Search

Background and Purpose: GAT107 ((3aR,4S,9bS)-4-(4-bromo-phenyl)-3a,4,5,9b-tetrahydro-3H-cyclopenta-[c]quinoline-8-sulfonamide) is a positive allosteric modulator (PAM) and agonist of α7 nicotinic acetylcholine receptors (nAChRs)that can cause a prolonged period of primed potentiation of acetylcholine responses after drug washout. NS6740 is a silent agonist of α7 nAChRs that has little or no efficacy for activating the ion channel but induces stable desensitization states, some of which can be converted into channel-active states by PAMs. Although GAT107 and NS6740 appear to stably induce different non-conducting states, both agents are effective treatment for inflammation and inflammatory pain models. We sought to better understand how both of these drugs that have opposite effects on channel activation could regulate signal transduction.Experimental Approach: Voltage-clamp experiments were conducted with α7 nAChRs expressed in Xenopus oocytes.Key Results: Long-lived sensitivity to a PAM or to an agonist was produced by NS6740 or GAT107 respectively. With sequential applications, these two drugs induced varying levels of persistent activation, which is a unique condition for a receptor that is known for rapid desensitization. The non-conducting states induced by NS6740 or GAT107 differ in their sensitivity to an α7 nAChR-selective antagonist and in how effectively they promote current.Conclusions& Implications: Our data suggest that the persistent currents represent a dynamic interconversion between different stable desensitized states and the PAM-inducible conducting states. However, the similarity of NS6740 and GAT107 effects on inflammation and pain suggests that the different stable non-conducting states have common activity on signal transduction.Linked Articles: This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc. [ABSTRACT FROM AUTHOR]

Background and Purpose: Orthosteric agonists and positive allosteric modulators (PAMs) of the α7 nicotinic ACh receptor (nAChR) represent novel therapeutic approaches for pain modulation. Moreover, compounds with dual function as allosteric agonists and PAMs, known as ago-PAMs, add further regulation of receptor function.Experimental Approach: Initial studies examined the α7 ago-PAM, GAT107, in the formalin, complete Freund's adjuvant (CFA), LPS inflammatory pain models, the chronic constriction injury neuropathic pain model and the tail flick and hot plate acute thermal nociceptive assays. Additional studies examined the locus of action of GAT107 and immunohistochemical markers in the dorsal horn of the spinal cord in the CFA model.Key Results: Complementary pharmacological and genetic approaches confirmed that the dose-dependent antinociceptive effects of GAT107 were mediated through α7 nAChR. However, GAT107 was inactive in the tail flick and hot plate assays. In addition, GAT107 blocked conditioned place aversion elicited by acetic acid injection. Furthermore, intrathecal, but not intraplantar, injections of GAT107 reversed nociception in the CFA model, suggesting a spinal component of action. Immunohistochemical evaluation revealed an increase in the expression of astrocyte-specific glial fibrillary acidic protein and phosphorylated p38MAPK within the spinal cords of mice treated with CFA, which was attenuated by intrathecal GAT107 treatment. Importantly, GAT107 did not elicit motor impairment and continued to produce antinociceptive effects after subchronic administration in both phases of the formalin test.Conclusions and Implications: Collectively, these results provide the first proof of principle that α7 ago-PAMs represent an effective pharmacological strategy for treating inflammatory and neuropathic pain. [ABSTRACT FROM AUTHOR]

Abstract: We report here an efficient and expeditious microwave-assisted synthesis of cyclopentadiene ring-fused tetrahydroquinolines using the three-component Povarov reaction catalyzed by indium (III) chloride. This method has an advantage of shorter reaction time (10–15min) with high and reproducible yields (up to 90%) and is suitable for parallel library synthesis. The optimization process is reported and the results from the microwave route are compared with those of the conventional synthetic route. In almost all cases, the microwave acceleration consistently provided improved yields favoring the cis-diastereomer. [Copyright &y& Elsevier]

Supplemental oxygen therapy with supraphysiological concentrations of oxygen (hyperoxia; >21% O2) is a life-saving intervention for patients experiencing respiratory distress. However, prolonged exposure to hyperoxia can compromise bacterial clearance processes, due to oxidative stress-mediated impairment of macrophages, contributing to the increased susceptibility to pulmonary infections. This study reports that the activation of the α7 nicotinic acetylcholine receptor (α7nAChR) with the delete allosteric agonistic-positive allosteric modulator, GAT107, decreases the bacterial burden in mouse lungs by improving hyperoxia-induced lung redox imbalance. The incubation of RAW 264.7 cells with GAT107 (3.3 µM) rescues hyperoxia-compromised phagocytic functions in cultured macrophages, RAW 264.7 cells, and primary bone marrow-derived macrophages. Similarly, GAT107 (3.3 µM) also attenuated oxidative stress in hyperoxia-exposed macrophages, which prevents oxidation and hyper-polymerization of phagosome filamentous actin (F-actin) from oxidation. Furthermore, GAT107 (3.3 µM) increases the (1) activity of superoxide dismutase 1; (2) activation of Nrf2 and (3) the expression of heme oxygenase-1 (HO-1) in macrophages exposed to hyperoxia. Overall, these data suggest that the novel α7nAChR compound, GAT107, could be used to improve host defense functions in patients, such as those with COVID-19, who are exposed to prolonged periods of hyperoxia. [ABSTRACT FROM AUTHOR]

The α7 nicotinic acetylcholine receptors (nAChRs) are uniquely sensitive to selective positive allosteric modulators (PAMs), which increase the efficiency of channel activation to a level greater than that of other nAChRs. Although PAMs must work in concert with "orthosteric" agonists, compounds such as GAT107 ((3aR,4S,9bS)-4-(4-bromophenyl)-3a,4,5,9b-tetra-hydro-3H-cyclopenta[c]quinoline-8-sulfonamide) have the combined properties of agonists and PAMs (ago-PAM) and produce very effective channel activation (direct allosteric activation (DAA)) by operating at two distinct sites in the absence of added agonist. One site is likely to be the same transmembrane site where PAMs like PNU-120596 function. We show that the other site, required for direct activation, is likely to be solvent-accessible at the extracellular domain vestibule. We identify key attributes of molecules in this family that are able to act at the DAA site through variation at the aryl ring substituent of the tetrahydroquinoline ring system and with two different classes of competitive antagonists of DAA. Analyses of molecular features of effective allosteric agonists allow us to propose a binding model for theDAAsite, featuring a largely non-polar pocket accessed from the extracellular vestibule with an important role for Asp-101. This hypothesis is supported with data from site-directed mutants. Future refinement of the model and the characterization of specific GAT107 analogs will allow us to define critical structural elements that can be mapped onto the receptor surface for an improved understanding of this novel way to target α7 nAChR therapeutically. [ABSTRACT FROM AUTHOR]

The α7 nicotinic acetylcholine receptor (nAChR) is a promising drug target for a number of neurological disorders including chronic pain and inflammatory diseases. Since α7 can function as a ligand-gated ion channel, drug development initially focused on ligands that were selective activators of the α7 ion channel. However, the best α7 drugs for chronic pain and inflammation indications may not be ion channel activators but rather “silent agonists”, which bind to the receptor but preferentially induce non-conducting states that modulate signal transduction in non-neuronal cells. One such compound is NS6740. We show that NS6740 selectively induces prolonged desensitization of α7 nAChRs. There are two forms of α7 desensitization that can be distinguished by their sensitivity to the positive allosteric modulators (PAMs). At high concentrations, NS6740 preferentially induces PAM-insensitive desensitization, which over the course of several minutes reverts to the sensitive form. NS6740 was tested in several pain models after in vivo administration in the mouse. Although it had no effects in acute thermal pain, NS6740 induced significant dose- and time-dependent antinociceptive activity in formalin- and acetic acid-induced nociceptive behaviors as well as in the chronic constrictive nerve injury (CCI) model for neuropathic pain. The antinociceptive activity of NS6740 in these models was α7-dependent. In addition, NS6740 administration reversed pain-induced aversion, an important affective component of pain. The time and concentration dependence of the effects were consistent with NS6740 induction of PAM-insensitive non-conducting states, suggesting that signal transduction required for analgesia is accomplished by α7 receptors in that conformation. [ABSTRACT FROM AUTHOR]

GAT107, the (+)-enantiomer of racemic 4-(4-bromophenyl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinoline-8-sulfonamide, is a strong positive allosteric modulator (PAM) of α7 nicotinic acetylcholine receptor (nAChR) activation by orthosteric agonists with intrinsic allosteric agonist activities. The direct activation produced by GAT107 in electrophysiological studies is observed only as long as GAT107 is freely diffusible in solution, although the potentiating activity primed by GAT107 can persist for over 30min after drug washout. Direct activation is sensitive to α7 nAChR antagonistmethyllycaconitine, although the primed potentiation is not. The data are consistent with GAT107 activity arising from two different sites. We show that the coupling between PAMs and the binding of orthosteric ligands requires tryptophan 55 (Trp-55), which is located at the subunit interface on the complementary surface of the orthosteric binding site. Mutations of Trp-55 increase the direct activation produced by GAT107 and reduce or prevent the synergy between allosteric and orthosteric binding sites, so that thesemutants can also be directly activated by other PAMs such as PNU-120596 and TQS, which do not activate wild-type α7 in the absence of orthosteric agonists. We identify Tyr-93 as an essential element for orthosteric activation, because Y93C mutants are insensitive to orthosteric agonists but respond to GAT107. Our data show that both orthosteric and allosteric activation of α7 nAChR require cooperative activity at the interface between the subunits in the extracellular domain.These cooperative effects rely on key aromatic residues, and although muta-tions of Trp-55 reduce the restraints placed on the requirement for orthosteric agonists, Tyr-93 can conduct both orthosteric activation and desensitization among the subunits. [ABSTRACT FROM AUTHOR]

Recently, α7 nicotinic acetylcholine receptors (nAChRs), primarily activated by binding of orthosteric agonists, represent a target for anti-inflammatory and analgesic drug development. These receptors may also be modulated by positive allosteric modulators (PAMs), ago-allosteric ligands (ago-PAMs), and α7-silent agonists. Activation of α7 nAChRs has been reported to increase the brain levels of endogenous ligands for nuclear peroxisome proliferator-activated receptors type-α (PPAR-α), palmitoylethanolamide (PEA) and oleoylethanolamide (OEA), in a Ca 2 + -dependent manner. Here, we investigated potential crosstalk between α7 nAChR and PPAR-α, using the formalin test, a mouse model of tonic pain. Using pharmacological and genetic approaches, we found that PNU282987, a full α7 agonist, attenuated formalin-induced nociceptive behavior in α7-dependent manner. Interestingly, the selective PPAR-α antagonist GW6471 blocked the antinociceptive effects of PNU282987, but did not alter the antinociceptive responses evoked by the α7 nAChR PAM PNU120596, ago-PAM GAT107, and silent agonist NS6740. Moreover, GW6471 administered systemically or spinally, but not via the intraplantar surface of the formalin-injected paw blocked PNU282987-induced antinociception. Conversely, exogenous administration of the naturally occurring PPAR-α agonist PEA potentiated the antinociceptive effects of PNU282987. In contrast, the cannabinoid CB 1 antagonist rimonabant and the CB 2 antagonist SR144528 failed to reverse the antinociceptive effects of PNU282987. These findings suggest that PPAR-α plays a key role in a putative antinociceptive α7 nicotinic signaling pathway. [ABSTRACT FROM AUTHOR]

• Expression of RIC3 is dynamically regulated following immune activation. • Similar expression dynamics suggest similar function for RIC3 and α7 nAChR. • RIC3 is required for the cholinergic anti-inflammatory pathway. • RIC3 and α7 nAChR expression increases in some MS patients. • Expression of RIC3 and of the α7 nAChR encoding gene is correlated in MS patients. Nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels having many functions including inflammation control, as part of the cholinergic anti-inflammatory pathway. Genome wide association studies implicated RIC3, a chaperone of nAChRs, in multiple sclerosis (MS), a neuroinflammatory disease. To understand the involvement of RIC3 in inflammatory diseases we examined its expression, regulation, and function in activated immune cells. Our results show that immune activation leads to dynamic changes in RIC3 expression, in a mouse model of MS and in human lymphocytes and macrophages. We also show similarities in the expression dynamics of RIC3 and CHRNA7, encoding for the α7 nAChR subunit. Homomeric α7 nAChRs were shown to mediate the anti-inflammatory effects of cholinergic agonists. Thus, similarity in expression dynamics between RIC3 and CHRNA7 is suggestive of functional concordance. Indeed, siRNA mediated silencing of RIC3 in a mouse macrophage cell line eliminates the anti-inflammatory effects of cholinergic agonists. Furthermore, we show increased average expression of RIC3 and CHRNA7 in lymphocytes from MS patients, and a strong correlation between expression levels of these two genes in MS patients but not in healthy donors. Together, our results are consistent with a role for RIC3 and for the mechanisms regulating its expression in inflammatory processes and in neuroinflammatory diseases. [ABSTRACT FROM AUTHOR]