Discovery of a β-Hairpin Octapeptide, c[Pro-Arg-Phe-Phe-Dap-Ala-Phe-DPro], Mimetic of Agouti-Related Protein(87–132) [AGRP(87–132)] with Equipotent Mouse Melanocortin-4 Receptor (mMC4R) Antagonist Pharmacology

Abstract

Agouti-related protein (AGRP) is a potent orexigenic peptide that antagonizes the melanocortin-3 and -4 receptors (MC3R and MC4R). While the C-terminal domain of AGRP, AGRP(87-132), is equipotent to the full-length peptide, further truncation decreases potency at the MC3R and MC4R. Herein, we report AGRP-derived peptides designed to mimic the active β-hairpin secondary structure that contains the hypothesized Arg-Phe-Phe pharmacophore. The most potent scaffold, c[Pro-Arg-Phe-Phe-Asn-Ala-Phe-DPro], comprised the hexa-peptide β-hairpin loop from AGRP cyclized through a DPro-Pro motif. A 20 compound library was synthesized from this scaffold for further structure-activity relationship studies. The most potent peptide from this library was an Asn to diaminopropionic acid substitution that possessed sub-nM antagonist activity at the mMC4R and was greater than 160-fold selective for the mMC4R versus the mMC3R. The reported ligands may serve as probes to characterize the melanocortin receptors in vivo and leads in the development of novel therapeutics.

Structure–Activity Relationship Studies on a Macrocyclic Agouti-Related Protein (AGRP) Scaffold Reveal Agouti Signaling Protein (ASP) Residue Substitutions Maintain Melanocortin-4 Receptor Antagonist Potency and Result in Inverse Agonist Pharmacology at the Melanocortin-5 Receptor

“…54 The most potent substitution, Asn to diaminopropionic acid (Dap), resulted in a ligand that was equipotent to the C-terminal domain of AGRP at the MC4R. 54 It was previously hypothesized that the active loop Asn residue in AGRP may be in close proximity to an Asp residue in the MC4R (Asp 189 ), and that incorporation of basic residues into AGRP analogues at this position might form a novel salt bridge with the MC4R. 47 While the increased potency observed for basic substitutions supported the postulated salt bridge, the increased potency for Gly suggested other residues might also be incorporated without diminishing potency.…”

“…54 A macrocyclic scaffold containing the active hexapeptide loop of ARGP cyclized through a DPro-Pro motif, c[Pro-Arg-Phe-Phe-Asn-Ala-Phe-DPro], was 50-fold less potent at the MC4R compared to AGRP. 54 Further structure-activity relationship studies at the Asn position indicated basic residues (diaminopropionic acid [Dap], diaminobutyric acid [Dab], Orn, Lys, Arg) or Gly increased MC4R potency.…”

“…54 A macrocyclic scaffold containing the active hexapeptide loop of ARGP cyclized through a DPro-Pro motif, c[Pro-Arg-Phe-Phe-Asn-Ala-Phe-DPro], was 50-fold less potent at the MC4R compared to AGRP. 54 Further structure-activity relationship studies at the Asn position indicated basic residues (diaminopropionic acid [Dap], diaminobutyric acid [Dab], Orn, Lys, Arg) or Gly increased MC4R potency. The most potent substitution, the Asn to Dap replacement, resulted in a sub-nanomolar potent MC4R antagonist that was 160-fold selective for the MC4R over the MC3R, induced partial MC1R activation at 100 μM concentrations, and did not possess agonist activity at the MC5R.…”

“…The most potent substitution, the Asn to Dap replacement, resulted in a sub-nanomolar potent MC4R antagonist that was 160-fold selective for the MC4R over the MC3R, induced partial MC1R activation at 100 μM concentrations, and did not possess agonist activity at the MC5R. 54 To further investigate this scaffold, a series of truncated macrocycles were synthesized containing either the Asn or Dap residues and replacing the Arg-Phe-Phe tripeptide antagonist sequence with the melanocortin agonist tetrapeptide His-DPhe-Arg-Trp or tripeptide DPhe-Arg-Trp sequences. 55 This study examined if the AGRP DPro-Pro loop scaffold was amendable to substitutions that may result in melanocortin agonist activity as weight-management therapeutic leads.…”

Bench-top to clinical therapies: A review of melanocortin ligands from 1954 to 2016

“…Scaffolds reported by Ericson et al . were AGRP macrocyclic derivatives [133]. The antagonist reported by Lensing et al .…”