Synthesis and Evaluation of GAT-1 Selective PET Probes

Abstract

Positron emission tomography (PET) is a high resolution, non-invasive functional imaging technique used for observation of biochemical processes in vivo. This thesis describes work towards using this imaging modality to better understand the processes of the major inhibitory neurotransmitter gamma(γ)-amino butyric acid (GABA). The GABAergic system is understood to be involved in many neurological diseases and disorders including Alzheimer’s disease, epilepsy, schizophrenia, autism and depression. In the GABAergic system, GABA receptors and transporters are targets for drug development as well as investigation as biomarkers. While imaging of the post-synaptic GABAergic receptor subtype GABAA is possible with the use of [11C]flumazenil ([11C]FMZ), there is no complimentary PET radiotracer enabling in vivo imaging of GABA in pre-synaptic neurons. Located pre-synaptically and in high abundance in areas such as the cortex and basal ganglia, GABA Transporter subtype 1 (GAT-1) presents as an interesting target for pre-synaptic GABA imaging. Studies have shown an increase in GAT-1 expression, especially in the cortex, in the schizophrenic brain. GAT-1 is responsible for removal of GABA from the synaptic cleft, thereby ending its action on post-synaptic GABA receptors. Tiagabine was approved by the FDA in 1997 and remains the only GAT-1 inhibitor available today. Many structure activity relationship (SAR) studies have sought to improve the selectivity and affinity of tiagabine for GAT-1. Some research has been successful in the radiolabeling of these compounds for imaging for preclinical evaluation, but as of yet, no radiotracer has proven successful or advanced to widespread clinical use. To address this urgent need in functional neuroimaging, this work is focused upon developing a GAT-1 selective, blood-brain barrier (BBB) permeable radiotracer for use in PET imaging of the GABAergic system. The synthesis and preliminary evaluation of a number of novel GAT-1 selective PET tracers is described herein. Scaffolds with high affinity and excellent selectivity for GAT-1 were used as leads for adaptation into new PET radiotracers. Successful [18F]radiofluorination of one scaffold was promising, but subsequent in vivo evaluation indicated poor BBB permeability. Additional studies using carbon-11 to label additional small molecule GABA uptake inhibitors were undertaken to test our hypothesis that low brain uptake was due to the highly polar zwitterionic nipecotic acid moiety. As the acid group is necessary for high affinity binding to GAT-1, it cannot be simply removed or esterified. Therefore, future efforts in GAT-1 radiotracer design will need to investigate carboxylic acid bioisosteres in an attempt to improve brain uptake while maintaining affinity for GAT-1. Our preliminary efforts in this direction have focused upon a GAT-1 inhibitor bearing a thiazolyl bioisostere. Synthesis and preliminary evaluation of the new radiotracer are also described in this thesis, and the reasonable affinity for GAT-1 and excellent brain uptake show good promise for 2nd generation radiotracers going forward.