Abstract View

Objectives: Significant advances have been made over the last several decades toward the development of specific PET radiotracers to study various tauopathies such as Alzheimer’s disease (AD) and progressive supranuclear palsy (PSP).^1-5 These diseases are characterized by the misfolding of tau protein into different isoforms, which differ based on the number of repeated (R) units (i.e., 3R/4R – AD; 3R – Pick’s Disease; 4R – PSP). Therefore, there is an urgent need for novel radiotracers which can bind specific isoforms of tau to precisely diagnose and track the progression of different tauopathies with high specificity. Our laboratories and others have developed several PET tracers for imaging non-AD tauopathies, however, an optimized 4R-tau PET tracer is still needed.^6-9 Our lab recently developed Z-3272 which showed selective binding to 4R-tau in vitro. However, [¹¹C]Z-3272 exhibited rapid metabolism which gave rise to brain-penetrant radiometabolites.¹⁰

To improve the metabolic stability of Z-3272, analog generation was performed computationally in SmallWorld,¹¹ a powerful chemical similarity search tool. This in silico screening resulted in the identification of methyl (1-(5-(8-methoxynaphthalen-2-yl)pyridin-2-yl)piperidin-4-yl)carbamate, hereto referred to as 18A-Me. This work describes the automated synthesis of [¹¹C]18A-Me for preclinical evaluation.

Methods: [¹¹C]Methyl iodide was prepared from cyclotron-produced [¹¹C]CO₂ using a GE TracerLab FX2 C. [¹¹C]MeI was passed through and trapped in the HPLC loop which was pre-loaded with a solution of 0.4 mg of the des-methyl precursor methyl (1-(5-(8-hydroxynaphthalen-2-yl)pyridin-2-yl)piperidin-4-yl)carbamate (18A) and 1.0 equiv. of t-BuOK in 80 µL of DMF. The mixture was allowed to react for 5 min “in loop”, before being injected onto a semi-preparative HPLC column (LUNA C18(2), 250 x 10 mm; 5 mL/min; 0.1M NH₄HCO₂ 65% MeCN: 35% H₂O, pH = 5) for purification. The product was collected and diluted (20 mL HHasdfH₂O with 2 mL 0.8% NaHCO₃) and reformulated using solid-phase extraction (tC18) resulting in a final dose of 10% ethanol in saline. Radiochemical purity, molar activity and chemical purity were measured using analytical HPLC (LUNA C18(2), 150 x 4.6 mm; 1 mL/min; gradient: 0.1M NH₄HCO₂ 65% MeCN: 35% H₂O, pH = 5).

Results: We report the radiosynthesis of a new 4R-tau PET tracer candidate [¹¹C]18A-Me (n = 3) with a radiochemical yield (RCY) of 18% ± 7% (from starting [¹¹C]CO₂), and total synthesis time of ~40 minutes. The product was isolated and formulated consistently in >99% radiochemical purity (RCP) and molar activities of 341 GBq/µmol ± 104 GBq/µmol.

Conclusions: [¹¹C]18A-Me was successfully produced with high RCY and RCP suitable for preclinical use on a commercial automated radiosynthesis unit. In vitro tau binding results and preclinical PET imaging studies to further characterize [¹¹C]18A-Me as a 4R-tau-selective radiotracer are underway and will be reported.

References:

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3.     Vasdev, N.; et al. Ligand-Based Design of [¹⁸F]OXD-2314 for PET Imaging in Non-Alzheimer’s Disease Tauopathies. Nat. Comm. 2024, 15, 5109 – 5121.

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