AbstractThe process of liver colonization in colorectal cancer remains poorly characterized. Here, we addressed the role of microRNA (miRNA) dysregulation in metastasis. We first compared miRNA expression profiles between colorectal cancer cell lines with different metastatic properties and then identified target proteins of the dysregulated miRNAs to establish their functions and prognostic value. We found that 38 miRNAs were differentially expressed between highly metastatic (KM12SM/SW620) and poorly metastatic (KM12C/SW480) cancer cell lines. After initial validation, we determined that three miRNAs (miR‐424‐3p, −503, and −1292) were overexpressed in metastatic colorectal cancer cell lines and human samples. Stable transduction of non‐metastatic cells with each of the three miRNAs promoted metastatic properties in culture and increased liver colonization in vivo. Moreover, miR‐424‐3p and miR‐1292 were associated with poor prognosis in human patients. A quantitative proteomic analysis of colorectal cancer cells transfected with miR‐424‐3p, miR‐503, or miR‐1292 identified alterations in 149, 129, or 121 proteins, respectively, with an extensive overlap of the target proteins of the three miRNAs. Importantly, down‐regulation of two of these shared target proteins, CKB and UBA2, increased cell adhesion and proliferation in colorectal cancer cells. The capacity of distinct miRNAs to regulate the same mRNAs boosts the capacity of miRNAs to regulate cancer metastasis and underscores the necessity of targeting multiple miRNAs for effective cancer therapy. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.