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]<b>18</b>). The unlabeled compound was stable in rat liver microsomes and selective versus other ADP-activated receptors. [³H]<b>18</b> displayed high-affinity binding to membrane preparations recombinantly expressing the human P2Y₁₂R (<i>K</i>_D = 4.57 nM), showing very low nonspecific binding. Radioligand binding assays were established and employed to characterize P2Y₁₂Rs natively expressed in human platelet (<i>K</i>_D = 2.53 nM), rat brain cortex (<i>K</i>_D = 5.35 nM), and mouse microglial cell preparations (<i>K</i>_D = 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.