[18F]Amylovis as a Potential PET Probe for β-Amyloid Plaque: Synthesis, In Silico, In vitro and In vivo Evaluations

Abstract

Background: Alzheimer's disease (AD) is the most common form of dementia. Neuroimaging methods have widened the horizons for AD diagnosis and therapy. The goals of this work are the synthesis of 2-(3-fluoropropyl)-6-methoxynaphthalene (5) and its [18F]-radiolabeled counterpart ([18F]Amylovis), the in silico and in vitro comparative evaluations of [18F]Amylovis and [11C]Pittsburg compound B (PIB) and the in vivo preclinical evaluation of [18F]Amylovis in transgenic and wild mice.

Methods: Iron-catalysis cross coupling reaction, followed by fluorination and radiofluorination steps were carried out to obtain 5 and 18F-Amylovis. Protein/Aß plaques binding, biodistribution, PET/CT Imaging and immunohistochemical studies were conducted in healthy/transgenic mice.

Results: The synthesis of 5 was successful obtained. Comparative in silico studies predicting that 5 should have affinity to the Aβ-peptide, mainly through π-π interactions. According to a dynamic simulation study the ligand-Aβ peptide complexes are stable in simulation-time (ΔG = -5.31 kcal/mol). [18F]Amylovis was obtained with satisfactory yield, high radiochemical purity and specific activity. The [18F]Amylovis log Poct/PBS value suggests its potential ability for crossing the blood brain barrier (BBB). According to in vitro assays, [18F]Amylovis has an adequate stability in time. Higher affinity to Aβ plaques were found for [18F]Amylovis (Kd 0.16 nmol/L) than PIB (Kd 8.86 nmol/L) in brain serial sections of 3xTg-AD mice. Biodistribution in healthy mice showed that [18F]Amylovis crosses the BBB with rapid uptake (7 %ID/g at 5 min) and good washout (0.11±0.03 %ID/g at 60 min). Comparative PET dynamic studies of [18F]Amylovis in healthy and transgenic APPSwe/PS1dE9 mice, revealed a significant high uptake in the mice model.

Conclusion: The in silico, in vitro and in vivo results justify that [18F]Amylovis should be studied as a promissory PET imaging agent to detect the presence of Aβ senile plaques.

Keywords: Alzheimer's disease diagnosis; docking and dynamic simulations; fluorine-18; iron cross-coupling reaction; positron emission tomography; β-amyloid probe..

Scheme 1

General procedure for the synthesis of 2-(3-fluoropropyl)-6-methoxynaphthalene (5) and [¹⁸F]Amylovis. All reported yields correspond to purified products.

Fig. (1)

A: Flexible docking of 5 (at 5 Å distance): the π-π hydrophobic interactions were located between the naphthyl ring of 5 and the phenyl ring of the F19 and F20 residues. B: Flexible docking of PIB (at 5 Å distance): the H-bond was located between nitrogen atom of the benzothiazole ring of PIB and hydrogen atom of -NH₂ group of K16, and also a π-π hydrophobic interaction was located between the phenyl rings of PIB and F19.

Fig. (2)

Molecular Dynamics (MD) study II of 2-(3-fluoropropyl)-6-methoxynaphthalene (5). A: Evolution of different conformations of the ligand-Aβ_1-42 peptide complex (at 10 Å) during different times. B: Root Mean Square Deviation (RMSD) of 5 position with respect to the Aβ peptide. C: Van der Waals (red) and electrostatic (green) energies for ligand-Aβ peptide complex at each time interval during MD simulations.

Fig. (3)

HPLC chromatograms of 5 (A) and [¹⁸F]Amylovis (B) after purification with C18 light-SPE cartridge, retention times: 8.5 and 8.7 min, respectively, and a purity > 99% for both compounds. Chromatographic conditions: Phenomenex Luna C18 (250 x 4.6 mm, 5 µm) column; mobile phase: AcN:H₂O (75:25); flow rate: 1 mL/min.

Fig. (4)

Biodistribution of [¹⁸F]Amylovis in healthy Balb/C male mice (10-12 week-old and 28.5±2.0 g). %ID/g (percentage injected dose per gram tissue).

Fig. (5)

Comparative standardized uptake value relative (SUVr) curves (cortex/cerebellum) of [¹⁸F]Amylovis in healthy (control) vs APPSwe/PS1dE9 mice up to 130 min. Each point represents mean values ± standard deviation (n=4).

Fig. (6)

I. Comparison of standardized uptake values in regard to the brain stem, SUVr, of [¹⁸F]Amylovis in some brain regions between control healthy and APPSwe/PS1dE9 mice (n=3). II. Representative microphotograph of the prefrontal cortex (A, F), hippocampus (B, G), striatum (C, H), thalamus (D, I) and cerebellum (E, J). Set of images A-E corresponds to healthy animals and F-J to transgenic APPSwe/PS1dE9 mice (scale bar = 400 µm).