Eupolyphaga sinensis Walker is an important insect used in Chinese traditional medicine. In this study, we purified a 72-kDa anticancer protein, designated as EPS72, from this species using ammonium sulfate precipitation, ultrafiltration, CM Sepharose Fast Flow cation exchange, Q Sepharose High Performance (HP) anion exchange, Butyl Sepharose HP hydrophobic chromatography, and Superdex 75 gel filtration chromatographic techniques. EPS72 exhibited a potent anticancer activity against the human lung cancer A549 cell line (IC50, 18.76 μg/mL). Further study showed that EPS72 could induce A549 cell detachment and apoptosis, inhibit cell adhesion to fibronectin and collagen IV, and restrain cell migration and invasion. Moreover, EPS72 significantly decreased the expression of β1-integrin. This study suggests that EPS72 could potentially be developed as a novel anticancer therapeutic agent due to its possible antimetastatic activity.