Synthesis of high-specific-radioactivity 4- and 6-[18F]fluorometaraminol-PET tracers for the adrenergic nervous system of the heart.

Fluorine-18- (t(1/2) 109.8 min) and carbon-11 (t(1/2) 20.4 min)-labeled norepinephrine analogues have been found previously to be useful positron-emission-tomography (PET) radioligands to map adrenergic nerve terminals of the heart. Metaraminol ((1R,2S)-2-amino-1-(3-hydroxyphenyl)-1-propanol) is a metabolically stable structural analogue of norepinephrine and possesses high affinity towards the norepinephrine transporter and the vesicular monoamine transporter. This paper presents the radiosynthesis of new positron-emission-tomography halogeno analogues of metaraminol labeled with high specific radioactivity. Firstly, fluorine-18-labeled 4-fluorometaraminol (4-[18F]FMR or (1R,2S)-2-amino-1-(4-[18F]fluoro-3-hydroxyphenyl)-1-propanol) and its three other stereoisomers were prepared based on the following key steps: (a) condensation of the corresponding no-carrier-added labeled fluorobenzaldehyde with nitroethane, and (b) HPLC (C18 and chiral) resolution of the diastereomeric product mixture into the four individual enantiomers. Secondly, the corresponding 6-fluoro analogues, fluorine-18-labeled 6-fluorometaraminol (6-[18F]FMR or (1R,2S)-2-amino-1-(2-[18F]fluoro-5-hydroxyphenyl)-1-propanol) and its three other enantiomers, were prepared in an analogous way. Typically, 0.48-0.55 GBq of 4-[18F]FMR and 0.14-0.15 GBq of 6-[18F]FMR could be obtained after 120-160 min total synthesis time, with a specific radioactivity of 56-106 GBq/micromol. Furthermore, the synthesis of racemic 4-fluorometaraminol and 6-fluorometaraminol as reference compounds was performed. as well as independent chiral syntheses of the optically active (1R,2S) enantiomers. For the chiral syntheses, the key step was an electrophilic fluorination with acetyl hypofluorite of (1R,2S)-configurated organometallic derivatives of metaraminol. Tissue distribution studies in rats suggested that both 4-[18F]FMR and 6-[18F]FMR display similar affinity towards the presynaptic adrenergic nerve terminal in the heart. From a practical point of view, 4-[18F]FMR appeared to be the more attractive candidate for future PET investigations, due to higher radiochemical yields.

Semiselective POCE NMR spectroscopy.

A new scheme is proposed to edit separately glutamate C(3) and C(4) resonances of (1)H bound to (13)C, in order to resolve these two signals which overlap at intermediate magnetic fields (1.5 T-3 T), commonly available for human brain studies. The two edited spectra are obtained by combining the individual acquisitions from a four-scan measurement in two different ways. The four acquisitions correspond to the two steps of the classical POCE scheme combined with another two-scan module, where the relative phases of the C(3) and C(4) (1)H resonances are manipulated using zero quantum and double quantum coherence pathways. This new technique exhibits the same sensitivity as POCE and allows the (13)C labeling of C(3) and C(4) glutamate from [1-(13)C]glucose to be monitored separately in the rat brain at 3 T.

General method to label antisense oligonucleotides with radioactive halogens for pharmacological and imaging studies.

Evaluation of oligonucleotides for biomedical applications requires different in vivo and in vitro approaches (pharmacokinetics, biodistribution, macro- and microimaging, metabolism,.), that are performed with different radioisotopes according to the temporal and spatial resolution needed. A method to introduce radioactive isotopes of halogens (fluorine, bromine, and iodine) in a small and stable molecule has been developed. Radiosynthons can then be conjugated with any given oligonucleotide in one step to create the appropriate radiotracer. This general radiolabeling procedure for oligonucleotides is efficient to synthesize (18)F-, (76)Br-, and (125)I-oligonucleotides for biological needs. Applications of the method to biodistribution, metabolism, in vivo and ex vivo imaging of (125)I- and (18)F-labeled oligonucleotides are reported.

Early N-acetylaspartate depletion is a marker of neuronal dysfunction in rats and primates chronically treated with the mitochondrial toxin 3-nitropropionic acid.

N-acetylaspartate (NAA) quantification by 1H-magnetic resonance spectroscopy has been commonly used to assess in vivo neuronal loss in neurodegenerative disorders. Here. the authors used ex vivo and in vivo 1H-magnetic resonance spectroscopy in rat and primate models of progressive striatal degeneration induced by the mitochondrial toxin 3-nitropropionate (3NP) to determine whether early NAA depletions could also be associated with neuronal dysfunction. In rats that were treated for 3 days with 3NP and had motor symptoms, the authors found a significant decrease in NAA concentrations, specifically restricted to the striatum. No cell loss or dying cells were found at this stage in these animals. After 5 days of 3NP treatment, a further decrease in striatal NAA concentrations was observed in association with the occurrence of dying neurons in the dorsolateral striatum. In 3NP-treated primates, a similar striatal-selective and early decrease in NAA concentrations was observed after only a few weeks of neurotoxic treatment, without any sign of ongoing cell death. This early decrease in striatal NAA was partially reversed after 4 weeks of 3NP withdrawal. These results demonstrate that early NAA depletions reflect a reversible state of neuronal dysfunction preceding cell degeneration and suggest that in vivo quantification of NAA 1H-magnetic resonance spectroscopy may become a valuable tool for assessing early neuronal dysfunction and the effects of potential neuroprotective therapies in neurodegenerative disorders.

Carbon-11 epidepride: a suitable radioligand for PET investigation of striatal and extrastriatal dopamine D2 receptors.

Epidepride [(S)-(-)-N-([1-ethyl-2-pyrrolidinyl]methyl)-5-iodo-2,3-dimethoxybenza mide] binds with a picomolar affinity (Ki = 24 pM) to the dopamine D2 receptor. Iodine-123-labeled epidepride has been used previously to study striatal and extrastriatal dopamine D2 receptors with single photon emission computed tomography (SPECT). Our aim was to label epidepride with carbon-11 for comparative quantitative studies between positron emission tomography (PET) and SPECT. Epidepride was synthesized from its bromo-analogue FLB 457 via the corresponding trimethyl-tin derivative. In an alternative synthetic pathway, the corresponding substituted benzoic acid was reacted with the optically pure aminomethylpyrrolidine-derivative. Demethylation of epidepride gave the desmethyl-derivative, which was reacted with [11C]methyl triflate. Total radiochemical yield was 40-50% within a total synthesis time of 30 min. The specific radioactivity at the end of synthesis was 37-111 GBq/micromol (1,000-3,000 Ci/mmol). Human postmortem whole-hemisphere autoradiography demonstrated dense binding in the caudate putamen, and also in extrastriatal areas such as the thalamus and the neocortex. The binding was inhibited by unlabeled raclopride. PET studies in a cynomolgus monkey demonstrated high uptake in the striatum and in several extrastriatal regions. At 90 min after injection, uptake in the striatum, thalamus and neocortex was about 11, 4, and 2 times higher than in the cerebellum, respectively. Pretreatment experiment with unlabeled raclopride (1 mg/kg) inhibited 50-70% of [11C]epidepride binding. The fraction of unchanged [11C]epidepride in monkey plasma determined by a gradient high performance liquid chromatography (HPLC) method was about 30% of the total radioactivity at 30 min after injection of [11C]epidepride. The availability of [11C]epidepride allows the PET-verification of the data obtained from quantitation studies with SPECT.

Synthesis and nicotinic acetylcholine receptor in vivo binding properties of 2-fluoro-3-[2(S)-2-azetidinylmethoxy]pyridine: a new positron emission tomography ligand for nicotinic receptors.

The lead compound of a new series of 3-pyridyl ethers, the azetidine derivative A-85380 (3-[(S)-2-azetidinylmethoxy]pyridine), is a potent and selective ligand for the human alpha4beta2 nicotinic acetylcholine receptor (nAChR) subtype. In vitro, the fluoro derivative of A-85380 (2-fluoro-3-[(S)-2-azetidinylmethoxy]pyridine or F-A-85380) competitively displaced [3H]cytisine or [3H]epibatidine with Ki values of 48 and 46 pM, respectively. F-A-85380 has been labeled with the positron emitter fluorine-18 (t1/2 (half-life) = 110 min) by no-carrier-added nucleophilic aromatic substitution by K[18F]F-K222 complex with (3-[2(S)-N-(tert-butoxycarbonyl)-2-azetidinylmethoxy]pyridin-2-yl) tri methylammonium trifluoromethanesulfonate as a highly efficient labeling precursor, followed by TFA removal of the Boc protective group. The total synthesis time was 50-53 min from the end of cyclotron fluorine-18 production (EOB). Radiochemical yields, with respect to initial [18F]fluoride ion radioactivity, were 68-72% (decay-corrected) and 49-52% (non-decay-corrected), and the specific radioactivities at EOB were 4-7 Ci/micromol (148-259 GBq/micromol). In vivo characterization of [18F]F-A-85380 showed promising properties for PET imaging of central nAChRs. This compound does not bind in vivo to alpha7 nicotinic or 5HT3 receptors. Moreover, its cerebral uptake can be modulated by the synaptic concentration of the endogenous ligand acetylcholine. The preliminary PET experiments in baboons with [18F]F-A-85380 show an accumulation of the radiotracer in the brain within 60 min. In the thalamus, a nAChR-rich area, uptake of radioactivity reached a maximum at 60 min (4% I.D./100 mL of tissue). [18F]F-A-85380 appears to be a suitable radioligand for brain imaging nAChRs with PET.

Synthesis of a fluorine-18 labeled derivative of epibatidine for in vivo nicotinic acetylcholine receptor PET imaging.

Epibatidine (exo-2-(2'-chloro-5'-pyridyl)-7-azabicyclo[2.2.1]heptane), a natural compound isolated from the skin of the Ecuadorian poison frog Epipedobates tricolor, is the most potent nicotinic acetylcholine receptor (nAChR) agonist reported to date. In order to visualize and quantify in vivo these receptors in human brain using Positron Emission Tomography (PET), [18F]norchlorofluoroepibatidine (exo-2-(2'-[18F]fluoro-5'-pyridyl)-7-azabicyclo[2.2.1]heptane), a fluorine-18 (t(1/2): 110 min) radiolabeled derivative of epibatidine has been designed. The corresponding 2'-bromo-, 2'-iodo- and 2'-nitro exo-2-(5'-pyridyl)-7-azabicyclo[2.2.1]heptane analogues as labeling precursors, as well as norchlorofluoroepibatidine as a reference compound have been synthesized by reductive, stereoselective, palladium-catalyzed Heck-type coupling between an N-Boc protected azanorbornene and the corresponding halopyridine. [18F]Norchlorofluoroepibatidine has been radiolabeled with fluorine-18 by nucleophilic aromatic substitution from the corresponding Boc-protected halo- and nitro precursors using [18F]FK-K222 complex in DMSO by conventional heating (at 150-180 degrees C for 10 min) or microwave activations (at 100 Watt, for 1 to 2.5 min), followed by TFA-removal of the protective group. Typically, using the microwave activation procedure, 60-80 mCi (2.22-2.96 GBq) of pure [18F]norchlorofluoroepibatidine could be obtained in less than 2 h (110-115 min) from the bromo labeling precursor, with specific radioactivities of 1.5-2.5 Ci/micromol (55.5-92.5 GBq/micromol) calculated for End of Bombardment. The preliminary PET experiments in baboon (Papio papio) with [18F]norchlorofluoroepibatidine show a high uptake and a rapid accumulation of the radiotracer into the brain within 30 min. In the thalamus, a nAChR rich area, uptake of radioactivity reached a maximum at 40 min (10% I.D./100 mL tissue). The ratio of radioactivity thalamus/cerebellum (the latter being a nAChR poor area) was 2 at 40 min and increased with time, up to 4.3 at 160 min. Its specific regiodistribution and its high ratio of specific-to-nonspecific binding confirm the ideal profile of [18F]norchlorofluoroepibatidine as a suitable radioligand for PET imaging of nAChRs in the brain.

Magnetization transfer characteristics in atherosclerotic plaque components assessed by adapted binomial preparation pulses.

Increasing the contrast between atheromatous plaque components is a major issue in cardiovascular MRI research. It would allow one to identify unstable plaque by differentiating the lipid core associated with vulnerability, from the fibrous cap, considered as a factor of stability. T2 and diffusion-weighted imaging have already provided satisfying results. Magnetization transfer (MT) between restricted protons Hr and free-water protons Hf could achieve a different contrast related to collagen and lipoprotein macromolecules present in the fibrous cap and lipid core, respectively. The purpose of this work was to evaluate in vitro the MT effect produced by adapted T2-selective 1-3-3-1 binomial pulses on isolated samples of atheromatous arteries at 3 T. A method based on simulation was used in order to improve the MT specificity: it is shown that 50% 1-3-3-1 pulses (the percentage indicating the level of Hr saturation) allow an estimation of T2r, the Hr T2. Using this technique, magnetization transfer was observed for the first time in atherosclerotic plaque components, an effect more pronounced for the fibrous cap and media than for the lipid core and adventitia. The T2r estimation gave values ranging from 20 to 25 micros for the four samples. This preliminary study provides a basis for establishing an MT imaging sequence of atheromatous arteries, by using 50% 1-3-3-1 pulses calibrated for saturating protons with a 20 micros T2. This MT protocol should be further compared to T2 and diffusion-weighted imaging.