Synthesis and Evaluation of the First Fluorescent Antagonists of the Human P2Y(2) Receptor Based on AR-C118925

[Image: see text] The human P2Y(2) receptor (hP2Y(2)R) is a G-protein-coupled receptor that shows promise as a therapeutic target for many important conditions, including for antimetastatic cancer and more recently for idiopathic pulmonary fibrosis. As such, there is a need for new hP2Y(2)R antagonists and molecular probes to study this receptor. Herein, we report the development of a new series of non-nucleotide hP2Y(2)R antagonists, based on the known, non-nucleotide hP2Y(2)R antagonist AR-C118925 (1), leading to the discovery of a series of fluorescent ligands containing different linkers and fluorophores. One of these conjugates, 98, displayed micromolar affinity for hP2Y(2)R (pK(d) = 6.32 ± 0.10, n = 17) in a bioluminescence-energy-transfer (BRET) assay. Confocal microscopy with this ligand revealed displaceable membrane labeling of astrocytoma cells expressing untagged hP2Y(2)R. These properties make 98 one of the first tools for studying hP2Y(2)R distribution and organization.