Strategies for the labeling of halogen-substituted peroxisome proliferator-activated receptor gamma ligands: potential positron emission tomography and single photon emission computed tomography imaging agents

Well-known as an important regulator of lipid metabolism and adipocyte differentiation, the peroxisome proliferator-activated receptor gamma (PPARgamma) also has potential use as a target for antitumor therapy in certain cancers. To develop agents for radionuclide imaging PPARgamma in vivo, we synthesized fluorine, bromine, and iodine-substituted analogs (1-3) of a high-affinity benzophenone-tyrosine PPARgamma ligand; all three analogs retain very high affinity for the PPARgamma receptor. In preparation for the synthesis of these PPARgamma ligands in radiolabeled form, we have synthesized two types of precursors: (a) an aryltributylstannane (9), from which the bromine and iodine-substituted analogs (2 and 3) can readily be prepared by electrophilic destannylation, and (b) three diaryliodonium tosylate derivatives (12a-c), precursors for nucleophilic aromatic fluorination using fluoride ion. Conditions were developed whereby the thiophenyliodonium tosylate (12c) underwent nucleophilic aromatic substitution with fluoride ion, efficiently and in short reaction times, to produce the desired fluorine-substituted target compound 1. These reactions laid the groundwork for producing these three PPARgamma ligands in radiolabeled form; in addition, our use of diaryliodonium ion precursors for aromatic fluorination in this series provides an example that should encourage application of this approach for radiofluorination of more complicated radiopharmaceuticals.     

Using single photon emission tomography (SPECT) and positron emission tomography (PET) to trace the distribution of muscarinic acetylcholine receptor (MACHR) binding radioligands

Two [18F] labeled ligands for the mAChR were prepared and evaluated in rodents and nonhuman primates. The properties of both compounds, one an agonist and the other an antagonist, were consistent with M2 subtype specificity.     

Synthesis and biological activity of fluoro-substituted pyrrolo[2,3-d]pyrimidines: the development of potential positron emission tomography imaging agents for the corticotropin-releasing hormone type 1 receptor

A series of fluoro-substituted 4-(dialkylamino)pyrrolo[2,3-d]pyrimidines was synthesized and their binding affinity for corticotropin-releasing hormone type 1 receptor (CRHR1) was investigated. Compounds 11a and 11b possessed very high CRHR1 affinity (Ki=3.5, 0.91 nM, respectively). They are promising candidates for the development of 18F-containing nonpeptide PET radioligands for CRHR1.     

Synthesis and radiolabeling of 111In-core-cross linked polymeric micelle-octreotide for near-infrared fluoroscopy and single photon emission computed tomography imaging

The objective of this study was the development of a dual-modality imaging device, namely ¹¹¹In-core-cross-linked polymeric micelle (CCPM)-octreotide, for neuroendocrine tumor detection, using near-infrared fluoroscopy (NIRF) and single photon emission computed tomography (SPECT). The tumor targeting ability of the ¹¹¹In-labeled CCPM-octreotide was evaluated in a tumor mouse model. SPECT/CT, NIRF and gamma imaging results showed high tumor uptake of ¹¹¹In-labeled CCPM-octreotide. In contrast, there was a much lower signal in the same mouse model injected with ¹¹¹In-labeled CCPM. The high accumulation of ¹¹¹In-labeled CCPM-octreotide in U87 tumor was reduced after co-injection with an excess amount of CCPM-octreotide. These results suggested CCPM-octreotide’s potential applications in tumor diagnosis, drug delivery and molecular imaging.     

The baboon model under anesthesia for in vivo cerebral blood flow studies using single photon emission computed tomographic (SPECT) techniques.

Single photon emission computed tomography of the brain can be useful in animal experimentation directed toward cerebral conditions. A well established and understood baboon model, necessarily under anesthesia, could be especially valuable in such investigations. Six normal baboons were studied under various anesthetic agents and their combinations: ketamine, thiopentone, pentobarbitone, and halothane. Cerebral blood flow (CBF) studies were performed with 99mTc-HMPAO. CBF effects from various anesthesia were detected, requiring careful choice of the anesthesia for cerebral investigations.     

Synthesis and evaluation of indium-111-labeled imidazothiadiazole sulfonamide derivative for single photon emission computed tomography imaging targeting carbonic anhydrase-IX

Carbonic anhydrase-IX (CA-IX) is a zinc enzyme overexpressed in the hypoxic regions of many types of solid tumors; therefore, in vivo imaging of CA-IX may contribute to cancer diagnosis. In this study, we newly designed and synthesized an ¹¹¹In-labeled CA-IX imaging agent based on an imidazothiadiazole sulfonamide (IS) scaffold conjugated with a chelating moiety, DO3A ([¹¹¹In]DO3A-IS1), and evaluated its utility for imaging of CA-IX high-expressing tumors. [¹¹¹In]DO3A-IS1 was successfully synthesized at a 76% radiochemical yield by reacting its precursor with ¹¹¹InCl₃ in acetate buffer. In in vitro assays, [¹¹¹In]DO3A-IS1 showed marked stability in murine plasma and greater binding to CA-IX high-expressing (HT-29) cells (118 ± 21% initial dose/mg protein) than CA-IX low-expressing (MDA-MB-231) cells (1.4 ± 0.3% initial dose/mg protein). Moreover, in an in vivo biodistribution assay, [¹¹¹In]DO3A-IS1 showed marked accumulation in the HT-29 tumor (8.71 ± 1.41% injected dose/g at 24 h postinjection). In addition, in a single photon emission computed tomography (SPECT) study, [¹¹¹In]DO3A-IS1 clearly and selectively visualized the HT-29 tumor as compared with the MDA-MB-231 tumor. These results indicate that [¹¹¹In]DO3A-IS1 may serve as a useful SPECT imaging agent with the novel scaffold targeting CA-IX.     

Physiological imaging of the lung: single-photon-emission computed tomography (SPECT).

Emission tomography provides three-dimensional, quantitative images of the distribution of radiotracers used to mark physiological, metabolic, or pathological processes. Quantitative single photon emission computed tomography (SPECT) requires correction for the image-degrading effects due to photon attenuation and scatter. Phantom experiments have shown that radioactive concentrations can be assessed within some percentage of the true value when relevant corrections are applied. SPECT is widely spread, and radiotracers are available that are easy to use and comparably inexpensive. Compared with other methods, SPECT suffers from a lower spatial resolution, and the time required for image acquisition is longer than for some alternative methods. In contrast to some other methods, SPECT allows simultaneous imaging of more than one process, e.g., both regional blood flow and ventilation, for the whole lung. SPECT has been used to explore the influence of posture and clinical interventions on the spatial distribution of lung blood flow and ventilation. Lung blood flow is typically imaged using macroaggregates of albumin. Both radioactive gases and particulate aerosols labeled with radioactivity have been used for imaging of regional ventilation. However, all radiotracers are not equally suited for quantitative measurements; all have specific advantages and limitations. With SPECT, both blood flow and ventilation can be marked with radiotracers that remain fixed in the lung tissue, which allows tracer administration during conditions different from those at image registration. All SPECT methods have specific features that result from the used radiotracer, the manner in which it is administered, and how images are registered and analyzed.     

Long-term estrogen therapy and 5-HT(2A) receptor binding in postmenopausal women; a single photon emission tomography (SPET) study

Variation in estrogen level is reported by some to affect brain maturation and memory. The neurobiological basis for this may include modulation of the serotonergic system. No neuroimaging studies have directly examined the effect of extended estrogen therapy (ET), on the 5-HT(2A) receptor in human brain. We investigated the effect of long-term ET on cortical 5-HT(2A) receptor availability in postmenopausal women. In a cross-sectional study, we compared cortical 5-HT(2A) receptor availability in 17 postmenopausal ERT-naive women and 17 long-term oophorectomised estrogen-users, age- and IQ-matched using single photon emission tomography and the selective 5-HT(2A) receptor ligand (123)I-5-I-R91150. Also, we used the Revised Wechsler Memory Scale to relate memory function to 5-HT(2A) receptor availability. Never-users had significantly higher 5-HT(2A) receptor availability than estrogen-users in hippocampus (1.17 vs. 1.11, respectively, p=0.02), although this did not remain significant after correction for multiple comparisons. Hippocampal 5-HT(2A) receptor availability correlated negatively with verbal and general memory and delayed recall (r=-0.45, p=0.01; r=-0.40, p=0.02; r=-0.36, p=0.04). Right superior temporal 5-HT(2A) receptor availability correlated negatively with verbal memory (r=-0.36, p=0.04). In estrogen-users, receptor availability correlated negatively with verbal and general memory (r=-0.70, p=0.002; r=-0.69, p=0.002); and in never-users, receptor availability negatively correlated with attention and concentration (r=-0.54, p=0.02). Long-term ET may be associated with lower 5-HT(2A) receptor availability in hippocampus. This may reflect increased activity within the serotonergic pathway leading to down-regulation of post-synaptic receptor. Also, increased availability of the 5-HT(2A) receptor in hippocampus is associated with poorer memory function.     

Synthesis and binding affinity of a fluorine-substituted peroxisome proliferator-activated gamma (PPARgamma) ligand as a potential positron emission tomography (PET) imaging agent

The peroxisome proliferator-activated receptor gamma (PPARgamma) is an important regulator of lipid metabolism and the differentiation of pre-adipocytes. Thus, imaging PPARgamma in vivo using positron-emission tomography (PET) might be useful in assessing lipid metabolism disorders and identifying tumor cell differentiation. A fluorine-substituted PPARgamma ligand from tyrosine-benzophenone class, compound 1, has a very high affinity for PPARgamma receptor (Ki = 0.14 nM). To develop this compound as a PPARgamma PET imaging agent, we investigated synthetic routes suitable for its labeling with the short-lived PET radionuclide fluorine-18 (t1/2 = 110 min). To obtain the high specific activity material needed for receptor imaging with this isotope, reactions need to proceed efficiently, within a short time, starting from fluoride ion at the tracer level. The most promising approach involves introduction of fluorine into a suitable benzophenone precursor, followed by efficient coupling of this intermediate with the heterocyclic tyrosine component using a copper-catalyzed Ullmann-type condensation.     

Synthesis of [18F]-labeled (6-fluorohexyl)triphenylphosphonium cation as a potential agent for myocardial imaging using positron emission tomography

Lipophilic cations such as phosphonium salts penetrate the hydrophobic barriers of the plasma and mitochondrial membranes and accumulate in mitochondria in response to the negative inner-transmembrane potentials. Thus, as newly developed noninvasive imaging agents, [(18)F]-labeled phosphonium salts may serve as molecular "voltage sensor" probes to investigate the role of mitochondria, particularly in myocardial disease. The present study reports the radiosynthesis of (6-fluorohexyl)triphenylphosphonium salt (3) as a potential agent for myocardial imaging by using positron emission tomography (PET). The reference compound of (6-[(18)F]fluorohexyl)triphenylphosphonium salt ([(18)F]3) was synthesized with 74% yield via three-step nucleophilic substitution reactions. The reference compound was radiolabeled via two-step nucleophilic substitution reactions of no-carrier-added [(18)F]fluoride with the precursor hexane-1,6-diyl bis(4-methylbenzenesulfonate) in the presence of Kryptofix 2.2.2 and K(2)CO(3). The radiolabeled compound was synthesized with 15-20% yield. The radiochemical purity was >98% by analytical HPLC, and the specific activity was >6.10-6.47 TBq/μmol. The cellular uptake assay showed preferential uptake of [(18)F]3 in cardiomyocytes. The results of biodistribution and micro-PET imaging studies of [(18)F]3 in mice and rats showed preferential accumulation in the myocardium. The results suggest that this compound would be a promising candidate for myocardial imaging.     

A new single-photon emission computed tomography (SPECT) imaging agent for serotonin transporters: [125I]Flip-IDAM, (2-((2-((dimethylamino)methyl)-4-iodophenyl)thio)phenyl)methanol

New ligands for in vivo brain imaging of serotonin transporter (SERT) with single photon emission tomography (SPECT) were prepared and evaluated. An efficient synthesis and radiolabeling of a biphenylthiol, FLIP-IDAM, 4, was accomplished. The affinity of FLIP-IDAM was evaluated by an in vitro inhibitory binding assay using [¹²⁵I]-IDAM as radioligand in rat brain tissue homogenates (K_i = 0.03 nM). New [¹²⁵I]Flip-IDAM exhibited excellent binding affinity to SERT binding sites with a high hypothalamus to cerebellum ratio of 4 at 30 min post iv injection. The faster in vivo kinetics for brain uptake and a rapid washout from non-specific regions provide excellent signal to noise ratio. This new agent, when labeled with ¹²³I, may be a useful imaging agent for mapping SERT binding sites in the human brain.     

18F Labeled benzimidazole derivatives as potential radiotracer for positron emission tomography (PET) tumor imaging

This article reported the synthesis and bioevaluation of two [¹⁸F] labeled benzimidazole derivatives, 4-(5-(2-[¹⁸F] fluoro-4-nitrobenzamido)-1-methyl-1H-benzimidazol-2-yl) butanoic acid ([¹⁸F] FNBMBBA, [¹⁸F]a1) and 3-(2-fluoroethyl)-7-methyl-2-propyl-3H-benzimidazole-5-carboxylic acid ([¹⁸F] FEMPBBA, [¹⁸F]b1) for PET tumor imaging. The preparation [¹⁸F] FEMPBBA was completed in 1 h with overall radiochemical yield of 50–60% (without decay corrected). Biodistribution assay in S180 tumor bearing mice of both compounds were carried out, and the results are both meaningful. [¹⁸F] FEMPBBA which can be taken as a revision of [¹⁸F] FNBMBBA got an excellent result, and has significant advantages in some aspects compared with L-[¹⁸F] FET and [¹⁸F]-FDG in the same animal model, especially in tumor/brain uptake ratio. The tumor/brain uptake ratio of [¹⁸F] FEMPBBA gets to 4.81, 7.15, and 9.8 at 30 min, 60 min and 120 min, and is much higher than that of L-[¹⁸F] FET (2.54, 2.92 and 2.95) and [¹⁸F]-FDG (0.61, 1.02, 1.33) at the same time point. The tumor/muscle and tumor/blood uptake ratio of [¹⁸F] FEMPBBA is also higher than that of L-[¹⁸F] FET at 30 min and 60 min. This result indicates compound [¹⁸F] FEMPBBA is a promising radiotracer for PET tumor imaging.     

Design and docking studies of [diethylenetriaminepentaacetic acid-(amino acid)2] with acetylcholine receptor as a molecular imaging agent for single-photon emission computed tomographic application

The acetylcholine receptor is an essential link between the brain and the muscles, so it is a sensitive location for attack. In this study, some reversible [diethylenetriaminepentaacetic acid-(amino acid)2] have been docked computationally to the active site of the acetylcholine receptor. The induced fit method was employed to perform the automolecular docking for these systems. The result of docking studies generated thermodynamic properties, such as free energy of bindings (Glide score) and their weak electrostatic interactions. On the basis of these results, scintigraphic imaging studies were performed in mice. Among the radiotracers evaluated in this study, compound derived from 5-hydroxytryptophan/tryptophan exhibited remarkable localization in the brain, whereas radiotracer derived from l-histidine shows moderate accumulation in the brain. Preliminary studies with these amino acid-based ligands are encouraging to carrying out further in vivo experiments for targeted imaging.     

CRHR1 Receptor binding and lipophilicity of pyrrolopyrimidines, potential nonpeptide corticotropin-releasing hormone type 1 receptor antagonists.

A series of compounds related to N-butyl-N-ethyl[2,5,6-trimethyl-7-(2,4,6-trimethylphenyl)pyrrolo[2,3-d]pyrimidin-4-yl]amine (1, antalarmin) have been prepared and evaluated for their CRHR1 binding affinity as the initial step in the development of selective high affinity hydrophilic nonpeptide corticotropin-releasing hormone type 1 receptor (CRHR1) antagonists. Calculated log P (Clog P) values were used to evaluate the rank order of hydrophilicity for these analogues. Introducing oxygenated functionalities (delta-hydroxy or bis-beta-ethereal) into 1 gave more hydrophilic compounds, which had good affinity for the receptor. Introducing an amino group or shortening the alkyl side chain was detrimental to CRHR1 affinity. The alcohol 4-[ethyl[2,5,6-trimethyl-7-(2,4,6-trimethylphenyl)pyrrolo[2,3-d]pyrimidin-4-yl]amino]butan-1-ol (3), bearing a terminal hydroxyl group on an N-alkyl side-chain, showed the highest CRHR1 binding affinity among these compounds (K(i)=0.68 nM), and is one of the highest affinity CRHR1 ligands known. Compounds 3-5, and 8, which are likely to be less lipophilic than 1, have high CRHR1 affinity and may be valuable probes to further study the CRH system.     

Human stresscopin and stresscopin-related peptide are selective ligands for the type 2 corticotropin-releasing hormone receptor

Adaptive stress responses mediated by the endocrine, autonomic, cardiovascular and immune systems are essential for the survival of the individual. Initial stress-induced responses provide a vital short-term metabolic lift, but prolonged or inappropriate exposure to stress can compromise homeostasis thereby leading to disease. This 'fight-or-flight' response is characterized by the activation of the corticotropin-releasing hormone (CRH)-adrenocorticotropin-glucocorticoid axis, mediated by the type 1 CRH receptor. In contrast, the type 2 CRH receptor mediates the stress-coping responses during the recovery phase of stress. We identified human stresscopin (SCP) and stresscopin-related peptide (SRP) as specific ligands for the type 2 CRH receptor. The genes encoding these peptides were expressed in diverse peripheral tissues as well as in the central nervous system. Treatment with SCP or SRP suppressed food intake, delayed gastric emptying and decreased heat-induced edema. Thus SCP and SRP might represent endogenous ligands for maintaining homeostasis after stress, and could allow the design of drugs to ameliorate stress-related diseases.