Loss of p53 in esophageal squamous cell carcinoma and the correlation with survival: analyses of gene mutations, protein expression, and loss of heterozygosity in Japanese patients.

BACKGROUND: A high frequency of p53 protein expression or gene mutation has been reported in the early stages of esophageal squamous cell carcinoma (ESCC), and thus loss of p53 function is thought to be very important in esophageal carcinogenesis. However, there is controversy surrounding the correlation between p53 dysfunction and ESCC tumor progression. The complexity arises from the different modalities, such as mutation analysis, immunohistochemistry, and the detection of loss of heterozygosity (LOH) at the p53 genomic locus. METHODS: In this study, we comprehensively analyzed p53 gene mutation, p53 protein expression, and LOH at 17p13 in 94 surgically resected Japanese cases of ESCC. RESULTS: The frequency of p53 gene mutation was 60.6%. The rate of positive p53 protein expression was 56.4%. The frequency of LOH at 17p13 was 67.5%. There was a statistically significant correlation between the presence of a gene mutation and LOH, whereas, there was no significant correlation between gene mutation and protein expression. CONCLUSIONS: Despite the importance of loss of p53 function in esophageal carcinogenesis, none of the examined parameters, either singly or combined, correlated with overall survival. Taken together, p53 function is a primary target for esophageal carcinogenesis but there is no apparent correlation with the malignant phenotype in ESCC.

Recent results of therapy for scirrhous gastric cancer.

The prognosis of patients with scirrhous gastric cancer (SGC) is extremely poor. However, recent advances in therapeutic strategies against SGC, using effective anticancer drugs, have prolonged the survival of patients with SGC. This paper reviews the recent therapeutic outcomes of this type of gastric cancer and introduces a new treatment protocol for SGC.

Free jejunal graft reconstruction after resection of neck cancers: our surgical technique.

Locally advanced carcinomas arising in the hypopharynx, cervical esophagus, or thyroid are traditionally treated by resection of the hypopharynx and cervical esophagus. Various methods of reconstruction aiming to achieve safety and functionality have been reported, including the myocutaneous flap and the free jejunal graft. With advances in microscopic surgery, the free jejunal transplant is now used dominantly; however, this procedure is not without major risks. In this review we examine the short- and long-term complications of this procedure. We also describe our technique of free jejunal graft reconstruction after pharyngoesophagectomy and total laryngectomy with definitive tracheostomy. We used free jejunal graft reconstruction after resection with hypopharyngeal cancer or thyroid cancers in 22 patients. Twenty-one of these patients acquired a good quality of life, but one died after loss of the jejunal graft. Thus, using a free jejunal graft for reconstruction of the hypopharynx or cervical esophagus can be very useful in improving the quality of life of patients.

Protein and gene expression of tumor-associated antigen RCAS1 in esophageal squamous cell carcinoma.

RCAS1 (receptor-binding cancer antigen expressed on SiSo cells) is expressed on the tumor cell membrane and induces apoptosis on infiltrated immune lymphocytes. RCAS1 has been reported to correlate with the escape of tumor cells from host immune surveillance, and with poor prognosis. However, the clinical importance of RCAS1 protein and gene expression in esophageal squamous cell carcinoma (ESCC) has not been well investigated. In the present study, RCAS1 gene and protein expression levels were evaluated and compared with clinical findings in 67 patients with ESCC. Expression levels of RCAS1 and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) messenger RNAs (mRNAs) from tumors and non-cancerous epithelia were analyzed quantitatively by real-time reverse transcriptase polymerase chain reaction (RT-PCR). RCAS1 protein expression was analyzed by immunohistochemistry. The mean RCAS1/GAPDH ratio of tumors (0.7) was not different from that of non-cancerous epithelia (0.7, p=0.715). RCAS1 immunoreactivity was detected in 19 tumors (28.4%). The mean RCAS1/GAPDH ratio of tumors with RCAS1 protein positive (0.6) did not differ from tumors without RCAS1 expression (0.8, p=0.131). RCAS1 gene and protein expressions did not correlate with tumor size, depth of invasion, lymph node metastasis, or patient prognosis. Thus, RCAS1 gene or protein expression may not correlate with tumor progression in ESCC.

Detection of cancer cells in the peripheral blood of gastric cancer patients.

The existence of occult metastasis in peripheral blood has been reported in various tumors. However, in gastric cancer (GC), this metastasis has not been well analyzed. In the present study, to identify circulating cancer cells in patients with GC, peripheral blood samples from GC patients were investigated. Total RNA was extracted from 1.5 ml peripheral blood from 55 patients with GC, from 34 non-cancer patients, and from 10 healthy volunteers. Carcinoembryonic antigen (CEA), cytokeratin 19 (CK19), and 20 (CK20) messenger RNA (mRNA) were used as probes to detect GC cells in the blood samples using real-time reverse transcriptase polymerase chain reaction (RT-PCR). CEA and CK19 mRNA expression were not detected in the 40 healthy volunteers and non-cancer patients, while 2 of the 40 showed CK20 mRNA expression. In 55 patients with GC, CK19 mRNA was not detected and CEA mRNA was detected in only one case (1.8%) with stage IV. While CK20 mRNA expression was observed in 15 cases (27.3%) and even in stage I, 8 of 24 (33.3%) showed CK20 mRNA expression. Thus, the specificity of CK20 marker may be low. Even though the sensitivity of CEA marker is low, CEA may be a more reliable marker than cytokeratins for detection of cancer cells in GC patient's peripheral blood.

Expression of the murine double minute gene 2 oncoprotein in esophageal squamous cell carcinoma as a novel marker for lack of response to chemoradiotreatment.

The protein product of the murine double minute gene 2 (MDM2) negatively controls the work of the p53 protein and the retinoblastoma protein (pRB). Recent studies have found that MDM2 expression correlates with chemoresistance of tumor cells. In the present study, the correlation between MDM2 expression and chemoradioresistance was evaluated in patients with esophageal squamous cell carcinoma (ESCC). The immunoreactivities of p53, pRB, and MDM2 were evaluated in 148 surgically resected ESCC by using monoclonal antibodies. The disease-free survival of 107 surviving patients who underwent curative surgery was compared among groups with positive and negative expressions of p53, pRB, and MDM2. High immunoreactivities of p53, pRB, and MDM2 were detected in 47.3%, 64.2%, and 32.4% of cases, respectively. In 107 patients undergoing curative surgery, pRB losses and MDM2 overexpression were found to be poor prognostic factors by univariate analysis. In multivariate analysis, only MDM2 expression was determined to be a prognostic factor independent of the tumor stage. Moreover, we found that MDM2 expression correlates with short survival of patients undergoing postoperative adjuvant chemoradiotherapy. In patients without adjuvant therapy, however, the MDM2 status did not correlate with patient survival. The present results indicate that MDM2 expression may be not only a prognostic marker for patients with ESCC, but also a novel marker for predicting a lack of response to chemoradiotreatment.

Lymph node metastasis of gastric cancer: comparison of Union International Contra Cancer and Japanese systems.

BACKGROUND: The pN classification of gastric cancer (GC) in the Japanese system (Japanese Gastric Cancer Association; JGCA) is based on the site and distance of metastatic nodes from the primary tumour. Union International Contra Cancer (UICC) has recently proposed a classification system based on the number of nodes involved (TNM-1997). The aim of the present study is to assess which classification system is more suitable for providing a prognosis in advanced GC with lymph node metastasis. METHODS: A total of 224 patients who underwent curative gastrectomy (R0: UICC-TNM and Resection A and B: JGCA) and D2 lymphadenectomy between 1990 and 1999, and diagnosed as pT2, pT3 and pT4 GC were enrolled. Patients were followed until the end of 2002. The disease-free survival rates of patients were compared between the two-stage systems (UICC-TNM and JGCA). RESULTS: Using the JGCA system, there was a significant difference between the two survival curves (pN0 and pN1, P = 0.025; pN1 and pN2, P < 0.001; pN2 and pN3, P = 0.031), but the 5-year survival rate of 27 pN2 patients (32.7%) was not significantly different from that of 14 pN3 patients (34.3%, P = 0.994) using the UICC-TNM. In 47 patients with JGCA pN2, the 5-year survival rate of 18 patients with UICC-TNM pN1 (42.9%) was not significantly different from that of 18 patients with UICC-TNM pN2 (25.2%, P = 0.422) or from that of 11 patients with UICC-TNM pN3 (24.2%; P = 0.383). CONCLUSIONS: The JGCA system is more suitable for estimating the prognosis of Japanese patients with advanced GC than the UICC-TNM.

Impact of dendritic cell vaccines pulsed with Wilms' tumour-1 peptide antigen on the survival of patients with advanced non-small cell lung cancers.

PURPOSE: Dendritic cell (DC)-based vaccines have been expected to serve as new therapeutic approaches for advanced non-small cell lung cancers (NSCLCs); however, their clinical outcomes have not been fully elucidated. We report a single-centre clinical study analysing factors affecting the survival of patients with advanced NSCLCs who received DC vaccines pulsed with or without Wilms' tumour protein-1 (WT1) peptide. METHODS: Among 62 patients with previously treated inoperable or postoperatively relapsed NSCLCs who met the inclusion criteria, DCs from 47 (76%) patients who showed HLA-A2402/0201/0206 were pulsed with one or more corresponding WT1 peptide antigens. DC vaccines were intradermally injected biweekly. RESULTS: Clinical responses based on response evaluation criteria in solid tumours (RECIST) were found in 31 (50%) patients at 3 months after the first DC vaccine (complete response: 1 (1.6%), partial response: 4 (6.5%), stable disease: 26 (41.9%)). Median survival time was 27 months (82% in 1 year and 54% in 2 years) from initial diagnosis, and that was 12 months (48% in 1 year and 22% in 2 years) from the first DC vaccination. Importantly, multivariate analyses revealed that only two factors, blood haemoglobin and the use of WT1 peptides, significantly affected the overall survival of patients from both initial diagnosis and first vaccination. CONCLUSIONS: This study is the first to suggest that DC vaccines pulsed with WT1 may hold a significant impact to prolong the overall survival of patients with advanced NSCLCs.

Dendritic cell-based immunotherapy targeting synthesized peptides for advanced biliary tract cancer.

BACKGROUND: The aim of this retrospective study was to clarify the safety and efficacy of dendritic cell (DC)-based immunotherapy targeting synthesized peptides, Wilms tumor 1 (WT1) and Mucin 1, cell surface associated (MUC1) for biliary tract cancers (BTCs). METHODS: Sixty-five patients who had nonresectable, recurrent, or metastatic BTCs and received the DC-based immunotherapy were selected for the study. DCs were pulsed with WT1 and/or MUC1. The adverse events (AEs) and clinical responses were examined. RESULTS: No serious treatment-related AEs were observed. Median survival time (MST) from diagnosis and from the first vaccination was 18.5 and 7.2 months, respectively. By multivariate Cox proportional hazard analysis, the significant independent factors were found to be (1) combined chemotherapy, (2) albumin level ≥4.0 g/dL before vaccination, (3) C-reactive protein level <0.5 mg/dL before vaccination, and (4) fever after vaccination. The MST from the first vaccination with or without chemotherapy was 8.2 and 5.3 months, respectively (P = 0.016), and MST for the patients with prognostic nutritional index ≥40 and <40 was 8.1 and 5.0 months, respectively (P = 0.023). CONCLUSIONS: Although a small uncontrolled nonrandomized study, DC-based immunotherapy for BTCs was safe and produced a clinical response for the patients who underwent chemotherapy and maintained a good nutrition status.

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

BACKGROUND: The 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. METHODS: Total 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. RESULTS: In 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. CONCLUSION: The 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.

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-beta (TGF-beta), 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.

Quantitative analysis of heparanase gene expression in esophageal squamous cell carcinoma.

BACKGROUND: Heparan sulfate proteoglycans, the main components of the extracellular matrix, are recognized as important components of signal transduction and play an important role in tumor progression. Heparanase (hep) degrades heparan sulfate proteoglycans, but the clinical importance of hep is unclear. In this study, we investigated the clinicopathologic importance of hep messenger RNA (mRNA) expression in esophageal squamous cell carcinoma (ESCC). METHODS: Fresh tumors and noncancerous epithelia were obtained from 57 ESCC patients after esophagectomy. Expression levels of hep and glyceraldehyde-3-phosphate dehydrogenase mRNA were quantitatively analyzed by real-time reverse transcriptase-polymerase chain reaction. Apoptotic cancer cells and microvessel density were evaluated immunohistochemically. RESULTS: The relative hep mRNA expression level (hep:glyceraldehyde-3-phosphate dehydrogenase ratio) in ESCC was lower than in noncancerous tissue (P <.001). Tumor hep expression decreased according to tumor progression and correlated with the occurrence of apoptotic cancer cells, but not with tumor microvessel density. Moreover, low hep expression correlated with poor patient survival. CONCLUSIONS: Reduced hep mRNA expression might result in abnormal cell growth and correlate with ESCC progression.

Gene expression levels of cytokines in peritoneal washings from patients with gastric cancer.

Peritoneal seeding is frequently detected in patients with gastric cancer. The peritoneal cavity is a compartment in which the immunologic host-tumor interaction can occur. Here, we investigated the gene expression levels of cytokines, and compared these gene expressions with the progression of gastric cancer. Total RNA was extracted from 50 ml of peritoneal washings from 78 patients with gastric cancer and 11 noncancerous patients. The gene expression levels of transforming growth factor-beta (TGF-beta), interleukin (IL)-2, IL-6 and IL-12 were analyzed by real-time reverse transcription-polymerase chain reaction. The gene expression levels of TGF-beta and IL-12 in 16 patients with peritoneal seeding (peritoneal metastasis or free cancer cells) did not differ from those in controls (n = 11) and stage I and II (n = 43) patients. However, the relative gene expression level of IL-2 in patients with peritoneal seeding (0.9) was lower than that in controls (1.4, p = 0.066) and stage I and II patients (1.4, p = 0.036). In contrast, the relative gene expression level of IL-6 in patients with peritoneal seeding (3.3) was higher than that in control (2.4, p = 0.064) and stage I and II patients (2.6, p = 0.065). Low IL-2 gene and high IL-6 gene expressions in the peritoneal cavity may correlate with cancer development in the peritoneal cavity in patients with gastric cancer.