PROJECT 2: Development of 4R-Tau PET Radioligands for Imaging Tauopathies ABSTRACT The goal of Project 2 is to develop a positron emission tomography (PET) radiopharmaceutical useful for imaging 4-repeat (4R) tauopathies, which include progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), and frontotemporal lobar degeneration (FTLD-tau). Several useful PET agents for imaging mixed 3R/4R- tau aggregates in Alzheimer’s disease (AD) have been reported, but none of these agents has proven highly sensitive for imaging 4R-tau in non-AD 4R-tauopathies, and higher affinity 4R-tau radioligands are needed to improve specific binding signal in vivo. Novel disease modifying therapies that target tau offer new treatment possibilities for patients with different tauopathies. Key to a successful therapeutic strategy will be the ability to discriminate patients with different tauopathies and to assess the efficacy of anti-tau treatments. The proposed research in this project will pursue parallel tracks of lead compound identification and optimization, working closely with the Medicinal Chemistry and Radiochemistry Core (MCRC) to identify new 4R-tau candidate radioligands for subsequent evaluation in Project 2. Binding studies in tissue specimens of 4R-tauopathies and other proteinopathies will characterize the sensitivity and specificity of candidate radioligands for binding to aggregated 4R-tau, identifying the most promising leads to advance into in vivo studies in rodents and non- human primates. The MCRC will prepare a series of analogs from two lead series: Z-indoles and substituted- indazoles for in vitro binding assays in Project 2 and further define the structure-activity relationship of the analogs. The MCRC will employ its computational resources to help guide the selection of new analogs to be purchased from commercial sources or synthesized by the MCRC, and subsequently evaluated in Project 2. In vitro binding assays conducted in Project 2 will test the predictive power of the in silico molecular similarity studies conducted by the MCRC and help refine binding site features that most influence radioligand-4R-tau high affinity binding interactions. Project 2 will employ a staged approach using both in vitro and in vivo assay methods to characterize and evaluate candidate 4R-tau PET imaging agents provided by the MCRC, identifying the most promising 4R-tau agents for subsequent evaluation in first-in-human studies of PSP and FTLD-tau subjects in the Clinical Core.