Design, Synthesis, and Biological Evaluation of New Cyclic Melanotropin Peptide Analogues Selective for the Human Melanocortin-4 Receptor

Abstract

Intensive efforts have been made to develop potent and selective ligands for certain human melanocortin receptors as possible treatments for obesity and sexual dysfunction due to the role of these receptors in feeding behavior, energy homeostasis, sexual function, etc. A number of novel α-MSH analogues were designed and synthesized primarily on the basis of our previous MTII NMR structure. In these peptide analogues, a disulfide or lactam bridge between residues at positions 5 and 8 was used as a conformational constraint to enhance the β-turn spanning His 6 and D-Phe 7 , while the pharmacophore group in Arg 8 was mimicked via N α -alkylation of residues 8 or 9 with the guanidinylbutyl group. Biological assays for binding affinities and adenylate cyclase activities for the hMC1R, hMC3R, hMC4R, and hMC5R showed that three analogues have good binding affinity for the hMC4R (0.7-4.1 nM), but have no binding affinity up to 10 μM at the other three melanocortin receptors. Interestingly, the three hMC4R selective analogues display only 50% binding efficiency, suggesting there is allosteric modulation of the melanocortin-4 receptor. These analogues were found to act as antagonists of the hMC4R. This result represents a discovery of very selective peptide-based antagonists for the hMC4R. The high selectivity may be due to the strong conformational constraint via ring contraction as compared to MTII, and the rigid conformation preferred by these new ligands allows them to recognize only the hMC4R, but not to activate the second messenger. The MTII NMR structure-based design thus not only examined the structural model of melanocortin ligands, but also yielded new biologically unique α-MSH analogues.

Conformational constraint in protein ligand design and the inconsistency of binding entropy

“…[2][3][4][5][6][7] Adding constraint can improve target selectivity; namely, favoring the binding to one protein or receptor over others that share a high degree of homology or ligand recognition properties. [8][9][10][11][12] This is closely aligned with another advantage, the potential for establishing the bioactive conformation of the unconstrained parent compound. [13][14][15] In the case of peptide and peptidomimetic structure, additional reported benefits of adding organic constraints or effecting cyclization include increased resistance to proteases and in vivo metabolic stability, [16][17][18][19][20][21][22][23] as well as improved cell membrane permeability [24][25][26][27][28][29] (although the latter may be a contingent rather than a necessary property of cyclic peptides 30,31 ).…”

Allosteric ligands for G protein-coupled receptors: A novel strategy with attractive therapeutic opportunities

“…The binding affinity and the AC activity of the peptides under investigation were evaluated for the various human MCRs. Peptide 8 was characterized by a good binding affinity toward the hMC4R (IC 50 5 1.8 nM) without exhibiting any affinity for the other MCR subtypes up to 10 À5 M. 197 Worth noting, the compound behaved as an antagonist with a 50% binding efficiency when compared with that of the reference melanocortin receptor agonist MTII. Thus, the authors suggested an allosteric molecular interaction between 8 and the hMC4R, which was corroborated by assaying MTII in the presence of antagonists.…”

“…However, the allosteric modulation of MCRs by exogenous ligands has been evidenced only within a small group of conformationally constrained peptides, exemplified by compound 8 (Fig. 9), 197 designed and tested following a structural NMR study on cyclic melanocortin agonists and antagonists. The binding affinity and the AC activity of the peptides under investigation were evaluated for the various human MCRs.…”

Approaches to the rational design of selective melanocortin receptor antagonists

“…Recently, we have discovered allosteric ligands for the hMC4R ( 25 and 26 , Table 4) [65]. These ligands were designed as structural analogs with the arginine side chain moiety in different relationships to the other pharmacophore side chain groups for the MCRs [65].…”