METTL3-mediated m6A modification of CDCA7 mRNA promotes COAD progression.

BACKGROUND: Colon adenocarcinoma (COAD) represents a frequent malignant tumor of the digestive system with high mortality and poor prognosis. As a prevalent internal mRNA modification in eukaryotic cells, N6-methyladenosine (m6A) has been reported to participate in tumor malignancy. This study is designed to explore the role and mechanism of Methyltransferase-like 3 (METTL3) in the progression of COAD. METHODS: In this research, the GEPIA database was applied to analyze the relationship between COAD and cell division cycle-associated protein 7 (CDCA7) or METTL3. Cell viability, cell cycle progression, apoptosis, migration, and invasion were detected by Cell Counting Kit-8 (CCK-8), flow cytometry, transwell assays. The glycolysis level was detected via specific kits. CDCA7, E-cadherin, N-cadherin, and METTL3 protein levels were determined by western blot assay. The biological role of CDCA7 on COAD tumor growth was examined by the xenograft tumor model in vivo. After RBPsuite analysis, the interaction between METTL3 and CDCA7 was verified by methylated RNA immunoprecipitation (MeRIP). RESULTS: METTL3 and CDCA7 were highly expressed in COAD tissues and cells. Furthermore, the silencing of CDCA7 hindered COAD cell proliferation, migration, invasion, glycolysis, EMT, and promoted apoptosis in vitro, as well as retarded tumor growth in vivo. At the molecular level, METTL3 might enhance the stability of CDCA7 mRNA via m6A methylation. CONCLUSION: METTL3 contributes to the malignant progression of COAD cells partly by regulating the stability of CDCA7 mRNA, providing a promising therapeutic target for COAD treatment.

Circ_0007422 Knockdown Inhibits Tumor Property and Immune Escape of Colorectal Cancer by Decreasing PDL1 Expression in a miR-1256-Dependent Manner.

Circular RNAs (circRNAs) are a group of important molecules involved in the progression of various cancers, including colorectal cancer (CRC). Here, we aim to investigate the role and molecular mechanism of circ_0007422 in regulating CRC malignant progression. The expression levels of circ_0007422, miR-1256, and PDL1 were detected by qRT-PCR. Cell viability, proliferation, apoptosis, invasion, and self-replication ability were analyzed by CCK-8, EdU, flow cytometry, transwell, and spheroid formation experiments, respectively. Protein levels were determined by western blotting assay. CRC cells were co-cultured with CD8 + T cells, phytohemagglutinin-stimulated peripheral blood mononuclear cells (PBMCs), or cytokine-induced killer (CIK) cells in vitro, and CD8 + T-cell apoptosis, IFN-γ and TNF-α levels, and survival rate of CRC cells were analyzed to reveal the role of circ_0007422 in antitumor immunity. The relationship between miR-1256 and circ_0007422 or PDL1 was identified by a dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. A xenograft tumor model was established to verify the function of circ_0007422 in tumor growth in vivo. Immunohistochemistry (IHC) assay was used to detect positive expression rates of Ki67, E-cadherin, N-cadherin, and PDL1 expression in primary tumors from CRC cells. Circ_0007422 was upregulated in CRC tissues and cells and its knockdown inhibited proliferation, invasion, self-replication ability, and immune escape and promoted apoptosis of CRC cells. Additionally, circ_0007422 bound to miR-1256, which was identified to target PDL1. MiR-1256 inhibition reversed the effects of circ_0007422 knockdown on the tumor properties and immune escape of CRC cells. Moreover, miR-1256 introduction interacted with PDL1 to suppress proliferation, invasion, self-replication ability, and immune escape and promote apoptosis of CRC cells. Further, circ_0007422 knockdown hampered tumorigenesis of CRC cells in vivo. Circ_0007422 knockdown inhibited tumor property and immune escape of colorectal cancer through the miR-1256/PDL1 pathway, providing a potential novel therapeutic target for CRC.

Role of Long Noncoding RNA Regulator of Reprogramming in Colon Cancer Progression via Epidermal Growth Factor Receptor Signaling.

Background: Long intergenic noncoding RNA regulator of reprogramming (linc-ROR) is a novel long noncoding RNA that exhibits significant effects on cancer progression. This research presented that linc-ROR had a crucial part in promoting biological characteristics associated with worse prognosis in colon cancer. Method: Bioinformatics analysis was performed to predict signaling pathways related to linc-ROR. In addition, western blot, quantitative reverse transcription-polymerase chain reaction, RNA-pulldown, cell proliferation assays, colony formation assays, wound healing assays, and transwell assays were applied to detect the role and regulation of particular molecules. Results: Our results showed that the knockdown of linc-ROR reduced cell invasion, proliferative ability, and migration in colon cancer. Further evaluation verified that downregulating linc-ROR inhibited the activation of epidermal growth factor receptor (EGFR) signaling. In addition, cbl-b, a kind of E3 ubiquitin ligase that increases the degradation of EGFR, was found to be a potential linc-ROR target. Conclusions: Based on our findings, it was presented that linc-ROR served a role as a tumor-promoting factor via repressing the ubiquitination and degradation of EGFR signaling, which indicated that it could be a possible prognostic marker and therapeutic target for colon cancer.

Inhibition of the growth of hepatocellular carcinoma cells through fibroblast growth factor 18 suppressed by miR-139.

Hepatocellular carcinoma (HCC) is a highly malignant tumor and one of the most common causes for human cancer-related deaths. Fibroblast growth factor 18 (FGF18) is overexpressed in many types of cancer, and is associated with cell proliferation, invasion and angiogenesis. miR-139 has recently been reported as a tumor suppressor in various types of cancer and it can regulate many tumor-related genes, however its association with FGF18 expression in HCC has not been reported and thus remains unknown. In the present study, to explore the potential regulation mechanism of miR-139 with FGF18 in HCC, HCC tissues and cell lines were used. The results revealed that FGF18 was highly expressed in HCC tissues and cells, however miR-139 was lowly expressed. FGF18 was demonstrated to be a direct target of miR-139. Furthermore, the suppressive effect of miR-139 on FGF18 and in turn on proliferation, apoptosis, invasion, migration and tumor-induced angiogenesis of HCC cells was investigated. FGF18 was suggested as a prognostic biomarker and therapeutic target in HCC patients and miR-139 may be a promising strategy used in HCC treatment via the suppression of FGF18.

HBx-induced miR-21 suppresses cell apoptosis in hepatocellular carcinoma by targeting interleukin-12.

Hepatitis B virus (HBV) X protein (HBx) plays a key role in the initiation and progression of HBV infection­induced hepatocellular carcinoma (HCC). Oncogenic microRNA-21 (miR-21) can be modulated by HBx protein in HCC. However, critical regulator genes in the pathway of HBx-induced miR-21 in HCC remain unclear. This study aimed to investigate the role of HBx-induced miR-21 in the apoptosis of HCC cells. In the study, interleukin-12 (IL-12) was demonstrated as a direct target of miR-21 by dual­luciferase report assay, and miR-21 was highly expressed in HCC cells (HepG2 and HepG2 2.2.15) compared to L02 cells, but IL-12 was weakly expressed as detected by real-time quantitative PCR (RT-qPCR). Furthermore, miR-21 mimics, inhibitor, HBx-targeted siRNA, and the HBx overexpression vector (pHBx) were used to observe the regulatory effects of HBx-induced miR-21 via IL-12, and cell apoptosis was assessed. The results showed that overexpression of HBx resulted in the inhibition of IL-12. A high level of miR-21 resulted in a significant increase in proliferation and a decrease in IL-12 expression. Inhibition of miR-21 resulted in a significant increase in apoptosis and increased IL-12 expression. The results suggest that HCC cell apoptosis was suppressed at least partially through HBx-induced miR-21 by targeting IL-12.

Inhibition of tumor metastasis in vivo by combination of paclitaxel and hyaluronic acid.

There is little information about the inhibitory effects of hyaluronic acid with paclitaxel on tumor metastasis and ascites formation. The aim of present study is to determine whether the combined administration of hyaluronic acid (HA) with paclitaxel can produce additional or synergistic therapeutic effects in the control of Lewis lung carcinoma (LLC) migration and ascites formation of U14 cervical tumor. The anti-metastasis effect of hyaluronic acid alone, paclitaxel alone and paclitaxel-hyaluronic acid combination were examined in a mouse model bearing LLC cells. i.v. Paclitaxel-hyaluronic acid, in the ratio of 1:3 (w/w), significantly reduced the migration of LLC cells in a synergistic fashion. In the experiment with mice bearing U14 tumor cells, i.p. paclitaxel-hyaluronic acid markedly improved the life span of mice and significantly decreased ascites formation when compared to paclitaxel alone or hyalunonan alone. We also analyzed the serum proteome of mice with lung neoplasm before and after treatment with paclitaxel-hyaluronic acid. Proteomic analysis on LLC mice was studied using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry. The results showed that i.v. administration of hyaluronic acid in combination with paclitaxel had significantly up-regulated the expression of vitamin D(3) binding proteins (DBP), which is a macrophage-stimulating activator. In conclusion, the in vivo anti-metastasis effect was associated with the synergistic therapeutic effects of hyaluronic acid-paclitaxel combination and a significant promotion of immune proteins expression. These results demonstrated that the combination of paclitaxel and hyaluronic acid may be an effective way to inhibit tumor migration.

A fluorescence study of sodium hyaluronate/surfactant interactions in aqueous media.

The interactions between sodium hyaluronate, an anionic polysaccharide, with surfactants (anionic and nonionic) were investigated using pyrene fluorescence measurement methods. The change of micropolarity produced by the interaction was monitored by the measurement of emission intensity ratio between the first and third bands (I1/I3), and the intensity ratio of the excimer and the third vibration monomer band (I(E)/I(M)). Because the hydrophilic heads on the SDS were attracted by the domains formed by the hydroxyl groups of hyaluronate, the I1/I3 ratio was reduced by the addition of hyaluronate at lower than 0.06% of sodium dodecyl sulfate (SDS) concentration. No aggregation was observed between hyaluronate and nonionic surfactants (Tween-80 and Cremophor EL) in the whole concentration range studied. At a higher concentration of surfactant, the I1/I3 ratio of hyaluronate/surfactant was influenced by the addition of saccharide (glucose, lactose, or mannitol). However, the effect of saccharide could be reduced by the addition of salt.