Involvement of clusterin expression in the refractory response of pancreatic cancer cells to a MEK inhibitor.

Constitutive activation of the mitogen-activated protein kinase (MAPK) signaling pathway is essential for tumorigenesis of pancreatic ductal adenocarcinoma (PDAC). To date, however, almost all clinical trials of inhibitor targeting this pathway have failed to improve the outcome of patients with PDAC. We found that implanted MIA Paca2, a human PDAC cell line sensitive to a MAPK inhibitor, PD0325901, became refractory within a week after treatment. By comparing the expression profiles of MIA Paca2 before and after acquisition of the refractoriness to PD0325901, we identified clusterin (CLU) as a candidate gene involved. CLU was shown to be induced immediately after treatment with PD0325901 or expressed primarily in more than half of PDAC cell lines, enhancing cell viability by escaping from apoptosis. A combination of PD0325901 and CLU downregulation was found to synergistically or additively reduce the proliferation of PDAC cells. In surgically resected PDAC tissues, overexpression of CLU in cancer cells was observed immunohistochemically in approximately half of the cases studied. Collectively, our findings highlight the mechanisms responsible for the rapid refractory response to MEK inhibitor in PDAC cells, suggesting a novel therapeutic strategy that could be applicable to patients with PDAC using inhibitor targeting the MAPK signaling pathway and CLU.

Downregulation of ZNF395 Drives Progression of Pancreatic Ductal Adenocarcinoma through Enhancement of Growth Potential.

BACKGROUND: Progression of pancreatic intraepithelial neoplasia (PanIN) to invasive carcinoma is a critical factor impacting the prognosis of patients with pancreatic tumors. However, the molecular mechanisms involved are not fully understood. We have reported that the process frequently involves loss of chromosome 8p, causing downregulation of DUSP4, thus conferring invasive ability on cancer cells. Here, we focus on ZNF395, whose expression was also found to be decreased by 8p loss and was predicted to be a growth suppressor gene. METHODS: Pancreatic cancer cell lines inducibly expressing ZNF395 were established to assess the functional significance of ZNF395 in pancreatic carcinogenesis. Immunohistochemistry was also performed to analyze the expression levels of ZNF395 in pancreatic cancer tissues. RESULTS: Induction of ZNF395 in pancreatic cancer cells resulted in marked activation of JNK and suppression of their proliferation through a delay in cell cycle progression. Immunohistochemistry revealed that ZNF395 was expressed ubiquitously in both normal pancreatic ducts and PanINs but was significantly reduced in invasive cancers, especially those showing poor differentiation. CONCLUSION: ZNF395 acts as a novel tumor suppressor gene. Its downregulation caused by 8p loss in intraepithelial cells accelerates their proliferation through dysregulation of the cell cycle, leading to progression to invasive cancer.

Altered glycosylation associated with dedifferentiation of hepatocellular carcinoma: a lectin microarray-based study.

BACKGROUND: Altered glycosylation associated with hepatocellular carcinoma (HCC) is well documented. However, few reports have investigated the association between dedifferentiation and glycosylation. Therefore, the aim of this study was to analyze glycosylation associated with dedifferentiation of HCC within the same nodule and to investigate glycosyltransferase related to the glycosylation. METHODS: We analyzed resected HCC specimens (n = 50) using lectin microarray to comprehensively and sensitively analyze glycan profiles, and identify changes to glycosylation between well- and moderately-differentiated components within the same nodule. Moreover, we performed immunohistochemical staining of mannosyl(α-1,3-)-glycoprotein ß-1,2-N-acetylglucosaminyltransferase (MGAT1), which is an essential glycosyltransferase that converts high-mannose glycans to complex- or hybrid-type N-glycans. RESULTS: Four lectins from Narcissus pseudonarcissus agglutinin (NPA), Concanavalin A, Galanthus nivalis agglutinin, and Calystegia sepium agglutinin were significantly elevated in moderately-differentiated components of HCC compared with well-differentiated components, and all lectins showed binding specificity to high-mannose glycans. Therefore, these structures were represented to a greater extent in moderately-differentiated components than in well-differentiated ones. Immunohistochemical staining revealed significantly increased NPA expression and decreased MGAT1 expression in moderately-differentiated components. Low MGAT1 expression in moderately-differentiated components of tumors was associated with intrahepatic metastasis and had tendency for poor prognosis. CONCLUSION: Dedifferentiation of well-differentiated HCC is associated with an increase in high-mannose glycans. MGAT1 may play a role in the dedifferentiation of HCC.

Fucosylation is associated with the malignant transformation of intraductal papillary mucinous neoplasms: a lectin microarray-based study.

PURPOSE: Intraductal papillary mucinous neoplasm (IPMN) is an intraductal mucin-producing pancreatic neoplasm with the potential for malignant transformation. Changes in glycans expressed on the cell surface and glycotransferases play important roles in malignant transformation. We conducted this study to analyze glycan alterations in IPMNs by using a lectin microarray and to identify the factors associated with altered glycans and their relationships with malignant transformation. METHODS: Using a lectin microarray, we evaluated glycan expression in 22 samples of IPMN with carcinoma, obtained from curative resections performed in our department. We also used immunohistochemistry to investigate fucosyltransferase 8 (Fut 8) protein expression, which is associated with glycan alterations in IPMNs. RESULTS: The lectin microarray demonstrated that only two lectins, Aleuria aurantia lectin (AAL) and Aspergillus oryzae L-fucose-specific lectin (AOL), which bind to fucose, exhibited significant sequential increases from normal pancreatic duct to adenoma and carcinoma. Similarly, Fut 8 protein expression, which is associated with AAL and AOL, sequentially and significantly increased from the normal pancreatic duct to adenoma and carcinoma. CONCLUSIONS: Lectin microarray analysis suggested that fucosylation is associated with the malignant transformation of IPMNs.

Operative timing of laparoscopic cholecystectomy for acute cholecystitis in a Japanese institute.

BACKGROUND AND OBJECTIVES: In patients with acute cholecystitis who cannot undergo early laparoscopic cholecystectomy (within 72 hours), 6 weeks to 12 weeks after onset is widely considered the optimal timing for delayed laparoscopic cholecystectomy. However, there has been no clear consensus about it. We aimed to determine optimal timing for delayed laparoscopic cholecystectomy for acute cholecystitis. METHODS: Medical records of 100 patients who underwent standard laparoscopic cholecystectomy were reviewed retrospectively. Patients were divided into group 1, patients undergoing laparoscopic cholecystectomy within 72 hours of onset; group 2, between 4 days to 14 days; group 3, between 3 weeks to 6 weeks; group 4, >6 weeks. RESULTS: No significant differences existed between groups in conversion rate to open surgery, operation time, blood loss, or postoperative morbidity, and hospital stay. However, total hospital stay in groups 1 and 2 was significantly shorter than that in groups 3 and 4 (P<.01). In addition, the total hospital stay in group 3 was also significantly shorter than that in group 4 (P<.01). CONCLUSIONS: Best timing of laparoscopic cholecystectomy for acute cholecystitis may be within 72 hours, and the delayed timing of laparoscopic cholecystectomy in patients who cannot undergo early laparoscopic cholecystectomy is probably as soon as possible after they can tolerate laparoscopic cholecystectomy.

Gluteal muscle metastasis with peritoneal dissemination from gastric cancer during postoperative adjuvant chemotherapy: a case report.

BACKGROUND: Skeletal muscle metastasis from gastric cancer is rare and has a poor prognosis. We reported a case of gluteal muscle metastasis with peritoneal dissemination from gastric cancer during postoperative adjuvant chemotherapy. CASE PRESENTATION: A 64-year-old man with gastric cancer underwent distal gastrectomy with D2 lymph node resection. The pathological diagnosis was poorly differentiated adenocarcinoma and signet cell carcinoma, T3N3bM0, Stage IIIC. Metastases were found in all regional lymph nodes, except 11p. The resection margin was negative. S-1 plus docetaxel therapy was administered as postoperative adjuvant chemotherapy. Six month post-operation, the patient presented with right gluteal muscle tenderness and abdominal distension. Computed tomography revealed a solid mass in the right gluteal muscle, a disseminated nodule on the abdominal wall, and massive ascites. Pathological examination of the gluteal muscle revealed signet cell carcinoma, similar to the resected gastric cancer. The tumor was diagnosed as gastric cancer metastases. Ascites cytology was class V. Thereafter, the patient underwent one course of capecitabine plus cisplatin combined with trastuzumab. Radiation therapy was also administered to relieve the pain of gluteal muscle metastasis. However, chemoradiotherapy was ineffective, and the patient died 2 months after the recurrence. CONCLUSIONS: Skeletal muscle metastasis and peritoneal dissemination during adjuvant chemotherapy indicated a poor prognosis.

Groove pancreatic carcinoma.

The groove area is localized between the head of the pancreas, the duodenum, and the common bile duct. Differentiating of groove pancreatitis from pancreatic carcinoma is often difficult. Herein, we report a 54-year-old woman with groove pancreatic adenocarcinoma presenting epigastralgia, jaundice, and vomiting. The diagnosis was confirmed by computed tomography (CT), endoscopic retrograde cholangiopancreatography (ERCP), and endoscopic biopsy. The patient underwent pancreatoduodenectomy. Microscopically, well-differentiated adenocarcinoma was mainly located in Santorini's duct, but there was no invasion to the main pancreatic duct. The patient followed a satisfactory post operative course. She is doing well without recurrence 15 months after the surgery. It is very difficult to differentiate groove pancreatic carcinoma from groove pancreatitis. To avoid unnecessary surgical treatment, endoscopic biopsy and observation of the duodenum are useful for diagnosis. However, keeping in mind the differential diagnosis of pancreatic head carcinoma is necessary.

Effect of pre-firing compression on the prevention of pancreatic fistula in distal pancreatectomy.

BACKGROUND: Postoperative pancreatic fistula (POPF) is a major complication of distal pancreatectomy (DP). Several procedures for resection and closure of the pancreas have been proposed; however, the rate of POPF remains high. The aims of this study were to investigate the relationship between perioperative factors and POPF and to clarify the advantages of pre-firing compression of the pancreas in the DP. PATIENTS AND METHOD: From 2008 to 2016, records of 75 patients who underwent DP were retrospectively reviewed. The relationship between the perioperative factors and clinically relevant POPF was investigated. RESULTS: Univariate analysis showed that body mass index, thickness of the pancreas, and pre-firing compression were significantly related with clinically relevant POPF. Multivariate analysis showed that the pre-firing compression was an independent factor of clinically relevant POPF (OR = 44.31, 95%CI = 3.394-578.3, P = 0.004). CONCLUSIONS: Pre-firing compression of the pancreas can prevent clinically relevant POPF in DP.

Laparoscopic caudal approach for a small hepatocellular carcinoma located in the paracaval portion of a cirrhotic liver.

Resection of a tumor located in the paracaval portion of the liver, without sacrificing other parts of the liver, is surgically challenging. Here, we describe using a laparoscopic caudal approach for a small hepatocellular carcinoma located in the paracaval portion. It is the first case using the laparoscopic caudal approach to be reported in the literature. Compared with the conventional approaches, this procedure can minimize both surgical invasiveness and the volume of liver parenchyma to be resected. In addition, the laparoscopic view is suitable for this procedure. The laparoscopic caudal approach can be a safe procedure with minimal invasiveness for a small paracaval tumor, particularly in patients with liver cirrhosis.

Benefit of laparoscopic liver resection in high body mass index patients.

AIM: To explore the impact of body mass index (BMI) on surgical outcomes in patients undergoing laparoscopic liver resection (LLR). METHODS: From January 2010 to February 2015, sixty-eight patients who underwent primary partial liver resection in our institute were retrospectively reviewed. Surgical outcomes of LLR were compared with those of open liver resection (OLR). In addition, we analyzed associations with BMI and surgical outcomes. RESULTS: Among 68 patients, thirty-nine patients underwent LLR and 29 were performed OLR. Significant difference in operation time, blood loss, and postoperative hospital stay was observed. There were no significant differences in mortality and morbidity in two groups. Twenty-two patients (32.4%) were classified as obese (BMI ≥ 25). A statistically significant correlation was observed between BMI and operation time, between BMI and blood loss in OLR, but not in LLR. The operation time and blood loss of OLR were significantly higher than that of LLR in obese patients. Open liver resection and BMI were independent predictors for prolonged operation time and increased blood loss in multivariate analysis. CONCLUSION: The present study demonstrated that BMI had influenced to surgical outcomes of OLR. LLR was less influenced by BMI and had great benefit in obese patients.

Clinical Utility of the Difficulty Scoring System for Predicting Surgical Time of Laparoscopic Liver Resection.

INTRODUCTION: Laparoscopic liver resection (LLR) has several problems, such as technical complexity and patient and tumor factors. A difficulty scoring system to preoperatively evaluate the difficulty of LLR was established at the second International Consensus Conference on Laparoscopic Liver Resection. The aim of this study was to explore the clinical usefulness of the difficulty scoring system for LLR. PATIENTS AND METHODS: From January 2010 to June 2015, the records of 78 patients who had undergone LLR were retrospectively reviewed. The patients' data were used to assign a difficulty score, and the correlations between scores and surgical outcomes were investigated. RESULTS: The mean surgical time was 256 minutes, and the mean blood loss was 168 mL. The difficulty score significantly correlated with surgical time and blood loss, but not with morbidity and postoperative hospital stay. Multivariate analysis showed that the difficulty score was an independent predictor of prolonged surgical time. CONCLUSIONS: The difficulty score was found to be an effective predictor of surgical time for LLR.

Biotin-rich intranuclear inclusions in morule-lacking adenocarcinoma of the gallbladder: a new category of "neoplastic/non-morular" lesions.

Biotin-rich intranuclear inclusions, also called "optically clear nuclei," are observed in various neoplastic and non-neoplastic lesions, including pregnancy-related endometrium and benign and malignant neoplasms with morular structures. A recent study reported that lesions with biotin-rich intranuclear inclusions can be classified as "(non-neoplastic) pregnancy-related endometrial" and as "(neoplastic) morular" category. In the present report, we describe two cases of well-differentiated adenocarcinoma of the gallbladder in which biotin-rich intranuclear inclusions were found without morular structures. Immunohistochemically, as reported previously, the intranuclear inclusions were positive for biotin and two biotin-binding enzymes (pyruvic acid carboxylase and propionyl CoA carboxylase). Intranuclear expression of beta-catenin was also observed in neoplastic cells with and without intranuclear inclusion. We also detected a frame shift mutation of APC gene in one case but no mutation of beta-catenin gene in both cases. Although intranuclear expression of beta-catenin by mutation of APC gene might contribute to carcinogenesis in our cases, the relationships among intranuclear expressions of beta-catenin, biotin, biotin-binding enzymes and intranuclear inclusions remain unclear. Our cases are the first neoplastic lesions with biotin-rich intranuclear inclusions that lacked morular structures. We propose a new "neoplastic/non-morular" category for lesions with biotin-rich intranuclear inclusions.

Surgical Outcomes of Laparoscopic Liver Resection in Elderly Patients: A Comparative Study From a Single Center.

The life expectancy continues to gradually increase worldwide. Laparoscopic liver resection (LLR) was recently reported to be a relatively safe procedure for treatment of liver disease because of the development of new techniques and instruments. The aim of this study was to evaluate the surgical outcomes of elderly patients who underwent LLR. Of these 61 patients who underwent LLR in our institute from January 2010 through April 2014, 21 were aged 75 years and above (group E), and 40 were aged below 75 years (group NE). Patient characteristics were retrospectively analyzed between the 2 groups. The results showed that patient characteristics were similar between the 2 groups, although the incidence of hypertension was significantly greater among patients in group E. There were no significant differences in the incidence of postoperative complications or length of hospital stay between the 2 groups. In conclusion, LLR for elderly patients was a feasible procedure compared with nonelderly patients.

Duodenal gangliocytic paraganglioma treated with endoscopic hemostasis and resection.

Gangliocytic paragangliomas are exceedingly rare tumors that arise in close proximity to the papilla of Vater. There are few reports of the endoscopic resection of duodenal gangliocytic paraganglioma. A 61-year-old woman was admitted with a complaint of melena. Endoscopic examination revealed a pedunculated submucosal tumor with erosion in the third portion of the duodenum. Hemostasis, using a gold probe, was performed. Nine days later, we successfully resected the tumor, using endoscopic polypectomy. To determine the depth of tumor invasion, endoscopic ultrasonography was used. The size of the tumor was 3.0 x 2.5 x 1.0 cm. A total of 25 cases of duodenal gangliocytic paraganglioma have been reported in Japan. Generally, this tumor is considered benign. However, resection was performed in many patients because preoperative diagnosis was impossible. In Japan, no previous studies have reported using endoscopic hemostasis, to our knowledge. Our patient is the fourth in Japan to be treated by endoscopic resection. We report on our patient, with a review of the literature.

Protease-activating-receptor-2 is frequently expressed in papillary adenocarcinoma of the gallbladder.

Gallbladder carcinoma is one of the most devastating malignant tumors in Japan. An important risk factor for gallbladder carcinoma is pancreaticobiliary maljunction (PBM), which allows reciprocal reflux of bile and pancreatic juice. Protease-activated-receptor-2 (PAR-2), which is activated by trypsin, may be a key molecule in the process of carcinogenesis in the gallbladder epithelium. We investigated the relation between the expression of PAR-2 and clinicopathological findings in gallbladder carcinoma. The study group comprised 58 patients with gallbladder carcinoma. PAR-2 expression was identified by immunohistochemical staining of all tumor specimens. PAR-2 was expressed in cancerous gallbladder epithelium in 37 of 58 patients (64%). PAR-2 expression occurred more frequently in papillary adenocarcinoma (15 of 16 patients, 94%) than in non-papillary types (20 of 42 patients, 48%, p=0.005). Neither lymphatic invasion (p=0.03) nor venous invasion (p=0.009) occurred more frequently in gallbladder carcinoma with PAR-2 than in that without PAR-2. PAR-2 expression was not directly related to PBM (p=0.46). Papillary adenocarcinoma was associated with polypoid growth (p=0.01), PBM (p=0.01), decreased invasion to lymphatic (p=0.007) and venous vessels (p=0.005), lower T-factor (p<0.001), and lower clinical stage (p=0.02). PAR-2 is frequently expressed in papillary adenocarcinoma of the gallbladder. Trypsin may play an important role for carcinogenesis of the gallbladder through PAR-2 signaling.

Factors influencing recurrence after surgical treatment for T2 gallbladder carcinoma.

BACKGROUND/AIMS: The overall outcome of T2 gallbladder carcinoma has not been favorable, although there is a modest hope for long-term survival after radical resection. The aim of this study was to examine factors influencing postoperative disease-free survival of patients with T2 gallbladder carcinoma to clarify optimal treatment. METHODOLOGY: Of 53 patients with gallbladder carcinoma who had undergone surgical resection from 1985 to 2000, 22 had T2 carcinoma histologically proved. The significance of variables for disease-free survival was examined retrospectively by the Kaplan-Meier method and the log-rank test. RESULTS: There were 16 patients with stage II (T2N0M0), 6 with stage III (T2N1M0) disease. Eleven patients were treated by extended cholecystectomy with resection of the extrahepatic bile duct, 10 patients underwent extended cholecystectomy without resection of the extrahepatic bile duct, and 1 patient underwent cholecystectomy. All patients underwent lymph node dissection in the hepatoduodenal ligament, below the pancreatic head, and along the common hepatic artery. Lymph node metastasis was present in 6 patients. Lymphatic, venous, and perineural invasions were found in 9, 4, and 4 patients, respectively. The absence of lymphatic invasion was a significant factor related to good postoperative disease-free survival (5-year disease-free survival rate, 88.9% vs. 31.3% in the presence of lymphatic invasion). Lymph node, venous, or perineural invasion, and surgical procedure were not significant factors to good postoperative disease-free survival. CONCLUSIONS: For patients with T2 gallbladder carcinoma, the presence of lymphatic invasion is an unfavorable prognostic indicator that calls for additional treatment after radical surgery.

Clinical significance of combined pancreas and portal vein resection in surgery for pancreatic adenocarcinoma.

BACKGROUND/AIMS: Surgical resection remains the only potentially curative treatment for pancreatic adenocarcinoma for which the resectability and prognosis are still poor. The aim of the present study was to evaluate the efficacy of portal vein resection for pancreatic adenocarcinoma. METHODOLOGY: Between August 1983 and December 2000, 69 patients with pancreatic ductal cell carcinoma underwent resection in our department; 22 of the 69 had combined resection of the pancreas and portal vein. When the pancreas could not be separated from the portal vein, the vein was judged to be invaded by cancer and resected. RESULTS: The mortality rate for portal vein resection was 4.5%, which was similar to that in 47 patients with no resection of the portal vein (2.1%). Postoperative histologic analysis showed that 8 (37%) of the patients who underwent portal vein resection did not have cancer invasion to the portal vein, and 3 of them remain disease free to date. The 3-year survival rate of patients undergoing portal vein resection was 21.3%, and that of patients without portal vein resection was 20.0%. CONCLUSIONS: Resection of the portal vein in cases of pancreatic ductal cell carcinoma has no adverse affect on long-term survival for selected patients.

Mammalian target of rapamycin signaling activation patterns in pancreatic neuroendocrine tumors.

BACKGROUND: Phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (mTOR) pathway dysregulation has been implicated in the development of various human cancers. However, expression of mTOR cascade components in pancreatic neuroendocrine tumors (PNETs) has not been fully explored. The aim of this study was to assess the expression of mTOR pathway in PNETs using immunohistochemistry. METHODS: From December 1984 to April 2012, we surgically treated 42 patients with PNETs. We used immunohistochemistry to evaluate expression of mTOR, phosphorylated mTOR (p-mTOR), p70S6 kinase (S6K), phosphorylated S6 ribosomal protein (p-S6rp), eukaryotic initiation factor 4E-binding protein 1 (4E-BP1), and phosphorylated 4E-BP1 (p-4E-BP1) in the resected specimens. The relation between the expression of these molecules and clinicopathological characteristics was investigated. RESULTS: We identified the expression of mTOR (28.6%), p-mTOR (52.4%), S6K (52.4%), p-S6rp (40.5%), 4E-BP1 (81.0%), and p-4E-BP1 (26.2%) in PNETs. The expression of mTOR, p-mTOR, S6K, and p-S6rp was significantly associated with tumor invasion, proliferation, and an advanced-stage. Particularly, the expression of p-mTOR was related to clinically relevant factors such as tumor size, vascular invasion, extrapancreatic invasion, lymph node and/or distant metastasis, mitotic count, and European Neuroendocrine Tumor Society TNM staging as well as the 2004 and 2010 World Health Organization (WHO) classification. In addition, p-S6rp expression was related to vascular invasion, extrapancreatic invasion, lymph node and distant metastasis, mitotic count, and the 2010 WHO classification. In contrast, no significant relation between 4E-BP1 activation and clinicopathological factors was observed. The expression of p-mTOR was strongly correlated with that of p-S6rp (r = 0.474, P = 0.002). CONCLUSIONS: Our results suggest that activation of the mTOR/S6K signaling pathway plays a significant role in tumorigenesis and progression of PNET.

Risk factors of liver failure after right-sided hepatectomy.

BACKGROUND: To prevent hepatic failure after major hepatectomy, it is important to assess preoperative factors related to liver failure. METHODS: We examined 80 patients who underwent right-sided hepatectomy. Hyperbilirubinemia, uncontrolled ascites, and prolonged postoperative hospital stay were defined as liver failure after hepatectomy, and these 3 factors were evaluated in relation to clinicopathological and surgical factors. RESULTS: In the 80 patients, hyperbilirubinemia was observed in 10 (12.7%) patients, uncontrolled ascites in 18 (22.5%) patients, and prolonged hospital stay after surgery in 39 (48.8%) patients. Multivariate analyses identified platelet count as a risk factor of hyperbilirubinemia, uncontrolled ascites, and prolonged postoperative hospital stay, and the ratio of remnant liver volume to body surface area (RLV/BSA ratio) as an additional risk factor of hyperbilirubinemia and prolonged postoperative hospital stay. CONCLUSIONS: Platelet count and RLV/BSA ratio are useful risk factors for prediction of liver failure after right-sided hepatectomy.