Aggressive Hepatocellular Carcinoma Research Articles

Cyclin A2/E1 activation defines a hepatocellular carcinoma subclass with a rearrangement signature of replication stress

Cyclins A2 and E1 regulate the cell cycle by promoting S phase entry and progression. Here, we identify a hepatocellular carcinoma (HCC) subgroup exhibiting cyclin activation through various mechanisms including hepatitis B virus (HBV) and adeno-associated virus type 2 (AAV2) insertions, enhancer hijacking and recurrent CCNA2 fusions. Cyclin A2 or E1 alterations define a homogenous entity of aggressive HCC, mostly developed in non-cirrhotic patients, characterized by a transcriptional activation of E2F and ATR pathways and a high frequency of RB1 and PTEN inactivation. Cyclin-driven HCC display a unique signature of structural rearrangements with hundreds of tandem duplications and templated insertions frequently activating TERT promoter. These rearrangements, strongly enriched in early-replicated active chromatin regions, are consistent with a break-induced replication mechanism. Pan-cancer analysis reveals a similar signature in BRCA1-mutated breast and ovarian cancers. Together, this analysis reveals a new poor prognosis HCC entity and a rearrangement signature related to replication stress.

Indian Hedgehog links obesity to development of hepatocellular carcinoma.

Obesity increases the risk of hepatocellular carcinoma (HCC), but precise identification and characterization of druggable oncogenic pathways that contribute to the progression of NAFLD to HCC, and hence to the increased incidence and aggressiveness of HCC in obese individuals is lacking. In this regard, we demonstrate that the Indian Hedgehog (Ihh) signaling pathway is upregulated in the fatty livers of mice consuming a high fat diet, and furthermore sustained in HCC tumors specifically within the context of a NAFLD microenvironment. Using a diet-induced mouse model of HCC wherein only obese mice develop HCC, targeted ablation of hepatocyte-secreted Ihh results in a decreased tumor burden and lower grade tumors. Ihh activation regulates the transdifferentiation of ciliated stellate cells and proliferation of Epcam+ ductal cells to promote fibrosis. Mechanistically, increased expression of hitherto uncharacterized effectors of Hh pathway, namely Myc and Tgf-β2 is critical to the observed physiology. This pro-tumorigenic response is driven by increased expression of Wnt5a to effect a poorly-differentiated and invasive tumor phenotype. Wnt5a secreted from activated stellate cells act on Ror2-expressing hepatocytes. We further demonstrate that Wnt5a expression is also elevated in poorly-differentiated HCC cells, suggesting that these ligands are also able to function in an autocrine positive feedback manner to sustain poorly-differentiated tumors. Taken together, our study provides a mechanistic understanding for how Ihh signaling promotes HCC tumorigenesis specifically in obese mice. We propose that therapeutic targeting of the Hh pathway offers benefit for patients with dietary / NAFLD-driven steatotic HCC.

High Keratin 8/18 Ratio Predicts Aggressive Hepatocellular Cancer Phenotype

BACKGROUND & AIMS: Steatohepatitis (SH) and SH-associated hepatocellular carcinoma (HCC) are of considerable clinical significance. SH is morphologically characterized by steatosis, liver cell ballooning, cytoplasmic aggregates termed Mallory-Denk bodies (MDBs), inflammation, and fibrosis at late stage. Disturbance of the keratin cytoskeleton and aggregation of keratins (KRTs) are essential for MDB formation. METHODS: We analyzed livers of aged Krt18−/− mice that spontaneously developed in the majority of cases SH-associated HCC independent of sex. Interestingly, the hepatic lipid profile in Krt18−/− mice, which accumulate KRT8, closely resembles human SH lipid profiles and shows that the excess of KRT8 over KRT18 determines the likelihood to develop SH-associated HCC linked with enhanced lipogenesis. RESULTS: Our analysis of the genetic profile of Krt18−/− mice with 26 human hepatoma cell lines and with data sets of >300 patients with HCC, where Krt18−/− gene signatures matched human HCC. Interestingly, a high KRT8/18 ratio is associated with an aggressive HCC phenotype. CONCLUSIONS: We can prove that intermediate filaments and their binding partners are tightly linked to hepatic lipid metabolism and to hepatocarcinogenesis. We suggest KRT8/18 ratio as a novel HCC biomarker for HCC.

A20 Enhances the Radiosensitivity of Hepatocellular Carcinoma Cells to 60Co-γ Ionizing Radiation

The radioresistance of hepatocellular carcinoma (HCC) cells is a critical obstacle for effectively applying radiotherapy (RT) in HCC treatment. NF-κB, an important transcription factor, can influence critical cell fate decisions by promoting cell survival or anti-apoptosis in response to cell-stress, e.g. chemotherapies or ionizing radiation (IR). A20, also named as tumor necrosis factor α induced protein 3 (TNFAIP3 ), is a dominant negative regulator of NF-κB pathway and its functions in HCC are largely unknown. Cell culture and adenovirus vectors L-02, HepG2, MHCC-97H (a highly aggressive HCC cell line) and MHCC-97L (a lowly aggressive HCC cell line) cells were purchased from cell resources center of Chinese Academy of Medical Sciences (Beijing, China). The adenovirus vectors containing full length sequence of A20 and its A20 siRNA was purchased from Vigene Company (Jinan, Shandong Province, China). The HCC specimens and the adjacent non-tumor tissues were obtained by routinely surgical operations and preserved by our lab. The collection of tissues and the study protocol was all approved by the Ethics Committee of Xi’an Jiaotong University School of Medicine, and with the informed consent of patients. Paired HCC and non-tumorous tissues (40 paired) were obtained from surgical resections from February 2015 to April 2016. Using such methods as qPCR; Western Blot analysis; 60Co-γ ionizing radiation and colony formation; TUNEL reaction; Apoptosis analysis; Transwell analysis; In vivo tumor growth. Higher expression of A20 was detected in hepatic non-tumor cell line or clinical specimens compared with HCC cell lines or clinical specimens. A20 decreased the expression of proteins mediating cellular stress/injury response or epithelial-mesenchymal transition (EMT) process. Overexpression of A20 via adenovirus enhanced the effect of 60Co-γ ionizing radiation (IR) on HCC cells’ injury, e.g. G2/M arrest or DNA double strands break (DSB). Moreover, A20 also enhanced the in vitro or in vivo survival inhibiting of HCC cells induced by IR. Our results showed A20 downregulates the protein level of cellular stress/injury response regulators and by then potentiate the susceptibility of HCC cell line to 60Co-γ IR. Overexpression of A20 via an adenoviral vector can be a useful strategy for increasing tumor radiosensitivity.

C-Reactive Protein and Platelet-Lymphocyte Ratio as Potential Tumor Markers in Low-Alpha-Fetoprotein Hepatocellular Carcinoma

The hepatocellular carcinoma (HCC) tumor marker alpha-fetoprotein (AFP) is only elevated in about half of the HCC patients, limiting its usefulness in following the effects of therapy or screening. New markers are needed. It has been previously noted that the inflammation markers C-reactive protein (CRP) and platelet-lymphocyte ratio (PLR) are prognostically important and may reflect HCC aggressiveness. We therefore examined these 2 markers in a low-AFP HCC cohort and found that for HCCs > 2 cm, both markers significantly rise with an increasing maximum tumor diameter (MTD). We calculated the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and Youden index value for each marker, and their area-under-the-curve values for each MTD group. Patients were dichotomized into 2 groups based on the CRP and PLR from the receiver-operating characteristic curve analysis. In the logistic regression models of the 4 different MTD patient groups, CRP and PLR levels were statistically significant to estimate MTD in univariate logistic regression models of MTD groups > 2 cm. CRP and PLR were then combined, and the combination was statistically significant to estimate MTD groups of 3-, 4-, and 5-cm cutoffs. CRP and PLR thus have potential as tumor markers for low-AFP HCC patients, and possibly for screening.

Acute Liver Rejection After Exposure to Nivolumab in Treatment of Metastatic Hepatocellular Carcinoma

72 year old man with an orthotropic liver transplantation for hepatitis C with hepatocellular carcinoma (HCC) in 2012, complicated by metastatic HCC with a solitary lung lesion in 2015. Patient was initially treated with stereotactic body radiation with progression of disease in 2017 despite systemic therapy with sorafenib and regorafenib. Due to limited treatment options for progressive lung metastasis and excellent liver graft function, treatment with nivolumab was initiated. On routine monitoring, liver tests became elevated ten weeks after exposure to nivolumab with suspicion for either acute rejection or immune mediated hepatitis. Empiric treatment with oral steroids and an increased tacrolimus trough level to 5 ng/mL from 2-3 ng/ml failed to improve liver tests. A liver biopsy revealed lymphomonocytic infiltrate with endothelitis consistent with moderate acute rejection with Rejection Activity Index of 5. C4D staining was negative. Intravenous immunoglobulin 1 g/kg and methylprednisone 500 mg were employed without a durable response. Thymoglobulin 1.5 mg/kg/day with cumulative dose 6 mg/kg was started for steroid-resistant rejection with good response. Patient's liver enzymes trended down and normalized with addition of mycophenolate mofetil to tacrolimus to maintain remission. Recurrence rate of HCC is approximately 10% in five years in post-transplant population where transplant was performed due to HCC. Despite promising data in advanced HCC of immune checkpoint inhibitors (CPI) like nivolumab, an anti-PD-1 checkpoint inhibitor, these agents have not been studied in liver transplant recipients due to possibility of rejection and fatal liver failure. Current data regarding acute liver rejection caused by CPIs is scant and heterogenous. Most of the fatal rejections are found in the pediatrics population with aggressive fibrolamellar HCC and acute rejection happened within one week. There is a single case report of complete cancer remission with pembrolizumab, for metastatic HCC after live transplant. Here we report a case of acute liver rejection ten weeks after exposure to nivolumab that was resistant to steroids and IVIG. In this case, aggressive second line treatment with thymoglobulin was proven to be effective in the treatment of acute rejection. Factors that contribute to acute graft rejection with CPI use remain unknown and there is a need to develop regimens that optimize tumor response and minimize the risk of allograft rejection.2337_A Figure 1. Alanine aminotransferase/serum glutamic pyruvic transaminase trend2337_B Figure 2. Total bilirubin trend

Molecular Scoring of Hepatocellular Carcinoma for Predicting Metastatic Recurrence and Requirements of Systemic Chemotherapy.

Hepatocellular carcinoma (HCC) causes one of the most frequent cancer-related deaths; an HCC subset shows rapid progression that affects survival. We clarify molecular features of aggressive HCC, and establish a molecular scoring system that predicts metastasis after curative treatment. In total, 125 HCCs were examined for TP53, CTNNB1, and TERT promoter mutation, methylation of 8 tumor suppressor genes, and 3 repetitive DNA sequences to estimate promoter hypermethylation and global hypomethylation. A fractional allelic loss (FAL) was calculated to represent chromosomal instability through microsatellite analysis. Molecular subclasses were determined using corresponding and hierarchical clustering analyses. Next, twenty-five HCC patients who underwent liver transplantation were analyzed for associations between molecular characteristics and metastatic recurrence; survival analyses were validated using a publicly available dataset of 376 HCC cases from the Cancer Genome Atlas (TCGA). An HCC subtype characterized by TP53 mutation, high FAL, and global hypomethylation was associated with aggressive tumor characteristics, like vascular invasion; CTNNB1 mutation was a feature of the less-progressive phenotype. A number of molecular risk factors, including TP53 mutation, high FAL, significant global hypomethylation, and absence of CTNNB1 mutation, were noted to predict shorter recurrence-free survival in patients who underwent liver transplantation (p = 0.0090 by log-rank test). These findings were validated in a cohort of resected HCC cases from TCGA (p = 0.0076). We concluded that molecular risks determined by common genetic and epigenetic alterations could predict metastatic recurrence after curative treatments, and could be a marker for considering systemic therapy for HCC patients.

HSCs-derived COMP drives hepatocellular carcinoma progression by activating MEK/ERK and PI3K/AKT signaling pathways

BackgroundCartilage oligomeric matrix protein (COMP) is known to promote fibrosis in skin, lung and liver. Emerging evidence shows that COMP plays critical roles in tumor development, including breast cancer, colon cancer and hepatocellular carcinoma (HCC). Nevertheless, the role of COMP in HCC proliferation and metastasis and its underlying mechanisms remain fully unclear.MethodsSerum COMP was determined by ELISA. Cell Counting Kit-8 and plate colony formation were performed to evaluate cell proliferation. Wound healing and transwell assays were used to determine migration and invasion of HCC cells. Western blotting and immunofluorescence were carried out for detection of epithelial-to-mesenchymal transition (EMT) markers and MMPs in HCC cells. The in vivo role of COMP was evaluated using mouse models. We also measured effects of hepatic stellate cells (HSCs)-conditioned medium (CM) on HCC progression using transwell coculture system.ResultsHere, we found that serum COMP levels in HCC patients were significantly higher than those in healthy controls. Accordingly, high serum COMP levels in HCC patients significantly correlated with malignant clinical characteristics and poor clinical outcomes. Next, we investigated that recombinant human COMP protein (rCOMP) treatment resulted in increased abilities of proliferation, invasion and migration of HCC cells. Furthermore, rCOMP treatment enhanced proliferative and metastatic colonization of HCC cells in vivo. Mechanistically, CD36 receptor played an essential role in COMP-mediated HCC cell proliferation and metastasis. Functionally, COMP/CD36 signaling caused phosphorylation of ERK and AKT, resulting in the upregulation of tumor-progressive genes such as EMT markers, MMP-2/9, Slug and Twist in HCC cells. Interestingly, we revealed that COMP was secreted by HSCs. CM of LX2 cells with COMP knockdown showed weaker effects on the activation of MEK/ERK and PI3K/AKT signaling pathways in HCC cells compared to control CM.ConclusionsOur findings indicated that HSCs-derived COMP collaborated with CD36 and subsequently played an essential role in MEK/ERK and PI3K/AKT-mediated HCC progression. COMP might act as a promising target for the diagnosis and treatment of aggressive HCC.

4Mu Decreases CD47 Expression on Hepatic Cancer Stem Cells and Primes a Potent Antitumor T Cell Response Induced by Interleukin-12.

The tumor microenvironment (TME) represents a complex interplay between different cellular components, including tumor cells and cancer stem cells (CSCs), with the associated stroma; such interaction promotes tumor immune escape and sustains tumor growth. Several experimental approaches for cancer therapy are focused on TME remodeling, resulting in increased antitumor effects. We previously demonstrated that the hyaluronan synthesis inhibitor 4-methylumbelliferone (4Mu) decreases liver fibrosis and induces antitumor activity in hepatocellular carcinoma (HCC). In this work, 4Mu, in combination with an adenovirus encoding interleukin-12 genes (AdIL-12), elicited a potent antitumor effect and significantly prolonged animal survival (p< 0.05) in an orthotopic HCC model established in fibrotic livers. In assessing the presence of CSCs, we found reduced mRNA levels of CD133+, CD90+, EpCAM+, CD44+, and CD13+ CSC markers within HCC tumors (p< 0.01). Additionally, 4Mu downregulated the expression of the CSC marker CD47+ on HCC cells, promoted phagocytosis by antigen-presenting cells, and, combined with Ad-IL12, elicited a potent cytotoxic-specific Tcell response. Finally, animal survival was increased when CD133low HCC cells, generated upon 4Mu treatment, were injected in a metastatic HCC model. In conclusion, the combined strategy ameliorates HCC aggressiveness by targeting CSCs and as a result of the induction of anticancer immunity.

Epithelial to mesenchymal transition is involved in ethanol promoted hepatocellular carcinoma cells metastasis and stemness

Hepatocellular Carcinoma (HCC) is a malignant tumor with high rate of relapse and metastasis. Ethanol is a well-known risk factor for HCC; it promotes the progression and aggressiveness of HCC. However, the underlying mechanism remains unclear. In clinic studies, we showed that alcohol consumption is positively correlated with TNM stage and vessel invasion; HCC patients with chronic drinking history had faster progression rate and poorer prognosis compared to non-drinkers. In experimental models, ethanol exposure enhanced the metastasis, and invasion of HCC cells. Ethanol exposure increased cancer stem cells (CSC) population and enhanced stemness of HCC cells in vitro and in vivo. Mechanically, we found that ethanol exposure induced epithelial to mesenchymal transition (EMT) through activating Wnt/β-catenin signaling pathway in HCC cells. We further demonstrated that β-catenin siRNA or salinomycin (an inhibitor of Wnt/β-catenin pathway) partially rescued ethanol-induced EMT. In conclusion, this study suggested that ethanol exposure promotes the metastasis and stemness of HCC cells by inducing EMT.

PO-007 Investigating the hyperactivation of ERK5 signalling in skin cancer

IntroductionAcyl-Coenzyme A dehydrogenase (ACADM) is an enzyme responsible for degrading medium-chain fatty acids found in dairy foods and body fats. Mutations in the ACADM gene results in the accumulation of non-degraded or partially degraded medium chain fatty acids in tissues, leading to eventual liver and brain damage. Therefore, investigating the clinical relevance and functional role of ACADM in hepatocellular carcinoma (HCC) tumorigenesis can help us understand better the role of beta-oxidation genes in HCC.Material and methodsACADM mRNA expression in human HCC cell lines and tissues was determined by quantitative real time PCR and clinicopathological analysis was performed. Subsequent functional characterisation of ACADM in HCC was determined with cell migration, invasion and soft agar assays after the stable knockdown of ACADM in HCC cell lines.Results and discussionsACADM was downregulated in 64% of HCC cases examined. The overall ACADM mRNA level was significantly down-regulated in HCC tissues when compared with non-tumorous liver tissues and negatively correlated with the aggressiveness of HCC cell lines. Clinicopathological analysis revealed that ACADM under-expression was significantly correlated with bigger tumour size, venous invasion, late stage and poor differentiation in HCC. Stable knockdown clones of ACADM established in immortalised liver cell line and HCC cell lines showed elevated level of cellular triglyceride and lipid content. Functional analyses with cell migration, invasion and soft agar assays illustrated that when ACADM activity was suppressed, cells displayed increased abilities of migration, invasion and anchorage independent growth. Our findings further found that caveolin-1 (Cav1) acts as an upstream negative regulator of ACADM. In Cav1 knockdown cells, ACADM was upregulated at the transcriptional and translational levels. Cav1 and ACADM expressions showed a significant negative correlation in HCC tissues and cell lines.ConclusionCav1 has been reported to be a metastasis promoter in HCC. Here, we delineate a new molecular pathway in which oncogenic Cav1 downregulates ACADM leading to the dysregulation of beta-oxidation in HCC. Our findings also reveal the tight association between ACADM under-expression with the aggressive HCC clinical features suggesting targeting Cav1, negative regulator of ACADM, as a therapeutic intervention in HCC.

Low expression of WW domain-containing oxidoreductase associates with hepatocellular carcinoma aggressiveness and recurrence after curative resection.

WW domain‐containing oxidoreductase (WWOX), which has a protein‐interaction domain and is regarded to be a tumor suppressor, has been known to play an important role in anti‐angiogenesis and cancer progression. This study aimed to investigate prognostic values of WWOX expression in hepatocellular carcinoma (HCC) patients after hepatectomy. Additionally, we intended to formulate a valuable prognostic nomogram for HCCs. 182 HCC patients who underwent hepatectomy from January 2009 to January 2010 were enrolled in our study. qRT‐PCR, Western blot, and immunohistochemistry on tissue microarrays were then used to determine the expression levels of WWOX. An evaluation of the role of WWOX expression levels in the prognosis and outcome of patients was established. A decrease in the expression of WWOX was found when compared to adjacent tumor‐free tissues, which led to worse overall survival (OS) and recurrence‐free survival (RFS) and, therefore, was considered as an independent negative factor in the prognosis of HCC. Two nomograms, comprising WWOX, alpha‐fetoprotein (AFP), tumor size, and γ‐glutamyltransferase (γ‐GT), were constructed to obtain superior discriminatory abilities than conventional staging systems in terms of C‐index and clinical net benefit on decision curve analysis (DCA) for OS and RFS. Our data suggest that WWOX expression is strongly related to HCC post‐resection aggressiveness and recurrence. Additional advanced and accurate predictive model through the incorporation of WWOX into nomogram could help predict OS or RFS for HCC patients.

Macroscopic Portal Vein Thrombosis in HCC Patients.

Macroscopic portal vein invasion (PVT) by hepatocellular carcinoma (HCC) in the liver is one of the most important negative prognostic factors for HCC patients. The characteristics of a large cohort of such patients were examined. We found that the percent of patients with PVT significantly increased with increasing maximum tumor diameter (MTD), from 13.7% with tumors of MTD <5cm to 56.4% with tumors of MTD >10cm. There were similar numbers of HCC patients with very large tumors with and without PVT. Thus, MTD alone was insufficient to explain the presence of PVT, as were high AFP levels, since less than 50% of high AFP patients had PVT. However, the percent of patients with PVT was also found to significantly increase with increasing blood alpha-fetoprotein (AFP) levels and tumor multifocality. A logistic regression model that included these 3 factors together showed an odds ratio of 17.9 for the combination of MTD>5.0cm plus tumor multifocality plus elevated AFP, compared to low levels of these 3 parameters. The presence or absence of macroscopic PVT may therefore represent different HCC aggressiveness phenotypes, as judged by a significant increase in tumor multifocality and AFP levels in the PVT positive patients. Factors in addition to MTD and AFP must also contribute to PVT development.

Systematic review with meta-analysis: recurrence of hepatocellular carcinoma following direct-acting antiviral therapy.

Although studies suggest decreased incident hepatocellular carcinoma (HCC) after direct-acting antivirals (DAA), data are conflicting regarding HCC recurrence and aggressiveness in patients who have a history of HCC with complete response. Characterize HCC recurrence patterns after DAA therapy. Two reviewers searched MEDLINE and SCOPUS from January 2015 to December 2017 and identified studies evaluating HCC recurrence patterns following DAA therapy. A pooled estimate was calculated using the DerSimonian and Laird method for a random effects model. The study was conducted in accordance with PRISMA guidelines. Among 24 studies (n=1820 patients), the proportion of patients with HCC recurrence following DAA therapy ranged from 0% to 59% (pooled estimate 24.4%; 95% CI: 18.4%-30.4%). Among 11 full text manuscripts, pooled HCC recurrence was 21.9% (95% CI: 16.2%-28.3%). Factors associated with recurrence included history of prior HCC recurrence and a shorter interval between HCC complete response and DAA initiation. Nine studies comparing DAA-treated and interferon-treated or untreated patients found similar recurrence among DAA-treated patients. Most (77.8%) patients with HCC recurrence were detected at an early tumour stage, of whom 64.7% received curative treatment. Study limitations included heterogeneous cohorts, potential misclassification of HCC absence prior to DAA, ascertainment bias for recurrence, and short durations of follow-up. Current data suggest acceptable HCC recurrence rates after DAA therapy, particularly if DAA therapy is delayed at least 6months after HCC complete response. However, data characterising HCC recurrence after DAA therapy are of limited quality, highlighting the need for high quality prospective studies.

Triclosan treatment decreased the antitumor effect of sorafenib on hepatocellular carcinoma cells.

BackgroundTriclosan is a widely applied antimicrobial agent which affects the endocrine system and homeostasis; it may also promote the cirrhosis and hepatocellular carcinoma (HCC) growth in a mice model. The exact roles of triclosan in regulating human hepatocellular carcinoma development and treatment remain unknown.MethodsMHCC97-H, a highly aggressive HCC cell line, was treated with indicated concentration of triclosan or sorafenib. The expression of drug-resistance genes was examined by qPCR. The clearance or metabolism of sorafenib was determined by liquid chromatograph-mass spectrometer/mass spectrometer (LC-MS/MS). MTT assay was used to examine the MHCC97-H cell proliferation. Nude mice were used to exam the anti-tumor effect of sorafenib on subcutaneous and intrahepatic growth of MHCC97-H cells.ResultsIn the present study, triclosan could induce the expression of drug-resistance genes in MHCC97-H cells (a highly aggressive HCC cell line), accelerate the clearance of sorafenib, and attenuate the anti-proliferation effect of this molecular targeted agent in MHCC97-H cells. Triclosan decreased the antitumor effect of sorafenib on subcutaneous and intrahepatic growth of MHCC97-H in nude mice.ConclusionBy discovering the fact that triclosan treatment enhances sorafenib resistance in HCC cells, this work suggests exposure of triclosan is detrimental to HCC patients during chemotherapy.

Analysis of aggressiveness factors in hepatocellular carcinoma patients undergoing transarterial chemoembolization.

AIMTo investigate novel predictors of survival in hepatocellular carcinoma (HCC) patients following transarterial chemoembolization (TACE).METHODSOne hundred sixty seven patients with un-resectable HCC were retrospectively analyzed to identify factors that might contribute to their HCC biology and aggressiveness. We correlated routine laboratory results (total bilirubin, AST, ALKP, GGTP, albumin etc.) to maximum tumor diameter, number of tumor nodules, portal vein thrombosis and blood alpha-fetoprotein levels. These 4 parameters were previously combined to form an aggressiveness index (AgI). We used The Wilcoxon rank-sum (Mann-Whitney), to test the correlation between the AgI categories and liver function parameters. The Cox proportional hazards model was applied to evaluate the categories of AgI associated with overall survival.RESULTSThe AgI was strongly correlated with survival in this novel patient population. Three year survival probability for AgI > or < 4 was 42.4% vs 61.8%; P < 0.0863 respectively. Several factors independently correlated with AgI using univariate multiple logistic regression of AgI with 8 laboratory parameters. Lower albumin levels had an OR of 2.56 (95%CI: 1.120-5.863 P < 0.026), elevated Alkaline phosphatase and gamma glutamyl transpeptidase (GGTP) had ORs of 1.01 (95%CI: 1.003-1.026, P < 0.017) and 0.99 (95%CI: 0.99-1.00, P < 0.053) respectively. In a Cox proportional hazard model combining mortality for AgI score and liver function parameters, only GGTP levels and the AgI were independently associated with survival. An AgI > 4 had HR for mortality of 2.18 (95%CI: 1.108-4.310, P < 0.024). GGTP’s single unit change had a HR for mortality of 1.003 (95%CI: 1.001-1.006, P < 0.016). These were considered in the final multivariate model with the total cohort. An AgI > 4 had a HR for mortality of 2.26 (95%CI: 1.184-4.327, P < 0.016). GGTP had a HR of 1.003 (95%CI: 1.001-1.004, P < 0.001).CONCLUSIONOur study validates the AgI in a new population with un-resectable HCC patients undergoing TACE. The analysis establishes a correlation between GGTP and the AgI.

Aggressive hepatocellular carcinoma occurring during direct antiviral therapy

Aggressive hepatocellular carcinoma occurring during direct antiviral therapy

TIPE‑2 suppresses growth and aggressiveness of hepatocellular carcinoma cells through downregulation of the phosphoinositide 3‑kinase/AKT signaling pathway.

Rapid proliferation and migration are the main features of hepatocellular carcinoma (HCC) cells, which serve an essential role in carcinogenesis and are a hallmark of cancer therapy resistance. Previous studies have reported that tumor necrosis factor-α-induced protein-8 like-2 (TIPE-2) is involved in cancer initiation and the progression of HCC. The present study aimed to clarify the role of TIPE-2 in HCC carcinogenesis, growth and aggressiveness. The effects of TIPE-2 on HCC were determined using colony forming and cell cycle analyses. Cell apoptosis, and growth and aggressiveness of HCC cells, were investigated following TIPE-2 treatment. Treatment with TIPE-2 markedly suppressed HCC cell proliferation and increased the number of cells in S phase of the cell cycle. The results demonstrated that TIPE-2 significantly inhibited growth, migration and invasion of HCC cells via the downregulation of tumor metastasis-associated genes. Flow cytometric analysis indicated that TIPE-2 promoted apoptosis of HCC cells via regulation of apoptosis-associated gene transcription. In addition, TIPE-2 administration downregulated the expression of phosphoinositide 3-kinase (PI3K) and protein kinase B (AKT) in HCC cells. In addition, TIPE-2 selectively decreased neuroblastoma Ras viral oncogene and p27 expression in HCC cells. In vivo assays revealed that TIPE-2 significantly inhibited tumor growth and prolonged animal survival by promoting apoptosis of tumor cells. The results of the present study indicated that TIPE-2 acts as an inhibitor of HCC cell growth and aggressiveness, and promotes apoptosis, thus suggesting that TIPE-2 may inhibit the metastasis-associated PI3K/AKT signaling cascade and may arrest the tumor cell cycle. These findings provide a potential molecular mechanism by which TIPE-2 promotes apoptosis of HCC cells.

Unusual presentation of a hepatocellular carcinoma as a potential late side effect of radiotherapy in a patient treated for Wilms tumor in childhood

BackgroundThe development of a second primary tumor is a potential late side effect of radiotherapy. Particularly, an increased risk of secondary cancers, mostly of digestive or breast origin, has been observed in patients treated with high-dose radiotherapy for Wilms tumor (WT) in childhood. However, hepatocellular carcinoma (HCC) has been very rarely described as a potentially radiotherapy-induced tumor. We describe the case of a patient with an aggressive HCC 50 years after the treatment of a WT.Case presentationA 49-year old man, treated at the age of 6 weeks for a right WT by a right nephrectomy and adjuvant radiotherapy, presented with a right abdominal mass. Imaging demonstrated a 100-mm tumor invading the inferior segment of the right liver, the right colon and the right psoas muscle. The patient had no previous history of liver disease, nor of alcohol consumption, and hepatitis serologies were negatives. Biopsy demonstrated a poorly differentiated tumor of unknown origin. A panel of tumor markers was negative. Explorative surgery has been performed allowing en bloc R0 tumor resection, including resection of segments VI and VII of the liver, right hemicolectomy and resection of the anterior sheet of the right psoas muscle. Pathological examination revealed a poorly differentiated HCC. No signs of cirrhosis or chronic liver disease were observed in the non-tumor liver. Twenty weeks after surgery, the patient developed a multifocal tumor recurrence that was treated with intra-arterial 90Yttrium radioembolization.ConclusionIn this case, the absence of risk factors for HCC, such as cirrhosis, viral hepatitis and chronic liver disease, highly suggests the development of HCC to be related to previous high-dose radiation therapy given for a right WT to a field involving the inferior part of the liver. This observation shows radiotherapy to/near the liver, particularly in childhood, to be a potential risk factor for HCC, stressing the need for a long-term specific follow-up in patients irradiated in childhood.

Targeting DTL induces cell cycle arrest and senescence and suppresses cell growth and colony formation through TPX2 inhibition in human hepatocellular carcinoma cells.

BackgroundHepatocellular carcinoma (HCC) has an increasing incidence and high mortality. Surgical operation is not a comprehensive strategy for liver cancer. Moreover, tolerating systemic chemotherapy is difficult for patients with HCC because hepatic function is often impaired due to underlying cirrhosis. Therefore, a comprehensive strategy for cancer treatment should be developed. DTL (Cdc10-dependent transcript 2) is a critical regulator of cell cycle progression and genomic stability. In our previous study, the upregulation of DTL expression in aggressive HCC correlated positively with tumor grade and poor patient survival. We hypothesize that targeting DTL may provide a novel therapeutic strategy for liver cancer. DTL small interference RNAs were used to knock down DTL protein expression.MethodsA clonogenic assay, immunostaining, double thymidine block, imaging flow cytometry analysis, and a tumor spheroid formation assay were used to analyze the role of DTL in tumor cell growth, cell cycle progression, micronucleation, ploidy, and tumorigenicity.ResultsOur results demonstrated that targeting DTL reduced cell cycle regulators and chromosome segregation genes, resulting in increased cell micronucleation. DTL depletion inhibited liver cancer cell growth, increased senescence, and reduced tumorigenesis. DTL depletion resulted in the disruption of the mitotic proteins cyclin B, CDK1, securin, seprase, Aurora A, and Aurora B as well as the upregulation of the cell cycle arrest gene p21. A rescue assay indicated that DTL should be targeted through TPX2 downregulation for cancer cell growth inhibition. Moreover, DTL silencing inhibited the growth of patient-derived primary cultured HCC cells.ConclusionOur study results indicate that DTL is a potential novel target gene for treating liver cancer through liver cancer cell senescence induction. Furthermore, our results provide insights into molecular mechanisms for targeting DTL in liver cancer cells. The results also indicate several other starting points for future preclinical and clinical studies on liver cancer treatment.