Seiya Saito

MicroRNA-200b Regulates Cell Proliferation, Invasion, and Migration by Directly Targeting ZEB2 in Gastric Carcinoma

BackgroundThe microRNA-200 (miR-200) family has been reported to induce epithelial differentiation and suppress epithelial–mesenchymal transition (EMT) by inhibiting translation of zinc finger E-box-binding homeobox (ZEB) 1 and 2 mRNAs in several types of cancers. This study aimed to clarify the role of miR-200b in regulating EMT and promoting cellular proliferation, invasion, and migration in gastric cancer.MethodsThe relationships among the expression levels of miR-200b, ZEB1 and ZEB2, and E-cadherin mRNAs were analyzed by quantitative real-time reverse transcription–polymerase chain reaction in frozen tissue samples from 40 gastric cancer patients who underwent gastrectomy from 2008 to 2010. The effects of miR-200b on EMT in gastric cancer cells in vitro were also analyzed.ResultsDiffuse histologic type, depth of tumor, tumor size, lymph node metastasis, and lymphatic invasion were significantly higher in the low-miR-200b expression group compared with the high expression group. There was a strong correlation between the levels of miR-200b, and ZEB2 and E-cadherin mRNAs in gastric cancer patients. Upregulation of miR-200b in gastric cancer cells changed the cell morphology from fibroblast- to epithelial-like, associated with localization of E-cadherin to the plasma membrane. ZEB2 mRNA levels fell, while E-cadherin expression levels increased in gastric cells overexpressing miR-200b, associated with significantly reduced cellular proliferation, and inhibition of cellular migration and invasion.ConclusionsmiR-200b regulates ZEB2 expression and thus controls metastasis in gastric cancer.

Overexpression of microRNA-223 regulates the ubiquitin ligase FBXW7 in oesophageal squamous cell carcinoma

Background:F-box and WD repeat domain-containing 7 (FBXW7) is a cell cycle regulatory gene whose protein product ubiquitinates positive cell cycle regulators such as c-Myc, cyclin E, and c-Jun, thereby acting as a tumour-suppressor gene. This study focused on microRNA-223 (miR-223), which is a candidate regulator of FBXW7 mRNA. The aim of this study was to clarify the clinical significance of miR-223 and FBXW7 in oesophageal squamous cell carcinoma (ESCC) patients, and to elucidate the mechanism by which FBXW7 is regulated by miR-223.Methods:The expression levels of miR-223 and the expression of FBXW7 protein was examined using 109 resected specimens to determine the clinicopathological significance. We also investigated the role of miR-223 in the regulation of FBXW7 expression in ESCC cell lines in an in vitro analysis.Results:We found that miR-223 expression was significantly higher in cancerous tissues than in the corresponding normal tissues. There was a significant inverse relationship between the expression levels of miR-223 and FBXW7 protein. Moreover, patients with high miR-223 expression demonstrated a significantly poorer prognosis than those with low expression. On the basis of a series of gain-of-function and loss-of-function studies in vitro, we identified FBXW7 as a functional downstream target of miR-223.Conclusion:Our present study indicates that high expression of miR-223 had a significant adverse impact on the survival of ESCC patients through repression of the function of FBXW7.

Serum microRNA-21 is a novel biomarker in patients with esophageal squamous cell carcinoma

It was recently revealed that microRNAs (miRNAs) can stably exist in serum and may affect the pathogenesis of several diseases. However, there are few reports that have demonstrated the significance of miRNAs in the serum of patients with esophageal squamous cell carcinoma (ESCC). Thus, the aims of this study were to clarify the status of miRNA‐21 in serum of ESCC patients and to reveal the usefulness of this molecule as a biomarker.

Induction chemotherapy with docetaxel/cisplatin/5-fluorouracil for patients with node-positive esophageal cancer.

Background: Despite improvements in the surgical management of esophageal cancer, the prognosis of patients with lymph node metastases is still unsatisfactory. Recently, survival benefit of neoadjuvant or induction chemotherapy for patients with esophageal cancer has been highlighted. Methods: Efficacy and toxicity of induction chemotherapy for esophageal cancer were reviewed. In addition, our experience on modified docetaxel/cisplatin/5-FU (DCF) as induction chemotherapy was also demonstrated. The modified DCF consisted of 60 mg/m2 of docetaxel on day 1, and 350 mg/m2 of 5-FU and 6 mg/m2 of cisplatin on days 1–5. Two courses have been administered as induction chemotherapy in 51 patients with node-positive esophageal cancer. Response was evaluated by RECIST v1.0 and changes in standardized uptake value by 18F-fluorodeoxyglucose positron emission tomography. Results: Induction chemotherapy may be beneficial for node-positive esophageal cancer, although the consensus has not yet been established. A regimen of induction chemotherapy should have a high response rate and cisplatin/5-FU may be underpowered as an induction setting. DCF can be a candidate for the regimen of induction chemotherapy for esophageal cancer, although severe adverse events have been reported. Several modified regimens to reduce the toxicity have been reported. The response rate of our series was 61% and a significant decrease in standardized uptake values was observed after the induction chemotherapy. Although high-grade neutropenia was still observed with this regimen, neither treatment-related death nor delay in the following treatment was observed. Conclusions: Modified DCF can be a regimen of induction chemotherapy for node-positive esophageal cancer because of its high efficacy, although an adequate care for severe neutropenia is needed.

CD44s signals the acquisition of the mesenchymal phenotype required for anchorage-independent cell survival in hepatocellular carcinoma

Background:Circulating tumour cells (CTCs) have an important role in metastatic processes, but details of their basic characteristics remain elusive. We hypothesised that CD44-expressing CTCs show a mesenchymal phenotype and high potential for survival in hepatocellular carcinoma (HCC).Methods:Circulating CD44+CD90+ cells, previously shown to be tumour-initiating cells, were sorted from human blood and their genetic characteristics were compared with those of tumour cells from primary tissues. The mechanism underlying the high survival potential of CD44-expressing cells in the circulatory system was investigated in vitro.Results:CD44+CD90+ cells in the blood acquired epithelial–mesenchymal transition, and CD44 expression remarkably increased from the tissue to the blood. In Li7 and HLE cells, the CD44high population showed higher anoikis resistance and sphere-forming ability than did the CD44low population. This difference was found to be attributed to the upregulation of Twist1 and Akt signal in the CD44high population. Twist1 knockdown showed remarkable reduction in anoikis resistance, sphere formation, and Akt signal in HLE cells. In addition, mesenchymal markers and CD44s expression were downregulated in the Twist1 knockdown.Conclusions:CD44s symbolises the acquisition of a mesenchymal phenotype regulating anchorage-independent capacity. CD44s-expressing tumour cells in peripheral blood are clinically important therapeutic targets in HCC.

Usefulness of Transcription–Reverse Transcription Concerted Reaction Method for Detecting Circulating Tumor Cells in Patients With Colorectal Cancer

PurposeThe CellSearch system (Veridex, LLC) is useful for detecting circulating tumor cells (CTCs) in various carcinomas, including colorectal cancer (CRC); however, there are some problems associated with its clinical use. A transcription–reverse transcription concerted reaction (TRC) method, which is a PCR-based technique producing more stable and reliable results, because it is a more simplified process compared with the conventional techniques, has been introduced for detecting micrometastasis in some carcinomas. We aimed to demonstrate the effectiveness of TRC method in the CTC detection.MethodsWe compared the two methods for the sensitivity for CTC detection using the colon cancer cell line and 42 whole-blood samples from patients with advanced or metastatic CRC. Furthermore, 25 patients with metastatic CRC were enrolled to investigate the correlation between CTC detection and prognosis in both methods.ResultsThe sensitivity of the TRC method was similar to that of the CellSearch system. The overall survival rate was significantly worse in the patients diagnosed as CTC-positive by the TRC method than in those diagnosed as CTC-negative; this finding was similar to the prognosis indicated by the CellSearch system. However, clinically, the TRC method could detect CTCs more rapidly and at a reduced cost compared with the CellSearch system.ConclusionsThe TRC method seems to be a useful alternative to the CellSearch system for clinically detecting CTCs in patients with metastatic CRC.

RPN2 expression predicts response to docetaxel in oesophageal squamous cell carcinoma

Background:Neoadjuvant chemotherapy – often using docetaxel in various combinatorial regimens – is a standard treatment choice for advanced oesophageal squamous cell carcinoma (ESCC) in Japan. However, no useful markers exist that predict docetaxel’s effects on ESCC. Ribophorin II (RPN2) silencing, which reduces glycosylation of P-glycoproteins and decreases membrane localisation, promotes docetaxel-dependent apoptosis. We investigated whether RPN2 expression in ESCC biopsy specimens could be a predictive biomarker in docetaxel-based neoadjuvant chemotherapy.Methods:We evaluated RPN2 expression immunohistochemically in biopsy specimens from 79 patients with node-positive ESCC, who received docetaxel-based adjuvant chemotherapy, and compared clinical and pathological responses between the RPN2-positive and RPN2-negative groups. We also studied susceptibility of RPN2-suppressed ESCC cells to docetaxel.Results:The RPN2-negative group had better clinical and pathological responses to docetaxel than the RPN2-positive group. We also found RPN2 suppression to alter docetaxel susceptibility in vitro.Conclusion:Expression of RPN2 in biopsy specimens could be a useful predictive marker for response to docetaxel-based neoadjuvant chemotherapy in ESCC.

Extensive lymphatic spread of cancer cells in patients with thoracic esophageal squamous cell carcinoma: detection of CEA-mRNA in the three-field lymph nodes.

The aim of this study is to clarify the extent of lymphatic spread of cancer cells using a novel genetic test to examine patients with thoracic esophageal squamous cell carcinoma (ESCC).

Epithelial-mesenchymal transition in gastroenterological cancer

Epithelial-mesenchymal transition (EMT) was fi rst reported as an essential process in embryonic cells and later showed that cancer cells, regardless of the context, exhibited a similar phenomenon that was crucial for tumor progression. Epithelial cells lose their adhesive characteristic capacity which is necessary for their functions but gain a mesenchymal phenotype. This change from epithelial to the mesenchymal phenotype of cancer cells makes it diffi cult to understand the mechanism underlying cancer biology and tumor progression. A number of transcription factors involved in tumor cell EMT and microRNA-regulated EMT have been reported. This review discussed recent fi ndings and new players in EMT in gastrointestinal cancers. Since the molecular mechanisms of tumor progression are sometimes context-dependent, the recent fi ndings of EMT have been reviewed in a context-dependent manner.

Abstract 1155: Serum microRNA-21 is a novel biomarker in patients with esophageal squamous cell carcinoma

Backgrounds and objective: Esophageal squamous cell carcinoma (ESCC) is one of the most difficult malignancies to be cured among gastrointestinal tract cancers. Recently, it was revealed that microRNAs stably exist in serum and may affect pathogenesis of several diseases. However, there are only a few reports that have demonstrated the significance of microRNA (mir) in the serum of patients with ESCC. Accordingly, the aims of this study were to clarify the status of mir-21 in the serum of ESCC patients and to reveal the usefulness of this molecule as a biomarker. Patients and methods: Serum samples were obtained from 52 patients diagnosed with ESCC, as well as from 17 “control” patients diagnosed with gallstones, without active inflammation, in Kumamoto University Hospital from 2008 to 2010. Pre-treatment serum samples were collected during diagnostic studies. Thirty-two ESCC patients received induction chemotherapy using the Docetaxel/ cisplatin /5-Fluorouracil (DCF) regimen (60 mg/m 2 Docetaxel on day 1; 350 mg/m 2 5-Fluorouracil and 6 mg/m 2 cisplatin on days 1-5; 2 courses) after being diagnosed with node-positive ESCC. Twenty-four pairs of serum samples before and after chemotherapy were collected. The expression of miR-21 was determined by qRT-PCR using TaqMan microRNA assay Kits ® (Applied Biosystems, USA). Results: The results of qRT-PCR analysis indicated that the ESCC patients had significantly higher levels of serum miR-21 than the control patients (P = 0.007). The value of area under the receiver operating characteristic curve was 0.885 for microRNA-21. When the patients were divided into two groups according to tumor size (≥40mm vs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1155. doi:10.1158/1538-7445.AM2011-1155