Background & Aims Nearly 20% of the global cancer burden can be linked to infectious agents. Fusobacterium nucleatum promotes tumor formation by epithelial cells via unclear mechanisms. We aimed to identify microRNAs (miRNAs) induced by F nucleatum and evaluate their ability to promote colorectal carcinogenesis in mice. Methods Colorectal cancer (CRC) cell lines were incubated with F nucleatum or control reagents and analyzed in proliferation and would healing assays. HCT116, HT29, LoVo, and SW480 CRC cell lines were incubated with F nucleatum or phosphate buffer saline (PBS control) and analyzed for miRNA expression patterns and in chromatin immunoprecipitation assays. Cells were incubated with miRNAs mimics, control sequences, or small interfering (si) RNAs; expression of reporter constructs was measured in luciferase assays. CRC cells were incubated with F nucleatum or PBS and injected into BALB/C nude mice; growth of xenograft tumors was measured. C57BL APCmin/+, C57BL miR21a−/−, and C57BL mice with full-length miR21a (controls) were given F nucleatum by gavage; some mice were given azoxymethane (AOM) and dextran sodium sulfate (DSS) to induce colitis and colon tumors. Intestinal tissues were collected and tumors were counted. Serum samples from mice were analyzed for cytokine levels by ELISAs. We performed in situ hybridization analyses to detect enrichment of F nucleatum in CRC cells. F nucleatum DNA in 90 tumor and matched non-tumor tissues from patients in China were explored for the expression correlation analysis; levels in 125 tumor tissues from patients in Japan were compared with their survival times. Results F nucleatum increased proliferation and invasive activities of CRC cell lines, compared with control cells. CRC cell lines infected with F nucleatum formed larger tumors, more rapidly, in nude mice than uninfected cells. APCmin/+ mice gavaged with F nucleatum developed significantly more colorectal tumors than mice given PBS and had shorter survival times. We found several inflammatory factors to be significantly increased in serum from mice given F nucleatum (interleukin 17F [IL17F], IL21, IL22, and MIP3A). We found 50 miRNAs to be significantly upregulated and 52 miRNAs to be significantly downregulated in CRCs incubated with F nucleatum vs PBS; levels of miR21 increased by the greatest amount (more than 4-fold). Inhibitors of miR21 prevented F nucleatum from inducing cell proliferation and invasion in culture. miR21a−/− mice had a later appearance of fecal blood and diarrhea after administration of AOM and DSS, and had longer survival times, compared with control mice. The colorectum of miR21a−/− mice had fewer tumors, of smaller size, and the miR21a−/− mice survived longer than control mice. We found RASA1, which encodes a RAS GTPase, to be one of the target genes consistently downregulated in cells that overexpressed miR21 and upregulated in cells exposed to miR21 inhibitors. Infection of cells with F nucleatum increased expression of miR21 by activating TLR4 signaling to MYD88, leadi...

Purpose Colorectal cancer (CRC) is one of the most common malignancies worldwide. Recently, a novel circular RNA, ciRS-7, was proposed to be a potential miR-7 sponge. Since miR-7 regulates the expression of several important drivers of CRC, we analyzed the clinical significance of ciRS-7 in CRC patients. Experimental Design Initially, we evaluated the expression levels of ciRS-7 in a training cohort comprising of 153 primary CRC tissues and 44 matched normal mucosae. We subsequently confirmed its clinical relevance in an independent validation cohort (n=165), and evaluated the effect of ciRS-7 on miR-7, and its targets genes EGFR and RAF1. Functional analysis were performed in cell lines and an animal model to support clinical findings. Results Our data revealed that ciRS-7 was significantly up-regulated in CRC tissues compared with matched normal mucosae (P=0.0018), and its overexpression was associated with poor patient survival (P=0.0224 and 0.0061 in the training and validation cohorts, respectively). Multivariate survival analysis revealed that ciRS-7 emerged as an independent risk factor for overall survival (P=0.0656 and 0.0324 in the training and validation cohorts, respectively). Overexpression of ciRS-7 in HCT116 and HT29 cells led to blocking of tumor suppressive effects of miR-7 and resulted in a more aggressive oncogenic phenotype, and ciRS-7 overexpression permitted inhibition of miR-7 and subsequent activation of EGFR and RAF1 oncogenes. Conclusions CiRS-7 is a promising prognostic biomarker in CRC patients and may serve as a therapeutic target for reducingEGFR-RAF1 activity in CRC patients.

Rationale : Colorectal cancer (CRC) is a malignant tumor with the third highest morbidity rate among all cancers. Driven by the host's genetic makeup and environmental exposures, the gut microbiome and its metabolites have been implicated as the causes and regulators of CRC pathogenesis. We assessed human fecal samples as noninvasive and unbiased surrogates to catalog the gut microbiota and metabolome in patients with CRC. Methods : Fecal samples collected from CRC patients (CRC group, n = 50) and healthy volunteers (H group, n = 50) were subjected to microbiome (16S rRNA gene sequencing) and metabolome (gas chromatography-mass spectrometry, GC-MS) analyses. The datasets were analyzed individually and integrated for combined analysis using various bioinformatics approaches. Results : Fecal metabolomic analysis led to the identification of 164 metabolites spread across 40 metabolic pathways in both groups. In addition, there were 42 and 17 metabolites specific to the H and CRC groups, respectively. Sequencing of microbial diversity revealed 1084 operational taxonomic units (OTUs) across the two groups, and there was less species diversity in the CRC group than in the H group. Seventy-six discriminatory OTUs were identified for the microbiota of H volunteers and CRC patients. Integrated analysis correlated CRC-associated microbes with metabolites, such as polyamines (cadaverine and putrescine). Conclusions : Our results provide substantial evidence of a novel interplay between the gut microbiome and metabolome (i.e., polyamines), which is drastically perturbed in CRC. Microbe-associated metabolites can be used as diagnostic biomarkers in therapeutic explorations.

BackgroundEmerging evidence suggests a potential relationship between gut microbiota and the host response to chemotherapeutic drugs including 5-fluorouracil (5-Fu). Fusobacterium nucleatum (Fn) has been linked to the initiation and progression of colorectal cancer (CRC). Unfortunately, little was known about the relationship between Fn infection and chemotherapeutic efficacy. Here, we investigate the potential relationship between Fn infection and chemotherapeutic efficacy of 5-Fu in CRC.MethodsDifferentially expressed genes of CRC cell lines induced by Fn infection were analyzed based on a whole genome microarray analysis Then, we explored the relationship between upregulation of BIRC3 induced by Fn infection and chemoresistance to 5-Fu in vitro and in vivo. Furthermore, we dissected the mechanisms involved in Fn-induced BIRC3 expression. Finally, we investigated the clinical relevance of Fn infection, BIRC3 protein expression and chemoresistance to 5-Fu treatment in CRC patients.ResultsBIRC3 was the most upregulated gene induced by Fn infection via the TLR4/NF-κB pathway in CRC cells; Fn infection reduced the chemosensitivity of CRC cells to 5-Fu through upregulation of BIRC3 in vitro and in vivo. High Fn abundance correlated with chemoresistance in advanced CRC patients who received standard 5-Fu-based adjuvant chemotherapy after radical surgery.ConclusionsOur evidence suggests that Fn and BIRC3 may serve as promising therapeutic targets for reducing chemoresistance to 5-Fu treatment in advanced CRC.Electronic supplementary materialThe online version of this article (10.1186/s13046-018-0985-y) contains supplementary material, which is available to authorized users.

Yes-associated protein (YAP), the downstream effecter of the Hippo-signaling pathway as well as cyclic adenosine monophosphate response element-binding protein (CREB), has been linked to hepatocarcinogenesis. However, little is known about whether and how YAP and CREB interact with each other. In this study, we found that YAP-CREB interaction is critical for liver cancer cell survival and maintenance of transformative phenotypes, both in vitro and in vivo. Moreover, both CREB and YAP proteins are highly expressed in a subset of human liver cancer samples and are closely correlated. Mechanistically, CREB promotes YAP transcriptional output through binding to 2608/ 2439, a novel region from the YAP promoter. By contrast, YAP promotes protein stabilization of CREB through interaction with mitogen-activated protein kinase 14 (MAPK14/p38) and beta-transducin repeat containing E3 ubiquitin protein ligase (BTRC). Gain-of-function and loss-of-function studies demonstrated that phosphorylation of CREB by MAPK14/p38 at ser133 ultimately leads to its degradation. Such effects can be enhanced by BTRC through phosphorylation of MAPK14/p38 at Thr180/Tyr182. However, YAP negatively controls phosphorylation of MAPK14/p38 through inhibition of BTRC expression. Conclusion: There is a novel positive autoregulatory feedback loop underlying the interaction between YAP and CREB in liver cancer, suggesting that YAP and CREB form a nexus to integrate the protein kinase A, Hippo/ YAP, and MAPK14/p38 pathways in cancer cells and thus may be helpful in the development of effective diagnosis and treatment strategies against liver cancer. (HEPATOLOGY 2013;58:1011-1020 L iver cancer is the fifth-most common cancer worldwide and the third-leading cause of cancer death. 1 The treatment options for these hepatic malignancies are extremely limited, mainly because the mechanisms of pathogenesis of these cancers are not completely known. Recently, the dysfunctional Hippo/ Yes-associated protein (YAP)-signaling pathway has been linked to hepatocarcinogenesis. 2 Transgenic mice with liver-targeted YAP overexpression demonstrated a dramatic increase in liver size and eventually developed tumors. 3 In addition, clinical studies revealed that YAP was overexpressed in 62% of hepatocellular carcinoma (HCC) patients and was an independent predictor associated with poor disease-free survival and overall

BackgroundEmerging evidence suggests that PIWI-interacting RNAs (piRNAs) may be important epigenetic regulators of gene expression in human cancers; however, their functional and clinical significance in colorectal cancer (CRC) remains unknown.MethodsWe performed piRNA expression profiling in paired cancer and normal tissues through small RNA-sequencing. The clinical significance of candidate piRNAs was investigated, and independently validated in 771 CRC patients from three independent cohorts. The biological function of piRNAs was characterized in cell lines, followed by identification and validation of downstream target genes in CRC tissues.ResultsWe identified piR-1245 as a novel and frequently overexpressed noncoding RNA in CRC, and its expression significantly correlated with advanced and metastatic disease. Patients with high piR-1245 expression experienced significantly shorter overall survival, and multivariate analysis identified its expression to serve as an independent prognostic biomarker in CRC. Functionally, piR-1245 acts as an oncogene and promotes tumor progression, and gene expression profiling results identified a panel of downstream target-genes involved in regulating cell survival pathway. Based upon piRNA:mRNA sequence complementarity, we identified a panel of tumor suppressor genes (ATF3, BTG1, DUSP1, FAS,NFKBIA, UPP1, SESN2, TP53INP1 and MDX1) as direct targets of piR-1245, and successfully validated an inverse correlation between their expression and piR-1245 in CRC.ConclusionsWe for the first time have identified the role for a PIWI-interacting noncoding RNA, piR-1245, as a novel oncogene and a potential prognostic biomarker in colorectal cancer.Electronic supplementary materialThe online version of this article (10.1186/s12943-018-0767-3) contains supplementary material, which is available to authorized users.

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