Potential Target For Gene Therapy Research Articles

CSN6 Promotes the Migration and Invasion of Cervical Cancer Cells by Inhibiting Autophagic Degradation of Cathepsin L.

CSN6 is one subunit of the highly conserved constitutive photomorphogenesis 9 (COP9) signalosome (CSN), which is overexpressed in many types of cancers, and has received great attention as a regulator of the degradation of cancer-related proteins, suggesting its importance in oncogenic activity. CSN6 has been shown to be overexpressed in cervical cancer (CC) and associated with CC development. CC remains to be one of the most aggressive cancers affecting women. Cathepsin L (CTSL), significantly associated with the autophagy, plays a critical role in degradation of extracellular matrix for metastasis. However, the detailed biological functions of CSN6 on CTSL in CC metastasis have not been well clarified. Our data has shown that CSN6 and CTSL are positively correlated. The overexpression of CSN6 and CTSL might be a strong indicator for CC enhanced aggressiveness. CSN6 could suppress the degradation of CTSL, then facilitated the migration and invasion of CC cells. Interestingly, our results indicated that autophagy is essential for decreasing CTSL, while CSN6 could inhibit the autophagy ability of CC cells. In addition, blocking of the mammalian target of rapamycin (mTOR) pathway reversed CSN6-mediated autophagy inhibition. We further demonstrated that CSN6 positively regulated CTSL expression through an autophagy-lysosomal system. Taken together, we concluded that CSN6 might promote the migration and invasion of cervical cancer cells by inhibiting autophagic degradation of CTSL and serve as a potential gene therapy target for the treatment of CC metastasis.

Histone demethylase KDM3B regulates the transcriptional network of cell-cycle genes in hepatocarcinoma HepG2 cells

Hepatocellular carcinoma (HCC) is the fifth most common cancer and the third most lethal cancer worldwide. Although gene mutations associated with HCC development have been intensively studied, how epigenetic factors specifically modulate the functional properties of HCC by regulating target gene expression is unclear. Here we demonstrated the overexpression of KDM3B in liver tissue of HCC patients using public RNA-seq data. Ablation of KDM3B by CRISPR/Cas9 retarded the cell cycle and proliferation of hepatocarcinoma HepG2 cells. Approximately 30% of KDM3B knockout cells exhibited mitotic spindle multipolarity as a chromosome instability (CIN) phenotype. RNA-seq analysis of KDM3B knockout revealed significantly down-regulated expression of cell cycle related genes, especially cell proliferation factor CDC123. Furthermore, the expression level of Cyclin D1 was reduced in KDM3B knockout by proteosomal degradation without any change in the expression of CCND1, which encodes Cyclin D1. The results implicate KDM3B as a crucial epigenetic factor in cell cycle regulation that manipulates chromatin dynamics and transcription in HCC, and identifies a potential gene therapy target for effective treatment of HCC.

Otx2 enhances transdifferentiation of Müller cells-derived retinal stem cells into photoreceptor-like cells.

Retinal Müller glial cells have the potential of neurogenic retinal progenitor cells, and could reprogram into retinal‐specific cell types such as photoreceptor cells. How to promote the differentiation of Müller cells into photoreceptor cells represents a promising therapy strategy for retinal degeneration diseases. This study aimed to enhance the transdifferentiation of rat Müller cells‐derived retinal stem cells (MC‐RSCs) into photoreceptor‐like cells and explore the signalling mechanism. We dedifferentiated rat Müller cells into MC‐RSCs which were infected with Otx2 overexpression lentivirus or control. The positive rate of photoreceptor‐like cells among MC‐RSCs treated with Otx2 overexpression lentivirus was significantly higher compared to control. Furthermore, pre‐treatment with Crx siRNA, Nrl siRNA, or GSK‐3 inhibitor SB‐216763 reduced the positive rate of photoreceptor‐like cells among MC‐RSCs treated with Otx2 overexpression lentivirus. Finally, Otx2 induced photoreceptor precursor cells were injected into subretinal space of N‐methyl‐N‐nitrosourea induced rat model of retinal degeneration and partially recovered retinal degeneration in the rats. In conclusion, Otx2 enhances transdifferentiation of MC‐RSCs into photoreceptor‐like cells and this is associated with the inhibition of Wnt signalling. Otx2 is a potential target for gene therapy of retinal degenerative diseases.

Inhibiting GIT1 reduces the growth, invasion, and angiogenesis of osteosarcoma.

BackgroundGIT1, a scaffold protein with ubiquitous multi-domain, is involved in many cellular processes. In recent years, it was proved that GIT1 participated in various tumors’ growth or metastasis. However, the biological function of GIT1 in osteosarcoma is still unclear. In this study, we aimed to investigate the role and mechanism of GIT1 in osteosarcoma.Materials and methodsHuman osteosarcoma tissues were obtained to investigate the distribution of GIT1. Adequate osteosarcoma cells were stably infected with lentivirus to knockdown GIT1 level and then was used to carry out cell invasion and vascular endothelial growth factor (VEGF) assay in vitro. Orthotopic femoral osteosarcoma model was constructed to investigate the growth, invasion, and angiogenesis in vivo. Western blot was used to detect extracellular signal-regulated kinase (ERK1/2) activation and hypoxia-inducible factor-1 (HIF-1α) expression.ResultsIn this study, we found that GIT1 was distributed in human osteosarcoma tissues and highly expressed in osteosarcoma (OS) cells. Knockdown of GIT1 inhibited cell invasion and VEGF release in vitro and suppressed tumor growth, invasion, and angiogenesis in vivo. Furthermore, knockdown of GIT1 substantially downregulated the protein levels of p-ERK and HIF-1α in OST cells and inhibition of p-ERK by PD98059 could significantly decrease the expression of HIF-1α and concentration of VEGF in GIT1-shRNA-treated cells.ConclusionGIT1 knockdown can effectively inhibit the growth, invasion, and angiogenesis of osteosarcoma. Thus, GIT1 might act as an oncogenic factor in osteosarcoma and could be a potential molecular target for osteosarcoma gene therapy.

The regulatory role of Annexin 3 in a nude mouse bearing a subcutaneous xenograft of MDA-MB-231 human breast carcinoma

The following study investigated the effects of Annexin A3 (ANXA3) on breast cancer biological behavior in vivo, using nude mouse model bearing a subcutaneous tumor. A total of 18 female nude mice were randomly divided into three groups (n = 6): negative control group which was inoculated with MDA-MB-231 cells, blank control group which was inoculated with MDA-MB-231-NC cells, and the transfection group which was inoculated with MDA-MB-231-Sh cells. The experiment lasted for 4 weeks, during which mice conditions, diet and defecation were monitored on a daily basis. Body weight, as well as tumor diameters, which were assessed using standard caliper method, were measured once a week. In vivo imaging was performed to detect the activity of transplanted tumors. H&E staining was used to analyze the histological structure of tumor tissues in three groups, while flow cytometry and fluorescent RT-PCR were performed to measure cell proliferation and the expression of ANXA3 mRNA. Briefly, significantly slower tumor growth and tumor activity were observed in the transfection group compared to negative and blank controls, while the tumor weight and volume in this group were also significantly lower compared to the other two groups (P < 0.01). Sparse tumor cells accompanied with massive fibrous connective tissue proliferation, and lower new blood vessels formation were observed in transfection group compared to other groups. Moreover, mRNA and protein levels of ANXA3 were significantly lower in transfection group compared to the other two groups (P < 0.01). In addition, lower proliferation index and higher G0/1 cell count were observed in transfection group compared to negative and blank controls (P < 0.01). To sum up, these results suggested that ANXA3 silencing regulates the proliferation and inhibits the growth of MDA-MB-231 breast cancer cells. Consequently, ANXA3 might be used as a potential target for gene therapy in breast cancer.

The emerging role of miR-19 in glioma.

Glioma has been regarded as the most common, highly proliferative and invasive brain tumour. Advances in research of miRNAs in glioma are toward further understanding of the pathogenesis of glioma. MiR‐19, a member of miR‐17~92 cluster, was reported to play an oncogenic role in tumourigenesis. Here we review the identified data about the effect of miR‐19 on proliferation, apoptosis, migration and invasion of glioma cells, the target genes regulated by miR‐19, and correlation of miR‐19 with the sensitivity of glioma cells to chemotherapy and radiotherapy. It is concluded that miR‐19 plays an important role in the pathogenesis of glioma and can be a potential target for gene therapy of glioma.

Isolation of T cell receptors targeting recurrent neoantigens in hematological malignancies

Mutation-derived neoantigens represent an important class of tumour-specific, tumour rejection antigens, and are attractive targets for TCR gene therapy of cancer. The majority of such mutations are patient-specific and targeting these requires a fully personalized approach. However, some mutations are found recurrently among cancer patients, and represent potential targets for neoantigen-specific TCR gene therapy that is more widely applicable. Therefore, we have investigated if some cancer mutations found recurrently in hematological malignancies encode immunogenic neoantigens presented by common European Caucasoid HLA class I alleles and can form targets for TCR gene therapy. We initially focused on identifying HLA class I neoepitopes derived from calreticulin (CALR) exon 9 mutations, found in ~ 80% of JAK2wt myeloproliferative neoplasms (MPN). Based on MHC class I peptide predictions, a number of peptides derived from mutant CALR (mCALR) were predicted to bind to HLA-A*03:01 and HLA-B*07:02. However, using mass spectrometry and ex vivo pMHC multimer staining of PBMC from MPN patients with CALR exon 9 mutations, we found no evidence that these peptides were naturally processed and presented on the surface of mCALR-expressing target cells. We next developed a protocol utilizing pMHC multimers to isolate CD8+ T cells from healthy human donor PBMC that are specific for mCALR and additional putative neoepitopes found recurrently in hematological malignancies. Using this approach, CD8+ T cells specific for HLA-A*03:01- and HLA-B*07:02-presented mCALR peptides and an HLA-A*11:01-presented mutant FBXW7 (mFBXW7) peptide were successfully isolated. TCRs isolated from mCALR-specific CD8+ T cell populations were not able to recognize target cells engineered to express mCALR. In contrast, mFBXW7-specific CD8+ T cells were able to recognize target cells engineered to express mFBXW7. In conclusion, while we found no evidence for mCALR derived neoepitope presentation in the context of the HLA class I alleles studied, our data suggests that the recurrent pR465H mutation in FBXW7 may encode an HLA-A*11:01 presented neoepitope, and warrants further investigation as a target for T cell based immunotherapy of cancer.

Correlation between S100A11 and the TGF-β1/SMAD4 pathway and its effects on the proliferation and apoptosis of pancreatic cancer cell line PANC-1.

S100A11 as a S100 protein family member has been documented to play dual-direction regulation over cancer cell proliferation. We explored the role of S100A11 in the proliferation and apoptosis of pancreatic cancer cell line PANC-1 and the potential mechanisms involving the TGF-β1/SMAD4/p21 pathway. S100A11 and TGF-β1 protein expressions in 30 paraffin-embedded specimens were evaluated by immunohistochemistry. S100A11 and TGF-β1 expression in PANC-1 cell line was suppressed using small interfering RNA (siRNA), respectively. Subsequently, pancreatic cancer cell apoptosis was measured by Cell Counting Kit-8 and flow cytometry, and S100A11 and TGF-β1/SMAD4/p21 pathway proteins and genes were detected with Western blotting and quantitative polymerase chain reaction (qPCR). S100A11 cytoplasmic/nuclear protein translocation was examined using NE-PER® cytoplasm/nuclear protein extraction in cells interfered with TGF-β1 siRNA. Our results showed that S100A11 expression was positively correlated with TGF-β1 expression in pancreatic cancerous tissue. Silencing TGF-β1 down-regulated intracellular P21WAF1 expression by 90%, blocked S100A11 from cytoplasm entering nucleus, and enhanced cell proliferation. Silencing S100A11 down-regulated intracellular P21 expression and promoted cell apoptosis without significantly changing TGF-β1 and SMAD4 expression. Our findings revealed that S100A11 and TGF-β1/SMAD4 signaling pathway were related but mutually independent in regulating PANC-1 cells proliferation and apoptosis. Other independent mechanisms might be involved in S100A11's regulation of pancreatic cell growth. S100A11 could be a potential gene therapy target for pancreatic cancer.

Targeting cysteine-rich angiogenic inducer-61 by antibody immunotherapy suppresses growth and migration of non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is the most frequent type of human lung cancer; lung cancer is responsible for the highest rates of cancer-associated mortality in the world. Cysteine-rich angiogenic inducer-61 (CYR-61) has been identified as a tumorigenesis-, development- and metastasis-related gene, and is reported to enhance proliferation, migration and invasion through hepatocyte growth factor (HGF)-induced scattering and the metastasis-inducing HGF/Met signaling pathway in tumor cells and xenograft models. CYR-61 is a protein that promotes human lung cancer cell metastasis and is closely related to the patient's prognosis in NSCLC. The purpose of the present study was to investigate whether CYR-61 may serve as a dual potential target for gene therapy of human NSCLC. In the present study, an antibody targeted against CYR-61 (anti-CYR-61) was constructed and the therapeutic effects and underlying mechanism of this antibody in NSCLC cells and mice with NSCLC was investigated. It was observed that NSCLC cell viability, migration and invasion were inhibited while cell apoptosis was induced by the neutralization of CYR-61 protein by anti-CYR-61. Western blotting demonstrated that extracellular signal-regulated kinase (ERK) and protein kinase B (AKT) expression levels in NSCLC cells were decreased following treatment with anti-CYR-61. In addition, it was observed that inhibition of NSCLC cell viability was achieved by the suppression of the epithelial-mesenchymal transition signaling pathway. ERK and AKT phosphorylation levels were downregulated in NSCLC cells and tumors following anti-CYR-61 treatment. Analysis of a murine model indicated that tumor growth was inhibited and tumor metastasis was significantly suppressed (P<0.01) following anti-CYR-61 treatment for CYR-61. In conclusion, CYR-61 may serve as a potential target for gene therapy for the treatment of human NSCLC.

The effect of downregulation of Stathmin gene on biological behaviors of U373 and U87-MG glioblastoma cells

BackgroundStathmin as a critical protein involved in microtubule polymerization, is necessary for survival of cancer cells. However, extremely little is known about Stathmin in glioblastoma. So, this study was designed to elucidate the function of Stathmin gene in the tumorigenesis and progression of glioblastoma cells.MethodThe lentiviral interference vector pLV3-si-Stathmin targeting Stathmin gene and the control vector pLV3-NC were established for the co-transfection of 293T cells together with the helper plasmids. Viral titer was determined via limiting dilution assay. Then pLV3-si-Stathmin and pLV3-NC were stably co-transfected into U373 and U87-MG glioblastoma cells. Expression levels of Stathmin protein in each group were determined by using Western Blot, and the proliferation and migration ability of the cells with downregulated Stathmin were evaluated through CCK8 assay and transwell invasion assay, respectively. Cell cycles and cell apoptosis were detected with flow cytometry. Finally, the effect of Stathmin in tumor formation was determined in nude mice.ResultDNA sequencing and viral titer assay indicated that the lentiviral interference vector was successfully established with a viral titer of 4 × 108 TU/ml. According to the results from Western Blotting, Stathmin protein expression level decreased significantly in the U373 and U87-MG cells after transfected with pLV3-si-Stathmin, respectively, compared with those transfected with pLV3-NC. In glioblastoma cells, the cell proliferation and migration were greatly inhibited after the downregulation of Stathmin protein. Flow cytometry showed that much more cells were arrested in G2/M phasein Stathmin downregulated group, compared with the non-transfection group and NC group. But Stathmin downregulation did not induce significant cell apoptosis. Tumor formation assay in nude mice showed that tumor formation was delayed after Stathmin downregulation, with a reduction in both tumor formation rate and tumor growth velocity.ConclusionStathmin downregulation affected the biological behaviors of U373 and U87-MG glioblastoma cells, inhibiting the proliferation and migration of tumor cells. Stathmin gene may serve as a potential target in gene therapy for glioblastoma.

Role of taurine upregulated gene 1 as a predictor of poor outcome in osteosarcoma.

Previous studies have found that long noncoding RNA taurine upregulated gene 1 (TUG1) can regulate osteosarcoma cells apoptosis and proliferation; the aim of this study was to investigate the clinical significance of TUG1 in osteosarcoma. The expression of TUG1 was detected by real-time and quantitative polymerase chain reaction assay in 94 pairs of tumor tissues and corresponding noncancerous bone tissues of osteosarcoma patients. Its correlations with clinicopathologic features were analyzed, and the significance of TUG1 as a prognostic factor was determined. This study shows that the expression of TUG1 in osteosarcoma tissues was significantly higher than that in adjacent normal bone tissues. Upregulation of TUG1 was significantly correlated with the larger tumor size and advanced tumor-node-metastases stage of osteosarcoma patients. Kaplan-Meier curve showed a decreased overall survival time of osteosarcoma patients with high TUG1 expression. Moreover, univariate and multivariate analyses suggested that low-expression level of TUG1 was an independent poor prognostic indicator for osteosarcoma patients. In conclusion, our data support TUG1 as a potential prognostic predictor and gene therapy target with its high expression in tumor tissues and its association with carcinogenesis and progression in osteosarcoma.

Adeno-associated viral vector-mediated ATPase gene transfer alleviates asthmatic airway inflammation in OVA-sensitized mice model

Abstract Bronchial asthma is a chronic inflammatory disorder of the airway characterized by airway hyperresponsiveness (AHR), pulmonary eosinophilia, elevated serum IgE, and dysregulated T helper (Th) 2 cytokines. It is becoming urgent to identify target molecules for the preventive and therapeutic purpose. ATP accumulation in bronchoalveolar lavage fluid (BALF) of asthmatic subjects indicates that extracellular ATP and the downstream responses are involved. ATP is tightly regulated by CD39, a nucleoside triphosphate diphosphohydrolase (ATPase) encoded by entpd1. Upregulated BALF ATP levels and decreased CD39 RNA were accompanied with asthmatic cardinal features in ovalbumin (OVA)-sensitized mice. Recombinant adeno-associated virus (rAAV) has been developed to carry exogenous genes with safe and low toxic therapeutic potential. The different serotypes and tropisms of AAVs benefit tissue specific gene targeting. This study aims to determine whether lung-targeting rAAV carrying CD39 (AAV/CD39) is able to reduce extracellular ATP in BALF and inhibit the OVA-induced allergic inflammation in an asthmatic murine model. rAAV vectors were administrated into the lungs of mice 3 days before the first OVA exposure. The data indicate that ATP levels in BALF were downregulated accompanied with rescued CD39 expression. The administration of AAV/CD39 was able to suppress AHR, pulmonary eosinophilia, and the production of IL-5 and IL-13 from OVA-activated mediastinal lymph nodes (MLNs) cells. IL-33 in BALF was also reduced. These data indicate that the attenuation of asthmatic airway inflammation may affect innate lymphoid cells (ILC) 2 activation. Thus, ATP level could be a potential target for asthma gene therapy.

Downregulation of TACC3 inhibits tumor growth and migration in osteosarcoma cells through regulation of the NF-κB signaling pathway.

TACC3, a member of the transforming acidic coiled-coil protein (TACC) family, is a multifunctional protein that is involved in various biological functions, including proliferation and differentiation of tumor cells, cancer progression and metastasis. The aims of the present study were to examine whether TACC3 expression is associated with the proliferation and migration of osteosarcoma (OS) cells and to investigate the potential underlying molecular mechanisms of TACC3 in OS. First, the levels of mRNA and protein expression in OS cell lines by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively were examined. Second, the effects of TACC3 knockdown and overexpression on the proliferative, migratory and invasive capacities of OS cells were investigated. Finally, western blot analysis was employed to detect the potential mechanism of TACC3 in osteosarcoma. TACC3 expression was significantly increased in osteosarcoma tissues and cell lines, compared to matched controls. The knockdown of TACC3 was able to significantly inhibit the proliferation, migration and invasion of osteosarcoma cells, whereas the overexpression of TACC3 was able to promote cell proliferation and migration. Mechanistically, TACC3 may promote the migration and invasion of osteosarcoma cells via through nuclear factor-κB signaling. These data suggest that TACC3 has an important part in the progression of osteosarcoma and may serve as a potential target for gene therapy.

The ubiquitin ligase COP1 regulates cell cycle and apoptosis by affecting p53 function in human breast cancer cell lines.

The E3 ubiquitin ligase constitutive photomorphogenic 1 (COP1) mediates cell survival, growth, and development, and interacts with the tumor suppressor protein p53 to induce its ubiquitination and degradation. Recent studies reported that COP1 overexpression is associated with increased cell proliferation, transformation, and disease progression in a variety of cancer types. In this study, we investigated whether COP1 regulates p53-mediated cell cycle arrest and apoptosis in human breast cancer cell lines. We downregulated COP1 expression using lentiviral particles expressing short hairpin RNA (shRNA) targeting COP1 and measured the effects of the knockdown in three different breast cancer cell lines. COP1 silencing resulted in p53 activation, which induced the expression of p21 and p53-upregulated modulator of apoptosis (PUMA) expression, and reduced the levels of cyclin-dependent kinase 2 (CDK2). Notably, knockdown of COP1 was associated with cell cycle arrest during the G0/G1 phase. The COP1-mediated degradation of p53 regulates cancer cell growth and apoptosis. Our results indicate that COP1 regulates human breast cancer cell proliferation and apoptosis in a p53-dependent manner. These findings suggest that COP1 might be a promising potential target for breast cancer-related gene therapy.

Down-regulated REIC expression in lung carcinogenesis: a molecular target for gene therapy.

REIC (Reduced Expression in Immortalized Cells) gene is down-regulated in immortalized cells, compared with the normal parental counterparts. Its encoding protein could inhibit colony formation, tumor growth, and induce apoptosis. To investigate the roles of REIC expression in lung cancer, we examined REIC expression in lung cancer cells and tissues by RT-PCR or Western blot, and observed the effects of both recombinant REIC exposure and REIC overexpression on the aggressive phenotypes of lung cancer cells. It was found that the demethylation of REIC promoter by 5-Aza-dC could reserve its mRNA expression in lung cancer cells (P<0.05). There was a lower REIC mRNA expression in lung cancer than that in matched normal tissue (P<0.05). Recombinant REIC treatment enhanced the proliferation of lung cancer cells (P<0.05), but versa for REIC overexpression (P<0.05). Both recombinant REIC treatment and REIC overexpression induced apoptosis, and inhibited the migration and invasion of SQ-5 and KJ cells (P<0.05). Immunohistochemically, there was a positive correlation between REIC and Caspase-3 expression in lung cancer (P<0.05). According to Kaplan-Meier plotter, REIC mRNA overexpression was found to positively correlate with overall, progression-free and post- progression survival rates of lung cancer patients (P<0.05), even stratified by sex, histological subtyping, grading, TNM staging, chemotherapy, radiotherapy, or smoking. These findings suggested that the down-regulated REIC expression might be involved in lung carcinogenesis due to its promoter methylation. Both recombinant REIC exposure and REIC overexpression might reverse the aggressive phenotypes of lung cancer cells. REIC may be employed as a potential target of gene therapy for lung cancer.

Anti-apoptosis Effect of Decoy Receptor 3 in Cholangiocarcinoma Cell Line TFK-1.

Background:Decoy receptor 3 (DcR3) is a protein with anti-apoptotic effect that belongs to the tumor necrosis factor receptor superfamily. DcR3 is highly expressed in a variety of malignant tumors including cholangiocarcinoma and its expression was found to be related to the clinical stage, the invasion, and the metastasis of the tumor. This in vitro study aimed to investigate the effect of downregulated expression of DcR3 on cell viability, cell apoptosis, and cell cycle in cholangiocarcinoma cell line TFK-1.Methods:Three different cell lines were cultured: human cholangiocarcinoma TFK-1, human biliary epithelial carcinoma HuCCT-1, and human cholangiocarcinoma RBE. The cholangiocarcinoma cell line with the highest expression of DcR3 was selected for further investigation. The expression of DcR3 was silenced/knocked down by transfection with DcR3-siRNA in the selected cell line. Various biological phenotype parameters such as cell viability, apoptosis, and cell cycle were observed.Results:The mRNA and protein levels of DcR3 were measured in the three cell lines, and TFK-1 was selected. After the treatment with DcR3-siRNA for 48 h, DcR3 mRNA and protein expression in the treatment group were 38.45% (P < 0.01) and 48.03% (P < 0.05) of that of the control, respectively. It was found that the cell viability decreased to 61.87% of the control group (P < 0.01) after the downregulation of DcR3 in cholangiocarcinoma cell line TFK-1 by transfection with DcR3-siRNA, while the percentage of apoptotic cells was 2.98 times as compared with the control group (P < 0.05). Compared with the control group the ratio of G0/G1 increased, and the ratio of G2/M decreased in the treatment group. However, the differences were not statistically significant.Conclusions:The effect of DcR3 on the growth and apoptosis of cholangiocarcinoma has been demonstrated. DcR3 is not only a predictive marker for malignant tumor but it is also likely to be a potential target for cancer gene therapy. Further studies should focus on exploring the binding ligand of DcR3, the signaling pathway involved, and the molecular mechanism for the regulation of DcR3 expression in cholangiocarcinoma.

Identification of Potential Biomarkers Involved in Gastric Cancer Through Integrated Analysis of Non-Coding RNA Associated Competing Endogenous RNAs Network.

Gastric cancer is one of the most common malignant tumors worldwide. Increasing studies have indicated that non-coding RNAs (ncRNAs) play critical roles in cancer progression. They have shown great potential to be useful markers and therapeutic targets. Bioinformatics analysis was conducted to detect mRNA and ncRNAs' expression changes between tumor samples and adjacent non-tumor samples. The gene expression profiles were obtained from the national center of biotechnology information gene expression omnibus (GEO). The differentially expressed genes (DEGs) were identified through fold change filtering. The interaction between lncRNAs and miRNAs were predicted by DIANA-LncBase, and the interaction between mRNAs and miRNAs were predicted by miRTarBase. Gene ontology (GO) enrichment analysis and pathway analysis were performed using standard enrichment computational methods. An LncRNA-miRNA-mRNA regulation network was constructed based on the gene expression profiles to identify hub genes. A protein-protein interaction (PPI) network was constructed based on STRING database. Survival analysis sourced from The Cancer Genome Atlas (TCGA) data was performed and the log-rank test was conducted to confirm the relationship between gene expression and risk of gastric cancer. With a threshold of p-value < 0.05 and absolute value of fold change (FC) > 2, differentially expressed genes including 70 miRNAs, 3266 mRNAs (4206 probe IDs) and 174 lncRNAs (188 probe IDs) were screened. After the results of predicted interactions and DEGs were intersected, 114 mRNAs, 56 miRNAs, and 68 lncRNAs were selected. GO enrichment analysis and pathway analysis were performed for the 114 mRNAs. A PPI network including 61 nodes and 91 edges was constructed for the selected mRNAs and ncRNAs. Survival analysis was performed for the weighted genes in the network and showed that KRAS, TRAF7, SUCLG2-AS1, IGF1R, UBE2B, AQP4-AS1, LINC00284, LINC01122, and RGMB were closely related with the overall survival coupled with highrisk, and LUM, UBE2D1, HNRNPU, and TOP2A were closely related with the overall survival coupled with lowrisk in gastric cancer. Our study indicated that KRAS, TRAF7, SUCLG2-AS1, IGF1R, UBE2B, AQP4-AS1, LINC00284, LINC01122, RGMB, LUM, UBE2D1, HNRNPU, and TOP2A might be potential targets for gene therapy for human gastric cancer.

Up-regulated expression of miR-155 in human colonic cancer.

The aim of this study is to investigate the expression of miR-155 in colonic cancer tissue and to assess the potential predictive value of miR-155 in colonic cancer patients. From March to September of 2011, we included 57 patients with primary colonic cancer who underwent curative surgical resection. Total RNAs were extracted from colonic cancer tissues and adjacent normal tissues. Then the expression of miR-155 in colonic cancer and paracancerous tissues was investigated using real time quantitative reverse transcription-polymerase chain reaction. And the relationship between miR-155 expression level and the clinical, pathological parameters of colonic cancer was analyzed. The relative expression level of miR-155 in colonic cancer tissues was significantly higher than that in paracancerous tissues and related to tumor-node-metastasis staging, tumor invasion, metastasis, and differentiation. The expression of miR-155 is up-regulated in colonic cancer tissues. MiR-155 may acts as proto-oncogenes involved in carcinogenesis, development, and invasion of colon cancer making it a potential target for gene therapy of colon cancer.

Neuropilin-1 promotes primary liver cancer progression by potentiating the activity of hepatic stellate cells.

As a co-receptor for a variety of cytokines, neuropilin-1 (NRP-1) is detectable in primary liver cancer (PLC) cells. Previous studies determined that silencing of NRP-1 expression attenuated the proliferation, migration and invasion of PLC cells. An increasing number of studies have highlighted the crucial role of the tumor microenvironment in the pathogenesis of cancer. Hepatic stellate cells (HSCs) are one of the major interstitial cell types present in the liver tumor microenvironment, and can promote the proliferation, migration and invasion of PLC cells. It remains unknown whether NRP-1 can promote PLC progression by potentiating the activity of HSCs. In the present study, the expression of NRP-1, and its co-expression with platelet-derived growth factor receptor-β, in HSCs was detected via immunofluorescence. LX2 HSCs were transfected with NRP-1 short hairpin RNA lentiviral vectors and their proliferation was observed. The proliferation, migration and invasion of HepG2 cells co-cultured with LX2 cells were also observed. Finally, LX2 and HepG2 cells were co-injected into nude mice as subcutaneous xenografts, and the tumor growth and α-smooth muscle actin expression levels were observed. NRP-1 knockdown attenuated LX2 cell activation, with concomitant downregulation of HepG2 cell proliferation, migration and invasion (P<0.05). Thus, silencing of NRP-1 expression may inhibit the activation of HSCs, as well as the proliferation, migration and invasion of PLC cells. The mechanism underlying the inhibition of PLC cell progression is possibly mediated by the inhibition of HSC activation, reduction of transforming growth factor-β1 levels in the conditioned medium and downregulation of extracellular signal-related kinase activity in PLC cells. Thus, NRP-1 could be regarded as a potential gene therapy target for PLC.

BTBD7 silencing inhibited epithelial- mesenchymal transition (EMT) via regulating Slug expression in human salivary adenoid cystic carcinoma.

Epithelial-mesenchymal transition (EMT) is a complicated process that has been implicated in cancer progression and metastasis as well as the formation of many tissues and organs. BTB/POZ domain-containing protein 7 (BTBD7) is reported to regulate transcriptional factors and involved in the process of invasion and metastasis of some malignant tumors. Additionally, our preliminary studies have confirmed that BTBD7 expression was significantly correlated with Slug expression and poor prognosis of primary salivary adenoid cystic carcinoma (SACC). On this basis, this study further investigated function of BTBD7 in the invasion and metastasis of SACC in vitro, which may be a possible target of gene therapy in the future. The expression of BTBD7 and Slug were both examined in SACC-LM and SACC-83 cell lines by immunofluorescence staining. High invasive SACC-LM cells were transfected with BTBD7 siRNA and the expression levels of BTBD7 and Slug were detected in both gene and protein levels by qRT-PCR and western blot analysis. Assays were performed to survey cell migration, invasion and proliferation capabilities with BTBD7 silencing. BTBD7 and Slug proteins were detected in SACC-LM and SACC-83 cell lines. BTBD7 silencing down-regulated the expression of Slug and MMP9 meanwhile up-regulated the expression of E-cadherin in SACC-LM cells, the migration and invasion abilities of cells were obviously suppressed but with no influence on cell proliferation. BTBD7 silencing inhibited EMT through regulation of Slug expression in SACC-LM cells and might act as a potential molecular target for gene therapy of SACC.