Characterization of optically resolved 9-fluoropropyl-dihydrotetrabenazine as a potential PET imaging agent targeting vesicular monoamine transporters

Abstract

Labeling derivatives of dihydrotetrabenazine (DTBZ) with F-18 (T(1/2)=110 min) instead of C-11 (T(1/2)=20 min) would improve their utility and availability for imaging vesicular monoamine transporters (VMAT2) in clinical settings. The successful synthesis, reported previously, of two novel 9-fluoroalkyl(+/-)-DTBZ ligands prompted us to study the optically resolved active ligand 9-fluoropropyl-(+)-DTBZ (FP-(+)-DTBZ), which may have more promising characteristics. The inhibition constant (K(i)) estimated for FP-(+)-DTBZ (using [(3)H](+/-)-DTBZ as the labeled ligand in rat striatal homogenates) showed a lower value as compared to the racemic FP-(+/-)-DTBZ (0.10+/-0.01 vs 0.19+/-0.04 nM). The inactive isomer, FP-(-)-DTBZ, displayed a much lower binding affinity with a K(i) value >3000 nM. Biodistribution studies in mice after an iv injection of [(18)F]FP-(+)-DTBZ exhibited a ratio of striatum (ST, target) to cerebellum (CB, background) of 4.51 at 30 min postinjection, which is a higher value than previously obtained with the racemic ligand [(18)F]FP-(+/-)-DTBZ (ST/CB=2.95). Brain extraction at 30 min after the tracer injection in mice showed that >95% of the radioactivity corresponded to the parent, nonmetabolized, compound remaining in the ST, suggesting that the tracer has an excellent in vivo stability. Furthermore, localization of the tracer in the brain examined with ex vivo autoradiography displayed a typical distribution pattern consistent with VMAT2 sites. The highest labeling was observed in monoaminergic neuron regions (caudate putamen, olfactory tubercle, nucleus accumbens, substantia nigra, dorsal raphe and locus coerules). We also tested the selective labeling of this tracer at the dopamine neurons in unilateral-lesioned mice (treated with 6-hydroxydopamine). When [(18)F]FP-(+)-DTBZ and [(125)I]IPT ((N-(3'-iodopropen-2'-yl)-2-beta-carbomethoxy-3-beta-(4-chlorophenyl)tropane, a selective marker for dopamine transporters (DATs) in dopaminergic neurons) were simultaneously injected into lesioned mice, we observed an excellent correlation (r=0.95) for these tracers. From these findings, we conclude that [(18)F]FP-(+)-DTBZ is a sensitive and selective tracer for VMAT2 binding sites and it may be useful for in vivo evaluation of diseases relating to changes of monoamine neuronal integrity.

Fig. 1

Separation of four enantiomers of FP-(±)-DTBZ on a chiral AD column with hexanes/isopropranol (9/1) as the eluting solvent (flow rate of 1 mL/min). The ratio of two major enantiomers (peak 3 & 4) to the minor ones (peak 1 & 2) is 5:1. The optical resolution of FP-(+)-DTBZ reached 98% (shown as peak 3) and the inactive isomer FP-(−)-DTBZ showed a major peak 4 (90%) with a small contamination of peak 1 (10%).

Fig. 2

Ex vivo autoradiography of a normal mouse brain revealed the anatomical localization of VMAT sites labeled by [¹⁸F]FP-(+)-DTBZ. In this experiment 500 μCi [¹⁸F]FP-(+)-DTBZ was injected into a normal ICR mouse and sacrificed at 30 minutes post-injection. The high-density sites clearly reflected the regional distribution of monoaminergic neurons in the brain: CPu, caudate putamen; OT, olfactory tubercle; Ac, nucleus accumbens; Hy, hypothalamic nucleus; SN, substantia nigra; DR, dorsal raphe; MR, median raphe; LC, locus coeruleus.

Fig. 3

Ex vivo autoradiography (30 minutes postinjection) of [¹⁸F]FP-(+)-DTBZ clearly differentiate the lesioned (L, indicated by an arrow) from the un-lesioned side (N) in the brains of 6-OH-DA unilaterally lesioned mice. 300 μCi [¹⁸F]FP-(+)-DTBZ was injected and animals were sacrificed at 30 minutes post-injection.