The novel synthesized naltrexone-related MOR antagonist AT-99 counteracts dopamine releasing and behavioral depressant morphine-induced effects

Abstract

The development of new μ-opioid receptor (MOR) antagonists has been stimulated by the opioid overdose crisis. Our earlier in silico investigations on ligand-MOR receptor interactions indicated that the ligand cis-equatorial conformation of the amine and phenol is the most likely orientation observed within the MOR receptor. Here, we synthesized and characterized AT-99 [3-(1,3-dimethyl-cis-5-(methyl(phenethyl)amino)cyclohexyl)phenol] as a new in vivo and in vitro MOR antagonist. AT-99 effectively blocked MOR-mediated G protein activation by the agonists fentanyl and DAMGO, similarly to the reference compound, the opioid antagonist naloxone (NAL). Moreover, AT-99 behaves as a neutral MOR antagonist since it failed to stimulate [³⁵S]GTPγS binding, but it dose-dependently inhibited DAMGO-induced [³⁵S]GTPγS binding. While 0.1 μM NAL significantly reduced DAMGO potency, AT-99 produced a comparable effect only at the highest concentration tested. Furthermore, in radioligand competition binding assays, AT-99 fully displaced specific [³H]DAMGO binding in a concentration-dependent manner, although with lower affinity than NAL. In vivo, AT-99 (1 or 3 mg/kg i.v.) dose-dependently reduced the increase of dialysate dopamine (DA) in the nucleus accumbens (NAc) shell induced by morphine (1 mg/kg i.v.). Notably, AT-99 at the highest dose tested counteracted the reduction of behavioral rating scale (BRS) induced by morphine. Altogether, these data indicate that, although AT-99 interacts with the MOR relatively weakly, it displays interesting MOR antagonist properties, and as such it might serve as a scaffold to develop more potent MOR antagonists.