Brox, Regine and Milanos, Lampros and Saleh, Noureldin and Baumeister, Paul and Buschauer, Armin and Hofmann, Dagmar and Heinrich, Markus R. and Clark, Timothy and Tschammerl, Nuska (2018) Molecular Mechanisms of Biased and Probe-Dependent Signaling at CXC-Motif Chemokine Receptor CXCR3 Induced by Negative Allosteric Modulators. MOLECULAR PHARMACOLOGY, 93 (4). pp. 309-322. ISSN 0026-895X, 1521-0111
Full text not available from this repository. (Request a copy)Abstract
Our recent explorations of allosteric modulators with improved properties resulted in the identification of two biased negative allosteric modulators, BD103 (N-1-{[3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[2,3-d]pyrinni-din2yl]ethyl}-4-(4-fluorobutoxy)-N-[(1 -methylpiperidin-4-yl)methyl}]butanannide) and BD064 (5-[(N-{1-[3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[2,3-d]pyrinnidin-2-yl]ethyl-2-[4-fluoro-3-(trifluoromethyl)phenyl]acetannido)methyl]-2-fluorophenyl}boronic acid), that exhibited probe-dependent inhibition of CXC-motif chemokine receptor CXCR3 signaling. With the intention to elucidate the structural mechanisms underlying their selectivity and probe dependence, we used site-directed mutagenesis combined with homology modeling and docking to identify amino acids of CXCR3 that contribute to modulator binding, signaling, and transmission of cooperativity. With the use of allosteric radioligand RAMX3 ([H-3]N-{1-[3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[2,3-d]pyrinnidin-2-yl]ethyl}-2-[4-fluoro-3-(trifluoronnethyl)phenyl]-N-[(1-methylpiperidin-4-yl)methyl] acetamide), we identified that F131(3.32) and Y308(7.43) contribute specifically to the binding pocket of BD064, whereas D186(4.60) solely participates in the stabilization of binding conformation of BD103. The influence of mutations on the ability of negative allosteric modulators to inhibit chemokine-mediated activation (CXCL11 and CXCL10) was assessed with the bioluminescence resonance energy transfer-based cAMP and beta-arrestin recruitment assay. Obtained data revealed complex molecular mechanisms governing biased and probe-dependent signaling at CXCR3. In particular, F131(3.32), S304(7.39), and Y308(7.43) emerged as key residues for the compounds to modulate the chemokine response. Notably, D186(4.60), W268(6.48), and S304(7.39) turned out to play a role in signal pathway selectivity of CXCL10, as mutations of these residues led to a G protein-active but beta-arrestin-inactive conformation. These diverse effects of mutations suggest the existence of ligand- and pathway-specific receptor conformations and give new insights in the sophisticated signaling machinery between allosteric ligands, chemokines, and their receptors, which can provide a powerful platform for the development of new allosteric drugs with improved pharmacological properties.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | PROTEIN-COUPLED RECEPTORS; PHARMACOLOGICAL CHARACTERIZATION; FUNCTIONAL SELECTIVITY; BINDING-SITE; LIGANDS; ACTIVATION; EXPRESSION; PATHWAYS; AGONISM; BETA-ARRESTIN-1; |
| Subjects: | 500 Science > 540 Chemistry & allied sciences |
| Divisions: | Chemistry and Pharmacy > Institute of Pharmacy > Pharmaceutical/Medicinal Chemistry II (Prof. Buschauer) |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 05 Mar 2020 08:49 |
| Last Modified: | 05 Mar 2020 08:49 |
| URI: | https://pred.uni-regensburg.de/id/eprint/14754 |
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