This project aimed to explore novel anticancer therapeutics from products of parasite since several data related to benefits from the T. spiralis infection have been documented. We validated antitumor activity of T. spiralis infective larval extract and extricate the parasite-derived-antitumor peptide. We found that larval extract exerted antitumor activity to three types of carcinoma cells including hepatocellular carcinoma HepG2, ovarian cancer SK-OV-3, and lung adenocarcinoma A549. Interestingly, it displayed the most antitumor effect to HepG2 cells. Using proteomic and bioinformatic approaches, three putative anticancer peptides were identified from T. spiralis infective larval extract. One of these peptides showed a dose-dependent-anti-HepG2 effect by inducing ROS accumulation, leading to inhibition of the cell proliferation. Our data indicate potential application of the larval extract-derived antitumor peptide as a complementary agent for human hepatoma treatment and highlight a positive aspect of parasite apart from its deleterious effect.