Evaluation in vitro and in animals of a new 11C-labeled PET radioligand for metabotropic glutamate receptors 1 in brain

Abstract

Purpose: Two allosteric modulators of the group I metabotropic glutamate receptors (mGluR1 and mGluR5) were evaluated as positron emission tomography (PET) radioligands for mGluR1.

Methods: LY2428703, a full mGluR1 antagonist (IC(50) 8.9 nM) and partial mGluR5 antagonist (IC(50) 118 nM), and LSN2606428, a full mGluR1 and mGluR5 antagonist (IC(50) 35.3 nM and 10.2 nM, respectively) were successfully labeled with (11)C and evaluated as radioligands for mGluR1. The pharmacology of LY2428703 was comprehensively assessed in vitro and in vivo, and its biodistribution was investigated by liquid chromatography-mass spectrometry/mass spectrometry, and by PET imaging in the rat. In contrast, LSN2606428 was only evaluated in vitro; further evaluation was stopped due to its unfavorable pharmacological properties and binding affinity.

Results: (11)C-LY2428703 showed promising characteristics, including: (1) high potency for binding to human mGluR1 (IC(50) 8.9 nM) with no significant affinity for other human mGlu receptors (mGluR2 through mGluR8); (2) binding to brain displaceable by administration of an mGluR1 antagonist; (3) only one major radiometabolite in both plasma and brain, with a negligible brain concentration (with 3.5 % of the total radioactivity in cerebellum) and no receptor affinity; (4) a large specific and displaceable signal in the mGluR1-rich cerebellum with no significant in vivo affinity for mGluR5, as shown by PET studies in rats; and (5) lack of substrate behavior for efflux transporters at the blood-brain barrier, as shown by PET studies conducted in wild-type and knockout mice.

Conclusion: (11)C-LY2428703, a new PET radioligand for mGluR1 quantification, displayed promising characteristics both in vitro and in vivo in rodents.

Figure 1

Reaction schemes and chemical structures of ¹¹C-LSN2606428 (top) and ¹¹C-LY2428703 (bottom).

Figure 2

Time course of cerebellar (●) and cortical (○) tracer levels of LY2428703. The differential uptake between the target-rich cerebellum and the frontal cortex indicates specific binding at all time points.

Figure 3

Receptor occupancy calculations of LY2428703 for mGluR1 and mGluR5. While mGluR1 occupancy increases with increasing doses of LY2428703, little to no mGluR5 occupancy was found up to doses of 60 mg/kg.

Figure 4

¹¹C-LY2428703 PET studies in a rat at baseline (A) and after blocking using LY2212157 (10 mg/kg) (B). The strongest signal at baseline was in the cerebellum (arrow), which disappeared after blockade. The rest of the brain (area inside the black circle) showed a faint uptake at baseline, which disappeared after blockade. The residual activity was non-specific binding. Please note that the two areas of high uptake visible at both baseline and during the blocked study in the anterior part of the head represent nonspecific uptake in the orbital regions.

Figure 5

(A) Baseline (black symbols; n=10) and blocked (LY2212157, 10 mg/kg, white symbols; n=4) curves for cerebellum (●) and forebrain (■) in rats. (B) Forebrain and cerebellum curves of a displacement study in rat (n=1). The displacer (LY2212157, 10 mg/kg) was administered 15 minutes after the radiotracer injection, which rapidly decreased cerebellar activity (○) to the level of forebrain activity (●).

Figure 6

Baseline (black symbols; n=10) and mGluR5 blocked (MTEP, 5 mg/kg, white symbols; n=2) curves for the cerebellum (●) and forebrain (■) in rats. Forebrain uptake was not affected by pretreatment with an mGluR5 antagonist.

Figure 7

Time-activity curves in the cerebellum (●) and the forebrain (■) of wild type mice (black symbols; n=5) and knock-out mice (white symbols) for Pg-p (n=4) (A) and BCRP (n=4) (B). Very similar results were obtained with triple knock-out mice (n=4) (data not shown).