Identification of Distinct Molecular Patterns and a Four-Gene Signature in Colon Cancer Based on Invasion-Related Genes.

BACKGROUND: The pathological stage of colon cancer cannot accurately predict recurrence, and to date, no gene expression characteristics have been demonstrated to be reliable for prognostic stratification in clinical practice, perhaps because colon cancer is a heterogeneous disease. The purpose was to establish a comprehensive molecular classification and prognostic marker for colon cancer based on invasion-related expression profiling. METHODS: From the Gene Expression Omnibus (GEO) database, we collected two microarray datasets of colon cancer samples, and another dataset was obtained from The Cancer Genome Atlas (TCGA). Differentially expressed genes (DEGs) further underwent univariate analysis, least absolute shrinkage, selection operator (LASSO) regression analysis, and multivariate Cox survival analysis to screen prognosis-associated feature genes, which were further verified with test datasets. RESULTS: Two molecular subtypes (C1 and C2) were identified based on invasion-related genes in the colon cancer samples in TCGA training dataset, and C2 had a good prognosis. Moreover, C1 was more sensitive to immunotherapy. A total of 1,514 invasion-related genes, specifically 124 downregulated genes and 1,390 upregulated genes in C1 and C2, were identified as DEGs. A four-gene prognostic signature was identified and validated, and colon cancer patients were stratified into a high-risk group and a low-risk group. Multivariate regression analyses and a nomogram indicated that the four-gene signature developed in this study was an independent predictive factor and had a relatively good predictive capability when adjusting for other clinical factors. CONCLUSION: This research provided novel insights into the mechanisms underlying invasion and offered a novel biomarker of a poor prognosis in colon cancer patients.

LINC01123 Promotes the Progression of Colorectal Cancer via miR-625-5p/LASP1 Axis.

Background: Evidence from previous investigations points to a rising trend in the incidence of colorectal cancer (CRC) worldwide. The mortality resulting from this cancer is high. Unlike nonsmall cell lung cancer for which LINC01123 has been investigated, there are few reports on how this long noncoding RNA (lncRNA) regulates CRC. Materials and Methods: The authors evaluated the expression of LINC01123 in CRC tissues by quantitative real-time polymerase chain reaction. Its impact on cancer cell behavior was analyzed with cell counting kit-8 (CCK-8), colony formation, and Transwell invasion assays. To establish the mechanisms of LINC01123 in CRC they carried out RIP and luciferase reporter assays. Results: The results show that LINC01123 expression is abnormally elevated in CRC tissues and cell lines. High LINC01123 expression closely correlates with poor prognosis, advanced TNM stage, and lymph-node metastasis. The authors also show that knockdown of LINC01123 inhibits proliferation and invasion in CRC cells. In mechanism, it is revealed that LINC01123 may function as competitive endogenous RNA (ceRNA) against miR-625-5p to promote LIM and SH3 protein 1 (LASP1) expression. Conclusions: The data indicate that high LINC01123 exerts its oncogenic roles by regulating the miR-625-5p/LASP1 axis in CRC progression.

Up-regulation of FAT4 enhances the chemosensitivity of colorectal cancer cells treated by 5-FU.

BACKGROUND: Colorectal cancer (CRC) has high mortality, and 5-fluorouracil (5-FU) is a common clinical chemotherapeutic drug. The current study aimed to investigate the role of FAT4 in chemosensitivity of CRC cells treated by 5-FU. METHODS: The immunohistochemistry and qRT-PCR was conducted to measure the FAT4 expression in CRC and adjacent tissues. The FAT4 expression was determined by qRT-PCR and Western blot, comparison of FAT expression between normal and several CRC cell lines was then made, so as to identify cell lines with the highest (LS174T) and the lowest (SW-620) expressions of FAT4. The effects of 5-FU stimulation at various doses on cell viability were determined by CCK-8, and the level of FAT4 was also measured. After FAT4 knockdown in LS174T or FAT4 overexpression in SW-620 with or without pretreatment of 5-FU (30 µg/mL), cell growth, colony formation, cell migration and invasion, angiogenesis were determined by flow cytometry, wound-healing, transwell assay and tube formation assay, respectively. The expression levels of epithelial-mesenchymal transition (EMT) markers were detected by qRT-PCR and Western blot. RESULTS: FAT4 was down-regulated in CRC tissues and cells, cell viability of CRC cells was decreased. The level of FAT4 was increased with the increase of 5-FU concentrations. Moreover, 5-FU stimulation increased FAT4 expression, and reduced cell proliferation, migration, invasion, angiogenesis and cell EMT process, furthermore, such effects of 5-FU stimulation could be enhanced by FAT4 overexpression but reversed by FAT4 knockdown. CONCLUSIONS: Upregulation of FAT4 could increase the sensitivity of CRC cells to 5-FU.

Knockdown of KLK12 inhibits viability and induces apoptosis in human colorectal cancer HT-29 cell line.

Kallikrein­related peptidase 12 (KLK12) is overexpressed in cancer tissues including gastric, breast and prostate cancer. However, the role of KLK12 in colorectal cancer is not fully understood. In the present study, the level of KLK12 was determined by performing reverse transcription­polymerase chain reaction (RT­qPCR) in colorectal cancer tissues and cell lines. Lipofectamine® 2000 was used to transfect HT­29 cells to overexpress and knockdown KLK12. Cell viability, migration, invasion and apoptosis were detected by MTT, wound healing, Transwell and flow cytometry assays, respectively. The mRNA and protein expression levels of EMT­associated proteins, apoptosis­associated proteins, phosphorylated adenosine monophosphate­activated protein kinase (p­AMPK) and phosphorylated mammalian target of rapamycin (p­mTOR) were determined by RT­qPCR and western blot analysis. It was identified that the KLK12 mRNA levels were increased significantly in colorectal cancer tissues and cell lines. KLK12 small interfering RNA inhibited cell viability, migration and invasion. Furthermore, epithelial­mesenchymal transition (EMT)­associated proteins were altered by siKLK12. Cell apoptosis was induced by KLK12 downregulation, which was demonstrated by the changes in apoptosis­associated proteins; however, KLK12 overexpression produced the opposite effect. SiKLK12 enhanced the expression of p­AMPK and suppressed the expression of p­mTOR, while KLK12 overexpression had the opposite effect. Promotion of KLK12 overexpression­induced cell viability was reversed by 5­aminoimidazole­4­carboxamide ribonucleotide, an activator of the AMPK signaling pathway, and rapamycin, a specific inhibitor of the mTOR signaling pathway. Taken together, the results of the present study indicated that KLK12 was overexpressed in colorectal cancer and may regulate cell behavior, potentially via the AMPK and mTOR pathways.

Effect of STC2 gene silencing on colorectal cancer cells.

Stanniocalcin 2 (STC2), a secretory glycoprotein hormone, regulates many biological processes including cell proliferation, apoptosis, tumorigenesis and atherosclerosis. However, the effect of STC2 on proliferation, migration and epithelial­mesenchymal transition (EMT) progression in human colorectal cancer (CRC) cells remains poorly understood. The expression level of STC2 was determined by quantitative real­time polymerase chain reaction (qPCR) and western blot analysis. Cell Counting Kit­8 (CCK­8) was used to detect the viability of SW480 cells. The invasion and migration of cells were identified by wound healing and Transwell assays. The mRNA and protein expression levels of ß­catenin, matrix metalloproteinase (MMP)­2, MMP­9, E­cadherin and vimentin were assessed by qPCR and western blot analysis. In the present study, it was demonstrated that STC2 was highly expressed in the CRC cell lines. After silencing of STC2, the cell viability, migration and invasion were significantly reduced. Silencing of STC2 in the CRC Sw480 cells increased the expression of E­cadherin and decreased the expression of vimentin, MMP­2 and MMP­9, compared to those in the normal and empty vector group. Furthermore, the expression of ß­catenin in the STC2 gene silenced group was suppressed, and the expression of ß­catenin was reversed by Wnt activator, SB216763. These results demonstrated that STC2 participates in the development and progression of CRC by promoting CRC cell proliferation, survival and migration and activating the Wnt/ß­catenin signaling pathway.