Transition In Hepatocellular Carcinoma Research Articles

Deciphering the cross-talk between miRNA and tight junctions in hepatocellular carcinoma

The most common type of liver cancer is hepatocellular carcinoma (HCC) causes a lower survival rate even after systemic treatment. Previous studies have shown evidence that various molecular and epigenetic mechanisms are involved in the transition of HCC from normal liver cells. Epigenetics plays an important role in maintaining genomic stability in normal liver cells. Apart from the mutation of genes, epigenetic factors are involved in HCC progression. miRNA tends to be a major epigenetic factor involved in regulating major cell cycle pathways. miRNA regulates the HCC progression by inhibiting the major apoptotic pathways and favors angiogenesis and tumor microenvironment. Apart from regulating major pathways, miRNA appears to regulate tight junction integrity. Tight junction proteins appear to be strong barrier proteins involved in cell adhesion and integrity. Disturbance in cell adhesion and integrity leads to the major dysregulation of cell cycle resulting in cancer progression. Moreover, dysregulation of tight junction integrity was observed in the pathogenesis of HCC. The regulation of tight junction proteins via miRNA were studied in various diseases. Here, we discussed the regulation of tight junction proteins via miRNA in HCC progression. This review may focus on the significance of miRNA in cellular function and its role in regulating tight junction proteins that impact in HCC progression which opens a new approach to develop a new diagnostic marker for an early detection of HCC and novel therapeutic approach against HCC.

Abstract 1637: EPHB4 is a key regulator of G1/S transition in hepatocellular carcinoma

Abstract Effective therapeutic regimens against advanced hepatocellular carcinoma (HCCs) have recently improved but it remains very limited. Targeting receptor tyrosine kinases (RTKs) has proven to one of the main ways of achieving better survival outcome in HCC patients, yet the precise mechanism for this success remains to be elusive. We have systematically investigated the expression profiles of RTKs transcriptomic datasets i.e. The Cancer Genome Atlas - Liver hepatocellular carcinoma (TCGA-LIHC) (n=371) and our in-house cohort of paired i.e. HKU-QMH (n=41), and identified EPHB4 as one of the most abundantly expressed and frequently mutated RTKs in human HCCs. EPHB4 expression was found to be significantly correlated with Alpha Fetoprotein (AFP) expression, a factor commonly used in the diagnosis of HCCs. Furthermore, EphB4 has been demonstrated to play a regulatory role in VEGF/VEGFR-mediated angiogenesis, critical targets for current successful drugs. We confirmed abundant membrane EphB4 expression in HCC clinical specimens and human HCC cell line models using immunohistochemistry. Clinicopathological correlation analysis of EPHB4 mRNA expression revealed that its overexpression is associated with aggressive tumor features including presence of tumor microsatellite, liver invasion, venous invasion, and absence of tumor encapsulation. Functionally, stably knocking down EPHB4 (shEPHB4) resulted in a strong reduction of proliferation and self-renewal rates of multiple HCC cell lines in vitro. It significantly reduced tumor growth rates in subcutaneous and liver orthotopic models in vivo. Furthermore, limiting dilution assay showed EPHB4 plays a role in tumor initiation property of HCC cells. Mechanistically, we performed Gene Set Enrichment Analysis (GSEA) of transcriptome profiles for shEPHB4 with respective controls and found that knocking down EPHB4 causes HCC cells undergo drastic changes in several cell cycle checkpoints including G1/S transition. Using thymidine blockade, we further confirmed that knockdown of significantly hinders the transition of HCC cells from G1 to S phase of cell cycle. Our data suggests a strong link between EphB4 and cell cycle. Conclusion: We found EphB4 plays a functional role in regulating G1/S transition in human HCCs and understanding regulatory role of EphB4 on cell cycle progression may provide mechanistic insights into the how to effectively target HCC in the clinic. Citation Format: Abdullah Husain, Elley Y. Chiu, Hoi-Tang Ma, Daniel W. Ho, Karen M. Sze, Lo-Kong Chan, Irene O. Ng. EPHB4 is a key regulator of G1/S transition in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1637.

Influence of C60 Nanofilm on the Expression of Selected Markers of Mesenchymal–Epithelial Transition in Hepatocellular Carcinoma

Influence of C60 Nanofilm on the Expression of Selected Markers of Mesenchymal–Epithelial Transition in Hepatocellular Carcinoma

Actinidia chinensis Planch. root extract attenuates hypoxia-induced hepatic stellate cell activation and epithelial–mesenchymal transition in hepatocellular carcinoma by inhibiting inflammatory cytokine secretion

IntroductionHepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide. Several studies have demonstrated the potency of Chinese herbs in protecting the liver from inflammation, hypoxia, fibrosis, and cancer. The roots of Actinidia chinensis Planch. exhibit strong anti-tumour effects; however, their mechanisms of action in HCC are not clearly understood. MethodsTo investigate the molecular mechanisms underlying the anti-cancer effect of A chinensis Planch. root extract (acRoot) in HCC and hepatic stellate cells (HSCs). The HSC line LX2 and HCC cell line MHCC97H were cultured under oxygen-rich or oxygen-deprived conditions and then exposed to various acRoot concentrations for different periods. Subsequently, cell viability, cytotoxicity, and enzyme-linked immunosorbent assays and western blotting were conducted to assess the effects of acRoot treatment. ResultsacRoot decreased the number of viable LX2 and MHCC97H cells in a concentration- and time-dependent manner. Under hypoxic and/or co-culture conditions, pro-inflammatory cytokine expression was up-regulated in both cell lines; however, the expression levels were significantly decreased following treatment with acRoot. Additionally, acRoot inhibited hypoxia-induced activation of LX2 cells and inflammation and epithelial–mesenchymal transition (EMT) in MHCC97H cells. Moreover, acRoot inhibited cross-talk between LX2 and MHCC97H cells, further inhibiting cellular activation, inflammation, and EMT. ConclusionThese results show the therapeutic potential of acRoot as an anti-cancer, anti-inflammatory, and anti-fibrotic agent, shedding new light on the medicinal properties of Chinese herbs.

B4GALNT1 promotes carcinogenesis by regulating epithelial-mesenchymal transition in hepatocellular carcinoma based on pan-cancer analysis.

β-1,4-N-Acetyl galactosaminyltransferase1 (B4GALNT1) has been reported to play important roles in tumor progression and metastasis. Herein, we investigate the oncogenic roles of B4GALNT1 for hepatocellular carcinoma (HCC) based on immune microenvironment. The Cancer Genome Atlas database and Genotype-Tissue Expression, cBioportal, ImmuCellAI, TIMER2 and other databases were searched to analyze the expression and clinical applications of B4GALNT1 in liver cancer patients. Kaplan-Meier survival analysis, Cox regression analysis, Kyoto Encyclopedia of Genes and Genomes and Gene Ontology enrichment analysis were utilized. Moreover, western blot assay, immune histochemistry staining, Cell Counting Kit-8 (CCK-8) assay, invasion and migration assay were performed to evaluate the function of B4GALNT1 in HCC. B4GALNT1 is overexpressed in 14 tumors, and the mRNA expression levels of B4GALNT1 were remarkably elevated in most tumor types, including HCC. In addition, B4GALNT1 expression was an independent prognostic factor, and a low expression of B4GALNT1 showed a better overall survival and disease-specific recurrence-free survival in patients with HCC. Gene set variation analysis indicated that B4GALNT1 presented a positive correlation with the epithelial-mesenchymal transition (EMT) pathway in HCC. B4GALNT1 expression was closely associated with immune activation genes in the HCC microenvironment. Moreover, B4GALNT1 expression was higher in HCC tissue than that in surrounding tissues. B4GALNT1 promoted the expression of apoptosis-related or EMT-related proteins and then decreased the expression of Bcl-2 and Bcl-xl in HCC cells, suggesting that B4GALNT1 knockdown significantly inhibited the proliferation and invasion of HCC cells. B4GALNT1 may promote HCC development through regulating the EMT pathway, which suggests that B4GALNT1 may serve as a promising predictive biomarker and a potential therapeutic target for HCC.

High expression of embryonic stem cell marker SSEA3 confers poor prognosis and promotes epithelial mesenchymal transition in hepatocellular carcinoma

BackgroundMalignant cells may arise from dedifferentiation of mature cells and acquire features of the progenitor cells. Definitive endoderm from which liver is derived, expresses glycosphingolipids (GSLs) such as stage-specific embryonic antigen 3 (SSEA3), Globo H, and stage-specific embryonic antigen 4 (SSEA4). Herein, we evaluated the potential prognosis value of the three GSLs and biological functions of SSEA3 in hepatocellular carcinoma (HCC). MethodsThe expression of SSEA3, Globo H, and SSEA4 in tumor tissues obtained from 328 patients with resectable HCC was examined by immunohistochemistry staining. Epithelial mesenchymal transition (EMT) and their related genes were analyzed by transwell assay and qRT-PCR, respectively. ResultsKaplan Meier survival analysis showed significantly shorter relapse-free survival (RFS) for those with higher expression of SSEA3 (p < 0.001), Globo H (p < 0.001), and SSEA4 (p = 0.005) and worse overall survival (OS) for those with high expression of either SSEA3 (p < 0.001) or SSEA4 (p = 0.01). Furthermore, multivariable Cox regression analysis identified the SSEA3 as an independent predictor for RFS (HR: 2.68, 95% CI: 1.93–3.72, p < 0.001) and OS (HR: 2.99, 95% CI: 1.81–4.96, p < 0.001) in HCC. Additionally, SSEA3-ceramide enhanced the EMT of HCC cells, as reflected by its ability to increase migration, invasion and upregulate the expression of CDH2, vimentin, fibronectin, and MMP2, along with ZEB1. Moreover, ZEB1 silencing abrogated the EMT-enhancing effects of SSEA3-ceramide. ConclusionsHigher expression of SSEA3 was an independent predictor for RFS and OS in HCC and promoted EMT of HCC via upregulation of ZEB1.

SAT-216 - MAP17 promotes an epithelial-mesenchymal-amoeboid transition in hepatocellular carcinoma by switching one-carbon metabolism

SAT-216 - MAP17 promotes an epithelial-mesenchymal-amoeboid transition in hepatocellular carcinoma by switching one-carbon metabolism

SMYD3 induces sorafenib resistance by activating SMAD2/3-mediated epithelial-mesenchymal transition in hepatocellular carcinoma

Drug resistance prominently hampers the effects of systemic therapy of sorafenib to hepatocellular carcinoma (HCC). Epigenetics have critical regulatory roles in drug resistance. However, the contributions of histone methylatransferase SET and MYND domain containing 3 (SMYD3) to sorafenib resistance in HCC remain largely unknown. Here, using our established sorafenib-resistant HCC cell and xenograft models, we found SMYD3 was markedly elevated in sorafenib-resistant tumors and cells. Functionally, loss- and gain-of-function studies showed that SMYD3 promoted the migration, invasion, metastasis and stemness of sorafenib-resistant HCC cells. Mechanistically, SMYD3 is required for SMAD2/3-mediated epithelial-mesenchymal transition (EMT) in sorafenib-resistant HCC cells by interacting with SMAD2/3 and epigenetically promoting the expression of SOX4, ZEB1, SNAIL1 and MMP9 genes. In summary, our data demonstrate that targeting SMYD3 is an effective approach to overcome sorafenib resistance in HCC.

Isorhamnetin exerts anti-tumor activity in DEN + CCl4-induced HCC mice.

Hepatocellular carcinoma (HCC) is the most prevalent type of liver cancer and the main cause of cancer death globally. The use of medicinal herbs as chemotherapeutic agents in cancer treatment is receiving attention as they possess no or minimum side effects. Isorhamnetin (IRN), a flavonoid, has been under attention for its anti-inflammatory and anti-proliferative properties in a number of cancers, including colorectal, skin, and lung cancers. However, the in vivo mechanism of isorhamnetin to suppress liver cancer has yet to be explored. HCC was induced by N-diethylnitrosamine (DEN) and carbon tetrachloride (CCL4) in Swiss albino mice. Isorhamnetin (100mg/kg body weight) was given to examine its anti-tumor properties in HCC mice model. Histological analysis and liver function assays were performed to assess changes in liver anatomy. Probable molecular pathways were explored using immunoblot, qPCR, ELISA, and immunohistochemistry techniques. Isorhamnetin inhibited various pro-inflammatory cytokines to suppress cancer-inducing inflammation. Additionally, it regulated Akt and MAPKs to suppress Nrf2 signaling. Isorhamnetin activated PPAR-γ and autophagy while suppressing cell cycle progression in DEN + CCl4-administered mice. Additionally, isorhamnetin regulated various signaling pathways to suppress cell proliferation, metabolism, and epithelial-mesenchymal transition in HCC. Regulating diverse cellular signaling pathways makes isorhamnetin a better anti-cancer chemotherapeutic candidate in HCC. Importantly, the anti-TNF-α properties of isorhamnetin could prove it a valuable therapeutic agent in sorafenib-resistant HCC patients. Additionally, anti-TGF-β properties of isorhamnetin could be utilized to reduce the EMT-inducing side effects of doxorubicin.

Size-optimized simvastatin-loaded TPGS modified lipid nanocapsules for targeting epithelial-to-mesenchymal transition in hepatocellular carcinoma: Role of PTEN/AKT signaling

ABSTRACT Objectives Novel D-α-tocopherol polyethylene glycol 1000 succinate (TPGS) modified lipid nanocapsules (LNC) were prepared with the aim of improving the effectiveness of simvastatin (SIM) in hepatocellular carcinoma (HCC). The present study, therefore, sought to investigate the effect of size-optimized SIM-loaded LNC on epithelial-to-mesenchymal transition (EMT) in HCC, providing insights on the implication of phosphatase and tensin homolog (PTEN)/protein kinase B (AKT) axis. Methods Two optimized SIM-loaded LNCs with particle sizes 25 nm (SIM-LNC25) and 50 nm (SIM-LNC50) were prepared and biodistribution studies were performed. The anticancer effect of the prepared LNC was evaluated both in vitro and in vivo. The anti-migratory potential and EMT suppression through PTEN/AKT axis modulation were also explored. Results SIM-LNC50 was superior to SIM-LNC25 in both in vitro and in vivo experiments, as evidenced by cytotoxicity assays, tumor histopathology, and enhanced apoptosis. SIM-LNC50 also alleviated the migratory potential of HCC cells. Moreover, EMT markers implied a transition of tumor cells toward the epithelial rather than the mesenchymal phenotype both in vitro and in vivo. PTEN/AKT axis modulation was also evident with SIM-LNC50. Conclusion The present study, therefore, suggests the efficacy of the 50 nm particles in SIM-loaded LNC in HCC by targeting EMT via modulating the PTEN/AKT signaling axis.

XPO1 intensifies sorafenib resistance by stabilizing acetylation of NPM1 and enhancing epithelial-mesenchymal transition in hepatocellular carcinoma

Emerging studies have suggested that exportin-1 (XPO1) plays a pivotal role in hepatocellular carcinoma (HCC). However, the underlying mechanism of XPO1 in HCC sorafenib resistance remains enigmatic. The expression of XPO1 in HCC tumor tissues and sorafenib-resistant (SR) cells were analyzed by bioinformatics analysis, immunohistochemistry (IHC) and Western blotting. The interaction mechanism between XPO1 and Nucleophosmin (NPM1) was investigated by immunoprecipitation (IP), Mass-spectrometric (MS) analysis, immunofluorescence colocalization, CRISPR/CAS9 technology and RNA-seq. Analyses were also conducted on KPT-8602 and sorafenib's combined therapeutic effect. Our findings unraveled that the XPO1 overexpression was observed in HCC, and correlated with poorer survival. Knockdown of XPO1 inhibited the migration and proliferation of HCC cells, and also reduced the resistance of HCC cells to sorafenib. Mechanistically, XPO1 interacted with the C-terminus of NPM1 and mediated the acetylation of NPM1 at lysine 54 to maintain sorafenib resistance. XPO1 was bound to Vimentin, resulting in the epithelial-mesenchymal transition (EMT) progression in sorafenib-resistant cells. KPT-8602 in combination with sorafenib suppressed the tumor growth. These results highlighted the therapeutic value of targeting XPO1 in overcoming sorafenib resistance. The combinational treatment of KPT-8602 and sorafenib might be an improved therapeutic option.

Retrograde regulation of mitochondrial fission and epithelial to mesenchymal transition in hepatocellular carcinoma by GCN5L1.

Metabolic reprogram is crucial to support cancer cell growth and movement as well as determine cell fate. Mitochondrial protein acetylation regulates mitochondrial metabolism, which is relevant to cancer cell migration and invasion. The functional role of mitochondrial protein acetylation on cancer cell migration remains unclear. General control of amino acid synthesis 5 like-1(GCN5L1), as the regulator of mitochondrial protein acetylation, functions on metabolic reprogramming in mouse livers. In this study, we find that GCN5L1 expression is significantly decreased in metastatic HCC tissues. Loss of GCN5L1 promotes reactive oxygen species (ROS) generation through enhanced fatty acid oxidation (FAO), followed by activation of cellular ERK and DRP1 to promote mitochondrial fission and epithelia to mesenchymal transition (EMT) to boost cell migration. Moreover, palmitate and carnitine-stimulated FAO promotes mitochondrial fission and EMT gene expression to activate HCC cell migration. On the other hand, increased cellular acetyl-CoA level, the product of FAO, enhances HCC cell migration. Taken together, our finding uncovers the metastasis suppressor role as well as the underlying mechanism of GCN5L1 in HCC and also provides evidence of FAO retrograde control of HCC metastasis.

Overexpression of CLEC5A inhibits cell proliferation and metastasis and reverses epithelial-mesenchymal transition in hepatocellular carcinoma

To evaluate the effects of CLEC5A expression level on cell proliferation, migration and invasion and epithelial-mesenchymal transition (EMT) in hepatocellular carcinoma (HCC) and explore the role of CLEC5A in the tumorigenesis and progression of HCC. The expression level of CLEC5A was detected in 50 pairs of HCC and adjacent tissues using immunohistochemical staining, and its association with clinicopathological parameters of HCC patients was analyzed. Cultured HCC cell line SK-HEP-1 was transfected with a lentiviral vector overexpressing CLEC5A, and the transfection efficiency was verified using real-time fluorescence quantitative PCR and Western blotting. The changes in proliferation, migration and invasion abilities of the transfected cells were analyzed using CCK-8, 5-ethynyl-29-deoxyuridine (EdU) and Transwell assays, and EMT of the cells was determined using Western blotting. The protein expression level of CLEC5A was significantly lower in HCC tissues than in the adjacent tissues (P < 0.001). The expression level of CLEC5A was significantly correlated with tumor size (P=0.008), tumor number (P=0.010), histological differentiation (P=0.016), microvascular invasion (P=0.024) and BCLC stage (P=0.040). In SK-HEP-1 cells, overexpression of CLEC5A obviously inhibited the cell proliferation, migration and invasion and reversed EMT phenotype of the cells. CLEC5A is a potential HCC suppressor gene and may serve as a promising therapeutic target for HCC.

The Natural Compound Dehydrocrenatidine Attenuates Nicotine-Induced Stemness and Epithelial-Mesenchymal Transition in Hepatocellular Carcinoma by Regulating a7nAChR-Jak2 Signaling Pathways.

Background Exposure to nicotine has been observed associated with tumor progression, metastasis, and therapy resistance of many cancers. Hepatocellular carcinoma (HCC) is one major cancer related to the liver and the most difficult to treat malignancies worldwide. The underlying mechanism of nicotine in the stimulation of HCC tumorigenesis is still not studied well. Methods Classically, nicotine binds to nicotinic acetylcholine receptors (nAChRs) and induces many downstream cancer-associated signaling pathways. Big data analysis is used to explore the importance of a7nAChR-Jak2 axis in the progression of hepatocellular carcinoma. Bioinformatic analysis was performed to determine gene associated with a7nAChR-Jak2 axis of HCC patients. Biological importance of a7nAChR-Jak2 axis was investigated in vitro (Hun7 and HepG2 cell lines), and athymic nude mouse models bearing HepG2-HCC cells xenografts were established in vivo. Result We found that nicotine exposure stimulated the HCC tumorigenicity by inducing the expression of one of the key nAChRs subunit that is α7nAChR as well as the expression of Janus kinase (JAK)-2. In both the in vitro and in vivo studies, the reduced overexpression of α7nAChR and increased sensitization of HCC towards treatment is observed with dehydrocrenatidine (DHCT), a novel and potent JAK family kinase inhibitor. Interestingly, DHCT treatment results in the reduction of the epithelial-mesenchymal transition process which leads to a significant reduction of clonogenicity, migratory, and invasive ability of HCC cells. Moreover, DHCT treatment also inhibits the cancer stem cell phenotype by inhibiting the tumor-sphere formation and reducing the number of ALDH1+ cells population in nicotine-stimulated HCC cells. Conclusions Taken together, the presented results indicate the positive effect of inhibition of nicotine induced overexpression of α7nAChR and JAK2, unique to HCC. Thus, these findings suggest the nicotine effect on HCC progression via α7nAChR-mediated JAK2 signaling pathways, and DHCT treatment enhances the therapeutic potential of HCC patients via overcoming/reversing the effect of nicotine in HCC patients.

Cyclin-Dependent Kinase 4 is expected to be a therapeutic target for hepatocellular carcinoma metastasis using integrated bioinformatic analysis

ABSTRACT Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related mortality worldwide. HCC cells possess biological characteristics of high invasion and metastasis. In this respect, to prevent cancer cell invasion and metastasis and early active intervention, we herein screened through the TCGA database for further prognostic analysis including overall survival and disease-free survival . The Kaplan-Meier curve suggested that Cyclin-Dependent Kinase 4 (CDK4) might be an independent prognostic factor for HCC. Moreover, we performed mRNA expression analysis to measure CDK4 levels in normal liver tissues and HCC tissues, and immunohistochemistry analysis to detect protein level of CDK4 in Non-tumor tissue and HCC tissues . Our findings indicated that the expression of CDK4 was significantly higher in tumor tissues compared with Non-tumor tissue in HCC, which increased from HCC stage 1 to 3. Furthermore, the results of transwell-assay indicated that knocking down CDK4 significantly suppresses the invasion and migration of HCC cells, and the results of bioinformatics analysis revealed that genes closely associated with CDK4 are potentially worthy of further investigation. Additionally, the results of Western Blot indicated CDK4 regulates epithelial mesenchymal transition in HCC,and CDK4 appears to regulate EMT and HCC progression via the Wnt/β-catenin pathway. Collectively, this study found the key target gene through bioinformatic analysis and further functional validation through cell experiments. In particular, CDK4 is anticipated to become a crucial hub gene to snipe the metastasis of cancer cells in HCC. Abbreviations: Hepatocellular carcinoma (HCC);Cyclin-Dependent Kinase 4(CDK4);Genomic Data Commons (GDC); genes; EC, Endometrial cancer; GEO, gene expression omnibus; GO, Gene Ontology; GSEA, Gene set enrichment analysis; KEGG, Database; TCGA, The Cancer Genome Atlas; TSGs, tumor suppressor genes;epithelial mesenchymal transition (EMT).

Pinin Induces Epithelial-to-Mesenchymal Transition in Hepatocellular Carcinoma by Regulating m6A Modification.

Pinin is a moonlighting protein localized in desmosomes and nucleus. It could promote the growth of hepatocellular carcinoma. Whether this protein can induce epithelial-to-mesenchymal transition (EMT) and malignant progression in HCC is unknown. This work found that Pinin prompts EMT in vitro and in vivo. Further mechanism study found that Pinin increases the level of N6-methyladenosine (m6A) modification of RNA by interacting with METTL3, which in turn induces snail1 expression. These findings suggest that Pinin induces EMT by regulating m6A modification and, thus, could be a potential anticancer target for HCC therapy.

Downregulation of long noncoding RNA T1NCR inhibits cell proliferation, invasion, and epithelial-mesenchymal transition of hepatocellular carcinoma

Accumulating reports have identified that long non-coding RNAs (IncRNAs) function as key regulators of tumor initiation and progression. The aim of the current study was to determine the clinical significance and functional role of TINCR in hepatocellular carcinoma (HCC). In the present study, the level of IncRNA TINCR expression was significantly upregulated in HCC tissues compared to adjacent normal tissues. Higher levels of IncRNA TINCR expression were significantly correlated with tumor size and vascular invasion of HCC patients. LncRNA TINCR knockdown inhibited cell proliferation ability, increased the proportion of G1 phase cells, reduced the proportion of S phase cells, and suppressed cell invasion of HCC in vitro. Additionally, IncRNA TINCR knockdown inhibited the HCC cell epithelial-mesenchymal transition (EMT) phenomenon by upregulating E-cadherin and reducing N-cadherin expression. We demonstrated that knockdown of IncRNA reduced tumor growth in vivo. Thus, these results indicated that IncRNA TINCR exhibits a tumor oncogenic role in HCC and inhibition of IncRNA TINCR might serve as a therapeutic target for HCC.

Preparation of an anti-NEK2 monoclonal antibody and its application in liver cancer

BackgroundNever in mitosis gene-A (NIMA)-related expressed kinase 2 (NEK2) is a serine/threonine protein kinase regulated by the cell cycle. The purpose of this study was to obtain NEK2 protein to prepare an anti-NEK2 monoclonal antibody (mAb) and explore the application of the anti-NEK2 mAb of therapeutic and diagnostic in hepatocellular carcinoma (HCC).ResultsThe NEK2 gene sequence was cloned from the normal liver cell line HL7702, and the full-length NEK2 gene sequence was cloned into the prokaryotic expression vector pET30a and transformed into Escherichia coli BL21 (DE3) cells. The recombinant fusion protein was obtained under optimized conditions and injected in BALB/c mice to prepare an anti-NEK2 mAb. By screening, we obtained a stable hybridoma cell line named 3A3 that could stably secrete anti-NEK2 mAb. Anti-NEK2 3A3 mAb was purified from ascites fluid. The isotype was IgG1, and the affinity constant (Kaff) was 6.0 × 108 L/mol. Western blot, indirect enzyme-linked immunosorbent assay (iELISA), immunofluorescence and immunocytochemical analyses showed that the mAb could specifically recognize the NEK2 protein. MTT assays showed that the mAb 3A3 could inhibit the proliferation of HCC cells. KEGG pathway analysis showed that NEK2 might affected pathways of the cell cycle. Moreover, NEK2-related genes were mainly enriched in the S and G2 phases and might act as tumor-promoting genes by regulating the S/G2 phase transition of HCC cells.ConclusionsAn anti-NEK2 mAb with high potency, high affinity and high specificity was prepared by prokaryotic expression system in this study and may be used in the establishment of ELISA detection kits and targeted treatment of liver cancer.

Fluid shear stress activates YAP to promote epithelial-mesenchymal transition in hepatocellular carcinoma.

Epithelial–mesenchymal transition (EMT) mediated by fluid shear stress (FSS) in the tumor microenvironment plays an important role in driving metastasis of the malignant tumor. As a mechanotransducer, Yes‐associated protein (YAP) is known to translocate into the nucleus to initiate transcription of genes involved in cell proliferation upon extracellular biophysical stimuli. Here, we showed that FSS facilitated cytoskeleton rearrangement in hepatocellular carcinoma cells, which led to the release of YAP from its binding partner, integrin β subunit, in the cytomembrane. Moreover, we found that upregulation of guanine nucleotide exchange factor (GEF)‐H1, a microtubule‐associated Rho GEF, is a critical step in the FSS‐induced translocation of YAP. Nuclear YAP activated the expression of the EMT‐regulating transcription factor SNAI1, but suppressed the expression of N6‐methyladenosine (m6A) modulators; together, this promoted the expression of EMT‐related genes. We also observed that FSS‐treated HepG2 cells showed markedly increased tumorigenesis and metastasis in vivo. Collectively, our findings unravel the underlying molecular processes by which FSS induces translocation of YAP from the cytomembrane to the nucleus, contributes to EMT and enhances metastasis in hepatocellular carcinoma.

Role of LncRNAs in the Epithelial-Mesenchymal Transition in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a type of primary liver cancer with a high incidence and mortality rate. HCC develops insidiously, and most newly diagnosed cases are in the middle and advanced stages. The epithelial-mesenchymal transition (EMT) is a vital mechanism underlying metastasis in patients with advanced HCC. EMT is a multistep and complex procedure. The promotion and inhibition of EMT directly affect the migration and invasion of HCC. LncRNAs are involved in the epigenetic modification of genes, regulation of gene transcription, and posttranslational modification of proteins. LncRNAs also play important roles in regulating EMT progression in HCC and are promising biomarkers and therapeutic targets. This review focused on summarizing the mechanism by which lncRNAs regulate EMT in HCC. In particular, lncRNAs were reported to primarily act as RNA sponges, and the regulation of EMT involves major signaling pathways. Finally, we reviewed the mechanisms by which lncRNAs are involved in drug resistance and discussed the clinical prospects and potential challenges of utilizing lncRNAs to treat HCC.