MCH receptor peptide agonists and antagonists

Melanin-concentrating hormone (MCH) is an important neuropeptide hormone involved in multiple physiological processes. Peptide derivatives of MCH have been developed as tools to aid research including potent radioligands, receptor selective agonists, and potent antagonists. These tools have been used to further understand the role of MCH in physiology, primarily in rodents. However, the tools could also help elucidate the role for MCHR1 and MCHR2 in mediating MCH signaling in higher species.     

The discovery and optimization of pyrimidinone-containing MCH R1 antagonists

Optimization of a series of constrained melanin-concentrating hormone receptor 1 (MCH R1) antagonists has provided compounds with potent and selective MCH R1 activity. Details of the optimization process are provided and the use of one of the compounds in an animal model of diet-induced obesity is presented.     

Antidepressant,anxiolytic and anorectic effects of a melanin-concentrating hormone-1 receptor antagonist

Melanin concentrating hormone (MCH) is an orexigenic hypothalamic neuropeptide, which plays an important role in the complex regulation of energy balance and body weight. Here we show that SNAP-7941, a selective, high-affinity MCH1 receptor (MCH1-R) antagonist, inhibited food intake stimulated by central administration of MCH, reduced consumption of palatable food, and, after chronic administration to rats with diet-induced obesity, resulted in a marked, sustained decrease in body weight. In addition, after mapping the binding sites for [(3)H]SNAP-7941 in rat brain, we evaluated its effects in a series of behavioral models. SNAP-7941 produced effects similar to clinically used antidepressants and anxiolytics in three animal models of depression/anxiety: the rat forced-swim test, rat social interaction and guinea pig maternal-separation vocalization tests. Given these observations, an MCH1-R antagonist may be useful not only in the management of obesity but also as a treatment for depression and/or anxiety.     

Discovery of tetralin ureas as potent melanin concentrating hormone 1 receptor antagonists

Melanin concentrating hormone (MCH) plays an important role in the regulation of food intake and energy balance in mammals. MCH-1 receptor (MCH1R) deficient mice are lean and resistant to diet-induced obesity. As such, MCH1R antagonists are believed to have potential as possible treatments for obesity. The discovery of a novel class of tetralin ureas as potent MCH1R antagonists is described herein.     

Design and synthesis of substituted quinolines as novel and selective melanin concentrating hormone antagonists as anti-obesity agents

A novel series of substituted quinoline analogs were designed and synthesized as potent and selective melanin concentrating hormone (MCH) antagonists. These analogs show potent (nM) activity (12a-k) with a moderate selectivity. Conversely, the conformationally constrained thienopyrimidinone analogs (18a-g) showed improved activity in MCH-1R and selectivity over 5HT2C.     

Redundancy of a functional melanocortin 1 receptor in the anti-inflammatory actions of melanocortin peptides: studies in the recessive yellow (e/e) mouse suggest an important role for melanocortin 3 receptor

The issue of which melanocortin receptor (MC-R) is responsible for the anti-inflammatory effects of melanocortin peptides is still a matter of debate. Here we have addressed this aspect using a dual pharmacological and genetic approach, taking advantage of the recent characterization of more selective agonists/antagonists at MC1 and MC3-R as well as of the existence of a naturally defective MC1-R mouse strain, the recessive yellow (e/e) mouse. RT-PCR and ultrastructural analyses showed the presence of MC3-R mRNA and protein in peritoneal macrophages (M phi) collected from recessive yellow (e/e) mice and wild-type mice. This receptor was functional as Mphi incubation (30 min) with melanocortin peptides led to accumulation of cAMP, an effect abrogated by the MC3/4-R antagonist SHU9119, but not by the selective MC4-R antagonist HS024. In vitro M phi activation, determined as release of the CXC chemokine KC and IL-1 beta, was inhibited by the more selective MC3-R agonist gamma(2)-melanocyte stimulating hormone but not by the selective MC1-R agonist MS05. Systemic treatment of mice with a panel of melanocortin peptides inhibited IL-1 beta release and PMN accumulation elicited by urate crystals in the murine peritoneal cavity. MS05 failed to inhibit any of the inflammatory parameters either in wild-type or recessive yellow (e/e) mice. SHU9119 prevented the inhibitory actions of gamma(2)-melanocyte stimulating hormone both in vitro and in vivo while HS024 was inactive in vivo. In conclusion, agonism at MC3-R expressed on peritoneal M phi leads to inhibition of experimental nonimmune peritonitis in both wild-type and recessive yellow (e/e) mice.     

Differential regulation of melanin-concentrating hormone and orexin genes in the agouti-related protein/melanocortin-4 receptor system

Agouti protein and agouti-related protein (AGRP) antagonize alpha-melanocyte-stimulating hormone that binds to and activates the melanocortin-4 receptor (MC4-R) in the hypothalamus, thereby stimulating food intake. Melanin-concentrating hormone (MCH) and orexin are orexigenic peptides that specifically are synthesized in the lateral hypothalamus. MCH gene expression was augmented in A(y)/a (agouti) mice which overexpress agouti protein, but orexin mRNA was not. AGRP administered intracerebroventricularly into wild-type rats augmented MCH but not orexin gene expression. Also, SHU9119, a peptidergic antagonist of MC4-R, increased only MCH mRNA. These findings indicate that interruption of signaling at MC4-R activates the MCH but not the orexin gene. The biosyntheses of MCH and orexin are regulated through different pathways.     

Building a MCHR1 homology model provides insight into the receptor-antagonist contacts that are important for the development of new anti-obesity agents

Melanin-concentrating hormone (MCH) regulates feeding and energy homeostasis through interaction with its receptor, the melanin-concentrating receptor 1 (MCHR1), making it a target in the treatment of obesity. Molecular modeling and docking studies were performed in order to find a binding model for the docking of two new series of MCHR1 antagonists to the receptor. Results suggested interactions between the ligands and two glutamines (Gln5.42 and Gln6.55) not conserved in many of the GPCRs family members. Histamine 3 receptor (HRH3) presents two apolar residues in the aforementioned positions and the available biological data against this receptor supported the role of the two glutamines in the binding of antagonists to the MCHR1. This knowledge could be useful in the development of new, more active and more selective MCHR1 antagonists.     

Expression of receptors for melanin-concentrating hormone (MCH) in different tissues and cell lines

Melanin-concentrating hormone (MCH) is a potent orexigenic neuropeptide and a physiological antagonist of alpha-melanocyte-stimulating hormone (alpha-MSH) in the brain as well as at peripheral sites, including the pigmentary systems of specific vertebrates. Two receptor subtypes for MCH, MCH-R1 and MCH-R2, have been cloned, but other receptor subtypes are likely to exist. Based on our own data and the current literature, we have compared the expression of different receptors for MCH in various mammalian cell lines and tissues. Summarizing all data currently available, we conclude that the two cloned MCH receptors, MCH-R1 and MCH-R2, exhibit differences in their expression pattern, although MCH-R1 is generally colocalized in all tissues where MCH-R2 expression is found. It appears that MCH-R1 is more abundant and has a wider distribution pattern than MCH-R2. Other hypothetical MCH-R subtypes may be expressed in specific tissues, e.g., in the pigment cell system.     

Interaction of melanin-concentrating hormone (MCH), neuropeptide E-I (NEI), neuropeptide G-E (NGE), and alpha-MSH with melanocortin and MCH receptors on mouse B16 melanoma cells.

Melanin-concentrating hormone (MCH) and alpha-melanocyte-stimulating hormone (alpha-MSH) are known to exhibit mostly functionally antagonistic, but in some cases agonistic activities, e.g., in pigment cells and in the brain. Neuropeptide E-I (NEI) displays functional MCH-antagonist and MSH-agonist activity in different behavioral paradigms; the role of neuropeptide G-E (NGE) is not known. This study addressed the question of possible molecular interactions between alpha-MSH, MCH and the MCH-precursor-derived peptides NEI and NGE at the level of the pigment cell MCH receptor subtype (MCH-Rpc) and the different melanocortin (MC) receptors. Radioreceptor assays using [125I]MCH, [125l]alpha-MSH and [125I]NEI as radioligands and bioassays were performed with MCI-R-positive and MC1-R-negative mouse B16 melanoma cells and with COS cells expressing the different MC receptors. The IC50s of alpha-MSH and NEI or NGE for [125I]MCH displacement from mouse MCH-Rpc were 80-fold and, respectively, >300-fold higher than that of MCH, and the IC50s for MCH and NEI or NGE for [125I]alpha-MSH displacement from mouse MC1-R were 50,000-fold and >200,000-fold higher than that of alpha-MSH. No high-affinity binding sites for NEI were detected on B16 melanoma cells and there was no significant displacement of [1251]alpha-MSH by MCH, NEI or NGE with MC3-R, MC4-R and MC5-R expressed in COS cells. At concentrations of 100 nM to 10 microM, however, MCH, NEI and NGE induced cAMP formation and melanin synthesis which could be blocked by agouti protein or inhibitors of adenylate cyclase or protein kinase A. This shows that mammalian MCH-precursor-derived peptides may mimic MSH signalling via MC1-R activation at relatively high, but physiologically still relevant concentrations, as e.g. found in autocrine/paracrine signalling mechanisms.     

SVK14 cells express an MCH binding site different from the MCH1 or MCH2 receptor

Melanin-concentrating hormone (MCH) is a cyclic peptide, mainly involved in the regulation of skin pigmentation in teleosts and feeding behavior in mammals. The human keratinocyte SVK14 cell line has been previously shown to express binding sites for the MCH analog [125I]-[Phe13,3-iodo-Tyr19]MCH. We report here that: (1) this binding site similarly recognized [125I]-[3-iodo-Tyr13]MCH; (2) its pharmacological profile clearly differed from those observed at the two human MCH receptor subtypes, MCH1-R and MCH2-R; (3) MCH did not induce any effect on second messenger systems (including cAMP, calcium, and MAP kinase signaling pathways), and (4) no mRNAs corresponding to the MCH receptors were found. In conclusion, the binding site characterized in the SVK14 cell line is distinct from the MCH1 and MCH2 receptors and deserves therefore further investigation.     

Discovery and SAR of 4-amino-2-biarylbutylurea MCH 1 receptor antagonists through solid-phase parallel synthesis

-4-Amino-2-arylbutylbenzamides such as 1 were identified as micromolar MCH 1 receptor (MCH1R) antagonists via screening using a scintillation proximity assay based on [125I]-MCH binding to recombinant, human MCH1R. Subsequent lead optimization efforts using solid-phase parallel synthesis resulted in the defined structure-activity relationships and the identification of 4-amino-2-biarylbutylureas, such as 11g, as potent single digit nanomolar MCH1R antagonists.     

Identification of melanin-concentrating hormone receptor and its impact on drug discovery

The neuropeptide melanin-concentrating hormone (MCH) was originally isolated from the pituitary of salmon, in which it causes skin paling. MCH is also found abundantly in mammalian neurons, and has been detected in the lateral hypothalamus and zona incerta, brain regions that are at the center of feeding behavior. Acute central administration of MCH leads to a rapid and significant increase in food intake, while MCH expression changes in states of altered energy balance, such as fasting and obesity. Furthermore, MCH knockout mice tend toward hypophagia and leanness. In 1999, we and four other groups identified an orphan G-protein-coupled receptor (GPCR) as a specific receptor for MCH (MCH-1 receptor). Although a second MCH receptor (MCH-2 receptor) was isolated in humans, it was found to be non-functional or encode a non-functional pseudogene in non-human species, including rodents. The discovery of these MCH receptors permitted the launch of a broad array of drug screening efforts and three MCH-1 receptor antagonists were identified to reduce food intake and body weight. Interestingly, some antagonists unexpectedly produced evidence that blockade of these receptors has antidepressant and anxiolytic activities. The expressions of the MCH receptors, which have been implicated in regulating emotion, stress and motivation, make MCH an excellent candidate for integrating the various homeostatic stimuli necessary for maintaining the proper conditions of energy metabolism and other physiological functions. Finally, the speed at which MCH receptor studies have been undertaken exemplifies the impact that this deorphanized GPCR will have on setting the stage for more detailed physiological studies.     

Identification and characterization of a selective radioligand for melanin-concentrating hormone 1-receptor (MCH1R)

We have developed and characterized [³⁵S]4a as a potent and selective radioligand for melanin-concentrating hormone 1-receptor (MCH1R). Compound [³⁵S]4a showed appreciable specific signals in brain slices prepared from wild-type mice but not from MCH1R deficient mice, confirming the specificity and utility of [³⁵S]4a as a selective MCH1R radioligand for ex vivo receptor occupancy assays.     

Bicyclic[4.1.0]heptanes as phenyl replacements for melanin concentrating hormone receptor antagonists

Melanin concentrating hormone (MCH) receptor antagonists have been proposed as potential treatments of obesity. MCH receptor antagonists with a biphenylamine subunit have been reported previously at Schering-Plough. Herein, we report the discovery of bicyclo[4.1.0]heptanes as replacements for the middle phenyl ring of the biphenylamine moiety in order to eliminate its potential mutagenic liability. Structure-activity relationships in this series were found to be very similar to those of the original biphenylamine series, suggesting that the two series have similar binding modes.     

The kinin system mediates hyperalgesia through the inducible bradykinin B1 receptor subtype: evidence in various experimental animal models of type 1 and type 2 diabetic neuropathy

Both insulin-dependent (type 1) and insulin-independent (type 2) diabetes are complex disorders characterized by symptomatic glucose intolerance due to either defective insulin secretion, insulin action or both. Unchecked hyperglycemia leads to a series of complications among which is painful diabetic neuropathy, for which the kinin system has been implicated. Here, we review and compare the profile of several experimental models of type 1 and 2 diabetes (chemically induced versus gene-prone) and the incidence of diabetic neuropathy upon aging. We discuss the efficacy of selective antagonists of the inducible bradykinin B1 receptor (BKB1-R) subtype against hyperalgesia assessed by various nociceptive tests. In either gene-prone models of type 1 and 2 diabetes, the incidence of hyperalgesia mostly precedes the development of hyperglycemia. The administration of insulin, achieving euglycemia, does not reverse hyperalgesia. Treatment with a selective BKB1-R antagonist does not affect basal nociception in most normal control rats, whereas it induces a significant time- and dose-dependent attenuation of hyperalgesia, or even restores nociceptive responses, in experimental diabetic neuropathy models. Diabetic hyperalgesia is absent in streptozotocin-induced type 1 diabetic BKB1-R knockout mice. Thus, selective antagonism of the inducible BKB1-R subtype may constitute a novel and potential therapeutic approach for the treatment of painful diabetic neuropathy.     

EXPRESSION OF RECEPTORS FOR MELANIN-CONCENTRATING HORMONE (MCH) IN DIFFERENT TISSUES AND CELL LINES

ABSTRACT

Melanin-concentrating hormone (MCH) is a potent orexigenic neuropeptide and a physiological antagonist of α-melanocyte-stimulating hormone (α-MSH) in the brain as well as at peripheral sites, including the pigmentary systems of specific vertebrates. Two receptor subtypes for MCH, MCH-R₁ and MCH-R₂, have been cloned, but other receptor subtypes are likely to exist. Based on our own data and the current literature, we have compared the expression of different receptors for MCH in various mammalian cell lines and tissues. Summarizing all data currently available, we conclude that the two cloned MCH receptors, MCH-R₁ and MCH-R₂, exhibit differences in their expression pattern, although MCH-R₁ is generally colocalized in all tissues where MCH-R₂ expression is found. It appears that MCH-R₁ is more abundant and has a wider distribution pattern than MCH-R₂. Other hypothetical MCH-R subtypes may be expressed in specific tissues, e.g., in the pigment cell system.     

Structure-activity relationship studies of melanin-concentrating hormone (MCH)-related peptide ligands at SLC-1, the human MCH receptor

Melanin-concentrating hormone (MCH) is a cyclic nonadecapeptide involved in the regulation of feeding behavior, which acts through a G protein-coupled receptor (SLC-1) inhibiting adenylcyclase activity. In this study, 57 analogues of MCH were investigated on the recently cloned human MCH receptor stably expressed in HEK293 cells, on both the inhibition of forskolin-stimulated cAMP production and guanosine-5'-O-(3-[(35)S]thiotriphosphate ([(35)S]- GTPgammaS) binding. The dodecapeptide MCH-(6-17) (MCH ring between Cys(7) and Cys(16), with a single extra amino acid at the N terminus (Arg(6)) and at the C terminus (Trp(17))) was found to be the minimal sequence required for a full and potent agonistic response on cAMP formation and [(35)S]- GTPgammaS binding. We Ala-scanned this dodecapeptide and found that only 3 of 8 amino acids of the ring, namely Met(8), Arg(11), and Tyr(13), were essential to elicit full and potent responses in both tests. Deletions inside the ring led either to inactivity or to poor antagonists with potencies in the micromolar range. Cys(7) and Cys(16) were substituted by Asp and Lys or one of their analogues, in an attempt to replace the disulfide bridge by an amide bond. However, those modifications were deleterious for agonistic activity. In [(35)S]- GTPgammaS binding, these compounds behaved as weak antagonists (K(B) 1-4 microm). Finally, substitution in MCH-(6-17) of 6 out of 12 amino acids by non-natural residues and concomitant replacement of the disulfide bond by an amide bond led to three compounds with potent antagonistic properties (K(B) = 0.1-0.2 microm). Exploitation of these structure-activity relationships should open the way to the design of short and stable MCH peptide antagonists.     

INTERACTION OF MELANIN-CONCENTRATING HORMONE (MCH), NEUROPEPTIDE E-I (NEI), NEUROPEPTIDE G-E (NGE), AND α-MSH WITH MELANOCORTIN AND MCH RECEPTORS ON MOUSE B16 MELANOMA CELLS

Melanin-concentrating hormone (MCH) and α-melanocyte-stimulating hormone (α-MSH) are known to exhibit mostly functionally antagonistic, but in some cases agonistic activities, e.g., in pigment cells and in the brain. Neuropeptide E-I (NEI) displays functional MCH-antagonist and MSH-agonist activity in different behavioral paradigms; the role of neuropeptide G-E (NGE) is not known. This study addressed the question of possible molecular interactions between α-MSH, MCH and the MCH-precursor-derived peptides NEI and NGE at the level of the pigment cell MCH receptor subtype (MCH-R_pc) and the different melanocortin (MC) receptors. Radioreceptor assays using [¹²⁵I]MCH, [¹²⁵I]α-MSH and [¹²⁵I]NEI as radioligands and bioassays were performed with MC1-R-positive and MC1-R-negative mouse B16 melanoma cells and with COS cells expressing the different MC receptors. The IC₅₀s of α-MSH and NEI or NGE for [¹²⁵I]MCH displacement from mouse MCH-R_pc were 80-fold and, respectively, > 300-fold higher than that of MCH, and the IC₅₀s for MCH and NEI or NGE for [¹²⁵I]α-MSH displacement from mouse MC1-R were 50,000-fold and > 200,000-fold higher than that of α-MSH. No high-affinity binding sites for NEI were detected on B16 melanoma cells and there was no significant displacement of [¹²⁵I]α-MSH by MCH, NEI or NGE with MC3-R, MC4-R and MC5-R expressed in COS cells. At concentrations of 100 nM to 10 μM, however, MCH, NEI and NGE induced cAMP formation and melanin synthesis which could be blocked by agouti protein or inhibitors of adenylate cyclase or protein kinase A. This shows that mammalian MCH-precursor-derived peptides may mimic MSH signalling via MC1-R activation at relatively high, but physiologically still relevant concentrations, as e.g. found in autocrine/paracrine signalling mechanisms.     

Design and optimization of quinazoline derivatives as melanin concentrating hormone receptor 1 (MCHR1) antagonists

Melanin concentrating hormone (MCH) is an important mediator of energy homeostasis and plays a role in metabolic and CNS disorders. The modeling-supported design, synthesis and multi-parameter optimization (biological activity, solubility, metabolic stability, hERG) of novel quinazoline derivatives as MCHR1 antagonists are described. The in vivo proof of principle for weight loss with a lead compound from this series is exemplified. Clusters of refined hMCHR1 homology models derived from the X-ray structure of the β2-adrenergic receptor, including extracellular loops, were developed and used to guide the design.