Fully Automated Synthesis of Novel TSPO PET Imaging Ligand [18F]Fluoroethyltemazepam.

INTRODUCTION: Benzodiazepines, including temazepam are described as TSPO antagonists. In fact, TSPO was initially described as a peripheral benzodiazepine receptor (PBR) with a secondary binding site for diazepam. TSPO is a potential imaging target of neuroinflammation because there is an amplification of the expression of this receptor. OBJECTIVES: Herein, we developed a novel fluorinated benzodiazepine ligand, [18F]Fluoroethyltemazepam ([18F]F-FETEM), for positron emission tomography (PET) imaging of translocator protein (18 kDa). METHODS: [18F]F-FETEM was radiolabelled with an automated synthesizer via a one-pot procedure. We conducted a [18F]F-aliphatic nucleophilic substitution of a tosylated precursor followed by purification on C18 and Alumina N SPE cartridges. Quality control tests was also carried out. RESULTS: We obtained 2.0-3.0% decay-uncorrected radiochemical activity yield (3.7% decay-corrected) within the whole synthesis time about 33 min. The radiochemical purity of [18F]F-FETEM was over 90% by TLC analysis. CONCLUSIONS: This automated procedure may be used as basis for future production of [18F]F-FETEM for preclinical PET imaging studies.

Synthesis and evaluation of novel potent TSPO PET ligands with 2-phenylpyrazolo[1,5-a]pyrimidin-3-yl acetamide.

Translocator protein (TSPO) expression is closely related with neuroinflammation and neuronal damage which might cause several central nervous system diseases. Herein, a series of TSPO ligands (11a-c and 13a-d) with a 2-phenylpyrazolo[1,5-a]pyrimidin-3-yl acetamide structure were prepared and evaluated via an in vitro binding assay. Most of the novel ligands exhibited a nano-molar affinity for TSPO, which was better than that of DPA-714. Particularly, 11a exhibited a subnano-molar TSPO binding affinity with suitable lipophilicity for in vivo brain studies. After radiolabeling with fluorine-18, [18F]11a was used for a dynamic positron emission tomography (PET) study in a rat LPS-induced neuroinflammation model; the inflammatory lesion was clearly visualized with a superior target-to-background ratio compared to [18F]DPA-714. An immunohistochemical examination of the dissected brains confirmed that the uptake location of [18F]11a in the PET study was consistent with a positively activated microglia region. This study proved that [18F]11a could be employed as a potential PET tracer for detecting neuroinflammation and could give possibility for diagnosis of other diseases, such as cancers related with TSPO expression.

Evaluation of 18F-labeled exendin(9-39) derivatives targeting glucagon-like peptide-1 receptor for pancreatic ß-cell imaging.

ß-cell mass (BCM) is known to be decreased in subjects with type-2 diabetes (T2D). Quantitative analysis for BCM would be useful for understanding how T2D progresses and how BCM affects treatment efficacy and for earlier diagnosis of T2D and development of new therapeutic strategies. However, a noninvasive method to measure BCM has not yet been developed. We developed four 18F-labeled exendin(9-39) derivatives for ß-cell imaging by PET: [18F]FB9-Ex(9-39), [18F]FB12-Ex(9-39), [18F]FB27-Ex(9-39), and [18F]FB40-Ex(9-39). Affinity to the glucagon-like peptide-1 receptor (GLP-1R) was evaluated with dispersed islet cells of ddY mice. Uptake of exendin(9-39) derivatives in the pancreas as well as in other organs was evaluated by a biodistribution study. Small-animal PET study was performed after injecting [18F]FB40-Ex(9-39). FB40-Ex(9-39) showed moderate affinity to the GLP-1R. Among all of the derivatives, [18F]FB40-Ex(9-39) resulted in the highest uptake of radioactivity in the pancreas 30 min after injection. Moreover, it showed significantly less radioactivity accumulated in the liver and kidney, resulting in an overall increase in the pancreas-to-organ ratio. In the PET imaging study, pancreas was visualized at 30 min after injection of [18F]FB40-Ex(9-39). [18F]FB40-Ex(9-39) met the basic requirements for an imaging probe for GLP-1R in pancreatic ß-cells. Further enhancement of pancreatic uptake and specific binding to GLP-1R will lead to a clear visualization of pancreatic ß-cells.

Synthesis of PET probe O6-[(3-[11C]methyl)benzyl]guanine by Pd0-mediated rapid C-[11C]methylation toward imaging DNA repair protein O6-methylguanine-DNA methyltransferase in glioblastoma.

O6-Benzylguanine (O6-BG) is a substrate of O6-methylguanine-DNA methyltransferase (MGMT), which is involved in drug resistance of chemotherapy in the majority of glioblastoma multiform. For clinical diagnosis, it is hoped that the MGMT expression level could be determined by a noninvasive method to understand the detailed biological properties of MGMT-specific tumors. We synthesized 11C-labeled O6-[(3-methyl)benzyl]guanine ([11C]mMeBG) as a positron emission tomography probe. Thus, a mixed amine-protected stannyl precursor, N9-(tert-butoxycarbonyl)-O6-[3-(tributylstannyl)benzyl]-N2-(trifluoroacetyl)guanine, was subjected to rapid C-[11C]methylation under [11C]CH3I/[Pd2(dba)3]/P(o-CH3C6H4)3/CuCl/K2CO3 in NMP, followed by quick deprotection with LiOH/H2O, giving [11C]mMeBG with total radioactivity of 1.34GBq and ≥99% radiochemical and chemical purities.

Simplified one-pot 18F-labeling of biomolecules with in situ generated fluorothiophosphate synthons in high molar activity.

Rationale: Conventional 18F-labeling methods that demand substrate pre-modification or lengthy radiosynthesis procedures have impeded the visualization and translation of numerous biomolecules, as biomarkers or ligands, using modern positron emission tomography techniques in vivo. Moreover, 18F-labeled biomolecules in high molar activity (Am) that are indispensable for sensitive imaging could be only achieved under strict labeling conditions. Methods: Herein, 18F-labeled fluorothiophosphate (FTP) synthons in high Am have been generated rapidly in situ in reaction solutions with < 5% water via nucleophilic substitution by wet [18F]F-, which required minimal processing from cyclotron target water. Results: Various 18F-labeled FTP synthons have been prepared in 30 sec at room temperature with high radiochemical yields > 75% (isolated, non-decay-corrected). FTP synthons with unsaturated hydrocarbon or activated ester group can conjugate with typical small molecules, peptides, proteins, and metallic nanoparticles. 337-517 GBq µmol-1 Am has been achieved for 18F-labeled c(RGDyK) peptide using an automatic module with 37-74 GBq initial activity. Conclusion: The combination of high 18F-fluorination efficiency of FTP synthons and following mild conjugation condition provides a universal simplified one-pot 18F-labeling method for broad unmodified biomolecular substrates.

18F-Labelled pyrrolopyrimidines reveal brain leucine-rich repeat kinase 2 expression implicated in Parkinson's disease.

18F-Labelled pyrrolopyrimidines were synthesized and evaluated as positron emission tomography (PET) probes to determine leucine-rich repeat kinase 2 (LRRK2) expression in the brain. With pyrrolopyrimidine derivative PF-06447475 as the lead compound, two in vivo-stable 18F-labelled pyrrolopyrimidines ([18F]1 and [18F]2) were synthesized automatically at radiochemical yields 8-10% (non-decay-corrected) with molar activities of 0.95 and 0.5 GBq/µmol, respectively. The measured Kd of 6.90 nM for 1 and 14.27 nM for 2 demonstrated high affinities for LRRK2. The LRRK2 G2019S mice had higher uptakes (P < 0.01) of [18F]1 in the olfactory bulb, striatum, and hippocampus than WT mice during microPET/CT imaging, consistent with immunohistology results of LRRK2 distribution. [11C]CFT microPET/CT imaging demonstrated a lower expression of dopamine transporter in LRRK2 G2019S mice. Parkinson's disease-like deficits in dopamine transporter synthesis and cognitive declines were noticed along with LRRK2 expression increase in the olfactory bulb, striatum, and hippocampus. Therefore, 18F-labelled pyrrolopyrimidines can reflect real-time LRRK2 expression changes implicated in Parkinson's disease, which paves the way for LRRK2-related neurodegenerative precise therapy.