Non-invasive diagnostic biomarkers may contribute to an early identification of gastric cancer (GC) and improve the clinical management. Unfortunately, no sensitive and specific screening biomarkers ...are available yet and the currently available approaches are limited by the nature of the disease. GC is a heterogenic disease with various distinct genetic and epigenetic events that occur during the multifactorial cascade of carcinogenesis. MicroRNAs (miRNAs) are commonly deregulated in gastric mucosa during the
infection and in stepwise manner from chronic gastritis, through preneoplastic conditions such as atrophic gastritis and intestinal metaplasia, to early dysplasia and invasive cancer. Identification of miRNAs in blood in 2008 led to a great interest on miRNA-based diagnostic, prognostic biomarkers in GC. In this review, we provide the most recent systematic review on the existing studies related to miRNAs as diagnostic biomarkers for GC. Here, we systematically evaluate 75 studies related to differential expression of circulating miRNAs in GC patients and provide novel view on various heterogenic aspects of the existing data and summarize the methodological differences. Finally, we highlight several important aspects crucial to improve the future translational and clinical research in the field.
Abstract
Background
Assessment of the disease activity in inflammatory bowel disease (IBD) is essential for adequate treatment management and reliable noninvasive biomarkers for verification of ...mucosal healing are still needed. MicroRNAs (miRNAs) are differentially expressed in IBD and cancer. We aimed to evaluate the potential of circulating and fecal miRNAs as diagnostic biomarkers for IBD.
Methods
In this proof-of-principle study we used 2 independent patient cohorts. Testing cohort (n = 96) included serum and fecal samples from controls (n = 35) and IBD patients (n = 61) including 43 patients with Crohn′s disease (CD), 18 with ulcerative colitis (UC) with an active disease (n = 38), or in remission (n = 23). Validation cohort included fecal samples from patients with calprotectin/endoscopy-confirmed active disease (n = 30) or in remission (n = 15). Target-based approach (miR-16, miR-21, miR-155, and miR-223) has been used to evaluate miRNA expression.
Results
Sera samples from IBD patients showed higher level of miR-16, miR-21, and miR-223, but not miR-155, compared to controls and was higher in CD than in UC patients. Much stronger miRNA expression changes were observed in feces from IBD patients for all studied miRNAs with highest expression of miR-155 and miR-223 in testing and validation cohorts. MiRNA expression correlated with clinical remission, however, only fecal but not circulating miRNAs, correlated with surrogate parameters such as fecal calprotectin or C-reactive protein.
Conclusions
Our data provide a novel evidence for differential expression level of fecal miRNAs in IBD. We demonstrate that miRNAs in feces correlate with disease activity and may be considered as potential tool for the further biomarker research in IBD.
10.1093/ibd/izy046_video1
izy046.video1
5794822319001
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.
Colorectal cancer (CRC) is the second leading cause of cancer-related deaths, but currently available noninvasive screening programs have achieved only a modest decrease in mortality. MicroRNAs ...(miRNA) play an important role in a wide array of biological processes and are commonly dysregulated in neoplasia. We aimed to evaluate the feasibility of fecal miRNAs as biomarkers for colorectal neoplasia screening.
Total RNA was extracted from freshly collected stool samples from 8 healthy volunteers and 29 samples collected via fecal occult blood testing from subjects with normal colonoscopies, colon adenomas, and CRCs. miRNA expression analyses were done with TaqMan quantitative reverse transcription-PCR for a subset of miRNAs. Illumina miRNA microarray profiling was done to evaluate the differences in expression patterns between normal colonic mucosa tissues and stool samples from healthy subjects.
We efficiently extracted miRNAs from stool specimens using our developed protocol. Data from independent experiments showed high reproducibility for miRNA extraction and expression. miRNA expression patterns were similar in stool specimens among healthy volunteers, and reproducible in stool samples that were collected serially in time from the same individuals. miRNA expression profiles from 29 patients showed higher expression of miR-21 and miR-106a in patients with adenomas and CRCs compared with individuals free of colorectal neoplasia.
Our data indicate that miRNAs could be extracted from stool easily and reproducibly. The stools of patients with colorectal neoplasms have unique and identifiable patterns of miRNA expression.
Fecal miRNAs may be an excellent candidate for the development of a noninvasive screening test for colorectal neoplasms.
Scope
Diet is amongst the most crucial factors contributing to the multistep process of carcinogenesis. The role of exogenous microRNAs (miRNAs) is still debatable. In this proof‐of‐principle work, ...the presence of miRNAs in a variety of foods, its stability to processing, and detectability in GI mucosa and feces are studied and the effect of short‐term diet on human‐ or plant‐derived miRNAs in feces and blood is examined.
Methods and results
Animal and plant miRNAs are detected in all foods irrespective of processing. Animal‐derived foods showed the highest miRNA level and the lowest is found in cheese and milk. The impact of the short‐term vegetarian or meat‐rich diet on blood and feces miRNA is evaluated in healthy subjects using qPCR and Affymetrix profiling. Diet is not associated with changes in ultraconserved miRNAs. However, a vegetarian diet is associated with an increase of miR‐168 in feces but not in blood. Overall, plant miR‐168 is detectable in normal GI mucosa and in colorectal cancer.
Conclusions
Food provides a great source of miRNAs and diet may be associated with changes in xenomiRs. Plant‐derived miR‐168 is ubiquitously present in feces, normal mucosa, and cancer. Further studies are needed to evaluate the functional interaction between diet‐derived miRNAs and GI tract.
Exogenous microRNAs are suggested to trigger gastrointestinal diseases. Here, it is shown that food may provide a great source of microRNAs. The diet challenge described in this study is associated with an increase in plant microRNA miR‐168 in feces while no changes in ultraconserved microRNAs are observed. Detectability of miR‐168 in gastric and colon mucosa, colorectal cancer, and peritoneal fluid suggests its functional role.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Pancreatic cancer (PCA) is an aggressive tumor that associates with high mortality rates. Majority of PCA patients are diagnosed usually at late tumor stages when the therapeutic options are limited. ...MicroRNAs (miRNA) are involved in tumor development and are commonly dysregulated in PCA. As a proof-of-principle study, we aimed to evaluate the potential of fecal miRNAs as biomarkers for pancreatic cancer.
Total RNA was extracted from feces using Qiagen's miRNA Mini Kit. For miRNA expression analyses we selected a subset of 7 miRNAs that are frequently dysregulated in PCA (miR-21, -143, -155, -196a, -210, -216a, -375). Subsequently, expression levels of these miRNAs were determined in fecal samples from controls (n = 15), chronic pancreatitis (n = 15) and PCA patients (n = 15) using quantitative TaqMan-PCR assays.
All selected miRNAs were detectable in fecal samples with high reproducibility. Four of seven miRNAs (miR-216a, -196a, -143 und -155) were detected at lower concentrations in feces of PCA patients when compared to controls (p<0.05). Analysis of fecal miRNA expression in controls and patients with chronic pancreatitis and PCA revealed that the expression of miR-216a, -196a, -143 und -155 were highest in controls and lowest in PCA. The expression of the remaining three miRNAs (miR-21, -210 and -375) remained unchanged among controls and the patients with either chronic pancreatitis or PCA.
Our data provide novel evidence for the differential expression of miRNAs in feces of patients with PCA. If successfully validated in large-scale prospective studies, the fecal miRNA biomarkers may offer novel tools for PCA screening research.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Recent evidence suggests that several dietary polyphenols may exert their chemopreventive effect through epigenetic modifications. Curcumin is one of the most widely studied dietary chemopreventive ...agents for colon cancer prevention, however, its effects on epigenetic alterations, particularly DNA methylation, remain unclear. Using systematic genome-wide approaches, we aimed to elucidate the effect of curcumin on DNA methylation alterations in colorectal cancer cells.
To evaluate the effect of curcumin on DNA methylation, three CRC cell lines, HCT116, HT29 and RKO, were treated with curcumin. 5-aza-2'-deoxycytidine (5-aza-CdR) and trichostatin A treated cells were used as positive and negative controls for DNA methylation changes, respectively. Methylation status of LINE-1 repeat elements, DNA promoter methylation microarrays and gene expression arrays were used to assess global methylation and gene expression changes. Validation was performed using independent microarrays, quantitative bisulfite pyrosequencing, and qPCR.
As expected, genome-wide methylation microarrays revealed significant DNA hypomethylation in 5-aza-CdR-treated cells (mean β-values of 0.12), however, non-significant changes in mean β-values were observed in curcumin-treated cells. In comparison to mock-treated cells, curcumin-induced DNA methylation alterations occurred in a time-dependent manner. In contrast to the generalized, non-specific global hypomethylation observed with 5-aza-CdR, curcumin treatment resulted in methylation changes at selected, partially-methylated loci, instead of fully-methylated CpG sites. DNA methylation alterations were supported by corresponding changes in gene expression at both up- and down-regulated genes in various CRC cell lines.
Our data provide previously unrecognized evidence for curcumin-mediated DNA methylation alterations as a potential mechanism of colon cancer chemoprevention. In contrast to non-specific global hypomethylation induced by 5-aza-CdR, curcumin-induced methylation changes occurred only in a subset of partially-methylated genes, which provides additional mechanistic insights into the potent chemopreventive effect of this dietary nutraceutical.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Gastric carcinogenesis is associated with alterations of microRNAs (miRNAs) and reversal of these alterations may be a crucial element in cancer prevention. Here we evaluate the influence of H. ...pylori eradication, low-dose aspirin (LDA), non-steroidal anti-inflammatory drugs (NSAIDs) and proton-pump inhibitors (PPI) on modification of inflammatory mucosal miRNAs miR-155 and miR-223 in Helicobacter pylori-infected and non-infected subjects. The study was performed in two parts: 1) interventional study in 20 healthy subjects with and without H. pylori infection or following eradication (each n = 10) where LDA (100 mg) was given daily for 7 days; 2) prospective case-control observational study (n = 188). MiR-155 and miR-223 expression was strongly linked to H. pylori-infection and in short-term view showed a trend for reversal after eradication. Daily LDA as well as regular NSAIDs showed no influence on miRNAs expression both in healthy subjects and patients, while regular PPI intake was associated with lower miR-155 expression in antrum of patients with chronic gastritis independent of density of neutrophils and mononuclear infiltrate. In summary, PPI but not LDA or NSAIDs were associated with modification of inflammatory miRNAs miR-155 and miR-223 in an H. pylori dependent manner. The functional role of inflammatory miR-155 and miR-223 in understanding of H. pylori-related diseases needs further evaluation.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Helicobacter pylori (H. pylori) infection has been considered as the main causal factor in gastric carcinogenesis, but other bacterial species may also play an important role in pathophysiology of ...gastric cancer. The aim of the study was to explore the link between gastric cancer prognosis and the mucosal microbial community in tumorous and adjacent gastric tissue. The bacterial profile was analysed using 16S sequencing (V1-V2 region). Microbial differences were mostly characterized by lower relative abundances of H. pylori in tumorous gastric tissues. Bacterial community and outcome data analysis revealed the genus Fusobacterium and Prevotella significantly associated with worse overall survival in gastric cancer patients. In particular, Fusobacterium was associated with significant increase in hazard ratio in both univariable and multivariable analysis and independently validated using TCMA data. Phylogenetic biodiversity of Fusobacterium species in the stomach revealed F. periodonticum as the most prevalent in healthy subjects, while F. nucleatum was most abundant in patients with gastric cancer. Bacterial community network analysis in gastric cancer suggests substantial complexity and a strong interplay between F. nucleatum and Prevotella. In summary, mucosal microbial community in the stomach was associated with worse overall survival in gastric cancer patients. Strongest negative impact on prognosis was linked to the abundance of F. nucleatum in tumorous specimens, suggesting its translational relevance in management of gastric cancer patients.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Biomarkers for surveillance, diagnosis and predictionof prognosis in patients with hepatocellular carcinoma（HCC） are currently not ready for introduction intoclinical practice because of limited ...sensitivity andspecificity. Especially for the early detection of smallHCC novel biomarkers are needed to improve thecurrent effectiveness of screening performed byultrasound. The use of high-throughput technologiesin hepatocellular research allows to identify moleculesinvolved in the complex pathways in hepatocarcinogenesis.Several invasive and non-invasive biomarkershave been identified already and have been evaluatedin different clinical settings. Gene signatures withprognostic potential have been identified by geneexpression profiling from tumor tissue. However, asingle ＂all-in-one＂ biomarker that fits all-surveillance,diagnosis, prediction of prognosis-has not been foundso far. The future of biomarkers most probably lies in acombination of non-invasive biomarkers, imaging andclinical parameters in a surveillance setting. Molecularprofiling of tumorous and non-tumorous liver tissuemay allow a prediction of prognosis for the individualpatient and hopefully clear the way for individualtreatment approaches. This article gives an overviewon current developments in biomarker research inHCC with a focus on currently available and novelbiomarkers, in particular on microRNA.
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