Selective, Non-nucleotidic Radiotracer for P2Y12 Receptors: Design, Synthesis, Characterization, and Imaging of Brain Slices

The G protein-coupled, ADP-activated P2Y12 receptor (P2Y12R) expressed by microglial cells is involved in neuroinflammation constituting a promising biomarker. Here, we designed and characterized a potent and selective non-nucleotidic P2Y12-antagonist radioligand, [3H]PSB-22219 ([3H]18). The unlabeled compound was stable in rat liver microsomes and selective versus other ADP-activated receptors. [3H]18 displayed high-affinity binding to membrane preparations recombinantly expressing the human P2Y12R (KD = 4.57 nM), showing very low nonspecific binding. Radioligand binding assays were established and employed to characterize P2Y12Rs natively expressed in human platelet (KD = 2.53 nM), rat brain cortex (KD = 5.35 nM), and mouse microglial cell preparations (KD = 269 nM), with microglia showing extraordinarily high P2Y12R expression. Autoradiography studies allowed the visualization of human P2Y12R overexpression in the brain of a humanized rat model. The new radioligand is expected to become a useful pharmacological tool that will contribute to the development of therapeutics and radiodiagnostics targeting brain P2Y12Rs.