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内容記述 |
Introduction: Monoacylglycerol lipase (MAGL) is a 33 kDa cytosolic serine hydrolase that is widely distributed in the central nervous system and peripheral tissues. Inhibition of MAGL in the brain elevates levels of 2-arachidonoylglycerol and leads to decreased pro-inflammatory prostaglandin. Therefore, MAGL is considered a potential therapeutic target for treating neuropsychiatric disorders, cancer, chronic pain, and inflammation. In this study, we aimed to evaluate a novel class of spirocyclic cyclobutane-lactam scaffolds [18F](R)-6 (codenamed [18F]MAGL-2304) and [18F](S)-6 (codenamed [18F]MAGL-2305) as positron emission tomography (PET)radioligands for imaging of MAGL.Methods: Compounds (R)-6 and (S)-6 and the corresponding precursors 11 were synthesized from commercially available compound 1. The potency and selectivity of (R)-6 and (S)-6 were assessed in human MAGL screening assays. Activity-based protein profiling (ABPP) assays were employed to assess the selectivity of (R)-6 and (S)-6 against other serine hydrolases in mouse brain lysate. In silico properties of compounds (R)-6 and (S)-6 were predicted by ChemDraw 21.0. [18F](R)-6 and [18F](S)-6 were prepared via a Cu-mediated fluorination reaction. In vitro autoradiography experiments were performed on sagittal rat brain, as well as on brain sections from wild-type (WT) and MAGL knockout (KO) mice. PET imaging studies of [18F](S)-6 were performed in Sprague−Dawley (SD) rats. The in vitro stability of [18F](S)-6 was evaluated in serums and liver microsomes from humans and various animal species. Radiometabolite analysis was conducted in the brain and plasma of SD rats. Ex vivo whole-body distribution experiments were carried out in ddY mice at five time points (5, 15, 30, 60 and 120 min) following [18F](S)-6 injection.Results: Compounds (R)-6 and (S)-6 were synthesized with overall yields of 5.1% and 28% over four steps, respectively (Figure 1A). (R)-6 demonstrated an IC50 of 18.57±1.09 nM, while (S)-6 exhibited superior potency with an IC50 values of 1.59±1.08 nM. Both compounds showed no significant inhibition of other lipid hydrolases. LogD values for (R)-6 and (S)-6 were 2.81 and 2.80, respectively, with predicted topological polar surface areas (tPSAs) of 67.87 (Figure 1B). [18F](R)-6 and [18F](S)-6 were synthesized with non-decay corrected radiochemical yields of 13% and 9%, respectively (Figure 1C). In vitro autoradiography demonstrated that [18F](R)-6 and [18F](S)-6 accumulated in the MAGL-rich regions, such as the cortex, hippocampus, striatum, and thalamus. Blocking experiments with KML29 led to a significant reduction of radioactivity (Figure 1D). In MAGL KO mice, [18F](S)-6 uptake was minimal, while wild-type mouse brain showed heterogeneous (Figure 1E). PET imaging study of [18F](S)-6 under baseline conditions exhibited a heterogeneous distribution and the blocking experiments led to moderate reduction in radioactivity uptake (Figure 1F). P-gp inhibition studies suggest that [18F](S)-6 may constitute a P-gp substrate (Figure 1G). [18F](S)-6 exhibited high in vitro stability in serums and liver microsomes as well as formulated in PBS (Figure 1H&1I). In vivo metabolic stability showed the unchanged form of parent [18F](S)-6 constituted 98.6% in the brain and 42.3% in plasma (Figure 1J). Ex vivo biodistribution revealed hepatobiliary and urinary elimination pathways of [18F](S)-6 (Figure 1K).Conclusions: We synthesized and characterized two 18F-labeled MAGL PET ligands, [18F](R)-6 and [18F](S)-6 based on a novel spirocyclic system. (S)-6 (IC50 = 1.6 nM) showed superior binding affinity (IC50 = 1.6 nM) and selectivity than (R)-6. In vitro autoradiography showed [18F](S)-6 demonstrated heterogeneous distribution and specific binding to MAGL-rich brain regions PET imaging studies encompassing P-gp inhibition indicate [18F](S)-6 is subject to P-gp efflux at the BBB. Structural optimization of the spirocyclic MAGL PET ligands is currently underway. |
