that patients with HER-2 upregulation had higher recurrence rates and shorter survival times. Moreover, HER-2 upregulation was identified as an independent prognostic factor following surgery for T3 gastric adenocarcinoma. Kurokawa et al (8) conducted a large-scale multicenter study to evaluate the prognostic implications of HER-2 expression in gastric cancer. The research demonstrated that patients with HER-2 positivity had markedly poorer survival outcomes compared with HER-2-negative individuals. Cheng et al (26) pooled 30 studies (n=9,945) and showed HER-2 upregulation was associated with worse OS in both resectable (HR, 1.56; 95% CI, 1.32-1.85) and advanced (HR, 1.70; 95% CI, 1.23-2.35) gastric cancer, supporting HER-2 positivity as an unfavorable prognostic biomarker. However, Kung et al (9) reported no marked differences in OS or DFS between patients with HER-2-negative and HER-2-positive gastric cancer after surgical resection (n=334), suggesting that HER-2 is not an independent prognostic factor. Notably, HER-2-positive tumors were associated with a higher risk of distant lymphatic recurrence. The present study identified HER-2 expression, tumor size, tumor invasion, lymph node metastasis, TNM stage, PNI and LVI as factors associated with reduced DFS in patients with HER-2-positive gastric cancer. Importantly, HER-2 expression, lymph node metastasis and PNI were identified as independent prognostic factors influencing DFS in patients with gastric cancer, which is consistent with the findings of Zhang et al (27). Additionally, the present study indicated that tumor size, tumor invasion, lymph node metastasis, PNI and LVI were associated with reduced OS time; however, no factors, including HER-2 protein expression, were independent prognostic factors in HER-2-positive gastric cancer. This result is in line with previous research by Fisher et al (28). Although individual studies report inconsistent prognostic implications of HER-2 in gastric cancer, most investigations and meta-analyses support HER-2 positivity as a marker of poor survival (7-9,26,27). Endogenous HER-2 upregulation drives an aggressive phenotype, largely through preferential formation of HER2/HER3 heterodimers that constitutively activate the PI3K/Akt and MAPK pathways, thereby promoting proliferation, angiogenesis, epithelial-mesenchymal transition and chemoresistance (29).

HER-2 status in clinical practice is primarily assessed by IHC and FISH. Although FISH is considered the gold standard owing to its high sensitivity, specificity and reproducibility, its widespread use is limited by technical complexity, specialized equipment requirements and high cost. IHC is a commonly used method to evaluate HER-2 protein expression due to its cost-effectiveness, rapid detection, and broad applicability in gastric and breast cancer (30,31). In the present study, the IHC method was employed for the initial screening of HER-2-positive gastric cancer. For cases with ambiguous IHC interpretation results, the FISH method was subsequently utilized for confirmation, which not only reduced testing costs but also enhanced the accuracy of the results. For the first time, HER-2-positive gastric cancer was stratified by protein expression level to evaluate its association with clinicopathological features and survival. These findings provide evidence to guide prognostic assessment and targeted therapy in this patient population.

In the present study, HER-2 protein expression was not significantly associated with sex, age, tumor differentiation, TNM stage, Lauren classification, tumor size, depth of invasion or lymph node status. However, HER-2-positive tumors, particularly IHC 3+ cases, were more frequently characterized by perineural and lymphovascular invasion. Although the current TNM classification does not include LVI and PNI, these factors are widely recognized by researchers as being linked to the aggressive biological behavior of tumors (32-34). LVI refers to the invasion, destruction or presence of tumor cells within the lumen of small blood vessels and lymphatic vessels in and around the tumor area under microscopic examination (35). A previous study found that LVI is a prognostic factor for gastric cancer. Positive LVI in patients with gastric cancer was linked to malignancy, deeper invasion, lymph node metastasis and a worse prognosis (36). Wu et al (37) showed that LVI correlates with deeper invasion, greater nodal burden and lower overall survival rate. PNI, an independent dissemination pathway, is a well-established predictor of early recurrence and poor prognosis in gastric cancer. Patients with gastric cancer positive for PNI have higher rates of postoperative local recurrence and lower OS rates (38). Sun et al (39) reported that, in patients with locally advanced gastric cancer receiving neoadjuvant therapy followed by surgery, PNI and lymph node metastasis were independent prognostic factors. These findings underscore that both LVI and PNI are key determinants of survival in gastric cancer. Patients with both perineural and lymphovascular invasion in gastric cancer face an increased risk of lymph node metastasis and a poorer prognosis (34). Studies show that nerves, together with blood vessels and lymphatic vessels, collectively constitute the external microenvironment for tumor growth; their interaction with tumor cells promotes tumorigenesis and progression (40,41). A study also showed that VEGF expression is upregulated in gastric cancer tissues. By promoting angiogenesis and generating structurally immature vessels with defective basement membranes, VEGF facilitates tumor cell intravasation and hematogenous dissemination (42). PNI is currently considered a special mode of tumor metastasis; it involves complex interactions between tumor cells, stromal cells, surrounding immune cells and nerve fibers (40,43). Current understanding of the biological mechanisms underlying LET/PNI in gastric cancer is limited, and the interaction with the HER-2 gene is even more poorly understood. This gap highlights the imperative for future, in-depth research to elucidate these clinically important mechanisms.

Patients with HER-2 IHC 3+ gastric cancer may exhibit unfavorable biological characteristics associated with increased recurrence and risk of metastasis. The present study found that HER-2 IHC 3+ patients had significantly shorter median DFS times compared with HER-2 2+ patients. Additionally, the DFS rates at 1, 3 and 5 years post-surgery were notably lower for patients with HER-2 3+ gastric cancer compared with patients with HER-2 2+ gastric cancer, indicating a heightened vulnerability to postoperative recurrence and metastasis. Although 1-, 3- and 5-year OS rates did not differ significantly between patients with HER-2 IHC 3+ and 2+ gastric cancer, survival trends suggested a potential disadvantage for HER-2 IHC 3+ cases, a pattern that may become more pronounced with extended follow-up.

In conclusion, patients with HER-2 IHC 3+ gastric cancer demonstrate elevated rates of post-surgical recurrence and metastasis, resulting in a reduced OS time. It is advisable to maintain vigilant monitoring and implement intensified postoperative surveillance protocols for this patient group to detect signs of tumor recurrence and metastasis promptly. Furthermore, the potential efficacy of anti-HER-2 therapy in enhancing survival outcomes among these individuals warrants additional investigation.

Limitations include the single-center, retrospective design and selection of only surgically resectable cases, limiting generalizability. Potential bias from HER-2 heterogeneity across Lauren subtypes and the absence of dynamic HER-2 monitoring may further affect interpretation. Larger, multi-center prospective studies with serial HER-2 assessments are needed.

In conclusion, HER-2 is a significant negative prognostic factor associated with poorer DFS, although data on OS remain limited. HER-2 IHC 3+ gastric cancer exhibits a higher prevalence of PNI and LVI, as well as an elevated risk of postoperative recurrence and metastasis.

Acknowledgements

Not applicable.

Funding

Funding: The present study was supported by the Startup Fund for Scientific Research, Fujian Medical University (grant no. 2022QH1276).

Availability of data and materials

The data generated in the present study may be requested from the corresponding author.

Authors' contributions

HS contributed substantially to study conception, design, data acquisition, analysis, and interpretation, and wrote the main manuscript. SL conceived the research idea and critically revised the manuscript. HS and HH confirm the authenticity of all the raw data. XX contributed to data acquisition through active participation in both data analysis and interpretation, and co-wrote the first draft. HH prepared the figures and tables, involving data selection and organization for visual presentation, and contributed to manuscript revision. HH was also actively engaged in the interpretation of pathological data, playing a crucial role in elucidating the pathological findings and their correlation with the clinical data. All authors commented on previous versions of the manuscript. All authors read and approved the final version of the manuscript.

Ethics approval and consent to participate

All procedures involving human participants were performed according to the ethical standards of the institutional and national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. The study was approved by the Institutional Research Ethics Committee of Zhangzhou Hospital Affiliated with Fujian Medical University (approval no. 2025LWB426). As a retrospective study, the requirement for informed consent was waived by the ethics committee.

Patient consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

References