Effects In Hepatocellular Carcinoma Cells Research Articles

Natural Product Auraptene Targets SLC7A11 for Degradation and Induces Hepatocellular Carcinoma Ferroptosis.

The natural product auraptene can influence tumor cell proliferation and invasion, but its effect on hepatocellular carcinoma (HCC) cells is unknown. Here, we report that auraptene can exert anti-tumor effects in HCC cells via inhibition of cell proliferation and ferroptosis induction. Auraptene treatment induces total ROS and lipid ROS production in HCC cells to initiate ferroptosis. The cell death or cell growth inhibition of HCC cells induced by auraptene can be eliminated by the ROS scavenger NAC or GSH and ferroptosis inhibitor ferrostatin-1 or Deferoxamine Mesylate (DFO). Mechanistically, the key ferroptosis defense protein SLC7A11 is targeted for ubiquitin-proteasomal degradation by auraptene, resulting in ferroptosis of HCC cells. Importantly, low doses of auraptene can sensitize HCC cells to ferroptosis induced by RSL3 and cystine deprivation. These findings demonstrate a critical mechanism by which auraptene exhibits anti-HCC effects via ferroptosis induction and provides a possible therapeutic strategy for HCC by using auraptene or in combination with other ferroptosis inducers.

Anticancer Activity of Delta-Tocotrienol in Human Hepatocarcinoma: Involvement of Autophagy Induction.

(1) Hepatocellular carcinoma (HCC) is the predominant form of primary liver cancer. Surgical resection, tumor ablation, and liver transplantation are curative treatments indicated for early-stage HCC. The management of intermediate and advanced stages of pathology is based on the use of systemic therapies which often show important side effects. Vitamin E-derivative tocotrienols (TTs) play antitumoral properties in different tumors. Here, we analyzed the activity of delta-TT (δ-TT) on HCC human cell lines. (2) We analyzed the ability of δ-TT to trigger apoptosis, to induce oxidative stress, autophagy, and mitophagy in HepG2 cell line. We evaluated the correlation between the activation of autophagy with the ability of δ-TT to induce cell death. (3) The data obtained demonstrate that δ-TT exerts an antiproliferative and proapoptotic effect in HCC cells. Furthermore, δ-TT induces the release of mitochondrial ROS and causes a structural and functional alteration of the mitochondria compatible with a fission process. Finally, δ-TT triggers selective autophagy process removing dysfunctional mitochondria. Inhibition of autophagy reversed the cytotoxic action of δ-TT. (4) Our results demonstrate that δ-TT through the activation of autophagy could represent a potential new approach in the treatment of advanced HCC.

Hsa_circ_0000518 stimulates the malignant progression of hepatocellular carcinoma via regulating ITGA5 to activate the Warburg effect

Studies have shown that the abnormal expression of circular RNA (circRNA) is inextricably linked to hepatocellular carcinoma (HCC). Recently, hsa_circ_0000518 (circ_0000518) was discovered in many cancer progressions. However, its function in HCC is still unclear. Through GEO database analysis combined with gene expression detection of HCC related clinical samples and cell lines, we identified that circ_0000518 was abnormally overexpressed in HCC. Cell and animal model experiments jointly indicated that circ_0000518 can stimulate HCC cell proliferation, migration, invasion and suppress apoptosis. Furthermore, we also found that knocking down the circ_0000518 could inhibit the Warburg effect in HCC cells. Mechanistically, circ_0000518 was found to be primarily localized in the cytoplasm, and sponge hsa-miR-326 (miR-326) promoted integrin alpha 5 (ITGA5) expression. In addition, circ_0000518 could enhance the stability of HuR-mediated ITGA5 mRNA, thereby activating the Warburg effect. In conclusion, this study elucidated that circ_0000518 was a cancer-promoting circRNA, which could enhance ITGA5 expression through competing endogenous RNAs (ceRNA) and RNA Binding Protein (RBP) mechanisms, thus facilitating the development of HCC. It provides a meaningful diagnostic and therapeutic target for HCC.

Cannabidiol reverts the malignant phenotype of hepatocellular carcinoma cells via the GPR55/TP53/MAPK axis

Cannabidiol (CBD) has antioxidant and anti-inflammatory activities. However, the anti-tumor effect of CBD on hepatocellular carcinoma (HCC) remains unclear. Here, we investigated whether CBD displays anti-tumorigenic effects in HCC cells and whether it could reduce tumorigenesis and metastases in vivo. First, this study treated HCC cells with different concentrations of CBD, followed by analyzing the changes in the proliferative, apoptotic, migratory and invasive abilities. The effects of CBD on the growth and metastasis of HCC cells in vivo were verified by tumorigenesis and metastasis assays. Subsequently, the target genes of CBD were predicted through the SwissTarget website and the genes differentially expressed in cells after CBD treatment were analyzed by microarray for intersection. The enrichment of the pathways after CBD treatment was analyzed by KEGG enrichment analysis, followed by western blot validation. Finally, rescue assays were used to validate the functions of genes as well as pathways in the growth and metastasis of HCC cells. A significant weakening of the ability of HCC cells to grow and metastasize in vitro and in vivo was observed upon CBD treatment. Mechanistically, CBD reduced GRP55 expression in HCC cells, along with increased TP53 expression and blocked MAPK signaling activation. In CBD-treated cells, the anti-tumor of HCC cells was restored after overexpression of GRP55 or deletion of TP53. CBD inhibits the MAPK signaling activation and increases the TP53 expression by downregulating GRP55 in HCC cells, thereby suppressing the growth and metastasis of HCC cells.

Dihydroartemisinin breaks the positive feedback loop of YAP1 and GLUT1-mediated aerobic glycolysis to boost the CD8+ effector T cells in hepatocellular carcinoma

Aerobic glycolysis is a hallmark of hepatocellular carcinoma (HCC). Dihydroartemisinin (DHA) exhibits antitumor activity towards liver cancer. Our previous studies have shown that DHA inhibits the Warburg effect in HCC cells. However, the mechanism still needs to be clarified. Our study aimed to elucidate the interaction between YAP1 and GLUT1-mediated aerobic glycolysis in HCC cells and focused on the underlying mechanisms of DHA inhibiting aerobic glycolysis in HCC cells. In this study, we confirmed that inhibition of YAP1 expression lowers GLUT1-mediated aerobic glycolysis in HCC cells and enhances the activity of CD8+T cells in the tumor niche. Then, we found that DHA was bound to cellular YAP1 in HCC cells. YAP1 knockdown inhibited GLUT1-mediated aerobic glycolysis, whereas YAP1 overexpression promoted GLUT1-mediated aerobic glycolysis in HCC cells. Notably, liver-specific Yap1 knockout by AAV8-TBG-Cre suppressed HIF-1α and GLUT1 expression in tumors but not para-tumors in DEN/TCPOBOP-induced HCC mice. Even more crucial is that YAP1 forms a positive feedback loop with GLUT1-mediated aerobic glycolysis, which is associated with HIF-1α in HCC cells. Finally, DHA reduced GLUT1-aerobic glycolysis in HCC cells through YAP1 and prevented the binding of YAP1 and HIF-1α. Collectively, our study revealed the mechanism of DHA inhibiting glycolysis in HCC cells from a perspective of a positive feedback loop involving YAP1 and GLUT1 mediated-aerobic glycolysis and provided a feasible therapeutic strategy for targeting enhanced aerobic glycolysis in HCC.

MiR-1972 inhibits hepatocellular carcinoma proliferation by targeting GZMH-mediated DNA replication in the cell cycle.

To understand the regulatory roles of miR-1972 and GZMH in hepatocellular carcinoma (HCC) and explore their potential as therapeutic biomarkers. In vitro verification of the regulation of malignant cell behavior by differential expression of miR-1972 in HCC cells. The GSE113996 dataset was studied using weighted gene co-expression network analysis (WGCNA) and differential expressed genes respectively to identify the key prognostic gene GZMH and assess the effect of its differential expression on the prognosis of the patient. Finally, the regulation of GZMH expression by miR-1972 was verified, and the effect of their combination on HCC cell behavior was analyzed. Inhibition of miR-1972 can reduce cell proliferation, migration, and invasion, while overexpression of miR-1972 has the opposite effect in HCC cells. According to the data, a positive prognosis for HCC was linked with higher GZMH expression. Interestingly, miR-1972 was observed to reverse-regulate the expression of GZMH. Besides, the combined regulation of GZMH and miR-1972 has been discovered to affect the cell growth, invasive capacity, and migratory potential of HCC cells, especially the cell cycle arrest in the G2 phase. miR-1972 regulates the malignant behavior of HCC cells, especially cell proliferation, by regulating GZMH expression.

Aspongopus chinensis Dallas induces pro-apoptotic and cell cycle arresting effects in hepatocellular carcinoma cells by modulating miRNA and mRNA expression

Aspongopus chinensis Dallas is a traditional Chinese medicinal insect with several anticancer properties can inhibit cancer cell growth, by inhibiting cell division, autophagy and cell cycle. However, the precise therapeutics effects and mechanisms of this insect on liver cancer are still unknown. This study examined the inhibitory influence of A. chinensis on the proliferation of hepatocellular carcinoma (HCC) cells and explore the underlying mechanism using high-throughput sequencing. The results showed that A. chinensis substantially reduced the viability of Hep G2 cells. A total of 33 miRNAs were found to be upregulated, while 43 miRNAs were downregulated. Additionally, 754 mRNAs were upregulated and 863 mRNAs were downregulated. Significant enrichment of differentially expressed genes was observed in signaling pathways related to tumor cell growth, cell cycle regulation, and apoptosis. Differentially expressed miRNAs exhibited a targeting relationship with various target genes, including ARC, HSPA6, C11orf86, and others. Hence, cell cycle and apoptosis were identified by flow cytometry. These findings indicate that A. chinensis impeded cell cycle advancement, halted the cell cycle in the G0/G1 and S stages, and stimulated apoptosis. Finally, mouse experiments confirmed that A. chinensis significantly inhibits tumor growth in vivo. Therefore, our findings indicate that A. chinensis has a notable suppressive impact on the proliferation of HCC cells. The potential mechanism of action could involve the regulation of mRNA expression via miRNA, ultimately leading to cell cycle arrest and apoptosis. The results offer a scientific foundation for the advancement and application of A. chinensis in the management of HCC.

Circ_0026134 regulates the miR-1270/GRB2 pathway to affect the radiosensitivity of hepatoma cells

Objective: To investigate whether circular RNA 0026134 (circ_0026134) affects the radiosensitivity of hepatoma cells by regulating the miR-1270/growth factor receptor-bound protein 2 (GRB2) pathway. Methods: Real-time quantitative PCR (RT-qPCR) was used to detect the expression levels of circ_0026134, miR-1270, and GRB2 in liver cancer tissues and cells. Bioinformatics analysis, a dual-luciferase gene reporter assay, RT-qPCR, and western blot were used to analyze the targeting relationships between circ_0026134 and miR-1270 and miR-1270 and GRB2. The effects of circ_0026134, miR-1270, and GRB2 expression combined with 6 Gy on the proliferation, invasion, migration, and apoptosis of Huh7 and SK-HEP-1 cells were detected by a cell counting kit, a transwell assay, a scratch assay, and flow cytometry. The tumorigenesis experiment was used to detect the effect of silencing circ_0026134 in nude mice. Measurement data are expressed as the mean ± standard deviation. The independent sample t-test was used for comparison between two groups, and the one-way analysis of variance and SNK-q test were used for comparison between multiple groups. P < 0.05 was considered statistically significant. Results: The expression levels of circ_0026134 and GRB2, Huh7, and SK-HEP-1 cells in liver cancer tissues were significantly increased, while the expression levels of miR-1270 were significantly decreased (P < 0.05). The expression of circ_0026134 in Huh7 and SK-HEP-1 decreased significantly after radiotherapy (P < 0.05). circ_0026134 binds directly to miR-1270 and negatively regulates miR-1270 expression (P < 0.05). miR-1270 binds directly to GRB2 and negatively regulates GRB2 expression (P < 0.05). 6 Gy radiation significantly inhibited the proliferation, migration, and invasion of Huh7 and SK-HEP-1 cells and induced apoptosis (P < 0.05). Silencing circ_0026134 or overexpression of miR-1270 significantly enhanced the anti-proliferation, anti-migration, invasion, and pro-apoptosis effects of 6 Gy treatment on hepatoma cells (P < 0.05). Inhibition of miR-1270 significantly weakened the effects of silencing circ_0026134 combined with 6 Gy radiation on proliferation, migration, invasion, and apoptosis of hepatoma cells (P < 0.05). Overexpression of GRB2 significantly weakened the effects of miR-1270 overexpression combined with 6 Gy radiation on proliferation, migration, invasion, and apoptosis of hepatoma cells (P < 0.05). circ_0026134 knockdown significantly delayed tumor growth in vivo (P < 0.05). Conclusion: Silencing circ_0026134 strengthens radiation treatment's anti-proliferation, anti-migration, invasion, and pro-apoptotic effects in hepatoma cells by negatively regulating the miR-1270/GRB2 pathway, thereby enhancing radiosensitivity.

Harmine Hydrochloride Induces G2/M Cell Cycle Arrest and Apoptosis in SK-Hep1 Hepatocellular Carcinoma Cells by Regulating Mitogen-Activated Protein Kinases and the PI3K/AKT Pathway.

Liver cancer is a globally common form of cancer. Thus, novel drugs derived from natural products are needed to reduce the side effects of chemotherapy. The present study aimed to analyze the anticancer properties and effects of harmine hydrochloride (HMH), a water-soluble metabolite of harmine that can be easily absorbed into tissues, in treating liver cancer cells. HMH dose-dependently inhibited cell growth, migration, invasion, and colony formation in SK-Hep1 cells. It also induced G2/M arrest by reducing the expression of p-cdc2, cyclin B1, and Rb (G2/M phase regulatory proteins) in a dose-dependent manner. HMH treatment reduced the expression of caspase-9, caspase-3, PARP, and Bcl-2 and increased the expression of Bax (a proapoptotic protein). Moreover, it increased the production of reactive oxygen species and decreased the intracellular uptake of rhodamine 123 due to mitochondrial dysfunction because of oxidative stress. HMH treatment also upregulated the phosphorylation of JNK, p38, and FOXO3a in SK-Hep1 cells and downregulated the PI3K/AKT signaling pathway. Our findings suggest that HMH may activate the compounds responsible for anticancer effects in hepatocellular carcinoma cells.

Matrine inhibits hepatocellular carcinoma cell malignancy through the circ_0013290/miR-139-5p/MMP16 pathway.

Previous studies have shown the anticancer effect of Matrine on hepatocellular carcinoma (HCC); however, the underlying mechanism is still indistinct. The expression of circular RNA_0013290 (circ_0013290), microRNA-139-5p (miR-139-5p), matrix metallopeptidase 16 (MMP16), CyclinD1 and N-cadherin was analyzed by quantitative real-time polymerase chain reaction, Western blotting or immunohistochemistry assay. Cell viability, proliferation, apoptosis, invasion and tube formation were analyzed by cell counting kit-8, 5-Ethynyl-2'-deoxyuridine, flow cytometry analysis, transwell invasion and tube formation assays, respectively. The associations among circ_0013290, miR-139-5p and MMP16 were predicted by starbase online database, and identified by dual-luciferase reporter and RNA pull-down assays. A xenograft mouse model assay was conducted to disclose the effects of circ_0013290 and Matrine on tumor tumorigenesis in vivo. Circ_0013290 and MMP16 expression were significantly upregulated, while miR-139-5p was downregulated in HCC tissues and cells compared with the matched normal liver tissues and cells. Matrine treatment inhibited HCC cell proliferation, invasion and tube formation but induced cell apoptosis, accompanied by the decrease of CyclinD1 and N-cadherin expression; however, these effects were counteracted when circ_0013290 expression was increased. MiR-139-5p depletion or MMP16 introduction relieved Matrine-induced effects in HCC cells. The regulation of circ_0013290 toward HCC cell processes involved MMP16. With respect to the mechanism, circ_0013290 acted as a miR-139-5p sponge, and miR-139-5p targeted MMP16 in HCC cells. Besides, circ_0013290 regulated MMP16 expression through miR-139-5p. Further, circ_0013290 depletion enhanced the inhibitory effects of Matrine on tumor tumorigenesis. Matrine inhibited HCC cell malignancy through the circ_0013290/miR-139-5p/MMP16 pathway, suggesting that Matrine is a potential therapeutic agent for HCC.

FOXP2 suppresses the proliferation, invasion, and aerobic glycolysis of hepatocellular carcinoma cells by regulating the KDM5A/FBP1 axis.

The Warburg effect is the preference of cancer cells to use glycolysis rather than oxidative phosphorylation to generate energy. Accumulating evidence suggests that aerobic glycolysis is widespread in hepatocellular carcinoma (HCC) and closely related to tumorigenesis. The purpose of this study was to investigate the role and mechanism of forkhead box P2 (FOXP2) in aerobic glycolysis and tumorigenesis in HCC. Here, we found that FOXP2 was lower expressed in HCC tissues and cells than in nontumor tissues and normal hepatocytes. Overexpression of FOXP2 suppressed cell proliferation and invasion of HCC cells and promoted cell apoptosis in vitro, and hindered the growth of mouse xenograft tumors in vivo. Further researches showed that FOXP2 inhibited the Warburg effect in HCC cells. Moreover, we demonstrated that FOXP2 up-regulated the expression of fructose-1, 6-diphosphatase (FBP1), and the inhibitory effect of FOXP2 on glycolysis was dependent on FBP1. Mechanistically, as a transcription factor, FOXP2 negatively regulated the transcription of lysine-specific demethylase 5A (KDM5A), and then blocked KDM5A-induced H3K4me3 demethylation in FBP1 promoter region, thereby promoting the expression of FBP1. Consistently, overexpressing KDM5A or silencing FBP1 effectively reversed the inhibitory effect of FOXP2 on HCC progression. Together, our findings revealed the mechanistic role of the FOXP2/KDM5A/FBP1 axis in glycolysis and malignant progression of HCC cells, providing a potential molecular target for the therapy of HCC.

Long noncoding RNA CASC11 suppresses sorafenib-triggered ferroptosis via stabilizing SLC7A11 mRNA in hepatocellular carcinoma cells

As a frontline treatment for patients with advanced hepatocellular carcinoma (HCC), sorafenib is an effective drug approved by the Food and Drug Administration (FDA). Ferroptosis, a newly defined programmed cell death process with the hallmark of the accumulation of iron-dependent lipid peroxides, can be induced by sorafenib treatment. Our previous study identified oncogenic roles of long noncoding RNA (lncRNA) Cancer susceptibility candidate 11 (CASC11) in HCC progression. However, the relationship between CASC11 and sorafenib-induced ferroptosis in HCC remains unclear. In the present study, we aim to investigate the role of CASC11 in sorafenib-induced ferroptosis in HCC cell lines and determine the involved molecular mechanisms. Here, we demonstrated that sorafenib decreased CASCL11 expression. Knockdown of CASC11 enhanced sorafenib-induced ferroptosis, while overexpression of CASC11 exerted the opposite effect in HCC cells. Moreover, CASC11 led to the accumulation of intracellular malondialdehyde (MDA), lipid reactive oxygen species (ROS) and Fe2+ while depleting glutathione (GSH), thereby suppressing sorafenib-induced ferroptosis and cell death. Ferrostatin-1 (Ferr-1), a ferroptosis inhibitor, reversed the enhanced anticancer effect of sorafenib caused by the silence of CASC11 in HCC cells. Mechanistically, CASC11 upregulated the expression of solute carrier family 7 member 11 (SLC7A11) which is critical for ferroptosis inhibition. CASC11 associated with and stabilized SLC7A11 mRNA. In summary, our data revealed, for the first time, that CASC11 inhibits the sorafenib-induced ferroptosis in HCC cells via regulating SLC7A11, providing a new basis for clinical therapeutic strategies for patients with HCC.

Salvigenin Suppresses Hepatocellular Carcinoma Glycolysis and Chemoresistance Through Inactivating the PI3K/AKT/GSK-3β Pathway

Salvigenin is a Trimethoxylated Flavone enriched in Scutellariae Barbatae Herba and Scutellariae Radix and is demonstrated to have anti-tumor properties in colon cancer. Notwithstanding, the function and mechanism of Salvigenin in hepatocellular carcinoma (HCC) are less well studied. Different doses of Salvigenin were taken to treat HCC cells. Cell viability, colony formation ability, cell migration, invasion, apoptosis, glucose uptake, and lactate production levels were detected. As shown by the data, Salvigenin concentration dependently dampened HCC cell proliferation, migration, and invasion, weakened glycolysis by abating glucose uptake and lactate generation, and suppressed the profiles of glycolytic enzymes. Moreover, Salvigenin strengthened HCC cells’ sensitivity to 5-fluorouracil (5-FU) and attenuated HCC 5-FU-resistant cells’ resistance to 5-FU. Through network pharmacological analysis, we found Salvigenin potentially regulates PI3K/AKT pathway. As shown by the data, Salvigenin repressed the phosphorylated levels of PI3K, AKT, and GSK-3β. The PI3K activator 740Y-P induced PI3K/AKT/GSK-3β pathway activation and promotive effects in HCC cells. However, Salvigenin substantially weakened 740Y-P-mediated effects. In-vivo assay revealed that Salvigenin hampered the growth and promoted apoptosis of HCC cells in nude mice. Collectively, Salvigenin impedes the aerobic glycolysis and 5-FU chemoresistance of HCC cells by dampening the PI3K/AKT/GSK-3β pathway.

Yupingfeng San exhibits anticancer effect in hepatocellular carcinoma cells via the MAPK pathway revealed by HTS2 technology.

Yupingfeng San (YPFS) is a classic rousing prescription in Chinese medicine, with widly clinical application and remarkably curative effect. It consists of three herbs named Astragalus mongholicus Bunge (Huangqi), Atractylodes rubra Dekker (Baizhu) and Saposhnikovia divaricata (Turcz.) Schischk. (Fangfeng), and has a variety of pharmacological activities including immune regulation, antioxidant, anti-tumor, regulation of cytokines, etc. AIM OF THE STUDY: It has been proved that YPFS exerts its anti-tumor effect through enhancing the systemic and local immune responses in tumor patients, moreover, it has the direct tumor-suppressing effect and can reduce the adverse reactions caused by radiotherapy and chemotherapy drugs. Therefore, in this study, we explored the potential anti-HCC mechanism of YPFS based on HTS2 technology and system pharmacology, aiming to provide a scientific basis for the clinical application of YPFS and a new strategy for Chinese medicine research. In this study, systems pharmacology plus high throughput sequencing-based high throughput screening(HTS2) technology, and experimental validation were used to investigate the therapeutic mechanisms and the chemical basis of YPFS in HCC treatment. Firstly, the potential therapeutic targets and signaling pathways of YPFS in the treatment of HCC were obtained through systems pharmacology. Subsequently, HTS2 technology combined with PPI network analysis were used to reveal potential therapeutic targets. Finally, the anti-HCC effects and underlying mechanisms of YPFS were further verified in vitro in human hepatocellular carcinoma cell lines. Moreover, the possible chemical basis was explored by drug target verification and molecular docking technology. In total, 183 active ingredients were predicted by YPFS screening and 49 anti-HCC targets were further identified. Most of these targets were enriched into the "MAPK pathway", and the expression of 37 genes was significantly changed after herb treatment. Among them, 5 key targets, including VEGFA, GRB2, JUN, PDGFRB and CDC42, were predicted by protein-protein interaction (PPI) network analysis. According to our results, YPFS inhibited the proliferation, induced the apoptosis and caused cell cycle arrest of HCC cells. In addition, YPFS significantly reduced P38 gene expression. Fangfeng, one of the three herbs in YPFS, significantly down-regulated the expression of more target genes than that of the other two herbs. Lastly, as revealed by molecular docking analysis, 4'-O-glucosyl-5-O-methylvisamminol, an active ingredient identified in Fangfeng, showed a high affinity for P38. Taken together, this study shows that YPFS possesses the activities of anti-proliferation and pro-apoptosis in treating HCC, which are achieved by inhibiting the MAPK signaling pathway. P38 is one of the critical targets of YPFS in treating HCC, which may be directly bound and inhibited by 4'-O-glucosyl-5-O-methylvisamminol, a compound derived from YPFS.

Combination of Vitamin C and Lenvatinib potentiates antitumor effects in hepatocellular carcinoma cells in vitro.

Lenvatinib has become a first-line drug in the treatment of advanced hepatocellular carcinoma (HCC). Investigating its use in combination with other agents is of great significance to improve the sensitivity and durable response of Lenvatinib in advanced HCC patients. Vitamin C (L-ascorbic acid, ascorbate, VC) is an important natural antioxidant, which has been reported to show suppressive effects in cancer treatment. Here, we investigated the effect of the combination of VC and Lenvatinib in HCC cells in vitro. We found that treatment of VC alone significantly inhibited the proliferation, migration and invasion in HCC cells. Additionally, VC was strongly synergistic with Lenvatinib in inhibition of the proliferative, migratory and invasive capacities of HCC cells in vitro. In conclusion, our results demonstrate that the combination of VC and Lenvatinib has synergistic antitumor activities against HCC cells, providing a promising therapeutic strategy to improve the prognosis of HCC patients.

METTL5 stabilizes c-Myc by facilitating USP5 translation to reprogram glucose metabolism and promote hepatocellular carcinoma progression.

Hepatocellular carcinoma (HCC) is one of the most prevalent cancers in the world, with a high likelihood of metastasis and a dismal prognosis. The reprogramming of glucose metabolism is critical in the development of HCC. The Warburg effect has recently been confirmed to occur in a variety of cancers, including HCC. However, little is known about the molecular biological mechanisms underlying the Warburg effect in HCC cells. In this study, we sought to better understand how methyltransferase 5, N6-adenosine (METTL5) controls the development of HCC and the Warburg effect. In the current study, quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of METTL5 in HCC tissues and cell lines. Several different cell models and animal models were established to determine the role of METTL5 in glucose metabolism reprogramming and the underlying molecular mechanism of HCC. Glutathione-S-transferase pulldown, coimmunoprecipitation, RNA sequencing, non-targeted metabolomics, polysome profiling, and luciferase reporter assays were performed to investigate the molecular mechanisms of METTL5 in HCC cells. We discovered that METTL5 drove glucose metabolic reprogramming to promote the proliferation and metastasis of HCC. Mechanistically, upregulation of METTL5 promoted c-Myc stability and thus activated its downstream glycolytic genes lactate dehydrogenase A (LDHA), enolase 1 (ENO1), triosephosphate isomerase 1 (TPI1), solute carrier family 2 member 1 (SLC2A1), and pyruvate kinase M2 (PKM2). The c-Box and ubiquitin binding domain (UBA) regions of ubiquitin specific peptidase 5 (USP5) binded to c-Myc protein and inhibited K48-linked polyubiquitination of c-Myc. Further study revealed that METTL5 controled the USP5 translation process, which in turn regulated the ubiquitination of c-Myc. Furthermore, we identified cAMP responsive element binding protein 1 (CREB1)/P300 as a critical transcriptional regulator of METTL5 that promoted the transcription of METTL5 in HCC. In patient-derived tumor xenograft (PDX) models, adenovirus-mediated knockout of METTL5 had a good antitumor effect and prolonged the survival of PDX-bearing mice. These findings point to a novel mechanism by which CREB1/P300-METTL5-USP5-c-Myc controls abnormal glucose metabolism and promotes tumor growth, suggesting that METTL5 is a potential therapeutic target and prognostic biomarker for HCC.

LncRNA NUTM2A-AS1 aggravates the progression of hepatocellular carcinoma by activating the miR-186-5p/KLF7-mediated Wnt/beta-catenin pathway.

Emerging evidence has uncovered that noncoding RNAs (ncRNAs) contribute to the development of hepatocellular carcinoma (HCC). Nevertheless, the functions of the majority of long ncRNAs (lncRNAs) in HCC are unknown. Here, we intend to probe the function of lncRNA NUTM2A-AS1 in the evolvement of HCC and the related mechanism. Expression levels of lncRNA NUTM2A-AS1, miR-186-5p and KLF7 mRNA in HCC tissues and adjacent non-tumor tissues were monitored. Gain- or loss-of-function assays were utilized to investigate the biological functions of lncRNA NUTM2A-AS1, miR-186-5p and KLF7 in HCC cell lines (including HCCLM3 and Huh7). Western blot was implemented for the detection of the epithelial-mesenchymal transition (EMT)-related proteins (including E-cadherin, Vimentin and Snail), KLF7, Wnt, β-catenin, and stemness-related proteins (Nanog, OCT4, YKL40, and CD133). Furthermore, the targeted associations between lncRNA NUTM2A-AS1, miR-186-5p, and KLF7 were verified by bioinformatics analysis, dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. As a result, lncRNA NUTM2A-AS1 and KLF7 profiles were heightened in the HCC tissues versus adjacent normal tissues, while miR-186-5p had the opposite expression tendency. Up-regulation of lncRNA NUTM2A-AS1 was related to tumor size, advanced tumor stage, and lymph node metastasis of HCC patients. Functionally, overexpression of lncRNA NUTM2A-AS1 heightened HCC cells' growth, invasion, EMT, and stemness and repressed their apoptosis by activating the Wnt/β-catenin pathway. In contrast, up-regulation of miR-186-5p or inhibition of KLF7 had reverse effects. In vivo, lncRNA NUTM2A-AS1 overexpression facilitated tumor growth and EMT, accompanied by declined miR-186-5p levels and enhanced KLF7 expression. The mechanistic studies revealed that miR-186-5p served as a common target of lncRNA NUTM2A-AS1 and KLF7. As hinted by the rescue experiments, NUTM2A-AS1 partly abated miR-186-5p-mediated anti-tumor effects in HCC cells, whereas KLF7 knockdown reversed the promotive effects of NUTM2A-AS1. LncRNA NUTM2A-AS1 accelerated the evolution of HCC by up-regulating the KLF7/Wnt/beta-catenin pathway through sponging miR-186-5p.

The Elevated Circ_0067835 Could Accelerate Cell Proliferation and Metastasis via miR-1236-3p/Twist2 Axis in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a malignant cancer with leading mortality worldwide. Circ_0067835 is a circRNA which plays an important role in various kinds of tumor, while the potential functions of circ_0067835 in HCC remains unclear. In this study, our results of microarray and real-time PCR (RT-PCR) showed that it was obviously elevated in human HCC tumor tissues and HCC cell lines. Inhibition of circ_0067835 restrained cell proliferation and migration in vitro. Furthermore, miR-1236-3p was decreased in tumor samples, and it was indicated to be a target of circ_0067835. Moreover, Twist2 was established to be elevated in HCC tissues, and we identified it as the direct target of miR-1236-3p. Finally, we found that knockdown of miR-1236-3p could reverse the circ_0067835 inhibition effects in HCC cells. In conclusion, our study demonstrated that circ_0067835 contributed to promoting hepatocellular carcinoma cell proliferation and metastasis through downregulating miR-1236-3p expression and then elevating Twist2 expression, which might provide a new vision for HCC patients.

Hepatocellular cancer cell lines, Hep-3B and Hep-G2 display the pleiotropic response to resveratrol and berberine

PurposeHuman carcinoma cells with different p53 status exposed to a combination of bioactive substances, resveratrol and berberine, revealed different responses in cell viability via p53-dependant apoptosis pathway activation. Materials and methodsUsing 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, we investigated various and opposing effects in hepatocellular carcinoma cells, Hep-G2 and Hep-3B with different p53-status. ResultsCells decreased in viability after treatment with dose-dependent concentrations of resveratrol and berberine. Hep-3B p53 mutants were more sensitive in comparison to the p53 wild type Hep-G2 cell line. A synergistic effect was observed after treatment of Hep-3B cells with a combination of resveratrol/berberine ratios in favor of resveratrol (2:1, 3:1). The results suggest that an effective concentration of berberine, in the presence of resveratrol, could be decreased even to 50% (half the IC50 for berberine) in cancer treatment. Combined treatment with berberine and resveratrol, at the investigated concentrations and fractions, significantly reduces the viability of wild type p53 Hep-G2 and null p53-mutant Hep-3B cells by 20% and 40%, respectively. ConclusionsStronger toxic effects on viability and proliferation were observed in Hep-3B cells what is consistent with the assumptions that null p53-mutants activate apoptosis canonical pathway. In conclusion, p53 status in human hepatocellular cancer cell lines modulates responses to plant-derived therapies.

ANKFN1 plays both protumorigenic and metastatic roles in hepatocellular carcinoma

Ankyrin repeat and fibronectin type III domain containing 1 (ANKFN1) is reported to be involved in human height and developmental abnormalities, but the expression profile and molecular function of ANKFN1 in hepatocellular carcinoma (HCC) remain unknown. This study aimed to evaluate the clinical significance and biological function of ANKFN1 in HCC and investigate whether ANKFN1 can be used for differential diagnosis in HCC. Here, we showed that ANKFN1 was upregulated in 126 tumor tissues compared with adjacent nontumorous tissues in HCC patients. The upregulation of ANKFN1 in HCC was associated with cirrhosis, alpha-fetoprotein (AFP) levels and poor prognosis. Moreover, silencing ANKFN1 expression suppressed HCC cell proliferation, migration, invasion, and metastasis in vitro and subcutaneous tumorigenesis in vivo. However, ANKFN1 overexpression promoted HCC proliferation and metastasis in an orthotopic liver transplantation model and attenuated the above biological effects in HCC cells. ANKFN1 significantly affected HCC cell proliferation by inducing G1/S transition and cell apoptosis. Mechanistically, we demonstrated that ANKFN1 promoted cell proliferation, migration, and invasion via activation of the cyclin D1/Cdk4/Cdk6 pathway by stimulating the MEK1/2-ERK1/2 pathway. Moreover, ANKFN1-induced cell proliferation, migration, and invasion were partially reversed by ERK1/2 inhibitors. Taken together, our results indicate that ANKFN1 promotes HCC cell proliferation and metastasis by activating the MEK1/2-ERK1/2 signaling pathway. Our work also suggests that ANKFN1 is a potential therapeutic target for HCC.