Synthesis and Evaluation of PET Radiotracers for the Presynaptic High-affinity Choline Transporter

1041 Objectives Acetylcholine (ACh) is essential for autonomic function, motor control, memory and learning, and dysfunction of cholinergic signaling plays a critical role in several neurodegenerative diseases. Current cholinergic PET radiotracers such as N-[11C]methylpiperidiny-4-yl propionate ([11C]PMP: acetylcholinesterase) and [18F]fluoro-ethoxybenzovesamicol ([18F]FEOBV: vesicular acetylcholine transporter) have provided valuable information on the integrity of cholinergic synapses in PD, but they are not capable of quantifying the level at which the synapses are working. As a new approach to studying the functional status of the cholinergic neurons, we propose the in vivo imaging of the presynaptic high-affinity choline transporter, CHT-1. The transport of choline across the synaptic membrane by the CHT-1 is the rate-limiting determinant in ACh biosynthesis; the functional status of the transporter is regulated by shuttling of transporter molecules from the cytosol to the synaptic membrane. In vivo radioligands for the CHT-1 have not been available due to the lack of high affinity ligands that can readily cross the blood-brain barrier. To address the need for a CHT- 1 radiotracer, the goals of this project are the synthesis and in vivo evaluation of [11C]ML352, a potential PET radiotracer based upon a scaffold with high affinity for CHT-1 (Ennis et al., ACS Chem. Neurosci. 2015;6:417-27). Methods The lead compound, ML352, was discovered by high throughput screening and identified as a high affinity (Ki = 92 nM) and selective inhibitor of CHT-1. Pharmacokinetic studies demonstrated metabolic stability and good CNS penetration (Ennis et al., 2015). [11C]ML352 was synthesized from the desmethyl piperidinic precursor by treatment with [11C]methyl triflate in an automated radiochemistry synthesis module. Results [11C]ML352 was produced in excellent radiochemical yield (105 mCi from 3 Ci of [11C]CO2, corresponding to a non-corrected radiochemical yield of 3.5%), radiochemical purity (>99%) and specific activity (7777 Ci/mmol). Product identification was confirmed by HPLC co-injection with the unlabeled reference standard and the radiotracer was formulated in an isotonic solution of ethanolic saline suitable for in vivo use in pre-clinical PET imaging studies. Preliminary PET imaging studies of [11C]ML352 in CHT-1 overexpressing mice and nonhuman primates are ongoing to explore in vivo functional neuroimaging of CHT-1. Conclusions The synthesis of [11C]ML352, the first brain-permeant PET radiotracer for the CHT-1 transporter, has been completed. The feasibility of imaging CHT-1 activity using [11C]ML352 PET is underway. Acknowledgements: Financial support for this work from the NIH (1P50NS091856-01) and the Dystonia Medical Research Foundation is gratefully acknowledged.