Serotonin Transporter Imaging Research Articles

A novel serotonin transporter ligand: (5-Iodo-2-(2-dimethylaminomethylphenoxy)-benzyl alcohol

The serotonin transporters (SERT) are the primary binding sites for selective serotonin reuptake inhibitors, commonly used antidepressants such as fluoxetine, sertraline, and paroxetine. Imaging of SERT with positron emission tomography and single photon emission computed tomography in humans would provide a useful tool for understanding how alterations of this system are related to depressive illnesses and other psychiatric disorders. In this article the synthesis and characterization of [ 125I]ODAM [(5-iodo-2-(2-dimethylaminomethylphenoxy)-benzyl alcohol, 9)] as an imaging agent in the evaluation of central nervous system SERT are reported. A new reaction scheme was developed for the preparation of compound 9, ODAM, and the corresponding tri-n-butyltin derivative 10. Upon reacting 10 with hydrogen peroxide and sodium[ 125I]iodide, the radiolabeled [ 125I]9 was obtained in good yield (94% yield, radiochemical purity >95%). In an initial binding study using cortical membrane homogenates of rat brain, ODAM displayed a good binding affinity with a value of K i = 2.8 ± 0.88 nM. Using LLC-PK 1 cells specifically expressing the individual transporter (i.e. dopamine [DAT], norepinephrine [NET], and SERT, respectively), ODAM showed a strong inhibition on SERT (K i = 0.12 ± 0.02 nM). Inhibition constants for the other two transporters were lower (K i = 3.9 ± 0.7 μM and 20.0 ± 1.9 nM for DAT and NET, respectively). Initial biodistribution study in rats after an intravenous (IV) injection of [ 125I]ODAM showed a rapid brain uptake and washout (2.03, 1.49, 0.79, 0.27, and 0.07% dose/organ at 2, 30, 60, 120, and 240 min, respectively). The hypothalamus region where the serotonin neurons are located exhibited a high specific uptake. Ratios of hypothalamus-cerebellum/cerebellum based on percent dose per gram of these two regions showed values of 0.35, 0.86, 0.86, 0.63, and 0.34 at 2, 30, 60, 120, and 240 min, post-IV injection, respectively. The specific uptake in hypothalamus can be effectively blocked by pretreatment of known SERT ligands. The results suggest that this novel ligand displays desirable in vitro and in vivo properties as a potential SERT imaging agent.

Single-photon emission tomography imaging of serotonin transporters in the nonhuman primate brain with [(123)I]ODAM.

We have described previously a selective serotonin transporter (SERT) radioligand, [(123)I]IDAM. We now report a similarly potent, but more stable IDAM derivative, 5-iodo-2-[2-[(dimethylamino)methyl]phenoxy]benzyl alcohol ([(123)I]ODAM). The imaging characteristics of this radioligand were studied and compared against [(123)I]IDAM. Dynamic sequences of single-photon emission tomography (SPET) scans were obtained on three female baboons after injection of 375 MBq of [(123)I]ODAM. Displacing doses (1 mg/kg) of the selective SERT ligand (+)McN5652 were administered 120 min after injection of [(123)I]ODAM. Total integrated brain uptake of [(123)I]ODAM was about 30% higher than [(123)I]IDAM. After 60-120 min, the regional distribution of tracer within the brain reflected the characteristic distribution of SERT. Peak specific binding in the midbrain occurred 120 min after injection, with an equilibrium midbrain to cerebellar ratio of 1. 50+/-0.08, which was slightly lower than the value for [(123)I]IDAM (1.80+/- 0.13). Both the binding kinetics and the metabolism of [(123)I]ODAM were slower than those of [(123)I]IDAM. Following injection of a competing SERT ligand, (+)McN5652, the tracer exhibited washout from areas with high concentrations of SERT, with a dissociation kinetic rate constant k(off)=0.0085+/-0.0028 min(-1) in the midbrain. Similar studies using nisoxetine and methylphenidate showed no displacement, consistent with its low binding affinity to norepinephrine and dopamine transporters, respectively. These results suggest that [(123)I]ODAM is suitable for selective SPET imaging of SERT in the primate brain, with higher uptake and slower kinetics and metabolism than [(123)I]IDAM, but also a slightly lower selectivity for SERT.

Single-photon emission tomography imaging of serotonin transporters in the non-human primate brain with the selective radioligand [(123)I]IDAM.

A new radioligand, 5-iodo-2-[[2-2-[(dimethylamino)methyl]phenyl]thio]benzyl alcohol ([(123)I]IDAM), has been developed for selective single-photon emission tomography (SPET) imaging of SERT. In vitro binding studies suggest a high selectivity of IDAM for SERT (K(i)=0.097 nM), with considerably lower affinities for norepinephrine and dopamine transporters (NET K(i)= 234 nM and DAT K(i)>10 microM, respectively). In this study the biodistribution of SERT in the baboon brain was investigated in vivo using [(123)I]IDAM and SPET imaging. Dynamic sequences of SPET scans were performed on three female baboons (Papio anubis) after injection of 555 MBq of [(123)I]IDAM. Displacing doses (1 mg/kg) of the selective SERT ligand (+)McN5652 were administered 90-120 min after injection of [(123)I]IDAM. Similar studies were performed using a NET inhibitor, nisoxetine, and a DAT blocker, methylphenidate. After 60-120 min, the regional distribution of tracer within the brain reflected the characteristic distribution of SERT, with the highest uptake in the midbrain area (hypothalamus, raphe nucleus, substantia nigra), and the lowest uptake in the cerebellum (an area presumed free of SERT). Peak specific binding in the midbrain occurred at 120 min, with a ratio to the cerebellum of 1.80+/-0.13. At 30 min, 85% of the radioactivity in the blood was metabolite. Following injection of a competing SERT ligand, (+)McN5652, the tracer exhibited rapid washout from areas with high concentrations of SERT (dissociation rate constant in the midbrain, averaged over three baboons, k(off)=0. 025+/-0.002 min(-1)), while the cerebellar activity distribution was undisturbed (washout rate 0.0059+/- 0.0003 min(-1)). Calculation of tracer washout rate pixel-by-pixel enabled the generation of parametric images of the dissociation rate constant. Similar studies using nisoxetine and methylphenidate had no effect on the distribution of [(123)I]IDAM in the brain. These results suggest that [(123)I]IDAM is suitable for selective SPET imaging of SERT in the primate brain, with high contrast, favorable kinetics, and negligible binding to either NET or DAT.

In vivo imaging of serotonin transporters with [99mTc]TRODAT-1 in nonhuman primates.

[99mTc]TRODAT-1 was the first 99mTc-labeled imaging agent to show specific binding to dopamine transporters (DAT) in the striatum (STR) of human brain. Additionally, in vitro binding and autoradiographic experiments demonstrated that this tracer also binds to serotonin transporters (SERT) in the midbrain/hypothalamus (MB) area. In this study, [99mTc]TRODAT-1 was investigated as a potentially useful ligand to image SERT in the MB of living brain. A total of eight single-photon emission tomography (SPET) scans were performed in two baboons (Papio anubis) after intravenous (i.v.) injection of 740 MBq (20 mCi) of [99mTc]TRODAT-1 using a triple-head gamma camera equipped with ultra-high-resolution fan-beam collimators (scan time: 0-210 min). In four blocking studies, baboons were pretreated with (+)McN5652 (1 mg/kg, i.v.) or methylphenidate (1 mg/kg, i.v.) to specifically block SERT or DAT, respectively. After co-registration with magnetic resonance images of the same baboon, a region of interest analysis was performed using predefined templates to calculate specific uptake in the midbrain area and the striatum, with the cerebellum as the background region [(MB-CB)/CB, (STR-CB)/CB]. Additionally, two PET scans of the same baboons were performed after i.v. injections of 74-111 MBq (2-3 mCi) of [11C](+)McN5652 to identify the SERT sites. In [99mTc]TRODAT-1/SPET scans, the SERT sites in the MB region were clearly visualized. Semiquantitative analysis revealed a specific uptake in MB ([MB-CB]/CB) of 0.30+/-0.02, which was decreased to 0. 040+/-0.005 after pretreatment with nonradioactive (+)McN5652, a selective SERT ligand. Pretreatment with methylphenidate reduced the specific binding of [99mTc]TRODAT-1 to DAT sites [(STR-CB)/CB] from 2.45+/-0.13 to 0.32+/-0.04 without any effect on its binding to SERT sites [(MB-CB)/CB], which was confirmed by the co-registration of the [11C](+)McN5652/PET scans. This preliminary study suggests that specific binding of [99mTc]TRODAT-1 to SERT sites can be detected by in vivo SPET imaging despite the low target to background ratio. These findings provide impetus for further development of similar compounds with improved binding affinity and selectivity to SERT sites.

Iodine-123 labelled nor-beta-CIT binds to the serotonin transporter in vivo as assessed by biodistribution studies in rats.

Iodine-123 labelled 2beta-carbomethoxy-3beta-4-iodophenylnortropane (nor-beta-CIT), a radioiodinated cocaine analogue, was evaluated as an agent for the in vivo labelling of serotonin transporters by biodistribution studies in rats. Intravenous injection of [123I]nor-beta-CIT resulted in high accumulation of radioactivity in brain areas with high densities of serotonin (hypothalamus) and dopamine transporters (striatum), although the binding was less pronounced in the hypothalamus. While binding of [123I]nor-beta-CIT in the hypothalamus was blocked significantly by fluvoxamine (a selective serotonin transporter blocker) but not by GBR12,909 (a selective dopamine transporter blocker), the opposite was observed in the striatum. The results of this study indicate that [123I]nor-beta-CIT, although not being a selective radioligand, binds specifically to serotonin transporters in the hypothalamus in vivo and thus suggest that [123I]nor-beta-CIT promises to be a suitable radioligand for single-photon emission tomography imaging of serotonin transporters in humans.

A Comparison of SPM’96 with ROI Analysis in Serotonin Transporter Imaging

A Comparison of SPM’96 with ROI Analysis in Serotonin Transporter Imaging

In vitro and in vivo characterisation of nor-?-CIT: a potential radioligand for visualisation of the serotonin transporter in the brain

Radiolabelled 2beta-carbomethoxy-3beta-(4-iodophenyl)tropane (beta-CIT) has been used in clinical studies for the imaging of dopamine and serotonin transporters with single-photon emission tomography (SPET). 2beta-Carbomethoxy-3beta-(4-iodophenyl)nortropane (nor-beta-CIT) is a des-methyl analogue of beta-CIT, which in vitro has tenfold higher affinity (IC50=0.36 nM) to the serotonin transporter than beta-CIT (IC50=4.2 nM). Nor-beta-CIT may thus be a useful radioligand for imaging of the serotonin transporter. In the present study iodine-125 and carbon-11 labelled nor-beta-CIT were prepared for in vitro autoradiographic studies on post-mortem human brain cryosections and for in vivo positron emission tomography (PET) studies in Cynomolgus monkeys. Whole hemisphere autoradiography with [125I]nor-beta-CIT demonstrated high binding in the striatum, the thalamus and cortical regions of the human brain. Addition of a high concentration (1 microM) of citalopram inhibited binding in the thalamus and the neocortex, but not in the striatum. In PET studies with [11C]nor-beta-CIT there was rapid uptake of radioactivity in the monkey brain (6% of injected dose at 15 min) and high accumulation of radioactivity in the striatum, thalamus and neocortex. Thalamus to cerebellum and cortex to cerebellum ratios were 2.5 and 1.8 at 60 min, respectively. The ratios obtained with [11C]nor-beta-CIT were 20%-40% higher than those previously obtained with [11C]beta-CIT. Radioactivity in the thalamus and the neocortex but not in the striatum was displaceable with citalopram (5 mg/kg). In conclusion, nor-beta-CIT binds to the serotonin transporter in the primate brain in vitro and in vivo and has potential for PET and SPET imaging of the serotonin transporter in human brain.

SPECT imaging of dopamine and serotonin transporters with [123I]beta-CIT. Binding kinetics in the human brain.

Single photon emission computerized tomography (SPECT) studies in non-human primates have previously shown that the cocaine derivative [123I]-2-beta-carbomethoxy-3-beta-(4-iodophenyl)-tropane ([123I]beta-CIT) labels dopamine transporters in the striatum and serotonin transporters in the hypothalamus-midbrain area. Here, we report on the regional kinetic uptake of [123I]beta-CIT in the brain of 4 normal volunteers and 2 patients with Parkinson's disease. In healthy subjects striatal activity increased slowly to reach peak values at about 20 hours post injection. In the hypothalamus-midbrain area peak activities were observed at about 4 hours with a slow decrease thereafter. Low activity was observed in cortical and cerebellar areas. The striatal to cerebellar ratio was about 4 after 5 hours and 9 after 20 hours. In 2 patients with idiopathic Parkinson's disease striatal activity was markedly decreased while the activity in hypothalamus-midbrain areas was only mildly diminished. Uptake into cortical and cerebellar areas appeared to be unchanged in Parkinson's disease. Consequently, in Parkinson's disease the striatal to cerebellar ratio was decreased to values around 2.5 after 20 hours. These preliminary methodological studies suggest that [123I]beta-CIT is a useful SPECT ligand for studying dopamine and possibly also serotonin transporters in the living human brain.

SPECT imaging of dopamine and serotonin transporters with [123I]β‐CIT: Pharmacological characterization of brain uptake in nonhuman primates

Single photon emission computed tomography (SPECT) studies of regional kinetic uptake and pharmacological specificity of [123I]methyl 3 beta-(4-iodophenyl) tropane-2 beta-carboxylate ([123I]beta-CIT) were performed in nonhuman primates (n = 41). In control experiments, activity was concentrated in striatum and in hypothalamic/midbrain regions. Striatal uptake increased for 140-180 min and displayed stable levels thereafter. Striatal to cerebellar activity ratios were 7.3 +/- 0.9 (mean +/- SEM) at 300 min. About 75% of striatal uptake was displaceable by injection of nonradioactive beta-CIT. Hypothalamic/midbrain activity reached maximal levels at approximately 45 min. A slow washout phase followed this peak activity. Activities in frontal, occipital, and cerebellar regions were characterized by an early peak (20-30 min), followed by rapid washout. Displacement studies demonstrated that striatal uptake was associated with dopamine (DA) transporters, as it was displaced by GBR 12909, a selective DA uptake inhibitor, but not by citalopram, a selective serotonin (5-HT) uptake inhibitor. The inverse was true in the hypothalamic/midbrain area, suggesting that the uptake in this area was associated primarily with 5-HT transporters. Maprotiline, a selective norepinephrine uptake inhibitor, did not affect [123I]beta-CIT uptake. In vivo site occupancy ED50 values of cocaine, 2 beta-carbomethoxy-3 beta-(4-fluorophenyl)tropane (CFT), and beta-CIT were measured in the striatum with a stepwise displacement paradigm. In vivo ED50 values correlated strongly with in vitro IC50 values for binding to DA transporters. Infusion of high dose of L-DOPA (250 mumol/kg) failed to displace striatal [123I]beta-CIT binding, suggesting that the binding would not be affected by L-DOPA administration in Parkinsonian patients. However, studies performed with injection of d-amphetamine indirectly suggested that high synaptic levels of DA may compete with [123I]beta-CIT binding. These studies suggest that [123I]beta-CIT will be a useful SPECT tracer of DA and 5-HT transporters in living human brain.

125I]5-Iodo-6-nitro-2-piperazinylquinoline: a potent and selective ligand for the serotonin uptake complex

Two novel ligands with 4′ substitution on the Phenyl Ring B of biphenylthiol, 5-chloro-2-(2′-((dimethylamino)methyl)-4′-iodophenylthio)benzenamine (7) and 2-(2′-((dimethylamino)methyl)-4′-methoxyphenylthio)-5-iodobenzenamine (8), were prepared and tested as potential serotonin transporter (SERT) imaging agents. The new ligands displayed extremely high binding affinities to SERT (Ki=0.22±0.09 and 0.11±0.04 nM, respectively), with very low binding affinities to dopamine and norepinephrine transporters (Ki>1000 nM). The corresponding [125I]7 and [125I]8 were successfully prepared from tri-n-butyltin derivatives. They showed good brain uptakes and prolonged retention after intravenous injection in rats (brain uptake was 1.77% and 0.98% dose/g for [125I]7, and 0.92% and 0.29% dose/g for [125I]8, at 2 and 120 min, respectively). Significantly, [125I]7 showed excellent uptake and prolonged retention in the hypothalamus, where SERT concentration was highest. The hypothalamus/cerebellum (HY/CB) ratios (target/background ratios) were 4.24, 7.10, 8.24 and 12.6 at 2, 4, 6 and 12 h, respectively. The HY/CB ratios for [125I]8 were 3.97, 5.57 and 5.06 at 1, 2 and 4 h, respectively. Adding the 4′-iodo group to the Phenyl Ring B of Compound (7) appeared to reduce the rate of clearance from the brain, and kinetics favored uptake and retention in the hypothalamus. The localization of [125I]7 in the hypothalamus region in the rat brain could be blocked by pretreatment with (+)McN5652, escitalopram and ADAM (2), which are all selective SERT ligands (at 2 mg/kg iv, 5 min pretreatment). Ex vivo autoradiograms of rat brain sections (at 4 h after intravenous injection of [125I]7) showed intense labeling in regions of the brain known to have high SERT density. The excellent selective uptake and retention in the hypothalamus region suggest that [123I]7 is a potential lead compound for developing new imaging agents targeting SERT-binding sites with single-photon emission computed tomography.