Jean N. DaSilva

In vivo evaluation of [11C]-and [18F]-labelled cocaine analogues as potential dopamine transporter ligands for positron emission tomography☆

Four analogues of the potent dopamine transporter ligand, WIN 35,428, were radiolabelled with 11C and 18F at the 2-beta-carboxy position for evaluation as potential ligands for imaging dopamine uptake sites by positron emission tomography (PET) namely, methyl (1R-2-exo-3-exo)-8- methyl-3-(4-methylphenyl)-8-azabicyclo[3.2.1]octane-2-carboxylate (RTI-32), its 4-chlorophenyl analogue (RTI-31), 2-fluoroethyl (1R-2-exo-3-exo)-8-methyl-3-(4-methylphenyl)-8-azabicyclo[3.2.1]octane-2 - carboxylate (FETT) and its 4-chlorophenyl analogue (FECT). Upon intravenous injection in rats, all four radiotracers displayed preferential accumulation of radioactivity in regions known to contain high concentrations of dopamine uptake sites. Competition studies with two of the analogues, [11C]RTI-32 and [18F]FETT, demonstrated that, for both radiotracers, binding was saturable and displayed the appropriate pharmacology as potential PET ligands for imaging the dopamine transporter. Striatum to cerebellar ratios for [11C]RTI-32 (at 90 min post-injection) and [18F]FETT (at 120 min post-injection) were 27 and 21, respectively.

Synthesis of two radiofluorinated cocaine analogues using distilled 2-[18F]fluoroethyl bromide

Two fluorinated congeners of cocaine, 2-fluoroethyl (1R-2-exo-3-exo)-8-methyl-3-(4-methylphenyl)-8-azabicyclo[3.2.1]octane- 2-carboxylate (FETT) and its 4-chlorophenyl analogue (FECT) were synthesized. Radiolabelling with 18F was achieved by O-[18F]fluoroalkylation of the corresponding carboxylic acid salts with distilled 2-[18F]fluoroethyl bromide in DMF. After HPLC purification, yields of radiochemically pure, formulated products were 22-30% (not corrected for decay) in a synthesis time of 60-70 min. The use of distilled 2-[18F]fluoroethyl bromide was indispensable for the reliable production of pure products.

Neuromodulation of frontal and temporal cortex by intravenous d-fenfluramine: an [15O]H2O PET study in humans

This study assessed the modulatory effect of a serotonergic agonist, d-fenfluramine, on localized neuronal firing as indexed by changes in regional cerebral blood flow (rCBF). Previously, we reported the effect of oral d, l-fenfluramine on neuronal activity as measured by change in [18F]fluorodeoxyglucose uptake. Improvements in the current study include: a more specific serotonin agonist, d-fenfluramine; a more reliable administration route, intravenous; and a one session paradigm made possible with the radiotracer [15O]H2O. Changes in relative rCBF (P < 0.001) were observed: increases within the frontal cortex bilaterally and decreases within the temporal cortex bilaterally, and left thalamus. Other significant findings were elevated cortisol and growth hormone; increased euphoria and panic symptoms and decreased tiredness. These results support further investigation with intravenous d-fenfluramine to study the net functional effects of serotonergic stimulation in health and illness.

Derivatives of way 100635 as potential imaging agents for 5-ht1a receptors: syntheses, radiosyntheses, and in vitro and in vivo evaluation

Analogues of the potent and selective 5-HT1A ligand, WAY 100635, were synthesized and examined as potential candidates for imaging 5-HT1A receptors by positron emission tomography (PET). Several of the analogues displayed nanomolar affinity for the 5-HT1A receptor, comparable to WAY 100635. Three of these were examined in a model of human liver metabolism vis-à-vis WAY 100635. All showed a markedly lower propensity for amide hydrolysis than WAY 100635. Radiolabelling of these three potential PET radiotracers with carbon-11 was readily achieved from [11C]-iodomethane, and the newly synthesized radioligands were tested in vivo in rats for binding to 5-HT1A receptors. Whereas two of the ligands failed to bind to 5-HT1A receptors in vivo, one was successful. The latter, [11C]-7 [4-(2-methoxyphenyl)-1-[2-[N-(2-pyridinyl)-2-bicyclo[2.2.2]octanec arboxamido]ethyl]-piperazine], showed good brain penetration, hippocampal:cerebellar ratios of 10:1 at 45 min postinjection. Blocking studies with a variety of drugs demonstrated that the binding of [11C]-7 in vivo was selective for 5-HT1A receptors. [11C]-7 is a promising candidate as a ligand for imaging 5-HT1A receptors by PET.

The D2 receptor occupancy profile of loxapine determined using PET

Positron emission tomography (PET) studies of typical neuroleptics suggest that 60% to 80% of striatal D2 occupancy may be sufficient for optimal clinical treatment of psychosis. Therefore, striatal D2 occupancy may be used as an index to determine the optimal dose range. Toward this end, we determined the in vivo D2 profile of loxapine, using [11C]-raclopride and PET. Seven patients selected from a clinical population were scanned while taking steady-state oral loxapine from 10 to 100 mg/day. Their D2 receptor occupancy was estimated by comparing them to age-matched data from neuroleptic-naive patients. The D2 receptor occupancy ranged from 52% to 90%, and there was a very strong relationship between dose and D2 occupancy, suggesting that 15 to 30 mg/day of loxapine would produce, the putatively optimal, 60% to 80% striatal D2 blockade. This dose range is much lower than that used in most clinical settings and points to the potential efficacy of loxapine at lower doses.

[18F]fluoroalkyl analogues of the potent 5-HT1A antagonist WAY 100635: Radiosyntheses and in vivo evaluation

Two analogues of the potent 5-HT1A antagonist WAY 100635 have been synthesized and radiolabelled with 18F, namely N-[2-[4-(2-2-[18F] fluoroethoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclohe xan e carboxamide ([18F]FEC) and N-[2-[4-(2-3-[18F] fluoropropoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cycloh exa ne carboxamide ([18F]FPC). Biodistribution studies in rats showed selective uptake of both radiotracers in regions known to be rich in 5-HT1A receptors following i.v. injection. The ratio of radioactivity in hippocampus to that in the cerebellum was 5.5 (for [18F]FEC) and 7.5 (for [18F]FPC) at 60 min postinjection. Regional brain heterogeneity of radioactivity could be abolished by pretreatment with WAY 100635 and FPC but was unaffected by pretreatment with a variety of drugs including ketanserin, sulpiride, and SCH 23390. These results are compared vis-a-vis with those obtained using [11C]WAY 100635 to evaluate [18F]FEC and [18F]FPC as potential radiotracers for imaging 5-HT1A receptors by positron emission tomography.

Dopamine D1 agonist R-[11C]SKF 82957: synthesis and in vivo characterization in rats

The active enantiomer R-SKF 82957 was labeled with 11C by N-[11C]methylation of the full dopamine (D1) agonist R-SKF 81297, using [11C]methyl iodide in the presence of N-ethyldiisopropylamine, in high specific activity, radiochemical purity and yields. Compared with the D1 agonist R/S-[11C]SKF 82957, R-[11C]SKF 82957 showed higher binding in the D1 rich regions, such as striatum and olfactory tubercles (approximately 1.7 times), thereby improving the tissue contrast. R-[11C]SKF 82957 exhibited high in vivo binding selectivity for D1 receptors in rats, because only high doses of D1 competitors, but not D2 or serotonin (5-HT2) blockers, significantly reduced the radioactivity levels in all brain areas. No labeled metabolites were detected in rat brain. These results indicate that R-[11C]SKF 82957 will provide more sensitive measurements of D1 receptors in in vivo studies than the racemic mixture.

Synthesis and autoradiographic localization of the dopamine D-1 agonists [11C]SKF 75670 and [11C]SKF 82957 as potential PET radioligands.

The high affinity benzazepine D1 agonists SKF 75670 and SKF 82957 were labeled with 11C by N-[11C]methylation of SKF 38393 and SKF 81297, respectively, using [11C]methyl iodide in the presence of N-ethyldiisopropylamine. Both radiotracers were purified using a semi-preparative cation exchange HPLC column. Radiochemical yields of 20-75% were obtained (from [11C]methyl iodide, decay-corrected) with a synthesis time of 30-35 min from EOB. The specific activities were 700-2500 Ci/mmol (25.9-92.5 GBq/mumol) at EOS, and the radiochemical purities were > 99%. Autoradiographic studies showed selective binding for both tracers in rat brain regions rich in D1 receptors such as the caudate-putamen, nucleus accumbens, olfactory tubercles and substantia nigra. [11C]SKF 75670 and [11C]SKF 82957 are thus potential PET radioligands for the functional high-affinity state of D1 receptors.

In vivo binding of [11C]SKF 75670 and [11C]SKF 82957 in rat brain: Two dopamine D-1 receptor agonist ligands

The high affinity benzazepine D1 agonists SKF 75670 and SKF 82957 labeled with C-11 were evaluated in vivo in rats as potential radioligands for imaging dopamine D1 receptors with positron emission tomography (PET). Their in vivo pharmacological profile revealed selective binding for both tracers in rat brain regions rich in D1 receptors such as the caudate-putamen. The more lipophilic [11C]SKF 82957 (6-chloro-[11C]SKF 75670) showed a higher brain uptake (more than 2-fold up to 30 min), higher specific uptake in the striatum and higher signal-to-noise ratio (striatum-to-cerebellum = 3.2 +/- 0.4 for [11C]SKF 75670 and 9.7 +/- 2.5 for [11C]SKF 82957 at 60 min post-injection) as compared to [11C]SKF 75670. Both radiotracers exhibited high specificity and selectivity for D1 receptors, since only D1 competitors but not the D2 antagonist sulpiride or the 5-HT2 antagonist ritanserin reduced significantly their binding the striatum with [11C]SKF 75670 or the striatum and olfactory tubercles with [11C]SKF 82957. Previous reports have shown that only D1 agonists can recognize the functional high-affinity state from the low-affinity state of D1 receptors. [11C]SKF 75670 and especially [11C]SKF 82957 are D1 agonist radioligands that can potentially be used to study in vivo the functional high-affinity state of D1 receptors using PET.

Chapter 51 - Measurement of [ 11 C]Radopride Binding Using a Bolus plus Infusion Protocol

The ratio of basal ganglia to cerebellar uptake of [ 11 C]-raclopride is often used as an index of dopamine D 2 receptor density. This chapter describes a bolus plus constant infusion technique and a data analysis approach that provides reliable estimates of the bound-to-free radioligand ratio. The technique does not require arterial blood sampling or correction for metabolites and protein binding. This method is first developed by computer simulation then it is applied to positron emission tomography studies of 12 control subjects and to 15 receptor occupancy studies. A nonlinear curve-fitting method is used to obtain the free and bound time–activity curves. Then the differential equation is solved by describing the transport of radioligand between the free and bound compartments, but without assuming equilibrium conditions. Because of this, the data analysis technique applies equally well to bolus injection studies. Computer simulations indicate that the free receptor index obtained with the method presented in this chapter is not sensitive to changes in cerebral blood flow when applied to both the bolus and the bolus plus infusion methods. The bolus plus infusion is a simple and reliable technique for clinical research.