1. Structure-based identification and characterisation of structurally novel human P2X7 receptor antagonists.
- Author
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Caseley EA, Muench SP, Fishwick CW, and Jiang LH
- Subjects
- Adenosine Triphosphate analogs & derivatives, Adenosine Triphosphate chemistry, Adenosine Triphosphate metabolism, Adenosine Triphosphate pharmacology, Amides chemistry, Amides metabolism, Animals, Anti-Inflammatory Agents, Non-Steroidal chemistry, Anti-Inflammatory Agents, Non-Steroidal metabolism, Anti-Inflammatory Agents, Non-Steroidal pharmacology, Binding Sites, Calcium Signaling drug effects, HEK293 Cells, Hepatocytes cytology, Hepatocytes metabolism, Humans, Image Processing, Computer-Assisted, Indoles chemistry, Indoles metabolism, Ligands, Microscopy, Fluorescence, Patch-Clamp Techniques, Purinergic P2X Receptor Antagonists chemistry, Purinergic P2X Receptor Antagonists metabolism, Rats, Receptors, Purinergic P2X3 chemistry, Receptors, Purinergic P2X3 genetics, Receptors, Purinergic P2X3 metabolism, Receptors, Purinergic P2X4 chemistry, Receptors, Purinergic P2X4 genetics, Receptors, Purinergic P2X4 metabolism, Receptors, Purinergic P2X7 chemistry, Receptors, Purinergic P2X7 genetics, Recombinant Proteins chemistry, Recombinant Proteins metabolism, Single-Cell Analysis, Small Molecule Libraries, Structure-Activity Relationship, Thiophenes chemistry, Thiophenes metabolism, Amides pharmacology, Hepatocytes drug effects, Indoles pharmacology, Models, Molecular, Purinergic P2X Receptor Antagonists pharmacology, Receptors, Purinergic P2X7 metabolism, Thiophenes pharmacology
- Abstract
The P2X7 receptor (P2X7R) plays an important role in diverse conditions associated with tissue damage and inflammation, meaning that the human P2X7R (hP2X7R) is an attractive therapeutic target. The crystal structures of the zebrafish P2X4R in the closed and ATP-bound open states provide an unprecedented opportunity for structure-guided identification of new ligands. The present study performed virtual screening of ∼100,000 structurally diverse compounds against the ATP-binding pocket in the hP2X7R. This identified three compounds (C23, C40 and C60) out of 73 top-ranked compounds by testing against hP2X7R-mediated Ca(2+) responses. These compounds were further characterised using Ca(2+) imaging, patch-clamp current recording, YO-PRO-1 uptake and propidium iodide cell death assays. All three compounds inhibited BzATP-induced Ca(2+) responses concentration-dependently with IC50s of 5.1±0.3μM, 4.8±0.8μM and 3.2±0.2μM, respectively. C23 and C40 inhibited BzATP-induced currents in a reversible and concentration-dependent manner, with IC50s of 0.35±0.3μM and 1.2±0.1μM, respectively, but surprisingly C60 did not affect BzATP-induced currents up to 100μM. They suppressed BzATP-induced YO-PRO-1 uptake with IC50s of 1.8±0.9μM, 1.0±0.1μM and 0.8±0.2μM, respectively. Furthermore, these three compounds strongly protected against ATP-induced cell death. Among them, C40 and C60 exhibited strong specificity towards the hP2X7R over the hP2X4R and rP2X3R. In conclusion, our study reports the identification of three novel hP2X7R antagonists with micromolar potency for the first time using a structure-based approach, including the first P2X7R antagonist with preferential inhibition of large pore formation., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2016
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