Dopamine Transporter Imaging Agent Research Articles

Synthesis and initial evaluation of radioiodine-labelled deuterated tropane derivatives targeting dopamine transporter

The dopamine transporter (DAT) is closely related to a variety of neurological disorders including Parkinson’s disease (PD) and other neurodegenerative diseases. In vivo imaging of DAT with radio-labelled tracers has become a powerful technique in related disorders. The radioiodine-labelled tropane derivative [123I]FP-CIT ([123I]1a) is widely used in clinical single photon emission computed tomography (SPECT) imaging as a DAT imaging agent. To develop more metabolically stable DAT radioligands for accurate imaging, this work compared two novel deuterated tropane derivatives ([131I]1c-d) with non-deuterated tropane derivatives ([131I]1a-b). [131I]1a-d were obtained in high radiochemical purity (RCP) above 99 % with molar activities of 7.0–10.0 GBq/μmol. The [131I]1a and [131I]1c exhibited relatively higher affinity to DAT (Ki: 2.0–3.12 nM) than [131I]1b and [131I]1d. Biodistribution results showed that [131I]1c consistently exhibited a higher ratio of the target to non-target (striatum/cerebellum) than [131I]1a. Furthermore, metabolism studies indicated that the in vivo metabolic stability of [131I]1c was superior to that of [131I]1a. Ex vivo autoradiography showed that [131I]1c selectively localized on DAT-rich striatal regions and the specific signal could be blocked by DAT inhibitor. These results indicated that [131I]1c might be a potential probe for DAT SPECT imaging in the brain.

Development and validation of high-performance liquid chromatography method for analysis of β-CCT and its related substances

Compound 2β-carbomethoxy-3β-(4-chlorophenyl)tropane (β-CCT) is a key intermediate for the synthesis of some clinical dopamine transporter (DAT) imaging agents. Potential impurities from synthesis process of β-CCT and degradation during storage might have detrimental effect on the final imaging agents. Thus, it is necessary to guarantee the quality of β-CCT. In this study, a rapid, sensitive and accurate high-performance liquid chromatography (HPLC) method was developed and validated for the analysis of β-CCT and its related substances. The chromatographic separation was achieved on a reverse-phase phenomenex™ Gemini C18 column with an isocratic mobile phase consisted of methanol, water and TFA (30:70:0.1 v/v/v). The flow rate was 1.0 mL/min at 30 °C and samples were monitored at 220 nm. The method was validated concerning system suitability, linearity, accuracy, precision, specificity, robustness and stability. The limit of detection (LOD) and the limit of quantification (LOQ) of β-CCT were 0.5 and 1.5 μg/mL, respectively. The linearity range of β-CCT was 1.5–450 μg/mL with a good linear correlation coefficient (R 2 = 0.9999) between the peak response and concentration. Specificity investigation through forced degradation experiments displayed that β-CCT was stable in acidic, thermal and photolytic degradation conditions, but significantly unstable in alkaline and oxidative conditions. With the developed chromatographic method, possible impurity α-CCT from synthetic process and potential degradation products could be well separated from β-CCT. Good recovery and precision were manifested in the assay method. These results indicated that the present method would be suitable for not only the quality assurance of β-CCT in regular production sample assays but also the monitoring and determination of its related substances.

Optimization of the Radiosynthesis of the Dopamine Transporter Imaging Agent of 11C-β-CFT

Optimization of the Radiosynthesis of the Dopamine Transporter Imaging Agent of 11C-β-CFT

18F-FPEB PET/CT Shows mGluR5 Upregulation in Parkinson's Disease.

Dopamine and glutamate reciprocally regulate each other in some of the neurocircuits affected by Parkinson's disease (PD). The objective of this pilot study was to explore relationships between these neurotransmitter systems with positron emission tomography. The sample consisted of nine patients with PD and eight healthy volunteers (HVs). Dynamic images of the brain were acquired after the IV administration of ∼370 MBq (10 mCi) of [11 C]PE2i, a dopamine transporter (DaT) imaging agent, and ∼185 MBq (∼5 mCi) of [18 F]FPEB, a selective metabotropic glutamate receptor 5 (mGluR5) antagonist. Multiple volumes of interest were semiautomatically placed on contemporaneously acquired MRI scans. Nondisplaceable binding potentials (BPND ) were calculated with the Logan reference tissue model using cerebellar white matter as the reference region. The findings showed that average [18 F]FPEB BPND values were slightly more than 20% higher in PD than HVs in several mesocortical regions, including the bilateral putamen (P = .01), hippocampus (P = .02), and amygdala (P = .05). Average [11 C]PE2i BPND was significantly reduced by about half or more in patients with PD in the bilateral caudate (P < .001) and putamen (P < .001). mGluR5 seems upregulated in strategic dopaminergic brain regions adversely affected by PD. The findings seem to confirm that DaT tracers are better discriminatory biomarkers for diagnosing PD; however, mGluR5 tracers might deserve further exploration as potential biomarkers of response in clinical trials.

Preparation of 99mTc-TRODAT-1 with high labeling yield in boiling water bath: A new formulation

A new formulation for preparation of (99m)Tc-labeled tropane derivative, (99m)Tc-TRODAT-1, which is useful as a potential CNS dopamine transporter imaging agent, was evaluated and characterized. Preparation of (99m)Tc-TRODAT-1 was attained previously by a formulation in which vial has to be autoclaved at 121 °C for 30 min. It is highly desirable to further improve the preparation method by developing a simplified one vial formulation which will be labeled in boiling water bath (95 °C) for 15 min and a high labeling yield will be achieved. A formulation contained 10 μg of TRODAT-1, 20 μg tricine, 40 μg SnCl2 and 20mg manitol was prepared. Labeling was performed at 95 °C for 15 min and radiochemical analysis involved ITLC and HPLC methods. The stability of radioconjugate was checked in the presence of human serum at 37 °C up to 24h. (99m)Tc-TRODAT-1 was prepared with a radiochemical purity of more than 95% and specific activity of 64.3 MBq/nmol. Biodistribution studies of this new formulation in rats revealed similar regional brain distribution as compared with those obtained with the previous preparation in which brain uptake was high in striatum and striatum to cerebellum ratio was high. Requiring no autoclave facility for labeling, this new formulation will significantly improve the using feasibility of this radiopharmaceutical in clinic.

Performance of a high‐sensitivity dedicated cardiac SPECT scanner for striatal uptake quantification in the brain based on analysis of projection data

The authors have previously reported the advantages of high-sensitivity single-photon emission computed tomography (SPECT) systems for imaging structures located deep inside the brain. DaTscan (Isoflupane I-123) is a dopamine transporter (DaT) imaging agent that has shown potential for early detection of Parkinson disease (PD), as well as for monitoring progression of the disease. Realizing the full potential of DaTscan requires efficient estimation of striatal uptake from SPECT images. They have evaluated two SPECT systems, a conventional dual-head gamma camera with low-energy high-resolution collimators (conventional) and a dedicated high-sensitivity multidetector cardiac imaging system (dedicated) for imaging tasks related to PD. Cramer-Rao bounds (CRB) on precision of estimates of striatal and background activity concentrations were calculated from high-count, separate acquisitions of the compartments (right striata, left striata, background) of a striatal phantom. CRB on striatal and background activity concentration were calculated from essentially noise-free projection datasets, synthesized by scaling and summing the compartment projection datasets, for a range of total detected counts. They also calculated variances of estimates of specific-to-nonspecific binding ratios (BR) and asymmetry indices from these values using propagation of error analysis, as well as the precision of measuring changes in BR on the order of the average annual decline in early PD. Under typical clinical conditions, the conventional camera detected 2 M counts while the dedicated camera detected 12 M counts. Assuming a normal BR of 5, the standard deviation of BR estimates was 0.042 and 0.021 for the conventional and dedicated system, respectively. For an 8% decrease to BR = 4.6, the signal-to-noise ratio were 6.8 (conventional) and 13.3 (dedicated); for a 5% decrease, they were 4.2 (conventional) and 8.3 (dedicated). This implies that PD can be detected earlier with the dedicated system than with the conventional system; therefore, earlier identification of PD progression should be possible with the high-sensitivity dedicated SPECT camera.

Synthesis and evaluation of novel tropane derivatives as potential PET imaging agents for the dopamine transporter

A novel series of tropane derivatives containing a fluorinated tertiary amino or amide at the 2β position was synthesized, labeled with the positron-emitter fluorine-18 (t1/2=109.8min), and tested as potential in vivo dopamine transporter (DAT) imaging agents. The corresponding chlorinated analogs were prepared and employed as precursors for radiolabeling leading to the fluorine-18-labeled derivatives via a one-step nucleophilic aliphatic substitution reaction. In vitro binding results showed that the 2β-amino compounds 6b, 6d and 7b displayed moderately high affinities to DAT (Ki<10nM). Biodistribution studies of [18F]6b and [18F]6d showed that the brain uptakes in rats were low. This is likely due to their low lipophilicities. Further structural modifications of these tropane derivatives will be needed to improve their in vivo properties as DAT imaging agents.

Ex vivo and in vivo Evaluation of [18F]PR04.MZ in Rodents: A Selective Dopamine Transporter Imaging Agent

N-4-Fluorobut-2-yn-1-yl-2beta-carbomethoxy-3beta-phenyltropane (PR04.MZ) has been developed as dopamine transporter (DAT) ligand for molecular imaging. It contains a terminally fluorinated, conformationally constrained nitrogen substituent that is well suited for the introduction of fluorine-18. The present report describes the pharmacological characterisation of [18F]PR04.MZ. The ligand shows an IC50 value of 2 nM against human DAT, whereas the IC50 value against human serotonin transporter and human noradrenalin transporter are lower (110 nM and 22 nM, respectively). Furthermore, its ex vivo organ distribution, its binding profile in the rat brain and reversibility of binding were examined. A muPET study illuminates a fast kinetic profile and specific binding to rat DAT.

Dual-isotope single-photon emission computed tomography for dopamine and serotonin transporters in normal and parkinsonian monkey brains

Parkinson's disease (PD) affects both dopaminergic and serotonergic systems. In this study, we simultaneously evaluated dopamine and serotonin transporters in primates using dual-isotope single-photon emission computed tomography (SPECT) imaging and compared the results with traditional single-isotope imaging. Four healthy and one 6-OHDA-induced PD monkeys were used for this study. SPECT was performed over 4 h after individual or simultaneous injection of [(99m)Tc]TRODAT-1 (a dopamine transporter imaging agent) and [(123)I]ADAM (a serotonin transporter imaging agent). The results showed that the image quality and uptake ratios in different brain regions were comparable between single- and dual-isotope studies. The striatal [(99m)Tc]TRODAT-1 uptake in the PD monkey was markedly lower than that in normal monkeys. The uptake of [(123)I]ADAM in the midbrain of the PD monkey was comparable to that in the normal monkeys, but there were decreased uptakes in the thalamus and striatum of the PD monkey. Our results suggest that dual-isotope SPECT using [(99m)Tc]TRODAT-1 and [(123)I]ADAM can simultaneously evaluate changes in dopaminergic and serotonergic systems in a PD model.

A one-step automated high-radiochemical-yield synthesis of 18F-FECNT from mesylate precursor

2 β-Carbomethoxy-3 β-(4-chlorophenyl)-8-(2-[ 18F]fluoroethyl)nortropane ( 18F-FECNT) is a potential dopamine transporter imaging agent. In this article, a new mesylate precursor was prepared and a one-step automated synthesis of 18F-FECNT was developed. The mesylate precursor ( 4) was synthesized from 2 β-carbomethoxy-3 β-(4-chlorophenyl)tropane ( 1) by N-demethylation, hydroxyethylation followed by mesylation at a total yield of 47%. [ 18F]fluorination was performed by heating 4 mg mesylate precursor and K[ 18F] in 1 ml acetonitrile at 90 °C for 20 min. The crude 18F-FECNT was obtained with a radiolabeling yield of 48%. After purification by preparative high performance liquid chromatography (HPLC), the final 18F-FECNT product was obtained with a radiochemical purity of 98.4% and a decay corrected radiochemical yield of 33±9% (and the uncorrected radiochemical yield was 19±5%, n=5). The duration of the total procedure was 80–90 min.

Clinical acceptance of a molecular imaging agent: a long march with [99mTc]TRODAT

In the past 10 years, significant progress has been made in using a technetium-99m dopamine transporter imaging agent, [99mTc]TRODAT, for routine clinical studies. Developing a molecular imaging agent from bench to the bedside is more than a simple scientific venture. Currently, Taiwan is the only place where [99mTc]TRODAT is approved for routine clinical use in the diagnosis of Parkinson's disease. The trials and tribulations of developing [99mTc]TRODAT for routine clinical use in Taiwan provide an interesting case study in how to (critics may say, how not to) develop a molecular imaging agent.

Diastereoselective synthesis of polyfunctionalized piperidines as precursors of dopamine transporter imaging agents

The design and synthesis of a new kind of precursors of radiolabelled tracer agents for dopamine transporters imaging is described. The concept is based on the possibility of introducing the radioelement far from the sites of recognition and on the fact that only the labelled marker could cross the blood brain barrier to reach the dopamine transporters. For this purpose, a polyfunctionalized piperidine was synthesized bearing two aldehyde functions in the configuration allowing the formation of a tropane analog after the introduction of complexing moieties and complexation of a radioactive metal.

Biological evaluation of a technetium-99m-labeled integrated tropane-BAT and its piperidine congener as potential dopamine transporter imaging agents

Introduction Recently, we have reported modification of 99mTc-TRODAT-1 by integrating the N2S2 metal chelating unit and the tropane skeleton. Results of a preliminary biodistribution study in rats were promising with respect to brain uptake. The present report deals with the further biological characterization of the 99mTc-labelled integrated TRODAT derivatives ( 99mTc-TropaBAT and 99mTc-norchloro-TropaBAT) and with the synthesis and biological evaluation of a novel 99mTc-labelled piperidine-based derivative ( 99mTc-PipBAT). Methods Biodistribution of all radiolabelled complexes was studied in normal mice. A more detailed ex vivo intracerebral distribution study of the two 99mTc-TropaBAT complexes was additionally performed in normal rats. Autoradiography of brain sections of normal mice (with or without pretreatment with FP-β-CIT or haloperidol) and rats was performed. Affinity for the dopamine transporter (DAT) was also assessed in vitro in the presence or absence of cocaine. Results Both 99mTc-TropaBAT complexes show a slightly higher brain uptake than 99mTc-TRODAT-1, but the striatum/cerebellum activity ratio is less favourable. Nevertheless, significant striatal uptake was detected after ex vivo autoradiography, but this uptake was also observed after pretreatment with FP-β-CIT. Unexpectedly, no striatal uptake was detected after in vitro incubation of mouse brain sections with the tracer agents. For 99mTc-PipBAT, neither brain uptake nor in vitro striatal uptake was found. Conclusion Both 99mTc-TropaBAT complexes exhibit similar diffusion into brain as 99mTc-TRODAT-1, and ex vivo autoradiography shows significant striatal uptake. However, the inferior striatum/cerebellum activity ratio, the striatal uptake in mice pretreated with FP-β-CIT or haloperidol, and the lack of striatal uptake during in vitro incubation prove that the DAT is not targeted. Brain uptake disappears when the tropane skeleton is replaced by a piperidine ring, and also in this case no striatal uptake is found in vitro.

Greater availability of brain dopamine transporters in major depression shown by [99m Tc]TRODAT-1 SPECT imaging.

Studies of laboratory animals have shown that administration of antidepressants of all pharmacological classes produces changes in dopamine transporter binding affinity. These observations suggest that dopamine transporter function may play a critical role in the pathophysiology of depression. The present study was an examination of the availability of brain dopamine transporter sites in patients with major depression and in healthy comparison subjects. Single photon emission computed tomographic (SPECT) brain scans were acquired for 15 drug-free depressed patients and 46 age- and gender-matched healthy comparison subjects by using [(99m)Tc]TRODAT-1, a selective dopamine transporter imaging agent. Specific regions of interest in the basal ganglia and supratentorial areas of the brain were examined. Specific uptake values of dopamine transporter [(99m)Tc]TRODAT-1 binding affinity were calculated from the SPECT scan data, and the values for the patients and healthy subjects were compared. The specific uptake values of [(99m)Tc]TRODAT-1 binding were significantly higher in the right anterior putamen (23%), right posterior putamen (36%), left posterior putamen (18%), and left caudate nucleus (12%) of the patients than in the comparison subjects. These differences persisted when the data were further analyzed according to gender and age. Dopamine transporter affinity may be higher than normal in the basal ganglia of depressed patients. These findings suggest that dopamine function may be altered in depression and may also be a mechanism of antidepressant activity.

Preparation of [99mTc]TRODAT‐1 involving a simple purification method

AbstractDirect preparation of the novel dopamine transporter imaging agent [99mTc]TRODAT‐1 results in uncertainty concerning the labelling efficiency. Without purification of the tracer, the statistically inefficient labelling yield becomes apparent in an inadequate radiochemical purity. The aim of the present work was to improve the radiochemical purity of this tracer by including a facile, on‐the‐spot purification step in the labelling process.Kits containing 100 μg free ligand resulted in an average labelling efficiency of 91.6% (S.D. = 5.15%, n = 32), while an average of 73.0% (S.D.= 6.76%, n = 22) was obtained for the 50 μg ligand series. Introduction of solid‐phase extraction into the preparation process allowed the quantitative removal of hydrophilic species from both series of reaction mixtures. An average radiochemical purity of 98% was achieved for the purified tracer. Copyright © 2003 John Wiley &amp; Sons, Ltd.

A second-generation 99m technetium single photon emission computed tomography agent that provides in vivo images of the dopamine transporter in primate brain.

The dopamine transporter (DAT), located presynaptically on dopamine neurons, provides a marker for Parkinson's disease (Pd) and attention deficit hyperactivity disorder (ADHD). In ADHD, DAT density levels are elevated, while in Pd these levels are depleted. The depletion of DAT levels also corresponds with the loss of dopamine. We now describe the design, synthesis, biology, and SPECT imaging in nonhuman primates of second-generation (99m)technetium-based tropane ligands that bind potently and selectively to the DAT. We demonstrate that improved selectivity and biological stability allows sufficient agent to enter the brain and label the DAT in vivo to provide a quantitative measure of DAT density in nonhuman primates. We introduce FLUORATEC (N-[(2-((3'-N'-propyl-(1"R)-3"alpha-(4-fluorophenyl)tropane-2"beta-1-propanoyl)(2-mercaptoethyl)amino)acetyl)-2-aminoethanethiolato]technetium(V) oxide), a DAT imaging agent that has emerged from these studies and is now in phase 1 clinical trials in the U.S.

Biodistribution study of [ 99mTc] TRODAT-1 alone or combined with other dopaminergic drugs in mice with macroautoradiography

A 99mTc labeled tropane derivative, [ 99mTc] TRODAT-1 (2 β-(( N, N′-bis(2-mercaptoethyl) ethylene diamino)methyl), 3 β-(4-chlorophenyl) tropane), is a potential dopamine transporter (DAT) imaging agent for the central nervous system. To better understand the binding localization of [ 99mTc] TRODAT-1 both in the brain and the body, whole-body macroautoradiography (WBAR) was used in this study. The effect of DAT competing drugs, such as levadopa ( l-DOPA), N-methyl-2 β-carbomethoxy-3 β-(4fluorophenyl)tropane (CFT, WIN 35,428) and methylphenidate, on the biodistribution of [ 99mTc] TRODAT-1 were also included in this study. Doses of 150 MBq [ 99mTc] TRODAT-1 were injected into normal male ICR mice through the caudal veins. For comparison, mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), l-DOPA, methylphenidate and CFT, respectively, were also investigated under the similar protocols. One and a half hours after [ 99mTc] TRODAT-1 injection, the mice were sacrificed. Whole-body autoradiography was performed immediately after sacrifice. Both frontal and sagittal sections showed that the liver and mucosa of stomach had the highest uptake of [ 99mTc] TRODAT-1. Other binding sites included the periphery of the spinal cord and the epithelium of the intestine. In the brain, autoradiographic imaging obtained from frontal sections showed symmetrical uptakes of [ 99mTc] TRODAT-1 in bilateral striata. Remaining binding sites include olfactory bulbs, thyroid gland, and salivary gland. The autoradiographic imaging obtained from sagittal sections showed a similar biodistribution. Mice treated with MPTP or l-DOPA showed no significant difference in the uptake of [ 99mTc] TRODAT-1 in bilateral striata, as compared to those of the control. In CFT or methylphenidate-treated mice, DAT binding sites were almost completely inhibited. These data showed that [ 99mTc] TRODAT-1 has potential clinical use for neurological investigation, such as Parkinson's and similar diseases.

Development of Tc-99m labeled tropanes: TRODAT-1, as a dopamine transporter imaging agent

Advances in the technetium chemistry have significantly enhanced the development of Tc-99m radiopharmaceuticals for clinical use. Efforts in designing novel Tc-99m labeled receptor or site-specific imaging agents using [Tc vO] +3N 2S 2 core have produced useful imaging agents for single photo emission computed tomography (SPECT). The success in developing Tc-99m TRODAT-1 for CNS dopamine transporters (DAT) serves as the first example of Tc-99m site-specific imaging of human brain. New innovative Tc-99m labeled site-specific agents are particularly suitable to develop simple and useful routine diagnostic procedures.

Synthesis and biological evaluation of two novel DAT-binding technetium complexes containing a piperidine based analogue of cocaine

Two new technetium complexes containing a piperidine template have been synthesized and evaluated as possible leads for the development of dopamine transporter (DAT) imaging agents. Binding data for the corresponding rhenium complexes containing either a monoaminomonoamide (MAMA') or a diaminodithiol (DADT) chelating unit exhibited significant affinity for the DAT. Initial biodistribution studies in rats revealed only a low brain uptake.

Development of dopamine transporter imaging agents for the diagnosis of Parkinson's disease

Development of dopamine transporter imaging agents for the diagnosis of Parkinson's disease