Synthesis, Radiosynthesis, in Vitro and in Vivo Evaluation of the GABA_A-Benzodiazepine Receptor Agonist [¹¹C]RO6899880

The aim of this study is to synthesize and evaluate the GABA_A-benzodiazepine receptor agonist radiotracer (<i>S</i>)-10-chloro-1-(3-(methoxy-¹¹C)pyrrolidine-1-carbonyl)-3-phenyl-6,7-dihydro-4<i>H</i>-pyrido[2,1-<i>a</i>]isoquinolin-4-one ([¹¹C]RO6899880) using <i>in vitro</i> and <i>in vivo</i> experiments to determine its suitability for human positron emission tomography (PET) neuroimaging studies. RO6899880 and its desmethyl precursor were synthesized over multiple steps in 3% and 2% yields, respectively. Reaction of the precursor with [¹¹C]CH₃I in DMF with NaOH at 23 °C for 5 min followed by HPLC purification and formulation produced [¹¹C]RO6899880 with a radiochemical yield of 4.1 ± 1.6% (nondecay corrected), radiochemical purity >95% and molar activities of 6.40 ± 0.63 GBq/μmol (<i>n</i> = 3). Preliminary autoradiography with [³H]RO6899880 was carried out in brain tissues from rat, healthy human, patients with Alzheimer's disease and chronic traumatic encephalopathy, and aligned with the GABA_A antagonist radiotracer [³H]flumazenil. PET imaging in rats revealed modest brain permeability, moderate clearance, and distribution across brain regions with higher uptake in cortical areas. However, polar radiometabolites were found in both plasma and brain homogenates. PET imaging of [¹¹C]RO6899880 in higher species including radiometabolism studies are needed prior to human translation.