Anatoly Mazurov

Selective α7 Nicotinic Acetylcholine Receptor Ligands

Neuronal nicotinic acetylcholine receptors (nAChRs) are ligand gated ion channels of broad distribution and structural heterogeneity. Their functional diversity demonstrated involvement in a variety of neuronal processes (e.g., sensory gating and cognitive function) and generated great interest in them as targets for therapeutic intervention in a number of neuropathological conditions and diseases. In order to control distinct nicotinic functions pharmacologically, it is important to design ligands that selectively interact with distinct receptor subtypes in such a way as to maximize the therapeutic effect and minimize the adverse effects. The α7 nAChR, a CNS subtype, has been the most intensively studied nAChR in recent years. Selective α7 nAChR agonists have been developed as potential candidates for the treatment of schizophrenia, cognitive disorders (including Alzheimers disease), and inflammation. Despite early concerns that the rapid desensitization property of the α7 nAChR would limit their therapeutic potential, several have already been advanced to clinical trials (e.g., PH-399733, Pfizer; MEM 3454, Memory Pharmaceuticals/Roche). Further development of allosteric modulators and pharmaceutically relevant antagonists might expand the therapeutic potential of compounds that target α7 nAChRs. In this review we briefly describe the structure and function of the α7 nAChR and its in vitro and in vivo pharmacology, discuss the clinical relevance of these efforts, and review the current progress in α7 ligand development.

An α7 Nicotinic Acetylcholine Receptor-Selective Agonist Reduces Weight Gain and Metabolic Changes in a Mouse Model of Diabetes

Type 2 diabetes has become a pervasive public health problem. The etiology of the disease has not been fully defined but appears to involve abnormalities in peripheral and central nervous system pathways, as well as prominent inflammatory components. Because nicotinic acetylcholine receptors (nAChRs) are known to interact with anti-inflammatory pathways and have been implicated in control of appetite and body weight, as well as lipid and energy metabolism, we examined their role in modulating biological parameters associated with the disease. In a model of type 2 diabetes, the homozygous leptin-resistant db/db obese mouse, we measured the effects of a novel α7 nAChR-selective agonist [5-methyl-N-[2-(pyridin-3-ylmethyl)-1-azabicyclo[2.2.2]oct-3-yl]thiophene-2-carboxamide (TC-7020)] on body mass, glucose and lipid metabolism, and proinflammatory cytokines. Oral administration of TC-7020 reduced weight gain and food intake, reduced elevated glucose and glycated hemoglobin levels, and lowered elevated plasma levels of triglycerides and the proinflammatory cytokine tumor necrosis factor-α. These changes were reversed by the α7-selective antagonist methyllycaconitine, confirming the involvement of α7 nAChRs. Prevention of weight gain, decreased food intake, and normalization of glucose levels were also blocked by the Janus kinase 2 (JAK2) inhibitor α-cyano-(3,4-dihydroxy)-N-benzylcinnamide (AG-490), suggesting that these effects involve linkage of α7 nAChRs to the JAK2-signal transducer and activator of transcription 3 signaling pathway. The results show that α7 nAChRs play a central role in regulating biological parameters associated with diabetes and support the potential of targeting these receptors as a new therapeutic strategy for treatment.

Discovery and Development of α7 Nicotinic Acetylcholine Receptor Modulators

Efforts in the design and discovery of selective α7 nicotinic acetylcholine receptor (nAChR) agonists were both facilitated and hampered by observation that ligands can show dual affinity at both the 5-HT3 receptor (5-HT3R) and nAChR. 1 The 5-HT3R and nAChRs are both part of the Cys-loop superfamily of ligand-gated ion channels. Further, there is significant sequencehomology between 5-HT3R and α7nAChR inthe ligand recognition domain. 2 Previously reported potent α7 nAChR agonists lacked selectivity versus 5-HT3R, 3 and antagonist activity at 5-HT3R often translated into agonism at α7 nAChR. The crossover in affinity might be explained by pharmacophoric elements common to both 5-HT3R and α7 nAChR: a basic amine (protonated at physiological pH) provides for a cation� π interaction; a hydrogen-bond acceptor, e.g., a carbonyl group, forms a hydrogen bond; and aromatic moieties participate in π� π interactions. 4 In view of reported side effects, i.e., constipation, asymptomatic electrocardiogram changes, and arrhythmias, associated with 5-HT3R antagonists, 5 efforts of research groups were focused on design of ligands specifically interacting with only the α7 nAChR to maximize the therapeutic effect and minimize the adverse effects. Over the past 10 years, drug discovery efforts significantly expanded the quantity and quality of selective α7 nAChR ligands. Those have been summarized in several excellent reviews. 6� 8 Despite early skepticism centered around the rapid desensitization of the α7 nAChR and the hypothesis that agonists might not be functional agonists in vivo, 10 of them have already been advanced to clinical trials. In this review, we highlight the most advanced and characterized (especiallyinvivo)selective α7nAChRorthostericandallosteric α7 nAChR modulators. 1. AGONISTS

Traceless synthesis of benzimidazoles on solid support.

Traceless solid-phase syntheses of benzimidazoles and 5-(benzimidazol-2-yl)benzimidazoles on 2-(4-formyl-3-methoxyphenoxy)ethyl polystyrene are described. No auxiliary functional groups are left in the products after ultimate cleavage and cyclization.

Discovery of (2S,3R)-N-[2-(pyridin-3-ylmethyl)-1-azabicyclo[2.2.2]oct-3-yl]benzo[b]furan-2-carboxamide (TC-5619), a selective α7 nicotinic acetylcholine receptor agonist, for the treatment of cognitive disorders.

(2S,3R)-N-[2-(Pyridin-3-ylmethyl)-1-azabicyclo[2.2.2]oct-3-yl]benzo[b]furan-2-carboxamide (7a, TC-5619), a novel selective agonist of the α7 neuronal nicotinic acetylcholine receptor, has been identified as a promising drug candidate for the treatment of cognitive impairment associated with neurological disorders. 7a demonstrated more than a thousand-fold separation between the affinities for the α7 and α4β2 receptor subtypes and had no detectable effects on muscle or ganglionic nicotinic receptor subtypes, indicating a marked selectivity for the central nervous system over the peripheral nervous system. Results obtained from homology modeling and docking explain the observed selectivity. 7a had positive effects across cognitive, positive, and negative symptoms of schizophrenia in animal models and was additive or synergistic with the antipsychotic clozapine. Compound 7a, as an augmentation therapy to the standard treatment with antipsychotics, demonstrated encouraging results on measures of negative symptoms and cognitive dysfunction in schizophrenia and was well tolerated in a phase II clinical proof of concept trial in patients with schizophrenia.

3D molecular descriptors important for clinical success.

The pharmacokinetic and safety profiles of clinical drug candidates are greatly influenced by their requisite physicochemical properties. In particular, it has been shown that 2D molecular descriptors such as fraction of Sp3 carbon atoms (Fsp3) and number of stereo centers correlate with clinical success. Using the proteomic off-target hit rate of nicotinic ligands, we found that shape-based 3D descriptors such as the radius of gyration and shadow indices discriminate off-target promiscuity better than do Fsp3 and the number of stereo centers. We have deduced the relevant descriptor values required for a ligand to be nonpromiscuous. Investigating the MDL Drug Data Report (MDDR) database as compounds move from the preclinical stage toward the market, we have found that these shape-based 3D descriptors predict clinical success of compounds at preclinical and phase1 stages vs compounds withdrawn from the market better than do Fsp3 and LogD. Further, these computed 3D molecular descriptors correlate well with experimentally observed solubility, which is among well-known physicochemical properties that drive clinical success. We also found that about 84% of launched drugs satisfy either Shadow index or Fsp3 criteria, whereas withdrawn and discontinued compounds fail to meet the same criteria. Our studies suggest that spherical compounds (rather than their elongated counterparts) with a minimal number of aromatic rings may exhibit a high propensity to advance from clinical trials to market.

Discovery of 3-(5-chloro-2-furoyl)-3,7-diazabicyclo[3.3.0]octane (TC-6683, AZD1446), a novel highly selective α4β2 nicotinic acetylcholine receptor agonist for the treatment of cognitive disorders.

Diversification of essential nicotinic cholinergic pharmacophoric elements, i.e., cationic center and hydrogen bond acceptor, resulted in the discovery of novel potent α4β2 nAChR selective agonists comprising a series of N-acyldiazabicycles. Core characteristics of the series are an exocyclic carbonyl moiety as a hydrogen bond acceptor and endocyclic secondary amino group. These features are positioned at optimal distance and with optimal relative spatial orientation to provide near optimal interactions with the receptor. A novel potent and highly selective α4β2 nAChR agonist 3-(5-chloro-2-furoyl)-3,7-diazabicyclo[3.3.0]octane (56, TC-6683, AZD1446) with favorable pharmaceutical properties and in vivo efficacy in animal models has been identified as a potential treatment for cognitive deficits associated with psychiatric or neurological conditions and is currently being progressed to phase 2 clinical trials as a treatment for Alzheimers disease.

Analgesic effects mediated by neuronal nicotinic acetylcholine receptor agonists: Correlation with desensitization of α4β2* receptors

Nicotinic α4β2* agonists are known to be effective in a variety of preclinical pain models, but the underlying mechanisms of analgesic action are not well-understood. In the present study, we characterized activation and desensitization properties for a set of seventeen novel α4β2*-selective agonists that display druggable physical and pharmacokinetic attributes, and correlated the in vitro pharmacology results to efficacies observed in a mouse formalin model of analgesia. ABT-894 and Sazetidine-A, two compounds known to be effective in the formalin assay, were included for comparison. The set of compounds displayed a range of activities at human (α4β2)(2)β2 (HS-α4β2), (α4β2)(2)α5 (α4β2α5) and (α4β2)(2)α4 (LS-α4β2) receptors. We report the novel finding that desensitization of α4β2* receptors may drive part of the antinociceptive outcome. Our molecular modeling approaches revealed that when receptor desensitization rather than activation activitiesat α4β2* receptors are considered, there is a better correlation between analgesia scores and combined in vitro properties. Our results suggest that although all three α4β2 subtypes assessed are involved, it is desensitization of α4β2α5 receptors that plays a more prominent role in the antinociceptive action of nicotinic compounds. For modulation of Phase I responses, correlations are significantly improved from an r(2) value of 0.53 to 0.67 and 0.66 when HS- and LS-α4β2 DC(50) values are considered, respectively. More profoundly, considering the DC(50) at α4β2α5 takes the r(2) from 0.53 to 0.70. For Phase II analgesia scores, adding HS- or LS-α4β2 desensitization potencies did not improve the correlations significantly. Considering the α4β2α5 DC(50) value significantly increased the r(2) from 0.70 to 0.79 for Phase II, and strongly suggested a more prominent role for α4β2α5 nAChRs in the modulation of pain in the formalin assay. The present studies demonstrate that compounds which are more potent at desensitization of α4β2* receptors display better analgesia scores in the formalin test. Consideration of desensitization propertiesat α4β2* receptors, especially at α4β2α5, in multiple linear regression analyses significantly improves correlations with efficacies of analgesia. Thus, α4β2* nicotinic acetylcholine receptor desensitization may contribute to efficacy in the mediation of pain, and represent a mechanism for analgesic effects mediated by nicotinic agonists.

Docking studies of benzylidene anabaseine interactions with α7 nicotinic acetylcholine receptor (nAChR) and acetylcholine binding proteins (AChBPs): application to the design of related α7 selective ligands.

AChBPs isolated from Lymnaea stagnalis (Ls), Aplysia californica (Ac) and Bulinus truncatus (Bt) have been extensively used as structural prototypes to understand the molecular mechanisms that underlie ligand-interactions with nAChRs [1]. Here, we describe docking studies on interactions of benzylidene anabaseine analogs with AChBPs and α7 nAChR. Results reveal that docking of these compounds using Glide software accurately reproduces experimentally-observed binding modes of DMXBA and of its active metabolite, in the binding pocket of Ac. In addition to the well-known nicotinic pharmacophore (positive charge, hydrogen-bond acceptor, and hydrophobic aromatic groups), a hydrogen-bond donor feature contributes to binding of these compounds to Ac, Bt, and the α7 nAChR. This is consistent with benzylidene anabaseine analogs with OH and NH(2) functional groups showing the highest binding affinity of these congeners, and the position of the ligand shown in previous X-ray crystallographic studies of ligand-Ac complexes. In the predicted ligand-Ls complex, by contrast, the ligand OH group acts as hydrogen-bond acceptor. We have applied our structural findings to optimizing the design of novel spirodiazepine and spiroimidazoline quinuclidine series. Binding and functional studies revealed that these hydrogen-bond donor containing compounds exhibit improved affinity and selectivity for the α7 nAChR subtype and demonstrate partial agonism. The gain in affinity is also due to conformational restriction, tighter hydrophobic enclosures, and stronger cation-π interactions. The use of AChBPs structure as a surrogate to predict binding affinity to α7 nAChR has also been investigated. On the whole, we found that molecular docking into Ls binding site generally scores better than when a α7 homology model, Bt or Ac crystal structure is used.

Discovery of novel α7 nicotinic acetylcholine receptor ligands via pharmacophoric and docking studies of benzylidene anabaseine analogs

Based on pharmacophore elucidation and docking studies on interactions of benzylidene anabaseine analogs with AChBPs and α7 nAChR, novel spirodiazepine and spiroimidazoline quinuclidine series have been designed. Binding studies revealed that some of hydrogen-bond donor containing compounds exhibit improved affinity and selectivity for the α7 nAChR subtype in comparison with most potent metabolite of GTS-21, 3-(4-hydroxy-2-methoxybenzylidene)-anabaseine. Hydrophobicity and rigidity of the ligand also contribute into its binding affinity. We also describe alternative pharmacophoric features equidistant from the carbonyl oxygen atom of the conserved Trp-148 of the principal face, which may be exploited to further design diverse focused libraries targeting the α7 nAChR.