Loss of tumor suppressor IGFBP4 drives epigenetic reprogramming in hepatic carcinogenesis.

Genomic sequencing of hepatocellular carcinoma (HCC) uncovers a paucity of actionable mutations, underscoring the necessity to exploit epigenetic vulnerabilities for therapeutics. In HCC, EZH2-mediated H3K27me3 represents a major oncogenic chromatin modification, but how it modulates the therapeutic vulnerability of signaling pathways remains unknown. Here, we show EZH2 acts antagonistically to AKT signaling in maintaining H3K27 methylome through epigenetic silencing of IGFBP4. ChIP-seq revealed enrichment of Ezh2/H3K27me3 at silenced loci in HBx-transgenic mouse-derived HCCs, including Igfbp4 whose down-regulation significantly correlated with EZH2 overexpression and poor survivals of HCC patients. Functional characterizations demonstrated potent growth- and invasion-suppressive functions of IGFBP4, which was associated with transcriptomic alterations leading to deregulation of multiple signaling pathways. Mechanistically, IGFBP4 stimulated AKT/EZH2 phosphorylation to abrogate H3K27me3-mediated silencing, forming a reciprocal feedback loop that suppressed core transcription factor networks (FOXA1/HNF1A/HNF4A/KLF9/NR1H4) for normal liver homeostasis. Consequently, the in vivo tumorigenicity of IGFBP4-silenced HCC cells was vulnerable to pharmacological inhibition of EZH2, but not AKT. Our study unveils chromatin regulation of a novel liver tumor suppressor IGFBP4, which constitutes an AKT-EZH2 reciprocal loop in driving H3K27me3-mediated epigenetic reprogramming. Defining the aberrant chromatin landscape of HCC sheds light into the mechanistic basis of effective EZH2-targeted inhibition.

Percutaneous Portal Vein Embolization Using a Simplified Sheathless 18-Gauge Trocar Needle Approach: Review of Efficacy and Safety.

PURPOSE: Portal vein (PV) embolization (PVE) is traditionally performed via a PV sheath with selective embolization of PV branches. Here, the efficacy and safety of PVE with the use of only an 18-gauge needle is reported. MATERIALS AND METHODS: Consecutive patients who underwent PVE from 2009 through 2017 were retrospectively reviewed. Forty-five patients (mean age, 60 y ± 7.6; 38 men) underwent 45 PVE procedures. Hepatocellular carcinoma, cholangiocarcinoma, and metastases accounted for 26 (58%), 13 (29%), and 6 (13%) patients, respectively. PVE was performed by puncturing a branch of right PV with an 18-gauge needle under US guidance. Via the same needle, direct portography was performed, followed by PVE with an N-butyl cyanoacrylate/Lipiodol mixture. Percentage increase of future liver remnant (FLR) volume and increase in ratio of FLR to total liver volume were estimated as measures of efficacy. Complications were reported according to Society of Interventional Radiology classification. Fluoroscopy time, procedure time, and dose-area product (DAP) were recorded. RESULTS: Technical success rate was 100%. The median DAP, fluoroscopy time, and procedure time were 74,387 mGy·cm2 (IQR, 90,349 mGy·cm2), 3.5 min (IQR, 2.10 min), and 24 min (IQR, 10.5 min). Among the 23 patients with complete CT volumetry data, mean increase in the ratio of FLR to total liver volume and percentage increase of FLR volume were 12.5% ± 7.7 and 50% ± 33, respectively. There were 3 minor complications (asymptomatic nonocclusive emboli in FLR) and 3 major complications (1 hepatic vein emboli, 1 subphrenic collection, and 1 hepatic infarct). CONCLUSIONS: PVE via a sheathless 18-gauge needle approach is feasible, with satisfactory FLR hypertrophy.

The CCCTC-binding factor (CTCF)-forkhead box protein M1 axis regulates tumour growth and metastasis in hepatocellular carcinoma.

CCCTC-binding factor (CTCF) is a DNA-binding protein that interacts with a large number of highly divergent target sequences throughout the genome. It is implicated in a variety of functions, including chromatin organization and transcriptional control. The functional role of CTCF in tumour pathogenesis remains elusive. We showed that CTCF is frequently upregulated in a subset of primary hepatocellular carcinomas (HCCs) as compared with non-tumoural liver. Overexpression of CTCF was associated with shorter disease-free survival of patients. Short hairpin RNA (shRNA)-mediated suppression of CTCF inhibited cell proliferation, motility and invasiveness in HCC cell lines; these effects were correlated with prominent reductions in the expression of telomerase reverse transcriptase (TERT), the shelterin complex member telomerase repeat-binding factor 1, and forkhead box protein M1 (FOXM1). In contrast, upregulation of CTCF was positively correlated with FOXM1 and TERT expression in clinical HCC biopsies. Depletion of CTCF resulted in reduced motility and invasiveness in HCC cells that could be reversed by ectopic expression of FOXM1, suggesting that FOXM1 is one of the important downstream effectors of CTCF in HCC. Reporter gene analysis suggested that depletion of CTCF is associated with reduced FOXM1 and TERT promoter activity. Chromatin immunoprecipitation (ChIP)-polymerase chain reaction (PCR) analysis further revealed occupancy of the FOXM1 promoter by CTCF in vivo. Importantly, depletion of CTCF by shRNA significantly inhibited tumour progression and metastasis in HCC mouse models. Our work uncovered a novel functional role of CTCF in HCC pathogenesis, which suggests that targeting CTCF could be further explored as a potential therapeutic strategy for HCC. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Novel biomarkers GEP/ABCB5 regulate response to adjuvant transarterial chemoembolization after curative hepatectomy for hepatocellular carcinoma.

BACKGROUND: Transarterial chemoembolization (TACE) is the most commonly used adjuvant therapy for hepatocellular carcinoma (HCC) after curative resection. Responses to TACE are variable due to tumor and patient heterogeneity. We had previously demonstrated that expression of Granulin-epithelin precursor (GEP) and ATP-dependent binding cassette (ABC)B5 in liver cancer stem cells was associated with chemoresistance. The present study aimed to evaluate the association between GEP/ABCB5 expression and response to adjuvant TACE after curative resection for HCC. METHODS: Patients received adjuvant TACE after curative resection for HCC and patients received curative resection alone were identified from a prospectively collected database. Clinical samples were retrieved for biomarker analysis. Patients were categorized into 3 risk groups according to their GEP/ABCB5 status for survival analysis: low (GEP-/ABCB5-), intermediate (either GEP+/ABCB5- or GEP-/ABCB5+) and high (GEP+/ABCB5+). Early recurrence (recurrence within 2 years after resection) and disease-free survival were analyzed. RESULTS: Clinical samples from 44 patients who had followed-up for more than 2 years were retrieved for further biomarker analysis. Among them, 18 received adjuvant TACE and 26 received surgery alone. Patients with adjuvant TACE in the intermediate risk group was associated with significantly better overall survival and 2-year disease-free survival than those who had surgery alone (P = 0.036 and P = 0.011, respectively). Adjuvant TACE did not offer any significant differences in the early recurrence rate, 2-year disease-free survival and overall survival for patients in low and high risk groups. CONCLUSIONS: Adjuvant TACE can only provide survival benefits for patients in the intermediate risk group (either GEP+/ABCB5- or GEP-/ABCB5+). A larger clinical study is warranted to confirm its role in patient selection for adjuvant TACE.

Liver stiffness measurement predicts high-grade post-hepatectomy liver failure: A prospective cohort study.

BACKGROUND AND AIM: Liver stiffness measurement using transient elastography appears to be an excellent tool for detection of liver fibrosis and cirrhosis with high accuracy. The aim of this study is to evaluate the efficacy of preoperative liver stiffness measurement in predicting post-hepatectomy liver failure. METHODS: A prospective cohort study of all consecutive patients undergoing hepatectomy for hepatocellular carcinoma from February 2010 to August 2014 was studied. All patients received detailed preoperative assessments including liver stiffness measurement. The primary outcome was post-hepatectomy liver failure according to the International Study Group of Liver Surgery definition. RESULTS: A total of 255 patients were included. Liver stiffness measurement showed significant correlation with grade B or C post-hepatectomy liver failure. (P = 0.003) Using the cutoff at 12 kPa, liver stiffness measurement had a sensitivity of 52.4% and specificity of 73.3% in predication of high-grade (grade B or C) post-hepatectomy liver failure. Liver stiffness measurement > 12 kPa was also an independent prognostic factor for both high-grade post-hepatectomy liver failure and major postoperative complications by multivariate analysis. The diagnostic accuracy was better in patients without right lobe tumor with an area under the receiver operating characteristic of 0.83 compared with an area under the receiver operating characteristic of only 0.62 in patients with right lobe tumor. CONCLUSIONS: Liver stiffness measurement using Fibroscan is good to predict high-grade post-hepatectomy liver failure especially in patients without right lobe tumor.

A CCRK-EZH2 epigenetic circuitry drives hepatocarcinogenesis and associates with tumor recurrence and poor survival of patients.

BACKGROUND & AIMS: Aberrant chromatin modification is a key feature of hepatocellular carcinoma (HCC), which is characterized by strong sexual dimorphism. Both enhancer of zeste homolog 2 (EZH2) and cell cycle-related kinase (CCRK) contribute to hepatocarcinogenesis, yet whether the two oncogenic factors have functional crosstalk is unknown. METHODS: Cellular proliferation and tumorigenicity upon transgenic expression and RNA interference were determined by colony formation and soft agar assays, xenograft, orthotopic and diethylnitrosamine-induced HCC models. Gene regulation was assessed by chromatin immunoprecipitation, site-directed mutagenesis, luciferase reporter, co-immunoprecipitation and expression analyses. Protein levels in clinical specimens were correlated with clinicopathological parameters and patient survival rates. RESULTS: Ectopic CCRK expression in immortalized human liver cells increased EZH2 and histone H3 lysine 27 trimethylation (H3K27me3) to stimulate proliferation and tumor formation. Conversely, knockdown of CCRK reduced EZH2/H3K27me3 levels and decreased HCC cell growth, which could be rescued by EZH2 over-expression. Mechanistically, GSK-3ß phosphorylation by CCRK activated a ß-catenin/TCF/E2F1/EZH2 transcriptional feedback loop to epigenetically enhance androgen receptor (AR) signaling. Simultaneously, the phosphorylation of AKT/EZH2 by CCRK facilitated the co-occupancy of CCRK promoter by EZH2-AR and its subsequent transcriptional activation, thus forming a self-reinforcing circuitry. Lentiviral-mediated knockdown of CCRK, which abrogated the phosphorylation-transcriptional network, prevented diethylnitrosamine-induced tumorigenicity. More importantly, the hyperactivation of the CCRK-EZH2 circuitry in human HCCs correlated with tumor recurrence and poor survival. CONCLUSIONS: These findings uncover an epigenetic vicious cycle in hepatocarcinogenesis that operates through reciprocal regulation of CCRK and EZH2, providing novel therapeutic strategy for HCC.

Cell cycle-related kinase mediates viral-host signalling to promote hepatitis B virus-associated hepatocarcinogenesis.

BACKGROUND: Androgen receptor (AR) signalling contributes to male predominance in hepatocellular carcinoma (HCC), which is more pronounced in HBV-endemic areas. Cell cycle-related kinase (CCRK) is essential for AR-induced hepatocarcinogenesis but its molecular function in HBV-associated HCC remains obscure. OBJECTIVE: To determine the molecular function of CCRK in HBV-associated HCC. DESIGN: Transcriptional regulation was assessed by chromatin immunoprecipitation, promoter mutation and luciferase reporter assays. Hepatocellular proliferation and tumourigenesis were examined by colony formation, soft agar assays and using HBV X protein (HBx) transgenic mice with low-dose exposure to diethylnitrosamine. Protein expressions were examined in clinical samples and correlated with patient survival by log-rank Mantel-Cox test. RESULTS: Overexpression of CCRK, but not its kinase-defective mutant, activated ß-catenin/T cell factor signalling through phosphorylation of glycogen synthase kinase-3ß (GSK-3ß) at Ser9, led to upregulation of AR transcriptional activity and, subsequently, expression of HBx. The viral transactivator in turn induced CCRK expression through enhanced AR signalling, thus forming a positive regulatory loop. RNA interference silencing of CCRK, which suppressed the CCRK/GSK-3ß/ß-catenin/AR regulatory loop, significantly suppressed HBx-induced hepatocellular proliferation (p=0.001) and transformation (p<0.001) and remarkably reduced >80% diethylnitrosamine-mediated hepatocarcinogenesis in HBx transgenic mice. Finally, patients with HBV-associated HCC with concordant overexpression of CCRK, GSK-3ß phosphorylation at Ser9, active dephosphorylated ß-catenin and AR phosphorylation at Ser81 had poorer overall (HR=31.26, p<0.0001) and disease-free (HR=3.60, p<0.01) survival rates. CONCLUSIONS: Our findings highlight the critical role of CCRK in a self-reinforcing circuitry that regulates HBV-associated hepatocarcinogenesis. Further characterisation of this intricate viral-host signalling may provide new prognostic biomarkers and therapeutic targets for HCC treatment.

SIRT2 overexpression in hepatocellular carcinoma mediates epithelial to mesenchymal transition by protein kinase B/glycogen synthase kinase-3ß/ß-catenin signaling.

UNLABELLED: Sirtuin 1 (SIRT1) has been implicated in telomere maintenance and the growth of hepatocellular carcinoma (HCC). Nevertheless, the role of other sirtuins in the pathogenesis of HCC remains elusive. We found that sirtuin 2 (SIRT2), another member of the sirtuin family, also contributes to cell motility and invasiveness of HCC. SIRT2 is up-regulated in HCC cell lines and in a subset of human HCC tissues (23/45). Up-regulations of SIRT2 in primary HCC tumors were significantly correlated with the presence of microscopic vascular invasion (P = 0.001), a more advanced tumor stage (P = 0.004), and shorter overall survival (P = 0.0499). Functional studies by short hairpin RNA-mediated suppression of SIRT2 expression in HCC cell lines revealed significant inhibition of motility and invasiveness. Depletion of SIRT2 also led to the regression of epithelial-mesenchymal transition (EMT) phenotypes, whereas the ectopic expression of SIRT2 in the immortalized hepatocyte cell line L02 promoted cell motility and invasiveness. Mechanistic studies revealed that SIRT2 regulates the deacetylation and activation of protein kinase B, which subsequently impinges on the glycogen synthase kinase-3ß/ß-catenin signaling pathway to regulate EMT. CONCLUSIONS: Our findings have uncovered a novel role for SIRT2 in HCC metastasis, and provide a rationale to explore the use of sirtuin inhibitors in HCC therapy. (HEPATOLOGY 2013;).

Liver stiffness measurement by transient elastography as a predictor on posthepatectomy outcomes.

BACKGROUND: Liver fibrosis and cirrhosis are well-known risk factors for morbidity after hepatectomy. Liver stiffness measurement (LSM) using transient elastography is a new method for detection of hepatic fibrosis and cirrhosis with high accuracy. Whether LSM can predict posthepatectomy outcomes has not been studied. METHODS: This was a prospective cohort study in which consecutive patients underwent hepatectomy for various indications from February 2010 to July 2011. All patients received detailed preoperative assessments including LSM and indocyanine green (ICG) clearance test. The primary outcome was major postoperative complication. RESULTS: One hundred five patients with a mean age of 59 years were included; 75 (71.4%) had chronic viral hepatitis and 76 (72.4%) had hepatocellular carcinoma. Thirty-four patients (32.4%) received major hepatectomy. The median ICG retention rate at 15 minutes was 4.2 (0.1%-32%) and the median LSM was 9.4 (3.3-75 kPa). For posthepatectomy outcomes, only LSM but not ICG showed significant correlation with major postoperative complications on receiver operating characteristic curves, with area under the curve of 0.79 (P < 0.001). Using the calculated cutoff at 12.0 kPa, LSM had sensitivity of 85.7% and specificity of 71.8% in the prediction of major postoperative complications. It was also an independent prognostic factor for major postoperative complications by multivariate analysis. The operative blood loss and transfusion rate were also significantly higher in patients with LSM >12.0 kPa. CONCLUSIONS: High LSM (>12.0 kPa) predicted worse posthepatectomy outcomes. Preoperative LSM was better than ICG test in the prediction of major postoperative complications. It was a useful preoperative investigation for risk stratification before hepatectomy.

Percutaneous radiofrequency ablation versus surgical radiofrequency ablation for malignant liver tumours: the long-term results.

BACKGROUND: Radiofrequency ablation (RFA) has been used to treat hepatocellular carcinoma (HCC) and liver metastases for more than 10 years with promising early outcomes. Preliminary results comparing percutaneous and surgical approaches have shown no difference in short-term outcomes. In this study, the longer-term outcomes were presented. METHODS: Patients with liver malignancies treated by RFA were prospectively studied from 2003 to 2011. Post-ablation assessment by computed tomography (CT) scan and serum biochemistry was performed at regular intervals. Recurrence rates and long-term survival were analysed. RESULTS: A total of 233 patients with liver malignancies (75.5% HCC and 24.5% liver metastases) were analysed. Three RFA approaches were used (percutaneous 58.4%, laparoscopic 9.4% and open 32.2%). The median follow-up time was 29 months. Complete ablation was achieved in 83.7%, with no difference between the two approaches. More wound and chest complications were observed in the surgical group. Intra-hepatic recurrences were observed in 69.5%; extra-hepatic recurrences were detected in 22.3%, with no difference between the two groups. There was no statistical difference between the two approaches in overall 1-, 3- and 5-year survival. CONCLUSION: An extended period of follow-up in patients with liver malignancies showed that RFA is an effective treatment. No difference was demonstrated between the percutaneous and surgical approach, in terms of recurrence and survival.

Long-term results of laparoscopic hepatectomy versus open hepatectomy for hepatocellular carcinoma: a case-matched analysis.

BACKGROUND: Laparoscopic hepatectomy (LH) is established as a safe and feasible treatment option for liver tumors. However, whether the adoption of laparoscopic approach for malignant tumors, such as hepatocellular carcinoma (HCC), will compromise the long-term result is still unknown. This study was designed to evaluate the long-term results of LH compared with a cohort of case-matched open hepatectomy (OH). METHODS: Thirty-three patients who underwent LH for HCC in our institution between June 2004 and March 2010 were recruited. A group of 50 patients who underwent OH for HCC during the same period was identified by matching to magnitude of operation, size of tumor, site of tumor, and the absence of concomitant local ablation or major procedure. The perioperative outcomes, disease recurrence, and survival of the two groups of patients were determined and compared. RESULTS: LH resulted in less operative complications (6.1% vs. 24%, P = 0.033) and shorter median hospital stay (5 vs. 7 days, P < 0.0005) but required longer operative time compared with OH (225 vs. 195 min, P = 0.019). There was no difference between LH and OH in recurrence rate (45.5% vs. 38%, P = 0.499). The 1-, 3-, and 5-year overall survival were 86.9%, 81.8%, and 76% for LH and 98%, 80.6%, and 76.1% for OH respectively (P = 0.646). The 1-, 3-, and 5-year disease-free survival were 78.8%, 51%, and 45.3% for LH and 69.2%, 55.9%, and 55.9% for OH, respectively (P = 0.849). CONCLUSIONS: Compared with OH, LH for HCC has similar long-term outcomes, but it has short-term advantages of less operative complications and shorter hospital stay.

Modification of right hepatectomy results in improvement outcome: a retrospective comparative study.

OBJECTIVE: To evaluate any change in the operative and survival outcomes in patients undergoing a right hepatectomy after adoption of the no-clamp technique using a radiofrequency dissecting sealer (TissueLink™) in liver resection. METHODS: In all, 58 consecutive patients who underwent a right hepatectomy from July 2003 to December 2007 (Group 1) were compared with 66 consecutive patients who underwent a right hepatectomy from January 1999 to June 2003 (Group 2). In group 1, a liver transection was performed with a cavitron ultrasonic surgical aspirator (CUSA) and TissueLink™ without hilar clamping whereas in group 2, a liver transection was performed with CUSA and diathermy with routine continuous hilar clamping. RESULTS: For the operative outcomes, there was significantly less blood loss (median 450 vs. 900 ml, P < 0.001) in group 1. The complication rate was also significantly lower in group 1 (22.4% vs. 47.0%, P = 0.004). In subgroup analysis for patients with hepatocellular carcinoma (HCC), the overall survival rate was significantly better in group 1; 1-, 3- and 5-year survival rates were 78%, 72% and 57% in group 1 vs. 72%, 44% and 39% in group 2, respectively (P = 0.048). CONCLUSIONS: When compared with the retrospective cohort, a right hepatectomy utilizing TissueLink™ without hilar clamping was feasible with potential benefits in surgical outcomes.

Resection margin in laparoscopic hepatectomy: a comparative study between wedge resection and anatomic left lateral sectionectomy.

BACKGROUND: Experience from open hepatectomy shows that anatomic liver resection achieves a better resection margin than wedge resection. In recent years, laparoscopic hepatectomy has increasingly been performed in patients with liver pathology including malignant lesions. Wedge resection (WR) and left lateral sectionectomy (LLS), which also represent non-anatomic and anatomic resection respectively, are the two most common types of laparoscopic hepatectomy performed. The aim of the present study was to compare the two types of laparoscopic hepatectomy with emphasis on resection margin. METHODS: Between November 2003 and July 2009, 44 consecutive patients who underwent laparoscopic hepatectomy were identified and retrospectively reviewed. The WR and LLS group of patients were compared in terms of operative outcomes, pathological findings, recurrence patterns and survival. RESULTS: Out of the 44 patients, 21 underwent LLS and 23 a WR. The two groups of patients were comparable in demographics. The two groups did not differ in conversion rate, blood loss, blood transfusion, mortality, morbidity and post-operative length of stay. The LLS group patients had significantly larger liver lesions, wider resection margin and less sub-centimetre margins. In patients with malignant liver lesions, there was no difference between the two groups in incidence of intra-hepatic recurrence and 3-year overall and disease-free survival. CONCLUSION: Operative outcomes are similar between laparoscopic WR and LLS. However, WR is less reliable than LLS in achieving a resection margin of more than 1 cm. Larger studies involving more patients with longer follow-up are warranted to determine the impact of the resection margin on intra-hepatic recurrence and survival.

Successful robotic extirpation of diaphragmatic seeding of hepatocellular carcinoma after previous rupture.

A 51-year-old man who was a hepatitis B carrier presented with ruptured hepatocellular carcinoma (HCC). Hepatic arterial embolization was performed for control of bleeding which was followed by staged open left lateral sectionectomy for tumor removal. Pathology confirmed a 3.5 cm ruptured subcapsular HCC in a cirrhotic liver with clear resection margin. However, the alpha-fetoprotein (AFP) increased from 14 to 72 µg/L after 7 months. A 1.7 × 0.8 cm nodule at left subdiaphragmatic region abutting on the spleen but no intrahepatic lesion was seen on computed tomography (CT). Dual tracer positron emission tomography suggested the nodule was a HCC seeding with no other recurrent tumor noted. Robotic exploration was offered to patient with the possibility of splenectomy. During operation, the nodule was adherent to the diaphragm with no splenic involvement. The lesion was locally excised. The diaphragmatic defect was closed with non-absorbable suture. Recovery was uneventful and the patient was discharged on postoperative day 4. Pathology confirmed HCC cells infiltrating to skeletal muscle and fibrous tissue. The resection margin was clear. Post-operatively AFP normalised. Serial abdominal CT and ultrasound revealed no evidence of recurrent disease. Patient had a disease-free survival of 47 months after excision of tumor seeding.

ER-residential Nogo-B accelerates NAFLD-associated HCC mediated by metabolic reprogramming of oxLDL lipophagy.

Non-alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome that elevates the risk of hepatocellular carcinoma (HCC). Although alteration of lipid metabolism has been increasingly recognized as a hallmark of cancer cells, the deregulated metabolic modulation of HCC cells in the NAFLD progression remains obscure. Here, we discovers an endoplasmic reticulum-residential protein, Nogo-B, as a highly expressed metabolic modulator in both murine and human NAFLD-associated HCCs, which accelerates high-fat, high-carbohydrate diet-induced metabolic dysfunction and tumorigenicity. Mechanistically, CD36-mediated oxLDL uptake triggers CEBPß expression to directly upregulate Nogo-B, which interacts with ATG5 to promote lipophagy leading to lysophosphatidic acid-enhanced YAP oncogenic activity. This CD36-Nogo-B-YAP pathway consequently reprograms oxLDL metabolism and induces carcinogenetic signaling for NAFLD-associated HCCs. Targeting the Nogo-B pathway may represent a therapeutic strategy for HCC arising from the metabolic syndrome.

Aberrant enhancer hypomethylation contributes to hepatic carcinogenesis through global transcriptional reprogramming.

Hepatocellular carcinomas (HCC) exhibit distinct promoter hypermethylation patterns, but the epigenetic regulation and function of transcriptional enhancers remain unclear. Here, our affinity- and bisulfite-based whole-genome sequencing analyses reveal global enhancer hypomethylation in human HCCs. Integrative epigenomic characterization further pinpoints a recurrent hypomethylated enhancer of CCAAT/enhancer-binding protein-beta (C/EBPß) which correlates with C/EBPß over-expression and poorer prognosis of patients. Demethylation of C/EBPß enhancer reactivates a self-reinforcing enhancer-target loop via direct transcriptional up-regulation of enhancer RNA. Conversely, deletion of this enhancer via CRISPR/Cas9 reduces C/EBPß expression and its genome-wide co-occupancy with BRD4 at H3K27ac-marked enhancers and super-enhancers, leading to drastic suppression of driver oncogenes and HCC tumorigenicity. Hepatitis B X protein transgenic mouse model of HCC recapitulates this paradigm, as C/ebpß enhancer hypomethylation associates with oncogenic activation in early tumorigenesis. These results support a causal link between aberrant enhancer hypomethylation and C/EBPß over-expression, thereby contributing to hepatocarcinogenesis through global transcriptional reprogramming.

Elevated perioperative transaminase level predicts intrahepatic recurrence in hepatitis B-related hepatocellular carcinoma after curative hepatectomy.

OBJECTIVE: We aimed to evaluate the role of elevated perioperative alanine aminotransferase (ALT) as a surrogate marker of hepatitis activity in determining the risk of recurrence and survival in hepatitis B-related hepatocellular carcinoma (HCC) after curative hepatectomy. METHODS: A retrospective review of the hepatectomy database was performed and 142 patients were found who had hepatitis B-related HCC from January 2001 to March 2006. Their ALT levels preoperatively and 1 month, 3 months, and 6 months postoperatively were recorded. The risk factors for recurrence and prognostic factors of survival were analysed. RESULTS: An elevated perioperative ALT level (p = 0.021), multiple tumour nodules in the resected specimen (p < 0.001), and a tumour size greater than 5 cm (p = 0.001) were significant independent risk factors for tumour recurrence. The latter two factors were also independent prognostic factors for overall survival and disease-free survival. An elevated ALT level was an independent prognostic factor for disease-free survival (p = 0.025). CONCLUSION: An elevated perioperative ALT level, which reflects increased hepatitis activity, is an independent risk factor for intrahepatic recurrence of hepatitis B-related HCC. It is also associated with a poorer disease-free survival rate.

Pattern of disease recurrence and its implications for postoperative surveillance after curative hepatectomy for hepatocellular carcinoma: experience from a single center.

BACKGROUND: Hepatectomy is a widely accepted curative treatment for hepatocellular carcinoma (HCC). However, the disease frequently recurs after a curative hepatectomy. The objective of this study is to provide a better understanding of the pattern of disease recurrence and the risk factors involved so as to improve the postoperative surveillance. METHODS: A retrospective study for all patients receiving hepatectomy for HCC between 2003 and 2014 was performed. Emphasis was made on the timing and pattern of recurrent disease, and type of treatment given. RESULTS: There were 506 patients in the study. Median follow-up was 43.7 months. The 1-, 3-, 5-, 10-year overall and disease free survival were 89.5%, 74.1%, 63.9%, 49.0% and 69.5%, 54.3%, 43.4%, 30.9% respectively. Recurrent disease occurred in 267 patients, 47.2% occurred within 9 months of hepatectomy and 80.1% recurred only in liver. Median survival was shorter for recurrence occurring within 9 months compared with those occurring between 10 months and 2 years postoperatively (36.2 vs. 65.7 months, P<0.01) whilst less curative treatment was offered for patients with early (within 9 months) intrahepatic alone recurrence (22.2% vs. 51.7%, P<0.01). Multivariate analysis revealed tumor size >3.5 cm and history of rupture were risk factors for recurrence within 9 months. CONCLUSIONS: These findings suggest that recurrent diseases are common after curative hepatectomy for HCC and most recurrences occur in the remnant liver. Since almost half of recurrences occurred within first 9 months after hepatectomy, a more stringent postoperative surveillance with target imaging of liver in this period is needed. Early diagnosis of recurrent disease and curative retreatment hopefully can bring about a longer survival.

An inflammatory-CCRK circuitry drives mTORC1-dependent metabolic and immunosuppressive reprogramming in obesity-associated hepatocellular carcinoma.

Obesity increases the risk of hepatocellular carcinoma (HCC) especially in men, but the molecular mechanism remains obscure. Here, we show that an androgen receptor (AR)-driven oncogene, cell cycle-related kinase (CCRK), collaborates with obesity-induced pro-inflammatory signaling to promote non-alcoholic steatohepatitis (NASH)-related hepatocarcinogenesis. Lentivirus-mediated Ccrk ablation in liver of male mice fed with high-fat high-carbohydrate diet abrogates not only obesity-associated lipid accumulation, glucose intolerance and insulin resistance, but also HCC development. Mechanistically, CCRK fuels a feedforward loop by inducing STAT3-AR promoter co-occupancy and transcriptional up-regulation, which in turn activates mTORC1/4E-BP1/S6K/SREBP1 cascades via GSK3ß phosphorylation. Moreover, hepatic CCRK induction in transgenic mice stimulates mTORC1-dependent G-csf expression to enhance polymorphonuclear myeloid-derived suppressor cell recruitment and tumorigenicity. Finally, the STAT3-AR-CCRK-mTORC1 pathway components are concordantly over-expressed in human NASH-associated HCCs. These findings unveil the dual roles of an inflammatory-CCRK circuitry in driving metabolic and immunosuppressive reprogramming through mTORC1 activation, thereby establishing a pro-tumorigenic microenvironment for HCC development.