Synthesis, ex vivo evaluation, and radiolabeling of potent 1,5-diphenylpyrrolidin-2-one cannabinoid subtype-1 receptor ligands as candidates for in vivo imaging

Abstract

We have reported that [methyl- (11)C] (3 R,5 R)-5-(3-methoxyphenyl)-3-[(R)-1-phenylethylamino]-1-(4-trifluoromethylphenyl)pyrrolidin-2-one ([(11)C] 8, [(11)C]MePPEP) binds with high selectivity to cannabinoid type-1 (CB 1) receptors in monkey brain in vivo. We now describe the synthesis of 8 and four analogues, namely, the 4-fluorophenyl (16, FMePPEP), 3-fluoromethoxy (20, FMPEP), 3-fluoromethoxy- d 2 (21, FMPEP- d 2), and 3-fluoroethoxy analogues (22, FEPEP), and report their activity in an ex vivo model designed to identify compounds suitable for use as positron emission tomography (PET) ligands. These ligands exhibited high, selective potency at CB 1 receptors in vitro (K b < 1 nM). Each ligand (30 microg/kg, iv) was injected into rats under baseline and pretreatment conditions (3, rimonabant, 10 mg/kg, iv) and quantified at later times in frontal cortex ex vivo with liquid chromatography-mass spectrometry (LC-MS) detection. Maximal ligand uptakes were high (22.6-48.0 ng/g). Under pretreatment, maximal brain uptakes were greatly reduced (6.5-17.3 ng/g). Since each ligand readily entered brain and bound with high selectivity to CB 1 receptors, we then established and here describe methods for producing [(11)C] 8, [(11)C] 16, and [(18)F] 20- 22 in adequate activities for evaluation as candidate PET radioligands in vivo.

Figure 1

Structures of some CB₁ receptor agonists. Asterisks mark positions of radiolabel.

Figure 2

Structures of 3 and candidate CB₁ receptor PET radioligands. Asterisks mark positions of radiolabel.

Figure 3

Time course evaluation in the frontal cerebral cortex of male Sprague Dawley rats after administration (30 μg/kg, i.v.) of 8 (Panel A), 16 (Panel B), 20 (Panel C) and 22 (Panel D) under baseline and pretreated with 3 (10 mg/kg, i.v.) 15 min before ligand injection. Key: (░), baseline; (▓), pretreatment.

Scheme 1

Synthesis of 8, 16 and 20–22). Reagents and conditions: a) 4-Aminobenzotrifluoride, ethyl pyruvate, AcOH; b) conc. HCl, AcOH; c) (R)-(+)-1-phenylethylamine, CH₂Cl₂; d) (R)-1-(4-fluoro-phenyl)-ethylamine, CH₂Cl₂; e) NaBH₃CN, AcOH; f) pyridinium hydrochloride, heat; g) Cs₂CO₃, DMF, fluoromethyl tosylate; h) Cs₂CO₃, DMF, fluoromethyl tosylate-d₂; i) Cs₂CO₃, DMF, F(CH₂)₂Br.

Scheme 2

Radiosynthesis of [¹¹C]8 and [¹¹C]16. Reagents, conditions and yields: a) “Loop”, Precursor (18 or 19), DMF, 0.5M (n-Bu)₄NOH in MeOH, [¹¹C]MeI, rt, ∼ 3 min, RCY = 2.5 ± 1.1% (8, n = 57) and 16.5% ([¹¹C]16, n = 2) decay-corrected from [¹¹C]carbon dioxide.

Scheme 3

Radiosynthesis of [¹⁸F]20–22. Reagents, conditions and yields: a) [¹⁸F]fluoride ion-K 2.2.2-K⁺ complex, solvent (MeCN for CH₂Br_2, CD₂Br₂ and o-DCB for bromo(m)ethyl tosylate), Δ; b) Precursor (18), Cs₂CO₃, DMF, Δ; c) Precursor (18), DMF, (n-Bu)₄NOH in MeOH, decay-corrected RCY’s = 5.92 ± 1.34% ([¹⁸F]20; n = 3), 7.93 ± 2.48% ([¹⁸F]21; n = 6) and 7.92 ± 2.16% ([¹⁸F]22; n = 6) from [¹⁸F]fluoride ion.