Ryuichi Karashima

MicroRNA-21 Regulates the Proliferation and Invasion in Esophageal Squamous Cell Carcinoma

Purpose: MicroRNAs are ∼22 nucleotide noncoding RNA molecules that posttranscriptionally regulate gene expression. The aim of this study was (a) to determine a role of microRNA-21 in esophageal squamous cell carcinoma and (b) to elucidate the regulation of the programmed cell death 4 (PDCD4) gene by microRNA-21. Experimental Design: MicroRNA-21 expression was investigated in 20 matched normal esophageal epitheliums and esophageal squamous cell carcinomas and seven esophageal squamous cell carcinoma cell lines (TE6, TE8, TE10, TE11, TE12, TE14, KYSE30) by TaqMan quantitative real-time PCR and in situ hybridization. To evaluate the role of microRNA-21, cell proliferation and invasion were analyzed with anti–microRNA-21–transfected cells. In addition, the regulation of PDCD4 by microRNA-21 was elucidated to identify the mechanisms of this regulation. Results: Of 20 paired samples, 18 cancer tissues overexpressed microRNA-21 in comparison with matched normal epitheliums. Specifically, patients with lymph node metastasis or venous invasion showed significantly high expression of microRNA-21. In situ hybridization for microRNA-21 showed strong positive staining in paraffin-embedded esophageal squamous cell carcinoma tissues. All seven esophageal squamous cell carcinoma cell lines also overexpressed microRNA-21, and anti–microRNA-21–transfected cells showed significant reduction in cellular proliferation and invasion. The PDCD4 protein levels in esophageal squamous cell carcinoma cells have an inverse correlation with microRNA-21 expression. Anti–microRNA-21–transfected cells increased PDCD4 protein expression without changing the PDCD4 mRNA level and increased a luciferase-reporter activity containing the PDCD4-3′ untranslated region construct. Conclusions: MicroRNA-21 targets PDCD4 at the posttranscriptional level and regulates cell proliferation and invasion in esophageal squamous cell carcinoma. It may serve as a novel therapeutic target in esophageal squamous cell carcinoma.

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.

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.

Lymphatic vessel invasion detected by the D2-40 monoclonal antibody is an independent prognostic factor in node-negative esophageal squamous cell carcinoma

D2‐40 staining has been reported to be useful for both identifying lymphatic vessel invasion (LVI) and counting lymphatic vessel density (LVD) in various cancers. The aim of this study was to clarify the prognostic significance of D2‐40 staining in patients with esophageal squamous cell carcinoma (ESCC).

Influence of preoperative docetaxel, cisplatin, and 5-fluorouracil on the incidence of complications after esophagectomy for resectable advanced esophageal cancer.

A limited number of patients with resectable advanced esophageal cancer can be cured by surgery alone. Although a regimen that consists of docetaxel, cisplatin, and 5-fluorouracil (DCF) is a potential preoperative chemotherapy (PCT) option for squamous cell carcinoma of the esophagus, the influence of DCF on subsequent esophagectomies remains unclear. A total of 80 patients who received preoperative DCF chemotherapy, and 174 patients who did not receive any preoperative treatment were retrospectively analyzed. There were no treatment-related deaths. No delays in surgery due to adverse events related to DCF were reported. Although patients who received PCT had significantly more advanced cancers and worse preoperative conditions, the incidence rates of complications did not increase. Although the frequency of severe complications was significantly higher in patients who received PCT, this treatment was not an independent factor for the occurrence of severe complications. PCT with DCF did not negatively affect subsequent esophagectomies with regard to the frequency of complications.

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).

Abstract 5150: JMJD3 suppresses progression of colorectal carcinoma by regulating cell cycle and anti-apoptosis

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Background: JMJD3 has been known to play important roles in transcriptional regulation and cell proliferation by demethylating histone3 lysin27 (H3K27) in various cancers. However, the mechanism underlying JMJD3-mediated transcriptional regulation and the relation with prognosis were unclear in colorectal carcinoma. Here, we investigated the expression of JMJD3 in clinical samples and the function of JMJD3 in colorectal carcinoma cell lines. Methods: Quantitative real-time PCR were performed to check the expression of JMJD3 mRNA using the carcinoma and normal colorectal tissue in 14 colorectal carcinoma patients who underwent a curative surgery. In vitro analysis, we investigated the expression of JMJD3 in colorectal carcinoma cell lines by Western Blotting, and we knocked down the expression of JMJD3 by using small interfering RNA in Colo201 and colo320. The influence of JMJD3 on cell growth, apoptosis and cell cycle was assessed by growth assay, apoptosis assay and cell cycle analysis. We checked the changes of cell cycle related genes by real time PCR. Results: In clinical samples, the expression of JMJD3 in carcinoma tissue was significantly lower than in normal tissue (P=0.02). In vitro analysis, we knocked down the expression of JMJD3 by siRNA targeting JMJD3 and checked the reduction on mRNA and protein level. Knockdown of JMJD3 significantly increased cell proliferation (P=0.02), reduced apoptosis (P=0.003), and increased G2/M phase population (P = 0.05) which was detected by FACS. Moreover, knockdown of JMJD3 significantly decreased the expression of p15INK4B in quantitative real-time PCR. Conclusion: We showed the expression of JMJD3 in carcinoma tissue was lower than in normal tissue and demonstrated that the down-regulation of JMJD3 resulted in the increase of cell proliferation, acceleration of cell cycle and anti-apoptosis in vitro analysis. These results suggest that JMJD3 is a tumor suppressor gene for colorectal carcinoma. Note: This abstract was not presented at the meeting. Citation Format: Ryuma Tokunaga, Shigeki Nakagawa, Yasuo Sakamoto, Ryuichi Karashima, Satoshi Ida, Yu Imamura, Takatsugu Ishimoto, Shiro Iwagami, Yoshifumi Baba, Yuji Miyamoto, Naoya Yoshida, Hideo Baba. JMJD3 suppresses progression of colorectal carcinoma by regulating cell cycle and anti-apoptosis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5150. doi:10.1158/1538-7445.AM2014-5150

Erratum to: Advantages of FDG-PET/CT over CT alone in the preoperative assessment of lymph node metastasis in patients with esophageal cancer.

Two inaccurate technical descriptions appeared in above titled article. In the “Imaging studies” subsection, the dose of intravenously injected F-FDG was misdescribed as 370 MBq (10 mCi). Actual dose of F-FDG was 185–296 MBq (5–8 mCi). In addition, injection rate of nonionic contrast medium during CT imaging was misdescribed as 2.5 ml/s, and accurate injection rate was 2.0 ml/s. The authors sincerely apologize for these errors.

Diagnostic laparoscopic biopsy for intraabdominal tumors.

Improvements in imaging technology have resulted in an increase in the incidental detection of intraabdominal tumors. Diagnostic computed tomography (CT)- and ultrasound (US)-guided biopsy, while minimally invasive, often provides specimens that are insufficient for histological evaluation. Moreover, it can be difficult to perform because the location and size of the tumor. In such cases, laparoscopic biopsy is useful because it is less invasive than laparotomy, but more reliable than imaging-guided biopsy, to obtain a sufficient specimen, regardless of the location and size of the tumor. We report a series of seven patients who underwent laparoscopic biopsy of intraabdominal tumors of unknown origin. There were no cases of conversion to laparotomy and all patients were able to resume oral intake on postoperative day 1. There were no intraoperative or postoperative complications. Thus, laparoscopic biopsy for a tumor of unknown origin is useful and minimally invasive.

Abstract 3861: Prognostic impact of KRAS copy number amplification in esophagogastric junction adenocarcinoma

Background: The incidence of esophageal adenocarcinoma (EAC) or esophagogastric junction adenocarcinoma (EGJA) has been dramatically increasing. KRAS oncogenic alterations activate multiple pathways in various types of cancer. A recent comprehensive genetic study has reported that around 20% of EAC harbor KRAS copy number gain (Dulak AM et al, 2012, Nat Genet). However, the prognostic role KRAS amplification has not been well examined in EAC/EGJA. Methods: DNA was extracted from formalin-fixed paraffin-embedded tissue of 145 EGJA (Siewert types I-III). KRAS copy number was detected by real-time quantitative PCR, and KRAS copy number gain was defined as KRAS/RNase P (as a reference) ratio ≥ 2.5. The impact of KRAS amplification was analyzed in relation to clinicopathological factors, and patient outcomes [disease free survival (DFS), and overall survival (OS)]. Multivariate cox proportional hazards model was performed adjusting clinicopathological factors. Results: KRAS amplification was observed in 40 (33.1%) cases. KRAS amplification was significantly associated with the lymph node metastasis (P = 0.0145), and was significantly associated with poorer patient outcome (P = 0.0355 in DFS, and P = 0.0067 in OS). In multivariate analysis, KRAS amplification was an independent prognostic factor for OS [multivatiate hazard ratio (HR) = 2.53; 95% confidence interval (CI), 1.22-5.25, P = 0.014] and DFS [multivatiate hazard ratio (HR) = 1.94; 95% confidence interval (CI), 1.04-3.56, P = 0.0382]. Conclusions: KRAS copy number gain may be a useful prognostic marker in EGJA. Citation Format: Kenichi Nakamura, Yu Imamura, Ryuma Tokunaga, Mayuko Ohuchi, Yuki Kiyozumi, Keisuke Kosumi, Daisuke Izumi, Kazuto Harada, Junji Kurashige, Ryuichi Karashima, Yukiharu Hiyoshi, Shiro Iwagami, Yoshifumi Baba, Yasuo Sakamoto, Yuji Miyamoto, Naoya Yoshida, Hiroshi Saeki, Eiji Oki, Yoshihiko Maehara, Hideo Baba. Prognostic impact of KRAS copy number amplification in esophagogastric junction adenocarcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3861. doi:10.1158/1538-7445.AM2015-3861