CircDCAF8 promotes the progression of hepatocellular carcinoma through miR-217/NAP1L1 Axis, and induces angiogenesis and regorafenib resistance via exosome-mediated transfer.

BACKGROUND: Circular RNAs (circRNAs), which are a new type of single-stranded circular RNA, have significant involvement in progression of many diseases, including tumors. Currently, multiple circRNAs have been identified in hepatocellular carcinoma (HCC). Our study aims to investigate the function and mechanism of circDCAF8 in HCC. METHODS: The expression of circDCAF8 (hsa_circ_0014879) in HCC and para-carcinoma tissue samples was determined using quantitative real-time polymerase chain reaction (qRT-PCR). The biological function of circDCAF8 in HCC was confirmed by experiments conducted both in vitro and in vivo. And the relationship between circDCAF8, miR-217 and NAP1L1 was predicted by database and verified using qRT-PCR, RNA-binding protein immunoprecipitation (RIP) and dual-luciferase reporter assays. Exosomes isolated from HCC cells were utilized to assess the connection of exosomal circDCAF8 with HCC angiogenesis and regorafenib resistance. RESULTS: CircDCAF8 is upregulated in HCC tissues and cell lines, and is linked to an unfavourable prognosis for HCC patients. Functionally, circDCAF8 was proved to facilitate proliferation, migration, invasion and Epithelial-Mesenchymal Transformation (EMT) in HCC cells. Animal examinations also validated the tumor-promoting characteristics of circDCAF8 on HCC. Besides, exosomal circDCAF8 promoted angiogenesis in HUVECs. Mechanistically, circDCAF8 interacted with miR-217 and NAP1L1 was a downstream protein of miR-217. CircDCAF8 promoted NAP1L1 expression by sponging miR-217. In addition, exosomes may transfer circDCAF8 from regorafenib-resistant HCC cells to sensitive cells, where it would confer a resistant phenotype. CONCLUSION: CircDCAF8 facilitates HCC proliferation and metastasis via the miR-217/NAP1L1 axis. Meanwhile, circDCAF8 can promote angiogenesis and drive resistance to regorafenib, making it a viable therapeutic target for HCC patients.

M2 macrophage-secreted exosomes promote metastasis and increase vascular permeability in hepatocellular carcinoma.

BACKGROUND: Metastasis is a key feature of malignant tumors and significantly contributes to their high mortality, particularly in hepatocellular carcinoma (HCC). Therefore, it is imperative to explore the mechanism of tumor metastasis. Recently, tumor-associated macrophages (TAMs) have been demonstrated to promote tumor progression, while TAM-derived molecules involved in HCC metastasis warrant further investigation. METHODS: THP-1 was treated with IL-4 (Interleukin-4) and IL-13 (Interleukin-13) for M2 polarized macrophages. Exosomes derived from M2 macrophages were characterized. Then, HCC cells or human umbilical vein endothelial cells (HUVECs) were co-cultured with M2 macrophages or treated with M2 macrophage-secreted exosomes. Next, Transwell®, Scratch assay, tube formation, and endothelial permeability assays were performed. Moreover, RT-PCR, western blotting, immunofluorescence, and ELISA were used to assess mRNA and protein expression levels. Finally, the miRNA expression profiles of exosomes derived from M2 and M0 macrophages were analyzed. RESULTS: M2 macrophage infiltration was correlated with metastasis and a poor prognosis in HCC patients. M2-derived exosomes were absorbed by HCC and HUVEC cells and promoted the epithelial-mesenchymal transition (EMT), vascular permeability, and angiogenesis. Notably, MiR-23a-3p levels were significantly higher in M2-derived exosomes and hnRNPA1 mediated miR-23a-3p packaging into exosomes. Phosphatase and tensin homolog (PTEN) and tight junction protein 1 (TJP1) were the targets of miR-23a-3p, as confirmed by luciferase reporter assays. Lastly, HCC cells co-cultured with M2-derived exosomes secreted more GM-CSF, VEGF, G-CSF, MCP-1, and IL-4, which in turn further recruited M2 macrophages. CONCLUSIONS: Our findings suggest that M2 macrophage-derived miR-23a-3p enhances HCC metastasis by promoting EMT and angiogenesis, as well as increasing vascular permeability. Video Abstract.

M2 Macrophage-Derived Exosomes Facilitate HCC Metastasis by Transferring αM ß2 Integrin to Tumor Cells.

BACKGROUND AND AIMS: The development and progression of hepatocellular carcinoma (HCC) is dependent on its local microenvironment. Tumor-associated macrophages (TAMs) are deemed a key factor for the tumor microenvironment and attribute to contribute to tumor aggressiveness. However, the detailed mechanism underlying the pro-metastatic effect of TAMs on HCC remains undefined. APPROACH AND RESULTS: The present study proved that TAMs were enriched in HCC. TAMs were characterized by an M2-polarized phenotype and accelerated the migratory potential of HCC cells in vitro and in vivo. Furthermore, we found that M2-derived exosomes induced TAM-mediated pro-migratory activity. With the use of mass spectrometry, we identified that integrin, αM ß2 (CD11b/CD18), was notably specific and efficient in M2 macrophage-derived exosomes (M2 exos). Blocking either CD11b and/or CD18 elicited a significant decrease in M2 exos-mediated HCC cell metastasis. Mechanistically, M2 exos mediated an intercellular transfer of the CD11b/CD18, activating the matrix metalloproteinase-9 signaling pathway in recipient HCC cells to support tumor migration. CONCLUSIONS: Collectively, the exosome-mediated transfer of functional CD11b/CD18 protein from TAMs to tumor cells may have the potency to boost the migratory potential of HCC cells, thus providing insights into the mechanism of tumor metastasis.

5-Hydroxytryptamine Receptor 1D Aggravates Hepatocellular Carcinoma Progression Through FoxO6 in AKT-Dependent and Independent Manners.

Serotonin and its receptors have been shown to play critical regulatory roles in cancer biology. Nevertheless, the contributions of 5-hydroxytryptamine 1D (5-HT1D), an indispensable member of the serotonergic system, to hepatocellular carcinoma (HCC) remain unknown. The present study demonstrated that the 5-HT1D expression level was significantly up-regulated in HCC tissues and cell lines. The 5-HT1D expression level was closely correlated with unfavorable clinicopathological characteristics. Survival analyses show that elevated 5-HT1D expression level predicts poor overall survival and high recurrence probability in HCC patients. Functional studies revealed that 5-HT1D significantly promoted HCC proliferation, epithelial-mesenchymal transition, and metastasis in vitro and in vivo. Mechanistically, 5-HT1D could stabilize PIK3R1 by inhibiting its ubiquitin-mediated degradation. The interaction between 5-HT1D and phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) enhanced the expression of FoxO6 through the PI3K/Akt signaling pathway; FoxO6 could also be directly transcriptionally activated by 5-HT1D in an Akt-independent manner. MicroRNA-599 was found to be an upstream suppressive modulator of 5-HT1D. Additionally, 5-HT1D could attenuate tryptophan hydroxylase 1 expression through the PI3K/Akt/cut-like homeobox 1 axis in HCC. Conclusion: Herein, we uncovered the potent oncogenic effect of 5-HT1D on HCC by interacting with PIK3R1 to activate the PI3K/Akt/FoxO6 pathway, and provided a potential therapeutic target for HCC.

circLARP4 induces cellular senescence through regulating miR-761/RUNX3/p53/p21 signaling in hepatocellular carcinoma.

Circular RNAs (circRNAs), a novel class of non-coding RNAs, have emerged as indispensable modulators in human malignancies. Aberrant cellular senescence is a phenotype observed in various cancers. The association of circRNAs with cellular senescence in tumors is yet to determined. Here, we investigated the role of circLARP4 in cellular senescence and cell proliferation in hepatocellular carcinoma (HCC). Downregulated circLARP4 level was observed in HCC tissues and cell lines. Low expression level of circLARP4 independently predicted poor survival outcome. Gain-of-function and loss-of-function assays demonstrated that circLARP4 suppressed HCC cell proliferation, mediated cell cycle arrest and induced senescence in vitro. Levels of p53 and p21, 2 key regulatory molecules in cellular senescence, were increased in circLARP4-overexpressed HCC cells and decreased in circLARP4-silenced HCC cells. In vivo experiments further confirmed the tumor-suppressing activity of circLARP4. Further mechanistic studies showed that circLARP4 dampened HCC progression by sponging miR-761, thereby promoting the expression level of RUNX3 and activating the downstream p53/p21 signaling. Our study revealed the role of circLARP4/miR-761/RUNX3/p53/p21 signaling in HCC progression, providing a potential survival predictor and therapeutic candidate for HCC.

PRDM8 exhibits antitumor activities toward hepatocellular carcinoma by targeting NAP1L1.

Hepatocellular carcinoma (HCC) is a major leading cause of cancer mortality worldwide. PRDI-BF1 and RIZ homology domain containing 8 (PRDM8) is a key regulator in neural development and testis steroidogenesis; however, its role in liver carcinogenesis remains to be investigated. In this study, PRDM8 was found to be down-regulated in HCC, which was linked with shorter recurrence-free survival. Lentiviral-based overexpression and knockdown approaches showed that PRDM8 inhibited HCC cell proliferation, migration, and invasion. PRDM8 caused G1/S cell cycle arrest and induced apoptosis. An in vivo tumor model confirmed the antitumor role of PRDM8 in HCC growth and metastasis. Mechanistic study showed that PRDM8 suppressed the PI3K/AKT/mTOR signaling cascade through the regulation of nucleosome assembly protein 1-like 1 (NAP1L1). Conclusion: PRDM8 as a functional tumor suppressor is frequently down-regulated in HCC. Through regulating NAP1L1, PRDM8 inhibits PI3K/AKT/mTOR signaling in HCC. PRDM8 is a potential target for therapies of HCC. (Hepatology 2018).

MicroRNA-873 Promotes Cell Proliferation, Migration, and Invasion by Directly Targeting TSLC1 in Hepatocellular Carcinoma.

BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and has the third highest mortality rate among all cancers. MicroRNAs are a class of endogenous, single-stranded short noncoding RNAs. The purpose of this study was to study the role of microRNA-873 in HCC. METHODS: The expression of miRNA-873 and tumor suppressor in lung cancer 1 (TSLC1) in HCC tissues and cell lines was detected by real-time quantitative RT-PCR (RT-qPCR) or western blot. A CCK-8 assay was used to examine cell proliferation; flow cytometry was used to assess the cell cycle; the Transwell migration assay was used to test for metastasis. Luciferase assays were performed to assess whether TSLC1 was a novel target of miRNA-873. RESULTS: We showed that miRNA-873 was upregulated in HCC tissues and cell lines compared with the normal control. Knockdown of miRNA-873 inhibited the growth and metastasis of HepG2 and accelerated G1 phase arrest, while overexpression of miRNA-873 had the opposite effect. The dual-luciferase reporter assays revealed that TSLC1 was a novel target of miRNA-873. Further study showed that TSLC1 was decreased in HCC tissues and cell lines. There was a negative correlation between the expression levels of TSLC1 and miRNA-873. The effect of miRNA-873 overexpression was neutralized by TSLC1. We also found that miRNA-873 activated the PI3K/AKT/mTOR signaling pathway and promoted HCC. CONCLUSIONS: Our data demonstrated that miRNA-873 promoted HCC progression by targeting TSLC1 and provided a new target for the therapy of HCC.

A Meta-Analysis of the Diagnostic Accuracy of Circular RNAs in Digestive System Malignancy.

BACKGROUND/AIMS: Circular RNAs (circRNAs), a novel class of noncoding RNAs, have been found to be dysregulated in various cancers. However, the clinical application value of these circRNAs in digestive system cancers remains to be clarified. We aimed to comprehensively explore the potential role of circRNAs as diagnostic indicators in digestive system malignancies. METHODS: Relevant studies were systematically retrieved from PubMed, Web of Science and the Cochrane Library. The data that were required to complete 2 × 2 contingency tables were obtained from the included studies. Stratified analyses by cancer type, sample size and publication year were performed. RESULTS: Thirteen studies with 2,276 individuals were included in the meta-analysis. The pooled sensitivity and specificity of circRNAs in the diagnosis of digestive system malignancy were 0.72 [95% confidence interval (CI): 0.65-0.77] and 0.77 (95% CI: 0.72-0.81), respectively. The overall positive likelihood ratio was 3.09 (95% CI: 2.64-3.62), and the overall negative likelihood ratio was 0.37 (95% CI: 0.31-0.44). The pooled diagnostic odds ratio was 8.38 (95% CI: 6.86-10.25), and the overall area under the curve was 0.81 (95% CI: 0.77-0.84), indicating good discriminative ability of circRNAs as biomarkers for digestive system malignancy. CONCLUSION: circRNAs distinguish patients with digestive system cancer from controls with relatively high diagnostic accuracy. circRNAs may be used as potential biomarkers for the diagnosis of digestive system malignancy.

Role of tumor-associated macrophages in hepatocellular carcinoma: impact, mechanism, and therapy.

Hepatocellular carcinoma (HCC) is a highly frequent malignancy worldwide. The occurrence and progression of HCC is a complex process closely related to the polarization of tumor-associated macrophages (TAMs) in the tumor microenvironment (TME). The polarization of TAMs is affected by a variety of signaling pathways and surrounding cells. Evidence has shown that TAMs play a crucial role in HCC, through its interaction with other immune cells in the TME. This review summarizes the origin and phenotypic polarization of TAMs, their potential impacts on HCC, and their mechanisms and potential targets for HCC immunotherapy.

Derivation and validation of machine learning models for preoperative estimation of microvascular invasion risk in hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) represents a considerable burden to patients and health systems. Microvascular invasion (MVI) is a significant risk factor for HCC recurrence and survival after hepatectomy. We aimed to establish a preoperative MVI prediction model based on readily available clinical and radiographic characteristics using machine learning algorithms. Methods: Two independent cohorts of patients with HCC who underwent hepatectomy were included in the analysis and divided into a derivation set (466 patients), an internal validation set (182 patients), and an external validation set (140 patients). Least absolute shrinkage and selection operator (LASSO) analysis was used to optimize variable selection. We constructed the MVI prediction model using several machine learning algorithms, including logistic regression, k-nearest neighbors, support vector machine, decision tree, random forest, extreme gradient boosting, and neural network. Performance of the model was assessed in terms of discrimination, calibration, and clinical usefulness. Results: The three most significant variables associated with MVI-α-fetoprotein, protein induced by vitamin K absence or antagonist-II, and tumor size-were identified by the LASSO analysis. Among the machine learning algorithms, the logistic regression model achieved the largest area under the receiver operating characteristic curve and was presented in the form of a user-friendly, online calculator. The concordance (C)-statistic of the model was 0.745 [95% confidence interval (CI): 0.701-0.790] for the derivation set, 0.771 (95% CI: 0.703-0.839) for the internal validation set, and 0.812 (95% CI: 0.734-0.891) for the external validation set. The Hosmer-Lemeshow calibration test and calibration plot indicated a good fit for all 3 data sets. Decision curve analysis showed the model was clinically useful. Conclusions: This study provided a convenient and explainable approach for MVI prediction before surgical intervention. Our model may assist clinicians in determining the optimal therapeutic modality and facilitate precision medicine for HCC.

Hypoxia-associated circPRDM4 promotes immune escape via HIF-1α regulation of PD-L1 in hepatocellular carcinoma.

BACKGROUND: Hypoxia is a hallmark of cancer, and is closely intertwined with tumor immune evasion. Circular RNAs (circRNAs) have been implicated in tumor response to immune checkpoint blockades. However, hypoxia-associated circRNAs that orchestrate the association between hypoxia and response to immunotherapy remain poorly understood. Here, we aimed to determine the roles of hypoxia-associated circRNAs in immune escape of hepatocellular carcinoma (HCC) cells. METHODS: Differentially expressed hypoxia-associated circRNAs were determined using high-throughput sequencing technology. HCC patients treated with PD-1 blockade were enrolled to assess the clinical significance of circPRDM4. RT-qPCR, western blotting, flow cytometry, T cell-mediated tumor cell killing assay, and enzyme linked immunosorbent assay were used to investigate the roles of circPRDM4 in immune escape of HCC cells in vitro. Patient-derived xenograft mouse models and adoptive human tumor infiltrating lymphocyte-CD8+ T cell transfer were adopted to evaluate the effects of circPRDM4 in vivo. RNA pull-down, mass spectrometry, RNA immunoprecipitation, chromatin immunoprecipitation, chromatin isolation by RNA purification, dual-luciferase reporter assays, dot blotting, DNA in situ hybridization, and immunoprecipitation were utilized to examine the interaction between circPRDM4, HIF-1α, and CD274 promoter. RESULTS: We identified circPRDM4 as a hypoxia-associated circRNA in HCC. circPRDM4 was upregulated in responders to PD-1 blockade and associated with therapeutic efficacy. In vitro and in vivo experiments showed that circPRDM4 induced PD-L1 expression and promoted CD8+ T cell-mediated immune escape under hypoxic conditions. Mechanistically, circPRDM4 acted as a scaffold to recruit HIF-1α onto CD274 promoter, and cemented their interaction, ultimately promoting the HIF-1α-mediated transactivation of PD-L1. CONCLUSIONS: These findings illustrated that circPRDM4 promoted immune escape of HCC cells by facilitating the recruitment of HIF-1α onto the promoter of CD274 under hypoxia, thereby inhibiting CD8+ T cell infiltration in the tumor microenvironment. This work may provide a novel prognostic biomarker and therapeutic candidate for HCC immunotherapy.

E2F1-mediated GINS2 transcriptional activation promotes tumor progression through PI3K/AKT/mTOR pathway in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) has high morbidity and mortality rates. It is therefore imperative to study the underlying mechanism of HCC to identify potential prognostic biomarkers and therapeutic targets. Recently, GINS2 has been identified to be a cancer-promoting gene in different cancer types. Nevertheless, the exact mechanism of GINS2 in HCC remains to be elucidated. To systematically explore the significance of GINS2, we first assessed the relative expression of GINS2 in pan-cancers based on data obtained from the HCCDB, TIMER, and TCGA databases. Then, we explored the clinical significance of GINS2 in HCC through Kaplan-Meier method as well as univariate and multivariate cox regression analysis. Additionally, functional enrichment analysis of GINS2 was done through GO, KEGG, PPI network, and immune cell infiltration analyses. Functional experiments were also conducted to investigate the biological significance of GINS2 in HCC cell lines. Our research revealed that GINS2 is involved in HCC progression and highlighted its potential value as a crucial diagnostic and therapeutic target for HCC.

Development and Validation of a Novel Model to Predict Regional Lymph Node Metastasis in Patients With Hepatocellular Carcinoma.

BACKGROUND: The evaluation of the nodal status of hepatocellular carcinoma (HCC) is a classic but controversial topic. This study aimed to investigate the incidence of lymph node metastasis (LNM), explore the role of lymph node dissection (LND), and develop and validate a novel model to predict LNM in patients with HCC, not other specified (NOS). METHODS: The study cohort was taken from the Surveillance, Epidemiology, and End Results database. The annual percent change (APC) was calculated using the Joinpoint regression. Survival analyses adopted the competing risk model. The nomogram was constructed based on the least absolute shrinkage and selection operator (LASSO) logistic regression algorithm and validated by calibration curves. The area under the receiver operating characteristic curve (AUROC) was obtained to compare prognostic performance. Decision curve and clinical impact curve analyses were introduced to examine the clinical value of the models. RESULTS: A total of 8,829 patients were finally enrolled in this study, and 1,346 (15.2%) patients received LND. The LND rate showed no noticeable fluctuation in the last decade, with an APC of 0.5% (P=0.593). LNM was identified in 56 (4.2%) patients and confirmed an independent prognostic factor of HCC patients (P=0.005). There were 2,497 lymph nodes retrieved, and 93 (3.7%) of them were positive. After propensity score matching, LND indicated no direct oncologic benefit and did not worsen competing risks. Moreover, an increased number of lymph nodes retrieved could not improve prognoses. 1,346 patients with LND were further randomly divided into the training and validation sets with the ratio of 1:1. Race, tumor size, clinical T stage, extrahepatic bile duct invasion, and tumor grade were independent risk factors for LNM. The constructed model was well calibrated and showed good discrimination power and net benefits in clinical practice. CONCLUSION: LNM is an independent prognostic factor in HCC, but routine LND seems to be unnecessary in HCC patients. The constructed model could predict the presence of LNM in HCC patients with good performance, which is meaningful to patient stratification and individual treatment strategies optimization.

CircPAK1 promotes the progression of hepatocellular carcinoma via modulation of YAP nucleus localization by interacting with 14-3-3ζ.

BACKGROUND: Circular RNA (circRNA), a new class of non-coding RNA, has obvious correlations with the occurrence and development of many diseases, including tumors. This study aimed to investigate the potential roles of circPAK1 in hepatocellular carcinoma (HCC). METHODS: High-throughput sequencing was performed on 3 pairs of HCC and matched normal tissues to determine the upregulated circRNAs. The expression level of circPAK1 was detected by qRT-PCR in HCC and paired with normal liver tissue samples. The effects of circPAK1 on proliferation, invasion, metastasis and apoptosis of HCC cells were evaluated by in vitro and in vivo experiments. We also constructed Chitosan/si-circPAK1 (CS/si-circPAK1) nanocomplexes using Chitosan material to evaluate its in vivo therapeutic effect on HCC. High-throughput sequencing, RNA-sequencing, RNA probe pull-down, RNA immunoprecipitation and Co-Immunoprecipitation assays were performed to explore the relationship between circPAK1, 14-3-3ζ, p-LATS1 and YAP. Exosomes isolated from lenvatinib-resistant HCC cell lines were used to evaluate the relationship between exosomal circPAK1 and lenvatinib resistance. RESULTS: CircPAK1, a novel circRNA, is highly expressed in HCC tumor tissues and cell lines as well as correlated with poor outcomes in HCC patients. Functionally, circPAK1 knockdown inhibited HCC cell proliferation, migration, invasion and angiogenesis while circPAK1 overexpression promoted HCC progression. The tumor-promoting phenotypes of circPAK1 on HCC were also confirmed by animal experiments. Importantly, the application of CS/si-circPAK1 nanocomplexes showed a better therapeutic effect on tumor growth and metastasis. Mechanistically, circPAK1 enhanced HCC progression by inactivating the Hippo signaling pathway, and this kind of inactivation is based on its competitively binding of 14-3-3 ζ with YAP, which weakens the recruitment and cytoplasmic fixation of 14-3-3 ζ to YAP, thus promoting YAP nucleus localization. Additionally, circPAK1 could be transported by exosomes from lenvatinib-resistant cells to sensitive cells and induce lenvatinib resistance of receipt cells. CONCLUSION: CircPAK1 exerts its oncogenic function by competitively binding 14-3-3 ζ with YAP, thus promoting YAP nucleus localization, leading to the inactivation of a Hippo signaling pathway. Exosomal circPAK1 may drive resistance to lenvatinib, providing a potential therapeutic target for HCC patients.

Epidemiological and Clinical Characteristics of Five Rare Pathological Subtypes of Hepatocellular Carcinoma.

Background: Hepatocellular carcinoma (HCC) is a highly heterogeneous tumor with several rare pathological subtypes and which is still poorly understood. This study aimed to describe the epidemiological and clinical spectrum of five rare HCC subtypes and develop a competing risk nomogram for cancer-specific survival prediction. Methods: The study cohort was recruited from the Surveillance, Epidemiology, and End Results database. The clinicopathological data of 50,218 patients histologically diagnosed with classic HCC and five rare subtypes (ICD-O-3 Histology Code = 8170/3-8175/3) between 2004 and 2018 were reviewed. The annual percent change (APC) was calculated utilizing Joinpoint regression. The nomogram was developed based on multivariable competing risk survival analyses. Akaike information criterion, Bayesian information criterion, C-index, calibration curve, and area under the receiver operating characteristic curve were obtained to evaluate the prognostic performance. A decision curve analysis was introduced to examine the clinical value of the models. Results: Despite scirrhous carcinoma, which showed a decreasing trend (APC = -6.8%, P = 0.025), the morbidity of other rare subtypes remained stable from 2004 to 2018. The incidence-based mortality was plateau in all subtypes during the period. Clear cell carcinoma is the most common subtype (n = 551, 1.1%), followed by subtypes of fibrolamellar (n = 241, 0.5%), scirrhous (n = 82, 0.2%), spindle cell (n = 61, 0.1%), and pleomorphic (n = 17, ~0%). The patients with fibrolamellar carcinoma were younger and more likely to have a non-cirrhotic liver and better prognoses. Scirrhous carcinoma shared almost the same macro-clinical characteristics and outcomes as the classic HCC. Clear cell carcinoma tended to occur in the Asia-Pacific elderly male population, and more than half of them were large HCC (Size>5cm). Sarcomatoid (including spindle cell and pleomorphic) carcinoma was associated with a larger tumor size, poorer differentiation, and more dismal prognoses. The pathological subtype, T stage, M stage, surgery, alpha-fetoprotein, and cancer history were confirmed as the independent predictors in patients with rare subtypes. The nomogram showed good calibration, discrimination, and net benefits in clinical practice. Conclusion: The rare subtypes had unique clinicopathological features and biological behaviors compared with the classic HCC. Our findings could provide a valuable reference for clinicians. The constructed nomogram could predict the prognoses with good performance, which is meaningful to individualized management.

Promising role of liver transplantation in patients with combined hepatocellular-cholangiocarcinoma: a propensity score matching analysis.

Background: Combined hepatocellular-cholangiocarcinoma (CHC) is a rare but vital heterogeneous histological subtype of primary liver cancer (PLC) with no standardized treatment strategy. This study aimed to preliminarily investigate the role of liver transplantation (LT) in CHC and develop a novel risk scoring model (RSM) to evaluate the benefits of transplantation. Methods: The study cohort was taken from the Surveillance, Epidemiology, and End Results database. The annual percent change (APC) in incidence or ratio was calculated utilizing the Joinpoint regression. Propensity score matching (PSM) was introduced to reduce the selection bias between groups. A novel RSM was developed based on the independent prognostic factors identified by the Cox regression model. The predictive performance of the RSM was compared with the Milan Criteria and the University of California, San Francisco (UCSF) Criteria, respectively. Results: A total of 223 CHC patients were enrolled, and 60 (26.9%) of them received LT. The incidence-based mortality did not decrease between 2004 and 2015 (APC =1.7%, P=0.195). Although LT was considered an independent protective predictor for CHC, it showed a declining ratio from 33.3% in 2004 to 15.4% in 2015 (APC =-8.9%, P=0.012). The LT recipients had better outcomes than others who underwent hepatectomy or local destruction (P<0.05). Compared with other subtypes of PLC, the post-transplantation prognoses of CHC patients were similar to those with hepatocellular carcinoma (P>0.05) but significantly better than those with intrahepatic cholangiocarcinoma (ICC) (P<0.05). Based on the RSM (vascular invasion: 1 point; tumor size >2 cm: 1 point; multiple tumors: 2 points), patients were stratified into two prognostic subgroups: the low-risk (scoring ≤2) and the high-risk (scoring >2 or extrahepatic metastasis) groups. Patients in the low-risk group were more likely to benefit from LT. The predictive performance of the RSM outperformed the Milan and UCSF Criteria in both the training and validation sets. Conclusions: Therapeutic strategies for CHC should be further improved. Patients with CHC should also be considered potential LT candidates. The novel RSM could be helpful to stratify patients and assist clinical decision-making.

Immune checkpoint inhibitors plus capecitabine and oxaliplatin in unresectable or advanced biliary tract cancer patients: A retrospective study.

Objective: Immune checkpoint inhibitors (ICIs) have recently been increasingly used in cancer treatment, whereas their clinical application in biliary tract cancer (BTC) patients is uncommon. This study aimed to evaluate the efficacy and safety of ICIs plus capecitabine and oxaliplatin (CAPOX) in the treatment of BTC patients. Methods: This retrospective study reviewed 26 unresectable or advanced BTC patients who received ICIs plus CAPOX. The treatment continued until disease progression, uncontrollable adverse event (AE) occurrence, intolerable toxicity occurrence, or voluntary withdrawal. Results: The median treatment cycles were 5.5 [interquartile range (IQR): 3.8-8.0]. Complete response, partial response, stable disease, and progressive disease rates were 0.0%, 46.2%, 23.1%, and 30.8%, respectively. Objective response rate (ORR) and disease control rate (DCR) were 46.2% and 69.2%, correspondingly. Regarding survival, the median progression-free survival (PFS) and overall survival (OS) were 6.1 (95% CI: 4.4-7.7) months and 16.5 (95% CI: 5.0-28.0) months; moreover, the 1-year PFS and OS rates were 21.5% and 54.3%, respectively. An Eastern Cooperative Oncology Group (ECOG) score of 1-3 (vs. 0) was associated with declined DCR, PFS, and OS (all p < 0.050). The most common AEs of ICIs plus CAPOX were thrombocytopenia (61.5%), neutropenia (26.9%), and reactive cutaneous capillary endothelial proliferation (RCCEP) (23.1%). Moreover, 13 (50.0%) patients suffered from grade 3-4 AEs, including thrombocytopenia (50.0%), neutropenia (7.7%), liver dysfunction (7.7%), and RCCEP (3.8%). Notably, the majority of AEs were controllable. Conclusion: ICIs plus CAPOX chemotherapy exhibit a good efficacy and a manageable safety profile in the treatment of patients with unresectable or advanced BTC.

FER Regulated by miR-206 Promotes Hepatocellular Carcinoma Progression via NF-κB Signaling.

OBJECTIVES: Feline sarcoma-related protein (FER) is known to play a critical regulatory role in several carcinomas. However, the exact biological function of FER in hepatocellular carcinoma (HCC) still needs to be investigated. The primary objective of this research was to investigate the unknown function and molecular mechanisms of FER in HCC. MATERIALS AND METHODS: The expression level of FER in HCC tissue samples and cells was examined by RT-qPCR, immunohistochemistry and western blot. Cellular and animal experiments were used to explore the effect of FER on the proliferative and metastatic capacities of HCC cells. The crosstalk between FER and NF-κB signaling was explored by western blot. The upstream factors that regulate FER were evaluated through dual-luciferase experiments and western blot assays. RESULTS: FER was overexpressed in HCC specimens and HCC cell lines. FER expression levels were positively associated with unfavorable clinicopathological characteristics. The higher the expression of FER was, the worse the overall survival of HCC patients was. The results of loss-of-function and gain-of-function experiments indicated that knockdown of FER decreased, while overexpression of FER increased, the proliferation, invasion and metastasis of HCC cells in vitro and in vivo. Mechanistically, we found that FER activated the NF-κB signaling pathway and stimulated epithelial-to-mesenchymal transition (EMT). We also found that FER was directly regulated by miR-206, and the downregulation of miR-206 was associated with proliferation and metastatic progression in HCC. CONCLUSIONS: The present research was the first to reveal that a decrease in miR-206 levels results in an increase in FER expression in HCC, leading to enhanced cell growth and metastatic abilities via activation of the NF-κB signaling pathway.

Morbidity, Prognostic Factors, and Competing Risk Nomogram for Combined Hepatocellular-Cholangiocarcinoma.

BACKGROUND: Combined hepatocellular-cholangiocarcinoma (CHC) is a rare and heterogeneous histological subtype of primary liver cancer, which is still poorly understood. This study aimed to describe the epidemiological and clinical features, investigate the prognostic indicators, and develop a competing risk nomogram for CHC. METHODS: The study cohort was taken from the Surveillance, Epidemiology, and End Results database. The annual percent change (APC) in incidence was calculated using the joinpoint regression. The nomogram was developed based on multivariate competing risk survival analyses and validated by calibration curves. Akaike information criterion, Bayesian information criterion, Harrell's C-index, and area under the receiver operating characteristic curves were obtained to compare prognostic performance. Decision curve analysis was introduced to examine the clinical value of the models. RESULTS: The overall incidence of CHC was 0.062 per 100,000 individuals in 2004 and 0.081 per 100,000 individuals in 2018, with an APC of 1.0% (P > 0.05). CHC displayed intermediate clinicopathological features of hepatocellular carcinoma and intrahepatic cholangiocarcinoma. Race, tumor size, vascular invasion, extrahepatic invasion, distant metastasis, grade, surgery, and Metavir stage were confirmed as the independent predictors of cancer-specific survival. The constructed nomogram was well calibrated, which showed better discrimination power and higher net benefits than the current American Joint Committee on Cancer staging system. Patients with liver transplantation had better survival than those with hepatectomy, especially patients within the Milan Criteria (P=0.022 and P=0.015). There was no survival difference between liver transplantation and hepatectomy in patients beyond the Milan Criteria (P=0.340). CONCLUSION: The morbidity of CHC remained stable between 2004 and 2018. The constructed nomogram could predict the prognosis with good performance, which was meaningful to individual treatment strategies optimization. CHC patients should also be considered as potential liver transplantation recipients, especially those within the Milan Criteria, but the finding still needs more evidence to be further confirmed.

STK39 enhances the progression of Cholangiocarcinoma via PI3K/AKT pathway.

Serine/threonine kinase 39 (STK39) is overexpressed in various tumor tissues and plays an essential role in tumor progression. In this study, we investigated the clinical value, as well as the potential functions and mechanisms of STK39 in cholangiocarcinoma (CCA). The results showed that STK39 was overexpressed in CCA and negatively associated with the prognosis of patients with CCA. Functionally, STK39 knockdown suppressed cell proliferation, migration, and invasion, while STK39 overexpression facilitated tumor aggressiveness. The tumor-promoting effects of STK39 in CCA were also validated by in vivo experiments. Mechanistically, RNA-seq analysis identified that STK39 enhanced the progression of CCA by activating PI3K/AKT signaling pathway. Furthermore, overexpression of STK39 could induce gemcitabine resistance in CCA cells. Moreover, the increased expression of STK39 may be mediated by the dysregulation of miR-26a-5p. In summary, STK39 could be served as a valuable prognostic candidate and a potential therapeutic target of CCA.