Osamu Nakahara

Carcinogenesis of Intraductal Papillary Mucinous Neoplasm of the Pancreas: Loss of MicroRNA-101 Promotes Overexpression of Histone Methyltransferase EZH2

BackgroundThe mechanisms of IPMN carcinogenesis are as yet unclear. This study aimed to determine whether expression of EZH2 promotes neoplastic progression of IPMN and PDCA, and to elucidate regulation of EZH2 expression by miR-101.MethodsEZH2 mRNA and protein expression were investigated in 8 human pancreatic cancer cell lines by PCR and western blotting. Pre-miR-101 and anti-miR-101 were transfected into pancreatic cancer cells to elucidate EZH2 regulation by miR-101. To evaluate whether EZH2 modulates malignant progression of IPMN, EZH2 expression in IPMN was examined by immunohistochemistry. Next, we collected malignant and benign cells from FFPE samples of IPMNs using laser capture microdissection and extracted the RNA. miR-101 expression in IPMN was assessed using real-time PCR.ResultsAll pancreatic cancer cell lines expressed EZH2 mRNA and protein. The induction of miR-101 by transfection of pre-miR-101 in MIA PaCa-2 was closely related to a reduction in EZH2 protein production compared with control, whereas there was little difference in the expression of EZH2 mRNA. Anti-miR-101 transfected pancreatic cancer cells showed an increase in EZH2 protein, while the level of EZH2 mRNA was not elevated. Immunohistochemistry revealed that the expression of EZH2 was significantly higher in malignant than benign IPMN. Expression of miR-101 was significantly lower in malignant IPMN than benign IPMN.ConclusionsMiR-101 targets EZH2 at the posttranscriptional level, and loss of miR-101 could be a trigger for the adenomacarcinoma sequence of IPMN by upregulation of EZH2. This study suggests miR-101–EZH2 blockade as a potential therapeutic target in IPMN carcinogenesis.

Is an estimation of physiologic ability and surgical stress able to predict operative morbidity after pancreaticoduodenectomy

BackgroundMortality rates after pancreaticoduodenectomy (PD) are below 4% in high volume centers, although morbidity rates still remain high. Therefore, it is important to clarify a predictor associated with operative morbidity after PD. The estimation of physiologic ability and surgical stress (E-PASS) score has been developed for comparative audit in general surgical patients.ObjectiveTo evaluate whether E-PASS scoring system could predict the occurrence of complications after PD.MethodsWe performed retrospective analysis of 69 patients (42.0% pancreatic cancer, 31.9% bile duct cancer, and others) who underwent PD using the E-PASS as a predictor of morbidity. Correlations between the incidence rates of postoperative complications and the preoperative risk score (PRS), surgical stress score (SSS) and comprehensive risk score (CRS) of the E-PASS scoring system were evaluated.ResultsOf the 69 patients 30 (43.5%) experienced a total of 54 postoperative complications. All E-PASS scores, especially PRS and CRS were significantly higher in the patients with postoperative complications than in the patients without complication. The complication rate gradually increased as the PRS, SSS and CRS score increased. Under receiver operating characteristic analysis, if a cut-off point of CRS was 0.75, sensitivity and specificity for the prediction of operative morbidity after PD was 80.0 and 79.5%, respectively. Neoadjuvant chemotherapy and intraoperative radiation therapy (IORT) did not influenced on operative morbidity after PD.ConclusionE-PASS scoring system is useful to evaluate for morbidity after PD. Neoadjuvant chemotherapy and IORT could be adapted without significant extra risk for surgical complication.

5-fluorouracil intra-arterial infusion combined with systemic gemcitabine for unresectable pancreatic cancer.

Objectives: The aim of this study was to define assessment of response and adverse events of the combination chemotherapy of 5-fluorouracil (5-FU) pancreatic and hepatic arterial continuous infusion and systemic gemcitabine administration for unresectable pancreatic cancer. Methods: We treated 24 chemotherapy-naive patients with unresectable pancreatic cancer. To prevent gastroduodenal injury from 5-FU infusion, the catheter was placed to allow the distribution of 5-FU to the pancreatic tumor and the liver after occlusion of the gastric and pancreaticoduodenal arteries. 5-FU was administered at a dose of 250 mg/d on days 1 to 5 every week as a continuous arterial infusion. Gemcitabine was infused intravenously at a dose of 1000 mg once weekly for 3 consecutive weeks of every 4 weeks. Results: The partial response rate was 20.8% (5 of 24), although there was no case of complete response. Fourteen cases (58.3%) were stable disease, and 5 cases (20.8%) were progressive disease. The most common toxicities were hematological and gastrointestinal events. No patients died of adverse effects using this chemotherapy. Gastric and/or duodenal ulcers occurred because of 5-FU intra-arterial infusion. Catheter-related cholangitis occurred in patients with biliary drainage for obstructive jaundice. Median survival time was 14 months, with a 50.9% 1-year survival rate, although patients with performance status 2 and multiple organ metastases had a poor prognosis. Conclusions: This combination chemotherapy was well tolerated and seemed to be effective for patients with unresectable pancreatic cancer.

Joint Symptoms : A Practical Problem of Anastrozole

BackgroundAnastrozole and tamoxifen have mild toxicity. However, we noticed that more patients treated with anastrozole complained of joint symptoms than expected. In particular, digital stiffness as is seen with rheumatoid arthritis is a problem. Some clinical trials of anastrozole in Europe and the United States reported musculoskeletal disorders as adverse events, however, joint symptoms were not described in detail.Patients and MethodsAt our clinic from August 2001 to March 2005, 53 postmenopausal women with estrogen receptor-positive breast cancer were treated with anastrozole. We calculated the incidence and classified the grade of joint symptoms by interviewing patients. We also investigated the patients’ characteristics and their relevance to joint symptoms.ResultsOf 53 patients, 14 patients (26%) had joint symptoms (13 patients with digital stiffness and 3 patients with arthralgias of wrist and shoulders). Joint symptoms tended to occur in the patients who had previously undergone chemotherapy; however, there has no relationship between prior hormonal therapy and joint symptoms. Seven patients who discontinued anastrozole treatment showed improved symptoms. Five patients with grade 1 digital stiffness continued anastrozole treatment without additional treatment. Two patients with grade 1 digital stiffness, who took a Chinese herbal medicine showed improved symptoms and continued anastrozole treatment.ConclusionBenefits to the patients may possibly be lost by discontinuation of anastrozole or changing to tamoxifen since the clinical superiority of anastrozole to tamoxifen has been reported. We should continue anastrozole in patients with low grade symptoms, while ensuring that patients are aware of the toxicity of anastrozole.

Cancer cells spread through lymph vessels in the submucosal layer of the common bile duct in gallbladder carcinoma.

INTRODUCTION In the present study, we performed immunohistochemical staining with a lymphatic epithelium-specific marker, D2-40, to analyze the status of lymphatic spreading in the hepatoduodenal ligament in T2 gallbladder carcinoma (GC). METHODS One hundred and eighty-six paraffin-embedded specimens from 15 T2 GC patients were reviewed. RESULTS Lymph vessels lined with D2-40 were visualized in the submucosal layer of the common bile duct in all cases. In 3 of 15 patients, clusters of cancer cells were identified in the submucosal lymph vessels of the extrahepatic bile duct, and this lymphatic invasion of cancer cells failed to be detected with only conventional hematoxylin-eosin staining. The frequency of the invasion to the submucosal lymph vessels in T2 GC correlated with presence of microscopic invasion to hepatoduodenal ligament and perineural invasion. CONCLUSION There were lymph vessels in the submucosal layer of the common bile duct, and cancer cells can spread through these channels in addition to the large lymph vessels in subserosal layer around the extrahepatic bile duct in GC. The present results would support the concept of en bloc resection of the extrahepatic bile duct in curative resection for T2 GC.

EZH2 is associated with malignant behavior in pancreatic IPMN via p27Kip1 downregulation.

Background The epigenetic mechanism of tumorigenesis in pancreatic intraductal papillary mucinous neoplasm (IPMN) remains largely unknown. The aim of this study is to examine the role of enhancer of zeste homologue 2 (EZH2) alteration in pancreatic IPMN progression. Methods Fifty-four surgically resected pancreatic IPMN specimens, including a total of 181 lesions (normal duct in 48, adenoma in 50, borderline atypia in 53, carcinoma in situ (CIS) in 19, and invasive carcinoma in 11) were analyzed by immunohistochemical staining (EZH2, Ki-67, p27Kip1). Using paraffin embedded sections, total RNA was successfully extracted from 20 IPMN lesions (borderline IPMN in 9, CIS in 6, invasive carcinoma in 5) and 7 pancreatic normal ducts, and then levels of EZH2 and p27Kip1 mRNA were analyzed by real time PCR. Results In immunohistochemical analysis, cell proliferative activity revealed by Ki-67 positive nuclei was increased during IPMN progression (normal duct<adenoma<borderline atypia<CIS ≈ invasive carcinoma). EZH2 expression displayed a similar pattern (normal duct<adenoma<borderline atypia<CIS ≈ invasive carcinoma) with cell proliferative activity. EZH2 expression in malignant (CIS and invasive carcinoma) IPMNs was significantly higher than that in adenoma and borderline-atypia IPMNs. EZH2 expression level in IPMN lesions was positively correlated with the Ki-67 positive nuclear ratio (p<0.0001). EZH2-positive cells in malignant IPMN did not express p27Kip1. EZH2 mRNA expressions in malignant lesions were significantly higher than those in benign lesions (p<0.0001). In contrast, p27Kip1 mRNA in malignant lesions was significantly decreased compared to those in benign lesion (p<0.05), and there was an inverse correlation between EZH2 and p27Kip1 mRNA levels (p = 0.0109). Conclusion EZH2 is associated with the accelerated cell proliferation and malignant step in pancreatic IPMN via the downregulation of p27Kip1.

Clinical significance of dihydropyrimidine dehydrogenase and thymidylate synthase expression in patients with pancreatic cancer

BackgroundLittle is known about the clinical significance of TS and DPD in pancreatic cancer. We aimed to evaluate TS and DPD expression levels in not only pancreatic cancer but also surrounding normal pancreatic tissues to assess the clinical implications of the expression of TS and DPD in this study.Patients and methodsPancreatic cancer and normal pancreatic tissues were obtained from 18 patients with pancreatic cancer who underwent pancreatic resection to measure TS and DPD activities. The TS and DPD activities were determined by enzyme-linked immunosorbent assay using non-fixed fresh-frozen specimens.ResultsPancreatic cancer tissues had significantly higher DPD and TS enzyme activities than surrounding normal tissue. Anaplastic ductal carcinoma had higher DPD and TS activities than the other histological types. Patients with high DPD in this study demonstrated poorer prognosis than those with low DPD. On the other hand, there was no statistically significant difference in survival between the high and the low TS groups.ConclusionsThe efficacy of 5-FU may be lower in pancreatic cancer tissue than in normal tissue because DPD activity is upregulated in pancreatic cancer tissue compared to normal pancreatic tissue. It is necessary to develop an effective 5-FU delivery system and/or 5-FU combined with an inhibitor for DPD that can be used when 5-FU must be administered to patients with pancreatic cancer. High DPD activity may be a prognostic factor in patients with pancreatic cancer.

Cutaneous metastases secondary to pancreatic cancer

AIM To evaluate prognoses after cutaneous metastases, derived from pancreatic cancer. METHODS We treated two patients with cutaneous metastases from pancreatic cancer. We reviewed 40 reported patients in addition to our cases and analyzed clinical features of cutaneous metastases from pancreatic cancer. RESULTS The median survival time (MST) was 5 mo after diagnoses of cutaneous metastases. The cumulative 2-year survival rate was 3.5%. The most frequent site of cutaneous metastases was the umbilicus. The MST of patients who were treated with chemotherapy or chemoradiotherapy (CRT) was 6.5 mo, which was statistically longer in comparison to patients without treatment. Prognoses of cutaneous metastases are similar to other metastatic sites from pancreatic cancer. Receiving chemotherapy or CRT was the only prognostic factor of cutaneous metastases from pancreatic cancer. CONCLUSION The prognoses of cutaneous metastases are similar to other metastatic pancreatic cancers. Receiving chemotherapy or CRT was the only prognostic factor of cutaneous metastases from pancreatic cancer.

NO donor and MEK inhibitor synergistically inhibit proliferation and invasion of cancer cells

Nitric oxide (NO) shows tumoricidal activity. We had previously reported that NO downregulates the phosphatidylinositol-3-kinase/Akt pathway, but upregulates the MEK/ERK pathway downstream of growth factor signaling. We hypothesized that NO donor and MEK inhibitor in combination synergistically inhibit the viability of cancer cells compared to either NO donor or MEK inhibitor alone. We determined the effects of S-nitrosoglutathione (GSNO, NO-donor) and U0126 (MEK inhibitor) on insulin-like growth factor-I (IGF-I) and epidermal growth factor (EGF) signaling, proliferation and invasion in cancer cell lines. GSNO inhibits phosphorylation of IGF-I receptor (IGF-IR), EGF receptor (EGFR) and Akt, but upregulates ERK1/2 phosphorylation in MIAPaCa-2 and HCT-116 cells after stimulation by IGF-I and EGF. On the other hand, U0126 inhibits phosphorylation of ERK1/2, but upregulates phosphorylation of IGF-IR and EGFR in MIAPaCa-2 and HCT-116 cells. The combination of GSNO and U0126 downregulates phosphorylation of IGF-IR, EGFR, Akt and ERK1/2 after stimulation by IGF-I and EGF. GSNO as well as U0126, inhibits the proliferation of MIAPaCa-2, HCT-116, Panc-1, MCF-7, HT-29 and AGS cells in a dose-dependent manner. GSNO and U0126 in combination synergistically inhibit proliferation and invasion of cancer cells. These results indicate that the combined treatment of NO donor and MEK inhibitor may be promising in cancer therapy.

Risk factors associated with delayed haemorrhage after pancreatic resection

BACKGROUND Delayed haemorrhage (DH) is a life-threatening complication of pancreatic resection (PR) and the mortality rate for DH is very high. However, the risk factors and prognostic factors associated with DH are rarely evaluated. METHODS A pancreatic resection was performed on 457 patients. Delayed haemorrhage was defined as bleeding from the surgical site ≥ 5 days after PR. Risk factors for DH were assessed according to demographics and pathological and operative parameters. Prognostic factors after DH were evaluated for the shock index (heart rate/systolic blood pressure) and systemic inflammatory response syndrome (SIRS) scores. RESULTS Of the 457 patients, 11 (2.4%) experienced DH after PR. Logistic regression analysis showed that age >60 years and a diagnosis of malignant disease were risk factors for DH. The shock index and SIRS scores at the onset of DH were significantly higher in patients who died as compared with those patients that survived (P < 0.05). DISCUSSION PR-associated DH carries an increased risk for patients aged >60 years with malignant disease. Prognostic factors were a shock index score ≥ 0.7 and SIRS at the onset of DH.