Colon cancer is the second leading cause of cancer‑related mortality worldwide, and the prognosis of advanced colon cancer has remained poor in recent years. Galectin‑9 (Gal‑9) is a tandem‑repeat type galectin that has recently been shown to exert antiproliferative effects on various types of cancer cells. The present study aimed to assess the effects of Gal‑9 on human colon and colorectal cancer cells in vitro and in vivo, as well as to evaluate the microRNAs (miRNAs/miRs) associated with the antitumor effects of Gal‑9. We examined the ability of Gal‑9 to inhibit cell proliferation via apoptosis, and the effects of Gal‑9 on cell cycle‑related molecules in various human colon and colorectal cancer cell lines. In addition, Gal‑9‑mediated changes in activated tyrosine kinase receptors and angiogenic molecules were assessed using protein array chips in colon and colorectal cancer cells. Moreover, miRNA array analysis was performed to examine Gal‑9‑induced miRNA expression profiles. We also elucidated if Gal‑9 inhibited tumor growth in a murine in vivo model. We found that Gal‑9 suppressed the cell proliferation of colon cancer cell lines in vitro and in vivo. Our data further revealed that Gal‑9 increased caspase‑cleaved keratin 18 levels in Gal‑9‑treated colon cancer cells. In addition, Gal‑9 enhanced the phosphorylation of ALK, DDR1, and EphA10 proteins. Furthermore, the miRNA expression levels, such as miR‑1246, miR‑15b‑5p, and miR‑1237, were markedly altered by Gal‑9 treatment in vitro and in vivo. In conclusion, Gal‑9 suppresses the cell proliferation of human colon cancer by inducing apoptosis, and these findings suggest that Gal‑9 can be a potential therapeutic target in the treatment of colon cancer.Keywords:apoptosis; caspase‑cleaved keratin 18; colon cancer; galectin‑9; microRNAs.