Synthesis and in vitro evaluation of novel compounds and discovery of a promising iodine-125 radioligand for purinergic P2X7 receptor (P2X7R)

We discovered a highly potent P2X7R-specific radioligand [125I]1c which we used to establish a screening platform for new P2X7R compounds. [Display omitted] •A series of P2X7R ligands were designed and synthesized.•The fluorescence assay identified that iodide compound 1c has high potency and specificity for P2X7R.•The radiosynthesis of [125I]1c was achieved with good radiochemical yield (44 ± 12 %, n = 3) and high radiochemical purity (>95 %).•The radioligand [125I]1c was used to establish a screening platform for new P2X7R compounds. The purinergic P2X ligand-gated ion channel 7 receptor (P2X7R) plays a critical role in various inflammatory processes and other diseases. Fast determination of compounds P2X7R binding potency and discovery of a promise PET radiotracer for imaging P2X7R require a P2X7R suitable radioligand for radioactive competitive binding assay. Herin, we designed and synthesized thirteen new P2X7R ligands and determined the in vitro binding potency. The fluorescence screening assay identified the iodide compound 1c with high potency and specificity toward P2X7R with an IC50 of 0.25 ± 0.05 nM. Therefore, 1c was 125I-labeled to afford [125I]1c with a good radiochemical yield (44 ± 12 %, n = 3) and high radiochemical purity (>95 %). Radioligand saturation binding assay showed that [125I]1c specifically bound to human P2X7R with high affinity (Kd = 1.68 nM and Bmax = 94 fmol/mg). A radioactive high throughput binding assay using [125I]1c for our new compounds demonstrated that the imidazole compounds 1b, 1c, and 1d exhibited high inhibition for >70 %, while the analogues of GSK314181A exhibited low inhibition for