Eosinophils are multifunctional leukocytes that are involved in innate and adaptive immune responses through the expression of various receptors and mediators. Previously, we showed that human eosinophils and T cells shared cytotoxic activities against tumor cells that involved the γ-δ TCR and cell–cell contact. In this study, we investigated the molecules involved in eosinophil–tumor cell interactions. Given the role of IL-18 in cell adhesion and in protecting against colon cancer, we evaluated its role in eosinophil-mediated cytotoxicity against Colo-205, a human colon carcinoma cell line. We found that human eosinophils exerted dose-and time-dependent tumoricidal activity against Colo-205 cells. Neutralization of IL-18 significantly reduced eosinophil-mediated Colo-205 apoptosis and inhibited cell–cell adhesion. Moreover, addition of rIL-18 led to upregulation of CD11a and ICAM-1 adhesion molecules, which were involved in the contact between eosinophils and Colo-205 cells. Our results indicated that IL-18 was involved in the eosinophil-mediated death of Colo-205 by facilitating contact between effector and target cells. These data underscored the involvement of an additional mediator in eosinophil-mediated antitumor cytotoxicity. Our findings support existing evidence that eosinophils could play a beneficial role in the context of colon cancer.