Design and Synthesis of N-(3,3-Diphenylpropenyl)alkanamides as a Novel Class of High-Affinity MT 2 -Selective Melatonin Receptor Ligands Annalida Bedini, ² Gilberto Spadoni,* ,² Giuseppe Gatti, ² Simone Lucarini, ² Giorgio Tarzia, ² Silvia Rivara, ‡ Simone Lorenzi, ‡ Alessio Lodola, ‡ Marco Mor, ‡ Valeria Lucini, § Marilou Pannacci, § and Francesco Scaglione § Istituto di Chimica Farmaceutica e Tossicologica, UniVersita ` degli Studi di Urbino “Carlo Bo”, Piazza Rinascimento 6, 61029 Urbino, Italy, Dipartimento Farmaceutico, UniVersita ` degli Studi di Parma, V.le G. P. Usberti 27/A Campus UniVersitario, 43100 Parma, Italy, and Dipartimento di Farmacologia, Chemioterapia e Tossicologia Medica, UniVersita ` degli Studi di Milano, Via VanVitelli 32, 20129 Milano, Italy ReceiVed July 20, 2006 A novel series of melatonin receptor ligands was discovered by opening the cyclic scaffolds of known classes of high affinity melatonin receptor antagonists, while retaining the pharmacophore elements postulated by previously described 3D-QSAR and receptor models. Compounds belonging to the classes of 2,3- and [3,3-diphenylprop(en)yl]alkanamides and of o- or [(m-benzyl)phenyl]ethyl-alkanamides were synthesized and tested on MT 1 and MT 2 receptors. The class of 3,3-diphenyl-propenyl-alkanamides was the most interesting one, with compounds having MT 2 receptor affinity similar to that of MLT, remarkable MT 2 selectivity, and partial agonist or antagonist behavior. In particular, the (E)-m-methoxy cyclobutanecar- boxamido derivative 18f and the di-(m-methoxy) acetamido one, 18g, have sub-nM affinity for the MT 2 subtype, with more than 100-fold selectivity over MT 1 , 18f being an antagonist and 18g a partial agonist on GTPγS test. Docking of 18g into a previously developed MT 2 receptor model showed a binding scheme consistent with that of other antagonists. The MT 2 expected binding affinities of the new compounds were calculated by a previously developed 3D-QSAR CoMFA model, giving satisfactory predictions. Introduction Melatonin (N-acetyl-5-methoxytryptamine, MLT, 1) is a neurohormone primarily secreted by the pineal gland at night in all species. 1 The circadian pattern of MLT secretion, coupled with the localization of specific MLT binding sites in the brain region associated with the “biological clock”, suggests that MLT may play an important role in modulation of the sleep-wake cycle and of circadian rhythms in humans. 2 Some MLT receptor agonists, with improved properties in comparison to MLT, are now in clinical trials for treatment of insomnia or circadian rhythm sleep disorders, 3-5 and ramelteon (TAK-375) has been recently approved for insomnia. 6 Other effects of MLT described in the literature include its anti-inflammatory, 7 pain modulatory, 8 retinal, 9 vascular, 10 antitumor, 11 and antioxidant 12 properties. A remarkable efficacy of MLT in animal models of focal cerebral ischemia had also been reported, suggesting the hormone as a candidate neuroprotective drug for human stroke. 13 However, the functions of MLT in mammals are still a matter of investigation, and more rigorous clinical studies are needed to demonstrate the potential benefits of MLT assumption and to rule out the possibility of toxic effects. Most physiological MLT effects result from the activation of high affinity G protein-coupled receptors. MT 1 and MT 2 receptors 14-16 have been found in mammals, including humans, and subsequently cloned. A third subtype (Mel 1c ), first cloned from Xenopous laeVis, has been found only in nonmammalian species. 14 In addition to these high-affinity MLT receptors (K i = 0.1 nM), another low-affinity MLT binding site, termed MT 3 (K i = 60 nM), has recently been characterized as a melatonin- sensitive form of the human enzyme quinone reductase 2. 17 Whereas it is known that MT 1 and MT 2 receptors are expressed both centrally (suprachiasmatic nucleus, cortex, pars tuberalis, etc.) and peripherally (kidney, adipocytes, retina, blood vessels, etc.), 18 the physiological roles of these receptors are not as yet well defined. There is evidence that MT 1 receptors might be implicated in the sleep promoting effects of MLT 19,20 and in mediating vasoconstriction, 21 whereas MT 2 receptors appear to play a major role in the resynchronizing activity of MLT 19,22 and in mediating vasodilation. Moreover, an antidepressive effect has been reported for the antagonist luzindole in a mice model, being ascribed to its selective action at the MT 2 receptor. 23 However, an accurate characterization of MLT receptors-mediated functions in native tissues can only be made by using subtype-selective ligands. Therefore, a substantial share of current drug discovery efforts in the melatonin area is being directed toward the development of subtype-selective MLT receptor agonists and antagonists, which will be valuable tools in understanding the role of this enigmatic hormone in health and disease. In the course of our studies, various modifications of the MLT structure had been examined to determine which structural features are required for receptor affinity, intrinsic activity, and/ or subtype selectivity at MLT receptors. 24,25 However, marked subtype selectivity is still a challenge, and only recently this field has registered some important advances, leading to the identification of a small number of selective compounds. 26 While only a few examples of MT 1 27 and MT 3 28 ligands have been reported, the majority of subtype-selective compounds behave as MT 2 receptor antagonists. These compounds belong to different structural classes (Figure 1) and display various degrees of binding affinity and selectivity. A consistent structural motif found in most of these MLT ligands is the presence of a lipophilic substituent, which can be located out-of-the-plane of their core nucleus (i.e., the indole ring in luzindole or the tetralin scaffold in 4P-PDOT) in a position corresponding to positions 1 and 2 of the indole in MLT, and we had hypothesized that this arrangement confers selectivity for the MT 2 receptor and * To whom correspondence should be addressed. Tel.: ++39 0722 303323. Fax: ++39 0722 303313. E-mail: gilberto@uniurb.it. ² Universita ` degli Studi di Urbino “Carlo Bo”. ‡ Universita ` degli Studi di Parma. § Universita ` degli Studi di Milano. 7393 J. Med. Chem. 2006, 49, 7393-7403 10.1021/jm060850a CCC: $33.50 © 2006 American Chemical Society Published on Web 11/15/2006