PERITUMORAL BUDDING AS A PREDICTOR FOR LYMPH NODE METASTASES IN COLORECTAL CARCINOMAS: WHAT IS THE IMPORTANCE?

TL;DR

Tumor budding in colorectal cancer is linked to lymph node metastases and could improve patient prognosis and treatment decisions.

Contribution

This study demonstrates that tumor budding is a significant predictor of lymph node metastases in colorectal cancer patients.

Findings

Tumor budding was present in 63.39% of patients and was significantly associated with lymph node metastases.

High budding scores were observed in 51.55% of cases with tumor budding.

Tumor budding complements TNM staging in predicting metastasis risk.

Abstract

Microscopic analysis of tumor budding (TB) may be an essential predictive tool for regional lymph node metastases in colorectal cancer, especially among patients in intermediate stages, who exhibit considerable prognostic variability.

The aim of this study was to assess the predictive power of BT regarding the presence of lymph node metastases and its association with other characteristics related to colorectal carcinoma progression.

This is a cross-sectional, retrospective study with a quantitative approach, focusing on the review of medical records and histopathological reports of patients who underwent oncologic surgery for colorectal cancer.

A total of 153 patient records were examined, with a predominance of the 61-70 age group and a male majority (50.98%). Adenocarcinoma not otherwise specified was the most common histological type (60.78%), with the majority exhibiting moderate differentiation (87.58%). From the total sample, 97 cases (63.39%) exhibited TB, with 51.55% classified as a high budding score. Invasion of adipose tissue/subserosa was the most prevalent, occurring in 46.41% of cases. Regional lymph node metastases and angiolymphatic invasion were observed in 66 and 101 patients, respectively. Cross-tabulation analysis showed a statistically significant association between TB and lymph node metastasis (p<0.05).

The relationship between TB and lymph node metastasis highlights the significance of this histological factor in the risk stratification and prognosis of patients with colorectal cancer, complementing TNM staging. Therefore, the assessment of tumor budding is crucial in histopathological reports, potentially influencing additional therapeutic decisions.

Figures

Distribution of patients with colorectal cancer by age groups (n=153).

Relationship between lymph node involvement and tumor budding.ROC: receiver operating characteristic; AUC: area under the curve.

The blue curve shows the test performance; the dashed gray line shows a test with no discriminative ability. The area under the curve of 0.622 indicates moderate test accuracy.

Full text

INTRODUCTION

Colorectal cancer (CRC) is the second-most prevalent cancer in both sexes in Brazil, excluding non-melanoma skin cancer, and is associated with significant morbidity and mortality11. The prognosis of CRC is largely based on the disease stage according to the TNM classification12. However, there is significant variability in outcomes and prognosis among patients at identical stages, especially in intermediate stages (II and III)16. Therefore, additional strategies to stratify the risk of recurrence and metastasis, aiming to enhance the treatment of CRC patients, may provide beneficial alternatives through the utilization of the tumor budding (TB) marker14.

TB is a phenomenon where single isolated cells or small clusters of up to four cells are present at the invasive front of the tumor, indicating the tumor’s capacity to infiltrate the extracellular matrix and lymphatic vessels to reach regional lymph nodes19. This process has been associated with worse prognosis, including higher local recurrence, distant metastases, and reduced survival in CRC patients16. In 2016, the International Tumor Budding Consensus Conference (ITBCC) standardized TB assessment in CRC patients into three distinct groups based on the number of buds identified under microscopy:

1. Low TB group: a maximum of four buds in the assessment field (hotspot area);
2. Intermediate TB group: between five and nine buds;
3. High TB group: 10 or more buds8.

The aim of this study was to assess the predictive power of TB regarding the presence of lymph node metastases, and other characteristics of tumor progression in CRC, in a Brazilian university hospital.

METHODS

This is an observational, retrospective study with a quantitative approach, conducted in Curitiba (PR), Brazil. Medical records from the Mackenzie Evangelical University Hospital (HUEM) were examined, along with histopathological results of resected surgical specimens from patients who underwent oncological surgery for CRC between November 2019 and June 2022. Patients with missing data in medical records and/or incomplete histopathological analyses were excluded. Multiple variables, such as sex, age, type/grade of histological neoplasia, depth of invasion, angiolymphatic embolization, lymph node metastases, and TB assessment, were considered.

For analysis, the data were exported to a Microsoft Office Excel spreadsheet, where descriptive statistics were used, including absolute numbers and percentages. The chi-squared test was performed when applicable with a significance level of <0.05. The study was approved by the HUEM Research Ethics Committee (nº 70071623.7.0000.0103), in accordance with Resolution no. 466/12 of the National Research Council, which regulates scientific research in human subjects. The data obtained were compared with the literature on the subject.

RESULTS

From November 2019 to June 2022, 153 CRC cases were recorded at the hospital’s high-complexity center that met the inclusion criteria for this study. The distribution between sexes was similar, with a slight predominance of males, accounting for 50.98% of the cases. Age ranged from 35 to 94 years, as shown in Figure 1, with a mean of 62.76±12.02 and with 30.07% predominantly in the age group of 61-70 years. Figure 1 shows the distribution of CRC patients by age group (n=153).

Figure 1. Distribution of patients with colorectal cancer by age groups (n=153).

Adenocarcinoma not otherwise specified (NOS) was the most prevalent histological type, present in 93 patients (60.78%) of the cases. Table 1 shows the degree of tumor differentiation, with the majority classified as moderately differentiated, totaling 134 cases (87.58%).

Table 1. Distribution of colorectal cancer differentiation degree (n=153).Differentiation degreen%Moderately differentiated13487.58Poorly differentiated1610.45Well-differentiated31.96Total153100

Table 2 shows that in 66 patients (43.13%), at least one metastasis in regional lymph nodes was detected in the surgical specimen.

Table 2. Distribution of colorectal cancer patients according to lymph node involvement (n=153).Lymph node involvementn%Not involved8757.14Involved6642.86Total153100

The analysis of tumor invasion depth (pathological staging T) of the samples showed a significant number of cases classified as pT3 (46.41%), characterized by tumor cell invasion into the pericolic adipose tissue, followed by pT4a (30.07%), which includes infiltration to the serosal surface. Additionally, the presence of pT2 (16.69%), characterized by the infiltration of the muscularis propria, was also particularly noteworthy. The relationship between tumor depth and lymph node metastasis presence was analyzed and presented in Table 3. There was a statistically significant association between tumor invasiveness and the probability of lymph node involvement (p<0.05).

Table 3. Tissue invasion depth (T) in colorectal cancer (n=153).Tissue infiltrationpTn%Lymph node metastasis (n)p-valueMucosapTis10.650 SubmucosapT142.60 Muscularis propriapT22416.693 Adipose tissuepT37146.4133 SerosapT4a4630.0727 Adjacent organspT4b74.583 Total-153100660.0032

Angiolymphatic embolization was examined and found in 101 patients (66.01%). However, in 53 cases (34.64%), it was not identified.

TB identification was described in 97 cases (63.39%), classified into low, intermediate, and high scores, as shown in Table 4. Over half of these cases had the highest score.

Table 4. Distribution of budding score according to classification into high, intermediate, or low (n=97).Budding scoren%High5051.55Intermediate2626.80Low2121.65Total97100

The association between TB and lymph node metastasis was considered significant (p<0.05) since 51 of the 66 patients with lymph node metastasis had TB. Conversely, most patients without lymph node involvement had positive TB (Figure 2). Furthermore, the sensitivity and specificity of peritumoral budding for regional lymph node involvement were 77.27% and 47.13%, respectively, with a positive predictive value of 52.58%.

Figure 2. Relationship between lymph node involvement and tumor budding.ROC: receiver operating characteristic; AUC: area under the curve.

Additionally, the ROC (receiver operating characteristic) curve analysis for TB on lymph node involvement in CRC patients revealed an area under the curve (AUC) of 0.622, as shown in Figure 3. This value indicates moderate accuracy of TB in distinguishing between cases with and without lymph node involvement. Moreover, likelihood ratios were calculated, with a positive likelihood ratio of approximately 1.46 suggesting that a positive TB test result increases the probability of lymph node involvement by approximately 46%. In contrast, a negative likelihood ratio of approximately 0.46 indicates that a negative result reduces the probability of lymph node involvement by approximately 46%.

Figure 3. The blue curve shows the test performance; the dashed gray line shows a test with no discriminative ability. The area under the curve of 0.622 indicates moderate test accuracy.

Finally, TB was associated with lymph node metastasis, histological grade, invasion depth, and angiolymphatic embolization. However, the latter exhibited no statistical significance, as shown in Table 5.

Table 5. Age, sex, lymph node involvement, angiolymphatic invasion, histological grade, and invasion depth in CRC patients with and without tumor budding (n=153). Tumor buddingp-valuePresent (n)Absent (n)9756Age (mean)62.6562.860.877Sex (n)Male 5227 Female 4530 Lymph node metastasis (n)Present 51150.0034^†^ Absent 4641Angiolymphatic invasion (n)Present 68330.2192^†^ Absent 2923Histological grade (n)Well-differentiated 030.0131^†^ Moderately differentiated 9044Poorly differentiated 79Invasion depthMucosa010.068^†^ Submucosa04Muscularis propria159Adipose tissue4526Serosa3214Adjacent organs52Student’s t-test; ^†^χ2 test.

DISCUSSION

Most of the 153 patients with CRC surgically treated in a high-complexity center were male (50.98%). These findings align with current data from the World Health Organization (WHO) for Brazil, where CRC exhibits an almost equal distribution between sexes, with males accounting for 49.6% of the total CRC cases6 ^,^ 18. The American Cancer Society (ACS) estimates that in 2022, there were 80,690 new cases among males, compared to 70,340 new cases among females1.

The predominant age group in this study was between 61 and 70 years, with a mean age of 62 years. Age assessment is crucial as it is a risk factor for CRC development, with cases being more common in patients over 50 years and significantly increasing with each decade. The average age at diagnosis is 72 years for women and 68 years for men10. A Brazilian study of 521 cases found an average age of 63 years, with an affected range between the fifth and the eighth decades of life3.

The most widely used and gold-standard pathological staging system for CRC evaluation is the TNM system, recommended by the American Joint Committee on Cancer (AJCC), which considers tumor invasion depth (T); the number of compromised lymph nodes (N); and the presence or absence of distant metastases (M)12 ^,^ 20. Our study showed a prevalence of 71 (46.41%) cases with infiltration into the pericolic adipose tissue, corresponding to pT3, which is also described as predominant in the global literature3 ^,^ 4 ^,^ 17. Additionally, at least one lymph node metastasis was identified in 66 patients (42.86%). A similar value has been described in the literature13.

TB was identified in 97 (63.39%) cases, with the majority classified as high scores, corresponding to 51.55% of the results. Furthermore, a statistically significant association was found between peritumoral budding and the presence of lymph node metastasis (p<0.05), with 77.27% sensitivity.

Although the mechanism of TB formation in colorectal cancer is not yet fully understood, the predominant theory suggests that at least some types of budding represent an example of epithelial-mesenchymal transition. It suggests that during the epithelial-mesenchymal transition, tumor cells undergo changes that include the loss of cell adhesion molecules, cytoskeletal modifications, increased production of extracellular matrix components, resistance to apoptosis, and the ability to degrade the basement membrane7.

These described changes result in a cellular phenotype with greater migratory and invasive capacity, allowing tumor cells to detach from the primary tumor mass and disseminate to adjacent tissues, lymph nodes, and distant organs7. Therefore, TB is considered an early step in the neoplasm’s ability to invade lymphovascular spaces and metastasize. Additionally, TB can currently be used as an auxiliary in CRC risk stratification2 ^,^ 15 ^,^ 19.

In 46 (30.06%) patients, the presence of TB was detected without lymph node involvement; yet, this finding suggests increased tumor aggressiveness, being associated with infiltration depth and substantial loss of cellular differentiation. Histopathological detection is crucial in assessing patients with early stage CRC due to its association with the presence of lymph node metastases, indicating the need for additional resections or association with other complementary therapeutic approaches, such as adjuvant oncological therapy, and in metastatic cases, it is associated with a worse prognosis5 ^,^ 15.

Marx et al. have shown a statistically significant association between TB and lymphovascular invasion9. However, in this study, this association was not confirmed. Several factors may justify this outcome, including imprecise histological assessment, technical difficulties in histotechnical processing, insufficient macroscopic sampling, or the fact that these were initial cases and TB had not yet resulted in subsequent embolization.

The ITBCC recommends the assessment of TB using hematoxylin and eosin-stained histological slides, and a hotspot at the invasive tumor front (standard field size of 0.785 mm^2^). To assist this analysis, the consensus provides a table for standardizing this count across various microscopes8. According to ITBCC, TB should be included in CRC protocols and described in patients’ pathology reports due to its prognostic, clinical, and therapeutic value. Thus, this histopathological evaluation should always be described. In the present study, information was sourced from radical surgical specimens; nonetheless, TB identification is particularly important in endoscopic resections of early cases, such as polypectomies and mucosectomies8.

In recent years, there has been a growing increase in CRC diagnosis, with local resection surgery considered curative in the absence of metastases5. TNM classification remains the primary staging stratification method. However, evaluating other new prognostic predictors in CRC patients is uniquely important in therapeutic decision-making in oncology.

CONCLUSIONS

This study established a significant relationship between the presence of TB and lymph node metastasis, emphasizing the relevance of tumor budding as a detectable histological prognostic factor in patients with colorectal cancer.