RESUMEN
The central melanocortin-3 and melanocortin-4 receptors (MC3R, MC4R) are key regulators of body weight and energy homeostasis. Herein, the discovery and characterization of first-in-class small molecule melanocortin agonists with selectivity for the melanocortin-3 receptor over the melanocortin-4 receptor are reported. Identified via "unbiased" mixture-based high-throughput screening approaches, pharmacological evaluation of these pyrrolidine bis-cyclic guanidines resulted in nanomolar agonist activity at the melanocortin-3 receptor. The pharmacological profiles at the remaining melanocortin receptor subtypes tested indicated similar agonist potencies at both the melanocortin-1 and melanocortin-5 receptors and antagonist or micromolar agonist activities at the melanocortin-4 receptor. This group of small molecules represents a new area of chemical space for the melanocortin receptors with mixed receptor pharmacology profiles that may serve as novel lead compounds to modulate states of dysregulated energy balance.
Asunto(s)
Guanidina/metabolismo , Pirrolidinas/química , Receptor de Melanocortina Tipo 3/agonistas , Algoritmos , Animales , Evaluación Preclínica de Medicamentos , Metabolismo Energético/efectos de los fármacos , Guanidina/análogos & derivados , Guanidina/farmacología , Guanidina/uso terapéutico , Ensayos Analíticos de Alto Rendimiento , Humanos , Ratones , Ratones Noqueados , Isoformas de Proteínas/agonistas , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Pirrolidinas/metabolismo , Pirrolidinas/farmacología , Pirrolidinas/uso terapéutico , Receptor de Melanocortina Tipo 3/genética , Receptor de Melanocortina Tipo 3/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/metabolismo , Bibliotecas de Moléculas Pequeñas/farmacología , Bibliotecas de Moléculas Pequeñas/uso terapéutico , Relación Estructura-ActividadRESUMEN
The centrally expressed melanocortin-3 and -4 receptors (MC3R/MC4R) have been studied as possible targets for weight management therapies, with a preponderance of studies focusing on the MC4R. Herein, a novel tetrapeptide scaffold [Ac-Xaa1-Arg-(pI)DPhe-Xaa4-NH2] is reported. The scaffold was derived from results obtained from a MC3R mixture-based positional scanning campaign. From these results, a set of 48 tetrapeptides were designed and pharmacologically characterized at the mouse melanocortin-1, -3, -4, and -5 receptors. This resulted in the serendipitous discovery of nine compounds that were MC3R agonists (EC50 < 1000 nM) and MC4R antagonists (5.7 < pA2 < 7.8). The three most potent MC3R agonists, 18 [Ac-Arg-Arg-(pI)DPhe-Tic-NH2], 1 [Ac-His-Arg-(pI)DPhe-Tic-NH2], and 41 [Ac-Arg-Arg-(pI)DPhe-DNal(2')-NH2] were more potent (EC50 < 73 nM) than the melanocortin tetrapeptide Ac-His-DPhe-Arg-Trp-NH2. This template contains a sequentially reversed "Arg-(pI)DPhe" motif with respect to the classical "Phe-Arg" melanocortin signaling motif, which results in pharmacology that is first-in-class for the central melanocortin receptors.
Asunto(s)
Oligopéptidos/química , Oligopéptidos/farmacología , Receptor de Melanocortina Tipo 3/agonistas , Receptor de Melanocortina Tipo 4/antagonistas & inhibidores , Secuencia de Aminoácidos , Animales , Descubrimiento de Drogas , Ratones , Biblioteca de Péptidos , Receptor de Melanocortina Tipo 3/metabolismo , Receptor de Melanocortina Tipo 4/metabolismoRESUMEN
A novel series of endomorphin-1 (EM-1) and endomorphin-2 (EM-2) analogues was synthesized, incorporating chiral α-hydroxy-ß-phenylalanine (AHPBA), and/or Dmt(1)-Tic(2) at different positions. Pharmacological activity and metabolic stability of the series was assessed. Consistent with earlier studies of ß-amino acid substitution into endomorphins, multiple analogues incorporation AHPBA displayed high affinity for µ and δ opioid receptors (MOR and DOR, respectively) in radioligand competition binding assays, and an increased stability in rat brain membrane homogenates, notably Dmt-Tic-(2R,3S)AHPBA-Phe-NH2 (compound 26). Intracerebroventricular (i.c.v.) administration of 26 produced antinociception (ED50 value (and 95% confidence interval) = 1.98 (0.79-4.15) nmol, i.c.v.) in the mouse 55 °C warm-water tail-withdrawal assay, equivalent to morphine (2.35 (1.13-5.03) nmol, i.c.v.), but demonstrated DOR-selective antagonism in addition to non-selective opioid agonism. The antinociception of 26 was without locomotor activity or acute antinociceptive tolerance. This novel class of peptides adds to the potentially therapeutically relevant collection of previously reported EM analogues.
Asunto(s)
Dihidroxifenilalanina/análogos & derivados , Dihidroxifenilalanina/química , Oligopéptidos/síntesis química , Oligopéptidos/farmacología , Receptores Opioides delta/agonistas , Receptores Opioides delta/antagonistas & inhibidores , Receptores Opioides mu/agonistas , Animales , Células CHO , Cricetulus , Relación Dosis-Respuesta a Droga , Células HEK293 , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Conformación Molecular , Oligopéptidos/química , Ratas , Relación Estructura-ActividadRESUMEN
Human obesity has been linked to genetic factors and single nucleotide polymorphisms (SNPs). Melanocortin-4 receptor (MC4R) SNPs have been associated with up to 6% frequency in morbidly obese children and adults. A potential therapy for individuals possessing such genetic modifications is the identification of molecules that can restore proper receptor signaling and function. These compounds could serve as personalized medications improving quality of life issues as well as alleviating diseases symptoms associated with obesity including type 2 diabetes. Several hMC4 SNP receptors have been pharmacologically characterized in vitro to have a decreased, or a lack of response, to endogenous agonists such as α-, ß-, and γ2-melanocyte stimulating hormones (MSH) and adrenocorticotropin hormone (ACTH). Herein we report the use of a mixture based positional scanning combinatorial tetrapeptide library to discover molecules with nM full agonist potency and efficacy to the L106P, I69T, I102S, A219V, C271Y, and C271R hMC4Rs. The most potent compounds at all these hMC4R SNPs include Ac-His-(pI)DPhe-Tic-(pNO2)DPhe-NH2, Ac-His-(pCl)DPhe-Tic-(pNO2)DPhe-NH2, Ac-His-(pCl)DPhe-Arg-(pI)Phe-NH2, and Ac-Arg-(pCl)DPhe-Tic-(pNO2)DPhe-NH2, revealing new ligand pharmacophore models for melanocortin receptor drug design strategies.
Asunto(s)
Oligopéptidos/química , Receptor de Melanocortina Tipo 4/agonistas , Sustitución de Aminoácidos , Animales , Técnicas Químicas Combinatorias , Bases de Datos de Compuestos Químicos , Células HEK293 , Humanos , Ligandos , Ratones , Modelos Moleculares , Oligopéptidos/farmacología , Polimorfismo de Nucleótido Simple , Receptor de Melanocortina Tipo 4/genética , Relación Estructura-ActividadRESUMEN
ß-Sheet antimicrobial peptides (AMPs) are well recognized as promising candidates for the treatment of multidrug-resistant bacterial infections. To dissociate antimicrobial activity and hemolytic effect of ß-sheet AMPs, we hypothesize that N-methylation of the intramolecular hydrogen bond(s)-forming amides could improve their specificities for microbial cells over human erythrocytes. We utilized a model ß-sheet antimicrobial peptide, gramicidinâ S (GS), to study the N-methylation effects on the antimicrobial and hemolytic activities. We synthesized twelve N-methylated GS analogues by replacement of residues at the ß-strand and ß-turn regions with N-methyl amino acids, and tested their antimicrobial and hemolytic activities. Our experiments showed that the HC50 values increased fivefold compared with that of GS, when the internal hydrogen-bonded leucine residue was methylated. Neither hemolytic effect nor antimicrobial activity changed when proline alone was replaced with N-methylalanine in the ß-turn region. However, analogues containing N-methylleucine at ß-strand and N-methylalanine at ß-turn regions exhibited a fourfold increase in selectivity index compared to GS. We also examined the conformation of these N-methylated GS analogues using (1)Hâ NMR and circular dichroism (CD) spectroscopy in aqueous solution, and visualized the backbone structures and residue orientations using molecular dynamics simulations. The results show that N-methylation of the internal hydrogen bond-forming amide affected the conformation, backbone shape, and side chain orientation of GS.