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Medchemcomm. 2017 Aug 1;8(8):1659-1667. doi: 10.1039/C7MD00247E. Epub 2017 Jun 22.

Bitopic fluorescent antagonists of the A2A adenosine receptor based on pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine functionalized congeners.

Author information

1
Molecular Recognition Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bldg. 8A, Rm. B1A-19, Bethesda, MD 20892-0810, USA.
2
Inserm, CHU Lille, UMR-S 1172 - JPArc - Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer, Univ. Lille, F-59000 Lille, France.
3
Department of Pharmaceutical Chemistry, Telangana University, Nizamabad, Telangana, India 503322.
4
Department of Pharmacology, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, Wisconsin 53226, USA.
5
Unitat de Farmacologia, Departament Patologia i Terapeutica Experimental, Facultat de Medicina, IDIBELL, Universitat de Barcelona, 08907 L'Hospitalet de Llobregat, Spain.

Abstract

A pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine antagonist of the A2A adenosine receptor (AR) was functionalized as amine congeners, fluorescent conjugates and a sulfonate, and the A2AAR binding modes were predicted computationally. The optimal n-butyl spacer was incorporated into the following A2AAR-selective (Ki, nM) conjugates: BODIPY630/650 derivative 11 (MRS7396, 24.6) and AlexaFluor488 derivative 12 (MRS7416, 30.3). Flow cytometry of 12 in hA2AAR-expressing HEK-293 cells displayed saturable binding (low nonspecific) and inhibition by known A2AAR antagonists. Water-soluble sulfonate 13 was a highly potent (Ki = 6.2 nM) and selective A2AAR antagonist based on binding and functional assays. Docking and molecular dynamics simulations predicted the regions of interaction of the distal portions of these chain-extended ligands with the A2AAR. The BODIPY630/650 fluorophore of 11 was buried in a hydrophobic interhelical (TM1/TM7) region, while AlexaFluor488 of 12 associated with the hydrophilic extracellular loops. In conclusion, we have identified novel high affinity antagonist probes for A2AAR drug discovery and characterization.

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