Synthesis and characterization of a new Positron emission tomography probe for orexin 2 receptors neuroimaging

Abstract

The orexin receptors (OXRs) have been involved in multiple physiological and neuropsychiatric functions. Identification of PET imaging probes specifically targeting OXRs enables us to better understand the OX system. Seltorexant (JNJ-42847922) is a potent OX₂R antagonist with the potential to be an OX₂R PET imaging probe. Here, we describe the synthesis and characterization of [¹⁸F]Seltorexant as an OX₂R PET probe. The ex vivo autoradiography studies indicated the good binding specificity of [¹⁸F]Seltorexant. In vivo PET imaging of [¹⁸F]Seltorexant in rodents showed suitable BBB penetration with the highest brain uptake of %ID/cc = 3.4 at 2 min post-injection in mice. The regional brain biodistribution analysis and blocking studies showed that [¹⁸F]Seltorexant had good binding selectivity and specificity. However, pretreatment with unlabelled Seltorexant and P-gp competitor CsA observed significantly increased brain uptake of [¹⁸F]Seltorexant, indicating [¹⁸F]Seltorexant could interact P-gp at the blood-brain barrier. Our findings demonstrated that [¹⁸F]Seltorexant is a potential brain OX₂R PET imaging probe, which paves the way for new OX₂R PET probes development and OX system investigation.

Keywords: Imaging; Orexin receptors; PET; Radiotracer.

Figure 1.

Chemical structures and binding affinities of representative dual orexin receptor antagonists (DORA) and selective orexin 1/2 receptor antagonists.

Figure 2.

The pharmacological and physicochemical properties of Seltorexant. ^acLogP value of Seltorexant is calculated by ChemBioDraw 16.0; Log D values were quantified in n-octanol/phosphate buffer (pH 7.4) by the shake-flask method. ^bLetavic et al. reported the in vitro binding affinity of Seltorexant. ^cBonaventure et al. reported the pharmacokinetics of Seltorexant; single oral administration (p.o.) of Seltorexant at 30 mg/kg showed high concentration in rat brain.

Figure 3.

(A) Representative in vitro autoradiographic images of mice brains (sagittal); (B) the relative radioactive uptake in baseline and blocking. The baseline mice brain sections were treated with [¹⁸F]Seltorexant only, and the blocking mice brain sections were co-incubation with OX₂R antagonist compound 30 (10 μM). The gray value data were expressed as mean ± SD, n = 4; Asterisks indicate statistical significance. *p < 0.05, **p ≤ 0.01, and ***p ≤ 0.001.

Figure 4.

(A) Representative baseline and blocking PET/CT images of [¹⁸F]Seltorexant focus on the mice brain (0–60 min, the baseline is shown as horizontal, coronal, and sagittal plane; the blockings are shown as a sagittal plane); (B) the baseline and blocking time-activity curves of [¹⁸F]Seltorexant in the mice whole brain; (C) Time-radioactivity curves for [¹⁸F]Seltorexant in the hippocampus, cerebellum, thalamus, cortex, and striatum (n = 3); (D) the normalized baseline and blocking time-activity curves [¹⁸F]Seltorexant (normalized the brain uptake curves with the highest radioactivity in the blood at each time point).

Figure 5.

The representative baseline and Cyclosporin A pretreatment PET/CT images with [¹⁸F]Seltorexant focused on mice brain and time-activity curves of [¹⁸F]Seltorexant in the whole brain of mice.

Scheme 1.

Synthesis of precursor 4. Reagents and conditions: (i) NaNO₂, KI, in H₂O, 90 °C, 16 h; (ii) 2H-1,2,3-triazole, N1,N2-Dimethylethane-1,2-diamine, K₂CO₃, CuI, THF, 90 °C, 16 h; (iii) 2-(4,6-dimethylpyrimidin-2-yl)octahydropyrrolo[3,4-c]pyrrole, DIPEA, HATU, DMF, room temperature, 16 h.

Scheme 2.

Radiosynthesis of [¹⁸F]Seltorexant. Radiolabeling condition: 4 (precursor, 3.0 mg), DMSO, K₂₂₂, K₂CO₃, [¹⁸F]F⁻, 150°C, 20 min.