Some biological properties of new bifunctional conjugates designed for drug targeting were evaluated through in vitro experiments. Eight peptidylcyclodextrin compounds were used, which correspond to modified beta- or gamma-cyclodextrin (CD) grafted on neuropeptide substance P (SP) or a shorter derivative (SP(4-11)). Using anti-SP and anti-CD antibodies as molecular probes, we showed that the main structural features of the two moieties of these adducts were preserved. Binding experiments, using CHO cells expressing the human SP-specific NK1 receptor, demonstrated the functionality of all peptidylcyclodextrin derivatives, which exhibited IC50 values in a 10(-9)-10(-7) M range. All compounds were able to induce a pharmacological response, triggering phosphatidylinositol turnover with EC50 values in the same range as the natural ligand. Moreover, autoradiography analysis of rat spinal corn sections proved that [125I]SP binding was dose-dependently displaced by one selected compound (a gamma-CD-SP), showing a similar affinity of this adduct for the rat neurokinin 1 receptor. Our observations demonstrate that these peptidylcyclodextrins efficiently target NK1 receptor-expressing cells.