Tachyplesin is an antimicrobial peptide present in leukocytes of the horseshoe crab (Tachypleus tridentatus). In this study, a synthetic tachyplesin conjugated to the integrin homing domain RGD was tested for antitumor activity. The in vitro results showed that RGD-tachyplesin inhibited the proliferation of both cultured tumor and endothelial cells and reduced the colony formation of TSU prostate cancer cells. Staining with fluorescent probes of FITC-annexin V, JC-1, YO-PRO-1, and FITC-dextran indicated that RGD-tachyplesin could induce apoptosis in both tumor and endothelial cells. Western blotting showed that treatment of cells with RGD-tachyplesin could activate caspase 9, caspase 8, and caspase 3 and increase the expression of the Fas ligand, Fas-associated death domain, caspase 7, and caspase 6, suggesting that apoptotic molecules related to both mitochondrial and Fas-dependent pathways are involved in the induction of apoptosis. The in vivo studies indicated that the RGD-tachyplesin could inhibit the growth of tumors on the chorioallantoic membranes of chicken embryos and in syngenic mice.