PT - JOURNAL ARTICLE AU - Ahmed Haider AU - Luca Gobbib AU - Jian Ronga AU - Jiahui Chen AU - Tuo Shao AU - Catarina Raposo AU - Michael Honer Honer AU - Hazem Ahmed AU - Uwe Grether Grether AU - Steven Liang AU - Simon Ametamey TI - Evaluation of CB2 PET Radioligand <sup>18</sup>F RoSMA-18-d<sub>6</sub> in Non-Human Primates and Experimental Autoimmune Encephalomyelitis DP - 2021 May 01 TA - Journal of Nuclear Medicine PG - 2--2 VI - 62 IP - supplement 1 4099 - http://jnm.snmjournals.org/content/62/supplement_1/2.short 4100 - http://jnm.snmjournals.org/content/62/supplement_1/2.full SO - J Nucl Med2021 May 01; 62 AB - 2Objectives: The cannabinoid type 2 (CB2) receptor is an essential component of the highly regulated mammalian endocannabinoid system. In the central nervous system (CNS), CB2 receptor upregulation on activated microglial cells has been reported under neuroinflammatory conditions. As such, CB2 receptors have been suggested as a diagnostic target for patients suffering from neurodegeneration - including multiple sclerosis (MS). However, despite substantial efforts to develop a suitable CB2 positron emission tomography (PET) radioligand, a clinically established probe is currently lacking. We have recently identified a 2,5,6-trisubstituted pyridine derivative, codenamed [18F]RoSMA-18-d6, that exhibited an outstanding CB2 specificity and selectivity in rodents. Herein, the utility of [18F]RoSMA-18-d6 as a CB2 PET radioligand was evaluated in non-human primates (NHP). Further, we sought to assess whether a tritiated analog, [3H]RoSMA-18, can be used to detect a CB2 receptor upregulation in autoradiograms of experimental autoimmune encephalomyelitis (EAE), a validated mouse model of multiple sclerosis. Methods: The inhibitory constant (Ki) of RoSMA-18-d6 was determined by competitive radioligand binding assays. [18F]RoSMA-18-d6 was synthesized via nucleophilic substitution from the respective tosylate precursor. Microsome stability assays were performed to assess the stability of [18F]RoSMA-18-d6. In vitro autoradiography with [18F]RoSMA-18-d6 was performed on CB2-positive NHP spleen tissue sections, using the structurally unrelated CB2 ligands, CP55,940 (10 µM) and GW-405,833 (10 µM), as blockers. Similarly, in vitro autoradiography with [18F]RoSMA-18-d6 was carried out on sagittal NHP brain tissue sections under baseline and blockade conditions. PET experiments with [18F]RoSMA-18-d6 were conducted in NHP, thereby assessing time-activity curves in the spleen as well as the brain. To obtain high resolution autoradiograms, the tritiated analog, [3H]RoSMA-18, was synthesized and tested on spleen sections originating from CB2 knock-out mice and respective wild-type controls. Further, [3H]RoSMA-18 was employed to assess CB2 receptor upregulation in an EAE mouse model. Results: Radioligand binding assays revealed a Ki value of 0.6±0.3 nM (n=3). Unlike the non-deuterated version, [18F]RoSMA-18, [18F]RoSMA-18-d6 was stable against microsomal degradation. Further, in vitro autoradiograms unveiled significant specific binding of [18F]RoSMA-18-d6 to the NHP spleen. Of note, CP55,940 and GW-405,833 blockade resulted in a similar extend of signal reduction. As expected, no specific binding was detected in the healthy NHP brain. These in vitro findings were corroborated in PET studies with NHP, whereby specific spleen binding was observed, along with a fast washout from the brain. The tritiated analog, [3H]RoSMA-18, was obtained via tritium enriched Pd-catalyzed hydrogenation of the bis-olefin precursor at ambient temperature in a high radiochemical purity of 98.5%. The virtual lack of [3H]RoSMA-18 binding on CB2 knock-out spleen tissue sections elegantly confirmed the high tracer selectivity. By in vitro autoradiography with [3H]RoSMA-18 an increased radioligand binding was detected in the brain and spinal cords of EAE mice compared to the wild-type controls. Conclusions: [18F]RoSMA-18-d6 was successfully employed for the visualization and quantification of CB2 receptors in non-human primates. Our findings further indicate that [18F]RoSMA-18-d6 can be used to detect CB2 receptor upregulation in experimental MS. Clinical studies are warranted to assess whether CB2-targeted PET has the potential to improve contemporary diagnostic tools for MS patients.