Regulation Of Colorectal Cancer Research Articles

LINC00707 promotes cell proliferation and invasion of colorectal cancer via miR-206/FMNL2 axis.

Long non-coding RNAs (lncRNAs) have been verified to participate in the regulation of colorectal cancer (CRC). However, the role of LINC00707 in CRC still remains unknown. Here, we aim to study the role of LINC00707 in CRC. LINC00707 expression in 97 pairs of CRC tissues and adjacent normal tissues was determined by the quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). LINC00707 overexpression or knockdown in SW620 or HCT116 cells was achieved by lentivirus transfection. The proliferation and cell circle progression of established cells were detected by cell counting kit-8 (CCK-8) assay and flow cytometry, respectively. Cell invasion and migration abilities were studied by transwell assay. Dual-luciferase assay and Western blot was used to verify the underlying mechanism of LINC00707 in CRC. Nude mice were obtained to identify the in vivo function of LINC00707 in CRC. LINC00707 was significantly over-expressed in CRC tissues and cell lines. Up-regulation of LINC00707 promoted cell proliferation, cell cycle progression, invasion, and migration of SW620 cells. Conversely, down-regulation of LINC00707 reduced cell growth and metastasis of HCT116 cells. MiR-206 was verified as a direct target of LINC00707, and its function was inhibited by LINC00707. FMNL2 was a target for miR-206 in CRC cells. Meanwhile, LINC00707 promoted tumor growth of CRC in vivo. LINC00707 was up-regulated in CRC tissues and cells, which promoted cell proliferation and metastasis via sponging miR-206 to increase FMNL2 expression. This might provide a novel target for the biological treatment of CRC.

Corrigendum] Lipocalin2 inversely regulates TRAIL sensitivity through p38 MAPK‑mediated DR5 regulation in colorectal cancer.

Following the publication of this article, an interested reader drew to our attention that Fig. 1C contained an important flaw. The Figure shows a western blot for LCN2, DR4, DR5, and actin, and it was noted that the identical bands shown for actin were also featured in a paper by the same authors published in 2017 [Lipocalin 2 negatively regulates cell proliferation and epithelial to mesenchymal transition through changing metabolic gene expression in colorectal cancer. Kim SL, LeeST, MinIS, Park YR, Lee JH, Kim DG and KimSW: Cancer Sci108: 2176‑2186, 2017], except that the lanes for the cell linesHCT116 and SW620 were depicted the other way around in the International Journal of Oncology article. Upon investigating the matter with the authors, they were able to confirm that the lanes were labelled incorrectly in the Figure itself; moreover, the incorrect control bands were included with the Figure. The corrected version of Fig.1 is shown opposite, including the correct control data for Fig.1C. This error did not have an impact on the overall meaning of the paper, or on the reported conclusions of this study. The authors regret that this error was introduced into the printed version of the paper, and apologize to the readership for any inconvenience caused. [the original article was published in International Journal of Oncology 53: 2789‑2799, 2018; DOI: 10.3892/ijo.2018.4562].

Interplay between post-translational cyclooxygenase-2 modifications and the metabolic and proteomic profile in a colorectal cancer cohort.

BACKGROUNDColorectal cancer (CRC) is the second most common cause of cancer death worldwide. It is broadly described that cyclooxygenase-2 (COX-2) is mainly overexpressed in CRC but less is known regarding post-translational modifications of this enzyme that may regulate its activity, intracellular localization and stability. Since metabolic and proteomic profile analysis is essential for cancer prognosis and diagnosis, our hypothesis is that the analysis of correlations between these specific parameters and COX-2 state in tumors of a high number of CRC patients could be useful for the understanding of the basis of this cancer in humans.AIMTo analyze COX-2 regulation in colorectal cancer and to perform a detailed analysis of their metabolic and proteomic profile.METHODSBiopsies from both healthy and pathological colorectal tissues were taken under informed consent from patients during standard colonoscopy procedure in the University Hospital of Bellvitge (Barcelona, Spain) and Germans Trias i Pujol University Hospital (Campus Can Ruti) (Barcelona, Spain). Western blot analysis was used to determine COX-2 levels. Deglycosylation assays were performed in both cells and tumor samples incubating each sample with peptide N-glycosidase F (PNGase F). Prostaglandin E2 (PGE2) levels were determined using a specific ELISA. 1H high resolution magic angle spinning (HRMAS) analysis was performed using a Bruker AVIII 500 MHz spectrometer and proteomic analysis was performed in a nano-liquid chromatography-tandem mass spectrometer (nano LC-MS/MS) using a QExactive HF orbitrap MS.RESULTSOur data show that COX-2 has a differential expression profile in tumor tissue of CRC patients vs the adjacent non-tumor area, which correspond to a glycosylated and less active state of the protein. This fact was associated to a lesser PGE2 production in tumors. These results were corroborated in vitro performing deglycosylation assays in HT29 cell line where COX-2 protein profile was modified after PNGase F incubation, showing higher PGE2 levels. Moreover, HRMAS analysis indicated that tumor tissue has altered metabolic features vs non-tumor counterparts, presenting increased levels of certain metabolites such as taurine and phosphocholine and lower levels of lactate. In proteomic experiments, we detected an enlarged number of proteins in tumors that are mainly implicated in basic biological functions like mitochondrial activity, DNA/RNA processing, vesicular trafficking, metabolism, cytoskeleton and splicing.CONCLUSIONIn our colorectal cancer cohort, tumor tissue presents a differential COX-2 expression pattern with lower enzymatic activity that can be related to an altered metabolic and proteomic profile.

A Study on the Aberrant Methylation of Colorectal Cancer MLH1 Gene

The aim of this paper is to study the mechanism of MLH1 gene expression and regulation in colorectal cancer, and to clarify the impact of 5' end CpG island aberrant methylation on the regulation of MLH1 gene expression. RT-PCR assay was used to detect the expression of MLH1 gene mRNA in 10 colon cancer cell strains. 62 fresh colorectal adenocarcinoma samples and paired distal normal mucosa were selected in our hospital. They comprised 37 males and 25 females aging 32 to 67 years old with the mean of 43.6. The cellular RNA was extracted, and immunohistochemistry assay was used to detect the expression of MLH1 protein in colorectal cancer, and MSP method was utilized to examine the methylation status of the first exon of MLH1 gene in the tissues. In the 10 cell strains, only the MLH1 mRNAs of SW480 and Caco2 were expressed, and no MLH1 mRNA expression was detected in the other eight strains. The expression level of MLH1 mRNA in the 62 cases of cancer tissue was higher than that in the paired normal mucosa, and MLH1 protein was mainly expressed in the cytoplasm. The expression level of MLH1 protein in adenocarcinoma tissues was significantly higher than that in the paired normal tissue (P<0.01), while the methylation level of cancer tissues was statistically significantly lower than that of the paired normal mucosa (P<0.05). Aberrant DNA methylation dominates the regulation of the MLH1 gene expression changes in colorectal cancer. The results herein provide a theoretical basis for clarifying the differential expressions of MLH1 in colorectal cancer in vitro and in vivo.

Increased EWSAT1 expression promotes cell proliferation, invasion and epithelial-mesenchymal transition in colorectal cancer.

Long non-coding RNAs (lncRNAs) have recently been identified as crucial regulators in colorectal cancer (CRC) progression. The aim of the study is to investigate the clinical role and biological effects of long non-coding RNA EWSAT1 in CRC. The expression of lncRNA EWSAT1 was detected in 106 cases of fresh CRC tissues and matched adjacent normal tissues by qRT-PCR analyses. The Kaplan-Meier analysis and log-rank test were used to assess the association between lncRNA EWSAT1 expression and overall survival (OS) rate of CRC patients. Cell proliferation and invasion capacity were evaluated by CCK8 assay, colony formation, and transwell invasion assays. The protein expression was detected using western blot analysis. LncRNA EWSAT1 expression was abnormally higher in CRC tissues compared to matched adjacent normal tissues. Higher lncRNA EWSAT1 expression significantly associated with depth of invasion, lymph node metastasis, and advanced tumor-node-metastasis (TNM) stage in CRC patients. Patients with higher EWSAT1 expression exhibited shorter OS compared with patients with lower EWSAT1 expression. Furthermore, lncRNA EWSAT1 knockdown significantly suppressed cell proliferation and invasion in CRC. In addition, lncRNA EWSAT1 knockdown suppressed cell epithelial-mesenchymal transition (EMT) through reducing Snail1, Snail2, and N-cadherin expression, but increasing E-cadherin expression in CRC cells. Downregulation of lncRNA EWSAT1 suppressed cell proliferation and invasion of CRC, which indicated that EWSAT1 may be a potential target of CRC treatment.

Lipocalin 2 inversely regulates TRAIL sensitivity through p38 MAPK-mediated DR5 regulation in colorectal cancer.

TNF-related apoptosis-inducing ligand (TRAIL) induces apoptosis through death receptors (DRs)4 and/or 5expressed on the cell surface. Multiple clinical trials are underway to evaluate the antitumor activity of recombinant human TRAIL and agonistic antibodies to DR4 or DR5. However, their therapeutic potential is limited by the high frequency of cancer resistance. In this study, we provide evidence demonstrating the role of lipocalin2 (LCN2) in the TRAIL-mediated apoptosis of human colorectal cancer(CRC). By analyzing the mRNA expression data of 71CRC tissues from patients, we found that DR5 was preferentially expressed in CRC tissues with a low LCN2 expression level compared to tissues with a high LCN2 expression level. Moreover, we analyzed the association between DR5 and LCN2 expression and this analysis revealed that DR5 expression in CRC tended to be inversely associated with LCN2 expression. By contrast, no association was found between the DR4 and LCN2 expression levels. The expression patterns of LCN2 in human CRC cell lines also exhibited an inverse association with DR5 expression. The knockdown of LCN2 by siRNA in the TRAIL‑resistant CRC cells expressing high levels of LCN2 led to a significant increase in TRAIL-induced apoptosis through the upregulation of DR5 protein and mRNA expression. The mechanism through which LCN2 silencing sensitized the CRC cells to TRAIL was dependent on the extrinsic pathway of apoptosis. In addition, we identified that the knockdown of LCN2 enhanced the sensitivity of the cells to TRAIL through the p38 MAPK/CHOP-dependent upregulation of DR5. Taken together, the findings of this study suggest that LCN2 is responsible for TRAIL sensitivity and LCN2 may thus prove to be a promising target protein in DR-targeted CRC therapy.

Targeting epigenetic regulation in colorectal cancer (CRC) liver metastases

Targeting epigenetic regulation in colorectal cancer (CRC) liver metastases

Abstract 3321: Dysregulation of m6A RNA methylation regulators in colorectal cancer: Clinical implication as prognostic biomarkers and potential therapeutic targets

Abstract Purpose Among more than 100 known post-transcriptional modifications, N6-Methyladenosine (m6A) represents the most prevalent internal modification in mammalian mRNAs. This epigenomic modification is orchestrated by a distinct group of enzymes and cofactors, categorized as writers, erasers and readers. Writers consist of a multicomponent complex consisting of at least three proteins, namely methyltransferase-like 3 (METTL3), METTL14, and Wilms' tumor 1-associating protein (WTAP1). These m6A marks are erased by two RNA demethylases, fat mass and obesity-associated protein (FTO) and alkylated DNA repair protein alkB homolog 5 (ALKBH5). Lastly, YTHDF1/2/3 family genes constitute the ‘readers' that preferentially bind to RNA harboring m6A modification. Accumulating evidence suggests that m6a RNA methylation plays a crucial role in cancer progression. Herein, we for the first time elucidate the role of m6a regulators in colorectal cancer (CRC), in order to gain insights into their functional role and potential clinical significance in this malignancy. Experimental design Two large independent publicly available genome-wide mRNA expression datasets (N=509, TCGA and N=566, CIT) were used to study the expression of m6a RNA methylation regulators and their association with prognosis and gene expression based Consensus Molecular Subtypes (CMS) in CRC. Furthermore, we studied the tumorigenic properties of the RNA demethylase, FTO, by knockdown and overexpression strategies in CRC cells. Results The risk-score derived from a seven gene mRNA expression classifier consisting of METTL3, METTL14, WTAP, YTHDF1, YTHDF2, FTO and ALKBH5 was associated with poor disease-free survival in the training (TCGA) and validation (CIT) cohorts, with corresponding hazard ratios of 1.90 (95% CI: 1.24-2.92, p&amp;lt;0.002) and 1.86 (95% CI: 1.25-2.78, p&amp;lt;0.001) respectively. Furthermore, a high-m6a risk-score was significantly associated with a mesenchymal, CMS4 subtype, with FTO being the most overexpressed gene in CMS4 subtype of CRCs. Functionally, knockdown of the FTO gene in CMS4 cell lines (MDST8 and NCIH716) dramatically inhibited cellular proliferation, invasion and colony formation abilities. In contrast, overexpression of FTO in RKO cells resulted in enhanced proliferation, invasion and colony formation. Conclusions In summary, we for the first time report the dysregulation of m6a RNA methylation regulators in CRC, wherein we discovered their significant association with poor prognosis and a CMS4 CRC subtype, as well as elucidated the functional role of FTO in CRC cell lines. Besides the potential of utilizing this seven gene m6a regulator gene expression classifier to predict prognosis, our data suggest that targeting FTO might represent an attractive strategy for therapeutic intervention in patients with a mesenchymal CRC subtype with poor prognosis. Citation Format: Raju Kandimalla, Feng Gao, Chun Ruan, Michael Hsieh, Xin Wang, Ajay Goel. Dysregulation of m6A RNA methylation regulators in colorectal cancer: Clinical implication as prognostic biomarkers and potential therapeutic targets [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3321.

Abstract 1472: The role of the BAF complex in Wnt signaling-mediated transcriptional regulation in colorectal cancer

Abstract Recent genome- and exome-wide sequencing studies have revealed a close association between the epigenome and the pathogenesis of cancer. Not only were chromatin regulators found to be commonly mutated in a wide range of cancers, but a class of these regulators, the ATP-dependent chromatin remodelers, especially subunits of the mammalian BAF complex, were among the most frequently mutated group of genes across cancer types, mutated in over 20% of cancers. Among these subunits, loss of function mutations in ARID1A (AT-rich interactive domain-containing protein 1A) are most frequent across cancer types including 10% of colorectal cancer (CRC). Very interestingly, a recent publication described the pivotal role of the loss of ARID1A in driving CRC wherein its inactivation alone led to the formation of invasive adenocarcinomas in mice. However, surprisingly, in contrast to the expected tumor suppressive role of ARID1A in CRC, we observe that the knockout (KO) of ARID1A in CRC cell lines leads to impaired proliferation. Moreover, subcutaneous xenografts in SCID/SHO mice using ARID1A KO CRC cells did not form more aggressive tumors than their wild type counterparts. One of the most commonly occurring mutations in CRC is in the APC gene, which leads to hyperactive Wnt signaling. Thus CRC progression is typically highly dependent on this pathway. Notably, it is reported that loss of ARID1A in the context of APC mutations results in diminished tumor formation in mice and increased differentiation. This led us to hypothesize that ARID1A is required for Wnt signaling-mediated transcriptional (de)regulation in CRC. To uncover this further, we utilized several publically available ChIP-seq, RNA-seq and ATAC-seq datasets as well as our own ChIP-seq data for ARID1A in the HCT116 cell line. We observe a substantial co-localization of the BAF complex with TCF7L2, a downstream effector of the Wnt signaling pathway, suggesting cooperation between these factors. Further, these sites are also co-occupied by AP1 transcription factors. A crosstalk between the TCF7L2 and AP1 factors has been reported in intestinal tumors. From data available in the ARID1A KO system, we observe a downregulation of potential target genes which are co-occupied at neighboring sites by these factors. Therefore, we suggest that ARID1A plays an important role in the regulation of colorectal cancer relevant genes that are targets of TCF7L2/AP1, thus facilitating a pathway that is most commonly hijacked in colorectal cancer. Citation Format: Madhobi Sen, Feda H. Hamdan, Xin Wang, Jacobe Rapp, Steven A. Johnsen. The role of the BAF complex in Wnt signaling-mediated transcriptional regulation in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1472.

Integrated omics profiling identifies hypoxia-regulated genes in HCT116 colon cancer cells

Hypoxia is associated with poor prognosis in most solid tumors due to its multiple effects on therapy resistance, angiogenesis, apoptotic resistance, and tumor invasion/metastasis. Here we used a comprehensive omics profiling to investigate hypoxia-regulated gene expression in HCT116 colon cancer cells. Quantitative analyses of proteome and secretome were performed in HCT116 cells cultured under hypoxic or normoxic conditions. A total of 5700 proteins were quantified in proteome analysis and 722 proteins were quantified in secretome analysis. Both datasets were combined with the transcriptome and translatome datasets for further analysis. Verification of candidate proteins/genes in this integrated omics analysis was performed using immunoblotting and quantitative real-time RT-PCR analyses. We also performed polysome profiling to assess changes in translational efficiency of hypoxia-induced genes. Notably, several genes were differently regulated at the transcriptional and translational levels in HCT116 cells during hypoxia. Bioinformatics analysis suggested that hypoxia regulates translation of genes involved in extracellular matrix organization, extracellular exosomes, and protein processing in endoplasmic reticulum. Aberrations in these metabolic pathways appear to be correlated with an increased risk of tumor invasion/metastasis. Biological significanceThis study integrates transcriptome/translatome and proteome/secretome to analyze gene expression changes in human colon cancer cells under hypoxic conditions. Candidate proteins/genes in this integrated omics analysis were further validated by immunoblotting, quantitative real-time RT-PCR, and polysome profiling. The datasets would be useful to uncover the molecular mechanisms of hypoxia-induced gene regulation in colorectal cancer.

MiR-206 regulates 5-FU resistance by targeting Bcl-2 in colon cancer cells.

IntroductionPrevious studies have found that miRNAs play a key role in drug resistance. Multiple reports show that miRNAs act as regulators in colorectal cancer (CRC) cells, but the role of miR-206 in CRC is still not well understood. The current study aimed to explore the potential function of miR-206 in 5-FU resistance.MethodsTo indentify the role of miR-206 in 5-FU resistance, the expression of miR-206 was examined by real-time polymerase chain reaction (RT-PCR) in 5-FU-resistant (FR) CRC (HCT116/FR and RKO/FR) and their parental cell lines. miR-206 mimic was transfected to 5-FU-FR CRC, and the 5-FU sensitivity was detected by MTS and flow cytometry. Using miRNA target prediction software, we found that miR-206 could target the 3′ untranslated region (3′UTR) sequence of Bcl-2.ResultsmiR-206 was found to be downregulated in 5-FU-FR CRC in comparison with their parental cell lines, suggesting its crucial relevance for colon cancer biology. Downregulation of miR-206 promoted drug resistance and decreased apoptosis of parental cells, while overexpression of miR-206 promoted drug cytotoxicity and apoptosis of HCT116/FR cells. We also identified miR-206 targeting Bcl-2 directly in CRC, which is required for miR-206 mediated-5-FU resistance.ConclusionOur results show that miR-206 targets Bcl-2 to mediate chemoresistance, proliferation, and apoptosis in CRC. This study provides a novel promising candidate for colon cancer therapy.

Micro-managers of disease: the long and short of miRNA regulation in colorectal cancer

The expression of non-coding RNAs is often altered in cancer cells. In particular, microRNAs (miRNAs) have progressively emerged as instrumental regulators of cancer progression, metastatic development, cancer stem cell properties and treatment response (1).

Tumor-suppressive miR-145 co-repressed by TCF4-β-catenin and PRC2 complexes forms double-negative regulation loops with its negative regulators in colorectal cancer.

The frequently dysregulated Wnt/β-catenin signaling in different malignancies, by activation of its own or orchestration with other co-factors, regulates various oncogenic or tumor-suppressive genes. Among these genes, miRNAs, which are negative posttranscriptional regulators, are also embedded in the Wnt signaling network. Different from the Wnt-induced oncogenic miRNAs, the specific mechanism underlying the Wnt-repressed tumor-suppressive miRNAs is much less understood. In our study, firstly by analyzing a ChIP-seq dataset against TCF4, the core transcription factor for initiation of Wnt signaling in colorectal cancer (CRC) cells, we screened out several tumor-suppressive miRNAs potentially regulated by Wnt signaling. Then through siRNA-mediated knock-down tests and protein and chromatin immunoprecipitations, we found the TCF4-β-catenin complex can recruit the histone trimethylation complex PRC2 as a co-repressor while binding to the TCF4-binding element (TBE) in the promoter regions of miR-145, miR-132 and miR-212. Thus, upon Wnt signaling activation, the PRC2-mediated trimethylation of histone H3 at lysine 27 increases at these promoter regions, leading to decreased miRNA levels. Furthermore, we found that by targeting TCF4 and SUZ12, the key components of the negative regulation complexes, the tumor-suppressive miR-145 co-repressed by Wnt signaling and histone trimethylation, forms double-negative regulation loops with its negative regulators in CRC cells. And the inverse associations between miR-145 and its targets/negative regulators have also been demonstrated in nude mice and clinical samples. Collectively, we elucidated the detailed molecular mechanism of how dysregulated Wnt/β-catenin signaling and tumor-suppressive miRNAs reciprocally regulate each other in CRC cells.

E2F1 somatic mutation within miRNA target site impairs gene regulation in colorectal cancer.

BackgroundGenetic studies have largely concentrated on the impact of somatic mutations found in coding regions, and have neglected mutations outside of these. However, 3’ untranslated regions (3' UTR) mutations can also disrupt or create miRNA target sites, and trigger oncogene activation or tumor suppressor inactivation.MethodsWe used next-generation sequencing to widely screen for genetic alterations within predicted miRNA target sites of oncogenes associated with colorectal cancer, and evaluated the functional impact of a new somatic mutation. Target sequencing of 47 genes was performed for 29 primary colorectal tumor samples. For 71 independent samples, Sanger methodology was used to screen for E2F1 mutations in miRNA predicted target sites, and the functional impact of these mutations was evaluated by luciferase reporter assays.ResultsWe identified germline and somatic alterations in E2F1. Of the 100 samples evaluated, 3 had germline alterations at the MIR205-5p target site, while one had a somatic mutation at MIR136-5p target site. E2F1 gene expression was similar between normal and tumor tissues bearing the germline alteration; however, expression was increased 4-fold in tumor tissue that harbored a somatic mutation compared to that in normal tissue. Luciferase reporter assays revealed both germline and somatic alterations increased E2F1 activity relative to wild-type E2F1.ConclusionsWe demonstrated that somatic mutation within E2F1:MIR136-5p target site impairs miRNA-mediated regulation and leads to increased gene activity. We conclude that somatic mutations that disrupt miRNA target sites have the potential to impact gene regulation, highlighting an important mechanism of oncogene activation.

Identification of type I and type II inhibitors of c-Yes kinase using in silico and experimental techniques

c-Yes kinase is considered as one of the attractive targets for anti-cancer drug design. The DFG (Asp-Phe-Gly) motif present in most of the kinases will adopt active and inactive conformations, known as DFG-in and DFG-out and their inhibitors are classified into type I and type II, respectively. In the present study, two screening protocols were followed for identification of c-Yes kinase inhibitors. (i) Structure-based virtual screening (SBVS) and (ii) Structure-based (SB) and Pharmacophore-based (PB) tandem screening. In SBVS, the c-Yes kinase structure was obtained from homology modeling and seven ensembles with different active site scaffolds through molecular dynamics (MD) simulations. For SB-PB tandem screening, we modeled ligand bound active and inactive conformations. Physicochemical properties of inhibitors of Src kinase family and c-Yes kinase were used to prepare target focused libraries for screenings. Our screening procedure along with docking showed 520 probable hits in SBVS and tandem screening (120 and 400, respectively). Out of 5000 compounds identified from different computational methods, 2410 were examined using kinase inhibition assays. It includes 266 compounds (5.32%) identified from our method. We observed that 14 compounds (12%) are identified by the present method out of 168 that showed > 30% inhibition. Among them, three compounds are novel, unique, and showed good inhibition. Further, we have studied the binding of these compounds at the DFG-in and DFG-out conformations and reported the probable class (type I or type II). Hence, we suggest that these compounds could be novel drug leads for regulation of colorectal cancer.

Rho GDP-dissociation inhibitor α is a potential prognostic biomarker and controls telomere regulation in colorectal cancer.

Rho GDP‐dissociation inhibitor α (RhoGDIα) is an essential regulator for Rho GTPases. Although RhoGDIα may serve as an oncogene in colorectal cancer (CRC), the underlying mechanism is still unclear. We investigated the function, mechanism, and clinical significance of RhoGDIα in CRC progression. We founded that downregulation of RhoGDIα repressed CRC cell proliferation, motility, and invasion. Overexpression of RhoGDIα increased DNA damage response signals at telomeres, and led to telomere shortening in CRC cells, also being validated in 26 pairs of CRC tissues. Mechanistic studies revealed that RhoGDIα could promote telomeric repeat factor 1 (TRF1) expression through the phosphatidylinositol 3‐kinase–protein kinase B signal pathway. Moreover, RhoGDIα protein levels were strongly correlated with TRF1 in CRC tissues. A cohort of 297 CRC samples validated the positive relationship between RhoGDIα and TRF1, and revealed that RhoGDIα and TRF1 levels were negatively associated with CRC patients' survival. Taken together, our results suggest that RhoGDIα regulate TRF1 and telomere length and may be novel prognostic biomarkers in colorectal cancer.

Identification of differentially expressed genes and their upstream regulators in colorectal cancer.

To identify the differentially expressed genes (DEGs) and their transcription factors (TFs) in colorectal cancer (CRC). We performed an integrated analysis of microarray studies. Functional annotation and CRC-specific transcriptional regulatory network construction were performed. Expression of selected DEGs and TFs was verified with The Cancer Genome Atlas (TCGA) data sets and qRT-PCR. SurvMicro was used to analyze the correlation between the overall survival time of CRC patients and the expression of DEGs and TFs. Seven data sets were obtained and 2014 DEGs in CRC were identified. Pathways in cancer and fatty acid metabolism were significantly enriched pathways of upregulated and downregulated DEGs, respectively. Expression of five DEGs (RERGL, ESM1, CA1, ANGPTL7 and TMEFF2) and their five TFs (ZNF354C, ARID3A, NFIC, BRCA1 and ZEB1) was verified by TCGA data sets and qRT-PCR. Their expression in TCGA data sets was same as that in our integrated analysis. Only the expression of EMS1, NFIC, BRCA1 and ZEB1 was inconsistent with integrated analysis and TCGA data sets. Expression of RERGL and BRCA1 was significantly correlated with the overall survival time of CRC patients. These five DEGs may have roles in CRC regulated by their five upstream TFs, which may make a contribution in uncovering the mechanism and providing new strategy of diagnosis and therapies for CRC.

MicroRNAs in the etiology of colorectal cancer: pathways and clinical implications.

ABSTRACTMicroRNAs (miRNAs) are small single-stranded RNAs that repress mRNA translation and trigger mRNA degradation. Of the ∼1900 miRNA-encoding genes present in the human genome, ∼250 miRNAs are reported to have changes in abundance or altered functions in colorectal cancer. Thousands of studies have documented aberrant miRNA levels in colorectal cancer, with some miRNAs reported to actively regulate tumorigenesis. A recurrent phenomenon with miRNAs is their frequent participation in feedback loops, which probably serve to reinforce or magnify biological outcomes to manifest a particular cellular phenotype. Here, we review the roles of oncogenic miRNAs (oncomiRs), tumor suppressive miRNAs (anti-oncomiRs) and miRNA regulators in colorectal cancer. Given their stability in patient-derived samples and ease of detection with standard and novel techniques, we also discuss the potential use of miRNAs as biomarkers in the diagnosis of colorectal cancer and as prognostic indicators of this disease. MiRNAs also represent attractive candidates for targeted therapies because their function can be manipulated through the use of synthetic antagonists and miRNA mimics.

MicroRNA-375 inhibits colorectal cancer cells proliferation by downregulating JAK2/STAT3 and MAP3K8/ERK signaling pathways.

MicroRNA-375 is involved in many types of alimentary system cancers. Our previous studies showed that microRNA-375 was significantly down-regulated in carcinoma tissues compared with para-carcinoma tissues, which strongly indicates that microRNA-375 might suppress the occurrence and development of colorectal cancer. However, the mechanism underlying the microRNA-375 regulation in colorectal cancer remains unclear. In this study, we first sorted out jak2, map3k8 and atg7 as microRNA-375 targeted genes from multiple databases, and found that jak2, map3k8 and their downstream genes stat3 and erk were up-regulated in carcinoma tissues. Secondly, we over-expressed microRNA-375 in colorectal cancer cell lines (HCT116, Caco2 and HT29). Our results showed that in microRNA-375 over-expressing cells, JAK2/STAT3 and MAP3K8/ERK proteins were down-regulated, cell proliferation was inhibited, cell migration rate did not change. There was no significant difference on ATG7 expression between the control group and microRNA-375 over-expressing HT29/Caco2 cells, whereas microRNA-375 down-regulated ATG7 specifically in HCT116 cells. Finally, we demonstrated that expressing microRNA-375 suppressed tumor formation in nude mice. In conclusion, microRNA-375 might function as a tumor-repressive gene to inhibit cell proliferation, mainly through targeting both JAK2/STAT3 and MAP3K8/ERK signaling pathways in colorectal cancer. These findings suggest miR-375 as a promising diagnostic marker and a therapeutic drug for colorectal cancer.

Mechanistic evaluation and transcriptional signature of a glutathione S-transferase omega 1 inhibitor.

Glutathione S-transferase omega 1 (GSTO1) is an atypical GST isoform that is overexpressed in several cancers and has been implicated in drug resistance. Currently, no small-molecule drug targeting GSTO1 is under clinical development. Here we show that silencing of GSTO1 with siRNA significantly impairs cancer cell viability, validating GSTO1 as a potential new target in oncology. We report on the development and characterization of a series of chloroacetamide-containing potent GSTO1 inhibitors. Co-crystal structures of GSTO1 with our inhibitors demonstrate covalent binding to the active site cysteine. These potent GSTO1 inhibitors suppress cancer cell growth, enhance the cytotoxic effects of cisplatin and inhibit tumour growth in colon cancer models as single agent. Bru-seq-based transcription profiling unravelled novel roles for GSTO1 in cholesterol metabolism, oxidative and endoplasmic stress responses, cytoskeleton and cell migration. Our findings demonstrate the therapeutic utility of GSTO1 inhibitors as anticancer agents and identify the novel cellular pathways under GSTO1 regulation in colorectal cancer.