This study recruited 299 patients undergoing a proctectomy for rectal cancer within 10 cm from the anal verge measured with preoperatively rigid sigmoidoscopy or colonoscopy at Pusan Paik Hospital from January 1995 to December 2004. We excluded 100 patients with negative lymph node metastasis, distant metastasis found at diagnosis, history of preoperative adjuvant therapy and remaining local residual cancer left. We also excluded nine patients using postoperative adjuvant chemotherapy other than 5-fluorouracil (5-FL) or not being followed up, so 190 patients were finally included in this study. All surgery was performed by one surgeon, and it was open surgery based on a total mesorectal excision. An abdominoperineal resection was performed when the tumor distance was less than 4 cm from anal verge, invasion of the pelvic floor muscle caused by the tumor was suspected during the surgery or the distal resection margin was threatened by the tumor.

A combination of chemotherapy and radiotherapy was provided postoperatively, and its indication was patients with an involvement of the circumferential resection margin. However, it was optionally done when the number of metastatic lymph nodes was one to three in the postoperative pathological examination or when the patients were over 75 years or had heart disease. The postoperative adjuvant chemotherapy started within 2-4 weeks after the surgery and was given "Mayo regimen" for 5 days every month by bolus infusing 5-FL and leucovorin six times or by orally administering 5-FL for the elderly. Radiotherapy began on the first day of the third cycle of intravenous chemotherapy and at around four weeks after the surgery for the oral administration of chemotherapy. Fractionated irradiation of the whole pelvic cavity with a dose of 4,500-5,000 cGy was conducted. When the dose of intravenously administered chemotherapy and the dose of planned radiotherapy were accomplished fully, the patient was considered to have completed chemoradiation therapy.

After assessing the proximal resection margin, the distal resection margin, the integrity of the meorectal fascia, and the metastatic lymph nodes of the resected specimen through gross examination by the surgeon in the operating room, specimen was sent to pathology. The samples were fixed in 10% formalin solution for 24 hours, and lymph nodes were collected by dissecting them through manual palpation to examine lymph nodes of the mesorectum around the primary lesion and the proximal mesorectum. Special techniques, such as the fat-clearing method, were not used in the dissection of lymph nodes. Tissues showing firm texture under digital compression during the collection of lymph nodes were thought to be lymph nodes; lymph nodes with sizes of more than 3 mm were sectioned along the longitudinal axis, and those with sizes of less than 3 mm were used as a whole for hematoxilin-eosine staining and paraffin section manufacturing.

Lymph node stages were classified into N1 and N2 based on the sixth edition of the classification of the American Joint Committee on Cancer (AJCC) by using the number of metastatic lymph nodes and into N1a, N1b, N2a and N2b based on its seventh edition. To investigate the prognosis by the LNR of the number of metastatic lymph nodes to the number of collected and examined lymph nodes, its cut-off values were determined to be 0.21, 0.32, and 0.61 through recursive partitioning [16] using R 2.2.1 (R Development Core Team, Vienna, Austria). The patients were classified into four groups with the three cut-off values: group 1 (LNR1, n = 74) for LNRs of less than 0.21; group 2 (LNR2, n = 25) for LNRs of 0.21-0.32; group 3 (LNR3, n = 56) for LNRs of 0.32-0.61; and group 4 (LNR4, n = 35) for LNRs of over 0.61.

Postoperative follow-up included physical examination and blood tests such as serum carcinoembryonic antigen every three months for first 2 years and every six months, after 2 years respectively. Radiologic examination, like abdominopelvic computerized tomography (CT), chest X-ray and chest CT, was performed every six or twelve months for the first two years and then every 12 months. If any suspicious lesion was found on radiologic examination, a bone scan and positron emission tomography were additionally performed.

Local recurrence was defined as a recurrence newly found in the pelvic cavity or the perineum in the postoperative follow-up and confirmed by biopsy or by a continuous increase of the size in the radiologic examinations conducted at intervals of 3-6 months. Systemic recurrence of the disease was defined pathologically or on the basis of radiologic findings during follow-up. If the serum carcinoembryonic antigen level became higher but there was no abnormality on radiologic examination, the patient was not considered as having a recurrence. The period to recurrence or of survival was calculated from the date of surgery for rectal cancer to the date of the last visit to the outpatient clinic or the date of death. If recurrence was not observed by the last follow-up, regardless of death, the case was considered as no recurrence. Deaths due to causes other than rectal cancer were considered as deaths in the survival analysis.

Statistical analysis was performed with SPSS ver. 12.0 (SPSS Inc., Chicago, IL, USA). The means of the groups were compared with one-way ANOVA, and chi-squared test and Fisher's exact tests were used to compare the frequencies of the four LNR groups with the clinicopathological variables. The Kaplan-Meier method was used to estimate the rates of survival and recurrence. Log rank tests were used to compare patient groups with respect to the estimated survival and recurrence curves. For independent prognostic factors, a multivariate analysis using the Cox regression model was used to determine factors influencing recurrence and death. Factors with a P-value of less than 0.1 in the univariate analysis were included in the multivariate analysis. A P-value of less than 0.05 was considered to be statistically significant. By considering that N stages of TNM staging, two groups in the sixth edition and four groups in the seventh edition, the LNRs were also classified into two and four groups, respectively, and the two groups and the four groups of the N staging and the LNR were compared in the multivariate analysis for a more objective statistical analysis.

The median age of the patients was 57.0 years (range, 29 to 81 years) and the median follow-up period was 55 months (range, 6 to 145 months). Eighteen (9.4%) patients were followed up for less than five years. The median number of lymph nodes retrieved was 12 (range, 2 to 52), and the median number of metastatic lymph nodes was 3 (range, 1 to 32) (Table 1). The number of the collected lymph nodes was inversely correlated to the LNR (r = -0.085), but the correlation was not statistically significant (P = 0.245) (Fig. 1). Although the four groups divided through recursive partitioning did not show any significant differences in comparison of rates with various clinicopathological variables, LNR4 had higher rates of N2 and N2b (P = 0.000), and LNR1 had relatively higher rates of N1 and N1a, respectively, in the sixth and the seventh editions (P = 0.000). Histologic differentiation of LNR3 and LNR4 showed significantly lower grades compared to LNR1 and LNR2 (P = 0.009), and the proportions of patients with postoperative adjuvant therapy were not different among the groups divided by LNR (P = 0.361) (Table 2).

Thirty-four (18.5%) patients had a local recurrence, and 78 (41.0%) developed systemic recurrence during the follow-up period. Sixteen patients developed both local and systemic recurrence, and 90 (42.3%) died during follow-up. In the univariate analysis, N staging in the sixth and the seventh editions, LNR and preoperative carcinoembryonic antigen level were significant factors of survival and systemic recurrence, and only N staging in the seventh edition and LNR classified into two groups were significant factors of local recurrence. N staging in the sixth edition, LNR divided into four groups and the preoperative carcinoembryonic antigen level did not affect local recurrence significantly (Table 3). The five-year survival rate and the five-year systemic recurrence rate of LNR4 were 31.3% and 61.0%, respectively, compared to those of LNR1, 72.0% and 26.0%, respectively (P = 0.001, P = 0.016). In the multivariate analysis, the risks of death and systemic recurrence were 3.9 times (95% confidence interval [CI], 2.16 to 7.16) and 2.9 times (95% CI, 1.58 to 3.33) higher in LNR4 than in LNR1, respectively, and the risk factors for death and systemic recurrence were found to be the preoperative carcinoembryonic antigen level and the LNR classified into two groups or into four groups. Neither N staging based on the sixth nor on the seventh edition were independent risk factors (Table 4). Univariate analysis revealed that there was interaction between LNR and retrieval of less than 12 lymph nodes in relation to survival and to systemic recurrence (Figs. 2, 3).