NOCICEPTIN/ORPHANIN FQ RECEPTOR LIGANDS: PHARMACOLOGICAL STUDIES

The neuropeptide nociceptin/orphanin FQ (N/OFQ) selectively binds and activates the N/OFQ peptide (NOP) receptor. At cellular level N/OFQ inhibits cAMP accumulation and Ca2+ conductance and stimulates K+ currents. N/OFQ regulates several biological functions both at central (pain, locomotion, memory, emotional responses, food intake) and peripheral (airways, cardiovascular, genitourinary and gastrointestinal systems) sites. Potent and selective NOP ligands are now required for investigating the roles played by NOP receptors in pathophysiological studies and for firmly defining the therapeutic indications of NOP receptor ligands. A novel assay to screen NOP receptor ligands has been validated with a large panel of ligands: the Gaqi5 chimeric protein has been used to force the NOP receptor to signal through the Ca2+ pathway in CHO cells. [Ca2+]i levels were monitored using the fluorometer FlexStation II. Data are in general agreement with classical Gi driven assay systems. The NOP peptide partial agonist, ZP120 was extensively characterized in vitro using electrically stimulated isolated tissues (mouse and rat vas deferens) and in vivo with the tail withdrawal assay. The selective involvement of the NOP receptor in the actions of ZP120 has been demonstrated in NOP(-/-) mice studies. A detailed pharmacological characterization of the recently identified non-peptide antagonist Compound 24 has been performed. Moreover in the context of a SAR study on Compound 24, a novel NOP ligand named Compound 35 was identified. Compound 24 and Compound 35 bound the human recombinant NOP receptor expressed in CHOhNOP cell membranes with high affinity (pKi values 9.62 and 9.14, respectively). Our findings derived from functional studies on CHOhNOP and bioassay studies on native receptors demonstrated that Compound 24 and Compound 35 behave as potent, competitive and selective non-peptide NOP antagonists. Finally, the NOP antagonist properties of Compound 24 have been confirmed in vivo in the mouse tail withdrawal assay.