Melatonin receptors are seven transmembrane-spanning proteins belonging to the GPCR superfamily. In mammals, two melatonin receptor subtypes exist -MT 1 and MT 2 -encoded by the MTNR1A and MTNR1B genes respectively. The current review provides an update on melatonin receptors by the corresponding subcommittee of the International Union of Basic and Clinical Pharmacology. We will highlight recent developments of melatonin receptor ligands, including radioligands, and give an update on the latest phenotyping results of melatonin receptor knockout mice. The current status and perspectives of the structure of melatonin receptor will be summarized. The physiological importance of melatonin receptor dimers and biologically important and type 2 diabetes-associated genetic variants of melatonin receptors will be discussed. The role of melatonin receptors in physiology and disease will be further exemplified by their functions in the immune system and the CNS. Finally, antioxidant and free radical scavenger properties of melatonin and its relation to melatonin receptors will be critically addressed.

Numerous physiological functions of the pineal gland hormone melatonin are mediated via activation of two G-protein-coupled receptors, MT1 and MT2. The melatonergic drugs on the market, ramelteon and agomelatine, as well as the most advanced drug candidates under clinical evaluation, tasimelteon and TIK-301, are high-affinity nonselective MT1/MT2 agonists. A great number of MT2-selective ligands and, more recently, several MT1-selective agents have been reported to date. Herein, we review recent advances in the field focusing on high-affinity agonists and antagonists and those displaying selectivity toward MT1 and MT2 receptors. Moreover, the existing models of MT1 and MT2 receptors as well as the current status in the emerging field of melatonin receptor oligomerization are critically discussed. In addition to the already existing indications, such as insomnia, circadian sleep disorders, and depression, new potential therapeutic applications of melatonergic ligands including cardiovascular regulation, appetite control, tumor growth inhibition, and neurodegenerative diseases are presented.

The dengue virus (DENV) and West Nile Virus (WNV) NS2B-NS3 proteases are attractive targets for the development of dual-acting therapeutics against these arboviral pathogens. We present the synthesis and extensive biological evaluation of inhibitors that contain benzyl ethers of 4-hydroxyphenylglycine as non-natural peptidic building blocks synthesized via a copper-complex intermediate. A three-step optimization strategy, beginning with fragment growth of the C-terminal 4-hydroxyphenylglycine to the benzyloxy ether, followed by C- and N-terminal optimization, and finally fragment merging generated compounds with in vitro affinities in the low nanomolar range. The most promising derivative reached Ki values of 12 nM at the DENV-2 and 39 nM at the WNV proteases. Several of the newly discovered protease inhibitors yielded a significant reduction of dengue and West Nile virus titers in cell-based assays of virus replication, with an EC50 value of 3.4 μM at DENV-2 and 15.5 μM at WNV for the most active analogue.

Diverse muscarinic allosteric ligands exhibit greatest affinity toward the M 2 receptor subtype and lowest affinity toward M 5 . In this study, we evaluated the potencies with which two groups of highly M 2 /M 5 selective allosteric agents modulate the dissociation of [3 H]N-methylscopolamine from M 2 /M 5 chimeric and point-mutated receptors. These allosteric ligands included two alkane-bisammonium compounds and a series of caracurine V derivatives, which are structurally closely related to (but stereochemically different from) the prototype allosteric ligand alcuronium. Like alcuronium, the caracurine V and alkanebisammonium compounds displayed significantly increased affinities compared with M 5 toward the chimera that included the M 2 second outer loop (o2) plus surrounding regions. Unlike alcuronium, the compounds had enhanced affinities for a chimera with M 2 sequence in transmembrane region (TM) 7; sitedirected mutagenesis in wild-type and chimeric receptors indicated that the threonine residue at M 2 423 was entirely responsible for the sensitivity toward TM7. Subsequent studies demonstrated that this TM7 epitope is likewise present in the M 4 receptor, as M 4 436 serine. The M 2 423 threonine residue is near the M 2 419 asparagine identified previously to influence gallamine binding. These studies demonstrate that a stereochemical difference can be sufficient to translate into divergent epitope sensitivities. Nonetheless, these allosteric ligands seem to derive affinity from two main regions of the receptor: o2 plus flanking regions and o3/TM7. These two epitopes are sufficient to explain the M 2 /M 5 selectivity of the presently investigated compounds; this is the first time that the subtype selectivity of muscarinic allosteric agents has been completely accounted for by distinct receptor epitopes.The five subtypes of muscarinic acetylcholine receptors are members of the superfamily of G protein-coupled receptors. The binding site for acetylcholine and conventional agonists and antagonists on muscarinic receptors seems to be located within a pocket formed by the seven ␣-helical transmembrane domains characteristic of all G protein-coupled receptors (Hulme et al., 1990;Wess, 1993). There is a high degree of conservation of amino acid sequence in the regions that are considered to bind agonists and antagonists. This may be a reason for the difficulty in developing agonists and antagonists that are highly subtype selective.Another feature of muscarinic receptors is the presence of a second, allosteric, binding site (Lee and El-Fakahany, 1991;Lazareno and Birdsall, 1995;Tucek and Proska, 1995;Ellis, 1997;Christopoulos et al., 1998;Holzgrabe and Mohr, 1998). Several observations have suggested that muscarinic allosteric ligands bind to more extracellular and presumably less conserved regions of the receptor, which may allow for greater selectivities. All five muscarinic receptor subtypes are subject to allosteric modulation (Ellis et al., 1991), and a wide array of structurally very different allost...

No abstract

Allosteric effects on muscarinic acetylcholine M(2) receptors were examined in a series of bisquaternary salts of the Strychnos alkaloid caracurine V (6) and related iso-caracurine V, tetrahydrocaracurine V, and bisnortoxiferine ring systems. The compounds inhibited dissociation of the orthosteric antagonist [(3)H]N-methylscopolamine (NMS) from porcine cardiac M(2) receptors with EC(0.5,diss) values from 4 to 3270 nM. The majority of compounds hardly changed [(3)H]NMS equilibrium binding, indicating similar binding affinities in free and NMS-occupied M(2) receptors. The most potent agents were found in the caracurine V, iso-caracurine V, and tetrahydrocaracurine V series and carried nonpolar alkyl groups with a maximal chain length of three carbon atoms. 3D QSAR (CoMSIA) analysis explained the wide range of binding affinities by steric and electrostatic properties of the side chains. Furthermore, the findings suggest that the spatial orientation of the "caracurine" aromatic rings compared with the bisnortoxiferine ring skeleton is favorable to optimal allostere-receptor interactions.

Tamoxifen is used to prevent and treat estrogen receptorpositive (ER1) breast cancer (BC); however, its chronic use can increase uterine cancer risk and induce tamoxifen resistance. Novel melatonin-tamoxifen drug conjugates may be promising to treat BC and may help offset the adverse effects of tamoxifen usage alone due to the presence of melatonin. We synthesized and screened five drug conjugates (C2, C4, C5, C9, and C15 linked) for their effects on BC cell (MCF-7, tamoxifenresistant MCF-7, mouse mammary carcinoma, MDA-MB-231, and BT-549) viability, migration, and binding affinity to melatonin receptor 1 (MT1R) and estrogen receptor 1 (ESR1). C4 and C5 demonstrated the most favorable pharmacological characteristics with respect to binding profiles (affinity for ESR1 and MT1R) and their potency/efficacy to inhibit BC cell viability and migration in four phenotypically diverse invasive ductal BC cell lines. C4 and C5 were further assessed for their actions against tamoxifen-resistant MCF-7 cells and a patient-derived xenograft triple-negative BC cell line (TU-BcX-4IC) and for their mechanisms of action using selective mitogen-activated protein kinase kinase MEK1/2, MEK5, and phosphoinositide 3-kinase (PI3K) inhibitors. C4 and C5 inhibited tamoxifen-resistant MCF-7 cells with equal potency (IC 50 5 4-8 mM) and efficacy (∼90% inhibition of viability and migration) but demonstrated increased potency (IC 50 5 80-211 mM) and efficacy (∼140% inhibition) to inhibit migration versus cell viability (IC 50 5 181-304 mM; efficacy ∼80% inhibition) in TU-BcX-4IC cells. Unique pharmacokinetic profiles were observed, with C4 having greater bioavailability than C5. Further assessment of C4 and C5 demonstrates that they create novel pharmacophores within each BC cell that is context specific and involves MEK1/2/ pERK1/2, MEK5/pERK5, PI3K, and nuclear factor kB. These melatonin-tamoxifen drug conjugates show promise as novel anticancer drugs and further preclinical and clinical evaluation is warranted.

Melatonin is a hormone exerting its multiple actions mainly through two G-protein-coupled receptors MT(1) and MT(2). Exploring the physiological role of each of these subtypes requires subtype selective MT(1) and MT(2) ligands. While several MT(2)-selective ligands were developed in the 1990s, no selective agonists and antagonists for the MT(1) subtype were described. The present article reviews mela toninergic ligands developed in the current millennium focusing on subtype selective agents and on drug candidates. Notable compounds are the MT(1)-selective agonists 35 and 134, MT(1)-selective antagonists 117 and 131, MT(2)-selective agonists 58, 70, 79, 97 and 125, MT(2)-selective antagonists 27, 73 and 119, and the highly potent non-selective agonist 120. The non-selective agonists agomelatine 2, and ramelteon 87 are drug candidates as antidepressive agent and for the treatment of insomnia and circadian rhythm disfunction, respectively.