The novel role and function of LINC01235 in metastasis of gastric cancer cells by inducing epithelial-mesenchymal transition

LncRNAs play a vital role in the tumorigenesis of gastric cancer (GC). This study determined that LINC01235 expression has greater fold changes by analyzing TCGA RNA-Seq data. The qRT-PCR assay confirmed that LINC01235 is significantly over-expressed in GC cells and tissues. Additionally, the overall survival analysis showed that patients with a higher LINC01235 expression had a poorer prognosis than those with a lower LINC01235 expression. Univariate Cox regression analysis indicated that high LINC01235 expression is positively correlated with poor prognosis. Moreover, LINC01235 was an independent poor prognostic marker for GC in multivariate Cox analysis. In vitro assays suggested that LINC01235 knockdown suppresses GC cell migration and invasion. GSEA revealed that high LINC01235 expression is strongly enriched in the EMT pathway. Western blotting results revealed that LINC01235 silencing decreases the expression of EMT-induced proteins. In conclusion, LINC01235 can promote GC cell metastasis via EMT and function as a prognostic biomarker.     

Long non‐coding RNA LINC01225 promotes proliferation,invasion and migration of gastric cancer via Wnt/β‐catenin signalling pathway

Emerging evidence has classified the aberrant expression of long non‐coding RNAs (lncRNAs) as a basic signature of various malignancies including gastric cancer (GC). LINC01225 has been shown to act as a hepatocellular carcinoma‐related gene, with its expression pattern and biological function not clarified in GC. Here, we verified that LINC01225 was up‐regulated in tumour tissues and plasma of GC. Analysis with clinicopathological information suggested that up‐regulation of LINC01225 was associated with advanced disease and poorer overall survival. Receiver operating characteristic (ROC) analysis showed that plasma LINC01225 had a moderate accuracy for diagnosis of GC. In addition, knockdown of LINC01225 led to retardation of cell proliferation, invasion and migration, and overexpression of LINC01225 showed the opposite effects. Mechanistic investigations showed that LINC01225 silencing inhibited epithelial‐mesenchymal transition (EMT) process and attenuated Wnt/β‐catenin signalling of GC. Furthermore, ectopic expression of Wnt1 or suppression of GSK‐3β abolished the si‐LINC01225‐mediated suppression against EMT, thereby promoting cell proliferation, invasion and migration of GC. In conclusion, LINC01225 promotes the progression of GC through Wnt/β‐catenin signalling pathway, and it may serve as a potential target or strategy for diagnosis or treatment of GC.     

Long noncoding RNA ZEB1-AS1 attenuates ferroptosis of gastric cancer cells through modulating miR-429/BGN axis

Gastric cancer (GC) is the fifth utmost common malignant cancer type globally, in which ferroptosis acts a critical function in the progress of GC. Long noncoding RNA ZEB1-AS1 has been recognized in numerous cancers, but the role of ZEB1-AS1 in ferroptosis remains obscure. Hence, we investigated the efficacy of ZEB1-AS1 on ferroptosis of GC cells. The cell growth and viability were analyzed via cell counting kit assay and xenograft tumor model in vivo and in vitro, respectively. The RNA and protein expression were measured by qRT-PCR and western blot analysis assay, respectively. The levels of Fe²⁺, malondialdehyde (MDA), and lipid reactive oxygen species (ROS) were tested to determine ferroptosis. The erastin and RSL3 were used to induce ferroptosis. The mechanism was analyzed via luciferase reporter gene and RIP assays. The treatment of ferroptosis inducer Erastin and RSL3 suppressed the viability of GC cells and the ZEB1-AS1 overexpression rescued the phenotype in the cells. The levels of Fe²⁺, MDA, and ROS were enhanced through the depletion of ZEB1-AS1 in Erastin/RSL3 treated GC cells. ZEB1-AS1 directly sponged miR-429 in GC cells and miR-429 targeted BGN in GC cells, and the inhibition of miR-429 rescued ZEB1-AS1 depletion-inhibited BGN expression. We validated that miR-429 induced and BGN-repressed ferroptosis in cancer cells. The BGN overexpression and miR-429 suppression could reverse the efficacy of ZEB1-AS1 on proliferation and ferroptosis in cancer cells. The expression of ZEB1-AS1 and BGN was enhanced and miR-429 expression was decreased in clinical GC tissues. ZEB1-AS1 attenuated ferroptosis of cancer cells by modulating miR-429/BGN axis.     

The calcium pump PMCA4 prevents epithelial-mesenchymal transition by inhibiting NFATc1-ZEB1 pathway in gastric cancer

Epithelial-mesenchymal transition (EMT) is considered as the key mechanism involved in cancer metastasis. Several studies showed that various cell membrane calcium channels play different roles in cancer metastasis. In the present study, the potential role of ATPase plasma membrane Ca²⁺ transporting 4 (PMCA4) in regulating EMT in gastric cancer (GC) was investigated. GC patients who underwent radical surgery were enrolled in this study. In vitro human GC cell lines MKN45 and NCI-N87 were used, and MKN45 cells were injected in nude mice to evaluate tumor development. Our results showed that low PMCA4 expression was associated with advanced TNM stage and poor prognosis in GC patients. Knockdown of PMCA4 suppressed E-cadherin, grainyhead like 2 (GRHL2) and ovo-like 1 (OVOL1) expression, up-regulated vimentin expression, increased migration and invasion ability, and promoted the resistance to cytotoxic drug. Furthermore, GC cells displayed an elongated fibroblastoid morphology when PMCA4 was knockdown. PMCA4 overexpression resulted in an up-regulated E-cadherin expression and decreased migration and invasion ability. In vivo metastasis assay showed that PMCA4 overexpression resulted in a decreased incidence of lung metastasis. PMCA4 inhibition increased ZEB1 expression and nuclear accumulation of nuclear factor of activated T-cell isoform c1 (NFATc1). EMT induced by PMCA4 inhibition could be prevented by the knockdown of NFATc1 or ZEB1. In addition, cyclosporine A prevented EMT induced by PMCA4 inhibition by suppressing the NFATc1-ZEB1 pathway. Our data identified a novel mechanism in the regulation of EMT in GC, and provided a novel target in the treatment of EMT subtype in GC.     

ZEB1‐mediated vasculogenic mimicry formation associates with epithelial–mesenchymal transition and cancer stem cell phenotypes in prostate cancer

The zinc finger E‐box‐binding homeobox 1 (ZEB1) induced the epithelial–mesenchymal transition (EMT) and altered ZEB1 expression could lead to aggressive and cancer stem cell (CSC) phenotypes in various cancers. Tissue specimens from 96 prostate cancer patients were collected for immunohistochemistry and CD34/periodic acid–Schiff double staining. Prostate cancer cells were subjected to ZEB1 knockdown or overexpression and assessment of the effects on vasculogenic mimicry formation in vitro and in vivo. The underlying molecular events of ZEB1‐induced vasculogenic mimicry formation in prostate cancer were then explored. The data showed that the presence of VM and high ZEB1 expression was associated with higher Gleason score, TNM stage, and lymph node and distant metastases as well as with the expression of vimentin and CD133 in prostate cancer tissues. Furthermore, ZEB1 was required for VM formation and altered expression of EMT‐related and CSC‐associated proteins in prostate cancer cells in vitro and in vivo. ZEB1 also facilitated tumour cell migration, invasion and clonogenicity. In addition, the effects of ZEB1 in prostate cancer cells were mediated by Src signalling; that is PP2, a specific inhibitor of the Src signalling, dose dependently reduced the p‐Src⁵²⁷ level but not p‐Src⁴¹⁶ level, while ZEB1 knockdown also down‐regulated the level of p‐Src⁵²⁷ in PC3 and DU‐145 cells. PP2 treatment also significantly reduced the expression of VE‐cadherin, vimentin and CD133 in these prostate cancer cells. Src signalling mediated the effects of ZEB1 on VM formation and gene expression.     

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.     

RNA methylation-mediated LINC01559 suppresses colorectal cancer progression by regulating the miR-106b-5p/PTEN axis

Long noncoding RNAs (lncRNAs) regulate multiple biological effects in cancers. Recently, RNA methylation has been found to modify not only coding RNAs but also some noncoding RNAs. How RNA methylation affects lncRNAs to affect colorectal cancer (CRC) progression remains elusive. The expression of LINC01559 was explored through RNA sequencing, quantitative real-time PCR (qRT-PCR) and in situ hybridization (ISH). The preliminary exploration of its function was performed using Western blotting (WB) and immunohistochemistry (IHC). Functional experiments in vitro and in vivo were conducted to explore the biological functions of LINC01559 in CRC. The LINC01559/miR-106-5p/PTEN axis was verified through fluorescence in situ hybridization (FISH), luciferase assays, and rescue experiments. RIP-sequencing, m6A RNA immunoprecipitation (MeRIP) assays and bioinformatic analysis were conducted to determine the upstream mechanism of LINC01559. The results showed that LINC01559 was downregulated in CRC compared with normal controls. Lower expression of LINC01559 in CRC patients predicted a poor prognosis. In addition, PTEN was found to be positively correlated with LINC01559, and miR-106b-5p could be the link between LINC01559 and PTEN. Then, silencing LINC01559 restored the malignant phenotype of CRC cells, while cotransfection of miR-106b-5p inhibitor neutralized this effect. Mechanistically, we found abundant m6A modification sites on LINC01559. Then, we uncovered these sites as potential targets of METTL3 through experiments in vivo. The results revealed a negative functional regulation of the LINC01559/miR-106b-5p/PTEN axis in CRC progression and explored a new mechanism of METTL3-mediated m6A modification on LINC01559. These results elucidate a novel potential therapeutic target for CRC treatment.     

ZEB1 Enhances Transendothelial Migration and Represses the Epithelial Phenotype of Prostate Cancer Cells

Metastatic colonization involves cancer cell lodgment or adherence in the microvasculature and subsequent migration of those cells across the endothelium into a secondary organ site. To study this process further, we analyzed transendothelial migration of human PC-3 prostate cancer cells in vitro. We isolated a subpopulation of cells, TEM4-18, that crossed an endothelial barrier more efficiently, but surprisingly, were less invasive than parental PC-3 cells in other contexts in vitro. Importantly, TEM4-18 cells were more aggressive than PC-3 cells in a murine metastatic colonization model. Microarray and FACS analysis of these cells showed that the expression of many genes previously associated with leukocyte trafficking and cancer cell extravasation were either unchanged or down-regulated. Instead, TEM4-18 cells exhibited characteristic molecular markers of an epithelial-to-mesenchymal transition (EMT), including frank loss of E-cadherin expression and up-regulation of the E-cadherin repressor ZEB1. Silencing ZEB1 in TEM4-18 cells resulted in increased E-cadherin and reduced transendothelial migration. TEM4-18 cells also express N-cadherin, which was found to be necessary, but not sufficient for increased transendothelial migration. Our results extend the role of EMT in metastasis to transendothelial migration and implicate ZEB1 and N-cadherin in this process in prostate cancer cells.     

LINC00339 promotes gastric cancer progression by elevating DCP1A expression via inhibiting miR-377-3p

Up to date, the mechanism of gastric cancer (GC) development is poorly understood. This study was to demonstrate the effects of LINC00339 on GC progression. Here, we found that LINC00339 was overexpressed expressed in GC tissues and predicted poor outcome. By CCK8, colony formation and Transwell assays, we showed LINC00339 knockdown suppressed GC cell proliferation, migration, and invasion in vitro. Flow cytometry analysis (FACS) indicated that LINC00339 knockdown induced tumor cell apoptosis. Besides, we utilized the xenograft assay and found that LINC00339 depletion led to decreased tumor growth in vivo. Mechanistically, miR-377-3p was found to be inhibited by LINC00339. And LINC00339 suppressed miR-377-3p to upregulate DCP1A, which consequently promoted GC progression. In conclusion, LINC00339 promotes gastric cancer progression by elevating DCP1A expression via inhibiting miR-377-3p.     

Long non‐coding RNA ZEB2‐AS1 promotes the proliferation,metastasis and epithelial mesenchymal transition in triple‐negative breast cancer by epigenetically activating ZEB2

The triple‐negative breast cancer is the most malignant type of breast cancer. Its pathogenesis and prognosis remain poor despite the significant advances in breast cancer diagnosis and therapy. Meanwhile, long noncoding RNAs (LncRNAs) play a pivotal role in the progression of malignant tumors. In this study, we found that LncRNA‐ZEB2‐AS1 was dramatically up‐regulated in our breast cancer specimens and cells (MDA231), especially in metastatic tumor specimens and highly invasive cells, and high lncRNA‐ZEB2‐AS1 expression is associated with clinicopathologic features and short survival of breast cancer patients. LncRNA‐ZEB2‐AS1 promotes the proliferation and metastasis of MDA231 cells in SCID mice. Thus, it is regarded as an oncogene in triple‐negative breast cancer. It is mainly endo‐nuclear and situated near ZEB2, positively regulating ZEB2 expression and activating the epithelial mesenchymal transition via the PI3K/Akt/GSK3β/Zeb2 signaling pathway. Meanwhile, EGF‐induced F‐actin polymerization in MDA231 cells can be suppressed by reducing lncRNA‐ZEB2‐AS1 expression. The migration and invasion of triple‐negative breast cancer can be altered through cytoskeleton rearrangement. In summary, we demonstrated that lncRNA‐ZEB2‐AS1 is an important factor affecting the development of triple‐negative breast cancer and thus a potential oncogene target.