GnRH‐III has been shown to exert a cytotoxic effect on the GnRH‐R positive tumor cells. The chemotactic drug targeting (CDT) represents a new way for drug delivery approach based on selective chemoattractant guided targeting. The major goal of the present work was to develop and investigate various GnRH‐III derivatives as potential targeting moieties for CDT. The cell physiological effects (chemotaxis, adhesion, and signaling) induced by three native GnRHs (hGnRH‐I, cGnRH‐II, and lGnRH‐III) and nine GnRH‐III derivatives were evaluated in two model cells (Tetrahymena pyriformisand Mono Mac 6 human monocytes). According to our results, the native GnRH‐III elicited the highest chemoattractant and adhesion inducer activities of all synthesized peptides in micromolar concentrations in monocytes. With respect to chemoattraction, dimeric derivatives linked by a disulfide bridge ([GnRH‐III(C)]2) proved to be efficient in both model cells; furthermore, acetylation of the linker region ([GnRH‐III(Ac‐C)]2) could slightly improve the chemotactic and adhesion effects in monocytes. The length of the peptide and the type of N‐terminal amino acid could also determine the chemotactic and adhesion modulation potency of each fragment. The application of the chemoattractant GnRH‐III derivatives was accompanied by a significant activation of phosphatidylinositol 3‐kinase in both model cells. In summary, our work on low‐level differentiated model cells of tumors has proved that GnRH‐III and some of its synthetic derivatives are promising candidates to be applied in CDT: these compounds might act both as carrier, delivery unit, and antitumor agents. Copyright © 2012 European Peptide