Synthesis and Evaluation of ¹¹C-LY2795050 as a κ-Opioid Receptor Antagonist Radiotracer for PET Imaging

Abstract

Kappa opioid receptors (KOR) are believed to be involved in the pathophysiology of depression, anxiety disorders, drug abuse and alcoholism. To date, only one tracer, the kappa opioid receptor agonist [11C]GR103545, has been reported to be able to image KOR in primates. The goal of the present study was to synthesize the selective KOR antagonist [11C]LY2795050 and evaluate its potential as a PET tracer to image KOR in vivo. METHODS In vitro binding affinity of LY2795050 was measured in radioligand competition binding assays. Ex vivo experiments were conducted using microdosing of the unlabelled ligand in Sprague-Dawley rats, as well as wild-type and KOR knock-out mice, to assess the ligand’s potential as a tracer candidate. Imaging experiments with [11C]LY2795050 in monkeys were carried out on the Focus-220 PET scanner with arterial blood input function measurement. Binding parameters were determined with kinetic modeling analysis. RESULTS LY2795050 displays full antagonist activity and high binding affinity and selectivity for KOR. Microdosing studies in rodents and ex vivo analysis of tissue concentrations with LC/MS/MS identified LY2795050 as an appropriate tracer candidate able to provide specific binding signals in vivo. [11C]LY2795050 was prepared in an average yield of 12% and >99% radiochemical purity. In rhesus monkeys, [11C]LY2795050 displayed a moderate rate of peripheral metabolism, with ∼40% of parent compound remaining at 30 min postinjection. In the brain, [11C]LY2795050 displayed fast uptake kinetics (regional activity peak times < 20 min) and an uptake pattern consistent with the distribution of KOR in primates. Pretreatment with naloxone (1 mg/kg, iv) resulted in a uniform distribution of radioactivity. Further, specific binding of [11C]LY2795050 was reduced by the selective KOR antagonist LY2456302 in a dose-dependent manner. CONCLUSION [11C]LY2795050 displayed favorable pharmacokinetic properties and binding profiles in vivo, and therefore is a suitable ligand for imaging the KOR in primates. This newly developed KOR antagonist tracer has since been advanced to PET imaging of KOR in humans and constitutes the first successful KOR antagonist radiotracer.

Determination of the In Vivo Selectivity of a New κ-Opioid Receptor Antagonist PET Tracer 11C-LY2795050 in the Rhesus Monkey

“…As reported previously, the in vitro binding affinities ( K i ) of [ 11 C]LY2795050 were 0.72 and 25.8 nM, respectively, for KOR and MOR (16) at room temperature (22 °C). In our present work, in non-human primates, in vivo , ED 50 MOR was found to be 119 μg/kg based on occupancy studies with [ 11 C]carfentanil PET and blocking doses of LY2795050.…”

“…[ 11 C]LY2795050 was synthesized as previously described (16). Both chemical and radiochemical purity of the [ 11 C]LY2795050 final product were >95 %.…”

“…Twenty blood samples (0.5 mL) were drawn at 0.75, 1.5, 2.25, 3, 4, 5, 6, 8, 9, 12, 15, 20, 25, 30, 40, 50, 60, 75, 90, 105, and 120 min after the injection of [ 11 C]LY2795050. Samples drawn at 4, 12, 30, 60 (3.5 mL), and 90 min (1–5 mL) were also analyzed for radioactive metabolites according to published methods (16). Arterial samples were not taken for [ 11 C]carfentanil because there is an accepted reference region (22).…”

“…Our group recently reported a new KOR antagonist radioligand [ 11 C]LY2795050 (16). Radioligand competition assays using cloned human KOR, MOR and DOR indicated [ 11 C]LY2795050 displayed in vitro K i values of 0.72, 25.8, and 153 nM, respectively, for KOR, MOR and DOR.…”

Neuropeptide Receptors in Pain Circuitries: Useful Targets for CNS Imaging with Non-Peptide Ligands Suitable for PET?

“…On the analgesic route of pain circuits, however, not only µ opioid receptors but also κ-receptors appear to be of great relevance not only in relays of the spinothalamic tract conveying signals to higher centers but also as potential modulators of dopamine release in cortical and subcortical brain areas [240]. Most auspicious approach for κ receptor ligands was described in 2003 by Thomas et al [104] [241]. Simultaneously, entrapment in cellular compartments or activation of c-jun terminal kinase 1 (inductor of autophagosome formation) discussed to prolong the action of JDTic is not known for the shorter acting LY2795050 [242].…”