MicroRNAs (miRNAs) are small, non-coding RNAs that can function as oncogenes or tumor suppressors in human cancer. Recent reports have highlighted the oncogenic aspects of microRNA miR-125b. However, the clinical significance of miR-125b in gastrointestinal cancers has not been sufficiently investigated. To this end, we analyzed miR-125b expression in colorectal cancer cases. Quantitative RT-PCR was used to evaluate miR-125b expression in 89 colorectal cancer cases to determine the clinicopathological significance of miR-125b expression. The high miR-125b expression group showed a greater incidence of advanced tumor size and tumor invasion compared to the low miR-125b expression group (P<0.05). In addition, the high miR-125b expression group had a significantly poorer prognosis compared to the low expression group (P<0.05). Multivariate analysis indicated that high miR-125b expression was an independent prognostic factor for survival. Our analysis of miR-125b focused on the miR-125b/p53 pathway. In vitro assays revealed that overexpression of miR-125b repressed the endogenous level of p53 protein in human colorectal cancer cells. These data show that miR-125b is directly involved in cancer progression and is associated with poor prognosis in human colorectal cancer. Our findings suggest that miR-125b could be an important prognostic indicator for colorectal cancer patients.