Understanding ligand affinity and efficacy at mu and delta opioid receptors using cyclic pentapeptides

Chronic use of mu opioid drugs such as morphine causes neurochemical adaptations leading to tolerance and dependence. While the effects of morphine are mediated by mu‐opioid receptors, several studies have shown that antagonism or knockdown of delta‐opioid receptors can modulate development of tolerance and dependence. To test the hypothesis that delta‐opioid receptors contribute to chronic effects of mu opioids, we developed cyclic pentapeptides displaying both mu agonism and delta antagonism and equivalent affinity as tools to gain a better understanding of the functional mu‐delta interaction(s). Based on in silico modeling data at putative active and inactive conformations of the mu‐ and delta‐opioid receptors, several peptides containing a napthylalanine substitution were synthesized. These compounds displayed variable mu agonist/delta antagonist profiles. One candidate (Peptide 8), displayed full agonism at the mu receptor (99% stimulation versus mu agonist DAMGO) and antagonism at the delta receptor (Ke = 20 nM versus delta agonist SNC80) in the GTPγ 35 S assay. Peptide 8 bound with equipotent affinity (ranging between 1.2‐1.4 nM) to all opioid receptors stably expressed in C6 rat glioma cells. In SH‐SY5Y cells endogenously expressing mu‐ and delta‐opioid receptors, Peptide 8 was a mu agonist with greater potency but lower efficacy than DAMGO. Supported by DA007281, DA007267, DA03910, and DA04087