A novel carbon-11 PET radiotracer for imaging VAChT in the brain

Objectives Vesicular acetylcholine transporter (VAChT) plays an important role in mediating cholinergic transmission, and it is considered a specific biomarker for neurodegenerative diseases. Compound (-)-TZ6-59 showed high in vitro binding potency and selectivity towards VAChT (K_i-VAChT = 0.78 nM, K_i-σ1 = 990 nM, and K_{i- σ2} >10,000 nM). Here, we report the radiosynthesis and in vivo evaluation of (-)-[¹¹C]TZ6-59 to image VAChT.

Methods The synthesis of (-)-[¹¹C]TZ6-59 was accomplished by alkylating the Boc protected aniline precursor with [¹¹C]CH₃I, followed by the removal of the protective group. Biodistribution studies were performed in male SD rats (300-360 g). Rats were injected with ~300-400 µCi/150 µL of (-)-[¹¹C]TZ6-59 and euthanized at 5, 30 and 60 min post injection; the uptake (%ID/g) in each organ of interest was calculated. Blocking study was performed in rats pretreated with cold TZ4-3-76, a potent VAChT ligand at 2 mg/kg. Dynamic microPET imaging studies were performed in cynomolgus monkeys.

Results (-)-[¹¹C]TZ6-59 was synthesized in high chemical (>95%) and radiochemical (>99%) purities, and high specific activity (>0.5 Ci/µmol, EOB) with a yield of 20-25%. Biodistribution studies revealed that (-)-[¹¹C]TZ6-59 is able to cross the rat BBB and specifically accumulate in the VAChT-enriched striatal area; at 60 min, the uptake ratio of striatum vs. cerebellum reached ~5.5-fold. Pretreated with unlabeled TZ4-3-76, striatum uptake of labeled (-)-[¹¹C]TZ6-59 was reduced by ~48%. MicroPET imaging of monkey brains demonstrated that the highest uptake occurs in the striatal area with sufficient contrast ratios vs. reference regions.

Conclusions (-)-[¹¹C]TZ6-59 is a promising PET tracer for imaging VAChT in the brain. Further evaluations will be performed prior to seeking IND and RDRC approval for translational clinical validation in human beings.

Research Support 1R21NS061025-01A2 & 1R01NS075527-01.