Convergent genomic diversity and novel BCAA metabolism in intrahepatic cholangiocarcinoma.

BACKGROUND: Driver alterations may represent novel candidates for driver gene-guided therapy; however, intrahepatic cholangiocarcinoma (ICC) with multiple genomic aberrations makes them intractable. Therefore, the pathogenesis and metabolic changes of ICC need to be understood to develop new treatment strategies. We aimed to unravel the evolution of ICC and identify ICC-specific metabolic characteristics to investigate the metabolic pathway associated with ICC development using multiregional sampling to encompass the intra- and inter-tumoral heterogeneity. METHODS: We performed the genomic, transcriptomic, proteomic and metabolomic analysis of 39-77 ICC tumour samples and eleven normal samples. Further, we analysed their cell proliferation and viability. RESULTS: We demonstrated that intra-tumoral heterogeneity of ICCs with distinct driver genes per case exhibited neutral evolution, regardless of their tumour stage. Upregulation of BCAT1 and BCAT2 indicated the involvement of 'Val Leu Ile degradation pathway'. ICCs exhibit the accumulation of ubiquitous metabolites, such as branched-chain amino acids including valine, leucine, and isoleucine, to negatively affect cancer prognosis. We revealed that this metabolic pathway was almost ubiquitously altered in all cases with genomic diversity and might play important roles in tumour progression and overall survival. CONCLUSIONS: We propose a novel ICC onco-metabolic pathway that could enable the development of new therapeutic interventions.

Expression of mesenchymal markers vimentin and fibronectin: the clinical significance in esophageal squamous cell carcinoma.

BACKGROUND: The importance of mesenchymal characteristics has not been fully elucidated in esophageal cancer. METHODS: Ten normal and 77 tumor specimens were collected. Microarray analysis was performed to analyze the expression patterns of epithelial markers, mesenchymal markers, epithelial mesenchymal transition (EMT)-related genes and stem cell markers. RT-PCR analysis was conducted to confirm the results of microarray analysis. Immunohistochemical analysis was performed to verify the level of protein expression. Statistical analysis was performed to investigate the correlation between selected genes and clinicopathological factors. RESULTS: Microarray analysis showed that epithelial markers were significantly down-regulated whereas mesenchymal markers and EMT transcription factors were up-regulated in cancer cells. Two types of gene expression patterns were found in the clustering analysis, type 1 tumors and type 2 tumors. Type 1 tumor clusters did not reveal a fixed gene expression pattern whereas type 2 tumor clusters revealed up-regulation of mesenchymal markers EMT inducers and related genes. Vimentin and fibronectin were selected to distinguish between tumor types 1 and 2. Type 2 tumors showed significantly larger tumor sizes (p < 0.0001), wider ranges of lymph node metastasis (p = 0.0057), and a more severe clinical stage (p < 0.0001) than did type 1 tumors. The prognosis of patients with type 2 tumors was significantly worse than that of patients with type 1 tumors. Univariate and multivariate analyses revealed that classification of type 2 tumors was an independent prognostic factor. CONCLUSIONS: The analysis of mesenchymal markers in esophageal cancer is useful in distinguishing patients with a poor prognosis.

Clinical significance of PICT1 in patients of hepatocellular carcinoma with wild-type TP53.

BACKGROUND: TP53 is one of the most widely known cancer suppressor genes. Mutations in TP53 are ubiquitously observed in almost all cancers. Incidences of mutations range from ~15-70 % in patients with hepatocellular carcinoma (HCC). Moreover, patients with mutated TP53 have poorer prognoses than those with wild-type TP53; therefore, it would be beneficial to predict the prognosis of HCC patients with wild-type TP53. We previously reported that PICT1, coding a nucleolus protein, regulates TP53 through indirect association. METHODS: In this study, we examined PICT1 expression levels and the status of TP53 in 51 primary HCC tissues in order to determine the clinical significance of PICT1 expression and the function of PICT1 in HCC cells. RESULTS: We detected 6 mutations in the 51 samples. In 45 patients with wild-type TP53, those with high PICT1 expression (n = 11) had poorer prognoses than those with low PICT1 expression (n = 34), and there were no significant associations with other clinicopathological factors. According to gene set enrichment analysis, PICT1 expression was inversely correlated with the gene set of TP53. In vitro assays indicated that suppression of PICT1 expression caused an increase in TP53 expression, reduction in cell proliferation, and arrest at the G1 phase of the cell cycle in HCC cells expressing wild-type TP53. CONCLUSIONS: PICT1 should be a useful prognostic marker in HCC patients having wild-type TP53. Furthermore, PICT1 may become a promising therapeutic target because of its ability to increase the expression and activation of TP53.

Laparoscopic surgery minimizes the surgical manipulation of isolated tumor cells leading to decreased metastasis compared to open surgery for colorectal cancer.

PURPOSES: We focused on the possible benefits of laparoscopic surgery to protect against isolated tumor cells (ITC) generated by surgical manipulation in comparison to open surgery. METHODS: We performed conventional open surgery and laparoscopic surgery for 25 and 8 cases of colorectal cancer (CRC), respectively. We compared the presence of ITC in the peripheral blood (PB) immediately after surgery via quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) for a representative epithelial marker, carcinoembryonic antigen (CEA). RESULTS: In the 25 patients who underwent open surgery, 8 of the 10 cases with metastasis were positive for ITC in PB, while 13 of the 15 cases without metastasis were negative for ITC. Therefore, we validated that there was a significant clinical usefulness for the detection of ITC in the prediction of metastasis (p = 0.0024). We limited our subsequent analysis to the CRC cases with a Dukes stage of B or C to avoid differences due to the background, and we found that the positive ITC rate for metastasis was higher in the 19 patients who underwent open surgery (42.1 %) than in the 8 who underwent laparoscopic surgery (37.5 %). CONCLUSIONS: The short observation period, especially in the laparoscopic surgery group, and the inadequate number of cases limit the ability to draw any definitive conclusions; however, laparoscopic surgery appears to minimize the surgical manipulation, thus leading to reduced ITC from primary CRC compared with open surgery.

Downregulation of miR-144 is associated with colorectal cancer progression via activation of mTOR signaling pathway.

The mammalian target of rapamycin (mTOR) is a downstream integrator of essential pathways. mTOR signaling is frequently dysregulated in a variety of human cancers, and in silico analysis has revealed two miR-144 binding sites in the mTOR 3' untranslated region. We investigated the clinicopathologic magnitude of the mTOR pathway regulating microRNA, miR-144 in colorectal cancer (CRC) cases. The regulation of mTOR by miR-144 was examined with inhibitor miR-144-transfected cells. We also investigated changes in sensitivity to the mTOR inhibitor, rapamycin, in inhibitor miR-144-transfected cells. Quantitative RT-PCR was used to evaluate the clinicopathologic significance of miR-144 expression in 137 CRC. Furthermore, we assessed the correlation between CRC prognosis and the expression of 16 genes in the Akt/mTOR pathway. In vitro assays showed that mTOR is a direct target of miR-144, and downregulation of miR-144 facilitated proliferation of CRC cell line, HT29. In addition, the viability of HT29 cells with downregulated miR-144 expression was significantly reduced with rapamycin treatment. Low expression levels of miR-144 were associated with enhanced malignant potential such as venous invasion (P = 0.0013), liver metastasis (P = 0.08), liver recurrence (P = 0.0058) and poor prognosis (P = 0.0041). Multivariate analysis indicated that low miR-144 expression was an independent prognostic factor for survival. Among many genes consisting of the mTOR pathway, only high expression of Rictor was associated with poor prognosis of CRC. miR-144 is a meaningful prognostic marker. Downregulation of miR-144 leads to poor prognosis of CRC patients via activation of the mTOR signaling pathway.

MicroRNA-10b is a prognostic indicator in colorectal cancer and confers resistance to the chemotherapeutic agent 5-fluorouracil in colorectal cancer cells.

PURPOSE: Recent evidence has shown that altered patterns of microRNA (miRNA) expression correlate with various human cancers. We investigated the clinical significance of miR-10b and its involvement in chemotherapeutic resistance to 5-fluorouracil (5-FU), which is a key component of common chemotherapy regimens in colorectal cancer. METHODS: Quantitative RT-PCR was used to evaluate the clinicopathologic significance of miR-10b expression in 88 colorectal cancer cases. We also investigated the chemotherapeutic sensitivity to 5-FU in miR-10b-overexpressing colorectal cancer cells. To explore the mechanism of chemoresistance in miR-10b transfected cells, we examined whether miR-10b inhibits the pro-apoptotic BH3-only Bcl-2 family member BIM(BCL2L11), a key mediator of chemotherapy-induced cell death. RESULTS: High level miR-10b expression was found to be significantly associated with high incidence of lymphatic invasion (P = 0.0257) and poor prognosis (P = 0.0057). Multivariate analysis indicated that high miR-10b expression is an independent prognostic factor for survival. In vitro studies revealed that miR-10b directly inhibits pro-apoptotic BIM, and the overexpression of miR-10b confers chemoresistance in colorectal cancer cells to 5-FU. CONCLUSIONS: MiR-10b is a novel prognostic marker in colorectal cancer. Moreover, the expression of miR-10b is a potential indicator of chemosensitivity to the common 5-FU-based chemotherapy regimen.

MicroRNA-372 is associated with poor prognosis in colorectal cancer.

OBJECTIVE: MicroRNA-372 (miR-372) is reportedly shown to be an oncogene in human testicular germ cell tumors and gastric cancers, but its expression in colorectal cancer (CRC) is not yet determined. This study investigated the clinical significance of miR-372 expression in CRC. METHODS: qRT-PCR was used to evaluate miR-372 in 144 CRC patients, and large tumor suppressor 2 (LATS2) expression was also examined as the likely target gene of miR-372. In vitroassays were performed to evaluate the biological function of miR-372. RESULTS: Multivariate analysis indicated that high miR-372 expression was an independent prognostic factor (p = 0.006). High miR-372 expression was associated with synchronous liver metastasis (p = 0.035). We found an inverse relationship between miR-372 and LATS2 by qRT-PCR (p = 0.007) and immunohistochemistry (p = 0.042) using CRC tissue samples. Furthermore, pre-miR-372 led to a decrease in the LATS2 protein and an increase in proliferative activity of LoVo cells. We also found a significant association between low LATS2 expression and liver metastasis (p = 0.042). CONCLUSIONS: This study suggested that miR-372 was a novel independent prognostic factor in CRC. Our data suggest that LATS2 may serve as one of the target genes of miR-372 in clinical CRC tissues.

"Peripancreatic strands appearance" in pancreatic body and tail carcinoma: evaluation by multi-detector CT with pathological correlation.

OBJECTIVES: In pancreatic body and tail carcinoma, "peripancreatic strands appearance" is frequently seen on multidetector CT (MDCT). The purpose of this study was to clarify the pathological and clinical implications of peripancreatic strands appearance. METHODS: We retrospectively evaluated MDCT images in 17 patients with pancreatic body and tail carcinoma who underwent surgical resection. Peripancreatic strands appearance was defined as the strands structure deriving from the primary lesion and associated with increased CT attenuation of surrounding adipose tissues. All CT examinations were performed by contrast-enhanced MDCT with a multiplanar reformation technique. RESULTS: Peripancreatic strands appearance was detected on MDCT in 13 (76%) patients. The maximum width of the peripancreatic strands seen on MDCT was 1.55 ± 0.36 mm (range, 1.0-2.5 mm). This CT finding was well correlated with extrapancreatic carcinoma invasion with marked fibrotic thickening of adipose tissue septa, including microvessels. This pathological finding was confirmed in all 13 patients with positive CT finding whereas it was not confirmed in the 4 patients with negative CT finding. CONCLUSION: Peripancreatic strands appearance on MDCT in pancreatic body and tail carcinoma reflects extrapancreatic carcinoma invasion with marked fibrotic thickening of adipose tissue septa. This CT finding would indicate the property of carcinoma aggressiveness.

Review: Single nucleotide polymorphisms associated with the oncogenesis of colorectal cancer.

Colorectal cancer (CRC) is provoked by interactions between genetic and environmental factors. We herein review the incidence and the mechanisms of action of the reported single nucleotide polymorphisms (SNPs) in the oncogenesis of CRC. More than 15 reports have studied the SNPs at 8q24, which are associated with the incidence of CRC as well as prostate cancer. We have also reported a SNP at the 10p14 locus, and the risks of other loci for CRC oncogenesis. With regard to the underlying mechanism for CRC, 8q24 is a locus of the long-enhancer site for MYC, which could determine the efficacy of MYC transcription. We suggest that 8q24 SNPs might be associated with the progression of CRC cases mediated by MYC expression. In this report, we summarize the published studies of the genetic background SNPs in the oncogenesis of CRC. The level of risk (most studies indicated less than a twofold increase) for CRC was lower than anticipated. Therefore, it is necessary to identify bona fide SNPs that precisely predict the risk for CRC. Alternatively, it is important to consider other factors, such as environmental or lifestyle-related factors, for the future prevention of CRC.

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.

Two-stage laparoscopic resection of colon cancer and metastatic liver tumour.

We report herein the case of 70-year-old woman in whom colon cancer and a synchronous metastatic liver tumour were successfully resected laparoscopically. The tumours were treated in two stages. Both post-operative courses were uneventful, and there has been no recurrence during the 8 months since the second procedure.

FOXC2 is a novel prognostic factor in human esophageal squamous cell carcinoma.

BACKGROUND: FOXC2 has been implicated in cancer progression through its induction of epithelial-to-mesenchymal transition. We analyzed the clinical significance of FOXC2 in esophageal cancer cases, in which early distant metastasis or invasion to nearby organs is an obstacle to treatment. METHODS: Quantitative reverse transcriptase-polymerase chain reaction was used to evaluate FOXC2 mRNA expression in 70 esophageal cancer cases to determine the clinicopathologic significance of FOXC2 expression. Furthermore, we examined associations between FOXC2 expression and matrix metalloproteinases 2 (MMP2) and matrix metalloproteinases 9 (MMP9). We also performed in vitro invasion and migration assays for FOXC2-suppressed esophageal cancer cells. RESULTS: In clinicopathologic analysis, the high-FOXC2 expression group showed a higher incidence of advanced tumor stage, lymph node metastasis, and lymphatic invasion than the low-FOXC2 expression group (P < 0.05). In particular, tumor stage exhibited the most remarkable difference (P < 0.0001). Expression of MMP2 and MMP9 was far higher in the high-FOXC2 expression group. Furthermore, the high-FOXC2 expression group had a significantly poorer prognosis than did the low expression group (P = 0.006). Multivariate analysis indicated that high FOXC2 expression was an independent prognostic factor for survival. Suppression of FOXC2 expression altered the invasive and the migratory ability of esophageal cancer cells in vitro. CONCLUSIONS: Our findings suggest that FOXC2 could be an important prognostic indicator for esophageal cancer patients. FOXC2 is directly involved in cancer progression and is associated with poor prognosis in esophageal cancer cases.

Surgical outcomes of totally extraperitoneal repair for inguinal hernia: A retrospective multicenter propensity score-matched study.

BACKGROUND: Laparoscopic surgical approaches, including total extraperitoneal repair (TEP), have been widely accepted for inguinal hernia repair in Japan. However, there are limited data regarding recurrence after TEP in Japan, given the limited versatility of this procedure. This study retrospectively evaluated the rates of hernia recurrence after TEP and open mesh repair at multiple Japanese centers. METHODS: This retrospective study evaluated 1917 patients who underwent inguinal hernia repair at 32 institutions in the Oita prefecture between January 2014 and December 2015. Eligible patients were grouped according to whether they underwent TEP (1011 patients) or open mesh repair (636 patients). Propensity score matching was performed 1:1 (total: 1076 patients, 538 patients from each group). The outcomes of interest were recurrence, morbidity, and postoperative recovery. RESULTS: The TEP and open mesh repair groups had similar baseline characteristics. After propensity score matching, there was no significant difference between the two groups in terms of recurrence rate (TEP: 0.5% vs open mesh repair: 1.0%, P = .375). However, the TEP group had significantly longer operating times (median: 70.2 min vs 65.0 min, P < .001), significantly less blood loss (0-5.1 mL vs 0-20.4 mL, P < .001), and significantly shorter postoperative hospital stays (median: 5.0 days vs 6.4 days, P < .001). The overall incidences of morbidity were 6.2% in the TEP group and 7.2% in the open mesh repair group (P = .535). CONCLUSION: This multicenter retrospective study with propensity score matching revealed that the recurrence rates were similarly low for TEP and open mesh repair of inguinal hernia. Thus, a well-trained surgical team could use TEP as a standard procedure.

Clinical and genetic analysis in a family with familial renal glucosuria: Identification of an N101K mutation in the sodium-glucose cotransporter 2 encoded by a solute carrier family 5 member 2 gene.

We report the identification of a mutation in the solute carrier family 5 member 2 (SLC5A2) gene, which encodes sodium-glucose cotransporter 2, in a family with familial renal glucosuria. The proband was a 26-year-old Japanese man referred to the diabetes division with repeated glucosuria without hyperglycemia. His mother, uncle and grandfather also had a history of glucosuria. A heterozygous missense mutation (c.303T>A:p.N101K) in SLC5A2 was identified in the patient and his mother, but not in 200 chromosomes from 100 healthy and unrelated individuals, or in 3,408 Japanese individuals in the Tohoku Medical Megabank. Furthermore, bioinformatics software predicted that this lesion would be pathogenic. We infer that the mutation led to clinically relevant sodium-glucose cotransporter 2 dysfunction. The patient showed no symptoms of hypoglycemia, but continuous glucose monitoring confirmed asymptomatic hypoglycemia.

Reovirus inhibits the peritoneal dissemination of pancreatic cancer cells in an immunocompetent animal model.

The prognosis of pancreatic cancer with peritoneal dissemination has not improved. The aim of this study was to clarify whether oncolytic reovirus is effective against the peritoneal dissemination of pancreatic cancer in an immunocompetent animal model. The hamster pancreatic cancer cells HaP-T1 were inoculated into the peritoneal cavity of the hamster and reovirus (1x10(8) plaque-forming units) was administered into the peritoneal cavity on days 1, 3, 5 and 7 after HaP-T1 inoculations. The number and weight of the disseminated nodules in each group were recorded. Reovirus protein in the disseminated nodules was examined by immunohistochemical staining. The tumor volumes of peritoneal dissemination in the treatment group were significantly less than those in the control group (p<0.05). In addition, the amount of ascites was decreased in the treatment group in comparison to the control group. Immunohistochemical examination revealed that reovirus replication was seen only in the disseminated nodules but not in surrounding normal tissues. There were no serious side effects observed in this study. These data suggested that intraperitoneal administration of reovirus might be an effective form of oncolytic viral therapy for peritoneal dissemination of pancreatic cancer.

Increased heat-shock protein 90 expression contributes to impaired adaptive cytoprotection in the gastric mucosa of portal hypertensive rats.

BACKGROUND AND AIMS: Portal hypertensive (PHT) gastropathy results in an increased susceptibility to damage. Adaptive cytoprotection against ethanol-induced damage is impaired in the gastric mucosa of rats with portal hypertension. Excessive nitric oxide (NO) production occurs in portal hypertension and is mediated in part via heat-shock protein (Hsp)90 production. The aim of this study was to investigate the relation between adaptive cytoprotection after exposure to ethanol and gastric expression of Hsp90 in PHT rats. METHODS: Portal hypertension was induced in rats by staged portal vein occlusion. Adaptive cytoprotection to 70% ethanol was evaluated by assessing the injury index of the gastric mucosa with or without pretreatment with 10% ethanol. Expression of Hsp90 mRNA was evaluated by real-time polymerase chain reaction, and expression of Hsp90 protein was evaluated by western blotting. The effect of Hsp90 inhibition in PHT rats was evaluated by administration of geldanamycin. RESULTS: Gastric Hsp90 mRNA expression in PHT rats was significantly less than that in sham-operated (SO) controls. However, after 10% ethanol pretreatment, Hsp90 mRNA expression was significantly greater in PHT rats than in SO controls. In PHT rats, gastric Hsp90 protein expression after 10% ethanol pretreatment was significantly greater than that without the pretreatment. However, the pretreatment had no effect on the injury index compared to SO rats. Administration of geldanamycin prior to 10% ethanol pretreatment significantly decreased the injury index in response to 70% ethanol in the PHT rats. CONCLUSIONS: These results show that 10% ethanol pretreatment markedly increases gastric Hsp90 expression in PHT rats. Excessive production of Hsp90 may contribute impaired adaptive cytoprotection.

Ampullary cancer and preoperative jaundice: possible indication of the minimal surgery.

BACKGROUND/AIMS: Obstructive jaundice is common symptom of carcinoma of the ampulla of Vater. In the present study we evaluated association between preoperative jaundice and prognosis and possible indication of the minimal surgery. METHODOLOGY: Clinicopathological features of 50 patients who underwent curative resection were examined retrospectively and statistically analyzed. RESULTS: Thirty-two of the 50 patients (64%) had preoperative jaundice, and these patients had poorer survival compared with these without jaundice (5 year survival 57.2% vs. 100%, p < 0.01). Preoperative jaundice, serum carbohydrate antigen 19-9 level, pancreatic invasion, lymphatic invasion, venous invasion, perineural invasion, duodenal invasion, margin status, lymph node metastasis, and T stage were shown to be significant prognostic factors. In the 18 patients without preoperative jaundice, lymphatic invasion was significantly related to lymph node metastasis. In addition, 12 of the 14 patients without jaundice and lymphatic invasion were diagnosed as T1N0 (stage IA) disease, and all the tumors were papillary or well-differentiated adenocarcinoma. CONCLUSIONS: Preoperative jaundice may reflect advanced-stage in case of ampullary cancer. Radical surgery may not be necessary in patients without preoperative jaundice if the tumor is diagnosed as a papillary or well-differentiated adenocarcinoma without lymphatic invasion.