Disease Progression In Hepatocellular Carcinoma Research Articles

AHSA1 Regulates Hepatocellular Carcinoma Progression via the TGF-β/Akt-Cyclin D1/CDK6 Pathway.

Activator of heat shock protein 90 (HSP90) ATPase Activity 1 (AHSA1) regulates proliferation, apoptosis, migration, and invasion of osteosarcoma and hepatocellular carcinoma (HCC). However, the novel mechanism of AHSA1 in the tumor biology of hepatocellular carcinoma (HCC) remains unclear. We analyzed AHSA1 expression in 85 pairs of clinical samples of HCC and the Cancer Genome Atlas database. The role of AHSA1 in HCC was proved by cell proliferation, colony formation, migration, cell cycle analysis in vitro, xenograft models and tumor metastasis assay in vivo, and bioinformatics. High AHSA1 expression was demonstrated in HCC and associated with invasive depth, clinical stage, and poor overall survival of patients. UnivariateCox analysis confirmed that AHSA1 was an independent prognostic factor for patients with HCC. Meanwhile, AHSA1 upregulation promoted cell proliferation, colony formation, and cell migration in vitro and tumor cell proliferation and metastasis of HCC cells in vivo. AHSA1 upregulation increased the cell cycle transition from G1 to S phase by increasing the expression of cyclinD1, cyclinD3, and cyclin-dependent kinase 6(CD). Transforming growth factor beta 1 (TGF-β1)-induced protein kinase B (Akt) signaling regulated the expression of downstream targets, including cyclinD1. AHSA1 expression was closely correlated with the expression of TGF-β, Akt, cyclinD1, cyclinD3, and CDK6 using the Gene Expression Profiling Interactive Analysis database. AHSA1 upregulation participated in HCC progression by regulating TGF-β/Akt-cyclinD1/CDK6 signaling. AHSA1 might serve as a biomarker for predicting the clinical outcome of patients with HCC. It is vital in tumor metastasis and disease progression of HCC and may facilitate the development of clinical intervention strategies against HCC.

Dysregulated sphingolipid metabolism contributes to NASH-HCC disease progression

Background: Nonalcoholic fatty liver disease (NAFLD) is the fastest-rising cause of end-stage liver disease for liver transplantation. The underlying mechanisms of NAFLD disease progression from nonalcoholic steatohepatitis (NASH) to hepatocellular carcinoma (HCC) remain unclear. It has been well-accepted that inflammation and dysregulation of sphingolipid metabolism are two important contributors to NASH progression. Recent advances in gene profiling technologies have significantly improved the current understanding of NASH-HCC pathogenesis. However, the key pathway changes underlying liver fibrosis and dysregulation of sphingolipid metabolism haven’t been fully elucidated and are the focus of this study. Methods: DIMOND NASH mouse model (C57BL/6J and 129S1/SvlmJ mixed background) was used in this study. Male mice (21 to 24-week-old) were fed with WDSW (Western diet, Harlan, TD88137, and a high fructose-glucose solution, 23.1g/L D-fructose plus 18.9g/L D-glucose) or standard chow diet ad libitum for six months or one year. At the end of the study, liver tissues were harvested for total RNA isolation using Trizol. Gene profiles were analyzed using the NanoString nCounter® Gene Expression Fibrosis V2 panel with sphingolipid metabolism pathways. The differentially expressed genes (DEGs) between groups were determined using Rosalind. QIAGEN Ingenuity Pathway Analysis (IPA) was used to analyze the DEGs. Human HCC patient data were downloaded from The Cancer Genome Atlas Program (TCGA). The sphingolipid profiles in the serum and liver were determined using LC-MS/MS. Results: All the mice developed NASH and HCC after feeding with WDSW for 6 months and 1 year, respectively. Bioinformatic analysis identified 191 DEGs in the NASH group and 250 DEGs in HCC group compared to the corresponding controls. The IPA analysis revealed that the major pathways related to inflammation and fibrosis were upregulated in both NASH and HCC mice. The expression levels of key genes related to ceramide synthesis and metabolism [serine palmitoyltransferase long chain base subunit 2 (Sptlc2), ceramide synthase 6 (Cers6), N-acylsphingosine amidohydrolase 2 (Asah2)] and sphingosine-1-phosphate receptor 2 (S1PR2) were significantly upregulated in NASH mice. In HCC group, Cers5, Cers6, sphingomyelin synthase 1 (Sgms1), Asah1, and S1PR3 were significantly increased. The expression levels of sphingosine kinase 2 and S1P lyase 1 were decreased both in NASH and HCC. Analysis of the TCGA data set showed that CERS5, CERS6, SPTLC2, ASAH1 and SGMS1 were highly expressed in HCC compared to healthy controls. The LC-MS/MS data also showed significant changes in sphingolipid profiles in serum and liver NASH and HCC. Conclusion: Our study suggests that dysregulated sphingolipid metabolism is associated with NASH and HCC disease progression. Further validation and research are needed for the potential application of sphingolipid profiles as diagnostic or prognostic markers in the clinic. This study was supported by VA Merit Award I01BX004033; Research Career Scientist Award (IK6BX004477); VA ShEEP grants (1IS1BX005517-01 and 1 IS1 BX004777-01) National Institutes of Health Grant R01 DK104893, R01DK-057543 and R21 AA026629-01. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Identification and Characterization of an Ageing-Associated 13-lncRNA Signature That Predicts Prognosis and Immunotherapy in Hepatocellular Carcinoma.

In cancer pathology, cell senescence not only alters cell function but also reshapes the immune microenvironments in tumours. However, the association between cell senescence, tumour microenvironment, and disease progression of hepatocellular carcinoma (HCC) is yet to be fully understood. Therefore, the role of cell senescence-related genes and long noncoding RNAs (lncRNAs) in evaluating the clinical prognosis and immune cell infiltration (ICI) of HCC patients requires further investigation. The limma R package was utilised to investigate differentially expressed genes according to the multiomics data. The CIBERSORT R package was utilised to assess ICI, and unsupervised cluster analysis was conducted using the R software's ConsensusClusterPlus package. A polygenic prognostic model of lncRNAs was constructed by conducting univariate and least absolute shrinkage and selection operator (Lasso) cox proportional-hazards regression analyses. The time-dependent receiver operating characteristic (ROC) curves were used for validation. We utilised the survminer R package to evaluate the tumour mutational burden (TMB). Moreover, the gene set enrichment analysis (GSEA) helped in pathway enrichment analysis, and the immune infiltration level of the model was evaluated using the IMvigor210 cohort. The identification of 36 prognosis-related genes was achieved based on their differential expression between healthy and liver cancer tissues. Liver cancer individuals were categorised into 3 independent senescence subtypes using the gene list, revealing considerable survival differences (variations). We observed that the prognosis of patients in the ARG-ST2 subtype was substantially better as compared to that in the ARG-ST3 subtype. Differences were observed in gene expression profiles among the three subtypes, with the differentially expressed genes predominantly associated with cell cycle control. The enrichment of upregulated genes in the ARG-ST3 subtype was observed in pathways related to biological processes, for instance, organelle fission, nuclear division, and chromosome recombination. ICI in the ARG-ST1 and ARG-ST2 subtypes, with relatively better prognosis, was substantially higher as compared to the ARG-ST3 subtype. Furthermore, a risk-score model, which can be employed as a reliable prognostic factor in an independent manner for individuals suffering from liver cancer, was constructed based on 13 cell senescence-related lncRNAs (MIR99AHG, LINC01224, LINC01138, SLC25A30AS1, AC006369.2, SOCS2AS1, LINC01063, AC006037.2, USP2AS1, FGF14AS2, LINC01116, KIF25AS1, and AC002511.2). The individuals with higher risk scores had noticeably poor prognoses in contrast with those having low-risk scores. Moreover, increased levels of TMB and ICI were observed in individuals with low-risk scores and gaining more benefit from immune checkpoint therapy. Cell senescence is an essential factor in HCC onset and progression. We identified 13 senescence-related lncRNAs as HCC prognostic markers, which can help understand their function in the onset and progression of HCC and guide clinical diagnosis and treatment.

Impact of Traditional Chinese Medicine on Prognosis of Patients with Hepatocellular Carcinoma: A Retrospective Cohort Study.

In a previous study, we found that traditional Chinese medicine (TCM) alleviated the clinical symptoms and improved the quality of life (QoL) in patients with hepatocellular carcinoma (HCC). A cohort was continuously followed up to determine the impact of the TCM adjuvant therapies on the prognosis of HCC after conventional treatments. We did a retrospective monocentric cohort study including 175 eligible patients. The participants who received TCM adjuvant therapies were termed as TCM group. For the purpose of stratification analysis, the patients who received TCM adjuvant therapies over 3 months per year were further classified into the high frequency group, while the rest of the TCM users were classified into the low frequency group. Non-users were recorded as the control group. The primary outcome was overall survival (OS) and the secondary outcome was the mean progression-free survival (mPFS) primarily introduced in this study, referring to the time from initial diagnosis to the latest progression over the number of disease progressions. Analyses used Cox proportional hazards and Kaplan-Meier (K-M) methods, adjusted for stratification factors. Until June 30, 2021, 56 patients survived, 21 patients were lost to follow-up, and 98 patients died from the disease. Each disease progression of every individual was recorded, and most of the PFS was within 1 year. The baseline data of the allocated groups were balanced, the result revealed that TCM adjuvant therapies might have little influence on OS (P = .129). However, the 1, 3, and 5-year progression-free survival rates of the patients in TCM and control group were 68.75%, 37.50%; 25.00%, 8.33% and 8.33%, 2.08%, respectively, indicating TCM use significantly extended the mPFS, and decreased the risk of disease progression by a factor of 0.676 (P = .006). In the patients with BCLC stage B HCC, compared with controls, a 37-month median OS advantage in the high frequency group was noted (P = .045); and the high frequency of TCM use significantly suppressed disease progression (P = .001). The present study revealed that TCM adjuvant therapies could postpone disease progression in HCC. Furthermore, using TCM over 3 months per year might extend OS in patients with intermediate HCC.

Whole genome analysis of hepatitis B virus before and during long-term therapy in chronic infected patients: Molecular characterization, impact on treatment and liver disease progression.

Hepatitis B virus (HBV) infection remains a serious public health concern worldwide despite the availability of an efficient vaccine and the major improvements in antiviral treatments. The aim of the present study is to analyze the mutational profile of the HBV whole genome in ETV non-responder chronic HBV patients, in order to investigate antiviral drug resistance, immune escape, and liver disease progression to Liver Cirrhosis (LC) or Hepatocellular Carcinoma (HCC). Blood samples were collected from five chronic hepatitis B patients. For each patient, two plasma samples were collected, before and during the treatment. Whole genome sequencing was performed using Sanger technology. Phylogenetic analysis comparing the studied sequences with reference ones was used for genotyping. The mutational profile was analyzed by comparison with the reference sequence M32138. Genotyping showed that the studied strains belong to subgenotypes D1, D7, and D8. The mutational analysis showed high genetic variability. In the RT region of the polymerase gene, 28 amino acid (aa) mutations were detected. The most significant mutations were the pattern rtL180M + rtS202G + rtM204V, which confer treatment resistance. In the S gene, 35 mutations were detected namely sP120T, sT126S, sG130R, sY134F, sS193L, sI195M, and sL216stop were previously described to lead to vaccine, immunotherapy, and/or diagnosis escape. In the C gene, 34 mutations were found. In particular, cG1764A, cC1766G/T, cT1768A, and cC1773T in the BCP; cG1896A and cG1899A in the precore region and cT12S, cE64D, cA80T, and cP130Q in the core region were associated with disease progression to LC and/or HCC. Other mutations were associated with viral replication increase including cT1753V, cG1764A/T, cC1766G/T, cT1768A, and cC1788G in the BCP as well as cG1896A and cG1899A in the precore region. In the X gene, 30 aa substitutions were detected, of which substitutions xT36D, xP46S, xA47T, xI88F, xA102V, xI127T, xK130M, xV131I, and xF132Y were previously described to lead to LC and/or HCC disease progression. In conclusion, our results show high genetic variability in the long-term treatment of chronic HBV patients causing several effects. This could contribute to guiding national efforts to optimize relevant HBV treatment management in order to achieve the global hepatitis elimination goal by 2030.

Glucometabolic reprogramming: From trigger to therapeutic target in hepatocellular carcinoma.

Glucose, the central macronutrient, releases energy as ATP through carbon bond oxidation and supports various physiological functions of living organisms. Hepatocarcinogenesis relies on the bioenergetic advantage conferred by glucometabolic reprogramming. The exploitation of reformed metabolism induces a uniquely inert environment conducive to survival and renders the hepatocellular carcinoma (HCC) cells the extraordinary ability to thrive even in the nutrient-poor tumor microenvironment. The rewired metabolism also confers a defensive barrier which protects the HCC cells from environmental stress and immune surveillance. Additionally, targeted interventions against key players of HCC metabolic and signaling pathways provide promising prospects for tumor therapy. The active search for novel drugs based on innovative mutation targets is warranted in the future for effectively treating advanced HCC and the preoperative downstage. This article aims to review the regulatory mechanisms and therapeutic value of glucometabolic reprogramming on the disease progression of HCC, to gain insights into basic and clinical research.

Changes in immune profile affect disease progression in hepatocellular carcinoma.

Objective: Hepatocellular carcinoma (HCC) as a chronic liver condition is largely associated with immune responses. Previous studies have revealed that different subsets of lymphocytes play fundamental roles in controlling or improving the development and outcome of solid tumors like HCC. Hence, this study aimed to investigate whether immune system changes were related to disease development in HCC patients. Methods: Peripheral blood mononuclear cells were isolated from 30 HCC patients and 30 healthy volunteers using Ficoll density centrifugation. The isolated cells were stained with different primary antibodies and percentages of different immune cells were determined by flow cytometry. Results: HCC patients indicated significant reductions in the numbers of CD4+ cells, Tbet+IFNγ+cells, and GATA+IL-4+cells in peripheral blood in comparison with healthy individuals (p < 0.05). There was no significant change in IL-17+RORγt+cells between patient and healthy groups. In contrast, Foxp3+CD127lowcell frequency was significantly higher in patients than healthy subjects (p < 0.0001). The numbers of Th1, Th2, and Th17 cells were significantly lower in HCC patients than healthy control (p < 0.0001), although the reduction in Th2 cell numbers was not statistically significant. On the contrary, Treg percentage showed a significant increase in patients compared to healthy subjects (p < 0.0001). Other data revealed that Th1, Th2, and Th17 cell frequencies were significantly higher in healthy individuals than patients with different TNM stages of HCC, with the exception of Th2 in patients with stage II HCC (p < 0.01–0.05). Treg percentage was significantly increased in patients with different TNM stages (p < 0.0001). Among all CD4+ T cells, the frequency of Th2 cell was significantly associated with TNM stages of HCC (p < 0.05). Conclusion: Our data provide further evidence to show that immune changes may participate in determining HCC progression and disease outcome. However, it should be mentioned that more investigations are needed to clarify our results and explain possible impacts of other immune cells on the pathogenesis of HCC.

Acquired Resistance to Antiangiogenic Therapies in Hepatocellular Carcinoma Is Mediated by Yes-Associated Protein 1 Activation and Transient Expansion of Stem-Like Cancer Cells.

Induction of neoangiogenesis is a hallmark feature during disease progression of hepatocellular carcinoma (HCC). Antiangiogenetic compounds represent a mainstay of therapeutic approaches; however, development of chemoresistance is observed in the majority of patients. Recent findings suggest that tumor‐initiating cells (TICs) may play a key role in acquisition of resistance, but the exact relevance for HCC in this process remains to be defined. Primary and established hepatoma cell lines were exposed to long‐term sorafenib treatment to model acquisition of resistance. Treatment effects on TICs were estimated by sphere‐forming capacity in vitro, tumorigenicity in vivo, and flow cytometry. Adaptive molecular changes were assessed by whole transcriptome analyses. Compensatory mechanisms of resistance were identified and directly evaluated. Sustained antiproliferative effect following sorafenib treatment was observed in three of six HCC cell lines and was followed by rapid regrowth, thereby mimicking responses observed in patients. Resistant cells showed induction in sphere forming in vitro and tumor‐initiating capacity in vivo as well as increased number of side population and epithelial cell adhesion molecule‐positive cells. Conversely, sensitive cell lines showed consistent reduction of TIC properties. Gene sets associated with resistance and poor prognosis, including Hippo/yes‐associated protein (YAP), were identified. Western blot and immunohistochemistry confirmed increased levels of YAP. Combined treatment of sorafenib and specific YAP inhibitor consistently revealed synergistic antioncogenic effects in resistant cell lines. Conclusion: Resistance to antiangiogenic therapy might be driven by transient expansion of TICs and activation of compensatory pro‐oncogenic signaling pathways, including YAP. Specific targeting of TICs might be an effective therapeutic strategy to overcome resistance in HCC.

Increased Serum Pentraxin 3 Levels are Associated with Poor Prognosis of Hepatitis B Virus-Related Hepatocellular Carcinoma.

BackgroundHepatocellular carcinoma (HCC) is a major cause of liver-related mortality. Serum pentraxin 3 (PTX3) has been revealed to be associated with the development of hepatitis B virus (HBV)-related HCC. This study evaluated whether serum PTX3 is related to the survival of HBV-related HCC patients.MethodsOne hundred and seven patients with HBV-related HCC were included. Baseline serum PTX3 levels were quantified using quantitative immunoassay. The HCC patients were followed-up for a median of 24 months and divided into high serum PTX3 level and low PTX3 level groups according to the baseline serum PTX3 levels. The overall survivals of the HBV-related HCC patients according to the serum PTX3 levels were compared. Factors potentially influencing the prognosis of the patients with HBV-related HCC were analyzed.ResultsHCC patients with high serum PTX3 levels [PTX3 > 9.25ng/mL (n=85)] had a shorter overall survival time than HCC patients with low serum PTX3 levels [PTX3 ≤ 9.25ng/mL (n=22)] (P = 0.049). HCC patients with serum PTX3 levels between >9.25ng/mL and ≤9.25ng/mL had significant difference in HCC histology grade. Multivariate analysis showed that PTX3 level was an independent risk factor related to the overall survival of HCC patients (hazard ratio: 1.058, 95% confidence interval: 1.031–1.085, P <0.001).ConclusionThese results support the involvement of PTX3 in the disease progression of HCC and suggest the potential of using serum PTX3 levels as a biomarker for the prognostic prediction of HBV-related HCC patients.

Tspan5 promotes epithelial-mesenchymal transition and tumour metastasis of hepatocellular carcinoma by activating Notch signalling.

Hepatocellular carcinoma (HCC) is one of the most lethal cancers worldwide due to a high rate of tumour metastasis and disease recurrence. In physiological conditions, tetraspanins interact with specific partner proteins in tetraspanin‐enriched microdomains and regulate their subcellular localization and function. However, the function of Tspan5 in pathological processes, particularly in cancer biology and its clinical significance, are still unclear. Here, we describe that a high expression of Tspan5 is significantly associated with some clinicopathological features including invasive length, vascular invasion, clinical stage and poor overall survival of HCC patients. Alterations of Tspan5 expression by lentivirus transductions in HCC cells demonstrated that Tspan5 promotes wound healing and cell migration in vitro and tumour metastasis of HCC cells in vivo. Mechanistic studies revealed that Tspan5 promoted cell migration and tumour metastasis by increasing the enzymatic maturation of ADAM10 and activating Notch signalling via the increase of the cleavage of the Notch1 receptor catalysed by the γ‐secretase complex. Activation of Notch signalling by Tspan5 was shown further to enhance the epithelial–mesenchymal transition (EMT) and actin skeleton rearrangement of tumour cells. In clinical HCC samples, Tspan5 expression is strongly correlated with many key molecules acting in Notch signalling and EMT, highlighting the role of Tspan5 in the regulation of Notch signalling, EMT and tumour metastasis of HCC. Our findings provide new insights into the mechanism of tumour metastasis and disease progression of HCC and may facilitate the development of novel clinical intervention strategies against HCC.

Abstract 991: Platelets inhibit hepatocellular carcinoma growth in the context of nonalcoholic fatty liver disease

Abstract Purpose: Non-alcoholic fatty liver disease (NAFLD) is a major risk factor for hepatocellular carcinoma (HCC) in the US and globally. Platelets have been recognized as an important element of immune system and recently, platelets have been shown to be activated and aggerate in the livers of mice and humans with NAFLD. However, the role of platelets in HCC disease progression in NAFLD is still unknown. Methods: In this study, mice underwent an intrahepatic injection of an HCC cell line and were fed a methionine choline deficient (MCD) diet to generate an HCC NAFLD mouse model. The mouse model was then used to interrogate the interplay between platelets, immune responses and HCC growth within a liver with NAFLD. Anti-GP1bα or clopidogrel were used to deplete or inhibit platelet function and effects on tumor growth and immune cell activation were studied. Platelets express leukotriene receptors, and the possible involvement of leukotrienes was tested with zileuton, a leukotriene inhibitor. Finally, antibody dependent T cell depletion was used to identify the effector cells controlling HCC tumor development. Results: NAFLD mice with HCC treated with clopidogrel showed a close to 3-fold increase in tumor weight compared to vehicle control (50.6g vs 147.8g P&amp;lt;0.05) as well as tumor-to-liver weight ratio (0.05 vs 0.13 P&amp;lt;0.05). CD4+ and CD8+ T-cells from the livers of NAFLD mice showed a 2- to 3-fold increase in CD69+ T-cells compared to non-NAFLD mice (24.2% vs 9.1% P&amp;lt;0.05 in CD4+ and 23.7% vs 8.9% P&amp;lt;0.05 in CD8+, respectively). Interestingly, in NAFLD mice treated with clopidogrel, there was a decrease in CD69+ compared to NAFLD mice treated with control in both CD4+ (18.3% vs 24.2% P&amp;lt;0.05, respectively) and CD8+ (15.32% vs 23.7% P&amp;lt;0.05, respectively) T-cells. Consistently, when NAFLD mice were depleted of platelets with anti-GP1bα, there was a significant decreased in T cell activation compared to IgG control. Similarly, when NAFLD mice were treated with zileuton, there was a decrease in IFN-gamma production in liver isolated CD4+ (5.4% vs 3.1% P=0.12) and CD8+ (23.7% vs 16.7% P&amp;lt;0.05) T-cells compared to vehicle control treated mice. Finally, the clopidogrel treatment increased HCC tumor-to-liver weight ratio was abolished in mice treated with anti-CD8 (0.29 vs 0.23 P&amp;gt;0.05) but not in mice treated with anti-CD4 (0.11 vs 0.16 P&amp;lt;0.05). Conclusion: The results of this study show that in NAFLD livers, platelets increase immune cell activation and inhibit HCC in a CD8+ T-cell dependent manner. The results also suggest that leukotrienes may play a key role in modulating this response, but further studies are required to better characterize this observation. Citation Format: John C. McVey, Chi Ma, Laurence Diggs, Qiong Fu, Tim F. Greten. Platelets inhibit hepatocellular carcinoma growth in the context of nonalcoholic fatty liver disease [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 991.

Human Telomerase Reverse Transcriptase Gene Promoter Mutation in Serum of Patients with Hepatocellular Carcinoma

Background: Mutations in the human telomerase reverse transcriptase (hTERT) gene promoter have been reported in hepatocellular carcinoma (HCC); however, analyses of these mutations in liquid biopsies have been technically difficult because of the high GC content of the regions of interest within this promoter. We evaluated the feasibility and prognostic value of hTERT promoter mutations identified in circulating cell-free DNA (cfDNA) from the serum of patients with HCC. Objective: A cohort of HCC patients (n = 36) who were curatively treated by surgical resection between June 2003 and September 2014 were enrolled in this study. Methods: The presence of hTERT promoter mutations in cfDNA from the patients’ serum was analyzed via modified droplet digital polymerase chain reaction, and associations were sought between specific promoter mutations and patients’ disease-free survival (DFS). Results: The G>A hTERT mutation at –124 bp was detected in the serum of 25 patients (69%). Although no marked differences were observed between the characteristics of the serum mutation-positive and serum mutation-negative patient groups, the DFS of patients with the mutation was significantly shorter than that of the serum mutation-negative patients (p = 0.02). Among 18 clinicopathologic and background liver factors examined, the presence of the –124 bp G>A mutation was an independent and significant predictor of patients’ DFS (hazard ratio = 3.01, 95% confidence interval 1.11–10.5, p = 0.03) in multivariate analyses. Conclusions: The –124 bp G>A hTERT promoter mutation was observed in the serum of 69% of HCC patients who underwent surgical resection and was an independent predictor of disease progression in HCC.

Soluble AXL as a marker of disease progression and survival in melanoma.

Receptor tyrosine kinase AXL is a one-pass transmembrane protein upregulated in cancers and associated with lower survival and therapy resistance. AXL can be cleaved by the A Disintegrin and Metalloproteinases (ADAM)10 and ADAM17, yielding a soluble version of the protein. Elevated soluble AXL (sAXL) has been reported to be associated with disease progression in hepatocellular carcinoma, renal cancer, neurofibromatosis type 1 and inflammatory diseases. In the present work, we analyzed sAXL levels in blood from melanoma patients and showed that sAXL increases with disease progression. Additionally, increased sAXL levels were found correlated with shorter two-year survival in stage IV patients treated with ipilimumab. Furthermore, we showed that sAXL levels were related to the percentage of cells expressing AXL in resected melanoma lymph node metastases. This finding was verified in vitro, where sAXL levels in the cell media corresponded to AXL expression in the cells. AXL inhibition using the small-molecular inhibitor BGB324 reduced sAXL levels, while the cellular expression was elevated through increased protein stability. Our findings signify that quantification of sAXL blood levels is a simple and easily assessable method to determine cellular AXL levels and should be further evaluated for its use as a biomarker of disease progression and treatment response.

Molecular mechanisms of circular RNAs, transforming growth factor-β, and long noncoding RNAs in hepatocellular carcinoma.

At the heart of hepatocellular carcinoma (HCC) lies disruption of signaling pathways at the level of molecules, genes, and cells. Non‐coding RNAs (ncRNAs) have been implicated in the disease progression of HCC. For instance, dysregulated expression of circular RNAs (circRNAs) has been observed in patients with HCC. As such, these RNAs are potential therapeutic targets and diagnostic markers for HCC. Long non‐coding RNAs (lncRNAs), a type of ncRNA, have also been recognized to participate in the initiation and progression of HCC. Transforming growth factor‐beta (TGF‐β) is another element which is now recognized to play crucial roles in HCC. It has been implicated in many biological processes such as survival, immune surveillance, and cell proliferation. In HCC, TGF‐β promotes disease progression by two mechanisms: an intrinsic signaling pathway and the extrinsic pathway. Through these pathways, it modulates various microenvironment factors such as inflammatory mediators and fibroblasts. An interesting yet‐to‐be resolved concept is whether the HCC‐promoting role of TGF‐β pathways is limited to a subset of HCC patients or it is involved in the whole process of HCC development. This review summarizes recent advancements to highlight the roles of circRNAs, lncRNAs, and TGF‐β in HCC.

Safety of Combined Yttrium-90 Radioembolization and Immune Checkpoint Inhibitor Immunotherapy for Hepatocellular Carcinoma.

To investigate the safety of yttrium-90 radioembolization in combination with checkpoint inhibitor immunotherapy for the treatment of hepatocellular carcinoma (HCC). This single-center retrospective study included 26 consecutive patients with HCC who received checkpoint inhibitor immunotherapy within 90 days of radioembolization from April 2015 to May 2018. Patients had preserved liver function (Child-Pugh scores A-B7) and either advanced HCC due to macrovascular invasion or limited extrahepatic disease (21 patients) or aggressive intermediate stage HCC that resulted in earlier incorporation of systemic immunotherapy (5 patients). Clinical documentation, laboratory results, and imaging results at 1- and 3-month follow-up intervals were reviewed to assess treatment-related adverse events and treatment responses. The median follow-up period after radioembolization was 7.8 months (95% confidence interval [CI], 5.6-11.8). There were no early (30-day) mortality or grades 3/4 hepatobiliary or immunotherapy-related toxicities. Delayed grades 3/4 hepatobiliary toxicities (1-3 months) occurred in 2 patients in the setting of HCC disease progression. One patient developed pneumonitis. The median overall survival from first immunotherapy was 17.2 months (95% CI, 10.9-23.4). The median overall survival from first radioembolization was 16.5 months (95% CI, 6.6-26.4). From first radioembolization, time to tumor progression was 5.7 months (95% CI, 4.2-7.2), and progression-free survival was 5.7 months (95% CI, 4.3-7.1). Radioembolization combined with checkpoint inhibitor immunotherapy in cases of HCC appears to be safe and causes limited treatment-related toxicity. Future prospective studies are needed to identify the optimal combination treatment protocols and evaluate the efficacy of combination therapy.

Aberrant histone deacetylase 1 expression upregulates vimentin expression via an NF‑κB‑dependent pathway in hepatocellular carcinoma

Aberrantly elevated expression levels of histone deacetylase 1 (HDAC1) and vimentin are closely associated with disease progression in hepatocellular carcinoma (HCC). It was previously demonstrated that knocking down expression of HDAC1 resulted in a concurrent decrease in the expression levels of vimentin. However, a causal link between these two proteins has not yet been demonstrated, to the best of our knowledge. In the present study, the association between HDAC1 and vimentin was investigated using an HDAC1 overexpression platform. HDAC1 and vimentin were significantly increased in HCC cells, and HDAC1 overexpression enhanced vimentin mRNA and protein expression levels in an HDAC1 dose-dependent manner. Subsequently, truncation and mutation of a vimentin promoter demonstrated that HDAC1-induced vimentin expression was dependent on a nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) binding site in the vimentin promoter sequence. Furthermore, HDAC1 induced vimentin expression by promoting NF-κB translocation between the cytoplasm and the nucleus, as opposed to modulating the total expression level of vimentin directly. The data in the present study demonstrated that HDAC1 is overexpressed in HCC and that HDAC1 may upregulate vimentin expression through the NF-κB signaling pathway, thus demonstrating a causal link between HDAC1 and vimentin in HCC, and may provide valuable information in understanding the pathogenesis of HCC.

CCL15 Recruits Suppressive Monocytes to Facilitate Immune Escape and Disease Progression in Hepatocellular Carcinoma.

Chemokines play a key role in orchestrating the recruitment and positioning of myeloid cells within the tumor microenvironment. However, the tropism regulation and functions of these cells in hepatocellular carcinoma (HCC) are not completely understood. Herein, by scrutinizing the expression of all chemokines in HCC cell lines and tissues, we found that CCL15 was the most abundantly expressed chemokine in human HCC. Further analyses showed that CCL15 expression was regulated by genetic, epigenetic, and microenvironmental factors, and negatively correlated with patient clinical outcome. In addition to promoting tumor invasion in an autocrine manner, CCL15 specifically recruited CCR1+ cells toward HCC invasive margin, approximately 80% of which were CD14+ monocytes. Clinically, a high density of marginal CCR1+ CD14+ monocytes positively correlated with CCL15 expression and was an independent index for dismal survival. Functionally, these tumor-educated monocytes directly accelerated tumor invasion and metastasis through bursting various pro-tumor factors and activating signal transducer and activator of transcription 1/3, extracellular signal-regulated kinase 1/2, and v-akt murine thymoma viral oncogene homolog signaling in HCC cells. Meanwhile, tumor-derived CCR1+ CD14+ monocytes expressed significantly higher levels of programmed cell death-ligand 1, B7-H3, and T-cell immunoglobulin domain and mucin domain-3 that may lead to immune suppression. Transcriptome sequencing confirmed that tumor-infiltrating CCR1+ CD14+ monocytes were reprogrammed to upregulate immune checkpoints, immune tolerogenic metabolic enzymes (indoleamine and arginase), inflammatory/pro-angiogenic cytokines, matrix remodeling proteases, and inflammatory chemokines. Orthotopic animal models confirmed that CCL15-CCR1 axis forested an inflammatory microenvironment enriched with CCR1+ monocytes and led to increased metastatic potential of HCC cells. Conclusion: A complex tumor-promoting inflammatory microenvironment was shaped by CCL15-CCR1 axis in human HCC. Blockade of CCL15-CCR1 axis in HCC could be an effective anticancer therapy.

Transcriptomic analysis of hepatocellular carcinoma reveals molecular features of disease progression and tumor immune biology

Hepatocellular carcinoma (HCC) develops in the context of chronic inflammatory liver disease and has an extremely poor prognosis. An immunosuppressive tumor microenvironment may contribute to therapeutic failure in metastatic HCC. Here, we identified unique molecular signatures pertaining to HCC disease progression and tumor immunity by analyzing genome-wide RNA-Seq data derived from HCC patient tumors and non-tumor cirrhotic tissues. Unsupervised clustering of gene expression data revealed a gradual suppression of local tumor immunity that coincided with disease progression, indicating an increasingly immunosuppressive tumor environment during HCC disease advancement. IHC examination of the spatial distribution of CD8+ T cells in tumors revealed distinct intra- and peri-tumoral subsets. Differential gene expression analysis revealed an 85-gene signature that was significantly upregulated in the peri-tumoral CD8+ T cell-excluded tumors. Notably, this signature was highly enriched with components of underlying extracellular matrix, fibrosis, and epithelial–mesenchymal transition (EMT). Further analysis condensed this signature to a core set of 23 genes that are associated with CD8+ T cell localization, and were prospectively validated in an independent cohort of HCC specimens. These findings suggest a potential association between elevated fibrosis, possibly modulated by TGF-β, PDGFR, SHH or Notch pathway, and the T cell-excluded immune phenotype. Indeed, targeting fibrosis using a TGF-β neutralizing antibody in the STAM™ model of murine HCC, we found that ameliorating the fibrotic environment could facilitate redistribution of CD8+ lymphocytes into tumors. Our results provide a strong rationale for utilizing immunotherapies in HCC earlier during treatment, potentially in combination with anti-fibrotic therapies.

Phosphorylation of SMC1A promotes hepatocellular carcinoma cell proliferation and migration.

Structural maintenance of chromosomes protein 1A (SMC1A) has been implicated in the development of a variety of cancer types. However, its role in hepatocellular carcinoma remains unknown. In this study, we found that phosphorylated SMC1A was highly expressed in HepG2 and Bel7402 cells when compared with other cancer cell lines. Furthermore, SMC1A knockdown dramatically reduced HepG2 and Bel7402 cell proliferation and migration. Re-expressing phosphomimetic mutants S957DS966D significantly enhanced the proliferation and migration of SMC1A knockdown HepG2 and Bel7402 cells. In addition, phosphorylated SMC1A promotes hepatocellular carcinoma cells growth in vivo. Importantly, the expression of phosphorylated SMC1A was significantly higher in human hepatocellular carcinomacells when compared to peri-tumor benign hepatocytes, and its overexpression was significantly associated with worse prognostic outcomes. These observations suggest that phosphorylation of SMC1A is a vital event in tumorigenesis and disease progression in hepatocellular carcinoma thus necessitating further investigation.

DANCR Acts as a Diagnostic Biomarker and Promotes Tumor Growth and Metastasis in Hepatocellular Carcinoma.

Long non-coding RNA differentiation antagonizing non-protein coding RNA (DANCR) is considered to be an oncogene in many cancer types. However, the role of DANCR in diagnosis and progression of hepatocellular carcinoma (HCC) is still unknown. In this study, 146 participants including healthy volunteers (HVs) and patients with chronic hepatitis B (CHB), cirrhosis and HCC were recruited. Firstly, quantitative reverse transcription-PCR analysis showed that DANCR was up-regulated in tumor tissues and plasma of patients with HCC, and its expression was highly correlated with microvascular and liver capsule invasion of HCC. Receiver operating characteristic analysis showed that plasma DANCR exhibited significantly increased discriminatory power for differentiating patients with HCC from HVs and non-HCC patients compared to alpha fetoprotein, which has been used as a biomarker for HCC diagnosis. Furthermore, knockdown of DANCR repressed the β-catenin pathway and inhibited HCC cell proliferation and metastasis both in vitro and in vivo. This study revealed, for the first time, the importance of DANCR in clinical diagnosis and disease progression of HCC.