Expression of hepatic Wnt5a and its clinicopathological features in patients with hepatocellular carcinoma.

BACKGROUD: Wingless-type MMTV integration site family member 5a (Wnt5a) is involved in carcinogenesis. However, little data are available in Wnt5a signaling with hepatocellular carcinoma (HCC). In the present study, we investigated the expression of hepatic Wnt5a in HCC and the role of Wnt5a in HCC progression and outcome. METHODS: Wnt5a expression and cellular distribution in HCCs and their matched paracancerous tissues from 87 patients were analyzed with tissue microarray and immunohistochemistry and compared with hepatic Wnt3a signaling. Wnt5a expression was categorized into low or high based on immunohistochemistry. Overall survival rate of HCC patients was estimated in correlation with the hepatic Wnt5a level using Kaplan-Meier method; the survival difference between patients with low and those with high Wnt5a was compared with log-rank test; and prognostic analysis was carried out with Cox regression. RESULTS: Total incidence of Wnt5a expression in the HCC tissues was 70.1%, which was significantly lower (χ2 = 13.585, P < 0.001) than that in their paracancerous tissues (88.5%). Significant difference of Wnt5a intensity was found between HCC and their paracancerous tissues (Z = 8.463, P < 0.001). Wnt5a intensity was inversely correlated with Wnt3a signaling (r = -0.402, P < 0.001) in HCC tissues. A decrease of Wnt5a expression in relation to the clinical staging from stage I to IV and low or no staining at advanced HCC were observed. Wnt5a level was related to periportal embolus (χ2 = 11.069, P < 0.001), TNM staging (χ2 = 8.852, P < 0.05), 5-year survival (χ2 = 4.961, P < 0.05), and confirmed as an independent prognosis factor of HCC patients (hazard ratio: 1.957; 95% confidence interval: 1.109-3.456; P < 0.05). CONCLUSIONS: The decrease of hepatic Wnt5a signaling is associated with HCC progression and poor prognosis.

Abnormality of Wnt3a expression as novel specific biomarker for diagnosis and differentiation of hepatocellular carcinoma.

The member 3a of Wingless-type MMTV integration site family (Wnt3a) as an oncogene is overexpressed in many kinds of tumors with a worse outcome. However, the mechanism and alteration of Wnt3a expression in hepatocellular carcinoma (HCC) have not been clarified. In this study, the levels of Wnt3a expression were investigated in 80 HCC tissues or sera of 186 patients with chronic liver diseases. The incidence of hepatic Wnt3a expression in HCC tissues was 96.25 % and significantly higher (χ (2) = 48.818, P < 0.001) than that in their surrounding tissues (46.25 %). The higher level (>800 ng/L) of circulating Wnt3a expression was found in 92.5 % HCC patients and significantly related (P < 0.05) to alpha-fetoprotein (AFP) level, liver cirrhosis, hepatitis B virus infection, poor differentiation, tumor node metastasis, and extra-hepatic metastasis. The level of Wnt3a expression in HCC patients was obviously higher (P < 0.001) than that in any group of cases with benign liver diseases. The diagnostic specificity or the area under the receiver operating characteristic curve was 94.34 % or 0.994 in Wnt3a and 69.81 % or 0.710 in AFP for HCC, respectively. The present data suggested that Wnt3a expression associated with tumor progression should be a novel specific biomarker for diagnosis and differentiation of HCC.

Diagnostic and prognostic significance of secretory clusterin expression in patients with hepatocellular carcinoma.

The upregulation of secretory clusterin (sCLU) is associated with tumor progression by contributing to angiogenesis, chemo-resistance, cell survival, and metastasis. However, its diagnostic or prognostic values for hepatocellular carcinoma (HCC) still remain to be clarified. The average serum sCLU level analyzed by an enzyme-linked immunosorbent assay was significantly higher (P < 0.001) in HCC patients than that in any of cases with cirrhosis, chronic hepatitis, or healthy control. The area under receiver operating characteristic curve and diagnostic sensitivity were 0.75 and 74.7 % in sCLU, and 0.74 and 58.7 % in α-fetoprotein (AFP), respectively. The combining detections of sCLU and AFP rose up to 90.7 % for HCC diagnosis. In liver, sCLU by immunohistochemistry was significantly higher (P < 0.001) in the HCC (77.3 %) group than that in their para-cancerous group (33.3 %). Abnormal serum or tissue sCLU expression was closely associated with tumor-node-metastasis (TNM) classification of malignant tumors and lymph node metastasis, as an independent prognosis factor (hazard ratio, 2.287; 95 % confidence interval, 1.044-5.007; P = 0.039), and higher sCLU expression significantly correlated (χ (2) = 4.252, P = 0.039) with poor survival of HCC patients analyzed by multivariate Cox regression or Kaplan-Meier method, suggesting that abnormal sCLU expression associated with tumor progression could be a potential diagnostic and prognostic biomarker for HCC.

Glypican-3 is a biomarker and a therapeutic target of hepatocellular carcinoma.

BACKGROUND: The carcinogenesis of hepatocellular carcinoma (HCC) is a multi-factorial, multi-step and complex process. Early diagnosis and effective treatments are of utmost importance. This review summarized the recent studies of oncofetal glypican-3 (GPC-3), a membrane-associated heparan sulfate proteoglycan, in the diagnosis and treatment of HCC. DATA SOURCES: English-language reports published from June 2001 to September 2014 were searched from MEDLINE. The key words searched included: GPC-3, biomarker, target and HCC. The sensitivity, specificity, positive and negative predictive values were extracted, and the effect of GPC-3 targeted therapy on HCC was also evaluated. RESULTS: GPC-3 plays a crucial role in HCC cell proliferation and metastasis. It mediates oncogenesis involving signaling pathways during hepatocyte malignant transformation. GPC-3 expression is increased in atypical hyperplasia and cancerous tissues. GPC-3 levels in HCC patients are related to HBV infection, TNM stage, periportal cancerous embolus, and extrahepatic metastasis. The diagnostic accuracy of the combination of serum GPC-3 and alpha-fetoprotein in HCC is up to 94.3%. Down-regulation of GPC-3 with specific siRNA or anti-GPC-3 antibody alters cell migration, metastasis and invasion behaviors. The nude mice xenograft tumor growth is inhibited by silencing GPC-3 gene transcription. CONCLUSION: Oncofetal GPC-3 is a highly specific biomarker for the diagnosis of HCC and a promising target molecule for HCC gene therapy.

[Down-regulated clusterin expression enhances sensitivity of hepatoma cells to anti-cancer drugs].

OBJECTIVE: To investigate the relationship between and underlying mechanistic pathway of clusterin (CLU) and chemo-resistance ofhepatocellular carcinoma (HCC) cells. METHODS: CLU protein expression in HCC cell lines (Hep3B, SMMC7721, PLC, and HepG2) and HepG2/ADM cells was quantified by western blotting. Four short-hairpin (sh)RNAs designed to block CLU-mRNA were generated, screened by RT-PCR, and transfected into the cells to determine effects of CLU on cell viability and apoptosis. Effects of CLU blockade on drug efflux pump activity were measured by flow cytometry. RESULTS: CLU was found to be over-expressed in HCC cell lines and HepG2/ADM cells. The four shRNAs inhibited CLU-mRNA as follows (vs. levels in untransfected cells): shRNA-1: 73.68% (q =23.011, P < 0.01), shRNA-2: 39.26% (q =11.991, P < 0.01), shRNA-3: 62.36% (q =19.392, P < 0.01), and shRNA-4: 55.35% (q =17.149, P < 0.01). shRNA-mediated depletion of CLU led to increased sensitivity to anti-cancer drugs and increased doxorubicin-induced apoptosis in HepG2/ADM cells, as evidenced by the apoptosis ratio of the shRNA-1 group of 39.28% vs. the apoptosis ratio of the untransfected control group of 4.92%. Silencing of CLU also decreased drug etflux pump activity, and the level of MDR1/P-gp expression was significantly reduced (shRNA-1 group vs.untransfected control group: q =14.604, P < 0.01). CONCLUSION: CLU repression may enhance sensitivity of HCC cells to anti-cancers drugs and represents a potential molecular-target for reversal of multidrug-resistant HCC.

Clinicopathological, metastatic and prognostic features of stage IV esophageal adenocarcinoma versus squamous cell carcinoma: a SEER database analysis.

BACKGROUND: It is important to note that although the current treatment for advanced esophageal cancer (EC) has made great technological advances, patients' 5-year survival rates do not appear to be encouraging. Therefore, understanding the clinicopathological features and metastasis patterns of the patients with stage IV EC, combined with the prognosis of these patients, can aid in choosing the optimal treatment plan. It is well known that esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EAC) are the two most common pathological types. The aim of this study is to examine and compare the clinicopathological features and metastatic modes of stage IV ESCC and EAC, as well as their prognosis and survival. METHODS: Based on the Surveillance, Epidemiology, and End Results (SEER) database, we assessed the characteristics of ESCCs and EACs associated with prognosis using the Kaplan-Meier survival analysis, and the Cox regression model. Furthermore, the clinical data of 217 patients with stage IV ESCC and EAC in the Department of Gastroenterology of the Second Affiliated Hospital of Nantong University between 2014 and 2016 were reviewed. RESULTS: A total of 3,707 cases treated between 2010 and 2016 were included. The incidence of EAC in the United States is much higher than that of ESCC. Common metastasis patterns were lungs only, liver only, bones only, and lung & liver. The multivariate Cox analysis showed that treatment mode and metastasis patterns were independent risk factors affecting the overall survival (OS) time of patients (stage IV ESCC & EAC). EAC patients with only lung metastases may have a longer survival if chose treatment options that included surgery. In the external cohort, a total of 217 cases were included. The prevalence of ESCC is much higher than that of EAC, and the common metastasis patterns are liver only, lung only, and liver & lung. The multivariate Cox analysis showed that treatment mode was independent risk factor affecting the OS time of patients (stage IV ESCC & EAC). EAC patients treated with surgery combined with chemoradiotherapy may have a better prognosis. CONCLUSIONS: In general, the prognosis of patients with stage IV ESCC and EAC are poor. However, surgery was found to significantly improve the OS time of patients with stage IV EAC in this study.

Delivering Singlet Oxygen in Dark Condition With an Anthracene-Functionalized Semiconducting Compound for Enhanced Phototheranostics.

Photodynamic therapy (PDT) utilizes the photogeneration of reactive oxygen species (ROS) with high cytotoxicity to kill cancer cells, holding great promise for cancer treatment. Fractionated delivery of singlet oxygen (1O2) is a wise approach to relieving hypoxia, thus enhancing the therapeutic efficacy. In this article, an anthracene-functionalized semiconducting compound (DPPA) has been designed and synthesized. With irradiation, the compound is able to undergo efficient intersystem crossing (ISC) and non-radioactive decay for photodynamic/photothermal synergistic therapy. In addition, the anthracene module is able to capture and release 1O2 reversibly with or without irradiation. DPPA nanoparticles (NPs) obtained by nanoprecipitation with DSPE-PEG exhibit considerable high phototoxicity on human kidney cancer cells (A498), and the half maximum inhibitory concentration (IC50) is 15.8 µg/ml. Furthermore, an in vivo study demonstrates that complete tumor suppression was observed when the mice were administered DPPA NPs with the help of laser, compared with the control and dark groups. The H&E analysis of the normal tissues (the heart, liver, spleen, lungs, and kidney) indicates that such NPs cause no side effects, indicating the biosafety of DPPA NPs. The results provide a strategy to design a heavy-atom-free photosensitizer for photothermal and fractionated PDT against kidney tumors.

Oncogenic Wnt3a: A Candidate Specific Marker and Novel Molecular Target for Hepatocellular Carcinoma.

Background and aim: It is of the utmost importance for the specific diagnosis and effective therapy of hepatocellular carcinoma (HCC). Abnormality of oncogenic Wingless-type MMTV integration site family member 3a (Wnt3a) has been associated with progression of HCC. In this study, we aimed to evaluate Wnt3a as a novel biomarker and target for HCC. Methods: Circulating Wnt3a levels were quantitatively detected in a cohort of chronic liver diseases by an enzyme-linked immune-absorbent assay (ELISA). Hepatic Wnt3a expression in HCC and para-cancerous tissues was analyzed by immunohistochemistry. Prognostic value of Wnt3a for HCC was discovered in the cohort from the Cancer Genome Atlas (TCGA). Dynamic alterations of Wnt3a levels were detected in the hepatocarcinogenesis model induced by 2-acetylaminofluorene. Effects of Wnt3a on biological behaviors were evaluated in vitro and in vivo based on Crispr/Cas9. Results: Up-regulated Wnt3a levels were observed in serum of HCC patients with high specificity and sensitivity for HCC diagnosis. Combination of Wnt3a and AFP could improve sensitivity to 93.9% in serological detection. In addition, Wnt3a expression in HCC tissues was significantly higher than that in para-cancerous tissues. The cohort of TCGA demonstrated that high Wnt3a expression led to a poor survival of HCC patients, especially in cases at advanced stages. Furthermore, the hepatocarcinogenesis model showed that Wnt3a dynamically increased in the development of HCC. Functionally, silencing Wnt3a by Crispr/Cas9 suppressed the proliferation, colony formation, induced cell cycle arrest of HCC cells by de-activating Wnt/ß-catenin pathway in vitro, and inhibited xenograft tumor growth in vivo. Conclusions: Oncogenic Wnt3a could be considered as a candidate biomarker and novel target for HCC.

Expression of oncofetal antigen glypican-3 associates significantly with poor prognosis in HBV-related hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with poor prognosis. However, its prognostic evaluation is still an urgent problem. The objectives of this present study were to investigate oncofetal antigen glypican-3 (GPC-3) expression in HCC and their match para-cancerous tissues by the array technology with immunohistochemistry and estimate its value as a novel prognostic marker for HCC. The incidence of GPC-3 expression was 95.7 % in the cancerous tissues with significantly higher (χ2 = 33.824, P < 0.001) than that in the para-cancerous tissues (52.2 %). Abnormal expression of GPC-3 in HCC tissues was markedly related to poor or moderate differentiation (P < 0.001), hepatitis B virus (HBV) infection (P = 0.004), periportal cancer embolus (P = 0.043), and tumor-node- metastasis staging (P = 0.038). According to the univariate and multivariate analysis, the overall survival of HCC patients with high GPC-3 level was significantly worse than those with low or without GPC-3 expression (P < 0.001), suggesting that abnormal GPC-3 expression should be an independent prognostic factor for HBV-related HCC patient's survival.

Oncogenic Wnt3a expression as an estimable prognostic marker for hepatocellular carcinoma.

AIM: To investigate member 3a of Wingless-type MMTV integration site family (Wnt3a) expression in cancerous and surrounding tissues and the relationship between clinicopathologic features of hepatocellular carcinoma (HCC) and Wnt3a expression. METHODS: Wnt3a expression and cellular distribution and clinicopathologic characteristics in cancerous tissue and matched surrounding tissues were analyzed in 80 HCC patients from January 2006 to August 2008 by tissue microarrays and immunohistochemistry. The overall and disease-free survival rates were estimated using the Kaplan-Meier method and compared with the log-rank test. The prognostic analysis was carried out with univariate and multivariate Cox regressions models. RESULTS: The incidence of oncogenic Wnt3a expression in the cancerous group was up to 96.25% (77 of 80), which was significantly higher (χ(2) = 48.818, P < 0.001) than that in the surrounding group (46.25%, 37 of 80). Brown Wnt3a staining gradually increased with clinical staging that showed very strong staining in advanced HCC. The clinicopathologic features of high Wnt3a expression in HCC were related to poorly-differentiated grade (χ(2) = 20.211, P < 0.001), liver cirrhosis (χ(2) = 8.467, P < 0.004), hepatitis B virus (HBV) infection (χ(2) = 12.957, P < 0.001), higher tumor-node-metastasis stage (χ(2) = 22.960, P < 0.001), and 5-year survival rate (χ(2) = 15.469, P < 0.001). CONCLUSION: Oncogenic Wnt3a expression associated with HBV infection and cirrhotic liver might be an independent prognostic factor for HCC.

Abnormal Expression of Golgi Protein 73 in Clinical Values and Their Role in HBV-Related Hepatocellular Carcinoma Diagnosis and Prognosis.

BACKGROUND: The up-regulation of hepatic Golgi protein 73 (GP73) is associated with the progression of hepatocellular carcinoma (HCC). However, the exact mechanism and clinical values of its diagnosis and prognosis still need to be clarified. OBJECTIVES: To investigate the clinical values of abnormal liver or circulating GP73 expression and their effect on HCC diagnosis and prognosis. MATERIALS AND METHODS: The expression of GP73 was investigated in 88 cancerous and self-control non-cancerous tissues using tissue microarrays with immunohisto- chemistry and was confirmed by Western blotting. Circulating GP73 levels were detected in the sera of 281 patients with liver diseases using enzyme-linked immunosorbent assay. RESULTS: The levels of circulating GP73 expression in the HCC group were higher than those in any group of benign liver diseases or controls. No significant difference was found between GP73 expression and patients' sex or age, tumor size, or AFP level except for those with hepatitis B virus (HBV) infection or distal metastasis (P < 0.05). The area under the receiver operating characteristic curve, sensitivity, and specificity for HCC diagnosis were 0.881, 78.34%, and 77.59% for GP73 levels over 70 µg/L or 0.754, 71.97%, and 84.48% for alpha-fetoprotein levels over 50 µg/L, respectively. The total incidence of GP73 plus alpha-fetoprotein was up to 87.26% for HCC. A positive GP73 result with brown particles was mainly located in the cytosol, with a few in the nucleus and none in the cell membrane, with abnormal expression in HCC tissues (480.7 ± 148.7) that was significantly higher (t = 10.730, P < 0.001) than those in their non-cancerous tissues (208.0 ± 66.1). The high GP73 expression in HCC was related to lymph node metastasis (χ(2) = 6.940, P = 0.008), gross classification (χ(2) = 6.311, P = 0.012), HBV (χ(2) = 4.803, P = 0.028), tumor node metastasis staging (χ(2) = 4.887, P = 0.027), and five-year survival (χ(2) = 5.206, P = 0.023). CONCLUSIONS: Abnormality of hepatic or circulating GP73 expression should be regarded as an emerging biomarker for HCC diagnosis and prognosis.