Cell Function Experiments Research Articles

The microRNA-381(miR-381)/Spindlin1(SPIN1) axis contributes to cell proliferation and invasion of colorectal cancer cells by regulating the Wnt/β-catenin pathway

ABSTRACT Our study aimed to investigate the clinical significance and biological functions of Spindlin1 (SPIN1) in colorectal cancer (CRC) tumorigenesis and progression, as well as the mechanism underlying its upregulation. The expression of SPIN1 was detected by immunohistochemistry and western blotting assays. Bioinformatics prediction and dual-luciferase reporter assays were used to determine whether microRNA-381 (miR-381) could target SPIN1. A series of cell functional experiments were performed to investigate whether the miR-381-mediated regulation of SPIN1 is involved in the progression and aggressiveness of CRC cells via the Wnt/β-catenin pathway. Our results showed that SPIN1 is frequently overexpressed in CRC tissues and cell lines, and its upregulation is positively correlated with disease progression and lymph node metastasis. Moreover, SPIN1 depletion suppresses cell growth, migration, and invasion through inactivation of the Wnt/β-catenin signaling pathway, which recapitulates the effects of miR-381 upregulation. Moreover, SPIN1 is a target gene of miR-381, and miR-381 is downregulated in CRC. Furthermore, the reintroduction of SPIN1 partially abolished the miR-381-mediated inhibitory effects in CRC cells. In summary, our data revealed that the miR-381/SPIN1 axis greatly contributes to CRC tumorigenesis by orchestrating the Wnt/β-catenin pathway, thereby representing actionable therapeutic targets for colorectal cancer patients.

A Novel Four-Gene Signature as a Potential Prognostic Biomarker for Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a common malignant tumor with high incidence and mortality rates. However, a reliable prognostic signature has not yet been confirmed. Thus, the purpose of the present study was to develop a biomarker with high specificity and sensitivity for the diagnosis and prognosis of patients with HCC. The mRNA expression profiles of HCC were obtained from the GSE19665, GSE41804, and TCGA databases. Subsequently, 193 differentially expressed genes (DEGs) were identified from the intersection of the data from the three datasets. Bioinformatics analysis showed that the identified DEGs are related to the cell cycle, oocyte meiosis, and p53 signaling pathway, among other factors, in cancers. A protein-protein interaction (PPI) and a functional analysis were performed to investigate the biological function of the DEGs and obtain the candidate genes using the MCODE of Cytoscape. The candidate genes were introduced into the TCGA database for survival analysis, and the four candidate genes that were hub genes and meaningful for survival were retained for further verification. We validated the gene and protein expression and determined the prognosis of our patient cohort. In addition, we evaluated the biological functions regulating tumor cell proliferation and metastasis in vitro. According to the ROC curve analysis of gene expression in clinical samples, it was found that the four genes can be used to predict the diagnosis. A survival analysis based on data from the TCGA database and clinical samples showed that the four genes may be used as biomarkers for providing prognoses for patients. The cell functional experiments revealed that these four genes were related to tumor proliferation, migration, and invasion. In conclusion, the genes identified in the present study could be used as markers to diagnose and predict the prognosis of patients with HCC and guide targeted therapy.

LncRNA FOXP4-AS1 Promotes the Progression of Esophageal Squamous Cell Carcinoma by Interacting With MLL2/H3K4me3 to Upregulate FOXP4.

Malignant tumors are a grave threat to human health. Esophageal squamous cell carcinoma (ESCC) is a common gastrointestinal malignant tumor. China has a high incidence of ESCC, and its morbidity and mortality are higher than the global average. Increasingly, studies have shown that long noncoding RNAs (lncRNAs) play a vital function in the occurrence and development of tumors. Although the biological function of FOXP4-AS1 has been demonstrated in various tumors, the potential molecular mechanism of FOXP4-AS1 in ESCC is still poorly understood. The expression of FOXP4 and FOXP4-AS1 was detected in ESCC by quantitative real-time PCR (qRT–PCR) or SP immunohistochemistry (IHC). shRNA was used to silence gene expression. Apoptosis, cell cycle, MTS, colony formation, invasion and migration assays were employed to explore the biological functions of FOXP4 and FOXP4-AS1. The potential molecular mechanism of FOXP4-AS1 in ESCC was determined by dual-luciferase reporter, RNA immunoprecipitation (RIP) and chromatin immunoprecipitation (ChIP). Here, we demonstrated that FOXP4-AS1 was significantly increased in ESCC tissues and cell lines, associated with lymph node metastasis and TNM staging. Cell function experiments showed that FOXP4-AS1 promoted the proliferation, invasion and migration ability of ESCC cells. The expression of FOXP4-AS1 and FOXP4 in ESCC tissues was positively correlated. Further research found that FOXP4-AS1, upregulated in ESCC, promotes FOXP4 expression by enriching MLL2 and H3K4me3 in the FOXP4 promoter through a “molecular scaffold”. Moreover, FOXP4, a transcription factor of β-catenin, promotes the transcription of β-catenin and ultimately leads to the malignant progression of ESCC. Finally, FOXP4-AS1 may be a new therapeutic target for ESCC.

CircRNA_0044556 diminishes the sensitivity of triple‑negative breast cancer cells to adriamycin by sponging miR‑145 and regulating NRAS.

CircRNAs are associated with adriamycin (ADM) resistance in triple-negative breast cancer (TNBC), but the mechanism is unknown. Reverse transcription-quantitative PCR was applied to quantify circular RNA (circRNA)_0044556, microRNA (miR)-145 and NRAS proto-oncogene, GTPase (NRAS) in TNBC tissues and cells with or without ADM treatment. Following ADM treatment, the effects of circRNA_0044556 on the viability, ADM resistance, apoptosis and migration of TNBC cells were investigated by cell function experiments (Cell Counting Kit-8, flow cytometry and Transwell assays). The targeting relationship between circRNA_0044556 and miR-145 was investigated via bioinformatics analysis, dual-luciferase reporter assay and RNA immunoprecipitation. The effects of the circRNA_0044556/miR-145 axis on the TNBC cells were revealed by rescue experiments. Correlations among circRNA_0044556, miR-145 and NRAS were analyzed by Pearson's correlation test. CircRNA_0044556 was highly expressed in TNBC tissues and cells with or without ADM-resistance. The overexpression of circRNA_0044556 promoted cell viability, ADM-resistance and migration, while inhibiting the apoptosis by sponging miR-145. Upregulation of miR-145 reversed the effects of circRNA_0044556 on the TNBC cells. CircRNA_0044556 was negatively correlated with miR-145 yet positively correlated with NRAS, the target gene of miR-145, in addition to the discovery suggesting the negative regulatory effects of circRNA_0044556 on miR-145. CircRNA_0044556 diminished the sensitivity of TNBC cells to ADM via the miR-145/NRAS axis.

Long non-coding RNA TRPM2-AS regulates microRNA miR-138-5p and PLAU (Plasminogen Activator, Urokinase) to promote the progression of gastric adenocarcinoma

ABSTRACT Gastric adenocarcinoma (GAC) is a common malignant tumor, accounting for 95% of gastric cancers. However, the effects and regulatory mechanisms of long non-coding RNA TRPM2-AS (TRPM2-AS) in GAC have not been fully explored. Our study investigates the action mechanism of TRPM2-AS in GAC. After performing quantitative Real-Time polymerase chain reaction or western blotting, we found that the levels of TRPM2-AS and Plasminogen Activator, Urokinase (PLAU) were upregulated in GAC, whereas the level of miR-138-5p was downregulated. Cell function experiments proved that silencing TRPM2-AS suppressed proliferation and migration and induced apoptosis in GAC cells. Bioinformatic analysis and luciferase assay identified the interaction between TRPM2-AS, miR-138-5p, and PLAU. In addition, the inhibitory effect of silencing TRPM2-AS on GAC cells could be partially relieved by PLAU overexpression. In conclusion, our study revealed that TRPM2-AS sponging miR-138-5p to upregulate PLAU could contribute to GAC progression, which might be useful for identifying biomarkers for GAC therapy. Abbreviation Gastric adenocarcinoma (GAC); Long non-coding RNA (lncRNA); lncRNA TRPM2 antisense RNA (TRPM2-AS); Plasminogen Activator, Urokinase (PLAU); Wild-type (WT); mutant (MUT).

CircRNF13 Promotes the Malignant Progression of Pancreatic Cancer through Targeting miR-139-5p/IGF1R Axis.

Background Mounting evidence has shown circular RNAs (circRNAs) play an important role in the initiation and progression of pancreatic cancer (PC). Meanwhile, circRNAs may serve as the biomarkers for the diagnosis, treatment, and prognosis of PC. Therefore, it is urgent to elucidate the function and underlying mechanism of circRNAs in the development of PC. Methods The Cancer-Specific CircRNA Database (CSCD), Circular RNA Interactome database (circinteractome database), and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) were used to verify the expression level of circRNF13 in PC cell lines. Fluorescence in situ hybridization (FISH) and RNase protection assay were used to detect the localization and structure of circRNF13. Then, cell functional experiments were employed to estimate the proliferated, migrated, and invasive abilities in PC. Furthermore, bioinformatic tools, luciferase dual reporter assay, and RT-qPCR were used to investigate the interaction among circRNF13, miR-139-5p, and IGF1R. Eventually, the rescue functional experiments were employed to confirm that circRNF13 targeted the miR-139-5p/IGF1R axis to participate in the development of PC. Results CircRNF13 was overexpressed in PC cell lines compared with the normal pancreatic duct cell line. Additionally, inhibition of circRNF13 impaired the proliferation, migration, and invasion of PC cells. CircRNF13 could serve as the molecular sponge of miR-139-5p to inhibit its association with IGF1R that eventually accelerated the malignant progression of PC. Conclusion CircRNF13 serves as a competitive endogenous RNA of IGF1R to inhibit the function of miR-139-5p that eventually reinforces the malignant phenotype of PC.

MicroRNA miR-145-5p regulates cell proliferation and cell migration in colon cancer by inhibiting chemokine (C-X-C motif) ligand 1 and integrin α2

ABSTRACT Colon cancer (CC), which has high morbidity and mortality, can be regulated by microRNAs. This study aimed to investigate the regulatory function of microRNA miR-145-5p in CC cells. Bioinformatics analysis was used to screen key genes in CC. The expression of miR-145-5p, chemokine (C-X-C motif) ligand 1 (CXCL1), and integrin α2 (ITGA2) in CC was confirmed by quantitative reverse transcription polymerase chain reaction and western blotting. After cell transfection, changes in proliferation and migration in CC cells were detected using the cell counting kit-8 (CCK-8), colony formation assay, and wound healing assay. A luciferase assay was conducted to confirm the interactome of miR-145-5p, CXCL1, and ITGA2 in CC cells. Bioinformatics analysis confirmed that CXCL1 and ITGA2 were key genes in CC. After performing several cell functional experiments, the results confirmed that upregulation of miR-145-5p attenuated proliferation and migration of CC cells. Luciferase assay and western blotting confirmed that CXCL1 and ITGA2 were targets of miR-145-5p, and their expression in CC could be suppressed by miR-145-5p. In conclusion, miR-145-5p is a tumor suppressor in CC and can inhibit the expression of CXCL1 and ITGA2.

ABAT targeted by miR-183-5p regulates cell functions in liver cancer.

Liver cancer triggers a considerable number of global deaths. This work focused on mechanisms as well as impacts of ABAT in liver cancer. Differentially expressed mRNAs in liver cancer were analyzed with The Cancer Genome Atlas (TCGA) database to determine and evaluate the prognostic significance of the target gene ABAT. ABAT was overexpressed to explore its effect on liver cancer. Furthermore, the targeted regulation between miR-183-5p and ABAT was verified through dual-luciferase method. The effects of their expression on liver cancer functions were detected by cell functional experiments like Cell Counting Kit-8 (CCK8), Transwell and flow cytometry. Lastly, the inhibitory effect of ABAT on the tumor was proved in nude mice in vivo. At tissue and cell levels, ABAT was inactivated in liver cancer, and liver cancer patients with lowly expressed ABAT had poor prognosis. Overexpressing ABAT could inhibit cancer cell behaviors, and suppress tumorigenesis in nude mice. Meanwhile, overexpressed ABAT could upregulate E-cadherin in liver cancer cells, while downregulate MMP-9, Vimentin, MMP-2, N-cadherin, Ki67. Of note, miR-183-5p was highly expressed in liver cancer tissue and cells, which could target and downregulate ABAT expression. It was indicated by rescue assay that lowly expressed miR-183-5p could repress functions of liver cancer cells, while such inhibitory effect could be recovered by ABAT silencing. Downstream of miR-183-5p, ABAT was targeted to mediate progression of liver cancer.

LINC00261 regulates EBF1 to suppress malignant progression of thyroid cancer.

We aimed to explore the role of LINC00261 in thyroid cancer (TC) and the potential regulatory mechanism. 40 cases of tumor tissues and adjacent tissues of TC patients were collected, and the expressions of LINC00261 and EBF1 were detected by quantitative real-time polymerase chain reaction (qRT-PCR), and the relationship between LINC00261 and the clinical pathological indicators and prognosis of TC patients were analyzed. Next, LINC00261 overexpression and knockdown cell models were constructed in TC cell lines BPH5-16 and K1, respectively. Cell counting kit-8 (CCK-8) and transwell migration were used to detect the impact of LINC00261 overexpression or silencing on cell proliferative and migration ability. The bioinformatics website was used to screen the possible target gene of LINC00261. qRT-PCR analysis showed that LINC00261 level was markedly reduced in TC tumor tissues, as well as corresponding cell lines. Retrospective analysis showed that low expression of LINC00261 was in positive correlation with the pathological stage, lymphatic and distant metastasis in patients with TC, meanwhile, the expression of LINC00261 was also in positive correlation with overall survival rate of TC patients. Bioinformatics analysis suggested that LINC00261 could target EBF1. Luciferase reporter gene experiment and qRT-PCR analysis suggested that LINC00261 could target EBF1 and that their expressions showed a negative correlation in TC tumor tissues and cells. Cell functional experiments confirmed that LINC00261 can inhibit the proliferative and migration ability of TC cells. Subsequently, the recovery experiment also suggested that silencing EBF1 could reverse the promotion effect of LINC00261 knockdown on the proliferative and migration ability of TC cells; while EBF1 overexpression could reverse the inhibition of LINC00261 on the proliferative and migration ability of TC cells. LINC00261 was markedly downregulated in TC tissues and cells. In addition, the level of LINC00261 was closely related to lymph node and distant metastasis, as well as the prognosis in TC patients. Moreover, LINC00261 could negatively regulate EBF1, thereby promoting the malignant progression of TC.

Hsa_Circ_0001947/MiR-661/DOK7 Axis Restrains Non-Small Cell Lung Cancer Development.

Hsa_circ_0001947 is associated with multiple cancers, but its function in non-small cell lung cancer (NSCLC) is ambiguous and needs further research. The targeting relationship among circ_0001947, miR-661, and downstream of tyrosine kinase 7 (DOK7) was predicted by database and further verified by dual-luciferase reporter assay, while their expressions in cancer tissues and cells were detected by quantitative real-time polymerase chain reaction (qRT-PCR). After transfection, cell biological behaviors and expressions of miRNAs, miR-661 and DOK7 were determined by cell function experiments and qRT-PCR, respectively. Circ_0001947 was low-expressed in NSCLC tissues and cells. Circ_0001947 knockdown intensified cell viability and proliferation, induced cell cycle arrest at S phase, suppressed apoptosis and evidently enhanced miR-510, miR-587, miR-661 and miR-942 levels, while circ_0001947 overexpression did the opposite. MiR-661 was a target gene of circ_0001947 that participated in the regulation of circ_0001947 on cell biological behaviors. Furthermore, DOK7, the target gene of miR-661, partly participated in the regulation of miR-661 on cell viability. Hsa_circ_0001947 acts as a sponge of miR-661 to repress NSCLC development by elevating the expression of DOK7.

Ultrasound microbubbles-mediated miR-216b affects MALAT1-miRNA axis in non-small cell lung cancer cells

MiR-216b is ectopically expressed in various cancers. Ultrasound microbubbles (UTMBs) are an effective method for miRNA delivery. This article mainly explored the involvement of lncRNA in the effects of UTMBs-mediated miR-216b on non-small cell lung cancer (NSCLC) progression. Expressions and relationship of miR-216b and MALAT1 were examined using quantitative real-time polymerase chain reaction (qRT-PCR), Pearson, TargetScan, and dual-luciferase reporter assay. After the transfection with liposome- or UTMBs-mediated miR-216b mimic (M) or MALAT1 overexpression plasmid alone or together, levels of miR-216b and MALAT1, cell biological behaviors, as well as expressions of apoptosis- and epithelial mesenchymal transition (EMT)-related markers were examined using qRT-PCR, cell functional experiments, and western blot. Besides, we used qRT-PCR to quantify the expressions of multiple downstream miRNAs of MALAT1. MiR-216b expression was weakened yet MALAT1 expression was enhanced in NSCLC tissues, and miR-216b was negatively bound to MALAT1. TargetScan analysis manifested that miR-216b, targeted by MALAT1, was down-regulated in NSCLC cells. UTMBs-mediated miR-216b M further intensified miR-216b level yet weakened cell biological behaviors. The inhibitory effect of UTMBs-mediated miR-216b M on cell biological behaviors and MALAT1 expression was greatly better relative to that of miR-216b M. Moreover, miR-216b restrained the cell biological behaviors by repressing MALAT1 expression. We further manifested that miR-216b facilitated the expressions of apoptosis-related markers, but restrained those of EMT-related markers by repressing MALAT1 expression. Moreover, UTMBs-mediated miR-216b M enhanced the expressions of downstream multiple miRNAs of MALAT1, but this tendency was reversed by co-transfection of overexpressed MALAT1 and miR-216b M.Collectively, UTMBs-mediated miR-216b M restrained NSCLC cell growth by modulating the MALAT1-miRNA axis.

SLC41A3 Exhibits as a Carcinoma Biomarker and Promoter in Liver Hepatocellular Carcinoma.

Liver Hepatocellular Carcinoma (LIHC) is the fifth widely occurred carcinoma, which is thought to be the second primary contributor of carcinoma-associated death. There are almost 788,000 death tolls worldwide. Solute carrier family 41 member 3 (SLC41A3) is a member of solute carrier family 41, and it is the key point of numerous researches. Our research attempted to explore the links between SLC41A3 and LIHC through public databases. Higher expression of SLC41A3 displayed an intimate association with higher pathological stages and poorer prognosis. GO and KEGG analysis revealed the possible regulatory pathways of SLC41A3. Additionally, we carried out cell functional experiments to determine the expression of SLC41A3 in the cell lines of LIHC, as well as the effects of its silence on cell proliferation, migration, and invasion. Our data showed that SLC41A3 was greatly increased in the cell lines of LIHC. Moreover, silencing SLC41A3 impeded LIHC cell proliferation, migration, and invasion in vitro. Collectively, our study demonstrated that highly expressed SLC41A3 was a probable indication of LIHC occurrence, and SLC41A3 could be regarded as a prospective target in the treatment of LIHC.

Prognostic value of SEC61G in lung adenocarcinoma: a comprehensive study based on bioinformatics and in vitro validation

BackgroundStudies have shown that the Sec61 gamma subunit (SEC61G) is overexpressed in several tumors and could serve as a potential prognostic marker. However, the correlation between SEC61G and lung adenocarcinoma (LUAD) remains unclear. In the current study, we aimed to demonstrate the prognostic value and potential biological function of the SEC61G gene in LUAD.MethodsPublic datasets were used for SEC61G expression analyses. The prognostic value of SEC61G in LUAD was investigated using the Kaplan–Meier survival and Cox analyses. The correlation between the methylation level of SEC61G and its mRNA expression was evaluated via cBioPortal. Additionally, MethSurv was used to determine the prognostic value of the SEC61G methylation levels in LUAD. Functional enrichment analysis was conducted to explore the potential mechanism of SEC61G. Also, single sample GSEA (ssGSEA) and TIMER online tool were applied to identify the correlation between SEC61G and immune filtration. Furthermore, cell functional experiments were conducted to verify the biological behavior of SEC61G in lung adenocarcinoma cells (LAC).ResultsSEC61G was upregulated in pan-cancers, including LUAD. High SEC61G expression was significantly correlated with worse prognosis in LUAD patients. Multivariate analysis demonstrated that high SEC61G expression was an independent prognostic factor in the TCGA cohort. (HR = 1.760 95% CI: 1.297–2.388, p < 0.001). The methylation level of SEC61G negatively correlated with the SEC61G expression (R = − 0.290, p < 0.001), and patients with low SEC61G methylation had worse overall survival. (p = 0.0014). Proliferation-associated terms such as cell cycle and cell division were significantly enriched in GO and KEGG analysis. Vitro experiments demonstrated that knockdown of SEC61G resulted in decreased cell proliferation, invasion and facilitated apoptosis in LAC. GSEA analysis found that SEC61G expression was associated with the E2F targets. Moreover, SEC61G expression was negatively correlated with the immune cell infiltration including CD4+ T cell, CD8+ T cell, B cell, macrophage, neutrophil, and dendritic cell.ConclusionOur study indicated that overexpression of SEC61G was significantly associated with poor prognosis of LUAD patients and the malignant phenotypes of LUAD cells, suggesting that it could be a novel prognostic biomarker and potential therapeutic target of LUAD.

LBX2-AS1 Activates FSTL3 by Binding to Transcription Factor RARα to Foster Proliferation, Migration, and Invasion of Thyroid Cancer.

Background: Thyroid cancer is a frequent endocrine tumor in women. It is of great significance to investigate the molecular mechanism of progression of thyroid cancer. Methods: Gene expression data set and clinical data were downloaded from The Cancer Genome Atlas database for differential expression analysis. The triplet of downstream transcription factors (TFs) and modulatory genes of target lncRNA in thyroid cancer was predicted by the lncMAP database. mRNA and protein expression of lncRNA LBX2-AS1, RARα, and FSTL3 were detected by qRT-PCR and western blot. The localization of lncRNA LBX2-AS1 in cells was tested by Fluorescence in situ hybridization assay. The RNA immunoprecipitation assay was applied to verify the binding relationship between lncRNA LBX2-AS1 and FSTL3. ChIP and dual-luciferase assays were used to prove the binding relationship between RARα and FSTL3. Cell function experiments were used to test cell proliferation, migration and invasion in each treatment group. The role of lncRNA LBX2-AS1 in thyroid cancer progression was also confirmed in nude mice. Results: Bioinformatics analysis indicated that lncRNA LBX2-AS1, RARα, FSTL3 were remarkably fostered in thyroid cancer tissue, and LBX2-AS1 was evidently correlated with clinical features. The LncMAP triplet prediction showed that LBX2-AS1 recruited TF RARα to modulate FSTL3. RIP assay confirmed that LBX2-AS1 was prominently enriched on RARα. ChIP and dual-luciferase report assays unveiled that RARα bound to the promoter region of FSTL3 and functioned as a TF. Cell function experiments uncovered that LBX2-AS1 boosted the progression of thyroid cancer. The rescue experiments showed that LBX2-AS1 recruited the TF RARα to hasten the transcription activity of FSTL3 and thus promoted the development of thyroid cancer. Conclusion: The integrative results demonstrated that LBX2-AS1 activated FSTL3 by binding to TF RARα to hasten proliferation, migration and invasion of thyroid cancer.

A Pan-Cancer Landscape of HOX-Related lncRNAs and Their Association With Prognosis and Tumor Microenvironment.

The highly conserved homology cassette family (HOX) as well as 18 referenced long non-coding antisense transcripts (HOXATs) play vital roles in the development of some cancers. Nevertheless, their expression patterns as well as their association with cancer prognosis and the tumor microenvironment (TME) in pan-cancers are still unclear. Here, based on public databases, the expression levels of HOXATs, their prognostic potentials, and correlation with tumor mutation burden (TMB), immune cell infiltration, immune subtype, immune response-related genes, and stemness scores corresponding to 33 tumor types were analyzed systematically using R language. The results of the analysis indicated that different cancer tissues show different HOXAT expression profiles. Further, HOXAT expression showed association with cancer prognosis and immune and stemness regulation. Gene set enrichment analysis also demonstrated that HOXATs participate in cancer- and immune-related pathways, and based on their expression levels, HOTAIRM1 and HOXB-AS1 showed potential involvement in oncogenesis as well as possible involvement in immune regulation across a variety of cancer types. Further investigation also confirmed a significantly higher expression of HOXB-AS1 in GBM than in lower grade glioma tissues. Importantly, in vitro cell function experiments indicated that HOXB-AS1 supports cancer stem cell and plays a fundamental role in glioma metastasis. In conclusion, our results provide valuable resources that can guide the investigation of the mechanisms related to the role of HOXATs in cancers as well as therapeutic analysis in this regard.

LINC00641 inhibits the proliferation and invasion of ovarian cancer cells by targeting miR-320a.

BackgroundOvarian cancer is a common malignancy of the female reproductive system, with one of the highest mortality rates among all malignant tumors. However, the pathogenesis of ovarian cancer has not been fully elucidated. This study investigated the role and molecular mechanism of LINC00641 in the development and progression of ovarian cancer.MethodsQuantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression level of LINC00641 in ovarian cancer tissue and adjacent normal tissue. Cell Counting Kit-8 (CCK-8), colony formation, and Transwell assays were used to detect the effects of LINC00641 overexpression on the proliferation and migration of ovarian cancer cells. Bioinformatics analysis and luciferase reporter gene assay were employed to detect the binding of LINC00641 to the downstream target molecule, microRNA-320a (miR-320a). Western blotting was used to determine the effect of miR-320a overexpression on the expression of proliferation-related proteins [Ki-67 and proliferating cell nuclear antigen (PCNA)] and invasion-related proteins (E-cadherin, N-cadherin, and vimentin) in overexpressed LINC00641 cells.ResultsqRT-PCR results showed that LINC00641 was under-expressed in ovarian cancer tissue compared to adjacent tissue. Cell function experiments showed that the overexpression of LINC00641 could significantly inhibit the proliferation and migration of ovarian cancer cells. The luciferase reporter gene assay showed that LINC00641 could bind to miR-320a, and the overexpression of LINC00641 could markedly inhibit the expression of miR-320a in ovarian cancer cells. Overexpression of miR-320a could significantly block the inhibitory effect of LINC00641 on the proliferation and migration of ovarian cancer cells.ConclusionsAs a tumor suppressor gene, LINC00641 can inhibit the proliferation and invasion of ovarian cancer cells by targeting miR-320a. The LINC00641/miR-320a axis may be a new target for the early diagnosis, treatment, or prognosis of ovarian cancer patients.

Therapeutic Potential of CUDC-907 (Fimepinostat) for Hepatocarcinoma Treatment Revealed by Tumor Spheroids-Based Drug Screening.

Background: Cancer is the second leading cause of death globally. However, most of the new anti-cancer agents screened by traditional drug screening methods fail in the clinic because of lack of efficacy. Choosing an appropriate in vitro tumor model is crucial for preclinical drug screening. In this study, we screened anti-hepatocarcinoma (HCC) drugs using a novel spheroid cell culture device. Methods: Four HCC cell lines were three-dimensionally (3D) cultured to screen 19 small molecular agents. 3D-cultured primary HCC cells and a tumor-bearing mouse model were used to verify the candidate anti-hepatocarcinoma agent. Cell function experiments and western blotting were conducted to explore the anti-hepatocarcinoma mechanism of the candidate agent. Results: We found that CUDC-907 can serve as a potent anti-hepatocarcinoma agent. The study data show that CUDC-907 (fimepinostat), a novel dual acting inhibitor of phosphoinositide 3-kinase (PI3K) and histone deacetylase (HDAC), has potent inhibitory effects on HCC cell lines and primary HCC cells in vitro, Animal studies have shown that CUDC-907 can also suppress HCC cells in vivo. Furthermore, we found that CUDC-907 inhibits the PI3K/AKT/mTOR pathway and downregulates the expression of c-Myc, leading to the suppression of HCC cells. Conclusion: Our results suggest that CUDC-907 can be a candidate anti-HCC drug, and the 3D in vitro drug screening method based on our novel spheroid culture device is promising for future drug screening efforts.

Knockdown of long non-coding RNA AGAP2-AS1 suppresses the proliferation and metastasis of glioma by targeting microRNA-497-5p

ABSTRACT Long non-coding RNA (LncRNA) AGAP2-AS1 has been demonstrated to involve in various malignancies. However, the expression and biological effect of AGAP2-AS1 on glioma remain enigmatic. We aimed to explore the effects of AGAP2-AS1 on glioma. Expressions and relationship of AGAP2-AS1 and microRNA-497-5p (miR-497-5p) in different grades of glioma tissues and cell lines as well as normal ones were assessed by quantitative real-time polymerase chain reaction (qRT-PCR), starBase, and dual-luciferase reporter assay. In addition, the effect of AGAP2-AS1 on the cell biological behaviors, epithelial-mesenchymal transition (EMT)-related markers, and miR-497-5p level was detected by cell functional experiments, western blot, and qRT-PCR. After transfection with miR-497-5p mimic (M), inhibitor (I), and AGAP2-AS1 knockdown, miR-497-5p level, cell biological behaviors, and EMT-related markers were detected again. AGAP2-AS1 expression was increased while miR-497-5p expression was decreased in glioma tissues and cell lines, and increase of AGAP2-AS1 expression or reduction of miR-497-5p expression was also correlated with clinicopathological grades of glioma. Furthermore, AGAP2-AS1 knockdown repressed cell biological behaviors and EMT-related markers expressions. Mechanically, AGAP2-AS1 targeted miR-497-5p and AGAP2-AS1 knockdown led to elevation of miR-497-5p expression. In addition, rescue experiments were conducted to validate the vital influence of miR-497-5p on the AGAP2-AS1-regulated proliferation and metastasis of glioma. AGAP2-AS1 may serve as an oncogene in the tumorigenesis of glioma by inhibiting miR-497-5p expression, showing its potential as a prognostic biomarker and a therapeutic target for glioma.

YTHDF1 promotes the proliferation, migration, and invasion of prostate cancer cells by regulating TRIM44

BackgroundProstate cancer (PCa) is one of the most common malignancies in men. YTHDF1 may play an important role in promoting PCa progression, but there is no reports to date on YTHDF1 function in PCa.ObjectiveThis study explored whether YTHDF1 could regulate TRIM44 in PCa cells.MethodsBy querying the TCGA database, we evaluated YTHDF1 expression in PCa, the OS and DFS of YTHDF1, and the correlation between YTHDF1 and TRIM44 in PCa. We constructed vectors to interfere with YTHDF1 expression and overexpress TRIM44 to examine the role of YTHDF1 and TRIM44 in PCa cells. Differentially expressed mRNAs were identified by mRNA sequencing. The levels of YTHDF1, TRIM44, LGR4, SGTA, DDX20, and FZD8 were measured by qRT-PCR and WB was used to determine YTHDF1 and TRIM44 expression. A CCK-8 assay was used to assess cell proliferation. A Transwell chamber assay was used measure cell migration and invasion ability.ResultsYTHDF1 was highly expressed in both Pca tissues and cells. PCa patient prognosis with high YTHDF1 expression was relatively poor. Cell function experiments showed that inhibiting YTHDF1 expression decreased cell proliferation, migration, and invasion. RNA sequencing analysis revealed that YTHDF1 may promote PCa cell proliferation, migration, and invasion by modulating TRIM44 expression. Cell function experiments further verified that YTHDF1 promoted PCa cell proliferation, migration, and invasion by regulating TRIM44.ConclusionsYTHDF1 enhances PCa cell proliferation, migration, and invasion by regulating TRIM44.

FAM87A as a Competing Endogenous RNA of miR-424-5p Suppresses Glioma Progression by Regulating PPM1H.

Far less has been unveiled about the functions of lncRNAs on cancers yet. Here, we reported that lncRNA FAM87A, as a ceRNA of miR-424-5p, played a vital role in glioma development. qRT-PCR result indicated that FAM87A was abnormally downregulated in glioma tissue and cells. Survival analysis suggested that the FAM87A expression was negatively correlated with the survival rate. Effects of FAM87A on human glioma cell lines were also analyzed by MTT, Edu, and transwell assays. FAM87A hastened proliferation and migration of glioma cells. MiR-424-5p, predicted target of FAM87A, was fostered in glioma, which was examined by qRT-PCR. A negative correlation was indicated between FAM87A and miR-424-5p. Results of bioinformatics, dual luciferase, and RIP assays unveiled that FAM87A and miR-424-5p act upon each other. In addition, miR-424-5p targeted 3′-UTR of PPM1H. Also, effects of miR-424-5p/FAM87A on glioma cells were identified via the cell function experiments. FAM87A suppressed PPM1H by binding to miR-424-5p competitively, thereby restraining cell proliferation, migration, and invasion. Collectively, these findings illuminated a new mechanism for glioma progression. Therefore, FAM87A may act as a feasible target for glioma treatment.