9 research outputs found

    ์„ค์น˜๋ฅ˜์˜ [18F]Mefway์—์„œ ๋‡Œ ์„ญ์ทจ์— ๊ธฐ์—ฌํ•˜๋Š” P-glycoprotein์˜ ํšจ๊ณผ

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    Dept. of Medicine/๋ฐ•์‚ฌI. INTRODUCTION Efflux transporters in brain capillary endothelial cells impede xenobiotics, including therapeutic drugs and PET radioligands. Of these, P-gp (P-glycoprotein) and Bcrp (Breast cancer resistance protein) are prominent efflux pumps. P-gp is colocalized with Bcrp on the luminal membranes of brain capillary endothelial cells, and they have partly overlapping substrate specificities that include a wide range of compounds. Thus, to investigate the effects of P-gp, it is required to evaluate the impact of Bcrp. The aim of this study was to determine whether the brain uptake of [18F]Mefway is influenced by the action of P-gp and Bcrp in rodents. II. MATERIALS AND METHODS To effectively suppress in vivo defluorination, fluconazole (OneFlu, JW Pharmaceutial, Seoul, Korea) was used. [18F]Mefway was applied to the pharmacological inhibition rats and genetically disrupted mice. In rats, blocking of P-gp activity was performed by tariquidar (TQD). For the pharmacological inhibition experiments male Sprague-Dawley (n = 3 for each group) rats were used. For genetic disruption, male Mdr1a/b(-/-), Bcrp1(-/-), Mdr1a/b(-/-)Bcrp1(-/-), and wild-type (WT) mice were used (n = 4 for each group). III. RESULTS Pretreatment of TQD results in 160% higher hippocampal uptake compared with control rats. In genetically disrupted mice, a maximal brain uptake value of 3.2 SUV in the triple knockout mice (Mdr1a/b(-/-)Bcrp1(-/-) was comparable to that of the double knockout mice (Mdr1a/b(-/-)) and two-fold those of the wild type and Bcrp1(-/-) knockout mice. The differences of binding values were statistically insignificant between control and experimental groups. The brain-to-plasma ratios for triple knockout mice were also 2-5 times higher than those for other groups. IV. CONCLUSION In PET, chemical knockout P-gp disruption induced considerable increase of brain uptake of [18F]Mefway compared with control group or wild type. This result was substantiated by ex vivo experiments showing high brain-to-plasma ratios for P-gp disrupted mice. In conclusion, [18F]Mefway is modulated by P-gp, and not by Bcrp in rodents. I. ๋ชฉ์  ๋‡Œ ๋ชจ์„ธํ˜ˆ๊ด€ ๋‚ดํ”ผ ์„ธํฌ์—์„œ ์œ ์ถœ ์šด๋ฐ˜์ž๋“ค์€ ์น˜๋ฃŒ ์•ฝ๋ฌผ๋“ค๊ณผ PET ๋ฐฉ์‚ฌ์„ฑ ๋ฆฌ๊ฐ„๋“œ๋“ค์„ ํฌํ•จํ•œ ์ƒ์ฒด์ด๋ฌผ์งˆ๋“ค์„ ๋ฐฉํ•ดํ•œ๋‹ค. ์ด๋“ค ์ค‘์—์„œ P-glycoprotein (P-gp)์™€ breast cancer resistance protein (Bcrp)๋Š” ๊ฐ€์žฅ ์ค‘์š”ํ•œ ์œ ์ถœ ์šด๋ฐ˜์ž๋“ค์ด๋‹ค. P-gp๋Š” ๋‡Œ ๋ชจ์„ธํ˜ˆ๊ด€ ๋‚ดํ”ผ ์„ธํฌ์˜ ๊ด€๊ฐ•๋‚ด๋ง‰์— Bcrp์™€ ํ•จ๊ป˜ ๋ฐœํ˜„์ด ๋˜์–ด ์žˆ์œผ๋ฉฐ ์ผ๋ถ€ ๊ธฐ์งˆ ํŠน์ด์„ฑ์„ ๊ณต์œ ํ•˜๊ณ  ์žˆ๋‹ค. ๋”ฐ๋ผ์„œ P-gp์˜ ํšจ๊ณผ๋ฅผ ์•Œ์•„๋ณด๊ธฐ ์œ„ํ•ด์„œ๋Š” Bcrp์˜ ํšจ๊ณผ๋ฅผ ์•Œ์•„๋ณผ ํ•„์š”์„ฑ์ด ์žˆ๋‹ค. ๋ณธ ์—ฐ๊ตฌ์˜ ๋ชฉ์ ์€ ์„ค์น˜๋ฅ˜์˜ [18F]Mefway์—์„œ P-glycoprotein (P-gp)์™€ breast cancer resistance protein (Bcrp)์˜ ๋‡Œ์„ญ์ทจ์— ๊ด€๋ จ๋œ ํšจ๊ณผ๋ฅผ ์•Œ์•„๋ณด๊ณ ์ž ํ•˜์˜€๋‹ค. II. ๋Œ€์ƒ ๋ฐ ๋ฐฉ๋ฒ• ํƒˆ๋ถˆ์†Œํ™”๋ฅผ ํšจ๊ณผ์ ์œผ๋กœ ์–ต์ œ์‹œํ‚ค๊ธฐ ์œ„ํ•˜์—ฌ fluconazole (OneFlu, JW Pharmaceutial, ์„œ์šธ, ๋Œ€ํ•œ๋ฏผ๊ตญ)๋ฅผ ์‚ฌ์šฉํ•˜์˜€๋‹ค. [18F]Mefway๋ฅผ ์•ฝ๋ฌผํ•™์ ์œผ๋กœ ์–ต์ œ์‹œํ‚จ ์ฅ์™€ ์œ ์ „์ ์œผ๋กœ ๋ณ€ํ˜•์‹œํ‚จ ์ƒ์ฅ์—๊ฒŒ ์ ์šฉ์„ ํ•˜์˜€๋‹ค. ์ฅ์—์„œ๋Š” tariquidar(TQD)๋ฅผ ์ด์šฉํ•˜์—ฌ P-gp ๊ธฐ๋Šฅ์„ ์–ต์ œ ์‹œ์ผฐ๋‹ค. ์•ฝ๋ฌผํ•™์ ์œผ๋กœ ์–ต์ œ์‹œํ‚ค๋Š” ์‹คํ—˜์„ ์œ„ํ•ด์„œ๋Š” ์ˆ˜์ปท Sprague-Dawley (๊ฐ ๊ทธ๋ฃน ๋‹น ์„ธ ๋งˆ๋ฆฌ) ์ฅ๋ฅผ ์‚ฌ์šฉํ•˜์˜€๋‹ค. ์œ ์ „์  ๋ณ€ํ˜•์„ ์œ„ํ•˜์—ฌ์„œ๋Š” ์ˆ˜์ปท Mdr1a/b(-/-), Bcrp1(-/-), Mdr1a/b(-/-)Bcrp1(-/-), ๊ทธ๋ฆฌ๊ณ  ๋Œ€์กฐ๊ตฐ ์ƒ์ฅ๊ฐ€ ์‚ฌ์šฉ๋˜์—ˆ๋‹ค (๊ฐ ๊ทธ๋ฃน ๋‹น ๋„ค ๋งˆ๋ฆฌ). III. ๊ฒฐ๊ณผ TQD๋ฅผ ์ „์ฒ˜์น˜ํ•จ์œผ๋กœ์จ ๋Œ€์กฐ๊ตฐ ์ฅ์™€ ๋น„๊ต ์‹œ ํ•ด๋งˆ์˜ ๋‡Œ ์„ญ์ทจ๊ฐ€ 160% ์ฆ๊ฐ€๋˜์—ˆ๋‹ค. ์œ ์ „์ ์œผ๋กœ ๋ณ€ํ˜•๋œ ์ƒ์ฅ์—์„œ, ์ตœ๋Œ€ ๋‡Œ ์„ญ์ทจ๊ฐ€ 3.2 SUV์ธ ์‚ผ์ค‘ ์œ ์ „์ž ๊ฒฐ์† ์ƒ์ฅ (tKO, Mdr1a/b(-/-)Bcrp1(-/-) ์™€ ๋น„๊ต ์‹œ ์ด์ค‘ ์œ ์ „์ž ๊ฒฐ์† ์ƒ์ฅ (dKO, Mdr1a/b(-/-)) ์™€ ์ตœ๋Œ€ ๋‡Œ ์„ญ์ทจ๊ฐ€ ๋น„์Šทํ•˜์˜€์œผ๋ฉฐ ๋Œ€์กฐ ๊ตฐ๊ณผ Bcrp1(-/-) ์œ ์ „์ž ๊ฒฐ์† ์ƒ์ฅ์™€ ๋น„๊ต ์‹œ ๋‘ ๋ฐฐ ๋†’์€ ์ตœ๋Œ€ ๋‡Œ ์„ญ์ทจ๋ฅผ ๋ณด์˜€๋‹ค. ๋Œ€์กฐ ๊ตฐ๊ณผ ์‹คํ—˜ ๊ตฐ๊ฐ„์˜ ๊ฒฐํ•ฉ ๊ฐ€์น˜์˜ ์ฐจ์ด๋Š” ํ†ต๊ณ„์ ์œผ๋กœ ์˜๋ฏธ๊ฐ€ ์—†๋Š” ๊ฒƒ์œผ๋กœ ๋‚˜ํƒ€๋‚ฌ๋‹ค. [18F]Mefway ๋†๋„์˜ ๋‡Œ์™€ ํ˜ˆ์žฅ๊ฐ„ ๋น„์œจ์€ ๋‹ค๋ฅธ ๊ทธ๋ฃน๊ณผ ๋น„๊ต ์‹œ ์‚ผ์ค‘ ์œ ์ „์ž ๊ฒฐ์† ์ƒ์ฅ์—์„œ 2.5๋ฐฐ ๋†’์€ ๊ฒƒ์œผ๋กœ ํ™•์ธ๋˜์—ˆ๋‹ค. IV. ๊ฒฐ๋ก  PET์—์„œ ํ™”ํ•™์ ์œผ๋กœ P-gp ๊ฒฐ์—ฌ๋œ ๊ฒฝ์šฐ ๋Œ€์กฐ๊ตฐ๊ณผ ๋น„๊ต์‹œ [18F]Mefway์˜ ๋‡Œ์—์„œ ์„ญ์ทจ๋„๊ฐ€ ๋งค์šฐ ๋งŽ์ด ์ฆ๊ฐ€๋˜์—ˆ๋‹ค. ์ด๋Ÿฌํ•œ ๊ฒฐ๊ณผ๋Š” P-gp ๊ฒฐ์† ์ƒ์ฅ์—์„œ๋„ ๋†’์€ ๋‡Œ-ํ˜ˆ์žฅ๊ฐ„ ๋น„์œจ์„ ๋ณด์ธ ์‹คํ—˜ ๊ฒฐ๊ณผ์—์„œ๋„ ์ž…์ฆ๋˜์—ˆ๋‹ค. ๊ฒฐ๋ก ์ ์œผ๋กœ [18F]Mefway๋Š” ์„ค์น˜๋ฅ˜์—์„œ Bcrp๊ฐ€ ์•„๋‹Œ P-gp์— ์˜ํ•ด์„œ ์กฐ์ ˆ์ด ๋œ๋‹ค.ope

    PET Imaging of the Serotoninergic 5-HT1A System

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    Assessment of Serotonergic Function by Radioligands and Microdialysis:focus on stress-related behaviour and antidepressant efficacy

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    Serotonine is een chemische stof die betrokken is bij het doorgeven van signalen in de hersenen en speelt o.a. een rol bij het regelen van de gemoedstoestand, zoals in depressie. De meeste antidepressiva zijn erop gericht om serotonine concentraties in het menselijk brein te verhogen, maar deze geneesmiddelen lijken bij een deel van de patiรซnten niet erg goed te werken. De reden hiervan is niet duidelijk. Daarom is het van belang om te kunnen meten wat er met de signaaloverdracht door serotonine gebeurt onder invloed van stress en na behandeling met antidepressiva. Een methode om dit op een non-invasieve manier te meten is met positron emissie tomografie (PET). De studies die in dit poefschrift worden beschreven betreffen de validatie van radioactief gemerkte chemische stoffen, of PET tracers, die de aanmaak van serotonine ([11C]5-HTP) en de gevoeligheid van 5-HT2A receptoren voor serotonine ([11C]MDL 100907) in de hersenen kunnen meten. De laatstgenoemde stof bleek zeer geschikt. Hiermee hebben we aangetoond dat 5-HT2A receptoren waarschijnlijk geen essentiรซle rol spelen bij het omgaan met stress (coping style) en de reactie hierop. Daarnaast hebben we gekeken of de effectiviteit van antidepressiva verbeterd kan worden door aan serotonine heropname remmers (SSRIs) een specifieke 5-HT2C receptor blokker toe te voegen. Door deze geneesmiddelcombinatie lijken de niveaus van zowel serotonine als dopamine, een neurotransmitter betrokken bij motivatie, in de hersenen te kunnen worden verhoogd. PET lijkt een veelbelovende methode om serotonine signaaloverdracht te meten. Toekomstig onderzoek zal moeten uitwijzen of andere onderdelen betrokken bij serotonine signaaloverdracht belangrijk zijn voor stressgerelateerd gedrag. Daarnaast is het van belang om de effecten van antidepressiva op serotonine signaaloverdracht te meten en deze te relateren aan veranderingen in gedrag en gemoedstoestand

    PET-imaging in depression and antidepressant therapies : focus on the serotonin system and the cerebral glucose metabolism

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    The main scope of the research summarised in this dissertation comprises the use of positron emission tomography to investigate the role of the serotonin transporter in depression and antidepressant therapies. Hereby, several studies were performed using the PET radiotracer [11C]DASB, which specifically targets the serotonin transporter. To allow qualitative and safe research with this radiotracer, the first research topic focussed on the optimization of the radiotracerโ€™s purification procedure and its quality control. Using this radiotracer, a first-in-dog study was carried out to investigate the radiotracerโ€™s distribution and to define the appropriate image quantification methods. Subsequently, this radiotracer was used to perform a dose-occupancy in the dog to estimate the optimal dosing regimen to treat dogs with behavioural disorders with escitalopram. A second part of the dissertation focuses on rats and the current position of repetitive transcranial magnetic stimulation (rTMS) in the rat. Hereby, several additional objectives were put forward. The first objective comprised the evaluation of the accuracy of a for rodents adapted human neuronavigation system to perform rTMS in the rat. A second objective was the investigation of the construct validity of two depression models in terms of altered regional glucose metabolism. This was investigated via a PET study using the radiotracer [18F]FDG. Finally, for the preferred depression model, which was the one based on chronic corticosterone injections, the scope was extended from the serotonin transporter to the serotonin 5-HT1A and 5-HT2A receptors to explore the role of the serotonin system in the pathophysiology of this depression model in the rat. For this purpose, three radiotracers were applied: [11C]DASB, [18F]MPPF, and [18F]altanserin. This allowed to image the serotonin transporters, the 5-HT1A receptors, and the 5-HT2A receptors, respectively

    Positron Emission Tomography Radiochemistry: Improved Methodology and a Novel PET Imaging Agent for the Dopamine D3 Receptor

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    Positron Emission Tomography (PET) imaging is a powerful non-invasive imaging modality used to quantify biochemical processes in vivo. The work described in this thesis encompasses two aspects of PET radiochemistry: (1) the development of new chemical methodology applying the principles of green chemistry to late-stage 18F-fluorination, and (2) developing a PET radioligand for the dopamine D3 receptor. (1) As a way to simplify FDA mandated quality control and residual solvent analysis prior to the release of PET radiopharmaceuticals we sought to apply the principles of green chemistry to 18F-radiochemistry. Green radiochemistry in late-stage stage 18F-fluorination with [18F]KF employs ethanol and ethanol/water mixtures as the only solvents used throughout the entire radiopharmaceutical production: azeotropic drying, nucleophilic fluorination, purification and formulation. This work developed a method in which using ethanol in place of acetonitrile is a viable option for azeotroptic drying of [18F]fluoride. Furthermore, it demonstrated that aliphatic 18F-fluorination reactions can proceed in an optimized 15% water, 85% ethanol mixture. Proof of concept was demonstrated through the synthesis of clinically relevant PET radiopharmaceuticals. The new methodology described challenges the long held belief that nucleophilic fluorination reactions cannot proceed in protic solvents or aqueous media, and encourages further exploration into the utility of green fluorine chemistry. (2) Dopamine is a monoamine neurotransmitter in the Central Nervous System (CNS), and its signaling is mediated by pre-synaptic and post-synaptic dopamine receptors. These are D1-like (D1 and D5) and D2-like (D2, D3, D4) receptors, which are G protein-coupled receptors (GPCRs) and are classified based on these two subfamilies due to sequence homology and pharmacological similarities. The D3 receptor is implicated in many neurological and neurodegenerative diseases and disorders, including substance abuse, depression and schizophrenia. Significant efforts have gone into developing ligands as therapeutics, and as positron emission tomography (PET) diagnostics at this receptor. Significant challenges to this pursuit include the high sequence homology between the D2 and D3 receptors, and the physicochemical properties associated with BBB permeability and in vivo stability, while maintaining selectivity at this receptor. The synthesis and initial evaluation of a series of new carbon-11 and fluorine-18 PET radioligands for the D3 receptor is described. These radioligands are based on known ligands with good selectivity for the D3 receptor (pramipexole and BP897). Due to stability issues when using pramipexole-containing scaffolds, BP897 derivatives containing a naphthamide moiety were developed and evaluated in vitro using rad brain autoradiography, and in vivo using rodent microPET imaging. A carbon-11 D3-selective compound was synthesized and identified as a promising candidate, owing to its high affinity for D3 receptors, selectivity for D3 over D2 receptors, and good blood-brain barrier (BBB) penetration. Further evaluation and optimization of the scaffold is merited prior to clinical translation in the future.PHDMedicinal ChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/144039/1/mestewar_1.pd

    Frontal lobe epilepsy, sleep and parasomnias.

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    A close relationship exists between sleep and epilepsy. While many forms of epilepsy may be influenced by the sleep-wake cycle, this phenomenon is particularly evident in frontal lobe epilepsy where affected individuals may experience seizures exclusively during sleep (nocturnal frontal lobe epilepsy, NFLE). In this thesis, three aspects of the relationship between sleep and frontal lobe epilepsy are examined. Firstly, serotonergic neurotransmission across the human sleep-wake cycle was studied using the novel PET ligand l8F-MPPF, a serotonergic 5HT)A receptor radioligand sensitive to endogenous serotonin release. Fourteen individuals with narcolepsy underwent 18F-MPPF PET scans during sleep and wakefulness. The study demonstrated a 13% increase in 18F-MPPF binding potential (p<0.01) during sleep, indicating a reduction in serotoninergic neurotransmission, in line with existing animal data. Secondly, the characterisation of benign, non-epileptic parasomnias and their distinction from nocturnal frontal lobe seizures was addressed in two studies. The first comprised an analysis of the historical features of these conditions, and included the development and validation of a clinical scale for the diagnosis of nocturnal events. The second comprised a detailed semiological analysis of a series of parasomnias recorded on video-EEG monitoring, and a statistical comparison with seizures in NFLE. Although similarities between NFLE and parasomnias were observed, the results provide an evidence base for the confident distinction of these disorders. Finally, the familial form of NFLE (autosomal dominant nocturnal frontal lobe epilepsy, ADNFLE) is associated with mutations in genes for nicotinic acetylcholine receptor subunits, but recognised mutations account for only a minority of reported cases. The last study presented here is a clinical and genetic analysis of two large families with an unusually severe ADNFLE phenotype. Affected individuals had refractory epilepsy and increased rates of mental retardation and psychiatric disorders and, in one family, linkage studies suggest a previously unrecognised underlying mechanism
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