Wen-Guang Wu

Whole-exome and targeted gene sequencing of gallbladder carcinoma identifies recurrent mutations in the ErbB pathway

Individuals with gallbladder carcinoma (GBC), the most aggressive malignancy of the biliary tract, have a poor prognosis. Here we report the identification of somatic mutations for GBC in 57 tumor-normal pairs through a combination of exome sequencing and ultra-deep sequencing of cancer-related genes. The mutation pattern is defined by a dominant prevalence of C>T mutations at TCN sites. Genes with a significant frequency (false discovery rate (FDR) < 0.05) of non-silent mutations include TP53 (47.1%), KRAS (7.8%) and ERBB3 (11.8%). Moreover, ErbB signaling (including EGFR, ERBB2, ERBB3, ERBB4 and their downstream genes) is the most extensively mutated pathway, affecting 36.8% (21/57) of the GBC samples. Multivariate analyses further show that cases with ErbB pathway mutations have a worse outcome (P = 0.001). These findings provide insight into the somatic mutational landscape in GBC and highlight the key role of the ErbB signaling pathway in GBC pathogenesis.

MALAT1 promotes the proliferation and metastasis of gallbladder cancer cells by activating the ERK/MAPK pathway.

Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), a long non-coding RNA (lncRNA), is associated with metastasis and is an independent prognostic factor for lung cancer. Recent studies have demonstrated that MALAT1 plays an important role in other malignancies. However, little is known about the role of MALAT1 in gallbladder carcinoma (GBC), which is the most common cancer of the biliary tract and has an extremely poor prognosis. In this study, we focused on the expression, biological functions and mechanism of MALAT1 in GBC and found that MALAT1 was significantly upregulated in GBC tissues compared with corresponding non-cancerous tissues. Knockdown of MALAT1 in GBC cell lines using lentivirus-mediated RNA interference significantly inhibited the proliferation and metastasis of the GBC cells both in vitro and in vivo. Furthermore, ERK/MAPK pathway was found to be inactivated in the GBC cell lines after MALAT1 knockdown. These results indicated that MALAT1 might serve as an oncogenic lncRNA that promotes proliferation and metastasis of GBC and activates the ERK/MAPK pathway

Evaluation of two inflammation-based prognostic scores in patients with resectable gallbladder carcinoma.

BackgroundSurvival after surgery for gallbladder cancer is generally poor. A number of inflammation-based prognostic scores have been established to help predict survival after surgery for several types of cancer. The objective of this study was to analyze and compare the utility of two inflammation-based prognostic scores, the Glasgow prognostic score (GPS) and the neutrophil-to-lymphocyte ratio (NLR), for predicting survival in patients with gallbladder cancer after surgery with curative intent.MethodsWe retrospectively reviewed the medical records of 85 patients with histologically confirmed, resectable gallbladder carcinoma (GBC), who were to receive curative surgery in our department. Univariate and multivariate analyses were performed to evaluate the relationship between the variables to overall survival (OS).ResultsA significant difference was detected in OS in patients with low and high GPS and NLR scores. Univariate analyses using clinicopathological characteristics revealed that tumor differentiation; tumor invasion; lymph node metastasis; tumor, node, metastasis classification system stage; positive margin status; combined common bile duct resection; serum levels of C-reactive protein, albumin, carbohydrate antigen 19-9 (CA19-9), carcinoembryonic antigen, and CA125; white blood cell count; and GPS and NLR were all associated with OS. Among these characteristics, multivariate analysis demonstrated that a high GPS was independently associated with poorer OS, together with tumor invasion, lymph node metastasis, and positive margin status.ConclusionsGPS is superior to NLR with respect to its prognostic value for patients with GBC after surgery with curative intent. GPS is not only associated with tumor progression but is also an independent marker of poor prognosis.

Overexpression of lysine specific demethylase 1 predicts worse prognosis in primary hepatocellular carcinoma patients

AIM To investigate the clinicopathological features and prognostic value of lysine specific demethylase 1 (LSD1) in hepatocellular carcinoma (HCC). METHODS We examined LSD1 expression in 60 paired liver cancer tissues and adjacent noncancerous tissues by quantitative real time polymerase chain reaction (qRT-PCR) and Western blotting. In addition, we analyzed LSD1 expression in 198 HCC samples by immunohistochemistry. The relationship between LSD1 expression, clinicopathological features and patient survival was investigated. RESULTS Immunohistochemistry, Western blotting, and qRT-PCR consistently confirmed LSD1 overexpression in HCC tissues compared to adjacent non-neoplastic tissues (P < 0.01). Additionally, immunostaining showed more LSD1-positive cells in the higher tumor stage (T3-4) and tumor grade (G3) than in the lower tumor stage (T1-2, P < 0.001) and tumor grade (G1-2, P < 0.001), respectively. Moreover, HCC patients with high LSD1 expression had significantly lower 5-year overall survival rates (P < 0.001) and lower 5-year disease-free survival rates (P < 0.001), respectively. A Cox proportional hazards model further demonstrated that LSD1 over-expression was an independent predictor of poor prognosis for both 5-year disease-free survival [hazards ratio (HR) = 1.426, 95%CI: 0.672-2.146, P < 0.001] and 5-year overall survival (HR = 2.456, 95%CI: 1.234-3.932, P < 0.001) in HCC. CONCLUSION Our data suggest for the first time that the overexpression of LSD1 protein in HCC tissues indicates tumor progression and predicts poor prognosis.

NLK is a key regulator of proliferation and migration in gallbladder carcinoma cells

Gallbladder cancer (GBC) is one of the most lethal neoplasm and is the fifth most common malignancy of gastrointestinal tract. The prognosis of gallbladder cancer is extremely terrible partially due to metastasis. Thus, understanding the molecular pathways controlling metastasis of this lethal disease may provide new targets for targeted therapeutic approach. In this study, we investigated the function of nemo-like kinase (NLK) in GBC growth and migration. Lentivirus-mediated siRNA was employed to alleviate the expression level of NLK in GBC cell lines (GBC-SD and SGC-996). Real-time PCR and western-blot analysis demonstrated that both mRNA and protein levels of NLK in GBC-SD and SGC-996 cells were decreased after infection with NLK-siRNA-expressing lentivirus (Lv-shNLK). The proliferation and in vitro tumorigenesis (colony formation) ability as well as migration of GBC-SD and SGC-996 cells with low NLK expression decreased significantly. Our results suggested that NLK is a key regulator involved in proliferation and migration of GBC, and it could be used as a potential therapeutic target for GBC.

A role for p21-activated kinase 7 in the development of gastric cancer.

p21‐activated kinase (PAK)7 (also known as PAK5) is a member of the group B PAK family of serine/threonine protein kinases, which are effectors of the small GTPases Rac and CDC42. PAK7 can promote neurite outgrowth, induce microtubule stabilization, and activate cell survival signaling pathways. However, the role of PAK7 in cancer is still poorly understood. Here, we showed that PAK7 expression was upregulated in different gastric cancer cell lines and gastric cancer tissues, as compared with human embryonic kidney 293 cells and adjacent normal tissues, respectively. The results suggested that PAK7 expression was related to gastric cancer progression. Thus, we employed lentivirus‐mediated small interfering RNA to inhibit PAK7 expression, to investigate the role of PAK7 in human gastric carcinogenesis. RNA interference efficiently downregulated expression of PAK7 in SGC‐7901 and MGC‐803 cells at both mRNA and protein levels. Knockdown of PAK7 inhibited human gastric cancer cell proliferation by inducing cell cycle arrest in G0/G1 phase, in concordance with the downregulation of CDK2, CDC25A, and cyclin D1. Our data suggest that PAK7 is a new hallmark of gastric cancer, in which PAK7 might contribute to gain of tumor growth potential, acting by affecting the expression of cell cycle regulators. Therefore, PAK7 may be an attractive candidate as a therapeutic target in gastric cancer.

Yes-associated protein 1 (YAP1) promotes human gallbladder tumor growth via activation of the AXL/MAPK pathway.

The transcriptional coactivator Yes-associated protein 1 (YAP1), a key regulator of cell proliferation and organ size in vertebrates, has been implicated in various malignancies. However, little is known about the expression and biological function of YAP1 in human gallbladder cancer (GBC). In this study we examined the clinical significance and biological functions of YAP1 in GBC and found that nuclear YAP1 and its target gene AXL were overexpressed in GBC tissues. We also observed a significant correlation between high YAP1 and AXL expression levels and worse prognosis. The depletion of YAP1 using lentivirus shRNAs significantly inhibited cell proliferation by inducing cell cycle arrest in S phase in concordance with the decrease of CDK2, CDC25A, and cyclin A, and resulted in increased cell apoptosis and invasive repression in GBC cell lines in vitro. Furthermore, knockdown of YAP1 also inhibited tumor growth in vivo. Additionally, we demonstrated that the activation of the AXL/MAPK pathway was involved in the oncogenic functions of YAP1 in GBC. These results demonstrated that YAP1 is a putative oncogene and represents a prognostic marker and potentially a novel therapeutic target for GBC.

Fibronectin promotes cell proliferation and invasion through mTOR signaling pathway activation in gallbladder cancer

Fibronectin (FN), a heterodimeric glycoprotein overexpressed in several types of tumors, has been implicated in cancer progression via the activation of integrin-mediated pro-oncogenic pathways. The FN level in human bile fluid is dramatically increased in malignant biliary diseases; however, FN expression and its biological functions in gallbladder cancer (GBC) remain unknown. In this study, we found that FN was overexpressed in GBC tissues and was associated with a worse prognosis in GBC patients. In vitro experimental studies indicated that exogenous FN significantly enhanced cell proliferation, invasion and active MMP-9 secretion in human GBC cell lines (GBC-SD and NOZ). Moreover, the key kinases of the mTOR signaling pathway, including FAK, Akt, mTOR and 4E-BP1, were markedly activated in a time-dependent manner in FN-treated GBC-SD and NOZ cells. The IHC statistical analyses validated that FN expression was positively correlated with the phosphorylation levels of the 4E-BP1 protein in GBC tissues. Furthermore, rapamycin, a specific inhibitor of mTOR, almost completely blocked FN-induced phosphorylation of 4E-BP1 and also partially abrogated the stimulatory effects of FN on GBC cell proliferation and invasion. In vivo, FN treatment significantly promoted the proliferation and metastasis of GBC cells and markedly activated Akt/mTOR/4E-BP1 signaling cascade. These findings demonstrate that FN may play a critical role in the modulation of cell proliferation and invasion via mTOR signaling pathway activation during GBC progression.

Primary closure versus T-tube drainage in laparoscopic common bile duct exploration: a meta-analysis of randomized clinical trials

PurposeTo compare the safety and effectiveness of primary closure with those of T-tube drainage in laparoscopic common bile duct exploration (LCBDE) for choledocholithiasis.MethodsA comprehensive search was performed in the PubMed, EmBase, and Cochrane Library databases. Only randomized controlled trials comparing primary closure with T-tube drainage in LCBDE were considered eligible for this meta-analysis. The analyzed outcome variables included postoperative mortality, overall morbidity, biliary complication rate, biliary leak rate, reoperation, operating time, postoperative hospital stay, time to abdominal drain removal, and retained stone. All calculations and statistical tests were performed using ReviewerManager 5.1.2 software.ResultsA total of 295 patients (148 patients with primary closure and 147 patients with T-tube drainage) from three trials were identified and analyzed. No deaths occurred in any of the trials. Primary closure showed significantly better results in terms of morbidity (risk ratio (RR), 0.51; 95% confidence interval (CI), 0.30 to 0.88), biliary complication without a combination of retained stone (RR, 0.44; 95% CI, 0.20 to 0.97), reoperation (RR, 0.16; 95% CI, 0.03 to 0.87), operating time (mean difference (MD), −20.72; 95% CI, −29.59 to −11.85), postoperative hospital stay (MD, −3.24; 95% CI, −3.96 to −2.52), and time to abdominal drainage removal (MD, −0.45; 95% CI, −0.86 to −0.04). Statistically significant differences were not found between the two methods in terms of biliary leak, biliary complication, and retained stones.ConclusionThe current meta-analysis indicates that primary closure of the common bile duct is safer and more effective than T-tube drainage for LCBDE. Therefore, we do not recommend routine performance of T-tube drainage in LCBDE.

Prognostic significance of nemo-like kinase (NLK) expression in patients with gallbladder cancer.

Nemo-like kinase (NLK), a serine/threonine protein kinase, has been implicated in tumor development and progression, and plays an important role in diverse signaling pathways by phosphorylating a variety of transcription factors. Recent studies demonstrated that altered expression of NLK was observed in various types of human cancers. However, the clinical significance of NLK expression in gallbladder cancer (GBC) remains largely unknown. In this study, we focused on the clinical significance of NLK in GBC, and found that nuclear NLK protein overexpression was frequently detected in GBC tissues. The overexpression of NLK was significantly correlated with histological grade, TNM stage, and perineural invasion. The results of Kaplan–Meier analysis indicated that a high expression level of NLK resulted in a significantly poorer prognosis of GBC patients (P = 0.002). Furthermore, multivariate Cox regression analysis showed that high NLK expression was an independent prognostic factor for GBC patients (HR = 3.077). In conclusion, overexpression of NLK is closely related to progression of GBC, and NLK could be used as a potential prognostic marker for GBC patients.