The long intergenic noncoding RNA UFC1, a target of MicroRNA 34a, interacts with the mRNA stabilizing protein HuR to increase levels of ß-catenin in HCC cells.

BACKGROUND & AIMS: Altered activities of long noncoding RNAs (lncRNAs) have been associated with cancer development. We investigated the mechanisms by which the long intergenic noncoding RNA UFC1 (lincRNA-UFC1) promotes progression of hepatocellular carcinoma (HCC), using human tissues and cell lines. METHODS: We used microarrays to compare expression profiles of lncRNAs in HCC samples and adjacent nontumor tissues (controls) from 7 patients. HCC and nontumor tissues were collected from 2006 through 2012 from patients in Guangzhou, China. We used quantitative real-time polymerase chain reaction to measure levels of lincRNA-UFC1 in tissues from 49 patients, and in situ hybridization to measure levels in samples from 131 patients; clinical data were collected from patients for up to 5 years. The lincRNA-UFC1 was expressed transgenically, or knocked down with short hairpin RNAs, in BEL-7402, SK-Hep1, Huh7, and MHCC-97H HCC cell lines; luciferase reporter and RNA immunoprecipitation and pull-down assays were performed. We also analyzed growth of xenograft tumors from these cells in BALB/c nude mice. RESULTS: Levels of the lincRNA-UFC1 were increased in HCC tissues compared with controls, and associated with tumor size, Barcelona Clinic Liver Cancer stage, and patient outcomes. Transgenic expression of the lincRNA-UFC1 in HCC cells promoted their proliferation and cell-cycle progression and inhibited apoptosis, whereas short hairpin RNA knockdown of lincRNA-UFC1 had opposite effects. Xenograft tumors grown from cells overexpressing lincRNA-UFC1 had larger mean volumes and weights, and formed more rapidly, than tumors grown from control cells. Tumors grown from lincRNA-UFC1 knockdown were smaller than controls. The lincRNA-UFC1 interacted directly with the messenger RNA (mRNA) stabilizing protein HuR (encoded by ELAVL1) to increase levels of ß-catenin mRNA (encoded by CTNNB1) and protein. Levels of lincRNA-UFC1 correlated with those of ß-catenin in HCC tissues. In contrast, there was a negative correlation between levels of microRNA 34a and lincRNA-UFC1 in HCC tissues; microRNA 34a reduced the stability of lincRNA-UFC1. CONCLUSIONS: The lincRNA-UFC1, a target of microRNA 34a, promotes proliferation and reduces apoptosis in HCC cells to promote growth of xenograft tumors in mice. It interacts directly with the mRNA stabilizing protein HuR to regulate levels of ß-catenin in HCC cells.

SNHG3 correlates with malignant status and poor prognosis in hepatocellular carcinoma.

Long noncoding RNAs (lncRNAs) have been found dysregulated in human disease, especially in cancer. Small nucleolar RNA host gene 3 (SNHG3) is an lncRNA whose potential function and mechanism in hepatocellular carcinoma (HCC) remain largely unknown. In the present study, we aimed to determine SNHG3 expression and its clinical significance in HCC. Our results showed that the expression level of SNHG3 was significantly upregulated in HCC tissues compared with paired noncancerous tissues from 51 HCC patients, as determined by quantitative real-time polymerase chain reaction (qRT-PCR; P < 0.001), which was consistent with the results of two independent HCC cohorts from The Cancer Genome Atlas (TCGA) and Oncomine databases (P < 0.0001 and P = 0.0325, respectively). These results were further confirmed in 144 paired paraffin-embedded HCC specimens by in situ hybridization assay (ISH). Furthermore, SNHG3 expression was significantly correlated with tumor size (P = 0.003), portal vein tumor thrombus (PVTT; P = 0.014), and relapse (P = 0.038). The high expression level of SNHG3 was markedly correlated with overall survival (OS; P < 0.0001), recurrence-free survival (RFS; P = 0.006), and disease-free survival (DFS; P < 0.0001). More importantly, multivariate analysis indicated that SNHG3 expression was an independent prognostic factor for HCC patients (P < 0.001). In conclusion, increased SNHG3 expression is associated with malignant status and poor prognosis in HCC patients.

Single-cell Transcriptomic Analysis Reveals an Immunosuppressive Network Between POSTN CAFs and ACKR1 ECs in TKI-resistant Lung Cancer.

BACKGROUND/AIM: Tyrosine kinase inhibitor (TKI) therapy, a principal treatment for advanced non-small cell lung cancer (NSCLC), frequently encounters the development of drug resistance. The tumor microenvironment (TME) plays a critical role in the progression of NSCLC, yet the relationship between endothelial cells (ECs) and cancer-associated fibroblasts (CAFs) subpopulations in TKI treatment resistance remains largely unexplored. MATERIALS AND METHODS: The BioProject database PRJNA591860 project was used to analyze scRNA-seq data including 49 advanced-stage NSCLC samples across three different time points: pre-targeted therapy (naïve), post-partial response (PR) to targeted therapy, and post-progressive disease (PD) stage. The data involved clustering stromal cells into multiple CAFs and ECs subpopulations. The abundance changes and functions of each cluster during TKI treatment were investigated by KEGG and GO analysis. Additionally, we identified specific transcription factors and metabolic pathways via DoRothEA and scMetabolism. Moreover, cell-cell communications between PD and PR stages were compared by CellChat. RESULTS: ECs and CAFs were clustered and annotated using 49 scRNA-seq samples. We identified seven ECs subpopulations, with OIT3 ECs showing enrichment in the PR phase with a drug-resistance phenotype, and ACKR1 ECs being prevalent in the PD phase with enhanced cell adhesion. Similarly, CAFs were clustered into 7 subpopulations. PLA2G2A CAFs were predominant in PR, whereas POSTN CAFs were prevalent in PD, characterized by an immunomodulatory phenotype and increased collagen secretion. CellChat analysis showed that ACKR1 ECs strongly interacted with macrophage through the CD39 pathway and POSTN CAFs secreted Tenascin-C (TNC) to promote the progression of epithelial cells, primarily malignant ones, in PD. CONCLUSION: This study reveals that POSTN CAFs and ACKR1 ECs are associated with resistance to TKI treatment, based on single-cell sequencing.

Single-cell transcriptomics reveal the microenvironment landscape of perfluorooctane sulfonate-induced liver injury in female mice.

Epidemic and animal studies have reported that perfluoroalkyl and polyfluoroalkyl substances (PFASs) are strongly associated with liver injury; however, to date, the effects of PFASs on the hepatic microenvironment remain largely unknown. In this study, we established perfluorooctane sulfonic acid (PFOS)-induced liver injury models by providing male and female C57BL/6 mice with water containing PFOS at varying doses for 4 weeks. Hematoxylin and eosin staining revealed that PFOS induced liver injury in both sexes. Elevated levels of serum aminotransferases including those of alanine aminotransferase and aspartate transaminase were detected in the serum of mice treated with PFOS. Female mice exhibited more severe liver injury than male mice. We collected the livers from female mice and performed single-cell RNA sequencing. In total, 36,529 cells were included and grouped into 10 major cell types: B cells, granulocytes, T cells, NK cells, monocytes, dendritic cells, macrophages, endothelial cells, fibroblasts, and hepatocytes. Osteoclast differentiation was upregulated and the T cell receptor signaling pathway was significantly downregulated in PFOS-treated livers. Further analyses revealed that among immune cell clusters in PFOS-treated livers, Tcf7+CD4+T cells were predominantly downregulated, whereas conventional dendritic cells and macrophages were upregulated. Among the fibroblast subpopulations, hepatic stellate cells were significantly enriched in PFOS-treated female mice. CellphoneDB analysis suggested that fibroblasts interact closely with endothelial cells. The major ligand-receptor pairs between fibroblasts and endothelial cells in PFOS-treated livers were Dpp4_Cxcl12, Ackr3_Cxcl12, and Flt1_complex_Vegfa. These genes are associated with directing cell migration and angiogenesis. Our study provides a general framework for understanding the microenvironment in the livers of female mice exposed to PFOS at the single-cell level.

UBE2T-mediated Akt ubiquitination and Akt/ß-catenin activation promotes hepatocellular carcinoma development by increasing pyrimidine metabolism.

The oncogene protein ubiquitin-conjugating enzyme E2T (UBE2T) is reported to be upregulated in hepatocellular carcinoma (HCC) and correlated with poor clinical outcomes of HCC patients. However, the underlying mechanism by which UBE2T exerts its oncogenic function in HCC remains largely unexplored. In this study, in vitro and in vivo experiments suggested that UBE2T promoted HCC development including proliferation and metastasis. GSEA analysis indicated that UBE2T was positively correlated with pyrimidine metabolism, and LC/MS-MS metabolomics profiling revealed that the key products of pyrimidine metabolism were significantly increased in UBE2T-overexpressing cells. UBE2T overexpression led to the upregulation of several key enzymes catalyzing de novo pyrimidine synthesis, including CAD, DHODH, and UMPS. Moreover, the utilization of leflunomide, a clinically approved DHODH inhibitor, blocked the effect of UBE2T in promoting HCC progression. Mechanistically, UBE2T increased Akt K63-mediated ubiquitination and Akt/ß-catenin signaling pathway activation. The disruption of UBE2T-mediated ubiquitination on Akt, including E2-enzyme-deficient mutation (C86A) of UBE2T and ubiquitination-site-deficient mutation (K8/14 R) of Akt impaired UBE2T's effect in upregulating CAD, DHODH, and UMPS. Importantly, we demonstrated that UBE2T was positively correlated with p-Akt, ß-catenin, CAD, DHODH, and UMPS in HCC tumor tissues. In summary, our study indicates that UBE2T increases pyrimidine metabolism by promoting Akt K63-linked ubiquitination, thus contributing to HCC development. This work provides a novel insight into HCC development and a potential therapeutic strategy for HCC patients.

Wnt/ß-catenin inhibitor ICG-001 enhances the antitumor efficacy of radiotherapy by increasing radiation-induced DNA damage and improving tumor immune microenvironment in hepatocellular carcinoma.

BACKGROUND AND PURPOSE: Radiotherapy (RT) has a promising anti-tumor effect depending on its effects on both cancer cells and tumor immune microenvironment (TIME). As one of the most common alterations in hepatocellular carcinoma (HCC), wnt/ß-catenin pathway activation, has been reported to induce radioresistance and suppressive TIME. In this study, we aim to explore the effect of wnt/ß-catenin inhibitor ICG-001 on radiosensitivity and RT-related TIME of HCC and the underlying mechanism. MATERIALS AND METHODS: C57BL/6 and nude mouse tumor models were used to evaluate the efficacy of different treatments on tumor growth, recurrence and mice survival. Flow cytometry was performed to assess tumor infiltrating lymphocytes (TILs). DNA damage response (DDR) and radioresistance was investigated by colony formation assays, γ-H2AX and micronuclei measurements. RESULTS: The addition of ICG-001 to RT exhibited better anti-tumor and survival-prolong efficacy in C57BL/6 than nude mice. TILs analysis revealed that ICG-001 plus RT boosted the infiltration and IFN-γ production of TIL CD8+ T cells, meanwhile reduced the number of Tregs. Moreover, mechanistic study demonstrated that ICG-001 increased the radiation-induced DDR of HCC cells by suppressing p53, thus leading to stronger activation of cGAS/STING pathway. Utilization of cGAS/STING pathway inhibitors impaired the therapeutic effect of combination therapy. Furthermore, combination therapy led to stronger immunologic memory and tumor relapse prevention. CONCLUSIONS: Our findings showed that ICG-001 displayed both local and systematic effects by increasing radiosensitivity and improving immunity in HCC, which indicated that ICG-001 might be a potential synergetic treatment for radiotherapy and radioimmunotherapy in HCC patients.

UBE2T-regulated H2AX monoubiquitination induces hepatocellular carcinoma radioresistance by facilitating CHK1 activation.

BACKGROUND: Radioresistance is the major obstacle in radiation therapy (RT) for hepatocellular carcinoma (HCC). Dysregulation of DNA damage response (DDR), which includes DNA repair and cell cycle checkpoints activation, leads to radioresistance and limits radiotherapy efficacy in HCC patients. However, the underlying mechanism have not been clearly understood. METHODS: We obtained 7 pairs of HCC tissues and corresponding non-tumor tissues, and UBE2T was identified as one of the most upregulated genes. The radioresistant role of UBE2T was examined by colony formation assays in vitro and xenograft tumor models in vivo. Comet assay, cell cycle flow cytometry and γH2AX foci measurement were used to investigate the mechanism by which UBE2T mediating DDR. Chromatin fractionation and immunofluorescence staining were used to assess cell cycle checkpoint kinase 1(CHK1) activation. Finally, we analyzed clinical data from HCC patients to verify the function of UBE2T. RESULTS: Here, we found that ubiquitin-conjugating enzyme E2T (UBE2T) was upregulated in HCC tissues, and the HCC patients with higher UBE2T levels exhibited poorer outcomes. Functional studies indicated that UBE2T increased HCC radioresistance in vitro and in vivo. Mechanistically, UBE2T-RNF8, was identified as the E2-E3 pair, physically bonded with and monoubiquitinated histone variant H2AX/γH2AX upon radiation exposure. UBE2T-regulated H2AX/γH2AX monoubiquitination facilitated phosphorylation of CHK1 for activation and CHK1 release from the chromatin to cytosol for degradation. The interruption of UBE2T-mediated monoubiquitination on H2AX/γH2AX, including E2-enzyme-deficient mutation (C86A) of UBE2T and monoubiquitination-site-deficient mutation (K119/120R) of H2AX, cannot effectively activate CHK1. Moreover, genetical and pharmacological inhibition of CHK1 impaired the radioresistant role of UBE2T in HCC. Furthermore, clinical data suggested that the HCC patients with higher UBE2T levels exhibited worse response to radiotherapy. CONCLUSION: Our results revealed a novel role of UBE2T-mediated H2AX/γH2AX monoubiquitination on facilitating cell cycle arrest activation to provide sufficient time for radiation-induced DNA repair, thus conferring HCC radioresistance. This study indicated that disrupting UBE2T-H2AX-CHK1 pathway maybe a promising potential strategy to overcome HCC radioresistance.

ATR inhibitor AZD6738 enhances the antitumor activity of radiotherapy and immune checkpoint inhibitors by potentiating the tumor immune microenvironment in hepatocellular carcinoma.

BACKGROUND: Radioimmunotherapy has a promising antitumor effect in hepatocellular carcinoma (HCC), depending on the regulatory effect of radiotherapy on tumor immune microenvironment. Ionizing radiation (IR)-induced DNA damage repair (DDR) pathway activation leads to the inhibition of immune microenvironment, thus impairing the antitumor effect of radioimmunotherapy. However, it is unclear whether inhibition of the DDR pathway can enhance the effect of radioimmunotherapy. In this study, we aim to explore the role of DDR inhibitor AZD6738 on the combination of radiotherapy and immune checkpoint inhibitors (ICIs) in HCC. METHODS: C57BL/6 mouse subcutaneous tumor model was used to evaluate the ability of different treatment regimens in tumor growth control and tumor recurrence inhibition. Effects of each treatment regimen on the alterations of immunophenotypes including the quantification, activation, proliferating ability, exhaustion marker expression, and memory status were assessed by flow cytometry. RESULTS: AZD6738 further increased radiotherapy-stimulated CD8+ T cell infiltration and activation and reverted the immunosuppressive effect of radiation on the number of Tregs in mice xenografts. Moreover, compared with radioimmunotherapy (radiotherapy plus anti-PD-L1 (Programmed death ligand 1)), the addition of AZD6738 boosted the infiltration, increased cell proliferation, enhanced interferon (IFN)-γ production ability of TIL (tumor-infiltrating lymphocyte) CD8+ T cells, and caused a decreasing trend in the number of TIL Tregs and exhausted T cells in mice xenografts. Thus, the tumor immune microenvironment was significantly improved. Meanwhile, triple therapy (AZD6738 plus radiotherapy plus anti-PD-L1) also induced a better immunophenotype than radioimmunotherapy in mice spleens. As a consequence, triple therapy displayed greater benefit in antitumor efficacy and mice survival than radioimmunotherapy. Mechanism study revealed that the synergistic antitumor effect of AZD6738 with radioimmunotherapy relied on the activation of cyclic GMP-AMP synthase /stimulator of interferon genes (cGAS/STING) signaling pathway. Furthermore, triple therapy led to stronger immunologic memory and lasting antitumor immunity than radioimmunotherapy, thus preventing tumor recurrence in mouse models. CONCLUSIONS: Our findings indicate that AZD6738 might be a potential synergistic treatment for radioimmunotherapy to control the proliferation of HCC cells, prolong survival, and prevent tumor recurrence in patients with HCC by improving the immune microenvironment.

Development and validation of a genomic mutation signature to predict response to PD-1 inhibitors in non-squamous NSCLC: a multicohort study

BACKGROUND: Genetic variations of some driver genes in non-small cell lung cancer (NSCLC) had shown potential impact on immune microenvironment and associated with response or resistance to programmed cell death protein 1 (PD-1) blockade immunotherapy. We therefore undertook an exploratory analysis to develop a genomic mutation signature (GMS) and predict the response to anti-PD-(L)1 therapy. METHODS: In this multicohort analysis, 316 patients with non-squamous NSCLC treated with anti-PD-(L)1 from three independent cohorts were included in our study. Tumor samples from the patients were molecularly profiled by MSK-IMPACT or whole exome sequencing. We developed a risk model named GMS based on the MSK training cohort (n=123). The predictive model was first validated in the separate internal MSK cohort (n=82) and then validated in an external cohort containing 111 patients from previously published clinical trials. RESULTS: A GMS risk model consisting of eight genes (TP53, KRAS, STK11, EGFR, PTPRD, KMT2C, SMAD4, and HGF) was generated to classify patients into high and low GMS groups in the training cohort. Patients with high GMS in the training cohort had longer progression-free survival (hazard ratio (HR) 0.41, 0.28-0.61, p<0.0001) and overall survival (HR 0.53, 0.32-0.89, p=0.0275) compared with low GMS. We noted equivalent findings in the internal validation cohort and in the external validation cohort. The GMS was demonstrated as an independent predictive factor for anti-PD-(L)1 therapy comparing with tumor mutational burden. Meanwhile, GMS showed undifferentiated predictive value in patients with different clinicopathological features. Notably, both GMS and PD-L1 were independent predictors and demonstrated poorly correlated; inclusion of PD-L1 with GMS further improved the predictive capacity for PD-1 blockade immunotherapy. CONCLUSIONS: Our study highlights the potential predictive value of GMS for immunotherapeutic benefit in non-squamous NSCLC. Besides, the combination of GMS and PD-L1 may serve as an optimal partner in guiding treatment decisions for anti-PD-(L)1 based therapy.

[Long noncoding RNA UFC1 promotes metastasis and invasion of hepatocellular carcinoma cells in vitro via GSK-3ß/ß-catenin axis].

OBJECTIVE: To explore the role of Long noncoding RNA UFC1 (lincRNA-UFC1) in modulating the metastasis and invasion of hepatocellular carcinoma (HCC) cells and the underlying mechanism. METHODS: Human HCC cell line Huh7 was infected with the lentiviral vector carrying lincRNA-UFC1 to obtain a cell line with lincRNA-UFC1 overexpression. A short hairpin RNA (shRNA) targeting lincRNA-UFC1 was delivered in human HCC BEL-7402 cells via a lentiviral vector to obtain a cell line with lincRNA-UFC1 knockdown. Expression levels of lincRNA-UFC1 in the two HCC cell lines were detected using real-time PCR, and the changes in the cell invasion and migration in response to lincRNA-UFC1 overexpression or knockdown were analyzed using Transwell and wound-healing assays. The expressions of GSK-3ß/ß-catenin-related proteins in the cells were detected with Western blotting. XAV-939, a GSK-3ß/ß-catenin inhibitor, was used for assessing the impact of lincRNAUFC1 overexpression on the invasion and migration of the HCC cells through Transwell and wound-healing assays. RESULTS: Overexpression of lincRNA-UFC1 significantly promoted the invasion and migration of Huh7 cells as compared with the control cells (P < 0.001), while lincRNA-UFC1 knockdown obviously suppressed the invasion and migration of BEL-7402 cells (P < 0.001). The results of Western blotting showed that the expressions of proteins associated with the cell invasion and migration, namely ß-catenin and P-GSK-3ß, were significantly upregulated in response to lincRNA-UFC1 overexpression, and were obviously lowered after lincRNA-UFC1 knockdown. Treatment of the cells with XAV-939 significantly reversed the effect of lincRNA-UFC1 overexpression on the cell invasion and migration (P < 0.001). CONCLUSIONS: lincRNA-UFC1 overexpresison promotes cell invasion and migration through the GSK-3ß/ß-catenin axis in HCC cells in vitro.

Knockdown of UACA inhibitsproliferation and invasion and promotes senescence of hepatocellular carcinoma cells.

Uveal autoantigen with coiled-coil domains and ankyrin repeats (UACA/Nucling), has been reported to be upregulated in various cancers. However, its expression and function have not been studied in hepatocellular carcinoma (HCC). In the present study, expression of UACA was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and the results revealed that UACA was upregulated in 23 cases of HCC compared with paired corresponding non-tumor liver tissues. In addition, the upregulation of UACA in HCC was further validated by analyzing the datasets from The Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) and GSE36376. Furthermore, knockdown of UACA suppressed the proliferative and invasive ability as well as inducing senescence of HCC cells. Besides, the expression level of UACA was positively associated with Hif1α (hypoxia-inducible factor 1α) in HCC datasets from TCGA-LIHC and GSE54236. Moreover, treatment with CoCl2 led to the increased expression and the localization alteration of UACA in HCC cells. In summary, UACA is upregulated in HCC and knockdown of UACA ameliorated malignant behaviors of HCC cells, and UACA was correlated with and under control of Hif1α.

Identification and Functional Characterization of Long Non-coding RNA MIR22HG as a Tumor Suppressor for Hepatocellular Carcinoma.

Long non-coding RNAs (lncRNAs) have recently been identified as critical regulators in tumor initiation and development. However, the function of lncRNAs in human hepatocellular carcinoma (HCC) remains largely unknown. Our study was designed to explore the biological function and clinical implication of lncRNA MIR22HG in HCC. Methods: We evaluated MIR22HG expression in 52-patient, 145-patient, TCGA, and GSE14520 HCC cohorts. The effects of MIR22HG on HCC were analyzed in terms of proliferation, invasion, and metastasis, both in vitro and in vivo. The mechanism of MIR22HG action was explored through bioinformatics, luciferase reporter, and RNA immunoprecipitation analyses. Results:MIR22HG expression was significantly down-regulated in 4 independent HCC cohorts compared to that in controls. Its low expression was associated with tumor progression and poor prognosis of patients with HCC. Forced expression of MIR22HG in HCC cells significantly suppressed proliferation, invasion, and metastasis in vitro and in vivo. Mechanistically, MIR22HG derived miR-22-3p to target high mobility group box 1 (HMGB1), thereby inactivating HMGB1 downstream pathways. Additionally, MIR22HG directly interacted with HuR and regulated its subcellular localization. MIR22HG competitively bound to human antigen R (HuR), resulting in weakened expression of HuR-stabilized oncogenes, such as ß-catenin. Furthermore, miR-22-3p suppression, HuR or HMGB1 overexpression rescued the inhibitory effects caused by MIR22HG overexpression. Conclusion: Our findings revealed that MIR22HG plays a key role in tumor progression by suppressing the proliferation, invasion, and metastasis of tumor cells, suggesting its potential role as a tumor suppressor and prognostic biomarker in HCC.

miR-139-5p inhibits aerobic glycolysis, cell proliferation, migration, and invasion in hepatocellular carcinoma via a reciprocal regulatory interaction with ETS1.

Cancer cells have metabolic features that allow them to preferentially metabolize glucose through aerobic glycolysis, providing them with a progression advantage. However, microRNA (miRNA) regulation of aerobic glycolysis in cancer cells has not been extensively investigated. We addressed this in the present study by examining the regulation of miR-139-5p on aerobic glycolysis of hepatocellular carcinoma (HCC) using clinical specimens, HCC cells, and a mouse xenograft model. We found that overexpressing miR-139-5p restrained aerobic glycolysis, suppressing proliferation, migration, and invasion in HCC cells. miR-139-5p regulated hexokinase 1 (HK1) and 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) expression by directly targeting the transcription factor ETS1, which bound to the promoters of the HK1 and PFKFB3 genes. miR-139-5p-induced aerobic glycolysis, proliferation, migration, and invasion were reversed by ETS1 overexpression, while ETS1 silencing induced the expression of miR-139-5p via a post-transcriptional regulation mode involving Drosha. miR-139-5p expression was reduced in HCC compared to para-carcinoma tissue, which was confirmed in The Cancer Genome Atlas and GSE54751 HCC cohorts. Notably, the lower expression of mir-139 was correlated with worse prognosis. These outcomes indicate that reciprocal regulatory interactions between miR-139-5p and ETS1 modulate aerobic glycolysis, proliferation, and metastasis in HCC cells, suggesting new targets for HCC treatment.

Up-regulation of LncRNA SNHG20 Predicts Poor Prognosis in Hepatocellular Carcinoma.

Recent studies indicated that long noncoding RNAs (lncRNAs) played important regulatory roles in carcinogenesis and cancer progression. However, the contribution of small nucleolar RNA host gene 20 (SNHG20) to cancer development remains largely unknown. The aim of the study is to investigate the expression of SNHG20 and its clinical significance in hepatocellular carcinoma (HCC). Our results showed that the expression of SNHG20 was remarkably up-regulated in HCC tissues compared with adjacent non-tumor liver tissues from 49 fresh HCC samples (cohort 1) detected by quantitative reverse-transcription polymerase chain reaction (qRT-PCR, P = 0.004). The results were confirmed in 144 formalin-fixed, paraffin-embedded HCC tissues (cohort 2) by in situ hybridization (ISH). Meanwhile, the expression of SNHG20 was associated with tumor size (P = 0.027 for cohort 1 and P = 0.046 for cohort 2) and clinical stage (P = 0.027 for cohort 1 and P = 0.028 for cohort 2). Importantly, patients with high expression of SNHG20 had a shorter overall survival (OS, P < 0.001) and disease-free survival (DFS, P < 0.001) than those with low expression of SNHG20. Univatiate and multivariate analysis showed that SNHG20 was a significant and independent prognostic predictor for OS of HCC patients (hazard ratio = 3.985, 95% CI = 1.981-8.017, P < 0.001). In addition, a total of 331 HCC patients' data from the Caner Genome Atlas project (TCGA) were used to validate our findings. Consistently, the results from TCGA HCC cohort demonstrated that SNHG20 were overexpressed in HCC tissues compared with non-tumor liver tissues (P < 0.001). Patients with higher expression levels of SNHG20 had poorer OS (P = 0.021) and DFS (P < 0.001). Functionally, knockdown of SNHG20 in SK-Hep-1 cells significantly inhibited cellular proliferation, migration, and invasion. In conclusion, SNHG20, up-regulated in patients with HCC, may serve as an independent prognostic predictor for HCC patients.

UBE2T promotes nasopharyngeal carcinoma cell proliferation, invasion, and metastasis by activating the AKT/GSK3ß/ß-catenin pathway.

Increasing evidence has shown that UBE2T plays an important role in genomic integrity and carcinogenesis; however, its role in nasopharyngeal carcinoma (NPC) has not been investigated. Here, we evaluated the clinicopathological significance of UBE2T in NPC and its underlying mechanisms. Using immunohistochemical analysis of UBE2T expression in NPC samples, we demonstrated that UBE2T is highly expressed in NPC tissues, which correlated with the T/M classification, skull invasion, and poor prognosis. The in vitro assay showed that UBE2T overexpression promoted proliferation, migration, and invasion of NPC cells, while UBE2T knockdown inhibited these processes. Consistent with our in vitro results, in vivo studies indicated that UBE2T overexpression promoted the growth of NPC xenografts and NPC cell metastasis. We found that UBE2T overexpression activated, whereas UBE2T knockdown inhibited, the AKT/GSK3ß/ß-catenin pathway. Moreover, the pathway-activation and in vitro pro-metastasis effects of UBE2T were blocked by the AKT inhibitor, MK-2206 2HCl. Additionally, UBE2T and p-GSK3 ß co-expressed in NPC samples by serial section, and their expressions are correlated. Collectively, our findings demonstrated that UBE2T is a possible diagnostic/prognostic biomarker for NPC and may promote the development and progression of NPC by activating the AKT/GSK3ß/ß-catenin pathway. Thus, UBE2T could serve as an alternative target for the treatment of NPC.

[miR-128a is up-regulated in hepatocellular carcinoma and promotes tumor cell proliferation by targeting RND3].

OBJECTIVE: To investigate the expression of microRNA-128a (miR-128a) and its role in hepatocellular carcinoma (HCC). METHODS: Nineteen pairs of fresh surgical specimens of HCC and adjacent tissues were examined for miR-128a expression using qRT-PCR. A miR-128a mimics or inhibitor was transfected into HCC cells, and the cell viability was analyzed by MTT assay. RND3, one of the potential targets of miR-128a, was predicted by bioinformatics software and demonstrated by dual luciferase reporter assay. The expression of RND3 after transfection was detected using qRT-PCR and Western blotting, and the cell cycle-related proteins were determined with Western blotting. RESULTS: The expression of miR-128a were significantly up-regulated in HCC tissues as compared to the adjacent tissues (P<0.05). In cultured HCC cells, miR-128a promoted the cell proliferation and resulted in down-regulated RND3 mRNA and protein expressions by targeting RND3' 3'UTR (P<0.05) and also in the down-regulation of cyclin B1, cyclin D1 and CDK4 protein expressions. CONCLUSION: miR-128a is up-regulated in HCC and promotes HCC cell proliferation by targeting RND3.

[Effect of casein kinase 2ß in esophageal carcinoma and its clinical significance].

OBJECTIVE: To investigate the expression of casein kinase 2ß in esophageal carcinoma tissues and analyze its clinical significance. METHODS: The expression of CK2ß in a tissue chip containing 60 normal esophageal mucosa and 60 colorectal cancer specimens were detected immunohistochemically. Ten human esophageal carcinoma and adjacent normal tissues were examined for the expression of CK2ß protein and mRNA using Western blotting and real-time quantitative PCR, respectively. RESULTS: A strong expression of CK2ß was found in 85.71% of the esophageal cancer tissues; 1.79% of the cancer tissues were negative for CK2ß expression, and 1.79% and 10.71% of the cancer tissues were weakly and moderately positive, respectively. In the normal mucosal tissues, 96.67% of the tissues were negative for CK2ß and 3.33% showed weak CK2ß expression, showing a significant difference in the distribution of CK2ß between normal and esophageal carcinoma tissues (P<0.001). The expression level of CK2ß in esophageal cancers was associated with the pathological stage (TNM) (P=0.010). Western blotting and real-time quantitative PCR also confirmed an increased CK2ß expression in the esophageal cancer tissues. CONCLUSION: The high expression of protein kinase CK2ß is closely related to the carcinogenesis and malignancy of esophageal cancer.

Increased expression of OCT4 is associated with low differentiation and tumor recurrence in human hepatocellular carcinoma.

Octamer binding transcription factor 4 (OCT4), a key transcription factor required to maintain self-renewal and pluripotency of human and mouse embryonic stem cells, has been recently identified to be associated with tumorigenesis and malignant transformation of many types of cancers. This study was to determine the roles of OCT4 in HCC recurrence and their impact on the clinical outcome of HCC patients. Western blot and immunohistochemical stains were used to detect the expression of OCT4 protein in 152 HCC tissues and 40 cirrhosis tissues, as well as in 6 human HCC cell lines and normal hepatocytes. OCT4 expression in HCC cell lines and tumor tissues was higher than in normal hepatocytes and cirrhosis tissues. Overexpression of OCT4 was significantly associated with low differentiation and tumor recurrence. Patients with elevated expression of OCT4 protein usually carried a poor overall survival and high recurrence rate. Multivariate analysis showed that OCT4 expression was an independent predictive factor for HCC patients survival. OCT4 might serve as a promising biomarker for the diagnosis of highly recurrent cases of HCC and could be used as a valuable indicator for predicting the prognosis of HCC.