Synthesis and Biological Evaluation of Enantiomerically Pure (R)- and (S)-[18F]OF-NB1 for Imaging the GluN2B Subunit-Containing NMDA Receptors.

GluN2B subunit-containing N-methyl-d-aspartate (NMDA) receptors have been implicated in various neurological disorders. Nonetheless, a validated fluorine-18 labeled positron emission tomography (PET) ligand for GluN2B imaging in the living human brain is currently lacking. The aim of this study was to develop a novel synthetic approach that allows an enantiomerically pure radiosynthesis of the previously reported PET radioligands (R)-[18F]OF-NB1 and (S)-[18F]OF-NB1 as well as to assess their in vitro and in vivo performance characteristics for imaging the GluN2B subunit-containing NMDA receptor in rodents. A novel synthetic approach was successfully developed, which allows for the enantiomerically pure radiosynthesis of (R)-[18F]OF-NB1 and (S)-[18F]OF-NB1 and the translation of the probe to the clinic. While both enantiomers were selective over sigma2 receptors in vitro and in vivo, (R)-[18F]OF-NB1 showed superior GluN2B subunit specificity by in vitro autoradiography and higher volumes of distribution in the rodent brain by small animal PET studies.

Evaluation of [18F]RoSMA-18-d6 as a CB2 PET Radioligand in Nonhuman Primates.

The cannabinoid type 2 receptor (CB2) has been implicated in a variety of central and peripheral inflammatory diseases, prompting significant interest in the development of CB2-targeted diagnostic and therapeutic agents. A validated positron emission tomography (PET) radioligand for imaging CB2 in the living human brain as well as in peripheral tissues is currently lacking. As part of our research program, we have recently identified the trisubstituted pyridine, [18F]RoSMA-18-d6, which proved to be highly suitable for in vitro and in vivo mapping of CB2 in rodents. The aim of this study was to assess the performance characteristics of [18F]RoSMA-18-d6 in nonhuman primates (NHPs) to pave the way for clinical translation. [18F]RoSMA-18-d6 was synthesized from the respective tosylate precursor according to previously reported procedures. In vitro autoradiograms with NHP spleen tissue sections revealed a high binding of [18F]RoSMA-18-d6 to the CB2-rich NHP spleen, which was significantly blocked by coincubation with the commercially available CB2 ligand, GW405833 (10 µM). In contrast, no specific binding was observed by in vitro autoradiography with NHP brain sections, which was in agreement with the notion of a CB2-deficient healthy mammalian brain. In vitro findings were corroborated by PET imaging experiments in NHPs, where [18F]RoSMA-18-d6 uptake in the spleen was dose-dependently attenuated with 1 and 5 mg/kg GW405833, while no specific brain signal was observed. Remarkably, we observed tracer uptake and retention in the NHP spinal cord, which was reduced by GW405833 blockade, pointing toward a potential utility of [18F]RoSMA-18-d6 in probing CB2-expressing cells in the bone marrow. If these observations are substantiated in NHP models of enhanced leukocyte proliferation in the bone marrow, [18F]RoSMA-18-d6 may serve as a valuable marker for hematopoietic activity in various pathologies. In conclusion, [18F]RoSMA-18-d6 proved to be a suitable PET radioligand for imaging CB2 in NHPs, supporting its translation to humans.

Synthesis and Biological Evaluation of Enantiomerically Pure (R)- and (S)-[18F]OF-NB1 for Imaging the GluN2B Subunit-Containing NMDA receptors.

GluN2B subunit-containing N-methyl-d-aspartate (NMDA) receptors have been implicated in various neurological disorders. Nonetheless, a validated fluorine-18 labeled positron emission tomography (PET) ligand for GluN2B imaging in the living human brain is currently lacking. As part of our PET ligand development program, we have recently reported on the preclinical evaluation of [18F]OF-NB1 - a GluN2B PET ligand with promising attributes for potential clinical translation. However, the further development of [18F]OF-NB1 is currently precluded by major limitations in the radiolabeling procedure. These limitations include the use of highly corrosive reactants and racemization during the radiosynthesis. As such, the aim of this study was to develop a synthetic approach that allows an enantiomerically pure radiosynthesis of (R)-[18F]OF-NB1 and (S)-[18F]OF-NB1, as well as to assess their in vitro and in vivo performance characteristics for imaging the GluN2B subunit-containing NMDA receptor in rodents. A two-step radiosynthesis involving radiofluorination of the boronic acid pinacol ester, followed by coupling to the 3-benzazepine core structure via reductive amination was employed. The new synthetic approach yielded enantiomerically pure (R)-[18F]OF-NB1 and (S)-[18F]OF-NB1, while concurrently circumventing the use of corrosive reactants. In vitro autoradiograms with mouse and rat brain sections revealed a higher selectivity of (R)-[18F]OF-NB1 over (S)-[18F]OFNB1 for GluN2B-rich brain regions. In concert with these observations, blockade studies with commercially available GluN2B antagonist, CP101606, showed a significant signal reduction, which was more pronounced for (R)-[18F]OF-NB1 than for (S)-[18F]OF-NB1. Conversely, blockade experiments with sigma2 ligand, FA10, did not result in a significant reduction of tracer binding for both enantiomers. PET imaging experiments with CD1 mice revealed a higher brain uptake and retention for (R)-[18F]OF-NB1, as assessed by visual inspection and volumes of distribution from Logan graphical analyses. In vivo blocking experiments with sigma2 ligand, FA10, did not result in a significant reduction of the brain signal for both enantiomers, thus corroborating the selectivity over sigma2 receptors. In conclusion, we have developed a novel synthetic approach that is suitable for upscale to human use and allows the enantiomerically pure radiosynthesis of (R)-[18F]OF-NB1 and (S)-[18F]OF-NB1. While both enantiomers were selective over sigma2 receptors in vitro and in vivo, (R)-[18F]OF-NB1 showed superior GluN2B subunit specificity by in vitro autoradiography and higher volumes of distribution in small animal PET studies.