1. Academic Validation
  2. Identification of aurintricarboxylic acid as a potent allosteric antagonist of P2X1 and P2X3 receptors

Identification of aurintricarboxylic acid as a potent allosteric antagonist of P2X1 and P2X3 receptors

  • Neuropharmacology. 2019 Nov 1;158:107749. doi: 10.1016/j.neuropharm.2019.107749.
Astrid S Obrecht 1 Nicole Urban 2 Michael Schaefer 2 Anni Röse 1 Achim Kless 3 Jannis E Meents 4 Angelika Lampert 4 Aliaa Abdelrahman 5 Christa E Müller 5 Günther Schmalzing 1 Ralf Hausmann 6
Affiliations

Affiliations

  • 1 Molecular Pharmacology, RWTH Aachen University, 52074, Aachen, Germany.
  • 2 Rudolf-Boehm-Institut for Pharmacology and Toxicology, University of Leipzig, 04107, Leipzig, Germany.
  • 3 Grünenthal Innovation, Department of Translational Science and Intelligence, Grünenthal GmbH, 52078, Aachen, Germany.
  • 4 Institute of Physiology, RWTH Aachen University, 52074, Aachen, Germany.
  • 5 PharmaCenter Bonn, Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, 53121, Bonn, Germany.
  • 6 Molecular Pharmacology, RWTH Aachen University, 52074, Aachen, Germany. Electronic address: [email protected].
Abstract

The homotrimeric P2X3 receptor, one of the seven members of the ATP-gated P2X Receptor family, plays a crucial role in sensory neurotransmission. P2X3 receptor antagonists have been identified as promising drugs to treat chronic cough and are suggested to offer pain relief in chronic pain such as neuropathic pain. Here, we analysed whether compounds affect P2X3 receptor activity by high-throughput screening of the Spectrum Collection of 2000 approved drugs, Natural Products and bioactive substances. We identified aurintricarboxylic acid (ATA) as a nanomolar-potency antagonist of P2X3 receptor-mediated responses. Two-electrode voltage clamp electrophysiology-based concentration-response analysis and selectivity profiling revealed that ATA strongly inhibits the rP2X1 and rP2X3 receptors (with IC50 values of 8.6 nM and 72.9 nM, respectively) and more weakly inhibits P2X2/3, P2X2, P2X4 or P2X7 receptors (IC50 values of 0.76 μM, 22 μM, 763 μM or 118 μM, respectively). Patch-clamp analysis of mouse DRG neurons revealed that ATA inhibited native P2X3 and P2X2/3 receptors to a similar extent than rat P2X3 and P2X2/3 receptors expressed in Xenopus oocytes. In a radioligand binding assay, up to 30 μM ATA did not compete with [3H]-ATP for rP2X3 receptor binding, indicating a non-competitive mechanism of action. Molecular docking studies, site-directed mutagenesis and concentration-response analysis revealed that ATA binds to the negative allosteric site of the hP2X3 receptor. In summary, ATA as a drug-like pharmacological tool compound is a nanomolar-potency, allosteric antagonist with selectivity towards αβ-methylene-ATP-sensitive P2X1 and P2X3 receptors.

Keywords

Allosteric P2X3 antagonist; Drug-like P2X3 antagonist; High-throughput drug screening; Ligand docking; P2X3 receptor.

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