The Ascaris suum nicotinic receptor, ACR-16, as a drug target: Four novel negative allosteric modulators from virtual screening

Abstract

Soil-transmitted helminth infections in humans and livestock cause significant debility, reduced productivity and economic losses globally. There are a limited number of effective anthelmintic drugs available for treating helminths infections, and their frequent use has led to the development of resistance in many parasite species. There is an urgent need for novel therapeutic drugs for treating these parasites. We have chosen the ACR-16 nicotinic acetylcholine receptor of Ascaris suum (Asu-ACR-16), as a drug target and have developed three-dimensional models of this transmembrane protein receptor to facilitate the search for new bioactive compounds. Using the human α7 nAChR chimeras and Torpedo marmorata nAChR for homology modeling, we defined orthosteric and allosteric binding sites on the Asu-ACR-16 receptor for virtual screening. We identified four ligands that bind to sites on Asu-ACR-16 and tested their activity using electrophysiological recording from Asu-ACR-16 receptors expressed in Xenopus oocytes. The four ligands were acetylcholine inhibitors (SB-277011-A, IC50, 3.12 ± 1.29 μM; (+)-butaclamol Cl, IC50, 9.85 ± 2.37 μM; fmoc-1, IC50, 10.00 ± 1.38 μM; fmoc-2, IC50, 16.67 ± 1.95 μM) that behaved like negative allosteric modulators. Our work illustrates a structure-based in silico screening method for seeking anthelmintic hits, which can then be tested electrophysiologically for further characterization.

(S)-5-ethynyl-anabasine, a novel compound, is a more potent agonist than other nicotine alkaloids on the nematode Asu -ACR-16 receptor

“…Three crystal structures of a human α7 nAChR chimera co-crystalized with ligands of different modes of action were used as templates (Table 1) to build three different bound-form models of the ECD- Asu- ACR-16 (Li et al., 2011, Huang et al., 2013, Zheng et al., 2016). Smiles strings of nicotine derivatives were obtained from the ZINC website (http://zinc.docking.org/search/structure) and converted to PDBQT format for our docking studies.…”

“…Smiles strings of nicotine derivatives were obtained from the ZINC website (http://zinc.docking.org/search/structure) and converted to PDBQT format for our docking studies. Docking of these ligands was performed at the orthosteric ligand-binding sites of agonist-bound, apo (no ligand) and antagonist-bound forms of the ECD -Asu -ACR-16 models using AutoDock Vina Software (Trott and Olson, 2010, Zheng et al., 2016). We investigated the intermolecular interactions in our models by comparing the intermolecular distances in the crystal structures and our models (≤ 1 Å difference).…”

“…were prepared using the previously described methods (Zheng et al., 2016). A cRNA mixture of 25 ng Asu-acr-16 and 5 ng Asu-ric-3 cRNA in 50 nL RNAse-free water was injected into de-folliculated Xenopus laevis oocytes (Ecocyte Bioscience, Austin, TX, USA).…”

“…Asu- ACR-16 is widely distributed in A. suum tissues but its physiological function remains to be determined (Abongwa et al., 2016, Zheng et al., 2016). As one of the recently characterized nematode parasitic nAChRs, Asu- ACR-16 is pharmacologically different to its host α7 nAChR and may be exploited as an anthelmintic drug target to counter resistance to cholinergic anthelmintics directed at other pharmacological types of nAChR (Holden-Dye et al., 2013, Zheng et al., 2016). …”

“…Five conserved aromatic residues and two hydrogen-bond interacting residues have orientations very close to corresponding residues in other nAChRs, facilitating our investigation of drug-receptor interactions of the Asu- ACR-16 receptor (Zheng et al., 2016). …”