Ability Of Colorectal Cancer Cells Research Articles

NIFK, a Potential Prognostic and Immunological Biomarker in Pan-- Cancer Analysis, Significantly Regulates Proliferation and Metastasis of Colorectal Cancer.

As a binding protein of Ki67, NIFK plays an important role in the mitosis of cells and is closely related to the progression of specific types of tumors. However, there is still a lack of systematic analysis of NIFK in pan-cancer and insufficient research to explore its role in human tumors. We systematically evaluated the pan-cancer expression and mutation of NIFK in human cancers using data from The Cancer Genome Atlas (TCGA) through large-scale bioinformatics analysis. In addition, we explored the pan-cancer immunological characteristics of NIFK, especially in colorectal adenocarcinoma (COAD). Furthermore, we used single-cell sequencing to analyze the expression of NIFK in different cells of COAD tissues and performed GO, KEGG, and gene set enrichment analysis of NIFK in COAD. Lastly, we evaluated the effects of NIFK knockdown on the colorectal cancer cell lines in in vitro experiment. We found that NIFK was overexpressed in almost all types of tumors and showed significant prognostic efficacy. Additionally, correlations between NIFK and specific immune features, such as immune cell infiltration, immune checkpoint genes, TMB, and MSI, suggest that NIFK may be used to guide immunotherapy. Subsequently, it was found that the expression of NIFK was significantly upregulated in tumor cells through single-cell sequencing analysis, and the NIFK gene was closely associated with tumor progression and immune therapy response. Finally, we further elucidated the role of NIFK in colorectal cancer and found that downregulation of NIFK expression could inhibit the proliferation, migration, and invasion ability of colorectal cancer cells. The results of this study demonstrated that NIFK, as a member of the pan-cancer genes, will serve as a biomarker and a potential therapeutic target for a range of cancer types, providing new insight into precision medicine.

LncRNA MEG3 inhibits the proliferation and migration abilities of colorectal cancer cells by competitively suppressing MiR-31 and reducing the binding of MiR-31 to target gene SFRP1.

To explore the potential mechanism of long-chain non-coding ribonucleic acid (lncRNA) maternal expression gene 3 (MEG3) in colorectal cancer (CRC). The relationship between MEG3 and miR-31 was detected by dual-luciferase assay. Quantitative polymerase chain reaction was utilized to determine the expression of MEG3 in CRC cell lines. Cell Counting Kit-8 assay was performed to detect cell proliferation. Transwell, cell scratch wound assay, and monoclonal proliferation assay were used to detect the proliferation, migration, and invasion of cells. In addition, cell motility was evaluated by detecting the expression of cellular pseudopodia protein α-actinin via immunofluorescence assay, and cell proliferation and motility were judged by determining the expressions of Ki-67, MMP2 and MMP9 via Western blotting. The effect of MEG3 and miR-31 on the development of colorectal cancer was verified by nude mouse tumor-bearing assay and HE staining. Transient transfection with MEG3 overexpression plasmid revealed that MEG3 inhibited the proliferation and motility of cells. The results of dual-luciferase assay showed that MEG3 could specifically inhibit the expression of miR-31, which inhibits the development of colorectal cancer. Transwell, cell scratch wound assay, and monoclonal proliferation experiment showed that miR-31 enhanced cell proliferation, migration and invasion. MEG3 overexpression plasmid was capable of reversing the proliferation and motility of CRC cells enhanced by miR-31. MEG3 can inhibit the proliferation and motility of CRC cells by competitively suppressing the binding of miR-31 to the target gene SFRP1, thus playing an inhibitory role in the pathogenesis of CRC.

Serine and arginine rich splicing factor1‑regulated microtubule interacting and trafficking domain containing1 affects colorectal cancer progression and ferroptosis.

As the third most common type of cancer globally, colorectal cancer (CC) is a prevalent digestive malignancy, with the second highest mortality rate among all types of cancer. It has been reported that microtubule interacting and trafficking domain containing 1 (MITD1) serves a pivotal role in the initiation and progression of diverse types of tumors. Nevertheless, the underlying mechanism of MITD1 in CC has not been previously investigated. The ENCORI and GEPIA databases were used to investigate the expression levels of MITD1 in patients with CC. Immunohistochemistry was used to detect the expression of MITD1 in cancer tissues obtained from patients with CC, while its mRNA and protein expression levels in CC cell lines were determined by reverse transcription-quantitative PCR and western blot analysis, respectively. Subsequently, MITD1 was knocked down in CC cells using an interference plasmid and Cell Counting Kit 8, colony formation, as well as EdU assays were performed to assess cell proliferation. Concurrently, wound healing and Transwell assays were performed to evaluate the migration and invasion abilities of CC cells. Lipid reactive oxygen species (ROS) levels were determined by BODIPY 581/591 C11 staining. In addition, the levels of oxidative stress markers and those of total iron were measured using the corresponding kits. Furthermore, the association between serine and arginine rich splicing factor 1 (SRSF1) and MITD1 was verified by RNA immunoprecipitation and actinomycin D experiments. Finally, to further uncover the mechanism of MITD1, SRSF1 was overexpressed and MITD1 was silenced in CC cells. The results demonstrated that the expression of MITD1 was abnormally elevated in CC tissues and CC cell lines. MITD1 silencing distinctly diminished CC cell viability, increased CC cell ferroptosis and attenuated their invasion and migration abilities. In addition, MITD1 knockdown significantly increased the expression of lipid ROS and total iron levels in CC cells. Additionally, the results showed that SRSF1 could stabilize MITD1 mRNA expression in CC cells. Finally, it was revealed that SRSF1 could regulate MITD1 and affect the progression of CC and ferroptosis via p53/solute carrier family 7 member 11 (SLC7A11)/glutathione peroxidase 4 (GPX4) signaling. Overall, the results of the current study indicated that SRSF1-regulated MITD1 could affect CC progression and ferroptosis, probably via the p53/SLC7A11/GPX4 signaling pathway.

SNAI2/FTH1P3/miR-218-5p Positive Feedback Loop Promotes Colorectal Cancer Metastasis.

Colorectal cancer (CRC) is a type of intestinal cancer that causes more than 600,000 deaths every year. Overcoming the problems of metastasis requires detailed studies to reveal the potential molecular mechanisms. This study aims to reveal the molecular mechanism of CRC metastasis involving non-coding RNA regulation. The expression profile of FTH1P3 was analyzed based on the data of TCGA-COAD patient cohorts. Q-PCR analysis was performed to validate the expression of FTH1P3 in colorectal cancer cells. JASPR was used to screen transcription factors of FTH1P3. q-ChIP analysis was used to validate the target between FTH1P3 and transcription factor. Scratch assay and transwell assay were used to evaluate the migration and invasion ability of colorectal cancer cells. FTH1P3 is highly expressed in CRC patient cohort. FTH1P3 induced migration and invasion of SW480 cell through regulating epithelial-mesenchymal transition (EMT). In addition, FTH1P3 is a direct target of SNAI2. SNAI2 promotes the expression of FTH1P3. Both FTH1P3 and SNAI2 were directly targeted and repressed by miR-218-5p. Interestingly, ectopic FTH1P3 caused a decreased miR-218-5p level and an elevated nucleic SNAI2 protein expression level. Of note, only ectopic SNAI2 protein resulted in a repressed miR-218-5p and an increased FTH1P3, whereas SNAI2 3'UTR failed to affect the expression of miR-218-5p and FTH1P3. SNAI2 transcriptionally activates FTH1P3 expression. Both SNAI2 and FTH1P3 are targets of miR-218-5p. SNAI2/FTH1P3/miR-218-5p form a positive feedback loop in the regulation of CRC metastasis.

SNHG25 promotes colorectal cancer metastasis by regulating MMP2.

LncRNA has been shown to play an important role in tumors, but the functions of most lncRNAs in colorectal cancer is not clear. By analyzing the transcriptome data of tumor tissues and adjacent tissues, we identified the lncRNA profiles that were abnormally expressed in colorectal cancer and selected the abnormally highly expressed lncRNA SNHG25 for further study. The functional assays showed that after knocking down SNHG25, the metastatic ability of colorectal cancer cells was significantly reduced. Western blot and immunofluorescence assays showed that inhibiting SNHG25 would affect the expression of Vimentin and E-Cadherin. In terms of mechanism, the results of RNA pull down assays, RNA immunoprecipitation (RIP) assays and dual luciferase reporter assays showed that SNHG25 could promote MMP2 expression by adsorbing miR-296-3p. In addition, chromatin immunoprecipitation (ChIP) assays and promoter luciferase reporter assays revealed that PAX5 could activate the transcription of SNHG25 in colorectal cancer cells. Our study proved that SNHG25 acts a pro-metastasis role in colorectal cancer, enriching the theory of the functions of lncRNA in colorectal cancer.

Circular RNA circWHSC1 facilitates colorectal cancer cell proliferation by targeting miR-130a-5p/zeb1 signaling in vitro and in vivo

Colorectal cancer is a prevalent cancer globally and has become a threaten of human health. Recently, circular RNAs (circRNAs) have been widely studied in the cancer area, and the function of circular RNA circWHSC1 has been identified in several cancers. However, the role of circWHSC1 in colorectal cancer remains elusive. In this study, we were interested in the effects of circWHSC1 on colorectal cancer progression. We found that level of circWHSC1 was elevated in colorectal cancer cells compared with normal colon epithelial cells. FISH assay further confirmed that circWHSC1 was mainly localized in cytoplasm. CircWHSC1 depletion repressed the viability of colorectal cancer cells. The colony formation number and Edu-positive colorectal cancer cells were inhibited by the depletion of circWHSC1, respectively. The knockdown of circWHSC1 promoted the apoptosis of colorectal cancer cells. The tumor growth of colorectal cancer cells in nude mice was attenuated by circWHSC1 silencing. Meanwhile, the invasion and migration ability of colorectal cancer cells was suppressed by circWHSC1 depletion. Mechanically, circWHSC1 targets miR-130a-5p to promote zeb1 expression in colorectal cancer cell. The depletion of circWHSC1 remarkably reduced the cell viability and Edu-positive colorectal cancer cells, and the miR-130a-5p inhibitor or zeb1 overexpression could restore the phenotypes. Furthermore, the tumor growth of colorectal cancer cells in nude mice was attenuated by circWHSC1 knockdown, while miR-130a-5p depletion or zeb1 overexpression reversed the effect in the model. Therefore, we concluded that Circular RNA circWHSC1 facilitated colorectal cancer cell proliferation by targeting miR-130a-5p/zeb1 signaling in vitro and in vivo.

Chalcone derivatives as novel, potent and selective inhibitors against human Notum: Structure–activity relationships and biological evaluations

Human Notum (hNotum) inhibitors could be used for treating Wnt signalling-associated diseases including colorectal cancer. Herein, two series of chalcone derivatives were designed and synthesized aiming to find selective and potent hNotum inhibitors. Structure–activity relationship (SAR) studies showed that 2-methoxyl and 5-bromine substitutions on A-ring significantly enhanced anti-hNotum effect, while 4′-ethoxyl and 3′-alkyl substitutions on B-ring were beneficial for hNotum inhibition. Among all tested chalcones, B11 displayed the most potent anti-Notum effect (IC50 = 3.6 nmol/L), good selectivity, excellent chemical stability and suitable metabolic stability. Further investigations showed that B11 acted as a competitive inhibitor of hNotum, while this agent (5 µmol/L) significantly weaken the migration abilities of colorectal cancer cells. Collectively, this study deciphers the SARs of chalcones as hNotum inhibitors and reports a novel and potent hNotum inhibitor with the anti-migration effect on colorectal cancer cells, which offers a promising lead compound to develop novel anti-cancer agents.

Construction of TRAF6 ubiquitin site 331 mutant colorectal cancer cell stable line and its effect on biological behavior of colorectal cancer cells

Objective: To investigate the effect of ubiquitin mutation at position 331 of tumor necrosis factor receptor related factor 6 (TRAF6) on the biological characteristics of colorectal cancer cells and its mechanism. Methods: lentivirus wild type (pCDH-3×FLAG-TRAF6) and mutation (pCDH-3×FLAG-TRAF6-331mut) of TRAF6 gene expression plasmid with green fluorescent protein tag were used to infect colorectal cancer cells SW480 and HCT116, respectively. The infection was observed by fluorescence microscope, and the expressions of TRAF6 and TRAF6-331mut in cells was detected by western blot. Cell counting kit-8 (CCK-8) and plate cloning test were used to detect the proliferation ability of colorectal cancer cells in TRAF6 group and TRAF6-331mut group, cell scratch test to detect cell migration, Transwell chamber test to detect cell migration and invasion, immunoprecipitation to detect the ubiquitination of TRAF6 and TRAF6-331mut with ubiquitinof lysine binding sites K48 and K63. Western blot was used to detect the effects of TRAF6 and TRAF6-331mut over expression on the nuclear factor kappa-B (NF-κB) and mitogen activated protein kinase mitogen-activated protein kinase (MAPK)/activating protein-1(AP-1) signal pathway. Results: The successful infection of colorectal cancer cells was observed under fluorescence microscope. Western blot detection showed that TRAF6 and TRAF6-331mut were successfully expressed in colorectal cancer cells. The results of CCK-8 assay showed that on the fourth day, the absorbance values of HCT116 and SW480 cells in TRAF6-331mut group were 1.89±0.39 and 1.88±0.24 respectively, which were lower than those in TRAF6 group (2.09±0.12 and 2.17±0.45, P=0.036 and P=0.011, respectively). The results of plate colony formation assay showed that the number of clones of HCT116 and SW480 cells in TRAF6-331mut group was 120±14 and 85±14 respectively, which was lower than those in TRAF6 group (190±21 and 125±13, P=0.001 and P=0.002, respectively). The results of cell scratch test showed that after 48 hours, the percentage of wound healing distance of HCT116 and SW480 cells in TRAF6-331mut group was (31±12)% and (33±14)%, respectively, which was lower than those in TRAF6 group [(43±13)% and (43±7)%, P=0.005 and 0.009, respectively]. The results of Transwell migration assay showed that the migration numbers of HCT116 and SW480 cells in TRAF6-331mut group were significantly lower than those in TRAF6 group (P<0.001 and P<0.002, respectively). The results of Transwell invasion assay showed that the number of membrane penetration of HCT116 and SW480 cells in TRAF6-331mut group was significantly lower than those in TRAF6 group (P=0.008 and P=0.009, respectively). The results of immunoprecipitation detection showed that the ubiquitin protein of K48 chain pulled by TRAF6-331mut was lower than that of wild type TRAF6 in 293T cells co-transfected with K48 (0.57±0.19), and the ubiquitin protein of K63 chain pulled down by TRAF6-331mut in 293T cells co-transfected with K63 was lower than that of wild type TRAF6 (0.89±0.08, P<0.001). Western blot assay showed that the protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-HCT116 cells were 0.63±0.08, 0.42±0.08 and 0.60±0.07 respectively, which were lower than those in TRAF6-HCT116 cells (P=0.002, P<0.001 and P<0.001, respectively). The expression level of AP-1 protein in TRAF6-HCT116 cells was 0.89±0.06, compared with that in TRAF6-HCT116 cells. The difference was not statistically significant (P>0.05). The protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-SW480 cells were 0.50±0.06, 0.51±0.04, 0.48±0.02, respectively, which were lower than those in TRAF6-SW480 cells (all P<0.001). There was no significant difference in AP-1 protein expression between TRAF6-331mut-SW480 cells and TRAF6-SW480 cells. Conclusion: The ubiquitin site mutation of TRAF6 gene at 331 may prevent the binding of TRAF6 and ubiquitin lysine sites K48 and K63, and then affect the expressions of proteins related to downstream NF-κB and MAPK/AP-1 signal pathways, and inhibit the proliferation, migration and invasion of colorectal cancer cells.

Ring finger protein 128 promotes, rather than inhibits, colorectal cancer progression by regulating the Hippo signaling pathway.

Colorectal cancer is a common malignancy of the gastrointestinal tract, and its incidence and mortality rates have increased in recent years. RNF128 is an E3 ubiquitin ligase that plays an important role as a suppressor gene or oncogene in various cancers, but its mechanism in colorectal cancer is not yet clear. The aim of this study was to investigate the role and mechanism of RNF128 in colorectal cancer. The expression of RNF128 in colorectal cancer tissues was assessed by immunohistochemistry and western blotting. The proliferation ability of colorectal cancer cells was measured by colony formation assay and CCK-8 assay, the migration and invasion ability of colorectal cancer cells was measured by wound healing assay and transwell assay, and the protein expression levels of the Hippo signaling pathway and its target gene were examined by western blotting. Immunoprecipitation was used to assess the interaction of RNF128 with MST. In vivo, a xenograft tumor model was used to detect the effect of RNF128 on tumor growth. At the tissue level, the expression level of RNF128 was significantly higher in colorectal cancer tissues than in adjacent normal tissues. In LoVo cells and HCT116 cells, the proliferation, migration and invasion abilities were significantly reduced with RNF128 knockdown. At the protein level, knockdown of RNF128 resulted in significant activation of the Hippo signaling pathway. In vivo experiments, the volume and weight of xenograft tumors in nude mice were significantly decreased compared with those in the normal control group with RNF128 knockdown. RNF128 promotes the malignant behaviors of colorectal cancer cells by inhibiting the Hippo signaling pathway, which may provide a new target for colorectal cancer prevention and treatment.

Effects of Salidroside Combined with Paclitaxel on Proliferation, Migration, and Epithelial Mesenchyme of Colorectal Cancer Cells.

Colorectal cancer (CRC) is a multifactorial disease and one of the most common malignancies worldwide. Salidroside (Sal) is a plant with a wide range of pharmacological effects and plays an important role in the treatment of many diseases, and is considered a new hope for the treatment of tumors. The purpose of this study was to investigate the effect of the combination of Sal and paclitaxel (Pac) on colorectal cancer cells and its mechanism of action. The effects of different mass concentrations of Sal, Pac, and the combination intervened in the cells for 48 h were examined using the CCK8 method. The inhibition rate was obtained, and the optimal concentration of the respective drug group was screened. The proliferative capacity of the respective group was obtained. Subsequently, the results of apoptosis, cloning, migration, invasion, and angiogenesis were observed through cell morphological analysis (shape observation and Hoechst staining), colony formation assay, cell scratching assay, Transwell, angiogenesis assay, and protein immunoblotting (Western blotting) to detect the expression of epithelial-mesenchymal transition (EMT)-associated proteins and PI3K pathway-associated proteins. Different concentrations of Sal, Pac, and the combined application had significant effects in inhibiting cells in a concentration-dependent manner. Compared with the control group, the Sal group, the Pac group, and the combination group significantly inhibited the clonal number, migration, invasion, and tube-forming ability of colorectal cancer cells. Besides, the combined application had a better effect than the Sal and Pac groups. The apoptosis level was up-regulated in all drug groups, and the up-regulation was more significant in the combination group. The expression of E-cad protein was up-regulated, the expression of N-cad and Vim protein was down-regulated, and the expression of PI3K and AKT phosphorylation was down-regulated in the respective group, and the difference was more significant in the combination group compared with the group of individual drugs. The combined application of Sal and Pac significantly can decrease the survival rate of colorectal cancer cells, and the mechanism may be correlated with the blocking of the PI3K/AKT pathway, thus inhibiting EMT.

Exploring the oncogenic roles of LINC00857 in pan-cancer.

Although aberrant LINC00857 expression may play a key role in oncogenesis, no research has analyzed the pan-cancer oncogenic roles of LINC00857, particularly in tumor immunology. Here, we integrated data from several databases to analyze the characteristics of LINC00857 in pan-cancer. We found that LINC00857 was overexpressed and correlated with a poor prognosis in a variety of cancers. Furthermore, high-expression of LINC00857 was negatively associated with immune cell infiltration and immune checkpoint gene expression. Notably, LINC00857 expression was negatively related to microsatellite instability and tumor mutation burden in colorectal cancer, implying poor reaction to immunotherapy when LINC00857 was highly expressed. Targeting LINC00857 could dramatically impair the proliferative ability of colorectal cancer cells. After RNA-sequencing in HCT116 cells, gene set enrichment analysis showed that LINC00857 may accelerate cancer progression by inhibiting the ferroptosis pathway and promoting glycolipid metabolism in colorectal cancer. Screening by weighted gene co-expression network analysis determined PIWIL4 as a target of LINC00857, which also performed an immunosuppressive role in colorectal cancer. Based on the structure of PIWIL4, a number of small molecule drugs were screened out by virtual screening and sensitivity analysis. In summary, LINC00857 expression was closely correlated with an immunosuppressive microenvironment and may be a novel diagnostic and prognostic biomarker for diverse cancers. The LINC00857/PIWIL4 axis may be predictive biomarkers for immunotherapy and valuable molecular targets for malignant tumors.

Alisol A attenuates malignant phenotypes of colorectal cancer cells by inactivating PI3K/Akt signaling.

Despite the advancement in the diagnosis and therapeutic strategies for colorectal cancer, the outcomes of patients with colorectal cancer remain unsatisfactory. Alisol A is a natural constituent of Alismatis rhizoma (zexie) and has demonstrated anti-cancer properties; however, the function of Alisol A in colorectal cancer is still unknown. In the present study, the effect of Alisol A on colorectal cancer progression was investigated. MTT and colony formation assays showed that treatment with Alisol A repressed colorectal cancer cell proliferation in a dose-dependent manner. Similarly, western blot analysis demonstrated that Alisol A upregulated E-cadherin protein expression levels, but downregulated N-cadherin and Vimentin protein expression levels in colorectal cancer cells. In addition, the number of cells in G0/G1 phase was enhanced, while that of S phase was reduced in Alisol A-treated colorectal cancer cells. Apoptosis and pyroptosis of colorectal cancer cells were stimulated following treatment with Alisol A. Alisol A suppressed the migration ability of colorectal cancer cells in a dose-dependent manner. Moreover, Alisol A increased the chemotherapeutic sensitivity of colorectal cancer cells to cisplatin. Mechanically, western blot analysis confirmed that Alisol A repressed the phosphorylation levels of PI3K, Akt and mTOR in colorectal cancer cells. The Akt activator, SC79 reversed the effect of Alisol A on colorectal cancer cell proliferation and apoptosis. In conclusion, Alisol A induced an inhibitory effect on colorectal cancer progression by inactivating PI3K/Akt signaling.

Long non-coding RNA LINC01123 promotes cell proliferation, migration and invasion via interacting with SRSF7 in colorectal cancer

BackgroundIncreasing evidences demonstrated that long non-coding RNAs (lncRNAs) participates in the occurrence and development of cancer. In this study, we explored the function and molecular mechanism of LINC01123 in colorectal cancer progression. MethodsAnalyze the expression level of LINC01123 in gastrointestinal tumors via TCGA database. Colorectal tumor tissues and normal tissues were collected to detect the expression of LINC01123 by RT-qPCR. CCK-8 assay, clone formation assay, transwell assay, and wound healing assays were used to explore the effects of LINC01123 on the proliferation, invasion and migration of colorectal cancer cells. Coomassie blue staining, RNA pull-down and mass spectrometry were used to screen the protein interacted with LINC01123. Xenograft model was used to explore the effect of LINC01123 in vivo. ResultsLINC01123 was up-regulated in colorectal cancer tumor tissues. The proliferation, invasion and migration ability of colorectal cancer cells were decreased significantly after LINC01123 knockdown, and it may inhibit its expression by interacted with SRSF7, thereby promoting colorectal cancer progression. ConclusionsLINC01123 can promote the proliferation, invasion and migration of colorectal cancer cells by regulating SRSF7, suggesting that it may be an important regulator of colorectal cancer progression.

Dehydroevodiamine inhibits lung metastasis by suppressing survival and metastatic abilities of colorectal cancer cells

BackgroundDespite the rising 5-year survival rate of colorectal cancer (CRC) patients, the survival rate decreases as the stage progress, and a low survival rate is highly associated with metastasis. PurposeThe purpose of our study is to investigate the effect of dehydroevodiamine (DHE) on the lung metastasis of CRC and the proliferation of CRC cells. Study designCell death was confirmed after DHE treatment on several CRC cell lines. The mechanism of cell cytotoxicity was found using flow cytometry. After that, the expression of the proteins or mRNAs related to the cell cytotoxicity was confirmed. Also, anti-metastatic ability of DHE in CRC cells was measured by checking the expression of Epithelial to Mesenchymal Transition (EMT) markers. Lung metastasis mouse model was established, and DHE was administered orally for 14 days. ResultsDHE suppressed the viability of HCT116, CT26, SW480, and LoVo cells. DHE treatment led to G2/M arrest via a reduction of cyclin B1/CDK1 and caspase-dependent apoptosis. It also induced autophagy by regulating LC3-II and beclin-1 expression. Additionally, migration and invasion of CRC cells were decreased by DHE through regulation of the expression of EMT markers. Oral administration of DHE could inhibit the lung metastasis of CT26 cells in an in vivo model. ConclusionOur study demonstrated that DHE has a potential therapeutic effect on colorectal cancer metastasis.

Exosome-Derived ADAM17 Promotes Liver Metastasis in Colorectal Cancer.

Exosomes derived from cancer cells are deemed important drivers of pre-metastatic niche formation at distant organs, but the underlying mechanisms of their effects remain largely unknow. Although the role of ADAM17 in cancer cells has been well studied, the secreted ADAM17 effects transported via exosomes are less understood. Herein, we show that the level of exosome-derived ADAM17 is elevated in the serum of patients with metastatic colorectal cancer as well as in metastatic colorectal cancer cells. Furthermore, exosomal ADAM17 was shown to promote the migratory ability of colorectal cancer cells by cleaving the E-cadherin junction. Moreover, exosomal ADAM17 overexpression as well as RNA interference results highlighted its function as a tumor metastasis-promoting factor in colorectal cancer in vitro and in vivo. Taken together, our current work suggests that exosomal ADAM17 is involved in pre-metastatic niche formation and may be utilized as a blood-based biomarker of colorectal cancer metastasis.

Expression and prognostic significance of CBX2 in colorectal cancer: database mining for CBX family members in malignancies and vitro analyses

BackgroundThe Chromobox (CBX) domain protein family, a core component of polycomb repressive complexes 1, is involved in transcriptional repression, cell differentiation, and program development by binding to methylated histone tails. Each CBX family member plays a distinct role in various biological processes through their own specific chromatin domains, due to differences in conserved sequences of the CBX proteins. It has been demonstrated that colorectal cancer (CRC) is a multiple-step biological evolutionary process, whereas the roles of the CBX family in CRC remain largely unclear.MethodsIn the present study, the expression and prognostic significance of the CBX family in CRC were systematically analyzed through a series of online databases, including Cancer Cell Line Encyclopedia (CCLE), Oncomine, Human Protein Atlas (HPA), and Gene Expression Profiling Interactive Analysis (GEPIA). For in vitro verification, we performed cell cloning, flow cytometry and transwell experiments to verify the proliferation and invasion ability of CRC cells after knocking down CBX2.ResultsMost CBX proteins were found to be highly expressed in CRC, but only the elevated expression of CBX2 could be associated with poor prognosis in patients with CRC. Further examination of the role of CBX2 in CRC was performed through several in vitro experiments. CBX2 was overexpressed in CRC cell lines via the CCLE database and the results were verified by RT-qPCR. Moreover, the knockdown of CBX2 significantly suppressed CRC cell proliferation and invasion. Furthermore, the downregulation of CBX2 was found to promote CRC cell apoptosis.ConclusionsBased on these findings, CBX2 may function as an oncogene and potential prognostic biomarker. Thus, the association between the abnormal expression of CBX2 and the initiation of CRC deserves further exploration.

Fusobacterium Nucleatum Promotes the Development of Colorectal Cancer by Activating a Cytochrome P450/Epoxyoctadecenoic Acid Axis via TLR4/Keap1/NRF2 Signaling.

Emerging research has revealed regulation of colorectal cancer metabolism by bacteria. Fusobacterium nucleatum (Fn) plays a crucial role in the development of colorectal cancer, however, whether Fn infection modifies metabolism in patients with colorectal cancer remains unknown. Here, LC-MS/MS-based lipidomics identified the upregulation of cytochrome P450 monooxygenases, primarily CYP2J2, and their mediated product 12,13-EpOME in patients with colorectal cancer tumors and mouse models, which increased the invasive and migratory ability of colorectal cancer cells in vivo and in vitro by regulating the epithelial-mesenchymal transition (EMT). Metagenomic sequencing indicated a positive correlation between increased levels of fecal Fn and serum 12,13-EpOME in patients with colorectal cancer. High levels of CYP2J2 in tumor tissues also correlated with high Fn levels and worse overall survival in patients with stage III/IV colorectal cancer. Moreover, Fn was found to activate TLR4/AKT signaling, downregulating Keap1 and increasing NRF2 to promote transcription of CYP2J2. Collectively, these data identify that Fn promotes EMT and metastasis in colorectal cancer by activating a TLR4/Keap1/NRF2 axis to increase CYP2J2 and 12,13-EpOME, which could serve as clinical biomarkers and therapeutic targets for Fn-infected patients with colorectal cancer. SIGNIFICANCE: This study uncovers a mechanism by which Fusobacterium nucleatum regulates colorectal cancer metabolism to drive metastasis, suggesting the potential biomarker and therapeutic utility of the CYP2J2/12,13-EpOME axis in Fn-infected patients.

ELK1‑mediated upregulation of lncRNA LBX2‑AS1 facilitates cell proliferation and invasion via regulating miR‑491‑5p/S100A11 axis in colorectal cancer.

The aim of the present study was to investigate the role and regulatory mechanism of LBX2 antisense RNA 1 (LBX2-AS1) in colorectal cancer. Firstly, LBX2-AS1 expression was detected using reverse transcription-quantitative PCR in colorectal cancer tissues and cells, and its prognostic and diagnostic efficacy was assessed in a colorectal cancer cohort (n=145). Subcellular fractionation assay of LBX2-AS1 was performed. Secondly, the effects of LBX2-AS1 and microRNA (miR)-491-5p on colorectal cancer cell proliferation, apoptosis, migration and invasion were investigated by a series of functional assays. Thirdly, RNA immunoprecipitation, dual-luciferase reporter and gain and loss of function assays were carried out to analyze the interactions between ETS transcription factor ELK1 (ELK1) and LBX2-AS1, as well as LBX2-AS1, miR-491-5p and S100A11. The results showed that LBX2-AS1 was upregulated both in colorectal cancer tissues and cells, which was distributed in the cytoplasm and nucleus of colorectal cancer cells. Clinically, high LBX2-AS1 expression could be an independent prognostic factor for colorectal cancer. Furthermore, relative operating characteristic curve analysis showed that LBX2-AS1 was a sensitive diagnostic marker for colorectal cancer. Highly expressed ELK1, as a transcription factor, could bind to the two conserved sites in the promoter region of LBX2-AS1, thereby activating the transcription of LBX2-AS1. Silencing LBX2-AS1 markedly inhibited proliferative, migratory and invasive abilities of colorectal cancer cells. miR-491-5p expression was downregulated, while S100A11 expression was upregulated in colorectal cancer tissues and cells. Dual-luciferase reporter assays confirmed that LBX2-AS1 could block S100A11 degradation via competitively binding to miR-491-5p. Furthermore, LBX2-AS1 overexpression could notably reverse the inhibitory effect of miR-491-5p on proliferation and invasion of colorectal cancer cells. Taken together, LBX2-AS1 induced by transcription factor ELK1 may facilitate colorectal cancer cell proliferation and invasion via regulation of the miR-491-5p/S100A11 axis. Thus, LBX2-AS1 could be an underlying prognostic and diagnostic marker for colorectal cancer.

In vivo and in vitro Experiment of E74-Like Factor 5 Overexpression Inhibiting the Biological Behavior of Colon Cancer Cells

To investigate the effect of E74-like factor 5 (ELF5) overexpression on the growth and invasion ability of colorectal cancer cells and its effect on tumor formation in nude mice. Human colorectal cancer SW480 and HT-29 cells were divided into 5 groups: the lentivirus (LV)- GFP group transfected with empty vector LV- GFP, the LV- ELF5 group transfected with recombinant LV- ELF5, the shRNA-NC group transfected with empty vector shRNA-NC, the shRNA- ELF5 group transfected with recombinant shRNA- ELF5, and the control group, not transfected with any vector. Seventy-two h after transfection, the cell supernatant containing lentivirus was collected. The mRNA expression level of ELF5 in each group was examined by real-time fluorescent quantitative PCR (RT-qPCR). The protein expression levels of ELF5, apoptosis-related cleaved Caspase-3/Caspase-3 and cleaved Caspase-9/Caspase-9, and invasion-related E-cadherin and N-cadherin were checked with Western blot. CCK-8 was used to check cell viability. Colony formation experiment was done to evaluate colony formation rate. Flow cytometry was used to assess cell apoptosis. Transwell migration assay was used to examine cell invasion. TUNEL assay was used to examine the apoptosis of tissues cells. Immunohistochemistry test was done to determine the expression of E-cadherin and N-cadherin in tissues. 20 BALB/c nude mice were put into 4 groups (5 in each group): LV- GFP group, shRNA-NC group, LV- ELF5 group, and shRNA- ELF5 group. Recombinant lentiviral SW480 cell supernatants were subcutaneously injected into nude mice to construct nude mice tumorigenesis models and the volume changes of transplanted tumors were monitored. On the 30th day, transplanted tumor tissues from the nude mice were extracted and the tumor mass was measured. Western blot was done to measure the expression of ELF4 protein in the transplanted tumors. TUNEL staining was used to check cell apoptosis in the tissues, and the positive expression of N-cadherin in the transplanted tumor was measured by immunohistochemical tests. Compared with the control group, there was no statistically significant difference in the indicators of the two cell lines in the LV- GFP group and shRNA-NC group. The results of Western blot and RT-qPCR showed that the ELF5 protein and mRNA of the LV- ELF5 group of the two cell lines were up-regulated ( P<0.05, compared with those of the LV- GFP group), and the ELF5 protein and mRNA of the shRNA- ELF5 group were down-regulated ( P<0.05). The ELF5 overexpression system and interference system were successfully constructed. Compared with the LV- GFP group, data from the LV- ELF5 group showed that cell viability and colony formation rate ( P<0.05) were reduced, SW480 and HT-29 cell apoptosis was promoted, cleaved Caspase-3/Caspase-3 and cleaved Caspase-9/Caspase-9 protein expression was up-regulated ( P<0.05), cell invasion was inhibited, and the expression of E-cadherin protein was up-regulated while the expression of N-cadherin protein was down-regulated ( P<0.05). After ELF5 interference, the above-mentioned expression of cells demonstrated an opposite trend ( P<0.05, comparing shRNA- ELF5 group with shRNA-NC group). In vivo experimental results indicated that ELF5 overexpression reduced tumor volume and tumor mass ( P<0.05), promoted cell apoptosis in tissues ( P<0.05), and inhibited N-cadherin protein expression ( P<0.05). When ELF5 expression was inhibited, the above mentioned experimental results showed the opposite trend. In vivo and in vitro experiments showed that ELF5 overexpression could promote the apoptosis of colorectal cancer cells and inhibit the growth and invasion of colorectal cancer cells.

Wnt7a Promotes the Occurrence and Development of Colorectal Adenocarcinoma.

ObjectiveThe expression of Wnt7a in colorectal cancer tissues and cell lines was analyzed, and the effect of Wnt7a on the proliferation of colorectal cancer cells was studied, so as to confirm the relationship between Wnt7a and the occurrence and development of colorectal cancer.Methods(1) Immunohistochemical method was used to compare the expression of Wnt7a in different tissues and its relationship with the clinicopathology of colorectal adenocarcinoma. (2) The expression levels of Wnt7a in colorectal cancer cell lines HT-29 and HCT 116 were detected by qRT-PCR. (3) The down-regulated Wnt7A expression vector was constructed, and the down-regulated Wnt7A expression cell line was established. The regeneration ability of cancer cells was detected by stem cell ball formation assay, and the influence of plate cloning assay on the proliferation ability of colorectal cancer cells was detected.Results(1) The positive rates of Wnt7a in normal colorectal mucosa, colorectal adenoma and colorectal adenocarcinoma tissues gradually increased,Wnt7a are closely related to the degree of colorectal adenocarcinoma differentiation, lymph node metastasis and Duke stage. (2) The expression level of Wnt7a in colorectal cancer cells was higher than that in normal colorectal epithelial cells. (3) The down-regulation of Wnt7A reduced the proliferation ability of colorectal cancer cells.ConclusionsWnt7a promotes the occurrence and development of colorectal adenocarcinoma.