PET Imaging of the Adenosine A2A Receptor in the Rotenone-Based Mouse Model of Parkinson's Disease with [18F]FESCH Synthesized by a Simplified Two-Step One-Pot Radiolabeling Strategy

Abstract

The adenosine A_2A receptor (A_2AR) is regarded as a particularly appropriate target for non-dopaminergic treatment of Parkinson's disease (PD). An increased A_2AR availability has been found in the human striatum at early stages of PD and in patients with PD and dyskinesias. The aim of this small animal positron emission tomography/magnetic resonance (PET/MR) imaging study was to investigate whether rotenone-treated mice reflect the aspect of striatal A_2AR upregulation in PD. For that purpose, we selected the known A_2AR-specific radiotracer [¹⁸F]FESCH and developed a simplified two-step one-pot radiosynthesis. PET images showed a high uptake of [¹⁸F]FESCH in the mouse striatum. Concomitantly, metabolism studies with [¹⁸F]FESCH revealed the presence of a brain-penetrant radiometabolite. In rotenone-treated mice, a slightly higher striatal A_2AR binding of [¹⁸F]FESCH was found. Nonetheless, the correlation between the increased A_2AR levels within the proposed PD animal model remains to be further investigated.

Keywords: PET imaging; Parkinson’s disease; [18F]FESCH; adenosine A2A receptor; rotenone-based mouse model; two-step one-pot radiosynthesis.

Figure 1

Molecular structures of the herein discussed radiotracers for positron emission tomography (PET) imaging of the adenosine A_2A receptor (A_2AR).

Scheme 1

Simplified two-step one-pot radiosynthesis of [¹⁸F]FESCH. Reagents and conditions: (First step) 2 mg ethane-1,2-diol bis(4-methylbenzenesulfonate) in 100 µL MeCN, ~ 3 GBq azeotropically dried K⁺/[¹⁸F]F^-/K₂₂₂-carbonate complex in 400 µL MeCN, 90 °C, 10 min; (Second step) 2.5 mg desmethyl SCH442416 pre-treated with 10 µL TBAOH_aq. (40%) in 490 µL MeCN at 90 °C for 10 min, directly added to 2-[¹⁸F]fluoroethyl tosylate, 120 °C, 10 min.

Figure 2

(A) Semi-preparative HPLC profile of the crude reaction mixture for isolation of [¹⁸F]FESCH (column: Reprosil-Pur 120 C18-AQ, 250 × 10 mm, particle size: 10 µm, eluent: 50% MeCN/20 mM NH₄OAc_aq., flow: 7 mL/min). (B) Analytical HPLC profile of the formulated radiotracer [¹⁸F]FESCH spiked with the non-radioactive reference FESCH (column: Reprosil-Pur 120 C18-AQ, 250 × 4.6 mm, particle size: 5 µm; eluent: 26-90-26% MeCN/20 mM NH₄OAc_aq., flow: 1 mL/min).

Figure 3

Representative autoradiographic images of transversal CD-1 mouse brain slices. (A) In vitro distribution of activity after incubation with 0.2 MBq/mL of [¹⁸F]FESCH (TB = total binding). (B) Non-specific binding (NB) of [¹⁸F]FESCH determined in the presence of 1 μM of FESCH or ZM241385, respectively, as blocking agents. (C) Nissl staining, St = Striatum, Cb = Cerebellum.

Figure 4

(A) Representative horizontal PET images of [¹⁸F]FESCH uptake (average 10–30 min) in the brain of healthy CD-1 mice under vehicle (15 min pre-injection of DMSO:Kolliphor^® EL:0.9% NaCl, 1:2:7, 5 µL/g) and blocking conditions (15 min pre-injection of tozadenant 2.5 mg/kg in DMSO:Kolliphor^® EL:0.9% NaCl, 1:2:7, 5 µL/g; red = striatum; yellow = cerebellum). (B) Averaged TACs of [¹⁸F]FESCH for vehicle (n = 4) and tozadenant pre-injected mice (n = 4) with SUVRs for striatum over cerebellum.

Figure 5

Representative in vivo metabolism study of CD-1 mouse plasma and brain samples at 15 min p.i. of [¹⁸F]FESCH (~ 17 MBq): Analytical radio-HPLC profiles of extracted (A) brain and (B) plasma sample (column: Reprosil-Pur 120 C18-AQ, 250 × 4.6 mm, particle size: 5 µm; eluent: 26-90-26% MeCN/20 mM NH₄OAc_aq., flow: 1 mL/min).

Figure 6

(A) Representative horizontal PET images of [¹⁸F]FESCH uptake (average 2–61 min) in the brain of control and rotenone-treated C57BL/6JRj mice (red = striatum; yellow = cerebellum). (B) Averaged TACs of [¹⁸F]FESCH for control (n = 5) and rotenone-treated mice (n = 7) with SUVs for striatum and SUVRs for striatum over cerebellum.

Figure 7

Mean fluorescence intensity of A_2AR staining on the striatum of coronal C57BL/6JRj mouse brain sections: (A) Control vs. rotenone-treated wild-type mice (n = 3, respectively); (B) Young (39 weeks) vs. old (71–76 weeks) A30P transgenic mice (n = 3, respectively).