Identification of stool miR-135b-5p as a non-invasive diaognostic biomarker in later tumor stage of colorectal cancer

Aims: Colorectal cancer (CRC) is one of the most common cancers. However, the effective and non-invasive diagnostic biomarkers for the detection of CRC metastasis remain unsatisfied. Here, we aimed to evaluate the diagnostic performance of stool miR-135b-5p (a potential biomarker) for metastasis of CRC patients.

Materials and methods: Serum and stool specimens from 77 patients with CRC and 29 normal controls were collected for miRNA purification. Real-time quantitative PCR was used for the relative quantification of miR-135b-5p expression. Receiver operating and characteristic (ROC) curve analysis was used to estimate the diagnostic performance of stool/serum miR-135b-5p for CRC metastasis. Dual-luciferase reporter assay was conducted to determine miR-135b-5p's target gene. Besides, a trans-well matrigel assay was performed to evaluate the invasion ability of HT-29 cell lines.

Key findings: Stool miR-135b-5p expression was dramatically up-regulated in CRC patients, and it effectively distinguished the CRC patients from normal controls with 74.1% of specificity and 96.5% of sensitivity. Moreover, stool miR-135b-5p exhibited a better diagnostic performance in distinguishing the different TNM stages of CRC patients than serum miR-135b-5p. In the molecular mechanism, our observations indicated that miR-135-5p directly targets the mRNA of ZNRF3, and then activates the Wnt pathway. Over-expression of ZNRF3 in HT-29 cells obviously reversed miR-135b-5p's effects on cell invasion and migration, indicating miR-135b-5p achieves its biological functions in a ZNRF3 dependent manner.

Significance: MiR-135b-5p may be a promising non-invasive biomarker for the diagnosis of CRC patients with TNM stage-III/IV and a potential candidate to develop an intervention strategy for colorectal cancer.