The miR-532-E2F1 feedback loop contributes to gastric cancer progression.

Gastric cancer (GC) ranks fourth in incidence and mortality worldwide, ascertaining the pathogenesis of GC is crucial for its treatment. E2F1, which regulates the transcription of genes encoding proteins involved in DNA repair, DNA replication, mitosis and survival of cancer patients, functions as a key regulator in GC progression. However, the underneath mechanism of these processes is not fully elucidated. Here, TCGA database analysis, microarray immunohistochemical technique and western blot showed that E2F1 was highly upregulated in clinical GC tissues and correlated with tumor malignancy. In vitro and in vivo assays confirmed the oncogenic function of E2F1. MiR-532 was decreased and negatively correlated with E2F1 in GC tissues. MiR-532 directly targeted and inhibited E2F1 expression, leading to the decrease of ASK1 and elevation of TXNIP, and affected proliferation, cell cycle, apoptosis and DNA damage in vitro and tumor growth in vivo. Moreover, E2F1 serves as a transcriptional repressor to suppress miR-532 expression and a double-negative feedback loop was formed between them. This study demonstrates the significant roles of the E2F1-miR-532 double-negative feedback loop in GC progression and may represent a potential target for GC therapy.

MicroRNA-656-3p inhibits colorectal cancer cell migration, invasion, and chemo-resistance by targeting sphingosine-1-phosphate phosphatase 1.

Colorectal cancer presents high rates of recurrence and metastasis, and the occurrence and progression and mechanism of its invasion and metastasis are not fully understood. The expression of miR-656-3p in patient samples and 10 cell lines were measured. Bioinformatic databases were used to predict miRNAs. Protein expressions were examined using Western blot. Transwell assay was used to measure cell migration and invasion. Transplanted tumor model in nude mice was established. Removal of the miR-656-3p by specific knocking-down of this gene promoted the chemo-resistance of colorectal cancer cells. Critically, we identified sphingosine-1-phosphate phosphatase 1 (SGPP1) as a downsteam target of the miR-656-3p, which we first obtained from 199 potential target genes from Targetscan, 200 genes from miRDB and 200 genes from DIANA, respectively. Then, we identified the interaction between SGPP1 and the miR-656-3p on 3' UTR of SGPP1 gene. Knockdown of SGPP1 greatly suppressed the tumor growth in vivo and epithelial mesenchymal transition process. miR-656-3p could regulate cell proliferation and chemoresistance in the colorectal cancer that associate to downstream target with SGPP1. Along with its downstream molecule, we would like to predict that the SGPP1 associated miR-656-3p could be used to develop early for early diagnostics for CRC oncogenesis.

A novel role of miR-326 in colorectal carcinoma by regulating E2F1 expression.

PURPOSE: Colorectal carcinoma (CRC) ranks third in incidence but second in mortality worldwide, ascertaining the pathogenesis of CRC is crucial for its treatment. Accumulating studies have shown that E2F1 is a key regulator in CRC progression, which regulates the transcription of genes engaged in DNA replication, mitosis and survival of cancer patients, however, the mechanism of these processes is not fully elucidated. METHODS: Here, we determined E2F1 expression in clinical CRC specimens by TCGA database analysis, Microarray immunohistochemical technique and Western blot, respectively. The expression of E2F1 was elevated in CRC tumor tissues, and the patients' total survival time was associated with the level of E2F1. Then the prediction software and meta-analysis were used to predict the miRNAs targeting E2F1. RT-qPCR, TCGA analysis and in situ hybridization experiments were utilized to determine the decreased miR-326 expression in CRC tumor tissues. Luciferase and Western blot assays determined that miR-326 directly targeted E2F1 in CRC cells. Next, CCK8, flow cytometry, Transwell and wound healing assays were used to determine the biological function of miR-326-E2F1 axis in vitro. RESULTS: miR-326 overexpression significantly inhibited the viability, invasion and migration and promoted the apoptosis of CRC cells, but overexpression of both E2F1 and miR-326 in turn increased cell viability, invasion and migration and decreased cell apoptosis. CONCLUSIONS: This study demonstrates the significant roles of miR-326-E2F1 in CRC progression and may represent a potential target for CRC therapy.

The Critical Role of the miR-21-MEG2 Axis in Colorectal Cancer.

Protein tyrosine phosphatase MEG2 (MEG2/PTPN9), a classic tyrosine-specific protein tyrosine phosphatase (PTP), is involved in the progression of liver, breast, and gastric cancers. However, the function and regulation of MEG2 in colorectal cancer (CRC) still remain unclear. In this study, we investigated the expression of MEG2 in CRC and found that MEG2 was downregulated in human CRC tissues compared to normal corresponding tissues. Moreover, in vivo and in vitro assays revealed that MEG2 plays a vital role in CRC cell proliferation, invasion, and apoptosis. In addition, mechanism analysis validated miR-21 as a direct regulator of MEG2, and miR-21 plays a critical role in promoting proliferation, invasion, and suppression of apoptosis in CRC by targeting MEG2. Taken together, this study demonstrates the significant role for miR-21 in regulating MEG2 in CRC and may represent a potential target for CRC therapy.

Lentivirus-mediated reversion-inducing cysteine-rich protein with Kazal motifs gene transfer suppresses pancreatic cancer invasion in vitro.

OBJECTIVE: To investigate the biological functions of reversion-inducing cysteine-rich protein with Kazal motifs (RECK) over-expression in pancreatic cancer cell line Panc-1. METHODS: This study was carried out in the Department of General Surgery, First Affiliated Hospital of Soochow University, Suzhou, Jiangsu, China from January to September 2009. The design of the study was to construct a recombinant lentivirus carrying the RECK gene (LV-RECK) to be used to infect Panc-1 cells, and investigate the biological functions of RECK in pancreatic cancer in vitro. The RECK expression was measured by real-time polymerase chain reaction (PCR) and Western blotting. Cell proliferation and apoptosis were examined by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry. The ability of invasion was detected using Transwell chambers. The expression and activity of matrix metalloproteinase (MMP)-2 and matrix metalloproteinase-9 (MMP-9) was measured by real-time PCR, Western blotting, and gelatin zymography. RESULTS: We successfully constructed LV-RECK and proved it can over-express the RECK gene in Panc-1 cells. The RECK over-expression potently suppressed the invasion ability of Panc-1 cells in vitro without affecting cell proliferation and apoptosis. The RECK over-expression potently inhibited the secretion and activity of MMP-2, and the secretion of MMP-9 without affecting the messenger ribonucleic acid, and protein expression of MMP-2 and MMP-9. CONCLUSION: The RECK gene exerts repressive effects on the invasion ability of the pancreatic cancer cell line Panc-1, and it may be a novel therapeutic target for pancreatic cancer.