Identification and characterization of pyrrolidine diastereoisomers as potent functional agonists and antagonists of the human melanocortin-4 receptor Chen Chen, a, * Wanlong Jiang, a Joe A. Tran, a Fabio C. Tucci, a,  Beth A. Fleck, b Stacy Markison, c Jenny Wen, d Ajay Madan, d Sam R. Hoare, b Alan C. Foster, c Dragan Marinkovic, a Caroline W. Chen, a Melissa Arellano a and John Saunders a,à a Department of Medicinal Chemistry, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA b Department of Pharmacology, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA c Department of Neuroscience, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA d Department of Preclinical Development, Neurocrine Biosciences, Inc., 12790 El Camino Real, San Diego, CA 92130, USA Received 25 September 2007; revised 30 October 2007; accepted 31 October 2007 Available online 4 November 2007 Abstract—A series of trans-4-phenylpyrrolidine-3-carboxamides were synthesized and characterized as potent ligands of the human melanocortin-4 receptor. Interestingly, a pair of diastereoisomers 13b displayed potent functional agonist and antagonist activity, respectively. Thus, the 3S,4R-pyrrolidine 13b-1 possessed a K i of 1.0 nM and an EC 50 of 3.8 nM, while its 3R,4S-isomer 13b-2 exhib- ited a K i of 4.7 and an IC 50 of 64 nM. Both compounds were highly selective over other melanocortin receptor subtypes. The MC4R agonist 13b-1 also demonstrated efficacy in a diet-induced obesity model in rats. Ó 2007 Elsevier Ltd. All rights reserved. The melanocortin receptor family belongs to the G-pro- tein-coupled receptor (GPCR) superfamily and consists of five receptor subtypes that are highly expressed in the brain and other tissues and are regulated by the endog- enous peptide agonists, melanocyte-stimulating hor- mones (a-, b-, and c-MSH) and adrenocorticotropic hormone (ACTH), and by the endogenous antagonists, agouti-protein and agouti-related protein. 1 The melano- cortin-4 receptor (MC4R) plays an important role in the regulation of feeding behavior, 2 and thus, a potent and selective MC4R agonist with brain penetration could perhaps be useful in the treatment of obesity. 3 Several small molecule MC4R agonists from different chemical classes have been discovered. 4 For example, pyrrolidine derivatives exemplified by 1 (Fig. 1) have been reported by Ujjainwalla to be potent and selective at MC4R. 5 We have previously reported that, by com- bining the trans-4-arylpyrrolidine-3-carbonyl moiety in 1 with the piperazinebenzylamine portion in 2, presented in a series of 3-aryl-2-methylpropionamide MC4R antagonists, 6 potent MC4R agonists such as 3 have been identified. 7 Here we report the optimization of this series of compounds and the in vitro characterization of the diastereoisomers 13b-1 and 13b-2 as a potent MC4R agonist and antagonist, respectively. In vivo studies of the agonist 13b-1 in feeding and obesity animal models are also discussed. The trans-N-isopropyl-4-arylpyrrolidin-3-ylcarboxylic esters 6 were synthesized from the corresponding methyl cinnamates and N-(trimethylsilylmethyl)-N-(methoxy- methyl)isopropylamine 5, which was prepared by the reaction of isopropylamine with trimethylsilylmethyl chloride 4, followed by formaldehyde and methanol, using a procedure similar to that described in the litera- ture. 8 Hydrolysis of 6 under basic conditions gave the 0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmcl.2007.10.115 Keywords: Melanocortin-4 receptor; Agonist; Antagonist; Potency; Stereochemistry; Pyrrolidine; Obesity; Animal model. * Corresponding author. Tel.: +1 858 617 7634; fax: +1 858 617 7602; e-mail: cchen@neurocrine.com   Present address: Department of Medicinal Chemistry, Tanabe Research Laboratories, USA, Inc., 4540 Towne Centre Ct., San Diego, CA 92121, USA. à Present address: Vertex Pharmaceuticals, Inc., 11010 Torreyana Road, San Diego, CA 92121, USA. Available online at www.sciencedirect.com Bioorganic & Medicinal Chemistry Letters 18 (2008) 129–136