RESUMEN

There is an urgent need for nonopioid treatments for chronic and neuropathic pain to provide effective alternatives amid the escalating opioid crisis. This study introduces novel compounds targeting the α9 nicotinic acetylcholine receptor (nAChR) subunit, which is crucial for pain regulation, inflammation, and inner ear functions. Specifically, it identifies novel substituted carbamoyl/amido/heteroaryl dialkylpiperazinium iodides as potent agonists selective for human α9 and α9α10 over α7 nAChRs, particularly compounds 3f, 3h, and 3j. Compound 3h (GAT2711) demonstrated a 230 nM potency as a full agonist at α9 nAChRs, being 340-fold selective over α7. Compound 3c was 10-fold selective for α9α10 over α9 nAChR. Compounds 2, 3f, and 3h inhibited ATP-induced interleukin-1ß release in THP-1 cells. The analgesic activity of 3h was fully retained in α7 knockout mice, suggesting that analgesic effects were potentially mediated through α9* nAChRs. Our findings provide a blueprint for developing α9*-specific therapeutics for pain.

Asunto(s)

Analgésicos , Inflamación , Piperazinas , Receptores Nicotínicos , Animales , Humanos , Masculino , Ratones , Analgésicos/farmacología , Analgésicos/química , Analgésicos/síntesis química , Analgésicos/uso terapéutico , Inflamación/tratamiento farmacológico , Ratones Noqueados , Agonistas Nicotínicos/farmacología , Agonistas Nicotínicos/química , Agonistas Nicotínicos/uso terapéutico , Agonistas Nicotínicos/síntesis química , Dolor/tratamiento farmacológico , Piperazinas/farmacología , Piperazinas/química , Piperazinas/síntesis química , Piperazinas/uso terapéutico , Receptores Nicotínicos/metabolismo , Sales (Química)/química , Sales (Química)/farmacología , Relación Estructura-Actividad , Yoduros/química

RESUMEN

The nicotinic acetylcholine receptor has served, since its biochemical identification in the 1970s, as a model of an allosteric ligand-gated ion channel mediating signal transition at the synapse. In recent years, the application of X-ray crystallography and high-resolution cryo-electron microscopy, together with molecular dynamic simulations of nicotinic receptors and homologs, have opened a new era in the understanding of channel gating by the neurotransmitter. They reveal, at atomic resolution, the diversity and flexibility of the multiple ligand-binding sites, including recently discovered allosteric modulatory sites distinct from the neurotransmitter orthosteric site, and the conformational dynamics of the activation process as a molecular switch linking these multiple sites. The model emerging from these studies paves the way for a new pharmacology based, first, upon the occurrence of an original mode of indirect allosteric modulation, distinct from a steric competition for a single and rigid binding site, and second, the design of drugs that specifically interact with privileged conformations of the receptor such as agonists, antagonists, and desensitizers. Research on nicotinic receptors is still at the forefront of understanding the mode of action of drugs on the nervous system.

Asunto(s)

Sitio Alostérico , Microscopía por Crioelectrón , Simulación de Dinámica Molecular , Receptores Nicotínicos , Transducción de Señal , Receptores Nicotínicos/metabolismo , Receptores Nicotínicos/química , Receptores Nicotínicos/genética , Regulación Alostérica , Humanos , Animales , Cristalografía por Rayos X , Sitios de Unión , Conformación Proteica , Ligandos , Modelos Moleculares , Multimerización de Proteína , Agonistas Nicotínicos/química , Agonistas Nicotínicos/farmacología , Agonistas Nicotínicos/metabolismo

RESUMEN

The quinuclidine scaffold has been extensively used for the development of nicotinic acetylcholine receptor (nAChR) agonists, with hydrophobic substituents at position 3 of the quinuclidine framework providing selectivity for α7 nAChRs. In this study, six new ligands (4-9) containing a 3-(pyridin-3-yloxy)quinuclidine moiety (ether quinuclidine) were synthesized to gain a better understanding of the structural-functional properties of ether quinuclidines. To evaluate the pharmacological activity of these ligands, two-electrode voltage-clamp and single-channel recordings were performed. Only ligand 4 activated α7 nAChR. Ligands 5 and 7 had no effects on α7 nAChR, but ligands 6, 8, and 9 potentiated the currents evoked by ACh. Ligand 6 was the most potent and efficacious of the potentiating ligands, with an estimated EC50 for potentiation of 12.6 ± 3.32 µM and a maximal potentiation of EC20 ACh responses of 850 ± 120%. Ligand 6 increased the maximal ACh responses without changing the kinetics of the current responses. At the single-channel level, the potentiation exerted by ligand 6 was evidenced in the low micromolar concentration range by the appearance of prolonged bursts of channel openings. Furthermore, computational studies revealed the preference of ligand 6 for an intersubunit site in the transmembrane domain and highlighted some putative key interactions that explain the different profiles of the synthesized ligands. Notably, Met276 in the 15' position of the transmembrane domain 2 almost abolished the effects of ligand 6 when mutated to Leu. We conclude that ligand 6 is a novel type I positive allosteric modulator (PAM-I) of α7 nAChR.

Asunto(s)

Éter , Receptores Nicotínicos , Ligandos , Regulación Alostérica , Receptor Nicotínico de Acetilcolina alfa 7/metabolismo , Agonistas Nicotínicos/farmacología , Agonistas Nicotínicos/química , Éteres de Etila , Éteres , Receptores Nicotínicos/metabolismo

RESUMEN

The affinity and thermodynamic parameters for the interactions of two naturally occurring neurotoxins, (+)-anatoxin-a and (-)-hosieine-A, with acetylcholine-binding protein were investigated using a fluorescence-quenching assay and isothermal titration calorimetry. The crystal structures of their complexes with acetylcholine-binding protein from Aplysia californica (AcAChBP) were determined and reveal details of molecular recognition in the orthosteric binding site. Comparisons treating AcAChBP as a surrogate for human α4ß2 and α7 nicotinic acetylcholine receptors (nAChRs) suggest that the molecular features involved in ligand recognition and affinity for the protein targets are conserved. The ligands exploit interactions with similar residues as the archetypal nAChR agonist nicotine, but with greater affinity. (-)-Hosieine-A in particular has a high affinity for AcAChBP driven by a favorable entropic contribution to binding. The ligand affinities help to rationalize the potent biological activity of these alkaloids. The structural data, together with comparisons with related molecules, suggest that there may be opportunities to extend the hosieine-A scaffold to incorporate new interactions with the complementary side of the orthosteric binding site. Such a strategy may guide the design of new entities to target human α4ß2 nAChR that may have therapeutic benefit.

Asunto(s)

Receptores Nicotínicos , Acetilcolina/química , Acetilcolina/metabolismo , Proteínas Portadoras/química , Cristalografía por Rayos X , Toxinas de Cianobacterias , Compuestos Heterocíclicos de 4 o más Anillos , Humanos , Ligandos , Agonistas Nicotínicos/química , Agonistas Nicotínicos/metabolismo , Agonistas Nicotínicos/farmacología , Receptores Nicotínicos/metabolismo , Tropanos

RESUMEN

Nicotinic acetylcholine receptors (nAChRs) play an important role in neurotransmission and are also involved in addiction and several disease states. There is significant interest in therapeutic targeting of nAChRs; however, achieving selectivity for one subtype over others has been a longstanding challenge, given the close structural similarities across the family. Here, we characterize binding interactions in the α3ß4 nAChR subtype via structure-function studies involving noncanonical amino acid mutagenesis and two-electrode voltage clamp electrophysiology. We establish comprehensive binding models for both the endogenous neurotransmitter ACh and the smoking cessation drug cytisine. We also use a panel of C(10)-substituted cytisine derivatives to probe the effects of subtle changes in the ligand structure on binding. By comparing our results to those obtained for the well-studied α4ß2 subtype, we identify several features of both the receptor and agonist structure that can be utilized to enhance selectivity for either α3ß4 or α4ß2. Finally, we characterize binding interactions of the α3ß4-selective partial agonist AT-1001 to determine factors that contribute to its selectivity. These results shed new light on the design of selective nAChR-targeted ligands and can be used to inform the design of improved therapies with minimized off-target effects.

Asunto(s)

Agonistas Nicotínicos , Receptores Nicotínicos , Sitios de Unión , Ligandos , Agonistas Nicotínicos/química , Receptores Nicotínicos/química

RESUMEN

A series of racemic benzofurans bearing N-methyl-2-pyrrolidinyl residue at C(2) or C(3) has been synthesized and tested for affinity at the α4ß2 and α3ß4 nicotine acetylcholine receptors (nAChRs). As previously reported for the benzodioxane based analogues, hydroxylation at proper position of benzene ring results in high α4ß2 nAChR affinity and α4ß2 vs. α3ß4 nAChR selectivity. 7-Hydroxy-N-methyl-2-pyrrolidinyl-1,4-benzodioxane (2) and its 7- and 5-amino benzodioxane analogues 3 and 4, which are all α4ß2 nAChR partial agonists, and 2-(N-methyl-2-pyrrolidinyl)-6-hydroxybenzofuran (12) were selected for functional characterization at the two α4ß2 stoichiometries, the high sensitivity (α4)2(ß2)3 and the low sensitivity (α4)3(ß2)2. The benzene pattern substitution, which had previously been found to control α4ß2 partial agonist activity and α4ß2 vs. α3ß4 selectivity, proved to be also involved in stoichiometry-selectivity. The 7-hydroxybenzodioxane derivative 2 selectively activates (α4)2(ß2)3 nAChR, which cannot be activated by its 5-amino analogue 4. A marginal structural modification, not altering the base pyrrolidinyl benzodioxane scaffold, resulted in opposite activity profiles at the two α4ß2 nAChR isoforms providing an interesting novel case study.

Asunto(s)

Benzofuranos , Receptores Nicotínicos , Benceno , Benzofuranos/farmacología , Ligandos , Agonistas Nicotínicos/química , Receptores Nicotínicos/química

RESUMEN

Nicotinic partial agonists provide an accepted aid for smoking cessation and thus contribute to decreasing tobacco-related disease. Improved drugs constitute a continued area of study. However, there remains no reductionist method to examine the cellular and subcellular pharmacokinetic properties of these compounds in living cells. Here, we developed new intensity-based drug-sensing fluorescent reporters (iDrugSnFRs) for the nicotinic partial agonists dianicline, cytisine, and two cytisine derivatives - 10-fluorocytisine and 9-bromo-10-ethylcytisine. We report the first atomic-scale structures of liganded periplasmic binding protein-based biosensors, accelerating development of iDrugSnFRs and also explaining the activation mechanism. The nicotinic iDrugSnFRs detect their drug partners in solution, as well as at the plasma membrane (PM) and in the endoplasmic reticulum (ER) of cell lines and mouse hippocampal neurons. At the PM, the speed of solution changes limits the growth and decay rates of the fluorescence response in almost all cases. In contrast, we found that rates of membrane crossing differ among these nicotinic drugs by >30-fold. The new nicotinic iDrugSnFRs provide insight into the real-time pharmacokinetic properties of nicotinic agonists and provide a methodology whereby iDrugSnFRs can inform both pharmaceutical neuroscience and addiction neuroscience.

Asunto(s)

Alcaloides/química , Azepinas/química , Compuestos Heterocíclicos de 4 o más Anillos/química , Agonistas Nicotínicos/química , Cese del Hábito de Fumar , Alcaloides/metabolismo , Animales , Azocinas/química , Azocinas/metabolismo , Fluorescencia , Humanos , Ligandos , Ratones , Quinolizinas/química , Quinolizinas/metabolismo

RESUMEN

The extracellular domain of the nicotinic acetylcholine receptor isoforms formed by three α4 and two ß2 subunits ((α4)3(ß2)2 nAChR) harbors two high-affinity "canonical" acetylcholine (ACh)-binding sites located in the two α4:ß2 intersubunit interfaces and a low-affinity "noncanonical" ACh-binding site located in the α4:α4 intersubunit interface. In this study, we used ACh, cytisine, and nicotine (which bind at both the α4:α4 and α4:ß2 interfaces), TC-2559 (which binds at the α4:ß2 but not at the α4:α4 interface), and 3-(2-chlorophenyl)-5-(5-methyl-1-(piperidin-4-yl)-1H-pyrrazol-4-yl)isoxazole (CMPI, which binds at the α4:α4 but not at the α4:ß2 interface), to investigate the binding and gating properties of CMPI at the α4:α4 interface. We recorded whole-cell currents from Xenopus laevis oocytes expressing (α4)3(ß2)2 nAChR in response to applications of these ligands, alone or in combination. The electrophysiological data were analyzed in the framework of a modified Monod-Wyman-Changeux allosteric activation model. We show that CMPI is a high-affinity, high-efficacy agonist at the α4:α4 binding site and that its weak direct activating effect is accounted for by its inability to productively interact with the α4:ß2 sites. The data presented here enhance our understanding of the functional contributions of ligand binding at the α4:α4 subunit interface to (α4)3(ß2)2 nAChR-channel gating. These findings support the potential use of α4:α4 specific ligands to increase the efficacy of the neurotransmitter ACh in conditions associated with decline in nAChRs activity in the brain.

Asunto(s)

Agonistas Nicotínicos , Receptores Nicotínicos , Sitio Alostérico , Animales , Benzamidas/química , Benzamidas/farmacología , Sitios de Unión , Ligandos , Agonistas Nicotínicos/química , Agonistas Nicotínicos/farmacología , Oocitos/metabolismo , Receptores Nicotínicos/química , Receptores Nicotínicos/metabolismo , Xenopus laevis

RESUMEN

In the context of naturally occurring nitrogen heterocycles, nicotine is a chiral alkaloid present in tobacco plants, which can target and stimulate nicotinic acetylcholine receptors (nAChRs), a class of ligand-gated ion channels commonly located throughout the human brain. Due to its well-known toxicity for humans, there is considerable interest in the development of synthetic analogues; in particular, conformationally restricted analogues of nicotine have emerged as promising drug molecules for selective nAChR-targeting ligands. In the present mini-review, we will describe the synthesis of the conformationally restricted analogues of nicotine involving one or more catalytic processes. In particular, we will follow a systematic approach as a function of the heteroarene structure, considering: (a) 2,3-annulated tricyclic derivatives; (b) 3,4-annulated tricyclic derivatives; (c) tetracyclic derivatives; and (d) other polycyclic derivatives. For each of them we will also consider, when carried out, biological studies on their activity for specific nAChR subunits.

Asunto(s)

Sistemas de Liberación de Medicamentos , Nicotina , Agonistas Nicotínicos , Animales , Humanos , Nicotina/análogos & derivados , Nicotina/síntesis química , Nicotina/química , Nicotina/uso terapéutico , Agonistas Nicotínicos/síntesis química , Agonistas Nicotínicos/química , Agonistas Nicotínicos/uso terapéutico

RESUMEN

The selectivity of α4ß2 nAChR agonists over the α3ß4 nicotinic receptor subtype, predominant in ganglia, primarily conditions their therapeutic range and it is still a complex and challenging issue for medicinal chemists and pharmacologists. Here, we investigate the determinants for such subtype selectivity in a series of more than forty α4ß2 ligands we have previously reported, docking them into the structures of the two human subtypes, recently determined by cryo-electron microscopy. They are all pyrrolidine based analogues of the well-known α4ß2 agonist N-methylprolinol pyridyl ether A-84543 and differ in the flexibility and pattern substitution of their aromatic portion. Indeed, the direct or water mediated interaction with hydrophilic residues of the relatively narrower ß2 minus side through the elements decorating the aromatic ring and the stabilization of the latter by facing to the not conserved ß2-Phe119 result as key distinctive features for the α4ß2 affinity. Consistently, these compounds show, despite the structural similarity, very different α4ß2 vs. α3ß4 selectivities, from modest to very high, which relate to rigidity/extensibility degree of the portion containing the aromatic ring and to substitutions at the latter. Furthermore, the structural rationalization of the rat vs. human differences of α4ß2 vs. α3ß4 selectivity ratios is here proposed.

Asunto(s)

Agonistas Nicotínicos/química , Receptores Nicotínicos/ultraestructura , Animales , Sitios de Unión , Microscopía por Crioelectrón/métodos , Bases de Datos Genéticas , Humanos , Ligandos , Simulación del Acoplamiento Molecular , Agonistas Nicotínicos/farmacología , Piridinas/química , Piridinas/farmacología , Pirrolidinas/química , Pirrolidinas/farmacología , Ratas , Receptores Nicotínicos/metabolismo , Relación Estructura-Actividad , Transmisión Sináptica/efectos de los fármacos

RESUMEN

We detail here distinctive departures from lead classical cholinesterase re-activators, the pyridinium aldoximes, to achieve rapid CNS penetration and reactivation of AChE in the CNS (brain and spinal cord). Such reactivation is consistent with these non-canonical re-activators enhancing survival parameters in both mice and macaques following exposure to organophosphates. Thus, the ideal cholinesterase re-activator should show minimal toxicity, limited inhibitory activity in the absence of an organophosphate, and rapid CNS penetration, in addition to its nucleophilic potential at the target, the conjugated AChE active center. These are structural properties directed to reactivity profiles at the conjugated AChE active center, reinforced by the pharmacokinetic and tissue disposition properties of the re-activator leads. In the case of nicotinic acetylcholine receptor (nAChR) agonists and antagonists, with the many existing receptor subtypes in mammals, we prioritize subtype selectivity in their design. In contrast to nicotine and its analogues that react with panoply of AChR subtypes, the substituted di-2-picolyl amine pyrimidines possess distinctive ionization characteristics reflecting in selectivity for the orthosteric site at the α7 subtypes of receptor. Here, entry to the CNS should be prioritized for the therapeutic objectives of the nicotinic agent influencing aberrant CNS activity in development or in the sequence of CNS ageing (longevity) in mammals, along with general peripheral activities controlling inflammation.

Asunto(s)

Acetilcolinesterasa/química , Reactivadores de la Colinesterasa/química , Diseño de Fármacos , Agonistas Nicotínicos/química , Antagonistas Nicotínicos/química , Receptores Nicotínicos/química , Acetilcolinesterasa/metabolismo , Animales , Reactivadores de la Colinesterasa/metabolismo , Humanos , Ligandos , Agonistas Nicotínicos/metabolismo , Antagonistas Nicotínicos/metabolismo , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Receptores Nicotínicos/metabolismo

RESUMEN

SARS-CoV-2 is the coronavirus that originated in Wuhan in December 2019 and has spread globally. Studies have shown that smokers are less likely to be diagnosed with or be hospitalized for COVID-19 but, once hospitalized, have higher odds for an adverse outcome. We have previously presented the potential interaction between SARS-CoV-2 Spike glycoprotein and nicotinic acetylcholine receptors (nAChRs), due to a "toxin-like" epitope on the Spike glycoprotein, with homology to a sequence of a snake venom toxin. This epitope coincides with the well-described cryptic epitope for the human anti-SARS-CoV antibody CR3022. In this study, we present the molecular complexes of both SARS-CoV and SARS-CoV-2 Spike glycoproteins, at their open or closed conformations, with the model of the human α7 nAChR. We found that all studied protein complexes' interface involves a large part of the "toxin-like" sequences of SARS-CoV and SARS-CoV-2 Spike glycoproteins and toxin binding site of human α7 nAChR. Our findings provide further support to the hypothesis about the protective role of nicotine and other cholinergic agonists. The potential therapeutic role of CR3022 and other similar monoclonal antibodies with increased affinity for SARS-CoV-2 Spike glycoprotein against the clinical effects originating from the dysregulated cholinergic pathway should be further explored.

Asunto(s)

COVID-19/virología , Epítopos , Nicotina/farmacología , SARS-CoV-2/química , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/química , Glicoproteína de la Espiga del Coronavirus/química , Receptor Nicotínico de Acetilcolina alfa 7/metabolismo , Secuencia de Aminoácidos , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Antivirales/química , Anticuerpos Antivirales/uso terapéutico , Sitios de Unión de Anticuerpos , COVID-19/metabolismo , COVID-19/prevención & control , Humanos , Modelos Moleculares , Agonistas Nicotínicos/química , Agonistas Nicotínicos/uso terapéutico , Sistema Colinérgico no Neuronal , Pandemias , Factores Protectores , Conformación Proteica , Homología de Secuencia , Transducción de Señal , Fumadores , Fumar , Venenos de Serpiente/química

RESUMEN

A series of diastereomeric 2-(2-pyrrolidinyl)-1,4-benzodioxanes bearing a small, hydrogen-bonding substituent at the 7-, 6-, or 5-position of benzodioxane have been studied for α4ß2 and α3ß4 nicotinic acetylcholine receptor affinity and activity. Analogous to C(5)H replacement with N and to a much greater extent than decoration at C(7), substitution at benzodioxane C(5) confers very high α4ß2/α3ß4 selectivity to the α4ß2 partial agonism. Docking into the two receptor structures recently determined by cryo-electron microscopy and site-directed mutagenesis at the minus ß2 side converge in indicating that the limited accommodation capacity of the ß2 pocket, compared to that of the ß4 pocket, makes substitution at C(5) rather than at more projecting C(7) position determinant for this pursued subtype selectivity.

Asunto(s)

Dioxanos/química , Agonistas Nicotínicos/química , Receptores Nicotínicos/química , Sitios de Unión , Microscopía por Crioelectrón , Dioxanos/síntesis química , Dioxanos/metabolismo , Humanos , Enlace de Hidrógeno , Simulación del Acoplamiento Molecular , Mutagénesis Sitio-Dirigida , Agonistas Nicotínicos/síntesis química , Agonistas Nicotínicos/metabolismo , Antagonistas Nicotínicos/química , Antagonistas Nicotínicos/metabolismo , Pirrolidinas/química , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Estereoisomerismo , Relación Estructura-Actividad

RESUMEN

Epibatidine is a potent analgetic agent with very high affinity for brain nicotinic acetylcholine receptors (nAChR). We determined the activity profiles of three epibatidine derivatives, RTI-36, RTI-76, and RTI-102, which have affinity for brain nAChR equivalent to that of epibatidine but reduced analgetic activity. RNAs coding for nAChR monomeric subunits and/or concatamers were injected into Xenopus oocytes to obtain receptors of defined subunit composition and stoichiometry. The epibatidine analogs produced protracted activation of high sensitivity (HS) α4- and α2-containing receptors with the stoichiometry of 2alpha:3beta subunits but not low sensitivity (LS) receptors with the reverse ratio of alpha and beta subunits. Although not strongly activated by the epibatidine analogs, LS α4- and α2-containing receptors were potently desensitized by the epibatidine analogs. In general, the responses of α4(2)ß2(2)α5 and ß3α4ß2α6ß2 receptors were similar to those of the HS α4ß2 receptors. RTI-36, the analog closest in structure to epibatidine, was the most efficacious of the three compounds, also effectively activating α7 and α3ß4 receptors, albeit with lower potency and less desensitizing effect. Although not the most efficacious agonist, RTI-76 was the most potent desensitizer of α4- and α2-containing receptors. RTI-102, a strong partial agonist for HS α4ß2 receptors, was effectively an antagonist for LS α4ß2 receptors. Our results highlight the importance of subunit stoichiometry and the presence or absence of specific accessory subunits for determining the activity of these drugs on brain nAChR, affecting the interpretation of in vivo studies since in most cases these structural details are not known. SIGNIFICANCE STATEMENT: Epibatidine and related compounds are potent ligands for the high-affinity nicotine receptors of the brain, which are therapeutic targets and mediators of nicotine addiction. Far from being a homogeneous population, these receptors are diverse in subunit composition and vary in subunit stoichiometry. We show the importance of these structural details for drug activity profiles, which present a challenge for the interpretation of in vivo experiments since conventional methods, such as in situ hybridization and immunohistochemistry, cannot illuminate these details.

Asunto(s)

Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Agonistas Nicotínicos/farmacología , Subunidades de Proteína/metabolismo , Piridinas/química , Receptores Nicotínicos/metabolismo , Animales , Animales Modificados Genéticamente , Encéfalo/metabolismo , Compuestos Bicíclicos Heterocíclicos con Puentes/química , Humanos , Estructura Molecular , Complejos Multiproteicos/metabolismo , Agonistas Nicotínicos/química , Subunidades de Proteína/genética , Receptores Nicotínicos/genética , Tropanos/química , Tropanos/farmacología , Xenopus/genética

RESUMEN

It has been almost 20 years since the discovery and crystallization of a structural surrogate, the Lymnaea stagnalis acetylcholine binding protein (Ls-AChBP), comprising the extracellular domain of the nicotinic acetylcholine receptors (nAChRs). Structural characterization of this soluble protein has increased our understanding of the requirements for agonist and antagonist interactions at the ligand recognition site of the nAChRs. Application can be extended to orthologs in the pentameric ligand-gated ion channel superfamily, encompassing receptors that depolarize or hyperpolarize upon neurotransmitter association. Despite the lack of transmembrane and intracellular motifs, the highly conserved binding or recognition loci have made these soluble binding proteins, and mutants derived from them, prototypic tools for molecular recognition and structural studies of pentameric ligand-gated ion channels. Targeting nAChRs has been a major goal as this family is associated with neurodegenerative diseases and disorders. Accordingly, the ligand binding site has played a key role to the development of selective ligands for modulation of this transmembrane proteins. In this review article, we cover both the potential and limitations of soluble surrogates, termed the AChBP family, in drug development. This article is part of the special issue on 'Contemporary Advances in Nicotine Neuropharmacology'.

Asunto(s)

Proteínas Portadoras/química , Diseño de Fármacos , Agonistas Nicotínicos/química , Antagonistas Nicotínicos/química , Receptores Nicotínicos/química , Secuencia de Aminoácidos , Animales , Sitios de Unión/efectos de los fármacos , Sitios de Unión/fisiología , Proteínas Portadoras/agonistas , Proteínas Portadoras/antagonistas & inhibidores , Proteínas Portadoras/metabolismo , Humanos , Agonistas Nicotínicos/metabolismo , Agonistas Nicotínicos/farmacología , Antagonistas Nicotínicos/metabolismo , Antagonistas Nicotínicos/farmacología , Estructura Secundaria de Proteína , Receptores Nicotínicos/metabolismo , Relación Estructura-Actividad

RESUMEN

By linking two N-methyl-N-carbocyclic quaternary ammonium groups to an azobenzene scaffold in meta- or para-positions we generated a series of photoswitchable neuromuscular ligands for which we coined the term "azocuroniums". These compounds switched between the (E)- and (Z)-isomers by light irradiation at 400-450 nm and 335-340 nm, respectively. Meta-azocuroniums were potent nicotinic ligands with a clear selectivity for the muscular nAChRs compared to neuronal α7 and α4ß2 subtypes, showed good solubility in physiologic media, negligible cell toxicity, and would not reach the CNS. Electrophysiological studies in muscle-type nAChRs expressed in Xenopus laevis oocytes showed that (E)-isomers were more potent than (Z)-forms. All meta-azocuroniums were neuromuscular blockers, with the exception of the pyrrolidine derivative that was an agonist. These new meta-azocuroniums, which can be modulated ad libitum by light, could be employed as photoswitchable muscle relaxants with fewer side effects for surgical interventions and as tools to better understand the pharmacology of muscle-type nAChRs.

Asunto(s)

Fármacos Neuromusculares/efectos de la radiación , Agonistas Nicotínicos/química , Receptores Nicotínicos/metabolismo , Animales , Compuestos Azo/química , Humanos , Isomerismo , Ligandos , Luz , Fármacos Neuromusculares/síntesis química , Agonistas Nicotínicos/efectos de la radiación , Oocitos , Compuestos de Amonio Cuaternario/química , Relación Estructura-Actividad , Xenopus laevis

Asunto(s)

Proteínas Adaptadoras Transductoras de Señales/uso terapéutico , Ansiedad/tratamiento farmacológico , Colinérgicos/uso terapéutico , Neuronas Colinérgicas/metabolismo , Cognición/efectos de los fármacos , Receptores Nicotínicos/metabolismo , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Ansiedad/metabolismo , Ansiedad/psicología , Colinérgicos/química , Colinérgicos/metabolismo , Neuronas Colinérgicas/efectos de los fármacos , Cognición/fisiología , Humanos , Agonistas Nicotínicos/química , Agonistas Nicotínicos/metabolismo , Agonistas Nicotínicos/uso terapéutico , Antagonistas Nicotínicos/química , Antagonistas Nicotínicos/metabolismo , Antagonistas Nicotínicos/uso terapéutico , Estructura Secundaria de Proteína , Receptores Nicotínicos/química

RESUMEN

Novel pyrrole derivatives were discovered as potent agonists of the niacin receptor, GPR109A. During the derivatization, compound 16 was found to be effective both in vitro and in vivo. The compound 16 exhibited a significant reduction of the non-esterified fatty acid in human GPR109A transgenic rats, and the duration of its in vivo efficacy was much longer than niacin.

Asunto(s)

Agonistas Nicotínicos/química , Pirroles/química , Receptores Acoplados a Proteínas G/agonistas , Animales , Diseño de Fármacos , Ácidos Grasos no Esterificados/metabolismo , Humanos , Lipólisis/efectos de los fármacos , Agonistas Nicotínicos/metabolismo , Agonistas Nicotínicos/farmacología , Pirroles/metabolismo , Pirroles/farmacología , Ratas , Ratas Transgénicas , Receptores Acoplados a Proteínas G/metabolismo , Relación Estructura-Actividad

RESUMEN

Neuronal nicotinic acetylcholine receptors (nAChRs) are crucial mediators of central presynaptic, postsynaptic, and extrasynaptic signaling, and they are implicated in a range of CNS disorders. The numerous nAChR subtypes are differentially expressed and mediate distinct functions throughout the CNS, and thus there is considerable interest in developing subtype-selective nAChR modulators, both for use as pharmacological tools and as putative therapeutics. α6ß2-containing (α6ß2*) nAChRs are highly expressed in and regulate the activity of midbrain dopaminergic neurons, which makes them attractive drug targets in several psychiatric and neurological diseases, including nicotine addiction and Parkinson's disease. This paper presents the preclinical characterization of AN317, a novel α6ß2* agonist exhibiting functional selectivity toward other nAChRs, including α4ß2, α3ß4 and α7 receptors. AN317 induced [3H]dopamine release from rat striatal synaptosomes and augmented dopaminergic neuron activity in substantia nigra pars compacta brain slices in Ca2+ imaging and electrophysiological assays. In line with this, AN317 alleviated the high-frequency tremors arising from reserpine-mediated dopamine depletion in rats. Finally, AN317 mediated significant protective effects on cultured rat mesencephalic neurons treated with the dopaminergic neurotoxin MPP+. AN317 displays good bioavailability and readily crosses the blood-brain barrier, which makes it a unique tool for both in vitro and in vivo studies of native α6ß2* receptors in the nigrostriatal system and other dopaminergic pathways. Altogether, these findings highlight the potential of selective α6ß2* nAChR activation as a treatment strategy for symptoms and possibly even deceleration of disease progression in neurodegenerative diseases such as Parkinson's disease.

Asunto(s)

Fármacos Neuroprotectores/farmacología , Agonistas Nicotínicos/farmacología , Receptores Nicotínicos/metabolismo , Potenciales de Acción/efectos de los fármacos , Animales , Señalización del Calcio/efectos de los fármacos , Dopamina/metabolismo , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Células HEK293 , Humanos , Masculino , Mesencéfalo/efectos de los fármacos , Mesencéfalo/embriología , Ratones , Fármacos Neuroprotectores/administración & dosificación , Fármacos Neuroprotectores/química , Fármacos Neuroprotectores/farmacocinética , Agonistas Nicotínicos/administración & dosificación , Agonistas Nicotínicos/química , Agonistas Nicotínicos/farmacocinética , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Técnicas de Placa-Clamp , Unión Proteica , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Receptores Nicotínicos/genética , Sinaptosomas/efectos de los fármacos , Sinaptosomas/metabolismo , Xenopus laevis

RESUMEN

A lack of selectivity of classical agonists for the nicotinic acetylcholine receptors (nAChR) has prompted us to identify and develop a distinct scaffold of α7 nAChR-selective ligands. Noncanonical 2,4,6-substituted pyrimidine analogues were framed around compound 40 for a structure-activity relationship study. The new lead compounds activate selectively the α7 nAChRs with EC50's between 30 and 140 nM in a PNU-120596-dependent, cell-based calcium influx assay. After characterizing the expanded lead landscape, we ranked the compounds for rapid activation using Xenopus oocytes expressing human α7 nAChR with a two-electrode voltage clamp. This approach enabled us to define the molecular determinants governing rapid activation, agonist potency, and desensitization of α7 nAChRs after exposure to pyrimidine analogues, thereby distinguishing this subclass of noncanonical agonists from previously defined types of agonists (agonists, partial agonists, silent agonists, and ago-PAMs). By NMR, we analyzed pKa values for ionization of lead candidates, demonstrating distinctive modes of interaction for this landscape of ligands.

Asunto(s)

Agonistas Nicotínicos/química , Agonistas Nicotínicos/farmacología , Receptor Nicotínico de Acetilcolina alfa 7/agonistas , Animales , Sitios de Unión , Simulación por Computador , Evaluación Preclínica de Medicamentos/métodos , Femenino , Humanos , Isoxazoles/farmacología , Espectroscopía de Resonancia Magnética , Neurotransmisores/metabolismo , Agonistas Nicotínicos/síntesis química , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Técnicas de Placa-Clamp , Compuestos de Fenilurea/farmacología , Relación Estructura-Actividad , Xenopus laevis , Receptor Nicotínico de Acetilcolina alfa 7/genética , Receptor Nicotínico de Acetilcolina alfa 7/metabolismo