Retracted: EphB3‐targeted regulation of miR‐149 in the migration and invasion of human colonic carcinoma HCT116 and SW620 cells

microRNAs play key roles during various crucial cell processes such as proliferation, migration, invasion and apoptosis. Also, microRNAs have been shown to possess oncogenic and tumor‐suppressive functions in human cancers. Here, we describe the regulation and function of miR‐149 in colorectal cancer cell lines. miR‐149 expression patterns were detected in human colorectal cell lines and tissue samples, and then focused on its role in regulation of cell growth, migration, invasion, and its target gene identification. Furthermore, the function of the target gene of miR‐149 was analyzed in vitro and in vivo. miR‐149 expression was downregulated in human colorectal cancer HCT116 and SW620 cell lines compared to the normal colon epithelial NCM460 cell line using quantitative real‐time polymerase chain reaction methods. Further studies indicated that introduction of miR‐149 was able to suppress cell migration and invasion. Then, EphB3 was identified as a direct target gene of miR‐149 in colorectal cancer cells. Moreover, experiments in vitro showed that knockdown expression of EphB3 could suppress cell proliferation and invasion, and ectopic expression of EphB3 restored the phenotypes of CRC cell lines transfected with miR149. In addition, silencing of EphB3 significantly affected cycle progression distribution and increased apoptosis in CRC cell lines. Finally, in vivo results demonstrated that knockdown of EphB3 by siRNA inhibited tumor growth. In conclusion,the important role of miR‐149 in colorectal cancer progression suggesting that miR‐149 may serve as a therapeutic target for colorectal cancer treatment.     

miR‐122 promotes metastasis of clear‐cell renal cell carcinoma by downregulating Dicer

Although overall downregulation of microRNAs (miRNAs) is a general feature of clear‐cell renal cell carcinoma (ccRCC), several miRNAs are consistently upregulated, among which miR‐122 was markedly increased in ccRCC tissues. Our study aims to determine the functional importance and underlying mechanism of miR‐122 in ccRCC metastasis. Here, we demonstrate that the expression of miR‐122 increased in ccRCC tissues, and higher miR‐122 expression was found in ccRCC tissues with metastatic disease than in those without metastasis. The increased miR‐122 levels were associated with poor metastasis‐free survival in ccRCC patients with localized disease. Dicer was validated as a direct functional target of miR‐122. Overexpression of miR‐122 promoted migration and invasion of ccRCC cells in vitro and metastatic behavior of ccRCC cells in vivo. Inhibition of miR‐122 attenuated this metastatic phenotype in vitro. Importantly, miR‐122 exerted its pro‐metastatic properties in ccRCC cells by downregulating Dicer and its downstream effector, the miR‐200 family, thereby inducing epithelial–mesenchymal transition (EMT). Our results suggest an important role of the miR‐122/Dicer/miR‐200s/EMT pathway in ccRCC metastasis. Furthermore, miR‐122 may serve as a biomarker for discriminating ccRCC with metastatic potential.     

Clinicopathological significance of the microRNA‐146a/WASP‐family verprolin‐homologous protein‐2 axis in gastric cancer

Gastric cancer (GC) is one of the most common malignancies, and cancer invasion and metastasis are the leading causes of cancer‐induced death in GC patients. WASP‐family verprolin‐homologous protein‐2 (WASF2), with a role controlling actin polymerization which is critical in the formation of membrane protrusions involved in cell migration and invasion, has been reported to possess cancer‐promoting effects in several cancers. However, data of WASF2's role in GC are relatively few and even contradictory. In this study, we analyzed WASF2 expression in GC tissues and their corresponding adjacent normal tissues. We found that WASF2 was upregulated in GC tissues and high level of WASF2 was associated with lymph node metastasis of GC. Through gain‐ and loss‐of‐function studies, WASF2 was shown to significantly increase GC cells migration and invasion, but had no effect on proliferation in vitro. Importantly, WASF2 was also found to enhance GC metastasis in vivo. Our previous research suggested that WASF2 was a direct target of microRNA‐146a (miR‐146a). Furthermore, we analyzed miR‐146a's level in GC tissues and their corresponding adjacent normal tissues. We found that miR‐146a was downregulated in GC tissues and low miR‐146a level was associated with advanced TNM stage and lymph node metastasis. The level of WASF2 in GC tissues was negatively correlated with miR‐146a expression and had inverse clinicopathologic features. The newly identified miR‐146a/WASF2 axis may provide a novel therapeutic target for GC.     

Retracted: Glabridin attenuates the migratory and invasive capacity of breast cancer cells by activating microRNA‐200c

Current treatments for breast cancer, a common malignancy in human females, are less than satisfactory because of high rates of metastasis. Glabridin (GLA), which acts through the FAK/ROS signaling pathway, has been used as an antioxidant and anti‐metastatic agent. However, little is known regarding the effect of microRNA (miRNA) on GLA's anti‐metastatic activity. The miRNA‐200 family, which is frequently expressed at low levels in triple negative breast cancers, inhibits metastasis by blocking the epithelial–mesenchymal transition. Here, we found that GLA attenuated the migratory and invasive capacity of breast cancer cells by activating miR‐200c. GLA induced the mesenchymal–epithelial transition in vitro and in vivo, as determined by increased expression of the epithelial marker, E‐cadherin, and decreased expression of the mesenchymal marker, vimentin. Overexpression of miR‐200c enhanced the expression of E‐cadherin and decreased the expression of vimentin. Furthermore, in MDA‐MB‐231 and BT‐549 breast cancer cells exposed to GLA, knockdown of miR‐200c blocked the GLA‐induced mesenchymal–epithelial transition and alleviated the GLA‐induced inhibition of migration and invasion. Thus, elevation of miR‐200c by GLA has considerable therapeutic potential for anti‐metastatic therapy for breast cancer patients.     

ATP‐P2Y2‐β‐catenin axis promotes cell invasion in breast cancer cells

Extracellular adenosine 5′‐triphosphate (ATP), secreted by living cancer cells or released by necrotic tumor cells, plays an important role in tumor invasion and metastasis. Our previous study demonstrated that ATP treatment in vitro could promote invasion in human prostate cancer cells via P2Y2, a preferred receptor for ATP, by enhancing EMT process. However, the pro‐invasion mechanisms of ATP and P2Y2 are still poorly studied in breast cancer. In this study, we found that P2Y2 was highly expressed in breast cancer cells and associated with human breast cancer metastasis. ATP could promote the in vitro invasion of breast cancer cells and enhance the expression of β‐catenin as well as its downstream target genes CD44, c‐Myc and cyclin D1, while P2Y2 knockdown attenuated above ATP‐driven events in vitro and in vivo. Furthermore, iCRT14, a β‐catenin/TCF complex inhibitor, could also suppress ATP‐driven migration and invasion in vitro. These results suggest that ATP promoted breast cancer cell invasion via P2Y2‐β‐catenin axis. Thus blockade of the ATP‐P2Y2‐β‐catenin axis could suppress the invasive and metastatic potential of breast cancer cells and may serve as potential targets for therapeutic interventions of breast cancer.     

Targeting liver sinusoidal endothelial cells with miR‐20a‐loaded nanoparticles reduces murine colon cancer metastasis to the liver

Phenotypic transformation of liver sinusoidal endothelial cells is one of the most important stages of liver metastasis progression. The miRNA effects on liver sinusoidal endothelial cells during liver metastasis have not yet been studied. Herein, whole genome analysis of miRNA expression in these cells during colorectal liver metastasis revealed repressed expression of microRNA‐20a. Importantly, downregulation of miR‐20a occurs in parallel with upregulation of its known protein targets. To restore normal miR‐20a levels in liver sinusoidal endothelial cells, we developed chondroitin sulfate‐sorbitan ester nanoparticles conjugated with miR‐20a in a delivery system that specifically targets liver sinusoidal endothelial cells. The restoration of normal mir‐20a levels in these cells induced downregulation of the expression of its protein targets, and this also resulted in a reduction of in vitro LSEC migration and a reduction of in vivo activation and tumor‐infiltrating capacity and ability of the tumor decreased by ～80% in a murine liver metastasis model.     

Flubendazole elicits anti‐metastatic effects in triple‐negative breast cancer via STAT3 inhibition

Tumor metastasis remains the cause of 90% of cancer‐related deaths. Cancer stem cells (CSC) are thought to be responsible for the aggressive and metastatic nature of triple‐negative breast cancers (TNBC), and new therapeutic strategies are being devised to target them. Flubendazole (FLU) is a widely used anthelmintic agent that also exhibits anticancer activity in several cancer types. The aim of this study was to characterize the mechanism of action of FLU on breast cancer stem cell (BCSC)‐like properties and metastasis in TNBC. FLU treatment caused a significant induction of apoptosis, accompanied by G2/M phase accumulation, caspase‐3/‐7 activation and the dysregulation of STAT3 activation in TNBC cells. The latter phenomenon was associated with impairment of cancer stem‐like traits, concomitant with a reduction in the CD24^low/CD44^high, CD24^high/CD49f^high subpopulation, ALDH1 activity and mammosphere formation. The BCSC‐enriched populations exhibited enhanced metastasis with higher STAT3 activation, while FLU administration inhibited tumor growth, angiogenesis and lung and liver metastasis, coinciding with decreased MMP‐2 and MMP‐9 levels in circulating blood. FLU kills not only rapid proliferating tumor cells but also effectively eradicates BCSC‐like cells in vitro and in vivo. Our findings warrant further investigation of FLU as a treatment for metastatic TNBC.     

Tumor‐suppressive microRNA‐218 inhibits cancer cell migration and invasion via targeting of LASP1 in prostate cancer

Our recent studies of the microRNA (miRNA) expression signature in prostate cancer (PCa) indicated that miRNA‐218 (miR‐218) was significantly downregulated in clinical specimens, suggesting that miR‐218 might act as a tumor‐suppressive miRNA in PCa. The aim of the present study was to investigate the functional significance of miR‐218 in PCa and to identify novel miR‐218‐regulated cancer pathways and target genes involved in PCa oncogenesis and metastasis. Restoration of miR‐218 in PCa cell lines (PC3 and DU145) revealed that this miRNA significantly inhibited cancer cell migration and invasion. Gene expression data and in silico analysis demonstrated that LIM and SH3 protein 1 (LASP1) is a potential target of miR‐218 regulation. LASP1 is a cytoskeletal scaffold protein that plays critical roles in cytoskeletal organization and cell migration. Luciferase reporter assays showed that miR‐218 directly regulated expression of LASP1. Moreover, downregulating the LASP1 gene significantly inhibited cell migration and invasion in cancer cells, and the expression of LASP1 was upregulated in cancer tissues. We conclude that loss of tumor‐suppressive miR‐218 enhanced cancer cell migration and invasion in PCa through direct regulation of LASP1. Our data on pathways regulated by tumor‐suppressive miR‐218 provide new insight into the potential mechanisms of PCa oncogenesis and metastasis.     

Co‐delivery of microRNA‐21 antisense oligonucleotides and gemcitabine using nanomedicine for pancreatic cancer therapy

Tumor metastasis occurs naturally in pancreatic cancer, and the efficacy of chemotherapy is usually poor. Precision medicine, combining downregulation of target genes with chemotherapy drugs, is expected to improve therapeutic effects. Therefore, we developed a combined therapy of microRNA‐21 antisense oligonucleotides (ASO‐miR‐21) and gemcitabine (Gem) using a targeted co‐delivery nanoparticle (NP) carrier and investigated the synergistic inhibitory effects on pancreatic cancer cells metastasis and growth. Polyethylene glycol–polyethylenimine–magnetic iron oxide NPs were used to co‐deliver ASO‐miR‐21 and Gem. An anti‐CD44v6 single‐chain variable fragment (scFv_CD44v6) was used to coat the particles to obtain active and targeted delivery. Our results showed that the downregulation of the oncogenic miR‐21 by ASO resulted in upregulation of the tumor‐suppressor genes PDCD4 and PTEN and the suppression of epithelial–mesenchymal transition, which inhibited the proliferation and induced the clonal formation, migration, and invasion of pancreatic cancer cells in vitro. The co‐delivery of ASO‐miR‐21 and Gem induced more cell apoptosis and inhibited the growth of pancreatic cancer cells to a greater extent than single ASO‐miR‐21 or Gem treatment in vitro. In animal tests, more scFv_CD44v6‐PEG‐polyethylenimine/ASO‐magnetic iron oxide NP/Gem accumulated at the tumor site than non‐targeted NPs and induced a potent inhibition of tumor proliferation and metastasis. Magnetic resonance imaging was used to observed tumor homing of NPs. These results imply that the combination of miR‐21 gene silencing and Gem therapy using an scFv‐functionalized NP carrier exerted synergistic antitumor effects on pancreatic cancer cells, which is a promising strategy for pancreatic cancer therapy.     

miR‐203 inhibits cell proliferation,invasion, and migration of non‐small‐cell lung cancer by downregulating RGS17

Involvement of the RGS17 oncogene in the promotion of non‐small‐cell lung cancer (NSCLC) has been reported, but the regulation mechanism in NSCLC remains unclear. MicroRNAs (miRNAs) negatively regulate gene expression, and their dysregulation has been implicated in tumorigenesis. To understand the role of miRNAs in Regulator of G Protein Signaling 17 (RGS17)‐induced NSCLC, we showed that miR‐203 was downregulated during tumorigenesis, and inhibited the proliferation and invasion of lung cancer cells. We then determined whether miR‐203 regulated NSCLC by targeting RGS17. To characterize the regulatory effect of miR‐203 on RGS17, we used lung cancer cell lines, A549 and Calu‐1, and the constructed miR‐203 and RGS17 overexpression vectors. The CCK8 kit was used to determine cell proliferation, and the Transwell® assay was used to measure cell invasion and migration. RT‐PCR, western blots, and immunofluorescence were used to analyze expression of miR‐203 and RGS17, and the luciferase reporter assay was used to examine the interaction between miR‐203 and RGS17. Nude mice were used to characterize in vivo tumor growth regulation. Expression of miR‐203 inhibited proliferation, invasion, and migration of lung cancer cell lines A549 and Calu‐1 by targeting RGS17. The regulatory effect of miR‐203 was inhibited after overexpression of RGS17. The luciferase reporter assay showed that miR‐203 downregulated RGS17 by direct integration into the 3′‐UTR of RGS17 mRNA. In vivo studies showed that expression of miR‐203 significantly inhibited growth of tumors. Taken together, the results suggested that expression of miR‐203 inhibited tumor growth and metastasis by targeting RGS17.     

Tumor‐suppressive microRNA‐223 inhibits cancer cell migration and invasion by targeting ITGA3/ITGB1 signaling in prostate cancer

Analysis of microRNA (miRNA) expression signatures in prostate cancer (PCa) and castration‐resistant PCa has revealed that miRNA‐223 is significantly downregulated in cancer tissues, suggesting that miR‐223 acts as a tumor‐suppressive miRNA by targeting oncogenes. The aim of this study was to investigate the functional roles of miR‐223 and identify downstream oncogenic targets regulated by miR‐223 in PCa cells. Functional studies of miR‐223 were carried out to investigate cell proliferation, migration, and invasion using PC3 and PC3M PCa cell lines. Restoration of miR‐223 significantly inhibited cancer cell migration and invasion in PCa cells. In silico database and genome‐wide gene expression analyses revealed that ITGA3 and ITGB1 were direct targets of miR‐223 regulation. Knockdown of ITGA3 and ITGB1 significantly inhibited cancer cell migration and invasion in PCa cells by regulating downstream signaling. Moreover, overexpression of ITGA3 and ITGB1 was observed in PCa clinical specimens. Thus, our data indicated that downregulation of miR‐223 enhanced ITGA3/ITGB1 signaling and contributed to cancer cell migration and invasion in PCa cells. Elucidation of the molecular pathways modulated by tumor‐suppressive miRNAs provides insights into the mechanisms of PCa progression and metastasis.     

Dual‐specificity tyrosine‐regulated kinase 2 is a suppressor and potential prognostic marker for liver metastasis of colorectal cancer

Colorectal cancer is a common cancer and a leading cause of cancer‐related death worldwide. The liver is a dominant metastatic site for patients with colorectal cancer. Molecular mechanisms that allow colorectal cancer cells to form liver metastases are largely unknown. Activation of epithelial–mesenchymal transition is the key step for metastasis of cancer cells. We recently reported that dual‐specificity tyrosine‐regulated kinase 2 (DYRK2) controls epithelial–mesenchymal transition in breast cancer and ovarian serous adenocarcinoma. The aim of this study is to clarify whether DYRK2 regulates liver metastases of colorectal cancer. We show that the ability of cell invasion and migration was abrogated in DYRK2‐overexpressing cells. In an in vivo xenograft model, liver metastatic lesions were markedly diminished by ectopic expression of DYRK2. Furthermore, we found that patients whose liver metastases expressed low DYRK2 levels had significantly worse overall and disease‐free survival. Given the findings that DYRK2 regulates cancer cell metastasis, we concluded that the expression status of DYRK2 could be a predictive marker for liver metastases of colorectal cancer.     

Expression of DIAPH1 is up‐regulated in colorectal cancer and its down‐regulation strongly reduces the metastatic capacity of colon carcinoma cells

In most cases, metastatic colorectal cancer is not curable, thus new approaches are necessary to identify novel targets for colorectal cancer therapy. Actin‐binding‐proteins (ABPs) directly regulate motility of metastasising tumor cells, and for cortactin an association with colon cancer metastasis has been already shown. However, as its depletion only incompletely inhibits metastasis, additional, more suitable cellular targets have to be identified. Here we analyzed expression of the ABPs, DIAPH1, VASP, N‐WASP, and fascin in comparison with cortactin and found that, besides cortactin, DIAPH1 was expressed with the highest frequency (63%) in colorectal cancer. As well as cortactin, DIAPH1 was not detectable in normal colon tissue and expression of both proteins was positively correlated with metastasis of colorectal cancer. To analyse the mechanistic role of DIAPH1 for metastasis of colon carcinoma cells in comparison with cortactin, expression of the proteins was stably down‐regulated in the human colon carcinoma cell lines HT‐29, HROC‐24 and HCT‐116. Analysis of metastasis of colon carcinoma cells in SCID mice revealed that depletion of DIAPH1 reduced metastasis 60‐fold and depletion of cortactin 16‐fold as compared with control cells. Most likely the stronger effect of DIAPH1 depletion on colon cancer metastasis is due to the fact that in vitro knock down of DIAPH1 impaired all steps of metastasis; adhesion, invasion and migration while down‐regulation of cortactin only reduced adhesion and invasion. This very strong reducing effect of DIAPH1 depletion on colon carcinoma cell metastasis makes the protein a promising therapeutic target for individualized colorectal cancer therapy.     

Cancer‐associated fibroblast‐derived Lumican promotes gastric cancer progression via the integrin β1‐FAK signaling pathway

Gastric cancer (GC) is one of the most frequent malignant tumors worldwide and is associated with high invasiveness, high metastasis and poor prognosis. Cancer‐associated fibroblasts (CAFs), residing around tumor cells in tumor stroma, are important modifiers of tumor initiation and progression. However, the molecular mechanisms by which CAF's modulate tumor development have yet not to be characterized in GC. Here we performed tissue assay analyses identifying that Lumican, an extracellular matrix protein, is highly expressed in human gastric CAFs and its expression is positively associated with depth of invasion, lymph node metastasis, TNM stage and poor survival rate of GC. Functional studies revealed that integrin β1‐FAK signaling pathways mediate the promoting effect of Lumican on GC cell proliferation, migration and invasion in vitro. In accordance with these observations, in GC cells co‐cultured with CAFs in which Lumican had been knocked down, decreased gastric tumor growth and metastasis in vivo was apparent. In summary, CAF‐derived Lumican contributes to tumorigenesis and metastasis of GC by activating the integrin β1‐FAK signaling pathway.     

Suppression of AKT expression by miR‐153 produced anti‐tumor activity in lung cancer

Lung cancer is one of the leading causes of cancer death worldwide. microRNAs have been shown to be a novel class of regulators in lung cancer. Here, we explored the role of miR‐153 in the pathogenesis of lung cancer and its therapeutic potential. miR‐153 was significantly decreased in lung cancer tissues than the adjacent tissues. The protein and mRNA levels of protein kinase B (AKT), which were shown to promote tumor growth, were both increased in lung cancer tissues than adjacent tissues. Overexpression of miR‐153 significantly inhibited AKT protein expression, which were abrogated by co‐transfection of AMO‐153, the specific inhibitor of miR‐153. Luciferase assay showed that transfection of miR‐153 markedly suppressed the fluorescent intensity of chimeric vectors carrying the 3'UTR of AKT1, while produced no effect on the mutant construct, indicating that AKT is regulated by miR‐153. Overexpression of miR‐153 significantly inhibited the proliferation and migration, and promoted apoptosis of cultured lung cancer cells in vitro, and suppressed the growth of xenograft tumors in vivo. Interestingly, lung cancer cells with lower endogenous miR‐153 expression are more sensitive to ectopic overexpressed miR‐153. The IC₅₀ of miR‐153 on lung cancer cells is positive correlated with the endogenous miR‐153 level, while negative correlated with AKT level. Knockdown of AKT expression suppressed lung cancer cell proliferation. In summary, miR‐153 exerted anti‐tumor activity in lung cancer by targeting on AKT. The sensitivity of lung cancer cells to miR‐153 is determined by its endogenous miR‐153 level.     

MicroRNA‐143‐3p inhibits colorectal cancer metastases by targeting ITGA6 and ASAP3

MicroRNAs, which regulate mRNAs, operate through a variety of signaling pathways to participate in the development of colorectal cancer (CRC). In this study, we found that microRNA (miR)‐143‐3p expression was significantly lower in both CRC and liver metastatic CRC tissues from liver compared with normal colonic tissues. Functional assays showed that miR‐143‐3p inhibited CRC cell invasion and migration in vitro. Using a bioinformatics approach, we identified miR‐143‐3p target mRNAs. Among the candidate targets, only the expression of integrin alpha 6 (ITGA6) and ArfGAP with the SH3 domain and ankyrin repeat and PH domain 3 (ASAP3) were significantly reduced by miR‐143‐3p mimics as examined by western blot, and the metastasis potential of CRC cells was attenuated by endogenous ITGA6 and ASAP3 knockdown, determined by migration and invasion assays. Both ITGA6 and ASAP3 were upregulated in CRC tissues compared to normal tissues. Analysis of the relationship between clinicopathological features and ITGA6/ASAP3 protein expression in 200 patients with CRC showed a significant difference in positive ITGA6 expression between the early stage (I + II) and the advanced stage (III + IV), and ASAP3 expression levels positively correlated with metastasis in the lymph nodes. These results indicate that miR‐143‐3p acts as an anti‐oncogene by downregulating ITGA6/ASAP3 protein expression and could offer new insight into potential therapeutic targets for CRC.