Monoacylglycerol lipase (MAGL) is one of the key enzymes in the endocannabinoid system. Inhibition of MAGL has been proposed as an attractive approach for the treatment of various diseases. In this study, we designed and successfully synthesized two series of piperazinyl pyrrolidin-2-one derivatives as novel reversible MAGL inhibitors. (R)-[(18)F]13 was identified through the preliminary evaluation of two carbon-11-labeled racemic structures [(11)C]11 and [(11)C]16. In dynamic positron-emission tomography (PET) scans, (R)-[(18)F]13 showed a heterogeneous distribution and matched the MAGL expression pattern in the mouse brain. High brain uptake and brain-to-blood ratio were achieved by (R)-[(18)F]13 in comparison with previously reported reversible MAGL PET radiotracers. Target occupancy studies with a therapeutic MAGL inhibitor revealed a dose-dependent reduction of (R)-[(18)F]13 accumulation in the mouse brain. These findings indicate that (R)-[(18)F]13 ([(18)F]YH149) is a highly promising PET probe for visualizing MAGL non-invasively in vivo and holds great potential to support drug development.
Objectives: Monoacylglycerol lipase (MAGL) is the key enzyme for degradation of the most abundant brain endocannabinoid, 2-ara-chidonoylglycerol (2-AG). It is considered as an attractive drug target for oncological and psychiatric diseases [1]. During the last decade, several irreversible MAGL PET ligands have been disclosed, however, reversible MAGL radiotracers with sufficient brain penetration are barely reported. To the best of our knowledge, [18F]T-401 is the only available reversible MAGL tracer with moderate brain uptake in mice. In this study, we synthesized a series of reversible MAGL inhibitors based on the piperazinyl pyrrolidin-2-one core structure [2,3]. The most potent compounds were radiolabeled with either C-11 or F-18. Their utilities as PET imaging agents were evaluated.
Methods: Novel series of piperazinyl pyrrolidin-2-one derivatives were designed and prepared via multi-step synthetic approaches. Their IC 50 values towards MAGL were determined by tracing the hydrolysis of natural substrate 2-AG. The most potent ligands YH132 and YH135 were labeled with C-11, and YH149 was labeled with F-18. In vitro autoradiography and PET imaging were performed in MAGL knockout and wild-type mice to evaluate their in vitro and in vivo specificity. [18F]YH149 was further evaluated in ex vivo biodistribution and drug-occupancy studies to assess its utility for imaging MAGL in the rodent brain.
Results: IC50 values of the novel MAGL inhibitors ranged from 3 nM to 10.7 M. YH132 (IC50 = 6 nM),YH135 (IC50 = 18 nM), andYH149 (IC50 = 12 nM) that possess IC 50 values in a nano-molar range for human MAGL were further evaluated as radioligands. Both [11C]YH132 and [11C]YH135 showed high accumulation in MAGL-rich brain regions in in vitro autoradiographic studies. In PET studies using MAGL knock-out and wild-type mice, [11C]YH132 revealed higher brain permeability and specificity than [11C]YH135. Therefore, [18F]YH149, the F-18 labeled analog of YH132, was synthesized via copper-mediated fluorination. [18F]YH149 demonstrated excellent in vitro and in vivo specificity towards MAGL. The averaged PET image (12.5-52.5 min p.i.) was in high accordance with MAGL expression pattern in the mouse brain. In ex vivo biodistribution in mice, [18F]YH149 exhibited significantly higher brain uptake than the benchmark radiotracer [18F]T-401 at 30 min post-injection [2]. Blocking effects were observed in the brain and MAGL-expressive peripheral organs using PF-06795071, an irreversible MAGL inhibitor. Furthermore, the drug occupancy study indicated that [18F]YH149 can be used to determine the D50 value of PF-06795071 in the mouse brain.
Conclusions: The fluorinated PET probe, [18F]YH149, revealed high in vitro/in vivo specificity towards MAGL and increased brain permeability compared to the previously reported reversible MAGL PET radiotracers. Preliminary results indicate that [18F]YH149 is a very promising PET tracer for imaging MAGL and capable to evaluate target occupancy in vivo
