Inhibited Colorectal Cancer Cell Growth Research Articles

The Role of p53-Mediated Signaling in the Therapeutic Response of Colorectal Cancer to 9F, a Spermine-Modified Naphthalene Diimide Derivative.

Colorectal cancer (CRC) is one of the most prevalent cancers due to its frequency and high rate of mortality. Polyamine-vectorized anticancer drugs possess multiple biological properties. Of these drugs, 9F has been shown to inhibit tumor growth and the metastasis of hepatocellular carcinoma. This current study aims to investigate the effects of 9F on CRC and determine its molecular mechanisms of action. Our findings demonstrate that 9F inhibits CRC cell growth by inducing apoptosis and cell cycle arrest, and suppresses migration, invasion and angiogenesis in vitro, resulting in the inhibition of tumor growth and metastasis in vivo. Based on RNA-seq data, further bioinformatic analyses suggest that 9F exerts its anticancer activities through p53 signaling, which is responsible for the altered expression of key regulators of the cell cycle, apoptosis, the epithelial-to-mesenchymal transition (EMT), and angiogenesis. In addition, 9F is more effective than amonafide against CRC. These results show that 9F can be considered as a potential strategy for CRC treatment.

Resibufogenin inhibited colorectal cancer cell growth and tumorigenesis through triggering ferroptosis and ROS production mediated by GPX4 inactivation.

Resibufogenin (RB) has been used for cancer treatment, but the underlying mechanisms are still unclear. This study aimed to investigate the effects of RB treatment on colorectal cancer (CRC) cells, and to determine the underlying mechanisms. The cell counting kit-8 assay was used to determine cell viability. Cell morphology was observed under light microscopy, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay was employed to detect cell apoptosis. Intracellular ferrous iron (Fe2+ ), malondialdehyde (MDA), glutathione (GSH), and reactive oxygen species levels were detected by using commercial iron assay kit, MDA assay kit, GSH assay kit, and 2,7-dichlorodihydrofluorescein diacetate probes, respectively. The protein expressions were determined by Western blot and immunohistochemistry. RB inhibited cell viability in the CRC cell lines (HT29 and SW480) in a dose- and time-dependent manner, and caused cytotoxicity to the normal colonic epithelial cell line (NCM460) at high dose. Similarly, RB induced morphological changes in CRC cells from normal to round shape, and promoted cell death. Of note, RB triggered oxidative stress and ferroptotic cell death in CRC cells, and only ferroptosis inhibitors (deferoxamine and ferrostatin-1), instead of inhibitors for other types of cell death (apoptosis, autophagy, and necroptosis), reversed the inhibitory effects of RB on CRC cell proliferation. Furthermore, glutathione peroxidase 4 (GPX4) was inactivated by RB treatment, and overexpression of GPX4 alleviated RB-induced oxidative cell death in CRC cells. Consistently, the in vivo experiments validated that RB also triggered oxidative stress, and inhibited CRC cells growth and tumorigenicity in mice models. RB can inhibit CRC cells growth and tumorigenesis by triggering ferroptotic cell death in a GPX4 inactivation-dependent manner.

Linc00662 Promotes Tumorigenesis and Progression by Regulating miR-497-5p/AVL9 Axis in Colorectal Cancer.

BackgroundRecently, multiple lines of evidence have demonstrated that linc00662 serves as an oncogene in various cancers. However, the exact mechanism of oncogenesis mediated by linc00662 in colorectal cancer (CRC) remains unknown. In this study, we aimed to explore the biological role of linc00662 in the regulation of CRC progression.MethodsBoth gene expression omnibus (GEO) and the cancer genome atlas (TCGA) datasets were used to evaluate the expression of linc00662. RT-qPCR was used to analyze the expression of linc00662, miR-497-5p, and AVL9 in CRC clinical samples and cell lines. Cell Counting Kit-8 (CCK-8), flow cytometry, transwell assay, and xenograft model were used to investigate the effect of linc00662 on CRC cell proliferation, cell cycle, and metastasis. Western blot analysis was used to analyze the expression of the epithelial-mesenchymal transition (EMT)-associated markers. Furthermore, bioinformatics analysis and mechanism assays were used to elucidate the underlying mechanism. Dual-luciferase reporter assays were used to analyze the regulatory relationships among linc00662, miR-497-5p, and AVL9.ResultsIn this study, we found that the expression of linc00662 was significantly upregulated in CRC tissues compared to normal tissues and positively correlated with tissue differentiation, T stage, and lymphatic metastasis. Further, our data showed that the expression of linc00662 was positively associated with lymph node metastasis, TMN stage, and poor-moderate differentiation. Patients with higher linc00662 expression level were more likely to have poorer overall survival. Knockdown of linc00662 inhibited CRC cell growth, induced cell apoptosis, triggered cell cycle arrest at G2/M phase, and suppressed cell migration and invasion through regulating the EMT pathway. Further, mechanistic studies revealed that knockdown of linc00662 significantly reduced the expression of AVL9, a direct target of miR-497-5p.ConclusionsLinc00662 was significantly upregulated in CRC, and mediated CRC progression and metastasis by competing with miR-497-5p to modulate the expression of AVL9. Therefore, our result sheds light on the potential application of linc00662 in CRC diagnosis and therapy.

RING Finger Protein 38 Mediates LIM Domain Binding 1 Degradation and Regulates Cell Growth in Colorectal Cancer.

Background and ObjectivesRING finger protein 38 (RNF38) has been reported to be involved in the tumorigenesis of several tumors, but its role in colorectal cancer (CRC) is still not investigated. In the present study, we aimed to investigate the effect of RNF38 in CRC cells.Materials and MethodsThe public tumor databases GEPIA and Kaplan-Meier Plotter were used to analyze RNF38 expression and patients’ overall survival in CRC. The qRT-PCR was carried out to assess the mRNA levels of RNF38 and LDB1. Western blot and co-immunoprecipitation were used to detect protein expression and ubiquitination. CCK-8 assay was performed to analyze CRC cell growth and viability.ResultsRNF38 was found downregulated in CRC tumor tissues and cell lines, and CRC patients with high RNF38 expression had a longer overall survival than patients with low RNF38 expression. Our further investigations showed that RNF38 interacted with LDB1, and downregulated LDB1 expression by inducing its polyubiquitination. Moreover, overexpression of RNF38 inhibited CRC cell growth but enforced LDB1 could significantly antagonize RNF38-induced cell growth inhibition in CRC cells. Additionally, RNF38/LDB1 axis was involved in the drug sensitivity of 5-FU to CRC cells.ConclusionOur studies suggested that RNF38 was functional in CRC cells, and downregulated CRC cell growth by inducing LDB1 polyubiquitination, which indicated that RNF38 could be as a novel target for CRC therapy.

Apigenin-mediated Alterations in Viability and Senescence of SW480 Colorectal Cancer Cells Persist in The Presence of L-thyroxine.

Deregulation of Thyroid Hormones (THs) system in Colorectal Cancer (CRC) suggests that these hormones may play roles in CRC pathogenesis. Flavonoids are polyphenolic compounds, which possess potent antitumor activities and interfere, albeit some of them, with all aspects of THs physiology. Whether the antitumor actions of flavonoids are affected by THs is unknown. Therefore, we investigated the effects of apigenin (Api), a well-known flavone, on some tumorigenic properties of SW480 CRC cells in the presence and absence of L-thyroxine (T4). Cell viability was assessed by MTT assay. Flow cytometry and DNA electrophoresis were used to evaluate cell death. Cell senescence was examined by in situ detection of β-galactosidase activity. Protein expression was assessed by antibody array technique. While T4 had minimal effects, Api reduced cell growth and senescence by induction of apoptosis. Expression of anti-apoptotic and pro-apoptotic proteins were differentially affected by Api and T4. Survivin, HSP60 and HTRA were the most expressed proteins by the cells. Almost all Api-induced effects persisted in the presence of T4. These data suggest that Api may inhibit CRC cell growth and progression through induction of apoptosis rather than cell necrosis or senescence. In addition, they suggest that T4 has minimal effects on CRC cell growth, and is not able to antagonize the anti-growth effects of Api. Regardless of the treatments, cells expressed high levels of survivin, HSP60 and HTRA, indicating that these proteins may play central roles in SW480 CRC cell immortality.

Evaluation of Anticancer Activity of Polyherbal extract of Terminalia Chebula, Phyllanthus emblica and Dimocarpus Longan on Caco-2 Colorectal Cancer Cell Lines

Medicinal plants are the most essential wellsprings of medication or life sparing medications for most of the World& #39;s populace for a huge number of years. Many plant species are used to treat the cancer diseases. The ethanolic extract of composite medicinal herbs fruits of Terminalia chebula, Phyllanthus emblica and seeds of Dimocarpus longan are used as a potent natural source of anticancer agent. The colorectal cancer cell lines (Caco-2) are inhibited by ethanolic extract through MTT assay method. The percentage of Caco-2 colorectal cancer cell growth inhibition was increased from 1.5% to 62.9%.

Overexpression of miR-106a enhances oxaliplatin sensitivity of colorectal cancer through regulation of FOXQ1.

Chemotherapy resistance poses a major challenge for the clinical treatment of colorectal cancer, therefore, the aim of the present study was to examine its underlying mechanisms. Reverse transcription-quantitative polymerase chain reaction and western blot analysis were used to determine the microRNA (miRNA)/mRNA and protein expression levels, respectively. A dual luciferase assay was conducted for verification of the interaction between miR-106a and 3′untranslated region (UTR) of Forkhead box Q1 (FOXQ1). Cell viability was assessed using an MTT assay. In the present study, it was demonstrated that miR-106a is involved in regulating oxaliplatin sensitivity of colorectal cancer. Transfection of miR-106a mimics slightly inhibited colorectal cancer cell growth and sensitized colorectal cancer cells to oxaliplatin exposure. In addition, miR-106a overexpression induced a decrease of FOXQ1 at mRNA and protein levels in colorectal cancer cells. The enhanced expression of miR-106a also increased the expression of Wnt target genes, including vascular endothelial growth factor-A and matrix metallopeptidase 2, which were reported to be regulated by FOXQ1. It was predicted and validated that miR-106a could repress FOXQ1 expression via direct binding to 3′UTR. Elevation of miR-106a and a decrease of FOXQ1 expression levels were detected in tumor tissues from patients with oxaliplatin-sensitive colorectal cancer, compared with patients with oxaliplatin-resistant colorectal cancer. Furthermore, there was a significant association between miR-106a and FOXQ1 mRNA levels. In conclusion, the present study demonstrated that miR-106a increased oxaliplatin sensitivity of colorectal cancer cells through direct repression of FOXQ1 expression.

RETRACTED: METTL14 Suppresses CRC Progression via Regulating N6-Methyladenosine-Dependent Primary miR-375 Processing

Epigenetic alterations contributed to human carcinogenesis immensely. N6-methyladenosine (m6A) is one of the most preventive and abundant modifications on RNA molecules present in eukaryotes. However, the biological function of m6A methylation in colorectal cancer (CRC) remains largely unclear. Here, we found that METTL14 was downregulated in CRC tissues and cell lines, and closely correlated with overall survival (OS). METTL14 knockdown significantly reduced m6A levels in total RNAs and promoted CRC cell growth and metastasis, whereas METTL14 overexpression markedly increased m6A levels in total RNA and inhibited CRC cell growth and metastasis. Furthermore, we demonstrated that miR-375 was a downstream target of METTL14. We also verified that METTL14 suppressed CRC cell growth via the miR-375/Yes-associated protein 1 (YAP1) pathway, as well as inhibited CRC cell migration and invasion through the miR-375/SP1 pathway. Taken together, our studies showed an important role for METTL14 in CRC progression and provided novel insight into m6A modification in CRC progression.

Polo-like kinase 3 inhibits glucose metabolism in colorectal cancer by targeting HSP90/STAT3/HK2 signaling

BackgroundPolo-like kinase 3 (PLK3) has been documented as a tumor suppressor in several types of malignancies. However, the role of PLK3 in colorectal cancer (CRC) progression and glucose metabolism remains to be known.MethodsThe expression of PLK3 in CRC tissues was determined by immunohistochemistry. Cells proliferation was examined by EdU, CCK-8 and in vivo analyses. Glucose metabolism was assessed by detecting lactate production, glucose uptake, mitochondrial respiration, extracellular acidification rate, oxygen consumption rate and ATP production. Chromatin immunoprecipitation, luciferase reporter assays and co-immunoprecipitation were performed to explore the signaling pathway. Specific targeting by miRNAs was determined by luciferase reporter assays and correlation with target protein expression.ResultsPLK3 was significantly downregulated in CRC tissues and its low expression was correlated with worse prognosis of patients. In vitro and in vivo experiments revealed that PLK3 contributed to growth inhibition of CRC cells. Furthermore, we demonstrated that PLK3 impeded glucose metabolism via targeting Hexokinase 2 (HK2) expression. Mechanically, PLK3 bound to Heat shock protein 90 (HSP90) and facilitated its degradation, which led to a significant decrease of phosphorylated STAT3. The downregulation of p-STAT3 further suppressed the transcriptional activation of HK2. Moreover, our investigations showed that PLK3 was directly targeted by miR-106b at post-transcriptional level in CRC cells.ConclusionThis study suggests that PLK3 inhibits glucose metabolism by targeting HSP90/STAT3/HK2 signaling and PLK3 may serve as a potential therapeutic target in colorectal cancer.

AMP‐kinase inhibitor dorsomorphin reduces the proliferation and migration behavior of colorectal cancer cells by targeting the AKT/mTOR pathway

Colorectal cancer (CRC) is among the leading causes of cancer-related mortality, despite extensive efforts in the identification of new treatment options. Hence, there is a need for the development of novel agents with therapeutic potential in treatment of CRC. Dorsomorphin has demonstrated antiproliferative activity against different malignancies. Here we have investigated the pharmaceutical potential of dorsomorphin in two-dimensional and three-dimensional cell-culture models of CRC. The antiproliferative, antimigratory, apoptotic activity and effect of this agent on cell cycle was evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, wound healing assay, and flow cytometry, respectively, while the expression of genes involved in Wnt/Pi3K pathways was assessed at mRNA and/or proteins by reverse transcription polymerase chain reaction (RT-PCR) or western blot. Dorsomorphin inhibited CRC cell growth by modulating the cyclinD1, surviving and p-Akt. This agent was able to reduce the migratory behaviors of CRC cells, compared to control cells, through perturbation of E-cadherin. Also our data showed that dorsomorphin enhanced the percentage of the cells in sub-G1 and induced apoptosis in both late/early stages, as detected by annexin V. Also the regulatory effect of dorsomorphin on oxidant/antioxidant balance was assessed by cellular reactive oxygen species (ROS) generation. In particular, these data showed that dorsomorphin markedly increased the ROS production in CRC cells. Our finding demonstrated that dorsomorphin antagonizes cell growth and migration, through perturbation of Akt/mTOR/Wnt pathways in CRC, supporting further studies on the therapeutic potential of this novel anticancer agent in treatment of CRC.

ARHGAP25: A negative regulator of colorectal cancer (CRC) metastasis via the Wnt/β-catenin pathway

Herein, we found ARHGAP25 was down-regulated in colon biopsies of patients with colorectal cancer (CRC). Gene set enrichment analysis (GSEA) also showed that ARHGAP25 was negatively correlated with Wnt/β-catenin activation. To study the role of ARHGAP25 in CRC and its possible mechanism, we established lentiviral-mediated ARHGAP25 overexpression in HCT116 and RKO cells along with siRNA-mediated ARHGAP25 knockdown in SW620 cells. The metastatic capacity of the CRC cell lines in vitro was assessed by measuring cell proliferation, migration and invasion. Additionally, expression of matrix metalloproteinases (MMP2, MMP7 and MMP9), EMT-associated factors (E-cadherin, ZEB1, Snail and Twist1) and β-catenin (a core part of Wnt/β-catenin signaling) was determined. XAV939, a Wnt/β-catenin inhibitor, was used for treatment. Our data suggests that ARHGAP25 overexpression significantly inhibits CRC cell growth, suppresses cell migration and invasion, and reduces expression of MMPs, EMT-associated factors and β-catenin. Our results suggest not only that ARHGAP25 has an anti-metastatic role in CRC cells, but also that the inactivation of the Wnt/β-catenin pathway is important in this process. Accordingly, siRNA-ARHGAP25 resulted in the opposite effect, favoring CRC metastasis in vitro, and activating the Wnt/β-catenin pathway in CRC cells. In addition, the siRNA-ARHGAP25 induced changes on cell proliferation, migration, and invasion were significantly reversed with XAV939 treatment. Importantly, the anti-metastatic effects of ARHGAP25 were further substantiated in a CRC lung metastasis xenograft model. We conclude that ARHGAP25 negatively regulates the metastatic potential of CRC cells via the Wnt/β-catenin pathway. Thus, our findings implicate ARHGAP25 as a potential therapeutic target in CRC metastasis.

MicroRNA-193a-3p suppresses the colorectal cancer cell proliferation and progression through downregulating the PLAU expression.

BackgroundColorectal cancer (CRC) is one of the leading causes of cancer-related death in China. Dysregulation of microRNAs (miRNAs) is involved in cancer development and progression. Our previous study showed an inverse relationship between miR-193a-3p expression and the prognosis of CRC. However, the exact biological functions of miR-193a-3p in CRC are still poorly understood. This study aimed to explore the role and mechanism of miR-193a-3p in CRC.MethodsReal-time PCR and Western blotting were used to examine the expression levels of RNA and protein, respectively. A dual luciferase assay was performed to validate predicted targets of miR-193a-3p. Loss and gain-of-function studies were carried out to reveal the effects and potential mechanism of the miR-193a-3p in the proliferation, metastasis and angiogenesis of CRC cells.ResultsThe expression levels of miR-193a-3p in human CRC cell lines were significantly decreased compared with that in normal colonic epithelium cell line. Furthermore, plasminogen activator urokinase (PLAU) was validated as a direct target gene of miR-193a-3p. Over-expression of miR-193a-3p inhibited proliferation, migration and angiogenesis of HT-29 cell, whereas forced expression of PLAU could rescue the inhibitory effects.ConclusionmiR-193a-3p might inhibit CRC cell growth, migration and angiogenesis partly through targeting PLAU. MiR-193a-3p/PLAU axis might provide a potent therapeutic opportunity for aggressive CRC.

Dual PI3K-BRD4 Inhibitor SF1126 Inhibits Colorectal Cancer Cell Growth in Vitro and in Vivo

Bromodomain-containing protein 4 (BRD4) and phosphatidylinositol 3-kinase (PI3K) are key oncogenic cascades in colorectal cancer (CRC). SF1126 is a novel and potent PI3K-BRD4 dual inhibitor. CRC cells and human colon epithelial cells were treated with SF1126. Cell survival was tested by MTT and soft agar colony formation assays. Cell proliferation was tested by BrdU ELISA method. Cell apoptosis was tested by a TUNEL staining method and Histone DNA ELISA. Western blotting was utilized to test the signaling proteins. A HT-29 xenograft mice model was established to study the anti-tumor activity of SF1126 in vivo. SF1126 potently inhibited the survival, proliferation, and progression of the cell cycle in an established CRC cell line (HT-29) and primary human colon cancer cells. Significant activation of apoptosis was detected in SF1126-treated CRC cells. In CRC cells, SF1126 blocked Akt-mammalian target of rapamycin (mTOR) complex1/2 signaling and downregulated BRD4 target proteins (Myc and cyclin D1). Further studies showed that SF1126 activated p38 signaling in CRC cells. In contrast, the p38 inhibitors or p38 short hairpin RNA inhibited SF1126-induced cytotoxicity and apoptosis in CRC cells. In vivo, subcutaneous administration of SF1126 significantly inhibited HT-29 xenograft tumor growth in nude mice. SF1126 inhibits CRC cell growth possibly by targeting PI3K-Akt-mTOR, BRD4, and p38 signaling.

NAA40 contributes to colorectal cancer growth by controlling PRMT5 expression

N-alpha-acetyltransferase 40 (NAA40) catalyzes the transfer of an acetyl moiety to the alpha-amino group of serine 1 (S1) on histones H4 and H2A. Our previous studies linked NAA40 and its corresponding N-terminal acetylation of histone H4 (N-acH4) to colorectal cancer (CRC). However, the role of NAA40 in CRC development was not investigated. Here, we show that NAA40 protein and mRNA levels are commonly increased in CRC primary tissues compared to non-malignant specimens. Importantly, depletion of NAA40 inhibits cell proliferation and survival of CRC cell lines and increases their sensitivity to 5-Fluorouracil (5-FU) treatment. Moreover, the absence of NAA40 significantly delays the growth of human CRC xenograft tumors. Intriguingly, we found that NAA40 knockdown and loss of N-acH4 reduce the levels of symmetric dimethylation of histone H4 (H4R3me2s) through transcriptional downregulation of protein arginine methyltransferase 5 (PRMT5). NAA40 depletion and subsequent repression of PRMT5 results in altered expression of key oncogenes and tumor suppressor genes leading to inhibition of CRC cell growth. Consistent with this, NAA40 mRNA levels correlate with those of PRMT5 in CRC patient tissues. Taken together, our results establish the oncogenic function of the epigenetic enzyme NAA40 in colon cancer and support its potential as a therapeutic target.

Bromelain inhibits the ability of colorectal cancer cells to proliferate via activation of ROS production and autophagy

Advanced colorectal cancer (CRC) survival rates are still low despite advances in cytotoxic and targeted therapies. The development of new effective or alternative therapies is therefore urgently needed. Bromelain, an extract of pineapple, was shown to have anticancer effects, but its mechanisms in CRC have not been fully explored. Therefore, the roles of bromelain in CRC progression were investigated using different CRC cell lines, a zebrafish model, and a xenograft mouse model. The anticancer mechanisms were explored by assessing the role of bromelain in inducing reactive oxygen species (ROS), superoxide, autophagosomes, and lysosomes. The role of bromelain in the induction of apoptosis was also assessed. It was found that bromelain inhibited CRC cell growth in cell lines and tumor growth in the zebrafish and xenograft mouse models. It also induced high levels of ROS and superoxide, plus autophagosome and lysosome formation. High levels of apoptosis were also induced, which were associated with elevated amounts of apoptotic proteins like apoptotic induction factor, Endo G, and caspases-3, -8, and -9 according to a qPCR analysis. In a Western blot analysis, increases in levels of ATG5/12, beclin, p62, and LC3 conversion rates were found after bromelain treatment. Levels of cleaved caspase-3, caspase-8, caspase-9, and poly(ADP ribose) polymerase (PARP)-1 increased after bromelain exposure. This study explored the role of bromelain in CRC while giving insights into its mechanisms of action. This compound can offer a cheap alternative to current therapies.

TIMP3 expression associates with prognosis in colorectal cancer and its novel arylsulfonamide inducer, MPT0B390, inhibits tumor growth, metastasis and angiogenesis.

Tissue inhibitors of metalloproteinase 3 (TIMP3) are a major endogenous inhibitor of matrix metalloproteinase (MMPs) that inhibit tumor growth, invasion, metastasis and angiogenesis. In this study, we found that TIMP3 expression is associated with positive prognosis of colorectal cancer (CRC) clinicopathologically. Therefore, we developed a series of arylsulfonamide derivatives as TIMP3 inducers in order to define potential colorectal cancer therapeutic agent. Among these, MPT0B390 was selected for anti-tumor, anti-metastasis, and anti-angiogenesis property determination.Methods: The relationship between TIMP3 expression and clinical pathological features in colorectal patients and cell lines were determined by immunohistochemistry, bioinformatics analysis and western blotting. The anti-tumor function was validated by using MTT, apoptosis pathway detection and in vivo xenograft model for tumor growth inhibition determination. The anti-metastatic function was validated using a transwell migration assay, and using in vivo lung metastasis and liver metastasis models. The mechanism of MPT0B390-induced TIMP3 expression was further tested using qPCR and Chromatin IP assay. The anti-angiogenesis function was examined by using transwell migration assay, and in vivo Matrigel plug assay.Results: After screening candidate compounds, we identified MPT0B390 as an effective inducer of TIMP3. We showed that MPT0B390 induces TIMP3 expression significantly and inhibits CRC cell growth in vitro and in vivo. By inducing TIMP3 expression, MPT0B390 can also exert its anti-metastasis effect to inhibit CRC cell migration and invasion and downregulates migration markers such as uPA, uPAR, and c-Met. Subsequent Chromatin immunoprecipitation assay revealed that MPT0B390 can significantly inhibit EZH2 expression as well as its binding to TIMP3 promoter region to regulate TIMP3 induction. In addition to the anti-tumor and anti-metastasis capability, MPT0B390 can also induce TIMP3 expression in endothelial cells to inhibit tumor angiogenesis.Conclusion: These data suggest the potential therapeutic applications of the TIMP3 inducer, MPT0B390, for colorectal cancer treatment.

Ubiquitin specific peptidase 5 regulates colorectal cancer cell growth by stabilizing Tu translation elongation factor

Ubiquitin specific peptidase 5 (USP5) is a ubiquitous expressed deubiquitinating enzyme (DUB). It has been shown involved in DNA repair, apoptosis, inflammation, and tumor cell growth. However, the function and molecular mechanism of USP5 in colorectal cancer (CRC) are still unclear. In the present study, we asked how it affected the growth of colorectal cancer cells.Methods: A shRNA-based high-content screening was performed to identify DUBs affecting the growth of CRC cells. CCK-8 assay and xenografts were used to assess CRC cell growth, survival and tumorigenesis. RT-qPCR, immunoblotting and immunohistochemistry were carried out to quantitate USP5 expression in CRC tissues and cell lines. Immunoprecipitation and mass spectrometry analysis were performed to identify USP5-interacting proteins. Cycloheximide chase was performed to assess Tu translation elongation factor (TUFM) stability. Dual luciferase reporter assay was utilized for USP5 promoter analysis.Results: We found that USP5 was highly expressed in a group of primary CRC tissues, and the increased USP5 was correlated with clinical stages and shorter overall survival. While USP5 knockdown effectively inhibited CRC cell growth, overexpressed USP5 promoted the growth of CRC cells and made them more resistant to doxorubicin (DOX). TUFM was discovered as a substrate of USP5. USP5 deubiquitinated TUFM and increased its level in CRC cells. Enforced expression of TUFM was able to alleviate the growth inhibition induced by USP5 knockdown. Further analyses showed that EBF transcription factor 1 (EBF1) was a major regulator for USP5 transcription, and DOX inhibited EBF1-USP5-TUFM axis in CRC cells.Conclusions: USP5 was required for CRC cells and promoted their growth and resistance to chemotherapeutics. TUFM was a USP5 deubiquitinating substrate that mediated the cellular effects of USP5. The transcription of USP5 was regulated by EBF1. Thus, targeting EBF1-USP5-TUFM axis is a potential novel strategy for CRC treatment.

MiR-10b suppresses the growth and metastasis of colorectal cancer cell by targeting FGF13.

MicroRNA (miRNA) plays important roles in the progression of different cancers. In this study, we investigated the precise role of miR-10b in the growth and metastasis of colorectal cancer (CRC) cell. The levels of miR-10b in CRC cell lines were detected using quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) assay. A series of functional assays, including cell proliferation, colony formation, wound healing and transwell invasion were conducted using miR-10b transfected cells. The expressions of E-cadherin and N-cadherin were assessed by immunofluorescence staining. Xenograft model and lung metastasis model were constructed to investigate the impact of miR-10b on the growth and metastasis of CRC cell in vivo. We revealed that miR-10b was markedly down-regulated in CRC. The up-regulation of miR-10b suppressed the growth, colony formation, migration and invasion of CRC cell. Furthermore, the xenograft model indicated that miR-10b inhibited CRC cell growth and lung metastasis in vivo by targeting fibroblast growth factor 13 (FGF13). In addition, we demonstrated that the overexpression of FGF13 rescued the suppressive effects of miR-10b on CRC cells growth, migration and invasion. Finally, the knockdown of FGF13 was able to mimic the inhibitory effects of miR-10b on the progression of CRC cells. These results demonstrate that miR-10b plays an important role in growth, migration and invasion of CRC by targeting FGF13.

Deguelin induces apoptosis in colorectal cancer cells by activating the p38 MAPK pathway.

ObjectivesDeguelin, a rotenoid extracted from Mundulea sericea (Leguminosae), exhibits antitumor effects on several types of human cancers. Due to the limited studies of deguelin on colorectal cancer (CRC), the present study was designed to investigate the antitumor effect of deguelin and to explore the underlying mechanism in CRC.Materials and methodsCell viability was assessed by the cell counting kit-8 (CCK-8) assay, and cell apoptosis was determined by the annexin v-propidium iodide staining using flow cytometry and Western blot in CRC cell lines after incubation with deguelin. The antitumor effect of deguelin was further evaluated in tumor xenograft models. Moreover, SB203580, a specific inhibitor of p38 MAPK, was used to confirm the involvement of p38 MAPK pathway in deguelin-induced apoptosis.ResultsDeguelin significantly inhibited cell proliferation and induced apoptosis in CRC cell lines (SW620 and RKO) in a time-dependent and dose-dependent manner. Western blot analysis also showed that the expression of proapoptotic proteins (cleaved caspase 3 and cleaved PARP) was upregulated, while that of antiapoptotic proteins (Bcl-2 and survivin) was downregulated after deguelin treatment in CRC cell lines. Moreover, oral administration of deguelin significantly suppressed tumor growth and induced apoptosis in subcutaneous xenograft mouse models without obvious toxicity. Additionally, Western blot revealed that deguelin-induced apoptosis might be regulated by the p38 MAPK pathway and inhibition of p38 MAPK could attenuate deguelin-induced proliferative inhibition and apoptosis in CRC cells.ConclusionCollectively, these results demonstrated that deguelin inhibited CRC cell growth by inducing apoptosis via activation of p38 MAPK pathway.

Long non-coding RNA TP73‑AS1 promotes colorectal cancer proliferation by acting as a ceRNA for miR‑103 to regulate PTEN expression

There is an increasing evidence that long non-coding RNAs (lncRNAs) play an important role in tumorigenesis and cancer progression. This study focused on the functional role of P73 antisense RNA 1T (TP73‑AS1), a lncRNA, in colorectal cancer (CRC). We found that TP73‑AS1 expression was significantly low in CRC tissues and cells, and high TP73‑AS1 expression was negatively associated with TNM stage, prognosis, overall survival, and disease-free survival in the CRC patients. Moreover, TP73‑AS1 overexpression dramatically inhibited CRC cell growth, promoted apoptosis, downregulated Bcl‑2 levels, and increased caspase‑3 expression. Furthermore, TP73‑AS1 expression levels were positively associated with PTEN levels in clinical CRC samples. As expected, TP73‑AS1 could upregulate PTEN expression in CRC cells. Mechanistically, PTEN was shown to be the target of miR‑103. Interestingly, TP73‑AS1 overexpression could increase PTEN expression through competitive binding to miR‑103. Functionally, our data show that such TP73‑AS1-induced PTEN expression through binding to miR‑103 facilitated CRC cell proliferation. Thus, we showed that TP73‑AS1 inhibits CRC cell growth by functioning as a ceRNA (competing endogenous RNAs) to regulate PTEN levels. Our findings provide new insights into the underlying molecular mechanisms of TP73‑AS1-mediated CRC.