SticholysinII (StII) is a protein purified from the sea anemone Stichodactyla helianthus. Based on its ability to form pores in membranes, we evaluated their effects in vitro and in vivo in tumor cells in order to study the antitumor properties of this toxin. In order to understand the cell pathways triggered upon membrane destabilization by StII, we have examined the mechanisms that mediate its cytotoxicity on tumor cells. StII's action on L1210, Raji, X63, B16, A549 tumor cells leads to an increase in cell volume and the release of cytoplasmic components. In the sticholysin-treated cells, an activation of MAPKinases ERK1/2 and p38 were observed. Furthermore, KN62, an inhibitor of CaMKII and Necrostatin, a potent necroptosis inhibitor reduced the cytotoxic activity of StII. Additionally, the integrity of actin cytoskeleton seems to be crucial to pore- formation, probably due to its involvement in the stabilization of the multimeric structure of StII. Combination of toxin with doxorubicin and vincristine provoked the potentiation on chemotherapeutic cytotoxicity. Anti-tumor effect in vivo was study in a subcutaneous X63 tumor model in Balb/c mice. The intra-tumoral injection of StII significantly reduced the tumor volume and the histopathological analysis not showed systemic toxicity. In summary, StII induces cell death by regulated necrosis that depends of the intracellular signalization via MAPKinases and CaMKII. The anti-tumor effect in vivo without systemic toxicity suggests a new antitumoral compound.