In vitro and in vivo characterization of two C-11-labeled pet tracers for vesicular acetylcholine transporter

Abstract

Purpose: The vesicular acetylcholine transporter (VAChT) is a specific biomarker for imaging presynaptic cholinergic neurons. Herein, two potent and selective (11)C-labeled VAChT inhibitors were evaluated in rodents and nonhuman primates for imaging VAChT in vivo.

Procedures: For both (-)-[(11)C]2 and (-)-[(11)C]6, biodistribution, autoradiography, and metabolism studies were performed in male Sprague Dawley rats. Positron emission tomography (PET) brain studies with (-)-[(11)C]2 were performed in adult male cynomolgus macaques; 2 h dynamic data was acquired, and the regions of interest were drawn by co-registration of the PET images with the MRI.

Results: The resolved enantiomers (-)-2 and (-)-6 were very potent and selective for VAChT in vitro (K i < 5 nM for VAChT with >35-fold selectivity for VAChT vs. σ receptors); both radioligands, (-)-[(11)C]2 and (-)-[(11)C]6, demonstrated high accumulation in the VAChT-enriched striatum of rats. (-)-[(11)C]2 had a higher striatum to cerebellum ratio of 2.4-fold at 60 min; at 30 min, striatal uptake reached 0.550 ± 0.086 %ID/g. Uptake was also specific and selective; following pretreatment with (±)-2, striatal uptake of (-)-[(11)C]2 in rats at 30 min decreased by 50 %, while pretreatment with a potent sigma ligand had no significant effect on striatal uptake in rats. In addition, (-)-[(11)C]2 displayed favorable in vivo stability in rat blood and brain. PET studies of (-)-[(11)C]2 in nonhuman primates indicate that it readily crosses the blood-brain barrier (BBB) and provides clear visualization of the striatum; striatal uptake reaches the maximum at 60 min, at which time the target to nontarget ratio reached ~2-fold.

Conclusions: The radioligand (-)-[(11)C]2 has high potential to be a suitable PET radioligand for imaging VAChT in the brain of living subjects.

Fig. 1

Structures of VAChT PET tracers.

Fig. 2

Regional brain uptake of (−)-[¹¹C]2 and (−)-[¹¹C]6 in SD rat. (−)-[¹¹C]2 displayed high accumulation in the striatum (STR) and fast washout in nontarget regions such as the cerebellum (CB), thalamus (THA), cortex (CTX), and hippocampus (HIPPO); the uptake ratio in STR versus CB reached 2.4-fold at 60 min p.i. (−)-[¹¹C]6 also showed the highest uptake in the striatum; however, the ratio was only 1.6-fold at 60 min p.i.

Fig. 3

Autoradiography study of (−)-[¹¹C]2 in male SD rat: 115 MBq of (−)-[¹¹C]2 was administrated into the rat by tail i.v. injection. The animals were euthanized at 30 min under anesthesia p.i. a AChE staining, b scanned brain slice before autoradiography counting, c InstantImager autoradiography. The caudate and putamen regions were noted with CPu in all images.

Fig. 4

A representative microPET image (a) and an averaged time-activity curve (b) of (−)-[¹¹C]2 in cynomolgus monkey brain. a. microPET images (left), co-registered images (middle), and MR images (right). High uptake of (−)-[¹¹C]2 in the striatum produced clear visualization of the tracer in the putamen and caudate. b. The averaged time-activity curve from three microPET scans demonstrated a good reproducibility for (−)-[¹¹C]2.