Molecular imaging of σ receptors: synthesis and evaluation of the potent σ₁ selective radioligand [¹⁸F]fluspidine

Neuroimaging of σ₁ receptors in the human brain has been proposed for the investigation of the pathophysiology of neurodegenerative and psychiatric diseases. However, there is a lack of suitable ¹⁸F-labelled PET radioligands for that purpose.

The selective σ₁ receptor ligand [¹⁸F]fluspidine (1′-benzyl-3-(2-[¹⁸F]fluoroethyl)-3H-spiro[[2]benzofuran-1,4′-piperidine]) was synthesized by nucleophilic ¹⁸F⁻ substitution of the tosyl precursor. In vitro receptor binding affinity and selectivity were assessed by radioligand competition in tissue homogenate and autoradiographic approaches. In female CD-1 mice, in vivo properties of [¹⁸F]fluspidine were evaluated by ex vivo brain section imaging and organ distribution of intravenously administered radiotracer. Target specificity was validated by organ distribution of [¹⁸F]fluspidine after treatment with 1 mg/kg i.p. of the σ receptor antagonist haloperidol or the emopamil binding protein (EBP) inhibitor tamoxifen. In vitro metabolic stability and in vivo metabolism were investigated by LC-MSⁿ and radio-HPLC analysis.

[¹⁸F]Fluspidine was obtained with a radiochemical yield of 35–45%, a radiochemical purity of ≥ 99.6% and a specific activity of 150–350 GBq/μmol (n = 6) within a total synthesis time of 90–120 min. In vitro, fluspidine bound specifically and with high affinity to σ₁ receptors (K _i = 0.59 nM). In mice, [¹⁸F]fluspidine rapidly accumulated in brain with uptake values of 3.9 and 4.7%ID/g and brain to blood ratios of 7 and 13 at 5 and 30 min after intravenous application of the radiotracer, respectively. By ex vivo autoradiography of brain slices, resemblance between binding site occupancy of [¹⁸F]fluspidine and the expression of σ₁ receptors was shown. The radiotracer uptake in the brain as well as in peripheral σ₁ receptor expressing organs was significantly inhibited by haloperidol but not by tamoxifen. Incubation with rat liver microsomes led to a fast biotransformation of fluspidine. After an incubation period of 30 min only 13% of the parent compound was left. Seven metabolites were identified by HPLC-UV and LC-MSⁿ techniques. However, [¹⁸F]fluspidine showed a higher metabolic stability in vivo. In plasma samples ∼ 94% of parent compound remained at 30 min and ∼ 67% at 60 min post-injection. Only one major radiometabolite was detected. None of the radiometabolites crossed the blood-brain barrier.

[¹⁸F]Fluspidine demonstrated favourable target affinity and specificity as well as metabolic stability both in vitro and in animal experiments. The in vivo properties of [¹⁸F]fluspidine offer a high potential of this radiotracer for neuroimaging and quantitation of σ₁ receptors in vivo.