MicroRNA‐1258, regulated by c‐Myb,inhibits growth and epithelial‐to‐mesenchymal transition phenotype via targeting SP1 in oral squamous cell carcinoma

The biological function and underlying mechanism of miR‐1258 has seldom been investigated in cancer progression, including in oral squamous cell carcinoma (OSCC). In the current study, we revealed that the expression level of miR‐1258 was significantly down‐regulated in OSCC tissues and cell lines. Restoration of miR‐1258 decreased OSCC cell growth and invasion. The luciferase and Western blot assays revealed that SP1 protein was a downstream target of miR‐1258. Overexpression of SP1 dismissed miR‐1258’s effect on cell growth and invasion. We also revealed that c‐Myb inhibited miR‐1258 by directly binding at its promoter. In addition, miR‐1258 inhibited PI3K/AKT and ERK signalling pathway activity. Taken together, these findings demonstrated that miR‐1258 may function as a tumour‐suppressive micorRNA in OSCC and suggested that miR‐1258 may be a potential therapeutic target for OSCC patients.     

MicroRNA‐876‐5p inhibits cell proliferation,migration and invasion by targeting c‐Met in osteosarcoma

Recently, aberrant expression of miR‐876‐5p has been reported to participate in the progression of several human cancers. However, the expression and function of miR‐876‐5p in osteosarcoma (OS) are still unknown. Here, we found that the expression of miR‐876‐5p was significantly down‐regulated in OS tissues compared to para‐cancerous tissues. Clinical association analysis indicated that underexpression of miR‐876‐5p was positively correlated with advanced clinical stage and poor differentiation. More importantly, OS patients with low miR‐876‐5p level had a significant shorter overall survival compared to miR‐876‐5p high‐expressing patients. In addition, gain‐ and loss‐of‐function experiments demonstrated that miR‐876‐5p restoration suppressed whereas miR‐876‐5p knockdown promoted cell proliferation, migration and invasion in both U2OS and MG63 cells. In vivo studies revealed that miR‐876‐5p overexpression inhibited tumour growth of OS in mice. Mechanistically, miR‐876‐5p reduced c‐Met abundance in OS cells and inversely correlated c‐Met expression in OS tissues. Herein, c‐Met was recognized as a direct target of miR‐876‐5p using luciferase reporter assay. Notably, c‐Met restoration rescued miR‐876‐5p attenuated the proliferation, migration and invasion of OS cells. In conclusion, these findings indicate that miR‐876‐5p may be used as a potential therapeutic target and promising biomarker for the diagnosis and prognosis of OS.     

MiR‐376a suppresses the proliferation and invasion of non‐small‐cell lung cancer by targeting c‐Myc

It has been reported that miR‐376a is involved in the formation and progression of several types of cancer. However, the expression and function of miR‐376a is still unknown in non‐small cell lung carcinomas (NSCLC). In this study, the expression of miR‐376a in NSCLC tissues and cell lines were examined by real‐time PCR, the effects of miR‐376a on cell proliferation, apoptosis and invasion were evaluated in vitro. Luciferase reporter assay was performed to identify the targets of miR‐376a. The results showed that miR‐376a was significantly downregulated in NSCLC tissues and cell lines. Restoration of miR‐376a in NSCLC cell line A549 significantly inhibited cell proliferation, increased cell apoptosis and suppressed cell invasion, compared with control‐transfected A549 cells. Luciferase reporter assay showed that c‐Myc, an oncogene that regulating cell survival, angiogenesis and metastasis, was a direct target of miR‐376a. Over‐expression of miR‐376a decreased the mRNA and protein levels of c‐Myc in A549 cells. In addition, upregulation of c‐Myc inhibited miR‐376a‐induced inhibition of cell proliferation and invasion in A549 cells. Therefore, our results indicate a tumor suppressor role of miR‐376a in NSCLC by targeting c‐Myc. miR‐376a may be a promising therapeutic target for NSCLC.     

MicroRNA‑224 promotes the sensitivity of osteosarcoma cells to cisplatin by targeting Rac1

Osteosarcoma is the most common primary bone tumour in children and adolescents. Accumulating evidence has shown that microRNAs (miRNAs) participate in the development of almost all types of cancer. Here, we investigated the role of miR‐224 in the development and progression of osteosarcoma. We demonstrated that miR‐224 was down‐regulated in osteosarcoma cell lines and tissues. Lower miR‐224 levels were correlated with shorter survivalin osteosarcoma patients. Furthermore, overexpression of miR‐224 suppressed osteosarcoma cell proliferation, migration and invasion and contributed to the increased sensitivity of MG‐63 cells to cisplatin. We identified Rac1 as a direct target gene of miR‐224 in osteosarcoma. Rac1 expression was up‐regulated in the osteosarcoma cell lines and tissues, and there was an inverse correlation between Rac1 and miR‐224 expression in osteosarcoma tissues. Furthermore, rescuing Rac1 expression decreased the sensitivity of miR‐224‐overexpressing MG‐63 cells to cisplatin. We also demonstrated that ectopic expression of Rac1 promoted the proliferation, migration and invasion of miR‐224‐overexpressing MG‐63 cells. These data suggest that miR‐224 plays a tumour suppressor role in the development of osteosarcoma and is related to the sensitivity of osteosarcoma to cisplatin.     

Long non‐coding RNA deleted in lymphocytic leukaemia 1 promotes hepatocellular carcinoma progression by sponging miR‐133a to regulate IGF‐1R expression

Long non‐coding RNA (lncRNA) deleted in lymphocytic leukaemia 1 (DLEU1) was reported to be involved in the occurrence and development of multiple cancers. However, the exact expression, biological function and underlying mechanism of DLEU1 in hepatocellular carcinoma (HCC) remain unclear. In this study, real‐time quantitative polymerase chain reaction (qRT‐PCR) in HCC tissues and cell lines revealed that DLEU1 expression was up‐regulated, and the increased DLEU1 was closely associated with advanced tumour‐node‐metastasis stage, vascular metastasis and poor overall survival. Function experiments showed that knockdown of DLEU1 significantly inhibited HCC cell proliferation, colony formation, migration and invasion, and suppressed epithelial to mesenchymal transition (EMT) process via increasing the expression of E‐cadherin and decreasing the expression of N‐cadherin and Vimentin. Luciferase reporter gene assay and RNA immunoprecipitation (RIP) assay demonstrated that DLEU1 could sponge miR‐133a. Moreover, miR‐133a inhibition significantly reversed the suppression effects of DLEU1 knockdown on HCC cells. Besides, we found that silenced DLEU1 significantly decreased insulin‐like growth factor 1 receptor (IGF‐1R) expression (a target of miR‐133a) and its downstream signal PI3K/AKT pathway in HCC cells, while miR‐133a inhibitor partially reversed this trend. Furthermore, DLEU1 knockdown impaired tumour growth in vivo by regulating miR‐133a/IGF‐1R axis. Collectively, these findings indicate that DLEU1 promoted HCC progression by sponging miR‐133a to regulate IGF‐1R expression. Deleted in lymphocytic leukaemia 1/miR‐133a/IGF‐1R axis may be a novel target for treatment of HCC.     

miR‐516a‐3p inhibits breast cancer cell growth and EMT by blocking the Pygo2/Wnt signalling pathway

miR‐516a‐3p has been reported to play a suppressive role in several types of human tumours. However, the expression level, biological function and fundamental mechanisms of miR‐516a‐3p in breast cancer remain unclear. In the present study, we found that miR‐516a‐3p expression was down‐regulated and Pygopus2 (Pygo2) expression was up‐regulated in human breast cancer tissues and cells. Through analysing the clinicopathological characteristics, we demonstrated that low miR‐516a‐3p expression or positive Pygo2 expression was a predictor of poor prognosis for patients with breast cancer. The results of a dual luciferase reporter assay and Western blot analysis indicated that Pygo2 was a target gene of miR‐516a‐3p. Moreover, overexpression of miR‐516a‐3p inhibited cell growth, migration and invasion as well as epithelial‐mesenchymal transition (EMT) of breast cancer cells, whereas reduced miR‐516a‐3p expression promoted breast cancer cell growth, migration, invasion and EMT. Furthermore, we showed that miR‐516a‐3p suppressed cell proliferation, metastasis and EMT of breast cancer cells by inhibiting Pygo2 expression. We confirmed that miR‐516a‐3p exerted an anti‐tumour effect by inhibiting the activation of the Wnt/β‐catenin pathway. Finally, xenograft tumour models were used to show that miR‐516a‐3p inhibited breast cancer cell growth and EMT via suppressing the Pygo2/Wnt signalling pathway. Taken together, these results show that miR‐516a‐3p inhibits breast cancer cell growth, metastasis and EMT by blocking the Pygo2/ Wnt/β‐catenin pathway.     

Upregulation of miR‐874‐3p and miR‐874‐5p inhibits epithelial ovarian cancer malignancy via SIK2

Based on miR‐874 expression levels in the GSE47841 microarray, we hypothesized that the mature products of miR‐874, miR‐874‐3p, or miR‐874‐5p, would inhibit epithelial ovarian cancer (EOC) cell proliferation, metastasis, and chemoresistance. We first examined miR‐874‐3p and miR‐874‐5p expression levels in primary EOC tumor tissue samples and found that they were significantly decreased. 3‐(4,5‐Dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide (MTT) cell proliferation and transwell assays revealed that miR‐874‐3p and miR‐874‐5p significantly inhibit EOC cell proliferation, migration, and invasion. Then, using MTT and soft agar assays of paclitaxel‐treated Caov3 and SKOV3 cells transfected with miR‐874‐3p and miR‐874‐5p, we found that miR‐874‐3p and miR‐874‐5p enhance EOC cell chemosensitivity. We then confirmed that serine/threonine‐protein kinase 2 (SIK2) was a target gene of miR‐874‐3p and miR‐874‐5p. Overall, the results of this study indicate that SIK2 expression can serve as a prognostic biomarker for EOC and that miR‐874‐3p and miR‐874‐5p have the potential to enhance clinical treatment of EOC.     

MiR‐139‐5p,miR‐940 and miR‐193a‐5p inhibit the growth of hepatocellular carcinoma by targeting SPOCK1

The study was aimed to screen out miRNAs with differential expression in hepatocellular carcinoma (HCC), and to explore the influence of the expressions of these miRNAs and their target gene on HCC cell proliferation, invasion and apoptosis. MiRNAs with differential expression in HCC were screened out by microarray analysis. The common target gene of these miRNAs (miR‐139‐5p, miR‐940 and miR‐193a‐5p) was screened out by analysing the target genes profile (acquired from Targetscan) of the three miRNAs. Expression levels of miRNAs and SPOCK1 were determined by quantitative real time polymerase chain reaction (qRT‐PCR). The target relationships were verified by dual luciferase reporter gene assay and RNA pull‐down assay. Through 3‐(4,5‐dimethyl‐2‐thiazolyl)‐2,5‐diphenyl‐2‐H‐tetrazolium bromide,thiazolyl blue tetrazolium bromide (MTT) and transwell assays and flow cytometry, HCC cell viability, invasion and apoptosis were determined. In vivo experiment was conducted in nude mice to investigate the influence of three miRNAs on tumour growth. Down‐regulation of miR‐139‐5p, miR‐940 and miR‐193a‐5p was found in HCC. Overexpression of these miRNAs suppressed HCC cell viability and invasion, promoted apoptosis and inhibited tumour growth. SPOCK1, the common target gene of miR‐139‐5p, miR‐940 and miR‐193a‐5p, was overexpressed in HCC. SPOCK1 overexpression promoted proliferation and invasion, and restrained apoptosis of HCC cells. MiR‐139‐5p, miR‐940 and miR‐193a‐5p inhibited HCC development through targeting SPOCK1.     

MiR‐139‐5p inhibits the tumorigenesis and progression of oral squamous carcinoma cells by targeting HOXA9

Our study sought to clarify the effects of microRNA‐139‐5p (miR‐139‐5p) in the tumorigenesis and progression of oral squamous cell carcinoma (OSCC) by regulating HOXA9. MiR‐139‐5p and HOXA9 expression in OSCC tissues, tumour adjacent tissues, OSCC cells and normal cells were tested by qRT‐PCR. SAS and CAL‐27 cell lines were selected in among four OSCC cell lines and then transfected with miR‐139‐5p mimics, pEGFP‐HOXA9 and cotransfected with miR‐139‐5p mimics + pEGFP‐HOXA9. We used MTT, colony formation, transwell and wound healing assays to analyse cell viability, proliferation, invasion and migration. The target relationship between miR‐139‐5p and HOXA9 was verified by luciferase reporter assay and Western blot, respectively. MiR‐139‐5p was down‐regulated, whereas HOXA9 was up‐regulated in OSCC tissues and cells. The proliferation, invasion and migration ability of SAS and CAL‐27 cells in miR‐139‐5p mimics group were significantly weaker than those in the control group and the miR‐NC group (P < 0.01). MiR‐139‐5p can negatively regulate HOXA9. The proliferation, invasion and migration of SAS and CAL‐27 cells in the miR‐139‐5p mimics + pEGFP‐HOXA9 group were not significantly different from those in the blank control and negative control groups (P > 0.05). Our results indicated that miR‐139‐5p could directly inhibit HOXA9, which might be a potential mechanism in inhibiting the proliferation, invasiveness and migration of OSCC cells.     

Tumor‐suppressive roles of ΔNp63β‐miR‐205 axis in epithelial–mesenchymal transition of oral squamous cell carcinoma via targeting ZEB1 and ZEB2

We previously revealed that epithelial‐to‐mesenchymal transition (EMT) was mediated by ΔNp63β, a splicing variant of ΔNp63, in oral squamous cell carcinoma (OSCC). Recent studies have highlighted the involvement of microRNA (miRNA) in EMT of cancer cells, though the mechanism remains unclear. To identify miRNAs responsible for ΔNp63β‐mediated EMT, miRNA microarray analyses were performed by ΔNp63β‐overexpression in OSCC cells; SQUU‐B, which lacks ΔNp63 expression and displays EMT phenotypes. miRNAs microarray analyses revealed miR‐205 was the most up‐regulated following ΔNp63β‐overexpression. In OSCC cells, miR‐205 expression was positively associated with ΔNp63 and negatively with zinc‐finger E‐box binding homeobox (ZEB) 1 and ZEB2, potential targets of miR‐205. miR‐205 overexpression by miR‐205 mimic transfection into SQUU‐B cells led to decreasing ZEB1, ZEB2, and mesenchymal markers, increasing epithelial markers, and reducing cell motilities, suggesting inhibition of EMT phenotype. Interestingly, the results opposite to this phenomenon were obtained by transfection of miR‐205 inhibitor into OSCC cells, which express ΔNp63 and miR‐205. Furthermore, target protector analyses revealed direct regulation by miR‐205 of ZEB1 and ZEB2 expression. These results showed tumor‐suppressive roles of ΔNp63β and miR‐205 by inhibiting EMT thorough modulating ZEB1 and ZEB2 expression in OSCC.     

MicroRNA‐29b‐3p suppresses oral squamous cell carcinoma cell migration and invasion via IL32/AKT signalling pathway

Oral squamous cell carcinoma (OSCC) is aggressive accompanied with poor prognosis. We previously isolated the most invasive cells resembling the invasive tumour front by microfluidic technology and explored their differentially expressed microRNAs (miRNAs) in our previous work. Here, we verified the miR‐29b‐3p as a guarder that suppressed migration and invasion of OSCC cells and was down‐regulated in the most invasive cells. Besides that, the invasion suppression role of miR‐29b‐3p was achieved through the IL32/AKT pathway. Thus, miR‐29b‐3p and IL32 might serve as therapeutic targets for blocking the progression and improving the outcome of OSCC.     

Downregulation of Microrna‐148a in Cancer‐Associated Fibroblasts from Oral Cancer Promotes Cancer Cell Migration and Invasion by Targeting Wnt10b

It is well established that crosstalk between cancer‐associated fibroblasts (CAFs) and cancer cells plays a critical role in the occurrence and development of oral squamous cell carcinoma (OSCC). The molecular mechanisms underlying such interaction, however, remain far from clear. Accumulating data have indicated that microRNAs involved in tumor microenvironment, particularly in CAFs, contribute to the activation of fibroblasts and metastasis of cancer cells. Here, we showed that miR‐148a was downregulated in CAFs compared with normal fibroblasts isolated from clinical OSCC tissue. Investigation of miR‐148a function in fibroblasts demonstrated that overexpression of miR‐148a in CAFs significantly impaired the migration and invasion of oral carcinoma cells (SCC‐25) by directly targeting WNT10B. Taken together, these data suggested that miR‐148a might be a novel candidate target for the treatment of OSCC.     

MicroRNA‐379 suppresses osteosarcoma progression by targeting PDK1

Osteosarcoma is the most common primary bone tumour. Increasing evidence has demonstrated the pathogenic role of microRNA (miRNAs) dysregulation in tumour development. miR‐379 was previously reported to function as an oncogenic or tumour‐suppressing miRNA in a tissue‐dependent manner. However, its function in osteosarcoma remains unknown. In this study, we found that the expression of miR‐379 was downregulated in osteosarcoma tissues and cell lines. Further functional characterization revealed that miR‐379 suppressed osteosarcoma cell proliferation and invasion in vitro and retarded the growth of osteosarcoma xenografts in vivo. Mechanistically, PDK1 was identified as the direct target of miR‐379 in osteosarcoma, in which PDK1 expression was up‐regulated and showed inverse correlation with miR‐379. Enforced expression of PDK1 promoted osteosarcoma cell proliferation and rescued the anti‐proliferative effect of miR‐379. These data suggest that miR‐379 could function as a tumour‐suppressing miRNA via targeting PDK1 in osteosarcoma.     

MicroRNA‐1271 suppresses the proliferation and invasion of colorectal cancer cells by regulating metadherin/Wnt signaling

Recent studies have reported an important role for microRNA‐1271 (miR‐1271) in tumorigenesis. However, the role of miR‐1271 in colorectal cancer remains unknown. Here, we found that miR‐1271 was significantly decreased in colorectal cancer tissues and cell lines. Overexpression of miR‐1271 inhibited cell proliferation, colony formation, cell invasion, and induced cell cycle arrest in colorectal cancer cells. Metadherin (MTDH) was identified as a target gene of miR‐1271. Moreover, miR‐1271 negatively regulated MTDH expression in colorectal cancer cells and reversely correlated with MTDH expression in colorectal cancer specimens. Additionally, miR‐1271 also regulated the activation of Wnt signaling in colorectal cancer cells. The restoration of MTDH expression significantly reversed the antitumor effect of miR‐1271 in colorectal cancer cells. These findings indicate an important role for miR‐1271/MTDH in the tumorigenesis of colorectal cancer, and suggest that miR‐1271 may be a novel therapeutic target for colorectal cancer.     

Plasma from pre‐eclamptic patients induces the expression of the anti‐angiogenic miR‐195‐5p in endothelial cells

We examined the effect of plasma incubation from preeclampsia pregnant on the antiangiogenic miR‐195‐5p expression. Higher miR‐195‐5p expression was found in cultures incubated with preeclampsia plasma compared to those incubated with healthy pregnant plasma. Next, as VEGF is a target of miR‐195‐5p we have quantified its expression by real‐time qPCR and ELISA. We found reduced VEGF levels in culture incubated with preeclampsia plasma. Therefore, we have concluded that the higher expression of miR‐195‐5p in endothelial cell cultures incubated with preeclampsia plasma may contribute to decreased expression of VEGFA (gene and protein) and increased antiangiogenic status in preeclampsia. Therefore, this miR may be an important target in preeclampsia.