The optimal number of lymph nodes examined for rectal cancer patients undergoing neoadjuvant long-course chemoradiotherapy

Introduction

Colorectal cancer (CRC) is one of the leading causes of cancer-related death around the world (1). Lymph node (LN) metastasis is the most common metastasis and the most important prognostic factor for CRC patients (2). Currently, the tumor node metastasis (TNM) staging system relies on the absolute number of positive lymph nodes (PLNs) to categorize N stage: pN0 (PLN = 0), pN1 (PLN = 1–3), and pN2 (PLN ≥4), and each N substage carries a different prognosis. The established guidelines, such as those outlined by the National Comprehensive Cancer Network (NCCN) and the 8^th American Joint Committee on Cancer (AJCC) Staging Manual, recommend the examination of a minimum of 12 LNs for accurate N staging in CRC (2,3). Prior investigations have underscored the significant association between the number of LNs examined and LN metastasis in CRC (4-6). Therefore, the appropriate number of LNs examined is vital for the precise assessment of prognosis and individual treatment plan of the CRC patients.

Rectal cancer, which accounts for approximately half of new CRC cases reported annually worldwide (1), is a huge challenge in clinical practice due to the difficulty in treatment and the tendency to recur (7). Currently, neoadjuvant chemoradiotherapy (NCRT) followed by total mesorectal excision (TME) has become the standard treatment for the T3–4/N+ rectal cancer patients (8). However, numerous studies have documented a reduction in the number of LNs examined and the number of metastatic LNs in rectal cancer patients as a consequence of NCRT (9-11). Mechera et al. reported that NCRT led to an average reduction of 3.9 LNs and a mean reduction of 0.7 metastatic LNs, in comparison to the patients who did not receive neoadjuvant therapy (12). Therefore, it is plausible that the current guidelines advocating a specific number of LNs to be examined in rectal cancer surgery may be inappropriate for the patients undergoing neoadjuvant therapy.

This study aims to ascertain the optimal minimal number of examined LN (ELN) for rectal cancer patients who have undergone neoadjuvant therapy, using data derived from Cancer Hospital, Chinese Academy of Medical Sciences/National Cancer Center (NCC) and the Surveillance, Epidemiology, and End Results (SEER) database. We present this article in accordance with the STROBE reporting checklist (available at https://jgo.amegroups.com/article/view/10.21037/jgo-2025-463/rc).

Methods

Patient selection

The NCC and SEER databases served as the training cohort and validation cohort in this study, respectively.

In the NCC cohort, patient enrollment was based on the following inclusion criteria: (I) rectal cancer patients who underwent long-course NCRT and received radical resection at the Cancer Hospital, Chinese Academy of Medical Sciences/NCC from January 2004 to December 2015; (II) primary rectal cancer located at or below ≤10 cm from the anal verge; (III) clinical stage T3–4 or any T combined with N+; (IV) histological confirmation of adenocarcinoma. Patients were excluded from the NCC cohort if they met any of the following exclusion criteria: (I) absence of radiation therapy or receipt of short-course radiation; (II) underwent local resection or palliative surgery; (III) histopathological diagnosis other than adenocarcinoma; (IV) classified as stage N1c; (V) presence of metastatic disease at the time of diagnosis or intraoperatively; (VI) incomplete TNM staging information; (VII) incomplete LN evaluation data; (VIII) diagnosis of a second primary cancer; (IX) history of hereditary CRC syndromes, such as Lynch syndrome and familial adenomatous polyposis; (X) patients who died within 1 months following surgery or experienced relapse or metastasis within 6 months post-surgery; (XI) patients aged less than 18 years or over 85 years; (XII) patients with inflammatory bowel disease. The primary endpoint for the NCC cohort was disease-free survival (DFS). Overall survival (OS) is defined as the time interval from the date of surgery to the death of the patient. DFS is defined as the time interval from the date of surgery to the first occurrence of tumor recurrence or metastasis.

In the SEER cohort, data were retrieved from the SEER database for the 2019 release, spanning the years 1975 to 2016. The SEER program was the largest publicly accessible cancer dataset, encompassing data on incidence, prevalence, and survival, and it covers approximately 28% of the United States population across multiple geographical regions (13,14). We collected the data from the patients diagnosed with rectal adenocarcinoma who underwent NCRT follow by surgery between January 1, 2010 and December 31, 2015. The exclusion criteria for the SEER cohort mirrored those of the NCC cohort. Additionally, patients with unknown causes of death were also excluded from the SEER cohort. Given the absence of recurrence and metastasis timing data in the SEER database, cancer-specific survival (CSS) was employed as the primary endpoint for this cohort. CSS is defined as the time interval from the date of diagnosis to the death of the patient due to tumor-related causes.

The study was conducted in accordance with the Declaration of Helsinki and its subsequent amendments. The study was approved by ethics board of Cancer Hospital, Chinese Academy of Medical Sciences (No. 25/678-5625) and individual consent for this retrospective analysis was waived.

Statistical analysis

The optimal cutoff point for ELN was determined using X-tile software (Version 3.6.1) (15). Survival curves were generated employing the Kaplan-Meier method, and the disparities in survival were assessed through the log-rank test. Prognostic risk factors were identified using univariate and multivariate Cox regression analyses. All statistical analyses were carried out utilizing SPSS software, version 25.0 (SPSS Inc., Chicago, IL, USA). A two-sided significance level of P<0.05 was adopted to denote statistical significance.

Results

Patient characteristics

A total of 6,634 patients were included in this study, comprising 391 cases from the NCC cohort and 6,243 cases from the SEER cohort.

In the NCC cohort, the age of the enrolled patients ranged from 23–82 years (median, 55.0 years); 263 (263/391, 67.3%) of the patients were males. The distribution of patients across postneoadjuvant therapy pathological tumor (ypT) stages was as follows: 80 (80/391, 20.5%) cases with ypT0–1, 84 (84/391, 21.5%) cases with ypT2, and 227 (227/391, 58.0%) cases with ypT3–4. Regarding postneoadjuvant therapy pathological node (ypN) status, there were 263 (263/391, 67.3%) patients classified as ypN0, 90 (90/391, 23.0%) as ypN1, and 38 (38/391, 9.7%) as ypN2. Table 1 provides a summary of the key demographics and the clinicopathologic data of the patients. The median number of ELNs per patient was 14.0 (range, 2–61). The distribution of the number of ELNs in these patients is shown in Figure 1.

Figure 1 Distribution of the number of ELNs in the NCC cohort. ELN, examined lymph node; NCC, National Cancer Center.

In the SEER cohort, the median age of the enrolled patients was 59.0 years (range: 18–85 years). Of the 6,243 patients in this cohort, 3,824 (3,824/6,243, 61.3%) were males, and 2,419 (2,419/6,243, 38.7%) were females. The majority of the patients were of white ethnicity (5,043/6,243). The distribution of ypN stages in this cohort included 4,386 (4,386/6,243, 70.3%) ypN0 patients, 1,326 (1,326/6,243, 21.2%) ypN1 patients, and 531 (531/6,243, 8.5%) ypN2 patients (Table 2). The median number of ELNs in these patients was 14.0 (range, 1–83). The distribution of the number of ELNs in the SEER cohort is shown in Figure 2.

Figure 2 Distribution of the number of ELNs in the SEER cohort. ELN, examined lymph node; SEER, Surveillance, Epidemiology, and End Results.

The current minimum LN examination criteria have no prognostic significance for rectal cancer patients undergoing NCRT

The current guideline recommends a minimum of ELNs was 12. We divided the 391 patients in the NCC cohort into two groups based on the current criterion: ELNs <12 (143/391, 36.6%) and ELNs ≥12 (248/391, 63.4%). Our analysis revealed that there is no significant difference between the DFS of the two groups, the 5-year DFS rate is 72.0% for ELN <12 group and 76.5% for ELN ≥12 group (P=0.10) (Figure 3A). We further validated the results in the SEER cohort. Consistently, the survival analysis results also indicated no significant difference between the two groups (P=0.10), with 5-year CSS rates standing at 81.2% and 83.1%, respectively (Figure 3B). These findings suggest that the current LN examination criteria may not be suitable for the rectal cancer patients who have undergone NCRT, and it is necessary to substitute a more appropriate one.

Figure 3 The survival curves for patients with ELN <12 and those with ELN ≥12. (A) The disease-free survival curves for patients with ELN <12 and those with ELN ≥12 in the NCC cohort; (B) The cancer-specific survival curves for patients with ELN <12 and those with ELN ≥12 in the SEER cohort. ELN, examined lymph node; NCC, National Cancer Center; SEER, Surveillance, Epidemiology, and End Results.

Determining the optimal number of ELNs for rectal cancer patients undergoing NCRT

Given the limitations of the current criteria, we conducted an analysis using the NCC cohort as a development cohort to identify more appropriate minimum ELNs in rectal cancer patients who have received neoadjuvant therapy. The X-tile software, an objective methodology, possesses the capability to not only divide the cohort into two distinct subsets but also to ascertain an optimal cutoff value through the utilization of the minimum P value approach. And then we determined that 7 should be the optimal cutoff value (Figure 4). In Figure 4, the left-hand panel of the illustration depicts a prominent triangle and a rectangle. The horizontal axis of the triangle illustrates a gradual increase in the count of individuals possessing low values, whereas the vertical axis denotes a corresponding rise in those with high values. A click anywhere within the confines of the rectangle facilitates the segmentation of the entire population into two distinct groups, maximizing the difference between them. The resultant cutoff value for this segmentation is clearly presented in the bar graph positioned to the right.

Figure 4 To determine the optimal number of ELNs by using the minimum P value. ELN, examined lymph node.

To validate the new cutoff value, hazard ratios (HRs) for DFS were calculated based on various cutoff points of ELNs in the NCC cohort. The HR was found to be the lowest when the cutoff point for ELN was set as 7 [HR =0.542, 95% confidence interval (CI): 0.311–0.944] (Table 3). A lower HR value indicates a reduced risk of disease progression.

DFS analysis based on the new cut-off value

Using the new cutoff value of ELN 7, we categorized the 391 patients in the NCC cohort into two groups: those with ELN <7 (42 cases) and ELN ≥7 (349 cases). The 5-year DFS rate of patients with ELN <7 was 65.1%, compared to 76.3% for those with ELN ≥7 (P=0.03) (Figure 5).

Figure 5 The disease-free survival curves for patients with ELN <7 and those with ELN ≥7 in the NCC cohort. ELN, examined lymph node; NCC, National Cancer Center.

Cox univariate and multivariate regression analyses were conducted to identify the risk factors for DFS in the NCC cohort. In the univariate analysis, the factors significantly correlated with DFS included extramural vascular invasion (EMVI), vascular invasion, differentiation grade, tumor regression grade (TRG), ypT, ypN, and ELN (7 as the cut-off value) (all P<0.05). Multivariate analysis further confirmed that EMVI (P=0.001), TRG (P=0.005), ypN (P<0.001), and ELN (7 as the cutoff value) (P<0.001) were significantly associated with DFS (Table 1).

OS analysis based on the new cut-off value in the NCC cohort

OS was utilized as a second endpoint in the NCC cohort to validate the new cutoff value for prognostic stratification. The results demonstrated a 5-year OS rate of 72.8% for patients with ELN <7, compared to 84.8% for those with ELN ≥7 (P=0.01) (Figure 6A). Interestingly, no significant OS difference was observed between the group with <12 ELNs and the group with ≥12 ELNs (5-year OS rate: 83.5% vs. 82.6%, P=0.46) (Figure 6B). These findings confirmed the validity of the new cutoff value 7.

Figure 6 The overall survival curves for patients with ELN <7 and those with ELN ≥7. (A) The overall survival curves for patients with ELN <7 and those with ELN ≥7 in the NCC cohort. (B) The overall survival curves for patients with ELN <12 and those with ELN ≥12 in the NCC cohort. ELN, examined lymph node; NCC, National Cancer Center.

External validation in the SEER database

Applying the cutoff value 7, the patients in the SEER cohort were also divided into two groups: those with ELN <7 (742 cases) and ELN ≥7 (5,501 cases). The 5-year CSS rate for patients with ELN <7 and those with ELN ≥7 was 79.1% and 83.0%, respectively (P=0.04) (Figure 7). The 5-year OS rate for patients with ELN <7 was 74.5% compared with that of 78.9% for those with ELN ≥7 (P=0.01). Cox regression revealed that age, marital status, race, perineural invasion, differentiation grade, T stage, N stage, and ELN (7 as the cutoff value) were significantly associated with CSS in univariate analysis. In multivariate analysis, these variables remained the significant predictors of CSS (Table 2).

Figure 7 The cancer-specific survival curves for patients with ELN <7 and those with ELN ≥7 in the SEER cohort. ELN, examined lymph node; SEER, Surveillance, Epidemiology, and End Results.

Discussion

Currently, NCRT has become the standard treatment for T3–4/N+ rectal cancer. Nevertheless, the current guidelines recommend a minimum of ELNs was 12 irrespective of whether the patients have undergone neoadjuvant therapy or not (16-18). This standard has been applied uniformly to all patients, but its appropriateness for those who have received neoadjuvant therapy has remained questionable (3,19,20).

Several factors influence the ELNs, including the patient characteristics, tumor-related factors, the surgical techniques, the pathologists’ sampling methods, and the impact of neoadjuvant therapy (11,21-24). Neoadjuvant therapy, particularly the long-course NCRT, can result in down-stage and reduce the local recurrence of rectal cancer (25), also can decrease the ELNs and PLNs in subsequent resections samples (10,11,26).

The goal of our study was to assess the suitability of the 12 ELNs criteria and to determine a more appropriate cutoff value for ELNs in the context of neoadjuvant therapy. Our analysis, based on two distinct cohorts—the NCC cohort and the SEER cohort—aimed to provide a comprehensive perspective. Our findings indicate that the current standard of examining at least 12 ELNs may not be suitable for the rectal cancer patients who have undergone neoadjuvant therapy. In both the NCC and SEER cohorts, we observed that there was no significant survival difference between the patients with <12 ELNs and those with ≥12 ELNs. This suggests that the current guideline for ELNs may not accurately reflect the prognosis of these patients. To solve this problem, we employed the X-tile software to objectively determine the optimal cutoff value for ELNs. 7 was identified as the new cutoff of ELNs based on the NCC cohort. We then found that it can differentiated the survival significantly. The patients with fewer than 7 ELNs had significantly worse survival rates compared to those with 7 or more ELNs in both NCC cohort and SEER cohort. Numerous previous studies have confirmed that the number of LNs retrieved decreases in rectal cancer patients who have undergone neoadjuvant therapy, which is consistent with our current recommendation of examining at least 7 LNs (9-11).

We employed two independent cohorts, NCC and SEER, representing different populations and settings, in our study. This approach enhances the generalizability of our findings. Furthermore, the validation of the new cutoff value through various methods and perspectives underscores its reliability.

Nevertheless, our study has limitations inherent to retrospective analyses, including the potential for selection bias. Additionally, incomplete data in the SEER database, such as details regarding recurrence, metastasis, and specific chemoradiotherapy regimens, distance from the anal verge, circumferential resection margin status, or R0/R1/R2 resection status, may introduce bias. Important pathological and prognostic variables such as EMVI, vascular invasion, circumferential resection margin (CRM) involvement, and TRG are not available in the SEER database, which can also lead to bias. Further research should focus on prospective, multi-institutional, and large-scale clinical studies to validate 7 as the recommended minimal number of ELNs in the context of neoadjuvant therapy.

Conclusions

Our study suggests that the current minimum of 12 ELNs may not be appropriate for the rectal cancer patients who have received neoadjuvant therapy. Instead, at least 7 ELNs can more accurately assess the prognosis and guide personalized treatment decisions. Further validation through robust clinical studies is warranted to implement this recommendation effectively in clinical practice.

Acknowledgments

None.

Footnote

Reporting Checklist: The authors have completed the STROBE reporting checklist. Available at https://jgo.amegroups.com/article/view/10.21037/jgo-2025-463/rc

Data Sharing Statement: Available at https://jgo.amegroups.com/article/view/10.21037/jgo-2025-463/dss

Peer Review File: Available at https://jgo.amegroups.com/article/view/10.21037/jgo-2025-463/prf

Funding: This study was funded by Chinese Academy of Medical Sciences (CAMS) Innovation Fund for Medical Sciences (CIFMS; Nos. 2021-I2M-1-021 and 2021-I2M-C&T-A-013); and National Natural Science Foundation of China (No. 81972317).

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://jgo.amegroups.com/article/view/10.21037/jgo-2025-463/coif). All authors report that this study was funded by CAMS Innovation Fund for Medical Sciences (CIFMS; Nos. 2021-I2M-1-021 and 2021-I2M-C&T-A-013); and National Natural Science Foundation of China (No. 81972317). The authors have no other conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. The study was conducted in accordance with the Declaration of Helsinki and its subsequent amendments. The study was approved by ethics board of Cancer Hospital, Chinese Academy of Medical Sciences (No. 25/678-5625) and individual consent for this retrospective analysis was waived.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

References

1. Bray F, Ferlay J, Soerjomataram I, et al. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 2018;68:394-424. [Crossref] [PubMed]
2. Benson AB, Venook AP, Al-Hawary MM, et al. Rectal Cancer, Version 2.2018, NCCN Clinical Practice Guidelines in Oncology. J Natl Compr Canc Netw 2018;16:874-901. [Crossref] [PubMed]
3. Marks JH, Valsdottir EB, Rather AA, et al. Fewer than 12 lymph nodes can be expected in a surgical specimen after high-dose chemoradiation therapy for rectal cancer. Dis Colon Rectum 2010;53:1023-9. [Crossref] [PubMed]
4. Grega M, Vjaclovský M, Chmelová R, et al. Lymph nodes detection and the mesorectal excision quality evaluation. Rozhl Chir 2019;98:200-6. [Crossref] [PubMed]
5. Parsons HM, Tuttle TM, Kuntz KM, et al. Association between lymph node evaluation for colon cancer and node positivity over the past 20 years. JAMA 2011;306:1089-97. [Crossref] [PubMed]
6. Törnroos A, Garvin S, Olsson H. The number of identified lymph node metastases increases continuously with increased total lymph node recovery in pT3 colon cancer. Acta Oncol 2009;48:1152-6. [Crossref] [PubMed]
7. Okoshi K, Kono E, Tomizawa Y, et al. Can rectal washout reduce anastomotic recurrence after anterior resection for rectal cancer? A review of the literature. Surg Today 2020;50:644-9. [Crossref] [PubMed]
8. Kapiteijn E, Marijnen CA, Nagtegaal ID, et al. Preoperative radiotherapy combined with total mesorectal excision for resectable rectal cancer. N Engl J Med 2001;345:638-46. [Crossref] [PubMed]
9. Tsai CJ, Crane CH, Skibber JM, et al. Number of lymph nodes examined and prognosis among pathologically lymph node-negative patients after preoperative chemoradiation therapy for rectal adenocarcinoma. Cancer 2011;117:3713-22. [Crossref] [PubMed]
10. Amajoyi R, Lee Y, Recio PJ, et al. Neoadjuvant therapy for rectal cancer decreases the number of lymph nodes harvested in operative specimens. Am J Surg 2013;205:289-92; discussion 292. [Crossref] [PubMed]
11. Miller ED, Robb BW, Cummings OW, et al. The effects of preoperative chemoradiotherapy on lymph node sampling in rectal cancer. Dis Colon Rectum 2012;55:1002-7. [Crossref] [PubMed]
12. Mechera R, Schuster T, Rosenberg R, et al. Lymph node yield after rectal resection in patients treated with neoadjuvant radiation for rectal cancer: A systematic review and meta-analysis. Eur J Cancer 2017;72:84-94. [Crossref] [PubMed]
13. Nattinger AB, McAuliffe TL, Schapira MM. Generalizability of the surveillance, epidemiology, and end results registry population: factors relevant to epidemiologic and health care research. J Clin Epidemiol 1997;50:939-45. [Crossref] [PubMed]
14. Warren JL, Klabunde CN, Schrag D, et al. Overview of the SEER-Medicare data: content, research applications, and generalizability to the United States elderly population. Med Care 2002;40:IV-3-18. [Crossref] [PubMed]
15. Camp RL, Dolled-Filhart M, Rimm DL. X-tile: a new bio-informatics tool for biomarker assessment and outcome-based cut-point optimization. Clin Cancer Res 2004;10:7252-9. [Crossref] [PubMed]
16. Compton CC, Greene FL. The staging of colorectal cancer: 2004 and beyond. CA Cancer J Clin 2004;54:295-308. [Crossref] [PubMed]
17. Compton CC, Fielding LP, Burgart LJ, et al. Prognostic factors in colorectal cancer. College of American Pathologists Consensus Statement 1999. Arch Pathol Lab Med 2000;124:979-94. [Crossref] [PubMed]
18. Sobin LH, Greene FL. TNM classification: clarification of number of regional lymph nodes for pNo. Cancer 2001;92:452. [Crossref] [PubMed]
19. Dossa F, Acuna SA, Rickles AS, et al. Association Between Adjuvant Chemotherapy and Overall Survival in Patients With Rectal Cancer and Pathological Complete Response After Neoadjuvant Chemotherapy and Resection. JAMA Oncol 2018;4:930-7. [Crossref] [PubMed]
20. Kim WR, Han YD, Cho MS, et al. Oncologic Impact of Fewer Than 12 Lymph Nodes in Patients Who Underwent Neoadjuvant Chemoradiation Followed by Total Mesorectal Excision for Locally Advanced Rectal Cancer. Medicine (Baltimore) 2015;94:e1133. [Crossref] [PubMed]
21. Mekenkamp LJ, van Krieken JH, Marijnen CA, et al. Lymph node retrieval in rectal cancer is dependent on many factors--the role of the tumor, the patient, the surgeon, the radiotherapist, and the pathologist. Am J Surg Pathol 2009;33:1547-53. [Crossref] [PubMed]
22. Thorn CC, Woodcock NP, Scott N, et al. What factors affect lymph node yield in surgery for rectal cancer? Colorectal Dis 2004;6:356-61. [Crossref] [PubMed]
23. Baxter NN, Morris AM, Rothenberger DA, et al. Impact of preoperative radiation for rectal cancer on subsequent lymph node evaluation: a population-based analysis. Int J Radiat Oncol Biol Phys 2005;61:426-31. [Crossref] [PubMed]
24. Lykke J, Roikjaer O, Jess P, et al. Tumour stage and preoperative chemoradiotherapy influence the lymph node yield in stages I-III rectal cancer: results from a prospective nationwide cohort study. Colorectal Dis 2014;16:O144-9. [Crossref] [PubMed]
25. Sauer R, Becker H, Hohenberger W, et al. Preoperative versus postoperative chemoradiotherapy for rectal cancer. N Engl J Med 2004;351:1731-40. [Crossref] [PubMed]
26. Ha YH, Jeong SY, Lim SB, et al. Influence of preoperative chemoradiotherapy on the number of lymph nodes retrieved in rectal cancer. Ann Surg 2010;252:336-40. [Crossref] [PubMed]