The Structural Basis for GTS-21 Selectivity between Human and Rat Nicotinic α7 Receptors

Abstract

The ␣7 nAChR-selective partial agonist 3-(2,4-dimethoxybenzylidene)anabaseine (GTS-21) is more efficacious and potent for rat receptors than for human ␣7 receptors. Four single amino acid differences exist between human and rat ␣7 in the agonist binding site, two in the C loop, and one each in the E and F loops. Reciprocal mutations were made in these three domains and evaluated in Xenopus laevis oocytes. Mutations in the C and F loops significantly increased the efficacy of GTS-21 for the human receptor mutants but not to the level of the wild-type, and reciprocal mutations in rat ␣7 did not decrease responses to GTS-21. Whereas mutations in the E loop alone were without effect, the E-and F-loop mutations together increased GTS-21 efficacy and potency for human receptors, but the EF mutations in the rat receptors decreased the GTS-21 potency without changing the efficacy. The only mutants that showed a full reversal of the efficacy differences between human and rat ␣7 contained complete exchange of all four sites in the C, E, and F loops or just the sites in the C and F loops. However, the reversal of the potency ratio seen with the EF mutants was not evident in the CEF mutants. Our data therefore indicate that the pharmacological differences between rat and human ␣7 receptors are caused by reciprocal differences in sites within and around the binding site. Specific features in the agonist molecule itself are also identified that interact with these structural features of the receptor agonist binding site.A crucial assumption for the translation of preclinical research from animal studies to human therapeutics is that receptor pharmacology will be consistent between species. That is, drugs shown to be useful based on their ability to work in animal (rodent) models would also have similar activity on human forms of the receptors. The neuronal ␣7-type nicotinic acetylcholine receptor (nAChR) has been identified as a potential target for the treatment of Alzheimer's disease (Lindstrom, 1997), and 3-(2,4-dimethoxybenzylidene) anabaseine (GTS-21; also called DMXBA), which selectively targets this receptor, has been shown to improve learning and memory in animal models of cholinergic hypofunction (Kem, 2000). This ␣7-selective partial agonist has also been shown to prevent the death of differentiated PC-12 cells that occurs after nerve growth factor removal and the death of cultured primary neurons that occurs after high levels of NMDA receptor activation (Martin et al., 1994;Shimohama et al., 1998). It is interesting that although GTS-21 was able to protect PC-12 cells from the cytotoxic effects of amyloid peptide exposure, it was not able to protect human-derived SK-N-SH cells from the same cytotoxic stress, although the GTS-21 4-hydroxy metabolite, 3-(4-hydroxy,2-methoxybenzylidene)anabaseine (4-OH-GTS-21), was cytoprotective in the same assay (Meyer et al., 1998a). A likely explanation for these observed differences in cytoprotective activity came from the observation that GTS-21 was far less efficacious ...

Neurotransmitter GABA Activates Muscle but Notα7 Nicotinic Receptors

“…It could be also possible that, although the overall orientations of GABA appear similar in a/« and a7/a7 interfaces, differences in specific potential interactions may be involved in the differential selectivity. Also, loop F, which could not be included in our homology models, may be involved in the differential actions of GABA because it has been shown to be a determinant for drug selectivity and partial agonism (Stokes et al, 2004;Hibbs et al, 2009). Alternatively, other residues located outside the binding site, but involved in coupling agonist binding to channel opening, may influence the capability of GABA to activate muscle, but not a7 nAChRs.…”

Heterogeneity of nicotinic acetylcholine receptor expression in the caudal nucleus of the solitary tract

“…Cytisine, a nicotine-like alkaloid and a potent agonist of rat β4* nAChRs (Luetje and Patrick, 1991), produced similar effects of facilitation of mEPSCs (n=23, Figures 3C). Cytisine is also known to activate the rat α7 nAChR subtype (Stokes et al, 2004). In the presence of 20 nM MLA, applications of cytisine produced somatic/dendritic responses in numerous experiments (n=18, Figures 6, bottom traces).…”

Alpha7 nicotinic acetylcholine receptor: A link between inflammation and neurodegeneration

“…In vitro, GTS-21 can protect neurons against damage induced by amyloid peptides, this is in agreement with previous studies suggesting that alpha7nAChRs can have a neuroprotective potential. GTS21 have three major adversities for clinical use: (1) GTS21 is not specific for alpha7nAChR and it affects other receptors including alpha4beta2-nAChRs (Gerzanich et al, 1995;Meyer et al, 1998;Stokes et al, 2004) and 5-HT 3A (Machu et al, 2001), (2) GTS21 has high affinity for the rodent receptor but it presents low affinity for the human alpha7nAChR (Gerzanich et al, 1995;Meyer et al, 1998;Stokes et al, 2004), and (3) GTS21 appears to have a limited brain penetration . These limitations appear to be overcome by a second generation of alpha7nAChR-agonists including 4OHGTS (3-(4-hydroxy, 2-methoxybenzylidene) anabaseine) (Meyer et al, 1998;Uteshev et al, 2003), compound that was developed to increase its affinity for human alpha7nAChR (Gerzanich et al, 1995).…”