Abstract

BACKGROUND/AIMS

This study aims to assess the demographic, histopathological, and immunohistochemical (IHC) features of appendiceal neuroendocrine tumors (aNETs) and investigate their associations with tumor location, grade, size, and pathological stage. The follow-up data on survival and surgical outcomes is utilized to improve risk stratification and management strategies.

MATERIALS AND METHODS

A retrospective analysis was conducted on 5,483 appendectomy specimens evaluated between 2010 and 2020 in a single tertiary center. Among 115 neoplastic lesions, 45 cases were confirmed as aNETs. Demographic data, histopathological parameters, and IHC markers were reviewed. Survival data were collected from national records.

RESULTS

The median patient age was 35, with a female-to-male ratio of 1.37. Most tumors were located in the distal appendix (70.5%) and graded as G1 (82.2%). Mean tumor diameter was 0.91 cm, and the median Ki-67 proliferation index was 1%. Tumor grade and size were significantly associated with both tumor location and pathological tumor stage. Diffuse and proximal tumors were more likely to be G2, larger in size, and in advanced stages (pT4). Lymphovascular invasion, perineural invasion, and surgical margin positivity were more frequent in higher pT stages. Chromogranin A negativity was observed exclusively in pT1 cases.

CONCLUSION

aNETs are typically small, well-differentiated tumors with indolent behavior. However, tumor location, size, grade, and invasion features are associated with pathological stage and may serve as prognostic indicators. These parameters should be considered collectively to optimize clinical decision-making and surgical planning.

INTRODUCTION

Appendiceal neuroendocrine tumors (aNETs) are the most commonly encountered neoplasms of the appendix, often diagnosed incidentally during histopathological evaluation following appendectomy for suspected acute appendicitis (AA).^1-3 Despite their relative rarity, aNETs account for 30-80% of all appendiceal neoplasms and typically exhibit indolent biological behavior.^4-6 Most are well-differentiated, measure less than 2 cm, and are located at the distal tip of the appendix.^7-10 As such, simple appendectomy is considered curative in the majority of low-risk cases.^{7, 11}

Current clinical guidelines by the European Neuroendocrine Tumor Society (ENETS) and the North American ETS (NANETS) recommend right hemicolectomy (RH) for tumors larger than 2 cm or in the presence of high-risk histopathological features such as mesoappendiceal invasion (MAI), lymphovascular invasion (LVI), high Ki-67 index, or positive surgical margins.^{11, 12} However, the appropriateness of RH remains a subject of ongoing debate, particularly for intermediate-sized tumors (1-2 cm) without additional risk factors.^{13, 14}

Although tumor size (TS), grade, and invasion characteristics have been associated with recurrence risk and survival in prior studies, real-world data evaluating these variables regarding long-term outcomes remains limited, particularly in patients managed conservatively.^{2, 5} Furthermore, the prognostic relevance of tumor localization within the appendix (a parameter not consistently addressed in major series) warrants further investigation.

This study aims to evaluate the demographic, histopathological, and immunohistochemical (IHC) characteristics of aNETs diagnosed over ten years at a single tertiary care institution. In addition to characterizing tumor features, the study seeks to explore potential associations between tumor location, grade, size, and pathological tumor (pT) and to examine clinical follow-up data focusing on survival and the need for further surgical intervention. Through these objectives, the study intends to contribute meaningful data toward refining risk stratification and management strategies in patients with aNETs.

MATERIALS AND METHODS

This study is a single-center, observational cohort study based on the retrospective analysis of appendectomy specimens collected between 2010 and 2020. The study was approved by the University of Health Sciences Türkiye, Non-Interventional Clinical Research Ethics Committee of İstanbul Training and Research Hospital (approval number: 99, date: 02.05.2025) and all data were obtained in accordance with the principles of the Declaration of Helsinki.

Case Selection and Data Collection

We included 5,483 appendectomy specimens evaluated in our institution’s pathology laboratory between April 1, 2010, and April 1, 2020. Pathology reports and the hospital information system were reviewed to identify 115 neoplastic lesions, of which 45 were confirmed as aNETs. Diagnoses were verified by reevaluating hematoxylin-eosin stained slides and IHC analyses.

Data were collected from the hospital’s electronic archive system, including detailed reviews of pathology reports, radiological imaging results, surgical, and oncological treatment records.

Tumor characteristics, including tumor diameter, location, histological grade (HG), pT, resection margin status, presence of mesoappendiceal, lymphovascular, perineural invasion (PNI), and concurrent histopathological findings (CHF) such as AA and diverticulitis, were recorded. Tumor grading was based on mitotic count (per 10 HPF/2 mm²) and Ki-67 proliferation index (PI). The evaluation criteria were guided by the current American Joint Committee on Cancer, Cancer Staging Manual.¹⁵

IHC staining results for synaptophysin (Syn), chromogranin A (CgA), and Ki-67 antibodies were recorded following re-evaluation. A staining threshold of  >10% was considered positive for Syn and CgA. The Ki-67 PI was determined manually by counting at least 500-2,000 cells in hotspot areas.

Clinical Data and Follow-up

Demographic data and overall survival (OS) status were also recorded. OS was defined as the time from diagnosis to death from any cause or last follow-up. The presence of distant metastasis was evaluated at diagnosis and five-year follow-up. Additionally, whether patients underwent further surgical treatment after diagnosis (such as RH) was also recorded. Survival information was obtained through the Ministry of Health’s Death Notification System.

Statistical Analysis

Data were analyzed using SPSS version 25.0 (IBM Corp., Armonk, NY, USA) and Microsoft Office Excel 365 (Microsoft, Redmond, WA, USA). Continuous variables were tested for normality using the Shapiro-Wilk test. Normally distributed variables are expressed as mean and standard deviation (SD) and compared using the Independent Samples t-test, whereas non-normally distributed variables are presented as medians and analyzed using the Independent Samples Kruskal-Wallis test. Categorical variables are reported as frequencies and percentages and compared using the chi-square test, with Bonferroni correction applied for multiple comparisons. Survival analyses were conducted using the Kaplan-Meier method. Deaths that occurred between the date of diagnosis and April 2025 (the end date of the study) were recorded. Patients lost to follow-up or alive at the study endpoint were censored. The differences between survival curves were evaluated using the log-rank test, with a p-value of <0.05 considered statistically significant.

RESULTS

This study included 45 patients with aNETs. The mean age was 36 years (SD = 15), ranging from 14 to 70 years, with a median of 35 years. Of these, 26 patients (57.8%) were women, and 19 (42.2%) were men, resulting in a women-to-men ratio of 1.37:1 (Table 1).

Tumors were most frequently located in the distal appendix (70.5%), followed by diffuse (15.9%) and proximal (13.6%) locations. The mean TS was 0.91 cm (SD = 0.89), ranging from 0.1 to 5 cm. Histologically, 82.2% of tumors were classified as grade 1 and 17.8% as grade 2. No grade 3 tumors were observed.

Pathological staging revealed that 40% of tumors were pT1, 46.7% were pT3, and 13.3% were pT4. Surgical margins were negative in 90.5% of cases and positive in 9.5%. LVI was present in 17.8% of patients, PNI in 15.6%, and MAI in 6.7%. CHF included AA in 68.9%, mucinous neoplasia in 6.7%, and diverticulosis, and fibrous obliteration in 4.4% of cases (Table 1).

All tumors were positive for Syn. CgA was positive in 86.7% and negative in 13.3% of cases. The mean Ki-67 PI was 2.0% (SD = 2.9), with a median of 1%.

During follow-up, 93.3% of patients were alive, while 6.7% (n=3) had died. Two patients died due to cardiovascular comorbidities, and one patient died of sepsis. OS analysis was performed using the Kaplan-Meier method. Although only three deaths occurred during follow-up, a survival curve was generated for illustration (Figure 1). No statistical comparison was made due to the limited number of events.

At the time of diagnosis and during follow-up, no patients (0%) exhibited evidence of distant metastasis. Additionally, no patients underwent RH as a planned post-diagnostic treatment. However, four patients (8.9%) were diagnosed incidentally during RH procedures that had been performed for other clinical indications. Descriptive characteristics of the patients are summarized in Table 1.

Correlation Between Tumor Location and Clinicopathological Features

Tumor location was significantly associated with HG (p=0.007), pT (p=0.045), and TS (p=0.042). Distal tumors were predominantly grade 1 (93.5%), while proximal (50.0%) and diffuse tumors (42.9%) demonstrated  grade 2 differentiation. Similarly, pT1 tumors were mostly localized distally (51.6%), whereas diffuse tumors were more frequently staged as pT4 (42.9%, Table 2).

The median TS varied significantly by location, being largest in diffusely located tumors (1.00 cm), followed by distal (0.70 cm) and proximal (0.35 cm) locations (p=0.007).

There was a borderline association between tumor location and positive surgical margins (p=0.060), with the highest rate observed in proximally located tumors (33.3%).

Other clinicopathological parameters, including sex, LVI, PNI, CHF, MAI, CgA expression and Ki-67 PI, were not significantly associated with tumor location (Table 2).

Correlation Between Pathological Tumor and Clinicopathological Parameters

pT was significantly associated with TS (p<0.001), Ki-67 PI (p=0.016), HG (p=0.003), LVI (p=0.038), and CgA expression (p=0.006) Table 3. TS and Ki-67 PI increased in higher pT stages, with median TSs of 0.30 cm, 0.80 cm, and 1.85 cm in pT1, pT3, and pT4 stages, respectively. Similarly, the median Ki-67 PI rose from 1.0% in pT1 and pT3 tumors to 5.5% in pT4 tumors. Grade 2 tumors were more frequently observed in advanced stages, accounting for 66.7% of pT4 tumors.

As shown in Table 3, LVI was absent in pT1 cases but present in 28.6% and 33.3% of pT3 and pT4 tumors, respectively. CgA negativity was exclusively observed in pT1 tumors (33.3%), whereas all pT3 and pT4 tumors were CgA positive.

There was a borderline association between pT and both PNI (p=0.054) and positive surgical margins (p=0.052), with rates of 33.3% in pT4 tumors for both parameters. No statistically significant associations were found between pT and age, sex, CHF, or MAI (Table 3).

DISCUSSION

This study evaluated the demographic, histopathological, and IHC characteristics of aNETs diagnosed over 10 years. Our main findings demonstrated that aNETs predominantly occur in middle-aged women, are most often localized at the distal tip of the appendix, and measure less than 1 cm. Most tumors were classified as WHO grade 1 with low Ki-67 PIs, and MAI was infrequent. Tumor location was significantly associated with HG, pT, and TS. OS outcomes were excellent, with a 93.3% survival rate during follow-up. These results are consistent with the existing literature and support current clinical management guidelines favoring simple appendectomy for small, well-differentiated tumors without adverse features.

In line with previous reports, the median age of our cohort was 35 years, with a slight female predominance (57.8%). These findings are consistent with several large series reporting median ages between 33 and 36.5 years and a female predominance ranging from approximately 54.5% to 64.5%.^{2, 3} These findings reinforce the understanding that aNETs typically affect young to middle-aged adults and exhibit a slight gender disparity. The demographic profile of our study thus mirrors the general epidemiological trends observed globally.

The predominant localization of tumors at the distal tip of the appendix (70.5%) and a mean TS of 0.91 cm were consistent with existing studies, which reported distal localization rates between 58.1% and 83.9% and TSs predominantly less than 1 cm in diameter.^{2, 10, 16} This distribution is clinically relevant, as tumor location and TS significantly impact surgical decision-making and prognosis.

HG revealed that most tumors were grade 1 (82.2%), corroborating findings from prior studies where grade 1 tumors accounted for 60.7% to 90.9% of cases.^{3, 17} The observed low Ki-67 PI further reinforced the indolent nature of these neoplasms. Importantly, MAI was relatively infrequent (6.7%), supporting the generally favorable prognosis of small, well-differentiated aNETs.

Tumor location showed significant associations with HG, pT, and TS. Distally located tumors were more often grade 1 and staged as pT1, while diffuse tumors exhibited a tendency towards higher grades and stages. These results are in line with findings from Okut and Karahan¹⁸ highlighting that proximally or diffusely located tumors warrant closer clinical scrutiny.

pT correlated strongly with TS, Ki-67 PI, HG, and invasion parameters, which echoesthe established understanding that deeper invasion, and larger TS are markers of more aggressive behavior.^{9, 19-21} LVI and PNIs, while relatively infrequent, were significantly associated with advanced pTs, aligning with international observations.^{6, 9}

All tumors showed positivity for Syn (100%), and 86.7%, were positive for CgA, comparable to prior reports where positivity rates for Syn and CgA exceeded 83%.10 The low Ki-67 PIs, with most cases under 3%, reflect the low proliferative activity that characterizes well-differentiated aNETs.

Notably, no patients in our cohort exhibited evidence of distant metastasis at the time of diagnosis or during the follow-up period. This finding aligns with most published series, which consistently report very low rates of metastatic spread in small, well-differentiated aNETs (typically <1%). Distant metastases are reported in a small subset of patients, often associated with high-grade or deeply invasive tumors.^{4, 22, 23} These results underscore the indolent behavior of aNETs and reinforce the appropriateness of conservative surgical management in low-risk cases.

The majority of patients in this cohort underwent only appendectomy, aligning with ENETS and NANETS guidelines, which recommend simple appendectomy for tumors smaller than 1 cm without high-risk features.^{11, 12} The necessity of RH remains debated for tumors between 1 and 2 cm, particularly in the absence of adverse features.¹⁷In our cohort, the observation of negative surgical margins in most cases (90.5%) and a low frequency of adverse histopathological features may support the safety of conservative management in selected patients, which aligns with current guideline recommendations.

Survival outcomes were favorable, with a 93.3% OS rate at last follow-up and no evidence of disease recurrence, comparable to other series reporting 5-year disease-free survival rates exceeding 90%.^{3, 11, 13, 19}

Study Limitations

This study has several limitations, primarily its retrospective single-center design, which may limit generalizability. The relatively small sample size and limited number of adverse events also precluded advanced multivariate analysis. Furthermore, although the follow-up duration was substantial, long-term outcomes (>5 years) remain unexplored.

Future Directions

Further prospective multicenter studies are needed to validate prognostic markers and improve surgical decision-making, especially for tumors measuring between 1 and 2 cm. Recent proposals suggest that INSM1 and other novel IHC markers could assist in more precise risk stratification.²⁴

Molecular profiling studies also offer a promising frontier for understanding tumor behavior and tailoring therapy, especially considering the biological heterogeneity observed among EC-cell and L-cell tumors.^{25, 26}

Moreover, with the rising trend toward non-operative management of AA, the potential risk of missing incidental aNETs merits close surveillance and possibly the development of preoperative predictive algorithms.^{16, 17, 27}

CONCLUSION

Our findings reinforce the generally indolent behavior of aNETs, characterized by small TS, distal location, low-grade histology, and favorable prognosis. These results support the current guidelines advocating for appendectomy as the definitive treatment in the majority of cases and highlight the need for vigilance in assessing histopathological features indicative of higher-risk disease. Continued research into the biological diversity of aNETs and longer-term follow-up studies are essential to optimize patient outcomes.