Epigenetically silenced miR-34b/c as a novel faecal-based screening marker for colorectal cancer

Abstract

Background:MicroRNAs are tiny non-coding small endogenous RNAs that regulate gene expression by translational repression, mRNA cleavage and mRNA inhibition. The aim of this study was to investigate the hypermethylation of miR-34b/c and miR-148a in colorectal cancer, and correlate this data to clinicopathological features. We also aimed to evaluate the hypermethylation of miR-34b/c in faeces specimens as a novel non-invasive faecal-DNA-based screening marker.Methods:The 5-aza-2′-deoxycytidine treatment and methylation-specific PCR were carried out to detect the hypermethylation of miR-34b/c and miR-148a.Results:The miR-34b/c hypermethylation was found in 97.5% (79 out of 82) of primary colorectal tumours, P=0.0110. In 75% (21 out of 28) of faecal specimens we found a hypermethylation of miR-34b/c while only in 16% (2 out of 12) of high-grade dysplasia. In addition, miR-148a was found to be hypermethylated in 65% (51 out of 78) of colorectal tumour tissues with no significant correlation to clinicopathological features. However, a trend with female gender and advanced age was found, P=0.083. We also observed a trend to lower survival rate in patients with miR-148a hypermethylation with 10-year survival probability: 48 vs 65%, P=0.561.Conclusions:These findings show that aberrant hypermethylation of miR-34b/c could be an ideal class of early screening marker, whereas miR-148a could serve as a disease progression follow-up marker.

Great potential of miRNAs as predictive and prognostic markers for cancer

“…Blood is regarded as an ideal type of sample for cancer biomarker discovery as it can be collected easily in a minimally invasive manner [22]. Some studies have also attempted to use miRNAs from feces as markers for gastrointestinal cancer [23,24]. The use of different sample types enables the discovery of novel markers under various constraints.…”

MicroRNAs as molecular markers for colon cancer Diagnostic screening in stool & blood

“…On the other hand, our data and others [25,27,28,[54][55][56][57][58][59][60][61][62][63][64][65][66] have shown that quantitative changes in the expression of few miRNA genes in stool or blood that are associated with colon cancer permit development of more sensitive and specific CRC molecular markers than those currently available on the market. In comparison to the commonly employed FOBT stool test, a noninvasive molecular and reliable test would particularly be more convenient as there would be no requirement for dietary restriction, or meticulous collection of samples, and thus a screening test would be acceptable to a broader segment of the population.…”

“…It should be emphasized that although not all of the shed cells in stool are derived from a tumor, data published by us and others [25,27,28,[56][57][58][59][60][61][62][63][64][65][66][67][68] have indicate that diagnostic miRNA gene expression profiles are associated with adequate number of exfoliated cancerous cells and enough transformed RNA is released in the stool, and also the availability of measurable amount of circulating. miRNA genes in blood (either cellular or extracellularly), which can be determined quantitatively by a sensitive technique such as PCR in spite of the presence of bacterial DNA, non-transformed RNA and other interfering substances.…”

“…In comparison to the commonly employed FOBT stool test, a noninvasive molecular and reliable test would particularly be more convenient as there would be no requirement for dietary restriction, or meticulous collection of samples, and thus a screening test would be acceptable to a broader segment of the population. Using stable molecules such as miRNAs that are not easily degradable when extracted from stool or blood and manipulated thereafter, a miRNA -approach for colon cancer is thus preferable to a transcriptomic mRNA-, mutation DNA-, epigeneticor a proteomic-based test [25,27,[55][56][57][58][59][60][61][62][63][64][65][66][67][68], particularly that we and others have shown that these stable, nondegradable miRNA molecules can be easily extracted from stool or from circulation in vitro using commercially available kits. Advantages and disadvantages of the in vivo and in vitro tests are presented in table 1.…”

“…Several of the miRNAs were shown by microarrays, NGS and RTqPCR in CRC tissue, cell culture lines, stool and blood to be related to colon cancer tumorigenesis [25,27,28,[54][55][56][57][58][59][60][64][65][66][67][68]86,87,94,98,114,123] and UC [25,95]. A study indicated that a combination of mRNA and miRNA expression signatures represent a broader approach for improving biomolecular classification of CRC [123].…”