Positron emission tomography imaging of (2R,3R)-5-[18F]fluoroethoxybenzovesamicol in rat and monkey brain: a radioligand for the vesicular acetylcholine transporter

doi:10.1016/j.nucmedbio.2009.02.007

Nuclear Medicine and Biology

Volume 36, Issue 5, July 2009, Pages 489-493

https://doi.org/10.1016/j.nucmedbio.2009.02.007 Get rights and content

Abstract

Introduction

The regional brain distribution of (2R,3R)-5-[¹⁸F]fluoroethoxy-benzovesamicol ((−)-[¹⁸F]FEOBV), a radioligand for the vesicular acetylcholine transporter (VAChT), was examined in vivo in mice, rats and rhesus monkeys.

Methods

Regional brain distributions of (−)-[¹⁸F]FEOBV in mice were determined using ex vivo dissection. MicroPET imaging was used to determine the regional brain pharmacokinetics of the radioligand in rat and rhesus monkey brains.

Results

In all three species, clear heterogeneous regional brain distributions were obtained, with the rank order of brain tissues (striatum>thalamus>cortex>cerebellum) consistent with the distribution of cholinergic nerve terminals containing the VAChT.

Conclusions

(−)-[¹⁸F]FEOBV remains a viable candidate for further development as an in vivo imaging agent for positron emission tomography (PET) studies of the VAChT in the human brain.

Introduction

The vesicular acetylcholine transporter (VAChT) is a protein uniquely located in presynaptic vesicles of cholinergic neurons and is responsible for the transport of the neurotransmitter acetylcholine into the storage vesicles [1]. As a specific biochemical function of the cholinergic neuron, it has been a potential target for development of in vivo radioligands [2], [3] for single-photon emission computed tomography (SPECT) or positron emission tomography (PET) imaging of the potential losses of cholinergic neurons in degenerative neurological diseases. Successful imaging of the VAChT in the human brain, including losses in disease including Alzheimer's, has been reported using a SPECT radioiodinated ligand 5-[¹²³I]iodobenzovesamicol ((−)-5-IBVM) [4], [5], [6], [7]. For potential use in PET, radioligands labeled with the positron-emitting radionuclides carbon-11 and fluorine-18 were reported as early as 1990 [8], [9] and there has been continued interest in further evaluation of both SPECT and PET radioligands for imaging of the VAChT binding site in the human brain [10], [11].

Most recently, Giboureau et al. [10] reported the in vivo evaluation in rats and primate brain of two potential PET radioligands, (2R,3R)-5-[¹⁸F]fluoroethoxybenzovesamicol ((−)-[¹⁸F]FEOBV, first reported in 1993 by Mulholland et al. [12]) and (2R,3R)-5-[¹⁸F]fluoropropoxybenzovesamicol (((−)-[¹⁸F]FPOBV). Using ex vivo dissection techniques for both radioligands, and in vivo PET imaging of [¹⁸F]FPOBV in monkey brain, these investigators concluded that both (−)-[¹⁸F]FEOBV and (−)-[¹⁸F]FPOBV were unsuitable as in vivo radioligands for the imaging for the VAChT in the mammalian brain. Their report directly contradicts our earlier and extensive rodent studies of (−)-[¹⁸F]FEOBV [13], which had consistently supported that it would be a viable candidate for further development as a PET imaging agent for human studies of the VAChT. As we have continued to pursue the development of (−)-[¹⁸F]FEOBV with the intention of eventual human applications, we felt it was necessary to revisit our in vivo studies of (−)-[¹⁸F]FEOBV to verify our earlier conclusions. We report here in vivo studies of (−)-[¹⁸F]FEOBV in mice, rats and nonhuman primates, including dynamic microPET imaging studies, which clearly demonstrate that (−)-[¹⁸F]FEOBV is indeed a successful imaging agent for the VAChT and is worthy of further development and implementation.

Section snippets

Synthesis of (−)-[¹⁸F]FEOBV

Syntheses of (−)-[¹⁸F]FEOBV and unlabeled (−)-FEOBV were done by slight modifications of the literature procedure [12]. Reaction of the tosyloxy precursor, (2R,3R)-5-(2-tosyloxyethoxy)benzovesamicol, with no-carrier-added [¹⁸F]fluoride ion in dimethylsulfoxide followed by purification using preparative high-pressure liquid chromatography provided the desired (−)-[¹⁸F]FEOBV with high radiochemical purity (>95%) and specific activities of 3500–13,300 Ci/mmol at the end of synthesis. For the

Regional mouse brain biodistribution and blocking studies

At 3 h after bolus intravenous injection of (−)-[¹⁸F]FEOBV, there was a clear heterogeneous distribution of radioactivity in the mouse brain (striatum>cortex>hippocampus>thalamus>cerebellum) consistent with the distribution of cholinergic synapses and VAChT. Administration of doses of cold (−)-FEOBV reduced the retention of radioactivity in regions of high VAChT concentrations (striatum and cortex) at the 5 and 10 μg/kg doses (Table 1).

Rat microPET imaging

Following bolus intravenous injection, (−)-[¹⁸F]FEOBV

Discussion

The synthesis of (−)-[¹⁸ F]FEOBV, in vitro characterization as a high affinity radioligand for the VAChT, and in vivo and ex vivo rodent studies had been reported from our laboratories more than a decade ago [12], [13]. From those studies, and others completed but not reported, we had concluded that (−)-[¹⁸F]FEOBV was a suitable candidate for further development into an in vivo PET imaging agent for the VAChT in the human brain, with potential applications in the study of cholinergic terminal

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Cited by (48)

Development and preclinical evaluation of [18F]FBVM as a new potent PET tracer for vesicular acetylcholine transporter
2022, European Journal of Medicinal Chemistry
Age-related neurodegenerative diseases have in common the occurrence of cognitive impairment, a highly incapacitating process that involves the cholinergic neurotransmission system. The vesicular acetylcholine transporter (VAChT) positron emission tomography (PET) tracer [¹⁸F]fluoroethoxybenzovesamicol ((−)-[¹⁸F]FEOBV) has recently demonstrated its high value to detect alterations of the cholinergic system in Alzheimer's disease, Parkinson's disease and dementia with Lewy body. We present here the development of the new vesamicol derivative tracer (−)-(R,R)-5-[¹⁸F]fluorobenzovesamicol ((−)[¹⁸F]FBVM) that we compared to (−)[¹⁸F]FEOBV in the same experimental conditions. We show that: i) in vitro affinity for the VAChT was 50-fold higher for (−)FBVM (Ki = 0.9 ± 0.3 nM) than for (−)FEOBV (Ki = 61 ± 2.8 nM); ii) in vivo in rats, a higher signal-to-noise specific brain uptake and a lower binding to plasma proteins and peripheral defluorination were obtained for (−)[¹⁸F]FBVM compared to (−)[¹⁸F]FEOBV. Our findings demonstrate that (−)[¹⁸F]FBVM is a highly promising PET imaging tracer which could be sufficiently sensitive to detect in humans the cholinergic denervation that occurs in brain areas having a low density of VAChT such as the cortex and hippocampus.
In vitro characterization of [3H]VAT in cells, animal and human brain tissues for vesicular acetylcholine transporter
2021, European Journal of Pharmacology
Citation Excerpt :
Deficiency of cholinergic neurons and synapses contributes to progressive cognitive dysfunction in AD, PD, Huntington disease (HD), and other neurodegenerative disorders (Bohnen and Albin, 2011; Bohnen et al., 2003, 2018; Braak et al., 2004; Ferreira-Vieira et al., 2016; Pirker et al., 2003; Ransome and Hannan, 2012; Suzuki et al., 2002). VAChT, which is responsible for translocating acetylcholine from the cytoplasm into vesicles, provides a reliable biomarker for cholinergic neurons under normal (Efange, 2000; Haense et al., 2012; Kilbourn et al., 2009; Mulholland et al., 1998; Petrou et al., 2014; Prado et al., 2013) and neurodegenerative conditions (Aghourian et al., 2017; Bohnen and Albin, 2011; Bohnen et al., 2009). Cholinergic striatal deficits with decreased VAChT expressions and ChAT activities were observed in progressive supranuclear palsy (PSP) (Javoy-Agid, 1994; Ruberg et al., 1985; Suzuki et al., 2002).
Vesicular acetylcholine transporter plays a crucial role in the cholinergic system, and its alterations is implicated in several neurodegenerative disorders. We recently developed a PET imaging tracer [¹⁸F]VAT to target VAChT in vivo with high affinity and selectivity. Here we report in vitro characterization of [³H]VAT, a tritiated counterpart of [¹⁸F]VAT. Using human VAChT-rich cell membrane extracts, a saturated binding curve was obtained for [³H]VAT with K_d = 6.5 nM and B_max = 22.89 pmol/mg protein. In the [³H]VAT competition-binding assay with a panel of CNS ligands, binding inhibition of [³H]VAT was observed using VAChT ligands, the K_i values ranged from 5.41 to 33.3 nM. No inhibition was detected using a panel of other CNS ligands. In vitro [³H]VAT autoradiography of rat brain sections showed strong signals in the striatum, moderate to high signals in vermis, thalamus, cortex, and hippocampus, and weak signals in cerebellum. Strong [³H]VAT ARG signals were also observed from striatal sections of normal nonhuman primates and human brains. Competitive ARG study with human striatal sections demonstrated strong ARG signals of [³H]VAT in caudate and putamen were blocked significantly by either VAChT ligand TZ659 or (−)-vesamicol, but not by the σ₁ receptor ligand Yun-122. ARG study also indicated that signal in the striatal sections from PSP human brains was lower than normal human brains. These data provide solid evidence supporting [¹⁸F]VAT as a suitable PET radiotracer for quantitative assessment of VAChT levels in vivo.
First-in-human imaging and kinetic analysis of vesicular acetylcholine transporter density in the heart using [18F]FEOBV PET
2021, Journal of Nuclear Cardiology
In contrast to cardiac sympathetic activity which can be assessed with established PET tracers, there are currently no suitable radioligands to measure cardiac parasympathetic (cholinergic) activity. A radioligand able to measure cardiac cholinergic activity would be an invaluable clinical and research tool since cholinergic dysfunction has been associated with a wide array of pathologies (e.g., chronic heart failure, myocardial infarction, arrythmias). [¹⁸F]Fluoroethoxybenzovesamicol (FEOBV) is a cholinergic radiotracer that has been extensively validated in the brain. Whether FEOBV PET can be used to assess cholinergic activity in the heart is not known. Hence, this study aimed to evaluate the properties of FEOBV for cardiac PET imaging and cholinergic activity mapping. PET data were collected for 40 minutes after injection of 230 ± 50 MBq of FEOBV in four healthy participants (1 female; Age: 37 ± 10; BMI: 25 ± 2). Dynamic LV time activity curves were fitted with Logan graphical, 1-tissue compartment, and 2-tissue compartment models, yielding similar distribution volume estimates for each participant. Our initial data show that FEOBV PET has favorable tracer kinetics for quantification of cholinergic activity and is a promising new method for assessing parasympathetic function in the heart.
Molecular Imaging: A New Frontier in Neurotoxicology
2017, Comprehensive Toxicology: Third Edition
Due to its complexity and protracted developmental time course, the nervous system is particularly sensitive to toxic insults. Even relatively minor insults that occur during development may result in severe adverse cognitive and behavioral effects that persist throughout life. New approaches in molecular imaging provide noninvasive biomarkers for visualizing neurotoxicity with high specificity and sensitivity in comparison to previous modalities. This chapter summarizes recent microPET findings using a variety of radiotracers on the developmental neurotoxicity induced by general anesthetics in both the rodent and nonhuman primate brain. MicroPET imaging is a very powerful too with which to repeatedly obtain from the same subjects non-invasive measurements of multiple biological processes in various organs.
Application of advanced preclinical models and methods in anesthetic neurotoxicity research
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Recently, there has been increasing concern regarding the potential of anesthetics to disturb the long-term function of the central nervous system (CNS). The field of anesthesia-related toxicology, therefore, has engaged multiple scientific disciplines and utilized a variety of pre-clinical research models in an attempt to identify the basic characteristics of the anesthetic agents that may produce acute and/or chronic adverse effects on the CNS. This review discusses how the application of advanced research approaches and models, such as the nonhuman primate, neural stem cell-derived organotypic slice cultures and/or organs-on-chips systems, can serve as translational models of infantile anesthetic exposure. Utilization of these models may expeditiously decrease the uncertainty in the risk posed to children by postnatal anesthetic exposure.
Synthesis and in vitro evaluation of 5-substituted benzovesamicol analogs containing N-substituted amides as potential positron emission tomography tracers for the vesicular acetylcholine transporter
2017, Bioorganic and Medicinal Chemistry
Citation Excerpt :
The tracer is a molecule that has been radiolabeled with a positron-emitting radionuclide, such as 11C and 18F. Several attempts at synthesizing a PET tracer for VAChT have been made but only a limited number have progressed to in vivo preclinical evaluations.17–24 All VAChT ligands synthesized thus far are structurally related to (±)-trans-2-(4-phenylpiperidino)-cyclohexanol (vesamicol, Fig. 1).
Herein, new ligands for the vesicular acetylcholine transporter (VAChT), based on a benzovesamicol scaffold, are presented. VAChT is acknowledged as a marker for cholinergic neurons and a positron emission tomography tracer for VAChT could serve as a tool for quantitative analysis of cholinergic neuronal density. With an easily accessible triflate precursor, aminocarbonylations were utilized to evaluate the chemical space around the C5 position on the tetrahydronaphthol ring. Synthesized ligands were evaluated for their affinity and selectivity for VAChT. Small, preferably aromatic, N-substituents proved to be more potent than larger substituents. Of the fifteen compounds synthesized, benzyl derivatives (±)-7i and (±)-7l had the highest affinities for VAChT. Compound (±)-7i was chosen to investigate the importance of stereochemistry for binding to VAChT and selectivity toward the σ₁ and σ₂ receptors. Enantiomeric resolution gave (+)-7i and (−)-7i, and the eutomer showed seven times better affinity. Although racemate (±)-7i was initially promising, the affinity of (−)-7i for VAChT was not better than 56.7 nM which precludes further preclinical evaluation. However, the nanomolar binding together with the ready synthesis of [¹¹C]-(±)-7i shows that (−)-7i can serve as a scaffold for future optimizations to provide improved ¹¹C-labelled VAChT PET tracers.

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^☆: This work was supported by grants from the National Institutes of Health (MH66506) and the Office of Science, US Department of Energy (DE-FG02-87ER60561).

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Nuclear Medicine and Biology

Abstract

Introduction

Methods

Results

Conclusions

Introduction

Section snippets

Synthesis of (−)-[¹⁸F]FEOBV

Regional mouse brain biodistribution and blocking studies

Rat microPET imaging

Discussion

References (14)

Acetylcholine transport, storage and release

Int Rev Neurobiol

Imaging of monoaminergic and cholinergic vesicular transporters in the brain

In vivo SPECT imaging of vesicular acetylcholine transporter using [(¹²³I)]-IBVM in early Alzheimer's disease

NeuroImage

Mouse brain distribution of a carbon-11 labeled vesamicol derivative: presynaptic marker of cholinergic neurons

Life Sciences

Positron emission tomographic studies of central cholinergic nerve terminals

Neurosci Lett

In vivo imaging of the vesicular acetylcholine transporter and the vesicular monoamine transporter

FASEB J

In vivo mapping of cholinergic terminals in normal aging, Alzheimer's disease, and Parkinson's disease

Ann Neurol

Cited by (48)

Development and preclinical evaluation of [<sup>18</sup>F]FBVM as a new potent PET tracer for vesicular acetylcholine transporter

In vitro characterization of [<sup>3</sup>H]VAT in cells, animal and human brain tissues for vesicular acetylcholine transporter

First-in-human imaging and kinetic analysis of vesicular acetylcholine transporter density in the heart using [<sup>18</sup>F]FEOBV PET

Molecular Imaging: A New Frontier in Neurotoxicology

Application of advanced preclinical models and methods in anesthetic neurotoxicity research

Synthesis and in vitro evaluation of 5-substituted benzovesamicol analogs containing N-substituted amides as potential positron emission tomography tracers for the vesicular acetylcholine transporter

Positron emission tomography imaging of (2R,3R)-5-[18F]fluoroethoxybenzovesamicol in rat and monkey brain: a radioligand for the vesicular acetylcholine transporter☆

Abstract

Introduction

Methods

Results

Conclusions

Introduction

Section snippets

Synthesis of (−)-[18F]FEOBV

Regional mouse brain biodistribution and blocking studies

Rat microPET imaging

Discussion

Int Rev Neurobiol

NeuroImage

Life Sciences

Neurosci Lett

In vivo imaging of the vesicular acetylcholine transporter and the vesicular monoamine transporter

FASEB J

In vivo mapping of cholinergic terminals in normal aging, Alzheimer's disease, and Parkinson's disease

Ann Neurol

Positron emission tomography imaging of (2R,3R)-5-[¹⁸F]fluoroethoxybenzovesamicol in rat and monkey brain: a radioligand for the vesicular acetylcholine transporter☆

Synthesis of (−)-[¹⁸F]FEOBV