Galectin-3 is a multifunctional β-galactoside‑binding lectin that is involved in multiple biological functions which are upregulated in malignancies, including cell growth, adhesion, proliferation, progression and metastasis, as well as apoptosis. A previous study has confirmed the roles of galecin-3 overexpression in the biological behavior of Eca109 human esophageal cancer (EC) cells. In the present study, small interfering (si)RNA-mediated galectin-3 silencing was performed to analyze the effects of decreased galectin-3 expression on the biological behavior of EC cells. Western blot and quantitative polymerase chain reaction analyses were utilized to confirm galectin-3 knockdown at the protein and mRNA level (P<0.05 vs. siRNA-control and untransfected groups). Cell proliferation was assessed using the Cell Counting Kit-8 assay. At 72 and 96 h after transfection, the proliferation of Eca109 cells in the siRNA-Gal-3 group was decreased compared with that in the siRNA-Control and untransfected groups (P<0.001 and P=0.004, respectively). Furthermore, Transwell assays demonstrated that inhibition of galecin-3 significantly reduced the migration and invasion of Eca109 cells compared with that in the other groups (P<0.05). Finally, apoptosis of Eca109 cells was detected using Annexin V/7-amino‑actinomycin double-staining and flow cytometric analysis. Galectin-3 knockdown significantly enhanced the apoptotic rate of Eca109 cells compared with that in the siRNA-control and untreated groups (P=0.031 and P=0.047, respectively). In conclusion, following successful knockdown of galecin-3 expression in Eca109 cells, the cell proliferation, migration and invasion were reduced, while the apoptosis was enhanced, which indicates that galectin silencing may represent a therapeutic strategy for EC.