123I-iododexetimide preferentially binds to the muscarinic receptor subtype M1 in vivo.

UNLABELLED: The muscarinic M1 receptor (M1R) is highly involved in cognition, and selective M1 agonists have procognitive properties. Loss of M1R has been found in postmortem brain tissue for several neuropsychiatric disorders and may be related to symptoms of cognitive dysfunction. (123)I-iododexetimide is used for imaging muscarinic acetylcholine receptors (mAchRs). Considering its high brain uptake and intense binding in M1R-rich brain areas, (123)I-iododexetimide may be an attractive radiopharmaceutical to image M1R. To date, the binding affinity and selectivity of (123)I-iododexetimide for the mAchR subtypes has not been characterized, nor has its brain distribution been studied intensively. Therefore, this study aimed to address these topics. METHODS: The in vitro affinity and selectivity of (127)I-iododexetimide (cold-labeled iododexetimide), as well as its functional antagonist properties (guanosine 5'-[γ-(35)S-thio]triphosphate [GTPγ(35)S] assay), were assessed on recombinant human M1R-M5R. Distributions of (127)I-iododexetimide and (123)I-iododexetimide in the brain were evaluated using liquid chromatography-mass spectrometry and storage phosphor imaging, respectively, ex vivo in rats, wild-type mice, and M1-M5 knock-out (KO) mice. Inhibition of (127)I-iododexetimide and (123)I-iododexetimide binding in M1R-rich brain areas by the M1R/M4R agonist xanomeline, or the antipsychotics olanzapine (M1R antagonist) and haloperidol (low M1R affinity), was assessed in rats ex vivo. RESULTS: In vitro, (127)I-iododexetimide displayed high affinity for M1R (pM range), with modest selectivity over other mAchRs. In biodistribution studies on rats, ex vivo (127)I-iododexetimide binding was much higher in M1R-rich brain areas, such as the cortex and striatum, than in cerebellum (devoid of M1Rs). In M1 KO mice, but not M2-M5 KO mice, (127)I-iododexetimide binding was strongly reduced in the frontal cortex compared with wild-type mice. Finally, acute administration of both an M1R/M4R agonist xanomeline and the M1R antagonist olanzapine was able to inhibit (123)I-iododexetimide ex vivo, and (123)I-iododexetimide binding in M1-rich brain areas in rats, whereas administration of haloperidol had no effect. CONCLUSION: The current results suggest that (123)I-iododexetimide preferentially binds to M1R in vivo and can be displaced by M1R ligands. (123)I-iododexetimide may therefore be a useful imaging tool as a way to further evaluate M1R changes in neuropsychiatric disorders, as a potential stratifying biomarker, or as a clinical target engagement biomarker to assess M1R.

Incremental utility of iodine-123 meta-iodobenzylguanidine imaging beyond established heart failure risk models.

BACKGROUND: Nuclear myocardial imaging with iodine-123 meta-iodobenzylguanidine ((123)I-mIBG) is approved for risk stratification of patients with systolic heart failure (HF). Whether (123)I-mIBG imaging provides incremental prognostic utility beyond established risk models remains unclear. METHODS AND RESULTS: In a multicenter study, 961 patients with moderate systolic HF underwent (123)I-mIBG imaging and were followed for cardiac death, progressive HF, or life-threatening arrhythmias over 2 years. We constructed 4 multivariable models, using variables from each of 4 published HF risk models, and patient-level scores were calculated both before and after adding the heart-to-mediastinum ratio (H/M) from (123)I-mIBG imaging. Incremental utility was evaluated by calculating integrated discrimination improvement (IDI), which quantifies the increase in probability of experiencing the primary end point after adding H/M to each model. The composite end point occurred in 25% of patients. After adding H/M, absolute IDI ranged from 2.1% to 3.0%, representing 33%-59% relative improvements in risk stratification. Of note, hazard ratios for H/M were remarkably similar between risk models (0.40-0.44 for predicting the composite end point, 0.10-0.18 for mortality; all P < .001). CONCLUSIONS: Despite notable differences in predictor variables, patient populations, and analytic techniques from which each model was initially derived, adding (123)I-mIBG data to HF risk models consistently identified patients at lower risk of experiencing adverse events.

Decreased ipsilateral [¹²³I]iododexetimide binding to cortical muscarinic receptors in unilaterally 6-hydroxydopamine lesioned rats.

INTRODUCTION: Dysfunction of the cholinergic neurotransmitter system is present in Parkinson's disease, Parkinson's disease related dementia and dementia with Lewy bodies, and is thought to contribute to cognitive deficits in these patients. In vivo imaging of the cholinergic system in these diseases may be of value to monitor central cholinergic disturbances and to select cases in which treatment with cholinesterase inhibitors could be beneficial. The muscarinic receptor tracer [(123)I]iododexetimide, predominantly reflecting M1 receptor binding, may be an appropriate tool for imaging of the cholinergic system by means of SPECT. In this study, we used [(123)I]iododexetimide to study the effects of a 6-hydroxydopamine lesion (an animal model of Parkinson's disease) on the muscarinic receptor availability in the rat brain. METHODS: Rats (n=5) were injected in vivo at 10-13 days after a confirmed unilateral 6-hydroxydopamine lesion. Muscarinic receptor availability was measured bilaterally in multiple brain areas on storage phosphor images by region of interest analysis. RESULTS: Autoradiography revealed a consistent and statistically significant lower [(123)I]iododexetimide binding in all examined neocortical areas on the ipsilateral side of the lesion as compared to the contralateral side. In hippocampal and subcortical areas, such asymmetry was not detected. CONCLUSIONS: This study suggests that evaluation of muscarinic receptor availability in dopamine depleted brains using [(123)I]iododexetimide is feasible. We conclude that 6-hydroxydopamine lesions induce a decrease of neocortical muscarinic receptor availability. We hypothesize that this arises from down regulation of muscarinic postsynaptic M1 receptors due to hyperactivation of the cortical cholinergic system in response to dopamine depletion. ADVANCES IN KNOWLEDGE: In rats, dopamine depletion provokes a decrease in neocortical muscarinic receptor availability, which is evaluable by [(123)I]iododexetimide imaging. IMPLICATIONS FOR PATIENT CARE: This study may further underline the role of a dysregulated muscarinic system in patients with Lewy body disorders.

Sequential 123I-iododexetimide scans in temporal lobe epilepsy: comparison with neuroimaging scans (MR imaging and 18F-FDG PET imaging).

PURPOSE: Muscarinic acetylcholine receptors (mAChRs) play an important role in the generation of seizures. Single-photon emission computed tomography (SPECT) with 12I-iododexetimide (IDEX) depicts tracer uptake by mAChRs. Our aims were to: (a) determine the optimum time for interictal IDEX SPECT imaging; (b) determine the accuracy of IDEX scans in the localisation of seizure foci when compared with video EEG and MR imaging in patients with temporal lobe epilepsy (TLE); (c) characterise the distribution of IDEX binding in the temporal lobes and (d) compare IDEX SPECT and 18F-fluorodeoxyglucose (FDG) positron emission tomography (PET) in identifying seizure foci. METHODS: We performed sequential scans using IDEX SPECT imaging at 0, 3, 6 and 24 h in 12 consecutive patients with refractory TLE undergoing assessment for epilepsy surgery. Visual and region of interest analyses of the mesial, lateral and polar regions of the temporal lobes were used to compare IDEX SPECT, FDG PET and MR imaging in seizure onset localisation. RESULTS: The 6-h IDEX scan (92%; kappa=0.83, p=0.003) was superior to the 0-h (36%; kappa=0.01, p>0.05), 3-h (55%; kappa=0.13, p>0.05) and 24-h IDEX scans in identifying the temporal lobe of seizure origin. The 6-h IDEX scan correctly predicted the temporal lobe of seizure origin in two patients who required intracranial EEG recordings to define the seizure onset. Reduced ligand binding was most marked at the temporal pole and mesial temporal structures. IDEX SPECT was superior to interictal FDG PET (75%; kappa=0.66, p=0.023) in seizure onset localisation. MR imaging was non-localising in two patients in whom it was normal and in another patient in whom there was bilateral symmetrical hippocampal atrophy. CONCLUSION: The 6-h IDEX SPECT scan is a viable alternative to FDG PET imaging in seizure onset localisation in TLE.

In vivo imaging of muscarinic cholinergic receptors in temporal lobe epilepsy with a new PET tracer: [76Br]4-bromodexetimide.

UNLABELLED: Muscarinic acetyl cholinergic receptors (mAChRs) may be involved in the pathophysiology of partial epilepsy. Previous experimental and imaging studies have reported medial temporal abnormalities of mAChR in patients with medial temporal lobe epilepsy (MTLE). Suitable radiotracers for mAChR are required to evaluate these disturbances in vivo using PET. Dexetimide is a specific mAChR antagonist that has been labeled recently with 76Br. This first study in humans focused on regional distribution and binding kinetics of [76Br]4-bromodexetimide (BDEX) in patients with MTLE. METHODS: Ten patients with well-lateralized MTLE had combined MRI, 18F-fluorodeoxyglucose (FDG) PET and 76Br-BDEX PET studies. Time-activity curves were generated in PET-defined regions of interest, including the medial, polar and lateral regions of the temporal lobe; the basal ganglia; the external and medial occipital cortex; and the white matter. RESULTS: The highest radioactivity concentration was observed in the basal ganglia and in the cortical regions, whereas radioactivity was lower in the white matter. On late images of PET studies, 76Br-BDEX uptake was statistically significantly decreased only in the medial temporal region ipsilateral to the seizure focus (1.37 +/-0.28, P < 0.01) as determined by FDG PET imaging, anatomic MRI and electroencephalogram correlation, compared with the contralateral medial temporal region (1.46 +/- 0.31). CONCLUSION: 76Br-BDEX concentration is reduced in the temporal lobe ipsilateral to the seizure focus in patients with MTLE. This preliminary study suggests that 76Br-BDEX is a suitable radiotracer for studies of mAChR in humans. Further studies are required to investigate the potential value of 76Br-BDEX PET in other neurological disorders with muscarinic disturbances.

A study of myocardial muscarinic receptors in streptozotocin-induced diabetic rats using iodine-123 N-methyl-4-iododexetimide.

In previous studies we have shown that iodine-123 N-methyl-4-iododexetimide ([123I]MIDEX) is a suitable single-photon emission tomography radiotracer for the characterisation of myocardial muscarinic acetylcholine receptors (m-AChR) in the normal state. It has been demonstrated that m-AChR are altered as a consequence of diabetes. The aim of the present study was to examine myocardial m-AChR density using [123I]MIDEX in streptozotocin (STZ)-induced diabetic rats. In vitro binding experiments were conducted on left and right ventricle and atrium homogenate membranes of 1-week, 5-week and 10-week STZ-induced diabetic and aged-matched normal rats. The m-AChR densities (Bmax values), as determined by saturation experiments with [123I]MIDEX, revealed no difference in left and right ventricles or atrium in 1-week and 5-week STZ-diabetic rats when compared with normal rats. However, the 10-week STZ-diabetic group revealed a 39% (P<0.001) decrease in m-AChR density in atrium with no change in left and right ventricles. The equilibrium dissociation constant (Kd values) was similar in all groups. In vitro binding autoradiography revealed a 40% decrease in m-AChR density in atrium in the same 10-week diabetic rats. No statistically significant difference was found in 1-week and 5-week diabetic rats compared with normals. Ex vivo autoradiography showed a 50% decrease in [123I]MIDEX uptake in atrium in 5-week diabetic rats and a 60% decrease in 10-week diabetic rats. These results demonstrate the ability of the single-photon agent [123I]MIDEX to measure in vitro and ex vivo alterations in myocardial m-AChR density observed in STZ-induced diabetic rats.

Influence of acetylcholine on binding of 4-[125I]iododexetimide to muscarinic brain receptors.

The distribution of nicotinic and muscarinic cholinergic receptors in the human brain in vivo has been successfully characterized using radiolabeled tracers and emission tomography. The effect of acetylcholine release into the synaptic cleft on receptor binding of these tracers has not yet been investigated. The present study examined the influence of acetylcholine on binding of 4-[125I]iododexetimide to muscarinic cholinergic receptors of porcine brain synaptosomes in vitro. 4-Iododexetimide is a subtype-unspecific muscarinic receptor antagonist with high affinity. Acetylcholine competed with 4-[125I]iododexetimide in a dose-dependent manner. A concentration of 500 microM acetylcholine inhibited 50% of total specific 4-[125I]iododexetimide binding to synaptosomes when both substances were given simultaneously. An 800 microM acetylcholine solution reduced total specific 4-[125I]iododexetimide binding by about 35%, when acetylcholine was given 60 min after incubation of synaptosomes with 4-[125I]iododexetimide. Variations in the synaptic acetylcholine concentration might influence muscarinic cholinergic receptor imaging in vivo using 4-[123I]iododexetimide. Conversely, 4-[123I]iododexetimide might be an appropriate molecule to investigate alterations of acetylcholine release into the synaptic cleft in vivo using single photon emission computed tomography.

Effect of partial volume correction on muscarinic cholinergic receptor imaging with single-photon emission tomography in patients with temporal lobe epilepsy.

Animal experiments and preliminary results in humans have indicated alterations of hippocampal muscarinic acetylcholine receptors (mAChR) in temporal lobe epilepsy. Patients with temporal lobe epilepsy often present with a reduction in hippocampal volume. The aim of this study was to investigate the influence of hippocampal atrophy on the quantification of mAChR with single photon emission tomography (SPET) in patients with temporal lobe epilepsy. Cerebral uptake of the muscarinic cholinergic antagonist [123I]4-iododexetimide (IDex) was investigated by SPET in patients suffering from temporal lobe epilepsy of unilateral (n=6) or predominantly unilateral (n=1) onset. Regions of interest were drawn on co-registered magnetic resonance images. Hippocampal volume was determined in these regions and was used to correct the SPET results for partial volume effects. A ratio of hippocampal IDex binding on the affected side to that on the unaffected side was used to detect changes in muscarinic cholinergic receptor density. Before partial volume correction a decrease in hippocampal IDex binding on the focus side was found in each patient. After partial volume no convincing differences remained. Our results indicate that the reduction in hippocampal IDex binding in patients with epilepsy is due to a decrease in hippocampal volume rather than to a decrease in receptor concentration.

Pharmacological characterization and positron emission tomography evaluation of 4-[76Br]bromodexetimide and 4-[76Br]bromolevetimide for investigations of central muscarinic cholinergic receptors.

4-[76Br]bromodexetimide and its inactive enantiomer 4-[76Br]bromolevetimide were prepared via electrophilic bromodesilylation using chloramine-T and no-carrier-added (NCA) [76Br]NH4. In vitro, Bmax measured on rat cortex membranes were 3.7 +/- 0.2 and < 0.07 pmol/mg protein for 4-[76Br]bromodexetimide and 4-[76Br]bromolevetimide, respectively. The kD of 4-[76Br]bromodexetimide was 1.9 +/- 0.3 nM. In vivo studies in rats showed specific uptake of 4-[76Br]bromodexetimide in cortex, striatum, thalamus and hippocampus. No specific uptake was observed with 4-[76Br]bromolevetimide. With [76Br]bromodexetimide, positron emission tomography (PET) studies in primates demonstrated a preferential accumulation of the radioactivity in the cortex and striatum which was displaced to the level of cerebellum by dexetimide. With 4-[76Br]bromolevetimide, the radioactivity concentrations in the cortex and striatum were similar to that of cerebellum.

Radiosynthesis of [123I]N-methyl-4-iododexetimide and [123I]N-methyl-4-iodolevetimide: in vitro and in vivo characterisation of binding to muscarinic receptors in the rat heart.

[123I]N-methyl-4-iododexetimide, [123I]MIDEX, and its pharmacologically inactive enantiomer [123I]N-methyl-4-iodolevetimide, [123I]MILEV were prepared via electrophilic iododesilylation using Chloramine-T as oxidising agent followed by N-methylation using methyl iodide. The radiotracers were purified with semi-preparative HPLC with radiochemical yields of 80 +/- 11% (n = 6). The average time of synthesis was 100 min with specific activity > 2000 Ci/mmol. In vitro, the binding of [123I]MIDEX, after addition of carrier, measured on homogenates of rat atrium was Bmax = 4.5 +/- 0.4 pmol/mg protein, Kd = 3.3 +/- 0.2 nM while in the ventricle Bmax = 2.3 +/- 0.2 pmol/mg protein, Kd = 4.0 +/- 1.4 nM. In vitro, the binding of [123I]MILEV was non-specific. The in vivo biodistribution of [123I]MIDEX showed high uptake in the atrium (3.2% ID/g) and left and right ventricles (2.2, 2.5% ID/g respectively) at 5 min followed by clearance. High heart-to-lung and moderate liver-to-lung ratios were obtained during 60 min. Radioactivity in the atrium and ventricles was reduced by pre-administration of the m-AChR antagonist MQNB (1 mg/kg). Pretreatment of rats with other m-AChR ligands, pirenzapine (M1), methoctramine (M2) and 4-DAMP (M3) also resulted in reduction of [123I]MIDEX uptake with methoctramine being the most potent. [123I]MIDEX distribution in the rat heart was not significantly inhibited by pre-administration of selective adrenergic drugs. The uptake was highly stereoselective since the inactive enantiomer, [123I]MILEV, demonstrated very low myocardial retention. The stability of [123I]MIDEX was evaluated by performing a metabolite study on atrium samples which revealed unchanged radiotracer 60 min postinjection. These results suggest that [123I]MIDEX may be a useful single photon agent for in vivo imaging of myocardial m-AChR in humans with [123I]MILEV offering the potential of assessing non-specific binding of the active tracer.

In vitro pharmacological properties of 4-bromodexetimide for muscarinic receptors.

The decrease of m-AChR density observed in neurodegenerative disorders has generated considerable interest in non-invasive mapping of muscarinic acetylcholine receptors (m-AChR) in the central nervous system. The aim of our study was to evaluate the selectivity of 4-bromodexetimide for the M1, M2, M3 and M4 m-AChR subtypes using in vitro binding analysis to determine the potential use of the bromine-76 labelled 4-bromodexetimide in the investigation of m-AChR subtypes in human brain with Positron Emission Tomography. Subtype selectivity of 4-bromodexetimide was determined in competition studies against tritiated subtype selective ligands using various rat or rabbit structure homogenates reflecting a single binding site and in optimal saturation and low non specific binding conditions. These conditions were reached for every subtype studied by analyzing the data from the saturation experiments of the tritiated ligands. 4-bromodexetimide displayed nanomolar affinities for the four m-AChR subtypes and a preferential selectivity for the M1 and M4 subtypes. The saturation analysis of [76Br]4-bromodexetimide, performed with rat cortex membranes showed high affinity for m-AChR receptors (Kd = 1.8 nM). As in vivo studies of [76Br]4-bromodexetimide showed preferential localization in the cortex and the striatum which are M1 and M4 rich structures and since it binds preferentially to the M1 and M4 subtypes, this radiotracer can still allow a combined subtype specific measurement of these muscarinic receptors.

Human dosimetry and biodistribution of iodine-123-iododexetimide: a SPECT imaging agent for cholinergic muscarinic neuroreceptors.

UNLABELLED: Iodine-123-iododexetimide (IDEX) has recently been used for SPECT imaging of muscarinic cholinergic neuroreceptors (mAChR) in humans. We report the human radiation dosimetry, whole-body and normal cerebral distribution of IDEX. METHODS: Serial whole-body planar and brain SPECT scans were performed over 24 hr in four normal subjects. Organ activity was calculated from attenuation-corrected geometric mean counts from ROIs drawn over visible organs. Thigh activity was used for background subtraction. Organ absorbed doses and effective dose were calculated using the MIRD schema. Brain SPECT was performed 6 hr postinjection in ten normal subjects. ROIs placed over cortical and subcortical structures were used to determine brain distribution. RESULTS: The effective dose was 24.7 microSv/MBq. An average of 54% of IDEX remained in the body background. Decay-corrected brain uptake was 6.9% of injected dose at 1 hr, 8.6% at 6 hr and 8.1% at 24 hr. Regional brain distribution showed high uptake in striatum and cortex with low activity in thalamus and cerebellum. At 6 hr, activity relative to striatum was 70% for frontal and parietal cortex, 102% for occipital cortex, 54% for thalamus and 11% for cerebellum. CONCLUSION: Iodine-123-IDEX produced high quality SPECT images with activity at 6 hr reflecting the known distribution of mAChR receptors. The favorable dosimetry of IDEX and high synthetic yield (50%-70%) suggest it to be a suitable agent for clinical studies.

Iodine-123 N-methyl-4-iododexetimide: a new radioligand for single-photon emission tomographic imaging of myocardial muscarinic receptors.

Cardiac muscarinic receptor ligands suitable for positron emission tomography have previously been characterised. Attempts to develop radioligands of these receptors suitable for single-photon emission tomographic (SPET) imaging have not been successful due to high lung retention and high non-specific binding of previously investigated potential tracers. The purpose of this study was to evaluate the biodistribution and in vivo imaging characteristics of a new radiopharmaceutical, [123I]N-methyl-4-iododexetimide. Biodistribution studies performed in rats showed high cardiac uptake (2.4% ID/g) 10 min after injection with a heart to lung activity ratio of 5:1. Specificity and stereoselectivity of cardiac binding were demonstrated using blocking experiments in rats. Dynamic imaging studies in anaesthetised greyhounds demonstrated rapid and high myocardial uptake and low lung binding with stable heart to lung activity ratios of > 2.5:1 between 10 and 30 min, making SPECT imaging feasible. Administration of an excess of an unlabelled muscarinic antagonist, methyl-quinuclidinyl benzylate rapidly displaced myocardial activity to background levels and the pharmacologically inactive enantiomer, [123I]N-methyl-4-iodolevetimide, had no detectable cardiac uptake, indicating specific and stereoselective muscarinic receptor binding. SPET revealed higher activity in the inferior than in the anterior wall, this being consistent with previously described regional variation of cardiac parasympathetic innervation. [123I]N-methyl-4-iododexetimide shows promise as an imaging agent for muscarinic receptor distribution in the heart and may be helpful in evaluating diverse cardiac diseases associated with altered muscarinic receptor function, including heart failure and diabetic heart disease.

Assessment of brain muscarinic acetylcholinergic receptors in living mice using a simple probe, [125I]-4-iododexetimide and [125I]-4-iodolevetimide.

This study describes assessment of brain muscarinic acetylcholinergic receptors in living mice using a single-crystal radiation detection system, the high-affinity antagonist [125I]-4-iododexetimide, and the inactive enantiomer [125I]-4-iodolevetimide. Kinetics of radioligand binding, as well as perturbation by atropine displacement, can be determined using this simple probe technique.

Decreased hippocampal muscarinic cholinergic receptor binding measured by 123I-iododexetimide and single-photon emission computed tomography in epilepsy.

Regional binding of 123I-iododexetimide, a muscarinic acetylcholine receptor antagonist, was measured in vivo in the temporal lobes of 4 patients with complex partial seizures using single-photon emission computed tomography. In the anterior hippocampus ipsilateral to the electrical focus, 123I-iododexetimide binding was decreased by 40 +/- 9% (mean +/- SD, p < 0.01) compared with the contralateral hippocampus; 123I-iododexetimide binding in other temporal lobe regions was symmetrical. The data indicate a regionally specific change of muscarinic acetylcholine receptor in anterior hippocampus in complex partial seizures of temporal lobe origin.

Imaging muscarinic cholinergic receptors in human brain in vivo with Spect, [123I]4-iododexetimide, and [123I]4-iodolevetimide.

A method to image muscarinic acetylcholine receptors (muscarinic receptors) noninvasively in human brain in vivo was developed using [123I]4-iododexetimide ([123I]IDex), [123I]4-iodolevetimide ([123I]ILev), and single photon emission computed tomography (SPECT). [123I]IDex is a high-affinity muscarinic receptor antagonist. [123I]ILev is its pharmacologically inactive enantiomer and measures nonspecific binding of [123I]IDex in vitro. Regional brain activity after tracer injection was measured in four young normal volunteers for 24 h. Regional [123I]IDex and [123I]ILev activities were correlated early after injection, but not after 1.5 h. [123I]IDex activity increased over 7-12 h in neocortex, neostriatum, and thalamus, but decreased immediately after the injection peak in cerebellum. [123I]IDex activity was highest in neostriatum, followed in rank order by neocortex, thalamus, and cerebellum. [123I]IDex activity correlated with muscarinic receptor concentrations in matching brain regions. In contrast, [123I]ILev activity decreased immediately after the injection peak in all brain regions and did not correspond to muscarinic receptor concentrations. [123I]IDex activity in neocortex and neostriatum during equilibrium was six to seven times higher than [123I]ILev activity. The data demonstrate that [123I]IDex binds specifically to muscarinic receptors in vivo, whereas [123I]ILev represents the nonspecific part of [123I]IDex binding. Subtraction of [123I]ILev from [123I]IDex images on a pixel-by-pixel basis therefore reflects specific [123I]IDex binding to muscarinic receptors. Owing to its high specific binding, [123I]IDex has the potential to measure small changes in muscarinic receptor characteristics in vivo with SPECT. The use of stereoisomerism directly to measure nonspecific binding of [123I]IDex in vivo may reduce complexity in modeling approaches to muscarinic acetylcholine receptors in human brain.

In vitro and in vivo characterization of 4-[125I]iododexetimide binding to muscarinic cholinergic receptors in the rat heart.

4-[125I]iododexetimide binding to muscarinic cholinergic receptors (mAChR) was evaluated in the rat heart. 4-[125I]iododexetimide displayed high in vitro affinity (Kd = 14.0 nM) for rat myocardial mAChR. In vivo, there was high accumulation of 4-[125I]iododexetimide in the rat atrium and ventricle which could be blocked by approximately 60% by preinjection of atropine. In contrast, accumulation of the radiolabeled stereoisomer, 4-[125I]iodolevetimide, was 63% lower than 4-[125I]iodolevetimide and was not blocked by atropine. The blood clearance of 4-[125I]iododexetimide was rapid, providing heart-to-blood ratios of up to 14:1; however, heart-to-lung and heart-to-liver ratios were below unity. The data indicate that 4-[125I]iododexetimide binds potently to rat mAChR. However, since nonspecific binding is relatively high, it is not clear whether iododexetimide labeled with 123I will be useful in SPECT imaging studies of myocardial mAChR. Further studies in humans are indicated.

In vivo biodistribution of two [18F]-labelled muscarinic cholinergic receptor ligands: 2-[18F]- and 4-[18F]-fluorodexetimide.

Two [18F]-labelled analogues of the potent muscarinic cholinergic receptor (m-AChR) antagonist, dexetimide, were evaluated as potential ligands for imaging m-AChR by positron emission tomography (PET). Intravenous administration of both 2-[18F]- or 4-[18F]-fluorodexetimide resulted in high brain uptake of radioactivity in mice. High binding levels were observed in m-AChR rich areas, such as cortex and striatum, with low levels in the receptor-poor cerebellum. Uptake of radioactivity was saturable and could be blocked by pre-administration of dexetimide or atropine. Drugs with different sites of action were ineffective at blocking receptor binding. The results indicate that both radiotracers are promising candidates for use in PET studies.

Positron labeled muscarinic acetylcholine receptor antagonist: 2- and 4-[18F]fluorodexetimide. Syntheses and biodistribution.

Two 18F-labeled analogues of dexetimides, 2-[18F]fluorodexetimide (2-FDEX) and 4-[18F]fluorodexetimide (4-FDEX), were prepared and evaluated in vivo as possible agents for the study of the muscarinic acetylcholine receptor (mAChR) with PET. Two synthetic approaches, a 2-step reductive alkylation procedure and a 4-step alkylation approach, were investigated. The alkylation approach with higher overall radiochemical yields was used to prepare 2- and 4-FDEX for biodistribution studies. The overall synthesis time for both compounds was 2.5 h and the overall radiochemical yield at end-of-synthesis was 12%. The specific activity was found to be greater than 600 mCi/mumol. Biodistribution studies of 2-FDEX in rats produced striatum-to-cerebellum and cortex-to-cerebellum ratios of 8.6 +/- 1.1 and 8.4 +/- 1.0 at 1 h after injection, and 12.1 +/- 2.1 and 10.7 +/- 2.2 at 3 h, respectively. Substantial radioactivity detected in bone indicated the in vivo defluorination of 2-FDEX. The striatum-to-cerebellum ratio for 4-FDEX was slightly lower at 1 h (5.9 +/- 0.9) but equally high at 3 h (12.3 +/- 2.0) when compared to 2-FDEX, and there was little bone uptake. The uptake of both 2-FDEX and 4-FDEX into mAChR rich brain regions (e.g. striatum, cortex) was blocked by a dose of dexetimide (5 mg/kg). Our results suggest 4-FDEX is a potential PET agent for study mAChR in vivo.

Synthesis and biological evaluation of [125I]- and [123I]-4-iododexetimide, a potent muscarinic cholinergic receptor antagonist.

A series of halogenated racemic analogues of dexetimide (1) was synthesized and their affinity for the muscarinic cholinergic receptor measured. One analogue, 4-iododexetimide (21), was efficiently labeled with 125I and 123I at high specific activity. In vitro binding studies and in vivo biodistribution studies suggest that 123I-labeled 21 may be useful for imaging muscarinic cholinergic receptors in the living human brain with single photon emission computed tomography.