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Previous studies demonstrated that free‑floating cancer cells (FFCCs) in the lymph node sinuses were of prognostic significance for colorectal and gastric cancer. The present study investigated the clinical significance of detecting FFCCs using Fast Red staining for cytokeratin in stage I/II non‑small cell lung cancer (NSCLC) patients and hilar lymph node positive NSCLC patients who underwent curative resection. Between 2002 and 2011, a total of 164 patients (including 22 hilar lymph node positive patients) were investigated. Resected lymph nodes were stained for cytokeratin using an anti‑cytokeratin antibody. In order to achieve a clear distinction from coal dust, an anti‑cytokeratin antibody was labeled with a secondary antibody conjugated with alkaline phosphatase, which was detected by a reaction with Fast Red/naphthol that produced a red color. Patients were considered to be positive for FFCCs (FFCCs+) if one or more than one free‑floating cytokeratin‑positive cell was detected in the lymph node sinuses, which could not be detected by hematoxylin and eosin staining. Among all 164 patients, a significant difference was observed in 5‑year relapse‑free survival (5Y‑RFS) rates, with 76.9 and 33.3% being achieved by FFCCs‑ and FFCCs+ patients, respectively (P<0.001). Similarly, the 5‑year overall survival (5Y‑OS) rate was significantly lower in FFCCs+ patients, with 86.6% being achieved by FFCCs‑ and 65.8% by FFCCs+ patients, respectively (P=0.014). Among 22 hilar lymph node‑positive patients, a significant difference was also observed in 5Y‑RFS, with 53.8 and 0.0% being achieved by FFCCs‑ and FFCCs+ patients, respectively (P=0.006). The 5Y‑OS tended to be lower in FFCCs+ patients, with 69.2 and 53.3% being achieved by FFCCs‑ and FFCCs+ patients, respectively (P=0.463). The findings of the present study suggested the presence of FFCCs in stage I/II NSCLC patients was associated with a poor prognosis. In addition, FFCCs in hilar lymph node‑positive patients may potential be a useful marker in foreseeing the recurrence of cancer.

Lymph nodes from patients with colorectal cancer were immunohistochemically stained for cytokeratin to investigate the relationship between the presence of occult neoplastic cells (ONCs) and recurrence/metastasis. A total of 80 patients with stage III/Dukes' C colorectal cancer were divided into 16 patients who developed recurrence/metastasis (recurrence group) and 64 patients without recurrence (non-recurrence group). ONCs were compared between the two groups with respect to i) single cells (≥ 3 floating ONCs), ii) clusters of cells (1 or more floating aggregates of 2-20 ONCs) and iii) single cells + clusters. When single cells were detected, the sensitivity for recurrence was 87.5% (14/16, p = 0.002), the positive predictive value (PPV) was 32.6% (14/43), the specificity was 54.7% (35/64) and the negative predictive value (NPV) was 94.6% (35/37). For clusters, the sensitivity was 87.5% (14/16, p<0.001), PPV was 41.2% (14/34), specificity was 68.8% (44/64) and NPV 95.7% (44/46). With single cells + clusters, the values were 87.5% (14/16, p<0.001), 48.3% (14/29), 76.6% (49/64) and 96.1% (49/51), respectively. These results suggest that the detection of single cells + clusters is a sensitive indicator of a high risk of recurrence/ metastasis, while ONCs are useful for identifying the low-risk group of patients with stage III colorectal cancer.

Background: Occult neoplastic cells (ONCs) are the tumor cells floating in the lymph node sinuses, distant from the primary tumor, and supposed to be one of most reliable marker of prognosis., Methods: We report here the case of a 52-year-old woman with a gastric cancer associated by numerous ONCs., Results: Postoperative examination of the stomach disclosed an advanced, poorly differentiated adenocarcinoma with frequent lymph node metastases. In addition to ONCs and occasional micrometastases, focal aggregates of ONCs, one of the possible intermediate lesions between the ONCs and the usual metastases, are also observed., Conclusions: In the present case, at least some of ONCs seem to form the microaggregates of tumor cells in lymph nodes, anchor in the sinuses, and grow up to the large tumorous lesion. Even if most of the ONCs were trapped and disappeared under the influence of tumor immunity, the detection of ONCs could be one of the reliable clues to estimate the prognosis.

This study was designed to examine the relationship between occult neoplastic cells (ONCs) inside and outside harvested lymph nodes (intranodal/extranodal ONCs) and local recurrence in 30 patients who underwent curative resection of primary colorectal cancer. Among 10 patients with colon cancer (Dukes' A=1, Dukes' B=6 and Dukes' C=3), intranodal ONCs were positive in 1 patient (10.0%) and negative in 9 patients (90.0%), while extranodal ONCs were negative in all 10 patients (100.0%). There were no significant differences between the detection of intranodal or extranodal ONCs. Among 20 patients with rectal cancer (Dukes' A=4, Dukes' B=2 and Dukes' C=14), intranodal ONCs were positive in 5 (25.0%) and negative in 15 (75.0%), while extranodal ONCs were positive in 3 (15.0%) and negative in 17 (85.0%). There were no significant differences between the detection of intranodal or extranodal ONCs. These results suggest that patients with rectal cancer and extranodal ONCs should be followed-up carefully as a high-risk group for pelvic local recurrence. However, the prevalence of extranodal and intranodal ONCs was almost similar.

Among 13 patients with recurrent colorectal cancer (recurrence group: RG), immunohistochemical expression of Topo-1 was high in 4 patients (30.8%) and low in 9 patients (69.2%), while the non-recurrence group (N-RG) (n=8) showed high expression in 1 patient (12.5%) and low expression in 7 patients (87.5%) (NS). Regarding immunohistochemical expression of Bax/ERCC-1, high Bax/low ERCC-1 expression was observed in 6 patients (46.2%) from the RG and other patterns of expression were seen in 7 patients (53.8%), while high Bax/low ERCC-1 expression was observed in 4 patients (50.0%) from the N-RG and other patterns were found in 4 patients (50.0%) (NS). PCR analysis of Topo-1 expression revealed high expression in 9 patients (75.0%) from the RG (n=12) and low expression in 3 patients (25.0%), while the N-RG (n=8) showed high expression in all 8 patients (100.0%) and low expression in none (NS). With respect to ERCC-1, PCR analysis revealed high expression in 7 patients (58.3%) from the RG (n=12) and low expression in 5 patients (41.7%), while the N-RG (n=8) showed high expression in 1 patient (12.5%) and low expression in 7 patients (87.5%) (p<0.05). These results suggest that tumor sensitivity to CPT-11 and platinum derivatives is similar in stage II colorectal cancer patients with ONCs.

The sensitivity of LN metastases to anticancer drugs such as CPT-11 and platinum agents was investigated by assessing Topo-1 and Bax/ERCC-1 expression in patients who had stage III/Dukes' C colorectal cancer with ONCs. In the recurrence group (RG) (n=21), immunohistochemical expression of Topo-1 was high in 8 patients (38.1%), and low in 13 patients (61.9%), while the non-recurrence group (N-RG) (n=12) showed high expression in 1 patient (8.3%) and low expression in 11 patients (91.7%) (not significant; N.S.). Regarding the immunohistochemical expression of Bax/ERCC-1, high Bax/low ERCC-1 expression was observed in 6 patients (28.6%) from the RG and other patterns of expression were seen in 15 patients (71.4%), while high Bax/low ERCC-1 expression level was observed in 3 patients (25.0%) from the N-RG and other patterns were found in 9 patients (75.0%) (N.S.). PCR analysis of Topo-1 expression in the RG (n=13) revealed high expression in 10 patients (76.9%) and low expression in 3 patients (23.1%), while the N-RG (n=3) showed high expression in 3 patients (100%) and low expression in none (N.S.). With respect to ERCC-1, PCR analysis of the RG (n=13) revealed high expression in 6 patients (46.2%) and low expression in 7 patients (53.8%), while the N-RG (n=3) showed high expression in 2 patients (66.7%) and low expression in 1 patient (33.3%) (N.S.). These results suggest that tumor sensitivity to CPT-11 and platinum derivatives is similar in stage III colorectal cancer patients with ONCs.

This study was designed to prospectively examine whether the presence of occult neoplastic cells (ONCs) in lymph nodes or positive high-risk (HR) criteria were related to the survival of patients with stage II/III colorectal cancer or gastric cancer. The 3-year relapse-free survival (3Y-RFS) rate was calculated for 79 patients who were registered during a 2-year period. The 3Y-RFS rate was 80.5% in patients without ONCs (n=54) and 84.3% in patients with ONCs (n=25; p=0.9089). Among patients who had stage II/III colorectal cancer, it was 89.0% (n=47) and 76.2% (n=15), respectively (p=0.4131). For patients with stage III colorectal cancer alone, it was 80.8% (n=24) and 62.5% (n=9), respectively (p=0.4006). The 3Y-RFS rate was respectively 88.1% and 77.6% for the HR patients (n=31) and low-risk (LR) patients (n=48) with stage II/III colorectal cancer or gastric cancer (p=0.5545). It was respectively 92.3% and 84.3% for the HR patients (n=20) and LR patients (n=42) with stage II/III colorectal cancer (p=0.5073). Also, the rate was respectively 80% and 76.2% for the HR patients (n=7) and LR patients (n=26) with stage III colorectal cancer alone (p=0.9506). These results indicate that the 3Y-RFS rate is lower in ONC-positive patients with stage II/III colorectal cancer, suggesting that ONCs may have an influence on survival.

We studied 42 patients, consisting of 11 stage I/II patients who were node-negative (LN-) and 31 stage II/III patients who were node-positive (LN+). In the LN- patients, detection of occult neoplastic cells (ONCs) in lymph nodes had a sensitivity of 0.0% (0/5), a false-positive (FP) rate of 33.3% (2/6), a specificity of 66.7% (4/6), a false-negative (FN) rate of 100% (5/5), a positive predictive value (PPV) of 0.0% (0/2), and a negative predictive value (NPV) of 44.4% (4/9). In the LN+ patients, the sensitivity of ONCs was 25.0% (5/20), FP rate was 36.4% (4/11), specificity was 63.6% (7/11), FN rate was 75.0% (15/20), PPV was 55.6% (5/9), and NPV was 31.8% (7/22). In LN- patients, positivity for at least 2 of the 3 high-risk criteria had a sensitivity of 20.0% (1/5), FP rate of 16.7% (1/6), specificity of 83.3% (5/6), FN rate of 80.0% (4/5), PPV of 50.0% (1/2), and NPV of 55.6% (5/9). In LN+ patients, these criteria had a sensitivity of 75.0% (15/20), FP rate of 9.1% (1/11), specificity of 90.9% (10/11), FN rate of 25.0% (5/20), PPV of 93.8% (15/16), and NPV of 66.7% (10/15). These results suggest that the high-risk criteria may be useful for predicting recurrence or metastasis in stage II/III lymph node-positive patients with esophageal cancer.

This study was designed to examine the relationship between the presence of occult neoplastic cells (ONCs) in lymph nodes (LNs) and survival in 238 patients with stage I/II LN-negative cancer of the breast, lung, or stomach. In addition, immunohistochemistry for TS and DPD was used to compare the 5-FU sensitivity of the primary tumor in ONC (+) patients. The 5-year relapse-free survival (RFS) rate of 215 ONC (-) patients and 23 ONC (+) patients was 95.2 and 82.6%, respectively (p=0.0107). The 5-year overall survival (OS) rate of the ONC (-) and (+) patients was 97.4 and 77.4%, respectively (p=0.0000). The 6 ONC (+) patients with recurrence showed high and low TS expression in 33.3% (2/6) and 66.7% (4/6), respectively, while high and low DPD expression was observed in 16.7% (1/6) and 83.3% (5/6), respectively. In the 17 ONC (+) patients without recurrence, the corresponding values were 64.7% (11/17), 35.3% (6/17), 29.4% (5/17), and 70.6% (12/17). Patients with a combination of high TS and low DPD expression accounted for 33.3% (2/6) of the ONC (+) patients with recurrence and 52.9% (9/17) of those without recurrence, showing no significant difference between the two groups. These results suggest that ONCs are associated with a lower survival rate and that ONC (+) patients are unlikely to respond to 5-FU+LV therapy.

The 5-year overall survival (OS) rates for patients without occult neoplastic cells (ONCs) were 43.0% in stage II (n=15), 52.2% in stage III (n=23), and 48.5% for stages II and III combined (n=38). For ONC-positive patients, the 5-year OS rates were 44.4% in stage II (n=7; p=0.88322), 11.3% in stage III (n=30; p=0.0006), and 17.5% for stages II and III combined (n=37; p=0.0019). Among the ONC(+) recurrence group (75.7%, 28/37), 42.9% (12/28) showed high TS expression in metastatic lymph nodes and 57.1% (16/28) showed low TS expression. In the case of DPD expression, 32.1% (9/28) showed high expression and 67.9% (19/28) showed low expression. Among the ONC(+) non-recurrence group (24.3%, 9/37), 66.7% (6/9) showed high TS expression and 33.3% (3/9) showed low TS expression, while high and low DPD expression was seen in 22.2% (2/9) and 77.8% (7/9), respectively. A combination of high TS and low DPD expression was found in 32.1% (9/28) of the recurrence group vs. 66.7% (6/9) of the non-recurrence group (p=0.070). These results suggest that ONCs are associated with OS. Unlike the non-recurrence group, the ONC(+) patients with recurrence of stage II/III node-positive gastric cancer are unlikely to respond to treatment with 5-FU + LV and may need combination chemotherapy based on L-OHP and/or CPT-11.

Many studies have shown a close relationship between the detection of occult neoplastic cells (ONCs) freely floating in lymph node sinuses remote from the primary tumor and the recurrence/metastasis of various malignancies. If 5 to 10 circulating ONCs form an aggregate of tumor cells, it may easily escape attack by the host immune system. Since such malignant micro-aggregates cannot pass through the circulation in the liver and lungs, which filter tumor cells from the blood, the aggregates may become anchored in these organs and cause metastasis. A hypothesis is proposed that the most important factor leading to distant metastasis/recurrence of stage I or II cancer without microscopic lymph node metastasis, so-called N0 local disease (N0-LD), is the release of malignant micro-aggregates consisting of many ONCs from the primary tumor. Distant metastasis/recurrence of N0-LD may occur when such infiltrating/proliferating cells enter target organs, especially the liver and lungs, in a viable state during their first pass through the circulation preoperatively and/or intraoperatively.

A 57-year-old woman presented to our hospital with macrohematuria. Renal cell carcinoma (RCC) of the left kidney with bilateral pulmonary and mediastinal lymph node metastases was diagnosed. Radical nephrectomy was performed. Pathological examination confirmed the diagnosis of RCC (pT2pN0M1), so immunotherapy was performed using interleukin-2 (IL-2). The mediastinal lymph node metastases disappeared completely after 5 months of immunotherapy. Investigation of immunological parameters showed an increase of CD8-positive T cells (CD8) and a decrease of CD14-positive cells (CD14), leading to a marked increase of the CD8/CD14 ratio. Although her pulmonary metastases are unchanged, the mediastinal metastases still show complete remission at 14 months postoperatively. The CD8/CD14 ratio may have potential as a new marker for monitoring the response of RCC to immunotherapy and selecting suitable patients for such therapy.

The correlation between detection of occult neo-plastic cells (ONCs) in lymph node sinuses and the recurrence/metastasis of primary breast, lung, esophageal, and gastric cancer was examined. Among patients with Stage I-III cancer treated by radical resection with dissection of at least 10 lymph nodes, 40 patients who suffered recurrence/metastasis within 5 years post-operatively and 94 patients who did not have recurrence within 5 years were randomly selected. A total of 1,340 lymph nodes were subjected to immunohistochemical staining for cytokeratin to identify ONCs. Then the sensitivity, specificity, positive predictive value, and negative predictive value of ONCs were determined for predicting the recurrence of each cancer. These parameters were respectively 40.0, 75.9, 62.4, and 55.8% for breast cancer, while the respective values were 50.0, 77.4, 68.9, and 60.8% for lung, 57.1, 64.3, 61.5, and 60.0% for esophageal, and 68.8, 65.0, 66.3, and 67.5% for gastric cancer. All of the parameters exceeded 65% for gastric cancer. ONCs also showed a high specificity for breast and lung cancer, but both the sensitivity and specificity were low for esophageal cancer.