Comparison of in Silico, Electrochemical, in Vitro and in Vivo Metabolism of a Homologous Series of (Radio)fluorinated σ₁ Receptor Ligands Designed for Positron Emission Tomography

Abstract

The imaging of σ receptors in the brain by fluorinated radiotracers will be used for the validation of σ receptors as drug targets as well as for differential diagnosis of diseases in the central nervous system. The biotransformation of four homologous fluorinated PET tracers 1'-benzyl-3-(ω-fluoromethyl to ω-fluorobutyl)-3H-spiro[2]benzofuran-1,4'-piperidine] ([ F]1-4) was investigated. In silico studies using fast metabolizer (FAME) software, electrochemical oxidations, in vitro studies with rat liver microsomes, and in vivo metabolism studies after application of the PET tracers [ F]1-4 to mice were performed. Combined liquid chromatography and mass spectrometry (HPLC-MS) analysis allowed structural identification of non-radioactive metabolites. Radio-HPLC and radio-TLC provided information about the presence of unchanged parent radiotracers and their radiometabolites. Radiometabolites were not found in the brain after application of [ F]2-4, but liver, plasma, and urine samples contained several radiometabolites. Less than 2 % of the injected dose of [ F]4 reached the brain, rendering [ F]4 less appropriate as a PET tracer than [ F]2 and [ F]3. Compounds [ F]2 and [ F]3 possess the most promising properties for imaging of σ receptors in the brain. High σ affinity (K =0.59 nm), low lipophilicity (logD =2.57), high brain penetration (4.6 % of injected dose after 30 min), and the absence of radiometabolites in the brain favor the fluoroethyl derivative [ F]2 slightly over the fluoropropyl derivative [ F]3 for human use.

In vitro and in vivo Human Metabolism of (S)-[18F]Fluspidine – A Radioligand for Imaging σ1 Receptors With Positron Emission Tomography (PET)

“…The metabolic stability of rac - 1 , ( S )- 1 , and ( R )- 1 have previously been studied in vitro using RLM in presence of NADPH. After 30 min, rac - 1 showed the lowest stability (∼13%) among the fluoroalkyl homologs tested (Wiese et al, 2016), whereas 73% of intact ( S )- 1 was still present after the same incubation time (Holl et al, 2013). Compared with our results from HLM (14%) this might suggest that ( S )- 1 has higher stability in rats than in human.…”

“…Metabolism rates have been reported for rac -[ 18 F] 1 or ( S )-[ 18 F] 1 in mice, pigs, and monkeys. In mouse plasma the fraction of unchanged rac -[ 18 F] 1 was 89 ± 3% at 30 min post injection (Wiese et al, 2016). For ( S )-[ 18 F] 1 it was shown that in plasma of piglets 37% of the radioligand remained unchanged at 30 min post injection (Brust et al, 2014a).…”

“…Deduced from the retention behaviors in the radio-HPLC, no radiometabolite appeared to have a higher lipophilicity than ( S )-[ 18 F] 1 . Taking into account that in mouse brain 98% of rac -[ 18 F] 1 remained unchanged at 60 min post injection (Wiese et al, 2016), the absence of radiometabolites in human brain is highly likely as well.…”

“…In a detailed study the metabolism of racemic fluspidine (rac- 1 ) as well as its fluoroalkyl homologs and the corresponding 18 F-radioligands have been investigated covering incubations with rat liver microsomes (RLM) and in vivo studies in mice, respectively (Wiese et al, 2016). Similarly, enantiomerically pure ( S )- 1 as well as ( R )- 1 have been investigated in vitro and structural elucidation with LC-MS has been conducted for metabolites formed by incubations with RLM under conditions for oxygenation (Holl et al, 2013).…”

Bridging from Brain to Tumor Imaging: (S)-(−)- and (R)-(+)-[18F]Fluspidine for Investigation of Sigma-1 Receptors in Tumor-Bearing Mice

“…In a subsequent pilot study we performed PET imaging with [ 18 F]fluspidine in orthotopically implanted brain tumors. In general, [ 18 F]fluspidine has been shown to be suitable for brain imaging [ 31 , 33 , 52 ] and thus, we hypothesize that these radioligands are suitable for imaging of Sig1R expressing brain tumors. Preliminary data from these experiments revealed a tumor-to-background SUV ratio (TBR) of >1 for early time points and thus visualization of the tumor in the mouse brain appeared to be feasible.…”