RESUMO
Monoacylglycerol lipase (MAGL) constitutes a serine hydrolase that orchestrates endocannabinoid homeostasis and exerts its function by catalyzing the degradation of 2-arachidonoylglycerol (2-AG) to arachidonic acid (AA). As such, selective inhibition of MAGL represents a potential therapeutic and diagnostic approach to various pathologies including neurodegenerative disorders, metabolic diseases and cancers. Based on a unique 4-piperidinyl azetidine diamide scaffold, we developed a reversible and peripheral-specific radiofluorinated MAGL PET ligand [18F]FEPAD. Pharmacokinetics and binding studies on [18F]FEPAD revealed its outstanding specificity and selectivity towards MAGL in brown adipose tissue (BAT) - a tissue that is known to be metabolically active. We employed [18F]FEPAD in PET studies to assess the abundancy of MAGL in BAT deposits of mice and found a remarkable degree of specific tracer binding in the BAT, which was confirmed by post-mortem tissue analysis. Given the negative regulation of endocannabinoids on the metabolic BAT activity, our study supports the concept that dysregulation of MAGL is likely linked to metabolic disorders. Further, we now provide a suitable imaging tool that allows non-invasive assessment of MAGL in BAT deposits, thereby paving the way for detailed mechanistic studies on the role of BAT in endocannabinoid system (ECS)-related pathologies.
Assuntos
Endocanabinoides , Monoacilglicerol Lipases , Endocanabinoides/metabolismo , Tecido Adiposo Marrom/diagnóstico por imagem , Tecido Adiposo Marrom/metabolismo , Tomografia por Emissão de Pósitrons/métodos , Ligantes , Inibidores Enzimáticos/farmacologiaRESUMO
Fatty acid amide hydrolase (FAAH) is a key enzyme in the endocannabinoid system. N-(3,4-Dimethylisoxazol-5-yl)piperazine-4-[4-(2-fluoro-4-[11C]methylphenyl)thiazol-2-yl]-1-carboxamide ([11C]DFMC) was developed as an irreversible-type positron emission tomography (PET) tracer for FAAH. Here, we attempted to noninvasively estimate rate constant k3 (rate of transfer to the specifically-bound compartment) as a direct index for FAAH in the rat brain. First, the two-tissue compartment model analysis including three parameters [K1-k3, two-tissue compartment model for the irreversible-type radiotracer (2TCMi)] in PET study with [11C]DFMC was conducted, which provided 0.21 ± 0.04 ml·cm-3·min-1 of the net uptake value (Ki), an indirect index for FAAH, in the FAAH-richest region (the cingulate cortex). Subsequently, to noninvasively estimate Ki value, the reference model analysis (Patlak graphical analysis reference model) was tried using a time-activity curve of the spinal cord. In that result, the noninvasive Ki value (KREF) was concisely estimated with high correlation (r > 0.95) to Ki values based on 2TCMi. Using estimated KREF value, we tried to obtain calculated-k3 based on previously defined equations. The calculated k3 was successfully estimated with high correlation (r = 0.95) to direct k3 in 2TCMi. Finally, the dose relationship study using calculated k3 demonstrated that in vivo ED50 value of [3-(3-carbamoylphenyl)phenyl] N-cyclohexylcarbamate, a major inhibitor of FAAH, was 66.4 µg/kg in rat brain. In conclusion, we proposed the calculated k3 as an alternative index corresponding to regional FAAH concentrations and suggested that PET with [11C]DFMC enables occupancy study for new pharmaceuticals targeting FAAH. SIGNIFICANCE STATEMENT: In the present study, we proposed calculated k3 as an alternative index corresponding with fatty acid amide hydrolase concentration. By using calculated k3, in vivo ED50 of [3-(3-carbamoylphenyl)phenyl] N-cyclohexylcarbamate was successfully estimated to be 66.4 µg/kg for rats. Thus, we demonstrated the pharmacological utility of positron emission tomography with N-(3,4-dimethylisoxazol-5-yl)piperazine-4-[4-(2-fluoro-4-[11C]methylphenyl)thiazol-2-yl]-1-carboxamide.
Assuntos
Amidoidrolases/metabolismo , Encéfalo/metabolismo , Carbamatos/farmacologia , Radioisótopos de Carbono/metabolismo , Tomografia por Emissão de Pósitrons/métodos , Roedores/metabolismo , Animais , Endocanabinoides/metabolismo , Masculino , Compostos Radiofarmacêuticos/metabolismo , Ratos , Ratos Sprague-Dawley , Distribuição Tecidual/fisiologiaRESUMO
Selective metabotropic glutamate receptor 2 (mGluR2) inhibitors have been demonstrated to show therapeutic effects by improving alleviating symptoms of schizophrenic patients in clinical studies. Herein we report the synthesis and preliminary evaluation of a 11C-labeled positron emission tomography (PET) tracer originating from a mGluR2 inhibitor, 3-(cyclopropylmethyl)-7-((4-(4-methoxyphenyl)piperidin-1-yl)methyl)-8-(trifluoromethyl)-[1,2,4]triazolo[4,3-a]pyridine (CMTP, 1a). [11C]CMTP ([11C]1a) was synthesized by O-[11C]methylation of desmethyl precursor 1b with [11C]methyl iodide in 19.7 ± 8.9% (n = 10) radiochemical yield (based on [11C]CO2) with >98% radiochemical purity and >74 GBq/µmol molar activity. Autoradiography study showed that [11C]1a possessed moderate in vitro specific binding to mGluR2 in the rat brain, with a heterogeneous distribution of radioactive accumulation in the mGluR2-rich brain tissue sections, such as the cerebral cortex and striatum. PET study indicated that [11C]1a was able to cross the blood-brain barrier and enter the brain, but had very low specific binding in the rat brain. Further optimization for the chemical structure of 1a is necessary to increase binding affinity to mGluR2 and then improve in vivo specific binding in brain.
Assuntos
Meios de Contraste/farmacologia , Piridinas/farmacologia , Compostos Radiofarmacêuticos/farmacologia , Receptores de Glutamato Metabotrópico/metabolismo , Triazóis/farmacologia , Animais , Encéfalo/metabolismo , Radioisótopos de Carbono/química , Meios de Contraste/síntese química , Meios de Contraste/metabolismo , Masculino , Tomografia por Emissão de Pósitrons , Piridinas/síntese química , Piridinas/metabolismo , Compostos Radiofarmacêuticos/síntese química , Compostos Radiofarmacêuticos/metabolismo , Ratos Sprague-Dawley , Triazóis/síntese química , Triazóis/metabolismoRESUMO
Selective DAAO inhibitors have demonstrated promising therapeutic effects in clinical studies, including clinically alleviating symptoms of schizophrenic patients and ameliorating cognitive function in Alzheimer's patients with early phase. Herein we report the synthesis and preliminary evaluation of a 11C-labeled positron emission tomography ligand based on a DAAO inhibitor, DAO-1903 (8). 11C-Isotopologue of 8 was prepared in high radiochemical yield with high radiochemical purity (>99%) and high molar activity (>37 GBq/µmol). In vitro autoradiography studies indicated that the ligand possessed high in vitro specific binding to DAAO, while in vivo dynamic PET studies demonstrated that [11C]8 failed to cross the blood-brain barrier possibly due to moderate brain efflux mechanism. Further chemical scaffold optimization is necessary to overcome limited brain permeability and improve specific binding.
Assuntos
Encéfalo/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos/química , Animais , D-Aminoácido Oxidase/antagonistas & inibidores , D-Aminoácido Oxidase/metabolismo , Relação Dose-Resposta a Droga , Humanos , Camundongos , Simulação de Acoplamento Molecular , Estrutura Molecular , Compostos Radiofarmacêuticos/farmacologia , Ratos , Ratos Sprague-Dawley , Relação Estrutura-AtividadeRESUMO
Metabotropic glutamate receptor 2 (mGluR2) has been suggested as a therapeutic target for treating schizophrenia-like symptoms arising from increased glutamate transmission in the human forebrain. However, no reliable positron emission tomography (PET) radiotracer allowing for in vivo visualization of mGluR2 in the human brain is currently available. In this study, we synthesized 4-(2-fluoro-4-[11C]methoxyphenyl)-5-((2-methylpyridin-4-yl)methoxy)picolinamide ([11C]1) and evaluated its potential as a PET tracer for imaging mGluR2 in the rodent brain. Compound 1, a negative allosteric modulator (NAM) of mGluR2, showed high in vitro binding affinity (IC50: 26â¯nM) for mGluR2 overexpressed in human cells. [11C]1 was synthesized by O-[11C]methylation of the phenol precursor 2 with [11C]methyl iodide. After the reaction, HPLC purification and formulation, [11C]1 of 7.4⯱â¯2.8â¯GBq (nâ¯=â¯8) was obtained from [11C]carbon dioxide of 22.5⯱â¯4.8â¯GBq (nâ¯=â¯8) with >99% radiochemical purity and 70⯱â¯32â¯GBq/µmol (nâ¯=â¯8) molar activity at the end of synthesis. In vitro autoradiography for rat brains showed that [11C]1 binding was heterogeneously distributed in the cerebral cortex, striatum, hippocampus, and cerebellum. This pattern is consistent with the regional distribution pattern of mGluR2 in the rodent brain. The radioactivity was significantly reduced by self- or MNI-137 (a mGluR2 NAM) blocking. Small-animal PET studies indicated a low in vivo specific binding of [11C]1 in the rat brain. The brain uptake was increased in a P-glycoprotein and breast cancer resistant protein double knockout mouse, when compared to a wild-type mouse. While [11C]1 presented limited potential as an in vivo PET tracer for mGluR2, we suggested that it can be used as a lead compound for developing new radiotracers with improved in vivo brain properties.
Assuntos
Encéfalo/diagnóstico por imagem , Ácidos Picolínicos/química , Tomografia por Emissão de Pósitrons , Receptores de Glutamato Metabotrópico/análise , Animais , Encéfalo/metabolismo , Radioisótopos de Carbono , Relação Dose-Resposta a Droga , Humanos , Masculino , Camundongos , Camundongos Endogâmicos , Camundongos Knockout , Estrutura Molecular , Ácidos Picolínicos/síntese química , Ácidos Picolínicos/farmacocinética , Traçadores Radioativos , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Distribuição TecidualRESUMO
Monoacylglycerol lipase (MAGL) is a major serine hydrolase that hydrolyses 2-arachidonoylglycerol (2-AG) into arachidonic acid (AA) and glycerol in the brain. Because 2-AG and AA are endogenous biologically active ligands in the brain, the inhibition of MAGL is an attractive therapeutic target for neurodegenerative diseases. In this study, to visualize MAGL via positron emission tomography (PET), we report a new carbon-11-labeled radiotracer, namely 1,1,1,3,3,3-hexafluoropropan-2-yl-3-(1-benzyl-1H-pyrazol-3-yl)azetidine-1-[11C]carboxylate ([11C]6). Compound 6 exhibited high in vitro binding affinity (IC50â¯=â¯0.41â¯nM) to MAGL in the brain with a suitable lipophilicity (cLogDâ¯=â¯3.29). [11C]6 was synthesized by reacting 1,1,1,3,3,3-hexafluoropropanol (7) with [11C]phosgene ([11C]COCl2), followed by a reaction with 3-(1-benzyl-1H-pyrazol-3-yl)azetidine hydrochloride (8), which resulted in a 15.0⯱â¯6.8% radiochemical yield (decay-corrected, nâ¯=â¯7) based on [11C]CO2 and a 45â¯min synthesis time from the end of bombardment. A biodistribution study in mice showed high uptake of radioactivity in MAGL-rich organs, including the lungs, heart, and kidneys. More than 90% of the total radioactivity was irreversibly bound in the brain homogenate of rats 5â¯min and 30â¯min after the radiotracer injection. PET summation images of rat brains showed high radioactivity in all brain regions. Pretreatment with 6 or MAGL-selective inhibitor JW642 significantly reduced the uptake of radioactivity in the brain. [11C]6 is a promising PET tracer which offers in vivo specific binding and selectivity for MAGL in rodent brains.
Assuntos
Carbamatos/química , Monoacilglicerol Lipases/síntese química , Animais , Radioisótopos de Carbono/metabolismo , RatosRESUMO
Monoacylglycerol lipase (MAGL) is a main regulator of the endocannabinoid system within the central nervous system (CNS). Recently, [11C]SAR127303 was developed as a promising radioligand for MAGL imaging. In this study, we aimed to quantify regional MAGL concentrations in the rat brain using positron emission tomography (PET) with [11C]SAR127303. An irreversible two-tissue compartment model (2-TCMi, k4â¯=â¯0) analysis was conducted to estimate quantitative parameters (k3, Ki2-TCMi, and λk3). These parameters were successfully obtained with high identifiability (<10 %COV) for the following regions ranked in order from highest to lowest: cingulate cortexâ¯>â¯striatumâ¯>â¯hippocampusâ¯>â¯thalamusâ¯>â¯cerebellumâ¯>â¯hypothalamusâ¯≈â¯pons. In vitro autoradiographs using [11C]SAR127303 showed a heterogeneous distribution of radioactivity, as seen in the PET images. The Ki2-TCMi and λk3 values correlated relatively highly with in vitro binding (râ¯>â¯0.4, Pâ¯<â¯0.005). The Ki2-TCMi values showed high correlation and low underestimation (<10%) compared with the slope of a Patlak plot analysis with linear regression (KiPatlak). In conclusion, we successfully estimated regional net uptake value of [11C]SAR127303 reflecting MAGL concentrations in rat brain regions for the first time.
Assuntos
Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Modelos Teóricos , Monoacilglicerol Lipases/metabolismo , Neuroimagem/métodos , Tomografia por Emissão de Pósitrons/métodos , Animais , Carbamatos , Radioisótopos de Carbono , Masculino , Ratos , Ratos Sprague-Dawley , SulfonamidasRESUMO
CEP-32496, also known as RXDX-105 or Agerafenib, is a new orally active inhibitor for the mutated v-raf murine sarcoma viral oncogene homolog B1 (BRAFV600E), which has attracted considerable attention in clinical trials for the treatment of human cancers. Here, we used carbon-11-labeled CEP-32496 ([11C]CEP-32496) as a positron emission tomography (PET) radiotracer to evaluate its pharmacokinetic properties and explore its potential for in vivo imaging. Following radiotracer synthesis, we performed in vitro binding assays and autoradiography of [11C]CEP-32496 in the A375 melanoma cell line and on tumor tissue sections from mice harboring the BRAFV600E mutation. These were followed by PET scans and biodistribution studies on nude mice bearing subcutaneous A375 cell-induced melanoma. [11C]CEP-32496 showed high binding affinity for BRAFV600E-positive A375 melanoma cells and densely accumulated in the respective tissue sections; this could be blocked by the BRAFV600E selective antagonist sorafenib and by unlabeled CEP-32496. The PET and biodistribution results revealed that [11C]CEP-32496 accumulated continuously but slowly into the tumor within a period of 0 to 60 minutes postinjection in A375-melanoma-bearing nude mice. Metabolite analysis showed high in vivo stability of [11C]CEP-32496 in plasma. Our results indicate that [11C]CEP-32496 has excellent specificity and affinity for the BRAFV600E mutation in vitro, while its noninvasive personalized diagnostic role needs to be studied further.
Assuntos
Melanoma/genética , Mutação/genética , Compostos de Fenilureia/farmacocinética , Proteínas Proto-Oncogênicas B-raf/genética , Quinazolinas/farmacocinética , Animais , Autorradiografia , Linhagem Celular Tumoral , Humanos , Lipídeos/química , Melanoma/sangue , Melanoma/urina , Camundongos Nus , Compostos de Fenilureia/sangue , Compostos de Fenilureia/química , Compostos de Fenilureia/urina , Quinazolinas/sangue , Quinazolinas/química , Quinazolinas/urina , Distribuição Tecidual , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Two novel radiotracers, namely, N-(4-[18F]fluorobenzyl)-N-methyl-2-(7-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide ([18F]5) and 2-(5-(4-[18F]fluorophenyl)-2-oxobenzo[d]oxazol-3(2H)-yl)-N-methyl-N-phenylacetamide ([18F]6), were developed for positron emission tomography (PET) imaging of translocator protein (18 kDa) (TSPO) in ischemic brain in this study. The two radiotracers with a [18F]fluorobenzene ring were derived from the corresponding [18F]fluoroethyl tracers [18F]7 and [18F]8 which underwent [18F]defluoroethylation in vivo easily. [18F]5 or [18F]6 was synthesized by the radiofluorination of the spirocyclic iodonium ylide precursor 10 or 17 with [18F]F- in 23 ± 10% (n = 7) or 56 ± 9% (n = 7) radiochemical yields (decay-corrected, based on [18F]F-). [18F]5 and [18F]6 showed high in vitro binding affinities (Ki = 0.70 nM and 5.9 nM) for TSPO and moderate lipophilicities (log D = 2.9 and 3.4). Low uptake of radioactivity for both radiotracers was observed in mouse bones. Metabolite analysis showed that the in vivo stability of [18F]5 and [18F]6 was improved in comparison to the parent radiotracers [18F]7 and [18F]8. In particular, no radiolabelled metabolite of [18F]5 was found in the mouse brains at 60 min after the radiotracer injection. PET studies with [18F]5 on ischemic rat brains revealed a higher binding potential (BPND = 3.42) and maximum uptake ratio (4.49) between the ipsilateral and contralateral sides. Thus, [18F]5 was shown to be a useful PET radiotracer for visualizing TSPO in neuroinflammation models.
Assuntos
Isquemia Encefálica/diagnóstico por imagem , Encéfalo/diagnóstico por imagem , Proteínas de Transporte/metabolismo , Fluorbenzenos/química , Tomografia por Emissão de Pósitrons/métodos , Receptores de GABA-A/metabolismo , Compostos de Espiro/química , Compostos de Espiro/síntese química , Animais , Encéfalo/metabolismo , Isquemia Encefálica/metabolismo , Técnicas de Química Sintética , Interações Hidrofóbicas e Hidrofílicas , Camundongos , Traçadores Radioativos , Radioquímica , Ratos , Compostos de Espiro/farmacocinética , Distribuição TecidualRESUMO
DAA1106 (N-(2,5-dimethoxybenzyl)-N-(5-fluoro-2-phenoxyphenyl)acetamide), is a potent and selective ligand for the translocator protein (18â¯kDa, TSPO) in brain mitochondrial fractions of rats and monkey (Kiâ¯=â¯0.043 and 0.188â¯nM, respectively). In this study, to translate [18F]DAA1106 for clinical studies, we performed automated syntheses of [18F]DAA1106 using the spirocyclic iodonium ylide (1) as a radiolabelling precursor and conducted preclinical studies including positron emission tomography (PET) imaging of TSPO in ischemic rat brains. Radiofluorination of the ylide precursor 1 with [18F]F-, followed by HPLC separation and formulation, produced the [18F]DAA1106 solution for injection in 6% average (nâ¯=â¯10) radiochemical yield (based on [18F]F-) with >98% radiochemical purity and molar activity of 60-100â¯GBq/µmol at the end of synthesis. The synthesis time was 87â¯min from the end of bombardment. The automated synthesis achieved [18F]DAA1106 with sufficient radioactivity available for preclinical and clinical use. Biodistribution study of [18F]DAA1106 showed a low uptake of radioactivity in the mouse bones. Metabolite analysis showed that >96% of total radioactivity in the mouse brain at 60â¯min after the radiotracer injection was unmetabolized [18F]DAA1106. PET study of ischemic rat brains visualized ischemic areas with a high uptake ratio (1.9⯱â¯0.3) compared with the contralateral side. We have provided evidence that [18F]DAA1106 could be routinely produced for clinical studies.
Assuntos
Acetamidas/síntese química , Radioisótopos de Flúor/química , Éteres Fenílicos/síntese química , Tomografia por Emissão de Pósitrons/métodos , Animais , Automação , Iodo/química , Iodo/metabolismo , Masculino , Camundongos , Ratos , Ratos Sprague-Dawley , Distribuição TecidualRESUMO
Cannabinoid receptor type 1 (CB1) is mainly expressed in the brain, as well as being expressed in functional relevant concentrations in various peripheral tissues. 1-(4-Chlorophenyl)-3-(3-(6-(pyrrolidin-1-yl)pyridin-2-yl)phenyl)urea (PSNCBAM-1, 1) was developed as a potent allosteric antagonist for CB1 and its oral administration led to reductions in the appetite and body weight of rats. Several analogs of 1 (compounds 2 and 3) were recently identified through a series of structure-activity relationship studies. Herein, we report the synthesis of radiolabeled analogs of these compounds using [11C]COCl2 and an evaluation of their potential as PET ligands for CB1 imaging using in vitro and in vivo techniques. [11C]2 and [11C]3 were successfully synthesized in two steps using [11C]COCl2. The radiochemical yields of [11C]2 and [11C]3 were 17±8% and 20±9% (decay-corrected to the end of bombardment, based on [11C]CO2). The specific activities of [11C]2 and [11C]3 were 42±36 and 37±13GBq/µmol, respectively. The results of an in vitro binding assay using brown adipose tissue (BAT) homogenate showed that the binding affinity of 2 for CB1 (KD=15.3µM) was much higher than that of 3 (KD=26.0µM). PET studies with [11C]2 showed a high uptake of radioactivity in BAT, which decreased in animals pretreated with AM281 (a selective antagonist for CB1). In conclusion, [11C]2 may be a useful PET ligand for imaging peripheral CB1 in BAT.
Assuntos
Compostos de Fenilureia/química , Tomografia por Emissão de Pósitrons , Piridinas/química , Receptor CB1 de Canabinoide/análise , Animais , Isótopos de Carbono , Relação Dose-Resposta a Droga , Ligantes , Estrutura Molecular , Compostos de Fenilureia/síntese química , Compostos de Fenilureia/farmacologia , Piridinas/síntese química , Piridinas/farmacologia , Ratos , Receptor CB1 de Canabinoide/antagonistas & inibidores , Receptor CB1 de Canabinoide/metabolismo , Relação Estrutura-AtividadeRESUMO
The purpose of this study was to synthesize a new positron emission tomography radiotracer, N-(4-tertiarybutylphenyl)-4-(3-chloropyridin-2-yl)tetrahydropyrazine-1(2H)-[11C]carboxamide ([11C]BCTC, [11C]1), and assess its in vivo binding to the transient receptor potential vanilloid subfamily member 1 (TRPV1) receptor in mice. [11C]BCTC was synthesized by reacting the hydrochloride of 4-tertiarybutylaniline (2·HCl) with [11C]phosgene ([11C]COCl2) to give isocyanate [11C]4, followed by reaction with 4-(3-chloropyridin-2-yl)tetrahydropyrazine (3). [11C]BCTC was obtained at a 16-20% radiochemical yield, based on the cyclotron-produced [11C]CO2 (decay-corrected to the end of bombardment), with >98% radiochemical purity and 65-110GBq/µmol specific activity at the end of synthesis. An ex vivo biodistribution study in mice confirmed the specific binding of [11C]BCTC to TRPV1 in the trigeminal nerve, which is a tissue with high TRPV1 expression.
Assuntos
Pirazinas/farmacocinética , Piridinas/farmacocinética , Canais de Cátion TRPV/química , Nervo Trigêmeo/química , Animais , Sítios de Ligação/efeitos dos fármacos , Isótopos de Carbono , Relação Dose-Resposta a Droga , Camundongos , Camundongos Endogâmicos , Estrutura Molecular , Tomografia por Emissão de Pósitrons , Pirazinas/síntese química , Pirazinas/química , Piridinas/síntese química , Piridinas/química , Traçadores Radioativos , Relação Estrutura-Atividade , Canais de Cátion TRPV/biossíntese , Distribuição TecidualRESUMO
The purpose of this study was to develop three new radiotracers, 1-(cyclopropylmethyl)-4-([11C/18F]substituted-phenyl)piperidin-1-yl-2-oxo-1,2-dihydropyridine-3-carbonitrile ([11C]1, [11C]2, and [18F]4), and to examine their specific bindings with metabotropic glutamate receptor subtype 2 (mGluR2) in rat brain sections by using in vitro autoradiography. These compounds were found to possess potent in vitro binding affinities (Ki: 8.0-34.1nM) for mGluR2 in rat brain homogenate. [11C]1, [11C]2, and [18F]4 were synthesized by [11C/18F]alkylation of the corresponding phenol precursors with [11C]methyl iodide or [18F]fluoroethyl bromide with >98% radiochemical purity and 80-130GBq/µmol specific activity at the end of synthesis. In vitro autoradiography indicated that these radiotracers showed heterogeneous specific bindings in mGluR2-rich brain regions, such as the cerebral cortex, striatum, hippocampus, and granular layer of the cerebellum.
Assuntos
Encéfalo/metabolismo , Compostos Radiofarmacêuticos/síntese química , Receptores de Glutamato Metabotrópico/metabolismo , Animais , Autorradiografia , Encéfalo/diagnóstico por imagem , Radioisótopos de Carbono/química , Radioisótopos de Flúor/química , Marcação por Isótopo , Cinética , Tomografia por Emissão de Pósitrons , Ligação Proteica , Compostos Radiofarmacêuticos/química , Ratos , Razão Sinal-RuídoRESUMO
Intracellular pH is a valuable index for predicting neuronal damage and injury. However, no PET probe is currently available for monitoring intracellular pH in vivo. In this study, we developed a new approach for visualizing the hydrolysis rate of monoacylglycerol lipase, which is widely distributed in neurons and astrocytes throughout the brain. This approach uses PET with the new radioprobe [11C]QST-0837 (1,1,1,3,3,3-hexafluoropropan-2-yl-3-(1-phenyl-1H-pyrazol-3-yl)azetidine-1-[11C]carboxylate), a covalent inhibitor containing an azetidine carbamate skeleton for monoacylglycerol lipase. The uptake and residence of this new radioprobe depends on the intracellular pH gradient, and we evaluated this with in silico, in vitro and in vivo assessments. Molecular dynamics simulations predicted that because the azetidine carbamate moiety is close to that of water molecules, the compound containing azetidine carbamate would be more easily hydrolyzed following binding to monoacylglycerol lipase than would its analogue containing a piperidine carbamate skeleton. Interestingly, it was difficult for monoacylglycerol lipase to hydrolyze the azetidine carbamate compound under weakly acidic (pH 6) conditions because of a change in the interactions with water molecules on the carbamate moiety of their complex. Subsequently, an in vitro assessment using rat brain homogenate to confirm the molecular dynamics simulation-predicted behaviour of the azetidine carbamate compound showed that [11C]QST-0837 reacted with monoacylglycerol lipase to yield an [11C]complex, which was hydrolyzed to liberate 11CO2 as a final product. Additionally, the 11CO2 liberation rate was slower at lower pH. Finally, to indicate the feasibility of estimating how the hydrolysis rate depends on intracellular pH in vivo, we performed a PET study with [11C]QST-0837 using ischaemic rats. In our proposed in vivo compartment model, the clearance rate of radioactivity from the brain reflected the rate of [11C]QST-0837 hydrolysis (clearance through the production of 11CO2) in the brain, which was lower in a remarkably hypoxic area than in the contralateral region. In conclusion, we indicated the potential for visualization of the intracellular pH gradient in the brain using PET imaging, although some limitations remain. This approach should permit further elucidation of the pathological mechanisms involved under acidic conditions in multiple CNS disorders.
RESUMO
Parkinson's disease (PD) is one of the most highly debilitating neurodegenerative disorders, which affects millions of people worldwide, and leucine-rich repeat kinase 2 (LRRK2) mutations have been involved in the pathogenesis of PD. Developing a potent LRRK2 positron emission tomography (PET) tracer would allow for in vivo visualization of LRRK2 distribution and expression in PD patients. In this work, we present the facile synthesis of two potent and selective LRRK2 radioligands [11C]3 ([11C]PF-06447475) and [18F]4 ([18F]PF-06455943). Both radioligands exhibited favorable brain uptake and specific bindings in rodent autoradiography and PET imaging studies. More importantly, [18F]4 demonstrated significantly higher brain uptake in the transgenic LRRK2-G2019S mutant and lipopolysaccharide (LPS)-injected mouse models. This work may serve as a roadmap for the future design of potent LRRK2 PET tracers.
Assuntos
Morfolinas , Nitrilas , Doença de Parkinson , Pirimidinas , Camundongos , Animais , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Leucina , Tomografia por Emissão de Pósitrons/métodos , Doença de Parkinson/metabolismo , MutaçãoRESUMO
Deposition of α-synuclein fibrils is implicated in Parkinson's disease (PD) and dementia with Lewy bodies (DLB), while in vivo detection of α-synuclein pathologies in these illnesses has been challenging. Here, we have developed a small-molecule ligand, C05-05, for visualizing α-synuclein deposits in the brains of living subjects. In vivo optical and positron emission tomography (PET) imaging of mouse and marmoset models demonstrated that C05-05 captured a dynamic propagation of fibrillogenesis along neural pathways, followed by disruptions of these structures. High-affinity binding of 18F-C05-05 to α-synuclein aggregates in human brain tissues was also proven by in vitro assays. Notably, PET-detectable 18F-C05-05 signals were intensified in the midbrains of PD and DLB patients as compared with healthy controls, providing the first demonstration of visualizing α-synuclein pathologies in these illnesses. Collectively, we propose a new imaging technology offering neuropathology-based translational assessments of PD and allied disorders toward diagnostic and therapeutic research and development.
Assuntos
Modelos Animais de Doenças , Doença por Corpos de Lewy , Doença de Parkinson , Tomografia por Emissão de Pósitrons , alfa-Sinucleína , Animais , alfa-Sinucleína/metabolismo , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Doença de Parkinson/diagnóstico por imagem , Humanos , Camundongos , Tomografia por Emissão de Pósitrons/métodos , Doença por Corpos de Lewy/metabolismo , Doença por Corpos de Lewy/patologia , Doença por Corpos de Lewy/diagnóstico por imagem , Callithrix , Masculino , Encéfalo/metabolismo , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Feminino , Idoso , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: A family of BF2-chelated tetraaryl-azadipyrromethenes was developed as non-porphyrin photosensitizers for photodynamic therapy. Among the developed photosensitizers, ADPM06 exhibited excellent photochemical and photophysical properties. Molecular imaging is a useful tool for photodynamic therapy planning and monitoring. Radiolabeled photosensitizers can efficiently address photosensitizer biodistribution, providing helpful information for photodynamic therapy planning. To evaluate the biodistribution of ADPM06 and predict its pharmacokinetics on photodynamic therapy with light irradiation immediately after administration, we synthesized [18F]ADPM06 and evaluated its in vivo properties. RESULTS: [18F]ADPM06 was automatically synthesized by Lewis acid-assisted isotopic 18F-19F exchange using ADPM06 and tin (IV) chloride at room temperature for 10 min. Radiolabeling was carried out using 0.4 µmol of ADPM06 and 200 µmol of tin (IV) chloride. The radiosynthesis time was approximately 60 min, and the radiochemical purity was > 95% at the end of the synthesis. The decay-corrected radiochemical yield from [18F]F- at the start of synthesis was 13 ± 2.7% (n = 5). In the biodistribution study of male ddY mice, radioactivity levels in the heart, lungs, liver, pancreas, spleen, kidney, small intestine, muscle, and brain gradually decreased over 120 min after the initial uptake. The mean radioactivity level in the thighbone was the highest among all organs investigated and increased for 120 min after injection. Upon co-injection with ADPM06, the radioactivity levels in the blood and brain significantly increased, whereas those in the heart, lung, liver, pancreas, kidney, small intestine, muscle, and thighbone of male ddY mice were not affected. In the metabolite analysis of the plasma at 30 min post-injection in female BALB/c-nu/nu mice, the percentage of radioactivity corresponding to [18F]ADPM06 was 76.3 ± 1.6% (n = 3). In a positron emission tomography study using MDA-MB-231-HTB-26 tumor-bearing mice (female BALB/c-nu/nu), radioactivity accumulated in the bone at a relatively high level and in the tumor at a moderate level for 60 min after injection. CONCLUSIONS: We synthesized [18F]ADPM06 using an automated 18F-labeling synthesizer and evaluated the initial uptake and pharmacokinetics of ADPM06 using biodistribution of [18F]ADPM06 in mice to guide photodynamic therapy with light irradiation.
RESUMO
Orexin 2 receptors (OX2R) represent a vital subtype of orexin receptors intricately involved in the regulation of wakefulness, arousal, and sleep-wake cycles. Despite their importance, there are currently no positron emission tomography (PET) tracers available for imaging the OX2R in vivo. Herein, we report [11C]1 ([11C]OX2-2201) and [11C]2 ([11C]OX2-2202) as novel PET ligands. Both compounds 1 (Ki = 3.6 nM) and 2 (Ki = 2.2 nM) have excellent binding affinity activities toward OX2R and target selectivity (OX2/OX1 > 600 folds). In vitro autoradiography in the rat brain suggested good to excellent in vitro binding specificity for [11C]1 and [11C]2. PET imaging in rat brains indicated that the low brain uptake of [11C]2 may be due to P-glycoprotein and/or breast cancer resistance protein efflux interaction and/or low passive permeability. Continuous effort in medicinal chemistry optimization is necessary to improve the brain permeability of this scaffold.
RESUMO
The aim of this study is to investigate the changes in expression of metabotropic glutamate (Glu) receptor subtype 1 (mGluR1), a key molecule involved in neuroexcitetoxicity, during excessive Glu release in the brain by PET imaging. An animal model of excessive Glu release in the brain was produced by intraperitoneally implanting an Alzet osmotic pump containing N-acetylcysteine (NAC), an activator of the cysteine/Glu antiporter, into the abdomen of rats. Basal Glu concentration in the brain was measured by microdialysis, which showed that basal Glu concentration in NAC-treated rats (0.31 µM) was higher than that in saline-treated rats (0.17 µM) at day 7 after the implantation of the osmotic pump. Similarly, PET studies with [11C]ITDM, a useful radioligand for mGluR1 imaging exhibited that the striatal binding potential (BPND) of [11C]ITDM for mGluR1 in PET assessments was increased in NAC-treated animals at day 7 after implantation (2.30) compared with before implantation (1.92). The dynamic changes in striatal BPND during the experimental period were highly correlated with basal Glu concentration. In conclusion, density of mGluR1 is rapidly upregulated by increases in basal Glu concentration, suggesting that mGluR1 might to be a potential biomarker of abnormal conditions in the brain.
Assuntos
Ácido Glutâmico , Receptores de Glutamato Metabotrópico , Acetilcisteína/farmacologia , Animais , Ácido Glutâmico/metabolismo , Ratos , Regulação para CimaRESUMO
The transmembrane α-amino-3-hydroxyl-5-methyl-4-isoxazolepropionic acid (AMPA) receptor regulatory protein γ-8 (TARP γ-8) constitutes an auxiliary subunit of AMPA receptors, which mediates various brain functions including learning and memory. TARP γ-8 has emerged as a promising therapeutic target for central nervous system disorders. Despite considerable efforts, previously reported TARP γ-8 PET radioligands, such as [11C]TARP-1903 and [11C]TARP-1811 series, were plagued by limited brain uptake and/or high nonspecific binding in vivo. Herein, we developed two novel 11C-labeled probes, [11C]8 and [11C]15 (also named as [11C]TARP-2105), of which the latter exhibited a reasonable brain uptake as well as specific binding toward TARP γ-8 both in vitro and in vivo, as confirmed by blocking experiments with the commercially available TARP γ-8 inhibitor, JNJ-55511118 in the TARP γ-8-rich hippocampus. Overall, [11C]15 exhibited promising tracer characteristics and proved to be a lead positron-emission tomography ligand for the non-invasive quantification of TARP γ-8 in the mammalian brain.