Xin-Rong Yang

Serum DKK1 as a protein biomarker for the diagnosis of hepatocellular carcinoma: a large-scale, multicentre study

BACKGROUNDnHepatocellular carcinoma (HCC) is prevalent worldwide and improvements in timely and effective diagnosis are needed. We assessed whether measurement of Dickkopf-1 (DKK1) in serum could improve diagnostic accuracy for HCC.nnnMETHODSnWe analysed data for patients with HCC, chronic hepatitis B virus (HBV) infection, liver cirrhosis, and healthy controls, recruited from two Chinese centres between December, 2008, and July, 2009. A validation cohort matched for age and sex was recruited from another centre in China between July, 2010, and June, 2011. DKK1 was measured in serum by ELISA by independent researchers who had no access to patients clinical information. We used receiver operating characteristics (ROC) to calculate diagnostic accuracy.nnnFINDINGSnWe assessed serum DKK1 in 831 participants: 424 with HCC, 98 with chronic HBV infection, 96 with cirrhosis, and 213 healthy controls. The validation cohort comprised 453 participants: 209 with HCC, 73 with chronic HBV infection, 72 with cirrhosis, and 99 healthy controls. Levels of DKK1 in serum were significantly higher in patients with HCC than in all controls. ROC curves showed the optimum diagnostic cutoff was 2·153 ng/mL (area under curve [AUC] 0·848 [95% CI 0·820-0·875], sensitivity 69·1%, and specificity 90·6% in the test cohort; 0·862 [0·825-0·899], 71·3%, and 87·2% in the validation cohort). Similar results were noted for early-stage HCC (0·865 [0·835-0·895], 70·9%, and 90·5% in the test cohort; 0·896 [0·846-0·947], 73·8%, and 87·2% in the validation cohort). Furthermore, DKK1 maintained diagnostic accuracy for patients with HCC who were α-fetoprotein (AFP) negative (0·841 [0·801-0·882], 70·4%, and 90·0% in the test cohort; 0·869 [0·815-0·923], 66·7%, and 87·2% in the validation cohort), including for patients with early-stage HCC (0·870 [0·829-0·911], 73·1%, and 90·0% in the test cohort; 0·893 [0·804-0·983], 72·2%, and 87·2% in the validation cohort), compared with all controls. Raised concentrations of DKK1 in serum could differentiate HCC from chronic HBV infection and cirrhosis (0·834 [0·798-0·871], 69·1%, and 84·7% in the test cohort; 0·873 [0·832-0·913], 71·3%, and 90·6% in the validation cohort). Moreover, measurement of DKK1 and AFP together improved diagnostic accuracy for HCC versus all controls compared with either test alone (0·889 [0·866-0·913], 73·3%, and 93·4% in the test cohort; 0·888 [0·856-0·920], 78·5%, and 87·2% in the validation cohort).nnnINTERPRETATIONnDKK1 could complement measurement of AFP in the diagnosis of HCC and improve identification of patients with AFP-negative HCC and distinguish HCC from non-malignant chronic liver diseases.nnnFUNDINGnNational Key Basic Research Programme of China, National Key Sci-Tech Special Projects of Infectious Diseases, National Natural Science Foundation of China, Research Fund for the Doctoral Programme of Higher Education of China.

Circulating stem cell–like epithelial cell adhesion molecule–positive tumor cells indicate poor prognosis of hepatocellular carcinoma after curative resection

Epithelial cell adhesion molecule–positive (EpCAM+) hepatocellular carcinoma (HCC) cells may constitute a tumor‐initiating subpopulation in tumorigenic cell lines and HCC specimens. In the present study, EpCAM+ circulating tumor cells (CTCs) were identified prospectively in HCC patients undergoing curative resection, and the prognostic significance and their stem cell–like characteristics were investigated further. Blood samples from 123 HCC patients were tested prior to resection and 1 month thereafter. CTCs were present in 66.67% of patients, and the cell count measured in 7.5 mL of blood (CTC7.5) ranged between 1 and 34. Fifty‐one patients had CTC7.5 of ≥2 preoperatively, and these patients developed tumor recurrence earlier than those with CTC7.5 of <2 CTCs (P < 0.001). A preoperative CTC7.5 of ≥2 was an independent prognostic factor for tumor recurrence (P < 0.001). Its prognostic significance also applied to patients with alpha‐fetoprotein (AFP) levels of ≤400 ng/mL or subgroups with low recurrence risk (all P < 0.05). A significant decrease of CTC‐positive rates (66.67% to 28.15%, P < 0.05) and CTC7.5 values (2.60 ± 0.43 to 1.00 ± 0.36, P < 0.05) was observed 1 month after resection. Patients with consistent CTC7.5 <2 had lower recurrence rates than those with values consistently ≥2 (15.5% versus 87.50%, P < 0.001). EpCAM+ CTCs displayed cancer stem cell biomarkers (CD133 and ABCG2), epithelial‐mesenchymal transition, Wnt pathway activation, high tumorigenic potential, and low apoptotic propensity. Conclusion: Stem cell–like phenotypes are observed in EpCAM+ CTCs, and a preoperative CTC7.5 of ≥2 is a novel predictor for tumor recurrence in HCC patients after surgery, especially in patient subgroups with AFP levels of ≤400 ng/mL or low tumor recurrence risk. EpCAM+ CTCs may serve as a real‐time parameter for monitoring treatment response and a therapeutic target in HCC recurrence. (HEPATOLOGY 2013)

Circulating tumor cells: advances in detection methods, biological issues, and clinical relevance

BackgroundCirculating tumor cells (CTCs) have long been considered a reflection of tumor aggressiveness. Hematogenous spreading of CTCs from a primary tumor is a crucial step in the metastasis cascade, which leads ultimately to the formation of overt metastases. However, owing to the rarity of CTCs in peripheral blood, detecting these cells requires methods combined with high sensitivity and specificity, which sets tremendous challenges for the implementation of these assays into clinical routine.MethodsGenerally, CTCs detection methods are composed of the following two steps: enrichment (isolation) process (morphological and immunological techniques) and detection (identification) process (cytometric and nucleic acid techniques), which may or may not be separate from enrichment. Genetic and molecular characterization of CTCs carried out by fluorescent in situ hybridization (FISH), comparative genomic hybridization (CGH), PCR-based techniques, and biomarker immunofluorescent staining extract more information about malignant profile, metastatic potential of CTCs, and the extent to which CTCs are genetically identical to the primary tumor.ResultsRecent technical advances made it possible to detect CTCs. The efficacy of circulating tumor cell (CTC) detection among patients with solid malignancy has been investigated, which shows great potential to become a tool for real-time parameter of prognosis and serve as an early marker to assess the therapeutic response in overt cancers. Improvements in detection and characterization of CTCs will hopefully lead to refinement of clinical management of cancer patients.ConclusionThis review addresses the majority of assays that have been published thus far, including the enrichment and detection steps and the markers used in these assays, accompanied by some biological issues of CTC and the results of clinical application harvested.

CD151 modulates expression of matrix metalloproteinase 9 and promotes neoangiogenesis and progression of hepatocellular carcinoma.

Tetraspanin CD151 is involved in several pathological activities associated with tumor progression, including neoangiogenesis. However, the role and molecular mechanism of CD151 in the neoangiogenesis of hepatocellular carcinoma (HCC) remain enigmatic. We found that the level of expression of matrix metalloproteinase 9 (MMP9) was positively associated with CD151 expression in HCC cells. We developed a zone‐by‐zone blockade and demonstrated that overexpression of CD151 in HCC cells facilitated MMP9 expression through a phosphatidylinositol‐3‐kinase (PI3K)/protein kinase B (Akt)/glycogen synthase kinase 3β (GSK‐3β)/Snail signaling pathway. In contrast, down‐regulation of CD151 expression impaired the ability of HCC cells to form microvessels in vitro and reduced their in vivo metastatic potential. In a clinical setting, a significant correlation of the expression of CD151 with MMP9 expression and with microvessel density (MVD) was revealed by Pearson correlation analysis of HCC patients. The postoperative 3‐, 5‐, and 7‐year overall survival rates of HCC patients with CD151high/MMP9high/MVDhigh were significantly lower than those of the CD151low/MMP9low/MVDlow group or groups in which only one or two of CD151, MMP9, and MVD were highly expressed. Cumulative recurrence rates were also highest in HCC patients with CD151high/MMP9high/MVDhigh in comparison with the other groups. Multivariate Cox proportional hazards analysis showed that the concomitant overexpression of CD151, MMP9, and MVD was an independent marker for predicting poor prognosis of HCC. Conclusion: Overexpression of CD151 up‐regulated the expression of MMP9 through the PI3K/Akt/GSK‐3β/Snail pathway. CD151‐dependent neoangiogenesis appeared to promote the progression of HCC, and this suggests that CD151 may be useful as a high‐priority therapeutic target for antiangiogenesis in HCC. HEPATOLOGY 2010

Systemic Immune-Inflammation Index Predicts Prognosis of Patients after Curative Resection for Hepatocellular Carcinoma

Purpose: We developed a novel systemic immune-inflammation index (SII) based on lymphocyte, neutrophil, and platelet counts and explored its prognostic value in hepatocellular carcinoma (HCC). Experimental Design: The SII was developed based on a retrospective study of 133 patients with HCC undergoing resection between 2005 and 2006, and validated in a prospective study of 123 patients enrolled from 2010 to 2011. The circulating tumor cell (CTC) level in the validation cohort was measured using the CellSearch system. Prediction accuracy was evaluated with area under the receiver operating characteristic curve (AUC). Results: An optimal cutoff point for the SII of 330 × 109 stratified the patients with HCC into high (≥330) and low SII (<330) groups in the training cohort. Univariate and multivariate analyses revealed the SII was an independent predictor for overall survival and relapse-free survival, and prognostic for patients with negative α-fetoprotein and Barcelona Clinic Liver Cancer stage 0+A. The AUCs of the SII for survival and recurrence were higher than other conventional clinical indices. An SII ≥ 330 was significantly associated with vascular invasion, large tumors, and early recurrence. CTC levels were significantly higher in the SII ≥ 330 group (1.71 ± 0.34 vs. 4.37 ± 1.04, P = 0.029). In patients with detectable CTCs, those with SII ≥ 330 had higher recurrence rates and shorter survival time than patients with SII < 330. Conclusion: The SII was a powerful prognostic indicator of poor outcome in patients with HCC and is a promising tool for HCC treatment strategy decisions. The dismal outcome in patients with high SII scores might be related to higher CTC levels. Clin Cancer Res; 20(23); 6212–22. ©2014 AACR.

MiR-146a enhances angiogenic activity of endothelial cells in hepatocellular carcinoma by promoting PDGFRA expression

Endothelial cells (ECs) are critical for angiogenesis, and microRNAs play important roles in this process. We investigated the regulatory role of microRNAs in ECs of hepatocellular carcinoma (HCC) by examining the microRNA expression profile of human umbilical vein endothelial cells (HUVECs) in the absence or presence of human HCC cells, and identified miR-146a as the most highly upregulated microRNA. Furthermore, we revealed that miR-146a promoted the expression of platelet-derived growth factor receptor α (PDGFRA) in HUVECs, and this process was mediated by BRCA1. Overexpression of PDGFRA in the ECs of HCC tissues was associated with microvascular invasion and predicted a poorer prognosis. These results suggest that miR-146a plays a key role in regulating the angiogenic activity of ECs in HCC through miR-146a-BRCA1-PDGFRA pathway. MiR-146a and PDGFRA may emerge as potential anti-angiogenic targets on ECs for HCC therapy.

High expression of Dickkopf-related protein 1 is related to lymphatic metastasis and indicates poor prognosis in intrahepatic cholangiocarcinoma patients after surgery

Dickkopf‐related protein 1 (DKK1) has been reported involved in metastasis and invasion in several tumors. This study sought to investigate the prognostic value of DKK1 in intrahepatic cholangiocarcinoma (ICC) and its role in promoting ICC metastasis.

A case study of the highly time-resolved evolution of aerosol chemical and optical properties in urban Shanghai, China

Characteristics of the chemical and optical properties of aerosols in urban Shanghai and their relationship were studied over a three-day period in October 2011. A suite of real-time instruments, including an Aerosol Time-Of-Flight Mass Spectrometer (ATOFMS), a Monitor for AeRosols and GAses (MARGA), a Cavity Ring Down Spectrometer (CRDS), a nephelometer and a Scanning Mobility Particle Sizer (SMPS), was employed to follow the quick changes of the aerosol properties within the 72 h sampling period. The origin of the air mass arriving in Shanghai during this period shifted from the East China Sea to the northwest area of China, offering a unique opportunity to observe the evolution of aerosols influenced by regional transport from the most polluted areas in China. According to the meteorological conditions and temporal characterizations of the chemical and optical properties, the sampling period was divided into three periods. During Period 1 (00:00–23:00 LT, 13 October), the aerosols in urban Shanghai were mainly fresh and the single scattering albedo varied negatively with the emission of elemental carbon, indicating that local sources dominated. Period 2 (23:00 LT on 13 October to 10:00 LT on 15 October) was impacted by regionally transported pollutants and had the highest particulate matter (PM) mass loading and the lowest particle acidity, characterized by large fractions of aged particles and high secondary ion (nitrate, sulfate and ammonium) mass concentrations. Comparison between ATOFMS particle acidity and quantitative particle acidity by MARGA indicated the significance of semi-quantitative calculation in ATOFMS. Two sub-periods were identified in Period 2 based on the scattering efficiency of PM 1 mass. Period 3 (from 10:00 LT on 15 October to 00:00 LT on 16 October) had a low PM1/PM10 ratio and a new particle formation event. The comparison of these sub-periods highlights the influence of particle mixing state on aerosol optical properties. We directly observed the influence of regionally transported pollutants on local aerosol properties and demonstrate that the PM mass extinction efficiency is largely determined by the mixing states of the aerosol.

Prognostic significance of Capn4 overexpression in intrahepatic cholangiocarcinoma.

Background Calpain small subunit 1 (Capn4) has been shown to correlate with the metastasis/invasion of hepatocellular carcinoma. This study aimed to investigate the role of Capn4 in intrahepatic cholangiocarcinoma (ICC). Methods Capn4 expression was measured in 33 ICC tissues by quantitative real-time polymerase chain reaction and western blot. The role of Capn4 in the migration, invasion and proliferation of ICC cells and matrix metalloproteinase 2 (MMP2) expression were assessed after Capn4 depletion by specific small interfering RNA. Capn4 expression was further examined by immunohistochemistry in a tissue microarray consisting of 140 ICC patients and 13 normal liver tissues, and the prognostic role of Capn4 in ICC was evaluated by Kaplan-Meier and Cox regression analyses. Results Capn4 expression was significantly higher in the ICC tissues compared to the peritumor tissues. Capn4 down-regulation impaired the migration/invasion ability of HCCC-9810 and QBC939 cells in vitro and decreased MMP2 expression. Capn4 overexpression significantly correlated with the presence of lymphatic metastasis of ICC (pu200a=u200a0.026) and the tumor-node-metastasis (TNM) stage (pu200a=u200a0.009). The postoperative 2- and 5-year overall survivals in patients with Capn4low were higher than those in the Capn4high group. The cumulative recurrence rate in patients with Capn4low was much lower than in the Capn4high group. Multivariate analysis showed that Capn4 overexpression was an independent prognostic marker in ICC. Conclusions Capn4 overexpression was implicated in ICC metastasis/invasion, and Capn4 overexpression may be used as a molecular therapeutic target for ICC.

Overexpression of galectin‐1 is associated with poor prognosis in human hepatocellular carcinoma following resection

Background and Aim:u2002 The high expression of the galectin‐1 predicts poor patient outcome in several tumors. The aim of this study was to investigate its prognostic value in patients with hepatocellular carcinoma (HCC) after resection.