RESUMO
SignificanceTirzepatide is a dual agonist of the glucose-dependent insulinotropic polypeptide receptor (GIPR) and the glucagon-like peptide-1 receptor (GLP-1R), which are incretin receptors that regulate carbohydrate metabolism. This investigational agent has proven superior to selective GLP-1R agonists in clinical trials in subjects with type 2 diabetes mellitus. Intriguingly, although tirzepatide closely resembles native GIP in how it activates the GIPR, it differs markedly from GLP-1 in its activation of the GLP-1R, resulting in less agonist-induced receptor desensitization. We report how cryogenic electron microscopy and molecular dynamics simulations inform the structural basis for the unique pharmacology of tirzepatide. These studies reveal the extent to which fatty acid modification, combined with amino acid sequence, determines the mode of action of a multireceptor agonist.
Assuntos
Diabetes Mellitus Tipo 2 , Receptores dos Hormônios Gastrointestinais , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Polipeptídeo Inibidor Gástrico/metabolismo , Polipeptídeo Inibidor Gástrico/farmacologia , Polipeptídeo Inibidor Gástrico/uso terapêutico , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Humanos , Incretinas/farmacologia , Receptores dos Hormônios Gastrointestinais/agonistas , Receptores dos Hormônios Gastrointestinais/metabolismo , Receptores dos Hormônios Gastrointestinais/uso terapêuticoRESUMO
OBJECTIVE: The central melanocortin system is essential for the regulation of food intake and body weight. Agouti-related protein (AgRP) is the sole orexigenic component of the central melanocortin system and is conserved across mammalian species. AgRP is currently known to be expressed exclusively in the mediobasal hypothalamus, and hypothalamic AgRP-expressing neurons are essential for feeding. Here we characterized a previously unknown population of AgRP cells in the mouse hindbrain. METHODS: Expression of AgRP in the hindbrain was investigated using gene expression analysis, single-cell RNA sequencing, immunofluorescent analysis and multiple transgenic mice with reporter expressions. Activation of AgRP neurons was achieved by Designer Receptors Exclusively Activated by Designer Drugs (DREADD) and by transcranial focal photo-stimulation using a step-function opsin with ultra-high light sensitivity (SOUL). RESULTS: AgRP expressing cells were present in the area postrema (AP) and the adjacent subpostrema area (SubP) and commissural nucleus of the solitary tract (cNTS) of the mouse hindbrain (termed AgRPHind herein). AgRPHind cells consisted of locally projecting neurons as well as tanycyte-like cells. Food deprivation stimulated hindbrain Agrp expression as well as neuronal activity of subsets of AgRPHind cells. In adult mice that lacked hypothalamic AgRP neurons, chemogenetic activation of AgRP neurons resulted in hyperphagia and weight gain. In addition, transcranial focal photo-stimulation of hindbrain AgRP cells increased food intake in adult mice with or without hypothalamic AgRP neurons. CONCLUSIONS: Our study indicates that the central melanocortin system in the hindbrain possesses an orexigenic component, and that AgRPHind neurons stimulate feeding independently of hypothalamic AgRP neurons.
Assuntos
Hipotálamo , Melanocortinas , Camundongos , Animais , Proteína Relacionada com Agouti/genética , Proteína Relacionada com Agouti/metabolismo , Hipotálamo/metabolismo , Camundongos Transgênicos , Melanocortinas/metabolismo , Rombencéfalo/metabolismo , Mamíferos/metabolismoRESUMO
GDF15 and its receptor GFRAL/RET form a non-homeostatic system that regulates food intake and body weight in preclinical species. Here, we describe a GDF15 analog, LY3463251, a potent agonist at the GFRAL/RET receptor with prolonged pharmacokinetics. In rodents and obese non-human primates, LY3463251 decreased food intake and body weight with no signs of malaise or emesis. In a first-in-human study in healthy participants, single subcutaneous LY3463251 injections showed a safety and pharmacokinetic profile supporting further clinical development with dose-dependent nausea and emesis in a subset of individuals. A subsequent 12-week multiple ascending dose study in overweight and obese participants showed that LY3463251 induced significant decreases in food intake and appetite scores associated with modest body weight reduction independent of nausea and emesis (clinicaltrials.gov: NCT03764774). These observations demonstrate that agonism of the GFRAL/RET system can modulate energy balance in humans, though the decrease in body weight is surprisingly modest, suggesting challenges in leveraging the GDF15 system for clinical weight-loss applications.
Assuntos
Obesidade , Redução de Peso , Animais , Humanos , Peso Corporal , Obesidade/tratamento farmacológico , Vômito , Fator 15 de Diferenciação de CrescimentoRESUMO
OBJECTIVE: Glucagon-like peptide-1 receptor agonists (GLP-1RAs) are effective medications to reduce appetite and body weight. These actions are centrally mediated; however, the neuronal substrates involved are poorly understood. METHODS: We employed a combination of neuroanatomical, genetic, and behavioral approaches in the mouse to investigate the involvement of caudal brainstem cholecystokinin-expressing neurons in the effect of the GLP-1RA exendin-4. We further confirmed key neuroanatomical findings in the non-human primate brain. RESULTS: We found that cholecystokinin-expressing neurons in the caudal brainstem are required for the anorectic and body weight-lowering effects of GLP-1RAs and for the induction of GLP-1RA-induced conditioned taste avoidance. We further show that, while cholecystokinin-expressing neurons are not a direct target for glucose-dependent insulinotropic peptide (GIP), GIP receptor activation results in a reduced recruitment of these GLP-1RA-responsive neurons and a selective reduction of conditioned taste avoidance. CONCLUSIONS: In addition to disclosing a neuronal population required for the full appetite- and body weight-lowering effect of GLP-1RAs, our data also provide a novel framework for understanding and ameliorating GLP-1RA-induced nausea - a major factor for withdrawal from treatment.
Assuntos
Colecistocinina/farmacologia , Polipeptídeo Inibidor Gástrico/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Animais , Apetite/efeitos dos fármacos , Depressores do Apetite/farmacologia , Glicemia/efeitos dos fármacos , Exenatida/farmacologia , Feminino , Glucagon/metabolismo , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Receptor do Peptídeo Semelhante ao Glucagon 1/fisiologia , Hipoglicemiantes/farmacologia , Insulina/farmacologia , Liraglutida/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Receptores dos Hormônios Gastrointestinais/metabolismoRESUMO
Here we show that central administration of pyroglutamylated arginine-phenylamine-amide peptide (QRFP/26RFa) increases both food intake and locomotor activity, without any significant effect on energy expenditure, thermogenesis or reward. Germline knock out of either of the mouse QRFP receptor orthologs, Gpr103a and Gpr103b, did not produce a metabolic phenotype. However, both receptors are required for the effect of centrally administered QRFP to increase feeding and locomotor activity. As central injection of QRFP activated orexin/hypocretin neurons in the lateral hypothalamus, we compared the action of QRFP and orexin on behaviour. Both peptides increased arousal and locomotor activity. However, while orexin increased consummatory behaviour, QRFP also affected other appetitive behaviours. Furthermore, the feeding but not the locomotor response to QRFP, was blocked by co-administration of an orexin receptor 1 antagonist. These results suggest that QRFP agonism induces both appetitive and consummatory behaviour, but only the latter is dependent on orexin/hypocretin receptor signalling.
Assuntos
Receptores de Orexina , Peptídeos , Receptores Acoplados a Proteínas G , Animais , Camundongos , Amidas , Compostos de Anilina , Arginina , Peptídeos e Proteínas de Sinalização Intercelular , Locomoção , Neuropeptídeos , Receptores de Orexina/metabolismo , Orexinas , Peptídeos/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Comportamento AlimentarRESUMO
The induction of nausea and emesis is a major barrier to maximizing the weight loss profile of obesity medications, and therefore, identifying mechanisms that improve tolerability could result in added therapeutic benefit. The development of peptide YY (PYY)-based approaches to treat obesity are no exception, as PYY receptor agonism is often accompanied by nausea and vomiting. Here, we sought to determine whether glucose-dependent insulinotropic polypeptide (GIP) receptor (GIPR) agonism reduces PYY-induced nausea-like behavior in mice. We found that central and peripheral administration of a GIPR agonist reduced conditioned taste avoidance (CTA) without affecting hypophagia mediated by a PYY analog. The receptors for GIP and PYY (Gipr and Npy2r) were found to be expressed by the same neurons in the area postrema (AP), a brainstem nucleus involved in detecting aversive stimuli. Peripheral administration of a GIPR agonist induced neuronal activation (cFos) in the AP. Further, whole-brain cFos analyses indicated that PYY-induced CTA was associated with augmented neuronal activity in the parabrachial nucleus (PBN), a brainstem nucleus that relays aversive/emetic signals to brain regions that control feeding behavior. Importantly, GIPR agonism reduced PYY-mediated neuronal activity in the PBN, providing a potential mechanistic explanation for how GIPR agonist treatment reduces PYY-induced nausea-like behavior. Together, the results of our study indicate a novel mechanism by which GIP-based therapeutics may have benefit in improving the tolerability of weight loss agents.
Assuntos
Fármacos Antiobesidade , Peptídeo YY , Receptores dos Hormônios Gastrointestinais , Animais , Fármacos Antiobesidade/efeitos adversos , Camundongos , Náusea/induzido quimicamente , Náusea/tratamento farmacológico , Obesidade/tratamento farmacológico , Peptídeo YY/efeitos adversos , Receptores dos Hormônios Gastrointestinais/agonistasRESUMO
The cytokine, GDF15, is produced in pathological states which cause cellular stress, including cancer. When over expressed, it causes dramatic weight reduction, suggesting a role in disease-related anorexia. Here, we demonstrate that the GDF15 receptor, GFRAL, is located in a subset of cholecystokinin neurons which span the area postrema and the nucleus of the tractus solitarius of the mouse. GDF15 activates GFRALAP/NTS neurons and supports conditioned taste and place aversions, while the anorexia it causes can be blocked by a monoclonal antibody directed at GFRAL or by disrupting CCK neuronal signalling. The cancer-therapeutic drug, cisplatin, induces the release of GDF15 and activates GFRALAP/NTS neurons, as well as causing significant reductions in food intake and body weight in mice. These metabolic effects of cisplatin are abolished by pre-treatment with the GFRAL monoclonal antibody. Our results suggest that GFRAL neutralising antibodies or antagonists may provide a co-treatment opportunity for patients undergoing chemotherapy.
Assuntos
Anorexia/genética , Tronco Encefálico/fisiologia , Fator 15 de Diferenciação de Crescimento/genética , Neurônios/fisiologia , Pica/genética , Transdução de Sinais , Animais , Colecistocinina/metabolismo , Fator 15 de Diferenciação de Crescimento/administração & dosagem , Fator 15 de Diferenciação de Crescimento/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/administração & dosagemRESUMO
Tirzepatide (LY3298176) is a dual GIP and GLP-1 receptor agonist under development for the treatment of type 2 diabetes mellitus (T2DM), obesity, and nonalcoholic steatohepatitis. Early phase trials in T2DM indicate that tirzepatide improves clinical outcomes beyond those achieved by a selective GLP-1 receptor agonist. Therefore, we hypothesized that the integrated potency and signaling properties of tirzepatide provide a unique pharmacological profile tailored for improving broad metabolic control. Here, we establish methodology for calculating occupancy of each receptor for clinically efficacious doses of the drug. This analysis reveals a greater degree of engagement of tirzepatide for the GIP receptor than the GLP-1 receptor, corroborating an imbalanced mechanism of action. Pharmacologically, signaling studies demonstrate that tirzepatide mimics the actions of native GIP at the GIP receptor but shows bias at the GLP-1 receptor to favor cAMP generation over ß-arrestin recruitment, coincident with a weaker ability to drive GLP-1 receptor internalization compared with GLP-1. Experiments in primary islets reveal ß-arrestin1 limits the insulin response to GLP-1, but not GIP or tirzepatide, suggesting that the biased agonism of tirzepatide enhances insulin secretion. Imbalance toward GIP receptor, combined with distinct signaling properties at the GLP-1 receptor, together may account for the promising efficacy of this investigational agent.
Assuntos
Glicemia/metabolismo , Polipeptídeo Inibidor Gástrico/farmacologia , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Hipoglicemiantes/farmacologia , Insulina/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Receptores dos Hormônios Gastrointestinais/agonistas , Animais , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Masculino , Camundongos , Camundongos Knockout , beta-Arrestina 1/fisiologiaRESUMO
Growth/differentiation factor-15 (GDF-15) is a distant member of the transforming growth factor ß (TGF-ß) superfamily and is widely expressed in multiple mammalian tissues. Its expression is highly regulated and is often induced in response to conditions associated with cellular stress. GDF15 serum levels have a strong association with many diseases, including inflammation, cancer, cardiovascular diseases, and obesity, and potentially serve as reliable predictor of disease progression. A functional role for GDF15 has been suggested in cancer, cardiovascular disease, kidney disease and metabolic disease. However, the knowledge of its pathophysiological function at the molecular level is still limited and requires more investigation. Recent identification of the endogenous receptor for GDF15 may provide additional insight in to its' molecular mechanisms and relationship to disease states.
RESUMO
The glucagon-like peptide-1 receptor (GLP-1R) is a class B G-protein coupled receptor (GPCR) that has proven to be an effective target for developing medicines that treat type 2 diabetes mellitus (T2DM). GLP-1R agonists improve T2DM by enhancing glucose-stimulated insulin secretion, delaying gastric transit, decreasing glucagon levels, and reducing body weight due to anorexigenic actions. The therapeutic successes of these agents helped inspire the design of new multifunctional molecules that are GLP-1R agonists but also activate receptors linked to pathways that enhance insulin sensitization and/or energy expenditure. Herein, these agents are discussed in the context of polypharmacological approaches that may enable even further improvement in treatment outcomes. Moreover, we revisit classical polypharmaceutical GPCR approaches and how they may be utilized for treatment of T2DM. To determine optimal combination regimens, changes in drug discovery practices are likely needed because compensatory mechanisms appear to underlie progression of T2DM and limit the ability of current therapies to induce disease regression or remission.
RESUMO
One approach of modern drug discovery is to identify agents that enhance or diminish signal transduction cascades in various cell types and tissues by modulating the activity of GPCRs. This strategy has resulted in the development of new medicines to treat many conditions, including cardiovascular disease, psychiatric disorders, HIV/AIDS, certain forms of cancer and Type 2 diabetes mellitus (T2DM). These successes justify further pursuit of GPCRs as disease targets and provide key learning that should help guide identifying future therapeutic agents. This report reviews the current landscape of GPCR drug discovery with emphasis on efforts aimed at developing new molecules for treating T2DM and obesity. We analyse historical efforts to generate GPCR-based drugs to treat metabolic disease in terms of causal factors leading to success and failure in this endeavour. LINKED ARTICLES: This article is part of a themed section on Molecular Pharmacology of GPCRs. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.21/issuetoc.
Assuntos
Descoberta de Drogas , Doenças Metabólicas/tratamento farmacológico , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Animais , Humanos , Doenças Metabólicas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
A series of 6-bicycloaryloxynicotinamides were identified as opioid receptor antagonists at mu, kappa, and delta receptors. Compounds in the 6-(2,3,4,5-tetrahydro-1H-benzo[c]azepin-7-yloxy)nicotinamide scaffold exhibited potent in vitro functional antagonism at all three receptors.
Assuntos
Amidas/química , Ácidos Carboxílicos/química , Éteres/química , Éteres/farmacologia , Antagonistas de Entorpecentes , Resinas Acrílicas/química , Éteres/síntese química , Estrutura Molecular , Receptores Opioides/metabolismo , Relação Estrutura-AtividadeRESUMO
The FATZO/Pco mouse is the result of a cross of the C57BL/6J and AKR/J strains. The crossing of these two strains and the selective inbreeding for obesity, insulin resistance and hyperglycemia has resulted in an inbred strain exhibiting obesity in the presumed presence of an intact leptin pathway. Routinely used rodent models for obesity and diabetes research have a monogenic defect in leptin signaling that initiates obesity. Given that obesity and its sequelae in humans are polygenic in nature and not associated with leptin signaling defects, the FATZO mouse may represent a more translatable rodent model for study of obesity and its associated metabolic disturbances. The FATZO mouse develops obesity spontaneously when fed a normal chow diet. Glucose intolerance with increased insulin levels are apparent in FATZO mice as young as 6 weeks of age. These progress to hyperglycemia/pre-diabetes and frank diabetes with decreasing insulin levels as they age. The disease in these mice is multi-faceted, similar to the metabolic syndrome apparent in obese individuals, and thus provides a long pre-diabetic state for determining the preventive value of new interventions. We have assessed the utility of this new model for the pre-clinical screening of agents to stop or slow progression of the metabolic syndrome to severe diabetes. Our assessment included: 1) characterization of the spontaneous development of disease, 2) comparison of metabolic disturbances of FATZO mice to control mice and 3) validation of the model with regard to the effectiveness of current and emerging anti-diabetic agents; rosiglitazone, metformin and semaglutide. CONCLUSION: Male FATZO mice spontaneously develop significant metabolic disease when compared to normal controls while maintaining hyperglycemia in the presence of high leptin levels and hyperinsulinemia. The disease condition responds to commonly used antidiabetic agents.
Assuntos
Glucose/metabolismo , Hipoglicemiantes/farmacologia , Tecido Adiposo/efeitos dos fármacos , Animais , Peso Corporal/efeitos dos fármacos , Modelos Animais de Doenças , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Homeostase/efeitos dos fármacos , Masculino , Camundongos , Triglicerídeos/sangueRESUMO
Obesity in many current pre-clinical animal models of obesity and diabetes is mediated by monogenic mutations; these are rarely associated with the development of human obesity. A new mouse model, the FATZO mouse, has been developed to provide polygenic obesity and a metabolic pattern of hyperglycemia and hyperinsulinemia, that support the presence of insulin resistance similar to metabolic disease in patients with insulin resistance/type 2 diabetes. The FATZO mouse resulted from a cross of C57BL/6J and AKR/J mice followed by selective inbreeding for obesity, increased insulin and hyperglycemia. Since many clinical studies have established a close link between higher body weight and the development of type 2 diabetes, we investigated whether time to progression to type 2 diabetes or disease severity in FATZO mice was dependent on weight gain in young animals. Our results indicate that lighter animals developed metabolic disturbances much slower and to a lesser magnitude than their heavier counterparts. Consumption of a diet containing high fat, accelerated weight gain in parallel with disease progression. A naturally occurring and significant variation in the body weight of FATZO offspring enables these mice to be identified as low, mid and high body weight groups at a young age. These weight groups remain into adulthood and correspond to slow, medium and accelerated development of type 2 diabetes. Thus, body weight inclusion criteria can optimize the FATZO model for studies of prevention, stabilization or treatment of type 2 diabetes.
Assuntos
Peso Corporal , Diabetes Mellitus Tipo 2/etiologia , Dieta Hiperlipídica/efeitos adversos , Obesidade/etiologia , Animais , Glicemia/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/fisiopatologia , Ingestão de Alimentos , Insulina/metabolismo , Leptina/metabolismo , Masculino , Camundongos , Obesidade/metabolismo , Obesidade/fisiopatologia , Pâncreas/metabolismoRESUMO
The increased prevalence of obesity and its cardiometabolic implications demonstrates the imperative to identify novel therapeutic targets able to effect meaningful metabolic changes in this population. Antibody-mediated targeting of fibroblast growth factor receptor 1c isoform (FGFR1c) has been shown to ameliorate hyperglycemia and protect from diet- and genetically-induced obesity in rodents and nonhuman primates. However, it is currently unknown which tissue(s) contribute to this glucose-lowering effect. Thus, to elucidate this effect, we treated euglycemic mice with H7, a monoclonal antibody that selectively targets FGFR1c, and used whole-body positron emission computed tomography with a glucose tracer (18F-fluorodeoxyglucose). Treatment with H7 increased basal glucose uptake in white adipose tissue (WAT), brown adipose tissue (BAT), the brain, and liver but reduced it in the quadriceps muscles. Consequentially, blood glucose was significantly reduced in response to treatment. Under insulin-stimulated conditions, the effects of H7 were maintained in WAT, BAT, liver, and muscle. Treatment with H7 decreased triglyceride (TG) content and increased adipose TG lipase content in white adipose tissue, while increasing activation of acetyl coenzyme A carboxylase, suggesting futile cycling of TGs, albeit favoring net hydrolysis. We demonstrated, in vitro, this is a direct effect of treatment in adipose tissue, as basal cellular respiration and glucose uptake were increased in response to treatment. Taken together, these data suggest that antibody-mediated targeting of FGFR1c exerts its powerful glucose-lowering efficacy primarily due to increased glucose uptake in adipose tissue.
Assuntos
Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Anticorpos Monoclonais/administração & dosagem , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Acetil-CoA Carboxilase/metabolismo , Animais , Anticorpos Monoclonais/uso terapêutico , Glicemia/análise , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Ativação Enzimática/efeitos dos fármacos , Glucose/metabolismo , Insulina/farmacologia , Lipase/análise , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Obesidade/metabolismo , Isoformas de Proteínas , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia , Triglicerídeos/metabolismoRESUMO
Growth/differentiation factor 15 (GDF15), also known as MIC-1, is a distant member of the transforming growth factor-ß (TGF-ß) superfamily and has been implicated in various biological functions, including cancer cachexia, renal and heart failure, atherosclerosis and metabolism. A connection between GDF15 and body-weight regulation was initially suggested on the basis of an observation that increasing GDF15 levels in serum correlated with weight loss in individuals with advanced prostate cancer. In animal models, overexpression of GDF15 leads to a lean phenotype, hypophagia and other improvements in metabolic parameters, suggesting that recombinant GDF15 protein could potentially be used in the treatment of obesity and type 2 diabetes. However, the signaling and mechanism of action of GDF15 are poorly understood owing to the absence of a clearly identified cognate receptor. Here we report that GDNF-family receptor α-like (GFRAL), an orphan member of the GFR-α family, is a high-affinity receptor for GDF15. GFRAL binds to GDF15 in vitro and is required for the metabolic actions of GDF15 with respect to body weight and food intake in vivo in mice. Gfral-/- mice were refractory to the effects of recombinant human GDF15 on body-weight, food-intake and glucose parameters. Blocking the interaction between GDF15 and GFRAL with a monoclonal antibody prevented the metabolic effects of GDF15 in rats. Gfral mRNA is highly expressed in the area postrema of mouse, rat and monkey, in accordance with previous reports implicating this region of the brain in the metabolic actions of GDF15 (refs. 4,5,6). Together, our data demonstrate that GFRAL is a receptor for GDF15 that mediates the metabolic effects of GDF15.
Assuntos
Área Postrema/metabolismo , Ingestão de Alimentos/efeitos dos fármacos , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/genética , Fator 15 de Diferenciação de Crescimento/farmacologia , Obesidade/metabolismo , Redução de Peso/efeitos dos fármacos , Animais , Encéfalo/metabolismo , Ingestão de Alimentos/genética , Citometria de Fluxo , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Células HEK293 , Humanos , Immunoblotting , Macaca fascicularis , Masculino , Camundongos , Camundongos Knockout , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Ressonância de Plasmônio de Superfície , Redução de Peso/genéticaRESUMO
A series of novel, disulfide-constrained human beta-melanocyte stimulating hormone (beta-MSH)-derived peptides were optimized for in vitro melanocortin-4 receptor (MC-4R) binding affinity, agonist efficacy, and selectivity. The most promising of these, analogue 18, was further studied in vivo using chronic rat food intake and body weight models.
Assuntos
Fármacos Antiobesidade/síntese química , Oligopeptídeos/síntese química , Receptor Tipo 4 de Melanocortina/agonistas , beta-MSH/química , Animais , Fármacos Antiobesidade/química , Fármacos Antiobesidade/farmacologia , Peso Corporal/efeitos dos fármacos , Linhagem Celular , Ingestão de Alimentos/efeitos dos fármacos , Humanos , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Ensaio Radioligante , Ratos , Relação Estrutura-AtividadeRESUMO
The melanocortin receptors have been implicated as potential targets for a number of important therapeutic indications, including inflammation, sexual dysfunction, and obesity. We identified compound 1, an arylpiperazine attached to the dipeptide H-d-Tic-d-p-Cl-Phe-OH, as a novel melanocortin subtype-4 receptor (MC4R) agonist through iterative directed screening of nonpeptidyl G-protein-coupled receptor biased libraries. Structure-activity relationship (SAR) studies demonstrated that substitutions at the ortho position of the aryl ring improved binding and functional potency. For example, the o-isopropyl-substituted compound 29 (K(i) = 720 nM) possessed 9-fold better binding affinity compared to the unsubstituted aryl ring (K(i) = 6600 nM). Sulfonamide 39 (K(i) = 220 nM) fills this space with a polar substituent, resulting in a further 2-fold improvement in binding affinity. The most potent compounds such as the diethylamine 44 (K(i) = 60 nM) contain a basic group at this position. Basic heterocycles such as the imidazole 50 (K(i) = 110 nM) were similarly effective. We also demonstrated good oral bioavailability for sulfonamide 39.
Assuntos
Piperazinas/síntese química , Receptor Tipo 4 de Melanocortina/agonistas , Animais , Ligação Competitiva , Disponibilidade Biológica , Linhagem Celular , AMP Cíclico/biossíntese , Humanos , Ligantes , Piperazinas/química , Piperazinas/farmacologia , Ensaio Radioligante , Ratos , Ratos Endogâmicos F344 , Receptor Tipo 4 de Melanocortina/metabolismo , Relação Estrutura-AtividadeRESUMO
Analogues of 3,4-dimethyl-4-(3-hydroxyphenyl)piperidines are high affinity inverse agonists for micro-, delta- and kappa-opioid receptors. To characterize inverse agonist binding, we synthesized a high specific activity radioligand from this series, [3H]LY515300 (3-[1-((3-cyclohexyl-[3,4-3H(2)])-3(R,S)-hydroxypropyl)-3(R),4(R)-dimethylpiperidin-4-yl]phenol). In membranes expressing cloned human opioid receptors, [3H]LY515300 binding was saturable and exhibited low nonspecific binding. [3H]LY515300 bound with high affinity to the micro- (K(d)=0.07 nM), delta- (K(d)=0.92 nM) and kappa-(K(d)=0.45 nM) opioid receptors. High affinity [3H]LY515300 binding to all opioid receptors was Na(+)-dependent, a characteristic of inverse agonists. Displacement by standard opioid compounds yielded K(i) values consistent with their known opioid receptor affinities. Autoradiographic localization of specific [3H]LY515300 binding in rat and guinea pig brain was high in areas known to express high levels of opioid (particularly micro-opioid receptor) binding sites including the caudate, nucleus accumbens, and nucleus tractus solitarius. Thus, [3H]LY515300 is the first radiolabeled opioid receptor inverse agonist useful for the study of opioid receptors in cell lines and native tissues.
Assuntos
Piperidinas/metabolismo , Receptores Opioides/agonistas , Receptores Opioides/metabolismo , Sódio/metabolismo , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Células CHO , Cricetinae , Relação Dose-Resposta a Droga , Cobaias , Humanos , Masculino , Piperidinas/química , Piperidinas/farmacologia , Ligação Proteica/efeitos dos fármacos , Ligação Proteica/fisiologia , Ratos , Ratos Sprague-Dawley , Sódio/farmacologia , Trítio/metabolismoRESUMO
Differences in the anorectic activity of morphinan (e.g., naltrexone) and 3,4-dimethyl-4-(3-hydroxyphenyl)piperidine (4PP) opioid receptor antagonists have been described. In an attempt to explain these differences, the influence of Na(+) on opioid binding affinity and functional activity of 4PP antagonists was compared to other opioid antagonists. The binding affinities of neutral antagonists were unaffected by the addition of Na(+), whereas that for the peptide, inverse agonist N,N-diallyl-Tyr-Aib-Aib-Phe-Leu-OH (ICI174864) was increased. Similarly, the binding affinities of the 4PP antagonist (3R,4R)-1-((S)-3-hydroxy-3-cyclohexylpropyl)-4-(3-hydroxyphenyl)-3,4-dimethyl-1-piperidine (LY255582) and other 4PP antagonists were increased in the presence of Na(+) with the greatest effects at the delta opioid receptor followed by the mu and kappa opioid receptors, respectively. Similar to ICI174864, 4PP antagonists were found to inhibit basal GTPgamma[(35)S] binding at the delta opioid receptor indicating inverse agonist activity. A correlation was observed between the binding affinities in the presence of Na(+), the inverse agonist potency, and the anorectic potency of 4PP antagonists. These data suggest that 4PP antagonists differ from morphinan antagonists in their inverse agonist activity and suggest a relationship between inverse agonism and anorectic activity.