Patrick J. Riss

CuI-Catalyzed 11C Carboxylation of Boronic Acid Esters: A Rapid and Convenient Entry to 11C-Labeled Carboxylic Acids, Esters, and Amides†

Rapid and direct: the carboxylation of boronic acid esters with (11)CO(2) provides [(11)C]carboxylic acids as a convenient entry into [(11)C]esters and [(11)C]amides. This conversion of boronates is tolerant to diverse functional groups (e.g., halo, nitro, or carbonyl).

Applications of positron emission tomography in animal models of neurological and neuropsychiatric disorders

Positron emission tomography (PET) provides dynamic images of the biodistribution of radioactive tracers in the brain. Through application of the principles of compartmental analysis, tracer uptake can be quantified in terms of specific physiological processes such as cerebral blood flow, cerebral metabolic rate, and the availability of receptors in brain. Whereas early PET studies in animal models of brain diseases were hampered by the limited spatial resolution of PET instruments, dedicated small-animal instruments now provide molecular images of rodent brain with resolution approaching 1mm, the theoretic limit of the method. Major applications of PET for brain research have consisted of studies of animal models of neurological disorders, notably Parkinsons disease (PD), Alzheimers disease (AD), and Huntingtons disease (HD), stroke, epilepsy and traumatic brain injury; these studies have particularly benefited from selective neurochemical lesion models (PD), and also transgenic rodent models (AD, HD). Due to their complex and uncertain pathophysiologies, corresponding models of neuropsychiatric disorders have proven more difficult to establish. Historically, there has been an emphasis on PET studies of dopamine transmission, as assessed with a range of tracers targeting dopamine synthesis, plasma membrane transporters, and receptor binding sites. However, notable recent breakthroughs in molecular imaging include the development of greatly improved tracers for subtypes of serotonin, cannabinoid, and metabotropic glutamate receptors, as well as noradrenaline transporters, amyloid-β and neuroinflammatory changes. This article reviews the considerable recent progress in preclinical PET and discusses applications relevant to a number of neurological and neuropsychiatric disorders in humans.

Synthesis and Initial in Vivo Studies with [11C]SB-216763: The First Radiolabeled Brain Penetrative Inhibitor of GSK-3

Quantifying glycogen synthase kinase-3 (GSK-3) activity in vivo using positron emission tomography (PET) imaging is of interest because dysregulation of GSK-3 is implicated in numerous diseases and neurological disorders for which GSK-3 inhibitors are being considered as therapeutic strategies. Previous PET radiotracers for GSK-3 have been reported, but none of the published examples cross the blood-brain barrier. Therefore, we have an ongoing interest in developing a brain penetrating radiotracer for GSK-3. To this end, we were interested in synthesis and preclinical evaluation of [(11)C]SB-216763, a high-affinity inhibitor of GSK-3 (K i = 9 nM; IC50 = 34 nM). Initial radiosyntheses of [(11)C]SB-216763 proved ineffective in our hands because of competing [3 + 3] sigmatropic shifts. Therefore, we have developed a novel one-pot two-step synthesis of [(11)C]SB-216763 from a 2,4-dimethoxybenzyl-protected maleimide precursor, which provided high specific activity [(11)C]SB-216763 in 1% noncorrected radiochemical yield (based upon [(11)C]CH3I) and 97-100% radiochemical purity (n = 7). Initial preclinical evaluation in rodent and nonhuman primate PET imaging studies revealed high initial brain uptake (peak rodent SUV = 2.5 @ 3 min postinjection; peak nonhuman primate SUV = 1.9 @ 5 min postinjection) followed by washout. Brain uptake was highest in thalamus, striatum, cortex, and cerebellum, areas known to be rich in GSK-3. These results make the arylindolemaleimide skeleton our lead scaffold for developing a PET radiotracer for quantification of GSK-3 density in vivo and ultimately translating it into clinical use.

Radiolabelling and preliminary evaluation of 68Ga-tetrapyrrole derivatives as potential tracers for PET.

Tetrapyrroles are multisided natural products which are of relevance in clinical medicine. Owing to their specific accumulation in tumour tissue, porphyrins, metalloporphyrins and chlorins have been used as in photodynamic therapy and optical imaging. Moreover, their specific uptake into inflammatory atheromatous plaques via LDL endocytosis has been reported. The present study is concerned with the synthesis of (68)Ga labelled porphyrin derivatives and an in vitro assessment of the utility of radiotracers in positron emission tomography. A set of five porphyrin derivatives were labelled using (68)Ga from a commercially obtained radionuclide generator. Dedicated post-processing of the generator eluate was conducted to allow for labelling in aqueous media and also under anhydrous conditions. Challenge studies and incubation in human serum confirmed the stability of the tracers. Plasma protein binding was investigated in order to confirm the presence of freely diffusible radioligand in plasma. A preliminary microPET study in a tumour-bearing rat resulted in a clear visualisation of the tumour.

A methodical 68Ga-labelling study of DO2A-(butyl-L-tyrosine)2 with cation-exchanger post-processed 68Ga: practical aspects of radiolabelling.

Positron emission tomography (PET) with (68)Ga is a fast-growing field in molecular imaging, both in research and in clinical routine. The availability of (68)Ga via the (68)Ge/(68)Ga radionuclide generator facilitates the development and production of radiopharmaceuticals independent of a cyclotron. The presented work shows a complete (68) Ga labelling study exemplified on [(68)Ga]DO2A-(butyl-L-tyrosine)(2), a potential tumour tracer for PET. A methodical sequence is followed to optimize the (68)Ga-labelling reaction. Practical aspects are described and the different parameters contributing to the labelling yield are demonstrated. The influence of temperature, time, amount of labelling precursor and pH value on the radiochemical yields is demonstrated. A conventional heating method is compared with microwave irradiation as an alternative labelling method. Finally, purification of (68)Ga-labelled compounds via solid-phase extraction and quality control is shown. The procedure described in this manuscript may serve as a guideline for optimizing (68)Ga labelling reactions.

Radiosynthesis and characterization of astemizole derivatives as lead compounds toward PET imaging of τ-pathology

Formation of neurofibrillary tangles, comprising of microtubule-associated tau protein, is a hallmark of a group of neurodegenerative diseases, including Alzheimers disease. In consequence, in vivo imaging of neurofibrillary tangles is a current focus of positron emission tomography research. Herein, development of an in vitro radioligand binding assay which uses synthetic aggregates as a model of neurofibrillary tangles is reported, together with evaluation of novel derivatives of the tau protein ligand astemizole.

Validation and quantification of [18F]altanserin binding in the rat brain using blood input and reference tissue modeling.

The 5-hydroxytryptamine type 2a (5-HT2A) selective radiotracer [18F]altanserin has been subjected to a quantitative micro-positron emission tomography study in Lister Hooded rats. Metabolite-corrected plasma input modeling was compared with reference tissue modeling using the cerebellum as reference tissue. [18F]altanserin showed sufficient brain uptake in a distribution pattern consistent with the known distribution of 5-HT2A receptors. Full binding saturation and displacement was documented, and no significant uptake of radioactive metabolites was detected in the brain. Blood input as well as reference tissue models were equally appropriate to describe the radiotracer kinetics. [18F]altanserin is suitable for quantification of 5-HT2A receptor availability in rats.

Synthesis and Evaluation of 18F-FE-PEO in Rodents: An 18F-Labeled Full Agonist for Opioid Receptor Imaging

We have investigated the opioid receptor (OR) agonist (20R)-4,5-α-epoxy-6-(2-18F-fluoroethoxy)-3-hydroxy-α,17-dimethyl-α-(2-phenyleth-1-yl)-6,14-ethenomorphinan-7-methanol (18F-FE-PEO) as a candidate OR PET ligand. This tracer is attractive because it combines 18F labeling, is suited to the slow kinetics of high-affinity ligands, and has agonist binding, which has been shown to be more sensitive to changes in OR occupation than is antagonist binding. Methods: Agonist potency and off-target binding were investigated in vitro, and autoradiographic studies on rat brain sections were used to assess binding patterns. Quantification of the tracer in vivo was investigated using small-animal PET in rats with blood sampling. Results: 18F-FE-PEO was obtained by direct nucleophilic radiofluorination and subsequent deprotection with a yield of 28% ± 15%, a specific activity of 52–224 MBq/nmol, and a radiochemical purity of more than 97% (90 min from end of bombardment). In vitro studies showed it to be a full agonist ligand, which selectively binds to OR with high affinity, although it is not selective to a single OR subtype (inhibition constant, 0.4–1.6 nM across OR subtypes). Autoradiography binding patterns were consistent with the known distribution of OR, although nondisplaceable signal typically constituted one third of the signal in OR-dense regions. Although metabolites were present in blood (∼40% of plasma radioactivity was nonparent 3 h after injection), no significant metabolite fraction was found in brain tissue, aiding PET quantification. A plasma input 2-tissue-compartment model provided good fits to the PET data, and regional distribution volumes from the latter correlated well with those from Logan plot analysis (r2 = 0.98). The cerebellum had the lowest distribution volume, but the time–activity curve data could not be adequately fitted with a 1-tissue-compartment model. Reference tissue models using the cerebellum as the reference region did not provide good fits to the data, so blood-based kinetic analysis is recommended. Conclusion: As the first 18F-labeled OR agonist ligand, 18F-FE-PEO is a useful addition to the existing OR ligand portfolio.

Preclinical Aspects of Nicotinic Acetylcholine Receptor Imaging

Recent developments in radiochemistry have opened new vistas for investigations of nicotinergic acetylcholine receptors (nAChRs) in living brain by positron emission tomography (PET) and by single photon emission computed tomography (SPECT). In parallel, dedicated instrumentation for molecular imaging in small animals has facilitated preclinical investigations in a number of models in which perturbations in nAChR signalling are implicated, notably Alzheimer’s disease and other neurodegenerative conditions, schizophrenia and other neuropsychiatric disorders, substance abuse and traumatic brain injury. The nAChRs are members of a family of ligand-gated ion channels composed of five subunits, most commonly occurring in the central nervous system as heteropentamers designated α4β2, with lesser amounts of the α7 homopentamer. We present a systematic review of preclinical findings with the diverse nAChR ligands which have been investigated to date. Molecular imaging of the α4β2 nAChR subtype by PET has been successfully achieved by 2-[18F]fluoro-A-85380. Newer agents such as (−)-[18F]flubatine permit quantitation of α4β2 receptors with PET recordings not exceeding 90 min, without the toxicity characteristic of earlier epibatidine derivatives. Imaging studies of α7 nAChRs have been hampered by inadequate pharmacological specificity of available ligands and by the low natural abundance of this receptor subtype in the brain. However, a continued search for optimal ligands is justified by the particular association of α7 nAChRs with aspects of cognitive function. We note that no molecular imaging ligands have been developed for α6-containing nAChRs, despite their importance for the psychopharmacology of nicotine actions in the basal ganglia. Finally, we review the competitive binding model, in which the availability of α4β2 binding sites is altered by competition from endogenous acetylcholine, noting that this approach has yet to be applied for monitoring acetylcholine release in disease models.

The DAT Ligand [18F]PR17.MZ Mirrors the in vivo Pharmacokinetic Profile of [11C]Cocaine with Significantly Improved Monoamine Transporter Selectivity

Positron emission tomography (PET) studies of the availability of the dopamine transporter (DAT) provide valuable insight into the presynaptic integrity of dopaminergic neurons in vivo. Noninvasive PET imaging thereby contributes to the understanding of basic neuronal mechanisms in psychiatric diseases and to the routine diagnosis of movement disorders. The development of appropriate positron-emitter-labeled DAT ligands, however, has been complicated by nonspecific binding, low selectivity for the DAT, and slow binding equilibria. Highly specific and selective DAT inhibitors are prone to slow accumulation in DAT-containing brain regions and slow binding equilibrium. These characteristics lead to exhaustively longlasting PET scans. Conversely, the nonspecific monoamine transporter inhibiting ligand ( )-cocaine has a rapid pharmacokinetic profile and a fast binding equilibrium, resulting in much shorter PET scans. Unfortunately, it suffers from low affinity and a nonselective binding profile with similar affinity to dopamine, serotonin, and noradrenalin transporters. In addition, the corresponding PET ligand ( )-N-[C]cocaine is only available in PET centers equipped with an in-house cyclotron, owing to the isotope’s short half-life of 20.3 min. Ideally, a DAT imaging agent for PET would combine moderate affinity, high DAT selectivity, low nonspecific binding, rapid accumulation in the DAT-containing brain regions followed by fast washout, and the advantageous half-life of a fluorine-18 label. We designed and synthesized a series of cocaine analogues with improved affinity and selectivity profiles relative to ( )-cocaine. These compounds generally allow labeling with carbon-11 as well as fluorine-18. Herein we report our discovery of a potent and selective F-labeled DAT ligand that provides cocaine-like pharmacokinetic properties. PR17.MZ, (1R,2S,3S,5S)-methyl-8-{[(1S,2S)-2-(fluoromethyl)cyclopropyl]methyl}-3-phenyl-8-azabicyclo[3.2.1]octane-2-carboxylate (1) (Figure 1) has been designed through the use of a conformational restriction-based approach. 5] Compound 1 provides a moderate inhibition potency of human DAT (hDAT; IC50 = 11 nm) which is 29-fold higher than that of ( )-cocaine (2). Its inhibition selectivity over the human serotonin transporter (SERT; IC50 = 1.4 mm) is 67-fold improved, whereas its inhibition selectivity over the human noradrenalin transporter (NET; IC50 = 175 nm) is 28-fold improved relative to 2. [3] A labeling precursor for direct nucleophilic radiofluorination was synthesized from (1S,2S)-cyclopropane-1,2-diyldimethanol (3) in five consecutive steps as shown in Scheme 1. 7] Desymmetrization of 3 with sodium hydride and p-methoxybenzyl chloride in N,N-dimethylformamide gave compound 4 in 92 % Figure 1. Molecular structures of PR17.MZ (1) and ( )-cocaine (2).