In Silico Finding of Key Interaction Mediated α3β4 and α7 Nicotinic Acetylcholine Receptor Ligand Selectivity of Quinuclidine-Triazole Chemotype

The selective binding of six (S)-quinuclidine-triazoles and their (R)-enantiomers to nicotinic acetylcholine receptor (nAChR) subtypes α3β4 and α7, respectively, were analyzed by in silico docking to provide the insight into the molecular basis for the observed stereospecific subtype discrimination. Homology modeling followed by molecular docking and molecular dynamics (MD) simulations revealed that unique amino acid residues in the complementary subunits of the nAChR subtypes are involved in subtype-specific selectivity profiles. In the complementary β4-subunit of the α3β4 nAChR binding pocket, non-conserved AspB173 through a salt bridge was found to be the key determinant for the α3β4 selectivity of the quinuclidine-triazole chemotype, explaining the 47–327-fold affinity of the (S)-enantiomers as compared to their (R)-enantiomer counterparts. Regarding the α7 nAChR subtype, the amino acids promoting a however significantly lower preference for the (R)-enantiomers were the conserved TyrA93, TrpA149 and TrpB55 residues. The non-conserved amino acid residue in the complementary subunit of nAChR subtypes appeared to play a significant role for the nAChR subtype-selective binding, particularly at the heteropentameric subtype, whereas the conserved amino acid residues in both principal and complementary subunits are essential for ligand potency and efficacy

Selectivity Optimization of Substituted 1,2,3-Triazoles as α7 Nicotinic Acetylcholine Receptor Agonists

Three series of substituted <i>anti</i>-1,2,3-triazoles (IND, PPRD, and QND), synthesized by cycloaddition from azide and alkyne building blocks, were designed to enhance selectivity and potency profiles of a lead α7 nicotinic acetylcholine receptor (α7-nAChR) agonist, TTIn-1. Designed compounds were synthesized and screened for affinity by a radioligand binding assay. Their functional characterization as agonists and antagonists was performed by fluorescence resonance energy transfer assay using cell lines expressing transfected cDNAs, α7-nAChRs, α4β2-nAChRs, and 5HT_3A receptors, and a fluorescence cell reporter. In the IND series, a tropane ring of TTIn-1, substituted at N1, was replaced by mono- and bicyclic amines to vary length and conformational flexibility of a carbon linker between nitrogen atom and N1 of the triazole. Compounds with a two-carbon atom linker optimized binding with K_d’s at the submicromolar level. Further modification at the hydrophobic indole of TTIn-1 was made in PPRD and QND series by fixing the amine center with the highest affinity building blocks in the IND series. Compounds from IND and PPRD series are selective as agonists for the α7-nAChRs over α4β2-nAChRs and 5HT_3A receptors. Lead compounds in the three series have EC₅₀’s between 28 and 260 nM. Based on the EC₅₀, affinity, and selectivity determined from the binding and cellular responses, two of the leads have been advanced to behavioral studies described in the companion article (DOI: 10.1021/acschemneuro.5b00059)

Cognitive Improvements in a Mouse Model with Substituted 1,2,3-Triazole Agonists for Nicotinic Acetylcholine Receptors

The α7 nicotinic acetylcholine receptor (nAChR) is a recognized drug target for dementias of aging and certain developmental disorders. Two selective and potent α7-nAChR agonists, winnowed from a list of 43 compounds characterized in a companion article (DOI: 10.1021/acschemneuro.5b00058), 5-((quinuclid-3-yl)-1<i>H</i>-1,2,3-triazol-4-yl)-1<i>H</i>-indole (IND8) and 3-(4-hydroxyphenyl-1,2,3-triazol-1-yl) quinuclidine (QND8), were evaluated for cognitive improvement in both short- and long-term memory. Tacrine, a centrally active acetylcholinesterase inhibitor, and PNU-282987, a congeneric α7 nAChR agonist, were employed as reference standards. Three behavioral tests, modified Y-maze, object recognition test (ORT), and water maze, were performed in scopolamine-induced amnesic mice. Intraperitoneal injection of these two compounds significantly improved the cognitive impairment in a modified Y-maze test (5 μmol/kg for IND8 and 10 μmol/kg for QND8), ORT (10 μmol/kg), and water maze test (25 μmol/kg). For delay induced memory deficit or natural memory loss in mice, IND8 and QND8 at 10 μmol/kg were able to enhance memory comparable to PNU-282987 when evaluated using ORT time delay model. Cognitive enhancement of IND8 and QND8 was mediated through α7-nAChRs as evidenced by its complete abolition after pretreatment with a selective α7-nAChR antagonist, methyllycaconitine. These data demonstrate that IND8 and QND8 and their congeners are potential candidates for treatment of cognitive disorders, and the substituted triazole series formed by cycloaddition of alkynes and azides warrant further preclinical optimization

Varying Chirality Across Nicotinic Acetylcholine Receptor Subtypes: Selective Binding of Quinuclidine Triazole Compounds

The novel quinuclidine <i>anti</i>-1,2,3-triazole derivatives <b>T1</b>–<b>T6</b> were designed based on the structure of <b>QND8</b>. The binding studies revealed that the stereochemistry at the C3 position of the quinuclidine scaffold plays an important role in the nAChR subtype selectivity. Whereas the (<i>R</i>)-enantiomers are selective to α7 over α4β2 (by factors of 44–225) and to a smaller degree over α3β4 (3–33), their (<i>S</i>)-counterparts prefer α3β4 over α4β2 (62–237) as well as over α7 (5–294). The (<i>R</i>)-derivatives were highly selective to α7 over α3β4 subtypes compared to (<i>RS</i>)- and (<i>R</i>)-<b>QND8</b>. The (<i>S</i>)-enantiomers are 5–10 times more selective to α4β2 than their (<i>R</i>) forms. The overall strongest affinity is observed for the (<i>S</i>)-enantiomer binding to α3β4 (<i>K</i>_i, 2.25–19.5 nM) followed by their (<i>R</i>)-counterpart binding to α7 (<i>K</i>_i, 22.5–117 nM), with a significantly weaker (<i>S</i>)-enantiomer binding to α4β2 (<i>K</i>_i, 414–1980 nM) still above the very weak respective (<i>R</i>)-analogue affinity (<i>K</i>_i, 5059–10436 nM)