Discovery of Polypharmacological Melanocortin-3 and -4 Receptor Probes and Identification of a 100-Fold Selective nM MC3R Agonist versus a μM MC4R Partial Agonist

Abstract

The centrally expressed melanocortin-3 and melanocortin-4 receptors (MC3R, MC4R) are established targets to treat diseases of positive and negative energy homeostasis. We previously reported (J. Med. Chem. 60:4342 2017) mixture-based positional scanning approaches to identify dual MC3R agonist and MC4R antagonist tetrapeptides. Herein, 46 tetrapeptides were chosen for MC3R agonist screening selectivity profiles, synthesized, and pharmacologically characterized at the mouse melanocortin-1, −3, −4, and −5 receptors. Substitutions to the tetrapeptide template were selected solely based on MC3R agonist potency from the mixture-based screen. This study resulted in the discovery of compound 42 (Ac-Val-Gln-(pI)DPhe-DTic-NH 2 ), a full MC3R agonist that is 100-fold selective for the MC3R over the µM MC4R partial agonist pharmacology. This compound represents a first-in-class MC3R-selective agonist. This ligand will serve as a useful in vivo molecular probe for the investigation of the roles of the MC3R and MC4R in diseases of dysregulated energy homeostasis.

Structure–Activity Relationships of the Tetrapeptide Ac-His-Arg-(pI)DPhe-Tic-NH2 at the Mouse Melanocortin Receptors: Modification at the (pI)DPhe Position Leads to mMC3R Versus mMC4R Selective Ligands

“…An antagonist pA 2 value of 7.3 was observed for KNS2-22-4 at the mMC4R (Figure 3). In prior studies, this compound was observed to possess nanomolar agonist potency at the MC3R (30–40 nM), partial agonist stimulation of the MC4R, and sub-micromolar antagonist potency at the MC4R (pA 2 of 6.6–7) [40,41], similar to the results in the present study.…”

“…The ( p Cl)DPhe-substituted KNS2-22-1 maintained similar potencies compared to KNS2-22-4 at the mMC1R and mMC5R, but was 9-fold less potent at the mMC3R (120 nM, Figure 2) and possessed an increased partial agonist response relative to NDP-MSH (70%, EC 50 = 280 nM) at the mMC4R (Figure 2) compared to KNS2-22-4 . The Ac-His-Arg-( p Cl)DPhe-Tic-NH 2 ( KNS2-22-1 ) tetrapeptide was previously reported to possess agonist pharmacology at the mMC3R (110 nM) and partial agonist activity at the mMC4R (EC 50 = 140 nM), similar to the values observed in the present study [41]. While similar potency relative to KNS2-22-4 was observed for the ( p F)DPhe-substituted KNS2-22-2 at the mMC1R, this substitution decreased potency at the mMC3R and mMC5R (30- and 14-fold as compared to KNS2-22-4 , respectively).…”

“…Thus, a DPhe-Arg motif resulted in full agonism at the MC4R and full to partial agonism at the MC3R accompanied by antagonist activity (dependent on the DPhe para -position). Due to the MC3R agonism and MC4R antagonism observed in the Ac-His-Arg-( p I)DPhe-Tic-NH 2 ligand [40,41], it was hypothesized that different substitutions at the DPhe para -position within this scaffold (possessing an Arg-DPhe motif and a Tic residue in position 4) may modulate MC4R agonist efficacy. The results in Table 3 demonstrate that the efficacy at the mMC4R was modulated by various para -substitutions.…”

“…Compared to the endogenous tetrapeptide melanocortin sequence (His-Phe-Arg-Trp), the new scaffold switched the Phe and Arg positions and incorporated a Tic residue in place of the Trp. A follow-up study utilized the most potent MC3R substitutions at each position within the tetrapeptide from the mixture-based positional scan, retaining the switched Phe and Arg positions (Arg or Gln were utilized in the second position, while ( p I)DPhe or ( p Cl)DPhe were substituted at the third position) [41]. A 100-fold selective MC3R versus MC4R agonist compound was identified (Ac-Val-Gln-( p I)DPhe-DTic-NH 2 ) that did not possess antagonist potency at the MC4R and only partially stimulated the MC4R (less than 50% efficacy of NDP-MSH) [41].…”

“…A follow-up study utilized the most potent MC3R substitutions at each position within the tetrapeptide from the mixture-based positional scan, retaining the switched Phe and Arg positions (Arg or Gln were utilized in the second position, while ( p I)DPhe or ( p Cl)DPhe were substituted at the third position) [41]. A 100-fold selective MC3R versus MC4R agonist compound was identified (Ac-Val-Gln-( p I)DPhe-DTic-NH 2 ) that did not possess antagonist potency at the MC4R and only partially stimulated the MC4R (less than 50% efficacy of NDP-MSH) [41]. Switching the Arg and Phe positions within the melanocortin tetrapeptide sequence may therefore lead to MC3R-selective ligands that may be further developed into probe and therapeutic lead compounds.…”