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Sorafenib remains the standard first-line treatment for advanced hepatocellular carcinoma (HCC), although other clinical trials are currently underway for treatments that show better curative effects. However, some patients are not sensitive to sorafenib. α-Mangostin, extracted from the pericarp of the mangosteen, which is widely used as a traditional medicine, has anticancer and anti-proliferative properties in various types of cancers, including HCC. In the present study, we found that combining sorafenib and α-Mangostin could be synergistically toxic to HCC both in vitro and in vivo. We then demonstrated that the combination of sorafenib and α-Mangostin enhances the inhibition of cell proliferation in HCC cell lines. Combination therapy leads directly to apoptosis. In xenograft mouse models, the in vivo safety and effectivity was confirmed by a reduction in tumor size after combination treatment. RNA sequencing and protein testing showed that the expression of LRRC8A and RNF181 genes and mTOR and MAPK pathways may be associated with the synergistic effect of the two drugs. In conclusion, our results highlight the synergistic effect of the combination of sorafenib and α-Mangostin, which indicates a potential treatment for advanced HCC for patients that are not sensitive to sorafenib therapy.

Circular RNAs (circRNA) have been reported as regulators involved in hepatocellular carcinoma (HCC), but their mechanism of activity remains unknown. This study performed quantitative reverse-transcription polymerase chain reaction to determine if circNFATC3 was downregulated in 46 paired HCC tissues and cell lines. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, apoptotic, and transwell assay proved that circNFATC3 can inhibit hepatoma cell proliferation, apoptosis, and migration/invasion in vitro. Mouse xenograft assay demonstrated that circNFATC3 suppressed tumor size and weight and reduced lung metastasis in vivo, and vice versa. The RNA-seq results showed that NFATC3 itself was the most significantly differentially expressed gene when circNFATC3 was manipulated, and bioinformatics and luciferase reporter assays verified circNFATC3 regulated the expression of NFATC3 by interacting with the hsa-miR-548I. Additionally, it was also indicated that the level of NFATC3 was downregulated in HCC patients also and was significantly correlated with the staging and prognosis of HCC. Moreover, both circNFATC3 and NFATC3 were shown to inhibit the phosphorylation of JNK, c-Jun, AKT, and mTOR signaling pathways. Overall, the circNFATC3 can sponge miR-548I to protect NFATC3 itself, then it regulates hepatoma cell function via the JNK, c-Jun, AKT, and mTOR signaling pathways, and the circNFATC3 can be a tumor-repressor on HCC.

Long noncoding RNAs (lncRNAs), such as LINC00462, HOTAIR, and MALAT1, are significantly upregulated in hepatocellular carcinoma (HCC) tissues. However, lncRNA expression in the serum of HCC patients is still unclear. To identify candidate lncRNAs for HCC diagnosis, we purified exosomal total RNA from the serum of healthy volunteers (controls) and hepatitis, cirrhosis, and HCC patients. To assess the function of lncRNAs, small interfering RNAs and overexpression vectors were designed and cell viability and cell apoptosis assays conducted. The exosomes of the control group had a larger number of lncRNAs with a high amount of alternative splicing compared to hepatic disease patients. qPCR assays showed that lnc-FAM72D-3, lnc-GPR89B-15, lncZEB2-19, and lnc-EPC1-4 are differentially expressed in HCC. Furthermore, the expression level of lnc-EPC1-4 correlated with age. While the expression levels of lnc-GPR89B-15 and lnc-EPC1-4 correlated with serum alpha-fetoprotein level. lnc-FAM72D-3 knockdown decreased cell viability and promoted cell apoptosis, indicating that lnc-FAM72D-3 functions as an oncogene in HCC. In contrast, lnc-EPC1-4 overexpression inhibited cell proliferation and induced cell apoptosis, indicating that it functions as a tumor suppressor gene. Collectively, these findings show that lnc-FAM72D-3 and lnc-EPC1-4 play a novel role that might contribute to hepatocarcinogenesis and identify potential candidate biomarkers for HCC diagnosis.

Cancer-associated fibroblasts (CAFs) play crucial roles in enhancing cell survival, proliferation, invasion, and metastasis. We previously showed that hepatocellular carcinoma-derived CAFs (H-CAFs) promoted proliferation of hepatocellular carcinoma (HCC) cells. This study aimed to further explore the role of CAFs in HCC epithelial-mesenchymal transition (EMT) and the underlying mechanism. High CAF density was significantly associated with liver cirrhosis, inferior clinicopathologic characteristics, elevated EMT-associated markers, and poorer survival in human HCC. Within HCC cells, EMT was induced after co-culture with H-CAFs. Secretomic analysis showed that IL-6 and HGF were the key EMT-stimulating cytokines secreted by H-CAFs. Proteomic analysis revealed that TG2 was significantly upregulated in HCC cells with EMT phenotypes. Overexpression of TG2 promoted EMT of HCC cells, and knockdown of TG2 remarkably attenuated the H-CAF-induced EMT. Furthermore, during EMT, TG2 expression was enhanced after HCC cells were stimulated by IL-6, but not HGF. Inhibition of the IL-6/STAT3 signaling decreased TG2 expression. The principal TG2 transcription control element and a potential STAT3 binding site were identified using promoter analysis. Hence, H-CAFs facilitates HCC cells EMT mediated by IL-6, which in turn activates IL-6/IL6R/STAT3 axis to promote TG2 expression.

As a novel immune-nutritional biomarker, the controlling nutritional status (CONUT) score has been reported to predict outcomes in cancer patients. We aimed to elucidate the prognostic value of preoperative CONUT score and construct a CONUT score-based nomogram to predict individual survival of patients with hepatitis B viral (HBV)-associated hepatocellular carcinoma (HCC) after curative hepatectomy. Preoperative CONUT score was retrospectively calculated in 380 HBV-associated HCC patients undergoing radical resection between 2006 and 2012. Patients were assigned to two groups: CONUT-low (

MALAT1-associated small cytoplasmic RNA (mascRNA) is a cytoplasmic tRNA-like small RNA derived from nucleus-located long noncoding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1). While MALAT1 was extensively studied and was found to function in multiple cellular processes, including tumorigenesis and tumor progression, the role of mascRNA was largely unknown. Here we show that mascRNA is upregulated in multiple cancer cell lines and hepatocellular carcinoma (HCC) clinical samples. Using HCC cells as model, we found that mascRNA and its parent lncRNA MALAT1 can both promote cell proliferation, migration, and invasion in vitro. Correspondingly, both of them can enhance the tumor growth in mice subcutaneous tumor model and can promote metastasis by tail intravenous injection of HCC cells. Furthermore, we revealed that mascRNA and MALAT1 can both activate ERK/MAPK signaling pathway, which regulates metastasis-related genes and may contribute to the aggressive phenotype of HCC cells. Our results indicate a coordination in function and mechanism of mascRNA and MALAT1 during development and progress of HCC, and provide a paradigm for deciphering tRNA-like structures and their parent transcripts in mammalian cells.

Hepatic ischemia-reperfusion injury (IRI) is the most common cause of liver damage leading to surgical failures in hepatectomy and liver transplantation. Extensive inflammatory reactions and oxidative responses are reported to be the major processes exacerbating IRI. The involvement of Yes-associated protein (YAP) in either process has been suggested, but the role and mechanism of YAP in IRI remain unclear. In this study, we constructed hepatocyte-specific YAP knockout (YAP-HKO) mice and induced a hepatic IRI model. Surprisingly, the amount of serum EVs decreased in YAP-HKO compared to WT mice during hepatic IRI. Then, we found that the activation of YAP increased EV secretion through F-actin by increasing membrane formation, while inhibiting the fusion of multivesicular body (MVB) and lysosomes in hepatocytes. Further, to explore the essential elements of YAP-induced EVs, we applied mass spectrometry and noticed CD47 was among the top targets highly expressed on hepatocyte-derived EVs. Thus, we enriched CD47+ EVs by microbeads and applied the isolated CD47+ EVs on IRI mice. We found ameliorated IRI symptoms after CD47+ EV treatment in these mice, and CD47+ EVs bound to CD172α on the surface of dendritic cells (DCs), which inhibited DC activation and the cascade of inflammatory responses. Our data showed that CD47-enriched EVs were released in a YAP-dependent manner by hepatocytes, which could inhibit DC activation and contribute to the amelioration of hepatic IRI. CD47+ EVs could be a potential strategy for treating hepatic IRI.

Hepatic ischemia-reperfusion (I/R) injury fundamentally influences the performance of aged liver grafts. The significance of mitophagy in the age dependence of sensitivity to I/R injury remains poorly understood. Here, we show that aging aggravated hepatic I/R injury with decreased mitophagy in mice. The enhancement of mitophagy resulted in significant protection against hepatic I/R injury. Parkin, an E3 ubiquitin ligase, was found depleted by I/R in aged livers. In oxygen-glucose deprivation reperfusion (OGD-Rep.)-treated L02 cells, parkin silencing impaired mitophagy and aggravated cell damage through a relative large mitochondrial membrane potential transition. The phosphorylation of the endoplasmic reticulum stress response protein EIF2α, which was also reduced in the aged liver, induced parkin expression both in vivo and vitro. Forty-six hepatic biopsy specimens from liver graft were collected 2 hours after complete revascularization, followed by immunohistochemical analyses. Parkin expression was negatively correlated to donor age and the peak level of aspartate aminotransferase within first week after liver transplantation. Our translational study demonstrates that aging aggravated hepatic I/R injury by impairing the age-dependent mitophagy function via an insufficient parkin expression and identifies a new strategy to evaluate the capacity of an aged liver graft in the process of I/R through the parkin expression.

Tong Zhang,1–3 Wei Liu,4 Wei Meng,1–3 Hui Zhao,1–3 Qing Yang,1–3 Shi-jie Gu,1–3 Cui-cui Xiao,4 Chang-chang Jia,4 Bin-sheng Fu1–3 1Department of Hepatic Surgery, Liver Transplantation Center, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, People’s Republic of China; 2Organ Transplantation Institute of Sun Yat-sen University, Guangzhou, People’s Republic of China; 3Organ Transplantation Research Center of Guangdong Province, Guangzhou, People’s Republic of China; 4Guangdong Key Laboratory of Liver Disease Research, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, People’s Republic of China Introduction: Hepatocellular carcinoma (HCC) accounts for more than 90% of primary liver cancer. Although great progress has been made on HCC molecular mechanism and therapy techniques, the prognosis of HCC patient is poor due to high metastasis and recurrence. Materials and methods: Expression of miR-542-3p was quantified by quantitative real-time PCR (qRT-PCR). The role of miR-542-3p in HCC metastasis was examined using transwell and 3D-culture assay. qRT-PCR, Western blotting and luciferase reporter assay were used to elucidate the mechanisms of miR-542-3p-mediated cancer metastasis.Results and Conclusion: In the research, we found that miR-542-3p is decreased in HCC cell lines and tissues, and downregulation of miR-542-3p enhances, while upregulation suppresses HCC cell invasion ability. Further assay demonstrated that miR-542-3p can directly target TGF-β1 3' untranslated region (3'UTR) to influence TGF-β/Smad signaling pathway, and suppression of miR-542-3p can hyperactivate TGF-β/Smad pathway and further to promote Epithelial-Mesenchyme Transition (EMT) and induce poor prognosis. Lastly, the clinical correlation analysis illustrated that miR-542-3p is negatively related with the activity of TGF-β1. In summary, our results find that miR-542-3p takes an important role on HCC progression and provide more evidence of microRNAs (miRNAs) for cancer therapy. Keywords: microRNA, HCC, TGF-β, EMT&nbsp

Background Autophagy involves in both prevention and promotion in cancer, and its role probably changed during tumor development. Defined the dynamic function of autophagy in cancer may advance precision diagnostics, treatment, and guide drug design. Autophagy related protein ULK1 is key regulator of autophagy, and its role in hepatocellular carcinoma (HCC) was still unclear. This study aims to investigate ULK1’s capacity along with other autophagic markers in predicting prognosis of HCC and explore position of these biomarkers in dynamic function of autophagy during HCC progression. Methods The expression of ULK1 and other autophagic marker (LC3B) were test by Tissue microarray-based immunohistochemistry in 156 operable HCC patients. Survival analysis and correlation analysis were used to analysis influence of ULK1 and combined biomarker on clinical characteristics and prognosis. Results The expression level of ULK1 was not related to all clinicopathological features, however, high expression of the ULK1 as well as LC3B overexpression suggested large tumor size (P = 0.035), high levels of serum AFP (P = 0.049), more frequency of node metastasis (P = 0.015), later TNM stage (P = 0.009). Survival analysis showed that ULK1 expression were negatively correlated with PFS rather than OS in HCC patients (P = 0.021), while LC3B were suggested to be negatively related with patients’ PFS, However, Simultaneous high expression of ULK1 and LC3B had a poorer 5-year overall survival (OS) rate (P = 0.002) and shorter 5-year progression free survival (PFS)(P = 0.003), Further multivariate analysis revealed that the two combined biomarkers were independent factors to predict the prognosis of OS and PFS in all patients, while ULK1 alone or LC3B alone were only an independent predict factor for OS or PFS respectively. Conclusion ULK1 were demonstrated to be an important prognostic factor for HCC patient, and it combined LC3B would improve prognosis assessment of the patients. Combined autophagic biomarkers would better represent dynamic stage of autophagy and It might provide a potential therapeutic way that how to interfere autophagy in HCC.

Hepatic ischemia-reperfusion injury (IRI) is a major cause of liver damage during liver transplantation and hepatectomy, and its amelioration is an urgent unmet clinical need. Excess inflammatory response and oxidative stress contributes to the injury and Yes-associated protein (YAP) in the Hippo signaling pathway has been proved participating in it. In this study, using hepatocyte-specific YAP knockout (YAP-HKO) by crossing Yapflox/flox mice with Alb-Cre mice, we surprisingly noticed in the serum of these mice, the amount of serum extracellular vesicles (EVs) decreased after IRI. Then we found that YAP from hepatocytes regulate EVs secretion through F-actin by increasing membrane formation, as well as inhibiting the fusion of multivesicular body (MVB) and lysosomes. We further explored the essential element of YAP-induced EVs by mass spectrometry, and noticed CD47 was one of the target that highly expressed on hepatocyte-derived EVs in YAP-dependent manner. In addition, we found that CD47+ EVs can inhibit dendritic cells (DCs) activation and inflammatory responses via binding to CD172α on the surface of DCs. Finally, we isolated CD47+ EVs in IRI model and found IRI being ameliorated after treatment, suggesting that CD47+ EVs could be a potential therapeutic strategy for the treating hepatic IRI. Our data showed that CD47-enriched EVs were released in a YAP dependent manner by hepatocytes, which could inhibit DCs activation and contribute to the amelioration of hepatic IRI. Funding Statement: National Key RD the China Postdoctoral Science Foundation, Grant/Award Number: 2020M672980; the National 13th Five-Year Science and Technology Plan Major Projects of China, Grant/Award Number: 2017ZX10203205-006-001; National Natural Science Foundation of China, Grant/Award Number: 81702393, 81770648, 81670601, 81570593, 81802897, 81802897, 81900886; Key Scientific and Technological Projects of Guangdong Province, China, Grant/Award Number: 2015B020226004, 2015A030312013, 2017A030311034; Guangdong Natural Science Foundation, China, Grant/Award Number: 2017A030310373; Science and Technology Planning Project of Guangdong Province, China, Grant/Award Number:2017B030314027, 2017B020209004, 20169013, 2017A020215178; Science and Technology Planning Project of Guangzhou, China, Grant/Award Number: 2014Y2-00200, 2014Y2-00544, 201607010024, 201604020001, 201508020262, 201400000001-3 2017. Declaration of Interests: The authors have declared no conflict of interest. Ethics Approval Statement: The study protocol was approved by the Clinical Ethics Review Board of the Third Affiliated Hospital of Sun Yat- sen University. Informed consent was obtained according to the Declaration of Helsinki. All animal studies were approved and monitored by the animal experimental ethics committee of the Third Affiliated hospital of Sun-Yat Sen University.

Background To clarify the effects of cylcin E1 expression on HCC tumor progression, we studied the expression of cyclin E1 and inhibitory efficacy of regorafenib and sorafenib in HCC cells, and investigated a potential therapy that combines regorafenib treatment with cyclin E1 inhibition. Methods Western blotting for caspase-3 and Hoechst 33225 staining was used to measure the expression level of apoptosis-related proteins under drug treatment. Results Our results showed that enhanced expression of cyclin E1 after transfection compromised apoptosis in HCC cells induced by regorafenib or sorafenib. Conversely, down-regulation of cyclin E1 gene expression or inhibition of cyclin E1 by the cyclin-dependent kinase (CDK) inhibitors dinaciclib (DIN) or flavopiridol sensitized HCC cells to regorafenib and sorafenib by inducing apoptosis. The expression of Mcl-1, which is modulated by STAT3, plays a key role in regulating the therapeutic effects of CDK inhibitors. Xenograft experiments conducted to test the efficacy of regorafenib combined with DIN showed dramatic tumor inhibitory effects due to induction of apoptosis. Our results suggested that the level of cyclin E1 expression in HCCs may be used as a pharmacodynamic biomarker to assess the antitumor effects of regorafenib or sorafenib. Conclusions Combining regorafenib and CDK inhibitors may enhance the clinical efficiency of the treatment of HCCs. Electronic supplementary material The online version of this article (10.1186/s12964-019-0398-3) contains supplementary material, which is available to authorized users.

Background Hepatocellular carcinoma (HCC) is the most common tumors in the worldwide, it develops resistance to radiotherapy during treatment, understanding the regulatory mechanisms of radioresistance generation is the urgent need for HCC therapy. Methods qRT-PCR, western blot and immunohistochemistry were used to examine MCM3 expression. MTT assay, colony formation assay, terminal deoxynucleotidyl transferase nick end labeling assay and In vivo xenograft assay were used to determine the effect of MCM3 on radioresistance. Gene set enrichment analysis, luciferase reporter assay, western blot and qRT-PCR were used to examine the effect of MCM3 on NF-κB pathway. Results We found DNA replication initiation protein Minichromosome Maintenance 3 (MCM3) was upregulated in HCC tissues and cells, patients with high MCM3 expression had poor outcome, it was an independent prognostic factor for HCC. Cells with high MCM3 expression or MCM3 overexpression increased the radioresistance determined by MTT assay, colony formation assay, TUNEL assay and orthotopic transplantation mouse model, while cells with low MCM3 expression or MCM3 knockdown reduced the radioresistance. Mechanism analysis showed MCM3 activated NF-κB pathway, characterized by increasing the nuclear translocation of p65, the expression of the downstream genes NF-κB pathway and the phosphorylation of IKK-β and IκBα. Inhibition of NF-κB in MCM3 overexpressing cells using small molecular inhibitor reduced the radioresistance, suggesting MCM3 increased radioresistance through activating NF-κB pathway. Moreover, we found MCM3 expression positively correlated with NF-κB pathway in clinic. Conclusions Our findings revealed that MCM3 promoted radioresistance through activating NF-κB pathway, strengthening the role of MCM subunits in the tumor progression and providing a new target for HCC therapy. Electronic supplementary material The online version of this article (10.1186/s13046-019-1241-9) contains supplementary material, which is available to authorized users.

α-Mangostin, a xanthone derivative from the pericarp of mangosteen, exerts both anti-proliferative and cytotoxic effects on various cancers. However, the effect of α-mangostin on hepatocellular carcinoma (HCC) cells has not yet been fully determined. Therefore, the present study investigated the anti-proliferative effect of α-mangostin on HCC cells. α-Mangostin inhibited the proliferation of HCC and malignantly transformed hepatic cells. In addition, α-mangostin induced G0/G1 phase cell cycle arrest. Furthermore, α-mangostin also reversed doxorubicin-resistance in the HCC cells induced by IL-6. Moreover, the percentage of CD133, CD44 and EpCAM positive stem-like HCC cells treated with α-mangostin was reduced. The expression of key stemness transcription factors, such as Bmi-1, c-Myc and Nanog, were also suppressed following α-mangostin treatment. In conclusion, α-mangostin has been shown to have anti-proliferative effects in HCC cells and malignantly transformed hepatic cells. This work discovered the potential effect of α-mangostin in HCC prevention and treatment.

Background. Inflammation is the major risk factor for the progression of hepatocellular carcinoma (HCC), and the nuclear factor-κB (NF-κB) signaling plays the central role in the inflammation process. However, the activated mechanism of NF-κB signaling in HCC is unclear. Methods. The expression of PHF5A is examined by qPCR, western blotting, and immunohistochemistry (IHC) assay. The potential of PHF5A (PHD-finger domain protein 5a) for migration and invasion is examined by wound healing and Transwell assay. Luciferase reporter assay, western blotting, and qPCR were applied to explore the mechanism by which PHF5A is involved in progression of HCC. Results. PHF5A was significantly upregulated in HCC tissues and cells. Downregulation of PHF5A inhibits the migration and invasion of HCC cells. Further study demonstrated that PHF5A is implicated in HCC progression through NF-κB signaling. In addition, blocking the NF-κB signaling can weaken the stimulatory effect of PHF5A on migration and invasion of HCC cells. Conclusion. PHF5A expression is upregulated in HCC tissues, and depletion of PHF5A inhibits the migration and invasion of HCC cells. Further experiments demonstrated that PHF5A is implicated in NF-κB signaling and knockdown of PHF5A downregulates the activity of NF-κB pathway to inhibit the tumor progression. The above results provide the evidence that PHF5A plays an indispensable role in progressive effect of NF-κB pathway in HCC and may be a novel therapeutic target of HCC.

BDH2 is a short-chain dehydrogenase/reductase family member involved in several biological and pathological processes, including the utilization of cytosolic ketone bodies, immunocyte regulation and tumor progression. In this study, we first revealed that BDH2 was downregulated in HCC tissues by qRT-PCR and immunohistochemistry analysis and that low BHD2 expression was significantly associated with poor overall survival, poor tumor differentiation, increased tumor size, venous invasion and an advanced BCLC stage. Moreover, the results of a univariate analysis and multivariate analysis revealed that BDH2 may be regarded as an independent prognostic marker. As a member of a gene family involved in ketone metabolism, BDH2 upregulated the level of β-HB in liver cells as well as the level of H3 histone acetylation. Functional analysis showed that BDH2 expression inhibited tumor cell growth, proliferation and migration. The results of the mechanistic analysis revealed that BDH2 induced mitochondrial apoptosis and inhibited autophagy through the unfolded protein response. Therefore, BDH2 may be a new HCC prognostic marker and a useful treatment target.

In attempts to delay tumor progression after surgery or minimally invasive local treatments, multidisciplinary strategies have been broadly studied in patients with hepatocellular carcinoma (HCC). The objective of this present study was to evaluate the efficacy of autologous transplantations of cytokine-induced killer (CIK) cells as an adjuvant therapy for patients with HCC. A total of 264 patients with HCC were enrolled in this retrospective study. Of these patients, 165 received either CIK cell therapy alone or as adjuvant therapy to surgery, transcatheter arterial chemoembolization (TACE), or TACE-based comprehensive treatments (CT). The remaining 99 patients received only surgery or TACE. Kaplan-Meier analysis and the Chi-squared test were used to analyze the overall survival (OS), progression-free survival (PFS), and clinical characteristics of the patients in the different treatment subgroups. Kaplan-Meier analysis suggested that patients in the Surgery+CIK group had a significantly improved OS compared with those in the other three groups (P < 0.001). Furthermore, patients who developed a fever after the CIK cell treatments manifested a likely better OS (P = 0.028). Subgroup analysis indicated that patients in the Surgery+CIK group likely had an improved PFS but a similar OS compared with the patients in the Surgery-alone group (P = 0.055 for PFS, and P = 0.746 for OS). Further subgroup analysis showed that the OS in both the TACE+CIK and CT+CIK groups was prolonged significantly compared with that in the TACE-alone group (P = 0.015 and P = 0.018, respectively). However, similar OS was observed between the TACE+CIK and CT+CIK groups (P = 0.686). Autologous transplantation of CIK cells as an adjuvant therapy was associated with better survival for patients with HCC, especially for those who had also undergone TACE. A fever reaction might be a potential event for assessing the curative effect of the CIK treatment.

Dysregulation of long non-coding RNAs is a newly identified mechanism for tumour progression. Previous studies have suggested that the nuclear factor of activated T cells (NFAT) gene plays a very important role in cancer growth and metastasis. However, lncNRON is a newly identified repressor of NFAT, and its function is largely unknown, especially in hepatocellular carcinoma (HCC). Therefore, the expression levels of lncNRON in 215 pairs of HCC tissue were evaluated by qRT-PCR, and its relationship to clinicopathological parameters, recurrence, and survival was analysed. Furthermore, stably overexpressing lncNRON cell lines were constructed and evaluated for cell phenotype. Finally, we detected epithelial-to-mesenchymal transition (EMT) proteins to determine the underlying mechanism involved in lncNRON function. We observed that lncNRON was downregulated in HCC tumour tissues; low lncNRON expression was associated with poor tumour differentiation and the presence of vascular tumour thrombus, which tended to result in poor clinical outcomes, as demonstrated by the recurrence rate and survival curves. Functional analysis showed that lncNRON overexpression impaired colony formation and cell viability and inhibited cell migration and invasion. A study using tumour-bearing mice showed that lncNRON markedly limited tumour growth and lung metastasis in vivo. Importantly, western blot analysis revealed that the expression of the EMT-related epithelial marker, E-cadherin, increased, whereas the expression of mesenchymal markers N-cadherin, snail, and vimentin was attenuated by lncNRON overexpression in HCC cells. Therefore, lower lncNRON expression indicates a poorer clinical outcome in HCC. LncNRON overexpression can suppress HCC growth and metastasis via inhibiting the EMT, and lncNRON may function as a new HCC prognostic marker.

Mature microRNA (miRNA) 34a-5p, which is a well-known tumor suppressor in hepatitis virus-associated hepatocellular carcinoma (HCC), plays an important role in cell processes, such as cell proliferation and apoptosis, and is therefore an optimal biomarker for future clinical use. However, the role of miRNA-34a-5p in chemoresistance has yet to be identified. In the present study, the expression of miRNA-34a-5p was assessed by an in situ hybridization assay in HCC tissues and was found to be significantly decreased compared with the pericarcinomatous areas of the tissue specimens, which consisted of samples obtained from 114 patients with HCC. High expression of miRNA-34a-5p was found to be associated with a favorable overall survival time in HCC patients. Functional tests performed by transfecting miRNA-34a-5p mimics or inhibitors into MHCC-97L cells illustrated that miRNA-34a-5p inhibited proliferation, elevated apoptosis and decreased chemoresistance to cisplatin in HCC cells. AXL is the direct target of miRNA-34a-5p, as confirmed by sequence analysis and luciferase assay. Transfection of the cells with small interfering RNA for AXL (siAXL) increased the apoptosis ratio of the MHCC-97L cell line. Transfection with siAXL led to similar biological behaviors in the MHCC-97L cells to those induced by ectopic expression of miRNA-34a-5p. Thus, it was concluded that miRNA-34a-5p enhanced the sensitivity of the cells to chemotherapy by targeting AXL in hepatocellular carcinoma. In addition, low expression of miRNA-34a-5p in HCC tissues yielded an unfavorable prognosis for patients with HCC that received radical surgery, due to the promotion of proliferation and an increase in chemoresistance in HCC cells.

Hepatocellular carcinoma (HCC) is the most common tumors in the worldwide, it develops resistance to radiotherapy during treatment, understanding the regulatory mechanisms of radioresistance generation is the urgent need for HCC therapy.qRT-PCR, western blot and immunohistochemistry were used to examine MCM3 expression. MTT assay, colony formation assay, terminal deoxynucleotidyl transferase nick end labeling assay and In vivo xenograft assay were used to determine the effect of MCM3 on radioresistance. Gene set enrichment analysis, luciferase reporter assay, western blot and qRT-PCR were used to examine the effect of MCM3 on NF-κB pathway.We found DNA replication initiation protein Minichromosome Maintenance 3 (MCM3) was upregulated in HCC tissues and cells, patients with high MCM3 expression had poor outcome, it was an independent prognostic factor for HCC. Cells with high MCM3 expression or MCM3 overexpression increased the radioresistance determined by MTT assay, colony formation assay, TUNEL assay and orthotopic transplantation mouse model, while cells with low MCM3 expression or MCM3 knockdown reduced the radioresistance. Mechanism analysis showed MCM3 activated NF-κB pathway, characterized by increasing the nuclear translocation of p65, the expression of the downstream genes NF-κB pathway and the phosphorylation of IKK-β and IκBα. Inhibition of NF-κB in MCM3 overexpressing cells using small molecular inhibitor reduced the radioresistance, suggesting MCM3 increased radioresistance through activating NF-κB pathway. Moreover, we found MCM3 expression positively correlated with NF-κB pathway in clinic.Our findings revealed that MCM3 promoted radioresistance through activating NF-κB pathway, strengthening the role of MCM subunits in the tumor progression and providing a new target for HCC therapy.

Zinc finger protein 687 (ZNF687), identified as a C2H2 zinc finger protein, has been found to be mutated and upregulated in giant cell tumor of bone and acute myeloid leukemia, suggesting an oncogenic role for ZNF687 in cancer. However, the clinical significance and precise role of ZNF687 in cancer progression are largely unknown. Herein, we report that ZNF687 was markedly upregulated in hepatocellular carcinoma (HCC) cell lines and HCC tissues, and was significantly correlated with relapse-free survival in HCC. ZNF687 overexpression greatly enhanced HCC cell capability for tumorsphere formation, invasion and chemoresistance in vitro, whereas inhibiting ZNF687 reduced these capabilities and inhibited HCC cell tumorigenic capability in vivo. Importantly, extreme limiting dilution analysis revealed that even 1 × 102 ZNF687-transduced cells could form tumors in vivo, indicating that ZNF687 contributes to HCC recurrence. Moreover, we demonstrate that ZNF687 transcriptionally upregulated the expression of the pluripotency-associated factors BMI1, OCT4 and NANOG by directly targeting their promoters. Therefore, our results suggest that ZNF687 has a promoter role in regulating HCC progression, which provides a potential therapeutic target for HCC in humans.

With the expansion of surgical criteria, the comparative efficacy between surgical resection (SR) and liver transplantation (LT) for hepatocellular carcinoma is inconclusive. This study aimed to develop a prognostic nomogram for predicting recurrence-free survival of hepatocellular carcinoma patients after resection and explored the possibility of using nomogram as treatment algorithm reference.From 2003 to 2012, 310 hepatocellular carcinoma patients within Hangzhou criteria undergoing resection or liver transplantation were included. Total tumor volume, albumin level, HBV DNA copies and portal hypertension were included for constructing the nomogram. The resection patients were stratified into low- and high-risk groups by the median nomogram score of 116. Independent risk factors were identified and a visually orientated nomogram was constructed using a Cox proportional hazards model to predict the recurrence risk for SR patients.The low-risk SR group had better outcomes compared with the high-risk SR group (3-year recurrence-free survival rate, 71.1% vs 35.9%; 3-year overall survival rate, 89.8% vs 78.9%, both P0.001). The high-risk SR group was associated with a worse recurrence-free survival rate but similar overall survival rate compared with the transplantation group (3-year recurrence-free survival rate, 35.9% vs 74.1%, P0.001; 3-year overall survival rate, 78.9% vs 79.6%, P0.05).This nomogram offers individualized recurrence risk evaluation for hepatocellular carcinoma patients within Hangzhou criteria receiving resection. Transplantation should be considered the first-line treatment for high-risk patients.

// Xiu-Rong Cai 1, 2, * , Xing Li 1, 2, * , Jin-Xiang Lin 1 , Tian-Tian Wang 1, 2 , Min Dong 1, 2 , Zhan-Hong Chen 1, 2 , Chang-Chang Jia 2, 3 , Ying-Fen Hong 1, 2 , Qu Lin 1, 2 and Xiang-Yuan Wu 1, 2 1 Department of Medical Oncology, the Third Affiliated Hospital of Sun Yat-sen University, Guangzhou 510630, People’s Republic of China 2 Guangdong provincial Key Laboratory of Liver Disease Research, Guangzhou 510630, People’s Republic of China 3 Cell-gene Therapy Translational Medicine Research Center, the Third Affiliated Hospital of Sun Yat-sen University, Guangzhou 510630, People’s Republic of China * These authors have contributed equally to this work Correspondence to: Xiang-Yuan Wu, email: wuxiangy@mail.sysu.edu.cn Qu Lin, email: linqu@mail.sysu.edu.cn Keywords: cytokine induced killer cell, liver cancer, adoptive cells therapy, immunotherapy, meta-analysis Received: June 13, 2016 Accepted: November 12, 2016 Published: February 17, 2017 ABSTRACT Background: High recurrence rate after curative treatment is the major problem for hepatocellular carcinoma (HCC). Cytokine-induced killer cells (CIKs) therapy was extensively studied among HCC patients. However, the value of CIKs therapy was controversial. A meta-analysis was performed to investigate the efficacy of adjuvant CIKs after invasive treatments among HCC patients. Methods: We searched online for literatures studying sequential CIKs therapy for HCC patients. Recurrence-free survival (RFS), progress-free survival (PFS) and overall survival (OS) were set as the main endpoints. Both overall and subgroup analysis were accomplished. Results: A total of 12 clinical trials with 1,387 patients were included. The pooled analysis showed a significant improvement of RFS, PFS and OS in CIK group (HR 0.56, 95% CI 0.47-0.67, p

Defects in natural killer (NK) cell function are necessary for tumor immune escape, but the underlying regulatory mechanisms in human cancers remain largely unknown. Here we show that fibroblasts derived from hepatocellular carcinoma (HCC) were significantly superior to foreskin-derived fibroblasts at inducing NK cell dysfunction, which is characterized by low expression of cytotoxic molecules and surface markers for cell activation, impaired production of cytokines, and decreased cytotoxicity against K562 cells in vitro. Our results also indicate that PGE2 and IDO, derived from activated fibroblasts, suppress the activation of NK cells and thereby create favorable conditions for tumor progression.

Prognosis of patients with advanced hepatocellular carcinoma (HCC) is under expectation. Life expectancy more than 3 months is one inclusion criteria for molecular targeted drugs in clinical trials. The main purpose of this research is to compare Model for End-Stage Liver Disease (MELD) and four MELD-based prognostic models in predicting the survival rate of advanced HCC patients. One hundred eighty-three patients with advanced HCC who were not amendable to standard anti-tumor therapy were retrospectively analyzed. Data were collected to classify patients according to MELD, Model for End-Stage Liver Disease with the incorporation of serum sodium (MELD-NA), Model for End-Stage Liver Disease to ascites and sodium (MELD-AS), integrated Model for End-Stage Liver Disease (iMELD), and Model for End-Stage Liver Disease to sodium (MESO) scores at diagnosis. 1-, 3-, and 6-month survivals were the end points used in the analysis. When predicting 1-month survival, MELD-AS, MELD, and MESO were the top 3 ranking staging systems. When predicting 3-month survival, area under the receiver operating characteristic curve (AUC) of MELD-AS is significantly higher than that of the other models (P

// Jia-Jie Chen 1,3,4,* , Nan Cai 1,3,* , Guan-Zhong Chen 1,2,* , Chang-Chang Jia 1,3 , Dong-Bo Qiu 2,3 , Cong Du 2,3 , Wei Liu 1,2 , Yang Yang 1 , Zi-Jie Long 4 and Qi Zhang 1,2,3 1 Organ Transplantation Center, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, People’s Republic of China 2 Guangdong Provincial Key Laboratory of Liver Disease Research, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, People’s Republic of China 3 Vaccine Research Institute of Sun Yat-Sen University, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, People’s Republic of China 4 Department of Hematology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, People’s Republic of China * These authors have contributed equally to this work Correspondence to: Qi Zhang, email: // Zi-Jie Long, email: // Keywords : pimozide, hepatic cancer stem-like cells, self-renewal, STAT3 signaling, STAT3 inhibitor Received : February 11, 2015 Accepted : May 13, 2015 Published : May 27, 2015 Abstract Drug repurposing is currently an important approach for accelerating drug discovery and development for clinical use. Hepatocellular carcinoma (HCC) presents drug resistance to chemotherapy, and the prognosis is poor due to the existence of liver cancer stem-like cells. In this study, we investigated the effect of the neuroleptic agent pimozide to inhibit stem-like cell maintenance and tumorigenicity in HCC. Our results showed that pimozide functioned as an anti-cancer drug in HCC cells or stem-like cells. Pimozide inhibited cell proliferation and sphere formation capacities in HCC cells by inducing G0/G1 phase cell cycle arrest, as well as inhibited HCC cell migration. Surprisingly, pimozide inhibited the maintenance and tumorigenicity of HCC stem-like cells, particularly the side population (SP) or CD133-positive cells, as evaluated by colony formation, sphere formation and transwell migration assays. Furthermore, pimozide was found to suppress STAT3 activity in HCC cells by attenuating STAT3-dependent luciferase activity and down-regulating the transcription levels of downstream genes of STAT3 signaling. Moreover, pimozide reversed the stem-like cell tumorigenic phenotypes induced by IL-6 treatment in HCC cells. Further, the antitumor effect of pimozide was also proved in the nude mice HCC xenograft model. In short, the anti-psychotic agent pimozide may act as a novel potential anti-tumor agent in treating advanced HCC.

Substantial evidence indicates that cancer-associated fibroblasts (CAFs) are critical components in the process of cancer progression. However, the role of CAFs in the immunopathogenesis of human cancer remains elusive. In this study, we demonstrate that purified colorectal carcinoma-derived fibroblasts exhibit activated phenotypes characterized by substantial α-smooth muscle actin expression. These CAFs sharply suppress natural killer (NK) cell functions in co-culture experiments. In contrast, normal skin fibroblasts had only a minimal effect on NK cell phenotype and function. Moreover, we demonstrated that prostaglandin E2 (PGE2) was released by fibroblasts in co-culture experiments. Thus, the functional modulation of NK cells by CAFs may represent a novel mechanism linking the pro-inflammatory response to immune tolerance within the tumor milieu.

e15615Background: Prognosis of patients with advanced HCC(aHCC) is very poor even though they receive loco-regional therapy. We have established a new prognostic system called SYSU system (variable...

e15606Background: Cancer-associated fibroblasts (CAFs) reportedly regulate tumor progression by inducing tumor-stromal interaction; however, the roles of CAFs in hepatocellular carcinoma (HCC) deve...