Masahide Ikeguchi

Prognostic Significance of Level and Number of Lymph Node Metastases in Patients with Gastric Cancer

BackgroundTo present data that provide some insight into the appropriateness of a nodal grouping category and its relation to survival in patients with gastric cancer.MethodsWe reviewed data of 777 patients with advanced gastric cancer who had undergone curative gastrectomy to investigate the prognostic significance of level and number of lymph node metastases.ResultsThe prognosis of patients with gastric cancer was well correlated with the level and number of lymph node metastases. Multivariate analysis indicated that the level and number of lymph node metastases were independent prognostic indicators. Moreover, the number of lymph node metastases was an independent prognostic factor in N1, N2, and N3 patients. The most statistically significant difference in disease-specific survival was observed at a threshold of 11 lymph node metastases, yielding a χ2 value of 42.88, a hazard ratio of 2.523, at a 95% confidence interval of 1.913, 3.329 (Pxa0<xa0.0001) by Cox proportional hazard model. On the basis of this result, patients were divided into two groups as follows: marked lymph node metastasis group (number of positive nodes ≥11) and slight lymph node metastasis group (number of positive nodes ≤10). The prognosis of patients with marked lymph node metastasis was statistically significantly worse than that with slight lymph node metastasis in N1, N2, and N3 patients.ConclusionsBoth level and number were indispensable for evaluating lymph node metastasis. Therefore, addition of the number of positive nodes to the N category defined by the Japanese Classification of Gastric Carcinoma may be a useful strategy in the N staging classification in gastric cancer.

Detection of Circulating Cancer Cells After a Gastrectomy for Gastric Cancer

PurposeIn this study, we evaluated the correlation between the postoperative detection of circulating cancer cells and the risk of recurrence in patients with gastric cancer.MethodsTotal RNA was extracted from 1.5u2009ml of peripheral blood from 59 patients with gastric cancer and 15 patients with cholecystolithiasis (control) before and after operation. Carcinoembryonic antigen (CEA) messenger RNA (mRNA) was used as a probe to detect gastric cancer cells in samples using a real-time reverse transcription–polymerase chain reaction (RT-PCR).ResultsCarcinoembryonic antigen mRNA-positive cells were not found in the peripheral blood of the control patients either before or after operation, nor in the peripheral blood of the gastric cancer patients before operation. However, CEA and mRNA-positive cells were detected in 46% of the patients just after a gastrectomy, though these circulating cancer cells disappeared from peripheral blood within 2 postoperative days. In 55 patients who underwent a curative operation, the risk for cancer recurrence (10/30; 33%) in 30 patients who did not show circulating cancer cells postoperatively was higher than that for cancer recurrence (3/25; 12%) in 25 patients with positive for circulating cancer cells (P = 0.064). As a result, the presence of blood circulating tumor cells just after surgery tends to correlate with a low rate of tumor recurrence in patients operated on for gastric cancer.ConclusionThese findings indicate that a gastrectomy may spread gastric cancer cells into the peripheral blood from primary tumors; however, such circulating cancer cells may be destroyed within a short time. The detection of circulating cancer cells may therefore be a marker for a possibly better prognosis in patients with gastric cancer.

Decreased NKG2D Expression on CD8+ T Cell Is Involved in Immune Evasion in Patients with Gastric Cancer

Purpose: Some studies suggest that the immunoreceptor NKG2D expression on CD8+ T cells is down-regulated and this reduction may be involved in immune evasion in cancer patients. The present study was designed to investigate NKG2D expression on CD8+ T lymphocytes and its relationship to immune evasion in gastric cancer patients. Experimental Design: NKG2D expression on both circulating and tumor-infiltrating CD8+ T cells was evaluated by multicolor flow cytometry. Soluble MHC class I chain-related gene A (MICA) in the sera was quantitated by ELISA. Transwell experiments were carried out to determine the effect of cancer cells on NKG2D expression. Results: NKG2D expression on circulating CD8+ T cells was down-regulated and significantly correlated with IFN-γ production in gastric cancer patients (r = 0.68; P = 0.007). NKG2D expression was closely related to undifferentiated cancer (P = 0.021) as was the depth of invasion (P = 0.012). There was no difference in soluble MICA between gastric cancer patients and normal controls. NKG2D expression on CD8+ T cells was remarkably reduced in the tissue of gastric cancer compared with peripheral blood (P = 0.046). Complete removal of tumor by surgery restored NKG2D expression on CD8+ T cells (P = 0.0049). Transwell experiments showed that this down-regulation was induced by direct contact between cancer cells and CD8+ T cells and that soluble factors did not affect the NKG2D expression. This phenomenon was blocked by the addition of anti-MICA antibodies. Conclusions: Decreased NKG2D expression may be one of the key mechanisms responsible for immune evasion by tumors in gastric cancer.

Interleukin-2 Gene Expression Is a New Biological Prognostic Marker in Hepatocellular Carcinomas

Background: Cytokines produced by tumor cells and tumor-infiltrating lymphocytes (TIL) appear to regulate tumor cell growth. The present study analyzed the correlation between local immune responses and cytokine gene expression levels in patients with primary hepatocellular carcinoma (HCC). Material and Methods: The gene expression levels of interleukin-2 (IL-2) and -12 (IL-12) were evaluated quantitatively by real-time reverse transcriptase polymerase chain reaction (RT-PCR) and compared with the density of CD8+ TIL detected by immunohistochemistry in 59 surgically resected HCCs. Results: IL-2 gene expression was detected in 33 (56%) and IL-12 gene expression in 39 (66%) of 59 HCCs. Tissueinfiltrating CD8+ T lymphocytes in tumors were significantly suppressed compared with noncancerous liver tissues. The CD8+ T cell density of tumors with IL-2 gene expression was higher than that of tumors without IL-2 gene expression (p = 0.002). However, such a correlation was not detected in tumors with or without expression of IL-12 genes. Patients with IL-2 positive tumors had a favorable prognosis. IL-2 gene expression was detected as an important prognostic factor independent of tumor stage. Conclusions: These findings indicate that in HCCs, tumor-infiltrating CD8+ T cells might be activated by IL-2 produced by TIL and IL-2 gene expression in tumors may be an important prognostic biomarker in HCC.

Prognostic significance of RCAS1 expression in relation to the infiltration of dendritic cells and lymphocytes in patients with esophageal carcinoma.

RCAS1 (receptor-binding cancer antigen expressed on SiSo cells) expression was determined in 107 esophageal carcinoma patients by immunohistochemical procedures and compared with tumor infiltrating lymphocyte (TIL) and dendritic cell (DC) infiltration to evaluate the effect of RCAS1 on immune responses in esophageal carcinoma. RCAS1 immunoreactivity was detected in 59 of 107 patients (55.1%). RCAS1 expression was significantly correlated with the depth of invasion, lymph node metastasis, and histologic stage. RCAS1 expression tended to be correlated with a lower TIL density in tumors with marked DC infiltration. The survival time for patients with RCAS1-negative tumors was significantly longer than that for patients with RCAS1-positive tumors. Especially, the prognosis was predicted by RCAS1 in cases with marked DC infiltration. Multivariate analysis revealed that RCAS1 expression was an independent prognostic factor. RCAS1 expression may play an important role in evading the immunological defense mechanisms in esophageal carcinoma.

Laparoscopic-assisted intraperitoneal chemotherapy for patients with scirrhous gastric cancer.

Background: The prognosis for patients with scirrhous gastric cancer (SGC) is extremely poor. To improve the patients’ prognosis, laparoscopic-assisted intraperitoneal chemotherapy (IPC) was introduced for SGC. In this study, we analyzed whether IPC reduced the number of cancer cells in the peritoneal cavity of patients or changed the gene expression levels of cytokines in the peritoneal cavity. We also investigated whether IPC improved the prognosis of patients with SGC. Methods: Total RNA was extracted from 50 ml of peritoneal wash from 11 SGC patients before and after cisplatin-based IPC. The gene expression levels of survivin, c-myc, transforming growth factor-β (TGF-β), interleukin-2 (IL-2), IL-6, and IL-12 were analyzed using real-time reverse transcription-polymerase chain reaction (RT-PCR) assays. Also, carcinoembrionic antigen (CEA) messenger RNA (mRNA) was used to identify the number of gastric cancer cells in peritoneal washes by the real-time RT-PCR method. The gene expression levels of cytokines and the number of cancer cells in the peritoneal cavity were compared before and after cisplatin-based IPC treatment. Results: Before IPC, the gene expression of IL-2 from peritoneal washes of patients was significantly suppressed compared to the controls (p = 0.029); however, other gene expression levels did not differ. In 7 cases, more than 90% of the cancer cells were removed from the peritoneal cavity after cisplatin-based IPC. These 7 cases were named the IPC effective group, and the remaining 4 cases were named the IPC ineffective group. In the IPC effective group, elevated IL-2 and IL-6 genes were detected in 5 (71%) and in 6 (86%) after IPC. The correlation between IPC effectiveness and elevated gene expression after IPC (IL-2: p = 0.137, and IL-6: p = 0.044) was observed. However, the 50% survival period of the IPC effective group (9 months) was not different from that of that of the IPC ineffective group (6 months, p = 0.267). Conclusion: IPC effectiveness may correlate with elevation of gene expression of inflammatory cytokines, such as IL-2 and IL-6 in the peritoneal cavity after IPC. However, the prognostic benefits of IPC for SGC patients remain unclear.

Allogeneic gastric cancer cell-dendritic cell hybrids induce tumor antigen (carcinoembryonic antigen) specific CD8+ T cells

The development of protocols for the ex vivo generation of dendritic cells (DCs) has led to intensive research of their potential use in immunotherapy. Accumulating results show the efficacy of this treatment on melanomas which are highly immunogenic. However, its efficacy remains unclear in other tumors. In this study, allogeneic gastric cancer cell–DC hybrids were used to determine the efficacy of this type of immunotherapy in gastric cancer. Fusion cells of DC and allogeneic gastric cancer cells were generated by polyethylene glycol (PEG) and electrofusion. These hybrids were used to induce tumor associated antigen (TAA) specific cytotoxic T lymphocytes (CTLs). The DCs were successfully fused with the allogeneic gastric cancer cells resulting in hybrid cells. These hybrid cells were functional as antigen-presenting cell because they induced allogeneic CD4+ T cells proliferation. CD8+ T cells stimulated by the MKN-45-DC hybrid cells were able to kill MKN-45 when used for immunization. The CTLs killed another gastric cancer cell line, MKN-1, as well as a melanoma cell line, 888mel, suggesting the recognition of a shared tumor antigen. MKN-45 specific CTLs can recognize carcinoembryonic antigen (CEA), indicating that the killing is due to tumor antigens as well as alloantigens. This approach suggests the possible use of allogeneic gastric cancer cell–DC hybrids in DC based immunotherapy for gastric cancer treatment.

Detection of cancer cells and gene expression of cytokines in the peritoneal cavity in patients with gastric cancer

BackgroundThe gene expression of the cytokines interleukin-2 (IL-2) and IL-10 in peritoneal washings was examined in relation to the presence of cancer cells in the peritoneal cavity in patients with gastric cancer.MethodsTotal RNA was extracted from 50-ml peritoneal wash samples from 124 patients (gastric cancer, n = 110; controls, n = 14). Carcinoembrionic antigen (CEA) messenger RNA (mRNA) was used to identify the number of cancer cells in peritoneal wash samples by a real-time reverse transcription-polymerase chain reaction (RT-PCR) method, which method was also used to assay the IL-2 and IL-10 gene expression levels.ResultsIn the 14 control samples, CEA mRNA was not detected, while CEA mRNA was detected in 2 of the 51 stage I gastric cancer patients. Thus, the specificity of this method for the detection of cancer cells in peritoneal wash samples was 97% (63/65). The CEA-based real-time RT-PCR method demonstrated greater prognostic impact than the traditional cytological method. IL-2 gene expression in peritoneal wash samples that were CEA mRNA-positive was suppressed compared with that in peritoneal wash samples that were CEA mRNA-negative, while IL-10 gene expression did not differ according to the CEA mRNA findings.ConclusionThe detection of small numbers of cancer cells in peritoneal wash samples from patients with advanced gastric cancer is a good marker for peritoneal metastatic recurrence. In the peritoneal cavity, cancer cells may escape from immune surveillance by controlling the expression of cytokines.