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Recently, mixed opioid/NOP agonists came to the spotlight for their favorable functional profiles and promising outcomes in clinical trials as novel analgesics. This study reports on two novel chimeric peptides incorporating the fragment Tyr-c[D-Lys-Phe-Phe]Asp-NH 2 ( RP-170 ), a cyclic peptide with high affinity for µ and κ opioid receptors (or MOP and KOP, respectively), conjugated with the peptide Ac-RYYRIK-NH 2 , a known ligand of the nociceptin/orphanin FQ receptor (NOP), yielding RP-170-RYYRIK-NH 2 ( KW-495 ) and RP-170-Gly 3 -RYYRIK-NH 2 ( KW-496 ). In vitro, the chimeric KW-496 gained affinity for KOP, hence becoming a dual KOP/MOP agonist, while KW-495 behaved as a mixed MOP/NOP agonist with low nM affinity. Hence, KW-495 was selected for further in vivo experiments. Intrathecal administration of this peptide in mice elicited antinociceptive effects in the hot-plate test; this action was sensitive to both the universal opioid receptor antagonist naloxone and the selective NOP antagonist SB-612111. The rotarod test revealed that KW-495 administration did not alter the mice motor coordination performance. Computational studies have been conducted on the two chimeras to investigate the structural determinants at the basis of the experimental activities, including any role of the Gly 3 spacer.

Our formerly described pentapeptide opioid analog Tyr-c[D-Lys-Phe-Phe-Asp]NH 2 (designated RP-170 ), showing high affinity for the mu (MOR) and kappa (KOR) opioid receptors, was much more stable than endomorphine-2 (EM-2) in the rat brain homogenate and displayed remarkable antinociceptive activity after central (intracerebroventricular) and peripheral (intravenous ) administration. In this report, we describe the further modification of this analog, which includes the incorporation of a β 3 -amino acid, ( R )- and ( S )-β 3 -Lys, instead of D-Lys in position 2. The influence of such replacement on the biological properties of the obtained analogs, Tyr-c[( R )-β 3 -Lys-Phe-Phe-Asp]NH 2 ( RP-171 ) and Tyr-c[( S )-β 3 -Lys-Phe-Phe-Asp]NH 2 , ( RP-172 ), was investigated in vitro. Receptor radiolabeled displacement and functional calcium mobilization assays were performed to measure binding affinity and receptor activation of the new analogs. The obtained data revealed that only one of the diastereoisomeric peptides, RP-171 , was able to selectively bind and activate MOR. Molecular modeling (docking and molecular dynamics (MD) simulations) suggests that both compounds should be accommodated in the MOR binding site. However, in the case of the inactive isomer RP-172 , fewer hydrogen bonds, as well as instability of the canonical ionic interaction to Asp 147 , could explain its very low MOR affinity.

Reducing the well-known side effects of opioids prescribed to treat chronic pain remains unresolved, despite extensive research in this field. Among several options to tackle this problem the synthesis of multifunctional compounds containing hybridized structures gained a lot of interest. Recently, extensively investigated are combinations of opioid agonist and antagonist pharmacophores embodied in a single molecule. To this end, agonism at the μ opioid receptor (MOR) with simultaneous antagonism at the δ opioid receptor (DOR) emerged as a promising avenue to obtaining novel analogs devoid of serious adverse effects associated with morphine-based analgesics. In this review we covered up-to-date research on the synthesis of peptide-based ligands with MOR agonist/DOR antagonist profile., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier Inc. All rights reserved.)

New analogs of the endogenous opioid agonist endomorphin-2 (EM-2, H-Tyr-Pro-Phe-Phe-NH 2 ) have been obtained by introducing modified tyrosines at the position 1 of the sequence. For all analogs, the cis/trans conformation ratio about the tyramine-Pro amide bond, lipophilicity, receptor affinities, and functional activities, have been determined. Among the novel derivatives, [Dmt(3'-Cl)] 1 EM-2 (4) stood out for its subnanomolar μ-opioid receptor affinity and potent agonist activity, superior to that of the parent peptide EM-2. Hybrid quantum mechanics/molecular mechanics docking computations supported the cis tyramine-Pro bioactive conformation, and allowed us to analyze the contribution of the substituents of the "message" tyramine to binding, highlighting the role of halogen-bonding in the higher receptor affinity of peptide 4., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)