Cook, B. E.; Pickel, T. C.; Nag, S.; Bolduc, P. N.; Beshr, R.; Forsberg Moren, A.; Muste, C.; Boscutti, G.; Jiang, D.; Yuan, L.; Datta, P.; Ochniewicz, P.; Khani Meynaq, Y.; Tang, S.-P.; Plisson, C.; Amatruda, M.; Zhang, Q.; DuBois, J. M.; Delavari, A.; Klein, S. K.; Polyak, I.; Shoroye, A.; Girmay, S.; Halldin, C.; Martarello, L.; Peterson, E. A.; Kaliszczak, M.

Determination of a drug\'s biodistribution is critical to ensure it reaches the target tissue of interest. This is particularly challenging in the brain where invasive sampling methods may not be possible. Here, a pretargeted imaging methodology is disclosed that utilizes bioorthogonal click chemistry to determine the distribution of an antisense oligonucleotide in the living brain following intrathecal dosing. A novel positron emission tomography (PET) tracer, [18F]BIO-687, bearing a click-reactive trans-cyclooctene (TCO) was discovered and tested in conjunction with a Malat1 antisense oligonucleotide (ASO) conjugated with a methyltetrazine (MeTz). PET imaging in rats demonstrated that the tracer possesses good kinetic properties for CNS imaging and can react to form a covalent linkage with high specificity to the MeTz-conjugated-ASO in vivo. Further, the amount of tracer reacted by cycloaddition with the Tz was determined to be dependent on the concentration of ASO-MeTz in tissue, as determined through comparison of the imaging signal with the LC-MS of the tissue homogenate. The approach was evaluated in cynomolgus monkeys, using both Malat1 and the MAPT ASO BIIB080, with PET imaging showing favorable tracer kinetics and specific binding to both ASOs in vivo. These results demonstrate that the tracer [18F]BIO-687 can image intrathecally-delivered ASO distribution in the brain, and future studies should leverage this technology to evaluate ASO distribution in human patients to study distribution.