In most carcinomas, invasion of malignant cells into surrounding tissues involves their molecular reprogramming as part of an epithelial-to-mesenchymal transition (EMT). Mutation of the APC gene in ...most colorectal carcinomas (CRCs) contributes to the nuclear translocation of the oncoprotein β-catenin that upon binding to T-cell and lymphoid enhancer (TCF-LEF) factors triggers an EMT and a proinvasive gene expression profile. A key inducer of EMT is the ZEB1 transcription factor whose expression promotes tumorigenesis and metastasis in carcinomas. As inhibitor of the epithelial phenotype, ZEB1 is never present in the epithelium of normal colon or the tumor center of CRCs where β-catenin remains membranous. We show here that ZEB1 is expressed by epithelial cells in intestinal tumors from human patients (familial adenomatous polyposis) and mouse models (APCMin/+) with germline mutations of APC that result in nuclear accumulation of β-catenin. However, ZEB1 is not expressed in the epithelium of hereditary forms of CRCs that carry wild-type APC and where β-catenin is excluded from the nucleus (Lynch syndrome). We found that β-catenin/TCF4 binds directly to the ZEB1 promoter and activates its transcription. Knockdown of β-catenin and TCF4 in APC-mutated CRC cells inhibited endogenous ZEB1, whereas forced translocation of β-catenin to the nucleus in APC-wild-type CRC cells induced de novo expression of ZEB1. Upregulation of MT1-MMP and LAMC2 by β-catenin/TCF4 has been linked to invasiveness in CRCs, and we show here that both proteins are activated by ZEB1 coexpressing with it in primary colorectal tumors with mutated APC. These results set ZEB1 as an effector of β-catenin/TCF4 signaling in EMT and tumor progression.
Development of robust prognostic and/or predictive biomarkers in patients with colorectal cancer (CRC) is imperative for advancing treatment strategies for this disease. We aimed to determine whether ...expression status of certain miRNAs might have prognostic/predictive value in CRC patients treated with conventional cytotoxic chemotherapies.
We studied a cohort of 273 CRC specimens from stage II/III patients treated with 5-fluorouracil-based adjuvant chemotherapy and stage IV patients subjected to 5-fluorouracil and oxaliplatin-based chemotherapy. In a screening set (n = 44), 13 of 21 candidate miRNAs were successfully quantified by multiplex quantitative RT-PCR. In the validation set comprising of the entire patient cohort, miR-148a expression status was assessed by quantitative RT-PCR, and its promoter methylation was quantified by bisulfite pyrosequencing. Lastly, we analyzed the associations between miR-148a expression and patient survival.
Among the candidate miRNAs studied, miR-148a expression was most significantly down-regulated in advanced CRC tissues. In stage III and IV CRC, low miR-148a expression was associated with significantly shorter disease free-survival (DFS), a worse therapeutic response, and poor overall survival (OS). Furthermore, miR-148a methylation status correlated inversely with its expression, and was associated with worse survival in stage IV CRC. In multivariate analysis, miR-148a expression was an independent prognostic/predictive biomarker for advanced CRC patients (DFS in stage III, low vs. high expression, HR 2.11; OS in stage IV, HR 1.93).
MiR-148a status has a prognostic/predictive value in advanced CRC patients treated with conventional chemotherapy, which has important clinical implications in improving therapeutic strategies and personalized management of this malignancy.
Cancer is a complex multistep process involving genetic and epigenetic changes that eventually result in the activation of oncogenic pathways and/or inactivation of tumor suppressor signals. During ...cancer progression, cancer cells acquire a number of hallmarks that promote tumor growth and invasion. A crucial mechanism by which carcinoma cells enhance their invasive capacity is the dissolution of intercellular adhesions and the acquisition of a more motile mesenchymal phenotype as part of an epithelial-to-mesenchymal transition (EMT). Although many transcription factors can trigger it, the full molecular reprogramming occurring during an EMT is mainly orchestrated by three major groups of transcription factors: the ZEB, Snail and Twist families. Upregulated expression of these EMT-activating transcription factors (EMT-ATFs) promotes tumor invasiveness in cell lines and xenograft mice models and has been associated with poor clinical prognosis in human cancers. Evidence accumulated in the last few years indicates that EMT-ATFs also regulate an expanding set of cancer cell capabilities beyond tumor invasion. Thus, EMT-ATFs have been shown to cooperate in oncogenic transformation, regulate cancer cell stemness, override safeguard programs against cancer like apoptosis and senescence, determine resistance to chemotherapy and promote tumor angiogenesis. This article reviews the expanding portfolio of functions played by EMT-ATFs in cancer progression.
Global downregulation of microRNAs (miRNA) is a common feature in colorectal cancer (CRC). Whereas CpG island hypermethylation constitutes a mechanism for miRNA silencing, this field largely remains ...unexplored. Herein, we describe the epigenetic regulation of miR-137 and its contribution to colorectal carcinogenesis. We determined the methylation status of miR-137 CpG island in a panel of six CRC cell lines and 409 colorectal tissues 21 normal colonic mucosa from healthy individuals (N-N), 160 primary CRC tissues and their corresponding normal mucosa (N-C), and 68 adenomas. TaqMan reverse transcription-PCR and in situ hybridization were used to analyze miR-137 expression. In vitro functional analysis of miR-137 was performed. Gene targets of miR-137 were identified using a combination of bioinformatic and transcriptomic approaches. We experimentally validated the miRNA:mRNA interactions. Methylation of the miR-137 CpG island was a cancer-specific event and was frequently observed in CRC cell lines (100%), adenomas (82.3%), and CRC (81.4%), but not in N-C (14.4%; P < 0.0001 for CRC) and N-N (4.7%; P < 0.0001 for CRC). Expression of miR-137 was restricted to the colonocytes in normal mucosa and inversely correlated with the level of methylation. Transfection of miR-137 precursor in CRC cells significantly inhibited cell proliferation. Gene expression profiling after miR-137 transfection discovered novel potential mRNA targets. We validated the interaction between miR-137 and LSD-1. Our data indicate that miR-137 acts as a tumor suppressor in the colon and is frequently silenced by promoter hypermethylation. Methylation silencing of miR-137 in colorectal adenomas suggests it to be an early event, which has prognostic and therapeutic implications.
Let-7 miRNAs comprise one of the largest and most highly expressed family of miRNAs among vertebrates, and is critical for promoting differentiation, regulating metabolism, inhibiting cellular ...proliferation, and repressing carcinogenesis in a variety of tissues. The large size of the Let-7 family of miRNAs has complicated the development of mutant animal models. Here we describe the comprehensive repression of all Let-7 miRNAs in the intestinal epithelium via low-level tissue-specific expression of the Lin28b RNA-binding protein and a conditional knockout of the MirLet7c-2/Mirlet7b locus. This ablation of Let-7 triggers the development of intestinal adenocarcinomas concomitant with reduced survival. Analysis of both mouse and human intestinal cancer specimens reveals that stem cell markers were significantly associated with loss of Let-7 miRNA expression, and that a number of Let-7 targets were elevated, including Hmga1 and Hmga2. Functional studies in 3-D enteroids revealed that Hmga2 is necessary and sufficient to mediate many characteristics of Let-7 depletion, namely accelerating cell cycle progression and enhancing a stem cell phenotype. In addition, inactivation of a single Hmga2 allele in the mouse intestine epithelium significantly represses tumorigenesis driven by Lin28b. In aggregate, we conclude that Let-7 depletion drives a stem cell phenotype and the development of intestinal cancer, primarily via Hmga2.
Patients with Crohn disease commonly have bowel strictures develop, which exhibit varying degrees of inflammation and fibrosis. Differentiation of the distinct inflammatory and fibrotic components of ...strictures is key for the optimization of therapeutic management and for the development of antifibrotic drugs. Cross-sectional imaging techniques, including ultrasound, CT, and MRI, allow evaluation of the full thickness of the bowel wall as well as extramural complications and associated mesenteric abnormalities. Although promising data have been reported for a range of novel imaging biomarkers for detection of fibrosis and quantification of the degree of fibrosis, these biomarkers lack sufficient validation and standardization for clinical use. Additional methods, including PET with emerging radiotracers, artificial intelligence, and radiomics, are also under investigation for stricture characterization. In this review, we highlight the clinical relevance of identifying fibrosis in Crohn disease, review the histopathologic aspects of strictures in Crohn disease, summarize the morphologic imaging findings of strictures, and explore contemporary developments in the use of cross-sectional imaging techniques for detecting and characterizing intestinal strictures, with attention given to emerging quantitative biomarkers.
Measurement of the component of fibrosis in Crohn's disease (CD) may have important therapeutic implications. The aim of this study was to characterize the Magnetic Resonance Imaging (MRI) findings ...that are differentially associated with the presence of fibrosis and those associated with inflammatory activity, using the pathological analysis of surgically resected intestinal lesions as reference standard.
MRI studies with identical imaging protocol of 41 CD patients who underwent elective bowel resection within 4 months before surgery were reviewed. MRI evaluated wall thickening, edema, ulcers, signal intensity at submucosa at 70 s and 7 min after gadolinium injection, stenosis, and pattern of enhancement in each phase of the dynamic study and changes on this pattern over time. Pathological inflammatory and fibrosis scores were classified into three grades of severity.
In all, 44 segments from 41 patients were analyzed. The pathological intensity of inflammation was associated with the following MRI parameters: hypersignal on T2 (P=0.02), mucosal enhancement (P=0.03), ulcerations (P=0.01), and blurred margins (P=0.05). The degree of fibrosis correlated with the percentage of enhancement gain (P<0.01), the pattern of enhancement at 7 min (P<0.01), and the presence of stenosis (P=0.05). Using percentage of enhancement gain, MRI is able to discriminate between mild-moderate and severe fibrosis deposition with a sensitivity of 0.94 and a specificity of 0.89.
MRI is accurate for detecting the presence of severe fibrosis in CD lesions on the basis of the enhancement pattern.
Early-onset colorectal cancer (CRC) represents a clinically distinct form of CRC that is often associated with a poor prognosis. Methylation levels of genomic repeats such as LINE-1 elements have ...been recognized as independent factors for increased cancer-related mortality. The methylation status of LINE-1 elements in early-onset CRC has not been analyzed previously.
We analyzed 343 CRC tissues and 32 normal colonic mucosa samples, including 2 independent cohorts of CRC diagnosed ≤ 50 years old (n=188), a group of sporadic CRC >50 years (MSS n=89; MSI n=46), and a group of Lynch syndrome CRCs (n=20). Tumor mismatch repair protein expression, microsatellite instability status, LINE-1 and MLH1 methylation, somatic BRAF V600E mutation, and germline MUTYH mutations were evaluated.
Mean LINE-1 methylation levels (± SD) in the five study groups were early-onset CRC, 56.6% (8.6); sporadic MSI, 67.1% (5.5); sporadic MSS, 65.1% (6.3); Lynch syndrome, 66.3% (4.5) and normal mucosa, 76.5% (1.5). Early-onset CRC had significantly lower LINE-1 methylation than any other group (p<0.0001). Compared to patients with <65% LINE-1 methylation in tumors, those with ≥ 65% LINE-1 methylation had significantly better overall survival (p=0.026, log rank test).
LINE-1 hypomethylation constitutes a potentially important feature of early-onset CRC, and suggests a distinct molecular subtype. Further studies are needed to assess the potential of LINE-1 methylation status as a prognostic biomarker for young people with CRC.
LIN28B is a homologue of LIN28 that induces pluripotency when expressed in conjunction with OCT4, SOX2, and KLF4 in somatic fibroblasts. LIN28B represses biogenesis of let-7 microRNAs and is ...implicated in both development and tumorigenesis. Recently, we have determined that LIN28B overexpression occurs in colon tumors. We conducted a comprehensive analysis of LIN28B protein expression in human colon adenocarcinomas. We found that LIN28B overexpression correlates with reduced patient survival and increased probability of tumor recurrence. To elucidate tumorigenic functions of LIN28B, we constitutively expressed LIN28B in colon cancer cells and evaluated tumor formation in vivo. Tumors with constitutive LIN28B expression exhibit increased expression of colonic stem cell markers LGR5 and PROM1, mucinous differentiation, and metastasis. Together, our findings point to a function for LIN28B in promoting colon tumor pathogenesis, especially metastasis.
Background Narrow-band imaging (NBI) is a novel technique that may represent an alternative method to chromoendoscopy (CE) for the detection of colitis-associated intraepithelial neoplasia (IN) in ...patients with long-standing inflammatory bowel disease (IBD). Objective To compare NBI with CE for the detection of IN. Design Prospective, randomized, crossover study. Setting Academic hospital. Patients Patients with clinically inactive colonic IBD (≥8 years). Intervention Patients underwent both CE and NBI in randomized order. Targeted biopsy specimens from abnormal areas were obtained. Pathological examination was regarded as the reference standard. Main Outcome Measurements Number of false-positive and true-positive lesions in patients undergoing CE and NBI were compared as well as the proportion of patients with missed IN lesions. Results Eighty patients were screened, of whom 20 were excluded. Mean ± standard deviation withdrawal time for CE was significantly longer than that for NBI (26.87 ± 9.89 minutes vs 15.74 ± 5.62 minutes, P < .01). Thirteen patients had at least 1 IN lesion on 1 of the examinations. In the per-lesion analysis, NBI resulted in a significantly inferior false-positive biopsy rate ( P = .001) and a similar true-positive rate. The percentage of missed IN lesions and patients was superior with NBI, albeit without reaching statistical significance. Limitations Lesions were sampled immediately after detection, which precluded the possibility of paired analysis. Conclusions NBI appears to be a less time-consuming and equally effective alternative to CE for the detection of IN. However, given the NBI lesion and patient miss rates, it cannot be recommended as the standard technique.
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