Evaluation of transporter-mediated hepatobiliary transport of newly developed 18F-labeled pitavastatin derivative,PTV-F1, in rats by PET imaging

Quantitative evaluations of the functions of uptake and efflux transporters directly in vivo is desired to understand an efficient hepatobiliary transport of substrate drugs. Pitavastatin is a substrate of organic anion transporting polypeptides (OATPs) and canalicular efflux transporters; thus, it can be a suitable probe for positron-emission tomography (PET) imaging of hepatic transporter functions. To characterize the performance of [¹⁸F]PTV-F1, an analogue of pitavastatin, we investigated the impact of rifampicin (a typical OATP inhibitor) coadministration or Bcrp (breast cancer resistance protein) knockout on [¹⁸F]PTV-F1 hepatic uptake and efflux in rats by PET imaging. After intravenous administration, [¹⁸F]PTV-F1 selectively accumulated in the liver, and the radioactivity detected in plasma, liver, and bile mainly derived from the parent PTV-F1 during the PET study (∼40 min). Coadministration of rifampicin largely decreased the hepatic uptake of [¹⁸F]PTV-F1 by 73%. Because of its lower clearance in rats, [¹⁸F]PTV-F1 is more sensitive for monitoring changes in hepatic OATP1B function that other previously reported OATP1B PET probes. Rifampicin coadministration also significantly decreased the biliary excretion of radioactivity by 65%. Bcrp knockout did not show a significant impact on its biliary excretion.[¹⁸F]PTV-F1 enables quantitative analysis of the hepatobiliary transport system for organic anions.     

Development of a potential PET probe for HDAC6 imaging in Alzheimer's disease

Although the epigenetic regulatory protein histone deacetylase 6 (HDAC6) has been recently implicated in the etiology of Alzheimer's disease (AD), little is known about the role of HDAC6 in the etiopathogenesis of AD and whether HDAC6 can be a potential therapeutic target for AD. Here, we performed positron emission tomography (PET) imaging in combination with histopathological analysis to better understand the underlying pathomechanisms of HDAC6 in AD. We first developed [¹⁸F]PB118 which was demonstrated as a valid HDAC6 radioligand with excellent brain penetration and high specificity to HDAC6. PET studies of [¹⁸F]PB118 in 5xFAD mice showed significantly increased radioactivity in the brain compared to WT animals, with more pronounced changes identified in the cortex and hippocampus. The translatability of this radiotracer for AD in a potential human use was supported by additional studies, including similar uptake profiles in non-human primates, an increase of HDAC6 in AD-related human postmortem hippocampal tissues by Western blotting protein analysis, and our ex vivo histopathological analysis of HDAC6 in postmortem brain tissues of our animals. Collectively, our findings show that HDAC6 may lead to AD by mechanisms that tend to affect brain regions particularly susceptible to AD through an association with amyloid pathology.     

Microfluidic radiosynthesis and biodistribution of [18F] 2‐(5‐fluoro‐pentyl)‐2‐methyl malonic acid

Microfluidics technology has emerged as a powerful tool for the radiosynthesis of positron emission tomography (PET) and single‐photon emission computed tomography radiolabeled compounds. In this work, we have exploited a continuous flow microfluidic system (Advion, Inc., USA) for the [¹⁸F]‐fluorine radiolabeling of the malonic acid derivative, [¹⁸F] 2‐(5‐fluoro‐pentyl)‐2‐methyl malonic acid ([¹⁸F]‐FPMA), also known as [¹⁸F]‐ML‐10, a radiotracer proposed as a potential apoptosis PET imaging agent. The radiosynthesis was developed using a new tosylated precursor. Radiofluorination was initially optimized by manual synthesis and served as a basis to optimize reaction parameters for the microfluidic radiosynthesis. Under optimized conditions, radio‐thin‐layer chromatography analysis showed 79% [¹⁸F]‐fluorine incorporation prior to hydrolysis and purification. Following hydrolysis, the [¹⁸F]‐FPMA was purified by C18 Sep‐Pak, and the final product was analyzed by radio‐HPLC (high‐performance liquid chromatography). This resulted in a decay‐corrected 60% radiochemical yield and ≥98% radiochemical purity. Biodistribution data demonstrated rapid blood clearance with less than 2% of intact [¹⁸F]‐FPMA radioactivity remaining in the circulation 60 min post‐injection. Most organs showed low accumulation of the radiotracer, and radioactivity was predominately cleared through kidneys (95% in 1 h). Radio‐HPLC analysis of plasma and urine samples showed a stable radiotracer at least up to 60 min post‐injection.     

Discovery of a highly specific 18F-labeled PET ligand for phosphodiesterase 10A enabled by novel spirocyclic iodonium ylide radiofluorination

As a member of cyclic nucleotide phosphodiesterase (PDE) enzyme family, PDE10A is in charge of the degradation of cyclic adenosine (cAMP) and guanosine monophosphates (cGMP). While PDE10A is primarily expressed in the medium spiny neurons of the striatum, it has been implicated in a variety of neurological disorders. Indeed, inhibition of PDE10A has proven to be of potential use for the treatment of central nervous system (CNS) pathologies caused by dysfunction of the basal ganglia–of which the striatum constitutes the largest component. A PDE10A-targeted positron emission tomography (PET) radioligand would enable a better assessment of the pathophysiologic role of PDE10A, as well as confirm the relationship between target occupancy and administrated dose of a given drug candidate, thus accelerating the development of effective PDE10A inhibitors. In this study, we designed and synthesized a novel ¹⁸F-aryl PDE10A PET radioligand, codenamed [¹⁸F]P10A-1910 ([¹⁸F]9), in high radiochemical yield and molar activity via spirocyclic iodonium ylide-mediated radiofluorination. [¹⁸F]9 possessed good in vitro binding affinity (IC₅₀ = 2.1 nmol/L) and selectivity towards PDE10A. Further, [¹⁸F]9 exhibited reasonable lipophilicity (logD = 3.50) and brain permeability (P_app > 10 × 10⁻⁶ cm/s in MDCK-MDR1 cells). PET imaging studies of [¹⁸F]9 revealed high striatal uptake and excellent in vivo specificity with reversible tracer kinetics. Preclinical studies in rodents revealed an improved plasma and brain stability of [¹⁸F]9 when compared to the current reference standard for PDE10A-targeted PET, [¹⁸F]MNI659. Further, dose–response experiments with a series of escalating doses of PDE10A inhibitor 1 in rhesus monkey brains confirmed the utility of [¹⁸F]9 for evaluating target occupancy in vivo in higher species. In conclusion, our results indicated that [¹⁸F]9 is a promising PDE10A PET radioligand for clinical translation.KEY WORDS: Phosphodiesterase 10A, PET radioligand, ¹⁸F, Spirocyclic iodonium ylide, Nonhuman primate, Target occupancy     

Synthesis and bioevaluation of 4‐chloro‐2‐tert‐butyl‐5‐[2‐[[1‐[2‐[18F]fluroethyl]‐1H‐1,2,3‐triazol‐4‐yl]methyl]phenylmethoxy]‐3(2H)‐pyridazinone as potential myocardial perfusion imaging agent with PET

This study reports the synthesis and characterization of 4‐chloro‐2‐tert‐butyl‐5‐[2‐[[1‐[2‐[¹⁸F]fluroethyl]‐1H‐1,2,3‐triazol‐4‐yl]methyl]phenylmethoxy]‐3(2H)‐pyridazinone ([¹⁸F]Fmp2) for myocardial perfusion imaging (MPI). The tosylate precursor and non‐radioactive compound [¹⁹F]Fmp2 were synthesized and characterized by infrared, ¹H‐NMR, ¹³C‐NMR, and mass spectra (MS). The radiotracer [¹⁸F]Fmp2 was obtained by one‐step nucleophilic substitution of tosyl with ¹⁸F, and evaluated as an MPI agent in vitro and in vivo. Starting from [¹⁸F]KF/K₂₂₂ solution, the typical decay‐corrected radiochemical yield (RCY) was 38 ± 8.8% with high radiochemical purity (>98%). The specific activity was calculated as 10 GBq/µmol at the end of synthesis determined by HPLC analysis. In the mice biodistribution, [¹⁸F]Fmp2 showed very high initial heart uptake (53.35 ± 5.47 %ID/g at 2 min after injection) and remarkable retention. The heart/liver, heart/lung, and heart/blood ratios were 7.98, 8.20, and 53.13, respectively at 2 min post‐injection. In the Positron Emission Tomography (PET) imaging study of Chinese mini‐swine, the standardized uptake value of the liver decreased modestly during the 2 h post‐injection, while the heart uptake and heart/liver ratios continued to increase with time. [¹⁸F]Fmp2 exhibited good stability, high heart uptake and low lung uptake in mice and Chinese mini‐swine. It may be worthy of further modification to improve liver clearance for MPI in the future.     

A New Highly Deuterated [18F]AV-45, [18F]D15FSP,for Imaging β-Amyloid Plaques in the Brain

[¹⁸F]AV-45 (florbetapir f18, Amyvid) is an FDA-approved PET imaging agent targeting Aβ plaques in the brain for diagnosis of Alzheimer’s disease (AD). Its metabolites led to a high background in the brain and large bone uptake of [¹⁸F]F^–, produced from dealkylation of the PEG chain. To slow down the in vivo metabolism, we report the design, synthesis, and evaluation of a highly deuterated derivative, [¹⁸F]D15FSP, and compared it with N-methyl-deuterated [¹⁸F]D3FSP and nondeuterated [¹⁸F]AV-45. D15FSP displayed excellent binding affinity (K_i = 7.52 nM) to Aβ aggregates. In vitro autoradiography of [¹⁸F]D15FSP, [¹⁸F]D3FSP, and [¹⁸F]AV-45 showed excellent binding to Aβ plaques in human AD brain sections. Biodistribution studies displayed lower bone uptake at 120 min for [¹⁸F]D15FSP compared to that for [¹⁸F]D3FSP and [¹⁸F]AV-45 (1.44 vs 4.23 and 4.03%ID/g, respectively). As the highly deuterated [¹⁸F]D15FSP displayed excellent Aβ binding affinity, high initial brain penetration, and lower bone retention, it might be suitable for PET imaging in detecting Aβ plaques.     

Synthesis of [18F]Fluoroalanine and [18F]Fluorotamoxifen for Imaging Breast Tumors

Abstract

To develop ligands for imaging breast tumors, [¹⁸F]fluoro analogue of tamoxifen and [¹⁸F]fluoro-alanine were radiosynthesized. In vivo biodistribution studies were performed in mammary tumor-bearing rats. In studies on the biodistribution of an [¹⁸F]fluoro analogue of tamoxifen, tumor uptake decreased when rats were pretreated with diethylstilbestrol (DES), suggesting that tracer uptake in tumors was receptor-mediated. An estrogen receptor assay indicated that tumors have a receptor density of 7.5 fmol/mg protein. Studies of the distribution of [¹⁸F]fluoroalanine in tissue showed that the tumor-to-tissue ratio increases as a function of time. Positron emission tomography (PET) images of tumor-bearing rats demonstrated that tumors can be visualized 1 h after rats are injected with an [¹⁸F]fluoro analogue of tamoxifen. PET imaging of pigs after injection of 10 mCi of [¹⁸F]fluoro analogue of tamoxifen showed uterine uptake that could be blocked by DES (50 mg). The findings suggest that both radiotracers are useful for imaging breast tumors.     

Radiosynthesis and preliminary evaluation of an 18F-labeled tubastatin A analog for PET imaging of histone deacetylase 6

Histone deacetylase 6 (HDAC6) is a unique member of the HDAC family because of its characteristics, namely, its cytoplasmic localization and ubiquitin binding. HDAC6 has been implicated in cancer metastasis and neurodegeneration. In the present study, we performed radiosynthesis and biological evaluation of a fluorine-18–labeled ligand [¹⁸F] 3 , which is an analog of the HDAC6-selective inhibitor tubastatin A, for positron emission tomography (PET) imaging. [¹⁸F] 3 was synthesized by a two-step reaction composed of ¹⁸F-fluorination and formation of a hydroxamic acid group. IC₅₀ values of 3 against HDAC1 and HDAC6 activities were 996 nM and 33.1 nM, respectively. A biodistribution study in mice demonstrated low brain uptake of [¹⁸F] 3 . Furthermore, bone radioactivity was stable at around 2% ID/g after injection, suggesting high tolerance to defluorination. Regarding metabolic stability, 70% of the compound was observed as the unchanged form at 30 minutes post injection in mouse plasma. A small animal PET study in mice showed that pretreatment with cyclosporine A had no effect on initial brain uptake of [¹⁸F] 3 , suggesting low brain uptake of [¹⁸F] 3 was not caused by the P-glycoprotein–mediated efflux. While PET imaging using [¹⁸F] 3 has a limitation with respect to neurodegenerative diseases, further studies evaluating its utility for certain cancers are worth evaluating.     

Synthesis and preliminary radiopharmacological characterisation of an 11C‐labelled azadipeptide nitrile as potential PET tracer for imaging of cysteine cathepsins

An O‐methyltyrosine‐containing azadipeptide nitrile was synthesised and investigated for its inhibitory activity towards cathepsins L, S, K, and B. Labelling with carbon‐11 was accomplished by reaction of the corresponding phenolic precursor with [¹¹C]methyl iodide starting from cyclotron‐produced [¹¹C]methane. Radiopharmacological evaluation of the resulting radiotracer in a mouse xenograft model derived from a mammary tumour cell line by small animal PET imaging indicates tumour targeting with complex pharmacokinetics. Radiotracer uptake in the tumour region was considerably lower under treatment with the nonradioactive reference compound and the epoxide‐based irreversible cysteine cathepsin inhibitor E64. The in vivo behaviour observed for this radiotracer largely confirms that of the corresponding ¹⁸F‐fluoroethylated analogue and suggests the limited suitability of azadipeptide nitriles for the imaging of tumour‐associated cysteine cathepsins despite target‐mediated uptake is evidenced.     

Biodistribution, pharmacokinetics and metabolism of interleukin-1 receptor antagonist (IL-1RA) using [¹⁸F]-IL1RA and PET imaging in rats

BACKGROUND AND PURPOSE

Positron emission tomography (PET) has the potential to improve our understanding of the preclinical pharmacokinetics and metabolism of therapeutic agents, and is easily translated to clinical studies in humans. However, studies involving proteins radiolabelled with clinically relevant PET isotopes are currently limited. Here we illustrate the potential of PET imaging in a preclinical study of the biodistribution and metabolism of ¹⁸F-labelled IL-1 receptor antagonist ([¹⁸F]IL-1RA) using a novel [¹⁸F]-radiolabelling technique.

EXPERIMENTAL APPROACH

IL-1RA was radiolabelled by reductive amination on lysine moieties with [¹⁸F]fluoroacetaldehyde. Sprague-Dawley rats were injected intravenously with [¹⁸F]IL-1RA and imaged with a PET camera for 2 h. For the study of IL-1RA metabolites by ex vivoγ-counting of samples, rats were killed 20 min, 1 h or 2 h after injection of [¹⁸F]IL-1RA.

KEY RESULTS

[¹⁸F]IL-1RA distribution into the major organs of interest was as follows: kidneys >> liver > lungs >> brain. In lungs and liver, [¹⁸F]IL-1RA uptake peaked within 1 min post-injection then decreased rapidly to reach a plateau from 10 min post-injection. In the brain, the uptake exhibited slower pharmacokinetics with a smaller post-injection peak and a plateau from 6 min onward. IL-1RA was rapidly metabolized and these metabolites represented ∼40% of total activity in plasma and ∼80% in urine, 20 min after injection.

CONCLUSIONS AND IMPLICATIONS

Preclinical PET imaging is a feasible method of assessing the biodistribution of new biological compounds of therapeutic interest rapidly. The biodistribution of [¹⁸F]IL-1RA reported here is in agreement with an earlier study suggesting low uptake in the normal brain, with rapid metabolism and excretion via the kidneys.     

Preclinical evaluation and pilot clinical study of [18F]AlF-labeled FAPI-tracer for PET imaging of cancer associated fibroblasts

In recent years, fibroblast activation protein (FAP) has emerged as an attractive target for the diagnosis and radiotherapy of cancers using FAP-specific radioligands. Herein, we aimed to design a novel ¹⁸F-labeled FAP tracer ([¹⁸F]AlF-P-FAPI) for FAP imaging and evaluated its potential for clinical application. The [¹⁸F]AlF-P-FAPI novel tracer was prepared in an automated manner within 42 min with a non-decay corrected radiochemical yield of 32 ± 6% (n = 8). Among A549-FAP cells, [¹⁸F]AlF-P-FAPI demonstrated specific uptake, rapid internalization, and low cellular efflux. Compared to the patent tracer [¹⁸F]FAPI-42, [¹⁸F]AlF-P-FAPI exhibited lower levels of cellular efflux in the A549-FAP cells and higher stability in vivo. Micro-PET imaging in the A549-FAP tumor model indicated higher specific tumor uptake of [¹⁸F]AlF-P-FAPI (7.0 ± 1.0% ID/g) compared to patent tracers [¹⁸F]FAPI-42 (3.2 ± 0.6% ID/g) and [⁶⁸Ga]Ga-FAPI-04 (2.7 ± 0.5% ID/g). Furthermore, in an initial diagnostic application in a patient with nasopharyngeal cancer, [¹⁸F]AlF-P-FAPI and [¹⁸F]FDG PET/CT showed comparable results for both primary tumors and lymph node metastases. These results suggest that [¹⁸F]AlF-P-FAPI can be conveniently prepared, with promising characteristics in the preclinical evaluation. The feasibility of FAP imaging was demonstrated using PET studies.     

Fluorine‐18 radiolabeling of a nitrophenyl sulfoxide and its evaluation in an SK‐RC‐52 model of tumor hypoxia

The significance of imaging hypoxia with the positron emission tomography ligand [¹⁸F]FMISO has been demonstrated in a variety of cancers. However, the slow kinetics of [¹⁸F]FMISO require a 2‐h delay between tracer administration and patient scanning. Labeled chloroethyl sulfoxides have shown faster kinetics and higher contrast than [¹⁸F]FMISO in a rat model of ischemic stroke. However, these nitrogen mustard analogues are unsuitable for routine production and use in humans. Here, we report on the synthesis and in vitro and in vivo evaluation of a novel sulfoxide, which contains an ester moiety for hydrolysis and subsequent trapping in hypoxic cells. Non‐decay corrected yields of radioactivity were 1.18 ± 0.24% (n = 27, 2.5 ± 0.5% decay corrected radiochemical yield) based on K[¹⁸F]F. The radiotracer did not show any defluorination and did not undergo metabolism in an in vitro assay using S9 liver fractions. Imaging studies using an SK‐RC‐52 tumor model in BALB/c nude mice have revealed that [¹⁸F]1 is retained in hypoxic tumors and has similar hypoxia selectivity to [¹⁸F]FMISO. Because of a three times faster clearance rate than [¹⁸F]FMISO from normoxic tissue, [¹⁸F]1 has emerged as a promising new radiotracer for hypoxia imaging.     

Radiosynthesis and in vivo study of [18F]1‐(2‐fluoroethyl)‐4‐[(4‐cyanophenoxy)methyl]piperidine: a promising new sigma‐1 receptor ligand

The novel sigma‐1 receptor PET radiotracer [¹⁸F]1‐(2‐fluoroethyl)‐4‐[(4‐cyanophenoxy)methyl]piperidine ([¹⁸F]WLS1.002, [¹⁸F]‐2) was synthesized (n=6) by heating the corresponding N‐ethylmesylate precursor in an anhydrous acetonitrile solution containing [¹⁸F]fluoride, Kryptofix K₂₂₂ and potassium carbonate for 15 min. Purification was accomplished by reverse‐phase HPLC methods, providing [¹⁸F]‐2 in 59±8% radiochemical yield (EOB), with specific activity of 2.89±0.80 Ci/µmol (EOS) and radiochemical purity of 98.3±2.1%. Rat biodistribution studies revealed relatively high uptake in many organs known to contain sigma‐1 receptors, including the lungs, kidney, heart, spleen, and brain. Good clearance from normal tissues was observed over time. Blocking studies (60 min) demonstrated high (>80%) specific binding of [¹⁸F]‐2 in the brain, with reduction also noted in other organs known to express these sites. Copyright © 2005 John Wiley & Sons, Ltd.     

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

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Synthesis,labelling and first evaluation of [18F]R91150 as a serotonin 5‐HT2A receptor antagonist for PET

In psychiatric disorders such as anxiety, depression and schizophrenia, 5‐HT_2A receptors play an important role. In order to investigate them in vivo there is an increasing interest in selective and high‐affinity radioligands for receptor binding studies using positron emission tomography (PET). Since available radioligands have disadvantages, R91150, which is a selective and high‐affinity ligand for 5‐HT_2A receptors, was labelled with fluorine‐18. This was accomplished in six steps via 4‐[¹⁸F]fluorophenol and 1‐(3‐bromopropoxy)‐4‐[¹⁸F]fluorobenzene within 190 min starting from no‐carrier‐added [¹⁸F]fluoride. The overall radiochemical yield was 3.8±2% and the specific activity was at least 335 GBq/µmol at the end of the synthesis. First ex vivo studies in mice proved the uptake of [¹⁸F]R91150 in the brain. Radiometabolite studies revealed no radiometabolites in the brain, whereas in the plasma at least two could be detected 30 min p.i. Further preclinical studies are encouraged to evaluate the potential of this new 5‐HT_2A ligand as a radiotracer for PET. Copyright © 2008 John Wiley & Sons, Ltd.     

Discovery and development of brain-penetrant 18F-labeled radioligands for neuroimaging of the sigma-2 receptors

We have discovered and synthesized a series of indole-based derivatives as novel sigma-2 (σ₂) receptor ligands. Two ligands with high σ₂ receptor affinity and subtype selectivity were then radiolabeled with F-18 in good radiochemical yields and purities, and evaluated in rodents. In biodistribution studies in male ICR mice, radioligand [¹⁸F]9, or 1-(4-(5,6-dimethoxyisoindolin-2-yl)butyl)-4-(2-[¹⁸F]fluoroethoxy)-1H-indole, was found to display high brain uptake and high brain-to-blood ratio. Pretreatment of animals with the selective σ₂ receptor ligand CM398 led to significant reductions in both brain uptake (29%–54%) and brain-to-blood ratio (60%–88%) of the radioligand in a dose-dependent manner, indicating high and saturable specific binding of [¹⁸F]9 to σ₂ receptors in the brain. Further, ex vivo autoradiography in male ICR mice demonstrated regionally heterogeneous specific binding of [¹⁸F]9 in the brain that is consistent with the distribution pattern of σ₂ receptors. Dynamic positron emission tomography imaging confirmed regionally distinct distribution and high levels of specific binding for [¹⁸F]9 in the rat brain, along with appropriate tissue kinetics. Taken together, results from our current study indicated the novel radioligand [¹⁸F]9 as the first highly specific and promising imaging agent for σ₂ receptors in the brain.     

Automated radiosynthesis of the adenosine A2A receptor-targeting radiotracer [18F]FLUDA

[¹⁸F] FLUDA is a selective radiotracer for in vivo imaging of the adenosine A_2A receptor (A_2AR) by positron emission tomography (PET). Promising preclinical results obtained by neuroimaging of mice and piglets suggest the translation of [¹⁸F] FLUDA to human PET studies. Thus, we report herein a remotely controlled automated radiosynthesis of [¹⁸F] FLUDA using a GE TRACERlab FX2 N radiosynthesizer. The radiotracer was obtained by a one-pot two-step radiofluorination procedure with a radiochemical yield of 9±1%, a radiochemical purity of ≥99%, and molar activities in the range of 69–333 GBq/μmol at the end of synthesis within a total synthesis time of approx. 95 min (n = 16). Altogether, we successfully established a reliable and reproducible procedure for the automated production of [¹⁸F] FLUDA.     

In vivo affinity of [<Superscript>18</Superscript>F]fallypride for striatal and extrastriatal dopamine D<Subscript>2</Subscript> receptors in nonhuman primates

Rationale [¹⁸F]Fallypride is a new and promising radiotracer, suitable for imaging D₂ receptors with Positron Emission Tomography (PET) in both striatal and extrastriatal regions. The high signal to noise ratio of [¹⁸F]fallypride has been attributed to its high affinity for D₂ receptors (K_D of 0.03 nM, measured in vitro at room temperature).Objectives We sought to further characterize this tracer in terms of its in vivo affinity, possible affinity differences between brain regions and dependence of in vitro affinity on temperature.Methods PET scans were performed in baboons over a wide range of concentrations to measure the in vivo K_D of [¹⁸F]fallypride in striatal and extrastriatal regions. Several analytical approaches were used, including nonlinear kinetic modeling and equilibrium methods. Also, in vitro assays were performed at 22 and 37°C.Results No significant differences in the in vivo K_D were detected between regions. In vivo K_D of [¹⁸F]fallypride was 0.22±0.05 nM in striatum, 0.17±0.05 nM in thalamus, and 0.21±0.07 nM in hippocampus. These values were intermediate between in vitro K_D measured at 22 (0.04±0.03 nM) and 37 degrees (2.03±1.07 nM).Conclusion The in vivo affinity of [¹⁸F]fallypride was not as high as previously estimated from in vitro values. This property might contribute to the favorable kinetic properties of the tracer. The in vivo affinity was similar between striatal and extrastriatal regions. This result indicates that the measured regional in vivo affinities of this tracer are not affected by putative regional differences in endogenous dopamine, and that [¹⁸F]fallypride is an appropriate tool to provide unbiased estimates of the occupancy of D₂ receptors by antipsychotic drugs in striatal and extrastriatal regions.     

Structure–activity relationship of pyrazol-4-yl-pyridine derivatives and identification of a radiofluorinated probe for imaging the muscarinic acetylcholine receptor M4

There is an accumulating body of evidence implicating the muscarinic acetylcholine receptor 4 (M₄) in schizophrenia and dementia with Lewy bodies, however, a clinically validated M₄ positron emission tomography (PET) radioligand is currently lacking. As such, the aim of this study was to develop a suitable M₄ PET ligand that allows the non-invasive visualization of M₄ in the brain. Structure–activity relationship studies of pyrazol-4-yl-pyridine derivates led to the discovery of target compound 12 ― a subtype-selective positive allosteric modulator (PAM). The radiofluorinated analogue, [¹⁸F]12, was synthesized in 28 ± 10% radiochemical yield, >37 GBq/μmol and an excellent radiochemical purity >99%. Initial in vitro autoradiograms on rodent brain sections were performed in the absence of carbachol and showed moderate specificity as well as a low selectivity of [¹⁸F]12 for the M₄-rich striatum. However, in the presence of carbachol, a significant increase in tracer binding was observed in the rat striatum, which was reduced by >60% under blocking conditions, thus indicating that orthosteric ligand interaction is required for efficient binding of [¹⁸F]12 to the allosteric site. Remarkably, however, the presence of carbachol was not required for high specific binding in the non-human primate (NHP) and human striatum, and did not further improve the specificity and selectivity of [¹⁸F]12 in higher species. These results pointed towards significant species-differences and paved the way for a preliminary PET study in NHP, where peak brain uptake of [¹⁸F]12 was found in the putamen and temporal cortex. In conclusion, we report on the identification and preclinical development of the first radiofluorinated M₄ PET radioligand with promising attributes. The availability of a clinically validated M₄ PET radioligand harbors potential to facilitate drug development and provide a useful diagnostic tool for non-invasive imaging.     

A Novel 18F-Labeled Imidazo[2,1-b]benzothiazole (IBT) for High-Contrast PET Imaging of β-Amyloid Plaques

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