Hepatocellular carcinoma (HCC) is known as a highly prevalent cancer with a poor prognosis and short survival time, despite intensive research and clinical efforts. Increasing numbers of studies have reported that microRNAs are involved in the malignant behavior of hepatocellular carcinoma cells via directly targeting multiple oncogenes or tumor suppressors. Here, we report that the expression of microRNA-493 (miR-493) is decreased in HCC cell lines and in tumor tissues. Overexpression of miR-493 in HCC cells dramatically inhibited cell proliferation and colony-formation in vitro and inhibited tumor formation of HCC cell xenografts in vivo. miR-493 also suppressed cell migration and invasion in HCC cell lines. Novel targets ANTXR1 and RSPO2 were confirmed to be suppressed by miR-493 directly, and overexpression of ANTXR1 and RSPO2 could restore tumorigenesis in miR-493 treated HCC cell. Moreover, Wnt/β-catenin signaling pathway, which was reported to be activated by ANTXR1 and RSPO2, was also inhibited by miR-493 overexpression and might be involved in anti-tumor function of miR-493. These findings suggest that miR-493 acts as a negative regulator in hepatocellular carcinoma progression and may be a potential therapeutic target for HCC.