Exploring Non-orthosteric Interactions with a Series of Potent and Selective A₃ Antagonists

Abstract

A library of potent and highly A 3 AR selective pyrimidine-based compounds was designed to explore non-orthosteric interactions within this receptor. Starting from a prototypical orthosteric A 3 AR antagonist (ISVY130), the structure-based design explored functionalized residues at the exocyclic amide L1 region and aimed to provide additional interactions outside the A 3 AR orthosteric site. The novel ligands were assembled through an efficient and succinct synthetic approach, resulting in compounds that retain the A 3 AR potent and selective profile while improving the solubility of the original scaffold. The experimentally demonstrated tolerability of the L1 region to structural functionalization was further assessed by molecular dynamics simulations, giving hints of the non-orthosteric interactions explored by these series. The results pave the way to explore newly functionalized A 3 AR ligands, including covalent drugs and molecular probes for diagnostic and delivery purposes.

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“…We tested homology models of the inactive hA 3 R that were either generated from the crystal structure of an antagonist bound to hA 2A R (PDB ID 3EML ) provided by Adenosineland web-service and from ref or based on the crystal structure of an antagonist bound to A 1 R (PDB ID 5UEN ) provided in ref (see Figure S1 and the Methods Section). The 100 ns MD simulations of the complexes between A17 and hA 3 R for all of these three homology models converged to structures with a root-mean-square deviation value of protein (p) Cα carbons of all ΤMs [RMSD p (Cα)] that were <∼2.1 Å compared to the starting structure.…”

“…In this study, we have investigated three different homology models of , Model 1 , ,, Model 2 and Model 3 . Homology models for Model 1 (available from refs , ) and Model 2 (available from ref ) were generated using bioinformatics/chemoinformatics tools and are based on the crystal structures of the inactive hA 2A R or inactive hA 1 R, respectively.…”

“…When experimental structures are not available, as in the case of hA 3 R, homology models can be generated from available experimental structures of closely related protein homologues , using bioinformatics tools. Prior to the resolution of their structures, homology models were used extensively to explore both agonist and antagonist binding and for structure-based drug design against the hA 1 R, , hA 2A R, and hA 2B R. , Homology models of the inactive hA 3 R ,− or intermediate-active hA 3 R ,,,,,, or active hA 3 R ,, have been generated using the experimental structures of inactive or intermediate or active hA 2A R or the active and inactive hA 1 R, combined with bioinformatics-/chemoinformatics-based tools. Optimization of the homology models of the hA 3 R has also been applied based on a multireceptor conformation (MRC) selection ,, and a ligand docking-guided model optimization. ,, …”

“…These comparisons produced three unique homology models of hA 3 R in complex with the reference ligand. One model was based on the structure of the inactive hA 2A R in complex with an antagonist, ,, the second model was based on the structure of the inactive hA 1 R in complex with an antagonist, and the third was based on a multistate AF2 method . We investigated the ability of these three models to predict the experimental relative binding free energies and relative RTs of the studied antagonists against the inactive hA 3 R. For the alchemical perturbation calculations of relative binding free energies, we applied TI/MD calculations .…”

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“…Fourteen Letters and two Viewpoints have been submitted by top researchers in the most relevant areas. Letters cover subjects such as discovery of novel tripeptides as integrin-linked kinase modulating agents (Javier Garcia-Marin, Juan J. Vaquero, Diego Rodrı́guez-Puyol, et al), peptide shuttles for brain delivery (Miquel Vila-Perelló, Vera Neves, et al), triazene hybrids for the treatment of melanoma (Maria Jesus Perry et al), SAR of oxindole–lactam hybrids as selective butyrylcholinesterase inhibitors (Marta Pineiro et al), 2-prenylated alkoxylated benzopyrans with PPARα/γ agonist activity (Nuria Cabedo, Diego Cortes, et al), triple PI3K/mTOR/PIM inhibitors (Joaquı́n Pastor et al), piperazinyl bicyclic derivatives as selective calcium channels ligands (Carmen Almansa et al), indole-containing pyrazino­[2,1- b ]­quinazoline-3,6-diones active against Plasmodium and trypanosomatids (Fátima Nogueira, Emı́lia Sousa, et al), evaluation of non-orthosteric interactions with potent and selective A 3 antagonists (José Brea, Hugo Gutiérrez-de-Terán, Eddy Sotelo, et al), steroid–quinoline hybrids to prevent protein aggregation (Artur M. S. Silva et al), the BASHY platform for the assembly of a fluorescent bortezomib–GV1001 conjugate (Uwe Pischel, Sandra N. Pinto, Pedro M. P. Gois, et al), pyridazin-3­(2 H )-one guanidine-based derivatives as DNA binders (Carmen Terán et al), compounds from Plectranthus mutabilis Codd. leaves as modulators of P-glycoprotein activity (Milica Pešić, Patrı́cia Rijo, et al), and imidazoline- and benzamidine-based trypanosome alternative oxidase inhibitors (Christophe Dardonville et al) .…”