Search

This study aims to investigate the effect of ergosterol peroxide(EP) on the apoptosis of human hepatocellular carcinoma and its mechanism of action. The cell viability of HepG2 and SK-Hep-1 cells with 0(blank control), 2.5, 5, 10, 20, 40, and 80 μmol·L~(-1) of EP after 24, 48, and 72 h of action was detected by using CCK-8 assay, and the half inhibitory concentrations(IC_(50)) at 24, 48, and 72 h were calculated. Formal experiments were performed to detect the effect of EP on intracellular reactive oxygen species(ROS) using DCFH-DA staining, the effect of EP on intracellular mitochondrial membrane potential using JC-1 staining, the number of apoptotic cells using Annexin V-FITC/PI double-staining after HepG2 cells were co-cultured with 0(blank control), 10, 20, 40 μmol·L~(-1) EP for 48 h. The effects of EP at different concentrations on apoptotic morphology were detected using AO/EB staining. The effects of different concentrations of EP on the protein expression of mitochondrial apoptosis pathway-related proteins B cell lymphoma 2(Bcl-2), cytochrome C(Cyt-C), Bcl-2-related X protein(Bax), caspase-3, cleaved caspase-3, caspase-9, and cleaved caspase-9 were examined by using Western blot. The results showed that different concentrations of EP could inhibit the proliferation of hepatocellular carcinoma with concentration-and time-dependent trends. Compared with the blank control group, the ROS level in the EP-treated group increased significantly(P<0.05). The mitochondrial membrane potential decreased significantly(P<0.05). The total apoptosis rate increased significantly(P<0.05). The expression of Bcl-2 protein was significantly down-regulated, and the expression of Cyt-C, Bax, cleaved caspase-9, and cleaved caspase-3 were significantly up-regulated(P<0.05). In summary, EP may inhibit the proliferation of hepatocellular carcinoma by modulating the mitochondria-mediated apoptosis pathway and induce apoptosis.

Background & Aims: We aimed to develop a Transformer-based deep learning (DL) network for prognostic stratification in hepatocellular carcinoma (HCC) patients undergoing RFA., Methods: A Swin Transformer DL network was trained to establish associations between magnetic resonance imaging (MRI) datasets and the ground truth of microvascular invasion (MVI) based on 696 surgical resection (SR) patients with solitary HCC ≤3 cm, and was validated in an external cohort (n = 180). The multiphase MRI-based DL risk outputs using an optimal threshold of .5 was employed as a MVI classifier for prognosis stratification in the RFA cohort (n = 180)., Results: Over 90% of all enrolled patients exhibited hepatitis B virus infection. Liver cirrhosis was significantly more prevalent in the RFA cohort compared to the SR cohort (72.2% vs. 44.1%, p < .001). The MVI risk outputs exhibited good performance (area under the curve values = .938 and .883) for predicting MVI in the training and validation cohort, respectively. The RFA patients at high risk of MVI classified by the MVI classifier demonstrated significantly lower recurrence-free survival (RFS) and overall survival rates at 1, 3 and 5 years compared to those classified as low risk (p < .001). Multivariate cox regression modelling of a-fetoprotein > 20 ng/mL [hazard ratio (HR) = 1.53; 95% confidence interval (95% CI): 1.02-2.33, p = .047], high risk of MVI (HR = 3.76; 95% CI: 2.40-5.88, p < .001) and unfavourable tumour location (HR = 2.15; 95% CI: 1.40-3.29, p = .001) yielded a c-index of .731 (bootstrapped 95% CI: .667-.778) for evaluating RFS after RFA. Among the three risk factors, MVI was the most powerful predictor for intrahepatic distance recurrence., Conclusions: The proposed MVI classifier can serve as a valuable imaging biomarker for prognostic stratification in early-stage HCC patients undergoing RFA., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Emerging evidence suggests that Rho GTPases play a crucial role in tumorigenesis and metastasis, but their involvement in the tumor microenvironment (TME) and prognosis of hepatocellular carcinoma (HCC) is not well understood., Methods: We aim to develop a tumor prognosis prediction system called the Rho GTPases-related gene score (RGPRG score) using Rho GTPase signaling genes and further bioinformatic analyses., Results: Our work found that HCC patients with a high RGPRG score had significantly worse survival and increased immunosuppressive cell fractions compared to those with a low RGPRG score. Single-cell cohort analysis revealed an immune-active TME in patients with a low RGPRG score, with strengthened communication from T/NK cells to other cells through MIF signaling networks. Targeting these alterations in TME, the patients with high RGPRG score have worse immunotherapeutic outcomes and decreased survival time in the immunotherapy cohort. Moreover, the RGPRG score was found to be correlated with survival in 27 other cancers. In vitro experiments confirmed that knockdown of the key Rho GTPase-signaling biomarker SFN significantly inhibited HCC cell proliferation, invasion, and migration., Conclusions: This study provides new insight into the TME features and clinical use of Rho GTPase gene pattern at the bulk-seq and single-cell level, which may contribute to guiding personalized treatment and improving clinical outcome in HCC., (© 2024. The Author(s).)

Introduction: Heat ablation is one of the key modalities in treating liver cancer, yet the residual cancer tissues suffering sublethal heat treatment possess a potential for increased malignancy. This study conducts a comprehensive analysis of cellular dynamics, metabolic shifts, and macrophage polarization within the tumor microenvironment following sublethal heat treatment., Methods: We observed significant acidification in tumor cell supernatants, attributed to increased lactic acid production. The study focused on how this pH shift, crucial in tumor progression and resistance, influences macrophage polarization, especially towards the M2 phenotype known for tumor-promoting functions. We also examined the upregulation of MCT1 expression post sublethal heat treatment and its primary role in lactic acid transport., Results: Notably, the study found minimal disparity in MCT1 expression between hepatocellular carcinoma patients and healthy liver tissues, highlighting the complexity of cancer biology. The research further revealed an intricate relationship between lactic acid, MCT1, and the inhibition of macrophage pyroptosis, offering significant insights for therapeutic strategies targeting the tumor immune environment. Post sublethal heat treatment, a reduction in paraspeckle under lactic acid exposure was observed, indicating diverse cellular impacts. Additionally, PKM2 was identified as a key molecule in this context, with decreased levels after sublethal heat treatment in the presence of lactic acid., Discussion: Collectively, these findings illuminate the intertwined mechanisms of sublethal heat treatments, metabolic alterations, and immune modulation in the tumor milieu, providing a deeper understanding of the complex interplay in cancer biology and treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Fan, Yang, Luo, Qu, Ye, Wang, Yan, Shu, Zhang and Liu.)

According to WHO 2019, Hepatocellular carcinoma (HCC) is the fourth highest cause of cancer death worldwide. More precise diagnostic models are needed to enhance early HCC and cirrhosis quick diagnosis, treatment, and survival. Breath biomarkers known as volatile organic compounds (VOCs) in exhaled air can be used to make rapid, precise, and painless diagnoses. Gas chromatography and mass spectrometry (GCMS) are utilized to diagnose HCC and cirrhosis VOCs. In this investigation, metabolically generated VOCs in breath samples (n = 35) of HCC, (n = 35) cirrhotic, and (n = 30) controls were detected via GCMS and SPME. Moreover, this study also aims to identify diagnostic VOCs for distinction among HCC and cirrhosis liver conditions, which are most closely related, and cause misleading during diagnosis. However, using gas chromatography-mass spectrometry (GC-MS) to quantify volatile organic compounds (VOCs) is time-consuming and error-prone since it requires an expert. To verify GC-MS data analysis, we present an in-house R-based array of machine learning models that applies deep learning pattern recognition to automatically discover VOCs from raw data, without human intervention. All-machine learning diagnostic model offers 80% sensitivity, 90% specificity, and 95% accuracy, with an AUC of 0.9586. Our results demonstrated the validity and utility of GCMS-SMPE in combination with innovative ML models for early detection of HCC and cirrhosis-specific VOCs considered as potential diagnostic breath biomarkers and showed differentiation among HCC and cirrhosis. With these useful insights, we can build handheld e-nose sensors to detect HCC and cirrhosis through breath analysis and this unique approach can help in diagnosis by reducing integration time and costs without compromising accuracy or consistency., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Ain Nazir et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Hepatocellular carcinoma (HCC) is a leading cause of mortality, with Hepatitis B virus (HBV) infection as a dominant etiology worldwide. Effective early detection strategies may facilitate curative therapies and improve survival. We investigated genomic aberrations in circulating tumor DNA (ctDNA) as potential diagnostic markers of HCC in HBV-infected patients., Methods: We identified early stage (BCLC 0-A) HCC cases (n = 21) and patients without HCC (n = 14) from a cohort of Asian patients with HBV, undergoing surveillance between 2013 and 2017. Circulating cell-free DNA was isolated from blood samples, and assayed by next-generation sequencing of 23 genes implicated in HCC pathogenesis. Somatic mutations were identified using a computational pipeline. Using area under the curve (AUC) in receiver operating characteristic (ROC) analysis, we evaluated gene alterations and clinical factors in an exploratory early HCC detection model., Results: Mutant ARID1A, CTNNB1, TP53 genes were increased in HCC cases vs. non-HCC patients (85.7% vs 42.9%, P = 0.011; 42.9% vs 0%, P = 0.005; 100% vs 71.4%, P = 0.019, respectively). Using these three genes, AUC for discriminating HCC from non-HCC patients was 0.844 (95% confidence interval [CI]: 0.7317-0.9553). When combining these genes with clinical factors in an exploratory early HCC detection model, AUC increased from 0.7415 (using clinical factors alone) to 0.9354 (P = 0.041)., Conclusion: Genomic aberrations in ctDNA were more prevalent in HBV-infected HCC patients compared with patients without HCC. Combining these alterations with clinical factors may identify HCC in HBV-infected patients at an early stage. These findings warrant validation in future studies., Competing Interests: Declaration of Competing Interest The authors have no conflict of interest to disclose., (Copyright © 2023. Published by Elsevier Masson SAS.)

Objective: To construct a survival prediction model for patients with TNM stage III hepatocellular carcinoma (HCC) to guide the clinical diagnosis and treatment of HCC patients and improve prognosis., Methods: Based on data from patients with stage III (AJCC 7th TNM stage) recorded by the American Institute of Cancer Research from 2010 to 2013, risk factors affecting the prognosis were screened by Cox univariate and multivariate regression, line plots was constructed, and the credibility of the model was verified by Boostrap method. ROC operating curves, calibration curves and DCA clinical decision curves were used to evaluate the model, and Kaplan-Meier was used for survival analysis was used to evaluate the efficacy of the model. External survival data from patients newly diagnosed with stage III hepatocellular carcinoma during 2014-2015 were used to validate and fit the model and to optimize the model., Results: Age > 75 years vs.18-53 years [HR = 1.502; 95%CI(1.134-1.990)], stage IIIC vs. Stage IIIA [HR = 1.930; 95%CI(1.509-2.470)], lobotomy vs. non-surgery [HR = 0.295; 95%CI(0.228-0.383)], radiotherapy vs. non-radiotherapy [HR = 0.481; 95%CI(0.373-0.619)], chemotherapy vs. Non-chemotherapy [HR = 0.443; 95%CI(0.381-0.515)], positive serum AFP before treatment vs. negative [HR = 1.667; 95%CI(1.356-2.049)], the above indicators are independent prognostic factors for patients with stage III hepatocellular carcinoma, and the P values for the above results were less than 0.05. A joint prediction model was constructed based on age, TNM stage, whether and how to operate, whether to receive radiotherapy, whether to receive chemotherapy, pre-treatment serum AFP status and liver fibrosis score. The consistency index of the improved prognosis model was 0.725., Conclusions: The traditional TNM staging has limitations for clinical diagnosis and treatment, while the Nomogram model modified by TNM staging has good predictive efficacy and clinical significance., (© 2023. The Author(s).)

Purpose: To report the efficacy and safety of postoperative adjuvant hepatic arterial infusion chemotherapy (HAIC) with 5-fluorouracil and oxaliplatin (FOLFOX) in hepatocellular carcinoma (HCC) patients with microvascular invasion (MVI)., Patients and Methods: In this randomized, open-label, multicenter trial, histologically confirmed HCC patients with MVI were randomly assigned (1:1) to receive adjuvant FOLFOX-HAIC (treatment group) or routine follow-up (control group). The primary end point was disease-free survival (DFS) by intention-to-treat (ITT) analysis while secondary end points were overall survival, recurrence rate, and safety., Results: Between June 2016 and August 2021, a total of 315 patients (ITT population) at five centers were randomly assigned to the treatment group (n = 157) or the control group (n = 158). In the ITT population, the median DFS was 20.3 months (95% CI, 10.4 to 30.3) in the treatment group versus 10.0 months (95% CI, 6.8 to 13.2) in the control group (hazard ratio, 0.59; 95% CI, 0.43 to 0.81; P = .001). The overall survival rates at 1 year, 2 years, and 3 years were 93.8% (95% CI, 89.8 to 98.1), 86.4% (95% CI, 80.0 to 93.2), and 80.4% (95% CI, 71.9 to 89.9) for the treatment group and 92.0% (95% CI, 87.6 to 96.7), 86.0% (95% CI, 79.9 to 92.6), and 74.9% (95% CI, 65.5 to 85.7) for the control group (hazard ratio, 0.64; 95% CI, 0.36 to 1.14; P = .130), respectively. The recurrence rates were 40.1% (63/157) in the treatment group and 55.7% (88/158) in the control group. Majority of the adverse events were grade 0-1 (83.8%), with no treatment-related death in both groups., Conclusion: Postoperative adjuvant HAIC with FOLFOX significantly improved the DFS benefits with acceptable toxicities in HCC patients with MVI.

Ethnopharmacological Relevance: Jianpi Huayu decoction (JHD) is a traditional Chinese medicinal preparation used to treat a variety of malignant tumors including HCC, although the underlying mechanism remains unknown. Exosomes in the tumor microenvironment mediate intercellular signaling among cancer cells, but precise contributions to hepatocellular carcinoma (HCC) progression are still elusive., Aim of the Study: In this work, the main objective was to examine the mechanisms underlying anti-tumor effects of JHD and the potential contributions of exosomal signaling., Materials and Methods: LC-MS/MS was used for quality control of JDH preparation, while nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM) and western blotting were used for verification of exosomes. In vitro assays included CCK8, wound healing assay, transwell invasion assay, qRT-PCR and western blotting were performed to investigate the effects of JHD on HCC cells and the molecular mechanism. Furthermore, the effects of JHD on subcutaneous tumor model of nude mice were also determined., Results: JHD inhibited the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of cultured HCC cells. Further, exosomes isolated from EMT-induced HCC cells promoted the migration, invasion and EMT of other cultured HCC cells, while exosomes isolated from EMT-induced HCC cells after JHD treatment had little effect. In addition, JHD reduced the expression of exosomal miR-23a-3p in cultured HCC cells. miR-23a-3p was significantly up-regulated in tumor compared with that in adjacent non-cancerous tissues of patients with HCC. HCC patients with high miR-23a-3p expression had poor overall survival after hepatectomy. Meanwhile, miR-23a-3p enhanced HCC cell proliferation, EMT, and expression of Smad signaling proteins. More importantly, overexpression of miR-23a-3p can reverse the inhibition of EMT and Smad signaling pathway caused by JHD treatment. In vivo assays, treatment with JHD also reduced the growth of HCC-derived tumors in nude mice, reduced the expression of miR-23a-3p in serum exosomes and the level of EMT in tumor cells., Conclusions: the antitumor effects of JHD on HCC are mediated at least in part by inhibition of EMT due to downregulation of exosome-mediated intercellular miR-23a-3p transfer and subsequent blockade of Smad signaling. Disrupting this exosomal miR-23a-3p/Smad signaling pathway may be an effective treatment., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Objective: This study was aimed at exploring the prognostic and clinicopathological roles of aspartate aminotransferase-to-lymphocyte ratio index (ALRI) in patients with hepatocellular carcinoma via a meta-analysis., Methods: The PubMed, Embase, Cochrane Library, Web of Science, China National Knowledge Infrastructure (CNKI), Wanfang, and VIP databases were comprehensively searched from inception to November 20, 2021. Pooled hazard ratio (HR) and corresponding 95% confidence interval (CI) were used to evaluate the relationship between ALRI and overall survival (OS) as well as progression-free survival (PFS) in patients with hepatocellular carcinoma. Odds ratio (OR) and the corresponding 95% CI were also used to investigate correlations between clinical factors and ALRI in patients with hepatocellular carcinoma., Results: A total of 3914 patients with hepatocellular carcinoma from eleven retrospective cohorts were included in this meta-analysis. The combined results revealed that patients with hepatocellular carcinoma with elevated ALRI tended to have unfavorable OS (HR 1.53 [95% CI 1.25-1.82]; P < 0.001). Pooled HRs revealed that high ALRI was an independent risk factor for inferior PFS in patients with hepatocellular carcinoma (HR 1.36 [95% CI 1.10-1.63]; P < 0.001). In addition, high ALRI was strongly associated with male sex (OR 1.32 [95% CI 1.02-1.70]; P = 0.035), presence of cirrhosis (OR 1.68 [95% CI 1.01-2.81]; P = 0.046), larger tumor size (OR 2.25 [95% CI 1.31-3.88]; P < 0.001), presence of portal vein tumor thrombus (OR 2.50 [95% CI 1.52-4.11]; P < 0.001), and distant metastasis (OR 1.72 [95% CI 1.05-2.82]; P = 0.031)., Conclusion: Elevated ALRI in patients with hepatocellular carcinoma predicted inferior survival outcomes and was strongly associated with some important features of hepatocellular carcinoma., Competing Interests: The authors declared that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Xiulan Peng et al.)

Liver cancer is one of the malignant tumors with the highest fatality rate in China and the 5-years survival rate is only 12.5%. Early detection to undertake early treatment can improve the survival rate of patients with liver cancer. Nowadays, the unsatisfactory performance of serum Alpha-fetoprotein (AFP) test, and the problems of insensitive to small lesions for ultrasound and exposure to nuclear radiation for CT, necessitate the urgency to explore novel diagnostic biomarkers of liver cancer. It has been demonstrated the presence of autoantibodies targeting tumor-associated antigens prior to clinic symptoms implied underlying early diagnostic value of malignancies. High specificity but low sensitivity of single autoantibodies such as the most reported anti-p53, anti-insulin like growth factor-Ⅱ mRNA binding protein, and anti-glucose regulated protein can be solved by combining different autoantibodies. However, the autoantibodies of different combinations vary in studies. Simultaneously, autoantibodies in combination with AFP facilitate further improving the detection rate of liver cancer. Nevertheless, the autoantibodies related to prognosis of liver cancer needs to be more studied in the near future.

Emerging evidence suggests that microRNAs (miRNAs) participate in hepatocellular carcinoma (HCC) progression. Nevertheless, the mechanism of miR-7-5p in HCC cells has not been researched. In the research, the underlying biological function of miR-7-5p and SPC24 in HCC was explored. qRT-PCR was performed to measure the miR-7-5p and SPC24 level in HCC tissues and cells. The effect of miR-7-5p on HCC progression was detected by performing CCK-8, BrdU, and transwell assay. The relationship between miR-7-5p and SPC24 was determined using luciferase and RNA pull-down assays. Our findings showed that miR-7-5p was downregulated in HCC whereas SPC24 was upregulated in HCC. It was also showed that miR-7-5p upregulation restricted malignant behaviors of HCC cells, but this inhibitory effect of miR-7-5p could be relieved by its target gene SPC24. In conclusion, this research suggested that by inhibiting SPC24, miR-7-5p could act as a tumor inhibitory factor in HCC., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Hepatocellular carcinoma (HCC) is a common cancer with poor prognosis. Due to the lack of effective biomarkers and its complex immune microenvironment, the effects of current HCC therapies are not ideal. In this study, we used the GSE57957 microarray data from Gene Expression Omnibus database to construct a co-expression network. The weighted gene co-expression network analysis and CIBERSORT algorithm, which quantifies cellular composition of immune cells, were used to identify modules related to immune cells. Four hub genes (EFTUD2, GAPDH, NOP56, PA2G4) were identified by co-expression network and protein-protein interactions network analysis. We examined these genes in TCGA database, and found that the four hub genes were highly expressed in tumor tissues in multiple HCC groups, and the expression levels were significantly correlated with patient survival time, pathological stage and tumor progression. On the other hand, methylation analysis showed that the up-regulation of EFTUD2, GAPDH, NOP56 might be due to the hypomethylation status of their promoters. Next, we investigated the correlations between the expression levels of four hub genes and tumor immune infiltration using Tumor Immune Estimation Resource (TIMER). Gene set variation analysis suggested that the four hub genes were associated with numerous pathways that affect tumor progression or immune microenvironment. Overall, our results showed that the four hub genes were closely related to tumor prognosis, and may serve as targets for treatment and diagnosis of HCC. In addition, the associations between these genes and immune infiltration enhanced our understanding of tumor immune environment and provided new directions for the development of drugs and the monitoring of tumor immune status., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zhou, Chen, Luo, Li, Jiang and Ou.)

Elongation factor Tu GTP binding domain containing 2 (EFTUD2), a spliceosomal GTPase, plays a pivotal role in multiple organ development and innate immune. It has been reported that EFTUD2 is a new host factor with activity against HCV infection. However, the role of EFTUD2 in solid tumors, including hepatocellular carcinoma (HCC), remains unexplored. In this study, we investigated the molecular function of EFTUD2 in HCC. Data from The Cancer Genome Atlas (TCGA) indicated an upregulation of EFTUD2 in HCC tissues compared to that in nontumor liver tissues. Immunohistochemical analysis performed on two independent HCC cohorts confirmed the upregulation of EFTUD2 in HCC tissues and further suggested that a high level of EFTUD2 expression predicted shorter overall and recurrence-free survival in HCC patients. Functional studies suggested that siRNA interference with EFTUD2 expression significantly suppressed cell viability, blocked cell cycle progression, facilitated tumor cell apoptosis, and inhibited metastasis, while the enhancement of EFTUD2 expression promoted the proliferation and migration of HCC cells both in vitro and in vivo. Surprisingly, we also found that the stable knockdown of EFTUD2 expression via lentivirus infection was lethal for HCC cells. This finding suggested that EFTUD2 was essential for maintaining the survival of HCC cells. Mechanistically, RNA sequencing and gene set enrichment analysis (GSEA) suggested that the gene sets of epithelial-mesenchymal transition (EMT) and the JAK/STAT3 pathway were enriched in EFTUD2-overexpressing cells. Further verification indicated that EFTUD2-overexpressing cells exhibited an EMT-like phenotype and had enhanced STAT3 activation, while the STAT3 inhibitor S3I-201 partially blocked these pro-malignant effects of EFTUD2 overexpression. In summary, we report EFTUD2 as a novel oncogene that helps to maintain the survival of HCC cells and promotes HCC progression through the activation of STAT3. The high level of expression of EFTUD2 in HCC tissues indicates shorter overall and recurrence-free survival in HCC patients.

Small recurrent hepatocellular carcinoma (HCC) can show atypical imaging patterns, and a specific diagnostic algorithm for HCC is lacking. This study aimed to better characterize postoperative recurrent HCCs <20 mm in size with gadoxetic acid-enhanced magnetic resonance imaging (MRI). We evaluated 373 newly developed nodules after hepatectomy in 204 HCC patients with chronic hepatitis B virus infection. The diagnostic performance of Liver Imaging Reporting and Data System (LI-RADS) version 2018 was calculated with gadoxetic acid-enhanced MRI to characterize recurrent HCC. Modified diagnostic algorithms were proposed by combining significant imaging biomarkers related to subcentimeter and 10-19 mm recurrence, and the algorithms were then compared with the LI-RADS system. A total of 256 recurrent HCCs (108 recurrent HCCs <10 mm in size; 148 recurrent HCCs 10-19 mm in size) were confirmed via histology or follow-up imaging. Nonrim arterial phase hyperenhancement (APHE) and 3 LI-RADS ancillary features (AFs; hepatobiliary phase hypointensity, mild-moderate T 2 hyperintensity, and restricted diffusion) were significantly related to recurrent HCCs <20 mm in size according to a multivariate analysis. For subcentimeter recurrence, combining at least 2 of the 3 AFs only achieved better specificity (sensitivity, 83.3%; specificity, 87.7%) than the LR-4 category (sensitivity, 88.9%, P = 0.21; specificity, 70.8%, P = 0.006). For 10-19 mm recurrences, combining nonrim APHE and at least 1 of the 3 AFs achieved only a significantly enhanced sensitivity of 85.1% but a lower specificity of 86.5% compared with the LR-5 category (sensitivity: 63.5%, P < 0.001; specificity: 94.2%, P = 0.13). In conclusion, the diagnostic algorithms for subcentimeter and 10-19 mm recurrent HCCs should be stratified. Combining at least 2 AFs demonstrated comparable sensitivity with significantly enhanced specificity compared with the LR-4 category for characterizing subcentimeter recurrence., (Copyright © 2019 by the American Association for the Study of Liver Diseases.)

Objectives: We aimed to develop a radiomics-based model derived from gadoxetic acid-enhanced MR images to preoperatively identify cytokeratin (CK) 19 status of hepatocellular carcinoma (HCC)., Methods: A cohort of 227 patients with single HCC was classified into a training set (n = 159) and a time-independent validated set (n = 68). A total of 647 radiomic features were extracted from multi-sequence MR images. The least absolute shrinkage and selection operator regression and decision tree methods were utilized for feature selection and radiomics signature construction. A multivariable logistic regression model incorporating clinico-radiological features and the fusion radiomics signature was built for prediction of CK19 status by evaluating area under curve (AUC)., Results: In the whole cohort, 57 patients were CK19 positive and 170 patients were CK19 negative. By combining 11 and 6 radiomic features extracted in arterial phase and hepatobiliary phase images, respectively, a fusion radiomics signature achieved AUCs of 0.951 and 0.822 in training and validation datasets. The final combined model integrated a-fetoprotein levels, arterial rim enhancement pattern, irregular tumor margin, and the fusion radiomics signature, with a sensitivity of 0.818 and specificity of 0.974 in the training cohort and that of 0.769 and 0.818 in the validated cohort. The nomogram based on the combined model showed satisfactory prediction performance in training (C-index 0.959) and validation (C-index 0.846) dataset., Conclusions: The combined model based on a fusion radiomics signature derived from arterial and hepatobiliary phase images of gadoxetic acid-enhanced MRI can be a reliable biomarker for CK19 status of HCC., Key Points: • Arterial rim enhancement pattern and irregular tumor margin on hepatobiliary phase on gadoxetic acid-enhanced MRI can be useful for evaluating CK19 status of HCC. • A radiomics-based model performed better than the clinico-radiological model both in training and validation datasets for predicting CK19 status of HCC. • The nomogram based on the fusion radiomics signature can be easily used for CK19 stratification of HCC.

Prevention and treatment options for hepatocellular carcinoma (HCC) are presently limited, underscoring the necessity for further elucidating molecular mechanisms underlying HCC development and identifying new prevention and therapeutic targets. Here, we demonstrate a unique protumorigenic niche in the livers of Ncoa5 +/ - mouse model of HCC, which is characterized by altered expression of a subset of genes including p21 WAF1/CIP1 and proinflammatory cytokine genes, increased putative hepatic progenitors, and expansions of activated and tissue-resident memory (TRM) CD8+ T lymphocytes, myeloid-derived suppressor cells (MDSCs), and alternatively activated M2 macrophages. Importantly, prophylactic metformin treatment reversed these characteristics including aberrant p21 WAF1/CIP1 expression and subsequently reduced HCC incidence in Ncoa5 +/- male mice. Heterozygous deletion of the p21 WAF1/CIP1 gene alleviated the key features associated with the protumorigenic niche in the livers of Ncoa5 +/- male mice. Moreover, transcriptomic analysis reveals that preneoplastic livers of Ncoa5 +/- mice are similar to the livers of nonalcoholic steatohepatitis patients as well as the adjacent noncancerous liver tissues of a subset of HCC patients with a relatively poor prognosis. Together, our results suggest that p21 WAF1/CIP1 overexpression is essential in the development of protumorigenic microenvironment induced by NCOA5 deficiency and metformin prevents HCC development via alleviating p21 WAF1/CIP1 overexpression and protumorigenic microenvironment.

Accumulating data suggest that metadherin (MTDH) may function as an oncogene. Our previous study showed that MTDH promotes hepatocellular carcinoma (HCC) metastasis via the epithelial-mesenchymal transition. In this study, we aim to further elucidate how MTDH promotes HCC metastasis. Using Co-immunoprecipitation (co-IP) and mass spectrometry, we found that MTDH can specifically bind to protein arginine methyltransferase 5 (PRMT5). Further functional assays revealed that PRMT5 overexpression promoted the proliferation and motility of HCC cells and that knockout of PRMT5 impeded the effect of MTDH. The immunohistochemistry assay/tissue microarray results showed that when MTDH was overexpressed in HCC cells, PRMT5 translocated from the nucleus to the cytoplasm, with the subsequent translocation of β-catenin from the cytoplasm to the nucleus and upregulation of the WNT-β-catenin signaling pathway. Further in vivo experiments suggested that PRMT5 and β-catenin played a pivotal role in MTDH-mediated HCC metastasis. We therefore concluded that the MTDH-PRMT5 complex promotes HCC metastasis by regulating the WNT-β-catenin signaling pathway., (© The Author(s) 2019. Published by Oxford University Press.)

FAM83A is part of an 8-member protein family of unknown function and is reported to be a cancer-promoting and treatment-resistance factor in several cancers. However, its role in hepatocellular carcinoma (HCC) remains unclear. Analysis of the Cancer Genome Atlas (TCGA) showed that FAM83A mRNA expression is upregulated in HCC, as are the protein expression levels in both HCC cell lines and tissues. Clinical data have demonstrated that high FAM83A expression is positively correlated with poor progression-free survival time, thus suggesting its cancer-promoting potential. Functional analyses showed that FAM83A overexpression promoted HCC cell migration and invasion in vitro and suppressed sorafenib sensitivity. Inhibiting FAM83A reversed these results. A pulmonary metastasis model further confirmed that FAM83A promoted HCC cell metastasis in vivo. Mechanistic analyses indicated that FAM83A activated the PI3K/AKT signaling pathway, its downstream c-JUN protein, and epithelial-to-mesenchymal transition (EMT)-related protein levels, including downregulation of E-cadherin and upregulation of Vimentin and N-cadherin. Interestingly, c-JUN induced FAM83A expression by directly binding to its promoter region and thus forming a positive-feedback loop for FAM83A/PI3K/AKT/c-JUN. In conclusion, we demonstrated that FAM83A, as a cancer-metastasis promoter, accelerates migration, invasion and metastasis by activating the PI3K/AKT/c-JUN pathway and inducing its self-expression via feedback, thus forming a FAM83A/PI3K/AKT/c-JUN positive-feedback loop to activate EMT signaling and finally promote HCC migration, invasion and metastasis., Competing Interests: Declaration of competing interest The authors declare no potential conflicts of interest., (Copyright © 2019 The Author(s). Published by Elsevier Masson SAS.. All rights reserved.)

PURPOSE This study aims to evaluate the safety and effectiveness of the percutaneous cryoablation for subcapsular hepatocellular carcinoma (HCC). METHODS A total of 57 patients with subcapsular (<1 cm form the liver edge) HCCs (68 lesions), who were treated with CT-guided percutaneous cryoablation in the Department of Interventional Radiology of our hospital between July 1, 2016 and September 1, 2018, were retrospectively included. Complete ablation rate, local tumor progression (LTP) and treatment-related complications were evaluated. Furthermore, the degree of intraoperative and postoperative pain was measured with the visual analog scale (VAS), and laboratory findings were compared before and after the procedure. RESULTS All patients successfully completed the treatment. The mean follow-up period was 12.8 months (range, 3-27 months), and the complete ablation rate was 97% (66/68). Local tumor progression occurred in 11 lesions (16.2%), and the 6-, 12- and 18-month cumulative LTP rates were 4.0%, 8.2% and 20.5%, respectively. Two patients (3.5%, 2/57) developed major complications, and 12 patients had minor complications (22.8%, 12/57). The mean VAS score during the operation was 1.65 points (range, 1-3 points). Postoperative pain worsened in 3 patients, and the VAS scores reached 4-5. Transient changes in biochemical and hematologic markers were observed. CONCLUSION Percutaneous cryoablation for subcapsular HCC is safe and effective, the procedure is simple and the patients suffer less pain.

Cancer stem cells (CSCs) are the key factor in determining cancer recurrence, metastasis, chemoresistance and patient prognosis in hepatocellular carcinoma (HCC). The role of miR-5188 in cancer stemness has never been documented. In this study, we investigated the clinical and biological roles of miR-5188 in HCC. Methods: MiRNA expression in HCC was analyzed by bioinformatics analysis and in situ hybridization. The biological effect of miR-5188 was demonstrated in both in vitro and in vivo studies through the ectopic expression of miR-5188. The target gene and molecular pathway of miR-5188 were characterized using bioinformatics tools, dual-luciferase reporter assays, gene knockdown, and rescue experiments. Results: MiR-5188 was shown to be upregulated and confer poor prognosis in HCC patient data from TCGA database. MiR-5188 was subsequently identified as a significant inducer of cancer stemness that promotes HCC pathogenesis. Specifically, the targeting of miR-5188 by its antagomir markedly prolonged the survival time of HCC-bearing mice and improved HCC cell chemosensitivity in vivo . Mechanistic analysis indicated that miR-5188 directly targets FOXO1, which interacts with β-catenin in the cytoplasm to reduce the nuclear translocation of β-catenin and promotes the activation of Wnt signaling and downstream tumor stemness, EMT, and c-Jun. Moreover, c-Jun transcriptionally activates miR-5188 expression, forming a positive feedback loop. Interestingly, the miR-5188-FOXO1/β-catenin-c-Jun feedback loop was induced by hepatitis X protein (HBX) through Wnt signaling and participated in the HBX-induced pathogenesis of HCC. Finally, analyses of transcriptomics data and our clinical data supported the significance of the abnormal expression of the miR-5188 pathway in HCC pathogenesis. Conclusions: These findings present the inhibition of miR-5188 as a novel strategy for the efficient elimination of CSCs to prevent tumor metastasis, recurrence and chemoresistance in patients with hepatocellular carcinoma. Our study highlights the importance of miR-5188 as a tumor stemness inducer that acts as a potential target for HCC treatment., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Purpose: The overexpression and knockdown of PLK4 were both reported to generate aneuploidy. Thus, we aimed to investigate whether genetic variants in PLK4 contribute to the development of hepatocellular carcinoma (HCC)., Methods: We evaluated associations of common variants in PLK4 and its promoter for the risk of HCC in our association study (1300 cases and 1344 controls). The genotype-tissue expression (GTEx) and The cancer genome atlas (TCGA) databases were used to quantify the expression of PLK4. Cell proliferation and migration affected by PLK4 in HCC were assessed in vitro. Drug susceptibility testing (DST) model was used to assess the sensibility of PLK4-activated HCC to CFI-400945, a small molecule inhibitor of PLK4., Results: Herein, we found a significant association between rs3811741, located in the PLK4 intron, and liver cancer risk (OR = 1.26, P = 9.81 × 10 -5 ). Although PLK4 expressed at lower levels in somatic tissues compared to the testis, the risk allele A of rs3811741 was associated with increased PLK4 expression in liver cancer tissues. Additionally, PLK4 high expression was remarkably associated with shortened survival of HCC (HR = 1.97, P = .001). Furthermore, overexpression of PLK4 promoted, while knockdown of PLK4 suppressed cancer cell proliferation, migration, and invasion. DST model demonstrated that CFI-400945 can effectively suppress rampant proliferation of HCC with highly expressed PLK4., Conclusion: Taken together, our study demonstrated that PLK4 is a susceptibility gene and plays an oncogenic role in HCC. Furthermore, we identified that PLK4 sensitives HCC to CFI-400945, which may be an ideal therapy target for HCC., (© 2019 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Propofol is one of the most extensively used intravenous anaesthetic agents, which has been found to improve the surgical intervention outcome of several types of cancer, including hepatocellular carcinoma (HCC). Additionally, in vitro and in vivo experiments have also indicated that propofol affects the biological behaviour of HCC. However, the underlying mechanisms of the surgical resection of HCC with propofol have not been fully understood. In the present study, we aimed to investigate the underlying mechanism of propofol inhibition of the growth and invasion of HCC cells. Our results showed that treatment with propofol suppressed the proliferation, invasion and migration of HCC in vitro. The subcutaneous xenograft tumour and orthotopic xenograft tumour experiments in nude mice showed that propofol significantly decreased tumour volumes, growth rates and the liver orthotopic xenograft tumour in vivo. Furthermore, the underlying mechanism investigations of the suppressive effects of propofol on HCC cells revealed that propofol treatment upregulated the expression levels of the candidate tumour suppressor miR-219-5p. Silencing of propofol-induced miR-219-5p using anti-miR-219-5p abrogated the inhibitory effects on the proliferation, migration and invasion of HCC cells exerted by propofol treatment. Additionally, we demonstrated that propofol reversed the epithelial-mesenchymal transition of Huh7 and SMMC7721 cells via miR-219-5p induction. The molecular mechanism behind these findings is that propofol-induced miR-219-5p inhibits HCC cell progression by targeting glypican-3 and subsequently results in the inhibition of Wnt/β-catenin signalling. Taken together, our study provides new insights into the advantages of the surgical intervention of HCC with propofol anaesthetization., (© 2019 Wiley Periodicals, Inc.)

Accumulating evidence suggests that long-noncoding RNA (lncRNA) plays important roles in hepatitis B virus (HBV) infections. However, the mechanism underlying how lncRNA regulate hepatocellular carcinoma process remains largely unknown. In this study we found that the expression of LINC01152 was significantly increased in HBV positive HCC tissues and cells and was induced by HBx in vitro. The overexpression of LINC01152 could increases HCC cell proliferation and promotes tumor formation in nude mice. Mechanistically, HBx could increase the transcription of LINC01152. Elevated LINC01152 binds to the promoter region of IL-23, promoting its transcriptional activity and upregulating the levels of Stat3 and p-Stat3. Our findings suggest that LINC01152 plays an important role in HBV-related hepatocellular carcinoma development and may serve as a therapeutic marker for hepatocellular carcinoma., (Copyright © 2019 Shanghai Jiaotong University School of Medicine. Published by Elsevier Masson SAS.. All rights reserved.)

Aim: To determine the enhanced computed tomography (CT) and magnetic resonance imaging (MRI) characteristics of combined hepatocellular carcinoma and cholangiocarcinoma (cHCC-CC) in Chinese patients., Materials and Methods: Patients with histopathologically proven cHCC-CC (n=54) were compared with hepatocellular carcinoma (HCC; n=41) and cholangiocellular carcinoma (CCC; n=41) patients. Clinical information was measured in all patients. Tumour size, tumour margins, signs of cirrhosis, pseudocapsule, capsular retraction, rim enhancement, intrahepatic biliary dilatation, portal vein thrombosis, upper abdominal lymphadenopathy, were assessed on CT and/or MRI. The dynamic pattern of enhancement was also assessed., Results: The majority (81.5%) of cHCC-CC patients had positive hepatitis B serology. The presence of cirrhosis and tumour blood vessels was comparable in cHCC-CC and HCC, but significantly lower in CCC (p>0.05). The presence of ill-defined margin and regional lymphadenopathy was comparable in cHCC-CC and CCC, but significantly lower in HCC (p>0.05). The pseudocapsule, capsular retraction, biliary dilatation, rim enhancement, and abnormal perfusion were significantly different between the three types of lesions, with cHCC-CC being intermediate between HCC and CCC. Nearly half of the cHCC-CC tumours (25; 46.3%) showed the wash-in and wash-out enhancement pattern; the gradual, persistent, and mixed patterns were seen in 12 (22.2%), 5 (9.3%), and 12 (22.2%) tumours, respectively., Conclusion: The majority of cHCC-CC tumours occur against a background of positive hepatitis B serology and cirrhosis. Imaging findings vary widely between cHCC-CC tumours. In the present series, the enhancement pattern of cHCC-CC tumours was HCC-like in most cases., (Copyright © 2019 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.)

Hepatocellular carcinoma (HCC) is one of the most common cancers and has a high mortality rate in less developed countries, especially in China. Galangin (GA), one of the most important and naturally active flavonoids, extracted primarily from the root of Alpinia officinarum Hance, has been demonstrated to be effective in the treatment of HCC. It is a substance with defensive actions and a broad range of biological properties, including inhibitory effects on bacteria, fungi, viruses, the control of hypertension and diabetes, and chemoprevention of several cancers. Experiments have shown that GA prevents HCC through multiple anti-cancer mechanisms, anti-genotoxic activity against environmental and dietary carcinogens; anti-proliferative effects through reversal of the Warburg effect in HCC; arrest of the cell cycle in the G0/G1 phase; induction of apoptosis via stimulation of reactive oxygen species (ROS), endoplasmic reticulum (ER) stress, and the mitochondrial-dependent apoptosis pathway; induction of autophagy; and inhibition of angiogenesis, metastasis, and multidrug resistance (MDR). In addition, synergistic effects with other chemotherapy drugs have been demonstrated. Therefore, this review is focused on the anti-HCC mechanisms of GA., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Thyroid metastasis from hepatocellular carcinoma (HCC) is very rare. We present a case of a 41-year-old man with a markedly elevated serum α-fetoprotein (2,297 ng/mL) and a left neck mass 6 months after a hepatolobectomy for a primary HCC. F-FDG PET/CT imaging showed a focal uptake in the left thyroid lobe corresponding to a mass. The patient underwent a left hemithyroidectomy. Subsequent pathology confirmed HCC metastasis to the thyroid.

Purpose: To build and validate multivariate normal tissue complication probability (NTCP) models for radiation-induced hepatic toxicity (RIHT) after stereotactic body radiation therapy (SBRT)., Methods: Eighty-five patients with hepatocellular carcinoma (HCC) in a phase II clinical trial were enroled. A progression of at least 1 or 2 points in the Child-Pugh (CP) score post-SBRT was classified as RIHT (≥1 or ≥2). NTCP models for RIHT (≥1 or ≥2) were developed using logistic regression. Nomograms for each model were formulated. The cut-off point of each independent dosimetric risk factor was obtained using receiver-operating characteristic (ROC) analysis. We used an independent cohort (101 patients) for model validation., Results: Twenty (23.5%) and 12 (14.2%) patients experienced RIHT (≥1) and RIHT (≥2), respectively. V 15 , VS 10 , and pretreatment CP (pre-CP) were the optimal predictors for RIHT (≥1 and ≥2) modelling. V 15 ≤33.1% and VS 10 ≥416.2 mL for RIHT (≥1), and V 15 ≤21.5% and VS 10 ≥621.8 mL for RIHT (≥2), were the cut-off points. Four NTCP models and their nomograms were generated. These models and nomograms showed good prediction performance (area under the curve (AUC), 0.83-0.89). Our NTCP model (RIHT ≥2) based on V 15 plus pre-CP performed well (AUC = 0.78) in a validation cohort., Conclusion: V 15 , VS 10 , and pre-CP are crucial predictors for RIHT (≥1 and ≥2). Our NTCP models and nomograms were conducive to obtain individual constraints for patients with HCC., Registration Number: ChiCTR-IIC-16008233., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Purpose: Hepatocellular carcinoma (HCC) after insufficient radiofrequency ablation (RFA) could induce epithelial-mesenchymal transition (EMT) in residual tumours, resulting in rapid and aggressive recurrence. However, the role of EMT-related Long noncoding RNAs (lncRNAs) in residual tumour progression remains unclear., Methods: Insufficient RFA was simulated in vitro by heating Huh7 cells in water bath at 47 °C, named as Huh7-H. Cell invasion, migration assays and wound healing assay were conducted for functional analysis. Cell proliferation was determined by CCK8 assay. Differential expression profile of EMT-related lncRNAs between Huh7-H and Huh7 was analysed by LncPath human EMT array, and validated by qRT-PCR. Gain/loss-of-function assays of selected lncRNA were conducted by over-expressing or silencing its expression., Results: Huh7-H presented characteristic EMT morphological changes. WB analysis showed significantly decreased E-cadherin in Huh7-H cells. Transwell assays indicated the abilities of Huh7-H cells in migration and invasion were evidently strengthened. A new lncRNA, FUNDC2P4, was identified by LncPath human EMT array to be significantly down-regulated in Huh7-H cells. In vitro studies showed overexpression of FUNDC2P4 inhibited proliferation, invasion and migration potential and up-regulated E-cadherin expression in SMMC-7721 cells, whereas silencing FUNDC2P4 promoted these potentials and down-regulated E-cadherin expression in Huh7 cells., Conclusions: We explored that lncRNA FUNDC2P4 down-regulation promoted EMT leading to tumour proliferation, invasion and migration by reducing E-cadherin expression in residual HCC after insufficient RFA in vitro. These results suggest that FUNDC2P4 may have potentially therapeutic value for prevention and treatment of HCC recurrence after RFA in the future.

Background: Amino acids are increasingly being recognized as important signaling molecules in the pathogenesis of many diseases. We aim to establish a reversed-phase high-performance liquid chromatography with fluorescence detector (RP-HPLC-FLD) method for determination and quantification of hepatoma-associated DL-amino acids and to explore the relationship between amino acid concentrations and hepatocellular carcinoma (HCC)., Methods: In this work, O-phthaldialdehyde (OPA) and N-isobutyryl-L-cysteine (NAC) served as the pre-column derivatization reagents which significantly shortened the detection time and improved the detection sensitivity. Chromatographic determination was achieved using a programmed gradient elution with a flow rate of 1.0 mL/min. The eluted solution was monitored by a fluorescence detector with an excitation wavelength at 350 nm and an emission wavelength at 450 nm. Under the optimum conditions, an excellent quantification of DL-threonine, alanine, tyrosine, valine, methionine and phenylalanine was achieved., Results: Total analysis time was shortened to less than 25 min for one plasma sample and the linearity, recovery, intra- and inter-day precision were all meet the detection requirements of the DL-amino acids enantiomers in human plasma samples., Conclusions: The developed method demonstrates that DL-threonine, alanine, tyrosine, valine and methionine obtained from HCC patients' plasma samples have a close relationship with HCC. The method would be a potentially alternative tool for DL-amino acids detection in clinical samples., (© 2018 by the Association of Clinical Scientists, Inc.)

Nuclear receptor coactivator 5 (NCOA5) plays important roles in the development of a variety of malignancies. However, the underlying mechanisms remain obscure. In this study, we successfully generated the NCOA5 knockout hepatocellular carcinoma (HCC) cells by CRISPR/Cas9 - mediated genome editing and found that knockout of NCOA5 inhibited the proliferation and tumor microsphere formation of HCC cells significantly. Moreover, the migration ability of NCOA5 knockout HCC cells declined. Mechanistic analyses indicated that knockout of NCOA5 can suppress the epithelial - mesenchymal transition (EMT) in HCC cells. In conclusion, our findings provide a mechanistic insight into the role of NCOA5 in HCC progression., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: The high incidence of recurrence and metastasis of hepatocellular carcinoma (HCC) necessitate the discovery of new predictive biomarkers of invasion and prognosis. Minichromosome maintenance complex component 6 (MCM6), which has been reported to up-regulate in multiple malignancies, was considered to be a novel diagnoses biomarker in HCC. However, its functional contributions and prognostic value remain unclear., Methods: The expression of MCM6 was analyzed in 70 HCC tissues and 5 HCC cell lines by immunohistochemistry and real-time RT-PCR. The roles of MCM6 in HCC cell proliferation, migration and invasion were explored by CCK8, Wound healing and Transwell assays, respectively. Western blotting and Immunofluorescence staining were conducted to detect the protein expressions of ERK signaling pathway and EMT-related markers. To verify the above findings in vivo, we established subcutaneous xenograft tumor and orthotopic xenograft tumor models in nude mice. Finally, Enzyme-linked immunosorbent assay was used to evaluate the serum MCM6 level., Results: MCM6 was significantly up-regulated in HCC tissues. Increased MCM6 expression was associated with aggressive clinicopathological features and worse prognosis in HCC patients. These results were consistent with our analyses of The Cancer Genome Atlas database (TCGA). Furthermore, knockdown of MCM6 significantly decreased proliferative and migratory/invasive capability of HCC cells in vitro, as well as decreased tumor volume, weight and the number of pulmonary metastases in vivo. Mechanistic analyses indicated that MCM6 promoted EMT and activated MEK/ERK signaling. More importantly, serum MCM6 levels in HCC patients were significantly higher than those in cirrhosis and healthy controls (P < 0.0001), and allowed distinguishing early recurrence with high accuracy (AUC = 0.773)., Conclusions: Our findings indicate that MCM6 predicts poor prognosis and promotes metastasis in HCC. Postoperative serum MCM6 level could be valuable to detect preclinical early recurrence, indicative of a need for more careful surveillance and aggressive therapeutic intervention.

Background: Forkhead box A1 (FOXA1) expression is associated with various types of tumors; however, the function and underlying mechanism of FOXA1 in the development of hepatocellular carcinoma (HCC) remains obscure., Methods: Here, we investigated the role of FOXA1 in the development of HCC by applying gene function gain and loss analysis to HepG2 and Hep3B cell lines, and comparing outcomes with those of clinical HCC samples., Results: Phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1), which encodes protein PI3Kp85 (p85), was identified as a FOXA1 target gene. Analyses of the mechanism and function revealed that FOXA1 suppresses hepatocellular carcinoma cell viability and motility by inhibiting PI3K/Akt signaling through direct inhibition of PIK3R1 transcription. Moreover, in clinical samples from male HCC patients, FOXA1 expression was much lower, whereas PI3Kp85 levels were much higher in tumor than in non-tumor tissues. Elevated PI3Kp85 is an unfavorable factor in HCC., Conclusions: As a tumor suppressor, FOXA1 targets PIK3R1 directly to inhibit PI3K/Akt signaling pathway, thus exerting a negative regulatory effect on proliferation, migration, and invasion of HCC in male patients.

Schizophrenic patients tend to have reduced incidence of some cancers due to the treatment of antipsychotic drugs with antitumor effects, such as chlorpromazine and trifluoperazine (TFP). Forkhead Box O1 (FOXO1) as tumor suppressor in many malignancies is often inactivated by nuclear export, which could be inhibited by TFP. However, the antitumor efficiency of TFP and related role of FOXO1 in hepatocellular carcinoma (HCC) are unclear. Thus, two HCC cell lines SMMC-7721 and Bel-7402 were treated with different concentrations of TFP and the IC50 was determined. We found that TFP could inhibit the vitality of two cell lines and induce cell cycle arrest at G0/G1. Meanwhile, the apoptosis was also increased and the ability of migration or invasion was found to be impaired by TFP. Interestingly, TFP reversed the cytoplasmic localization of FOXO1 to nuclear and increased its expression in nuclear, and increased the ratio of Bax/Bcl-2. However, knockdown of FOXO1 significantly abrogated the TFP-induced apoptosis by decreasing the Bcl-2 expression [corrected]. Furthermore, we found that TFP in vivo could effectively restrict the angiogenesis and tumor growth with reduced expression of VEGF, Bcl-2, and PCNA, and increased the nuclear localization of FOXO1, which indicated its antitumor role in HCC.

Background: Hepatocellular carcinoma (HCC) is the most common type of liver cancer worldwide. Previously, we reported that cadherin-17 (CDH17) and its related CDH17/β-catenin axis may be responsible for inducing HCC in a subset of patients exhibiting CDH17 over-expression. Here we aimed at obtaining a better understanding of the CDH17-related HCC biology and to obtain further indications for the design of targeted therapies in CDH17 over-expressing HCC patients., Results: We found that SPINK1 acts as a downstream effector of the CDH17/β-catenin axis in HCC. In addition, we found that SPINK1 expression exhibited a positive correlation with CDH17 expression in human HCCs and was over-expressed in up to 70% of the tumors. We identified SPINK1 as a downstream effector of the CDH17/β-catenin axis using a spectrum of in vitro assays, including gene expression modulation and inhibitor assays, bioinformatics analyses and luciferase reporter assays. These in vitro results were validated in primary human HCCs, including the observation that alteration in β-catenin expression (a core component of the CDH17/β-catenin axis) in tumors affects SPINK1 serum levels in HCC patients. Similar to CDH17, SPINK1 expression in HCC cells was found to be associated with specific tumor-related properties via activating the c-Raf/MEK/ERK pathway., Conclusions: Our current data substantiate our knowledge on the role of CDH17 in the biology of HCC and suggest that components of the CDH17/β-catenin axis may serve as therapeutic targets in CDH17 over-expressing HCC patients.

Hyperactivation of transforming growth factor-β (TGF-β) signaling pathway is a common feature of hepatocellular carcinoma (HCC) progression. However, the driver factors leading to enhanced TGF-β activity are not well characterized. Here, we explore the mechanisms that loss of Krüppel-like factor 4 (KLF4) exacerbates oncogenic TGF-β signaling in human HCC. The expression of KLF4 and TGF-β signaling components in primary HCC and their clinicopathologic relevance and significance was evaluated by using tissue microarray and immunohistochemistry. Cellular and molecular impacts of altered KLF4 expression and TGF-β signaling were determined using immunofluorescence, western blot, reverse-transcriptase PCR, chromatin immunoprecipitation and promoter reporter assays. Loss of KLF4 expression in primary HCC closely correlated with decreased Smad7 expression, increased p-Smad2/3 expression and independently predicts reduced overall and relapse-free survival after surgery. TGF-β signaling components were expressed in most HCC cells, and activation of TGF-β signaling promoted cell migration and invasion. Enforced KLF4 expression blocked TGF-β signal transduction and inhibited cell migration and invasion via activation of Smad7 transcription, whereas deletion of its C-terminal zinc-finger domain diminished this effect. KLF4 protein physically interacts with the Smad7 promoter. Promoter deletion and point mutation analyses revealed that a region between nucleotides -15 bp and -9 bp of the Smad7 promoter was required for the induction of Smad7 promoter activity by KLF4. Our data indicate that KLF4 suppresses oncogenic TGF-β signaling by activation of Smad7 transcription, and that loss of KLF4 expression in primary HCC may contribute to activation of oncogenic TGF-β signaling and subsequent tumor progression.

Type 2 Diabetes (T2D) is a risk factor for hepatocellular carcinoma (HCC). We have previously described that haploinsufficiency of nuclear receptor coactivator 5 (NCOA5) is a genetic defect linking glucose intolerance to HCC. Here we report identification and characterization of a single nucleotide variation (T445A) in NCOA5, causing an amino acid Thr to Ala substitution, in adjacent non-tumorous liver tissues derived from patients with concurrent HCC and T2D. By using Tet-On inducible expression cells, we show that ectopic expression of NCOA5wt suppressed proliferation of HCC cells via induction of G2/M arrest, while ectopic expression of NCOA5T445A had a significantly lesser effect compared to ectopic expression of NCOA5wt. Furthermore, ectopic expression of NCOA5wt increased the occurrence of DNA damage and cell senescence, whereas expression of NCOA5T445A partly lost this activity. Xenograft tumor model analysis demonstrated that ectopic NCOA5wt expression reduced HCC tumor growth and the T445A variation impairs its tumor growth inhibitory function. Collectively, our data show that the T445A variation impairs the ability of NCOA5 to inhibit growth of HCC, suggesting that this variation may have potential to increase susceptibility to HCC comorbid with T2D., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Drug resistance to sorafenib is common in patients with hepatocellular carcinoma(HCC). We examined the effects of a combination of sorafenib and fluvastatin on HCC using in vitro and in vivo models. The dual treatment induced apoptosis and reduced cellular viability in HCC more effectively than either drug alone. The combination treatment also inhibited activation of hepatic stellate cells, whereas single drug treatments did not. On a molecular level, combined treatment inhibited activation of the MAPK and NF-κB pathways via Toll-like receptor 4 in HCC cells. Combined treatment also inhibited expression of stromal cell-derived factor 1α in HCC cells, which further inhibited the MAPK pathway in hepatic stellate cells. These results suggest that a combination of sorafenib and fluvastatin may be a promising therapeutic strategy for patients with advanced HCC.

Purpose: Although the albumin to globulin ratio (AGR) has been proven to be a prognostic factor in several cancers, no studies have assessed its prognostic significance in hepatocellular carcinoma (HCC). Therefore, this study aimed to investigate the prognostic value of the pretreatment AGR in the survival in HCC patients., Methods: 150 patients were enrolled, who were confirmed of HCC from October 2008 to December 2012 in Nanfang Hospital of Southern Medical University. Demographic, clinical and laboratory data were obtained. Univariate and multivariate Cox regression analysis were used to investigate the association of clinicopathological parameters with HCC patients' survival., Results: Patients were divided into 2 groups: AGR?1.18 and AGR ?1.18. Patients in the high AGR (?1.18) group had longer overall survival (OS) than those in the low AGR (<1.18) group (60.16 vs 20.48 months, p<0.001). Univariate analysis showed that portal vein tumor thrombosis, grade of differentiation, extrahepatic metastasis, BCLC stage, AFP level and AGR at diagnosis were significantly associated with OS. Multivariate analysis revealed that AGR (p<0.001) and grade of differentiation (p=0.007) were independent prognostic factors for survival of HCC patients. In subgroup analysis based on age and Child-Pugh class, AGR remained a significant prognostic parameter., Conclusion: Low pretreatment AGR was significantly associated with shorter OS in HCC patients. The pretreatment AGR could be a useful and effective prognostic index for identifying patients with poor prognosis, even when patients have well-preserved liver reserve function.

Background: Several studies have found that Cathepsin B (CTSB) is up-regulated in many tumor types and facilitates tumor progression. However, the role of CTSB in hepatocellular carcinoma (HCC) progression remains unclear. This study was aimed at investigating the expression and role of CTSB in HCC in a large set of samples and cell lines (MHCC-97H and MHCC-97 L), and evaluating the clinical and prognostic significance of CTSB protein in patients with HCC., Methods: The expression of CTSB was examined in HCC tissue and cell lines by Western-blotting, Real-time PCR, and immunohistochemical staining. Wound healing assay and invasion assay were used to verify the effect of CTSB on the migration and invasion ability of HCC cell lines. Tumor formation assay in nude mice was used to analyze the effect of CTSB on the tumorigenicity of HCC cell lines., Results: The status of CTSB protein in carcinoma tissues is much higher than that in paracarcinoma tissues. The overall survival of the patients with high CTSB expression was significantly shorter than the low CTSB expression group. High CTSB expression was significantly correlated with advanced clinical staging, histological grade, and tumor recurrence. In vitro and in vivo experiments demonstrated that over-expression of CTSB in MHCC-97 L cells promoted cell invasion and tumor progression ability. Down-regulation of CTSB in MHCC-97H showed the opposite effects. These phenotypic changes caused by CTSB knockdown or over-expression correlated with expression of the matrix metallopeptidase MMP-9. Moreover, multivariate analysis suggested that CTSB expression might be an independent prognostic indicator for the survival of HCC patients after curative surgery., Conclusions: CTSB might be involved in the development and progression of HCC as an oncogene, and thereby may be a valuable prognostic marker for HCC patients.

Although protein kinase D (PKD) has been shown to contribute to invasion and metastasis in several types of cancer, the role of PKD in the epithelial mesenchymal transition (EMT) of hepatocellular carcinoma (HCC) has remained unclear. We found that PKD2 is up-regulated in HCC and is correlated with the metastasis of HCC. PKD2 positively regulated TNF-α-induced EMT and metastasis of HCC. Mechanistic studies revealed TNF-α-induced PKD2 activation is mediated by the formation of a TNFR1/TRAF2 complex. PKD2 bound directly to the p110α and p85 subunits of PI3K and promoted the PI3K/Akt/GSK-3β signaling cascade to stimulate EMT. In conclusion, our results have uncovered a novel role for the regulation of EMT and suggest inhibition of PKD2 as a potential therapeutic strategy for HCC.

Previous studies have reported the association between excess body mass index (BMI) and increased risk of hepatocellular carcinoma (HCC). However, whether BMI is associated with the prognosis and postoperative complications of HCC is still not clear.We searched PubMed and Embase for relevant studies published until the date of August 30, 2014. Additional studies were manually identified by searching reference lists of retrieved articles. Pooled hazard ratios (HRs) with 95% confidence intervals (95% CIs) for overall survival (OS), disease-free survival (DFS), and risk ratios (RRs) with 95% CIs for postoperative complications were calculated using random effects or fixed effects models according to heterogeneities between studies.A total of 14 studies were included in the present meta-analysis. The pooled results showed that excess BMI was not significantly associated with improved OS (HR = 0.94; 95% CI: 0.74-1.19, P = 0.588) or DFS (HR = 0.93; 95% CI: 0.79-1.10, P = 0.382). In addition, higher BMI was not associated with increased rate of a number of complications including ascites (RR = 1.25, 95% CI: 0.94-1.65, P = 0.119), bile leaks (RR = 1.22, 95% CI: 0.81-1.83, P = 0.345), and 30-day mortality (RR = 1.05, 95% CI: 0.57-1.96, P = 0.871). However, HCC patients with higher BMI had increased incidence of wound infections (RR = 2.17, 95% CI: 1.28-3.68, P = 0.004).BMI was not an independent prognostic factor for the evaluation of the prognosis in HCC patients, and it was not associated with postoperative complications except for wound infections that as significantly associated with higher BMI scores.

Background: MicroRNAs (miRNAs) can serve as tumor suppressors and might provide an efficient strategy for annihilating tumor cells. Nevertheless, the potential role of miR-325 in hepatocellular carcinoma (HCC) is still unknown., Methods: Using RT-PCR, immunoblots invasion assays and bioinformatics strategies, we investigated the potential role of miR-325 in HCC., Results: We showed that miR-325 was decreased and HMGB1 was increased in 99 patients with hepatocellular carcinoma. MiR-325 inhibition promoted cell invasion and proliferation, while miR-325 upregulation inhibited cell invasion and proliferation by using transwell and CCK8 assays. We further showed that HMGB1 might be a direct target of miR-325 and is negatively regulated by miR-325. Down-regulation of miR-325 predicts poor prognosis for HCC patients., Conclusions: These findings implied that miR-325 regulates cell invasion and proliferation via targeting HMGB1 and may be a potential prognostic marker for HCC., Virtual Slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/4655707031717989.

Tumor-initiating cells (TICs) are a subpopulation of chemoresistant tumor cells that have been shown to cause tumor recurrence. Targeting and eliminating of TICs are therefore priorities for the development of new therapeutic paradigms. Much promise lies in adoptive immunotherapy. Cytokine-induced killer (CIK) cells are heterogeneous ex vivo-expanded T lymphocytes, with a mixed T-NK phenotype. It represents a realistic new option in the field of hepatocellular carcinoma (HCC) immunotherapy. In the very recent years, large clinical trials demonstrated that CIK cells could improve the Progression Free Survival (PFS) and Overall Survival (OS) in patients with HCC. By the same time, several studies reported that CIK cells were capable of clearing cells with stemness features in lymphoma, melanoma, bone and soft-tissue sarcomas. Based on the findings above mentioned, we hypothesized that CIK cells could eliminate the tumor-initiating cells, improving the PFS and OS of patients with HCC when combined with radiofrequency ablation (RFA) or transcatheter arterial chemoembolization (TACE)., (Copyright © 2014. Published by Elsevier Ltd.)

Objectives: To investigate the expression and role of Cathepsin L (CTSL) in Hepatocellular carcinoma (HCC) tissue and cell line (MHCC-97H), and to evaluate the clinical and prognostic significance of CTSL protein in patients with HCC., Methods: The expression of CTSL was examined in HCC tissue and MHCC-97H cells by Western-blotting, Real-time PCR and immunohistochemical staining. Cell growth curve assay and colony formation assay were used to verify the effect of CTSL on the proliferation and tumor progression ability of MHCC-97H cells. Tumor formation assay in nude mice was used to analyze the effect of CTSL on the tumorigenicity of MHCC-97H cells., Results: The status of CTSL protein in carcinoma tissues is much higher than that in paracarcinoma tissues. The overall survival of the patients with high CTSL expression was significantly shorter than the low CTSL expression group. high CTSL expression was significantly correlated with advanced clinical staging, histological grade and tumor recurrence. In vitro experiments demonstrated that over-expression of CTSL in MHCC-97H cells promoted cell proliferation and tumor progression ability. Down-regulation of CTSL showed the opposite effects. Over-expression of CTSL increase the tumorigenicity of MHCC-97H cells by in vivo experiments. Moreover, multivariate analysis suggested that CTSL expression might be an independent prognostic indicator for the survival of HCC patients after curative surgery., Conclusions: CTSL might involve in the development and progression of HCC as a oncogene, and thereby may be a valuable prognostic marker for HCC patients.

Purpose: Modified Response Evaluation Criteria In Solid Tumors (mRECIST), developed by the American Association for the Study of Liver Diseases (AASLD) criteria measure changes in arterialized hepatocellular carcinoma (HCC) and aim at providing a common framework for the design of clinical trials. It still isn't determined whether mRECIST can be applied in routine clinical practice and whether mRECIST could estimate viable tumor correctly., Methods: We retrospectively analyzed data from patients subjected to transcatheter arterial chemoembolization (TACE) as initial treatment for advanced HCC in our institution. Not suitable for using mRECIST standard cases and the agreement in response between RECIST and mRECIST were assessed. Then we selected HCC patients who achieved complete response (CR) according to mRECIST, following PET-CT examinations. We also compared arterial enhanced computed tomography (CT) or magnetic resonance imaging (MRI) with positron emission tomography (PET)-CT examination and analyzed their correlation., Results: Out of 143 HCC patients, mRECIST evaluation appeared to be applicable for 128 (89.51%) assessable patients. In these 128 assessable patients, the objective response (OR) rates (complete/CR+partial response/PR) according to RECIST and mRECIST were 64.06% (82 of 128 patients) and 78.13% (100 of 128; p<0.001), respectively. Discordance in the response evaluations between the two methods was observed in 46 patients (35.94%) and was statistically significant (Kappa=0.491; p<0.001). The overall survival (OS) of patients who achieved an OR as assessed by mRECIST or by RECIST was significantly better than the survival of non-responding patients (stable disease/SD, or progressive disease/PD)., Conclusions: Although mRECIST criteria show a good correlation with prognosis, they demand strict requirements for patient selection and couldn't be useful as a tool for routine clinical practice. Furthermore, merely by means of contrast-enhanced CT or MRI, mRECIST couldn't estimate viable tumor sufficiently.

Hepatocellular carcinoma(HCC) is serious condition associated with a high morbidity and mortality. Therefore is an urgent need to develop novel noninvasive techniques for early diagnosis, particularly for patients with AFP-negative [AFP(-)] HCC. In this study, iTRAQ-MALDI-MS/MS was used to identify differentially expressed proteins in AFP(-) HBV-related HCC compared with non-cancerous hepatitis B virus (HBV) and healthy controls subjects.Serum was obtained from 18 patients with AFP(-) HBV-related HCC, 18 matched patients with HBV without HCC and 18 healthy control subjects. High abundance proteins were removed from serum and the differentially expressed proteins from the three groups were screened out using iTRAQ-MALDI-MS/MS. The Gene Ontology (GO) function and the interaction networks of differentially expressed proteins were then analyzed. A total of 24 expressed differential proteins associated with AFP(-) HBV-related HCC were screened out, 15 proteins were up-regulated and 9 down-regulated. The most common molecular function of the 24 differentially expressed proteins was enzyme inhibition. Interaction network of the 24 differentially expressed proteins showed that 14 proteins (C5, KNG1, FN1, LRG1, HRG, SERPINC1, CRP, APOB, SAA1, APCS, C4BPA, CFI, CFB and GSN) were central to the functional network. The expression levels of the GSN protein were down-regulated in AFP(-) HBV-related HCC subjects compared with healthy controls and the HBV group (p<0.01), consistent with the iTRAQ results.The 14 proteins from the serum of AFP(-) HBV-related HCC appeared at the fulcrum of the functional network and were differentially expressed compare to HBV and healthy controls suggesting a possible association with HCC progression.

Type 2 diabetes (T2D) and male gender are associated with hepatocellular carcinoma (HCC) development. We demonstrate that heterozygous deletion of the Ncoa5 gene causes spontaneous development of HCC exclusively in male mice. Tumor development is preceded by increased interleukin-6 (IL-6) expression, early-onset glucose intolerance, and progressive steatosis and dysplasia in livers. Blockading IL-6 overexpression averts glucose intolerance and partially deters HCC development. Moreover, reduced NCOA5 expression is associated with a fraction of human HCCs and HCCs with comorbid T2D. These findings suggest that NCOA5 is a haploinsufficient tumor suppressor and that NCOA5 deficiency increases susceptibility to both glucose intolerance and HCC, partially by increasing IL-6 expression. Thus, our findings open additional avenues for developing therapeutic approaches to combat these diseases., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Fibulin-3, originally identified in senescent and Werner syndrome fibroblasts, has been implicated in cell morphology, growth, adhesion and motility. Fibulin-3 exhibits both antitumor and oncogenic activities towards human cancers; however, the role of Fibulin-3 in hepatocellular carcinoma (HCC) remains elusive. In this study, we showed that both the mRNA and protein levels of Fibulin-3 were remarkably downregulated in HCC cell lines and fresh tissues. Immunohistochemical data revealed that Fibulin-3 was decreased in tumorous tissues in 67.1% (171/255) of cases compared to the corresponding adjacent nontumorous tissues. The results of statistical analysis indicated that low Fibulin-3 expression, defined by the receiver operating characteristic curve (ROC), was significantly associated with tumor differentiation (P=0.008), clinical stage (P=0.014) and serum AFP levels (P<0.01). Furthermore, Kaplan-Meier and multivariate analysis suggested that Fibulin-3 is an independent negative prognostic indicator for both overall (P<0.001) and recurrence-free (P=0.036) survival. In addition, an in vitro study demonstrated that knockdown of Fibulin-3 by siRNA markedly increased cell viability and promoted cell invasion in HCC cells. Collectively, our data suggest that Fibulin-3 exhibits antitumor effects towards HCC and serves as a biomarker of unfavorable prognosis for this deadly disease.

Objective: To determine the differentially expressed serum proteins in patients with hepatoma carcinoma and identify a putative diagnostic marker., Method: The isobaric tags for relative and absolute quantitation (iTRAQ) labeling method and LC-MALDI-TOF/TOF MS detection method were used to quantify serum proteins in hepatocellular carcinoma patients (n =20) and healthy individuals (n =20). Real-time reverse transcription-polymerase chain reaction was used to verify the differentially expressed proteins by analyzing the corresponding mRNA expression levels in the hepatic carcinoma and healthy hepatocyte samples, as well as in 30 pairs of patient-matched hepatic carcinoma and adjacent normal tissue samples. Western blot analysis was used to verify the protein expression in hepatic carcinoma cells., Result: Fifty-one proteins were significantly differentially expressed between the hepatic carcinoma group and healthy controls. The iTRAQ protein profile showed that the serum level of clusterin was significantly lower in hepatoma carcinoma patients. The mRNA level of clusterin was 20-fold lower in hepatic carcinoma cells than in healthy hepatocytes, and was 2.38-fold lower in hepatoma tissues than that in adjacent normal tissues. The clusterin protein levels were significantly lower in hepatic carcinoma cells (8.06 vs normal hepatocytes: 27.81; P less than 0.01)., Conclusion: The serum expression of clusterin is significantly decreased in both serum and tissues of hepatic carcinoma patients. The relationship between hepatic carcinoma and clusterin should be evaluated in future studies.