Identification of RC-33 as a potent and selective o1 receptor agonist potentiating NGFinduced neurite outgrowth in PC12 cells. Part 2 g-scale synthesis, physicochemical characterization and in vitro metabolic stability

Daniela Rossi, Annamaria Marra, Pietro Picconi, Massimo Serra, Simona Collina, Laura Catenacci, Milena Sorrenti, Marco Peviani, Daniela Curti ( University of Pavia), Erik Laurini, Maurizio Fermeglia, Sabrina Pricl ( University of Trieste), Stefania Bramb

Strong pharmacological evidences indicate that σ1 receptors are implicated in the pathophysiology  of all major CNS disorders. In the last years our research group has conducted extensive studies aimed at discovering novel σ1 ​ligands and we recently selected (R/S)-RC-33 as a novel potent and selective σ1  receptor agonist. As continuation of our work in this field, here we report our efforts in the development of this new σ1 receptor agonist. Initially, we investigated the binding of (R) and (S) enantiomers of RC-33 to the σ1 receptor by in silico experiments. The close values of the predicted affinity of (R)-RC-33 and (S)-RC-33 for the protein evidenced the non-stereoselective binding of RC-33 to the σ1 receptor; this, in turn, supported further development and characterization of RC-33 in its racemic form. Subsequently, we set-up a scaled-up, optimized  synthesis of (R/S)-RC-33 along with some compound characterization data (e.g., solubility in different media and solid state characterization by thermal analysis techniques). Finally, metabolic studies of RC-33 in different biological matrices (e.g., plasma, blood, and hepatic S9 fraction) of different species (e.g., rat, mouse, dog, and human) were performed. (R/S)-RC-33 is generally stable in all examined biological matrices, with the only exception of rat and human liver S9 fractions in the presence of NADPH. In such conditions, the compound is subjected to a relevant oxidative metabolism, with a degradation of approximately 65% in rat and 69% in human. Taken together, our results demonstrated that (R/S)-RC-33 is a highly potent, selective, metabolically stable σ1 agonist, a promising novel neuroprotective drug candidate.

Details of the key interactions detected in the equilibrated MD snapshots of (R)-RC-33 (A) and (S)-RC-33 (B) in complex with the σ1 receptor